Experimental Biology and Medicine

1) The mechanism by which ingestion of large amounts of nicotinic acid lowers serum cholesterol in rabbits has been investigated. The rate at which cholesterol is synthesized from acetate-1-C¹⁴ by liver slices from animals on control or cholesterol supplemented diets, with or without nicotinic acid., has been measured. 2) There is marked inhibition of the rate of cholesterol synthesis by liver slices from animals fed nicotinic acid on both control and supplemented diets. Since the principal detoxication product of large doses of nicotinic acid is nicotinuric acid, it is possible that this inhibition of cholesterol synthesis occurs as a direct result of competition of lipid synthesizing and detoxication systems for a limiting amount of Co A in the liver cell.

Polio virus was found to replicate in HeLa cells treated with critical concentrations of 2,4-dinitrophenol which markedly stimulated oxygen uptake and glucose utilization under aerobic conditions. The time required before a change in rate of respiration of infected cells became apparent was similar in absence or presence of DNP, indicating that this substance did not alter the latent period of virus synthesis. Apparently the uncoupling effect on aerobic phosphorylating systems is compensated for by increased glycolysis and the concomitant fixation of phosphorous by systems not effected by DNP.

Purified corticotropin and growth hormone both stimulate lipid mobilization from the epididymal fat depots of fasting mice. Hydrocortisone failed to induce lipid loss in intact mice and inhibited fat mobilization in adrenalectomized mice, indicating that the lipid mobilizing effect of corticotropin is not mediated by adrenal stimulation. Furthermore, inhibition of fat loss by hydrocortisone in adrenalectomized mice could be overcome by simultaneous administration of corticotropin. The results demonstrate an extra-adrenal action of corticotropin on fat mobilization in the intact animal similar to that induced by growth hormone.

Gastric secretion in the intact rat is inhibited by fat in the small intestine. This inhibitory activity is decreased when bile has been diverted previously from the small intestine. Under these same conditions, addition of bile salts to administered fat results in the same degree of inhibition as in the intact animal.

Ten normal subjects ingested equimolar amounts of mephenesin, mephenesin carbamate and methocarbamol, with intervals of at least one week between medications. Blood levels of drugs were followed for 4 hours, and in one case for 7 1/2 hours. In all subjects mephenesin carbamate gave higher peak values than mephenesin. Methocarbamol exceeded the other 2 drugs significantly in this regard.

A factor in normal chicken serum when added to heated or unheated immune turkey serum-antigen complexes caused fixation of guinea pig complement. The factor was labile at 56°, unstable at room or refrigerator temperatures but remained active for several weeks at —20°C It was found in the water insoluble fraction of serum and could be removed from diluted serum on kaolin.

An agglutination method for detecting antibodies against blood lymphocytes is described. By this method non-specific clumping of polymorphonuclear leucocytes and/or monocytes usually encountered in leucoagglutination tests is avoided. The humoral response following homografting in chickens was reinvestigated using this lymphoagglutination reaction. In 82 combinations of normal sera or sera from autografted chickens, and lymphocytes, lymphoagglutination was found in only 5 instances. On the other hand, 103 cases of positive agglutination were obtained in tests with 107 combinations of immune sera and normal lymphocytes. These immune sera, including one with titer of 1:1024, were from animals previously injected with pooled homologous spleens. Sera from animals injected with splenic cells or grafted with skin from one donor, also contained lymphoagglutinins against lymphocytes of spleen or skin donor chicken. These lymphoagglutinins also reacted in some instances with lymphocytes of chickens other than spleen or skin donor.

When different amounts of uniformly labeled L-threonine, L-valine, L-histidine, or glucose were administered intraperitoneally to young chicks previously fed complete purified diets or diets lacking different essential amino acids, the total percentage of administered activity recovered as C¹⁴O₂ during 14 hour experimental periods averaged 10% for threonine, 25% for histidine, 40% for valine, and 50% for glucose. There were large differences in the fraction of each amino acid incorporated into picric acid-precipitable protein. Peak C¹⁴O₂ production occurred about one hour after administration of glucose, threonine, or valine; 2 hours after administration of histidine. Neither previous diet (complete or lacking the amino acid injected) nor 10-fold variations in amounts of amino acids administered appeared to affect fractions oxidized or synthesized into proteins.

Comparative rates of passage of strontium and calcium from plasma into cerebrospinal fluid have been studied in man following intravenous injection of Sr⁸⁵ and Ca⁴⁵. Cerebrospinal fluid to plasma ratios of Sr⁸⁵ and Ca⁴⁵ were approximately the same except in one patient who showed a greater ratio for Ca⁴⁵. As a greater proportion of Sr⁸⁵ than of Ca⁴⁵ is unbound to protein, differential passage into the cerebrospinal fluid in favor of Ca⁴⁵ appears to take place. The data indicate that cerebrospinal fluid may be formed by secretion rather than ultrafiltration. Rate of formation of fluid has been calculated to be approximately 0.03 to 0.04 ml/hr/kg of body weight.

In a study of effects of antibiotics on multiplication of L cells in suspension culture, positive inhibition of multiplication was found when less than 1 μg/ml of any one of the following were added to the media: actinomycins B, B_V, or D_IV; cycloheximide; gliotoxin; hygromycin; mitomycin C; patulin; thiostrepton; and xanthomycin. Actinomycin B_V was the most active of the group tested as 1.5 mμg/ml was sufficient to cause a 50% reduction in growth of culture. While the shape of dose-response curves varied with different inhibitors, it was possible to use the linear portion as the basis of a method for quantitative estimation of concentration of inhibitor present. This method may be useful in determining the presence of cytotoxic metabolites formed in microbial fermentations.

