Effects Of Sinomenine On The Biological Function Of Hypertrophic Scar Fibroblasts

ObjectiveHyperplasia of the human skin or hyperplastic scar(HS)is a manifestation of excessive repair of human tissue cells after various kinds of burn trauma or surgery,causing local tissue thickening,hyperpigmentation,or even loss of pigment.In severe cases,limb function may be be affected by scar contracture.HS imposes a heavy burden on the patient's body and mind.Therefore,hyperplastic scar has been one of the key issues in the field of medical research.It has been reported that fibroblasts play an important role in scar formation.Fibroblasts have enhanced proliferative capacity,decreased apoptosis,increased collagen secretion,and increased extracellular matrix deposition,which ultimately leads to the formation of HS.Sinomenine(SIN),a single alkaloid extracted from the root of the Sinomenium acutum plant,is used for treatment of rheumatism and swelling of the joints.In addition,it has been reported that sinomenine can play a role in anti-renal interstitial fibrosis by inhibiting TGF-?1,serum laminin,and fibronectin.In this study,we tried to explore the role of sinomenium and its molecular mechanism for the proliferation of fibroblasts.MethodsHuman fibroblasts were treated with different concentrations ofsinomenium and collected at different time points.Cell viability was determined by MTT assay,cell cycle was determined by flow cytometry,apoptosis rate was determined by Hoechst 33258 assay,and cell migration rate was determined by Transwell assay.Finally,changes in related signal molecules were detected by Western blot assay.ResultsSIN has a proliferation inhibition effect on human fibroblasts in a dose-and time-dependent manner.Human fibroblasts were continuously treated at 25 ?M and 50 ?M for 12 hours,24 hours,36 hours and 48 hours,and the cell proliferation activity was determined by MTT assay.The proliferation inhibition rate of 24 h cell proliferation was 13.14±5.51% and 43.28±4.98%.In the case of sinomenine,the same concentration of SIN was applied to the fibroblasts and the cell cycle was detected by flow cytometry.It was found that the cells in the G1 phase increased,and the cells in the S phase decreased,compared with the control group,indicating a G1/S cell block phenomenon.The phenomenon became more obvious with the increase of the SIN dose.After Hoechst 33258 staining,it was found that compared with the blank control group,the cells after SIN treatment showed apoptosis promotion in a dose-dependent manner.The same number of Transwell experiments showed that the fibroblast cells experienced migration inhibition after the treatment with SIN.Western blot analysis showed that the TGF-?1 signal of human fibroblasts was inhibited by SIN treatment,and the expression of Cyclin D1,bcl-2 and MMP2 was observed.ConclusionsWe concluded that SIN inhibits the growth and migration of fibroblasts.The proliferation inhibition caused by the stimulation will cause apoptosis and reduce cell invasion ability.The inhibitory effect of SIN on HS fibroblast growth and migration may be through regulation of the TGF-?1 pathway.