实验性支气管哮喘豚鼠肺和内脏感觉传入系统蛋白激酶C的表达及神经生长因子的调节作用

The expression of protein kinase C and the regulatory effect of nerve growth factor in the lung and the visceral sensory afferent system of asthmatic guinea pigs

目的探讨支气管哮喘(简称哮喘)豚鼠肺及内脏感觉传入系统(C7～T5脊神经节及对应的脊髓后角)蛋白激酶C(PKC)的表达及神经生长因子(NGF)的调节作用。方法40只豚鼠按随机数字表法分为4组生理盐水对照组(A组)8只、单纯致敏对照组(B组)8只、哮喘组(C组)12只和NGF抗体组(D组)12只。用免疫组织化学方法检测各组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的免疫反应变化;采用Westernblot方法分别检测各组豚鼠肺组织、C7～T5脊神经节和对应的脊髓后角NGF与PKC的表达;采用LuzexF实时图像分析系统和凝胶成像分析系统分别对以上结果进行图像分析。结果(1)免疫组织化学A、B组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的平均吸光度(A)值分别为0.102±0.009、0.113±0.009、0.106±0.005、0.116±0.007,二者比较差异无统计学意义(P>0.05)。C组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的A值分别为0.215±0.014、0.176±0.010,C组与A组比较差异有统计学意义(P<0.01)。D组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的A值分别为0.140±0.008、0.130±0.011,D组与C组比较差异有统计学意义(P<0.01)。(2)Westernblot蛋白印迹C组肺组织、C7～T5脊神经节神经元及相应脊髓节段PKC蛋白表达量(其A相对值分别为1.51±0.01、1.40±0.03、2.22±0.02)与A组(其A相对值分别为0.51±0.02、0.43±0.01、0.92±0.02)和B组比较均明显增加,而D组PKC蛋白表达量(其A相对值分别为0.80±0.03、0.83±0.01、1.12±0.02)明显低于C组。结论PKC可能参与哮喘的发病过程,而NGF可上调哮喘豚鼠PKC的表达。

Objective To explore the expression of protein kinase C （PKC） and the regulatory effect of nerve growth factor（ NGF）in the lung and the visceral sensory afferent system（C7-T5 spinal ganglia and the corresponding posterior horn of the spinal cord） of asthmatic guinea pigs. Methods Forty guinea pigs were divided to four groups, a saline control group （ group A, n = 8 ）, a provocation alone control group （group B, n =8）, an asthmatic group （group C, n = 12） and an anti-NGF group （group D, n = 12）. The alterations of PKC immunoreactivity were investigated by means of immunohistochemistry in the C7-T5 spinal ganglia and the corresponding posterior horn of the spinal cord of all groups. The expressions of NGF and PKC were investigated by Western blot in the lung, CT-T5 spinal ganglia and the corresponding posterior horn of the spinal cord of all groups. The results were analyzed by the Luzex-F real time image analysis system and the gel imaging analysis system respectively. Results （1） Immunohistochemistry results： The absorbency （ A ） values of PKC were 0. 102 ± 0. 009, 0. 113 ± 0. 009, 0. 106 ± 0. 005 and 0. 116 ± 0. 007 in the C7-T5 spinal ganglia and the corresponding posterior horn of the spinal cord of group A and group B respectively. There was no significant difference between group A and group B （ P 〉 0. 05 ）. The A values of PKC were 0. 215 ± 0. 014 and 0. 176 ±0. 010 respectively in the C7-T5 spinal ganglia and the corresponding posterior horn of the spinal cord of group C, which were significantly different compared with group A （ P 〈 0. 01 ）. The A values of PKC were 0. 140 ±0. 008 and 0. 130 ±0. 011 respectively in the C7-T5 spinal ganglia and the corresponding posterior horn of the spinal cord of group D, which were significantly different compared with group C （ P 〈 0. 01 ）. （2） Western blot results： Compared with group A （ the relative A values were 0. 51 ± 0. 02,0.43 ± 0. 01 and 0. 92 ± 0. 02 respectively ） and group B, the expression of PKC （the relative A values were 1.51 ±0. 01,1.40 _±0. 03 and 2. 22 ±0. 02 respectively） increased markedly in the lung, C7-T5 spinal ganglia and the corresponding posterior horn of the spinal cord of group C; however, the expression of PKC of group D （ the relative A values were 0. 80 ± 0. 03,0. 83 ± 0. 01 and 1. 12 ± 0. 02 respectively） decreased markedly in comparison with group C. Conclusion The present results indicate that PKC might be involved in the pathogenesis of bronchial asthma, and NGF can upregulate the expression of PKC.