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Microchimerism and transmission of porcine endogenous retrovirus from a pig cell line or specific pathogen-free pig islets to mouse tissues and human cells during xenografts in nude mice.

AIMS/HYPOTHESIS: Pig islets could transmit porcine endogenous retroviruses (PERV) to diabetic patients. Our previous work showed that pig islets expressed low levels of PERV mRNA and were not likely to transmit PERV to human cells in vitro. The real risk of infection during pig tissue xenografts can only be evaluated by in vivo experiments. METHODS: Nude mice bearing tumours containing human 293 cells were grafted with specific pathogen-free pig islets or PERV-producing pig PK15 cells to determine whether pig cells could transmit PERV to mouse and human cells in vivo. Infection was monitored by PCR, long PCR, RT-PCR and long RT-PCR. As detection of PERV sequences could be due to the presence of residual pig cells, we looked for pig mitochondrial (mt) DNA. Quantitative PCR for PERV and pig mt DNA was done to compare the PERV-to-pig mt (P-to-M) ratio of each sample with the reference ratio for grafted pig cells. RESULTS: Among 78 mouse tissues from PK15-grafted mice, 54 and 72 were positive for gag and pig mt DNA, respectively. Human tumours developed in these mice were positive for PERV (78%) and pig mt (89%). The P-to-M ratios for mouse tissues and PERV-positive human tumours from PK15-grafted mice were higher than the ratio in PK15 cells. Among 41 tissues from pig islet cell-grafted mice, 7 were positive for PERV (3 lymph nodes, 1 kidney, 2 salivary glands, 1 ovary), and 14 were positive for pig mt DNA. Three of these samples (1 lymph node, 1 kidney and 1 salivary gland) were positive for gag DNA, but negative for pig mt DNA. One human tumour in these mice was positive for PERV DNA. P-to-M reference ratio in grafted islet cells was 0.05+/-0.03. The three PERV-positive lymph nodes contained 78 gag/3 mt copies (P-to-M: 26), 101 gag/3 mt copies (P-to-M: 34), and 4 gag/0 mt copies. The two PERV-positive salivary glands contained 14 gag/1 mt copies, and 28 gag/0 mt copies. The ovary and the kidney contained 46 gag/3 mt and 69 gag/0 mt copies, respectively. The PERV-positive human tumour contained 47 gag/3 mt copies. CONCLUSIONS/INTERPRETATION: Microchimerism and PERV transmission were frequently observed in both mouse and human tissues during grafting of pig PK15 cells into nude mice bearing human tumours, and sometimes during pig islet xenograft in this model. This strengthens the notion that there is a risk of transmitting PERV during pig islet xenograft.

Animals↗

The development of acid and pepsin (EC 3.4.23.1) secretory capacity in the pig; the effects of age and weaning. 1. Studies in anaesthetized pigs.

The development of gastric secretory capacity of hydrochloric acid and pepsin (EC 3.4.23.1) was studied in thirty-eight Large White X Landrace pigs from the litters of six sows (three pairs of two), 9-38 d of age. The pigs of each pair of sows were born within 24 h of each other. The pigs of a litter were paired according to sex and size and cross-fostered, i.e. one pig from each pair was allocated to each sow. One litter from each pair was reared entirely by the sow (milk-fed, MF) whereas the other litter was reared by the sow for 21 d, but was allowed access to solid food (210 g crude protein (nitrogen X 6.25)/kg) at 12 d and was entirely dependent on solid food after 21 d (creep-fed, CF). Following a 14-18 h fast, pigs were anaesthetized (Halothane-sodium pentobarbitone) and their stomachs perfused at a constant rate with Ringer solution. Gastric secretion was stimulated by intravenous infusion of betazole hydrochloride (Histalog) at 3 mg/kg per h for 2 h. Hydrochloric acid and pepsin were measured in the perfusate which was collected at 15-min intervals. There were significant positive correlations between stomach weight and body-weight for both MF and CF pigs. The slope of the regression line for CF pigs was significantly greater than that for MF pigs (P less than 0.01). There were significant positive correlations between maximal acid output and stomach weight for both MF and CF pigs. There were significant positive correlations between maximal pepsin output and stomach weight for both MF and CF pigs. The slope of the regression line for CF pigs was significantly different from that for MF pigs (P less than 0.01). There were also significant positive correlations between maximal pepsin output per unit stomach weight and stomach weight for both MF and CF pigs. The pattern of development of pepsin secretory capacity in both CF and MF pigs was different from that for acid secretion. Maximal outputs of acid per unit stomach weight for MF and CF pigs remained relatively constant. Maximal outputs of pepsin per unit stomach weight and per unit body-weight increased with age for both MF and CF pigs. The results indicate that pigs given access to solid food before weaning and weaned on to solid food have heavier stomachs and greater acid and pepsin secretory capacity than pigs fed entirely on sows' milk.

