多系统腺苷信号的调节对外周神经病理性疼痛的影响*

THE EFFECTS OF REGULATION OF ADENOSINE SIGNALS FROM MULTISYSTEM ON PERIPHERAL NEUROPATHIC PAIN*

摘要:

目的：探索孕酮调节多系统腺苷上下游信号对坐骨神经结扎慢性损伤（CCI）模型大鼠外周神经病理性疼痛 (NeP)的影响。方法：雄性SD大鼠32只随机分为4组：假手术组（Sham+溶剂组，Sham+VEH组）（8只）、模型组（CCI+VEH组）（8只）、腺苷A1受体（A1R）拮抗剂组（DPCPX组，2 mg·kg-1）（8只）、孕酮治疗组（PROG组，12 mg·kg-1）（8只）。后两组于术后第14 d开始连续给药14天，DPCPX组先后给予DPCPX和孕酮，PROG组只给与孕酮。术前1天、术后第7、14、21和28天检测大鼠的机械性缩足阈值（MWT）和热缩足阈值（TWL）。术后第28天行为学检测后测定血清睾酮含量；甲醛灌注后取L4,5背根神经节和对应脊髓节段进行5'-核苷酸酶（CD73）免疫组化染色；取L4,5背根神经节和对应脊髓背角、睾丸和终纹床核（BST），检测CD73或A1R的蛋白表达。结果：与Sham+VEH组相比，其他三组MWT和TWL在术后第14 d降至最低（P＜0.01），且这三组间差异无统计学意义；治疗7天后PROG组MWT和TWL都比CCI+VEH组显著升高（P＜0.05），维持至第28 d，DPCPX组与CCI+VEH组间无明显差异。治疗14天后，PROG组血清睾酮水平比CCI+VEH组明显回升（P＜0.05），可被A1R拮抗剂抑制（P＜0.05）；背根神经节和脊髓背角PROG组比CCI+VEH组CD73阳性细胞明显增多（P＜0.05），拮抗A1R干扰此作用（P＜0.05），其A1R蛋白也显著高于DPCPX组（P＜0.05）；孕酮治疗后睾丸和BST降低的CD73蛋白和A1R蛋白都明显升高（P＜0.05），但拮抗睾丸和BST A1R时只有A1R蛋白的改变受影响（P＜0.05）。结论：孕酮治疗外周NeP时，生殖内分泌系统靶器官睾丸和BST与背根神经节和脊髓背角的腺苷上游信号CD73和腺苷下游信号A1R的改变，可能参与了从机体整体水平改善外周NeP的过程，拮抗A1R影响此效应和这些信号变化，但不显著影响睾丸和BST的腺苷上游信号CD73。

Abstract:

Objective To investigate the effects of progesterone that regulated the upstream and downstream signals of adenosine from multisystem on peripheral neuropathic pain (NeP) of chronic constriction injury (CCI) model of rats. Methods 32 male SD rats were divided into 4 groups: sham-operated group (Sham+vehicle group, Sham+VEH group) (n=8), model group (CCI+VEH group) (n=8), adenosine A1 receptor (A1R) antagonist group (DPCPX group) (2 mg·kg-1) (n=8), progesterone treatment group (PROG group) (12 mg·kg-1) (n=8). Rats of the two latter groups were receiving drug administration for 14 days continuously from the 14th day after surgery, and DPCPX and progesterone were injected one by one in the DPCPX group, but only progesterone was given in the PROG group. All the rats were tested with mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) for five times which were one day before surgery, the 7th, 14th, 21th, and the 28th day after surgery. The serum testosterone content was detected after behavioral tests on the 28th day after surgery; the L4,5 dorsal root ganglion and corresponding spinal dorsal horn were obtained to do immumohistochemical staining for 5'-nucleotidase (CD73) after formaldehyde perfusion; the L4,5 dorsal root ganglion and corresponding spinal dorsal horn, testis, and the bed nuclei of the stria terminals (BST) were gained to detect the protein expression of CD73 or adenosine 1 receptor (A1R). Results Compared with Sham+VEH group, results of MWT and TWL of the other three groups declined to the lowest at 14th day after surgery, and there were no statistical significance between and among these three groups. After treatment for 7 days, outcomes of MWT and TWL of PROG group were higher than the corresponding CCI+VEH group (P＜0.05), that lasted to the 28th day after surgery, and there was no difference between DPCPX group and CCI+VEH group. After treatment for 14 days, the serum testosterone level of PROG group rised obviously compared to CCI+VEH group (P＜0.05), that could be suppresed by the antagonist of A1R (P＜0.05); the posotive CD73 cells of PROG group of the dorsal root ganglion and the spinal dorsal horn were significantly higher than CCI+VEH group (P＜0.05), that could be interferenced through antagonizing A1R (P＜0.05), and the A1R proteins of these groups were noteworthily more than DPCPX group (P＜0.05); meanwhile the depressed CD73 and A1R proteins of testis and BST were apparently elevated after the treatment of progesterone(P＜0.05), however only the alterations of A1R proteins were affected when A1R of testis and BST was antagonized. Conclusion The regulation of upstream signals CD73 and downstream signals A1R of adenosine of testis and BST that were target organs of the reproductive endocrine system, dorsal root ganglion, and spinal dorsal horn might participate in the improvement of the peripheral NeP from the overall level of the body when treated with progesterone. Though this influence and these changes could be affected when A1R was antagonized, the upstream signals CD73 of adenosine of testis and BST were not influenced evidently.