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Pathogenesis and genetics of anemia due to decreased red cell productionBy Nada HosnyUnder supervision of Dr.Hanaa Fahmy1Definition of AnemiaReduction in concentration of circulating Hemoglobin or oxygen carrying capacity of blood below the level that is expected for healthy persons of same age and sex in the same environment.Etiological classificationAnemia due to impaired red cell production.Anemia due to excessive red cell destruction (hemolytic anemia).Anemia due to excessive blood loss.Anemias caused by decreased red blood cell production have been grouped under the broad category of bone marrow failure, although their mechanisms and/or presentations may be markedly different. Anemia due to decreased productionClassificationA- AcquiredA- Aplastic anemia1- pluripotential stem cell failure2- Anemia of leukemia and MDS3- Anemia associated with BM infiltration4- Post-chemotherapyB- Erythroid Progenitor red cell failure1- Pure red cell aplasia2- Endocrine disorders3- Acquired sideroblastic anemiaC- Functional impairment of erythroid and other progenitors due to nutritional and other causes1- Megaloblastic anemia2- Iron deficiency anemia3- Anemia resulting from other nutritional deficiencies4- Anemia of chronic disease and inflammation5- Anemia of renal failure6- Anemia caused by chemical agents (Lead toxicity)7- Acquired thalassemias8- Erythropoietin Antibodies

B-Hereditary:A- Pluripotential stem cell failure:1- Fanconi Anemia2- Shwachman syndrome3- Dyskeratosis congenitaB- Erythroid progenitor cell failure1- Diamon blackfan syndrome2- Congenital dyserythropoietic syndromeC- Functional impairment of erythroid and other progenitors due to nutritional and other causes1- Megaloblastic anemia2- Inborn purine and pyramidine metabolism defects3- Disorders of iron metabolism4- Hereditary sideroblastic anemia5- Thalassemias

Anemias due to RBCs production Pure red cell aplasia, either congenital or acquiredBone marrow aplasia involving more than one cell lineAnemia related to ineffective erythropoiesis or disordered erythroid maturationAnemia caused by decreased erythropoietin availabilityAnemia caused by marrow replacement (malignancy, storage disease)Pure red cell aplasia ( PRCA)Definition :A group of disorders characterized by Severe anemiaReticulocytes or equal 2 populations of cells with different genotypes in 1 person who has developed from single fertilized ova.

D- immunoblotting for FANCD2There are several FA genes 12 complementation groups (FA-A-M) DNA damage (MMC) FA protein complex (A/C/E/F/G complex) monoubiquitination of FANCD2(Ub) targets D2 to DNA repair nuclear foci containing BRCA1, BRCA2, and RAD51, which are important in maintaining genomic stability by promoting homologous recombination repair.

E- other investigationsFlow cytometry to identify mutant geneElevated alpha fetoprotein independent of liver complications or android therapyWhen FA cells exposed to alkylating agent; they arrest at G2/M phase of cell cycle.PrognosisMost of patients develop early BM failure except thiose with biallelic BRAC2 mutation which are prone to (early onset AML, Brain tumors, Wilims tumors.Increased risk of:Solid malignant tumorsLiver tumorsAMLMDSHSCT now improved prognosis of FA dramatically with median survival reaching >30 years.Dyskeratosis Congenita (DC)Clinical featuresClassical DC is an inherited multisystem disease including Mucocutaneous and hematopoietic abnormalitiesCharacterized by the triad of abnormal skin pigmentation, nail dystrophy and mucosal leucoplakiaSomatic abnormalities of DC

Triad of DC

Clinical manifestations in DCOften appear during childhood.The skin pigmentation and nail changes typically appear first, usually by the age of 10 years. BM failure usually develops below the age of 20 years; 8090% of patients will have developed BM abnormalities by the age of 30 years. Cause of deathBM failure/immunodeficiency (~6070%), Pulmonary complications (~1015%) Malignancy (~10%).:Epithelial cancers of GI tractMDS/AML10% develop cancers before the age of 40

