PROTEOSE INTOXICATIONS AND INJURY OF BODY PROTEIN : I. THE METABOLISM OF FASTING DOGS FOLLOWING PROTEOSE INJECTIONS.

Proteose injections in dogs cause vomiting, diarrhea, temperature reactions, low blood pressure, prostration, and, after large doses, an excess of antithrombin with incoagulable blood. A single proteose injection, for example one-half a lethal dose, causes abrupt clinical reactions in a normal dog with apparent complete recovery within 24 to 48 hours. The nitrogen elimination curve in a fasting dog under such conditions shows a great rise in total urinary nitrogen. The apex of the curve usually falls during the second 24 hour period following the injection. This rise may be over 100 per cent increase above the mean base-line nitrogen level. It does not fall promptly to normal but declines slowly in 3 to 5 days or more toward the original base-line (Text-fig. 1). This speaks for a definite cell injury with destruction of considerable protein substance due to a single proteose injection. The disturbance of cell equilibrium is not rapidly or promptly restored to normal. A dog which has received previous proteose injections is partially immune or tolerant to subsequent injections of proteose. These dogs, as a rule, show less intense clinical reactions and less rise in the curve of nitrogen elimination following a unit dose of standard proteose as compared with normal or non-immune controls. The proteose used in these experiments was prepared as described from material obtained in cases of intestinal obstruction or of closed intestinal loops. These experiments explain the sharp rise in blood non-protein nitrogen which follows within a few hours the injection of a toxic proteose. They also point to the correct explanation of the high non-protein nitrogen of the blood found in intestinal obstruction or with closed intestinal loops.     

INTESTINAL OBSTRUCTION : V. PROTEOSE INTOXICATION.

A definite intoxication develops as a result of a closed intestinal loop and toxic material accumulates in the closed loops. Much evidence has been submitted to show that this loop poison causes the intoxication observed after producing a closed intestinal loop. Sufficient evidence has been presented to prove that the essential poison is present in these closed intestinal loops, and usually in concentrated form. Chemical study of the contents of closed intestinal loops shows that a single substance or group of substances possesses toxic properties. This resists autolysis and pancreatic and ereptic digestion. It is thrown out of solution by five volumes of alcohol or by half saturation with ammonium sulphate. It is readily soluble in water and is not injured by boiling. It is not removed by dialysis. The method of isolation excludes practically all substances except primary proteoses. The characteristic resistance to digestive enzymes suggests a heteroproteose. Proteose intoxication in dogs gives a picture identical with that described after poisoning with intestinal loop fluid: early salivation and vomiting, followed by diarrhea and prostration, fall in temperature and blood pressure, and finally death in collapse. Autopsy shows essentially a splanchnic paralysis and remarkable engorgement of liver and spleen, but especially of the mucosa of the duodenum and small intestine. The blood shows great concentration due to loss of fluid and may remain incoagulable because of an excess production of antithrombin. Proteoses escaping from the blood are excreted in the urine. This toxic proteose concerned in intestinal obstruction has not yet been isolated in the urine, but may be excreted by the kidneys. This probably explains the clinical improvement and lessened intoxication noted after transfusion. Experimental evidence points to a primary proteose as the essential poison concerned in the intoxication of closed intestinal loops and intestinal obstruction.     

INTESTINAL OBSTRUCTION : III. THE DEFENSIVE MECHANISM OF THE IMMUNIZED ANIMAL AGAINST DUODENAL LOOP POISON.

Dogs may be immunized against lethal doses of the duodenal loop poison by means of small doses of the loop fluid from dog or cat and by material obtained from human cases of intestinal obstruction. The immunity is transient and may disappear within a few weeks. Dogs immunized by repeated doses of loop fluid show a definite resistance against the intoxication of a closed duodenal loop and may survive twice the usual period. A dog that recovers from simple intestinal obstruction may possess a strong resistance to the intoxication of a closed duodenal loop, thus indicating a similar type of intoxication in the two conditions. The sera of immune dogs are inactive when incubated with duodenal loop fluid. The organ extracts and emulsions (liver, spleen, lung) of immune dogs rapidly destroy the loop poison during incubation in vitro. This destructive property is possessed by a clear filtrate of the digested immune organs, excluding adsorption, and is lost after long periods of incubation (twelve weeks). We are investigating the action of this immune organ extract to determine whether it can destroy the closed-loop poison in vivo and perhaps be of value in treatment.     

