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team-work that lies before us in this country. A relativelysimple analysis would show that there are many occupa-tions particularly suitable for the men and women whohave reached the age of retirement, according to theirintelligence and experience. For many the curse ofoneliness’ could be exorcised, and- the days and yearsrenewed with vigour and interest. The selection pro-cedure should of course be positive, based on capacitiesand not on disabilities. In dealing with the disabledthis system has been effective, indicating the type ofwork that the applicant can do rather than setting out therange of jobs for which he is unfit.The whole subject is a delicate one in which we have to

tread warily ; but it is urgent and serious enough to beregarded with clear eyes, unclouded by either misplacedfrivolity or befuddled sentimentality :

I have not writ this letter of diviningTo make a glory of thy silent pining,A triumph of thy mute and strange declining.

So spoke Alice Meynell in " A Letter from a Girl to herown Old Age " ; and the neglected Browning put the wholematter in a nutshell thus :

And the whole is well worth thinking o’erWhen autumn comes : which I mean to do

One day, as I said before. 4

BIOCHEMISTRY OF INFLAMMATION

VALY MENKINM.D. Harvard

ASSOCIATE PROFESSOR OF EXPERIMENTAL PATHOLOGY,TEMPLE UNIVERSITY PHILADELPHIA

INFLAMMATION is a fundamental process in higheranimals, which involves lymphatics, blood-vessels, andthe locally affected tissue, and is initiated by a disturb-ance in the fluid exchange. The normal capillaryfiltration equilibrium is upset, one important changebeing an increase in capillary permeability, which is

readily demonstrable by the seepage of material intro-duced into the circulation. Diazo dyes (such as trypanblue), ferric chloride, graphite particles, and bacteriawill readily concentrate from the circulation into acutelyinflamed foci (Menkin 1940a). Since this increasedcapillary permeability is an initial stage of inflammation,knowledge of its mechanism is essential to the under-

standing of subsequent developments.LEUKOTAXINE

The early work of Lewis (1927) postulated that theincreased capillary permeability in inflammation was

primarily due to the presence in weal fluid of a

hypothetical H-substance, which was presumed to behistamine. I have not been able to confirm this (Menkin1936) ; in fact, the following observation suggests theliberation into inflammatory exudate of a substancewhich seems to have no direct biological relation tohistamine (Menkin 1938a).

If inflammatory exudate is injected into the normal skinof a rabbit, and this is immediately followed by theintravenous injection of trypan blue, the dye rapidlyaccumulates in the treated area of the skin, indicatingthe probable presence in the exudate of a factor whichincreases permeability. This factor is absent fromblood-serum, and the reaction is not elicited by physio-logical saline. The active substance, "leukotaxine," isthermostable and diffusible through a ’ Cellophane’membrane. By treating inflammatory exudate withpyridine and then acetone to remove proteins a

crystalline-like active material can be recovered ina butyl alcohol mother liquor. Leukotaxine can also be

precipitated by treating the exudate with ammoniumsulphate to half saturation.

4. Robert Browning. By the Fireside.

Various methods have been devised to extract thisactive factor (Menkin 1940a). The present method, some.what modified from the original scheme, is as follows :Exudate

.’

Pyridine (I : 1)I

Centrifuge

SupernatantAcetone (1 : 1)

Supernatant

Dried on steam bath at about 70°CButyl alcohol (0-5 : 1 part of acetone supernatant)Stir for 24 hours

Concentrate to 1/10 vol. on steam bath at about 70°C

Partially dry by freezing with CO2 snow

Crystalline-like material (leukotaxine)The active factor, leukotaxine, is biuret-negative but

ninhydrin-positive. The Millon test is negative, but theAdamkiewicz colour-reaction for the indole nucleus ispositive. The xanthoproteic test for the phenyl group islikewise positive. Chemically, leukotaxine seems to bea relatively simple polypeptide to which there may beattached an unidentified prosthetic group, but thishas not yet been established. Studies on the enzymaticeffect of crystalline trypsin on an otherwise inert serumalbumin or whole serum support the view that the

biological activity may largely be confined to the

peptide linkage (Menkin 1938b). Finally, amino-nitrogenmeasurement before and after hydrolysis indicates thepossibility that the active principle be a relativelysimple peptide.

