Is body focus restricted to self-evaluation? Body focus in the evaluation of self and others

Aspects of the local immune response to nematode challenge were investigated in vivo in isolated loops of the upper small intestine of mature sheep that were immunised by repeated infections with Trichostrongylus colubriformis infective larvae (L3). Groups of 3 sheep were challenged either through the loop (Group 1) or orally (Group 2) with T. colubriformis L3, the third group served as unchallenged controls (Group 3). Nematode specific antibody levels, mast cell proteinase levels (SMCP) and larval migration inhibition (LMI) activity were determined in loop secretions for 4 weeks after challenge. The intestinal loops remained functional throughout the experiment. Groups 1 and 2 were re-challenged 2 weeks after the first challenge, and all 3 Groups were slaughtered 2 weeks later. Histopathological examination showed elevated numbers of globule leukocytes (GL) in both the nematode-challenged loop and unchallenged small intestine of Group 1 and small intestine of Group 2 indicating that nematode infections induce the local appearance of large numbers of GL. Oral, but not loop challenge caused increased antibody levels in loop secretions when compared to unchallenged controls. Only loop-challenged sheep showed a peak in loop fluid SMCP levels 10-13 days after the first challenge which coincided with a peak in numbers of mucosal GL. The isolated loops of all 3 groups showed highly elevated numbers of eosinophils when compared to the intact small intestine. Loop fluid of all 3 groups showed a high level of LMI activity reflecting the high level of nematode-resistance induced by the immunisation procedures. Sheep in Groups 1 and 2 were both able to expel challenge infections, and when compared to Group 3, showed higher blastogenic activity of unstimulated cells derived from a mesenteric lymph node in the region of the challenged part of the intestine. The present experiment showed that surgically constructed intestinal loops provided a model system by which the substantial changes associated with the local intestinal immune response to challenge with T. colubriformis could be investigated.     

Prenatal ethanol exposure: changes in regional brain catecholamine content following stress

Previous studies have shown that fetal ethanol exposure (FEE) may have long-term effects on the function of catecholaminergic neurons in different regions of the CNS. The present study is the first to examine the effects of FEE on regional brain catecholamine responses following acute stress (a single 60-min restraint stress), repeated stress (single periods of restraint stress on 1, 5, or 10 consecutive days), and recovery from stress (recovery for up to 60 min in the home cage following a single 60-min period of restraint stress). Both male and female offspring from FEE, pair-fed (PF), and ad libitum-fed control (C) groups were tested in adulthood to determine catecholamine content in the cortex, hypothalamus, and hippocampus. A single period of restraint reduced cortical norepinephrine (NE) content in FEE and PF animals compared with that in the cortex of C animals, and reduced hypothalamic NE content in FEE female offspring below that found in animals in all other groups. In contrast, hippocampal NE content was higher in FEE than in C animals following a single period of restraint; PF animals had intermediate levels of hippocampal NE and did not differ significantly from either FEE or C animals. Following repeated periods of restraint, cortical NE content was lower in FEE than in C animals; PF animals once again had intermediate levels of NE. Importantly, basal (nonstressed) NE content did not differ among groups in any brain area examined. In addition, several significant changes in regional brain catecholaminergic responses to acute stress were observed in animals across all treatment groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enhanced ligand-induced activation of EGF-receptor and overall tyrosine kinase and phospholipase C in colonocytes isolated from azoxymethane-treated rats

BACKGROUND/AIMS: In order to evaluate the role of epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha) in colorectal cancer, the present investigation examines changes in EGF and TGF-alpha-mediated activation of overall and EGF receptor (EGF-R) associated tyrosine kinase activity in isolated rat colonocytes after administration of the colonic carcinogen azoxymethane. METHODOLOGY: Five days after a single injection of azoxymethane (20 mg/kg) or saline solution to 3-4 month old Fischer-344 rats, colonocytes were isolated, exposed for 2 minutes to 1 x 10(8) M EGF and TGF-alpha, and assessed for overall and EGF-R associated tyrosine kinase and phospholipase C activity. RESULTS: In colonocytes isolated from control animals, incubation with EGF and TGF-alpha resulted in a small (21-35%) increase in overall tyr-k. However, a marked (113-127%) rise of this enzyme occurred in colonocytes from AOM-treated rats, when compared with the corresponding basal levels. These differences were even more pronounced in colonocytes isolated from the distal part of the colon, as regards to the proximal part. In addition, EGF and TGF-alpha activated EGF-R tyr-k by 40-60% in controls and by 84-85% in AOM-treated animals. Incubation of colonocytes with these growth factors also stimulated PLC activity (in controls by 120-150% and in AOM injected rats by 204-271%) when compared with corresponding basal values. CONCLUSIONS: We conclude that AOM enhances the responsiveness of colonocytes to EGF and TGF-alpha, which may be one of the mechanisms involved in colorectal carcinogenesis.     

