靶向μ-阿片受体的 RNA 干扰逆转了芬太尼对大鼠胰岛葡萄糖刺激胰岛素释放的抑制作用。

RNA interference targeting mu-opioid receptors reverses the inhibition of fentanyl on glucose-evoked insulin release of rat islets.

机构信息

Department of Anesthesiology, Shenzhen Shekou People's Hospital, Shenzhen, Guangdong 518067, China.

出版信息

Chin Med J (Engl). 2010 Dec;123(24):3652-7.

Abstract

BACKGROUND

Mu opioid receptor plays an important role in many physiological functions. Fentanyl is a widely used opioid receptor agonist for analgesia. This study was conducted to test the role of mu-opioid receptor on insulin release by determining whether fentanyl affected insulin release from freshly isolated rat pancreatic islets and if small interfering RNAs (siRNA) targeting mu-opioid receptor in the islets could knock down mu-opioid receptor expression.

METHODS

Islets were isolated from ripe SD rats' pancreas by common bile duct intraductal collagenase V digestion and purified by discontinuous Ficoll density gradient centrifugation. The siRNA knock-down of mu-opioid receptor mRNA and protein in islet cells was analyzed by semi-quantitative real time-PCR and Western blotting. After siRNA-transfection for 48 hours, the islets were co-cultured with fentanyl as follows: 0 ng/ml, 3 ng/ml and 30 ng/ml for 48 hours. Then glucose-evoked insulin release was performed. As a control, the insulin release was also analyzed in islets without siRNA-trasfection after being co-cultured with fentanyl for 48 hours.

RESULTS

After 48 hours of transfections, specific siRNA targeting of mu-opioid receptors produced significant reduction of mu-opioid receptor mRNA and protein (P < 0.01). Fentanyl significantly inhibited glucose-evoked insulin release in islets in a concentration dependent manner (P < 0.01). But after siRNA-transfection for 48 hours, the inhibition on glucose-evoked insulin release was reversed (P < 0.01).

CONCLUSIONS

RNA interference specifically reduces mu-opioid receptor mRNA and protein expression, leading to reversal of the fentanyl-induced inhibition on glucose-evoked insulin release of rat islets. The activation of opioid receptor induced by fentanyl functions to inhibit insulin release. The use of RNAi presents a promising tool for future research in diabetic mechanisms and a novel therapy for diabetes.

摘要

背景

μ 阿片受体在许多生理功能中发挥重要作用。芬太尼是一种广泛用于镇痛的阿片受体激动剂。本研究通过测定芬太尼是否影响新鲜分离的大鼠胰岛胰岛素释放，以及胰岛中小干扰 RNA（siRNA）是否可以敲低μ阿片受体，来检测μ阿片受体在胰岛素释放中的作用。

方法

采用胆总管胶原酶 V 消化和不连续 Ficoll 密度梯度离心法从成熟 SD 大鼠胰腺中分离胰岛，采用半定量实时 PCR 和 Western blot 分析胰岛细胞中 μ 阿片受体 mRNA 和蛋白的 siRNA 敲低。siRNA 转染 48 小时后，将胰岛与芬太尼共培养，浓度分别为 0ng/ml、3ng/ml 和 30ng/ml，共培养 48 小时，然后进行葡萄糖刺激的胰岛素释放。作为对照，未经 siRNA 转染的胰岛在与芬太尼共培养 48 小时后，也进行胰岛素释放分析。