神经生长因子对哮喘豚鼠内脏感觉传入部位MMP-2表达的影响

目的探讨基质金属蛋白酶-2(MMP-2)在哮喘豚鼠内脏感觉传入部位的表达及神经生长因子(NGF)对MMP-2的调节作用。方法应用免疫组织化学方法检测豚鼠内脏感觉传入部位MMP-2免疫反应的变化。用W estern b lot蛋白印迹方法检测豚鼠C7~T5段脊神经节和相应节段脊髓背角NGF和MMP-2的表达。用Luzex-F实时图像分析系统和凝胶成像分析系统分别对结果进行图像分析。结果①免疫组织化学结果显示:豚鼠C7~T5段脊神经节和相应节段脊髓背角内MMP-2阳性免疫反应产物的平均灰度值(0-深色,255-浅色),生理盐水组(175.50±4.67、166.47±6.20)与单纯致敏组(172.63±9.51、165.20±5.14)比较没有显著差异(P>0.05);哮喘组(139.87±4.88、125.93±4.49)明显低于生理盐水组和单纯致敏组(P<0.01);哮喘+NGF抗体组(鼻腔吸入NGF抗体,169.73±6.24、152.37±8.67)则明显高于哮喘组(P<0.01)。②W estern b lot结果显示:与生理盐水组和单纯致敏组比较,哮喘组豚鼠C7~T5段脊神经节及相应节段脊髓样品中MMP-2和NGF阳性产物的IDV(Integrated Density Value)与β-actin IDV的比值均明显升高(P<0.01),而哮喘+NGF抗体组则明显低于哮喘组(P<0.01),表明NGF可上调豚鼠内脏感觉传入部位MMP-2的表达。结论C7~T5段脊神经节以及相应节段脊髓背角内的MMP-2可能也参与哮喘的发病过程,NGF诱发MMP-2的表达可能是NGF参与哮喘发病的机制之一,NGF抗体抑制哮喘时MMP-2的表达有可能延缓气道重塑的发生,可能是治疗哮喘的新途径。

The Upregulative Effect of NGF on MMP-2 in the Viscerosensory Afferent Sites of the Asthmatic Guinea Pig

Objective To explore the expression of matrix metalloproteinase-2(MMP-2) and the regulatory effect of nerve growth factor(NGF) in the viscerosensory afferent sites of the asthmatic guinea pig.Methods The changes of MMP-2 immunoreactivity were investigated by means of immunohistochemistry in the viscerosensory afferent sites of the guinea pigs in different groups.The expression of NGF and MMP-2 were observed by Western blot in the C_(7)-T_(5) spinal ganglia and the corresponding spinal dorsal horn of all the groups.The results were analyzed by the Luzex-F real time image analysis system and the gel imaging analysis system respectively.Results ①Immunohistochemistry results showed that there was no obvious difference for the mean grey values(0-black to 255-white) of MMP-2 positive products in the C_(7)-T_(5) spinal ganglia and the corresponding spinal dorsal horn between normal saline control group(175.50±4.67,166.47±6.20)and simply sensitized group(172.63±9.51,165.20±5.14)(P0.05).The mean grey values of MMP-2 positive products decreased significantly in the C_(7)-T_(5) spinal ganglia(139.87±4.88)and the corresponding spinal dorsal horn(125.93±4.49)of the asthmatic guinea pigs compared with the normal saline control group and the simply sensitized group(P<0.01).However,the mean grey values were much higher in NGF antibody group(inhalation of NGF antibody into nasal cavity,169.73±6.24、152.37±8.67) than in asthmatic group(P<0.01).②Western blot results showed that compared with normal salinecontrol group andsimply sensitized group,the ratios between integrated density value(IDV) for MMP-2 or NGF protein band and IDV for β-actin protein band increased markedly in the C_(7)-T_(5) spinal ganglia and the corresponding spinal dorsal horn of asthmatic group(P<0.01).However,the ratios of IDV for NGF antibody group reduced markedly in comparison with asthmatic group(P<0.01),indicating that NGF can upregulate the expression of MMP-2 in the viscerosensory afferent sites.Conclusion The present results indicate that the MMP-2 in the C_(7)-T_(5) spinal ganglia and the corresponding spinal dorsal horn might be involved in the pathogenesis of the bronchial asthma,the overexpression of MMP-2 induced by NGF might be one of the mechanisms for the involvement of NGF in the pathogenesis of asthma,and inhibitory expression of MMP-2 by NGF antibody might inhibit airway remodeling and be a new route for the prevention and treatment of the bronchial asthma.