Volume 2, Issue No. 2- June 2006

Egyptian Hypertension Society The Egyptian Journal of Hypertension

and Cardiovascular Risk

Editor: Hussein H. Rizk, MD

Prof. of Cardiology - Cairo University

Senior Editorial Consultant: M. Mohsen Ibrahim, MD, FACC

Prof. of Cardiology - Cairo University

Senior Associate Editors: Wafaa El-Aroussy, MD Prof. of Cardiology Cairo University

Soliman Gharieb, MD Prof. of Cardiology Cairo University

Editorial Office: Ibtihag A. Hamdy, MD Omar S. Awaad, MD Iglal Ghobashy, MD Omar Y. El-Khashaab, MD M. Khairy Abdel Dayem, MD Omneya Nayel, MD M. Mokhtar Gomaa, MD Sherif El-Tobgy, MD Mohamed Hamed, MD Zeinab Ashour, MD

Assistant Editors: Mona A. Nour, MD

5338-1687ترقيم الدويل الContact Information: Contact person: Attn. Mrs. Rehab Mohamed Address: 1 El-Diwan Street, Garden City, 11519, Cairo-Egypt Tel.: (202) 794-8877, Fax: (202) 794-8879, e-mail: ehs@link.net Web site: www.ehs-egypt.net

Editorial Board

Abdel Moneim Ibrahim, MD Mahmoud Hassanein, MD Adel A. El-Sayed, MD Mohamed Amin Fikry, MD Adel Abdel Aziz El-Sayed, MD Mohamed Bayoume Sammoor, MD Adel Zaki, MD Mohamed El-Ganzoury, MD Ahmed Abdel Moneim, MD Mohamed Fahmy Abdel Azziz, MD Ahmed Amin Fahmy, MD Mohamed Ghoneim, MD Ahmed El-Hawary, MD Mohamed Hassan, MD Ahmed Rashed, MD Mohamed Helmy Abu Zeid, MD Aly Massoud, MD Mohamed Shataa, MD Aly Ramzy, MD Mohamed Sobhy, MD Amal Ayyoub, MD Mohgah Hammad, MD Dawlat Salem, MD Mohsen Abdel Hamid Gadallah, MD Detlev Ganten, MD Nabil Alluba, MD Essam Soliman Khedr, MD Nadia Selim, MD Ezz El-Din El-Sawy, MD Omar El-Khashaab, MD Fathy Maklady, MD Paul Whelton, MD Fawzia El-Demerdash, MD Qaies Abdel Dayem, MD Fouad El-Nawawi, MD Sabry Gohar, MD Hassan Abdel Rahman, MD Salah Naga, MD Heba Mansour, MD Salwa Roushdi, Md Helmy Abu Zeid, MD Samir Abu Zeid, MD Helmy Siragy, MD Samir Helmy Asaad, MD Ikram Sadek, MD Taghreed Gaefar, MD Khaled Sorour, MD Tarek El-Badawy, MD Lawrence Appel, MD Thomas Pickering, MD Maher Fouad, MD Wafeia Eteiba, MD Mahmoud Allam, MD Wagdy Ayad, MD Mahmoud El-Khayaal, MD Yehya Keshk, MD

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Table of Contents 1. Editorial: Ethics of Medical Practice

Mokhtar Gomaa, MD. 5

2. Gender differences in Egyptian elderly with heart failure El Banouby M H, Hamza S A, Gamal A and Adly N N

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3. Hypertension and Bone Mineral Density Among Elderly Patients. Rania M Ezzat, Hala S Sweed, A K Mortagy

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4. Relationship of circulating insulin-like growth factor-1 (IGF-1) and insulin like growth factor binding proteins-1 and 3 (IGFBP-1 and IGFBP-3) with Endothelin-1 (ET-1) in hypertensive patients with and without type 2 diabetes mellitus.

Hosam Ghanem, M.D, Mohamad S. Abd Al-Hamid, M.D, Adel Zalata, M.D and Sabry Gaad, M.D.

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5. Transesophageal echocardiography assessment of aortic initma media thickness in correlation to coronary artery disease .

Ghareeb M, MD, Gamal A, MD, Elhammady W, MD, Wadeaa B, MD, Essam T, M.Sc, Abdel Aziz A, MD.

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Editorial: Ethics of Medical Practice

Dr. Mokhtar Gomaa M.B.,B.Ch.;D.C.V.D.;M.D.

M.A.S.N.C.;F.R.C.P.(Ed.);F.E.S.C. Professor of Cardiology

Al-Azhar Faculty of Medicine

I was asked by Editor in chief professor Rizk to address the following subject in this editorial: The Legal-Ethical aspect of: 1- Physician-owned diagnostic or

therapeutic facilities. 2- Fee-sharing schemes. 3- Accepting gratis from industry, For educational purpose e.g. Journal publication. For a web site.

Addressing that subject needs introduction, since physicians as well as the public are not aware of it. Physicians are not aware of it because the subject is not given enough attention in the curriculum of the medical schools. At the same time there is no motivation to study the subject since there are no marks allocated to this course in the final degree (except for Al-Azhar Medical School). Another major factor is the lack of awareness of the patients and their relatives with regards to their rights. Over and above patients and their relatives tend to accept the problems happening after the physician's intervention as a natural outcome; so I will start by these general statements about the subject: Medicine is not a trade to be learned, but a profession to be entered1 . A profession is characterized by (a) a specialized body of knowledge that its members must learn, teach and expand and (b) a code of ethics and a duty of service that put patient care above self-interest2. The society granted the physicians the privilege of self-regulation3, this creates an obligation on the physicians and the whole medical society to stick to the ethics of medical practice. The ethics of medical practice developed over the years in different populations. The statement by Peabody, F.W.(1927) "The secret of the care of the

patient is in caring for the patient"1 remains a cornerstone in ethics of medical practice. Care of the patient means listening well to his problems, performing a through examination, asking for investigations relevant to his condition, having the most up-to-date information about his case - otherwise consult a specialist - advice the treatment modality which is solely in the patient's interest and keep everything confidential. (Francis Weld Peabody. The care of the patient, JAMA. 1927;88:877-82.) Medicine, law, and social values are not static. Re-examining the ethical tenets (principal or doctrine) of medical practice and their application in new circumstances is a necessary exercise. (Ethics Manual, fifth edition, Am.CP, 2005). The majority of ethics of medical practice is fundamental and timeless, however, there are major changes taking place in the societies that may influence the practice of the medical profession. Clinicians must be prepared to deal with these changes and affirm what is fundamental. The medical profession has long subscribed to a body of ethical statements developed primarily for the benefit of the patient. As a member of this profession, a physician must recognize responsibility to patients first and foremost, as well as to society, to other health professionals, and to oneself. The Principles adopted by the American Medical Association, the Egyptian medical Syndicate or the British Medical Council are not laws, but standards of conduct which define the essentials of honorable behavior for the physician. Back to the subject I must admit that reviewing the EMS code of ethics, The Ethics Manual of the ACP, and the BMC guide to ethical conduct and behavior, I found that

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these subjects are not given attention. Codes of ethics are put to face problems arising in the medical community concerned. Regarding the first point, the physicians-owned diagnostic or therapeutic facilities, can be divided into two categories, a) facilities that are considered now as an integral part of the basic clinical examination e.g. ECG for a cardiologist, X-Ray machine for chest physician or a small laboratory for a diabetologist (such facilities are necessary for the diagnosis and make things easier for the patient and doctor); b) facilities like echo machines and exercise testing machines which are not part of every diagnostic procedure may raise question marks in the mind of patients about the validity of the test especially if the results are normal or if the patient notices that it the test is repeated several times in a short interval. Use of such facilities is not covered by the known medical

ethics regulation code, however, I think this is a subject that needs to be studied. With regards to the free sharing scheme, we have to have a common consensus about the word sharing. If sharing means that the physicians share the medical activity, they are then entitled to share the fees. But if a physician sees a patient in his clinic or a hospital and refers the patient to another physician, a surgeon, a consultant, a laboratory or any diagnostic institution, he is not entitled to any fees for this referral, this is a subject of agreement in all ethical codes. Regarding the last question of accepting gratis from industry for educational purposes, I think this is not a subject of ethics of medical practice, since these donations are not given to a doctor, they are donated to an institution e.g. a society. Each society has its own regulations in accepting these donations and the donations are audited by government institutions.

References: 1 (Francis Weld Peabody. The care of the patient, JAMA. 1927;88:877-82.)

2 American Collage of Physicians Ethics Manual: Professionalism, 4,2005.

3 Medical Professionalism in the new Millennium: a physician charter. Annals of Intern. Med. 2002; 136: 243-6 [ PMID:11827500].

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Gender differences in Egyptian elderly with heart failure El Banouby M H1, Hamza S A1, Gamal A2 and

Adly N N1

Department of Geriatric Medicine1 and Department of Cardiology.2 Faculty of Medicine, Ain Shams University, Cairo, Egypt.

Abstract: Heart failure appears to have different characteristics between both genders. Considering those differences might have an impact in the future strategies in managing heart failure. This study is a descriptive comparative study aiming at detecting the gender differences among elderly with heart failure. 120 patients were subjected to comprehensive geriatric assessment, laboratory investigations [complete blood count, renal function tests, liver function tests, serum sodium, potassium, albumin, and thyroid functions] and echocardiography. Significant differences were found regarding the causes of heart failure. Males had more symptoms of low cardiac out put and nocturnal dyspnea and females had more fatigue, palpitation, and delirium. Females had more anemia and hyponatremia .The echocardiography showed that males had greater left ventricular end diastolic dimensions and more valvular affection. This study confirmed that considering gender differences is important when managing heart failure among elderly.

Introduction: Heart failure is a relatively common disorder. Its prevalence increases steeply with age(1).

The syndrome of heart failure has reached epidemic proportions throughout the world, and appears to have different characteristics in men and women (2).

Among elderly population the prevalence of heart failure markedly increased with age, but this increase is more obvious in females rather then males (3).

Epidemiological and clinical evidence has accumulated about differences in several aspects of cardiovascular diseases between males and females, including risk factors, response to therapy, quality of care and outcome(4). However, most work has concentrated on ischemic heart disease. Vaccrrino et al.,1999 emphasized that very little information is available on sex differences in the care and outcome of heart failure(5). Gender-related differences have been reported regarding cardiovascular structure, function, etiology of heart failure, symptoms, signs and echocardiographic findings. These may affect the prevalence, incidence, severity and psychological impact of the heart failure syndrome (6). Women with heart failure often have different clinical features than men, such as age of onset and comorbidities. Compared with males, females also demonstrate differences in remodeling and the response to injury, such

as volume or pressure overload and myocardial infarction(7).

The characteristics of the Egyptian elderly population are relatively different. There is a lack of studies about the gender differences in chronic diseases in general and in heart diseases in particular. For theses reasons this study was conducted aiming at describing the gender related differences among elderly Egyptians with heart failure. Methods: Research design: A descriptive comparative design was used in this study. Setting: The study was conducted in different wards of Ain Shams University hospital. Subjects: The study subjects included 120 patients with heart failure, (60 males & 60 females), aged 60 years or more. Heart failure diagnosed by the criteria in the Framingham study (8). Data collection:

Each participant was subjected to: -Comprhensive geriatric assessment including, demographic data, detailed history taking, complete clinical examination, mental status examination by the Arabic version (9) of Mini-Mental Status Examination (MMSE)(10), assessment of depression by the Arabic version (11) of the Geriatric Depression Scale (GDS) 15 items (12) and functional assessment performed by Activities of Daily Living (ADL)(13).

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- Laboratory investigations included complete blood count, renal function tests, liver function tests, Random Blood Sugar (RBS), serum sodium, potassium, albumin, and thyroid functions. -Electrocardiogram, chest X- ray and echocardiography. The study went through two phases: A pilot study: A pilot study was conducted among 10 patients to test the suitability of the setting, availability of the study population, applicability of the constructed tools that were used in data collection. The results of the pilot study were used as a guide for necessary modifications. -A definitive study: The data were collected from 120 elderly patients with heart failure, from different departments in Ain Shams University Hospital. Each session with a patient did not exceed 45 minutes to do a comprehensive geriatric assessment, then electro-cardiogram and chest X- ray were done, an appointment for echocardiography was scheduled and a blood sample was taken for the laboratory investigations. Data handling and statistical analysis: Data management and analysis were performed using SPSS version 11 software.

