“肺与大肠相表里”关系的生物学机制研究——大鼠肺、肠组织相关性的生理机制研究

【作者】 李立华；

【导师】 郭霞珍；

【作者基本信息】 北京中医药大学 ， 中医基础理论， 2012， 博士

【摘要】 肺与大肠表里相合关系的理论历经2000多年的临床实践检验,迄今为止仍然有重要的临床价值。在现代,人们运用现代科技方法来对这一理论的发生机制及内涵进行探讨,已经得到一些认识：认为肺与大肠之间的关系不仅仅是古代人们所发现的经络之间的相互络属关系,还得出了它们之间的胚胎同源性、神经物质相关性的认识。这些认识在一定程度上有效的指导我们把这一经典理论用之于临床,所以,我们将深入对肺与大肠的生物学发生机制进行探讨。1研究目的本课题围绕“肺合大肠”这一经典理论,概括总结“肺合大肠”的理论内涵,在该理论指导下,对胎鼠和成鼠的肺、肠组织之间的关系及相关物质表达进行探讨,通过分析肺、肠组织上的物质表达,以期从微观上得到肺与大肠相关的新的物质支持,进一步丰富“肺与大肠相表里”的内涵,为以后中医藏象理论的现代研究提供新的思路和方法。2研究内容2.1研究SD大鼠在胚胎时期肺、肠组织的组织发生学相关性。2.2研究SD大鼠在不同生理状态下肺、肠组织的生理机能相关性。3研究方法3.1大鼠肺、肠组织在组织发生学相关性的研究：分别选取囊状期、肺泡期、性成熟期这三个时期的胎鼠,每期胎鼠都以肺、气管、空肠、回肠、结肠、膀胱为研究对象,以α-actin. T淋巴细胞亚群(CD3+、CD4+、CD8+)、PAR-2为检测指标,使用免疫组织化学技术对这些研究对象上的物质进行观察,分析比较胚胎时期各个器官上的这三种物质的阳性细胞表达以及表达的差异,探讨胚胎期肺与大肠在组织发生学上的相关性。3.2大鼠肺、肠组织在生理机能相关性的研究：制备五组大鼠模型：分别为高氧组、低氧组、限食组、限水组、正常组,分别取这五组大鼠的肺、空肠、回肠、结肠组织为研究对象,用放射免疫法检测VIP、CCK、P物质这三个肺、肠相关的神经内分泌物质和酶联免疫吸附法(ELISA)检测N0、NOS,分别分析各模型组各个组织上这些物质的表达,探讨在不同的生理状态下,肺与大肠的相关关系的稳定性、肺与大肠的生理机能相关性。4研究结果4.1肺与大肠的组织发生学相关性研究结果4.1.1 a-actin的表达情况4.1.1.1在囊状期,肺、气管、空肠、回肠、结肠以及膀胱上均有a-actin的阳性细胞表达,且a-actin在肺与空肠、回肠、结肠上的表达在一个范围内,没有显著差别；而肺与气管、膀胱的表达具有显著差异,P<0.05。4.1.1.2在肺泡期,肺、气管、空肠、回肠、结肠以及膀胱上均有a-actin的阳性细胞表达,且a-actin在肺与回肠、结肠上的表达在一个范围内,没有显著差别；而肺与气管、空肠、膀胱的表达具有显著差异,P<0.05。4.1.1.3在性成熟期,肺、气管、空肠、回肠、结肠以及膀胱上均有a-actin的阳性细胞表达,且a-actin在肺与空肠、回肠、结肠上的表达在一个范围内,没有显著差别；而肺与气管、膀胱的表达具有显著差异,P<0.05。4.1.2 T淋巴细胞亚群的表达情况4.1.2.1在囊状期,肺、气管、空肠、回肠、结肠、膀胱上都有CD3+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD3+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD3+表达没有差异。4.1.2.2在肺泡期,肺、气管、空肠、回肠、结肠、膀胱上都有CD3+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD3+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD3+表达没有差异。4.1.2.3在性成熟期,肺、气管、空肠、回肠、结肠、膀胱上都有CD3+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD3+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD3+表达没有差异。4.1.2.4在囊状期,肺、气管、空肠、回肠、结肠、膀胱上都有CD4+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD4+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD4+表达没有差异。