The Effect Of Several Traditional Chinese Medicine On The Skin Fibroblasts

PurposeIn vitro culture of normal adult skin fibroblasts,and identify the fibroblasts,observe effects of Chuanxiongzine,matrine, Asiaticoside of normal adult skin fibroblast ceils and their autocrine transforming growth factor-β1 Discussion Asiaticoside,Chuanxiongzine,matrine on the role of fibroblast mechanismMethod1,Use tissue culture method to culture primary fibroblast with normal adult foreskin,2,Identification of fibroblasts:(1) Morphological observation:fibroblasts will be digested by 0.25%trypsin to produce single-cell suspension,cell climb tablets,ethanol fixation,cells were observed morphological features.(2) immunohistochemical identification:check the above-mentioned cell climb tablets,ethanol fixed,immunohistochemical determination of cell vimentin and keratin expression.3,Not add medicine containing 10%FCS DMEM culture medium for the control group,based Matrine,Chuanxiongzine, Asiaticoside of three drug groups,each with five different concentrations,add cultured skin fibroblasts were cultured within 24 hours,48 hours after the camera under inverted phase contrast microscope,and for MTT,then detect OD value at microplate reader4,Ibid grouped separately,after cultivate 24 hours,cell supernatants of TGF-β1 content was determined by ELISA.All statistics data are express by x±s,using one-way ANOVA (one-way NAOVA),compared with LSD testing,applicate SPSS11. 0 statistical software.Standard test P<0.05 was significant difference5,Iactate dehydrogenase(LDH) activity assay,ibid division, LDH in the cell supernatant of the control group was measured by an automatic biochemical analyzer,and the scope of its fluctuations was regard as the normal Detection range Established for the 95%confidence interval,make one-sided test,statistical analysisResult1,Human fibroblast cell morphology and identification:the light microscope showed that cells polygonal or long spindle after HE staining.Vimentin immunocytochemical staining showed that fibroblast cytoplasm stained dark brown,that show positive.2,Morphological changes of fibroblast,fibroblast of control group was long and narrow,shape microscope,nuclei were spindle-shaped,mostly arranged in parallel and have curvature,the central of Cell has a round or oval nucleus,some cells can be seen protruding out 2-3 protrusions of varying lengths.With the increase of culture time,cell number was increased and cell gap narrowed,cell distribute as streak. Asiaticoside experimental group,when the drug concentration was 0.25mg/ml,the ceils became shorter and float after the 24 hours.During the period,with the cultivate concentration increased,the number of adherent ceils was further reduce. the cell gap has become larger.When the drug concentration was 2.5 mg/ml,the cell was round,a large number of cells floated in the culture medium.Matrine experimental group when the drug concentration was 0.5 mg/ml at 24 hours a period of time,with the increase of the number cultivate thick,reduce the number of cells,cell protrusions Clostridium retraction,reduced contact with neighboring cells,the cell gap become larger,gradually form a round,off the wall.When the drug concentration of 5mg/ml, the 48 hours after the cell that is round,a large number of cells floating in the culture medium.Ligustrazine experimental group when the drug concentration of 0.5 mg/ml at 24 hours a period of time,with the increase in the number cultivate thick,reduce the number of cells,cell Clostridium neurite retraction,reduced contact with neighboring cells,the cell gap change large,shape gradually round off the wall.When the drug concentration of 5mg/ml,the 48 hours after the cell that is round,a large number of cells floating in the culture medium.3,cell supernatant of each group after 24 hours were checked, and cell supernatant TGF-β1 content was measured by Elisa.The OD value of TGF-β1 was very low.the each group has not significant difference with contronl group by statistical treatment.(P>0.05)At a certain Concentration,Asiaticoside,matrine, ligustrazine had no effect on LDH activity of fibroblasts.ConclusionAt a certain concentration,Asiaticoside,matrine, Ligustrazine can inhibit the normal skin fibroblast cells.But does not affect fibroblast autocrine TGF-β1,normal human skin content is very less.