Characterization of Wnt gene expression in murine skin: possible involvement of epidermis-derived Wnt-4 in cutaneous epithelial-mesenchymal interactions

M. nemestrina immunized with an apathogenic HIV-2 molecular clone (HIV-2KR) were protected from CD4 decline and disease upon challenge with HIV-2(287), after any immunizing virus could be detected. Higher but not lower inocula of HIV-2KR were protective against intravenous inoculation of either 10(5) or 10(1) TCID50 of HIV-2(287). Protected animals displayed substantial reductions in PBMC proviral burden (1-3 logs), viral titers (1-2 logs), and plasma viral RNA (2-4 logs) compared to unprotected or naive animals as early as 1 week postinfection. Plasma viral RNA became undetectable after 24 weeks in protected animals, but remained high in unprotected animals. No viral RNA was present in the spleen of the protected animal necropsied more than a year after challenge (though viral DNA was still present). No neutralizing responses could be demonstrated, but CTL activity was detected sooner and at higher levels after challenge in protected than in unprotected macaques. In this novel HIV-2 vaccine model, protection was clearly dose-dependent, and clearance of challenge virus RNA from the plasma did not require detectable ongoing replication of the immunizing virus at the time of challenge.     

Repeated isolation in the neonatal rat produces alterations in behavior and ventral striatal dopamine release in the juvenile after amphetamine challenge.

Rat pups were isolated from the mother and nest for 1 hr per day from Postnatal Day (PN) 2 to 9. At PN 27, rats were tested for behavioral responsiveness to 2.0 or 7.5 mg/kg amphetamine. Only isolated rats receiving the 7.5 mg/kg dose displayed increased activity scores, compared with nonisolated and nonhandled controls. Their increased activity is attributed to a slower latency to enter into stereotypy. In a second experiment, similarly treated groups were challenged by the 7.5 mg/kg dose during a session in which a microdialysis probe implanted in the ventral striatum was being perfused. The challenge drug elicited a much greater increase in dialysate dopamine in isolated vs nonisolated groups. Results are discussed with regard to dissociation between sensitized and subsensitized responses. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Selenoenzymes regulate the activity of leukocyte 5-lipoxygenase via the peroxide tone

The variation of the selenium status of leukocytes was used as a tool to investigate the influence of selenium-containing glutathione peroxidases on the formation of 5-lipoxygenase metabolites in vitro and ex vivo. Selenium-deficient rat basophilic leukemia cells had < 1% of control glutathione peroxidase activity and 35% of control phospholipid hydroperoxide-glutathione peroxidase activity. Upon stimulation, these cells released an 8-fold amount of lipoxygenase metabolites compared to controls. No (5S)-hydroperoxyeicosatetraenoic acid was detectable in whole cells; however, it was found in homogenates of selenium-deficient cells. Addition of 0.25 microgram/ml selenium to selenium-deficient cells restored control phospholipid hydroperoxide-glutathione peroxidase activity within 8 h, whereas glutathione peroxidase activity needed 7 days. 12 h after resupplementation, selenium-deficient cells had 3% glutathione peroxidase and 100% phospholipid hydroperoxide-glutathione peroxidase activity compared to controls. Resupplemented cells released control amounts of 5-lipoxygenase metabolites, indicating that restoration of phospholipid hydroperoxide-glutathione peroxidase activity is associated with a selenium-adequate leukotriene metabolism. Leukocytes that were isolated from selenium-deficient rats released a 7-fold amount of total lipoxygenase metabolites compared to cells from control animals. By injecting normally fed rats with 500 micrograms/kg selenium as Na2SeO3, leukocyte phospholipid hydroperoxide-glutathione peroxidase activity was raised 8-fold within 114 h compared to controls. Leukocytes from these animals produced significantly less lipoxygenase metabolites than controls. These findings indicate that phospholipid hydroperoxide-glutathione peroxidase activity is primarily responsible for the reduction of 5-hydroperoxyeicosate-traenoic acid and therefore governs the actual activity of leukocyte 5-lipoxygenase via regulating the tone of endogenous hydroperoxides.     