INTESTINAL OBSTRUCTION : VI. A STUDY OF NON-COAGULABLE NITROGEN OF THE BLOOD.

Intestinal obstruction, as a rule, is associated with an increasing amount of non-coagulable nitrogen in the blood. With acute intoxication the rise in non-coagulable nitrogen may be rapid and reach as high as three or even ten times normal. With more chronic intoxication there may be little or no rise in the blood non-coagulable nitrogen. Closed intestinal loops show exactly the same picture, and, when combined with obstruction, may give very high nitrogen readings. Acute proteose intoxication due to injection of a pure proteose will show a prompt rise in blood non-coagulable nitrogen, even an increase of 100 per cent within 3 or 4 hours. These intoxications also show a high blood content of creatinine and urea. The residual or undetermined nitrogen may be very high. A human case of intestinal obstruction with autopsy presents blood findings exactly similar to those observed in many animal experiments. Clinically the non-coagulable nitrogen of the blood may give information of value in intestinal obstruction. A high reading means a grave intoxication, but a low reading may be observed in some fatal cases and gives no assurance that a fatal intoxication may not supervene. The kidneys in practically all these experiments are normal in all respects. It is possible that protein or tissue destruction rather than impaired eliminative function is responsible for the rise in non-coagulable nitrogen of the blood in these acute intoxications. Transfusions of dextrose solutions often benefit intestinal obstruction, and may depress the level of the non-coagulable nitrogen in the blood. Some cases show no change in non-coagulable nitrogen following transfusions and diuresis, and, as a rule, such cases present the most severe intoxication.     

STUDIES ON THE BLOOD PROTEINS : II. THE ALBUMIN-GLOBULIN RATIO IN EXPERIMENTAL INTOXICATIONS AND INFECTIONS.

The intoxication which develops as the result of a simple obstruction or a closed intestinal loop is accompanied by definite changes in the coagulable proteins of the blood serum. These changes consist essentially in an alteration in the normal albumin-globulin ratio; the globulin fraction is greatly increased and at times the normal relation of the two fractions may show a complete inversion. The increase in the globulin content of the blood serum is most marked in the animals which show some of the complications met with in loop animals,—rupture of the loop and peritonitis. In the latter conditions especially, the globulin increase is rapid and large. We believe this reaction to be of diagnostic value in acute infections attended by the sudden liberation and absorption of a toxic exudate. Infections and intoxications produced by inflammatory irritants are also accompanied by a rise in the blood globulins. This observation suggests that tissue disintegration with absorption of toxic products is responsible for the changes noted, and that bacterial invasion is important only in as far as it gives rise to toxic substances. Animals which have developed a tolerance to proteose intoxication following the periodic injection of small doses of proteose do not show a globulin increase. These experiments do not support the view that the rise in globulins observed in these experimental conditions is an expression of a resistance or tolerance developed by the animal. From the experimental evidence it seems more probable that the alteration in the partition of the blood protein fractions is one of the results of the metabolic disturbance which has been shown to occur in these conditions.     

THE BLOOD CHLORIDES IN PROTEOSE INTOXICATION

Dogs injected with proteose recovered from the intestinal contents of animals with obstruction at different levels show no significant changes in the blood chlorides even with a fatal intoxication. After the intravenous injection of lethal and sublethal doses of Witte''s peptone there is little change in the chlorides. Autolyzing liver in the abdominal cavity produces no change in the blood chlorides even with a very great increase in the urea and nonprotein nitrogen. Proteose intoxication is probably not a factor in the characteristic fall in chlorides seen after intestinal and pyloric obstruction.     

INTESTINAL OBSTRUCTION : I. A STUDY OF A TOXIC SUBSTANCE PRODUCED IN CLOSED DUODENAL LOOPS.

