Immunomodulatory Effects And Mechanism Of IPEC-J2 Cells By Propionibacterium Acnes

ObjectivePropionibacterium acnes as the first colonization microbes in mammalian gut. The interactions between the intestinal cells and Propionibacterium acnes and its influence to the intestinal cells has not been studied. In this study, viable and inactivated Propionibacterium acnes were co-cultured with IPEC-J2 cells. Observing the biological effects on IPEC-J2 cells acted by viable and inactivated of Propionibacterium acnes. To investigate the role played by Propionibacterium acnes in the gut.MethodsThrough LDH assay to detect the membrane injury rate of IPEC-J2 cell after co-cultured with viable and inactivated of Propionibacterium acnes; Changes in cell morphology was observed under the upright microscope after HE staining; Detecting the m RNA expression changes of tight junction protein ZO-1 and occludin through Real-Time PCR.Using the MTS method and trypan blue staining to detect the activity and survival of IPEC-J2 cell after co-cultured with viable and inactivated of Propionibacterium acnes; Morphological changes were observed by inverted microscope; Detecting the m RNA expression changes of three kinds of TLR receptors, five kinds of proinflammatory cytokines and two kinds of anti-inflammatory cytokine by Real-Time PCR.ResultsViable Propionibacterium acnes play a certain injury on IPEC-J2 cell, and the extent of damage was increased accompanied with the increasing concentration; HE staining showed that high concentrations of viable Propionibacterium acnes can expand the membrane gap of IPEC-J2 cell; Real-Time PCR result showed that viable Propionibacterium acnes can significantly reduce ZO-1 m RNA expression; but it did not play an effect on the expression of occludin m RNA.High concentrations(1×108 CFU/m L)of viable Propionibacterium acnes can obviously lower the activity and survival of IPEC-J2 cell. Nuclear shrinkage, highlight can be observed in the cell under inverted microscope. Real-Time PCR result showed that viable Propionibacterium acnes can upregulated the expression of pro-inflammatory cytokines, but the expression of anti-inflammatory have not changed. We can detect the expression of IL-10 m RNA in the IPEC-J2 cell. At the same time, viable Propionibacterium acnes can raise the expression of TLR9 m RNA, but it did not affect the gene expression of TLR2 and TLR4. Inactivated Propionibacterium acnes didn’t have any effect on the above experiments. ConclusionIn summary, higher concentrations of viable Propionibacterium acnes can affect tight connection function of IPEC-J2 cells by downregulating the expression of ZO-1 m RNA. Upregulating the expression of proinflammatory cytokines to change the dynamic balance of pro and anti inflammatory cytokines in IPEC-J2 cells. The broader cell membrane space and imbalanced expression of inflammatory factors maybe can induced inflammatory response and eventually leads to cell death. This series of biological responses may be mediated by activation of TLR9 receptors. But the mechanism is not clear, more research is needed.