PROTEOLYTIC ENZYMES AND ANTI-ENZYMES OF NORMAL AND PATHOLOGICAL CEREBRO-SPINAL FLUIDS

The experiments which have been presented show that the spinal fluid occupies a unique position among the fluids which accumulate in serous cavities of the body. It contains normally neither proteolytic enzyme nor anti-enzyme, whereas blood serum, from which it is derived, exhibits both enzymotic and anti-enzymotic activity. In the blood anti-enzyme greatly predominates over enzyme, so that proteolysis does not occur, unless the anti-enzymotic power of the serum has been destroyed by the addition of acid. In pathological conditions both enzyme and anti-enzyme may make their appearance in the spinal fluid. With inflammations of other serous cavities of the body the anti-enzyme of the exuded serum as a rule preponderates over and restrains the activity of the proteolytic enzyme freed from leucocytes. On the other hand, in infection of the meninges with Diplococcus lanceolatus and with Streptococcus mucosus free proteolytic enzyme has been present in considerable amount in four of five fluids which have been tested. Free proteolytic enzyme has not been observed in the spinal fluid in cases of epidemic meningitis. The cases which have been studied demonstrate that in epidemic meningitis some anti-enzymotic action may be present in the early stages of the disease; but it tends to disappear rapidly so that anti-enzyme seems to be constantly at a low ebb. It is possible that the absence of anti-enzyme in normal spinal fluid, and the tendency for it to disappear so much more rapidly than in other inflammatory exudates, may explain in part the severity of acute meningeal infections. Non-inflammatory transudates into the subdural spaces differ from inflammatory exudates in that the inhibitory element of the blood serum accumulates, and this accumulation suggests an interference with the elimination of the antibody from the spinal fluid. Such interference is not evident in so-called serous meningitis. In content of anti-enzyme the spinal fluid of chronic conditions, such as tuberculous meningitis, apparently occupies an intermediate position between acute inflammation and serous effusion, and five of seven tuberculous fluids which were tested exhibited various degrees of anti-enzymotic action. Variations in content of enzyme and anti-enzyme, noted above, may depend upon the rapidity with which the fluid, carrying the elements mentioned, enters the spinal cavity, as well as upon the rate of their elimination from the spinal fluid. Subdural injection of large quantities of anti-meningitis serum (horse''s blood serum) does not increase the anti-enzymotic activity of fluids withdrawn twenty-four hours after its injection; disappearance of anti-enzyme being caused by rapid elimination of serum from the spinal fluid.     

AN EXPERIMENTAL STUDY OF THE INFLUENCE OF KIDNEY EXTRACTS AND OF THE SERUM OF ANIMALS WITH RENAL LESIONS UPON THE BLOOD PRESSURE

1. Extracts of the rabbit''s kidney injected into the rabbit cause a slight, increase in blood pressure which is barely more than that due to the mechanical effect of the injection. 2. Extracts of the dog''s kidney injected into the dog cause a decided fall in pressure; an equal fall may be caused by the dog''s urine. A series of control experiments indicates that the fall caused by the kidney extract may be due to the urinary salts which it contains. 3. Extracts of cat''s kidney cause a rise in pressure. As the cat''s urine causes a fall, this rise in pressure indicates the possibility of a kidney extract containing a pressor substance which cannot be influenced by the depressor substance of the urine. 4. Rabbit''s kidney, which in the rabbit produces a slight rise, when injected into the dog causes a drop comparable to that caused by the dog''s kidney itself. Similarly, the dog''s kidney, which injected into the dog causes a drop, produces in the rabbit a rise analogous to that produced by rabbit''s kidney. It is evident therefore that these pressor and depressor substances of the kidneys in question do not have a constant effect on all animals as do the extracts of the adrenal gland. 5. Extracts of kidneys which are the seat of various forms of nephritis cause the same effect as extracts of normal kidneys. 6. The serum of dogs with considerable reduction of kidney substance causes a slight fall in pressure; the serum of dogs with spontaneous nephritis gives divergent results, as does also the serum of rabbits with various forms of acute nephritis. The serum of dogs with chromate nephritis causes a slight rise, while that of dogs with uranium nephritis produces a sharp and decided fall in pressure. Although there is no uniformity in these results, their general character, and especially the experience with uranium and chromate sera of the dog, suggests that pressure-disturbing substances are present in the serum as the result of the kidney lesion. The very slight evidence of the constant presence of a pressor substance, however, offers little support to the theory that such a substance is furnished by the diseased kidney or is due to disturbances of metabolism caused by disease of the kidney.     

