基于天然胰腺脱细胞三维支架的外分泌胰腺体外实验

A natural pancreatic 3D bioscaffold platform for in vitro study on exocrine pancreas

目的 体外评估大鼠天然全胰腺脱细胞三维支架（3 D-APB）促进种植细胞增殖和分化的功能.方法 选取10只新鲜大鼠胰腺组织,利用改良灌注技术制备大鼠3D-APB,评估其生物相容性.实验分为四组：将体外培养好的AR42J种植在四组3D环境中,即空白对照组（无组织1640培养基）、细胞外基质（ECM）组（加入ECM成分的1640培养基）、聚乳酸-羟基乙酸共聚物（PLGA）组（加入PLGA支架的1640培养基）和3D-APB组（加入3D-APB的1640培养基）,分别观察4组细胞培养后第3、5、7、10天的增殖活性和细胞分化功能.结果 3D-APB在皮下埋植后引起有益的重塑反应,生物相容性好;MTT结果提示3D-APB组细胞增殖率高于其他3组,而凋亡率低于其余3组,差别具有统计学意义（P＜0.05）;蛋白印迹（Western blot）法提示3D-APB组PDX-1和PTF-1蛋白表达均高于其余3组,差别均具有统计学意义（均P＜0.05）;实时定量PCR（qRT-PCR）结果提示3D-APB组PDX-1和PTF-1基因表达含量均高于其余3组,差别均具有统计学意义（均P＜0.05）.结论 与目前常用的化学合成支架和天然成分支架相比,3D-APB支架更有利于促进细胞增殖及分化,更适合应用于再生医学领域.

Objective To evaluate the biochemical function of rat nature three-dimensional acellular pancreatic bioscaffold （3D-APB） in promoting cell proliferation and differentiation in vitro.Methods The fresh pancreas from 10 rats were perfused to prepare 3D-APB.The biocompatibility of 3D-APB was eva luated.The experiment was divided into 4 groups based on 4 kinds of three-dimensional media for AR42J culture,including blank control group,ECM group,PLGA group and 3D-APB group.We compared the proliferation and differentiation of AR42J pancreatic acinar cell at 3 days,5 days,7 days and 10 days after seeding among 4 groups.Results The 3D-APB could represent a biocompatible scaffold capable of integrating within host tissue.The proliferation rate of AR42J cells by MTT in 3D-APB was higher,while the apoptosis rate by flow cytometry was lower than those in other 3 media,which were all significantly different （P 〈 0.05）,respectively.The protein expression of PDX-1 and PTF-1 by western blot in 3D-APB group was greatly higher than those in other 3 groups （both P 〈 0.05）.The mRNA expression of PDX-1 and PTF-1 through qRT-PCR was significantly higher than those in other 3 groups （both P 〈 0.05）.Conclusion Compared with the commonly used chemical and natural scaffold at present,3D-APB could promote cell proliferation and differentiation,which was more appropriate for regenerative medicine.