Experimental Study About Influence Of The Recipe Of Supplementing Ying-wei On The Proliferation And Cell Cycle Of The Aging Human Skin Fibroblasts

Objective To observe the proliferation of the aging human skin fibroblasts(HSF) which are cultured in vitro by using the recipe of supplementing Ying-wei;and to observe the relationship between cell proliferation and differentconcentrations of the drug serum of supplementing Ying-wei; and discuss itspossible mechanism.Methods1Cell culture: Purchase of human skin fibroblast cell line,HSF are cultured invitro till it get aging, which are cultured in vitro separately in DMEM with10%FBS.2Preparation of drug-containing serum: The health of60male SD rats wererandomly divided into three groups, they are dose groups of differentconcentrations of the recipe of supplementing Ying-wei,20rats in each group.Different groups are poured into different liquids by gavage. Blood samples inaccordance with common law, separation, inactivated serum, and respectivelyprepared containing10%serum DMEM complete medium.3HSF grouping: The young control group, the aging control group,supplementing Ying-wei of high, medium and low-dose groups, and in each groupadd the corresponding serum to culture.4The detection of cell proliferation: to observe continuously the cells’changes of each group in the number through cell counting, and draw the growth curve of cell.5The determination of cell cycle: by using flow cytometry.6Immunocytochemistry detection: with the method of immunocytochemistryto detect the protein expression of PCNA, c-Fos and c-Myc of each group.Results1From HSF growth curve, we can see,in the same time, the order of thenumber of cells from high to low is high, medium and low dose group, youngcontrol group, aging control group.2By using flow cytometry to detect cell cycle, S phase percentage from highto low is high, medium and low dose group, young control group, aging controlgroup.3Immunocytochemistry detection: the young control group，s OD valuecompare with aging control group，s OD are significant difference (P<0.05), itshow that after a certain number of passages, normal human skin diploid cell enterinto the non-proliferative state which is called cellular senescence; HSF get aging.supplementing Ying-wei of each dose group，s OD values compare with agingcontrol group，s OD value is significant difference (P<0.05), it indicate that theprescription of supplementing Ying-wei can promote the proliferation of the agingskin fibroblast; the OD value of medium dose group compared with the OD valueof the low dose group is significant difference (P<0.05), the OD value of high dosegroup compared with the OD values of the medium and low dose groups aresignificant difference (P<0.05), which show that high, medium and low dosegroups are significant differences (P<0.05), it suggeste that the recipe ofsupplementing Ying-wei can promote the proliferation of the aging skin fibroblast,and which is increased with the increasing of drug serum concentrations of therecipe. 4The positive express of PCNA, c-Fos and c-Myc protein in each dose groupare increased.Conclusions. The recipe of supplementing Ying-wei can promote theproliferation of the aging skin fibroblast, and which is increased with theincreasing of drug serum concentrations of the recipe. The recipe ofsupplementing Ying-wei can defer skin get aging.