The Study On Mechanisms Concerning Viability Of Umbilical Cord MSCs And Correlated Preservation Solutions

BackgroundMesenchymal stem cells(MSCs) are multipotent stem cells which have self-renewal capacity, differentiation potential and immunomodulation property. UC-derived MSCs possess all features of MSCs and are broadly used in clinical settings, along with the advantages of more widely source, easy acquisition and ethics uninvolved. The procedures of UC-MSCs transplantation therapy include collection and culture of tissue, cells passaging and digesting and cell delivery. Those mentioned factors have significant influence on UC-MSCs’quality and therapeutic effect of post-transplantation. Among elements, preservation occupies the most critical impact regards to cell quality, however, the relevant systematic study remain absent lately.The fresh isolated stem cells are not allowed to infuse into human as for UC-MSCs based clinical transplantation therapy. Prior to infusion, Cells are required to preserved into injectable protective solution (e.g.0.9%Sodium Chloride Injection) with a fixed time. To date, commonly used clinical preservation solutions include0.9%Sodium Chloride Injection,5%Glucose Injection and1%Human Albumin liquid etc. However, the poor effects of clinical therapy associate with the reduced activity of UC-MSCs which causes of preserving cells within injections above. Yet, the uncleared causation of decreased vitality and lacking related treatment measures are demanded for further discuss.To sum up, based on mentioned above, this study firstly tested corresponding influence for UC-MSCs, with0.9%Sodium Chloride Injection,5%Glucose Injection, Plasmalyte A,1%Human Albumin liquid and5%Human Albumin liquid. Then different concentrations of human AB serum have been infused into preservation solution to observe the protective effect of serum on UC-MSCs. Finally, preservation solution of UC-MSCs has been concocted to minmize the injury on cell activity by mixing protective factor in the preservation solution according to the serum components. All these have provided the laboratorial basis for the UC-MSCs based clinical therapy of transplantation. Objective1.Evaluation of effects of existing conventional preservation medium on activity of UC-MSCs.2.Study of the effects of different concentrations of human AB serum on UC-MSCs when preserved.3.Concocting preservation solution of UC-MSCs in order to improve the preservation effect by adding protective factors into cells based on the components of serum.Methods1. The study on effects of conventional preservation medium of transplantation on in vitro biological characteristics of UC-MSCs.Resuspendinge the laboratory-cultured UC-MSCs on0.9%Sodium Chloride Injection,5%Glucose Injection, plasma Lyte A,1%Human Albumin solution and5%Human Albumin solution separately. Liquids are kept at4℃or24℃condition for2,4,6h to observe the cell viability, mortality of cell, the effects of cellular adherent ability after preservation of cells, as well as cell surface markers and osteogenic and adipogenic ability.2. The study on preservation effect of UC-MSCs against various serum concentration.Diverse concentrations of human AB serum were infused into0.9%Sodium Chloride Injection as graft preservation solution, under the condition of4℃and saves6,9,12hours to observe the cell viability, mortality of cell, the effects of cellular adherent ability at post-preservation, also the cell surface markers and osteogenic and adipogenic ability.3. Self-made UC-MSCs preservation solution via infusing protective factors into cells. Under the condition of4℃, to concoct the protection solution of UC-MSCs which achieves through increasing disparate protective factors of cells, and preserves6,9,12hours to observe the cell viability, mortality of cell, and the effects of cellular adherent ability after preservation, as well as cell surface markers and osteogenic and adipogenic ability.Result:1. Under the condition of UC-MSCs preserved in various commonly used clinical preservation solutions, although the ability of multiple differentiation potential and surface markers of cells complied with the standard of UC-MSCs, the cell viability and cell adherent ability decreased with time, meanwhile the activity of UC-MSCs reduced to81～53%, cell adherence fell to72%～38%after6hours.2. By saving UC-MSCs in variable concentrations of serum, the activity of UC-MSCs within2.5%serum concentration was86%at6hours later. Though activity dropped to77%after6hours, the effect of preservation was far beyond against0.9%Sodium Chloride Injection without infusion of serum. The viable cells remained competent of adherence, the potency of multiple differentiation potential and surface marker tests of cells still complied with the standard of UC-MSCs.3. The vitality of cells was90%by keeping UC-MSCs within self-made protection solution at6hours afterwards, and showed a obvious downtrend with88%at9hours and80%at12hours separately. The ability of adherence remained, and the capacity of multiple differentiation potential and surface markers of cells complied with the standard of UC-MSCs still.Conclusion1. The phenomenon of mass cell death and significant declined viability and adherence ability of cells was observed after6hours preservation of UC-MSCs within clinical protective solution.2. Clearly2.5%concentration of serum had a positive effect with regard to protection of cell in contrast with other serum concentration. However the outstanding decline of cell archaeus along with incremental concentration of serum might be caused by various serum-involved unknown components which are probably toxic to cells.3. The stability and viability of preserved UC-MSCs could be significant improved by using self-made protection solution, providing experimental basis for UC-MSCs based clinical transplantation.