Experimental Biology and Medicine

Normal washed human erythrocytes, when incubated in a buffered saline solution (pH 7.5-7.6) with T₄ (3.75 X 10-8 M), demonstrated increased production of 2, 3-DPG and glucose consumption during the first 30 min of incubation. However, lactate production and ATP levels were decreased. The mechanism is probably related to a direct effect of T₄ on the diphosphoglycerate mutase.

Noninfectious complement fixing antigens of Venezuelan equine encephalitis virus were prepared by chromatography of infected tissue culture fluids and subsequent inactivation of the purified product by exposure to ionizing radiation. The product was highly active, stable, and free of anti-complementary activity.

A malabsorption syndrome denoted by jejunal lipodystrophy, steatorrhea, and osteomalacia has been produced in adult cotton top marmosets (Saguinus oedipus) fed a high cholesterol, high coconut oil containing diet. The diet included sufficient amounts of all presently known nutritional essentials for this primate. The lipodystrophy was marked by huge collections of neutral fat in the absorptive cells of the jejunal villi. The fat filled cells apparently created an absorptive block which contributed to the development of steatorrhea and osteomalacia.

Lactating rats were killed after exposure to specific nursing intensities and suckling intervals to examine the effects of these factors on circulating leukocytes, plasma corticosterone, and mammary nucleic acids. Nursing durations of 0.25, 0.5, and 1 hr caused a 3-fold increase in plasma corticosterone when compared to nonnursed levels. In comparison, plasma corticosterone concentrations were similar to nonnursed control values after hours 2, 3, and 4 of nursing. In addition, exteroceptive stimuli associated with nursing provoked increases in plasma corticosterone that were comparable to values obtained from 0.25 or 0.5 hr of nursing. Circulating leukocytes decreased 24% during the first 0.5 hr of nursing with 10 pups; whereas mammary nucleic acid content increased 9%, suggesting an influx of leukocytes into mammary tissue. Histological examination of mammary glands revealed a striking increase in leukocytes within the alveolar lumina of glands within 0.5 hr after nursing. The results from this study indicate that nursing is a dynamic situation which is capable of provoking striking alterations in leukocytic and adrenocortical function.

The acute administration of ethanol causes the accumulation of hepatic triglycerides in male rats and to a greater extent in female rats. Pyrazole, injected prior to ethanol administration, completely prevents accumulation of hepatic triglycerides in male rats, whereas in female rats the same treatment with pyrazole only partially lowers accumulation of hepatic triglycerides. Glucose administration simultaneously with ethanol decreases the concentration of hepatic triglycerides equally in both sexes.

Direct and indirect immunofluorescent (FA) techniques were used to demonstrate membrane fluorescence (MF) in herpes simplex virus (HSV) type 1- and type 2-infected cells. Sorption procedures of antisera with heterologous virus-infected cells, or of HSV-infected cells with heterologous sera, enabled the preparation of fluorescein-conjugated anti-HSV sera specific for each of the two HSV types, as well as of HSV-infected cells containing type-specific membrane antigens. Using the indirect FA technique, it has been possible to demonstrate HSV antibodies in the IgG, IgA, or IgM serum fractions, and to detect HSV IgA antibodies in the genital secretions of women with recent HSV genital infections.

The ability to prepare fluorescein-conjugated sera specific for type 1 and type 2 HSV has permitted more specific typing of HSV isolates or clinical specimens from patients with HSV infections. Similarly, the preparation of HSV-infected cells with type-specific membrane antigens permits more finite serological typing of type 1 and type 2 antibodies in human sera or secretions. In addition, the results obtained provide further support for the working formula for the antigens of HSV: type 1 HSV = AC, type 2 HSV = BC, where A, B, or C may comprise more than one antigen.

The experiments presented clearly showed that the ATP activation of the neutral proteolytic activity is a widespread phenomenon and, therefore, a possible regulatory system for the catabolic phase of protein turnover.

Several investigators (1-3) have shown that in tissue slices the process of protein degradation at neutral pH can be significantly reduced by the uncoupling or inhibition of the oxidative phosphorylation. Complementary studies have indicated that the addition of adenosine-5′-triphosphate (ATP) to preparations of intracellular organelles (4) or to whole tissue homogenates (5, 6) greatly increase the rate of proteolysis, and that this activating effect is mediated by aspartic acid, presumably through the formation of an adenylate derivative (7). This activating effect of ATP was found to be widespread in the organs of the rat (6) and to be able to return to a normal level the reduced proteolytic activity of the liver of rats subjected to an acute stress (8).

All these studies indicate that not only is there a regulatory mechanism for the control of protein degradation at neutral pH, but also that the same mechanism may be involved in the correlation of the rate of protein degradation with the rate of protein synthesis during turnover. This possibility has increased the need to elucidate the nature of the proteolytic enzymes responsible for the neutral proteolytic activity and particularly, the need to identify and characterize the enzyme or enzymes activated by ATP.

This article describes an effort to determine the main types of proteolytic enzymes active in liver homogenates at pH 7.5 and describes their behavior toward ATP during 2 hr of autolysis. These studies have allowed the formulation of an hypothesis, which provides a mechanism for the control of neutral proteolysis and for the interaction of this process with the process of protein synthesis.

Materials and Methods. These studies were carried out with 10% liver homogenates made in 0.22 M phosphate buffer (pH 7.5)-0.2% Triton X-100. The amount of protein in the homogenate was determined according to the method of Lowry et al. (9) using bovine serum albumin as standard.

Studies with synthetic substrates and modified albumin. Qualitative studies were carried out with 50 mM solutions of the following synthetic substrates³: L-leucine amide, L-phenylalanine amide, glycyl-L-phenylalanine, glycyl-L-phenylalanine amide, glycyl-L-phenylalanyl-L-phenylalanine, L-phenylalanyl-L-phenylalanine, L-phenylalanyl-L-tyrosine, L-leucyl-L-tyrosine, L-tyrosyl-L-leucine, N-carbobenzoxy-glycyl-L-tyrosine, N-carbobenzoxy-L-glutamyl-L-tyrosine, and α-N-benzoyl-L-arginine amide. Two hundred microliter of a peptide solution were incubated for 2 hr at 37° with the same volume of a 4% homogenate made by diluting a 10% homogenate with 0.22 M phosphate buffer, pH 7.5. At this time 200 μl of 10% tricholoroacetic acid (TCA) were added, and 50 μl of the supernatant were spotted on a 20 X 10-in. sheet of Whatman No. 1 paper. For identification purposes a similar volume of a 5 mM reference solution of each one of the possible products of hydrolysis of the peptide in question was simultaneously spotted. The constituents in the spots were separated with a solvent mixture containing n-butanol: acetic acid: pyridine: water (15:3:10:12) by descending chromatography after 6 hr of equilibration. The formation of Ninhydrin-positive products other than the products of hydrolysis of the synthetic peptides was monitored by spotting the TCA supernatant of incubation mixtures in which the liver homogenate had been previously heated at 95° or in which the substrate solution had been substituted for phosphate buffer. The products were developed with a Ninhydrin spray and were identified by comparing their rate of migration with the rate of the appropriate reference solutions.