Χ2 statistic was used for categoric qualitative variables. Odds ratio along with 95% CI (confidence interval) were estimated in analysis of symptoms, signs and echogardiografic findings. Logistic regression analysis were carried out to evaluate the interaction between various independent factors on palpitation. Quantitative variable were described as mean + standard deviation (SD) and groups were compared using unpaired T test. All participants and their primary caregivers were informed about the aims and procedures of the study. Results: By studying the age distribution among both genders it was found that 63.3% of the males were in the age group (60-64 years), 30% were in the age group (65-74 years) and only 6.6% were > 75 years. While among females 50% were in the age group (65-74 years) and 40% were in the age (60-64 years) and 10% were >75 years . The difference was statistically significant (P= 0.038). Comparing the distribution of cardiovascular diseases between both groups revealed significant predominance of Ischemic Heart Disease (IHD) in males (P=0.002) and hypertension in females (P=0.025).

Table (1): The difference between males and females as regards symptoms of heart failure.

Symptoms Males (n=60)

Females (n=60)

Odds Ratio 95% C.I. P value

Exertional dysnea 100% (n=60) 100% (n=60) ---- -----

Orthopnea 86.7% (n=52) 80% (n=48) 1.625 0.612 – 4.316 0.327

PND 86.7% (n=52) 70% (n=42) 2.786 1.103 – 7.038 0.027*

Cyanosis 26.7% (n=16) 40% (n=24) 0.545 0.252 – 1.179 0.121

Dry cough 60% (n=36) 50% (n=30) 1.500 0.728 – 3.091 0.271 Daytime oliguria with

nocturia 36.7% (n=22) 46.7% (n=28) 0.662 0.319 – 1.373 0.267

Low COP 90% (n=54) 53.3% (n=32) 7.875 2.493- 21.071 < 0.001**

palpitation 56.7% (n=34) 96.7% (n=58) 0.045 0.010 – 0.202 < 0.001**

Chest pain 43.3% (n=26) 73.3% (n=44) 0.278 0.129 – 0.599 < 0.001**

Pain in right hypochon drium. 26.7% (n=16) 53.3% (n=32) 0.318 0.148 – 0.684 0.003**

Fatigue 43.3% (n=26) 83.3% (n=50) 0.153 0.065 - 0.358 < 0.001**

PND, paroxysmal nocturnal dyspnea; COP, cardiac out put

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Gender differences regarding the clinical picture of heart failure As shown in table (1) the differences between males and females regarding typical symptoms of heart failure, showed that males have significantly more low cardiac output symptoms (P=0.001) and PND(P=0.027). While females have significant more

palpitation (P<0.001), chest pain (P<0.001), fatigue(P<0.001) and right hypochondrial pain (P=0.003). Comparing atypical symptoms revealed significant predominance of anorexia (P=0.017) and delirium (P=0.011) in females . (table 2)

Table (2): Comparing atypical symptoms of heart failure in both genders

Symptoms Males (n=60)

Females (n=60)

Odds Ratio 95% C.I. P value

Delirium 6.7% (n=4) 23.3% (n=14) 0.235 0.072-0.762 0.011* Insomnia 83.3% (n=50) 83.3% (n=50) 1 0.383-2.612 1 Anorexia 60% (n=36) 80% (n=48) 0.375 0.166-0.849 0.017*

Gastro- intestinal disturbance 53.3% (n=32) 70% (n=42) 0.490 0.231-1.037 0.06

Logistic regression analysis for the estimation of the effect of different variables; arrhythmias (tachyarrhythmias, bradyarrhythmias, atrial fibrillation (AF), ventricular extrasystole), heart disease (IHD, cardiomyopathy, S3 gallop of heart failure), hyperthyroidism, hypothyroidism, anemia and valvular affection [mitral regurge (MR), aortic regurge (AR), tricuspid regurge (TR)] on palpitation, illustrated significant effect of AF (P=0.006), followed by low hemoglobin (Hb)

(P=0.027) on palpitation, while other variables had no significant effect on palpitation (p>0.05). By comparing signs of heart failure in both genders, hepatojugular reflux (P<0.001), central cynosis (P=0.001),S3 gallop(P=0.018), fine basal crepitations (P=0.020) and enlarged tender liver(P=0.027) were found to be significantly more in females.(table 3).

Table (3): Comparing signs of heart failure in both genders Signs of heart failure Males (n=60) Females (n=60) Odds Ratio 95% C.I. P value

Central cyanosis 3.3% (n=2) 23.3% (n=14) 0.113 0.025-0.524 0.001** Bilateral Lower limb oedema 80% (n=48) 66.7% (n=40) 2 0.872-4.585 0.099

Neck vein distension 53.3% (n=32) 73.3% (n=44) 0.416 0.193-0.83 0.023* Hepatojugular reflux 30% (n=18) 63.3% (n=38) 0.248 0.116-0.532 < 0.001** Fine basal crepitations 23.3% (n=14) 43.3% (n=26) 0.398 0.181-0.874 0.020*

S3 gallop 10% (n=6) 26.7% (n=16) 0.306 0.110-0.847 0.018* Enlarged tender liver 33.3% (n=20) 53.3% (n=32) 0.438 0.209-0.916 0.027*

N.B. pulmonary causes of central cyanosis were excluded The effect of heart failure on the mental, psychological and functional state Assessing cognitive impairment among the participants revealed that females showed significantly higher incidence of cognitive decline (P<0.001). Furthermore applying the GDS revealed that females were significantly more depressed (P<0.001). Functional decline was also significantly more in

females (P=0.003). Gender related differences regarding the laboratory investigations As shown in table (4) studying the laboratory result between both groups revealed only significant difference in hyponatremia (P<0.001) and anemia (P=0.010) with females predominance.

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Table (4): Laboratory differences in patients with heart failure

Lab. Males (n=60)

Females (n=60) P value

Hb< 12 g/dl 33.3% 56.7% 0.010* RBS≥ 200 mg/dl 26.7% 33.3% 0.426 BUN >22 mg/dl 70% 63.3% 0.439

Creatinine>1.4 mg/dl 35% 40% 0.572 AST >40 U/L 26.7% 16.7% 0.184 ALT >37 U/L 20% 10% 0.125

Total Bilirubin >1.2 mg/dl 20% 23.3% 0.658 Na < 136 mmol/L 13.3% 53.3% < 0.001**

< 3.5 mmol/L 10% 13.3% 2.653 K > 5.1 mmol/L 3.3% 10% 2.653 Albumin <2.6 g/dl 26.7% 43.3% 0.056

By studying Thyroid Stimulating Hormone (TSH), it was found that males had significantly more normal results than females, 65.6% in males and 42.4% in females P=(0.006), while females had more results of elevated TSH; in whom T4 was done and found to be low. So females were found to have hypothyroidism significantly

more often than males (28.8% of women versus 13.1% of men) (P=0.043). Other results of TSH; those who diagnosed as hyperthyroidism (low TSH and high T3), or those who diagnosed as sick euthyroid syndrome (low TSH and low T3) were of no significant difference in both genders.

Table (5): Echocardiographic findings in both men and women with heart failure

Echocardiographic findings Males (n=60)

Females (n=60)

Odds Ratio 95% C.I. P value

LVEDD >56mm 70% 50% 0.429 0.203-0.906 0.025* Hypokinesia 60% 50% 1.500 0.728-3.091 0.271

Diastolic impairment 56.7% 70% 0.560 0.264-1.189 0.130 Pulmonary hypertension 70% 56.7% 1.784 0.841-3.785 0.130

Ischaemic cardiomyopathy 16.7% 6.7% 2.800 0.826-9.490 0.088 LVEDD, left ventricle end diastolic dimension

Echocardiographic findings in both genders Comparing Echocardiographic finding in both males and females showed significant increase in left ventricular end diastolic dimension LVEDD>56mm in males (P=0.025) (table 5). Normal ejection fraction (EF ≥55%) was significantly more positive in females (20% of women versus 6.7% of men), while mild decrease in EF (45-54%) was equal in both genders (20%) and men had more positive results in moderate to severe affection of EF (<30%- 44%) but did not reach the level of significance (73.3% of men versus 56.7% of women, p = 0.056). Regarding valvular affection it was found that there were statistically significant differences

between males and females in cases of aortic regurge, tricuspid regurge, and mitral stenosis with males predominance. Discussion: Gender differences were found in cardiovascular diseases, which might have an impact on the prevention or management of those diseases. Comparing the two studied groups, significant differences were found in different aspects of assessment. By comparing the age group for both genders, most of the males were in younger age group then females. The older age of the females patients can be explained by the presence of estrogen (derived from aromatization of adrenal androstenedione) (14) although its

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level is much less then in younger generations. Estrogen appeared to have cardioprotective effect(15) . By studying the predisposing factors for heart failure, history of hypertension was higher in females. This findings confirmed that among those with heart failure females have more history of hypertension(5,16).As males were noticed to suffer more from Ischemic heart disease(17,18) so it was not surprising that the history of ischemic heart disease was higher in males. However cases with hypokinetic, akinetic, dyskinetic segment in echocardiography reported to be higher in men then women with no significant difference. Although not significant, this correlates with the finding that coronary artery disease was more common in males. It could be less severe in women, which explains why only significant difference with consequent more symptoms was seen in history not in echo findings. In the current study, comparison between both genders regarding the typical symptoms of heart failure revealed statistically significant difference in PND with male predominance while in chest pain and fatigue with female predominance. Those gender-related differences had been discussed before in other study (5) and nearly similar results were found although performed in different population. The same was found in low cardiac output (LCOP) symptoms, in form of claudication, blurring of vision, syncopes or coldness of extremities, were more in men than females. Although those data had not been confirmed previously, still we can say that those symptoms are non-specific and can be attributed to several causes rather than low cardiac output. The predominance of palpitation in females could be explained by logistic regression. Therefore the more the presence of AF and anemia in females could be responsible for the predominance of palpitation. Fatigue was expected to be associated mainly with LCOP symptoms, which were more common in males. However, as females sometimes pay more attention to

their physical well-being, the predominance of fatigue in females could be due to more perception of females to their symptoms. It is worth mentioning that predominance of fatigue in women remained significant after exclusion of patients with possible other causes for fatigue. Females patients with heart failure more likely to develop delirium(5) and our study had similar finding also. Concerning signs of heart failure, central cyanosis was seen more often in females. There was no available studies discussing this point. In the current study, by applying Mini-Mental Status Examination MMSE to study the current mental status of the patients, it was found that females had more cognitive impairment than men. This is in line with another study, (19) which investigated cognitive impairment in elderly heart failure patients and found that females have poorer scores in cognitive performance tests than males, using MMSE. The cognitive decline could be attributed to the increased risk of hypo-perfusion related cerebral ischemia (20)

Studying depression among elderly with heart failure is an important issue although it effect on patients with heart failure is not as clear as other cardiovascular disease e.g. IHD(21,22). However, higher level of depression in women than in men was found among patients with heart failure, and depression was explained by the limitation in physical activity only in men and not in women although limitation in physical activity are more pronounced in women than in men. This could be related to the underlying social cognitive factors, men perceive their symptoms as more psychologically invasive than women. Since among men a positive well being depends more on the strenuous activities they are able to perform(23). Regarding depression, in the current study it was found that females were more depressed than males. Concerning activities of daily living (ADL), this study showed that most of females were dependent, while most of men were independent. The predominance of dependent and assisted cases rather than

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independent cases in females can be explained by the predominance of fatigue in females. By comparing laboratory results of both genders, it was found that anemia is significantly more predominant in females. The same was found for hyponatremia that was more positive in women. Hyponatremia has been identified as a risk factor for increased morbidity and mortality in patients with congestive heart failure(24) and the presence of anemia was an independent marker of in-hospital mortality (25). Based on this result, it is of striking importance its predominance in females. By studying the thyroid profile it was found that men had significantly more normal results, while women had more results suggesting hypothyroidism. Assessment of the thyroid status is essential for elderly with cardiovascular disorder. Classic symptoms of hypothyroidism were less frequent in elderly. Moreover patients with history of hypothyroidism are at increased risk of developing cardiovascular disease. The cardiovascular manifestations of thyrotoxicosis are usually predominant(26) . Studying echocardiographic findings in the current study revealed increased LVEDD > 56 mm in males. On the other hand, normal EF (≥55%) was more in females. Actually those with normal EF represent patients with only diastolic dysfunction, and its predominance in females which has been previously reported in another study (27) was expected as myocardium appears able to adapt to marked pressure overload (by hypertrophy) to a greater extent in women than in men(by increase size and less hypertrophy)(28). This may explain the finding that females may have diastolic dysfunction more often than men. This predominance of cases with only diastolic dysfunction in females, attributed to more presence of hypertension as a cause of heart failure in females. It can be concluded from the current study that there are differences between both genders in all aspects of assessment. This may have its impact on the plane of prevention whether primary secondary or

tertiary. Female patients with any cardiovascular disease should receive more frequent doctor visits and close observation on developing heart failure. On the other hand males should be screened for risk factors of IHD and receive the proper management at the proper time. References: 1. British Heart Foundation database, 2002 2. Jessup M, Piña IL. Is it important to examine

gender differences in the epidemiology and outcome of severe heart failure? The Journal of Thoracic and cardiovascular surgery.2004; 127: 1247-1252.

