艾灸预处理对神经通路切断/损毁术后大鼠胃黏膜保护作用机制的研究
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摘要
目的：为探讨艾灸启动内源性保护信息对胃粘膜的保护效应与神经通路的关系,本研究选择性地切断/损毁实验动物神经通路中的脊髓、孤束核、迷走神经和腓总神经,应用组织形态学、生化及分子生物学等方法,观察艾灸对胃黏膜损伤的保护效应,并观察艾灸对胃黏膜保护相关内源性物质EGF、SS、IL-2及保护蛋白HSP-70表达的影响。
方法：SD大鼠随机分为7组,即A正常对照组；B模型对照组；C艾灸预处理+模型组；D艾灸预处理+模型+脊髓损毁组；E艾灸预处理+模型+孤束核损毁组；F艾灸预处理+模型+迷走神经切断组；G艾灸预处理+模型+腓总神经切断组。对涉及手术的各组大鼠按要求分别进行神经损毁或切断手术,术后采用无水酒精灌胃法制备大鼠胃黏膜损伤模型,以艾灸预处理足三里穴为被试因素,按Guth标准记录大鼠胃黏膜损伤指数,光镜下观察胃黏膜组织形态学变化；采用酶联免疫吸附法检测胃黏膜组织中EGF、SS、血清中IL-2的含量；采用酶联免疫吸附法(ELISA)和蛋白质印迹法(Western-blot)检测血清中和胃黏膜组织中热休克蛋白70(HSP-70)表达。
结果：
1.造模后,模型对照组大鼠胃黏膜损伤指数最高,正常组大鼠胃黏膜未见明显损伤,说明造模成功；艾灸预处理后,胃黏膜损伤大鼠的胃黏膜均有不同程度的修复,主要表现为胃黏膜损伤指数降低(P<0.01),胃黏膜上皮细胞脱落情况好转,排列较为整齐规则,胃黏膜上皮出现组织增生,胃腺细胞增殖,趋向正常,炎性细胞渐趋减少,毛细血管增生。
2.各神经被切断／损毁的艾灸预处理组胃黏膜损伤的保护效果差,胃黏膜损伤指数降低幅度较小(P<0.01,P<0.05),胃黏膜表面破损较严重,炎症细胞浸润较明显,细胞内可见充血。
3.艾灸预处理有上调胃黏膜组织中EGF、血清IL-2含量以及降低胃黏膜组织中SS含量的作用。但此作用部分受到了神经切断／损毁术的影响(P<0.01,P<0.05)。
4.经过艾灸预处理的各组血清中HSP-70含量有明显上升(P<0.01,P<0.05)；而各艾灸预处理+神经切断顺毁组与未经手术的艾灸预处理组血清中HSP-70含量没有变化(P>0.05)。
5.经过艾灸预处理的各组胃粘膜组织中HSP-70表达模型对照组均有不同程度的上升(P<0.01,P<0.05),而各艾灸预处理+神经切断/损毁组较未经手术的艾灸预处理组胃粘膜组织中HSP-70表达降低(P<0.01,P<0.05),模型对照组和正常对照组在此项指标上差异无统计学意义(P>0.05)。
结论：
1.艾灸预处理可明显改善胃黏膜损伤大鼠胃黏膜组织形态,降低胃黏膜损伤指数。各部位神经切断/损毁术对艾灸预处理降低胃黏膜损伤指数效应有明显干扰。
2.艾灸预处理上调胃黏膜组织中EGF、血清IL-2含量以及降低胃黏膜组织中SS含量来保护胃黏膜的作用部分受到神经调节的影响,证明神经通路是艾灸对胃黏膜损伤与保护因子表达的重要调节途径。
3.艾灸预处理有较好的保护胃粘膜损伤的作用,其作用机制与诱导HSP-70表达有关；艾灸对胃黏膜组织中HSP-70表达的促进作用信号传导主要是通过神经途径实现的。
4.脊髓、孤束核、腓总神经、迷走神经均是艾灸预处理对胃粘膜保护效应信号神经通路的一部分,均起到了信息传导作用。
Aim:In this study, perform selective neurotomy of several parts of rats'central nervous and peripheral nervous, and use the methods of tectology, biochemistry, and molecular biology, to observe the effect of pre-moxibustionat acupoint Zusanli(ST36) on acute gastric mucosal lesion. And endogenous EGF, SS, IL-2, and HSP-70were checked to explore the mechanism of pre-moxibustion at acupoint Zusanli(ST36) to defense gastric mucosal lesion, if it is ralated to neuroregulation or humoral coordination.
