| Hepatic fibrosis a pathologic process that various of factors to damagesof liver cannot be removed, which activate myofibroblast, caused imbalanceof collagen metabolism and reconstruction of liver tissue. Hepatic fibrosis isthe inevitable stage of various of hepatic disease proceeding to cirrhosis.Chronic hepatic disease caused by various of factors seriously threaten thehealth of people in our country. The hotspot of research is to seek effectiveanti hepatic fibrosis drug. Nowadays, many studies showed that hepaticstellate cells were the target of treatment of hepatic fibrosis, while there is nospecific therapy. Traditional Chinese medicine receives widely attention in theworld because its characteristics of simple, convenient, inexpensive andefficacy in anti hepatic fibrosis. After years of studies in clinical andexperiment, Professor Li Diangui showed that turbidity poison played a keyrole in the formation of hepatic fibrosis, and therapy with HuaZhuo Jiedu tohepatic fibrosis have achieved a good effect. But the mechanism of HuaZhuoJiedu remains unclear.Objective: Our experiment is to observe the expression of α-SMAmRNA, p38MAPK and JNK protein on the cellular level, adopting the serumpharmacology method, then discuss the mechanism of Huazhuo Jiedu formulain therapy to rat hepatic fibrosis in vitro, and offer molecular and biologytheoretical basis for possible therapeutic targets and pathways of HuazhuoJiedu formula, and offer modern pharmacological evidence for the mechanismof chronic hepatic disease turbidity poison from through formula effectassessment.Methods: Healthy adult SD rats，choose to produce the medicated serum,were randomly divided into four groups: normal control group, positive control group(Fufang Biejia Ruangan tablet group), Huazhuo Jiedu formulaequivalent and double dose group. Primitive hepatic stellate cells were dividedinto five groups: control group (group A), model group (group B), positivegroup (group C), Huazhuo Jiedu formula medicated serum equivalent (groupD1) and double group (group D2). Each group was treated with TGF-beta1except the control group, and then added respectively rat serum. Aftercultivate24,48,72hours, the proliferation of HSC was tested by MTTmethod. After cultivate48hours, the expression of α-SMA mRNA were testedby RT-PCR, p-p38MAPK and p-JNK protein were separately examined byWestern-blot.Results:1Inhibitory effect on proliferation of HSC: Compared with group A, group Bhad a significantly higher proliferation at24hours (P<0.05). Compared withgroup B, groups C, D1, D2had a significantly higher inhibitory effect onproliferation (all P <0.05). Compared with group C, group D2played anhigher inhibitory effect on proliferation of HSC (P<0.05), but there was nodifference between C and D1. Group D2had a significantly higher inhibitoryeffect on proliferation compared with group D1(P<0.05).After treated with48,72hours, compared with group A, group B showed higher proliferation(allP<0.05). Compared with group B, groups C, D1, D2had a significantly higherinhibitory effect on proliferation (all P<0.05). Compared with group C, GroupD1and D2showed higher proliferation (P<0.05). Group D2had asignificantly higher inhibitory effect on HSC than that of group D1(P<0.05).Inhibitory effect of group D1, D2on proliferation of HSC increased graduallywith time.2RT-PCR results: After48hours, compared with group A, the expression ofα-SMA mRNA in group B significantly increased (P<0.05).Compared withgroup B, the expression of α-SMA mRNA in group C, D1, D2significantlydecreased (all P<0.05).Compared with group C, group D2can significantlydecreased the expression of α-SMA mRNA, and the difference wasstatistically significant (P<0.05), while there was no difference between group C and D1(P>0.05). Compared with group D1, group D2decreased theexpression of α-SMA mRNA (P<0.05).3Western-blot results show:3.1The expression of p-p38protein: compared with group A, the expressionof p-p38protein in group B significantly increased (P<0.05). Compared withgroup B, the expression of p-p38protein in group C, D1, D2decreased (allP<0.05). Compared with group C, the expression of p-p38protein in group D2decreased (P<0.05).The expression of p-p38protein in group D1increasedcompared with group C, but there was no difference between group C and D1(P>0.05). The expression of p-p38protein in group D2decreased than that ingroup D1(P<0.05).3.2The expression of p-JNK protein: Compared with group A, the expressionof p-JNK protein in group B significantly increased (P<0.05). Compared withgroup B, the expression of p-JNK protein in group C, D1, D2decreasedsignificantly (all P<0.05). Compared with group C, the expression of p-JNKprotein in group D1and D2decreased, the difference was statisticallysignificant (all P<0.05). Compared with group D1, the expression of p-JNKprotein decreased in group D2, the difference was statistically significant(P<0.05).Conclusions:1Huazhuo Jiedu formula medicated serum has aninhibitory effect on proliferation of HSC stimulated with TGF-β1, and therewas a time-dose dependent manner.2Huazhuo Jiedu formula medicated serum can reduce the level ofα-SMA mRNA in HSC stimulated with TGF-β1, and inhibits activation ofHSC.3Huazhuo Jiedu formula medicated serum can reduce the expression ofp-p38, p-JNK protein in HSC stimulated with TGF-β1, and then interventp38MAPK and JNK signaling pathway. This may be one of mechanisms inanti-fibrotic treated with HuaZhuo Jiedu formula.4Huazhuo Jiedu formula medicated serum can intervent p38MAPK andJNK signaling pathway, decrease the expression of α-SMA in rats HSC, speculating that it is one of mechanisms of anti-fibrotic. |
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