Aging↗

Haptoglobin and pig-major acute protein are increased in pigs with postweaning multisystemic wasting syndrome (PMWS).

The objective of this study was to determine the serum concentration levels of selected acute phase proteins (APP), haptoglobin (HPT) and pig-major acute phase protein (pig-MAP), in postweaning multisystemic wasting syndrome (PMWS) affected pigs and PCV2-subclinically infected pigs. In a first study, a group of 15 eight-week-old conventional pigs from a PMWS affected farm were bled and a complete necropsy, histopathology and in situ hybridisation to detect PCV2 were performed. Based on the results, pigs were classified as suffering from PMWS (n = 10) or healthy animals (n = 5). In a second study, a group of 45 pigs from another PMWS affected farm were selected and bled at 3, 7, 12 and 28 weeks of age. The assessment of PCV2 infection status in these pigs was retrospectively done by PCV2 PCR in serum samples. Selected APP were measured in the serum of all studied pigs by means of radial immunodiffusion. Mean HPT and pig-MAP levels were significantly increased (p = 0.004 and p = 0.0006 respectively) in PMWS-affected pigs when compared to levels found in healthy pigs (2.52 +/- 0.88 mg/mL vs. 1.06 +/- 0.73 mg/mL for HPT and 3.81 +/- 1.53 mg/mL vs. 0.76 +/- 0.34 mg/mL for pig-MAP). In the second study, no significant difference in mean HPT and pig-MAP values were observed between PCV2 PCR positive and negative pigs of any age. However, both APP increased significantly with age in PCV2 PCR negative pigs. Altogether, the present results suggest that APP levels are significantly increased in pigs that develop PMWS, but not in animals with a PCV2 subclinical infection.

Acute-Phase Proteins↗

Effect of spray-dried plasma and lipopolysaccharide exposure on weaned pigs: I. Effects on the immune axis of weaned pigs.

A study was conducted with 20 weaned barrows (14 d, 4.98 +/- .21 kg) to determine the effect of spray-dried plasma (SDP) on the pig's immune response to a lipopolysaccharide (LPS) challenge. After weaning, pigs were fed a diet containing 0 or 7% SDP for 7 d. On d 6 postweaning, all pigs were fitted with a jugular catheter. On d 7 postweaning, the pigs were given an i.p. injection of either saline or LPS (150 microg/kg BW) followed by a 3-h blood collection every 15 min. Following blood collection, all pigs were killed and tissue was collected for mRNA analysis. Additionally, the small intestine was collected for measurement of villus height, crypt depth, and villus height:crypt depth ratio (VCR) at three sites (25, 50, and 75% of the total length). Feeding SDP resulted in reduced (P < 0.05) mRNA expression of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) mRNA in the adrenal gland, spleen, hypothalamus, pituitary gland, and liver. Additionally, expression of IL-6 mRNA was reduced (P < 0.05) in the spleen and pituitary gland for pigs fed SDP. For pigs fed the diet with SDP, LPS administration did not affect (P > 0.10) cytokine mRNA expression, whereas LPS reduced expression of TNF-alpha mRNA in the spleen and IL-1beta mRNA in the adrenal gland, spleen, and thymus for pigs fed the diet without SDP. For pigs fed the diet with SDP, LPS caused serum TNF-alpha to increase 150-fold compared to a 60-fold increase for pigs fed the diet without SDP. Similarly, interferon-gamma (IFN-gamma) increased 110-fold for pigs fed the diet with SDP compared to a 16-fold increase for pigs fed the diet without SDP. For pigs fed the diet with SDP, LPS caused major villus atrophy, whereas for pigs fed the diet without SDP, LPS had no effect on intestinal morphology. These results demonstrate that the basal activation of the immune system appears to be less for pigs fed the diet with SDP compared to pigs fed the diet without SDP after weaning. Additionally, for pigs fed the diet with SDP, there appeared to be an overresponse of the immune system following LPS administration, which resulted in major damage to the mucosa of the gastrointestinal tract.

Adrenal Glands↗

Spleen cells of non-obese diabetic mice fed with pig splenocytes display modified proliferation and reduced aggressiveness in vitro against pig islet cells.