Methods:SD rats were randomly divided into7groups:A(Control group), B(Model group), C(Pre-moxibustion group), D(Group of pre-moxibustion+transection of spinal cord), E(Group of pre-moxibustion+damage nucleus of solitary tract) and F(Group of pre-moxibustion+vagotomy), G(Group of pre-moxibustion+neurotomy of common peroneal nerve). Neurotomy of several parts of rats'central nervous and peripheral nervous were performed beforehand, and then pre-moxibustionat acupoint Zusanli(ST36) were carried out. After that acute gastric mucosal lesion models were made. At last, the changes of rats'gastric mucosa were observed under light microscope; UI index were counted by GUTH method; EGF,SS in gastric mucos andIL-2in serum were checked by ELISA method; HSP-70were checked by western-blot method in gastric mucos, ELISA method in serum.
Results:
1.After modeled, the UI of gastric mucosa of the model group was the highest and the lowest was the normal group. The great significantly difference between these two groups illustrated that model was successful. And in group of pre-moxibustion the gastric mucosa of rats, which was damaged, recovered in various degree (P<0.01), that appeared by the decreased UI, upturn of slough off epidermal cell, and tend to be normal, and so on.
2. The effect of pre-moxibustion at acupoint Zusanli(ST36) to defense gastric mucosal lesion in groups of neurotomy ws not that good than that in group of pre-moxibustion(P<0.01, P<0.05). There are some damage on gastric mucosa surface, presence of inflammatory cells, and hyperemia in cells.
3. Pre-moxibustion at acupoint Zusanli(ST36) can up-raise the contents of EGF in gastric mucosa,IL-2in serum, and reduce that of SS in gastric mucosa. But this effect were interfered by neurotomy(P<0.01, P<0.05).
4. Considerable damage appeared in gastric mucosa of model group; UI indexs, but the expression of HSP-70in serum were of group C, D, E, F and G were considerabley increased vs groupB(P<0.01, P<0.05). And the expression of HSP-70in serum operation groups had no difference with group C(P>0.05).
5. The the expression of HSP-70in gastric mucosa of group C, D, E, F, G were higher than group B(P<0.01, P<0.05), but that of operation groups were lower than group C, and the expression of HSP-70in gastric mucosa in group B and group A had no difference(P>0.05).
6. There are no difference of the expression of HSP-70in different neurotomy of different parts of nerve tract(nervus peroneus communis, spinal cord, nucleus of solitary tract and vagus)(P>0.05).
Conclusion:
1.Pre-moxibustion at acupoint Zusanli(ST36) can improve the histomorphology and depress the UI of gastric mucosa of model rats.
2.Neurotomy of different parts has obvious interfere in the depress effect of Pre-moxibustion at acupoint Zusanli(ST36) on the UI of gastric mucosa.
3.Neurotomy of different parts has obvious interfere in the effect of Pre-moxibustion at acupoint Zusanli(ST36) on the up-raise the contents of EGF in gastric mucosa, IL-2in serum, and reduce that of SS in gastric mucosa, proving that neuroregulation is important to the mechanism of pre-moxibustion.
4.defense effect of pre-moxibustion at acupoint Zusanli(ST36)on gastric mucosal lesion is related to its inducement to expression of HSP-70. And neuroregulation participate the effective channel of signal transduction.
