Study On Preparation And Effectiveness And Stability Of Umbilical Cord Mesenchymal Stem Cells Conditioned Medium

Mesenchymal stem cells(MSCs)research is one of stem cell science which is a frontier science of Medical field.MSCs were used to dealing with diseases which were difficult treated with lots of modern medicine,because they had unique and powerful ability of differentiation,immune regulation and anti-inflammatory.At present,MSCs were found having a good therapeutic effect in painless wounds and skin diseases.MSCs could promote skin wound healing through intradermal or subcutaneous local injections.The therapeutic effects of MSCs mainly through its autocrine and paracrine mechanism.However,the survival time of cells' was only about 24 h under the low temperature with preservation solution.With the extension of time,the apoptotic cells were continue to appeared,the vitality and effectiveness of cells' was declined,which limits its clinical application.Umbilical cordmesenchymal stem cells conditioned medium,MSCM,was used only to instead of MSCs in the treatment of refractory wounds and skin diseases,can also played a similar effect and applicated more convenient.In this study,explant method and collagenase digestion method,two umbilical cord separation methods,had been contrasted,and a standard technical procedure of collecting umbilical cord mesenchymal stem cells(hUC-MSCs)was established.A rich and high purity of MSCs can be obtained using explant method.BCA method was used to detect the protein content in the culture media of different generations and different starvation time of MSCs,and the maximum concentration of MSCM was obtained by starvation 72 h in the MSCs of P 4 generation.Research with CCK 8 multiplication experiment and scratch experiment in MSCM,we found that when the concentration of protein is 10?g/ml,which can play the best multiplication effect.Compared with other groups,it has significant difference(P< 0.05).The characteristics of the protein is unstable,easy to decompose.The research found that the MSCM of effective concentration with lower temperature,longer save the time by the different environmental temperature for a long period of time to save and detection.4 ? reservation can maintain about 2 weeks.After adding different concentration of trehalose,effective concentration can be maintained for a long time.4 ? reservation can maintain about 4 weeks,with the remarkable effect.In the comparision of different concentrations of trehalose,concentration of 0.7 M trehalose has the best preservation effects.Compared with other groups,it has significant difference(P< 0.05).In the experiment,we found that MSCM which made of freeze-dried powder,can effectively extend the storage time.Stored 4 ? can be maintained for about 8 weeks.The study for understanding how to use MSCM,how to extend the effective concentration of protein provides theory basis.