A new means of modifying xenogeneic reaction to pig islet cells, which involves pre-feeding with pig spleen cells, was investigated for the first time in the non-obese diabetic (NOD) mouse. Compared with controls, mice fed with pig spleen cells displayed much higher splenocyte proliferation in response to pig spleen and islet cells (p < 0.0001). This enhanced proliferation was specific for the species providing the fed cells. Positive relationships (p < 0.01) were found between increased splenocyte proliferation in response to pig spleen or islet cells and the number of cells per feeding or the number of daily feedings. Concomitantly, while co-incubation with splenocytes from control mice led to inhibition of both basal and stimulated insulin releases from pig islet cells (p < 0.001), this aggressiveness was abolished (p < 0.001) after co-culture with splenocytes from mice fed with pig spleen cells. The proliferative responses of splenocytes from fed or control mice to pig islet or spleen cells were abolished after removal of plastic-adherent cells, indicating that the major indirect pathway of T-cell activation was unchanged by pig spleen cell feeding. The main T-splenocyte subsets involved were restricted to MHC class II as they did not proliferate in the presence of monoclonal antibodies (mAbs) directed at I-A molecules. In mice fed with pig spleen cells, as well as in control mice, the blocking of CD4 + T cells with mAbs led to abolition of proliferation (p < 0.002), while the blocking of CD8 + led to a less marked effect. However, an increase in the blocking effect of anti-CD8 mAbs was noted in mice fed with pig spleen cells (p < 0.02). In control mice, the main splenocyte subset involved during proliferation in response to pig islet cells was Thl, since interferon gamma (IFNgamma) production increased significantly (p < 0.01) while that of interleukin-10 (IL-10) increased only slightly. The main change observed in mice fed with pig spleen cells was a marked increase in basal IL-10 production (p < 0.01) and the basal IL-10/IFNgamma ratio (p < 0.001). It seems likely that feeding with pig spleen cells shifted the Th1/Th2 balance towards a dominance of Th2-type class II-restricted CD4 + T cells, which may have been conducive to activating CD8 + suppressor T cells. In any event, oral administration of pig cells modified xenogeneic cellular response, which may have implications for xenografts of pig islets. In a more general sense, physiological feeding of cells from xenogeneic species would appear to have certain effects on the immune system.

Administration, Oral↗

Effects of proportion of pigs removed from a group and subsequent floor space on growth performance of finishing pigs.

The effects of the proportion of pigs removed from an established group and subsequent floor space on growth performance during the final 19 d of the finishing period were evaluated using 28 pens of mixed-sex crossbred pigs (mean initial BW = 113.4 +/- 0.57 kg; n = 1,456; approximately 52 pigs per pen). A randomized block design was used with four pig-removal treatments: 1) 0% of pigs removed [Control], 2) approximately 25% of pigs removed, 3) approximately 50% of pigs removed, and 4) approximately 50% of pigs removed and floor and feeder spaces/pig decreased to equal those of Control. A block consisted of four pens with the same number of pigs and sex ratio per pen and with similar initial BW. Pens within blocks were randomly allocated to treatment, and the heaviest animals were removed from Treatments 2, 3, and 4 at the start of the study. Group size and floor space/pig for Treatments 1, 2, 3, and 4 were 52 and 0.65 m(2), 39 and 0.87 m(2), 26 and 1.30 m(2), and 26 and 0.65 m(2), respectively. Each pen contained a six-place feeder that provided 212 cm of total trough space; however, only three-places were accessible to pigs on Treatment 4. Compared with Controls, removing 25 or 50% of pigs resulted in increased (P < 0.001) ADG by 20.6 and 21.0%, ADFI by 10.8 and 7.9%, and G:F by 7.7 and 14.3%, respectively. Average daily gain by pigs on Treatment 4 (50% removal rate and decreased floor and feeder spaces) was greater (P < 0.05) than that of the Controls, but lower (P < 0.05) than that of Treatment 3 pigs (50% removal rate, no adjustment in floor or feeder spaces). No differences were observed among treatments for either morbidity or mortality. These results indicate that removing 25 or 50% of the heaviest pigs from groups of finishing pigs increased growth rate of the remaining pigs, and that the improved performance was only partly due to increased floor and feeder spaces.

Adipose Tissue↗

Effect of spray-dried plasma and lipopolysaccharide exposure on weaned pigs: II. Effects on the hypothalamic-pituitary-adrenal axis of weaned pigs.