EXPERIMENTAL PLEURISY—RESOLUTION OF A FIBRINOUS EXUDATE

Fibrinous pleurisy produced by a sterile inflammatory irritant offers opportunity for study of the part taken by enzymes of leucocytes in the resolution of a fibrinous exudate. When turpentine is injected into the subcutaneous tissue of the dog, an abscess results, but when an equal quantity of turpentine is injected into the pleural cavity, there is abundant exudation of coagulable fluid and the serous surfaces are covered by a layer of fibrin. Accumulation of fluid which can be followed during life by percussion of the animal''s chest reaches a maximum at the end of three days, and then gradually subsides, so that at the end of six days, in most instances, the cavity contains no fluid. Fibrin, though diminished in amount at the time when fluid has been absorbed, is still present, and gradually disappears; at the end of two or three weeks the cavity has returned to the normal, save for a few organized adhesions. Turpentine injected into the right pleural cavity may cause serofibrinous pleurisy on the left side; this inflammation may reach a maximum intensity at a time when pleurisy on the right side is subsiding. During the early stage of inflammation fibrinous exudate, freed from the serum by washing in salt solution, undergoes digestion when suspended in an alkaline (0.2 per cent. sodium carbonate) or in an acid medium (0.2 per cent. acetic acid). At the end of five days, at a time when fluid is disappearing from the pleural cavity, digestion fails to occur in an alkaline medium, but occurs with much activity in the presence of acid. During the first stage of the inflammatory reaction, when fluid is abundant and the fibrin which is present digests in alkali, thus indicating the presence of leucoprotease, polynuclear leucocytes are very numerous in the meshes of the fibrin. In the second stage, the exuded fibrin contains only one enzyme digesting in the presence of acid. At this time polynuclear leucocytes have disappeared and only mononuclear cells are embedded in the fibrin. Products of proteolytic digestion, namely, peptone and albumose, absent in the exuded fluid during the first day or two days of inflammation, are present after three days and are found in less quantity at a later period. The exuded fluid does not at any stage of the inflammatory reaction lose it spower to inhibit both enzymes contained in the leucocytes. The exudate remains alkaline throughout the period of inflammation, but its alkalinity is less than that of the blood and diminishes slightly with the progress of inflammation. Since the acids, which in vitro favor the action of the enzyme, present alone during the second stage of the inflammatory reaction, do not occur in the body, the possibility has suggested itself that carbon-dioxide brings this enzyme into action. If carbon-dioxide is passed through normal salt solution in which strips of such fibrin are suspended, digestion is greatly hastened. The normal inhibition exerted by blood serum upon the enzyme is overcome by carbon-dioxide and in the presence of a small quantity of blood serum, carbon-dioxide causes greater enzymotic activity than in the presence of salt solution alone.     

ANGIOTONIN-ACTIVATOR,RENIN- AND ANGIOTONIN-INHIBITOR,AND THE MECHANISM OF ANGIOTONIN TACHYPHYLAXIS IN NORMAL,HYPERTENSIVE, AND NEPHRECTOMIZED ANIMALS

