优化海藻酸钠-壳聚糖多孔复合支架用于人脐带来源间充质干细胞的体外扩增
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摘要
目的通过制备及优化海藻酸钠-壳聚糖多孔复合支架用于人脐带来源的间充质干细胞体外扩增,为干细胞在组织工程中的应用与疾病治疗提供实验基础。方法通过制备海藻酸钠-壳聚糖多孔复合支架,并接枝多聚赖氨酸对其优化。然后,从产妇脐带中分离出间充质干细胞,接种在复合支架表面,监控其增殖情况。利用细胞计数法和CCK-8试剂盒,绘制细胞增殖曲线,并用β-半乳糖苷酶衰老检测试剂盒评估干细胞的衰老程度。结果成功制备并优化了海藻酸钠-壳聚糖复合支架,发现在接枝多聚赖氨酸的支架上接种间充质干细胞,细胞能够很好地附着,覆盖在复合支架表面,且能快速增殖。结论优化的海藻酸钠-壳聚糖多孔复合支架能为细胞增殖提供优良环境,具备体外扩增间充质干细胞的可行性,可作为组织工程应用的潜在材料。
Objective Alginate-chitosan porous composite scaffold is prepared and optimized to culture human umbilical cord-derived mesenchymal stem cells in order to provide the application of stem cells in tissue engineering and disease treatment.Methods We prepared porous composite scaffold, and was optimized by grafting polylysine on its surface. Next, mesenchymal stem cells were isolated from umbilical cord, seeded and serially cultured in the scaffolds. Cell growth and proliferation were monitored, and cell proliferation curves were made using cytometry and CCK-8 kit techniques. Moreover, stem cell ageing was evaluated by senescence β-Galactosidase staining kit. Results This technology successfully prepared and optimized the alginate-chitosan composite scaffold. The mesenchymal stem cells easily attached to and cover the surface of the scaffold grafted with polylysine. Moreover, the stem cells grew in agglomerates rapidly. Conclusions The alginate-chitosan scaffold provides a suitable environment for cell growth and has the feasibility for cell culture in vitro. This scaffold can be used as potential material for tissue engineering research and production.
Objective Alginate-chitosan porous composite scaffold is prepared and optimized to culture human umbilical cord-derived mesenchymal stem cells in order to provide the application of stem cells in tissue engineering and disease treatment.Methods We prepared porous composite scaffold, and was optimized by grafting polylysine on its surface. Next, mesenchymal stem cells were isolated from umbilical cord, seeded and serially cultured in the scaffolds. Cell growth and proliferation were monitored, and cell proliferation curves were made using cytometry and CCK-8 kit techniques. Moreover, stem cell ageing was evaluated by senescence β-Galactosidase staining kit. Results This technology successfully prepared and optimized the alginate-chitosan composite scaffold. The mesenchymal stem cells easily attached to and cover the surface of the scaffold grafted with polylysine. Moreover, the stem cells grew in agglomerates rapidly. Conclusions The alginate-chitosan scaffold provides a suitable environment for cell growth and has the feasibility for cell culture in vitro. This scaffold can be used as potential material for tissue engineering research and production.
引文
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[2] Bharadwaj S,Liu G,Shi Y,et al.Multipotential differentiation of human urine-derived stem cells:potential for therapeutic applications in urology[J].Stem Cells,2013,31:1840-1856.
[3] Reitinger S,Wissenwasser J,Kapferer W,et al.Electric impedance sensing in cell-substrates for rapid and selective multipotential differentiation capacity monitoring of human mesenchymal stem cells[J].Biosens Bioelectron,2012,34:63-69.
[4] Barberini DJ,Freitas NP,Magnoni MS,et al.Equine mesenchymal stem cells from bone marrow,adipose tissue and umbilical cord:immunophenotypic characterization and differentiation potential[J].Stem Cell Res Ther,2014,5:1-11.
[5] Michler JK,Hillmann A,Savkovic V,et al.Horse hair follicles:A novel dermal stem cell source for equine regenerative medicine[J].Cytometry A,2017,93,104-114.
[6] Elasfar RK,Kamal MM,Abd ER,et al.Obestatin can potentially differentiate Wharton's jelly mesenchymal stem cells into insulin-producing cells[J].Cell Tissue Res,2017,372:91-98.
[7] Lin DH,Biswas A,Choolani M,et al.Induction of immunogenic cell death in lymphoma cells by Wharton's jelly mesenchymal stem cell conditioned medium[J].Stem Cell Rev,2017,13:801-816.
[8] Tiet P,Berlin J.Exploiting homing abilities of cell carriers:targeted delivery of nanoparticles for cancer therapy[J].Biochem Pharmacol,2017,145:18-26.
[9] Yan WU,Jie YU,Zhang L,et al.Study on the immunoregulation of mesenchymal stem cells from patients with aplastic anemia[J].Chin J Pract Pediatrics,2009,24:786-790.
[10] Nojehdehian H,Moztarzadeh F,Baharvand H,et al.Preparation and surface characterization of poly-L-lysine-coated PLGA microsphere scaffolds containing retinoic acid for nerve tissue engineering:in vitro study[J].Colloids and Surfaces B:Biointerfaces,2009,73:23-29.
[11] Zheng JN,Xie HG,Yu WT,et al.Chitosan-g-MPEG-modified alginate/chitosan hydrogel microcapsules:a quantitative study of the effect of polymer architecture on the resistance to protein adsorption[J].Langmuir,2010,26:17156-17164.
[12] Jain D,Bar-Shalom D.Alginate drug delivery systems:application in context of pharmaceutical and biomedical research[J].Drug Dev Ind Pharm,2014,40:1576-1584.