4.1.2.5在肺泡期,肺、气管、空肠、回肠、结肠、膀胱上都有CD4+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD4+表达具有统计学意义,P<0.0 5,肺与回肠、结肠的CD4+表达没有差异。4.1.2.6在性成熟期,肺、气管、空肠、回肠、结肠、膀胱上都有CD4+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD4+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD4+表达没有差异。4.1.2.7在囊状期,在囊状期,肺、气管、空肠、回肠、结肠、膀胱上都有CD8+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD8+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD8+表达没有差异。4.1.2.8在肺泡期,肺、气管、空肠、回肠、结肠、膀胱上都有CD8+阳性细胞表达,并且与肺相比,气管、膀胱的CD8+表达具有统计学意义,P<0.05,肺与回肠、结肠、空肠的CD8+表达没有差异。4.1.2.9在性成熟期,肺、气管、空肠、回肠、结肠、膀胱上都有CD8+阳性细胞表达,并且与肺相比,气管、空肠、膀胱的CD8+表达具有统计学意义,P<0.05,肺与回肠、结肠的CD8+表达没有差异。4.1.3 PAR-2表达情况4.1.3.1在胎鼠的囊状期、肺泡期、性成熟期,肺、气管、空肠、回肠、结肠、膀胱上均有par-2的阳性细胞表达。4.1.3.2在囊状期,尽管检测的6个组织上都有par-2的阳性表达,但与肺相比,气管、空肠、膀胱的表达明显低于肺组织,P<0.05,具有统计学意义；回肠、结肠与肺的表达没有明显差异。4.1.3.3在肺泡期,肺、气管、空肠、回肠、结肠、膀胱上都有par-2阳性细胞表达,与肺相比,气管、空肠、膀胱明显低于肺组织,P<0.05,具有统计学意义；回肠、结肠没有显著差异。4.1.3.4在性成熟期,肺、气管、空肠、回肠、结肠、膀胱上都有par-2阳性细胞表达,与肺相比,气管、空肠、膀胱明显低于肺组织,P<0.05,具有统计学意义；回肠、结肠没有显著差异。4.2肺与大肠的生理机能相关性研究结果4.2.1低氧组：大鼠在造模过程中发现,大鼠体重逐渐减轻,行动不稳,嗜睡,反应迟钝。造模后第4天,粪便减少,粪质较硬。取材时发现低氧模型大鼠肺部有损伤,出现瘀血,肺组织颜色与正常组大鼠相比明显变暗。结肠部有少量硬质粪便聚集。指标检测结果发现：与肺相比,空肠上的VIP、CCK表达有显著差异,P<0.05,而回肠、结肠没有明显差别；与肺相比,空肠、回肠上的SP表达有明显差异,P<0.05,而结肠上的SP表达没有明显区别；与肺相比,空肠组织上的NO、NOS有显著差异,P<0.05,回肠、结肠没有明显差别。4.2.2高氧组：大鼠在造模及取材过程中发现,大鼠状态良好,没有显著变化。指标检测发现：与肺相比,空肠上的VIP、SP表达具有明显差异,P<0.05,回肠、结肠没有明显差别；肺、空肠、回肠、结肠上的CCK表达都没有明显差别；与肺相比,空肠上的N0、NOS表达具有明显差异,P<0.05,回肠、结肠没有明显差别。4.2.3限食组：大鼠身体逐渐消瘦,体重减轻,在取材前两日发现：毛发直立,弓背,易激惹。取材时发现肺组织没有明显变化,而结肠中粪便较硬。指标检测发现：与肺相比,空肠上的VIP、SP表达具有统计学意义,P<0.05,而回肠、结肠没有显著区别；与肺相比,回肠上的CCK表达具有统计学意义,与空肠、结肠没有明显区别。与肺相比,空肠上的NO、NOS表达具有明显差异,P<0.05,回肠、结肠没有明显差别。4.2.4限水组：大鼠体重没有明显变化,状态与正常组相比没有什么区别。取材时发现肺组织没有明显变化,结肠中粪便较多,便质坚硬。与肺相比,空肠上的VIP表达具有统计学意义,P<0.05,而回肠、结肠没有显著区别；肺、空肠、回肠、结肠上的CCK表达没有明显区别；与肺相比,空肠、回肠上的SP相比有明显差别,P<0.05,而与结肠没有。与肺相比,空肠上的NO、NOS表达具有明显差异,P<0.05,回肠、结肠没有明显差别。4.2.5正常组：整天状态良好。与肺相比,空肠上的VIP、CCK、SP有明显差别,P<0.05,回肠、结肠上的表达没有明显差别。与肺相比,空肠、结肠上的NO有明显差异,P<0.05,回肠没有明显差别；肺、空肠、回肠、结肠上的NOS表达都没有明显差异。5结论5.1在胚胎的发育过程中,肺与大肠(回肠、结肠)具有同源相关性,即组织发生学相关性。肺与大肠在胚胎发育过程中就具有粘膜免疫相关性的物质基础。5.2肺与大肠(回肠、结肠)具有生理机能相关性。更多还原

