【作者】 王江；

【导师】 周永学；

【作者基本信息】 成都中医药大学 ， 方剂学， 2013， 博士

【摘要】 目的：观察半夏泻心汤及其拆方对胃溃疡大鼠模型炎性因子、修复因子及Hsp27、Hsp70蛋白表达的影响,分析半夏泻心汤及拆方治疗胃溃疡的作用机制,探讨该方不同药群在治疗胃溃疡中的配伍意义,揭示寒热并用配伍方法的优势和作用靶点。方法：SD大鼠,雌雄各半,造模各组采用大承气汤结合辣椒/乙醇混合液灌胃及乙酸注射法建立胃溃疡寒热错杂证病证结合动物模型。胃溃疡手术24h后给予半夏泻心汤及拆方各组药物灌胃,28d后处死大鼠,光镜观察胃组织病理学变化,黄嘌呤氧化酶法测定胃组织SOD活性,硫代巴比妥酸法测定胃组织MDA含量,硝酸还原酶法测定血清NO含量,ELISA法检测胃组织TNF-α、IL-8、GAS、MTL及血清PAF、ET含量；免疫组化检测胃粘膜EGF、VEGF、bFGF、PDGF表达；RT-PCR检测胃组织HSP27、HSP70mRNA的表达。结果：①一般状态和行为表现：全方组、寒热并用组大鼠的精神状态、毛色、饮食、体重等情况较模型组有明显改善,且全方组、寒热并用组的一般状态要优于其它各组。②病理形态学观察：空白组、假手术组胃粘膜结构完整,无充血、水肿,上皮细胞呈单层柱状,排列整齐,基本未见缺损、脱落,固有层内未见炎性细胞浸润,腺体形状规则;模型组：可见胃粘膜变薄,病灶局部上皮细胞脱落,出现缺损,粘膜固有层有大量炎性细胞浸润、部分有淋巴滤泡形成,粘膜固有腺体数量明显减少、排列稀疏紊乱：给药治疗后,全方组及寒热并用组上述病理改变明显减轻,胃粘膜结构基本完整,上皮结构修复,粘膜固有腺体增生数量较多,排列整齐,呈囊状扩张,浆膜层完整,仅见少量炎性细胞浸润；其余各组仍可见炎性细胞浸润等炎症表现。③胃组织TNF-α、IL-8含量：与空白组比较,模型组大鼠胃组织TNF-α、IL-8含量显著升高(P<0.01)：与模型组比较,全方组、寒热并用组、寒凉组及阳性组TNF-α、IL-8含量显著降低(P<0.01)；与全方组比较,寒热并用组和寒凉组TNF-α、IL-8含量无差异(P>0.05);④胃组织SOD、 MDA含量：与空白组比较,模型组大鼠胃组织SOD含量显著降低(P<0.01),胃组织MDA含量显著升高(P<0.01)；与模型组比较,全方组、寒热并用组、温热组、补益组、温补组及寒补组大鼠胃组织SOD含量显著升高(P<0.01),MDA含量显著降低(P<0.01)：与全方组比较,寒凉组SOD含量有显著性差异(P<0.01),补益组和寒补组有差异(P<0.05),寒凉组、寒补组和阳性组MDA含量有显著性差异(P<0.01),补益组有差异(P<0.05)；⑤胃组织GAS、MTL含量：与空白组比较,模型组大鼠胃组织GAS、MTL含量显著升高(P<0.01)；与模型组比较,全方组、寒热并用组、寒补组及阳性组胃组织GAS、MTL含量显著降低(P<0.01)；与全方组比较,寒热并用组和阳性组GAS含量无显著性差异(P>0.05),寒热并用组、寒凉组和寒补组MTL含量无显著性差异(P>0.05)；⑥血清PAF、ET、NO含量：与空白组比较,模型组大鼠血清PAF、ET含量显著升高(P<0.01),血清NO含量显著降低(P<0.01)；与模型组比较,全方组、寒热并用组、温热组及温补组PAF、ET含量显著降低(P<0.01),寒凉组和寒补组PAF含量降低(P<0.05),补益组和寒补组ET含量降低(P<0.05),全方组、寒热并用组、温热组、补益组、温补组、寒补组及阳性组NO含量显著升高(P<0.01);与全方组比较，寒热并用组PAF含量无差异(P>0.05),寒热并用组及温热组ET含量无差异(P>0.05),寒热并用组、温热组和温补组NO含量无差异(P>0.05)。⑦胃粘膜EGF、VEGF、bFGF、PDGF表达：与空白组比较,模型组大鼠胃粘膜EGF、VEGF、bFGF、PDGF平均光密度值显著升高(P<0.01)；模型组比较,全方组、寒热并用组、补益组、温补组、寒补组及阳性组EGF、 EGF平均光密度值显著升高(P<0.01),温热组升高(P<0.05),而寒凉组无显著性差异(P>0.05)，全方组、寒热并用组、补益组、温补组、寒补组及阳性组bFGF、PDGF平均光密度值显著升高(P<0.01),而寒凉组、温热组无显著性差异(P>0.05)：与全方组比较,寒热并用组、补益组、温补组及寒补组EGF、VEGF、 bFGF、PDGF平均光密度值无差异(P>0.05),而寒凉组和温热组有显著性差异(P<0.01)。⑧胃组织HSP27、HSP70mRNA的表达：与空白组比较,模型组大鼠胃组织HSP27、HSP70mRNA表达显著增强(P<0.01)；与模型组比较,全方组、寒热并用组、温补组胃组织HSP27、HSP70mRNA表达显著增强(P<0.01),温热组、补益组、寒补组和阳性组大鼠胃组织HSP27、HSP70mRNA表达增强(P<0.05),血寒凉组则无显著性差异(P>0.05)：与全方组比较,寒热并用组无差异(P>0.05)。结论：①半夏泻心汤及其拆方可显著改善胃溃疡大鼠的一般行为表现及胃溃疡的炎症程度。②半夏泻心汤及其拆方对胃溃疡大鼠有明显的治疗作用,其作用机制包括抑制炎性因子的释放；提高机体的抗氧化能力并减轻脂质过氧化损伤；减轻胃酸对胃粘膜的侵蚀；协调胃肠运动;改善微循环；提高胃粘膜的修复能力；上调内源性保护蛋白HSP27、HSP70mRNA的表达。③半夏泻心汤方中不同属性的药物从不同方面对胃溃疡发挥治疗作用。寒凉组药物在降低胃组织中TNF-α、IL-8含量等方面作用显著。温热组药物在升高血清中NO含量,降低血清中PAF、ET含量等方面作用显著。补益组药物在升高胃粘膜EGF、VEGF、 bFGF、PDGF阳性表达方面作用显著。温补组药物升高胃组织中SOD含量,并降低MDA含量,上调内源性保护蛋白HSP27、HSP70mRNA的表达方面作用显著。寒补组药物降低胃组织中GAS、MTL含量等方面作用显著。④寒热并用配伍能消除单用寒凉或温热药的局限和不足,通过不同的途径和作用靶点来发挥对胃溃疡的治疗作用,在很多方面产生了协同增效作用。更多还原

