| Because of the similarity of hydrogel materials with extracellular matrix on construction and properties,the hydrogel materials were widely used in tissue engineering and regenerative medicine.And the encapsulation of stem cell by hydrogel materials,therefor using as a platform for stem cell 3D culture,has been expanding its application on tissue engineering and regenerative medicine.Gelatin is the partial degradation products of collagen and can be used as a kind of excellent natural hydrogel material for the application of cell culture and drug delivery.There are many researches about gelatin hydrogel microsphere with loading drugs,but less about encapsulation of stem cells,especially single cells,which can be used as a platform of cell loading to study cell 3D cell culture and their therapeutical effects.We made gelatin hydrogel microsphere of gelatin which diameter was about 1mm by using gelatin as the scaffold material and method of microsphere forming in organic phase and catalytic cross-linking by microbial Transglutaminase(mTG),at the same time,adding the bone marrow mesenchymal stem cells(MSC)derived from SD rat,achieving hydrogel microsphere with encapsulation of MSC.Hydrogel microspheres in different concentration had been made with different concentration of gelatin solution respectively,and the swelling rate and modulus of different concentration of gelatin hydrogel were investigated,as well as the internal and surface morphological structure of hydrogel microsphere.We discovered that the gelatin microsphere cross-linked by enzyme would shrink after the water-balance.And then,the cell viability and proliferation of MSC encapsulated within the microsphere were tested,showing that MSC would spread its antennae and grow in the microsphere.MSC would not proliferate but their viability could maintain for about one week,and most of MSC would gradually die in the following days.However,during one week's culture,some of the encapsulated MSC would migrate to the surface from the interior of hydrogel microsphere and grow by adherence as well as proliferation on the surface until the cell sheet completely coats the whole hydrogel microsphere.This interesting phenomenon was seldom reported and may be benefit for the research and application of cell-in-hydrogel microsphere.Besides,we also constructed a cell surface engineering system for hydrogel encapsulated single MSC,which based on the hydrogel formation by enzymatic crosslinking.Using biocompatible anchors for membrane(BAM),enzyme which could catalyze for the formation of a layer of hydrogel was introduced onto cell surface,and encapsulation of a single cell by hydrogel shell was produced.We made two systems of hydrogel encapsulation for single cells including HRP-alginate hydrogel and mTG-gelatin hydrogel system.The effect of this encapsulation method on cellular functions was also investigated,found that mTG-gelatin hydrogel system would provide some protection effects for encapsulated cells under adverse condition,such as hypoxia and nutrition-deficient environment,at the same time,this single MSC-gelatin hydrogel system also had a better therapeutic effect on myocardial infarction(MI)of SD rat.In conclusion,by using of gelatin material and the method of microsphere forming in organic phase and enzyme catalytic cross-linking,we made gelatin hydrogel microsphere and the MSC encapsulated microsphere successfully,Some physical properties of hydrogel microsphere and biological representation of MSC in microsphere were investigated thoroughly.At the same time,we discovered an interesting phenomenon that part of MSC migrated to the surface from interior of microsphere.Our research may provide several inspirations for the research of 3D culture of stem cells in gelatin hydrogel and cell microcapsule based on gelatin microsphere which related to the field of tissue engineering and regenerative medicine.On the other hand,we constructed a system of hydrogel encapsulated single MSC and detected the effect on various cellular functions,discovering the protective effect of mTG-gelatin hydrogel system on MSC under adverse condition and well therapeutic effect on MI.These results may provide a new feasible solution for related research and applications of cell therapy. |
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