Singh Lipi, Bakshi Dapinder K, Vasishta Rakesh Kumar, Arora Sunil Kumar, Majumdar Siddarth, Wig Jai Dev
Department of General Surgery, Postgraduate Institute of Medical Education and Research, Chandigarh, India.
Dig Dis Sci. 2008 Sep;53(9):2569-75. doi: 10.1007/s10620-007-0162-1. Epub 2008 Feb 2.
The acinar cell culture plays a very important role in research of pancreatic pathophysiology. The aim of this study was to establish a long-term culture of human (foetal) pancreatic acinar cells in standardized nutrient media with supplements. Acinar cells were prepared from pancreatic tissues obtained from aborted foetus (> or =35 weeks) with no prior pancreatic complications by collagenase digestion and cultured using different media and supplements. The purity and phenotype of acinar cells was confirmed by various staining techniques and FACS. The acinar cell proliferation was determined at different time intervals by Bromo-deoxyuridine (BrdU) incorporation, and metabolic enzyme activity was analysed. The acini could be cultured and maintained in Ham's F-12 K/M199 media in the presence of 5% BSA, 0.1 mg/ml STI, 10 ng/ml EGF, and 10% FCS with the same morphological appearance as that of freshly prepared for 12 days with maximum viability of 80-85% and formation of monolayer without extracellular matrix. A significant BrdU incorporation of acinar cells in primary culture was observed which was maximum (105%) at day four. Higher amylase and lipase activity was seen in freshly isolated acinar cells which decreased with time of the culture. The established human pancreatic acinar cell culture may act as an excellent model to study exocrine dysfunction or pancreatitis in response to acinar cell injury.
腺泡细胞培养在胰腺病理生理学研究中发挥着非常重要的作用。本研究的目的是在添加了补充剂的标准化营养培养基中建立人(胎儿)胰腺腺泡细胞的长期培养体系。通过胶原酶消化从无胰腺并发症的流产胎儿(≥35周)获得的胰腺组织中制备腺泡细胞,并使用不同的培养基和补充剂进行培养。通过各种染色技术和流式细胞术(FACS)确认腺泡细胞的纯度和表型。通过溴脱氧尿苷(BrdU)掺入法在不同时间间隔测定腺泡细胞增殖,并分析代谢酶活性。在含有5%牛血清白蛋白(BSA)、0.1mg/ml大豆胰蛋白酶抑制剂(STI)、10ng/ml表皮生长因子(EGF)和10%胎牛血清(FCS)的Ham's F-12K/M199培养基中,可以培养并维持腺泡,其形态与新鲜制备的相同,可维持12天,最大活力为80-85%,且无需细胞外基质即可形成单层。在原代培养中观察到腺泡细胞有显著的BrdU掺入,在第4天达到最大值(105%)。新鲜分离的腺泡细胞中淀粉酶和脂肪酶活性较高,随培养时间降低。所建立的人胰腺腺泡细胞培养体系可作为研究腺泡细胞损伤后外分泌功能障碍或胰腺炎的优良模型。