Closed duodenal loops may be made in dogs by ligatures placed just below the pancreatic duct and just beyond the duodenojejunal junction, together with a posterior gastro-enterostomy. These closed duodenal loop dogs die with symptoms like those of patients suffering from volvulus or high intestinal obstruction. This duodenal loop may simulate closely a volvulus in which there has been no vascular disturbance. Dogs with closed duodenal loops which have been washed out carefully survive a little longer on the average than animals with unwashed loops. The duration of life in the first instance is one to three days, with an average of about forty-eight hours. The dogs usually lose considerable fluid by vomiting and diarrhea. A weak pulse, low blood pressure and temperature are usually conspicuous in the last stages. Autopsy shows more or less splanchnic congestion which may be most marked in the mucosa of the upper small intestine. The peritoneum is usually clear and the closed loop may be distended with thin fluid, or collapsed, and contain only a small amount of pasty brown material. The mucosa of the loop may show ulceration and even perforation, but in the majority of cases it is intact and exhibits only a moderate congestion. Simple intestinal obstruction added to a closed duodenal loop does not modify the result in any manner, but it may hasten the fatal outcome. The liver plays no essential role as a protective agent against this poison, for a dog with an Eck fistula may live three days with a closed loop. A normal dog reacts to intraportal injection and to intravenous injection of the toxic substance in an identical manner. Drainage of this loop under certain conditions may not interfere with the general health over a period of weeks or months. Excision of the part of the duodenum included in this loop causes no disturbance. The material from the closed duodenal loops contains no bile, pancreatic juice, gastric juice, or split products from the food. It can be formed in no other way than by the activity of the intestinal mucosa and the growth of the intestinal bacteria. This material after dilution, autolysis, sterilization, and filtration produces a characteristic effect when introduced intravenously. When in toxic doses it causes a profound drop in blood pressure, general collapse, drop in temperature, salivation, vomiting, and profuse diarrhea, which is often blood-stained. Splanchnic congestion is the conspicuous feature at autopsy and shows especially in the villi of the duodenal and jejunal mucosæ. Adrenalin, during this period of low blood pressure and splanchnic congestion, will cause the usual reaction when given intravenously, but applied locally or given intravenously it causes no bleaching of the engorged intestinal mucosa. Secretin is not found in the duodenal loop fluid, and the loop material does not influence the pancreatic secretion. Intraportal injection of the toxic material gives a reaction similar to intravenous injection. Intraperitoneal and subcutaneous injections produce a relatively slow reaction which closely resembles the picture seen in the closed duodenal loop dog. In both cases there is a relatively slow absorption, but the splanchnic congestion and other findings, though less intense, are present in both groups. There seems, therefore, to be no escape from the conclusion that a poisonous substance is formed in this closed duodenal loop which is absorbed from it and causes intoxication and death. Injection of this toxic substance into a normal dog gives intoxication and a reaction more intense but similar to that developing in a closed-loop dog.     

PROTEOSE INTOXICATIONS AND INJURY OF BODY PROTEIN : V. THE INCREASE IN NON-PROTEIN NITROGEN OF THE BLOOD IN ACUTE INFLAMMATORY PROCESSES AND ACUTE INTOXICATIONS.

Sterile abscess formation in the dog is accompanied by a large increase in output of urinary nitrogen and also by a small but definite increase in the blood non-protein nitrogen. All this nitrogenous material of course is derived from body protein injury and autolysis. Septic inflammation in the dog (pleurisy, pneumonia, peritonitis, etc.) likewise shows a distinct rise in the blood non-protein nitrogen. This rise is not often so great as that frequently observed in the intoxication of intestinal obstruction. Many acute infections in man (septicemia, peritonitis, pneumonia, etc.) show a definite rise in the non-protein nitrogen and urea nitrogen of the blood; some cases show a very great rise above normal (over 100 mg. of non-protein nitrogen per 100 cc. of blood). There may be no anatomical change in the kidney beyond the familiar picture of cloudy swelling. This does not exclude the possibility of some transient functional derangement of the kidney epithelium. Certain obscure intoxications in man may show a considerable rise in the non-protein nitrogen of the blood, indicating a large amount of protein disintegration. These findings must be taken into account in any clinical analysis and interpretation of high non-protein nitrogen of the blood in pathological conditions.     

INTESTINAL OBSTRUCTION : IV. THE MECHANISM OF ABSORPTION FROM THE MUCOSA OF CLOSED DUODENAL LOOPS.