Progesterone injections administered during late pregnancy effectively inhibit lactation in the rat. Subcutaneous doses of 0.5 and 2 mg of progesterone in sesame oil twice daily between days 18 and 23 of gestation produced a greater inhibitory effect than 0.1 mg of progesterone similarly administered, particularly with regard to mortality and weight changes of the young, and the release of milk in nursing tests. The primary inhibition appears to rest on processes of milk ejection rather than milk secretion or the motivation to nurse.

Experiments with liver and kidneys were undertaken in an attempt to demonstrate organ-specific inhibitory factors analogous to epidermal “chalones.” Extracts and alcohol precipitates of extracts of liver and kidney were tested for inhibitory action on DNA synthesis in the named organs. Whole organ extracts and high concentration alcohol precipitates had marked inhibitory action, as high as 50%, on DNA synthesis in both organs. However, an organ-specific effect could not be demonstrated.

Results of determinations of NI and HI antibodies in sera of persons before and after immunization with A2/Aichi/68 and B/Mass/66 influenza viruses are described. A correlation between the magnitude of HI and NI titers was established. Similar results were obtained with inactivated whole virus and virus subunit vaccines. The titer increases resulting from vaccination were, in general, higher for HI than for NI antibodies in accordance with the nonlinear correlation between the two titers.

Coelomic parabiosis between mice differing in θ and/or H2 antigens was produced. Spleens of parabionts contained plaque-forming cells involved in the production of antibodies to the partner's antigens. No significant exchange of PFC between partners was noted. Sera of parabionts contained lytic antibodies to the partner's antigens as well as the animal's own antigens. Data supporting the mutual exchange of antibodies were presented. Evidence was obtained that anti-θ antibodies produced in one partner suppress immune response of the other partner whose circulation they entered.

Cyclic AMP, dibutyryl cyclic AMP, epinephrine, and theophylline inhibited growth of marrow colonies in vitro. The interpretation of this observation is uncertain since various adenine nucleotides had a similar effect and since several substances lowering intracellular cyclic AMP levels were unable to reverse the inhibitory effect of exogenous cyclic AMP.

This study was undertaken to determine interrelationships between early and late embryonic and adult CFU-S. Values for density, volume, seeding efficiency, self-renewal, sensitivity to tritiated thymidine suicide and response in plethorized recipients were determined and graphically displayed as a fingerprint of embryonic and adult CFU-S. The data demonstrate the usefulness of density and velocity separations in characterizing heterogeneous cell populations.

The enzyme neuraminidase destroys the biological activity of erythropoietin even when the hormone is combined with antierythropoietin antibody. It is concluded that the sialic acid moiety is not an important antigenic determinant in the combination of the hormone and its antibodies.

The chemotactically active peptide products from the action of cutaneous collagenase upon purified soluble and native collagen were isolated and partially purified. These leukotactic collagenolysis products were made up of at least eight polypeptides weighing between 30,000 and 1000 daltons and seven of these peptides were leukotactic in vivo at a concentration of 10-¹⁰ M.

Studies on the incidence of bactericidal antibodies to a Hemophilus influenzae, Type B, and an unencapsulated H. influenzae were carried out on sera from 130 healthy individuals, ranging in age from 13 to 78, and from 19 chronic bronchitics. Of the 130 sera from normal people only 2 (1.5%) lacked bactericidal antibodies to the H. influenzae, Type B, whereas 21 (16%) lacked bactericidal antibodies to the unencapsulated H. influenzae. All of the 19 sera from the chronic bronchitics were bactericidal for both Hemophilus isolates. All five sera from agammaglobulinemic patients who were on gamma globulin replacement therapy demonstrated bactericidal activity against the H. influenzae, Type B isolate.

Utilizing the indirect fluorescent antibody technique, no correlation was apparent between the presence or absence of bactericidal antibodies and antibody levels as measured by immunofluorescence.

In vitro renin release from surviving rat kidney slices, expressed in Gold-blatt units/mg of protein or as percentage of the total renin content of the slice, was studied under anaerobic and aerobic conditions and in the presence of arsenite/arsenate blockade. Renin content and renin release from slices from donor rats kept on a sodium deficient diet were three times larger than for slices from rats on a normal diet. The release, expressed as a percentage of the total renin content, was the same. No release occurred under metabolic blockade, and release was much larger under aerobic than anaerobic conditions. Both the quantity and the percentage of the total renin content which was released as well as the Qo₂ decreased with decreasing sodium concentrations in the media. Under some conditions an apparent net synthesis of renin occurred. In vitro renin release is metabolically dependent, related to the quantity of renin stored in the slice, and in some manner influenced by the presence of sodium chloride. In vitro release appears to be much larger than in vivo release and is probably always maximal for a given metabolic state.

The administration of a single dose of D-galactosamine (1.5 g/kg) results in hepatocellular necrosis and fatty liver in the rat. Hepatic triglyceride accumulation is not altered by administration of adenine, ATP, uridine, uridine nucleotides, or uridine di-phosphoglucose.

Protein synthesis in vivo is impaired as early as 4 hr after administration of D-galactosamine. Subsequently the expected post-Triton WR 1339 hypertriglyceridemia is reduced and beta-lipoproteins are decreased by electrophoretic analysis. These data suggest that impaired hepatic lipoprotein release, presumably due to inhibition of synthesis of the protein moiety of lipoprotein, is the mechanism of the D-galactosamine-induced fatty liver.

Rabbits which had been superovulated with FSH and HCG were compared with rabbits ovulated with HCG to assay sperm for capacitation. Inseminations with freshly ejaculated sperm 0 to 13 hr after an ovulating injection of HCG were compared with inseminations with capacitated sperm 13 hr after HCG. Superovulation increased the number of recovered ova from 8 to 22 and depressed percentage of ova recovery, but did not influence fertility significantly. Site of insemination did not influence ova recovery, but uterine inseminations resulted in higher fertility than oviducal inseminations whether they were performed with freshly ejaculated or with capacitated sperm. Inseminations with freshly ejaculated sperm at the time of injection of HCG resulted in more advanced ova development 34 hr after HCG than inseminations with capacitated sperm 13 hr after HCG. The data indicate that sperm may be assayed for capacitation with oviducal or with uterine inseminations into superovulated test rabbits. Lower fertility from oviducal than from uterine inseminations may provide a more discriminative assay.