1. Angiotonin does not exert its vasoconstrictor effect in the absence of a substance contained in red blood cells and serum which we have called "angiotonin-activator." A fraction has been separated from blood in which angiotonin-activator is concentrated. It contains little or no reninactivator. 2. Repeated intravenous injections of angiotonin into animals causes the pressor response gradually to lessen and finally to disappear (the phenomenon of tachyphylaxis), but much more slowly than when renin is injected. When the response to angiotonin is abolished, renin also fails to act. Large doses of renin reduce and finally abolish the responsiveness to angiotonin. Exhaustion of renin-activator in the blood abolishes the response to renin without abolishing the response to angiotonin. 3. Blood from animals made tachyphylactic by infusion of angiotonin contains greatly reduced amounts of angiotonin-activator. An inhibitor also appears in the blood. 4. Bilateral nephrectomy prolongs and greatly enhances the rise of arterial pressure following injection of angiotonin and renin. The enhancement reaches a maximum in from 24 to 30 hours after operation. Blood from these animals exhibits greatly increased ability to activate angiotonin and renin when tested in isolated perfused organs. Large amounts of angiotonin are required to reduce the amount of activator in their blood. Renin-activator is simultaneously but little affected. 5. Tranfusion of blood from an animal made tachyphylactic to angiotonin into a nephrectomized dog reduces the response of the latter to angiotonin. Angiotonin when added to the blood of the recipient of the transfusion and perfused through a rabbit''s ear also exhibits greatly reduced vasoconstrictor action. 6. Transfusion of normal blood in large amounts into nephrectomized or hypertensive dogs reduces the recipient''s response to renin. If renintachyphylaxis is established in the donor, transfusion abolishes the response to renin in the recipient. The blood from such animals exhibits greatly reduced vasoconstrictor properties when perfused through an isolated organ with renin or angiotonin. 7. Renin-tachyphylactic or normals dog''s blood does not reduce arterial pressure elevated by a single injection of renin into nephrectomized dogs. 8. Nephrectomized dogs exhibit the greatest pressor response to infusion of angiotonin and renin, normal animals the least, and hypertensive animals about midway between.     

THE LIBERATION OF RENIN BY PERFUSION OF KIDNEYS FOLLOWING REDUCTION OF PULSE PRESSURE

1. Isolated dogs'' kidneys have been perfused with defibrinated blood under hemodynamic conditions similar to those in the body. Under these circumstances blood flow, urine secretion, and oxygen consumption are well maintained, but urea clearance is low. Renal venous blood collected initially and at the end of 3 or more hours of perfusion exhibited no difference in vasoconstriction properties when perfused along with renin or renin-activator through an isolated rabbit''s ear. 2. Reduction of pulse pressure by constricting the renal artery may be performed without reducing mean pressure significantly. Impairment of urea clearance and rate of urine secretion follow, and oxygen consumption is slightly reduced. 3. After an hour or more of perfusion with reduced pulse pressure, gradual rise in mean renal arterial pressure distal to the clamp and reduction of blood flow occur. 4. Renal venous blood collected after about one hour of perfusion with reduced pulse pressure differs from that collected before reduction of pulse pressure in that it causes intense vasoconstriction when perfused with renin-activator through an isolated rabbit''s ear. 5. Perfusion of a dog''s hind leg under similar circumstances does not cause this change in the venous blood to occur.     

ENZYMES OF TUBERCULOUS TISSUE

Epithelioid cells which form the chief element of tuberculous tissue contain an enzyme which causes active digestion of proteid in an approximately neutral or in a weakly acid medium, but is inactive in the presence of weak alkali. This enzyme resembles that which occurs in the large mononuclear cells of an inflammatory exudate and is more active than the similar enzyme of parenchymatons organs such as the liver. The enzyme which digests in the presence of acid exhibits greatest activity at a time when caseation is beginning. With advance of caseation its activity diminishes so that tissue which has undergone almost complete caseation exhibits trivial evidence of the presence of enzyme. It is probable that complete caseation is followed by total disappearance of enzymes. Tuberculous tissue contains an enzyme capable of digesting proteid in the presence of alkali (leucoprotease) only during the early stages of its development. This enzyme, present at a time when the tissue contains numerous polynuclear leucocytes, quickly disappears so that when enzyme digesting in acid is still active, leucoprotease has disappeared. The serum of a tuberculous pleural exudate obtained by intrapleural inoculation with tubercle bacilli causes slight inhibition of the mixture of enzymes contained in tuberculous tissue shortly after inoculation. The serum of blood causes complete inhibition of the enzymes contained in the same tuberculous tissue. Analysis of this difference indicates that the exuded tuberculous serum, like the serum of the blood, inhibits proteolysis caused by leucoprotease, but fails to inhibit digestion caused by an enzyme acting in the presence of acid. In testing this property of the exuded tuberculous serum lymphatic gland has been used because suitable tuberculous tissue has not been available. The serum of the tuberculous pleural exudate produced experimentally not only fails to exert the anti-enzymotic power which is exhibited by the serum of the blood, but is itself capable of causing active digestion of coagulated proteid. Normal blood serum does not digest proteid and the serum of a sterile inflammatory exudate obtained by injection of turpentine into the pleural cavity has caused very little digestion. The tests which have been made indicate that loss of anti-enzymotic power and ability to cause proteolysis increase with the age of the exudate. The foregoing facts offer suggestions which may serve to explain in part the nature of the tubercle and the changes which occur within it. The so-called epithelioid cells of the tubercle resemble the large mononuclear phagocytes of inflammatory exudates and both contain an enzyme of the same character. It is not improbable that caseation which, like autolysis, is accompanied by disappearance of nuclei is in part dependent upon the presence in the cells of this active proteolytic enzyme which is for a time held in check. Injury to cells by products of the tubercle bacillus or partial anæmia, the result of imperfect vascularization of the tuberculous tissue, may have a part in rendering these cells susceptible to self-digestion. Changes which have been observed in serum of the tuberculous exudate show that the anti-enzymotic property of the normal blood may be absent in the exudate of a tuberculous lesion. This loss of anti-enzymotic action, perhaps referable to changes caused by products of the tubercle bacillus, may favor self-digestion of the enzyme-containing cells and diffusion of their enzyme.     