Cell and molecular biology

11%31%4%8%46%X-linked dyskerinAutosomal recessiveNOP10, NHP2, TERTAutosomal dominantTERT,TERCSporadic TINF2UnknownGenetics of Dyskeratosis Congenita4444DC mutations Gene Inheritance EffectsDKC1X-linkedDCTINF2AD Severe sporadicNOP1ARDC rareNHP2ARDC rareTERCADDC AA PF MDS TERTADDC AA PF MDS AML CLL others4545Main manifestation of 3 modes of inheritance is defective telomere maintenance which lead to abnormal telomere shortening and forced apoptosis.Also all hematopoietic progenitor cells numbers are found to be reduced in DC which give rise to pancytopenia. Anemia caused by decreased erythropoietin availability

The kidneys are responsible for approximately 90% of erythropoietin production in an individual. Individual with normal functioning kidneys : hypoxia EPO gene transcription increased RBC productionIn those with anemia of CKD : there is primary deficiency of EPO production by the type I interstitial fibroblasts, thereby leading to anemia. The anemia that develops is directly related to the amount of residual renal functionOther chronic diseases For example RA, severe trauma, heart disease, liver disease or DM . In these conditions, there is Decreased availability of ironRelatively decreased levels of EPOMild decrease in the lifespan of RBCs to 70-80 days Cytokines e.g. (IL-1 and IL-6), (TNF-alpha), which are believed to cause (destruction of RBC precursors / EPO receptors on progenitor cells.

SchwachmanDiamond syndromeIt is now recognized as an Autosomal recessive disorder characterized by exocrine pancreatic insufficiency (100%), bone marrow dysfunction (100%) and other somatic abnormalitiesSigns of pancreatic insufficiency (malabsorption, failure to thrive) are apparent early in infancy Other common somatic abnormalities include short stature (~70%), protuberant abdomen and an ichthyotic skin rash (~60%) and metaphyseal dysostosis ( ~75% )Other abnormalities include hepatomegaly, rib/thoracic cage abnormalities, syndactyly, cleft palate, dental dysplasia, ptosis and skin pigmentation.Haematological abnormalitiesIncludes neutropenia (~60%)Other cytopenias (pancytopenia: 20% ), myelodysplasia and leukaemic transformation (~25%). The age at which leukaemia develops varies widely from 1 to 43 years. AML is the commonest (M/F ratio = ~9:1).Cell and molecular biologyMajority (> 90%) of SDS patients have been found to have mutations in SDS gene on 7q11 . Its precise function is unknown It is predicted to have an important role in RNA metabolism and/or ribosome biogenesis.Several abnormalities in SDS cells have been observed, including haemopoietic stem and stromal defects, increased rates of apoptosis and short telomeres. Pearson syndromeIs a mitochondrial disease (caused by a deletion in mitochondrial DNA.) Characterized by sideroblastic anemia and exocrine pancreas dysfunction. Other clinical features are failure to thrive, pancreatic fibrosis with insulin-dependent diabetes and exocrine pancreatic deficiency, muscle and neurologic impairment, and, frequently, early death. It is usually fatal in infancy. The few patients who survive into adulthood ( 100 reported) often develop symptoms of Kearns-Sayre syndrome.KearnsSayre syndrome(abbreviatedKSS) also known asoculocraniosomatic disorderorOculocraniosomatic neuromuscular disorder withragged red fibersis amitochondrial myopathywith a typical onset before 20 years of age. KSS is a more severe syndromic variant ofchronic progressive external ophthalmoplegia(abbreviated CPEO), a syndrome that is characterized by isolated involvement of the muscles controlling eyelid movement (levator palpebrae, orbicularis oculi), and those controlling eye movement (extra-ocular muscles). This results in ptosis and ophthalmoplegia respectively. KSS involves a triad of the already described CPEO, as well as bilateral pigmentary retinopathy, andcardiac conduction abnormalities.53Aplastic anaemias (AA)Aplastic anaemia is defined as the presence of pancytopenia in the PB and a hypocellular BM in which normal haemopoietic marrow is replaced by fat cellsDiagnosis of AA requires at least 2 of the following in addition to a hypocellular marrow: (i) haemoglobin < 10 g/dL;(ii) platelet count < 100 109/L; and (iii) neutrophil count < 1.5 109/L. - Abnormal cells are not found in either the PB or the BM. - The diagnosis is based on the absence of cells, not the presence of any characteristic feature.Classes of AA