3. Kitzman DW,Gardin JM, Gottdiener JS, et al. Importance of heart failure with preserved systolic function in patients≥ 65 years of age. Am J Cardiol. 2001; 87: 413-419.

4. Wenger NK, Speroff L, Packard B. Cardiovascular health and disease in women. New Engl J Med.1993; 329:247-256.

5. Vaccrrino V, Chen YT, Wang Y, et al. Sex differences in the clinical care and outcomes of congestive heart failure in the elderly. Am Heart J.1999;138:835-842.

6. Wenger NK. Women's heart failure and heart failure therapies. Circulation.2002; 105: 1526-1528.

7. Silber DH. Heart failure in women. Current Women's Health Reports.2003; 3:104-109.

8. McKee PA,Castlli WP, Mc NamaraPM et al. The natural history of congestive heart failure, The Framingham study. N Engl J Med.1971;285:1441-1446.

9. El Okl MA. Prevalence of Alzheimer dementia and other causes of dementia in Egyptian elderly. MD thesis, Faculty of Medicine, Ain Shams University.2002.

10. Folstein MF, Folstein SE, McHug PR. Mini-mental state. A practical method for grading the cognitive state of patients for the clinicians of Psychiat. Res.1975; 12:189-198.

11. Shehata AS. Prevalence of depression among Egyptian geriatric community. Master thesis. Faculty of Medicine, Ain Shams University.1998.

12. Sheikh JI, Yesavage JA. Geriatric Depression Scale (GDS): Recent evidence and development of a shorter version. Clinical Gerontology : A Guide to Assessment and Intervention 165-173, NY: The Haworth Press, 1986.

13. Katz S, Ford AB, Moskowitz AW ,et al . Studies of illness in the aged. The index of ADL: a standardized measure of biological and psychosocial function. Journal of American Medical Association.1963; 185: 914-919.

14. Gruenewald DA and Matsumoto AM. Aging of the endocrine system. In:Hazzard W R , Ettinger WH ,Halter JB and Ouslander JG. eds:

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Principles of Geriatrics and Gerontology.1999; 949-965.

15. Voss MR, Stallone JN, Li M, Cornelussen RNM, et al. Gender differences in the expression of heat shock proteins: the effect of estrogen. Am J Physiol Heart Circ Physiol.2003; 285: H687-H692.

16. McMurray JJV, Mc Donagh TA, Morrison CE and Dargie HJ . Trends in hospitalization for heart failure in Scotland. European Heart Journal.1993; 14: 1158–1162.

17. Philbin EF, DiSalvo TG. Influence of race and gender on care process, resources use, and hospital- based outcomes in congestive heart failure. Am J Cardiol.1998; 82: 76–81.

18. Ho KKL, Pinsky JL, Kannel WB and Levy D. The epidemiology of heart failure: the Framingham study. J Am Coll Cardiol.1993; 22: 6A–13A.

19. Khater MS. Relationship between heart failure and cognitive function among elderly. Master degree thesis ,Faculty of Medicine, Ain Shams University.2003.

20. Pullicino PM, Hart J. Cognitive impairment in congestive heart failure?: Embolism vs hypoperfusion. Neurology. 2001; 57:1986-1992.

21. MacMahon KMA, Lip GYH. Psychological factors in heart failure: a review of the literature. Arch Intern Med. 2002;162:509-516.

22. Januzzi JL Jr, Stern TA, Pasternak RC, et al. The influence of anxiety and depression on outcomes of patients with coronary artery disease. Arch Intern Med. 2000;160:1913-1921.

23. Murberg TA, Bru E, Aarsland T, Svebak S. Functional status and depression among men and women with congestive heart failure. Int Journal Psychiatry medicine.1998; 28: 273-291.

24. Oren RM. Hyponatremia in congestive heart failure. Am J Cardiol. 2005;95: 2B-7B.

25. Sales AL, Villacorta H, Reis L, Mesquita ET. Anemia as a prognostic factor in a population hospitalized due to decompensated heart failure. Arq Bras Cardiol.2005 ; 84:237-240.

26. Hassani S and Hershman J M .Thyroid diseases .In:Hazzard W R , Ettinger W H ,Halter J B and Ouslander J G.eds: Principles of Geriatrics and Gerontology. 1999;973-989.

27. Lindenfeld J, Krause-Stenrauf H, Salerno J. Where are all the women with heart failure? J Am Coll Cardiol.1997; 30:1417-1419.

28. Tandon S, Hankins SR, Le Jemtel TH . Clinical Profile of Chronic Heart Failure in Elderly Women. Am J Geriatr Cardiol .2002;11: 318-323.

15

Hypertension and Bone Mineral Density Among Elderly Patients. Rania M Ezzat, Hala S Sweed, A K Mortagy

Department of Geriatrics, Faculty of Medicine, Ain-Shams University, Egypt

Abstract A case control study was designed to assess the effect of hypertension on Bone Mineral Density (BMD). 124 male and female participants attending the outpatient clinic and the Osteoporosis unit, Geriatrics department at Ain-Shams university Hospitals, were recruited for the study. The study included two groups; 62 hypertensive patients and another 62 non-hypertensive subjects as their controls. Cases and controls were matched for age and sex. Subjects with diseases or taking drugs affecting bone density were excluded. The 124 subjects were subjected to comprehensive geriatric assessment. The BMD of femur neck and lumbar spine was measured using Dual Energy X-ray Absortiometry. In addition serum calcium and urinary calcium excretion in 24 hours urine, were done to 41 cases and 44 controls. Results; Data analysis revealed lower bone mineral density among cases compared to controls yet not statistically significant. Also the study revealed that hypertensive subjects have a highly significant increased urinary calcium excretion compared to controls .Negative correlation between urinary calcium excretion and bone mineral density was evident in this current study, but it was only significant at neck femur of hypertensive male cases. Also negative correlation between disease duration and bone density was evident in this present study, but this correlation was only highly statistically significant at femur neck of hypertensive females. Conclusion; hypertension is associated with decreased bone mineral density and this problems needs further planning for prophylactic planning.. Introduction

Hypertension and osteoporosis are major health problems facing the elderly, both having serious consequences such as fractures, stroke and myocardial infarction which affect dependency and quality of life of elderly (Perez et al., 2003). A number of studies in both humans (Dinna 2001, Cappuccio et al 2002, Perez et al., 2003) as well as animal models of hypertension (Umemura et al., 1986, Cirillo et al., 1989, Galletti et al 1991) have suggested an association between essential hypertension and osteoporosis. It was proved that high blood pressure precedes and predicts the loss of bone mineral through a large prospective study done by Kazushi et al., 2001 Hypertension is associated with increased urinary calcium excretion which causes reduction in ionized calcium, this activates compensatory mechanism including increase in parathyroid hormone, this will cause increase in bone resorption and increase risk of osteoporosis (Cappuccio et al, 2002).

Aim of the work: To assess the effect of hypertension on bone mineral density. Methodology Participant selection criteria

62 hypertensive patients (31males, 31 females), 60 years old and over, and 62 matched controls(31males, 31 females) were recruited from the outpatient clinic and the osteoporosis unit at the department of geriatrics, Ain-Shams university hospitals. Hypertension was confirmed by persistent increase of systolic blood pressure ≥ 140 mmHg and/or increase diastolic blood pressure ≥ 90 mmHg based on the average of 2 or more readings taken at each of 2 or more visits (JVC VII,2003) Or presence of known history of hypertension with therapy intake. This is a case-control study Exclusion criteria: Participants with diseases affecting bone density such as chronic renal disease, chronic liver disease, & some endocrinal diseases as Cushing disease were excluded. Also participants taking drugs affecting BMD as corticosteroids, heparin, anti-convulsants & loop diuretics were excluded.

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Tools of Assessment: All participants were subjected to Compre-hensive geriatric assessment including; full history and physical examination, functional assessment, cognitive function assessment, and depression screening. Functional assessment was done using Activities of daily living questionnaire (ADL) (Katz et al., 1963), Arabic version (El-sherpiny et al., 2000), Basic activities of daily living assess the ability of the patient to complete basic self-care tasks (e.g. bathing- dressing- toileting- continence- feeding and transfer). Mini-mental status examination (MMSE) (Folstien et al., 1975), Arabic version (El-Okl et al., 2002) was used for assessment of cognitive function with a score less than 24/30 indicating cognitive impairment. Presence of depression was assessed using Geriatric depression scale 15 items (GDS-15) (Sheikh and Yesavage, 1986), arabic version (Metwally et al., 1998). The cut-off point is 5/15 for significant depressive symptomology.

Measurement of the bone mineral density of femur neck and lumbar spine was done for all participants using Dual Energy X-ray Absortiometry (LUNAR DPX). Dual Energy X-ray Absortiometry, or DEXA scanning, is currently the most widely used method to measure bone mineral density. For the test, a patient lies down on an examining table, and the scanner rapidly directs x-ray energy from two different sources towards the bone being examined in an alternating fashion at a set frequency. The mineral density of the patient's bone weakens, or prolongs the transmission of these two sources of x-ray energy through a filter onto a counter in a degree related to the amount of bone mass present. The greater

the bone mineral density, the greater the signal picked up by the photon counter.

Not all participants approved for blood and urine samples to be taken from them so, serum calcium and urinary calcium excretion in 24 hours urine were done to only 41 cases (20males, 21females) and 44 controls (20males, 24females). Data processing & statistical analysis: Data collected was revised, coded, tabulated & introduced to PC for statistical analysis. All data manipulation & analysis were performed using the 10th version of SPSS (Statistical Package for Social Sciences. Qualitative data is presented in form of frequency tables (numbers and percent), while quantitative data is presented in form of mean ± standard deviation and range. The statistical tests used included; Student t test, Pearson Correlation coefficient (r) test and Chi-square test. Results The studied sample was in the form of two groups; 62 hypertensive subjects (31 male,31 female) with a mean age of 67.87± 4.90 (females;65.22±2.99-males; 66± 3.11) and 62 subjects (31 male,31 female) as their controls with a mean age of 65.56 ± 4.5 (females;67.56±4.21-males; 68 55 ±3.22) . The mean body mass index (BMI) of the cases was 29.85±6.37 (females; 33.09± 6.62- males; 26.61±4.11) and that of the controls was 28.01±8.02 (females;30. 44±10.21 -males; 25.56±3.73).There was no statistically significant difference between cases and controls as regards body mass index (BMI), smoking, milk drinking, tea drinking and coffee drinking. The following tables show the relations concerning hypertension and bone mineral density;

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Table(1):Comparison between male cases and controls as regard Bone density and T-score of femur neck and lumbar spine / serum ca and urinary ca excretion

Male cases Male controls t p Femur neck Bone density T-score

0.84±0.14 -1.72±1.13

0.85±.14 -1.63±1.11

0.19 0.34

0.84 0.94

lumbar spine Bone density T-score

0.96±0.21 -1.78±1.36

0.98±0.24 -1.77±1.19

0.23 0.03

0.82 0.98

serum ca 8.73±.31 8.65±.36 0.78 0.43 Urinary calcium excretion 198±28.25 163.30±33.59 3.61 0.001** Table(2):Comparison between female cases and controls as regard Bone density and T-score of femur neck and lumbar spine / serum ca and urinary ca excretion

female cases female controls t p Femur neck Bone density T-score

0.79±0.11 -1.72±0.96

0.81±0.19 -1.57±1.47

0.40 0.49

0.68 0.62

lumbar spine Bone density T-score

0.88±0.18 -2.20±1.25

0.90±0.22 -2.09±1.59

0.55 0.28

0.58 0.78

serum ca 8.74±0.28 8.75±0.30 0.12 0.90 Urinary calcium excretion

201.38±39.88 159.87±39.46 3.50 0.001**

Table(3):Correlation between urinary ca excretion and disease duration and bone density and T- score of lumbar spine and femur neck among hypertensive male and female cases

Femur neck Lumbar spine T- score Bone density T- score Bone density

r P r p r p r p Male cases Urinary ca excretion

-0.47 0.03 * -0.46 0.04* -0.30 0.19 -0.23 0.31

Disease duration

-0.22 0.22 -0.27 0.13 -0.19 0.28 -0.22 0.23

Female cases Urinary ca excretion

-0.24 0.29 -.04 0.84 -0.02 0.93 -0.30 0.17

Disease duration

-0.46 0.008 ** -0.55 0.001 ** -0.05 0.77 -0.19 0.28

** Highly significant < 0.01 * Significant < 0.05

Measurement of bone mineral density was done for the 62 cases (31 males and 31 females) and the 62 controls (31 males and 31 females) but serum calcium and urinary ca excretion was only possible for 41 cases (20 males and 21 females) and 44 controls (20 males and 24 females) [ Those who accepted to give blood and urine samples].

