ZHANG Jianbo 1,2 , YANG Qian 1 , CHEN Jie 1 , MU Dezhi 3 , ZHANG Lin 3 , MAO Meng 3 , QU Yi 3
  • 1Institute of Organ Transplantation, Sichuan Provincial People’s Hospital, Chengdu Sichuan, 610072, P.R.China;;
  • 2Institute of Biochemistry and Molecular Biology, School of Forensic and Preclinical Medicine, Sichuan University;;
  • 3Department of Pediatrics, West China Second University Hospital, Sichuan University.;
Rat islet cell; Isolation and purification; Cell culture
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To set up an economic and effective method for islet isolation from rat, and thereby prove a laboratory protocol of animal model for cl inical islet transplantation. Methods Twenty-five adult male SD rats weighing 230-380 g were used as organ donor. In each of 5 repeated experiments, pancreatic islets of 5 animals were isolated by intraductal infusion of compound sodium chloride injection (CSCI), and subsequently, digested with low concentration (0.5 mg/mL)
of collagenase V solution. Islet purification was performed by using a discontinuous density gradient centrifugation that
was prepared with 27.0%, 23.0%, 20.5% and 11.0% of Ficoll 400. Islet yield and purity were determined by dithizon (DTZ)
stain, and propidium iodide (PI)/fluorescein diacetate (FDA) double stain was used to check viabil ity of islets. The endocrine secretory function was assessed by insul in secretion in either low (2.8 mmol/L) or high (25.0 mmol/L) glucose incubation after 3 days of culture in RPMI1640 media. Results Average islet digestion time of 5 experiments was (13.8 ± 1.6) min. Before purification, average isolated number was (5 626 ± 422) islets, and the number was significantly reduced to (2 914 ± 485) islets after purification (P  lt; 0.01). The average recovery rate was 51.6% ± 6.0%, and the average yield was (583 ± 97) islets/pancreas. The average purity and viabil ity of islets were 90.2% ± 3.4% and 81.6% ± 7.0%, respectively. After 3 days of culture, insul in secretion of the islets was (116.1 ± 17.4) EU/L in high glucose incubation, which was significantly higher than that of low glucose environment [(39.7 ± 7.5) EU/L, P  lt; 0.01)]. The average insul in stimulation index was 3.0 ± 0.4. Conclusion The islet isolation with the CSCI solution and digestion with low concentration of collagenase V decrease experimental cost and also have a beneficial effect on islet recovery and their function.

Citation： ZHANG Jianbo,YANG Qian,CHEN Jie,MU Dezhi,ZHANG Lin,MAO Meng,QU Yi. AN ECONOMIC AND EFFECTIVE METHOD FOR ISLET CELL ISOLATION FROM SD RATS. Chinese Journal of Reparative and Reconstructive Surgery, 2008, 22(5): 610-613. doi: Copy