Generation of lymphokine-activated killer activity in rodents but not in humans is nitric oxide dependent

The role of nitric oxide (NO) in the generation of lymphokine-activated killer (LAK) cells was investigated. Here we report that L-arginine analog NG-monomethyl-L-arginine (NMMA), a specific inhibitor of nitric oxide synthase, prevents LAK cell generation from cultured rat splenic cells. Accumulated NO endproduct nitrite (NO2-), as measured in the supernatants of rat splenic cells, correlated well with the generation of LAK cells. In contrast, cell proliferation induced by rIL-2 or by Con A was not affected by NMMA. Similarly, phenotypic expression of CD25 in rIL-2-stimulated cultures was unaffected. Furthermore, we could not observe differences in percentages of CD5-CD8+ cells (NK and LAK cell phenotype markers in rats) between rIL-2-stimulated cultures performed in the presence or absence of NMMA. LAK cell generation could no longer be blocked if NMMA was added to the rat cell cultures 24 hr after rIL-2 stimulation. To further confirm the role of NO in LAK cell generation, rat splenic cells were cultured in medium without L-arginine. Under such conditions rIL-2 could not induce LAK cell generation. Hemoglobin, which is a scavenger of NO, also inhibited LAK cell generation. Finally, addition of sodium nitroprusside (SNP) which releases NO in cultures was able to overcome blocking effects of NMMA. To attempt the identification of NO-producing cells, lysosomotropic agent, L-leucine methyl ester (LME), was used. Generation of LAK cell activity was virtually abolished in cell cultures treated with LME. Addition of SNP to cultures, however, sufficed to restore LAK cell generation. These results suggest that LAK cell precursors depend on a exogenous NO supply from other cell types in order to display their full cytotoxic potential. Similar results were also obtained by using mouse splenocytes as responder cells. In contrast, NMMA did not affect generation of LAK cells from human peripheral blood or spleen mononuclear cells.     

The chemotaxis system, but not chemotaxis, is essential for swarming motility in Escherichia coli

The chemotaxis system plays an essential role in swarm cell differentiation and motility. We show in this study that two (Tsr and Tar) of the four chemoreceptors in Escherichia coli can support swarming individually, but sensing their most powerful chemoattractants is not necessary. Conditions that abolish chemotaxis toward serine (presence of serine concentrations that saturate Tsr, or mutations in Tsr that destroy serine binding) have no effect on swarming. Similar results were obtained for the aspartate and maltose chemoreceptor Tar. We also show that although a mutation in the signaling domain of Tsr that inhibits CheA kinase abolishes swarming, nonchemotactic flagellar switch mutants can swarm. Our results suggest that during swarming, the chemoreceptors signal through the chemotaxis pathway and induce swarmer cell differentiation in response to signals other than their known chemoeffectors.     

Hippocampal atrophy is related to impaired memory, but not frontal functions in non-demented Parkinson's disease patients

We investigated the neuropsychological correlates of hippocampal atrophy in Parkinson's disease (PD) patients. The memory impaired PD patients had smaller hippocampi than other PD patients. The performance of PD patients in spatial working memory and attentional set-shifting correlated with the severity of motor defect, and not with hippocampal atrophy. Our results suggests that failure of verbal/visual memory may be related to hippocampal atrophy in Parkinson's disease. On the contrast, the defect in spatial working memory and attentional set-shifting may be sensitive to dysfunction of 'fronto-striatal' systems in PD patients.     

Perforin is essential for control of ectromelia virus but not related poxviruses in mice

Lack of perforin renders the relatively resistant mouse strain C57BL/6 highly susceptible to the natural mouse pathogen ectromelia virus, a cytopathic orthopoxvirus. This is indicated by increased mortality, elevated virus titers and pathology in liver and spleen, and increased levels of liver enzymes in blood. Cowpox virus on the other hand is more virulent in the presence of perforin than in its absence. An additional lack of granzyme A which together with perforin is a constituent of cytoplasmic granules from cytotoxic T cells increases the virulence of cowpox virus.     

Amniotic fluid lecithin/sphingomyelin ratio changes related to centrifugal force

Lecithin/sphingomyelin ratio (L/S ratio) in amniotic fluid has gained wide clinical acceptance as an index of fetal lung maturity. We determined L/S ratios of amniotic fluids centrifuged at various "g-forces." Our studies demonstrate that the L/S ratio value is highly dependent on the g-force used to prepare the fluid. We recommend standardization of the g-force, time, and temperature used in preparation of the amniotic fluid for L/S ratio determination.     