Ca2+ release from intracellular stores is an initial step in hypoxic pulmonary vasoconstriction of rat pulmonary artery resistance vessels

BACKGROUND: A reduction in oxygen tension in the lungs is believed to inhibit a voltage-dependent K+ (Kv) current, which is thought to result in membrane depolarization leading to hypoxic pulmonary vasoconstriction (HPV). However, the direct mechanism by which hypoxia inhibits Kv current is not understood. METHODS AND RESULTS: Experiments were performed on rat pulmonary artery resistance vessels and single smooth muscle cells isolated from these vessels to examine the role of Ca2+ release from intracellular stores in initiating HPV. In contractile experiments, hypoxic challenge of endothelium-denuded rat pulmonary artery resistance vessels caused either a sustained or transient contraction in Ca2+-containing or Ca2+-free solution, respectively (n=44 vessels from 11 animals). When the ring segments were treated with either thapsigargin (5 micromol/L), ryanodine (5 micromol/L), or cyclopiazonic acid (5 micromol/L) in Ca2+-containing or Ca2+-free solution, a significant increase in pulmonary arterial tone was observed (n=44 vessels from 11 animals). Subsequent hypoxic challenge in the presence of each agent produced no further increase in tone (n=44 vessels from 11 animals). In isolated pulmonary resistance artery cells loaded with fura 2, hypoxic challenge, thapsigargin, ryanodine, and cyclopiazonic acid resulted in a significant increase in [Ca2+]i (n=18 cells from 6 animals) and depolarization of the resting membrane potential (n=22 cells from 6 animals). However, with prior application of thapsigargin, ryanodine, or cyclopiazonic acid, a hypoxic challenge produced no further change in [Ca2+]i (n=18 from 6 animals) or membrane potential (n=22 from 6 animals). Finally, application of an anti-Kv1.5 antibody increased [Ca2+]i and caused membrane depolarization. Subsequent hypoxic challenge resulted in a further increase in [Ca2+]i with no effect on membrane potential (n=16 cells from 4 animals). CONCLUSIONS: In rat pulmonary artery resistance vessels, an initial event in HPV is a release of Ca2+ from intracellular stores. This rise in [Ca2+]i causes inhibition of voltage-dependent K+ channels (possibly Kv1.5), membrane depolarization, and an increase in pulmonary artery tone.     

Dynamic accumulation of neutrophils in lungs and visceral organs during early abdominal sepsis in the pig

Activation and accumulation of polymorphonuclear leukocytes (PMNs, neutrophils) in the lungs is considered an important mechanism in the pathogenesis of pulmonary dysfunction in association with sepsis. It probably constitutes only part of a general cellular response; and a corresponding reaction has been implicated in other organs during sepsis (e.g., the liver). In this experiment a model was developed that allows study of the dynamic PMN reaction in the lungs and visceral organs during early abdominal sepsis. The animals were divided into two groups. In the septic group (n = 8) a bacterial challenge was attempted through the intraperitoneal administration of Escherichia coli (1 x 10(11)/kg). Five animals served as controls. All animals in the septic group developed bacteremia, leukopenia, and a hypodynamic circulatory response. PMNs were selectively labeled with 111In-oxine. The activity over the organs was followed dynamically with a gamma camera. The animals subjected to peritonitis exhibited a significant increase in 111In-oxine activity (i.e., neutrophil trapping) in the lungs, compared to the controls at 40 minutes and onward during the observation period. A similar picture was seen over the liver and abdomen, with significance after 70 minutes. The findings in this study indicate that accumulation of PMNs is an early phenomenon not only in the lungs but also in the liver during the development of sepsis. The present model offers possibilities for further studies of the cellular reactions during sepsis.     

Increased training in an aversively motivated task attenuates the memory-impairing effects of posttraining N-methyl-D-aspartate-induced amygdala lesions.