【Abstract】 Objective:Observe the influence on banxiaxiexin decoction and diffenent herbal formulation of component about gastric ulcer rat model in respects of inflammatory factors, repair factor and Hsp27, Hsp70protein expression, and analysis the mechanism of action about the treatment effects of banxiaxiexin decoction and its Subdivisions, investigate the Significance of diffenent herbal formulation of component in banxiaxiexin decoction on treat gastric ulcer (GU), reveal the superiority and targets of compatibility of both cold and heat.Methods:Select male and female SD rats and establish the gastric ulcer animal model of mixed cold and heat syndromes. After24hours, all the rats except blank group and sham operation group were given of banxiaxiexin decoction and different herbal formulation of component, all rats were sacrificed after28days, observe rats" pathological morphology change of gastric tissue, test SOD activity in xanthine oxidase method, gastric tissue MDA content in thiobarbituric acid method, serum NO Content in nitrate reductase method, gastric tissue TNF-a, IL-8, GAS, MTL and serum PAF, ET content in ELISA method, gastric mucosal EGF, VEGF, bFGF, PDGF expression in immunohistochemical detection, HSP27, HSP70mRNA expressions of gastric tissue of rats in RT-PCR method.Result:①General behavior:The rats’spirit, diet, fur color and weight were signify-cantly improved by banxiaxiexin decoction group and compatibility of both cold and heat group;②Pathoio-gical observation:The blank group and sham operation group rats’ structure of gastric mucosa surface was complete, and there were no congestion, edema, the simple columnar epithelial cells were arranged in neat rows, basically no defect, shedding no inflammatory cell infiltration in the lamina propria, glands regular shape;The model group rats’structure of gastric mucosa surface was thinning, Focal epithelial cell shedding, the defect in the lamina propria massive infiltration of inflammatory cells, part of the formation of lymphoid follicles, lamina propria glands significantly reduce sparse arrangement disorder; After treatment, Significantly reduced banxiaxiexin decoction group and compatibility of both cold and heat group,these pathological changes, the basic integrity of the structure of gastric mucosa epithelial structure repair, a large number of lamina propria gland hyperplasia, arranged in neat rows, cystic expansion, serosal integrity, only to see a small amount of inflammatory cellsinfiltration; the rest of the group is still visible inflammatory cell infiltration and inflammation;③TNF-α、IL-8content of gastric tissue:Compared with blank group, the content ofTNF-α、IL-8were significantly elevated of model group rats (P<0.01); Compared with model group, the content of TNF-α、IL-8were significantly decreased of banxiaxiexin decoction group, compatibility of both cold and heat group, cold group and positive group (P<0.01); Compared with banxiaxiexin decoction group, the content ofTNF-α、IL-8were no significantly differenee(P>0.05);④SOD、MDA content of gastric tissue:Compared with blank group, the content ofSOD was significantly decreased of model group rats(P<0.01), the content of MDA were significantly elevated of model group rats (P<0.01); Compared with model group, the content of SOD was