| Hepatic stellate cells(HSCs) are the core participants in liver fibrosis development. 1, 25-(OH)2D3, the active form of vitamin D, has antiproliferative properties and antifibrotic potential, as well as a role in renal and lung fibrosis. Little is known about the role of 1,25-(OH)2D3 in liver and its involvement in liver fibrosis. Therefore, we investigated the antiproliferative and antifibrotic effects of 1,25-(OH)2D3in HSC-T6 in vitro.The development of liver fibrosis is characterised by an accumulation of extracellular matrix(ECM) with subsequent destruction of the normal liver archi-tecture, leading to liver cell dysfunction. Hepatic stellate cells(HSCs) play a critical role in the development of liver fibrosis, since they are responsible for excessive deposition of ECM proteins, predominantly type I collagen. Multiple signalling pathways are implicated in HSC activation and proliferation. The most potent mitogenic factor for HSCs is platelet-derived growth factor(PDGF). The transforming growth factor PI3K/Akt signalling pathway is one of the main stimulating factors for profibrotic ECM protein synthesis.Vitamin D is a prohormone that requires sequential enzy-matic modification of 25- and 1a-hydroxylation in the liver and kidney, respectively, leading to its maximal biological activity as1,25-(OH)2D3.apart from its calcium regulatory properties, Indeed, it has been shown that1,25-(OH)2D3regulates hundreds of different genes and plays a significant role in adjusting and controlling cell proliferation, differentiation and apoptosis in many normal and cancer cells and also has immunomodulatory anti-inflammatory effects. Recent studies have indicated that1, 1,25-(OH)2D3 also has an inhibitory effects on renal and lung fibrosis.Treatment with vitamin D led to the inhibition of lung fibroblastproliferation and showed an antifibrotic effect in murine lung and obstructed kidney models.There are currently few effective treatments for liver fibrosis. The main targets in exploring therapeutic possibilities are aimedat inhibiting the proliferation and biotransformation of HSCs or by inhibiting the transduction of the profibrotic PI3K/Akt signalling cascade. The latest study confirms that 1,25-(OH)2D3 can guard aginest carbon tetrachloride induced liver fibrosis in rat in vivo. However, the potential function of 1, 25(OH)2D3 on HSCs and the acquaintance of the precise molecular mechanism by 1,25-(OH)2D3are weakly characterized.Objective:In order to inspect the effect of 1,25-(OH)2D3on proliferation and apoptosis and its promising mechanism in HSCs line T6, We become conscious of the biochemical markers of apoptosis in T6 cells to verify if the apoptosis occurred after treatment T6 with 1,25-(OH)2D3and detected the expression levels of apoptosis-related proteins and the proteins related to the signal transduction pathways to find out the relationship between their changes and apoptosis in this task. Then make available for the experimental proof for 1,25-(OH)2D3 as a new anti-hepatic fibrosis drug.Methods:In vitro T6 cultured to the exponential growth phase with different concentrations of 1,25-(OH)2D3. The role of a certain time, cells inhibition rates and proliferation of T6 were detected with MTT assay; HSC-T6 cells apoptosis and cell cycle were detected by Flow cytometry technology(FCM). Employ the method of experimental scratch test cells’ ability to migrate. Rt-PCR and western blot was used to evaluate the activation as well as inhibition of signal molecules and expression of type I collagen.Results: 1,25-(OH)2D3 could effectively reduce the migration and inhibits its multiplication in a concentration-dependent along with time-dependent manner on HSC-T6.HSC-T6 cell cycle was stucked in G0/G1 phase, induce apoptosis, apoptosis rate with rising drug concentration and showed an promoting trend,The result showed that the inhibition of T6 at 48 h in 2.5×10-6mol/l group was significantly decreased than other groups(60.01±1.69 vs 64.11 ±1.69 vs 64.62 ± 1.19 vs 72.01 ± 1.15)(P < 0.05); enhance activated caspase-3 and caspase-9 expression obviously(P<0.05); It also can down-regulated of PI3K/Akt/m TOR pathway and intervention m RNA levels of collagenα1(I) and the type Icollagen protein expression.Conclusion:1. 1,25-(OH)2D3 significantly inhibited the proliferation of HSC-T6 In vitro experiments.2. 1,25-(OH)2D3 could notly induce apoptosis of HSC-T6 and the cell cycle were arrest in G0 / G1 phase.3. 1,25-(OH)2D3 significantly promot the activation of caspase-3 and caspase-9.4. 1,25-(OH)2D3 greatly inhibited the expression of type I collagen α1(I) m RNA and type I collagen.5. 1, 25-(OH) 2D3 may inhibit the HSCs proliferation and promoting apoptosis through the PI3 k /Akt/m TOR signal path. |
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