1) In leukemic strains of mice, lymphoid leukemia is known to occur less often and later in males than females. However, in our AK mice, males that were isolated early in life and did not fight died of spontaneous leukemia at essentially the same age and in the same proportion as females, whereas fighting males died significantly later and in a lower percentage. Hence the difference between sexes is not explained by sex hormones only. An interpretation is proposed, based on adrenal corticosteroid secretion. 2) Fighting activity should be taken into account as a factor in pathogenesis of mouse leukemia and in the difference of incidence and survival time between male and female mice.

(1) Doses of 0.25 mc of radioactive yttrium phosphate (Y⁹⁰PO₄) were injected into peritoneal cavity of Albino mice immediately following administration of 2 ml of 0.85% NaCl solution; on 3rd to 4th day 10⁵ cells of Krebs-2 Ascites Tumor were inoculated by same route into pre-treated animals and normal controls; their accumulated fluid was withdrawn after 3 or 4 days for estimate of its volume, tumor cell counts and smears; they were sacrificed for autopsies 1 or 2 days later. (2) Amount of ascitic fluid and extent of implant growth were considerably reduced in pretreated animals as compared with controls; free tumor cells in the fluid appeared enlarged, altered (distorted mitoses) and less viable, after transfer into new mice, than in controls; decrease of their concentration in the fluid was not as rapid and extensive as in animals treated with an isotope after inoculation in previous experiments. (3) Cell destruction was increased by injection of 0.1 mc into pretreated mice. (4) It was concluded that pretreatment with radioactive yttrium increased resistance of the peritoneum to invasion with tumor cells, thus decreasing implantation of these cells and exudation of the fluid. (5) It was presumed that the principle of this method may be eventually applied for prevention of “transcoelomic metastases” from some abdominal tumors, before or after their removal.

Dietary administration to rats of the anti-thyroid drug, propylthiouracil, prevented development of hypertension accompanying administration of hypertonic (1.75%) NaCl solution as sole drinking fluid. Propylthiouracil administration prevented not only the expected rise in systolic blood pressure but also cardiac and renal hypertrophy. Although the results suggest the possibility that thyroid gland may play a role in salt hypertension, they could be accounted for by extra-thyroidal effects of the drug.

Glucose-6-phosphatase (G-6-Pase) and acid phosphatase (Ac-Pase) activities have been followed histochemically in hyperfunctioning rabbit pancreatic B cells in cortisone diabetes and after administration of hypoglycemic sulfonylureas. Cortisone diabetic rabbit pancreas with B cell degranulation and glycogenic vacuolization of ductules and B cells showed no definite changes in B cell Ac-Pase or G-6-Pase activity. Hypoglycemic sulfonylureas administered over period adequate to degranulate B cells, seemed to diminish their G-6-Pase content without effecting Ac-Pase. Although B cell degranulation was, in each of these instances, indicative of increased insulin output, lack of alteration of histochemically demonstrable B cell Ac-Pase activity was not considered to be at variance with the view that this enzyme may be important for insulin synthesis. Inhibition of B cell G-6-Pase activity by chronic sulfonylurea administration could be the mechanism of pancreatic action of these drugs. However, the alternative explanation that reduced G-6-Pase activity was a secondary adaptive response cannot be entirely excluded.

Eight purine analogs were compared as to their effects on growth of Tetrahymena A certain degree of correlation was found between type of structural change present in the analog and ability of adenine, acetate, sterol or combinations of these compounds to reverse the inhibition. Analogs which cause “suicide” of cultures do not fit the pattern of other inhibitors.

Previous investigators observed clot lysis by tissue grown in plasma clots(14, 15). Our results indicate that an “activator” of plasminogen is released from KB cell cultures. Supernatant from MK cell cultures activated plasminogen and was also proteolytic in the absence of plasminogen. These cell cultures had a third property, inhibition of proteolysis. The activator described here resembles “tissue activator” in that it is not proteolytic in the absence of serum and can be dissolved by KSCN. It also resembles activator from excised rat lung(16) in that it is the size of microsomes. The interpretation that release of activator from cell culture is a metabolic process and not from cell breakdown seems warranted since only a small fraction of the activity found in the supernatant can be extracted from the cells.

A method based on release of an intracellular, lysozyme-like enzyme from lysed leukocytes was an acceptable substitute for microscopic cell counts in measuring tuberculin cytolysis of leukocytes from guinea pigs injected with heat-killed, human type. M. tuberculosis.

1) Fibers of acid-soluble collagen reconstituted with chondroitin sulfate showed sheet-like structures without indication of spacings. After treatment for 2 days with basal salt solution at pH 7.3 and 37°C, containing neither calcium nor phosphate ions, non-striated fibers were formed of 300-1500 Å thickness. If either calcium or phosphate ions were present partial transformation to the form with 640 Å spacings took place. If both of these ions were present this transformation occurred more rapidly and more completely. 2) These findings suggest that calcium and phosphate ions interact with sites on the collagen fiber where the breaking of electrostatic bonds catalyzes transformation to the 640 Å spaced fiber.

Feeding various bile acids to mice for 3 weeks had the following effects: 1. Cholic acid increased hepatic and intestinal cholesterol levels, but had no effect on kidney cholesterol. It decreased hepatic cholesterol synthesis rates, but had no effect on intestinal synthesis rates. 2. Hyodeoxycholic and lithocholic acids decreased liver cholesterol levels. Hyodeoxycholic acid had no effect on intestine and kidney cholesterol, or on intestinal cholesterol-x-C¹⁴. Both acids effected large increases in hepatic cholesterol synthesis. 3. Deoxycholic acid significantly decreased liver, small intestine and kidney cholesterol levels. It also decreased hepatic cholesterol synthesis. 4. Results of this study suggest that homeostatic control of cholesterol metabolism in tissues other than liver is independent of hepatic control.

Extracts of the stalk-median eminence area of beef or rat brain produce a specific release of ACTH in rats with hypothalamic lesions. The major portion of this effect is due to a corticotrophin-releasing factor. Ten to 20% of the activity of these extracts appears to be due to vasopressin.