STUDIES ON THE INACTIVATION OF VACCINE VIRUS AND THE ACTION OF CERTAIN SUBSTANCES UPON THE INFECTING POWER OF THE INACTIVATED VIRUS

Vaccine virus, obtained from testicular inoculation shows a high susceptibility to chloroform as compared with ether, toluene, 95 per cent alcohol and acetone. Vaccine virus, after treatment with an amount of chloroform sufficient to render it incapable or only barely capable of originating an eruption in the rabbit''s skin, produces a characteristic eruption when injected with the supernatant fluid of embryonic tissue or sarcoma tissue "cultures" or kieselguhr, substances all of which are markedly irritative to the rabbit''s skin. Reactivation of the chloroformed vaccine virus is not possible when chloroform has been added to it in such quantity that the injection of large amounts of the treated virus fails to cause an eruption. Whenever reactivation has been accomplished it has been possible to get a vaccine eruption of greater or less intensity by the injection of large amounts of the chloroformed vaccine alone. Embryo and chicken sarcoma "culture" fluids when injected intradermally make the skin susceptible to the localization of the virus introduced intravenously. The bearing of these experiment on the interpretation of Gye'' theory of cancer causation is discussed.     

THE EFFECT OF INCREASED ANTIPNEUMOCOCCAL IMMUNITY ON THE INCEPTION OF EXPERIMENTAL LOBAR PNEUMONIA IN THE DOG

In view of the finding that one or more attacks of pneumococcus lobar pneumonia experimentally induced in dogs failed to protect the animals against subsequent infection, an attempt was made to determine whether or not the dog''s antipneumococcal immunity could be enhanced to the degree of complete resistance to the experimental disease. To this end dogs were passively immunized by the intravenous injection of large quantities of both unconcentrated antipneumococcus horse serum and concentrated antibody solution and actively immunized by vaccination with killed and living cultures of pneumococci. None of these procedures were found to result in constant protection against the pulmonary infection. The disease, however, was of brief duration, the lesions of limited extent and usually sterile within 24 hours. A combination of active and passive immunization produced no better results. It was only when immune bodies and leucocytes were implanted with the infecting dose that prevention of infection was secured with any degree of constancy. Even under these conditions the lesion sometimes involved a considerable portion of a lobe. The factors involved in the inception of experimental lobar pneumonia are discussed and the bearing of this study on the prophylactic immunization of human beings against pneumococcus pneumonia is suggested.     