Lower bone mineral density and T score of lumbar spine and femur neck was found among male (table 1) and female (table 2) cases compared to their controls yet with no statistical significance difference by

T-test. There was highly statistical significant higher mean urinary ca excretion among male (table 1) and female (table 2) cases compared to their controls but there was no statistical significant difference between them as regard serum calcium.

When assessing the correlation between urinary ca excretion and bone density and T- score of lumbar spine and femur neck among hypertensive male and female cases (table 3), negative correlation was found but only statistically significant with femur neck among males only.

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Negative correlation was found between disease duration and bone density and T- score of lumbar spine and femur neck

among hypertensive male and female cases (table 3), but only statistically significant with femur neck among females only.

Table(4):Comparison between controlled and uncontrolled hypertensive males as regard T score and bone density of femur neck and lumbar spine / serum ca and urinary ca excretion

Controlled HTN males (n=14)

Uncontrolled HTN males (n=17)

t p

Femur neck Bone density T-score

0.82±.15

-1.84±1.28

0.85±.13

-1.62±1.02

0.61 0.52

0.54 0.60

lumbar spine Bone density T-score

1.04±.16

-1.58±1.42

0.94±.23

-1.93±1.32

1.32 0.70

0.19 0.48

serum ca 8.71±.36 8.75±.29 0.25 0.80 Urinary calcium excretion 200.75±23.69 197.50±32.18 0.24 0.80

Table(5):Comparison between controlled and uncontrolled hypertensive females as regard T score and bone density of femur neck and lumbar spine / serum ca and urinary ca excretion

Controlled HTN females (n=14)

Uncontrolled HTN females (n=17)

t p

Femur neck Bone density T-score

0.81±.07 -1.40±.62

0.77±.13 -2±1.11

0.99 1.79

0.32 0.08

lumbar spine Bone density T-score

0.89±.17 -2.05±1.04

0.86±.18 -2.39±1.40

0.55 0.71

0.58 0.47

serum ca 8.77±.22 8.73±.31 0.34 0.73 Urinary calcium excretion 187.00±49.15 210.23±31.92 1.32 0.20

Among the 62 hypertensive cases 28 where found to be controlled by therapy (14 males and 14 females) while the remaining 34 (17 males and 17 females), had high blood pressure whether uncontrolled by the taken therapy or diagnosed to be hypertensive by the current study. Lower bone mineral density and T score of lumbar spine and femur neck was found among male (table 4) and female (table 5) hypertensive uncontrolled cases compared to the controlled cases yet with no statistical significance difference by T-test and when comparing serum ca and urinary ca [done for 16 controlled hypertensive (8males, 8females) and 25 uncontrolled (12males, 13females)], no statistical significance difference was found (table4&5). Discussion

Our study revealed that, on analysing the data collected to find the effect of hypertension on bone mineral density of elderly, there is lower bone mineral density and T-score of lumbar spine and neck femur

among female and male cases compared to male and female controls yet with not statistical significant difference. Lack of statistical significant difference in the current study can be explained by small sample size. Studies done by Graham et al,1999 and Dinna,2001 revealed that hypertensive patients are more likley to develop bone demineralization(osteoporosis) . Results of a study done by Francesco et al,1999 also revealed that high blood pressure in elderly women is associated with increased bone loss at the femoral neck It was also proved with the results of a study done by Kazushi et al,2001 that bone mineral density in lumbar spine was significantly decreased in female hypertensive subjects compared with female normotensive controls. The association between hypertension and bone mineral density can be explained by the presence of evidence showing that hypertension might be linked to abnormalities of calcium metabolism including an increased

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calcium excretion which results in elevated parathyroid hormone level which will cause an increase in bone loss (Kazushi et al, 2001). The cause of hypercalcuria in hypertensive subjects can be explained by either central blood volume expansion (central blood volume hypothesis) or renal tubular disorder (renal calcium leak hypothesis) (Cappuccio et al, 2002). Actually, this hypercalcuria was evident by the present study, where it was revealed that hypertensive subjects have a highly significant increased urinary calcium excretion among female and male cases compared to their controls. This agreed with the results of studies done by Gennari et al , (1986) , Hvarfner et al , (1987), Tillman and Semple , (1988) , Andreas et al , (1990) , Brickman et al , (1990) , Papagalanis et al , (1991) , Strazzullo , (1991) , Eric et al , (1995) , Carl et al , (1996) ,Francesco et al ,(1999) , Graham et al , (1999) , Kazushi et al,(2001) and Perez et al , (2003) which revealed increased urinary calcium excretion in hypertensives compared to normal subjects. When comparing the level of serum calcium among the studied group, the study revealed that there is no difference between cases and controls as regard serum calcium level. This agreed with the results of a study done by Hvarfner et al ,(1987) , Papagalanis et al , (1991) , Reichel et al , (1992) , Eric et al , (1995),Strazzullo and Cappuccio , (1995) and Carl et al , (1996) , which revealed that hypertensive patients had a mean total calcium concentration in the serum which was not significantly different from the corresponding mean value in the normotensive group. This can be explained by the fact that, the hypercalciuria of hypertension can produce an increase in parathyroid serum level because of a decrease in the serum calcium level, so returning serum calcium level to normal (Perez et al, 2003). Negative correlation between urinary calcium and bone mineral density was evident in this current study, but it was only significant at femur neck of hypertensive males.

A study done by Strazzullo and Cappuccio,(1995) also revealed that an increase in 24 h urinary calcium excretion may be associated with a number of metabolic disorders ,including increased rate of bone resorption and a primary renal defect in calcium handling. Results of a study done by Allan et al, (1991) and Gail et al,(1994) also revealed that renal calcium excretion is a risk factor of osteoporosis. Also the study done by Amanda et al, (1995) revealed that urinary calcium excretion is associated with an increased bone loss at the hip site. The mechanism of the bone loss with higher urinary calcium excretion is likely to be related to higher rates of bone resorption because of hyperparathyrodism as a compensatory response (Carl et al, 1996 and Graham et al 1999). Negative correlation between disease duration and bone density was evident in this present study, but this correlation was only highly statistically significant at femur neck of hypertensive female cases. Francesco et al, (1999) revealed that sustained hypercalcuria in people with high blood pressure over many years, leads to an increased risk of bone mineral loss in elderly women. When assessing the relation between control of blood pressure and bone mineral density of hypertensive subjects, the study revealed lower bone mineral density and higher urinary calcium excretion among uncontrolled hypertensive female cases yet not statistically significant. But the opposite was evident among male cases. The higher urinary calcium and consequently the lower bone mineral density among hyprtensive controlled males can be explained by higher BMI among them compared to hypertensive uncontrolled males, as hypercalcuria increased with larger body mass (Young et al,2002).

A study done by Cappuccio et al, (1999) revealed that the rate of bone loss at the femoral neck increased with the level of blood pressure at base line. Also a study done by Francesco et al., (1999) revealed that blood pressure level was

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a significant predictor of bone mineral loss at the femoral neck . This agreed with the results of a study done by Eric et al, (1995) which revealed that there is a direct correlation between urinary calcium excretion and level of blood pressure. So, hypertension has a negative impact on BMD through increased urinary calcium excretion .Hypertension duration and control affect bone mineral density.

Conclusion From our study we concluded that lower bone mineral density of lumbar spine and neck femur among female and male cases compared to male and female controls. Also study revealed that hypertensive subjects have a highly significant increased urinary calcium excretion compared to normotensives. References: 1. Allan GN, Howard AM , David BC ,et al (1991):

Effect of salt restriction on urine hydroxyproline excretion in postmenopausal women. Arch Intern Med 151:757-759.

2. Amanda DR, Arthur C, Ian MD, et al (1995): A longitudinal study of the effect of sodium and calcium intakes on regional bone density in postmenopausal women. Am J Clin Nutr 62:740-745.

3. Andreas H, Reinhold B, Claes M, et al (1990): Relationships between calcium metabolic indices and blood pressure in patients with essential hypertensionas compared with a healthy population. J Hypertens 5:451-456.

4. Brickman AS, Nyby MD, Von Hungen K, et al (1990):Calcitropic hormones, platelet calcium and blood pressure in essential hypertension. Hypertension 16:515-522.

5. Cappuccio FP, Meilahn E, Zmuda JM and Cauley JA (1999):High blood pressure and bone mineral loss in elderly white women. Lancet 354:971-975.

6. Cappuccio FP, Kalaitzidis R, Duneclift S and Eastwood JB (2002):Unravelling the links between calcium excretion, salt intake, hypertension, kidney stones and bone metabolism. J Nephrol 13:169-177.

7. Carl GO, Rosanne BM, Janet D, et al (1996): Evidence for the relationship of calcium to blood pressure. Nutrition Reviews 12:356-381.

8. Cirillo M, Galletti F, Strazzullo P, et al (1989):On the pathogenetic mechansim of hypercalcuria in genetically hypertensive rats of the Milan strain. Am J Hypertens2:741-746.

9. Dinna NC (2001):The renal tubular Na-Cl co-transporter(NCCT):a potential genetic link

between blood pressure and bone density?. Nephrol Dial Transplant 16:691-694.

10. El-Okl MA , El-Banouby MH, El-Etribi MA, et al (2002):Prevalence of Alzheimer disease and other types of dementia in the elderly. MD Thesis. Ain Shams University: Geriatrics Department Library.

11. El-sherpiny MS, Mortagy AK and Fahy HM (2000): Prevalence of hypercho-lesterolemia among elderly people living in nursing houses in Cairo. MD Thesis. Ain Shams University: Geriatrics Department Library.

12. Eric WY, Cynthia DM, Scott H, et al (1995): Regulation of parathyroid hormone and vitamin D in essential hypertension. Am J Hypertens 8:957-964.

13. Francesco P, Elaine M, Joseph MZ and Jane AC (1999): High blood pressure and bone mineral loss in elderly white women :a prospective study. Lancet 354:971-975.

14. Folstein MF, Folstein, SE and McHugh PR (1975): Mini-Mental State: A practical method for grading the state of patients for the clinician, Journal of Psychiatric Research, 12: 189-198.

15. Gail AG, Elizabeth B-C, Sbaron E, et al (1994): Dietary sodium and bone mineral density: results of a 16-year follow-up study. Am Geria Soc 42:1050-1055.

16. Galletti F, Rutledge A and Triggle DJ (1991):Dietary sodium intake :influnce on calcium channels and urinary calcium excretion in spontaneously hypertensive rats. Biochem Pharmacol 41:893-896.

17. Gennari C,Nami R, Bianchini C, et al (1986): Renal excretion of calcium in human hypertension. Am J Nephrol 6:124-127.

18. Graham A, MacGregor and Cappuccio FP (1999): The kidney and essential hypertension: a link to osteoporosis? . J Hypertens 11:781-785.

19. Hvarfner A, Bergstorm R, Morlin C and Wide L (1987): Relations between calcium metabolic and blood pressure in patients with essential hypertension as compared in healthy population. J Hypertens 5:451-456.

20. Katz S, Ford AB, Moskowitz RW, et al (1963): Studies of illness in the aged. The index of ADL. Standardized measure of biological and psychosocial function. JAMA; 185:914-919

21. Kazushi T, Ichiro N and Yoshiaki M (2001): bone mineral density in women with essential hypertension. Am J Hypertension 14:704-707.

22. Metwally AS, El-Banouby MH, Mortagy AK and Ghanem M. (1998); Prevalence of depression among Egyptian geriatric community. Master Thesis. Ain Shams University: Geriatrics Department Library. Pp3-5

23. Papagalanis ND , Skopelitis P , Kourti A, et al (1991):Urine calcium excretion, nephrogenous cyclic-adenosine monophosphate and serum parathyroid hormone levels in patients with essential hypertension. Nephron 59:226-231.

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24. Perez C, Justo I, Silva J, et al (2003):Bone mass and bone modelling markers in hypertensive postmenopausal women. Journal Of Human Hypertension 17:107-110.

25. Reichel H, Leibethal R, Hense H-W , et al (1992):Disturbed calcium metabolism in subjects with elevated diastolic blood pressure. Clin Invest 70:748-751.

26. Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment Of High Blood Pressure (2003); Evidence from new hypertension trials. Hypertension 1:1-3

27. Sheikh JI, Yesavage JA (1986); Geriatric Depression Scale (GDS): Recent evidence and development of a shorter version. Clinical Gerontology : A Guide to Assessment and Intervention 165-173, NY: The Haworth Press.

28. Strazzullo P (1991): The renal calcium leak in primary hypertension : pathophysiological aspects and clinical implications . Nutr Metab Cardiovasc Dis 1:98-103.

29. Strazzullo P and Cappuccio FP (1995): Hypertension and Kidney stones : hypotheses and implications . Sem Nephrol 15: 519-529.

30. Tillman Dm and Semple PE(1988):Calcium and magnesium in essential hypertension. Clini Sci 75:395-402.

31. Umemura S, Smyth DD and Nicar M (1986):Altered calcium homeostasis in Dahl hypertensive rats:physiological and biochemical studies. J Hypertens 4:19-26.

32. Young J. H, Klag M. J, Muntner P, et al (2002):Blood Pressure and Decline in Kidney Function: Findings from the Systolic Hypertension in the Elderly Program (SHEP) J. Am. Soc. Nephrol 13(11): 2776 – 2782. Correspondence: Hala Samir Sweed Lecturer of Geriatric Medicine, Geriatrics Department, Faculty of Medicine, Ain Shams University. E-Mail: halasweed@yahoo.co.uk

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Relationship of circulating insulin-like growth factor-1 (IGF-1) and insulin like growth factor binding proteins-1 and 3 (IGFBP-1 and IGFBP-3) with Endothelin-1 (ET-1) in

hypertensive patients with and without type 2 diabetes mellitus Hosam Ghanem*,M.D, Mohamad S. Abd al-hamid**,M.D, Adel Zalata***,M.D and Sabry

Gaad, M.D. Clinical pathology*, Internal Medicine **, Biochemistry*** and Physiology Departments,

Mansoura University, Faculty of Medicine.

Abstract Background: The IGF system is involved in the underlying pathophysiological processes and development of cardiovascular diseases. Studies suggest a role for IGF-I as a mediator of the hypertrophic responses of vascular smooth muscle cells in hypertension. Insulin like growth factor binding protein-1 (IGFBP-1) and insulin- like growth factor binding protein-3 (IGFBP-3) are the best characterized circulating insulin-like growth factor binding proteins. The effects of circulating IGFs on the vasculature are largely modulated by IGFBPs which control their access to cell-surface IGF receptors. IGFBP-I has been proposed as the acute regulator of IGF bioavailability because of its metabolic regulation by glucoregulatory hormones. Endothelins have been shown to have mitogenic effects on endothelial cells and could be involved in growth processes in vitro and in vivo. Objective: To assess the state of IGF system (IGF-I, IGFBP-I and IGFBP-3) and endotholin-I and their interrelationship in patients suffering from hypertension with and without type 2 DM, a disease known to have a effect on the vasculature. Design: prospective controlled study. Setting: Internal Medicine and Clinical Pathology departments, Mansoura University. Subjects and methods: This study was conducted on 60 patients, 20 hypertensive patients, 20 hypertensive diabetic patients and 20 normotensive diabetic patients, in addition to 15 subjects age and gender matched as a healthy reference group. All members of the study were subjected to the recommended clinical assessment and diagnostic laboratory investigations. Plasma ET-I levels were evaluated by enzyme immunoassay. Serum IGF-I and IGFBP-I have been estimated using the ELISA method. Serum IGFBP-3 was measured using immunoradiometric assay (IRMA) method. Results: There was a significant increase in plasma endothelin-I in the three patient groups versus the healthy normotensive control group (0.86 ± 0.05, 2.65 ± 0.06, 2.4 ± 0.05 in hypertensive, hypertensive diabetic and normotensive diabetic respectively versus 0.7 ± 0.03 in healthy controls). Plasma endothelin-1 was significantly higher in normotensive diabetic and hypertensive diabetic groups versus the hypertensive group. . Serum IGF-1 was significantly lower in three patient groups versus the healthy normotensive control group (169.5 + 29.4, 129.5+26.5, and 153.5 + 37.5, in hypertensive, hypertensive diabetic, and normotensive diabetic respectively versus

199.8 + 30.5 in healthy controls). Serum IGF-1 was significantly lower in hypertensive diabetic group versus hypertensive group (129.5+26.5 versus 169.5 + 29.4 respectively, p < 0.05).The lowest level of IGF-I was detected in hypertensive diabetic group (129.5 ± 26.5). . On the other hand, there was a significant decrease in IGFBP-I and IGFBP-3 in the three studied patient groups versus the healthy control group, while their levels showed no significant difference in diabetic hypertensive group versus diabetic group There were negative significant correlation between IGF-1 and both diastolic and systolic blood pressure in the three studied patient groups. Also circulating IGFBP-I showed significant

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negative correlation with diastolic and systolic blood pressure in the three studied patient groups. In the present study, there is a significant negative correlation between ET-I and measured parameters of IGF system (r = - 0.64, p < 0.05 with IGF-1, r= -0.59, p <0.05 with IGFBP-1 and r = -0.61, p <0.05 with IGFBP-3 in hypertensive diabetic group). Conclusions: Patients suffering from hypertension with or without type 2 diabetes mellitus showed significant reduction in IGF-I, IGFBP-I and IGFBP-3, while there was a significant increase in endothelin-I level as compared to the healthy reference group. In addition, a novel significant negative correlation between ET-I and assayed parameter of the IGF system. These findings implicate the role of IGF system and Endothelin-I as predictor markers of cardiovascular risk in patients with hypertension and hypertensive diabetic patients. ET-I receptor antagonists and IGF-1 replacement may have a role in ameliorating this dysfunction. Key words: IGF-I, IGFBP-I , IGFBP-3, endothelin-I. and hypertension. Introduction: Microvascular disease is an important cause of morbidity and mortality in both IDDM and NIDDM. It can result in retinopathy, and it can contribute to foot pathology, diabetic cardiopathy and neuropathy (1). Despite this, the pathogenesis of diabetic microangiopathy is uncertain. The haemodynamic hypothesis argues that an early increase in precapillary blood flow and capillary hypertension precede the adaptive changes that lead to microvascular sclerosis (2). Hyperglycaemia per se modulates endothelial dependent vasodilation and elevated free fatty acids concentrations may result in endothelial dysfuncrion (3). Endothelin is a potent vasoconstrictor oligopeptide. Like other biologically active oligopeptides, it is converted into its biologically active form via two virtually inactive precursors, pre-proendothelin and proendothelin by means of two endopeptidases (4). The enzyme that transforms proendothelin (also called big

endothelin) into endothelin is designated as endothelin converting enzyme (ECE) and acts in a highly specific manner (5). By screening cDNA libraries for homologies, it was possible to identify two further isoforms of endothelin in humans (6). The original endothlein was named endothelin-1 (ET-1) and its gene is localized on chromosome 6 (7). The ET-2 gene is located on chromosome 1 and the ET-3 gene on chromosome 20 in humans (6). The effects of the endothelins (ETs) are mediated by at least two distinct receptors, type A and B (ETa and ETb), which binds ETs with different affinities. ETb binds all three ETs equally, whereas ETa binds ET-1 preferentially. There is substantial evidence that ETa receptors primarily mediate vasoconstrictive and mitogenitc effects on smooth muscle cells. Etb receptor expression was demonstrated primarily on endothelial cells. These receptors are thought to exert vasodilator effects by stimulating nitric oxide (NO) formation (8&9). The physiological relevance of endogenous generation of endothelin-1 in controlling blood pressure has been unclear (10). If basal generation of ET-1 contributes to resistance-vessel tone, then drugs that inhibit the generation or action of endothelin would be expected to cause vasodilatation and decrease blood pressure and might have potential therapeutic value in diseases associated with sustained peripheral vasoconstriction, such as hypertension and chronic heart failure (11). However, results of animal studies using ECE inhibitors and endothelin receptor antagonists (12, 13 & 14) have been contradictory. Type 2 diabetes is associated with a three to fivefold increased risk of macrovascular disease that cannot be completely explained by known risk factors. IGFs and IGFBP are increasingly being recognized as contributing to cardiovascular risk (15 & 16). In addition, there are well-established links between IGF and other surrogate markers for cardiovascular risk. IGF-I has been shown to correlate with LDL cholesterol in normal women (17), and elderly (18) and with apolipoproteins B and cardiovascular risk in normal subjects (19).

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There is also evidence that IGF system is involved in the underlying pathophysiological processes associated with cardiovascular disease. In vivo animal studies suggest a role for IGF-I as a mediator of the hypertrophic responses of vascular smooth muscle cells in hypertension (20), and implicate IGF-I as a mediator of cardiac hypertrophy in congestive cardiac failure (21). IGF-I is released into the blood mostly by the liver (22) and reaches the target cells in the classic endocrine manner (23). The secreted IGF-I leads to increased collagen synthesis and cell differentiation (24).

IGF binding proteins predominantly control the access of IGFs to tissues and cell surface IGF receptors. Of these IGFBP-I is the most likely candidate for acute regulation of IGF actions because of its acute downregulation by insulin and upregualtion by other glucoregulatory hormones and cytokines (25, 26, 27, 28 & 29).

Furthermore, transendothetial movement of IGFBP-I into the tissues is known to occur and to be enhanced by insulin (30). IGFBP-I thus provides an important link between intermediary metabolism and regulation of IGF actions (31

& 32). It therefore seems likely that in type 2 diabetes, the known dysregulation of IGF system would result in changes in IGF bioavailability that subsequently could predispose an affected individual to develop premature cardiovascular disease (31).

The aim of the present study was to assess the state of IGF system (IGF-I, IGFBPI and IGFBP-3) and endothelin-I and their interrelationship in hypertensive patients with and without type 2 diabetes mellitus. Subjects and methods: This study was conducted on 60 patients who were admitted to Internal Medicine Department (Mansoura University). Patients were diagnosed on clinical and laboratory grounds. All subjects had normal renal function. Table (1) summarizes the clinical characteristics of all subjects. They were classified into three groups: patients with hypertension only included 20 subjects, 20 hypertensive diabetic patients, and 20 diabetic normotensive patients in addition to 15 healthy control subjects. Reference group was carefully selected from donors attending the blood bank of Mansoura University Hospital.

Table (1): Clinical characteristics of all subjects. Healthy

controls Hypertensive Hypertensive

diabetic Normotenstiv diabetic

Number (male/female) 9/6 12/8 12/8 12/8 Mean age (year) 48.0 50.0 53.0 51.0 Fasting blood glucose (mean + SD) 81+27 90.8+12.8 152.1+37.2 160+39.5

Duration of diabetes (year) (means + SD) 10.9+1.2 9.9+0.9

Blood pressure (mmHg) a- Systolic (mean + SD) b- Diastolic (mean + SD) c- Mean B.P (mean + SD)

129.5+9.6 81+7.4 124+8

177.5+12.08 106+9.07 165.1+13.1

163+22.5 100.3+8.7 154.6+16.0

121+14.5 74.6+7.5 114+9

Serum creatinine (mg/dl) (mean + SD) 0.82+0.05 0.96+0.15 0.98+0.13 0.81+0.07

Samples: Fasting blood samples were obtained from a subcutaneous vein in the forearm, from every subject included in the research (patients or controls): (1) 3.0 ml blood were collected in an ice chilled polypropylene tubes containing EDTA (1mg/ml of blood) and aprotonin (500 KIU/ml

of blood) (2),centrifuged in cooling centrifuge at 6000 rpm for 20 minutes and the plasma separated and stored at -20 oC until extracted, and used for endothelin-I assay. (2) 1.0 ml on sodium fluoride. The plasma was used for glucose measurement. (3) 5.0 ml blood was drawn in a plain polypropylene tubes. Serum was separated

26

after centrifugation at 6000 rpm for 10 minutes, divided into aliquots and stored immediately at - 20 oC until used for measurement of IGF-I, IGFBP-I, IGFBP-3 and creatinine. Methods

• Blood glucose was determined by glucose oxidase method (33) and and creatinine by Jaffe reaction (BioMerieux, France).

• Plasma endothelin-I was determined by the kits supplied from R and D system 614 Mckinley place N.E minneapolits MN 55413 USA (34).

• The insulin-like growth factor-1 (IGF-I) was determined by enzyme-linked immunosorbent assay (ELISA) kit supplied from diagnostic system laboratories, Inc., Webster, Texas, USA. It is an enzymatically amplified "one step" sandwitch-type immunoassay. The assay includes a simple extraction step in which IGF-I is separated from its binding protein. This step is considered to be essential for accurate determination of IGF-I (35 & 36).