Leukotaxine not only increases capillary permeabilitybut also induces the migration of polymorphs throughthe capillary walls. This chemotactic property can bedemonstrated both in vitro and in vivo (Menkin 1940a).The liberation of leukotaxine is therefore a reasonableexplanation of the increased capillary permeability andmigration of leucocytes in inflammation. The yield ofleukotaxine from inflammatory exudates is relativelysmall, but its activity is well marked. Workers in

England have confirmed the presence of leukotaxinein such exudates (Cullumbine and Rydon 1946), using asomewhat different technique of extraction. Further,these workers have pointed out the abundance of leuko-taxine in the succus entericus of the rabbit. In Japan,Minami and Inugami (1940) have also found leukotaxinein inflammatory exudates.

Recent studies indicate that leukotaxine or a similarsubstance can sometimes be recovered from variousother body fluids, such as those in ovarian cysts, cranio-pharyngiomata, and gliomata (astroblastomata). It isconceivable that the formation of fluid in these

pathological conditions is partly due to the localproduction of leukotaxine.

Increased capillary permeability, caused by the libera-tion of leukotaxine, allows the free passage of plasmaproteins from the circulation into the extracapillaryspaces. Fibrinogen is converted into fibrin, with therelease of thrombokinase, after injury to cellularstructures. A network of fibrin is formed in tissuedistended with oedema, and lymphatic channels becomeplugged with fibrin. This early lymphatic obstructionprevents the ready dissemination of material or micro-organisms from a focus of acute inflammation. This

discovery adds to our understanding of the r6le ofinflammation in immunity (Menkin 1938c).

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LEUCOCYT08IS-PROMOTING FACTOR

Leukotaxine is responsible for the local migration ordiapedesis of leucocytes to the site of injury ; it doesnot affect -4-11f number of circulating leucocytes. Yet

leucocYL_, )’’ten accompanies inflammation or infection.Now, injection of inflammatory exudate from an animalor a man with leucocytosis into the blood-stream of anotherwise normal dog or guineapig can cause leucocytosis.It is reasonable to conclude that the inflammatoryexudate contains a leucocytosis-promoting factor liberatedby injured cells (Menkin 1940b, Menkin et al. 1946).This leucocytosis-promoting factor (L.P.F.) has, indeed,been found ; it is non-diffusible and thermolabile, andresembles a protein. It is obtained from the exudate bytreating it with ammonium sulphate to half saturationafter removal of the euglobulin fraction (Menkin 1940c1946a, Menkin and Kadish 1943), and seems to be eithera pseudoglobulin or at least closely associated with thatfraction of the exudate. Unpublished studies in collabora-tion with Gerald Cooper, of Duke University school ofmedicine, using a Tiselius apparatus, indicate that theL.P.F. is closely associated with the alpha globulins ofinflammatory exudates.The L.P.F. induces not only a discharge of immature

leucocytes from the bone-marrow but also hyperplasiaof granulocytes and megacaryocytes in the bone-marrow(Menkin 1943a). The presence of the L.P.F. isdemonstrable only in the serum of an animal with acuteinflammation, and not in a normal animal’s blood-serum. The liberation of the L.P.F. into inflammatoryexudate and its absorption into the circulation offera reasonable explanation of the leucocytosis associatedwith inflammation.The apparent and essential non-antigenicity of the

L.P.F. (Menkin 1944) and its property of stimulating theproduction of leucocytes in the bone-marrow suggestdefinite clinical applications, especially since it is wellknown that the prognosis of numerous infections dependslargely on the number of circulating leucocytes. Recentstudies at Duke University have demonstrated that theintravenous injection of canine L.P.F. into human beingsis both innocuous and effective. The doses used rangedfrom 18 to 230 mg. The injection of the material intohuman beings is followed by a discharge of immaturegranulocytes into the circulation (Menkin 1946a). Theclinical implications of these studies are obvious. Alreadya patient with a’persistent leucopenia has respondedfavourably for short periods to the intravenous injectionsof relatively low doses of the L.P.F. Similar studies arenow in progress.

NECROSIN

Inflammation is a manifestation of severe cellular

injury. Neither leukotaxine nor the L.P.F. per se can

reproduce that injury. The injury seems to be due to theliberation of a substance located in or associated with theeuglobulin fraction of the exudate, particularly whenthe exudate has an acid reaction when withdrawnfrom the site of inflammation. This substance hasbeen termed necrosin (Menkin 1943b).The injection of necrosin into the skin of a rabbit is

followed within several minutes by swelling and fusionof the collagenous bundles ; this type of response cannotbe induced by the injection of the euglobulin fraction ofliormal blood-serum. After a longer interval inflam-mation develops, with intense redness, oedema, andsuperficial necrosis. An abundance of leucocytes maybe found in such areas. Lymphatics are occluded byfibrin, and blood-vessels may be thrombosed. Theeuglobulin fraction of normal blood-serum does notinduce lymphatic obstruction, which in turn is evidenceof severe damage.The intravenous injection of necrosin may be followed