significantly elevated of banxiaxiexin decoction group, compatibility of both cold and heat group, heat group, tonic group, heat tonic group and cold tonic group (P<0.01),while the content of MDA was significantly decreased (P<0.01); Compared with banxiaxiexin decoction group, the content of SOD was significant difference of cold group (P<0.01),while it was difference of tonic group and cold tonic group (P<0.05), the content of MDA was significant difference of cold group, cold tonic group and positive group (P<0.01), while it was difference of tonic group(P<0.05);⑤GAS,MTL content of gastric tissue:Compared with blank group, the content of GAS,MTL were significantly elevated of model group rats (P<0.01); Compared with model group, the content of GAS,MTL were significantly decreased of banxiaxiexin decoction group, compatibility of both cold and heat group, cold tonic group and positive group (P<0.01); Compared with banxiaxiexin decoction group, the content of GAS was significantly decreased of compatibility of both cold and heat group, positive group (P>0.05), the content of MTL was significantly decreased of compatibility of both cold and heat group, cold group and cold tonic group (P>0.05).⑥PAF、ET、NO content of serum:Compared with blank group, the content of PAF、ET of serum were significantly elevated of model group rats while the content of NO of serum decreased obviously (P<0.01); Compared with model group, the content of PAF、ET of serum were significantly decreased of banxiaxiexin decoction group, compatibility of both cold and heat group, heat group, heat tonic group (P<0.01) while it was decreased of cold group and cold tonic group (P<0.05), the content of ET of serum were significantly decreased of tonic group and cold tonic group (P<0.05), the content of ET of serum were significantly elevated of banxiaxiexin decoction group, compatibility of both cold and heat group, heat group, tonic group,heat tonic group, cold tonic group and positive group (P<0.01); Compared with banxiaxiexin decoction group, the content of PAF of compatibility of both cold and heat group was no significant difference (P>0.05), the content of ET of compatibility of both cold and heat, heat group were no significant difference (P>0.05), the content of NO of compatibility of both cold and heat, heat group and heat tonic group were no significant difference (P>0.05);⑦Gastric mucosal EGF, VEGF, bFGF, PDGF expression:Compared with blank group, the average optical density of EGF, VEGF, bFGF, PDGF expression were significantly enhanced of model group rats(P<0.01); Compared with model group,the average optical density of EGF, VEGF expression were significantly enhanced of banxiaxiexin decoction group, compatibility of both cold and heat group, tonic group, heat tonic group, cold tonic group and positive group (P<0.01), while it was enhanced of heat group (P<0.05), cold group was no significant difference (P>0.05), the average optical density of