Treatment with heterologous and isologous antisera against antigenic materials containing spermatozoa prevented adherence of spermatozoa to ova in vitro. These antisera caused agglutination of spermatozoa; however, removal of the agglutinating activity of antisera by papain treatment did not remove the antiadhesive effect. Inhibition of spermatozoal attachment to ova was caused also by oviductal secretions and concentrated media from culture of reproductive tissues from female rabbits isoimmunized with semen.

Serotonin (5 -hydroxytrypta-mine) was found to produce a significant increase in radioiron incorporation in red cells of polycythemic mice. A linear dose-response relationship is reported. Anti-ESF produced a significant inhibition of the erythropoietic effects of 5-HT in polycythemic mice. However, residual erythropoietic activity was still seen even in the presence of antiserum to erythropoietin suggesting a secondary erythropoietic effect which is not erythropoietin dependent. Intrarenal infusions of 5-HT into dogs was found to produce a significant elevation in plasma erythropoietic activity.

Extracts of Burkitt's lymphoma cell cultures which had been previously shown to contain an inhibitor of herpes simplex virus in vitro were used to treat rabbits with ocular herpes infection. Topical instillation of the extract into the eyes of the rabbits cured or suppressed the progression of herpetic keratitis. Histopathologic studies of the eyes of treated and control animals confirmed the inhibition of keratitis and other evidence of herpes-related ocular inflammation.

Antisera to various purified preparations of the lymphocyte stimulant PHA have been prepared in rabbits. These antisera neutralized lymphocyte activation by PHA but were not toxic. Using such antisera, we were not able to detect any differences between the mitogenic components of different purified preparations of PHA, including the separated erythroagglutinating and nonerythroagglutinating fractions. Antisera prepared against purified fractions of PHA also inhibited the mitogenic effects of crude PHA extracts. These antisera also prevented lymphocyte activation by the closely related wax bean hemagglutinin but did not inhibit stimulation by concanavalin A, lentil extract, pokeweed mitogen, staphylococcal filtrate, or antithymocyte serum.

Macrophages exposed to chymotrypsin, trypsin, or pronase failed to ingest aggregated albumin or ferritin in vitro. Treatment with neuraminidase, RNase, albumin, or DFP-chymotrypsin had no apparent effect on macrophage pinocytosis. When cells treated with active chymotrypsin were subsequently maintained in culture, they recovered their ability to ingest protein. This recovery was inhibited by partial inhibition of protein synthesis with cycloheximide. Since chymotrypsin itself is not ingested, the results are interpreted to indicate the removal or inactivation of a surface factor required for pinocytosis that is generated by the cell.

X-Irradiation of microsomal particles prepared from pig thyroid homogenates by conventional methods with 100, 000 rads before incubation with radioactive leucine reduced protein synthesis as much as 60% of that of a nonirradiated control. The effect of the radiation was most pronounced at early incubation times (15-30 min) and at low concentrations of the supernatant fraction. β-Mercaptoethanol, which was necessary for protein synthesis, reduced the amount of X-ray damage to protein synthesis considerably when added before X-irradiation, but had minimal effect when added after X-irradiation.

Groups of male subjects in varying states of general physical condition were exercised on a bicycle ergometer at three work loads. Pre- and postexercise concentrations of serum pyruvate were estimated by a simple, rapid automated technique which utilizes LDH in the serum specimens. Postexercise concentrations of pyruvate increased with increasing work load. After a test exercise of 720 kpm/min for 30 min, serum pyruvate concentrations correlated significantly with several indices of general physical condition: maximal oxygen uptake, time to run 3 miles, and time to run 1000 m. The techniques described may provide the basis for a simple biochemical test of cardiovascular fitness.

Hyperbaric oxygen decreases nitrite-induced mortality and methemoglobinemia. The action of HPO in reducing methemoglobin levels is brought about by hindering the oxidation of hemoglobin by nitrite. HPO had no effect on the erythrocyte reductase systems in reducing methemoglobin.

Serum FSH was measured by radioimmunoassay in developing normal and androgenized male and female rats. Immature, 30-day-old, as well as mature, 60-day-old males had higher concentrations of this gonadotropin than females. Serum FSH declined in androgenized females (50 μg of testosterone propionate, fifth day of age) between days 20 and 30. The level remained low at 60 days but was at about the level of normal females at 1 year. The results support the conclusion that androgenization in the female does not produce the male pattern of high serum FSH.

Leukocytes incubated in vitro in plasma or serum, but not in saline, develop a potent procoagulant activity like tissue factor. This activity is not present in freshly collected leukocytes from normal individuals. The generation of activity is stimulated by gram-negative endotoxin.

The radioisotope precipitation test was performed with ¹⁴C and ⁵¹Cr labeled vaccinia antigens after primary smallpox vaccination. The test gave positive results with the sera of successfully vaccinated individuals for a longer time than the neutralization test.

Red blood cell survival was prolonged in hypophysectomized rats. While the rate of random hemolysis was decreased in some hypophysectomized hosts, in all directly injected and cross-transfused hypophysectomized rat hosts, there was a significant prolongation of the phase of senescent death. In contrast, RBC from hypophysectomized donors survived normally in normal hosts. These experiments are further evidence of a relationship between RBC aging and metabolic rate, and suggest an intimate involvement with the calorigenic hormones.

The mechanisms of zinc absorption and the effect of EDTA (ethylenediaminetetraacetic acid) on zinc absorption were studied at the intestinal digesta, intestinal mucosa, and blood plasma levels. Dialytic and gel nitration studies showed complexing of ⁶⁵Zn with free amino acids or EDTA-¹⁴C in the intestinal digesta. The EDTA-¹⁴C:⁶⁵Zn complex was bound to a zinc binding protein in the intestinal mucosa (mol wt 14,000-18,000) and to a plasma protein (mol wt 65,000-70,000), that had a greater affinity for zinc than the mucosal protein. The ratios obtained for the cpm EDTA-¹⁴C:cpm ⁶⁵Zn in the main peaks eluted from gel filtration columns (1.10 in intestinal digesta preparation, 1.26 in mucosa soluble fractions, and 1.26 in blood plasma) indicated strong binding of the EDTA-zinc complex. When EDTA was not present in the diet, ⁶⁵Zn complexed with amino acids in the intestinal digesta and with the same mucosal and blood proteins.

A mild diabetic-like state was induced with streptozotocin in rats prior to breeding and fetal and placental composition was examined on day 21 of pregnancy. Fetuses of untreated diabetic rats were larger than those of controls and had significantly more fat, less water, and more DNA. These alterations in fetal size and composition were prevented by maternal insulin treatment during pregnancy.