A study was conducted with 20 weaned barrows (14 d, 4.98 +/- 0.21 kg) to determine the effect of feeding spray-dried plasma (SDP) after weaning on the pig's stress response to a lipopolysaccharide (LPS) challenge. After weaning, pigs were fed a diet containing 0 or 7% SDP for 7 d. On d 6 after weaning, all pigs were nonsurgically fitted with a jugular catheter. On d 7 after weaning, the pigs were given i.p. injections of either saline or LPS (150 microg/kg BW) followed by serial blood collection every 15 min for a 3-h period. Following the 3-h blood collection, all pigs were killed and tissue was collected for mRNA analysis. Pig weight on d 7 after weaning was not affected by dietary treatment (P > 0.21). Pigs fed the diet with SDP had lower (P < 0.05) levels of hypothalamic corticotropin-releasing hormone (CRH) mRNA, pituitary gland CRH receptor mRNA, and adrenal gland adrenocorticotropin-releasing hormone (ACTH) receptor mRNA. Dietary treatment did not affect pituitary gland proopiomelanocortin (POMC) mRNA. No effect of LPS treatment was observed in any of the mRNA levels examined. For both serum ACTH and cortisol, there was a significant diet x LPS treatment interaction (P < 0.01) such that both the ACTH and cortisol responses to the LPS challenge were greater in the pigs fed the diet with SDP than in the pigs fed the diet without SDP. For pigs given the saline injection, diet did not affect basal serum cortisol concentration; however, basal serum ACTH concentration was lower in those pigs fed the diet with SDP (P < 0.0001). A diet x LPS treatment interaction (P < 0.024) was observed for adrenal gland mRNA expression for steroidogenic acute regulatory (StAR) protein such that the LPS-induced increase in StAR mRNA was greater in the pigs fed SDP than in pigs fed the diet without SDP. These results demonstrate that pigs fed a diet with SDP have an increased activation of the pituitary-adrenal axis following an LPS challenge compared to pigs fed a diet without SDP.

Animals↗

[Natural human anti-pig agglutinins. Role in the hyperacute rejection of xenogeneic graft in the pig-human donor-recipient combination].

Binding of human natural xenoantibodies to the graft endothelial cells is considered as one of the initial steps in the hyperacute rejection of a pig liver by human. In addition to binding to endothelial cells, human xenoantibodies also agglutinate pig red blood cells. We have therefore compared the anti-pig hemagglutinin titers with the cytotoxic activity to pig endothelial cells of a series of human sera. In order to determine the localisation of the human xenoantibodies fixation in the pig liver, we have studied by immunoperoxidase staining, deposits of IgG, IgM and IgA, using an ex vivo model of isolated pig liver perfused with human blood. Moreover, we have measured hemagglutinin titers in the perfusate along the perfusion. We have found a strong correlation between the anti-pig hemagglutinin titers and the anti-pig endothelial cell cytotoxic activity of human serum. Anti-pig hemagglutinin become undetectable in the perfusate within 4 minutes of perfusion of the pig liver with human blood. The sinusoids in the pig liver were the only structures with positive peroxidase staining. It is therefore suggested that human anti-pig hemagglutinins are in great part the same than human anti-pig endothelial cell cytotoxic antibodies. Pig red blood cells could be used to easily detect anti-pig xenoantibodies in human, and to adsorb xenoantibodies from the recipient before a xenograft.

Agglutinins↗

gamma-Aminobutyric acid inhibition of histamine-induced inositol phosphate formation in guinea-pig cerebellum: comparison with guinea-pig and rat cerebral cortex.