ON THE NATURE OF THE PRESSOR ACTION OF RENIN

1. Tachyphylaxis occurs when renin is repeatedly injected into dogs and cats regardless of whether they are normal, anesthetized, pithed, hepatectomized, suprarenalectomized, nephrectomized, or eviscerated. 2. The pressor response to renin in brief experiments is independent of the height of the arterial pressure or the presence of the suprarenals. Evisceration and large doses of ergotamine reduce the response. It is largely uninfluenced by pithing, intracisternal injection of renin, cocaine, strychnine, caffeine, and infusion of sodium bicarbonate or hydrochloric acid. It may be slightly increased by large blood transfusions or hepatectomy but the result is short lived. 3. There is no parallelism between the pressor responses to carotid sinus stimulation, adrenine, and tyramine on the one hand and renin on the other. 4. Section of the brain may be followed by depressor responses to renin. 5. Intracisternal injection of renin elicits no significant rise in blood pressure or other circulatory manifestations. 6. Continuous infusion of renin produces a prolonged rise of arterial pressure in normal and chronically suprarenalectomized dogs, but the pressure ultimately falls despite continued infusion. 7. Tachyphylaxis develops in the isolated rabbit''s ear perfused with blood and small doses of renin. The same blood perfused through a second ear causes no vasoconstriction when renin is added. Addition of renin-activator restores the ability of renin to cause constriction. 8. Renin alone causes no vasoconstriction when perfused with Ringer''s solution, but renin plus renin-activator restores activity. Tachyphylaxis does not develop when Ringer''s solution is employed instead of recirculating blood. 9. Blood from animals made tachyphylactic by repeated injections of renin is lacking in activator and also fails to cause vasoconstriction in the rabbit''s ear when renin and renin-activator are added. 10. Renin-activator is lost and tachyphylaxis develops more slowly during continuous infusion of renin. Blood pressure may fall after a period of renin infusion despite the pressure in the blood of excess renin. Injection of partially purified activator restores the activator content of the blood as demonstrated in the rabbit''s ear, but no rise in arterial pressure occurs.     

STUDIES ON THE MECHANISM OF RECOVERY IN PNEUMONIA DUE TO FRIEDL?DER'S BACILLUS : III. THE R?LE OF "SURFACE PHAGOCYTOSIS" IN THE DESTRUCTION OF THE MICROORGANISMS IN THE LUNG

Phagocytosis of encapsulated Friedländer''s bacilli has been demonstrated in the lungs of rats in the absence of both circulating and local antibody. The mechanism of phagocytosis independent of antibody has been shown to be due to the same surface factors that operate in the phagocytosis of Type I pneumococcus under similar conditions. Direct observation of the phagocytic process reveals that leucocytes in the lung can phagocyte unopsonized Friedländer''s bacilli only by trapping them against the surfaces of alveolar walls or bronchi, or by pinning them against the surfaces of adjacent leucocytes. Evidence is presented that Friedländer''s bacilli thus phagocyted are rapidly killed in the cytoplasm of the phagocytic cells. Reasons are discussed for the failure of prolonged chemotherapy to cure lung abscesses that not infrequently complicate the pneumonia due to Friedländer''s bacillus.     

THE LS-ANTIGEN OF VACCINIA : IV. CHEMICAL ANALYSIS OF LS AND THE EFFECT OF CHYMOTRYPSIN ON LS

Pure LS-antigen of vaccinia contains 15.8 per cent N and 50.6 per cent C. These analytical data together with the absence of lipids, phosphorus, nucleic acid, and glucosamine in preparations of the antigen confirm the protein nature of the substance. The action of proteolytic enzymes on LS offers further confirmation of the protein character of the antigen. Both the L- and S-activities of the antigen are destroyed by digestion with papain. The effects of crystalline chymotrypsin on LS-antigen are particularly interesting for, under proper conditions, this enzyme destroys the serological activity of the S-portion of the molecule without affecting the L-portion. This newly prepared degradation product of LS, called LS'''', contains the same amount of N as the native substance but unlike LS, it forms needle-shaped crystalloids.     

A STUDY OF THE ACTION OF ACONITIN ON THE MAMMALIAN HEART AND CIRCULATION

The action of aconitin on the dog''s heart, therefore, seems to consist in: 1. A stimulation of the inhibitory mechanism, especially of the centres in the medulla oblongata. 2. An increase in the irritability of the muscle of the auricle and the ventricle, which leads to independent contractions of one or both of these divisions and culminates in fibrillary contractions in the ventricle. The first of these is the only effect seen in the therapeutic use of the drug, and aconitin may, therefore, be considered to be indicated when it is desirable to stimulate the inhibitory centre without acting on the heart muscle. Of course it has a further effect on the circulation through the stimulation of the vaso-motor centre, but this would appear to be of minor importance.     