• Insulin-like growth binding protein -I (IGFBP-1) ELISA kit is an enzymatically amplified "two step" sandwich-type immunoassay (37) supplied from diagnostic systems laboratories Inc., Webster, Texas, USA.

• IGFBP-3 was measured using a two site immunoradiometric assay (IRMA) principle described by Miles et al., (38). The IRMA is a non-competitive assay in which the analyte to be measured is "sandwiched" between two antibodies. The first antibody is immobilized to the inside walls of the tubes. The detected sensitivity limit of the assay was 0.5 ng/ml. The other antibody is radiolabelled for detection (39, 40 & 41) (Diagnostic systems laboratories-Inc. Texas, USA).

Statistical Analysis: The data of this study were statistically analyzed using the SPSS/PC computer package version 5 on an IBM compatible personal computer (SPSS INC, Chicago IL). Unpaired students t-test was used to evaluate the difference in each variable between the two groups. P value <0.05 is considered significant. Also, pearson,s correlation coefficient (r) was used to study the correlation between plasma endothelin-1, IGF-I, IGFBP-3 and IGFBP-1 and other different studied parameters. Results: Results of this work are illustrated in tables 2 – 6. Plasma endothelin-1 was significantly higher in three patient groups versus the healthy normotensive control group (0.86 ± 0.05, 2.65 ± 0.06, 2.4 ± 0.05 in hypertensive, and hypertensive diabetic and normotensive diabetic respectively versus 0.7 ± 0.03 in healthy controls). Serum IGF-1 was significantly lower in three patient groups versus the healthy normotensive control group (169.5 + 29.4, 129.5+26.5, and 153.5 + 37.5 in hypertensive, hypertensive diabetic, and normotensive diabetic respectively versus 199.8 + 30.5 in healthy controls). IGFBP-3 and IGFBP-1 were significantly lower in three patient groups versus the healthy normotensive control group (Table 2). Fasting blood glucose was significantly higher in normotensive diabetic and hypertensive diabetic groups only versus healthy control group (Table 2) The highest level of systolic and diastolic blood pressures were detected in hypertensive group per se. Plasma endothelin-1 was significantly higher in normotensive diabetic and hypertensive diabetic groups versus the hypertensive group (2.4 ± 0.05, 2.65 ± 0.06 in normotensive diabetic and hypertensive diabetic respectively versus 0.86 ± 0.05 in hypertensive group). There was no significant difference in endothelin-1 in hypertensive diabetic group versus normotensive diabetic group. Serum IGF-1 was significantly lower in hypertensive diabetic group versus hypertensive group (129.5+26.5 versus 169.5 + 29.4

27

respectively, p < 0.05). IGFBP-3 and IGFBP-1 were not significantly different between normotensive diabetic group, hypertensive diabetic group versus hypertensive group (Table 3). There were negative significant correlation between IGF-1 and both diastolic and systolic blood pressure in the three studied patient groups. On the other hand, no significant correlation between IGF-1 and fasting blood glucose, IGFBP-3 and IGFBP-1 in the three studied patient groups (Table 4). There were negative significant correlation between IGFBP-1 and both diastolic and systolic blood pressure in the three studied patient groups. On the other hand, no

significant correlation between IGFBP-1 and fasting blood glucose in the three studied patient groups (Table 5). In the present study, there is a significant negative correlation between ET-I and measured parameters of IGF system (r = - 0.64 , p < 0.05 with IGF-1, r= -0.59 , p <0.05 with IGFBP-1 and r = -0.61, p <0.05 with IGFBP-3 in hypertensive diabetic group, r= -0.65, p <0.05 with IGF-1, r= -0.60, p <0.05 with IGFBP-1 and r = -0.63, p <0.05 with IGFBP-3 in hypertensive group, r= -0.70, p <0.05 with IGF-1, r= -0.65, p <0.05 with IGFBP-1 and r = -0.64, p <0.05 with IGFBP-3 in diabetic group (Table 6).

Table (2): Plasma levels of endothelin-1, IGF system in the three studied patient groups versus healthy controls.

Healthy controls Hypertensive Hypertensive

diabetic Normotenstive

diabetic Fasting blood glucose (mg/dl) P-value

81 + 27

90.8+15.8 >0.05

152.13 + 37.2 0.001

160 + 39.5 0.001

Plasma Endothelin-I (mg/ml) P-value

0.7 + 0.03 0.86 + 0.05 0.001

2.65+0.06 0.001

2.40 + 0.05 0.001

IGF-I (ug/L) P-value

199.8 + 30.5 169.5 + 29.4 0.001

129.5+26.5 0.001

153.5 + 37.5 0.001

IGFBP-3 (ug/L) p-value

2460.5 + 1706

2050 + 180 0.001

2130 + 180 0.001

2080 + 170 0.001

IGFBP-1 (ug/L) p-value

86.6 + 9.1 48.9 + 8.0 0.001

52 + 7.5 0.001

50.4 + 8.0 0.001

Data are presented as means (+ SD). P<0.05 is considered significant; p<0.01 is highly significant. P<0.001 is very highly significant. Table (3): Anova test for comparative studies of IGF system, endothelin, F.B.S., blood pressure between the different studied patient groups.

Hypertensive Hypertensive diabetic Normotensive diabetic D.B.P P1 P2 P3

106 + 9.1 100.33 + 8.76

0.001 0.001

74.58 + 7.5 0.001

S.B.P P1 P2 P3

177.5 + 12.1 163 + 22.5

0.01 0.001

121.7 + 14.5 0.001

F.B.S P1 P2 P3

90.8 + 15.8 152.13 + 37

0.01 0.003

160 + 39.6 0.001

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Hypertensive Hypertensive diabetic Normotensive diabetic Endothelin-1 P1 P2 P3

0.86 + 0.05 2.65 + 0.06

0.01 >0.05

2.4 + 0.04 0.001

IGF-I P1 P2 P3

169.5 + 29.4 129.5 + 26.5

< 0.05 > 0.05

153.5 + 32.5 > 0.05

IGFBP-3 P1 P2 P3

2050 + 160 2130 + 180 >0.05 >0.05

2080 + 170 > 0.05

IGFBP-I P1 P2 P3

48.9 + 8.0 52 + 7.5

> 0.05 > 0.05

50.4 + 8.0 > 0.05

P1: Hypertensive group versus diabetic group; P2: Hypertensive versus hypertensive diabetic; P3: Diabetic group versus hypertensive diabetic. Table (4): Correlation between IGF-I and other different studied parameters in the three patient groups.

Group Diastolic

blood pressure (mmHg)

Systolic blood

pressure (mmHg)

F.B.S (mg/dl) IGFBP-1 IGFBP-3

Hypertensive group (n=20) R P

- 0.601 < 0.05

- 0.53 < 0.05

0.14 >0.05

0.122 > 0.05

0.21 >0.05

Hypertensive diabetic (n=20) R P

- 0.68 < 0.05

- 0.64 < 0.05

0.17 >0.05

0.05 > 0.05

0.25 >0.05

Diabetic group (n=20) R P

- 0.70 < 0.05

- 0.65 < 0.05

0.15

>0.05

0.31

> 0.05

0.23

>0.05 Table (5): Correlation between IGFB-I and other different studied parameters in the three patient groups.

Diastolic blood pressure (mmHg

Systolic blood pressure (mmHg)

F.B.S (mg/dl)

Hypertensive (n=20) R P

- 0.63 < 0.05

- 0.56 < 0.05

0.171 > 0.05

Hypertensive diabetic (n=20) R P

- 0.69 < 0.05

- 0.65 < 0.05

0.13

> 0.05

Diabetic (n=20) R P

- 0.71 < 0.05

- 0.67 < 0.05

0.32

> 0.05

29

Table (6): Correlation between endothelin-1 and different studied parameters in the three patient groups.

Group Diastolic

blood pressure (mmHg)

Systolic blood

pressure (mmHg)

Fasting glucose (mg/dl)

IGF-I (ug/L)

IGFBP-I (ug/L)

IGFBP-3 (ug/L)

Hypertensive group (n=20) R P

- 0.214 > 0.05

- 0.308 > 0.05

0.545 > 0.05

-0.65 < 0.05

-0.60 < 0.05

-0.63 < 0.05

Hypertensive diabetic (n=20) R P

- 0.115 > 0.05

- 0.468 > 0.05

- 0.260 > 0.05

-0.64 <0.05

-0.59 < 0.05

-0.61 < 0.05

Diabetic group (n=20) R P

0.273 > 0.05

- 0.02 > 0.05

0.009 > 0.05

-0.70 <0.05

-0.65 < 0.05

-0.64 < 0.05

P<0.05 is considered significant; P<0.01 is highly significant. Discussion E-1A is mainly synthesized by the vascular endothelial cells and acts on the vascular smooth muscle cells. Because of it vasconostrictor and mitogenic effects. It plays a role in development of vascular diseases (42). Endothelin-1 could be produced not only by large vessels but also by small vessels such as retinal microvessels (2). Endothelin receptors are present in cultured rat aortic smooth muscule cells (43). Angoipathy is a major complication in diabetes (44 & 45). Endothelial cell damage is suspected to occur in diabetic patients and may be an important cause of angiopathy (46). Animal data on hemodynamic effects of systemic endothelin receptor antagonists are apparently contradictory (47). The physiological relevance of endogenous generation of endothelin-1 in controlling blood pressure has been unclear (10). If basal generation of ET-1 contributes to resistance-vessel tone, then drugs that inhibit the generation or action of endothelin would be expected to cause vasodilatation and decrease blood pressure and might have potential therapeutic value in diseases associated with sustained peripheral vasoconstriction, such as hypertension and chronic heart failure

(11). However, results of animal studies using ECE inhibitors and endothelin receptor antagonists (12, 13 & 14) have been contradictory. In human, systemic administration of an Eta/Etb anagonist causes peripheral vasodilatation and hypotension confirming that endogenous generation of endothelin plays a fundamental role in the maintenance of blood pressure (10 & 48). The data of this work revealed a very highly significant increase of plasma endothelin-1 in type2 (NIDDM) patients as compared with healthy control subjects (P<0.001) and hypertensive control ones (P<0.001) (table 2 and 3). On the other side, when comparing normotensive diabetic one group with hypertensive diabetic one, there was no significant difference (P>0.05) suggesting no effects of increased blood pressure on plasma level endothelin-1, reflects endothelial cell damage and may in turn be related to the complication of diabetes. The elevated endothelin-1 concentration did not appear to be due to secondary hypertension or renal disease, nor was any relationship found between endothelin levels and fasting blood glucose (table 6). The results are in agreement with those of (49 & 50).

30

The increased plasma endothelin-1 in patients with NIDDM could be attributable to chronic hyperinsulinaemia in these patients (51) and/or more endothelial cell damage or dysfunction in NIDDM than in non diabetic subjects (52 & 53). Also, a possible causal factor of the increased ET-1 in NIDDM could be hyperglycaemia because (54) have reported that an elevated glucose level will increase ET-1 production in aortic endothelial cells. Another possible explanation for increased plasma ET-1 level in NIDDM patients is impaired renal neural endopeptidase which is implicated in the degradation of many peptides. In a number of cardiovascular pathologies such as hypertension, the balance in the endothelial production n of vasodilating and vasoconstricting mediators is altered. The resulting apparent decrease in endothelium dependent relaxation is termed endothelial dysfunction (55). This is the first study to demonstrate close relationship between plasma endothelin and IGF system (IGF-1, IGFBP-1 and IGFBP-3) in hypertensive patients with and without type 2 diabetes. Heald et al., (31) found significant negative relationship between IGFBP-I and other known cardiovascular risk factors namely Body mass index (BMI), triglyceride concentrations and insulin level. IGFBPs have been clearly shown to modulate the actions of IGFs. IGFBP-I in particular is accepted as an acute modulator of IGF bioavailability (27). Heald et al., (56) found a close relationship between low IGFBP-1 levels and surrogate markers of cardiovascular risk in a non diabetic population. The data of this work revealed a very highly significant decrease of IGF-I, IGFBP-1 and IGFBP-3 in hypertensive diabetic group versus control group. This is in agreement with Heald et al., (31) who found markedly reduced IGFBP-I in diabetic patients with hypertension and Coni et al., (57) who found reduced level of IGF-I in patients with angina pectoris. The lowest level of IGF-I in this study was found in hypertensive diabetic group (table 2). Heald et al., (31) concluded that low circulating levels of IGFBP-1 are closely correlated with macro-vascular disease and hypertension in

type 2 diabetes. Obese subjects who have mutation associated with higher serum IGF-1 levels seem to be protective factors against cardiovascular complication (58). On the other side, when comparing normotensive diabetic group with hypertensive diabetic one, there was no significant difference (P>0.05) suggesting no effect of increased blood pressure on serum levels of IGF-1, IGFBP-1 and IGFBP-3 in diabetics. While comparing diabetic hypertensive group with sole hypertensive group, there was significant decrease of IGF-I but no significant difference (P>0.05) as regards level of IGFBP-1 and IGFBP-3. Circulating IGF-1 correlated negatively with diastolic blood pressure and systolic blood pressure (P<0.05) in three studied patient groups (table 4). Colao et al., (59) reported that IGF-1 and IGFBP-3 were negatively correlated with common cardiovascular risk factors, studied as total/HDL cholesterol ratio, and/or early atherosclerosis, studied as intima-media thickness at common carotid arteries. They concluded that IGF-1/IGFBP-3 axis plays a role in the pathogenesis of atherosclerosis. Circulating IGFBP-1 showed significant negative correlation with diastolic and systolic blood pressure (P<0.05) in the three studied patient groups. This is in agreement with Heald et al., (31) who found that IGFBP-1 correlated negatively with mean arterial pressure (p=0.002) and this relationship persisted when corrected for age, sex, BMI and insulin, c-peptide concentrations. Also, Heald et al., (31) found IGFBP-1 correlated negatively with systolic blood pressure (p<0.001) and diastolic blood pressure (p<0.00). The relationship between IGFBP-1 and hypertension was robust. Using multiple regression modeling 40% of the variance in IGFBP-1 was accounted for by three variables, mean arterial pressure, triglycerides and non-esterified fatty acids. It is probable that a low circulating IGFBP-1 concentration predates the development of macrovascular disease (31). Our current data suggest that low circulating IGFBP-1 and IGF-1 pattern persists in type 2 diabetic population and further predisposes affected