Inevitable AAMost cytotoxic drugs, penetrating ionizing radiation and radioactive substances concentrated in the marrow are capable of producing AA through their effects on actively dividing cells.Idiosyncratic acquired AAAcquired AA may occur spontaneously or appear following exposure to drugs or viruses that do not produce marrow failure in the great majority of persons exposed to these agents. The disease is characterized by its unpredictable onset and prolonged course, death usually occurring as a result of a deficiency of granulocytes or platelets and a failure in support measures.Drugs associated with idiosyncratic acquired AA.

Viruses associated with idiosyncratic acquired AA. Hepatitis A, B or CEpsteinBarr virus infection.Human immunodeficiency virus (HIV)

PathogenesisThe pathogenesis of AA remains unclear, but an autoimmune mechanism appears to be important. There may unidentified underlying genetic predisposition. There is some association of (HLA) DR2, specifically the DR15 split, with acquired AAThere is debate about whether the primary defect is in the haemopoietic stem cell itself or is the result of environmental factors, particularly immunological attackThere is evidence of both quantitative and qualitative stem cell defects in AA and increased apoptosis of remaining early haemopoietic progenitor cells. Not only do cytotoxic T lymphocytes release cytokines, such as interferon- and TNF-, that are inhibitory to haemopoietic progenitor cells, but TNF- also upregulates Fas antigen expression on CD34+ cells reduced survival of AA marrow progenitor cellsTelomere shorteningPatients with acquired AA show increased terminal restriction fragments (TRF) loss in leucocytes compared with age-matched control subjects, and the extent of the loss correlates with the duration of disease. In patients who make a full haemopoietic recovery, the rate of TRF loss returns to normal for ageing.HaematologyThe peripheral blood film shows pancytopenia without gross morphological abnormalities in the circulating cells. There maybe some macrocytosis of remaining red cells, usually with an absolute reticulocytopenia. WBCsGranulocytes toxic granulation of neutropenia.Monocytes reduced lymphocyte count reduced.Platelets reduced and of small and uniform size.The bone marrow aspirateIs normally easily obtained, typically with many fragments, which appear hypocellularThe cell trails are hypocellular, with relative increase in lymphocytes and plasma cells Remaining haemopoietic precursors are normal in appearance.In the early stages of AA haemophagocytosis may be prominent. In a high proportion of cases, the hypocellularity of the marrow is patchy, with quite extensive areas of cellular marrow remaining.Misleading ?? A trephine biopsy (sometimes more than one) is necessary to assess cellularity properly. The bone marrow trephine Shows the fat replacement of marrow with or without the remaining islands of cellularityNon-haemopoietic cells remain, sometimes giving the impression of a chronic inflammatory infiltrate.Reticulin is not increased.

Definition of disease severity of AA (Camitta scoring system)

Remember The most common mistake in the diagnosis of aplastic anaemia is to make the diagnosis on the basis of a bone marrow aspirate in the presence of pancytopenia without obtaining adequate trephine specimen. Cytogenetic analysis of the bone marrow should be performed DD of pancytopenia and a hypocellular bone marrow Hypocellular myelodysplastic syndromeHypocellular acute myeloid leukaemiaHypocellular acute lymphoblastic leukaemia.Hairy cell leukaemiaLymphomaMyelofibrosis Mycobacterial infections Anorexia nervosa or prolonged starvationClinical presentationNon-specific related to pancytopenia. Easy bruising or petechiae, bleedingFeeling tired from anaemia. Sometimes infection is the presenting featureNO {Spleen , Liver , LNs, Jaundice}Anemia caused by decreased erythropoietin availability

