Effects Of Endotoxin On The Expression Of Connective Tissue Growth Factor During Experimental Hepatic Fibrosis

【Objective】To investigate the temporospatial expression patterns of CTGF mRNA and protein in rat livers with chronic fibrogenesis and to expelore its association with intestinal endotoxemia .【Methods】The fibrotic model of rats was induced by subcutaneous injection of carbon tetrachloride(CCl4).Thirty-seven male Wistar rats(200g±20g)were randomly divided into 2 groups: control groups(n=5)and experimental groups(n=32). The latter then were randomly allocated into 4 equal groups(n=8 per group). At the end of the experiment, rats were killed under pyrogen-free technics for collection of blood and liver tissue. Paraformaldehyde-fixed liver samples were embedded in paraffin and stained with hematoxylin-eosin to assess liver injury and fibrosis; Liver fibrosis was scored by image analysis of stain of sirius red and the content of hydroxyproline; Endotoxemia was measured with a Limulus Amebocyte Lysate test kit ; The intensity of the expression of CTGF protein in liver were assessed by westen-blot and the cellular localization of CTGF mRNA was studied by in situ hybridization; For identification of myofibroblasts and activated hepatic stellate cells,α-smooth muscle actin (α-SMA) immunohistochemistry was adopted.【Results】1. The levels of plasma endotoxin were markedly higher in experiment groups than in control group. 2. Collagen in liver increased gradually after injection of carbontetrachloride for 2 weeks. 3. In situ hybridization revealed CTGF transcripts primarily in actived HSCs and myofibroblasts from 2 weeks to 8 weeks after injection of carbontetrachloride,but sporadic HSCs expression is predominant in earlier period,later positive distribute relatively localized in portal area,fibrous septa; partial vascular endothelial cells and smooth muscle cells began to express siganificantly CTGF mRNA after injection of carbontetrachloride for 6 weeks; then after 8 weeks, the part of proliferating ductular epithelial cells were another main source of CTGF mRNA,whereas no transcripts were found in normal livers. 4. Western-blot analysis showed enhanced expression of CTGF protein in liver fibrosis compared with controls. 5. There was a strong correlation between liver CTGF protein and plasma endotoxin as well as the degree of fibrosis index.【Conclusions】In rat liver fibrogenesis, the visible increase of plasma endotoxin is precede to that of CTGF in liver tissue and siganificantly correlation with it , suggests that intestinal endotoxemia may one of the factors leading to CTGF overexpression and promote hepatic fibrosis.