31

individuals to premature macrovascular disease. In non diabetic hypertensive humans, circulating IGF-1 levels independently predict 16% of the variability in left ventricular mass (60). In another study, a reduction in serum IGF-1 level in relation to bezafibrate treatment in post myocardial infraction patients was independently associated with agiographic measures of coronary artery disease regression (61). Furthermore, in acromegaly, a human model of IGF-1 excess (where world health organization criteria indicate that 37.5% patients have hypertension), blood pressure positively correlates with IGF-1 concentration (62). These findings strongly support an endocrine effect of IGF-1 in vivo (31). It is important, however to recognize that IGFBP-1 is capable of inhibiting endocrine, paracrine and autocrine IGF effects because of its ability to access the extracellular space and to associate with specific cell-surface integrins (63). Thus a high circulating level of IGFBP-1 will inhibit IGF effects whatever the cellular source of the IGF-1. An elevated concentration of IGFBP-1 would therefore serve to protect vascular tissues from the mitogenic potential of circulating IGFs whereas a consistently reduced level induced by the diabetic state per se, hyperinsulinemia or both would increase IGF availability to susceptible tissues. Our finding of decreased IGFBP-1 and IGFBP-3 levels in hypertensive patients with or without diabetes would increase macrovascular disease in these patients. This situation could be further worsened in type 2 diabetes, where reports of increased IGFBP-3 proteolysis (64) would further contribute to the exposure of the vascular compartment to the mitogenic actions of IGFs. Endothelins (ETs) have been shown to have mitogenic properties and could be involved in growth processes in vitro and in vivo (65). ETs have mitogenic effects on endothelial (66) mesangial (67), human ovarian carcinoma cells (68). An unexpected role of the ET system in embryonic and fetal development has been shown and appears

to be essential for cardiovascular and neural crest derived cell differentiation (69). Callera et al., (70) found that increased ET-1 induced oxidative stress in hypertensive rats. In the present study, there is a significant negative correlation between endothelin-1 and measured parameters IGF- system (IGF-1, IGFBP-1 and IGFBP-3) in the three studied patient groups (table 6). Since increased level of indothelin-1 and decreased levels of IGFBP-1 and IGFBP-3 have mitogenic effect on vascular wall. These findings in this study support the hypothesis that ET-1 and IGF system have a role in the development of cardiovascular disease. Conclusion: Significant plasma endothelin-1 elevation in hypertensive patients with and without type 2 diabetes may relate to endothelial cell damage and in turn be an important background factor in vascular complications in these subjects, on the other hand this study demonstrates that low circulating IGF-1, IGFBP-1 and IGFBP-3 may relate to vascular complications in hypertensive and hypertensive diabetic patients, also this study demonstrates the novel and important finding of close links and relationship between low circulating levels of IGF- system (IGF-1, IGFBP-1 and IGFBP-3) and elevated endothelin-1 in hypertensive patients and hypertension with type 2 diabetes.. These findings implicate the role of IGF system and Endothelin-I as predictor markers of cardiovascular risk in patients with hypertension and hypertensive diabetic patients,so ET-I receptor antagonists and IGF-1 replacement may have a role in ameliorating this dysfunction. Reference: 1. Nugent A. G, McGurk C, Hayse JR and

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بمستوى 3 ، 1- والبروتین المرتبط بعامل النمو شبیھ االنسولین1-عالقة عامل النمو شبیھ االنسولین حب ارتفاع ضغط الدم الشریانى المص المصابین بامرضى ال فى1-اندوسیلین

النوع الثانى من السكري الداءحب بمرض اغیر المصو

جادصبري . دو *** عادل زلطة. د و** عبد الحميد السيدمحمد. دو* ام غانمحس.د من أقسام

كلية طب المنصورةالفسيولوجي و *** الطبية الحيويةالكيمياء و ** الباطنة العامةو * اإلكلينيكيةالباثولوجيا

، 1نمو شبيه االنسولين والبروتين المرتبط بعامل ال 1 -األنسولينلقد وجد حديثاً أن عامل النمو شبيه هذا البحث بهدف دراسة حالـة عامـل النمـو شـبيه وقد أجرى . الدموية األوعيةلهم عالقة بتطور أمراض 3

والعالقة بينهم فـى 1- ومستوى االندوسلين 3 و 1 والبروتين المرتبط بعامل النمو شبيه االنسولين 1-االنسولينوقد شمل هذا البحـث .السكريغير المصاحب بمرض الداء المصاحب أو مرضى ارتفاع ضغط الدم الشريانى

وكانت المجموعات كمـا ) حالة15(باالضافة الى المجموعة الضابطة مجموعات 3م الى مهمريضاً وتم تقسي 60 :يلى

فقط وتـشمل عـشرين المجموعة االولى وهم المرضى الذين يعانون من ارتفاع ضغط الدم الشريانى - مريضاً

وهم المرضى الذين يعانون من ارتفاع ضغط الدم الشريانى المصاحب بمرض الـداء المجموعة الثانية - .السكري من النوع الثانى وتشمل عشرين مريضاً

واليعانون من من النوع الثانى السكرى وهم المرضى الذين يعانون من مرض الداء الثالثة المجموعة - . مريضاًالشريانى وتشمل عشرينالدم ارتفاع ضغط

. شخصاً من االصحاء الذين تم اتخاذهم كمجموعة ضابطة15افة الى المجموعة الضابطة باالض - :وقد تم أجراء االتى

والمجموعـة الـضابطة وهـم ضـى تم تشخيص الحاالت اكلينيكياً وتحليلياً وتم أخذ عينة دم من المر :صائمون لعمل التحاليل التالية

البـروتين و1-األنـسولين عامل النمو شـبيه و1-الندوسليناتركيز و وهى نسبة الكرياتينين والسكر بالدم 3-البروتين المرتبط بعامل النمو شبيه االنسولين و 1-بعامل النمو شبيه االنسولين المرتبط

ى مجموعات البحـث ف 1-االندوسلين مستوى في إحصائيةوبفحص نتائج البحث وجد ارتفاع ذو داللة النمـو مـل عاانخفاض ذو داللة احصائية فى مستوى وجد بينما الثالثة عند مقارنتها بنتائج المجموعة الضابطة

مجموعات البحـث الثالثـة عنـد في 3 ، 1 والبروتين المرتبط بعامل النمو شبيه االنسولين 1-شبيه االنسولين فقط عند مقـارنتهم بمرضـى السكري الداءمن مرض يعانون ذينلا كذا ومقارنتها بنتائج المجموعة الضابطة

له عالقة سلبية 1-وقد وجد أن عامل النمو شبيه االنسولين السكري الداءحب بمرض اضغط الدم المرتفع المص 1-احصائية بين مستوى االندوسلين داللة كما وجد عالقة سلبية ذات . داللة احصائية بضغط الدم الشريانى ذات

وقد وجد أنه ال يوجـد اخـتالف ذو داللـة .3 ، 1 به ة والبروتينات المرتبط 1-األنسولينمو شبيه وعامل الن عامل النمـو وأيضاً 3 ، 1 - والبروتين المرتبط بعامل النمو شبيه االنسولين 1-فى مستوى االندوسلين احصائية

فقـط عنـد مقـارنتهم الثانى من النوع يعانون من مرض الداء السكري في المرضي الذين 1-األنسولينشبيه .النوع الثانىمن بمرض الداء السكريبمرضي ضغط الدم المرتفع المصاحب

36

:فى ضوء هذه النتائج يمكن أن نستخلص االتىارتفاع ضغط الـدم الـشريانى ب في المرضي المصابين 1-يوجد ارتفاع ملحوظ فى مستوى االندوسلين

فى مستوى عامل النمو كما لوحظ انخفاض )ع الثانى والن ( السكري الداءحب بمرضى احب أو غير المص االمص وقد تكون هذه التغيرات عامالً مهماً فى حـدوث مـضاعفات 3 ،1به مرتبطةلوتينات ا ر و الب 1-شبيه االنسولين

باستخدام مضادات مستقبالت التقليل من هذه المضاعفات وذلك يمكن لذا فى هؤالء المرضى و االوعية الدموية 1-والعالج بعامل النمو شبيه االنسولين 1-نالندوسليا

37

Transesophageal echocardiography assessment of aortic initma media thickness in correlation to coronary artery disease

Ghareeb M (MD), Gamal A (MD), Elhammady W (MD), Wadeaa B (MD), Essam T (M.Sc), Abdel Aziz A (MD, cardiothoracic department), Cardiology department, Ain Shams University Hospital, Egypt.

BACKGROUND: The purpose of this study was to prove the hypothesis that atherosclerosis of the thoracic aorta assessed by intima media thickness using transesophageal echocardiography (TEE) is correlated with coronary artery disease. METHODS: The thoracic aortas of 70 consecutive patients (50 men and 20 women, mean age 47 +/- 10 years) undergoing elective coronary angiography were assessed by TEE. Aortic intima media thickness, defined as two parallel echogenic lines separated by a relatively hypoechoic space ( the double line pattern ) and atherosclerotic plaques defined as ≥ 3 mm thick focal hyperechogenic zones of the aortic intima and/or lumen irregularities with mobile structures or ulcerations. Correlation between coronary angiography results and TEE results was done. RESULTS: aortic intima media thickness in patients with Normal coronaries was 2.59 +/- 1.22, Single V. disease 3.71 +/- 0.412, Two V. disease 4.16 +/- 0.494, Three V. disease 4.56 +/- 0.697. The intima media thickness, maximum transverse cross-sectional plaque area, the maximum plaque depth and the total plaque number all correlated significantly with the presence of CAD, especially with its severity. Multivariate regression analysis showed that aortic plaques, hypertension and hypercholesterolaemia were significant predictors of CAD, but aortic plaques were the most significant predictor regardless age and sex.

