高压氧对脊髓损伤大鼠 NALP3炎性体表达的影响

Effects of hyperbaric oxygen on the expression of inflammosome NALP3 in the rats after spinal cord injury

目的：建立脊髓损伤（ spinal cord injury ， SCI ）大鼠模型，探讨高压氧（ hyperbaric oxygen ， HBO）对SCI大鼠核苷酸结合寡聚化结构域样受体3（NACHT－LRR－PYD－containing protein 3， NALP3）炎性体表达的影响，为SCI损伤及修复机制研究奠定基础。方法成年SD大鼠120只，体质量250～300 g，按数字表法随机分为4组：SCI组，使用NYU脊髓损伤专用撞击器，以25 gcm致伤能量损伤大鼠T10脊髓节段；SCI＋HBO组，SCI造模后辅助以HBO处理；假手术对照组（ SH组），仅打开T10节段椎板，显露脊髓，不造成SCI；SH＋HBO组，假手术处理后辅以HBO；各组分别于造模后12 h和1、3、7、14 d处死，取脊髓损伤节段组织检查NALP3炎性体的表达。结果用相对定量2－ΔΔCT法分析RT－PCR结果显示在1、3、7 d时SCI ＋HBO 组的NALP3、凋亡相关斑点样蛋白（ adaptor molecule apoptosis－associated speck－like protein， ASC）、半胱天冬酶－1（cysteine aspartic acidspecific protease ， caspase－1）的mRNA含量与相应的SCI组相比明显降低（P＜0．05或P＜0．01），表明HBO处理可降低大鼠SCI组织的NALP3炎性体的mRNA表达水平；用软件灰度分析Western－blot结果显示，SCI＋HBO组与SCI组相比NALP3的灰度值在1、3、7 d明显降低，ASC和caspase－1的灰度值与SCI组相比在3、7 d明显降低（ P＜0．05或P＜0．01），表明HBO处理可降低SCI组织的NALP3、ASC、capase－1蛋白的表达。结论 HBO处理可降低大鼠SCI组织的NALP3、ASC、caspase－1的mRNA和蛋白的表达水平，通过抑制NALP3炎性体的激活而减轻脊髓继发性损伤，保护损伤区周围残存的神经细胞。

Objective To investigate the effects of hyperbaric oxygen （ HBO ） on the expression of inflammosome NALP3 in rats after spinal cord injury , so as to lay a good foundation for the research on the mechanism involved in the treatment and recovery of spinal cord injury （ SCI ） .Methods One hundred and twenty SD rats , with a body weight from 250 g to 300 g were divided into 4 groups：the sham surgery group （ or the control group）, the spinal cord injury group（or the SCI group）, the SCI＋HBO group, the sham surgery＋HBO group.In the control group, the vertebral lamina in T10 was surgically opened, with the spinal cord exposed, but without SCI.In the SCI group, injury of T10 section was inflicted with a force of 25 gcm.For the SCI＋HBO group, the animals received HBO therapy following development of the SCI model .In the sham surgery ＋HBO group, the animals also received HBO therapy .The animals of the 4 groups were sacrificed respectively at h 12, d 1, d 3, d 7 and d 14, and the tissues in the injured sections of the spinal cord were collected to detect the expression of inflammosome NALP 3.Results RT-PCR detection indicated that the values of NALP3, ASC and caspase-1 were decreased significantly at d 3 and d 7, as compared with those of＆amp;nbsp;the SCI group （P〈0.05）.The expression of NALP3 mRNA in the spinal cord of the SCI ＋HBO group [1 d：1.26 ±0.17, 3 d：1.14 ±0.02, 7 d：1.12 ±0.12] was significantly lower than that of the SCI group [1 d：1.54 ±0.05, 3 d：1.48 ±0.14, 7 d：1.45 ±0.15], with statistical significance （P〈0.05）.Detection by Western-Blot demonstrated that the grey values of NALP 3 at d1, d 3 and d 7 in the SCI ＋HBO group were significantly decreased , when compared with those of the SCI group , and the grey values of ASC and caspase-1 at d 3 and d 7 were also significantly decreased, when compared with those of the SCI group （P〈0.05）.The expression of NALP3 in the spinal cord of the SCI ＋HBO group [1 d：0.51 ±0.01, 3 d：0.44 ±0.06, 7 d：0.42 ±0.07] was significantly lower than that of the SCI group [1 d：0.58 ±0.02, 3 d：0.55 ±0.08, 7 d：0.50 ±0.03], also with statistical significance （P〈0.05）.Conclusions HBO could compromise the mRNA expressions of NALP3, ASC and caspase-1 in the damaged spinal cord tissue rats .Secondary damage to the spinal cord could be alleviated through the inactivation of NALP 3 inflammosome by HBO.At the same time, HBO could protect the remaining peripheral neural cells in the injured spinal cord tissue and promote neural recovery after SCI .