1. DPN injection produces toxic effects in mice similar to those described for nicotinamide analogs. 2. Toxicity of injected DPN is influenced by environmental temperatures; both cold and hot environment enhances toxicity. 3. Simultaneous cortisone potentiates the effect of heat and cold. 4. These effects are discussed as they bear on inhibition of DPN-ase activity of reserpine, promazine and nicotinamide.

The effect of feeding either desiccated thyroid or propylthiouracil was studied to determine if the former drug resulted in an anticariogenic effect and the latter produced a cariogenic effect in adult rats to the same degree as in weanling animals. Propylthiouracil resulted in significantly more caries than were found in weanling animals, but desiccated thyroid did not reduce the caries experience. Similar histologic changes were found in both thyroid and salivary glands in the adult rats as were found previously in weanlings.

High compression pressures attained at below freezing temperatures have been studied for their effects on influenza virus hemagglutinin, infectivity and antigenicity. Influenza virus hemagglutinin from the virus strains tested varies markedly in its resistance to pressure inactivation.

The method of differential adsorption has been successfully applied to separation of lymphocytes from granulocytes in human blood. When whole blood was passed through a short column of siliconized glass wool, the granulocytes were almost quantitatively retained on the column, and uninjured lymphocytes in 30-89% yield appeared in the effluent blood.

Elevated levels of epinephrine are found immediately after administration of a lethal dose of endotoxin. Following this, plasma catechol amine concentration is only slightly above control levels. Prior to death, toxic levels of epinephrine are attained. Serum serotonin concentration falls immediately after administration of endotoxin and remains low throughout the shock state.

1. Sera from newborn infants were examined by gel diffusion against whole culture concentrate of type 3, Group A beta hemolytic streptococcus. One to 3 bands of precipitation were obtained with average of 2.2 bands/individual. In the first 3 to 4 months of life the average number of bands/individual diminished sharply to almost complete absence. From 4 to 11 months of age the average number of bands/individual was 0.13. On twelfth month the average number/infant rose to 0.6, and in adults an average of 2.45 was obtained. These results are interpreted on the basis of transplacental transfer of antibodies which are short-lived and reappear with subsequent streptococcal infection later in life. 2. Sera from 331 apparently normal adults were studied by the gel diffusion method and simultaneously assayed for antistreptolysin O. A close correlation was found between number of bands and ASO titers, not apparent when individual cases were compared. With each increase in number of bands there was a corresponding increase in ASO titer.

Papillary muscles of the cat subjected to various resting tensions exhibited morphologic changes, the severity of which could be correlated with degree of loading with loads of 3 g or greater. These changes were similar to those which occur post mortem in cardiac muscle and appeared whether the muscle was stimulated to contract or subjected to static tension. Morphologic alterations appeared in both isotonic and isometric preparations but appeared more severe in isotonic preparations. Rate and degree of failure of isotonically contracting muscles could be correlated with size of the resting load but such a correlation was not observed with muscles contracting under isometric conditions.

1. Virulent and attenuated strains of the 3 types of poliovirus have been shown to have different patterns of growth at high (39°C) temperatures when tested by means of agar overlay tissue culture technic. Attenuated strains (T marker) are inhibited and have a low EOP at this temperature, while virulent strains (T⁺) grow equally well at 35°, 37°, and 39°C. Some strains give an intermediate (I) pattern. 2. The thermal marker T-T⁺ was used to screen poliovirus strains isolated during a type I oral vaccine field trial, in an effort to determine: a) whether reversion to greater virulence had occurred as a result of human passage; and b) whether strains isolated from local flies were the vaccine virus which had been orally administered; or were naturally occurring wild strains. The results indicated that virus excreted by 2 of the 3 infected children consistently had thermal markers identical with that of the vaccine virus; one child, on the other hand, excreted virus which over a period of 29 days showed a shift to the thermal marker pattern of more virulent strains. Neurovirulence for monkeys was low for all strains isolated from children. One T strain isolated from flies had an EOP at 39°C in the same range as the LSc vaccine strain, while the 14 other fly strains tested all fell between this range and that of virulent strains. It was concluded that most of the fly strains recovered in the course of the field trial probably represented naturally occurring wild strains. 3. Comparison was made between the thermal (T-T⁺) marker, the EOP at low and high bicarbonate concentrations (d-d⁺ marker), and neurovirulence as tested by intracerebral inoculation of monkeys. The thermal marker correlated better with monkey virulence than did the d-d⁺ marker. 4. Use of the thermal in vitro marker should be of some practical value in determining degree of spread of attenuated poliovirus strains introduced into a community, and in assessing degree of stability of vaccine strains on human passage.

The 24-hour in vitro transfer of radiocadmium, Cd¹¹⁵, from plasma to human erythrocytes was investigated. Most of the isotope became erythrocyte-bound during the first 8 hours of incubation although subsequent kinetics were somewhat obscured by hemolysis. The magnitude of the shift approximated 30% of total cadmium, although the erythrocytes of one subject, who had recently hemorrhaged, exhibited a much greater affinity for the metal. These findings suggest the desirability of further studies to determine the value of cadmium as an indicator of erythrocyte survival.

Free coagulase can be measured adequately by either the tube-plasma method or the tellurite-glycine method. We were unable to establish any direct relationship between certain groups of phage-typable staphylococci and production of alpha, beta or delta hemolysins. Fibrinolytic activity was variable but 50% of Group d (non-typable) exhibited some activity.

1. Glucose utilization, phosphate uptake and reduced glutathione content of erythrocytes are significantly lowered following uretero-caval anastomosis and contralateral nephrectomy of the dog. 2. Above biochemical changes are of the same magnitude as following bilateral nephrectomy. Thus presence of hypertrophying renal tissue without an excretory outlet is not protective toward these alterations.

Oral administration of 100,000 I. U. of Vit. A acetate for 4 to 6 months significantly reduced the elevated serum cholesterol levels in atherosclerotic patients but had no effect on individuals with normal cholesterol levels.