Intoxication is evident in a drained duodenal loop whether it opens externally or into the jejunum and may be associated with more or less immunity which can be demonstrated after a period of days. Intoxication with a closed duodenal loop is identical whether the loop is left empty at operation or filled with a lethal dose of loop fluid. This again emphasizes the fact that absorption of the poison is essentially from the mucous membrane rather than from the contents of the closed loop. The intoxication of a closed duodenal loop is not modified by the presence of bile, pancreatic juice, or gastric secretion. Cessation of the normal flow of intestinal fluids which bathe the mucous membrane may be essentially responsible for the perverted activity of the mucosa and secretion of a poisonous material into the blood. Animals may be slightly more resistant to closed or drained loops during the warm months, which may be explained by the increased loss of body heat in the colder months. This indicates that cases of acute intestinal intoxication with subnormal temperature may be benefited by a generous supply of artificial heat.     

PROTEOSE INTOXICATIONS AND INJURY OF BODY PROTEIN : IV. THE METABOLISM OF DOGS WITH STERILE ABSCESS,PANCREATITIS, AND PLEURITIS.

Sterile abscess, pleuritis, and pancreatitis give a clinical reaction in the experimental animal very like the same acute inflammatory processes due to bacterial activity, provided the bacterial agents are limited to the initial location. The curve of urinary nitrogen excretion in the fasting dog shows the same precipitous and sustained rise in sterile and bacterial inflammatory reactions. This indicates that the same type of protein injury and autolysis in the body is produced by the sterile inflammatory reaction as by the bacterial reaction. It is assumed that the primary effect of the chemical agent or of the bacterial growth in the tissues is local cell injury or necrosis. This injured cell protoplasm undergoes prompt autolysis with escape of toxic protein split products. These toxic protein split products may be, in part at least, of the proteose group and are absorbed into the circulation, producing the familiar general reaction. The injury of body protein is obvious from the great increase in elimination of nitrogen in the urine and appears to be the same in sterile and in bacterial inflammation. The injurious agent in the sterile inflammation must be derived from the host protein, and we may assume with safety that much of the injurious material emanating from a septic inflammation must come from the host protein rather than from the bacteria. Acute sterile pancreatitis is one of the purest examples of an acute non-specific reaction where the intensity of the host''s intoxication may reach a maximum in 12 to 24 hours. We believe that fundamentally this reaction is very similar to that observed after the production of a sterile abscess or pleurisy. Non-specific intoxication must account for the sterile reactions described above. Septic inflammations show the same acute reaction and injury of body protein. The deduction is obvious—that a great part, at least, of the reaction in septic inflammation is truly non-specific and results from the primary injury of the host''s protein and cell autolysis.     

I. RENAL FUNCTION INFLUENCED BY INTESTINAL OBSTRUCTION

Associated with the intoxication of intestinal obstruction there exists a definite impairment of the excretory function of the kidneys. The degree of functional depression corresponds roughly with the intensity of the clinical intoxication. The decrease in the urea ratio and in the capacity of the kidneys to excrete sodium chloride is more marked than is the percentage decrease of phenolsulfonephthalein elimination. The great increase in the non-protein nitrogen of the blood usually observed in acute intestinal obstruction, which has hitherto been explained as being due entirely to an increased rate of protein catabolism, is due in part to retention of the products released from the injured cell protein. It is probable that the impaired renal function is due to direct action of the toxic substances upon the renal epithelium. The actual demonstration of this renal injury is perhaps the strongest evidence so far obtained to prove the presence of an actual toxic substance in the blood during intestinal obstruction. This obscure disability of the kidneys during the height of the intoxication of acute ileus should always be considered in the clinical management of this condition. It may also serve as a guide to indicate the degree of intoxication.     

IMMUNITY IN INTESTINAL OBSTRUCTION

1. There is no increased immunity or tolerance to intestinal obstruction after recovery from previous obstruction. 2. Dogs recovered from intestinal obstruction are not more resistant to injections of closed loop fluid than normal dogs. 3. Dogs injected with closed loop fluid are not more resistant to intestinal obstruction than normal dogs. 4. In dogs the normal variation in resistance both to intestinal obstruction and to the injection of closed loop fluid is large.     

