Study Of The Inhibition And Apoptosis Effect Of IL-10 Paracrined By Bone Marrow Derived Mesenchymal Stem Cells On Hepatic Stellate Cells

Liver disease is a worldwide disease. If all sorts of acute or chronic liver diseases can not get effective protection and treatment, they will tend to deteriorate further, and finally to the development of liver cirrhosis or liver cancer. Liver disease deterioration is a long and complicated process, during this period which obviously happens is the pathological change of liver fibrosis. Liver fibrosis is caused by tissue excessive repair with regeneration after pathologic lesions in the liver, at the same time hepatic stellate cells are activated and liver extracellular matrix synthesis is larger than its degradation, eventually it leds to the deposition of extracellular matrix. The present study shows that liver fibrosis is a reversible process. What’s more, the effective measure to prevent liver fibrosis is to suppress the activation of hepatic stellate cells and promote its apoptosis. Therefore, the research is of great significance to target on HSCs for the treatment of liver disease.Stem cell technology is in the frontier field of regenerative medicine research, and it provides a new way for a variety of diseases. Bone marrow mesenchymal stem cells have the ability of suppressing the activation of hepatic stellate cells and promoting its apoptosis, at the same time it has the characteristics of convenient acquisition、easy to cultivate、good stability and small side effects, but its mechanism has not yet been completely cleared. Interleukin-10 is mainly secreted by Th2 cells as a kind of important anti-inflammatory cytokine, it has strong inhibition of inflammation and the effect of macrophage function. It have been reported that bone marrow mesenchymal stem cells can also secrete a large number of Interleukin-10. However, whether it is the IL-10 paracrined by BMSCs that plays a role of treatment of liver injury is unclear. So, this study will explore and analyze the scientific problem.In our study, we adopt the non-perfusion method to isolate rat hepatic stellate cells, through the cell morphological observation, oil red O, hematoxylin staining and immunohistochemical to identify the separated and cultured HSCs. We use density gradient centrifugation method to isolate bone marrow mesenchymal stem cells from rat femur and tibia, adopt the cell morphological observation and immunohistochemical to identify the separated and cultured BMSCs, apply osteogenesis and adipogenesis differentiation experiment to measure its pluripotency. We collect the third generation of BMSCs culture supernatant, use the method of Elisa to detect IL-10 concentration in the culture supernatant. To analyze the inhibition and apoptosis effect of supernatant on HSCs, we directly apply the supernatant on HSCs for 24 h, 48 h and 72 h. Further more, we use Transwell culture experiment to confirm the effect of IL-10 secreted by BMSCs on HSCs.Experimental results show that: the separated and cultured HSCs present obvious astrocytes form, oil red O staining is positive, and it has the ability to express specific proteins Desmin, HSCs, α-SMA and GFAP. Separated and cultured BMSCs can express specific markers CD29, CD44 and CD45, and has great adipogenesis and osteogenesis differentiation ability. BMSCs culture supernatant of Elisa detection demonstrate that it secrets of IL-10. With the supernatant directly contacting on HSCs in 24 h, 48 h and 72 h, it can significantly inhibit HSCs and promote its apoptosis. Transwell experiment shows that IL-10 secreted by BMSCs can effectively suppress the secretion of the HSCs TGF-β1 and the expression of α-SMA, while promoting HSCs apoptosis protein expression of caspase-3.To sum up, this study preliminary shows that BMSCs supernatant can inhibits the proliferation of HSCs, IL-10 paracrined by BMSCs inhibits the proliferation and promote apoptosis of HSCs.IL-10 plays an important role in the process of fibrosis reduce and reverse, and is expected to become important drug for the treatment of liver fibrosis diseases.