The trachea of the living rabbit was observed during exposure to un-ionized and ionized N₂, O₂ and CO₂ using a separate airway for respiration. Positively charged CO₂ was found to be responsible for: reduction of ciliary activity, contracture of smooth muscle, ischemia and enhanced vulnerability to trauma.

1) A contaminant, subsequently identified as a virus of the mouse hepatitis group, appeared in Princeton weanlings during the 132nd intraperitoneal passage of an ascites tumor. The virus was regularly demonstrable in the ascitic fluid and was readily maintained in both Princeton and Swiss mice by serial transfer of fluid. In its presence, livers showed necrotic lesions and the ascitic fluid was altered in amount, consistency, and color. An artificial mixture of the 2 agents behaved in much the same way as the natural mixture. 2) The 2 mouse strains were equally susceptible to the tumor but not to the virus. Liver lesions produced by the virus alone were much more pronounced in Princeton mice. Presence of both agents facilitated migration of tumor cells to the liver and resulted in a marked enhancement of the hepatic reaction in Swiss mice.

Development of righting reflexes was studied in offspring of manganese-deficient and normal rats by 2 tests: (1) time required to turn from their backs to their feet and (2) ability to right themselves in air when dropped upside down. The reflexes concerned with the turn-over test developed at an earlier age than did those involved in the air-righting response. In both tests, however, the manganese-deficient young exhibited a marked delay in development of reflexes responsible for righting reactions.

The gastrocnemius muscles of 30-day-old guinea pigs subjected to a vit. E-deficient diet for 21 or 30 days undergo a statistically significant increase in collagen nitrogen and decrease in contractile protein nitrogen. Masseter muscles are more resistant to the effects of vit. E-lack on the protein components. Sarcoplasmic nitrogen is not altered by 21 or 30 day dietary deficiency regimens.

1. Serum triglycerides and cholesterol were decreased, and fecal bile acids and digitonin precipitable sterols were increased, after injection of heparin in 4 human subjects. 2. Evidence was discussed which indicates that this effect of heparin upon cholesterol metabolism is a secondary phenomenon resulting from maintained reduction of circulating triglycerides.

1) Average weights per inch of height for both sexes in older population, ages 65 to 94, have been presented. This is the first study of its kind to appear. It is based on a population sample of apparently healthy persons and the sample size is considered statistically adequate. 2) There exists a definite trend towards a decrease in average weight of successive 5 year age groups after age of 65. There were 2 reasons for this: 1) individual weight loss accompanying aging due to loss of body substance, 2) high mortality rate among obese which favors survival of a larger proportion of underweight people. 3) Height and survival were not related.

The effects of δl-testololactone, δ4-androstene 17α-ol-3-one-17-β-oic acid, and fluoromevalonic acid upon in vitro conversion of labeled acetate and labeled mevalonate into cholesterol were studied. All of these compounds inhibited conversion of acetate, while only the latter 2 inhibited conversion of mevalonate. Fluoromevalonic acid was the most potent inhibitor studied. Fluoroacetate, fluoride ions, and zinc ions had no effect on the mevalonate system. Of these substances only zinc ions inhibited conversion of acetate to cholesterol.

Nitrogen and phosphorus contents of large and small granules and cytoplasmic supernatant from livers of mice 1 1/2-2, 6-7, 10-11, and 18-21 months of age were determined and expressed on a per nucleus basis. There were increased nitrogen contents, and relatively smaller increases in phosphorus contents in each of these fractions in older animals. The results, viewed in the light of observations of Weinbach and Garbus (7), suggest that as an animal ages, its liver cells lose metabolic capacity, and, also, that an attempt is made to restore at least some aspects of it by forming more of the active sites within the cell. Cell fractions examined in older animals differed chemically from those in younger animals, however, as indicated by decreased P/N ratios.

Corn oil or lard from which tocopherol had been removed, promoted chick encephalomalacia, whereas coconut oil, butter, linseed oil, and cod liver oil did not produce symptoms, and olive oil had a questionable effect. Dietary combinations of 2% corn oil with either 8% coconut oil, lauric acid, myristic acid, or a mixture of saturated fatty acids like coconut oil significantly increased the incidence of encephalomalacia over 2% corn oil alone, while 6% linseed oil, cod liver oil, or oleic acid inhibited the effect of 4% corn oil. Olive oil, butter, fatty acids like butter, palmitic acid, and stearic acid had no net effects upon the incidence of encephalomalacia induced by corn oil. The intake of linoleic acid appears to be a primary factor in the etiology of encephalomalacia, but some of the other fatty acids may secondarily increase or decrease this effect.

A method is described for determining penicillin in poliomyelitis vaccine. This method consists of acidifying the vaccine, extracting the penicillin into diethyl ether, removal of ether by vacuum distillation and transfer of penicillin into a buffer. The penicillin is determined by standard paper chromatographic and bioautographic technics. This method is capable of detecting 0.001 unit of penicillin/ml finished vaccine.

Treatment of rabbits with tranylcypromine followed by the injection of reserpine produces central nervous system stimulation and a marked rise in rectal temperature. The effects produced by tranylcypromine are assumed to reflect the activity of this agent as an inhibitor of monoamine oxidase.

Ascites fluid induced in mice by intraperitoneal injection of Ehrlich tumor cells contained specific antibody to S. typhimurium when mice were immunized against this organism. The serum and ascites were identical in their anticomplementary properties and neither contained detectable complement. The serum had normal properdin levels. but the ascites did not contain detectable properdin under the assay conditions employed.