The effects of sex difference and of gonadectomy on the uptake of 7, 12-dimethylbenz[a]anthracene-³H and benz-[a]anthracene-³H by rat liver slices, hepatic DNA; and histones were studied. Liver slices from the intact male rat incorporated 7, 12 -dimethylbenz [a] anthracene-³H at a higher rate than benz[a]anthracene-³H. The uptake of 7,12-dimethylbenz[a]anthracene-³H by intact isolated liver nuclei increased after gonadectomy of the male rat, whereas the uptake of benz[a]anthracene-³H increased in liver nuclei from female rats after ovariectomy. Relatively small amounts of radioactive hydrocarbons were bound to DNA under the in vitro experimental conditions. Liver histones were found to bind radioactivity after incubation of liver slices with 7, 12-dimethylbenz[a]anthracene-³H and benz-[a]anthracene-³H. Gonadectomy led to an increase in the extent of binding of benz-[a]anthracene-³H to liver histones from both male and female rats. Uptake of 7, 12 -dimethylbenz [a] anthracene-³H by histones was increased only in liver of ovariectomized female rats.

The progesterone concentration in peripheral plasma of sham-operated guinea pigs increased significantly during the estrous cycle from a nondetectable concentration at estrus to a maximum concentration on day 10, followed by a rapid decline. In the presence of an IUFB in both uterine horns, the progesterone concentration did not increase after day 4 of the cycle. Over the next 8 days, a slow decline in concentration occurred. The length of the estrous cycle was significantly shorter in animals having IUFB than in sham-operated animals.

By progressively increasing the duration of an electrical stimulus, two barosensory and one chemosensory fiber groups have been illustrated in the CSN of the dog. Both barosensory fiber groups are more easily excited than the chemosensory fibers. In the cat, similar groups of fibers have been reported, but the chemosensory fibers are of intermediate excitability. All the fibers appear to exert their action on the sympathetic adrenergic system, since the response is blocked by bretylium, but not by atropine.

The effect on survival of mice innoculated with a splenotropic reticulum cell sarcoma was tested in splenectomized and mock-operated syngeneic mice. The splenectomized animals survived longer. Evidence was presented that this is probably due to rapid destruction of circulating cells, and that the tumor cells are protected by being sequestered in the spleen.

The steady state rate of gluconeogenesis in rat liver slices incubated in U-¹⁴C-alanine was linearly related to the extracellular alanine concentration over a range of 1-9 mg of alanine/100 ml. The rate of oxidation of alanine to CO₂ was also linearly related to the extracellular alanine concentration and the ratio of alanine oxidized to CO₂ to the alanine converted to glucose was constant with changes in the alanine concentration.

Appendix: CO₂ trapping efficiency correction. Since the quantity of labeled carbon dioxide found in the centerwell of closed incubation flasks following a test injection (step forcing) of Na₂ ¹⁴CO₃ increased exponentially with time, the trapping mechanism was a first order process (15). During incubation the liver slice produced carbon dioxide continuously, therefore, under experimental conditions the first order carbon dioxide trapping process was subjected to a ramp, rather than a step forcing. The equation, predicting the fraction of released ¹⁴CO₂ that would be trapped in the centerwell, is that of a ramp response of a first order process:

F = fraction of ¹⁴CO₂ recovered = 1/t[t-r(l-e^-t/r)], where: r is the time constant of 79 min determined from the test experiment; t is the incubation period of 60 min; e is the base of natural logarithms. The equation predicts that at the end of a 60-min incubation period only 30% of the total carbon dioxide released during that time must have been trapped in the centerwell:

F = 1/60 [60-79 (1-e ^-60/79)] =0.30. Therefore, the ¹⁴CO₂ data obtained must be corrected by multiplying by 3.33.

The fact that the two strains IS and BA in AL showed such a large difference in sensitivity to the same NDV antibody makes it seem likely that the disappearance of cell-associated viral hemagglutinins is due to a reaction at the cell membrane, not inside the cell. Since the HAI titer was approximately the same for IS as for BA, entry of antibody into the cell should cause equal inhibition. In the antibody-sensitive cell-virus systems, virus budding through the membrane probably is removed as fast as it is formed and it follows that maturation is exclusively at a site accessible to antibody. With virus strains where large amounts of cell associated hemagglutinins are formed, even in the presence of extracellular antibody, intracellular HA production seems likely. Evidence for association of influenza virus with internal membranes of the cell has been presented by Holland and Kiehn (8). Also it is of interest that ascites lymphoma cells infected with influenza are more resistant to antiviral immune cytolysis, early or late in infection, than are those infected with Sendai or NDV (9). Part of the increased resistance in CE might be attributed to small clumps of cells which were difficult to eliminate from the suspensions. For example, NDV-IS, highly sensitive in AL, was resistant in CE. However, Sendai was inhibited by about the same antibody concentration in either cell species and WSN was apparently more resistant in AL than in CE.

Resistance was overcome by increasing the antibody concentration. In CE, the strains NDV-IS, NDV-BA, and influenza, became sensitive to high antibody levels. An explanation of this phenomenon must await further investigation but concentration dependence suggests forced diffusion across a barrier. The alternative is entrance of small amounts of antibody into the cell through pinocytosis.

The action of antibody at 24 hr in causing disappearance of cell-associated HA does not seem related to cell disruption by the virus, at least those cases where dye exclusion remained effective or the inhibition was reversible.

Renal excretory patterns of sodium, potassium and water were studied in unoperated and hypophysectomized rats as functions of pretreatment with d-aldosterone and actinomycin D. Observations included:

1. A GFR-depressing function of aldosterone in the sodium-depleted hypophysectomized rat.

2. A glucocorticoid-like response to aldosterone in the sodium-depleted hypophysectomized rat.

3. A GFR-stimulating function of actinomycin D, which was reversible by a priori but not by a posteriori administration of aldosterone.

The sphingolipidoses are hereditary disorders, secondary to enzyme defects, in which lipids accumulate in brain and other organs. The present studies indicate that the abnormal lipid cytosomes are derived from the smooth portion of the endoplasmic reticulum, and that the mitochondria appear to be the organelle that is primarily associated with the enzyme defect. Preliminary studies of mitochondrial fractions from normal and Tay-Sachs disease brains showed that the mitochondrial fraction of normal brains contained 28.6% N-acetyl hexosaminidase A activity, while it was absent in that from Tay-Sachs disease brain.

Plasma, frozen and thawed lymphocytes, and purified Australia antigen fractions from various patients suffering from viral hepatitis were tested for an effect on PHA-stimulated lymphocytes isolated from normal individuals. The addition of these materials to lymphocyte cultures produced no significant change from that of PHA alone or PHA plus control materials from normal (nonhepatitis) patients.

Using stimulated rat peritoneal macrophages, a specific antimacrophage antibody has been prepared in rabbits. Immunofluorescent studies reveal that the antigen(s) is present in free, and absent in fixed, phagocytic cells in the rat. The antigen(s) has also been identified in free macrophages of guinea pigs, mice, and rabbits. Fixed macrophages in the same animal species are consistently negative. The results indicate that free macrophages are antigenically different from fixed phagocytic cells, and that the antigen(s) lacks species specificity.

