基于脑肠肽探讨功能性消化不良脾虚证的生物学机制

发布时间：2018-06-06 00:39

  本文选题：动物模型 + 功能性消化不良　； 参考：《北京中医药大学》2016年博士论文

【摘要】：目的探讨功能性消化不良中医核心证型—脾虚证的生物学机制,进一步阐释并完善中医脾虚证相关理论。方法理论方面,首先对功能性消化不良临床文献进行整理,探讨功能性消化不良中医证型的分布规律,并对出现频次较高的中医证型的症状表现进行分析。挖掘疾病的核心症状。在此基础上,对疾病主症—“胀”,“满”相关文献进行中医古籍整理,梳理“腹胀”的源流及中医病机。同时,对“腹胀”西医文献进行整理,探讨“腹胀”产生的信号通路,寻找其可能的生物学机制。研究还梳理了以“腹胀”,“腹满”为主症的中医疾病源流,寻找最符合腹胀型功能性消化不良的中医病名,并研究此疾病的中医学病机。研究另外对“痞满”相关的方剂及中草药文献进行整理,探讨治疗“痞满”的中医用药规律。进而指导功能性消化不良脾虚证的中医用药。以理论研究为基础,实验方面,首先建立功能性消化不良脾虚证动物模型。运用碘乙酰胺复合小平台劳倦因素造模法,制作功能性消化不良脾虚证动物模型,并观察动物一般状态,体重,饮食情况及胃病理改变,运用糖水偏好试验,球囊试验对功能性消化不良脾虚证进行评价。在模型建立的基础上,运用免疫组化,Elisa, Real-time PCR对大鼠胃,下丘脑及血清中的脑肠肽(Ghrelin, CCK, VIP, SS, SP, CGRP)进行检测。此外,运用中药香砂六君子加减方对模型进行干预,西药多潘立酮作为阳性药物对照。研究期间,记录大鼠饮食情况,并运用糖水偏好试验,球囊试验对药物干预疗效进行评价。同时也运用免疫组化,Elisa, Real-time PCR对各组大鼠胃,下丘脑及血清中的脑肠肽(Ghrelin, CCK, VIP, SS, SP, CGRP)进行检测。结果理论方面,通过对功能性消化不良临床文献进行整理发现,功能性消化不良中医证型出现频次较高的依次为：肝胃不和,脾胃虚弱,寒热错杂,饮食积滞,肝郁脾虚,肝郁气滞,脾胃虚寒,脾胃湿热证,且各中医证型的共有症状为“上腹胀”。以此为切入点,对“胀”,“满”相关文献进行中医古籍整理,研究发现,中医“腹胀”主要由气虚不运,寒,热,湿,痰,食,血积聚腹内而成,病位涉及脾,胃,肝,肾等脏。其形成的生物学机制可与脑肠肽类激素的改变有关。从疾病角度,中医“痞满”的腹胀部位与起病时间与腹胀型功能性消化不良最相似,可以作为此病的中医病名。而“痞满”的病机以脾气虚为本,食欲的减少与否是鉴别“痞满”虚实的标准。而“痞满”治疗方剂中,以理气,益气健脾,燥湿化痰类中药出现频次较高。出现频次最高的药物,与中药成方香砂六君子汤的组成具有高度一致性,因此香砂六君子汤可以作为本病的主方。实验方面,建立的碘乙酰胺及复合小平台模型大鼠,均出现饮食减少,糖水偏好降低,胃敏感性增高。但碘乙酰胺复合小平台组大鼠体重下降明显,并伴有倦怠等精神症状。因此,两模型符合功能性消化不良脾虚证标准,但碘乙酰胺复合小平台组脾虚证表征更加明显。生物学指标方面,Ghrelin, CCK, VIP在两种功能性消化不良脾虚证大鼠胃中分泌减少,循环血及下丘脑中含量亦减少,但下丘脑中mRNA表达水平相反,其原因,可能与循环血中进入下丘脑肽类激素含量减少,引起脑内负反馈调节,从而使模型组中的mRNA表达含量升高有关。另外,SS在两种功能性消化不良脾虚证大鼠胃及下丘脑中分泌增加,循环血中含量增加,并且以单纯施加碘乙酰胺组大鼠增加程度大。研究也发现,复合小平台组大鼠胃内SP及CGRP蛋白升高较为明显,血清中CGRP明显降低,其余无显著性差异。因此,碘乙酰胺及复合小平台大鼠共同表现为具有促进食欲及降低胃壁紧张度作用的脑肠肽Ghrelin, CCK, VIP分泌及循环减少,体内抑制性脑肠肽SS含量增加,这可能是功能性消化不良脾虚证的病机。两脾虚证模型在胃内SP与CGRP蛋白表达的差异,以及SS的表达含量的差异可能是两种脾虚亚型的机制所在。不能食而瘦的脾虚证,以痛阈降低,敏感性相对偏高更显著,不能食而肥的脾虚证,则主要表现为胃肠激素紊乱。对模型施加药物治疗。研究发现,健脾方剂香砂六君子加减方,可增加模型大鼠饮食量。同时,健脾中药还可显著提高组内大鼠糖水消耗率75%所占百分比。且呈现量效关系。胃肠敏感性方面,健脾中药香砂六君子汤可显著降低模型组大鼠胃高敏感性,呈量效相关性。研究显示,健脾方剂香砂六君子加减方有效改善功能性消化不良脾虚证食少,纳呆,腹胀三大主症。另外,对大鼠毛色不泽,神情倦怠脾虚次症也有改善作用。因此,健脾方剂香砂六君子加减方对功能性消化不良脾虚证有治疗作用,同时也验证了本次研究复制的碘乙酰胺模型以及碘乙酰胺复合小平台模型与功能性消化不良脾虚证动物模型高度一致。在生物学指标方面,健脾方剂香砂六君子加减方可增加Ghrelin, CCK, VIP的分泌,进而升高循环血,及下丘脑中的含量。同时可以降低胃黏膜,血清及下丘脑中的SS蛋白含量。对于SP与CGRP,药物治疗组与碘乙酰胺组之间比较,无显著性差异。而对于以食少而瘦的脾虚证模型一复合小平台模型组,香砂六君子加减方组胃黏膜中SP与CGRP蛋白较模型组降低,其中以SP变化较为明显,但香砂六君子加减方对血清及下丘脑中SP与CGRP改变不明显。研究发现,健脾方剂香砂六君子加减方对脾虚型功能性消化不良具有治疗作用,其作用靶点集中的胃黏膜上,通过调节脑肠肽的生成,进而改善脾虚型功能性消化不良的症状。研究也证明了健脾中药治疗功能性消化不良脾虚证的作用机制与调节脑肠肽的生成有关。结论综合以上研究发现,中医学认为的功能性消化不良以“上腹胀”作为主要症状,而以“上腹胀”作为主要症状的中医病名为“痞满”。“痞满”的病机以脾虚为本,饮食的减少,食欲的减退与否是判定邪实盛衰的标准。根据这一特点,我们建立了功能性消化不良脾虚证动物模型,进而探讨了消化不良型脾虚证的生物学机制。研究发现,碘乙酰胺及复合小平台站立法符合功能性消化不良脾虚证标准,而脑肠肽类激素含量的变化与消化不良型脾虚证密切相关。运用健脾中药对模型反证,发现健脾中药可调整脑肠肽类激素的生成,进而改善脾虚证症状。因此,本研究证明了功能性消化不良脾虚证可能的生物学机制是脑肠肽的改变,表现为具有胃肠促进及抑制作用的脑肠肽失衡。研究成果一定程度上拓宽了传统中医“脾虚”的理论。