1. If a subcutaneously implanted Yoshida sarcoma is sensitive to nitrogen mustard N-oxide (“NMO”), pre-treatment or short-time post-treatment with NMO produces remarkable metastases. The severity of these metastases following pre-treatment with NMO suggests a strong effect on the host. 2. Treatment of NMO-resistant tumors with NMO does not promote the development of metastases, even with pre-treatment or short-time post-treatment. This indicates that NMO-resistant tumors do not liberate cells to form metastases even when the host is under the influence of NMO. 3. Treatment of tumors with X rays produces essentially the same results as treatment of NMO-sensitive tumors with NMO does. In both instances, diminution of host resistance appears to be an important factor in promoting the development of metastases. 4. Pre-treatment of tumors with cortisone promotes the development of metastases; post-treatment, whether short-time or extended, has no such effect. Evidently the chief action of cortisone is on the host, increasing the ease with which metastases are formed. 5. Post-treatment with either NMO or X rays for an extended period inhibits the formation of metastases, and causes the tumor itself to regress. This observation suggests that carcinostatic agents should be used clinically only in large enough doses.

Active erythropoietic factor is demonstrated in plasma and urine of bled dogs. A very striking erythropoietic factor response is obtained when severe anemia is produced. Activity is partially destroyed by boiling and by prolonged storage in freezer.

The normal rabbit, treated with hypoglycemic doses of phenethylbiguanide, shows a significant increase in serum inorganic phosphate levels. In this respect the action of phenethylbiguanide differs from that of insulin and sulfonylurea oral hypoglycemic agents, and closely resembles that of structurally related diguanidine, Synthalin A. Increase in serum inorganic phosphorus levels is consistent with inhibition of oxidative enzyme systems of the citric acid cycle in presence of phenethylbiguanide, which suggests that inorganic phosphorus might escape into the blood stream due to decreased binding of high energy phosphorus.

In monkey kidney cultures increments in infectious 2060 could be correlated with the degree of observed cytopathology. The absence of glutamine or vitamins contained in Eagle' medium had some effect in lessening yield of infectious virus, but alterations in pH away from neutrality were more significant in reducing viral multiplication. Infectious 2060 was relatively unstable as evidenced by the inactivation rate at temperatures of 35°C and above and pH below 5.0 and above 7.1. JH virus and 2060 were shown to be related antigenically, but were not identical, as shown by reciprocal neutralization titrations with animal and human antisera. Additional instances of neutralization titer increments in man following symptoms of mild respiratory illness have been presented.

An operative technic has been described which causes development of congestive heart failure in the guinea pig within 8 to 12 days. Essentially the technic involves intrathoracic incision and restriction of aortic blood flow by ligature surrounding the aorta. Tissues derived from this preparation may be subjected to biochemical and enzymologic studies because of feasibility of preparing slices, homogenates, cell free suspensions and isolated particles. Preliminary biochemical work has been accomplished with this preparation.

1) Bishydroxycoumarin-induced hypoprothrombinemia may result in a slight delay of fibrin clot in isolated venous segments of anesthetized dogs. 2) The hemorrhagic state produced by the drug fails to prevent acceleration of intravascular clotting caused by intravenous infusion of fat emulsion. This clot acceleration, therefore, may not operate through the mechanism of prothrombin conversion. 3) Our experiments lend support to the concept that anticoagulation with coumarin-type drugs is not synonymous with antithrombotic effect.

These studies indicate that, in general, alpha deflections reflect net changes in calibre of precapillary vessels and beta and gamma deflections reflect net changes in calibre of postcapillary vessels and that these changes in pre- and postcapillary vessels are probably important in the regulation of peripheral circulation and flow of tissue fluid and lymph.

Serial cultivation of cells obtained from the human pituitary is described. Somatotropin, corticotropin, and gonadotropin have been detected in these cultures. Two of the cell lines have produced gonadotropin, both FSH and ICSH, when in suspension culture, and one, S4374-M, has been frozen and recultivated in both stationary- and suspension types of culture with media containing 10% of human, equine or bovine serum, with resumption of production of hormone. The gonadotropin-producing cells have the microscopical appearance of a chromophobe type of pituitary cell and have a characteristic appearance when stained by the iron-PAS method. These cells are heteroploid with a chromosome count in narrow range with modality at 79.

Immature AKD2F₁ mice were examined for possible estrogenic inhibition of Sarcoma 1. Large doses of estradiol benzoate inhibited growth of tumor in intact animals and in hysterectomized castrates. It was concluded that this estrogenic effect can be achieved in absence of uterus, and that the tumor-inhibitory properties of endometrial secretions act by a mechanism different from that of estrogen.

1) Starvation or glucose plus tolbutamide potentiates the diabetogenic activity of alloxan with increased mortality. 2) Glucose alone or glucose plus insulin protects against alloxan diabetes. 3) Different results obtained with insulin or tolbutamide cannot be explained on the basis of liver glycogen content, blood glucose, or theoretical status of pancreatic activity at time of alloxan injection.

1. Normal, adult Z(C3H) mice, connected in parabiotic union with Z(C3H) mice tolerant of Ce tissue, fail to reject homografts of Ce skin. 2. Lack of immunological responsiveness to Ce tissues persists following disruption of parabiotic union. 3. The possibility that failure of transplantation reaction is due to transfer of immunological tolerance to previously unconditioned adult recipient is presented. 4. The implications of these observations are discussed.

Dwarf mice and their normal siblings were given thyroxine, thyrotrophic and somatotropic hormones or physiological saline intraperitoneally. In the skin the mast cells were counted and concentration of hexosamine and hydroxyproline were determined. The number of mast cells did not show any significant variation in the different groups. Concentration of hexosamine in the skin of thyrotrophin-treated dwarf mice was significantly higher than concentration in control dwarf mice. Hexosamine values in the group of thyroxine-treated mice were higher than in the group of control mice. Hydroxyproline content was significantly higher in the skin of thyrotrophin-treated dwarf mice than in thyroxine-treated dwarf mice.

Pullets receiving testosterone propionate demonstrated a significant increase in red blood cell count and hematocrit and a decrease in spleen weight.