PROTEOSE INTOXICATIONS AND INJURY OF BODY PROTEIN : III. TOXIC PROTEIN CATABOLISM AND ITS INFLUENCE UPON THE NON-PROTEIN NITROGEN PARTITION OF THE BLOOD.

The acute intoxication following an injection of a toxic proteose is usually associated with a large increase (40 per cent or more) in the non-protein nitrogen of the blood. This increase is found chiefly in the blood urea nitrogen, but the amino and peptide nitrogens also may show small increases. The changes observed in the blood non-protein nitrogen are identical with those which follow the feeding of large amounts of meat (8). These facts indicate that the proteose intoxication causes an abnormally rapid autodigestion of tissue proteins, but that the nitrogenous end-products are, in chief part at least, the same that result from normal catabolism of food proteins. There is no evidence that the autolytic products play any part in causing the intoxication. The possibility of such a part and a resultant vicious circle is not excluded, but from the available facts the autolysis appears more as a result rather than cause of the intoxication. It appears possible that in disease or intoxication tissue catabolism may be enormously accelerated and yet yield the end-products of normal protein metabolism.     

II. RENAL FUNCTION INFLUENCED BY PROTEOSE INTOXICATION

The injection of the toxic proteose obtained from the contents of the obstructed small intestine causes a definite impairment of the eliminative function of the kidneys as shown by a decreased capacity to excrete urea, sodium chloride, and phenolsulfonephthalein. This involvement of the renal function is similar to that shown by the preceding report (1) to accompany the intoxication of intestinal obstruction. The observed depression of function is readily demonstrable even when large amounts of fluid and urea, dye, or salt are injected directly into the blood stream. There is in all probability a temporary injury of the kidney cells, since the most important extrarenal factors have been largely eliminated in the above experiments. There is no appreciable impairment of the renal function following the injection of a number of other proteose preparations from a variety of sources. This study affords new evidence in favor of the view that the function of an organ can be profoundly disturbed for a time without any demonstrable anatomical lesions. The repair of this type of injury promptly follows the disappearance of the intoxication and is functionally and anatomically perfect.     

THE EFFECT OF PROTEIN SPLIT PRODUCTS ON THE SERUM FERMENTS AND ANTIFERMENT : STUDIES ON FERMENT ACTION. XXVI.

1. In dogs the toxic effect of primary proteoses is usually associated with the following serum changes: (a) an increase in serum antiferment, with a following fall in titer; (b) some increase in serum protease; (c) an increase in serum lipase; (d) a decrease in serum proteoses and amino nitrogen. 2. Secondary proteoses produce (a) less marked changes in the antiferment titer; (b) a marked increase in serum protease; (c) an increase in serum proteoses; (d) only a slight change in serum lipase; (e) a primary decrease in amino nitrogen. 3. The peptone which we have used (prepared from dog muscle) caused (a) a change in antiferment titer similar to that produced by the primary proteoses; (b) a marked increase in serum protease; (c) only a slight increase in serum lipase; (d) a primary decrease in proteoses, followed by an increase later; (e) an increase in amino-acids. 4. A very small dose of peptone resulted in a decrease in antiferment titer, together with a primary decrease in serum protease. 5. The peptone preparation was non-toxic when introduced into the stomach or rectum, while the intestinal injection was followed by an immediate intoxication.     

THE ORIGIN AND STRUCTURE OF A FIBROUS TISSUE FORMED IN WOUND HEALING

1. In experimental wounds, made by removing various sized pieces of skin from the frog, there is a rapid coagulation of the blood plasma and lymph to form a coagulation tissue which fills the wound cavity. 2. The observations on the living animals show that the coagulation a grave intoxication, but a low reading may be observed in some fatal cases and gives no assurance that a fatal intoxication may not supervene. The kidneys in practically all these experiments are normal in all respects. It is possible that protein or tissue destruction rather than impaired eliminative function is responsible for the rise in non-coagulable nitrogen of the blood in these acute intoxications. Transfusions of dextrose solutions often benefit intestinal obstruction, and may depress the level of the non-coagulable nitrogen in the blood. Some cases show no change in non-coagulable nitrogen following transfusions and diuresis, and, as a rule, such cases present the most severe intoxication.     

THE MECHANISM OF ANAPHYLACTIC SHOCK : STUDIES ON FERMENT ACTION. XXIII.