Examined the effect of variations in the amount of preoperative training on the retention deficit produced by posttraining lesions of the amygdaloid complex (AC). Rats received 1, 10, or 20 training trials in a footshock-motivated retention escape task 7 days before receiving N-methyl-{d}-aspartate (NMDA) lesions of the AC. Inhibitory avoidance retention performance, which was measured 4 days postoperatively, indicated that increased training improved retention in AC-lesioned animals as well as in control animals. The retention performance of AC-lesioned animals was impaired when compared with that of controls; however, the impairment was partially attenuated by increased preoperative training. The finding that AC-lesioned animals displayed greater locomotor activity on the retention test compared with nonlesioned controls suggests that the increased activity may have contributed to the impaired inhibitory avoidance retention performance. Two days after the retention test, some of the AC-lesioned animals were subsequently trained on a continuous multiple-trial inhibitory avoidance response in the same apparatus. AC lesions did not block acquisition or retention of the task. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Dietary supplementation with carotenoids: effects on alpha-tocopherol levels and susceptibility of tissues to oxidative stress

The ability of dietary supplementation with carotenoids to protect chick tissues against oxidative stress in vitro was examined. Male Leghorn chicks were fed on diets supplemented (100 mg supplement/kg diet) with either beta-carotene, zeaxanthin (beta,beta-carotene-3,3'-diol), canthaxanthin (beta,beta-carotene-4,4'-dione) or alpha-tocopherol, or on a control diet, from 1 d old until 37 d of age. Tissues (liver, heart, skeletal muscle and plasma) were removed and assayed for total carotenoids and alpha-tocopherol content and portions subjected to oxidative stress by incubation of homogenates with cumene hydroperoxide and FeSo4. Animals receiving zeaxanthin and canthaxanthin had significantly greater carotenoid concentrations in liver, heart, muscle and plasma compared with untreated controls (P < 0.05); animals fed on diets supplemented with beta-carotene, or alpha-tocopherol did not have significantly different tissue carotenoid contents compared with untreated controls. alpha-Tocopherol supplementation elevated alpha-tocopherol levels in all tissues examined (P < 0.05). Supplementation with carotenoids did not affect tissue alpha-tocopherol levels, but beta-carotene lowered plasma alpha-tocopherol levels by 50% (P < 0.05). Incubation of plasma or tissue homogenates with oxidant stressors induced lipid peroxidation (production of thiobarbituric-acid reactive substances) in all tissues. Animals given alpha-tocopherol, beta-carotene or zeaxanthin had a reduced susceptibility to oxidant stress in liver compared with unsupplemented controls (P < 0.05), and alpha-tocopherol-supplemented animals had reduced susceptibility in skeletal muscle compared with tocopherol-supplemented animals had reduced susceptibility in skeletal muscle compared with unsupplemented controls (P < 0.05). Canthaxanthin supplementation did not influence the susceptibility to oxidant stress in any tissue examined. These results suggest that zeaxanthin, a carotenoid present in animal and human diets, may have significant activity as an antioxidant against oxidative stress in tissues.     

Temporal analyses of virus replication, immune responses, and efficacy in rhesus macaques immunized with a live, attenuated simian immunodeficiency virus vaccine

Despite evidence that live, attenuated simian immunodeficiency virus (SIV) vaccines can elicit potent protection against pathogenic SIV infection, detailed information on the replication kinetics of attenuated SIV in vivo is lacking. In this study, we measured SIV RNA in the plasma of 16 adult rhesus macaques immunized with a live, attenuated strain of SIV (SIVmac239Deltanef). To evaluate the relationship between replication of the vaccine virus and the onset of protection, four animals per group were challenged with pathogenic SIVmac251 at either 5, 10, 15, or 25 weeks after immunization. SIVmac239Deltanef replicated efficiently in the immunized macaques in the first few weeks after inoculation. SIV RNA was detected in the plasma of all animals by day 7 after inoculation, and peak levels of viremia (10(5) to 10(7) RNA copies/ml) occurred by 7 to 12 days. Following challenge, SIVmac251 was detected in all of the four animals challenged at 5 weeks, in two of four challenged at 10 weeks, in none of four challenged at 15 weeks, and one of four challenged at 25 weeks. One animal immunized with SIVmac239Deltanef and challenged at 10 weeks had evidence of disease progression in the absence of detectable SIVmac251. Although complete protection was not achieved at 5 weeks, a transient reduction in viremia (approximately 100-fold) occurred in the immunized macaques early after challenge compared to the nonimmunized controls. Two weeks after challenge, SIV RNA was also reduced in the lymph nodes of all immunized macaques compared with control animals. Taken together, these results indicate that host responses capable of reducing the viral load in plasma and lymph nodes were induced as early as 5 weeks after immunization with SIVmac239Deltanef, while more potent protection developed between 10 and 15 weeks. In further experiments, we found that resistance to SIVmac251 infection did not correlate with the presence of antibodies to SIV gp130 and p27 antigens and was achieved in the absence of significant neutralizing activity against the primary SIVmac251 challenge stock.     

