| Objective: Theaim of the study was toprepare PLGA mixed gel(PLGA-chitosan / PLG-sodium alginate nanoparticle blends), and seed adipose stem cells(ADSCs) on the gel as biodegradable scaffold to construct tissue engineering bladders and analysis of its advantages and disadvantages.Methods: 1. Poly-lactic-co-glycolic acid(PLGA), sodium alginate(SA) and chitosan(CS) were used to prepare PLGA-CS / PLGA- SA mixed gel by the double emulsion-solvent evaporation technique. Nanoparticle diameter and potential tests was to confirm the properties of the nanoparticles. Scanning electron micrographs(SEM) of dried mixed gel revealed the 3D micro-porous structure. 2. Getting bladder submucosa from pigs, and treated it with chemical detergents to remove cellular components. Detecting the bladder submucosa acellular matrix with HE staining and Masson trichrome staining methods. 3. The groin fat of SD rats was digested with collagen enzyme?to extract ADSCs. The 3-5 generations of ADSCs were used in experiment. Detect the cytotoxicity of composition such as sodium alginate, chitosan, Z1, sodium hyaluronate and gelatin to ADSCs by MTT method. We decided bladder submucosa acellular matrix(BSAM) as the control group, and PLGA mixed gel polymer as the test group. Both seeded the 3rd generation of ADSCs and cultured in culture dishes, and detected by HE staining when cultured 4 days and 7 days. Cell proliferation was observed in the two materials.Results:1. PLGA-CS polymers and PLGA- SA polymers were prepared by the double emulsion-solvent evaporation technique. The nanoparticles were detected by potential and particle size analyzer. The results showed that the particle diameter of PLGA-CS polymer nanoparticles was(202.2 ± 13.7) nm, and the potential was(14.41 ± 2.37) m V; the particle diameter of PLGA-SA polymer nanoparticles was(143.4 ± 5.2) nm, and the potential was(-24.72 ± 0.67) m V. The results close to thereference. 2. The results of HE staining and Masson trichromestaining showed that there were no residual cells in BSAM, and the fibers showed cross-linked structures. 3. After 24 hours of primary cultivation, a greatnumber of SD rats ADSCs became adherent and present short spindle or polygon in morphology. When the adherent cells fused to 80%- 90%, subculture the cells according to the ratio of 1:3 and take the third generation of ADSCs to detect by MTT assay. MTT assay showed that the proliferation of cells in material groups(CS group, SA group, gelatin group and Z1 group) were significantly better than the control groups(p<0.05). However, the groups of sodium hyaluronate present varying degree of influence on cell proliferation. HE staining showed that the three-dimensional structure of the PLGA-CS / PLGA-SAmixed gel scaffold add with gelatin and Z1, was more suitable than BSAM in promoting cells growth and proliferation.Conclusion: PLGA mixed gel modified by chitosan and alginate can be used as a three-dimensional scaffold, and scaffold seeded with ADSCs can promote cell growth and proliferation. By comparison, PLGA-CS / PLGA-SA mixed gel added with gelatin and Z1, had greatly enhanced cells growth and proliferation.This exploratory research provided a new alternative material for the tissue engineering to build a bladder. |
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