Enzymatic (1) and gas chromatographic technics (2) for the quantification of bile acids developed in the past 10 years, have resulted in the publication of vast data on the composition of bile. Almost without exception, bile for these studies has been obtained under nonphysiologic conditions, e.g., single sampling of duodenal bile from fasting humans, collections from tubes in the common duct proximal to the sphincter of Oddi, or from chronic fistula preparations. However, interruption of the enterohepatic circulation changes the composition of bile (1), concentrations of the constituents of bile vary with sampling technics (2), and the volume, total solutes and percentage concentration of constituents vary throughout a 24-hr period (3). We have modified technics, first used by Dragstedt et al. (4) and later by Preshaw and Grossman (5), in studies of pancreatic secretion to develop a canine preparation which allows repeated sampling of bile from an intact enterohepatic circulation.

Figure 1 is a diagrammatical representation of the method we have used to construct a duodenal pouch containing the intact sphincter of Oddi, draining through a Gregory cannula (6) into the duodenum opposite a Thomas cannula (7) in the reconstituted duodenum and extending through the abdominal wall.

Surgical technic. The upper abdomen is opened through a right paramedian incision. Vessels between the duodenum and the pancreas are carefully ligated with fine silk to mobilize a segment of duodenum 1.5 cm distal and proximal to the point of entry of the common duct. After ligation of the accessory pancreatic duct the duodenum is transected at points 1 cm and 3 cm proximal to the main pancreatic duct. The Gregory cannula is passed through the antemesenteric wall of the duodenal segment and proximal and distal ends of the pouch are closed in 2 layers.

The GVH disease in 1C3F₁ hybrid mice following 500 R X-irradiation and injection of normal parent strain spleen cells is mild in terms of mortality, signs, and tissue pathology. Presensitization of the donor with spleen cells of the reciprocal parent strain, however, results in a highly lethal GVH disease. The timing of presensitization before donor use is an important factor in the severity of the resulting GVH disease. Presensitization of the donor seems to require more than 3 days, is near maximum effectiveness at 7 days, persists at this level for many months, and gradually falls off through the life of the donor. After 24 months, sensitized donors still are more effective in producing GVH disease than controls of comparable age.

The biological activity of human urinary erythropoietin is completely neutralized when combined with serum from rabbits immunized with erythropoietin. Acidification to pH 5.0, followed by heating in a boiling water bath for 5 min, recovers essentially all the erythropoietic biological activity from such a combination. These data further support the immunologic nature of the reaction between erythropoietin and the neutralizing serum.

Relationship between hemolytic plaque formation and repopulation of spleen and thymus after X-irradiation was studied by using two substrains of CBA mice. The CBA/H and CBA/St mice generated almost equal numbers of hemolytic plaques in the spleen at 7 days after intraperitoneal injection of sheep red blood cells (SRBC). The CBA/St mice that received 400 R of wholebody irradiation, or 600 R of irradiation and injection of 5 X 10⁶ marrow cells, generated smaller numbers of plaques at days 22 and 29 than did CBA/H mice that received the same treatment. Bone marrows, thymuses, and spleens of sham-thymectomized and thymectomized CBA/H and CBA/St mice were almost exclusively repopulated with donortype cells by day 22.

Donor-type cells in the spleens of CBA/H mice had been “instructed” in the thymus, were seeded to the spleen, and responded to antigens, but the spleens of CBA/St mice which were also repopulated with donor marrow cells, had very small numbers of these instructed cells that were capable of responding to antigens.

These data show that the pulmonary vasopressor response to perfusates of acidified plasma or whole blood fractions suspended in LMD is directly related to the platelet concentration of the perfusate. The pressor response is not directly related to local vascular effects of lowered perfusate pH within the range 7.40-6.35. This response appears to be mediated by the release of vasoactive substances from platelets at the site of acid infusion. It is suggested that the effect of an increase in hydrogen ion concentration on platelets causes direct physi-cochemical alterations which activate and release pulmonary vasopressor materials.

Aerosols containing ⁹⁵Zr and ⁹⁵Nb at near-equilibrium were produced at 100, 250, 600, and 1100° to give particles of four different chemical compositions. When inhaled by mice, a differential metabolism of the radionuclides was sometimes observed, dependent upon the temperature of formation. At the two highest temperatures, the deposited particles were retained tenaciously in the lung and maintained the starting ⁹⁵Zr-⁹⁵Nb ratio of ~1.9. At 100°, the radionuclides were translocated from lung to skeleton with a ratio of ~1.3, indicative of an excess burden of ⁹⁵Zr. At 250° the pattern was similar but less pronounced. The differential patterns of retention resulted in significant differences in radiation dose pattern; at the lowest temperature the highest radiation doses were received by skeleton, lung, and liver, while at the higher temperatures the radiation dose was delivered almost exclusively to lung.

Carrageenan, dextran, phytohemagglutinin-P, alpha-amylase, acid phosphatase, polycytidylic acid, polyadenylic acid, polygalacturonic acid, poly-L-proline, poly-L-glutamic acid and others promote red cell sedimentation rates (ESR) when added to washed red cells. Rabbit antisera to red cells and serum promote the ESR when added to either whole blood or washed red cells. All of these various acceleratory responses, with the exception of those induced by enzymes, are inhibited by nonsteroidal anti-inflammatory drugs (NAIFD). Many other pharmacologic classes, with the exception of ethacrynic acid, probenecid, sodium cromoglycate and chlorothiazide, fail to display similar inhibitory effects. The elevated ESR is inhibited when drugs are added before, during, or after the addition of dextran. The ESR is inhibited by 2, 4-dinitrophenol, but not other metabolic inhibitors. Heavy metal ions and anerobiosis have no effect on dextran-induced ESR.

Agents accelerating the ESR induce acute inflammatory responses when inoculated locally into the hindpaws of rats.

Drugs whose inhibitory effects are less readily removed from dextran-treated red cells by repeated saline washings, e.g., indomethacin, have longer durations of action when tested in acute inflammatory reactions of intact animals against salicylic acid whose effects are removed more rapidly.

Mechanisms of action of accelerators or inhibitors of the ESR are unknown. The NAIFD are, for the most part, acidic. They may adhere to available positive charges on damaged or altered cell membranes and prevent their tendency to “stick,” adhere, aggregate, clump or pull together.

Although red cell aggregation is among the many convenient models for the assessment in vitro of well-known NAIFD, it fails, like many other isolated methods (3-5), to have predictive value for unknown agents. Many undisclosed chemical agents have been discovered to be extremely potent in the isolated ESR system. When given subsequently at varying dosages and routes of administration to animals, however, they fail (in our hands, at least) to display “anti-inflammatory” activity.

