Effects And Mechanism Of Indole-3-carbinol On Growth And Development And LPS Induced Intestinal Inflammation In Rabbits

Indole-3-carbinol（I3C） is a natural hydrolysate of glucosinolate,which has a good curative effect on cancer,obesity and inflammatory bowel disease.This study expounds on the effect and mechanism of I3C on the growth and LPS-induced small intestinal inflammation of Hyla meat rabbits through two parts:a growth experiment and an LPS challenge experiment.The growth test:The 50-day-old Hyla male rabbits（weighing 1.8±0.1 kg） were selected for the study.The pre-test period was 7 days.After the pre-test period,the meat rabbits were randomly divided into two groups:control group and I3C group,with 5 replicates in each group and 2 rabbits in each replicate.Meat rabbits were fed with basic diet,and were free to feed and water throughout the whole process.During the growth test,the meat rabbits in the I3C group took 40 mg·kg^-1·BW^-1I3C（I3C dissolved in 8%DMSO and corn oil solution）orally every day,while the meat rabbits in the control group took the same dose of solvent orally for a total of 14 days.All animals were slaughtered on the 14th day of the experiment.The results of the growth test showed that compared with the control group,I3C significantly decreased the F/G of meat rabbits（P<0.05）,promoted the synthesis of collagen genes COL1A1,COL1A2,COL4A1,COL4A2,COL5A2,COL6A2 and COL6A3 in jejunal mucosa,and the expression of collagen degrading enzyme gene inhibitors FSTL1,SERPINE1,SPARC and TIMP1（P<0.05）.Transcriptome showed that I3C significantly activated ECM receptor signaling pathway,protein digestion and absorption signaling pathway,the intestinal immune network for Ig A production signaling pathway and the staphylococcus aureus infection signaling pathway.There are both up-regulated and down-regulated genes on the Focal adhesion signaling pathway,PI3K-AKT signaling pathway and NF-κB signaling pathway.The serum metabonomics showed that I3C significantly increased the contents of serum amino acids,fatty acids and vitamins in meat rabbits.In addition,I3C exerted no significant effects on the expression of the signaling pathway-related genes AhR,LOC100328613,USF1,HSP90AB1,HSP90AA1,ARNT and ARNTL（P>0.05）.The LPS challenge test:The 50-day-old meat rabbits were randomly divided into three treatment groups according to body weight:control group,LPS group and LPS+I3C group,with 5 replicates in each group and 2 meat rabbits in each replicate.The pre-experiment period was 7 days,and the pilot period was 14 days.During this period,all meat rabbits in the treatment group were fed with basic diet.During the pilot test,the meat rabbits in LPS+I3C group took 40 mg·kg^-1I3C orally（I3C was first dissolved in DMSO and then dissolved in corn oil,the content of DMSO was8%）,and the control group and LPS group took the same dose of DMSO and corn oil solvent.On the 14th day of the experiment,the rabbits in LPS group and LPS+I3C group were intraperitoneally injected with 0.5 mg·kg^-1LPS,and the rabbits in control group were intraperitoneally injected with 0.5 mg·kg^-1normal saline.All rabbits were slaughtered after 4 hours of injection.The results of the LPS challenge experiment showed that I3C significantly inhibited the expression of pro-inflammatory genes TNF-α,IL-1β,IL-6,ICAM1,NOS2 and GRO-B（P<0.05）,reduced the swelling of jejunal villi but had no significant effect on histological morphological indexes such as jejunal villi height,villi width,crypt depth and villi height/crypt depth（P>0.05）.Transcriptome results showed that I3C alleviated the downregulation of exogenous biodegradation and metabolism,lipid metabolism,carbohydrate metabolism,cofactor and vitamin metabolism,and inhibited the NOD-like receptor signaling pathway and NF-κB signaling pathway,activated PPAR signal pathway in LPS treatment.The serum metabolome showed that the glycerol phospholipid metabolic signal pathway was enriched in the LPS challenge test,I3C improved the serum metabolites.Correlation analysis showed that lyso PC（20:5（5Z,8Z,11z,14z,17Z））and PC（16:0/24:1（15z））were significantly positively correlated with PPAR signal pathway,but negatively correlated with NOD-like and NF-κB signaling pathway,PE（20:2（11z,14z）/16:0）,PC（16:0/20:4（8Z,11z,14z,17Z））and PC（20:2（11z,14z）/14:0）contrast to them.At the same time,PPAR signal pathway have a negative correlation with NOD-like signal pathway,NF-κB signaling pathway,glycerophosphatidyl-choline metabolism and choline metabolism in cancer signaling pathway significantly（r<-0.4,P<0.05）,and positively correlated with amino acid transport and metabolism,carbohydrate transport and metabolism,energy production and conversion,and lipid transport and metabolism pathway（r>0.4,P<0.05）.The RT-PCR results showed that I3C significantly activated PPARA,PPARG,ME1,FABP1,CYP27A1 and CYP1A1 genes,and inhibited the expression of RIPK2 and NFKB1 genes induced by LPS（P<0.05）,which was consistent with the transcriptome results.In all,I3C increased the concentration of serum amino acids,vitamins and fatty acids,significantly reduced the F/G of meat rabbits through the interaction between PI3K-AKT pathway,Focal adhesion pathway and ECM receptor signal pathway,and enhanced Intestinal immune network for Ig A production signaling pathway and its related pathways.I3C alleviates LPS induced inflammatory factor expression and metabolic disorder,which is related to the activation of PPAR signaling pathway and the inhibition of NF-κB signaling pathway and NOD-like receptor signaling pathway.Some glycerophospholipids such as lyso PC（20:5（5Z,8Z,11z,14z,17Z））,PC（16:0/24:1（15z））,PE（20:2（11z,14z）/16:0）,PC（16:0/20:4（8Z,11z,14z,17Z））,PC（20:2（11z,14z）/14:0）can be used as biomarkers of inflammation and metabolic disorders,participate in the regulation of PPAR signaling pathway,NF-κB signaling pathway and NOD-like receptor signal pathway.