Changes with aging, sex and physical exercise in murine natural killer activity and antibody-dependent cellular cytotoxicity

Antibody-dependent cellular cytotoxicity (ADCC) and natural killer (NK) activity were measured in leukocytes from the axillary nodes, the spleen and the thymus of young (12 +/- 2 weeks) and aged (60 +/- 2 weeks) male and female BALB/c mice, which had performed an acute bout of exercise (moderate swimming until exhaustion) or a training exercise (90 min of moderate swimming each day for 20 days). The results show that NK and ADCC activity in sedentary mice (controls) were similar in young and aged animals. However, both kinds of exercise resulted in higher cytotoxicity values in aged mice than in young mice. Acute exercise did not have any effect on NK activity in young and aged mice, nor on ADCC activity in young mice as compared to controls, while training exercise stimulated both cytotoxicities in the two age groups. No correlations between serum corticosterone levels and NK or ADCC activity were found. Our results suggest that moderate training exercise improves both NK and ADCC activity during aging.     

Evaluation of yeast functional assay to detect p53 gene mutation in minute endoscopic samples of colonic mucosal neoplasia

Male and female C57BL/6 (B6) and DBA/2 (D2) mice were subjected to either acute or 5 days of repeated restraint in ventilated, 50 ml centrifuge tubes. Control animals were not disturbed. The acute restraint animals were killed immediately following 15, 30 or 60 min of restraint and blood collected for corticosterone (CORT) analysis. The results of the acute restraint procedure revealed a strain difference in time to peak CORT in plasma with D2 animals showing an earlier peak. The males of both strains evinced similar maximum response and similar to B6 females; however, the D2 females showed a 2-fold greater CORT response than did the B6 females. Repeated restraint consisted of 5 days of 12 h in the tubes. At the end of 5 days, the animals were weighted and adrenalectomized in preparation for determination of brain corticosteroid receptors. Upon sacrifice, brains, thymus, adrenals and blood were harvested, the last for corticosteroid binding globulin (CBG). Five days of repeated restraint produced body weight loss in both strains, with B6s less affected than D2s. Repeated restraint reduced the mass of the adrenals in the B6s only. Restraint also reduced the mass of the thymus in both strains and sexes, but to a greater extent in the B6s. Plasma CBG densities were also sensitive to restraint, but only in females, showing a restraint-related decrease. Repeated restraint had no effect on hippocampal glucocorticoid or mineralocorticoid receptors; however for the latter, we observed significant strain and sex effects with D2 having higher Bmax than B6 and females having higher Bmax than males. In the pituitary, glucocorticoid receptors (GR) were reduced by repeated restraint in males, but increased in females, especially in the B6. These findings lend preliminary evidence for involvement of sex and genetics as sources of individual differences in bioadaptation to stress.     

Characterization of two cDNA clones which encode O-methyltransferases for the methylation of both flavonoid and phenylpropanoid compounds

