一种改进的简便经济的大鼠胰腺腺泡细胞分离提纯方法

A simple economical modified method for isolating and purifying the rat pancreatic acinar cells

目的:探索一种简单、经济和稳定的胰腺腺泡分离纯化的新方法,并探讨其对细胞功能的影响.方法:16只SD大鼠随机分成改进方法组和内灌注方法组,戊巴比妥腹腔麻醉后,其中8只按改进方法迅速切取并剪碎胰腺组织后置于新鲜配制的细胞分离液中,37℃,CO2孵箱中孵育30min,过滤获得细胞悬液,Hanks液(不含钙离子)清洗后获得胰腺腺泡细胞,另外8只采用内灌注方法获取胰腺腺泡细胞,然后进行细胞计数、活力测定、纯化鉴定和细胞内钙离子测定.结果:改进方法所获得的胰腺腺泡数量[(5.0±0.7)×106]、纯度[(78±6)%]与内灌注方法所获得的胰腺腺泡数量[(4.7±0.9)×106]、纯度[(79±7)%]比较无统计学意义(P>0.05),而细胞活力[(93±4)%]显著高于内灌注方法[(84±3)%](P<0.05),细胞内钙离子浓度(3.62±0.54)显著低于内灌注方法(5.31±0.37)组(P<0.05).结论:改进方法是一种稳定高效且对细胞活力及功能影响较小的胰腺腺泡细胞分离提纯方法.

AIM： To develop a simple and economical modified method for isolating and purifying the rat pancreatic acinar cells and to study its effect on the cell function. METHODS： Sixteen SD rats were randomly divided into 2 groups, modified method group and intraductal perfusion method group （ n = 8 ）. In modified method group, the pancreatic tissues were obtained and shattered immediately after intraabdominal anesthesia, and then were incubated with cellular separating medium （ collagenase I, 0. 5 mg/mL） in 37℃, 50 mL/L CO2 for 30 min. The cell suspensions were obtained through two-time filtration and the pancreatic acinar cells were harvested after emumlation with Hanks without Ca^2＋. In the other group, the pancreatic acinar cells were taken by intraductal perfusion method. The cell count, cell activities and purification and intracellular calcium concentration were assessed. RESULTS： The cell count [ （5.0 ± 0.7 ） × 10^6 ] and purification [ （78 ± 6 ） % ] of the pancreatic acinar cells harvested by the modified method had no statistical difference compared with those by the intraductal perfusion method [ （4.7 ±0.9） × 10^6, （79 ± 7） %, P 〉 0. 05 ]. The cell activity [ （ 93 ± 4 ） % ] of modified method group was higher while the concentration of intracellular calcium （3.62 ± 0. 54 ） was lower than that of intraductal perfusion methed group [（84 ±3）%, 5.31 ±0.37, P 〈0.05]. CONCLUSION： Modified method is a simple, stable and effective method for isolating and purifying the pancreatic acinar cells with less affect on cell function.