From these data, it is apparent that whole activated pancreas, unlike pancreatic juice, contains a thermo-labile material which can reduce by 50% both viscosity and ability to form fibers of acid soluble collagen solutions. This activity is not affected by various trypsin inhibitors, nor is it duplicated by trypsin, elastase, chymotrypsin or hyaluronidase alone. This activity may be due to a new enzyme in the pancreas, a collagenase. In view of lack of activity of this enzyme upon serum and egg albumin, casein and gelatin, this enzyme would seem to be reasonably specific for acid soluble collagen.

Injection of certain bacterial polysaccharides into the peritoneal cavity of rats results in a local neutrophilia which can easily be determined and quantitated. Preliminary findings suggest the possibility of using this response to detect the presence of certain bacterial derivatives in solutions.

Administration of large amounts of iodine to hamsters which have hyperplastic goiters produced by an iodine-deficient diet or by a goitrogen results in a morphologic response which is different from that encountered when iodine is given in physiological amounts. An acute inflammatory reaction and a diminution of the accumulation of colloid are found, neither of which has been observed in animals treated with smaller amounts of iodine.

Virus synthesis, cytopathogenicity, and modification of virulence for chicks, of AEV in chick kidney cell culture after 10 serial passages, have been described. The cell culture virus has been identified by histopathologic changes in inoculated chicks and virus neutralization tests in a cell culture system. The possible application of the findings are discussed.

The RH strain of Toxoplasma gondii induces formation of necrotic plaques on monolayers of chick embryo fibroblasts overlaid with nutrient agar. Correlation between number of organisms plated and number of plaques produced indicates that each plaque is induced by a single toxoplasma. Survival of toxoplasmata in various media has been tested, and it has been found that Hanks BSS containing chick embryo extract in concentrations of 5 to 50% preserves 80-100% of plaque forming units (pfu) for 3 to 4 hours, 50-60% for 6 hours. Adsorption of pfu on monolayers is maximal after 70 to 90 minutes (Kc = 0.024), can be retarded by increased viscosity of the medium, speeded by centrifugation. Artificial lysis of chick embryo fibroblasts infected at low multiplicity yields populations with a predictable efficiency of plating of about 50% of the counted organisms.

1) In Group II type of preparation described herein (antral exclusion with 50% of acid peptic secreting tissue attached to antrum) no abettment of ulcer diathesis was noted. Ulcer was not observed at either neostoma. 2) When, however, a similar fundic pouch was attached to the duodenum (Group I preparation) ulcer occurred at this neostoma in 50% of dogs studied and interestingly not in the short duodenal segment, but just over on the jejunal side, an occurrence which suggests an increased gradient of susceptibility to peptic ulceration.

Alterations in rabbit aortae were brought about by intravenous administration of epinephrine. Microscopic examination revealed accumulation of metachromatic ground substance. The aortic content of hexosamine and water together with the uptake of ³⁵S sulfate was increased, whereas the amount of hydroxyproline was unaltered. The increase in the mucopolysaccharide/collagen ratio may indicate a reparative process.

1. A single subcutaneous injection of carrageenin in rats caused a local tissue response, which from the 1st through the 4th day could be defined as an inflammatory lesion in the form of an acute abscess, which later (5th through the 28th day) was gradually transformed into a polysaccharide-containing granuloma. 2. Weight data indicate a maximum exudative response after 24 hours.

Evidence is presented for antigenicity of TSH, ACTH, and growth hormone. Rabbit antisera to these hormones were prepared and then tested by means of a hemagglutination test in which antigen is coupled to red blood cells, by means of bisdiazotized benzidine. The antibodies to TSH showed extensive cross reactions with other pituitary hormones. The antibodies to ACTH appeared to be hormone specific but not species specific. Antibodies to bovine, porcine, and human growth hormones appeared to be species specific, and for the most part, hormone specific. The application of human growth hormone and ACTH antisera to detection and measurement of these hormones in human body fluids is in progress.

1) Because ingestion of neomycin results in hypocholesterolemia in human subjects, the effect of this antibiotic on lipids of serum and liver has been studied in rats. 2) Basal control diet or cholesterol rich diet, alone or added with 0.2% neomycin, has been fed to rats for 4 months. This prolonged administration of neomycin to rats was not associated with any significant changes in serum or liver levels of cholesterol and phospholipids.

Detectable quantities of GOT and GPT were found in 100% and in 89% respectively of samples of stimulated saliva from 81 patients. The saliva GOT content exceeded saliva GPT level in 91% of cases. Incubation of an aliquot of each saliva sample with glucose resulted in a pronounced increase in GPT and diminution in GOT. Comparison of saliva transaminase levels with those of concurrently collected samples of serum showed no significant correlation, thus signifying that the prime and immediate source of saliva transaminase is oral rather than systemic.

(1) Progesterone administration to rats and dogs with experimental hypertension and to humans with primary arterial hypertension resulted in a decline in blood pressure levels. Blood pressures increased once more in all cases after progesterone was discontinued. (2) In humans, but not necessarily in rats and dogs, blood pressure reduction would appear to have resulted from natruresis.

Right-to-left circulatory shunts were produced in 11 mongrel dogs by anastomosis of the left atrial appendage to the pulmonary artery. When injections of Kr⁸⁵ in solution were made proximal to the shunt the level of radioactivity in arterial blood drawn during the first 20 sec. after injection was always greater than 1400 cts./min. In dogs without shunts, however, similar injections resulted in arterial blood counts of 206/min. or less. The technic thus provides a new method for detection and localization of venoarterial shunts.

6-MP, a powerful metabolic antagonist to nucleic acid synthesis, prolongs survival time of skin homografts in rabbits when administered after grafting. Treatment must be given continuously to maintain graft viability. However, toxicity of 6-MP limits the length of time treatment may be given.

It was shown that a 1:1 cholesterol:sitosterol mixed crystal are formed not only in aqueous in vitro systems, but also in vivo.