The effects of permanent disruption of neuropeptide transmission on the induction (i.e., sensitization) and elicitation (i.e., challenge) phases of contact hypersensitivity (CHS) are described. BALB/c mice were chemically denervated of neuropeptide (i.e., tachykinin) containing sensory C fibers by an acute injection of capsaicin (50 mg/kg) on postnatal day (PND) 2 to 3. As young adults (PND 45-60), these mice and their control littermates were sensitized by topical application of 0.1% 2,4-dinitrofluorobenzene (DNFB) or vehicle. Treatment groups generated from this exposure regimen consisted of untreated, controls (O/O), denervated, controls (CAP/O), untreated, sensitized (O/DNFB), and denervated, sensitized (CAP/DNFB). The elicitation phase of CHS was evaluated in these animals by measuring ear thickness in response to a DNFB challenge. In DNFB-sensitized groups, ear thickness was significantly increased over controls but was additionally increased 2.4-fold in CAP/DNFB compared to O/DNFB mice. The induction phase of CHS was next assessed in young adult mice by measuring lymph node cell (LNC) proliferation. For this, mice were sensitized for 3 consecutive days before their draining, auricular nodes were removed. The LNC were dissociated and cultured for 24 h with tritiated thymidine to assess LNC proliferation. As expected, significantly higher numbers of LNC occurred in both DNFB-sensitized groups (CAP/DNFB, O/DNFB) compared to the unsensitized, controls (CAP/O, O/O). However, LNC proliferation in CAP/DNFB was significantly higher than O/DNFB animals. Flow cytometry on similarly exposed mice failed to demonstrate any significant difference in the population of CD4CD8 or CD3CD45R LNC cells from neuropeptide-denervated (CAP/O, CAP/DNFB) mice or their respective treatment mates (O/O, O/DNFB), suggesting that alterations in T or B cell populations did not underlie these changes. Finally, cytokine release from the LNC from these treatment groups was examined. For this, the auricular lymph nodes were removed from animals, 2 to 4 h after the animals were administered a single application of a sensitizing concentration (0.1%) of DNFB or acetone vehicle. LNC, dissociated from these nodes, were cultured for 24 h. The nutrient media was removed from these cultured cells and examined for the release of proinflammatory cytokines, interleukin (IL)-1beta, IL-2 and tumor necrosis factor (TNF)alpha, by ELISA. There were no significant increases in IL-2. However, IL-1beta release was significantly increased in CAP/DNFB mice over O/DNFB by 18-fold and by over 30-fold compare to O/O controls. Levels of TNFalpha were significantly increased in both O/DNFB and CAP/DNFB mice over the nonsensitized controls (O/O, CAP/O). CAP/DNFB values were approximately double that of O/DNFB. There was no significant difference in IL-1beta or TNFalpha release between the nonsensitized controls (O/O, CAP/O). Collectively, these data indicate that neuropeptide denervation by neonatal administration of capsaicin alters both the induction and elicitation phases CHS and may modify sensitivity to chemically induced CHS.     

Application of solid-phase microextraction and gas chromatography-mass spectrometry for the determination of chlorophenols in urine

Previously, we have demonstrated that chronic exposure to immobilization (IMO) did not modify the influence of catecholamines on active behaviour of rats in the holeboard, but clearly increased the role of these amines in the forced swimming test (FST). In the present experiment, it was studied whether or not chronic IMO altered the role of dopamine in the two tests. Adult male Sprague-Dawley rats were left either undisturbed or subjected daily to 2 h of IMO stress for 12 days. On the following day, half of the rats were administered saline and the others the dopamine antagonist haloperidol (0.5 mg/kg). Then the rats remained undisturbed in the animal room (controls) or were subjected to acute IMO for 2 h. Finally, all animals were exposed consecutively to the holeboard (4 min) and the FST (5 min). In non-chronically stressed rats, acute IMO depressed behaviour in the holeboard but not in the FST. In chronic IMO rats the inhibitory effect of acute IMO on holeboard activity was slightly reduced as compared to controls. Acute IMO increased struggling in rats previously exposed to chronic IMO but did not alter struggling in non-chronically stressed rats. Whereas the inhibition caused by haloperidol treatment in the active behaviour of rats in the holeboard was not altered by chronic IMO, the inhibitory effect of haloperidol in the active behaviour of rats in the FST was greater after chronic IMO, particularly in rats also subjected to acute IMO. These data suggest that chronic IMO stress potentiates the role of dopamine in a specific behavioural task such as the FST and adds support to the previously published data demonstrating enhanced behavioural and neurochemical responses to dopamine-related drugs after chronic stress.     

