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01.02.2013 | Original Article | Ausgabe 2/2013

Archives of Virology 2/2013

The role of the toll-like receptor TLR4 in hepatitis B virus-associated glomerulonephritis

Zeitschrift:
Archives of Virology > Ausgabe 2/2013
Autoren:
Yi Zhou, Nan Zhu, Xuan Wang, Ling Wang, Li-jie Gu, Wei-jie Yuan

Abstract

Toll-like receptor 4 (TLR4) plays an important role in innate immunity. The aim of our study was to detect the expression of TLR4 in HBV-associated glomerulonephritis (HBV-GN) and explore the pathogenesis of TLR4 in inhibition of HBV replication in kidney. Immunohistochemical methods were used to detect the distribution of immunoglobulin, HBsAg, HBcAg and TLR4. Because TLR4 was mainly distributed in renal tubules and interstitial spaces, we used human proximal tubular epithelial cells (HK-2) as target cells, which were cultured with HBV-DNA-positive serum. Cells were divided into four groups: A, a normal control group; B, an HBV-induced group; C, an HBV and 10 μg/ml LPS (TLR4-stimulating factor) group; and D, an HBV and 5 μg/ml CLI095 (TLR4 inhibitor) group. The morphology of HK-2 cells was observed by microscopy, and the expression of α-SMA was examined by immunofluorescence before and after culturing with HBV-DNA-positive serum. MTT was used to detect HK2 proliferation. The expression of TLR4 protein was detected by immunofluorescence and western-blotting assays, HBsAg and HBeAg levels in supernatants were measured by ELISA, and the expression of HBV-DNA was measured by semi-quantitative reverse transcription-PCR (RT-PCR). The immunohistochemical staining for TLR4 in the normal control group was negative. The expression of TLR4 in HBV-GN was significantly higher than in the HBV-antigen-positive and -negative primary glomerulonephritis (PGN) groups and was mainly distributed in renal tubules and the interstitial area, where the distribution of HBsAg had similar intensity. The expression of TLR4 was significantly associated with renal tubular and interstitial lesions. In an in vitro study with prolonged infection with HBV-DNA-positive serum, more irregularly shaped cells and fewer HK-2 were observed. The expression of α-SMA in the cytoplasm of HK-2 was significantly increased after HBV infection. Immunofluorescence results showed that almost no expression of TLR4 in HK2 cells cultured with normal serum (group A), while the expression of TLR4 was increased after HBV infection (group B). With increasing LPS concentration, the rate of the proliferation of HK2 cells, the levels of HBV-DNA in the supernatant, and the expression of HBsAg and HBeAg decreased. TLR4 was mainly expressed in HBV-GN in renal tubules and interstitial spaces and was significantly associated with renal tubular and interstitial lesions. The stimulation of TLR4 not only inhibited HBV replication but also inevitably induced immune injury to cells. Therefore, we speculate that TLR4 may be involved in an immune inflammatory reaction by inhibiting HBV replication in HK2 cells, which could have an antiviral effect during HBV infection in the kidney.

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