[Abstract]:Objective to explore the biological mechanism of the core syndrome of functional dyspepsia, the core syndrome of TCM - spleen deficiency syndrome, and to further explain and improve the theory of the syndrome of spleen deficiency in Chinese medicine. On the basis of the analysis of the symptoms of the disease, the core symptoms of the disease were excavated. On this basis, the literature of "bloating", "bloating" and the pathogenesis of traditional Chinese medicine were sorted out in the literature of "bloating" and "full", and the literature of "abdominal distention" was sorted out to explore the possible signal pathways of "abdominal distention". The study also combed the origin of TCM disease with abdominal distention and abdominal fullness as the main disease, looking for the name of traditional Chinese medicine which most conforms to abdominal distention and functional dyspepsia and studying the pathogenesis of traditional Chinese medicine. On the basis of theoretical research and experiment, the animal model of functional dyspepsia spleen deficiency syndrome was established, and the animal model of functional dyspepsia spleen deficiency syndrome was made by using the compound iiacetate compound small platform fatigue factor, and the general state of the animal was observed and the body was observed. Weight, diet and gastric pathological changes, using sugar water preference test and balloon test to evaluate the spleen deficiency of functional dyspepsia. On the basis of the establishment of the model, the brain gut peptide (Ghrelin, CCK, VIP, SS, SP, CGRP) in rats' stomach, hypothalamus and serum were detected by immunohistochemistry, Elisa and Real-time PCR. At the time of the study, the diet of rats was recorded, and the diet of rats was recorded during the study, and the sugar water preference test was used to evaluate the effect of the drug intervention on the effect of the drug intervention. At the same time, the Elisa and Real-time PCR were used in the stomach, the hypothalamus and the serum of the rats in each group. Ghrelin, CCK, VIP, SS, SP, CGRP). Results theoretically, by sorting out the clinical literature of functional dyspepsia, it was found that