Role of PI3K/Akt signaling pathway in penehyclidine hydrochloride-induced inhibition of autophagy in intestinal tissues in a rat model of limb ischemia-reperfusion

Abstract

Objective To investigate the role of phosphatidylinositol 3-kinase/serine-threonine kinase (PI3K/Akt) signaling pathway in penehyclidine hydrochloride-induced inhibition of autophagy in intestinal tissues in a rat model of limb ischemia-reperfusion. Methods Sixty-four healthy adult male SPF Sprague-Dawley rats, weighing 220-250 g, were randomly divided into 4 groups (n=16 each) using a random number table: sham operation group (group S), limb ischemia-reperfusion group (group LIR), penehyclidine hydrochloride group (group PHC), and penehyclidine hydrochloride + PI3K inhibitor 3-methyladenine group (group PM). The animals were anesthetized with sevoflurane, and bilateral femoral arteries were clamped for 3 h followed by 4 h of reperfusion in LIR, PHC and PM groups.At 15 min before reperfusion, normal saline 1 ml was injected intravenously in S and LIR groups, penehyclidine hydrochloride 0.15 mg/kg was injected intravenously in group PHC, and penehyclidine hydrochloride 0.15 mg/kg was injected intravenously, and 3-methyladenine 10 mg/kg was injected intraperitoneally at the same time in group PM.At 4 h of reperfusion, venous blood samples were collected for determination of serum lipopolysaccharide (LPS) and lactic dehydrogenase (LDH) levels.The animals were sacrificed after blood sampling, and the small intestine was removed for determination of the expression of Beclin-1, microtubule-associated protein 1 light chain 3 Ⅰ (LC3Ⅰ) and LC3Ⅱ in intestinal tissues by Western blot.The ratio of LC3Ⅱ expression to LC3Ⅰexpression (LC3Ⅱ/LC3Ⅰ) was calculated. Results Compared with group S, the serum LPS and LDH levels were significantly increased, the expression of Beclin-1 in intestinal tissues was up-regulated, LC3Ⅱ/LC3Ⅰ was increased (P 0.05). Compared with group LIR, the serum LPS and LDH levels were significantly decreased, the expression of Beclin-1 in intestinal tissues was down-regulated, LC3Ⅱ/LC3Ⅰ was decreased (P 0.05). Compared with group PHC, the serum LPS and LDH levels were significantly increased, the expression of Beclin-1 in intestinal tissues was up-regulated, LC3Ⅱ/LC3Ⅰ was increased (P<0.05 or 0.01), and autophagosomes were increased in group PM. Conclusion The mechanism by which penehyclidine hydrochloride inhibits autophagy in intestinal tissues is related to activation of PI3K/Akt signaling pathway in a rat model of limb ischemia-reperfusion. Key words: 1-Phosphatidylinositol 3-kinase; Protein-serine-threonine kinases; Cholinergic agonists; Extremities; Reperfusion injury; Intestines; Autophagy