The dextran- and carrageenan-induced ESR appears to be a rather specific method for screening NAIFD discovered by other methods and for the study of their probable mechanisms of action.

Human plasma deoxyribonuclease I is inhibited by many naturally occurring RNA samples and also by ribonucleic acid-like polynucleotides, viz., polyuridylic acid (5¹) or polyadenylic acid (5¹), while polyglutamic acid, another polyanion, was found inactive. The significance of this partial immobilization of DNase I by RNA is not clear.

U-29,409 (2,3-dihydro- (1-naphthyl)-4-(1H)-quinazolinone), a male antispermatogenic drug, exerts anti-arthritic effects in rats similar to those elicited by Cytoxan. However, it is less effective than the latter. Laboratory screening tests fail to show that U-29,409 has acute anti-inflammatory effects when given as a single dose or for 1 week prior to performing acute inflammatory reactions. In combination with Cytoxan, U-29,409 appears to exert a synergistic inhibitory effect on developing arthritis. Neither Cytoxan nor U-29,409 appears to exert significant inhibitory effects in rats with established disease. Both drugs, unlike nonsteroidal anti-inflammatory drugs, fail to display inhibitory effects in yeast-induced fever of rats.

Purified Australia antigen [Au(1)], conjugated with ¹²⁵I, was reacted with and quantitatively precipitated by human anti-Au(1) in the presence of heterologous antihuman IgG and “dilute” ammonium sulfate. The sensitivity of the devised radioim-munoprecipitation assay procedure proved to be up to 1000 times more sensitive for the detection of antigen and up to 20,000 times more sensitive for the detection of antibody when compared with either immunodiffusion or complement fixation assays. This method is extremely valuable not only in the quantitative detection of antigen and antibody but also in the delineation of naturally occurring antigen-antibody complexes.

The possible role of the biliary system in the pathogenesis of hypercholesterolemia observed in rats following bilateral hypothalamic injury involving the ventral medial nuclei, the fornices, and the medial portions of the lateral hypothalamus was investigated. The hypercholesterolemia was found not to be mediated by any abnormality in the flow of bile.

Human peripheral leukocytes from 15 subjects were separated into two fractions. One fraction contained most of the basophils and very few eosinophils. The other fraction contained all or almost all of the eosinophils but few basophils. The neutrophils were distributed in a random fashion between the two fractions.

The histamine in each fraction was measured and found to correlate only with the basophil leukocytes. Variations of eosinophil distribution had no effect on histamine levels. Lysis of neutrophils produced only minimal amounts of histamine. These results demonstrate that histamine found in human peripheral leukocytes is localized to the basophil leukocytes.

The vascular supply of the grossly viable portions of grafts of the MTG-B mouse mammary carcinoma was measured with ⁵⁹Fe-labeled red cells. The relative vascular space was constant, estimated as 1.5 X 10^-2 μl/mg in tumors yielding 50 to 1900 mg of “cleaned” tissue from which necrotic, cystic, or hemorrhagic areas had been removed. We have previously reported that the potential growth rate in the grossly viable portions of MTG-B tumors is also constant from the time tumors are small until near death of the host animals. It is postulated that an equilibrium between the viable portions of such tumors and their vascular supply is established when tumors are small. This equilibrium persists well into the period when tumors are large; and necrosis, hemorrhage, and cysts are common. It is thus further suggested that such degenerative changes become marked when the absolute number of new tumor cells born exceeds the capacity for new vascular tissue formation. However, the functional vascular supply of the regions from which most, if not all, new cells arise in large tumors of this type is apparently adequate to support the same metabolic and proliferative rates as in small tumors, despite the occurrence of widespread degeneration in adjacent areas. Insofar as the response to therapy depends on the relative vascular supply and proliferative rate, the sensitivity per unit viable tissue would be expected to be similar in large and small tumors of this type.

When germfree mice are fed a diet supplemented with cholic acid, the turnover of intestinal mucosal cells is reduced from 4.44 to 2.12 days, which is the turnover time in conventional mice. It is concluded that cholic acid, which is produced in conventional mice by hydrolyzing enzymes of bacterial origin, has a regulatory effect on intestinal mucosal cell turnover.

Precapillary sphincters are currently thought by most investigators to completely close or open in response to intrinsic and extrinsic stimuli. In vivo experiments, using the rat mesentery and a high magnification (up to X6500) image-splitting television microscope recording system, were designed to test this hypothesis. The results clearly, and quantitatively, demonstrate that precapillary sphincters, at least in rat mesentery, do not respond (i.e., constrict or dilate) to vasoactive substances in an all-or-none manner but in a dose-dependent, graded fashion.

In the presence of porcine tissue plasminogen activator, fibrinolysis is greatly enhanced in comparison with fibrinogenolysis. In contrast, other activators (UK, SK-activator) and plasmin showed no, or moderate, differences between lysis of fibrinogen and fibrin. The enhanced tissue activator effect is related to the activation of plasminogen. The observations add a specific mechanism of enhancement of fibrinolysis by tissue activator to that caused by the influence of inhibitors. The process may help to enhance localized fibrinolysis following tissue injury without affecting fibrinogen.

Phenobarbital causes no significant change in either serum bilirubin concentration or the rate of bilirubin turnover in homozygous Gunn rats. There is only minimal enhancement of bilirubin production from nonhemoglobin sources in the liver, in contrast to the large changes reported for normal animals. These studies indicate that phenobarbital does not stimulate the alternate pathways by which animals unable to conjugate bilirubin excrete bile pigment. Moreover, they raise intriguing questions concerning the production and excretion of the hepatic bilirubin fraction in the Gunn rat.

Complement depletion of rats by cobra venom factor had only minor and variable effects on the hyperacute form of EAE. It appears likely that the exudation of polymorphonuclear leukocytes and massive amounts of fibrin in this condition is not dependent on complement. However, it is not possible to eliminate all traces of complement with cobra factor. Therefore, it is theoretically possible that small amounts of complement, too little to be detected by hemolytic activity or gel diffusion, might accumulate in the EAE lesions during the 1- to 3-day period in which they evolved. However, inhibition of the neutrophil accumulation of nephrotoxic serum glomerulitis attested to the efficacy of the cobra factor. This internal control provided evidence against the binding of C3 and terminal components, at least for the last 3 hr of development of EAE lesions. As C3 levels fall within 4 hr of the initial injection, it is most probable that complement depletion spanned the time during which EAE lesions developed.

Passive transfer of EAE can be accomplished in a single day. The rapidity of lesion development makes it highly likely that there was effective depletion of complement during the entire disease. Nevertheless, complement depletion of recipients had little or no effect on the outcome, either in conventional passive EAE in which mononuclear cells predominate or in the new, neutrophilic variety in which polymorphonuclear leukocytes are prominent.