1) A histochemical study has been conducted of the distribution of glucose-6-phosphate dehydrogenase (G-6-PD), triphosphopyridine nucleotide diaphorase (TPND), lactic dehydrogenase (LD), diphosphopyridine nucleotide diaphorase (DPND) and succinic dehydrogenase (SD) in the pancreas of the rabbit. G-6-PD was found principally in B cells and duct epithelium; LD in duct epithelium, acinar tissue and A cells. TPND was present throughout, with greatest activity in B cells and in duct epithelium and good activity in other areas. SD activity was present throughout the pancreas with greatest staining in B cells and duct epithelium. The reactive structures for SD seemed to be mitochondrial while the other enzymes apparently are mostly extramitochondrial. 2) These findings demonstrate that glucose metabolism in B cells is predominantly via the hexose monophosphate shunt, and indicates a role for it in insulin synthesis. A relationship between TPNH generation by this pathway and availability of reduced glutathione or of free SH groups in enzymes is considered as a possible mechanism controlling B cell insulin output.

Chick skeletal muscle fibroblasts in monolayer cultures grow actively in homologous undialyzed serum supplemented with Medium 199. Dialysis of chicken serum removes a factor essential for growth and maintenance of these cells. The deficiency syndrome in cultures with dialyzed serum can be corrected by chicken serum dialysate, but not by inositol, cholesterol, vit. B₁₂, spleen nucleoprotein fraction, or by a more complex synthetic nutrient, NCTC 109. Proteosepeptone at 0.1% may be substituted for serum dialysate to restore full activity of whole serum.

1) In an attempt to contribute to an understanding of unusual radiation sensitivity exhibited by the mammalian oocyte, a non-dividing cell, fertility experiments were carried out to determine the effect of dose rate (fractionation, different intensities of continuous radiation). 2) Fractionation markedly reduced damage to fertility; and division of dose into 10-r fractions was more effective in this respect than division into 25-r fractions. Continuously exposed females were even less affected in breeding performance than females which had received fractionated irradiation, and the lower the dose rate the smaller was the deleterious effect on fertility. In all groups, production, in terms of number of females casting litters, remained at maximum level until beginning of a sudden steep decline that ended in sterility. It is for this reason that some experiments of other investigators, who have measured performance only in terms of the first postirradiation litter, have failed to show similar dose-rate effects. 3) The results indicate that some repair of radiation damage to oocytes can occur, and that repair is greater at lower dose rates.

1) Pentobarbital, chlorpromazine, phenaglycodol and meprobamate when injected i.v. into rabbits produced a synchronous (high voltage low frequency) EEG pattern. Physostigmine or d-amphetamine, in suitable doses, antagonized this pattern and evoked an alert (low voltage high frequency) EEG tracing. The dose of amphetamine necessary to antagonize synchronous EEG record generally paralleled the dose of neurosedative administered, especially in the case of chlorpromazine and pentobarbital. In contrast, low doses of physostigmine usually exerted its EEG effect regardless of dose of CNS depressant employed. This indicates that the EEG activation mechanism is more sensitive to cholinergic than to adrenergic drugs and supports the conclusion of several authors that the EEG alerting mechanism is fundamentally cholinergic in rabbits. 2) The evidence indicates that the EEG effects of chlorpromazine are not necessarily related to its peripheral adrenolytic actions.

Mortality is reduced by approximately 50% in mice injected with zymosan (1000 γ) or with thermostable antigens from Proteus morganii, Klebsiella pneumoniae, or Escherichia coli. Soluble antigens from Gram-positive organisms, however, are ineffective. The greatest reduction of mortality is obtained by means of a Typhoid-Paratyphoid vaccine.

Studies with Shigella dysenteriae and Escherichia coli strains revealed enhancement of bacterial susceptibility to normal human serum in vitro in the presence of low concentrations (0.1%) of broth, of glucose, or of certain amino acids known to play a prominent role in bacteria' metabolism. Dipeptides containing these amino acids were more effective than single amino acids. In contrast, metabolic inhibitors, including 5-methyltryptophan, fluorophenylalanine, and non-bactericidal concentrations of chloramphenicol, decreased bacterial susceptibility to serum. Such decrease also was obtained in the presence of, or following exposure to, relatively high concentrations (0.5%) of so-called basic amino acids. The latter effects were reversible by washing when bacteria had been exposed in the absence of nutrients, but they were irreversible following exposure in the presence of nutrients.

Of 37 organic nitrogeneous compounds tested, arginine was the most effective as a primary nitrogen source for 2 strains of Streptococcus bovis. Other compounds supporting growth were glutamine, citrulline, serine, histidine, and glucosamine. The majority of compounds tested, including most amino acids, failed to support growth. Another strain of S. bovis did not grow well on any one organic compound tested as a primary nitrogen source. Simple ammonium compounds, however, were extremely effective in serving as nitrogen source for all 3 strains.‡Growth in media containing ammonium salts was stimulated by acetate, and C¹⁴ labeled acetate was rapidly incorporated. No evidence was found for a mechanism involving degradation of arginine by resting cells or cell free extracts.

Repeated doses of ergotamine and hexamethonium had no effect on increase in plasma non-esterified fatty acid (NEFA) concentration which occurs during fasting. Dibenzyline was similarly without effect on this response. These findings suggest that the stimulus to increased NEFA mobilization during fasting may not be mediated by the adrenergic nervous system.

1. Newborn rabbits were injected with leucocytes obtained from adult rabbits which were prospective donors of lymph node cells. When antigen-incubated lymph node cells from these donors were transferred to young rabbits 1-2 1/2 months later, agglutinins failed to appear, or did so in low titer. This suppression of transferred lymph node cells indicated that tissues of newborn rabbits had reacted to antigens of leucocytes of donor animals. 2. Transfer of lymph node cells incubated in vitro with Shigella-trypsin filtrate to uninjected newborn rabbits (1-11 days of age) resulted in the appearance of anti-Shigella agglutinins.