1. gamma-Aminobutyric acid (GABA), 2 mM, inhibited basal accumulation of [3H]-inositol monophosphate ([3H]-IP1) in lithium-treated slices of guinea-pig cerebellum preincubated with [3H]-inositol. In contrast, 2 mM GABA stimulated the accumulation of [3H]-IP1 in rat cerebral cortical slices over a 60 min incubation period, but had no significant effect in slices of guinea-pig cerebral cortex. The estimated IC50 for the inhibitory action of GABA in guinea-pig cerebellar slices was 0.52 +/- 0.12 mM. 2. GABA inhibited histamine-induced [3H]-IP1 accumulation in guinea-pig cerebellar slices in a non-competitive manner. The best-fit value for the maximum level of inhibition was 74 +/- 6%. The estimated IC50 for GABA was 0.77 +/- 0.15 mM and was not significantly different from the IC50 for inhibition of the basal accumulation of [3H]-IP1. The response to histamine in guinea-pig and rat cerebral cortical slices was also inhibited by 2 mM GABA. 3. In guinea-pig cerebellar slices 2 mM GABA potentiated histamine-induced [3H]-inositol bisphosphate ([3H]-IP2) accumulation, whereas in both guinea-pig and rat cerebral cortex the effect was inhibition. 4. Isoguvacine and muscimol, GABAA-selective agonists, and (-)-baclofen, GABA(B)-selective, had no significant effect on basal or histamine-stimulated accumulation of [3H]-IPs in guinea-pig cerebellar slices. (-)-Baclofen had only a weak inhibitory effect on [3H]-IP1 accumulation in guinea-pig-cerebral cortex (16 +/- 6% inhibition with 10 microM (-)-baclofen), whereas in rat cerebral cortex (-)-baclofen mimicked the inhibitory effect of GABA. 5. Nipecotic acid (1 mM) had qualitatively similar effects to those of 2mm GABA in guinea-pig cerebellar slices. 6. The competitive GABA uptake inhibitors SK&F 89976-A, SK&F 100330-A and SK&F 100561-A were potent histamine H,-receptor antagonists, as indicated by the inhibition of [3H]-mepyramine binding to homogenates of guinea-pig cerebellum and cerebral cortex. 7. GABA (2 mM) caused a small inhibition (12 + 3%) of [3H]-inositol incorporation into total inositol phospholipids in guinea-pig cerebellar slices, as in rat cerebral cortical slices, whereas 0.2mm histamine caused a small stimulation (15 + 4%). In the presence of both GABA and histamine, [3H]-inositol incorporation was unchanged from basal (101 + 5%). 8. GABA also inhibited [3H]-IP1 formation induced by endothelin-1 in guinea-pig cerebellar slices and increased, but not significantly, the amount of [3H]-IP2 accumulated. This, taken with the inhibitory effect on basal and histamine-stimulated accumulation, suggests that the action of GABA in guinea-pig cerebellar slices may be non-selective and may not be exerted through a specific GABA receptor.

Animals↗

Obese pigs fed a high cholesterol diet from birth to 2 months are less susceptible than lean pigs to atherosclerosis.

Dietary cholesterol in infancy may alter cholesterol metabolism and the propensity to develop atherosclerosis. This study examined the effects of a 1% cholesterol diet (HC) vs. a no-cholesterol diet (NC) during the first 2 mo of life on pigs selectively bred for leanness or obesity. Three lean and three obese pigs received the no-cholesterol diet, and four lean and four obese pigs received the 1% cholesterol diet from d 1. Lean and obese pigs fed the no-cholesterol diet showed no increase in serum lipid concentrations, nor did they develop atherosclerosis. Obese pigs fed the 1% cholesterol diet developed significantly higher serum total cholesterol (TC) and high density lipoprotein cholesterol (HDL-C) at 35 d than lean pigs fed the 1% cholesterol diet. By d 55, only HDL-C remained significantly higher in the obese pigs, resulting in a higher (P < 0.1) TC/HDL-C ratio in the lean pigs. Atherosclerotic plaque formation in the aorta was more extensive in the lean pigs. Cholesterol synthesis measured in vivo and at termination was equally suppressed in lean and obese pigs fed the 1% cholesterol compared with pigs fed the no-cholesterol diet. We conclude that genetic differences in the response of these lean and obese pigs to a high cholesterol diet render obese pigs less susceptible to atherosclerosis despite higher serum TC concentrations. The persistent elevation of HDL-C in obese pigs is the most likely mechanism of protection.

Aging↗

Effect of chlortetracycline on the spread of R-100 plasmid-containing Escherichia coli BEL15R from experimentally infected pigs to uninfected pigs and chicks.

Swine from two herds with different histories of antibiotic exposure were fed diets containing 0 or 55 mg of chlortetracycline (CTC)/kg. One of five pigs in each herd-diet treatment group was infected orally with Escherichia coli strain BEL15R that was resistant to nalidixic acid (NA), chloramphenicol (C), streptomycin (S), sulfamethizole (TH) and tetracycline (TE). Effects of CTC on the quantity and duration of fecal shedding of E. coli BEL15R and on the transmission of strain BEL15R and its R-100 plasmid from infected pigs to uninfected pigs and chicks were determined. Quantity and duration of shedding were greater in infected antibiotic-herd pigs than in infected nonantibiotic-herd pigs. Feeding on CTC increased the duration of shedding in infected pigs from both herds. Strain BEL15R colonized and was shed in one uninfected antibiotic pig in each treatment group, but it did not colonize in any of the uninfected nonantibiotic-herd pigs or in the uninfected chicks. In vivo transfer of resistance to C, S, TH and TE occurred in the infected antibiotic-herd pigs but not in the infected nonantibiotic-herd pigs. Transfer of the R-100 plasmid occurred from the infected to the uninfected antibiotic-herd pigs and to the uninfected chicks housed near the antibiotic-herd pigs fed CTC, but not to the chicks housed with the antibiotic-herd pigs fed the control diet. No transfer of resistance occurred from the infected nonantibiotic-herd pigs fed either CTC or control diet.