Direct intraamniotic injections (ia) of the glucocorticoid dexamethasone produced low fetal mortality and a high incidence of cleft palate in rat fetuses. Administration of 120 μg/30 μl of the hormone 12 to 24 hr before palate closure (day 15 of pregnancy) resulted in 100% retarded skeletal development and 91% cleft palate in term fetuses. Subcutaneous maternal injections of 0.4 mg daily for 3 days prior to palate closure resulted in a similar incidence of palatal defects. Intraamniotic injections of dexamethasone and the nicotinamide antimetabolite 6-aminonicotinamide (6-AN) failed to reduce or prevent the development of congenital malformations, potentiating the production of full cleft palate.

Treatment of rat kidney cortex slices with phospholipase C at high concentrations leads to increased total tissue water, inulin space, reduced _a-aminoisobutyric acid (AIB) uptake as well as increased efflux. These observations are concluded to represent alterations in permeability characteristics of the slices. Treatment with doses of phospholipase C that cause no alteration in distribution of water or sodium is associated with reduction in potassium content of slices and reduction in AIB accumulation which is not associated with altered efflux. The relationships of the role of phospholipids to renal tubule transport systems for monovalent cations and amino acids are discussed.

A study of the hexose hexosamine, sialic acid, uronic acid, glycogen, and collagen fractions has been made on hypertrophic scars, nonhypertrophic scars, and normal skin. Hexose, hexosamine, sialic acid, uronic acid, and glycogen levels were significantly and consistently elevated in hypertrophic scars. These same components were slightly, but significantly, higher in the nonhypertrophic scars. Salt-soluble collagen of hypertrophic scars was significantly elevated, citrate-soluble and insoluble collagen fractions were not different in the three groups. From these data it is concluded that hypertrophic scars contain more glycoprotein and mucopolysaccharide and are more active metabolically.

Different forms of interferon therapy, (i) exogenous interferon; (ii) interferon inducers: poly I:C (homopolymer pair of polyriboinosinic acid and polyribocytidylic acid) and COAM (chlorite-oxidized oxyamylose); (iii) peritoneal macrophages treated with interferon; and (iv) peritoneal macrophages treated with interferon inducers (poly I:C and COAM), have been compared in their capacity to protect mice from a lethal intranasal challenge with vesicular stomatitis virus (VSV). All treatment regimens were given by the same route (intraperitoneally) at the same day (5 days after infection, at the time that the clinical symptoms appear) and at comparable doses. All treatment regimens afforded a partial protection. This protection was greatest with poly I:C-treated macrophages. There was no essential difference in the protection conferred by exogenous interferon, COAM, interferon-treated and COAM-treated macrophages. However, poly I:C was therapeutically less effective than poly I:C-treated macrophages and also less active than might have been expected from the titers of interferon that it induced endogenously.

Transformation of the established cell lines of Rauscher leukemia virus (RLV)-infected and uninfected rat embryo cells by polyoma virus is reported. Transformation was evidenced by morphological changes and presence of polyoma “T” antigens by means of complement-fixation test and fluorescent antibody staining technique. Increased transformation efficiency of polyoma virus was observed in rat embryo cells infected with RLV.

Heat-killed M. tuberculosum injected intraperitoneally in rats produced peritonitis as a primary lesion and polyarthritis as a remote secondary lesion. Rats given 0.25 mg of the adjuvant developed paw swelling 21 to 28 days after the injection. Those given 1 mg developed severe swelling in 12 to 15 days. Parameters investigated were peritoneal leukocyte population and cell count, plasma β-glucuronidase, inflammation units, paw volumes, and body weights.

A sharp increase in peritoneal leukocyte counts and percentage lymphocytes was observed prior to swelling in the hind limbs. Plasma inflammation units and β-glucuronidase increased sharply at about the time paw volume increased. Bone damage was less severe than in animals injected in foot or tail.

Comparative studies of protection of mice against infection with MM virus and levels of interferon induced by graded amounts of Escherichia coli endotoxin, statolon, maleic acid-divinyl ether copolymer, and polyinosinic-polycytidylic acid complex have been carried out. In every instance, both protection and interferon response were dose-dependent, but interferon titers did not correlate with protection. The degree of protection could not be correlated to any specified amount of interferon. Possible explanation of these results would include: (i) protective potency of interferon may differ with the type of inducer, (ii) different interferons are utilized for protection in different ways, or (iii) the protective effect of inducers used may be unrelated to interferon.

A mixture of an erythropoiesis stimulating factor (ESF) and an ESF-generating factor (EGF), isolated from the urine of a patient with paroxysmal nocturnal hemoglobinuria (PNH), was separated by selective membrane filtration. The optimum pH for activity of the EGF fraction from the urine of the PNH patient was about 7.4, the same as that previously found for the EGF from the urine of a patient with an anemia secondary to multiple myeloma (MM). The urine from the PNH patient contained an ESF that appeared larger than EGF, whereas the urine from the MM patient did not; the urine from both patients, however, contained an ESF that appeared smaller than EGF as previously noted.

Hepatic mitochondrial functional activities, as determined by ADP/O, respiratory control ratio, and phosphorylation rate with glutamate as substrate, were augmented in male Fischer rats between 22-32 weeks following ingestion of the carcinogen, 2-N-fluorenylacetamide (2-FAA). Histologically, the livers from the carcinogen-fed group all contained hyperplastic nodules characteristic of areas of regeneration; and hepatomas were found in one animal in each of the three periods studied. Previously, increased mitochondrial activities of this type were reported for animals treated with the hepatotoxins, hydrazine and ethanol, as well as during regeneration following partial hepatectomy. This stage of hepatocarcinogenesis following the long-term feeding of 2-FAA represents an additional instance wherein these mitochondrial functional activities were elevated coincident with significant liver damage. In this case, enhanced mitochondrial activities might be an expression of the presence of the hyperplastic nodules. Whether these increases were part of the pathological chain of events during carcinogenesis or exposed a phenomenon common to both carcinogenesis and regeneration could not be ascertained from these experiments.

Noncontacted factor XII-deficient human plasma was mixed with Celite, the protein adsorbed to the Celite was eluted with a concentrated salt solution, and then chromatographed on QAE-Sephadex A-50, followed by Sephadex G-200.

The first (excluded) peak obtained by anion exchange chromatography contained substances with physicochemical and biological properties indistinguishable from plasma kallikrein and “contact activation product.” Material obtained with the excluded peak enhanced vascular permeability, hydrolyzed arginine ester, liberated kinin from both fresh and heated plasma, corrected the prolonged clotting time of plasmas congenitally deficient in factors XII or XI, artificially depleted of XI, but did not enhance the clotting of plasmas deficient in factors IX, VIII, and X. On gel filtration the molecular weight of kallikrein was estimated to be about 90, 000 and that of the clot-promoting material was about 170,000. The latter was confirmed by sucrose density gradient ultracentrifugation.