by the formation of fluid in the chest cavities or by

haemorrhages all along the gastro-intestinal tract ; butmore constantly focal necrosis of the liver develops.Sometimes, after several intravenous injections of

necrosin, one finds a considerable deposition’of fat inthe liver ; at other times the hepatic tissue is strewnwith large amounts of glycogen. The injection ofnecrosin into the blood-stream is accompanied by arelatively small rise in the blood-sugar level (Menkin1946b), averaging about 20% after a single injection.The kidneys may also show foci of leucocytic infiltrationas a result of the introduction of necrosin into the

circulating blood. Such infiltrations are found inter-spersed among tubules that show fuzziness of their

epithelial lining and vacuolation of the cells. The

glomeruli at times have a curious colloid-like materialoccluding the capsular space of Bowman.

Necrosin is absent from normal blood, but may bepresent in that of an animal with inflammation (Menkin1943b). The fact that necrosin penetrates from an areaof injury into the blood-stream and then damages essen-tial organs may prove of clinical significance in focalinfections.

Necrosin is also lethal to mice, in contrast with otherprotein fractions of inflammatory exudate. Finally,in collaboration with Frederick Bernheim, of Duke

University, it has’ been found that necrosin exerts amild proteolytic effect.

PYREXIN

It was observed that the whole euglobulin fraction ofinflammatory exudate would not only reproduce thecellular injury of inflammation but also, when injectedintravenously, would lead to fever and leucopenia.This whole euglobulin fraction was at first’ namednecrosin (Menkin 1943b), and its presence in inflammatoryexudate was confirmed by Smith and Smith (1945) andTanturi et al. (1945). Subsequently, however, it hasbeen found that, whereas necrosin is thermolabile, its

components that induce fever and leucopenia are

thermostable.It was soon recognised that the euglobulin fraction of

the exudate had a component insoluble in the presenceof electrolytes. By differential solubility this componentwas dissociated from a true euglobulin which in turnbehaved like necrosin, damaging the skin of rabbitsand proving lethal to mice. After purification necrosinwas found to be essentially non-pyrogenic. It is truethat at times it is difficult to obtain clear-cut separationbetween the injury factor (necrosin) and the pyrogeniccomponent, which has been termed " pyrexin." Theintravenous injection of pyrexin induces fever in therabbit (Menkin 1943c, 1945). It is therefore reasonableto suppose that its absorption is the cause of the feverthat accompanies inflammation.

Pyrexin is thermostable. Boiling does not essentiallyalter its activity. It seems to be excreted, at least inpart, in the urine. Its exact chemical composition isunknown. The biuret reaction is positive only inminute trace. The xanthoproteic test is positive. The

ninhydrin test is positive, except when pyrexin isrecovered from urine. Pyrexin gives a positive Molischtest. Observations made in collaboration with FrederickBernheim, indicate that pyrexin may be a relativelysimple polypeptide. In view of the constant presence ofa carbohydrate grouping (as indicated by the positiveMolisch test), pyrexin may prove to be a glycopeptide.

LEUCOPENIC FACTOR

As mentioned above, the whole euglobulin fraction ofinflammatory exudate induces leucopenia. Vomiting,diarrhoea, and general apathy are also not infrequent.Studies have been undertaken to determine, if possible,the mechanism of leucopenia, which often accompaniesinflammation. An inflammatory exudate, particularly

662

if acid on withdrawal, tends to induce a transitory leuco-penia when injected in a normal dog. The same resultis obtained by injecting the whole euglobulin fraction ofthe exudate. There is evidence that this effect is notdue to necrosin but to a leucopenic factor closely, butapparently not invariably, associated with pyrexin.This leucopenic factor seems to be a polypeptide (Menkin1946c). The leucopenia appears to be due to a trappingof the leucocytes in lungs, liver, and spleen. Thisaccumulation of leucocytes may help to explainthe acute enlargement of the spleen in inflammatoryconditions (Menkin 1946d).

COMMENTS

The leucocytosis-promoting factor, necrosin, pyrexin,and the leucopenic factor can be recovered from aninflammatory exudate as follows :

These biochemical substances present in inflammatoryexudate help to explain some of the major sequences inthe development of inflammation. There are doubtlessother factors to be discovered. For instance, it has beenshown that any injured cell is potentially gluconeogenetic,a fact which may partly explain why inflammation exertsa bad influence on diabetes (Menkin 1941a, 1943d).It is also conceivable that the end-stage of inflammationor repair may depend on the liberation of one or moregrowth-promoting factors at the site of injury (Menkin1941b).

