| Lactobacillus reuteri I5007 is a beneficial lactobacilli present in the intestines of animals.To explore the effects of Lactobacillus reuteri I5007 on the intestinal morphology,intestinal immune function and miRNA expression profile of newborn piglets,and the signal pathways involved.The experiment selected 6 litters(nine litters per nest,more than 9 pigs were excluded)healthy 1-day-old × Rong newborn piglets,divided into 2 groups of 3 litters per group: the control group was fed 2 mL 0.1% per head per day Peptone solution;Lactobacillus reuteri group was fed with 2 mL of 0.1% peptone solution dissolved Lactobacillus reuteri per head per day.Starting from the first day of birth,feeding for 20 consecutive days.At 10 and 20 days of age,the pigs were weighed on an empty stomach,2 piglets were selected from each litter,6 pigs per day in each group(6 replicates),a total of 24 pigs,blood was slaughtered from the heart,and blood was collected to determine the inflammation in the serum Factor content;intercept duodenum,jejunum,ileum and cecum at the same location,determine the morphology of duodenum,jejunum and ileum,short-chain fatty acid content of the colon,microbial flora structure of the jejunum and cecum Factor expression and miRNA expression profile.At the end of the trial,the average daily weight gain was calculated and the diarrhea rate was counted.The results show:1)Compared with the control group,feeding Lactobacillus reuteri significantly increased the daily weight gain of piglets at 11-20 days of age(P<0.05),and significantly reduced the diarrhea rate of piglets at 1-20 days of age(P<0.05).2)Feeding Lactobacillus reuteri significantly increased the ratio of ileum villi height / crypt depth in 10-day-old piglets(P<0.05),but no significant difference in duodenal,jejunum,and ileum villi height and crypt depth influences.3)Lactobacillus reuteri feeding to 10 days of age has no significant effect on piglet colon short-chain fatty acids;feeding to 20 days of age has a tendency to increase colonic acetic acid content(P<0.1),but has no significant effect on other short-chain fatty acids influences.4)Feeding Lactobacillus reuteri significantly increased the 10-day-old cecum shannon index(P<0.05);increased the proportion of pachyphyta in the 20-day-old jejunum and cecum,and reduced the Proteobacteria in 10 and 20 The proportion of old jejunum and caecum;at the genus level,L.reuteri increases the proportion of jejunum and caecum Lactobacillus and decreases the proportion of jejunum streptococcus and caecum.5)After feeding Lactobacillus reuteri to 10 days of age,the mRNA expression of Occludin in the jejunum increased significantly(P<0.01),and the protein expression of ZO-1 and Claudin-1 increased significantly(P<0.05).Ileum tight junction protein expression has no significant effect.After feeding Lactobacillus reuteri to 20 days of age,the mRNA and protein expression of jejunum ZO-1 and Claudin-1 were significantly increased(P<0.01),but there was no significant effect on the expression of Occludin;The expression increased significantly(P<0.05);the expression of MUC-2mRNA in jejunum and ileum also increased significantly(P<0.01).6)After feeding Lactobacillus reuteri to 10 days of age,the levels of inflammatory factors IL-6,IL-10 and TNF-? in the serum of piglets increased significantly(P<0.01);mRNA expression of jejunum inflammatory factors was not seen Significant effect,only the protein expression of pro-inflammatory factor IL-1? was significantly down-regulate(P<0.05);the protein expression of pro-inflammatory factor IL-1? in the ileum was significantly down-regulated(P<0.05),and the expression of IL-6 mRNA was significantly down-regulated(P<0.01),the expression of anti-inflammatory factor TGF-?mRNA was significantly increased(P<0.05).After feeding Lactobacillus reuteri to 20 days of age,the content of IL-1? in serum was significantly reduced(P<0.01),and the content of IL-6 and TNF-? both showed a downward trend(P<0.1);pro-inflammatory in jejunum The protein expression levels of factors IL-1? and TNF-? were significantly down-regulated(P<0.05),and the mRNA expression levels of proinflammatory factors IL-1?,IL-6 and TNF-? in the ileum were significantly down-regulated(P<0.05).7)After feeding Lactobacillus reuteri to 10 days of age,the TLR-4 mRNA expression of the jejunum tends to be down-regulated,and the TLR-6 mRNA expression tends to be up-regulated(P<0.1);the ileal TLR-4 protein expression is significant Down-regulated(P<0.05),TLR-9 mRNA expression increased significantly(P<0.01).When L.reuteri was fed to 20 days of age,the jejunum TLR-9 mRNA expression level increased significantly(P<0.01);the ileum TLR-4 mRNA expression level decreased significantly(P<0.05),and the TLR-2 mRNA expression level increased significantly(P<0.01);8)High-throughput sequencing results showed that after feeding piglets with Lactobacillus reuteri,7 and 9 mi RNAs were significantly differentially expressed in the jejunum at 10 and 20 days of age,respectively,and only 1 miRNA was different at both days Expression,so there are 15 differentially expressed miRNAs;the ileum has 14 and 9 mi RNAs differentially expressed at 10 and 20 days of age,respectively,and 4 miRNAs are differentially expressed at both days of age,so there are 19 differential expressions Mi RNAs.And the differentially expressed miRNAs are mostly related to the body's immune regulation.Bioinformatics analysis revealed that 15 target genes of jejunum differentially expressed miRNAs were significantly enriched in 66 pathways(P<0.05),and 19 target genes of ileum 19 differentially expressed miRNAs were significantly enriched in 48 pathways(P<0.05),the two pathways with the most target genes in the gut include the PI3K-Akt and MAPK signaling pathways that are closely related to intestinal immunity.In summary,feeding Lactobacillus reuteri from newborn piglets can improve piglet growth performance,reduce diarrhea rate,and adjust intestinal tight junction proteins,inflammatory factors and Toll-like receptors by improving intestinal morphology and flora structure To improve the intestinal immune function.It was also found that L.reuteri can cause differential expression of intestinal miRNAs.L.reuteri may stimulate MAPK and PIK3-Akt signaling pathways through differentially expressed mi RNAs to affect the intestine Immune factors,which in turn affect the health of the piglet intestines. |
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