5.The nervus peroneus communis, spinal cord, nucleus of solitary tract and vagus are all parts of effective signal passage of pre-moxibustion at acupoint Zusanli(ST36) to gastric mucosal lesion.
方法：SD大鼠随机分为7组,即A正常对照组；B模型对照组；C艾灸预处理+模型组；D艾灸预处理+模型+脊髓损毁组；E艾灸预处理+模型+孤束核损毁组；F艾灸预处理+模型+迷走神经切断组；G艾灸预处理+模型+腓总神经切断组。对涉及手术的各组大鼠按要求分别进行神经损毁或切断手术,术后采用无水酒精灌胃法制备大鼠胃黏膜损伤模型,以艾灸预处理足三里穴为被试因素,按Guth标准记录大鼠胃黏膜损伤指数,光镜下观察胃黏膜组织形态学变化；采用酶联免疫吸附法检测胃黏膜组织中EGF、SS、血清中IL-2的含量；采用酶联免疫吸附法(ELISA)和蛋白质印迹法(Western-blot)检测血清中和胃黏膜组织中热休克蛋白70(HSP-70)表达。
结果：
1.造模后,模型对照组大鼠胃黏膜损伤指数最高,正常组大鼠胃黏膜未见明显损伤,说明造模成功；艾灸预处理后,胃黏膜损伤大鼠的胃黏膜均有不同程度的修复,主要表现为胃黏膜损伤指数降低(P<0.01),胃黏膜上皮细胞脱落情况好转,排列较为整齐规则,胃黏膜上皮出现组织增生,胃腺细胞增殖,趋向正常,炎性细胞渐趋减少,毛细血管增生。
2.各神经被切断／损毁的艾灸预处理组胃黏膜损伤的保护效果差,胃黏膜损伤指数降低幅度较小(P<0.01,P<0.05),胃黏膜表面破损较严重,炎症细胞浸润较明显,细胞内可见充血。
3.艾灸预处理有上调胃黏膜组织中EGF、血清IL-2含量以及降低胃黏膜组织中SS含量的作用。但此作用部分受到了神经切断／损毁术的影响(P<0.01,P<0.05)。
4.经过艾灸预处理的各组血清中HSP-70含量有明显上升(P<0.01,P<0.05)；而各艾灸预处理+神经切断顺毁组与未经手术的艾灸预处理组血清中HSP-70含量没有变化(P>0.05)。
5.经过艾灸预处理的各组胃粘膜组织中HSP-70表达模型对照组均有不同程度的上升(P<0.01,P<0.05),而各艾灸预处理+神经切断/损毁组较未经手术的艾灸预处理组胃粘膜组织中HSP-70表达降低(P<0.01,P<0.05),模型对照组和正常对照组在此项指标上差异无统计学意义(P>0.05)。
结论：
1.艾灸预处理可明显改善胃黏膜损伤大鼠胃黏膜组织形态,降低胃黏膜损伤指数。各部位神经切断/损毁术对艾灸预处理降低胃黏膜损伤指数效应有明显干扰。
2.艾灸预处理上调胃黏膜组织中EGF、血清IL-2含量以及降低胃黏膜组织中SS含量来保护胃黏膜的作用部分受到神经调节的影响,证明神经通路是艾灸对胃黏膜损伤与保护因子表达的重要调节途径。
3.艾灸预处理有较好的保护胃粘膜损伤的作用,其作用机制与诱导HSP-70表达有关；艾灸对胃黏膜组织中HSP-70表达的促进作用信号传导主要是通过神经途径实现的。
4.脊髓、孤束核、腓总神经、迷走神经均是艾灸预处理对胃粘膜保护效应信号神经通路的一部分,均起到了信息传导作用。
Aim:In this study, perform selective neurotomy of several parts of rats'central nervous and peripheral nervous, and use the methods of tectology, biochemistry, and molecular biology, to observe the effect of pre-moxibustionat acupoint Zusanli(ST36) on acute gastric mucosal lesion. And endogenous EGF, SS, IL-2, and HSP-70were checked to explore the mechanism of pre-moxibustion at acupoint Zusanli(ST36) to defense gastric mucosal lesion, if it is ralated to neuroregulation or humoral coordination.