SUMMARY

The altered chemistry of injured cells liberates intothe inflammatory exudate various biochemical substanceswhich are responsible for the clinical features of inflam.mation.

Five of such substances are described : leukotaxine,a leucocytosis-promoting factor, necrosin, pyrexin, anda leucopenic factor.

Leukotaxine increases local capillary permeability anddiapedesis of leucocytes.The leucocytosis-promoting factor induces a discharge

of immature leucocytes from, and hyperplasia of granu-locytes and megacaryocytes in, bone-marrow.

Necrosin exerts a necrotic effect on local cells and,after absorption into the blood-stream, on distant organs.

Pyrexin induces fever.The leucopenic factor induces leucopenia, the leucocytes

apparently being trapped in lungs, liver, and spleen.REFERENCES

Cullumbine, H., Rydon, H. N. (1946) Brit. J. exp. Path. 27, 33.Lewis, T. (1927) The Blood Vessels of the Human Skin and Their

Responses, London.Menkin, V. (1936) J. exp. Med. 64, 485.

— (1938a) Ibid, 67, 129.- (1938b) Ibid, p. 145.- (1938c) Physiol. Rev. 18, 366.- (1940a) Dynamics of Inflammation, New York and London.- (1940b) Amer. J. Path. 16, 13.- (1949c) Arch. Path. 30, 363.- (1941a) Amer. J. Physiol. 134, 517.- (1941b) Cancer Res. 1, 548.- (1943a) Amer. J. Path. 19, 1021.- (1943b) Arch. Path. 36, 269.- (1943c) Proc. Soc. exp. Biol., N.Y. 54, 184.— (1943d) Amer. J. Physiol. 138, 396.- (1944) Proc. Soc. exp. Biol., N.Y. 56, 219.- (1945) Arch. Path. 39, 28.- (1946a) Ibid, 41, 376.- (1946b) Amer. J. Physiol. 147, 379.- (1946c) Arch. Path. 41, 50.- (1946d) Ibid, 42, 154.- Kadish, M. A. (1943) Amer. J. med. Sci. 205, 363.- Mattison, M. D., Ulled, E. (1946) Proc. Soc. exp. Biol., N.Y.

61, 318.Minami, G., Inugami, K. (1940) Trans. Soc. path. jap. 30, 389.Smith, O. W., Smith. G. V. (1945) Proc. Soc. exp. Biol., N.Y. 59,116.Tanturi, C. A., Canepa, J. F., Banfi, R. F. (1945) Medicina, Buenos

Aires, 6, 143.

PHYSICAL BASIS OF CONTINUOUS

INTRAVENOUS INFUSIONS

J. N. WALTONM.B. Durh.

HOUSE-PHYSICIAN, CHILDREN’S DEPARTMENT, ROYAL VICTORIAINFIRMARY, NEWCASTLE-ON-TYNE

THE administration of intravenous fluid is now acommon procedure, and on occasions its use is a clinicalnecessity for correcting the dehydration and circulatoryfailure which accompany many diseases of infancy andearly childhood. A measure of agreement has beenreached on the nature and amount of fluid required indifferent conditions (Coller and Maddock 1935, A-rnott andYoung 1942), but in view of the technical difficultiesand the hazards of intravenous infusions it is surprisingthat so little attention has been given to the physicalfactors that govern and affect them. Those who watchover the " drip " know how capricious its behaviourmay be, and such apparent caprice may be a seriousmatter for the infant, who may be

" drowned " by amoderate excess of intravenous fluid. Is the behaviourof the " drip " due mainly to varying factors in the

patient, or to the essential nature of the apparatusused 7 This paper describes laboratory experimentsdesigned to furnish an answer to the second questionand suggests some of the practical measures whichfollow.

The apparatacs (fig. 1) used in the experiments consistsof two reservoirs a and b, a standard interceptor c, two.adjustable clips d and e for rubber tubing, a Batemanneedle f, and a measuring cylinder g. Except for themeasuring cylinder, which was necessary for theseexperiments, the apparatus was the standard instru-ments used in the children’s department at this hospital.The graduated 100 c.cm. cylinder b is a particular featureof this apparatus. It allows accurate measurement ofthe quantity of fluid administered, and it is also a safetydevice, for, even if the drip ran more quickly than wasexpected, no more than 100 C.CIll. of fluid would runthrough the apparatus.