| Purpose:By molecular biological technique and in vivo experiment,we observed the influence of UV irradiation on TGF-β/Smads signal and MMPs/TIMPs of skin photoaging rats to partly explain the function of degradation of extracellular matrix and further study the mechanism of skin photoaging. Taking Taohong Siwu Decoction as the exmaple,we studied the pathogenesis and the substance basis of blood stasis photoaging. The action of Taohong Siwu Decoction in the treatment and prevention of skin photoaging was also studied,hoping to get a full understanding and provide new theory basis for using TCM to treat and prevent skin photoaging.Material and Methods: 60 Kunming mice were randomly divided into 6 groups with 10 in each:blank group,model group,TCM low,middle and high dose groups,VE group. The blank group mice were given no operations besides shaving. The model group was given the normal saline. Taohong Siwu Decoction was given to TCM three groups with the corresponding doses. VE group was given vitamin E as the positive control. Except the blank group,the other groups were given ultraviolet irradiation 30 min before modeling. UVA+UVB was used to make the mice skin photoaging model. Each mice’s back skin was shaved by a area of 1.5×1.5cm,three times a week,totally 14 weeks. 14 weeks later,the model group mice showed symptoms like fatigue,reduced actions,dry skin with dark red ecchymosis. All these pathological changes met the criterion of skin photoaging. In 15 th week,all the mice were killed and ELISA method was used to detect the contents of IL-1 and TNF-αin the blood. RT-PCR method was used to detect the expressions of Smad2/3,MMP-1,MMP-3,MMP-12,TIMP-1 and TIMP-2 m RNA in skin tissues and Western-blot method was for the protein expressions of Smad2/3,MMP-1,MMP-3,MMP-12,TIMP-1 and TIMP-2 in skin.Results:1. ELISA method:Compared with the blank group,TNF-α and IL-1 expressions were obviously increased in model group with significance(P<0.01). Compared with the model group,TNF-α and IL-1 expressions in TCM three groups were obviously decreased with significance(P<0.01) and compared with the VE group,the expressions in low and middle dose groups were increased with significance(P<0.01)and there was no significance in the high dose group. 2.RT-PCR method:(1)Smad m RNA expression:(1)Smad-2 m RNA:Compared with the blank group,it was obviously decreased in the model group with significance(P<0.01). Compared with the model group,it was obviously increased in the TCM three dose groups(P<0.01). Compared with the VE group,it was decreased in the TCM low dose group with significance(P<0.01). It was obviously decreased in TCM middle dose group(P<0.05)and there was no significance in TCM high dose group.(2)Smad-3 m RNA expression:Compared with blank group,it was obviously decreased in model group(P<0.01). Compared with model group,it was obviously increased in TCM high dose group(P<0.01)and the difference had no significance in low dose group. Compared with the VE group,the expression was obviously decreased(P<0.01)in low dose group and the decrease in the low dose group had singificance(P<0.05). The difference in high dose group had no MMPs m RNA expression MMP-1 and MMP-3 m RNA expressions: There was a low expression in the blank group. Compared with the blank group,MMP-1 and MMP-3 m RNA expressions in the model group were obviously increased(P<0.01). Compared with the model group,the expressions in TCM low,middle and high dose groups were all obviously increased(P<0.01). Compared with the VE group,the expressions in TCM low and middle dose groups were increased(P<0.01). The difference in TCM high dose group had no significance. MMP-12 m RNA expression: It had a low level in skin tissue of the blank group mice. Compared with the blank group,the expression was obviously increased inthe model group(P<0.01)and compared with the model group,the expression in TCM three doses groups was obviously decreased(P<0.01). Compared with the VE group, the expression in low and middle dose groups was increased significantly(P<0.01)and the high dose group had no significance.(3)TIMPs m RNA expression:(1)TIMP-1 m RNA:Compared with the blank group,it was obviously decreased in the model group(P<0.01). Compared with the model group,it was increased significantly in TCM three doses groups(P<0.01). Compared with the VE group,it was obviously decreased in low and middle dose groups(P<0.01)and the high dose group had no significance.(2)TIMP-2 m RNA: Compared with the blank group,,it was obviously decreased in the model group(P<0.01). Compared with the model group,it was increased significantly in TCM three doses groups(P<0.01). Compared with the VE group,it was obviously decreased in low and middle dose groups(Results of Western blot).(4)Smad protein expression: Compared with the other groups,it was obviously higher in the blank group with significance(P<0.01). It was obviously lower in the model group than that of the other groups(P<0.01). The expression in TCM high dose group as well as the VE gorup was highest,which was higher than that of the model and middel(P<0.05) and low(P<0.01)dose groups. There was no significance between high dose group and VE group.(P<0.01)and the high dose group had no significance. MMPs protein expression(1)MMP-1 and MMP-3 protein expressions:Compared with the other groups,it was obviously lower in the skin of the blank group mice with significance(P<0.01). It was higher in the model group than that of the other groups(P<0.01). The expressions in low and middel doses groups were higher than that of the high dose and VE groups(P<0.01).(2)MMP-12 protien expression:Compared with the other groups,it was obviously lower in the blank group with significance(P<0.01). It was obviously lowerin the model group than that of the other groups(P<0.01). It was lowest in the TCM high dose and VE groups,which was lower than that of the model and TCM low and middle dose groups(P<0.01). There was no significance between high dose group and VE group.(5)TIMPs protein expression:(1)TIMP-1 protein expression: Compared with the other groups,there was no significance between the blank group and TCM high dose group(P<0.05).It was lowest in the model group and had significance compared with that of the other group(P<0.05). The expression in the VE and high dose groups was higher than that of the low and middle dose groups significantly(P<0.05). There was no significance between high dose group and VE group.(2)TIMP-2 protein expression: Compared with the other groups,it was highest in the blank group(P<0.05). Compared with the other groups,it was lowest in the model group. The expression in TCM high dose and VE groups was significantly increased than that of the model group(P<0.05). There was no significance between high dose group and VE group.Conclusion:It has certain clinical effect on skin photoaging by using Taohong Siwu Decoction according to the principle of activating the blood and resolving the stasis and the high dose has the optimal effect. Under the ultraviolet light,the skin tissue can produce IL-1 and TNF-α,which by a complex signal passway,resulting in MMPs expression. It can speed the degeneration of extracellular matrix,leading to skin photoaging. MMP-1 and MMP-3 have the most close relation with this. TIMPs is the specific inhibiting factor of MMPs and it imbalance has the relation with the skin photoaging. Taohong Siwu Decoction can obviously decrease the levels of MMP-1,MMP-3 and MMP-12 and m RNA and increase the levels of TIMP-1,TIMP-2 and m RNA so as to balance the MMP-TIMP to prevent the skin photoaging. UV can inhibit the TGF-βsignal transduction by inhibiting the Smad 2/3expression to speed the skin photoaging. Taohong Siwu Decoction may through increasing the Smad 2/3 expression to prevent the skin photoaing. |
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