Apoptosis occurs in granulosa cells but not cumulus cells in the atretic antral follicles in pig ovaries

The porcine antral follicles, 3-6 mm in diameter, were dissected from the ovaries of mature pigs, and then granulosa and cumulus cells were isolated from each follicle. In atretic follicles, high activity of neutral Ca2+/Mg2+-dependent endonuclease and DNA ladder formation, estimated by electrophoresis, were noted in granulosa cells but not in cumulus cells. Extremely low activity of the endonuclease and no DNA ladder formation were observed in both types of cells obtained from healthy follicles. Moreover, apoptotic cells were observed histochemically among granulosa cells only. A good correlation (r = 0.987) between the endonuclease activity of granulosa cells and the progesterone/estradiol ratio of follicular fluid in each follicle was found. These results suggest that apoptosis occurs in granulosa cells but not cumulus cells in the atretic antral follicles in pigs.     

Tachykinins may mediate capsaicin-induced, but not vagally induced motility in porcine antrum

Tachykinins are thought to be involved in extrinsic control of motility in the gastrointestinal tract. Using the isolated perfused porcine antrum with intact vagal innervation, we studied the effects of substance P, neurokinin A and capsaicin infusion, and electrical stimulation of the vagus nerves on antral motility without or with infusion of non-peptide antagonists for NK-1 receptors (CP96345) and NK-2 receptors (SR48968). Substance P and neurokinin A stimulated antral motility in a dose-dependent manner. The effect could be inhibited by atropine or a combination of the NK-1 and NK-2 receptor antagonists. Electrical stimulation of the vagus nerves and infusion of capsaicin (10(-5) M) stimulated antral motility. Vagally induced motility was not influenced by infusion of CP96345 and SR48968, whereas the effect of capsaicin was blocked. We conclude that tachykinins may be involved in regulation of antral motility through sensory nerves in the porcine antrum, but they do not seem to be involved in vagal regulation of antral motility.     

Atrial natriuretic peptide is not degraded by the lungs in humans

In an attempt to identify and quantify the sites of atrial natriuretic peptide (ANP) degradation, particularly the lungs, a new tracer method to study ANP metabolism in vivo in humans was developed and applied to patients with left ventricular dysfunction. Thirteen male, normotensive, cardiac patients with different degrees of left ventricular myocardial involvement were enrolled in the study. The study protocol required constant infusion (3 patients) or bolus injection (10 patients) of 125I-labeled ANP just upstream of the right atrium and blood sampling from different sites (pulmonary artery, aorta, inferior vena cava, and femoral vein) during the hemodynamic study. Data analysis was based on a kinetic model consisting of three blocks in series (right heart, lungs and left heart, and periphery) supplied by the same plasma flow (plasma cardiac output). Plasma levels of native ANP were measured with a sensitive and specific immunoradiometric assay method. ANP values measured in the aorta (163.9 +/- 144.8 pg/mL, n = 80) were superimposable on those measured in the pulmonary artery (161.8 +/- 136.5 pg/mL, n = 80). Negligible extraction of 125I-labeled ANP was found in the lungs and left heart block (on average 0.08 +/- 3.92%), whereas the peripheral block extraction (46.2 +/- 7.8%) accounted for almost total hormone removal from the blood (whole body extraction was 46.4 +/- 6.6%). ANP metabolic clearance rate (3.11 +/- 1.48, range 1.4-6.8 L/min) declined with the progression of left ventricular dysfunction (plasma cardiac output 3.46 +/- 1.08, range 1.2-5.7 L/min), and a close correlation between metabolic clearance rate and cardiac output was evident. Our data suggest that lungs do not extract, or extract only very small amounts, of labeled ANP administered iv to patients with different degrees of left ventricular myocardial involvement, and whole body extraction of labeled ANP remains relatively stable with the progression of disease, and the large reductions in clearance values observed in our patients can be ascribed mainly to the reductions in cardiac output.     

Discriminative inhibition is specific to the response-reinforcer association but not to the discriminative stimulus

In 4 experiments an instrumental contingency between a response and a reinforcer was introduced in the presence of a discriminative stimulus. Then a discriminative inhibitor (S delta) was established that signaled that the instrumental contingency would no longer operate in the presence of that discriminative stimulus, so that the S delta suppressed operant responding. The degree to which the S delta, s inhibitory properties transferred to different discriminative stimuli and different response-reinforcer associations was then explored. In Experiments 1 and 2 the S delta, s effects transferred perfectly to a 2nd discriminative stimulus, whereas the results of Experiments 3 and 4 were consistent with the hypothesis that the S delta, s inhibitory properties were specific to the original response-reinforcer association. The theoretical implications of these findings are discussed.     

