MMP is the most critical protease which is involved in

MMP9 is the most critical protease which is involved in the degradation of ECM. TIMP-1 is an important regulator in the synthesis and degradation of ECM [8,9]. Hepatic TIMP-1 expression significantly increases in patients with liver fibrosis [10]. Serum level of TIMP-1 expression is positively associated with liver fibrosis in CHB patients [11]. Moreover, the development of hepatic fibrosis is promoted by the induction of TIMP-1 overexpression [12]. IL-32, a multi-function cytokine, could induce a lot of cytokine expressions, for example, IL-1β, IL-6, IL-8, and TNF-α. [13]. IL-32 also activates AP-1, NF-κβ, and p38MAPK signal pathways [14,15]. Hepatic IL-32 expressions are increased with the severity of liver fibrosis [16,17]. However, the mechanism of IL-32 in the pathogenesis of liver fibrosis is not thoroughly clear. Previous researches report [18,19] that there are several AP-1 L-365,260 in the promoter sequence of TIMP-1. EBV could induce TIMP-1 expression by activating AP-1 pathway [18]. Taken together, we investigated the effect of IL-32 on TIMP-1 expression by HSCs in order to clarify the mechanism of IL-32 in the pathogenesis of hepatic fibrosis in vitro in the present study.
Materials and methods
Results
Discussion In the present study, we found that IL-32 could induce TIMP-1 expression by LX-2 cells at a dose-dependent manner. IL-32 could increase TIMP-1 promoter activity and induce TIMP-1 expression by activating AP-1 signal pathway. Furthermore, the increase of TIMP-1 expression could promote the migration of LX-2 cells. TIMP-1, a broad inhibitor of collagenases, plays a vital role in liver fibrogenesis by suppressing ECM degradation [21]. So we think that IL-32 might be involved in the pathogenesis of liver fibrosis by inducing TIMP-1 expression. It is reported [4,22] that liver fibrosis is initiated by hepatocyte damage, which results in the recruitment of inflammatory cells and the subsequent expressions of cytokines. Finally, HSCs are activated and migrate into injury sites. Viral hepatitis (chronic hepatitis B, chronic hepatitis C) is one of the most common causes, which could result in liver fibrosis. So we think that the pathogenesis from viral hepatitis to liver fibrosis might be as follows: HBV, HCV could induce a lot of expressions of cytokines/chemokines (IL-32, IP-10, MIG, etc.) by hepatocytes [17,23–26]. MIG and IP-10 could induce migration of leukocytes into liver. IL-32 could induce IL-1β, IL-6, IL-8, TNF-α expressions by immune cells [13,27]. In return, TNF-α, IL-1β could also induce IL-32 expression. As a result, the intrahepatic cytokine network events would be initiated after HBV or HCV infection. Viral hepatitis might happen. Recently, it is demonstrated that hepatic IL-32 expression are increased in patients with CHB or CHC [16,17]. Hepatic IL-32 expression is positively related to liver inflammation and liver fibrosis. In the present study, we found that IL-32 could induce TIMP-1 expression by LX-2 cells (a hepatic stellate cells line), and the increased TIMP-1 could promote the migration of LX-2 cells. TIMP-1 plays an important role in the synthesis and degradation of ECM by inhibiting MMP9 and suppressing degradation of ECM. Furthermore, TIMP-1 also promotes the development of hepatic fibrosis [12]. It is accepted that after the liver is injured, HSCs might migrate into the injured site and secrete collagen I and other ECM to repair liver cells. The sustained wound-healing response would result in excessive ECM deposition. Excessive ECM deposition is characteristic of liver fibrosis. Taken together the reports above, we believe that IL-32 plays important roles in the pathogenesis from viral hepatitis to liver fibrosis. It is generally accepted that the increased expression of TIMP-1 promotes liver fibrosis by inhibiting matrix degradation. The decreased expression of TIMP-1 is related to the increased matrix degradation and the resolution of liver fibrosis [28]. Serum and hepatic TIMP-1 expressions are increased in patients with hepatic fibrosis [9,29]. However, the role of TIMP-1 in hepatic fibrosis is not thoroughly elucidated and is controversial. On the one hand, TIMP-1 could promote HSCs migration. Mannaerts et al. [30] report that siRNA-mediated silencing of Igfbp3 (Insulin-like growth factor binding protein 3) in primary mouse HSCs strongly reduces cell migration. The reduced HSCs migration after lgfbp3 knock-down could be overcome by TIMP-1 protein treatment. On the other hand, TIMP-1 might reduce HSCs migration. Ramezani-Moghadam et al. [20] report that adiponectin reduces HSCs migration by promoting TIMP-1 secretion. They think that TIMP-1 might contribute to the anti-fibrotic effect of adiponectin. In the present study, we found that IL-32 could induce TIMP-1 expression by LX-2 cells (HSCs line), and the increased TIMP-1 expression could promote the migration of LX-2 cells. Our results were in accordance with the report of Mannaerts et al. We think that TIMP-1 might play an important role in the promotion of hepatic fibrosis. The reasons might be as follows: when liver is injured, HSCs would fastly migrate into the injured site, and secrete ECM to repair liver. After HSCs arrive at the injured site, these HSCs should consistently reside in the injured site and repair liver until the injured liver is completely repaired. During this time, HSCs should not be attracted and, migrate into other sites. So TIMP-1 might reduce HSCs migration as well as promote HSCs migration in order to repair liver injury. If liver is consistently damaged, HSCs would dramatically secrete ECM. As a result, liver fibrosis would form and liver would lose function.