Methods:SD rats were randomly divided into7groups:A(Control group), B(Model group), C(Pre-moxibustion group), D(Group of pre-moxibustion+transection of spinal cord), E(Group of pre-moxibustion+damage nucleus of solitary tract) and F(Group of pre-moxibustion+vagotomy), G(Group of pre-moxibustion+neurotomy of common peroneal nerve). Neurotomy of several parts of rats'central nervous and peripheral nervous were performed beforehand, and then pre-moxibustionat acupoint Zusanli(ST36) were carried out. After that acute gastric mucosal lesion models were made. At last, the changes of rats'gastric mucosa were observed under light microscope; UI index were counted by GUTH method; EGF,SS in gastric mucos andIL-2in serum were checked by ELISA method; HSP-70were checked by western-blot method in gastric mucos, ELISA method in serum.
Results:
1.After modeled, the UI of gastric mucosa of the model group was the highest and the lowest was the normal group. The great significantly difference between these two groups illustrated that model was successful. And in group of pre-moxibustion the gastric mucosa of rats, which was damaged, recovered in various degree (P<0.01), that appeared by the decreased UI, upturn of slough off epidermal cell, and tend to be normal, and so on.
2. The effect of pre-moxibustion at acupoint Zusanli(ST36) to defense gastric mucosal lesion in groups of neurotomy ws not that good than that in group of pre-moxibustion(P<0.01, P<0.05). There are some damage on gastric mucosa surface, presence of inflammatory cells, and hyperemia in cells.
3. Pre-moxibustion at acupoint Zusanli(ST36) can up-raise the contents of EGF in gastric mucosa,IL-2in serum, and reduce that of SS in gastric mucosa. But this effect were interfered by neurotomy(P<0.01, P<0.05).
4. Considerable damage appeared in gastric mucosa of model group; UI indexs, but the expression of HSP-70in serum were of group C, D, E, F and G were considerabley increased vs groupB(P<0.01, P<0.05). And the expression of HSP-70in serum operation groups had no difference with group C(P>0.05).
5. The the expression of HSP-70in gastric mucosa of group C, D, E, F, G were higher than group B(P<0.01, P<0.05), but that of operation groups were lower than group C, and the expression of HSP-70in gastric mucosa in group B and group A had no difference(P>0.05).
6. There are no difference of the expression of HSP-70in different neurotomy of different parts of nerve tract(nervus peroneus communis, spinal cord, nucleus of solitary tract and vagus)(P>0.05).
Conclusion:
1.Pre-moxibustion at acupoint Zusanli(ST36) can improve the histomorphology and depress the UI of gastric mucosa of model rats.
2.Neurotomy of different parts has obvious interfere in the depress effect of Pre-moxibustion at acupoint Zusanli(ST36) on the UI of gastric mucosa.
3.Neurotomy of different parts has obvious interfere in the effect of Pre-moxibustion at acupoint Zusanli(ST36) on the up-raise the contents of EGF in gastric mucosa, IL-2in serum, and reduce that of SS in gastric mucosa, proving that neuroregulation is important to the mechanism of pre-moxibustion.
4.defense effect of pre-moxibustion at acupoint Zusanli(ST36)on gastric mucosal lesion is related to its inducement to expression of HSP-70. And neuroregulation participate the effective channel of signal transduction.
5.The nervus peroneus communis, spinal cord, nucleus of solitary tract and vagus are all parts of effective signal passage of pre-moxibustion at acupoint Zusanli(ST36) to gastric mucosal lesion.
引文
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