Animals↗

Isolation of cilia-associated respiratory (CAR) bacillus from pigs and calves and experimental infection of gnotobiotic pigs and rodents.

Filamentous, gram-negative bacteria morphologically similar to cilia-associated respiratory (CAR) bacillus of rodents and rabbits were isolated from the tracheas of 5 pigs and 4 calves. All pigs but none of the calves had histologic lesions of chronic tracheitis. In silver-stained histologic sections, CAR bacilli were adhered to the tracheal epithelium of each pig but were not found in the calves. Like CAR bacillus of rats, the bacteria displayed gliding motility and grew only in cell culture or cell culture medium supplemented with fetal serum. Initially, all isolates were contaminated by Mycoplasma spp. This contamination was eliminated from 4 pig isolates by limiting dilutions, and mycoplasma-free isolates were used to intranasally inoculate gnotobiotic pigs and CAR bacillus-free mice and rats and to immunize guinea pigs. The gnotobiotic pigs remained healthy, and when they were necropsied 4 and 7 weeks after infection no macroscopic or microscopic lesions were found in the respiratory tract. However, CAR bacillus was isolated at both times from the nasal cavities and tracheas of inoculated pigs, and the ciliated tracheal epithelium of infected pigs necropsied 7 weeks after infection was colonized by low numbers of CAR bacillus-like bacteria. The rats and mice remained healthy through week 12 postinoculation, and evidence of short- or long-term colonization was not detected by histologic examination or culture. When used as primary antibody for immunohistochemical staining, sera from guinea pigs immunized with pig CAR bacillus specifically stained CAR bacilli colonizing the respiratory epithelium of naturally infected pigs, whereas sera collected prior to immunization failed to react with the bacteria. These results indicate that CAR bacilli are unlikely to be primary pathogens of pigs or cattle and that rodents do not act as reservoirs.

Animals↗

cDNA cloning and expression of guinea pig neutrophil attractant protein-1 (NAP-1). NAP-1 is highly conserved in guinea pig.

cDNA for neutrophil attractant protein-1 (NAP-1, also known as IL-8) was cloned from Con A-stimulated guinea pig spleen cells with human NAP-1 cDNA as a probe. Guinea pig NAP-1 cDNA is composed of 1433 bp with an open reading frame which encodes for a 101-amino-acid protein. Guinea pig NAP-1 had 70% amino acid sequence similarity to human NAP-1, which was much higher than a similarity between human and guinea pig monocyte chemoattractant protein-1 (MCP-1) (56%). Nucleotide sequence similarity within the coding region was 75%. To confirm its biological activity in guinea pig, recombinant guinea pig NAP-1 was expressed in COS-7 cells then purified. N-terminal sequence analysis gave two different N-termini at position 23 (Met) or 24 (Val). The two proteins showed their peak activity for guinea pig neutrophils at the concentration of 1 microgram/ml (10-7 M). Despite its high similarity to human NAP-1, the responsiveness of human neutrophils to guinea pig NAP-1 was minimum. Recombinant guinea pig NAP-1 caused strong neutrophil infiltration after intradermal injection into guinea pig skin. Since guinea pig is classified as a rodent, it was of interest to know whether human NAP-1 cDNA hybridizes to genomic DNA of other rodents such as mouse or rat, in which a NAP-1 homologue has not been found. Under low stringency conditions, human NAP-1 cDNA hybridized to human, rabbit, and guinea pig DNA, but not to mouse or rat DNA. Unlike NAP-1, human MCP-1 cDNA hybridized to genomic DNA of rabbit, guinea pig, mouse, and rat; MCP-1 cDNA have been cloned from these species. The apparent absence of a NAP-1 gene in mouse or rat makes this chemoattractant unique among the members of the protein family to which NAP-1 and MCP-1 belong.

Amino Acid Sequence↗

Pseudorabies virus is transmitted among vaccinated conventional pigs, but not among vaccinated SPF pigs.

