Effect Of The Function Changes Of Mitochondrion And Expression Of Pyroptosis Associated Protein On Lesions Of Liver Infected With HEV

In order to reveal the pathogenesis of hepatits E virus (HEV), the swine HEV infected Mongolian gerbil model has been constructed. According to real-time qPCR, histopathology, ultrastructural pathology, immunohistochemistry, imuno-electron microscope, western-blotting, ELISA, and the other methods, the distribution of HEV RNA and HEV antigen in serum, fecal, and tissues of Mongolian gerbil has been detected in-depth and systematically. And the effect of the structure and function of liver and the hepatocyte mitochondria of Mongolian gerbil which has been infected by HEV has been observed. What’s more, the location and the quantitative research of the key proteins of mitochondrial apoptosis pathway and pyroptosis have been conducted. The results are as follows.1. Real-time qPCR was used to detect the the distribution of HEV RNA in serum, fecal, and tissues of Mongolian gerbil that had been infected by HEV. The result showed that HEV RNA replicated in serum, fecal, and tissues in different levels at different time points. In intestine, HEV RNA could be detected for the longest time, and the copies of HEV in intestine were also the highest. According to the detection of HEV antigen expression, HEV antigen expressed at different levels. The HEV antigen expression in liver, spleen and intestine last the longest. HEV-ORF2was expressed at the highest amount at28dpi in liver, and21dpi in intestine. HEV-ORF3was expressed at the highest amount at14dpi in liver, and42dpi in intestine.2. The concentrations of transaminase (ALT and AST) in HEV infected group were significantly higher than control group from14dpi to42dpi (p＜0.01). The concentration of T-BIL was obviously higher than control group from7dpi to56dpi (p<0.05). The hepatocyte was swelling, and the cytoplasm was colored pale due to the loss of the cytoplasm. The disorder arrangement and necrosis of hepatocyte had been observed. There were lymphocytes infiltration in portal tract. According to the observation of electron microscope, the electron density and the count of organelle decreased in cytoplasma. The mitochondria swollen, decrease of cristae and the swollen of endoplasmic reticulum had also been observed. All of the changes caused by HEV infection demonstrated that the lesions of liver and hepatocyte mitochondria of Mongolian gerbil infected by HEV. However, there was no differences that the bases of CytB, CytC, and D-Loop genes between the inoculated group and control group.3. After infection of HEV, the expression of ATP synthetase obviously decreased (p<0.05). At21dpi,28dpi, and42dpi, the concentration of ATPase significantly decreased (p<0.01). The activity of SOD in inoculated group was significantly decreased (p＜0.01) and the concentration of MOD in inoculated group was significantly increased (p＜0.01). The expression of NOX4was significantly higher than control group (p<0.01) from14dpi to28dpi. The expression of CytC was lower than control (p<0.05) at the early stage infection. However, it increased at the middle and later stage of HEV infection.4. Due to the infection of HEV, the expression of Bax, Apaf-1, CytC, Caspase3and Caspase9increased significantly (p<0.05), and the ratio of Bcl-2/Bax decreased. The count of TUNEL positive cells was significantly increased (p<0.01). It revealed the activation of mitochondrial apoptosis pathway.5. The expression of Caspasel, IL-1β and IL-18in the hepatocyte of inoculated group was significantly increased than the control group (p<0.01). The pyroptosis pathway may be activated. There was no significant change of the expression of TNF-α during HEV infection. The expression of INFy was significantly higher (p<0.01). It demonstrated that HEV infection had caused the dynamic of anti-viral immunoreactions of Mongolian gerbil.In conclusion, during HEV infection, HEV RNA could replication and proliferation in serum, fecal, and the tissues and HEV antigen continued expressed in the tissues of the Mongolian gerbil. The infection of HEV caused the lesions of the structure and function of liver, induced the activation of mitochondrial apoptosis pathway and pyroptosis pathway, and activated the inflammation associated cytokine, which may be the pathogenesis of HEV infection.