Mediation by nitric oxide of TRH-, but not metoclopramide-stimulated TSH secretion in humans

In order to establish whether nitric oxide (NO) participates in the regulation of thyroid stimulating hormone (TSH) secretion in humans, seven normal men were treated with a placebo (normal saline) or the NO synthase inhibitor L-NAME, given at doses (40 micrograms kg-1 injected plus 50 micrograms kg-1 infused i.v.) previously found to be unable to change blood pressure. Experiments were carried out either in basal conditions or during stimulation of TSH secretion with an i.v. injection of 200 micrograms thyrotropin releasing hormone (TRH) or 10 mg of the dopaminergic antagonist metoclopramide (MCP). Administration of L-NAME did not change the basal secretion of TSH or the TSH response to MCP, but significantly reduced the TSH increase induced by TRH. These data fail to provide evidence of NO involvement in regulation of basal TSH secretion. NO also appears to be without effects on the dopaminergic control of TSH secretion. In contrast, the inhibitory effect of L-NAME on TRH-induced TSH secretion suggests the mediation by NO of the TSH-releasing action of TRH.     

Topography following keratoplasty in cats is not representative of that in humans

The excurrent duct system of the rat submandibular gland consists of a number of distinct segments. Using the direction of salivary flow as a reference point, these segments are, in order, intercalated duct, granular convoluted tubule, striated duct, excretory duct, main excretory duct (MED), and salivary bladder (which is an expanded portion of the MED). Because these ducts (with the exception of the MED and the salivary bladder) are encased in secretory endpieces, they are difficult to locate and to observe by scanning electron microscopy. A simple method has been devised to rid the gland of these obscuring endpieces so that the detailed architecture of the duct system can be examined. Rat submandibular glands were fixed initially by vascular perfusion with half-strength Karnovsky's fixative. The connective tissue capsule was removed from extirpated glands and the glands remained in fixative for varying lengths of time. For our purposes, a 30-minute immersion in the aldehyde mixture was optimum. After the sublingual gland was removed, the submandibular gland was softly struck with forceps having rounded tips, then shaken in fixative or buffer. The tissue that remained was postfixed in osmium tetroxide. This method results in the complete divestment of nonductular parenchyma from the rat submandibular gland, leaving the duct system clean and ready for microscopic examination.     

Abnormal mucosal glycoprotein synthesis in inflammatory bowel diseases is not related to cigarette smoking

Patients with ulcerative colitis are usually non- or ex-smokers in contrast to Crohn's disease where smoking is common. Abnormalities of quantity and quality of intestinal mucus have been postulated in the pathogenesis of these diseases. It is possible that smoking habit may exert its effects via changes in mucus in inflammatory bowel disease. We have therefore studied incorporation of N-acetylglucosamine into synthesized colonic mucin in explants from 85 controls with normal colonoscopic appearances and histology, including 27 smokers and 58 nonsmokers, 36 patients with ulcerative colitis and 19 with ileocolonic Crohn's disease over 24 h in tissue culture. Incorporation of N-acetylglucosamine into normal explants was 31.3 +/- (SD) 7.1 dpm/microgram biopsy protein, incorporation was increased in patients with active Crohn's disease (mean 41.2 +/- (SD) 10.4 dpm/microgram biopsy protein, p = 0.003), decreased in inactive ulcerative colitis (mean 24.1 +/- 7.8 dpm/microgram biopsy protein, p = 0.0006) but normal in active ulcerative colitis (mean 35.0 +/- 13.8 dpm/microgram biopsy protein, p = 0.44). No significant relationship was found between cigarette smoking habits and mucus synthesis in controls with normal mucosa (nonsmokers, n = 58, mean 31.0 +/- (SD) 7.52 dpm/microgram biopsy protein; smokers, n = 27, mean 31.8 +/- (SD) 6.1 dpm/microgram biopsy protein, p = 0.9). This study shows that mucus glycoprotein synthesis is reduced in inactive ulcerative colitis, rising to normal levels in active disease and that synthesis is increased in Crohn's disease. There is no effect of smoking on mucus synthesis by control biopsies suggesting that the differences seen in inflammatory bowel disease are not related to cigarette smoking.     

Responses of leptin to short-term fasting and refeeding in humans: a link with ketogenesis but not ketones themselves

We investigated the response of leptin to short-term fasting and refeeding in humans. A mild decline in subcutaneous adipocyte ob gene mRNA and a marked fall in serum leptin were observed after 36 and 60 h of fasting. The dynamics of the leptin decline and rise were further substantiated in a 6-day study consisting of a 36-h baseline period, followed by 36-h fast, and a subsequent refeeding with normal diet. Leptin began a steady decline from the baseline values after 12 h of fasting, reaching a nadir at 36 h. The subsequent restoration of normal food intake was associated with a prompt leptin rise and a return to baseline values 24 h later. When responses of leptin to fasting and refeeding were compared with that of glucose, insulin, fatty acids, and ketones, a reverse relationship between leptin and beta-OH-butyrate was found. Consequently, we tested whether the reciprocal responses represented a causal relationship between leptin and beta-OH-butyrate. Small amounts of infused glucose equal to the estimated contribution of gluconeogenesis, which was sufficient to prevent rise in ketogenesis, also prevented a fall in leptin. The infusion of beta-OH-butyrate to produce hyperketonemia of the same magnitude as after a 36-h fast had no effect on leptin. The study indicates that one of the adaptive physiological responses to fasting is a fall in serum leptin. Although the mediator that brings about this effect remains unknown, it appears to be neither insulin nor ketones.     