1. The serum ferments are practically unaltered by a primary injection of foreign protein. 2. During the course of sensitization the injection of the antigen is followed by the mobilization of a non-specific protease which increases in rapidity and intensity as the maximum period of sensitization is reached. 3. Acute shock is accompanied by: (a) The instantaneous mobilization of a large amount of non-specific protease; (b) a decrease in antiferment; (c) an increase in non-coagulable nitrogen of the serum; (d) an increase in amino-acids; (e) a primary decrease in serum proteoses. 4. Later there is a progressive increase in the non-coagulable nitrogen, in proteoses, and in serum lipase. 5. The acute intoxication is brought about by the cleavage of serum proteins (and proteoses) through the peptone stage by a non-specific protease. 6. The specific elements lie in the rapid, mobilization of this ferment and the colloidal serum changes which bring about the change in antiferment titer.     

SERUM CHANGES FOLLOWING KAOLIN INJECTIONS : STUDIES ON FERMENT ACTION. XXV.

1. The intoxication produced by the intravenous injection of inert substances such as kaolin is due to protein split products derived from the serum proteins. 2. The kaolin acts as an adsorbing medium for the serum antiferment, bringing about an alteration in the ferment-antiferment balance. 3. The intoxication is accompanied by an increase in serum protease, and of proteoses. 4. The serum lipase, the amino-acids, and the total non-coagulable nitrogen show relatively little change. 5. The antiferment shows an initial increase, followed by a loss.     

CHEMICAL CHANGES IN THE BLOOD OF THE DOG AFTER INTESTINAL OBSTRUCTION

A study of the non-protein nitrogen, urea nitrogen, uric acid, creatinine, amino-acid nitrogen, sugar, and chlorides of the blood and the CO₂-combining power of the plasma in normal dogs, and in dogs after different types of intestinal obstruction, is reported. Following ligation of the duodenum, ligation of the duodenum with gastroenterostomy, and ligation of the upper half of the ileum, a fall in chlorides and a rise in the non-protein nitrogen and urea nitrogen of the blood and in the CO₂-combining power of the plasma occur. The uric acid, creatinine, amino-acid nitrogen, and sugar show no significant changes. The fundamental change is a fall in chlorides followed by an alkalosis. The degree of alkalosis depends upon the rate of formation of carbonate, rate of excretion by the kidneys, and extent of neutralization of the carbonate by acid bodies formed during the intoxication. The fall in chlorides is probably due to a utilization of the chlorine ion in the course of the intoxication. It is suggested that this use of chlorine is a protective measure on the part of the body. There are indications that high intestinal obstruction should not be treated by the administration of alkalies. The urea nitrogen is a good index of the protein destruction. Ligation of the ileum at the ileocecal valve is followed by little increase in nitrogen and no change in the chlorides or CO₂-combining power of the plasma. The close similarity of the blood findings in intestinal obstruction, acute lobar pneumonia, and serum disease suggests that these widely different conditions may have a common chemical basis.     

INTESTINAL OBSTRUCTION : II. A STUDY OF A TOXIC SUBSTANCE PRODUCED BY THE MUCOSA OF CLOSED DUODENAL LOOPS.

The blood of closed duodenal loop dogs is not toxic to normal dogs. The blood of dogs that have been fatally poisoned with duodenal loop fluid is likewise non-toxic to normal dogs. The mucosa of closed or drained duodenal loops contains a toxic substance quite similar to the toxic material found in the lumen of the closed loops. This toxic substance is absorbed from the mucosa itself and not from the lumen of the drained loops. The same is probably true of the closed loops which have an intact mucosa. It seems highly probable that the poison is formed by the mucosa and is in great part absorbed directly from it by the blood. Normal intestinal mucosa contains no toxic substance nor can it neutralize in vitro the toxic substance produced in the closed loops. There is no evidence that the toxic material when given intravenously is excreted by the intestine or held by the intestinal mucosa in any demonstrable form. The toxic substance is not absorbed from the normal intestinal tract. Destruction of the mucosa in a closed loop by means of sodium fluoride prevents the formation of the toxic substance. This fact furnishes the final proof that the mucosa is the essential factor in the elaboration of the poisonous material.