Praziquantel With Celecoxib Alleviates Liver Fibrosis In Mice With Schistosomiasis

Schistosomiasis japonica is a beast of parasitic diseases. Its main pathological damage is the granulomatous reaction caused by deposition of a large number of eggs in liver and intestine tissues and advanced hepatic and intestinal fibrosis.The traditional treatment namely praziquantel insecticidal, can significantly reduce the continuous inflammatory reaction in the body, but it still can’t effectively control liver fibrosis.Fibrosis is the repair response to tissue damage caused by the inflammation and long-term repeated inflammation is a prerequisite to develop liver fibrosis. Cyclooxygenase 2 (COX-2) is an important enzyme which can catalyze arachidonic acid to prostaglandins and participate in the inflammation process. In recent years, the study found that COX-2 expression in the liver increased markly when liver fibrosis and cirrhosis occurred, while little expression in normal liver tissue. Therefore, we speculated that COX-2 may play a key role in the formation of liver fibrosis. Sustained and effective anti-inflammatory therapy is one of the treatments to prevent fibrosis progression. The COX-2 inhibitor celecoxib as one of non-steroidal anti-inflammatory drugs (NSAIDs) is widely used in clinic, and it has been confirmed to inhibit liver fibrosis in many animal models. So we established mice model withSchistosoma japonicum infection and treatment with celecoxib in order to explore the relationship between COX-2 and schistosomiasis and the therapeutic effect of COX-2 inhibitor in schistosomiasis.Five experimental groups were designed as the normal group (uninfected),6w infected,12w infected, praziquantel treatment group(PZQ), and praziquantel with celecoxib treatment group (PZQ+CLC). We used Automatic biochemical analyzer to test serum alanine aminotransferase (ALT), HE staining to evaluate the degree of liver inflammation, Masson staining to determine the collagen deposition, immunohistochemical methods to evaluate fibrosis related factors’expression, real-time fluorescent quantitative PCR to detect the transcription level of COX-2, inflammation and fibrosis related genes and some microRNAs. Our aim is to explore the COX-2 inhibitor celecoxib’s function and its related mechanisms in preventing the fibrosis in schistosomiasis.The main results of our studies were as follows:1. Compared with the 12w infected group, mice in PZQ+CLC group’s serum ALT level dropped significantly and tended to normal level, but no statistical difference was shown between PZQ group and PZQ+CLC group.2. Liver COX-2 mRNA expression was elevated after infected with schistosome. The transcription levels of COX-2 mRNA in 12w infected group were lower those in 6w infected group. Compared to 12w group and PZQ group, COX-2 expression in mice of PZQ+CLC group was significantly suppressed. Thus COX-2 may participate in hepatic lesions of schistosome infection.3. The liver HE staining results showed that a large number of eggs gathered near the portal area and around the eggs with a large number of cells such as fibroblasts, lymphocytes, eosinophils and macrophages infiltration were observed in 12w-infected group, while those in PZQ+CLC group and PZQ group were relatively few. And compared to PZQ group and infection control group, the single egg granuloma area in mice of PZQ+CLC group significantly decreased.4. The liver Masson staining results showed that, collagen fibers were not seen in the liver of normal mice. Compared with 12w group and PZQ group, the area of collagen fibers were decreased in PZQ+CLC group.5. Immunohistochemical staining showed that, compared to mice in 12w infected group and PZQ group, the expression levels of Collagen Ⅰ, Collagen Ⅲ, α-SMA, MMP-9 and TIMP-1 in the livers of mice in PZQ+CLC group were significantly decreased.6. RT-PCR’s results showed that, compared to 12w infected group and PZQ group, the mRNA transcription levels of fibrosis-related factors such as Col1α1, Col3α1, α-SMA, MMP-9 and TIMP-1 in PZQ+CLC group were significantly decreased, while the proportion of MMP-9/TIMP-1 was increased.7. RT-PCR’s results showed that liver proinflammatory cytokine including TNF-α, IL-6 and IL-1β mRNA levels of 6w infected group were significantly higher than those of normal group and 12w infected group. Compared with 12w infected group and PZQ group, PZQ+CLC group’s liver proinflammatory cytokines (TNF-α, IL-6, IL-1β) and profibrotic cytokines (TGF-β, IL-4, IL-13) mRNA levels were markly decreased.8. Finally, miR-155 and miR-146a mRNA levels in the livers of 6w infected group mice were significantly higher than those of normal group and 12w infected group. Compared with 12w infected group and PZQ group, miR-155 and miR-146a mRNA levels in the livers of PZQ+CLC mice group were decreased, and tended to be normal. MiR-199a and miR-101a mRNA levels in the livers of mice in 6w infected group were significantly lower than those in normal group and 12w infected group. Compared with 12w infected group and PZQ group, PZQ+CLC group’s liver miR-199a and miR-101a mRNA levels were increased.To sum up, this study has defined the COX-2 expression characteristics in schistosomiasis, and COX-2 inhibitor celecoxib with praziquantel can alleviate liver fibrosis caused by schistosomiasis. These experimental evidences provide further guide in schistosomiasis prevention and control in the field.