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The aim of the present study was to investigate the effect of receptor for advanced glycation end products (RAGE)-specific small interfering (si)RNA on the generation of proinflammatory cytokines in primary rat hepatic stellate cells (HSCs) and hepatic fibrotic (HF) rats. The RAGE-specific siRNA expression vector pAKD-GR126 was constructed, and then transfected into primary rat HSCs. Reverse transcription- quantitative polymerase chain reaction and western blot analyses were conducted to determine the mRNA and protein expression levels, respectively, of RAGE, tumor necrosis factor (TNF)-α and interleukin (IL)-6 in the primary HSCs. In addition, a CCl4-induced Sprague Dawley (SD) rat model of hepatic fibrosis was established, and pAKD-GR126 was injected into the SD rats via the tail vein. Serum TNF-α and IL-6 concentrations were determined using radioimmunoassay. The mRNA and protein expression levels of RAGE (mRNA, F=7.791; protein, F=36.513), TNF-α (mRNA, F=474.568; protein, F=123.500) and IL-6 (mRNA, F=203.463; protein, F=320.555) in the pAKD-GR126-transfected primary HSCs were significantly reduced compared with those in the control and pAKD-NC groups (P<0.05). Serum TNF-α and IL-6 levels in the low-, medium- and high-dose pAKD-GR126 treatment groups were reduced compared with those in the fibrotic model group (TNF-α, F=416.397; IL-6, F=1,716.659; P<0.05). In summary, the RAGE-specific siRNA was able to effectively suppress the generation of the proinflammatory cytokines TNF-α and IL-6 in primary rat HSCs and HF rats. [ABSTRACT FROM AUTHOR]

Specific small interfering RNAs (siRNAs) targeting receptor for advanced glycation end products (RAGE) inhibit the expression of RAGE, α-smooth muscle actin and type I collagen in the T6 hepatic stellate cells (HSCs), indicating that RAGE is important for the activation of HSCs and the expression of collagen. The present study aimed to investigate the effect of specific siRNAs targeting RAGE on the development of hepatic fibrosis (HF), using primary rat HSCs, which were isolated and cultured in vitro. The expression vectors for specific siRNAs targeting RAGE were constructed and transfected into primary rat HSCs. Untreated and nonspecific siRNA-transfected primary rat HSCs served as controls. The expression levels of RAGE, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), transforming growth factor-β1 (TGF-β1), connective tissue growth factor (CTGF), laminin (LN), hyaluronic acid (HA) and N-terminal procollagen III propeptide (PIIINP) in primary HSCs were detected by reverse transcription quantitative polymerase chain reaction and western blotting. The mRNA and 42 kD protein expression of RAGE in the pAKD-GR126-transfected primary HSCs were significantly downregulated compared with those in the untreated and the pAKD-negative control (NC)-transfected controls. The mRNA and protein expression levels of IL-6, TNF-α, TGF-β1, CTGF, LN, HA and PIIINP in the pAKD-GR126-transfected primary HSCs were also markedly downregulated compared with those in the untreated and pAKD-NC-transfected controls. Therefore, RAGE-specific siRNAs inhibited the expression of RAGE in primary rat HSCs and inhibited the development of HF. [ABSTRACT FROM AUTHOR]

Since the receptor for advanced glycation end products (RAGE)-ligand axis has been demonstrated to be important in fibrogenesis, rat models may be used to assess whether specific small interfering RNAs (siRNAs) that target RAGE are able to reduce the progression of hepatic fibrosis. However, the effect of RAGE-targeted siRNA on established hepatic fibrosis remains to be elucidated. In the present study, RAGE-specific siRNA expression vectors were constructed prior to the animal experiment. Sprague-Dawley rats were treated initially with olive oil (2 ml/kg) or 50% CCl4 (2 ml/kg; CCl4/olive oil=1:1) twice per week for six weeks to generate the fibrosis model. The rats were then treated with phosphate-buffered saline, a RAGE-specific siRNA expression vector, at different doses or a non-specific siRNA expression vector twice weekly via tail vein injection for up to six weeks, and were sacrificed at week two, four or six. Compared with the control groups, RAGE-specific siRNA therapy significantly decreased RAGE mRNA and protein expression in rat livers (P<0.01). Following six weeks of RAGE gene-silencing treatment, the liver function, which was assessed by analyzing serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total bilirubin (TBIL), improved to varying degrees (P<0.01). The expression of nuclear factor-κB (NF-κB) significantly decreased following RAGE gene-silencing therapy (P<0.01). In addition, the serum levels of inflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), and extracellular matrix (ECM) components, including hyaluronic acid (HA), laminin (LN) and procollagen type III (PCIII) also decreased (P<0.01). Furthermore, the expression of α-smooth muscle actin (α-SMA) and collagen I, which indicate the activation of hepatic stellate cells (HSCs), were downregulated following RAGE gene-silencing therapy (P<0.01). Furthermore, the inflammatory activity grade and fibrosis stage of rat livers also significantly improved compared with the control groups following RAGE gene-silencing therapy. Specific targeting of RAGE using siRNA may inhibit RAGE gene expression effectively in the rat hepatic fibrosis model and attenuate the progression of established hepatic fibrosis. This therapeutic effect may be mediated via inhibition of the expression of NF-κB. These findings suggest that RAGE may be a new target to prevent hepatic fibrosis. [ABSTRACT FROM AUTHOR]