CONCLUSIONS: This study suggests that detection of aortic intima media thickness and atherosclerotic aortic plaques is a useful marker of significant coronary artery disease The common denominator in cardiovascular disease whether, aortic, coronary, cerebral, or peripheral vascular disease is atherosclerosis. Atherosclerosis is a diffuse process that seems to begin in the aorta and spreads to its branches. (1) Previous roentgengraphic studies have attempted to correlate the presence of atherosclerotic aortic plaque and coronary artery disease. The presence of thoracic aortic plaque on chest X-ray film was associated with an increased risk of cardiovascular death. (2) However, the limited resolution of X-ray studies for aortic plaque resulted in the inability to detect plaque consistently and decreased the sensitivity and specificity of the chest roentgenogram as a marker for coronary artery disease. With improved non invasive imaging techniques now available, it is necessary to determine whether atherosclerosis of the aorta can be reliably detected and correlated with coronary artery disease in a clinically useful manner.Transesophageal echocardiography

offers high resolution imaging for the evaluation of thoracic aortic disease. This technique has been instrumental in identifying thoracic aortic aneurysms and dissections, potential aortic sources of embolism and abnormal vascular connections. (3-5) It has also been utilized to avoid vascular complications during coronary bypass surgery and intraaortic balloon pump placement.(6-8) The close proximity of the transesophageal probe to the thoracic aorta allows high frequency imaging and superior resolution of intraluminal structures,particu-larly atherosclerosis. Aim of The Work To prove the hypothesis that atherosclerosis of the thoracic aorta assessed by intima media thickness using transesophageal echocardiography is correlated to coronary artery disease. Patients and Methods This study is a prospective case-control study. It was conducted at Ain Shams University hospital on 70 patients who underwent coronary angiography for variable indications (angina either recent-onset, or post myocardial infarction (MI); recent

38

myocardial infarction (MI), or preoperative coronary angiography prior to valve replacement). The study group included 70 patients studied over the period from 3/2003 to 10/2003. Fifty of them had coronary artery disease documented by coronary angiography, and twenty of them had normal coronaries documented by coronary angiography.All subjects examined were informed about the procedure and an oral consent was taken regarding acceptance to participate in the study.All the patients were aged from 40-60 years of age. Each individual was subjected to the following: 1- Thorough history taking with special stress on the following: A) Risk factors including: i) Smoking: Cigarette smoking alone has also been shown to increase the IMT of many types of arteries. In a study of 184 cigarette smokers ( aged 44 +\- 9 years) for whom smoking was the only cardiovascular risk factor, the carotid, aorta and femoral IMTs were significantly larger then in 56 non-smokers matched for age and gender. Subjects were classified as:

1- Smokers: combining the ex-smokers & current smokers in one group. Exsmokers (if they had ceased smoking for >6 months), or current smokers. Information on the duration of smoking in years and the average number of cigarettes smoked per day for each current and former smoker was asked for. Calculating the smoking index, which equals the result of multiplication of the duration of smoking in years and the average number of cigarettes smoked per day. 2- Non-smokers: patients who never smoked cigarettes or any nicotine derivative. ii) Hypertension: Its duration, and if present; its control using anti-hypertensive drugs or not. iii) Diabetes Mellitus: Duration of the disease if present, its type (juvenile type I, or maturity onset type II) and its mode of control using oral hypoglycemic agent or insulin. iiii) Family history: Familial risk factors (hypertension, DM, and dyslipidemia) or vascular disease including

coronary artery disease or peripheral vascular disease. B) Coronary artery disease: History of angina either chronic stable form or unstable form of anginal pains and/or myocardial infarction. C) Peripheral arterial disease: History of intermittent claudication pain and/or color changes, atrophic changes (loss of hair, wasting, nail dryness…), or gangrene. D) History of exclusion criteria: Contraindication to TEE

• Esophgeal disease • Aortic aneurysm • Aortic coarctation • Aortic dissection • Previous aortic surgery • Collagen disease

2)Thorough clinical examination: (A) General examination with special emphasis on: i) Pulse: Both carotid, brachial, popliteal, and femoral and dorsalis pedis arteries were examined for rate, rhythm, volume, special character and equality of pulsations. (B) Local examination: The first and second heart sounds were auscultated for any abnormality. Also, the presence of additional sounds or murmurs was looked for 3) Laboratory investigations * Fasting blood sugar: In our study patients were divided into: - Diabetics: patients being diagnosed as diabetics by the, by history in patients known to be diabetics, being on medication or the patients with FBS > 140 mg/dl. - Non-diabetics not fulfilling the previous parameters. * Serum total cholesterol, and triglycerides levels: They were determined on a blood sample drawn after 14 hours fast. The normal ranges for our lab were as follows: Serum cholesterol 150-250 mg/dl Serum triglycerides 40-160mg/dl. Hypercholesterolemia is associated with increased carotid and femoral IMTs.

39

4) Coronary angiography: Coronary arteriography was done via the right femoral artery approach using the Seldinger’s technique. Left and right coronary arteriography were done in multiple projections (RAO 30, LAO 55, left lateral projection for left coronary system and RAO 45 and LAO 45 with caudal 15 for right coronary system) using left Judkin’s and right Judkin’s catheters respectively. The state of vessels was judged using multiple projections by at least two experienced observers. Stenotic coronary arteries were measured in the projection, in which the stenosis was judged to be the most severe. Coronary artery stenosis of greater than or equal to 50 percent of the luminal diameter was considered hemodynamically significant. Patients were classified as having one vessel, two vessel or three-vessel disease +/- left main affection. Ramus intermedius branch which is best visualized in left anterior oblique projection with caudal angulation(spider view), and also subdivided into three segments. According to this angiographic data the patients were divided into two groups Group A with normal coronary angiograms, and Group B if having significant coronary lesion(s). Then Group B patients were further classified according to the number of vessels affected; Patients with single vessel affection, patients with two vessels affected and patients with multivessel affection. 5)Transesophageal Echocardiography: The TEE probe that was used in this study was a GE Vingmed 5 MHz multiplane probe connected to a GE Vingmed Vivid 5 echocardiography system, which was connected to a central station where the off line analysis was done. 6) Statistical Analysis: All the data obtained from the patients were tabulated and statistically analyzed using an IBM compatible computer. Windows SPSS. Version 8. Program was used for analysis of data. Results : The study group included 70 consecutive patients who were subjected to coronary angiography for different reasons.

Patients were divided into two groups according angiographic findings: Group A (Controls): Included 20 patients with normal coronary angiograms. Group B (Cases): The patients were fasting for at least 6 hours before the examination. The oropharynx was aneasethized with topical Lidocaine spray and light intravenous sedation. The examination was performed with the patient lying in a supine and slightly lateral position. For imaging the entire thoracic aorta, the TEE probe was rotated posteriorly and advanced to the distal esophagus, the probe was then slowly withdrawn to scan the descending aorta and the aortic arch. It was rotated and advanced again to image the ascending aorta. All images were recorder on the Echopac central station for further offline assessment. The intima media thickness was measured by calculating the distance between the two parallel echogenic lines separated by a relatively hypoechoic space ( the double line pattern ).9 Three frozen images of three segments of the aorta, ascending, arch, and low descending aorta were recorded, and the average thickness was calculated for each side. And then the average IMT for the three segments was calculated. Ultrasound instrument gain settings were dependent on the ultrasound properties of the individual subject and set such that trailing of the interfaces was at a minimum. All scans were done by the same operator to avoid inter-observer variability. Included 50 patients with significant coronary lesions, which was defined as 50% or more luminal stenosis, visualized in their coronary angiograms. According to the number of vessels affected, they were further classified into patients with single vessel affection (n=26), patients with two vessel (n=18) and patients with multi vessel affection (n=6).

40

Group A Group B P value Number 20 70 NS Mean age 47 ± 8.9 52 ± 2.1 NS DM 6 (30%) 20 (30%) NS Sm 8 (40%) 35 (70%) NS Chol 202 ± 54 237 ± 42 HS P < 0.001 TG 140 ± 27 165 ± 51 S P < 0.05 Table (1) Demographic data. Studying the relationship between aortic intima media thickness and various risk factors of coronary artery disease, we found that there is a significant correlation between aortic intima media thickness and DM (P = 0.04), cholesterol (P = 0.0008), TG (P= 0.039) and smoking index ( P= 0.046), however a nonsignificant correlation was found between aortic intima media thickness and history of smoking (P= 0.087) and family history of CAD. Average aortic intima media thickness in the normal coronaries group was 2.59±1.23 mm, in the single vessel group 3.72±0.41 mm, in the two vessel group 4.16±0.59 mm, and in the three vessel group 4.57±0.69 with a P value of 0.0002. It is clear from the previous figures and with calculating the statistical significance using the ANOVA test that there is statistically significant difference among the previous readings. It is also evident that the aortic intima-media thickness increases with the increase in the number of the vessels affected.

Table (2) Comparison of aortic intima- media thickness among groups with different vessel affection Table 2 shows that the difference of AoIMT between the normal and CAD groups was significant, and the value of AoIMT in the CAD group was more than that in the normal group (P<0.001). The value of AoMT in the two- or three-vessel disease group was more than that in the one-vessel disease group (P<0.001). In our study finding aortic intima media thickness of the thoracic aortas on transesophageal echocardiography test had a sensitivity of 91.9% for obstructive coronary artery disease and a specificity of 76.9%. The positive predictive value of aortic plaque for obstructive coronary artery disease was 91.9% and the negative predictive value was 76.9%. The accuracy of transesophageal echocardiography as predictor of obstructive coronary artery disease was 88.0% in our study.

Ascending Arch Descending Average

Mean (SD) Mean (SD) Mean (SD) Mean (SD) Normal coronaries 2.39 1.25 2.63 1.27 2.76 1.286 2.59 1.22 Single V. disease 3.67 0.400 3.67 0.489 3.79 0.388 3.71 0.412 Two V. disease 4.078 0.477 4.16 0.507 4.24 0.535 4.16 0.494

Three V. disease 4.485 0.699 4.64 0.734 4.58 0.710 4.56 0.697 F. Ratio 20.40 15.52 14.36 17.753 P. Value P<0.001 P<0.001 P<0.001 P<0.001

Table (3) correlation of the Ao IMT in different areas of the aorta and the extent of coronary atherosclerotic disease represented by number of vessels affectedThe occurrence rates of thoracic aortic plaques were significantly different between the normal group and the one-, two- or three-vessel disease group. They were gradually increased from the normal group to the one-, two-, and three-vessel disease groups. Thoracic aortic plaques were found in all patients of the three-vessel group

With transesophageal echocardiography, observers could not only find atherosclerotic plaques, but also accurately evaluate the profile, constituent and steadiness of the plaques. Discussion Today, transesophageal echocardiogr-aphy has been widely used to assess thoracic

≥Three vessels

Two vessels Single vessel

No vessel

affection

4.5683 mm

4.1611 mm 3.7165 mm 2.5925 mm

Mean of I.M.T.

0.6971 0.4947 0.4124 1.2295 S.D.

17.753 F value P=0.0002 Significance

41

aortic atherosclerotic conditions for clinical diagnosis and research (11, 12). Coronary artery atherosclerosis resulting in coronary artery disease may not be an isolated disease condition. It quite often coexists with aortic atherosclerosis, which is one part of the widespread atherosclerosis in the group of large-middle arteries in patients. However their pathological basis is identical. Our results have shown that the thoracic aortic intima media thickness detection by transesophageal echocardiography appears to be a useful marker for predicting the presence of coronary artery disease with 91.9% sensitivity and 91.9% positive predictive value. These findings are consistent with the observations of Fazio et al (13), and further substantiate the instinctive relation between thoracic aortic plaque and coronary artery disease, and the concept that atherosclerosis is a generalized process involving predominantly arteries of large and medium size. These findings are in accord with the pathological process, where atherosclerosis occurs on the aorta at first and gradually becomes widespread to its branches (14, 15). In our study, the specificity and negative predictive value of thoracic aortic plaque predicting coronary artery disease were all 76.9%, which is lower than the results of Fazio et al (13) and Tribouilloy et al (16). This could be due to the high-risk people of this group, since they are different from the other study groups, and the number of normal cases is relatively less. These findings further substantiate the correlation between the occurrence of thoracic aortic plaque and the severity of coronary artery disease. From this correlation, we could know the approximate severity of coronary artery disease from thoracic aortic plaques detected by transesophageal echocardiography References: 1. Eggen DA, Solberg LA. Variation of

atherosclerosis with age. Lab Invest 1968;18:571-9.

2. Freedman DS, Newman WP III, Tracy HE, Vuors RE, Srinivasan SR, Webber LS, et al. Black-white differences in aortic fatty streaks in adolescence and early adulthood: the Bogalusa Heart Study. Circulation 1986;77:856-64.

3. Karalis UG. Chandrtsekaran K. Victor MF, Ross J Jr. Mintz GS. Recognition and embolic

potential of intraaortic atherosclerotic debris. J Am Coll Cardiol 1991:17:73-8.

4. Tunick PA. Perez JL, Kronzon I. Protruding atheromas in the thoracic aorta and systemic embolization. Ann Intern Med 1991;115:423-7.

5. Barzilai B. Marshall WG. Salfttz JE, et al. Avoidance of embolic complications by ultrasonic characterization of the ascending aorta. Circulation 1989:809 (suppl l):I- 275.

6. Hosoda Y. Wamtamabe M. Hirooka Y. Ohse Y, Tanaka A, Watanabe T. Significance of atherosclerotic changes of the ascending aorta during coronary by pass surgery with intraoperative detection by echography. J Cardiovasc Surg. 1991:3Z:301-6.

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