Immunocyte Populations Observed from Birth to Weaning in Blood, Spleen and Mesenteric Lymph Nodes of Piglets.

Susceptibility to pathogen infections and efficacy of vaccination highly depend on the immune status of the piglet. Here, we measured immunocytes in piglets from birth to weaning to elucidate how immunocyte populations change during development and are affected by weaning. Crossbred piglets were used. Suckling piglets were euthanized at 1, 7, 14, 21, 28 or 35 days old (3~4 piglets at each time point). In addition, seven piglets were weaned at 21 days old, with four being euthanized at 28 days old and the remaining at 35 days old. Piglet carcasses were dissected, and blood, mesenteric lymph nodes (MLN) and spleen were sampled. In total, seven antibodies were used to stain the immunocyte population. Dynamics of myeloid (CD3−SWC3+CD16+), natural killer (NK; CD3−SWC3−CD16+), killer T (CD3+CD8+), helper T (CD3+CD4+) and B (CD3−CD21+) cells were analyzed. Percentage of innate immunity cells such as myeloid cells declined (p < 0.05) from the first day after birth. In contrast, percentage of NK cells increased in piglets while they were still suckling. Killer T, helper T, and B cell populations increased around 2~3 weeks after birth. No significant differences in the populations of the evaluated cell types were observed between suckling and weaned piglets at least for 14 days post weaning.

Production, Absorption, and Blood Flow Dynamics of Short-Chain Fatty Acids Produced by Fermentation in Piglet Hindgut during the Suckling⁻Weaning Period.

Luminal short-chain fatty acids (SCFA) are rapidly absorbed from the intestine and subsequently utilized by the host as substrate for metabolic energy production. In pigs, the energy contribution of SCFA is thought to be 30⁻76%. However, since absorption and blood flow dynamics of SCFA in pigs, particularly during the suckling⁻weaning period, remain unclear, we aimed to elucidate these phenomena. Thirty-two piglets were used in the present work. Cecal vein blood and digesta, and portal and abdominal vein blood were sampled from suckling (7-, 14-, 21- and 28-day-old) and weaned (weaning at 21 and 28 days of age) piglets. Four piglets from each group were euthanized. SCFA concentrations in blood samples were analyzed by a highly sensitive gas chromatography-mass spectrometry technique. Age at weaning tended to affect SCFA absorption. For example, acetate and propionate concentrations in the cecal vein tended to be higher in piglets weaned at day 21 than at day 28. SCFA concentrations in the abdominal vein tended to differ from those in other veins. Mucosal gene expression analysis suggested that monocarboxylate transporter 1 and occludin were associated in absorption of SCFA from the lumen into the blood of piglets.

Gene expression profiles of CD4/CD8 double-positive T cells in porcine peripheral blood.

A characteristic subset of T cells, known as double positive T cells (DPTC) and expressing both cluster of differentiation 4 (CD4) and CD8, is observed in porcine peripheral blood. Previous studies suggested that DPTC might be memory cells. However, detailed phenotypes and functions of DPTC are yet to be fully elucidated and thus, the relatedness of DPTC with memory phenotypes remains unclear. In this study, DPTC gene expression profiles in peripheral blood were analyzed by DNA microarray in Experiment 1 and compared with those of CD4 single positive T cells (4SPTC) and CD8 single positive T cells (8SPTC). Expressions of IFNG, CCL5, NCK2, CCR2 and ITGB1 were higher than that of 4SPTC and 8SPTC. In contrast, expressions of CCR7 and SELL were lower than that of 4SPTC and 8SPTC. These results suggested that DPTC were either effector T cells or effector memory T cells (TEM ). Next, to determine whether DPTC were effector T cells or TEM , differences in the response of DPTC and 8SPTC against immunized/primed antigens were compared (Experiment 2). While DPTC showed quick elevation of IL2 and CD25 gene expressions against in vitro stimulation of primed/immunized antigens, 8SPTC did not. These results suggest that at least some DPTC likely belong to TEM .

Comparison of gene expression profiles of T cells in porcine colostrum and peripheral blood.