bFGF, PDGF expression were significantly enhanced of banxiaxiexin decoction group, compatibility of both cold and heat group, tonic group, heat tonic group, cold tonic group and positive group (P<0.01), cold group and heat group was no significant difference (P>0.05); Compared with banxiaxiexin decoction group, compatibility of both cold and heat group, tonic group, heat tonic group, cold tonic group were no significant difference (P>0.05), cold group and heat group were significant difference (P<0.01).⑧HSP27, HSP70mRNA expression of gastric tissue: Compared with blank group, the HSP27, HSP70mRNA expression of gastric tissue were significantly enhanced of model group rats (P<0.01); Compared with model group, the HSP27, HSP70mRNA expression of gastric tissue were significantly enhanced of banxiaxiexin decoction group, compatibility of both cold and heat group, Heat tonic group (P<0.01), while it is enhanced of Heat group, tonic group, Cold tonic group and positive group (P<0.05), cold group was no significant difference (P>0.05); Compared with banxiaxiexin decoction group, compatibility of both cold and heat group was no difference (P>0.05).Conclusion:①Banxiaxiexin decoction and diffenent herbal formulation of component can improve the gastric ulcer rat’s General behavior and reduce the inflammation degree of rats" gastric ulcer significantly.②Banxia(?)exin decoction and diffenent herbal formulation of component have obvious therapeutic effect on gastric ulcer. The mechanisms of action include inhibition of inflammatory factors release and improve the body’s antioxidant capacity and mitigate the damage of lipid peroxidation, alleviate gastric acid erosion of the gastric mucosa, coordination of gastrointestinal motility, improve microcirculation, improve gastric mucosal’s ability to repair, upregulation of endogenous protective protein HSP27, HSP70mRNA expression.③The different properties drug of banxiaxiexin decoction plays a therapeutic role from different aspects on gastric ulcer. Cold group of drugs play an important part on reducing the TNF-a, IL-8, GAS and MTL content of gastric tissue; Heat group of drugs play an important part on increasing the NO content of serum,reducing the PAF, ET content of serum, Tonic group of drugs play an important part on increasing the EGF, VEGF, bFGF and PDGF positive expression in gastric mucosa; Heat tonic group of drugs play an important part on increasing the SOD content of gastric tissue and reducing the MDA content of gastric tissue, upregulation of endogenous protective protein HSP27, HSP70mRNA expression; Cold tonic group of drugs play an important part on reducing the GAS, MTL content of gastric tissue;④After compatibility of both cold and heat drugs, the synergistic effect significant was reinforced. Compatibility of cold and heat drugs can reduce the limitations and deficiencies of cold or heat drugs and embody the Chinese medicine targeted therapeutic advantage.更多还原