MULTIPLE VIRUS INFECTION OF SINGLE HOST CELLS

Evidence is presented to show that two or more viruses can simultaneously manifest their characteristic activities within individual epithelial cells of the normal rabbit''s cornea. This evidence, together with that previously presented (1, 5, 6), makes plain that multiple virus infection of a single host cell can take place in corneal cells, in the cells of chick embryos, and in those of rabbit tumors, both benign (Shope''s papilloma) and malignant. Certain implications of the findings are discussed.     

ON EXTRACELLULAR AND INTRACELLULAR VENOM ACTIVATORS OF THE BLOOD,WITH ESPECIAL REFERENCE TO LECITHIN AND FATTY ACIDS AND THEIR COMPOUNDS

In normal serums of the majority of mammalian and avian blood there exists certain substances capable of activating venom hæmolysin. They are extractable from serum by means of ether, and are capable of conferring upon the originally non-activating serum a power to activate venom, when mixed with the latter. The ethereal extract consists of fatty acids, neutral fats and possibly also some ether soluble organic soaps. The fatty acids and soaps, especially of the oleinic series, acquire certain characteristics of complements in general, when they are mixed with serum. They are inactive without the venom in the mixture; they are inactivable with calcium chloride; they exhibit a tendency to go off in activity with age; they are inactive or only weakly active at 0° C., and they are extractable by ether. In testing the serum from which the ether soluble substances are removed, it is found that no venom activating property is left. Warm alcoholic extraction of such serum yields, however, a large quantity of lecithin. In the case of non-activating serums no venom activating fats appear in the ethereal extract. Lecithin exists in such serum in no less quantity than in the activating kind. The addition of oleinic acid or its soluble soaps to a non-activating serum, in a ratio which corresponds to the percentage of fatty acids or soaps contained in some of the easily activating serums, will make the serum highly active in regard to venom. In normal serum of dog there exists, besides the group of activators already mentioned, another kind of venom activators which has been identified as a lecithin compound acting in the manner of free lecithin. A very sharp differentiation of the hæmolysis produced by this activator and by the other groups of activators is obtained by means of calcium chloride, which is powerless against lecithin or lecithin compounds, but effective in removing the action of the latter. This lecithin containing proteid can be precipitated by half saturation with ammonium sulphate, but is perfectly soluble in water, and is not coagulated in neutral alkaline salt solutions upon boiling. Alcohol precipitates a proteid-like coagulum and extracts lecithin from it; ether does not extract lecithin from this compound. Non-activating serums do not contain any such lecithin compound. Lecithin contained in other serum proteids, mainly as lecithalbumin, and perhaps as contained in globulin, is not able to activate venom. This is true of all the serums with which I worked; it matters not whether these fractions (obtained with ammonium sulphate) belong to the most activating serum (dog) or to the non-activating serum (ox). The non-coagulable portion of all heated serum contains a venom activator of the nature of lecithin. This activator is contained in a non-coagulable proteid described by Howell which is identical with Chabrie''s albumon. As there is no ether-extractable lecithin in this portion of the serum, the activating property of heated serum must be due to this proteid compound of lecithin. That this lecithin proteid does not pre-exist in normal serum but is produced by the action of high temperature is true of all serums except that of the dog. In venom activation we know now that lecithin becomes reactive with venom when it is transformed from other proteid compounds into the non-coagulable form, the albumon. Howell''s view of the non-existence of the non-coagulable proteid in normal serum seems to receive a biological support from venom hæmolysis. Ovovitellin derived from hen''s egg is one of the best venom activators of the lecithin proteid type. The cause of venom susceptibility of various kinds of blood corpuscles does not depend upon the existence of lecithin in the corpuscles, but solely upon the amount of fatty acids, and perhaps, also, soaps and fats, contained in the corpuscles. The protection which calcium chloride gives against venom haemolysis is proof of the absence of lecithin activation. From the stroma of susceptible corpuscles fatty acids or some fats can be extracted with ether. After ethereal extraction the stroma becomes non-activating, while the extract contains fatty acids and some soaps or fats, which when added to venom-resistant corpuscles render the latter vulnerable to venom. The corpuscular solution of non-activating corpuscles does not contain enough fatty acids. The larger the amount of fatty acids and soaps in the corpuscles, the easier the cells undergo venom haemolysis. Lecithin exists in the strorna of all kinds of corpuscles, but in a form unavailable for venom activation. The somatic cytolytic processes caused by venom requires intracellular complements. The experiments performed on the cells of liver, kidney, testis and brain of the guinea-pig and rat indicate that the substances which act as complements are inactivable by calcium chloride.     