Fas antigen is expressed in human diseased muscles, but does not link to apoptosis

In an attempt to determine whether the Fas-Fas-ligand mediated apoptosis occurs in the pathologic process of human myopathies, we examined the expression of Fas antigen, Fas-ligand mRNA, and chromosomal DNA fragmentation in the muscle cells of several human muscle disorders. The Fas antigen of a whole molecule was expressed on the muscle fibers of patients with muscle wasting diseases. However, there was no evidence of an apoptotic process, nor Fas-ligand synthesis in the diseased muscle tissue. Therefore, the expression of Fas antigen on muscle fibers in diseased muscle might be related to unknown biological functions other than "apoptosis" in the process of muscle fiber injury.     

Fever in goldfish is induced by pyrogens but not by handling

OBJECTIVE: To determine whether end-tidal partial pressure of carbon dioxide (PETCO2) was a reliable estimate of PaCO2 in dogs undergoing thoracotomy. DESIGN: Case series. ANIMALS: 18 dogs that underwent thoracotomy. PROCEDURE: PaCO2 and PETCO2 were measured shortly after induction of anesthesia, while dogs were breathing spontaneously; 5 minutes prior to initial skin incision, while dogs were receiving intermittent positive-pressure ventilation (IPPV); 5, 30, and 60 minutes after the thoracic cavity was opened, while dogs were receiving IPPV; and after the thoracic cavity was closed and evacuated, when dogs were again breathing spontaneously. For each period, arterial-end-tidal difference in partial pressure of carbon dioxide (PaCO2-PETCO2) was compared with PaCO2-PETCO2 for the preceding period. RESULTS: Significant changes in PaCO2-PETCO2 from one period to the next were not detected except when values obtained 5 minutes after the thoracic cavity was opened were compared with values obtained 5 minutes before incision. The PaCO2-PETCO2 was not constant for individual dogs. CLINICAL IMPLICATIONS: PETCO2 was not a reliable indicator of adequacy of ventilation during thoracotomy in these dogs, because it differed greatly from PaCO2, and PaCO2-PETCO2 was not consistent.     

Prolonged fasting in humans results in diminished plasma choline concentrations but does not cause liver dysfunction

The potential antiatherogenic actions of the angiotensin II receptor antagonist, losartan were investigated in apolipoprotein (apo) E deficient mice, an animal model with severe hypercholesterolemia and extensive atherosclerosis. In these animals accelerated atherosclerosis is associated with increased lipid peroxidation which may play a crucial role in the build up of the atherosclerotic lesions. Administration of losartan (25mg/kg/d) to the apo E deficient mice for a 3-month period increased the plasma renin activity 3.5-fold compared to the placebo group. Losartan increased the resistance of LDL to CuSO4-induced oxidative modification as shown by a significant reduction in the LDL content of malondialdehyde by 55% compared to placebo, as well as by the prolongation of the lag time required for LDL oxidation, from 60 min in the placebo-treated mice to more than 140 min in the losartan-treated mice. Losartan reduced significantly the mean atherosclerotic lesion area by 80% compared to the placebo group. We conclude that losartan inhibits LDL lipid peroxidation in the apo E deficient mice and this effect may have an important role in the attenuation of the accelerated atherosclerosis.     

Ketoconazole attenuates the cortisol response but not the subjective effects of smoked cocaine in humans

Attenuation of hypothalamo-pituitary-adrenal (HPA) function in laboratory rodents has been found to reduce the reinforcing effects of cocaine. To examine whether attenuation of HPA function reduces the effects of cocaine in humans, one female and seven male 'crack' cocaine abusers were pretreated with three doses of ketoconazole (0, 600, 1200 mg), an inhibitor of adrenocorticoid biosynthesis, 1 h before receiving cocaine. Three doses of smoked cocaine (0, 12, 50 mg) were administered in counterbalanced order under each ketoconazole condition. Ketoconazole dose-dependently reduced cocaine-induced cortisol, but not adrenocorticotropin (ACTH) release, and attenuated the cocaine-induced increase in heart rate and blood pressure. Plasma ACTH levels were more predictive of blood pressure changes than either cocaine or cortisol levels. Suppression of cortisol secretion was not associated with a reduction in ratings of the subjective effects of cocaine. These results support a role for the HPA axis in the cardiovascular effects of cocaine, but do not support a role for the HPA axis in the subjective effects of cocaine. To the extent that self-administration can be predicted by subjective effects, these results further argue that the HPA axis does not play a critical role in cocaine self-administration by humans.