We conclude that prekallikrein and factor XI can be activated directly by massive exposure to an active surface. This activation does not seem to require factor XII. It is further concluded that the clot-promoting activity of contact activation product is due to factor XI. Finally, we could obtain no prekallikrein-activating material from factor XII-deficient plasma.

A differential assay is described for testing compounds with hamsters which allows one to separate anti-implantation and antimating behavior effects from influences exerted at the time of capacitation and fertilization. Such tests with chlormadinone and megestrol acetate emphasize the effects of these compounds on the later phenomenon. Demonstration that the first phase of rabbit sperm capacitation occurs readily in 12 hr in the hamster uterus indicates the usefulness of this animal for cross-species studies of capacitation.

Saline solutions of about 1 μg of each of seven of the eight collagen peptides released by the action of cutaneous collagenase upon soluble collagen were capable of affecting the aggregation of platelets. Thus, at least two important cellular responses to injury (i.e., PMN infiltration and platelet aggregation) can be demonstrated to be produced by the products of collagenolysis.

These findings on platelet aggregation reinforce the relative importance of collagenolysis product peptides in the overall process of inflammation.

A sensitive and reliable system to culture mouse lymphocytes stimulated to transform by phytohemagglutinin is described. This system permits the convenient use of a relatively rapid modified “acidinsoluble” method to assay nucleoside incorporation. Under the conditions described, stimulated splenic and peripheral lymphocytes demonstrated elevated RNA synthesis 18-24 hr prior to maximum DNA synthesis. Nucleoside incorporation of stimulated peripheral lymphocytes exceeded that of splenic cells and occurred earlier. The degree of transformation was dependent on cell density, phytohemagglutinin-P concentration, and serum level.

Rate of locomotion of human PMN is markedly enhanced by temperature between 32 and 42° (Q ₁₀ = 5.2). This rate is not significantly altered between pH 6.5 and 7.5. Beyond pH 7.6 there is a significant and rapid inhibition of motility.

Ethanol, 1 and 2%, inhibited the synthesis of aldosterone in beef adrenal tissue in vitro. The inhibition appears to occur between the conversion of corticosterone to aldosterone.

DL-a-Methyl aspartic acid and DL-threo-β-methyl aspartic acid have been shown to inhibit, in vitro, both urea synthesis and the conversion of L-citrulline-6-¹⁴C to ¹⁴C-urea. In the presence of glutamine as nitrogenous substrate, the conversion of citrulline to urea is inhibited while urea synthesis is not inhibited by these methyl aspartates. No inhibition of urea biosynthesis or of conversion of L-citrulline-6-¹⁴C to ¹⁴C-urea could be demonstrated in the isolated perfused rat liver using these inhibitors.

The effect of nicotine administration on regional myocardial blood flow was examined in normal hearts and after partial coronary artery occlusion. Under normal circumstances as well as after infusion of nicotine in normal hearts the subendocardial portion of the myocardium had a higher capillary flow than the subepicardial fraction. Partial coronary artery constriction alone did not alter this relationship; however, when it was followed by infusion of nicotine, a significant reduction in capillary flow in the inner portion of the myocardium compared to the outer part was observed.

Serotonin (10-500 μg) was infused into the pulmonary circulation of anesthetized, spontaneously breathing dogs and the effect on pulmonary hemodynamics [pulmonary artery pressure (P _pa), left atrial pressure (P _la), the pulmonary artery-left atrial pressure gradient (P _pa-P _la)] and pulmonary mechanics [pulmonary resistance (R _L) and compliance (C _L)] was determined. Pulmonary arterial hypertension, manifested by an increase in P _pa and P _pa-P_la with no change in P _la, was caused by small doses of serotonin (30-100 μg) which had no measurable effect on R _l and C _l. These findings are consistent with the hypothesis that serotonin causes pulmonary arterial hypertension as a result of active pulmonary vasoconstriction rather than as a result of changes in the mechanical properties of the lungs.

The incorporation of ³H-labeled methyl groups into DNA from normal and leukemic human leukocytes has been studied. The sp act of DNA were higher in leukocyte samples of acute lymphocytic leukemia (ALL) and acute myelocytic leukemic (AML) than in those of normal and chronic lymphocytic leukemia (CLL). The highest radioactivity was found in 5-methylcytosine with moderate radioactivity in thymine and low levels in adenine, 7-methylguanine, and cytosine.

The effect of zinc sulfate on collagen biosynthesis and fibroblast proliferation has been studied in a tissue culture model system using human skin fibroblasts. Addition of zinc sulfate to newly established (low density) cultures in concentrations of 10^-4 to 10^-8 M results in concentration-dependent cytotoxicity with inhibition of collagen biosynthesis, as well as cellular proliferation. Zinc sulfate at 10^-4 to 10^-7 M added to confluent (high density) cultures does not influence cellular proliferation, but results in selective inhibition of collagen biosynthesis at the 10^-4 M concentration. Lower concentrations of zinc sulfate (10^-5 to 10^-7 M) do not affect collagen biosynthesis in high density cultures. The results show that zinc does not have a direct acceleratory effect on either cellular proliferation or collagen biosynthesis in human skin fibroblasts in vitro.

Excised rabbit testes were observed to contract autorhythmically in vitro at a frequency of 2-3 times/min. When PGF_1a was added to the bathing media, an increase in rate and overall tone with a change in amplitude was observed. At lower concentrations of prostaglandin, the amplitude increased; but at higher levels (greater than 71 nM), amplitude was decreased. The response of the testicular capsule was dose dependent from 0.4 to 213 nM concentration of PGF_1a. When PGF_1a was added to a testicular preparation during the latent period, before contractions were initiated spontaneously, an overall increase in tone was observed at 14 nM concentration, and the appearance of contractions approximately 7 min after treatment. At 57 nM concentration, PGF_1a initiated a series of modulated contractions within 1-3 min that increased in amplitude with time. The available data now suggest that prostaglandins may modulate testicular contractions in vitro: one group inhibiting and a second group stimulating this activity.

The effect of high-fat vs high-carbohydrate diets on the relative role of the pentose cycle (PC) in glucose catabolism was studied in exercised and sedentary rats fed different carbohydrate sources. Expired ¹⁴CO₂ was measured after injection of ¹⁴C-glucose labeled at C-1 or C-6 or at all carbons, and data were fitted to an experimental model. Ad libitum fed Sprague-Dawley rats appeared to show a diurnal rhythm in the degree of utilization of the PC in glucose catabolism. Exercise of calorically restricted animals reduced the relative role of the PC; but altering the fat:carbohydrate ratio of the diet of ad libitum fed animals had no apparent effect. Under a variety of conditions, at least 25% of the glucose which entered relatively labile pools was selectively decar-boxylated at the C-1 position.