Whereas the reproduction ratio (R) of pseudorabies virus (PRV) in vaccinated specific pathogen free (SPF) pigs without maternally derived antibodies under experimental conditions has repeatedly been shown to be significantly below 1, R in vaccinated conventional pigs in the field with maternally derived antibodies was significantly above 1. To exclude the difference in husbandry conditions as a cause for this discrepancy, we quantified and compared the transmission of PRV in both groups under identical experimental conditions. Whereas none of the SPF sentinel pigs became infected (R=0, significantly<1), all conventional sentinel pigs did become infected (R=2.5, significantly>1). Moreover, only one SPF pigs shed virus in saliva, the mean cumulative titre being almost a 100-fold less than in conventional pigs (17 pigs, P=0.003). In addition, the mean proliferation of peripheral blood lymphocytes in response to PRV antigens was significantly higher in SPF pigs than in conventional pigs at all points studied (P<0.0001). Moreover, the virus-neutralising antibody titre after vaccination was significantly higher in SPF pigs than in conventional pigs. We conclude that the discrepancy in transmission between vaccinated SPF pigs and vaccinated conventional pigs cannot be attributed to the experimental conditions.

Animal Husbandry↗

Pig but not human interferon-gamma initiates human cell-mediated rejection of pig tissue in vivo.

Split-thickness pig skin was transplanted on severe combined immunodeficient mice so that pig dermal microvessels spontaneously inosculated with mouse microvessels and functioned to perfuse the grafts. Pig endothelial cells in the healed grafts constitutively expressed class I and class II major histocompatibility complex molecules. Major histocompatibility complex molecule expression could be further increased by intradermal injection of pig interferon-gamma (IFN-gamma) but not human IFN-gamma or tumor necrosis factor. Grafts injected with pig IFN-gamma also developed a sparse infiltrate of mouse neutrophils and eosinophils without evidence of injury. Introduction of human peripheral blood mononuclear cells into the animals by intraperitoneal inoculation resulted in sparse perivascular mononuclear cell infiltrates in the grafts confined to the pig dermis. Injection of pig skin grafts on mice that received human peripheral blood mononuclear cells with pig IFN-gamma (but not human IFN-gamma or heat-inactivated pig IFN-gamma) induced human CD4(+) and CD8(+) T cells and macrophages to more extensively infiltrate the pig skin grafts and injure pig dermal microvessels. These findings suggest that human T cell-mediated rejection of xenotransplanted pig organs may be prevented if cellular sources of pig interferon (e.g., passenger lymphocytes) are eliminated from the graft.

Animals↗

Digestion in the pig between 7 and 3 d of age. 2. The digestion of dry matter and the pH of digesta in pigs given milk and soya-bean proteins.

1. In two separate experiments, forty-four pigs weaned at 4--5 d of age were given diets containing milk or soya-bean proteins until slaughtered at 14, 28 or 35 d of age. 2. The retention times of digesta in the stomach and the entire gastro-intestinal tract did not differ between pigs given pelleted diets containing milk or soya-bean protein. 3. Digesta retention times in the stomach were shorter (61 v. 145 min) in pigs given a milk-protein diet in a liquid form than in pigs given the same diet in a pelleted form. The retention times in the whole gastro-intestinal tract were 42 . 8 v. 29 . 6 h on the respective diets. 4. The total retention time of digesta in the gastro-intestinal tract increased with age of pigs between 14--35 d of age when given pelleted diets but not when given a liquid diet. 5. The daily amount of digesta flowing through the anterior small intestine was unaffected by the source of protein. Greater endogenous secretions into the anterior small intestine were observed when pigs were fed ad lib. than when fed at 2 g nitrogen (45 d dry matter (DM)/kg live weight 0 . 75 per d. 6. The apparent digestion of DM to the ileum of pigs given milk, isolated soya-bean protein (ISP) (Promine D) or soya-bean meal (SBM) protein at 28 d of age was 0 . 826, 0 . 825 and 0 . 644 respectively. 7. The apparent digestion of DM to the ileum of pigs given ISP (Supro 610) significantly increased with age of pigs from 0 . 851 at 14 d of age to 0 . 883 at 35 d of age. No increase was observed for pigs given milk protein. 8. Differences in the digestibility of DM between protein sources were greater to the ileum than over the entire gastro-intestinal tract. The greatest increase in DM digestion from the ileum to the faeces occurred in pigs given SBM (0 . 644--0 . 874). 9. Neither the protein source nor age of pigs influenced gastric pH values, the mean value being 4 . 05. pH increased along the small intestine and was not affected by the protein source of age of pig.

Aging↗

Intestinal macromolecular transmission in underprivileged and unaffected newborn pigs: implication for survival of underprivileged pigs.