The kidneys are responsible for approximately 90% of erythropoietin production in an individual. Individual with normal functioning kidneys : hypoxia EPO gene transcription increased RBC productionIn those with anemia of CKD : there is primary deficiency of EPO production by the type I interstitial fibroblasts, thereby leading to anemia. The anemia that develops is directly related to the amount of residual renal functionOther chronic diseases For example RA, severe trauma, heart disease, liver disease or DM . In these conditions, there is Decreased availability of ironRelatively decreased levels of EPOMild decrease in the lifespan of RBCs to 70-80 days Cytokines e.g. (IL-1 and IL-6), (TNF-alpha), which are believed to cause (destruction of RBC precursors / EPO receptors on progenitor cells.

Anemia related to ineffective erythropoiesis or disordered erythroid maturation

ThalassemiasIron deficiency anemias Congenital dyserythropoietic anaemiasSideroblastic anemiaMegaloblastic anemia Thalassemias

Hereditary disorders that can result in moderate to severe anemiaBasic defect is reduced production of selected ( one or more) globin chainsNet result: imbalanced globin chain production excess globin chains tetramers ppt. in RBCs chronic hemolysis in BM and PBSeverity varies depending on type of mutation or deletion of the or globin chain

Alpha ThalassemiasResult from gene deletionsOne deletionSilent carrier; no clinical significanceTwo deletionsa Thal trait; mild hypochromic microcytic anemiaThree deletionsHgb H; variable severity, but less severe than Beta Thal MajorFour deletionsBarts Hgb; Hydrops Fetalis; In Utero or early neonatal deathBeta ThalassemiasResult from Point Mutations on genesSeverity depends on where the hit(s) lieb0-no b-globin synthesis; b+ reduced synthesisDisease results in an overproduction of a-globin chains, which precipitate in the cells and cause splenic sequestration of RBCsErythropoiesis increases, sometimes becomes extramedullaryPathogenesisReduced or nonexistent production of b-globinPoor oxygen-carrying capacity of RBCsFailure to thrive, poor brain developmentIncreased alpha globin production and precipitationRBC precursors are destroyed within the marrowIncreased splenic destruction of dysfunctional RBCsAnemia, jaundice, splenomegalyHyperplastic Bone MarrowIneffective erythropoiesisRBC precursors destroyedPoor bone growth, frontal bossing, bone painIncrease in extramedullary erythropoiesisIron overloadincreased absorption and transfusionsEndocrine disorders, Cardiomyopathy, Liver failureIRON DEFICIENCY ANEMIAMost common cause of anemiaIt develops when iron stores are depleted, level of circulating iron is reduced and there is insufficient iron available for erythropoiesis.ETIOLOGY: EXCESSIVE LOSS OF IRON (CHRONIC BLEEDING )MENORRHAGIA PEPTIC ULCERSTOMACH CANCERULCERATIVE COLITISINTESTINAL CANCERHAEMORRHOIDSDECREASED IRON INTAKEINCREASED IRON REQUIRMENT (JUVENILE AGE, PREGNANCY, LACTATION)DEFECTIVE IRON ABSORPTION

IRON DEFICENCY - STAGESPrelatent reduction in iron stores without reduced serum iron levelsHb (N), MCV (N), iron absorption (), transferin saturation (N), serum ferritin (), marrow iron ()Latentiron stores are exhausted, but the blood hemoglobin level remains normalHb (N), MCV (N), TIBC (), serum ferritin (), transferrin saturation (), marrow iron (absent)Iron deficiency anemiablood hemoglobin concentration falls below the lower limit of normalHb (), MCV (), TIBC (), serum ferritin (), transferrin saturation (), marrow iron (absent)