Correlation Study Between Gut Flora And Metabolic Pathways In Liver Regeneration

Part Ⅰ The different biological phases during liver regenerationBackgrounds:The strong capability of regeneration allows the liver to recover to the normal volume after subjected to inflammatative or surgical insult. Currently, partial hepatectomy (PH) induced liver regeneration (LR) model is widely used for research due to its simplicity and reproductivity. However, the coccurence of injurious and proliferative responses upon PH provides the difficulty for distinguishing between the injurious or proliferative contribution of investigated parameters. Therefore, this section was designed to discriminate the different biological processes of LR.Methods:Male SD rats were subjected to either 70% liver resection, sham operation or no treatment. Samples were collected over 14 day period. Blood was sampled for biochemical examination, and liver was collected for histopathlogical, immunohistochemical and mRNA expression analysis.Results:PHx rats demonstrated mild hepatic damage as reflected by ALT, AST, total bilirubin and histological results. The serum levels of above index was elevated immediately after PH, peaked within 24h and returned to that of normal control at 72h. PHx rats showed an enhanced cell proliferation during 30-48 h. Hepatic mRNA expression of TNF-α, IL-6, HGF and TGF-β was upregulated abruptly upon PH, and peaked at 24,24,12 and 72 h respectively.Conclusions:Based on the performance of injurious and proliferative index, the biological processes of LR could be divided into three phases:initiation (12-24h), propagation (30-48h) and terminition stages (3-14d).PartⅡ Variations of the gut flora community during liver regenerationBackground:Gut flora is proposed to exert an immense impact on liver regeneration. However the role of individual bacteria species is still obscure.Methods:High-throughout pyrosequencing was used to characterize the microbial profile during liver regeneratin. Functional analysis of microbial genes were performed using PICRUSt based on 16S rRNA data.Results:PHx did not cause changes in the diversity of gut flora. However, it caused fluctuating changes in gut microbiome, which paralleled the biological processes of LR. Briefly, the enhanced cell proliferation occurring within 30-48 h was associated with a decreased ratio of Firmicutes to Bacteroidetes reflected by a reduction in Ruminococcaceae and Lachnospiraceae (within phyla Firmicutes), while an increase in Bacteroidaceae, Rikenellaceae and Porphyromonadaceae (within phyla Bacteroidetes), which was indicative of a lean phenotype. Whereas, the microbiota derived from 12-24 h and 3-14 day were characterized by elevated F/B ratios, suggesting the differing energy extract behaviors of microbiota during the course of LR. Functional changes of the shifted microbiota revealed by Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) confirmed the pyrosequencing results. The microbiome derived from hour 12 rats showed overpresentation of "lipid metabolism", "carbohydrate metabolism" and "glycan biosynthesis and metabolism". In contrast, the microbiome derived from day 2 rats was functionally unique in "replication and repair", "amino acid metabolism" and "nucleoid metabolism".Conclusions:The alterations of intestinal bacteria was an adaptive measure to fulfill the varing demands of cell proliferation and host metabolism at different biological phases of LR.PartⅢ The alteration of microbial community was associated with the variation of liver regeneration-related metabolic responsesBackground:It is well known that an augmented enery demands is imposed on host during liver regeneration. As revealed in Part Ⅱ, as a "metabolic organ", gut flora altered its biological function as reflected by the changes in functional genes after PHx. However, whether the gut flora-mediated metabolic pathways contribute to the metabolic perturbation in LR is poorly documented.Methods:UPLC/MS and multivariate statistical analysis were utilized to establish the systemic metabolic responses to LR. Besides, metabolic pathway analysis of putative biomarkers was conducted with the Metabolomics Pathway Analysis (MetPA) based on pathway library from Kyoto Encyclopedia of Genes and Genomes (KEGG) for identification of the affected metabolic pathwayResults:Based on PCA and OPLS-DA models, three clusters of hour 12-24 groups, 30-48 groups and 3-14 days were clearly separated. A number of serum perturbations were observed.84 biomarker metabolites were identified to be differentially expressed between normal control and PHx rats. Of them,15 metabolites displayed fluatuating changes throught three phases of LR, including PE (20:4/14:0), PC (22:5/15:0), PC (20:3/15:0), LysoPC (18:4), L-octanoylcarnitine, bilirubin, L-urobilin, pantothenic acid, D-leucic acid, D-glucosaminide,2,3-diaminopropionic acid,1-methylhistamine, ribitol, pimmelic acid and hypotaurin. Upon examining the dynamic pattern of these 15 metabolite, the specific pathways including glycerophospholipid metabolism, taurine and hypotaurine metabolism were identified to be attributable to the systemic alterations in LR related-metabolism. Moreover, our data demonstrated that several key functional bacteria were strongly related to the perturbations of above pathways.Conclusions:Gut flora may play a central role in manipulating metabolic responses in LR.