【Abstract】 The theory"the lung pairs up with the large intestine" after 2000 years is still valuable.Now, people use modern technology to explore its pathogenesis and connotation .They know:The relationship between the lung and large intestine is not only mutual contact relationship between the meridian of the ancient people found, also believe that embryonic homology and the nerve substance between them. We will continue along the footsteps of these scientific biology of mechanism of the lung and the large intestine to explore.1 PurposeThe subject will around the theory" The lung pairs up with the large intestine", and explore the expression of the substance between the intestine and lung in fetal rat and mouse lung. Through analysising of the substances on the lung and intestine, we can support "The lung pairs up with the large intestine", and rich theory. It will provide new ideas and methods to study the theory of viscera.2 Content2.1 To study the relation between lung and intestine, of the occurrence of organization in the embryonic period.2.2 To study the function correlation in lung and intestine. 3 Method3.1 To study of embryonic homology in lung and intestine:We choose rat in periods of cysts, alveoli, sexual maturity. And then took Lung, esophagus, jejunum, ileum, colon and bladder, to observingα-actin, T-lymphocyte subsets (CD3+, CD4+, CD8+), PAR-2 in those organs.3.2 To study the nerve-endocrine substances in lung and intestine:We made model of physiological group, hypoxia group, hyperoxia group, diet restriction group, water restriction group, then took lung jejunum, ileum and colon.Observed the VIP, CCK, SP, NO and NOS. 4 Result4.1 Findings of lung and colon homology4.1.1 Expression of a-actin4.1.1.1 In cysts period, lung and trachea, bladder expression was significantly different (P <0.05). There are no significant differences in lung, jejunum, ileum and colon.4.1.1.2 In the alveoli period, lung, trachea, jejunum and bladder expression was significantly different (P<0.05).There are no significant differences in lung, ileum and colon.4.1.1.3 In sexual maturity period, lung and trachea, bladder expression was significantly different (P<0.05). There are no significant differences in lung, jejunum, ileum and colon. 4.1.2 The expression of T cells (CD3+CD4+CD8+) In cysts, the alveoli, the sexual maturity period, compare with lung, trachea, jejunum and bladder expression was significantly different (P<0.05).There are no significant differences in lung, ileum and colon.4.1.3 The expression of PAR-24.1.3.1 In cysts, the alveoli, the sexual maturity period, there are par-2 positive cells in the lung, trachea, jejunum, ileum, colon, bladder.4.1.3.2 In the cysts period, compared with lung, trachea, jejunum and bladder expression was significantly different (P<0.05), which are lower than lung. There are no significant differences in lung, ileum and colon.4.1.3.3 In the alveoli period, compared with lung, trachea, jejunum and bladder expression was significantly different (P<0.05), which are lower than lung. There are no significant differences in lung, ileum and colon.4.1.3.4 In the sexual maturity period, compared with lung, trachea, jejunum and bladder expression was significantly different (P<0.05), which are lower than lung. There are no significant differences in lung, ileum and colon.4.2 Substance-related in lung and intestine4.2.1 In hypoxia group, we found:Compared with lung, VIP,CCK,NO,NOS are significantly different in jejunum(P<0.05). There are no significant differences in lung, ileum and colon. Compared with lung, SP was significantly different in jejunum and ileum (P<0.05). There are no significant differences in lung and colon.4.2.2 In hyperoxia group, we found:Compared with lung, VIP,SP,NO,NOS are significantly different in jejunum(P<0.05). There are no significant differences in lung, ileum and colon. There are no significant differences in lung,jejunum, ileum and colon in CCK.4.2.3 In diet restriction group, we found:Compared with lung, VIP,SP,NO,NOS are significantly different in jejunum(P<0.05). There are no significant differences in lung, ileum and colon. Compared with lung, CCK was significantly different in ileum (P<0.05). There are no significant differences in lung, jejunum and colon.4.2.4 In water restriction group, we found:Compared with lung. VIP.NO. NOS are significantly different in jejunum(P<0.05). There are no significant differences in lung, ileum and colon. Compared with lung, SP was significantly different in jejunum and ileum (P<0.05). There is no significant differences in lung and colon.4.2.5 In physiological group, we found:Compared with lung, VIP,CCK,SP are significantly different in jejunum(P<0.05). There are no significant differences in lung, ileum and colon. Compared with lung. NO was significantly different in jejunum and colon (P<0.05). There is no significant differences in lung and ileum. There are no significant differences in lung, jejunum,ileum and ileum.5 Conclusion5.1 In the process of embryonic development, lung and the large intestine(Ileum、colon)have the same sources, that is lung and the large intestine have the same histogenesis. During the embryonic development, Lung and large intestine have the basis material of the mucosal immune.5.2 Lung and colon are closely related in terms of physiological function.更多还原