1. The preparation and metabolism of radiochemically pure aminonucleoside, 6-dimethylamino-9-(3′-amino-3′-deoxy-β-D-ribofuranosyl)-purine, labeled with tritium by the Wilzbach procedure, is described and discussed. Data are presented for the distribution of radioactivity in body fluids and urine as well as in kidney and liver nucleic acids. 2. Study of the chemical nature of tritium activity in an acid-soluble nucleotide fraction, prepared from pooled kidney tissue of rats rendered nephrotic with the tritiated aminonucleoside, indicates that the rat is capable of metabolizing the latter into labeled products utilizable in the biosynthesis of adenosine and cytidine nucleotides. 3. Labeling of these nucleotides, which cannot be accounted for in terms of an exchange mechanism involving the hydrogens of body water, is considered in terms of alternative metabolic pathways.

Serum of rabbits rendered tolerant by a brief schedule of endotoxin administration protects mice against the lethality of homologous or heterologous endotoxin and this protective effect is not attributable to antibody content of the serum.

The loss of potassium attending repeated administration of large doses of nine-alpha-fluorohydrocortisone (FF) to dogs depressed their glomerular filtration rate. In contrast, glomerular function showed a tendency to be augmented in a group of dogs that received supplemental potassium with the steroid. The ability of FF to increase renal tubular reabsorption of sodium was observed under conditions of both potassium depletion and repletion. In the former case sodium retention was enhanced by the reduction in filtration rate but in the latter case no significant retention of sodium occurred, despite its augmented tubular reabsorption.

The virus of herpes simplex was passed serially 20 times in cultures of feline renal cells. Multiplication of the virus was demonstrated when infectivity titers were measured in mice and in tissue culture. The adapted virus induced cytopathogenic changes in cell cultures of feline kidney, primary human amnion, human kidney, and FL cells. Several conditions which appear to influence the type of cytopathogenic change in cell cultures are discussed.

During a study of radiostrontium absorption in rats, a simple method was developed for determining its progress through the intestinal tract. The method should prove equally of value in studies using other radioactive substances. It is particularly useful for estimating the amount of radioisotope available for absorption in different segments of the tract at various times after oral administration.

Methods. Adult female rats of Wistar strain were given 3 μc of Sr⁸⁹Cl₂ in 0.75 ml water by stomach tube. One group had been fasted during 24 hours prior to experiment, while the other group was fed ad lib. Both groups were subsequently given water ad lib., but no food. They were killed in groups of 4 at 1/2, 1, 2, 4, 6, and 24 hours, and the intestinal tracts removed and laid out in a straight line. The radioactive material present in successive 1 cm lengths of the gut was determined using a collimated Geiger counting tube, a counting ratemeter, and graphic recorder. The gut can be counted continuously by placing it on moving paper of a kymograph with extender, fixed so that the paper is horizontal.

Results. Fig. 1 shows distribution of radiostrontium at different time intervals in representative animals from the fasted group. Fig. 2 shows similar data for the fed group. It is apparent that previous feeding increased the emptying of stomach and movement of the dose through duodenum and jejunum, but resulted in longer retention in the ileum. In the fed group, emptying of colon also occurred earlier. In all cases, distribution of the isotope was not uniform, but was concentrated in separated areas.

The method described using a radio-tracer as indicator of its progress in the gut has advantages over the bead methods of Elliott(1). Alvarez(2) and other use of inert indicators because there is no lag or separation of the tracer.

Plasmalogen content in brains of newborn rats is about 2 μmoles/g of wet tissue and represents about 10% of total phospholipid. During growth, and after major increase in brain weight, phospholipid composition shifts to one containing about 20 to 25% plasmalogen (12 μmoles/g). Maximal rate of accumulation of plasmalogen in brain occurs in 20- to 24-day-old rats.

Rate of absorption of Sr⁸⁹ from ligated loops of rat intestine, and rate of passage of Sr⁸⁹ through the intact intestinal tract were measured. Assuming only that absorption rate in ligated loops is reasonably the same as rate of absorption from that segment in the intact rat, the actual effective absorption was estimated. The highest initial rate of absorption occurred in the duodenum with jejunum, ileum, colon and stomach following in decreasing order. Since the radiostrontium passed rapidly through the duodenum and jejunum, the largest actual effective absorption occurred in the ileum (65%), with smaller contributions by the jejunum (17%), colon (8%), duodenum (7%), and stomach (2%). Two factors were found to limit Sr⁸⁹ absorption: (1) Movement of the isotope into gut segments having slower absorption rate, and (2) decreased absorption of Sr⁸⁹ in each gut loop with time.

In humans whose erythrocytes were tagged with Cr⁵¹, fecal excretion of the isotope was used to measure gastrointestinal blood loss. In 38 subjects, ingestion of 3 g of aspirin/day for 3 or 6 days increased fecal blood loss of 3.3 ml/day. Patients who had recently had upper gastrointestinal bleeding and patients who gave a history of gastrointestinal hemorrhage while taking salicylates did not differ in their response from subjects without such histories. Blood loss was smaller in subjects who received antacids every waking hour while taking aspirin than in those who did not receive antacids. Alkaline solution of aspirin produced the same effect as aspirin tablets.

Degenerative changes occur in agar-overlaid tissue culture cells which have been exposed to light in the presence of neutral red. Temporary modification of the ability to form poliovirus plaques may occur in such cells when exposure to light is insufficient to produce discernible changes in the cell sheet.

Large doses of testosterone blocked urinary electrolyte effects of desoxycorticosterone acetate (DCA) in adrenalectomized rats. The steroid reversed the Na/K response to DCA by simultaneously blocking Na retention and K loss. It did not affect Na/K in a manner opposing DCA action when given alone. These observations suggest competitive inhibition as the mechanism of effect for testosterone. Unlike progesterone, it was devoid of DCA-like effects on Na/K given alone, and it gave increasing DCA-blocking effect over a relatively wide range of doses.