高原缺氧条件对大鼠肠黏膜组织及缺氧诱导因子-1α、诱导型一氧化氮合酶表达的影响

Influence of plateau hypoxia on the tissue injury and expression of HIF-1α and iNOS in intestinal mucosa of rats

目的探讨高原缺氧条件对大鼠肠黏膜组织形态以及缺氧诱导因子(HIF)-1α和诱导型一氧化氮合酶(iNOS)表达的影响。方法将40只雄性Wistar大鼠均分为平原对照组及高原24h、48h、72h组,每组10只。平原对照组在常温环境下饲养1个月后处死,高原组在平原饲养1个月后于24h内急运至高原,建立高原缺氧模型,分别于到达高原后24、48、72h处死。分别在光镜和电镜下观察大鼠空肠上段的微观组织学变化,用图像测量软件测量绒毛高度、隐窝深度、黏膜厚度及绒毛表面积。免疫组化染色检测HIF-1α及iNOS在肠黏膜中的表达。结果高原24h组肠绒毛高度、隐窝深度、黏膜厚度及绒毛表面积分别为313.10±10.84、123.10±2.64、432.60±7.37μm和0.09±0.01mm2,与平原对照组(分别为322.00±12.68、127.70±7.24、447.50±21.93μm和0.10±0.01mm2)相比差异无统计学意义(P>0.05),高原48h组上述指标分别为279.00±9.80、101.10±7.11、376.40±19.05μm和0.06±0.01mm2高原72h组上述指标分别为235.80±19.54、76.10±7.58、311.90±18.38μm和0.05±0.01mm2,均显著低于平原对照组,差异有统计学意义(P<0.05),且高原72h组低于高原48h组(P<0.05)。高原缺氧组24、48、72h时组织中HIF-1α和iNOS的平均光密度值分别为0.24±0.03和0.16±0.05、0.34±0.02和0.25±0.03、0.45±0.03和0.37±0.02,均高于平原对照组(0.13±0.03和0.07±0.02),差异有统计学意义(P<0.05),且高原72h组高于48h组,48h组高于24h组(P<0.05)。高原24、48、72h组肠黏膜组织中HIF-1α及INOS的表达均呈正相关(r1=0.844,r2=0.831,r3=0.863,P<0.05)。结论高原缺氧条件可使肠黏膜产生组织损伤,并诱导肠黏膜中HIF-1α和iNOS蛋白的表达上调;HIF-1α可能参与了肠组织iNOS蛋白的表达过程和肠黏膜损伤。

Objective To study the influence of hypoxia at high altitude on the tissue injury and expression of hypoxia-inducible factor 1 （HIF-1α） and nitric oxide synthase （iNOS） in intestinal mucosa of rats. Methods Forty male Wistar rats weighing 250-300g were randomly divided into 4 groups （10 each）： plain control group, high altitude 24h, 48h and 72h group. Rats in plain control group were sacrificed after being fed under environment of plain for one month. Rats in high altitude groups were transported quickly to plateau area 24h after being fed on plain for one month to induce high altitude hypoxia model, and then were sacrificed in 24h, 48h and 72h, respectively. Gut mucosal morphology was observed under light microscope and/or electron microscope. Villous height, crypt depth, mucosal wall thickness and villous surface area were measured by Image Measurement Software. The expression of HIF-1α and iNOS were determined by immunohistochemistry （SP）. Results The mucosal villous height, crypt depth, mucosal thickness and villous surface area were respectively 313.10±10.84μm, 123.10±2.64μm, 432.60±7.37μm and 0.092±0.011mm2 in high altitude 24h group, and they were 322.00±12.68μm, 127.70±7.24μm, 447.50±21.93μm and 0.101±0.010mm2 in plain control group. No significant difference was found between the two groups （P0.05）. The corresponding indices in high altitude 48h and 72h groups （279.00±9.80μm, 101.10±7.11μm, 376.40±19.05μm and 0.063±0.013mm2, and 235.80±19.54μm, 76.10±7.58μm, 311.90±18.38μm and 0.047±0.008mm2, respectively） decreased obviously compared with those in plain control group （P0.05）. These morphological indices in high altitude 72h group were lower than those in high altitude 48h group. The mean optical density of iNOS and HIF-1α in high altitude 24h, 48h and 72h groups were 0.238±0.028 and 0.162±0.049, 0.339±0.024 and 0.247±0.029, and 0.445±0.032 and 0.365±0.022, respectirely, which were higher than those in plain control group （0.128±0.030 and 0.068±0.022, P0.05）. iNOS protein was positively correlated with HIF-1α in 3 high altitude groups （r1=0.844, r2=0.831, r3=0.863, respectively, P0.05）. Conclusion Plateau hypoxia may induce mucosal damage and up-regulated expression of HIF-1α and iNOS, implying that HIF-1α may play an important role in the expression of iNOS and intestinal mucosal injury.