THE EFFECT OF CERTAIN EXPERIMENTAL PROCEDURES ON THE ISLANDS OF LANGERHANS

Although the results of this study have been of a negative character, the conclusions that may be drawn from it have an important bearing on the true anatomical character of the islands. 1. Neither inanition nor the prolonged injection of secretin has any noteworthy effect upon the number, size, or structure of the islands of Langerhans in the dog''s pancreas. 2. The islands of Langerhans in the guinea pig''s pancreas are in no way altered in phlorizin diabetes. 3. The islands of Langerhans are not formed out of exhausted or degenerated acini, but develop from the ducts or acini with which they are often in direct continuity.     

ALLOTYPY OF RABBIT SERUM PROTEINS : II. RELATIONSHIPS BETWEEN VARIOUS ALLOTYPES: THEIR COMMON ANTIGENIC SPECIFICITY,THEIR DISTRIBUTION IN A SAMPLE POPULATION; GENETIC IMPLICATIONS

The relationships between six of the seven allotypes or families of allotypes (a, b, c, d, f, g) described in the preceding paper, have been studied from the standpoints of (1) their antigenic specificities, (2) their mutual influence on the limitation of their respective frequencies, and (3) their genetic control. Although the six different allotypes (or families) react quite differently with the rabbit antisera, at least five of them react identically with a guinea pig antiserum. Therefore, a large portion of the antigenic specificity of these allotypes, distinct from their allotypic specificity, is uniform in all the individuals of the rabbit species and is termed for this reason "isotypic specificity." In the early period of the rabbit''s life, allotypes may be found in the serum, which are not determined by the genotype of the individual, but are directly transmitted by the mother. The allotypes of the antigenic species of globulin studied in this paper, which were synthesized by the young animal, did not appear in its serum before a certain period of time. Allelic relationships between the genes which control allotypes were indicated by, (1) the absence of certain kinds of groupings of the allotypes, which limits the number of allotypic formulas in the population sample studied, (2) dosage effects, the concentration of certain allotypes (drawn from the penetration of the zones in gel tubes) being smaller in supposed heterozygotes than in supposed homozygotes, (3) the results of the analysis of the sera of a number of rabbits and of their parents. Eight of the different antigenic substances studied in this paper (allotype e excluded) appear to be allotypic forms of what would have been considered to be a uniform protein antigen. They may be classified as follows: a first group which contains two allotypes b and d and a family of two allotypes c and c'' apparently controlled by three allelic genes b c d, c and c'' being controlled by the same gene; a second group which contains two allotypes a and f and a family g, g'' apparently controlled by three allelic genes a f g. There are reasons to believe that this list is not complete, especially in the b c d group.     

FREE ANTIGEN AND ANTIBODY CIRCULATING TOGETHER IN LARGE AMOUNTS (HEMAGGLUTININ AND AGGLUTINOGEN IN THE BLOOD OF TRANSFUSED RABBITS)