Effects of high doses of beclomethasone dipropionate in bronchial asthma

Fifteen bovine herpesvirus-1 (BHV-1)-negative calves were vaccinated intramuscularly with 10(7.4) plaque-forming units of a double-deletion BHV-1 mutant (IBRV(NG)dltkdlgIII), and 6 remained as nonvaccinated controls. Thirty days after vaccination, the animals were challenged by nasal instillation of 10(8.2) CCID50 of a virulent BHV-1 strain (Cooper). The vaccinated calves were protected against wildtype virus challenge as demonstrated by clinical evaluation. Most of the vaccinates developed only a mild rhinitis (lasting an average of 6.5 days) with almost no systemic symptoms, whereas the controls developed a serious illness characterized by rhinitis (mean = 11.5 days), conjunctivitis, hyperthermia, apathy, loss of appetite, and dyspnea. The vaccinates also shed significantly less virus and for a shorter period of time (mean = 5.5 days) than the controls (mean = 9 days). Thirty days after vaccination, the vaccinates were negative in an anti-gIII specific blocking enzyme-linked immunosorbent assay (ELISA), despite the fact that most of them had developed neutralizing antibodies (serum neutralization titers ranging from 1:2 to 1:16). Seroconversion to gIII was detected as early as 7 days postinfection (dpi). Fourteen days after the challenge, all the animals exposed to wildtype BHV-1 had developed anti-gIII antibodies and were positive in this differential serologic test. Six controls plus 8 vaccinates kept in isolation were still positive to gIII when tested at 75 dpi. The use of the IBRV(NG)dltkdlgIII strain in conjunction with an anti-gIII specific blocking ELISA kit represents a powerful tool for BHV-1 control/eradication programs.     

Sucrosemia in untreated celiac disease: a potential screening test

During studies to develop serum tests of small intestinal permeability, we detected an unidentified disaccharide in HPLC traces of sera from untreated celiacs. This present study aimed to identify the disaccharide and determine whether the presence of the disaccharide in the serum after an oral challenge had potential as a simple screening test for celiac disease. The disaccharide was identified as sucrose by incubation studies of sera with disaccharidases. Twenty untreated celiacs, 15 treated celiacs, and 20 normal or dyspeptic controls were studied for the presence of sucrose in their serum after an oral load (8 g). The results in celiacs were compared with the presence of serum IgA endomysial antibodies. The 10 normal controls were also given a larger sucrose challenge (50 g). Ten of the untreated celiacs and 10 controls had their brush border disaccharidase activities measured. Sucrose eluted in the same position as the unidentified disaccharide in the HPLC trace and the latter could be removed by incubation with sucrase. All untreated celiacs but none of the treated celiacs had sucrose in their serum after the 8-g oral challenge. None of the controls had sucrose in their serum after the 8-g or 50-g challenges. Three untreated celiacs were IgA endomysial antibody negative as were all the treated cases. Brush border sucrase activity was low in untreated celiac disease. The presence of sucrose in the serum after an oral load shows promise as a noninvasive test for celiac disease.     

Regulation of hepatic glutaminase in the streptozotocin-induced diabetic rat

The liver of diabetic animals removes increased quantities of glutamine. We therefore examined factors that affect hepatic glutaminase activity in hepatocytes and mitochondria. Glutamine use, through glutaminase, was measured in isolated rat hepatocytes by monitoring the production of 14CO2 from [1-(14)C]glutamine. Hepatocytes from streptozotocin-induced diabetic rats use glutamine more rapidly than do hepatocytes from normal or insulin-maintained diabetic rats. Glutamine use in all of these hepatocytes was stimulated by glucagon and epinephrine. Glutaminase activity, assayed in broken mitochondrial membranes, was increased approximately 2.5-fold in diabetic rats. The sensitivity of glutaminase, measured in intact liver mitochondria, to phosphate was markedly left-shifted in mitochondria from diabetic rats compared with those from controls. In fact, glutaminase was increased 10-fold at 2.5 mmol/l phosphate compared with controls. This increased sensitivity of glutaminase to physiological concentrations of phosphate is characteristic of its hormonal activation. Therefore, activation of glutaminase plays a major role in diabetes and is as important as increases in its total enzyme amount in determining the increased glutamine uptake in diabetes.     

Liver and heart mitochondrial succinate dehydrogenase activity of newborn rats in anoxic hypoxia and starvation

Succinate dehydrogenase activity was determined in the liver and heart of newborn rats after 3 and 48 hours' exposure to anoxic hypoxia (10% O2) and after 48 hours' starvation. Control determinations were made on newborn animals of corresponding ages, full term foetuses (21 days), infantile (1 and 2 weeks) and full grown animals. Hypoxia for 3 h had no influence on succinate dehydrogenase activity at all in either the heart or liver mitochondria of the newborn animals. After 48 h no difference was observed in the liver between the hypoxic animals and the starved controls of the same age, though starvation itself had resulted in a significant increase in activity, as much as 42%. When liver mitochondrial succinate dehydrogenase in normal mitochondria was activated by preincubation mitochondria with the substrate, the activity increase obtained was greater than that resulting from starvation. The increase in activity in the heart of the hypoxic or starved animals was not significant (less than 10%).