OBJECTIVE To compare gene expression patterns of T cells in porcine colostrum and peripheral blood. ANIMALS 10 multiparous sows. PROCEDURES Cytotoxic and CD4-CD8 double-positive T cells were separated from porcine colostrum and peripheral blood. Total RNA was extracted. The cDNA prepared from RNA was amplified, labeled, fragmented, and competitively hybridized to DNA microarray slides. The DNA microarray data were validated by use of a real-time reverse-transcription PCR assay, and expression of the genes FOS, NFKBI, IFNG, CXCR6, CCR5, ITGB2, CCR7, and SELL was assessed. Finally, DNA microarray data were validated at the protein level by use of flow cytometry via expression of c-Fos and integrin ß-2. RESULTS Evaluation of gene expression profiles indicated that in contrast to results for peripheral blood, numerous cell-signaling pathways might be activated in colostrum. Profile analysis also revealed that FOS and NFKBI (genes of transcription factors) were involved in most cell-signaling pathways and that expression of these genes was significantly higher in colostral T cells than in peripheral blood T cells. Furthermore, CCR7 and SELL (genes of T-cell differentiation markers) in colostral T cells had expression patterns extremely similar to those found in effector or effector memory T cells. CONCLUSIONS AND CLINICAL RELEVANCE All or most of the T cells in colostrum had an effector-like phenotype and thus were more activated than those in peripheral blood. This gene expression profile would enable T cells to migrate to mammary glands, be secreted in colostrum, and likely contribute to passive immunity provided by sows to newborn pigs.

Influence of weaning age on the villous height and disaccharidase activities in the porcine small intestine.

Body weight gain after weaning is correlated with villous height and disaccharidase activity. This evidence suggests that the maintenance of the small intestinal structure and function after weaning is important for the growth of piglets. We demonstrated that the influence of weaning age was obtained by disaccharidase activities and villous height in eight sections of the porcine small intestine. Therefore, we designed three weaning ages (weaned at the ages of 14, 21 or 28 days) and the piglets were slaughtered after 7 or 14 days post-weaning. The remaining suckling piglets were slaughtered at the age of 1, 7, 14, 21 and 28 days. Four piglets were slaughtered at each event; therefore, 44 piglets were used in this study. Villous height and disaccharidase activities were measured in each section of the small intestine. Early weaning such as that at 14 days had severe influence on villous and disaccharidase activities. In particular, weaning of 14-day-olds did not result in maltase activity at least 2 weeks post-weaning. Accordingly, the weaning age of crossbred piglets is recommended to be at least 21 days or more on the basis of villous height and disaccharidase activity analyses.

Weaning Markedly Affects Transcriptome Profiles and Peyer's Patch Development in Piglet Ileum.

Transcriptome analyses were conducted on the ileal mucosa of 14- to 35-day-old piglets to investigate postnatal gut development during suckling and postweaning. The transcriptome profiles of 14-day-old suckling piglets showed a considerably higher number of differentially expressed genes than did those of 21-, 28-, and 35-day olds, indicating an intensive gut development during the first 14-21 postnatal days. In addition, the analysis of biological pathways indicated that Chemotaxis Leucocyte chemotaxis was the most significantly affected pathway in suckling piglets between 14 and 21 days of age. Weaning negatively affected pathways associated with acquired immunity, but positively affected those associated with innate immunity. Interestingly, pathway Chemotaxis Leucocyte chemotaxis was found positively affected when comparing 14- and 21-day-old suckling piglets, but negatively affected in 28-day-old piglets weaned at 21 days of age, when compared with 28-day-old suckling piglets. Genes CXCL13, SLA-DOA (MHC class II), ICAM1, VAV1, and VCAM1 were involved in the pathway Chemotaxis Leukocyte chemotaxis and they were found to significantly change between 14- and 21-day-old suckling piglets and between groups of suckling and weaned piglets. The expression of these genes significantly declined after weaning at 14, 21, and 28 days of age. This decline indicated that CXCL13, SLA-DOA, ICAM1, VAV1, and VCAM1 may be involved in the development of Peyer's patches (PP) because lower gene expression clearly corresponded with smaller areas of PP in the ileal mucosa of piglets. Moreover, weaning piglets prior to a period of intensive gut development, i.e., 14 days of age, caused significant adverse effects on the size of PP, which were not reverted even 14 days postweaning.

Decreasing traits of fecal immunoglobulin A in neonatal and weaning piglets.

The concentration of fecal secretory immunoglobulin A (sIgA) in neonate and weaning piglets was measured daily from 1 day after birth to 50 days of age. The concentration of fecal sIgA started from the level of 10(4) microg/g wet feces 1 day after birth and then increased to a maximal value of up to 10(5) microg/g within a few days of birth. The values constantly declined to between 10(1) and 10(2) microg/g for the next 10 days and were relatively constant until weaning. The level of sIgA in the feces remained very low until at least 50 days of age. The vulnerability of pre- or post-weaning piglets can be explained, at least in part, by this low level of sIgA in the intestine.