In rabbits transfused almost daily with the whole citrated blood of other rabbits, an extraordinary condition often develops, which manifests itself in an almost immediate clumping together of all the red cells in specimens of the shed blood. This clumping is due to one or more true agglutinins, of which the strength may be such as to cause clumping in a 1: 2,800 plasma dilution. The agglutinating principle circulates with the corpuscles against which it is effective; but under ordinary circumstances intravascular clumping fails to occur because the union of antigen and antibody can take place only at a temperature several degrees below that of the body. If the temperature is sufficiently lowered, as when a tourniquet is applied to the rabbit''s ear, intravascular clumping ensues. In defibrinated blood, gradually cooled, clumping is first noted as the temperature of 35°C. is approached; and at room temperature (22°) the corpuscles will often come together in a short time into a single, solid mass. At 0°C. the agglutination is still more marked. The reaction seems to be completely reversible, for when the blood is warmed again, the clumps break up and disappear at between 35° and 36°C. Cooling and warming with the resultant clumping and dissociation can be carried out many times on the same blood specimen without apparent change in the corpuscles or in the rapidity of the reaction. The response to temperature changes is extremely prompt. Once it has been elicited, the agglutinating principle may persist for a long time after the transfusions are stopped, in one instance it was still strong 133 days after the last transfusion. During this period the plethora was succeeded by a severe anemia, which in turn was recovered from. In many rabbits no agglutinin develops, and a continuance of the transfusions will not elicit it. Indeed, when present it tends to disappear if the transfusions are persisted in. In several of the animals in which the agglutinin was strongest, the plethora was suddenly succeeded by severe anemia, despite continued transfusions. The character of the temperature control of the agglutination, which somewhat resembles that of the hemolysin in paroxysmal hemoglobinuria, has led us to consider whether the blood destruction might not be due to accidental chilling of the animal. Efforts to induce a fall in the hemoglobin by placing the rabbit''s ear in ice water have as yet been unsuccessful. Thus far no adequate search for an hemolysin has been made. The object of the present paper has been to describe a condition in which large amounts of free antigen and antibody circulate together in the organism, and to demonstrate the factor which prevents their union, the results of which could easily be fatal. The causes of the condition will be dealt with in a subsequent communication.     

FUNCTIONAL AND METABOLIC PROPERTIES OF POLYMORPHONUCLEAR LEUCOCYTES : II. THE INFLUENCE OF A LIPOPOLYSACCHARIDE ENDOTOXIN

The effects of a purified bacterial lipopolysaccharide endotoxin on homogenous populations of rabbit polymorphonuclear leucocytes have been studied in vitro under defined conditions. Employing a 500-fold range of concentration (0.1 to 50.0 µg./ml.), it was shown that endotoxin enhanced the rate at which staphylococci were killed by leucocytes. The mechanism underlying the increased killing was found to be a direct stimulation of the phagocytic activity of the leucocyte and not mediated by the release of bactericidins or opsonins from the treated cells. In the presence of 10 per cent serum all concentrations of endotoxin enhanced phagocytosis, whereas at lower serum concentrations, the higher doses of lipopolysaccharide inhibited the phagocytic activity of the cells. Similar concentrations of endotoxin were capable of increasing the utilization of glucose and the production of lactic acid. Endotoxin treated leucocytes exhibited no change in oxygen consumption, and only a slight depression in glycogen synthesis. It appeared that endotoxin could interact and alter the functional and metabolic properties of leucocytes in the absence of serum. The demonstration of enhanced phagocytic activity of endotoxin-treated cells was dependent upon the particular opsonic requirements for the organism under study.     

Intensive Care of a Collapsed,Anorexic and Pyrexic Dog

A dog arrives at the practice in a collapsed state having been referred, with a history of anorexia and pyrexia. Stabilising the dog's condition is the first priority and is an area where the nurses take an active role. This case report, which formed part of the author's diploma casebook, charts the dog's progress     

ON THE RELATION OF THE CATALYTIC ACTIVITY OF THE BLOOD TO THE NUMBER OF RED BLOOD CELLS IN HEALTH,AND TO THE NUMBER OF WHITE BLOOD CELLS AND THE BODY TEMPERATURE IN PERITONITIS

The catalytic activity of the blood of normal rabbits varies almost directly with the volume and number of red blood cells. This explains to a certain extent at least why animals of the same general degree of nutrition, and of the same litter, should have about the same activity since they are likely to have the same number of red blood cells, and why healthy large animals should read high while small poorly nourished ones should read low. Accompanying the hyperpyrexia resulting from puncture of the corpus striatum of a rabbit''s brain, there is no change in either the catalytic activity of the blood or the white blood count. In experimentally produced peritonitis, the catalytic activity of the blood always rises, and is, therefore, absolutely independent of body temperature and white blood cells since one or both of these may rise, fall or remain stationary while the catalytic action increases.