Passive immunisation of the newborn pig depends upon intestinal macromolecular transmission to the blood, which ceases within 18 to 36 hours after birth in fed pigs. Macromolecular transmission in newborn, unsuckled pigs was tested by gavage feeding bovine serum albumin and fluorescein isothiocyanate-labelled dextran (molecular weight 70 kDa) together with sow colostrum. Serum levels measured four hours after feeding showed that underweight pigs under 1.0 kg, both apparently normal and weak, had a greater capacity for transmission than did apparently normal pigs over 1.0 kg (unaffected). Weak pigs 1.0 kg, or more, had a transmission equivalent to that of unaffected pigs, indicating that they may be otherwise normal animals affected by birth. Splayleg pigs and their clinically unaffected littermates also showed a greater capacity for transmission than unaffected pigs; splayleg apparently involves the entire litter, although the expression of the syndrome varies. The capacity for transmission in the unaffected pig increased with litter size and decreased with an increase in weight and age after birth. In addition, it was shown that transmission was greater in pigs with higher glucose, insulin and alpha-fetoprotein concentrations and lower haemoglobin, cortisol and alpha 2-macroglobulin f concentrations at birth. It was concluded that immature pigs have a greater capacity for intestinal macromolecular transmission at birth than unaffected pigs over 1.0 kg. Since the capacity for macromolecular transmission decreases with age, the probability of their survival depends very much on the husbandry practices in the herd.

Animals↗

Characterization of the muscarinic receptor subtype(s) mediating contraction of the guinea-pig lung strip and inhibition of acetylcholine release in the guinea-pig trachea with the selective muscarinic receptor antagonist tripitramine.

1. The muscarinic receptor subtypes mediating contraction of the guinea-pig lung strip and inhibition of the release of acetylcholine from cholinergic vagus nerve endings in the guinea-pig trachea in vitro have previously been characterized as M2-like, i.e. having antagonist affinity profiles that are qualitatively similar but quantitatively dissimilar compared to cardiac M2 receptors. The present study sought to establish definitely the identity of these receptor subtypes by using the selective muscarinic receptor antagonist, tripitramine. Guinea-pig atria and guinea-pig trachea (postjunctional contractile response) were included for reference. 2. It was found that tripitramine antagonized methacholine-induced contractions of the guinea-pig lung strip with pKB value of 8.76 +/- 0.05. Both the parallel shifts of the concentration-response curves and the slope of the Schild plot begin not significantly different from unity (when antagonist preincubation was for 2 h) indicated the involvement of a single population of receptors in the contractile response. From the pKB values obtained with tripitramine and a range of other selective muscarinic receptor antagonists (cf. Roffel et al., 1993), this single population of receptors can only be classified as M2-like. 3. Tripitramine antagonized methacholine-induced chronotropic and inotropic responses in guinea-pig right and left atria with apparent pKB values of 9.4-9.6. However, such values were only obtained when antagonist preincubation was relatively long and/or antagonist concentration relatively high (e.g with 1 h at 100 or 300 nM but 3 h at 30 nM). It thus appears that low concentrations of tripitramine do not readily equilibrate with M2 receptors in guinea-pig atria nor with M2-like receptors in the guinea-pig lung strip. 4. Tripitramine increased electrical field stimulation-induced cholinergic twitch contractions in guinea-pig trachea in concentrations of 0.3-100 nM, by blocking prejunctional muscarinic inhibitory autoreceptors; with higher concentrations, twitch contractions were progressively diminished, as a result of blocking postjunctional M3 receptors (apparent pKB value 6.07 +/- 0.15). The pEC20 value (-log concentration that increases twitch by 20% maximum) was 8.29 +/- 0.08, which would suggest that M4 receptors are involved in this response. 5. Oxotremorine-induced inhibition of the release of prelabelled [3H]-acetylcholine from guinea-pig trachea, under conditions where there is no auto-feedback, was blocked by tripitramine (2 h preincubation) with a pKB value of 8.56 +/- 0.06. The slope of the corresponding Schild plot was not significantly different from unity, which together with the parallel shifts of the concentration-response curves indicated the involvement of a single muscarinic receptor subtype. 6. Since the pKB value for tripitramine at prejunctional receptors in guinea-pig trachea is in between the affinities towards M2 and M4 receptors, correlation plots were constructed to compare the pKB values obtained with tripitramine and a range of other selective muscarinic receptor antagonists (cf. Kilbinger et al., 1995) to reported affinities at M1-M4 receptors. This showed rather similar distribution patterns of the data points around the line of equality in the case of M2 and M4 receptor subtypes. However, the correlation coefficient was markedly better for M2 (0.9667) than for M4 (0.5976). Since recent evidence suggests that M4 receptors are not expressed in cholinergic nerves from guinea-pig trachea, it is concluded that prejunctional muscarinic autoinhibitory receptors in this tissue exhibit an atypical M2 type character, with a pharmacological profile distinct from cardiac M2 receptors.

Acetylcholine↗