the frequency of TCM syndrome of functional dyspepsia was higher in sequence: liver and stomach disharmony, spleen and stomach weakness, cold and heat, stagnation of liver depression, stagnation of liver depression, stagnation of liver depression, deficiency of spleen and stomach, damp heat syndrome of spleen and stomach, The common symptoms of TCM syndrome type are "upper abdominal distention". As a breakthrough point, the literature of "bloating" and "full" related literature of traditional Chinese medicine is arranged. It is found that "abdominal distention" mainly consists of qi deficiency, cold, heat, dampness, phlegm, food and the accumulation of blood in the abdomen. The biological mechanism of the disease is related to the spleen, stomach, liver and kidney. The biological mechanism of its formation can be related to the brain intestine. From the point of view of disease, the abdominal distention parts of the Chinese medicine "ruffian" and the onset time and abdominal distension type functional dyspepsia are the most similar, which can be used as the name of the disease of traditional Chinese medicine. In addition, the frequency of traditional Chinese medicine, which has the highest frequency, is highly consistent with the composition of the decoction of six Jun Zi Decoction of Chinese herbal medicine. Therefore, the fragrant sand six gentleman soup can be used as the main recipe of this disease. The preference of sugar and water decreased and the sensitivity of stomach increased. However, the weight decline of rats in the IIb compound small platform group was obvious and accompanied by mental symptoms such as burnout. Therefore, the two model was in line with the standard of functional dyspepsia, but the characterization of spleen deficiency syndrome in the IIB complex small platform group was more obvious. The biological indexes, Ghrelin, CCK and VIP were two kinds of work. In the rats with dyspeptic dyspepsia, the secretion of the stomach in the stomach was reduced, and the content of circulating blood and hypothalamus decreased, but the expression level of mRNA in the hypothalamus was opposite, which may be related to the decrease in the content of peptide hormone in the hypothalamus and the adjustment of the negative feedback in the brain, which is related to the increase of mRNA expression in the model group. In addition, SS is two The secretion of gastric and hypothalamus increased in the rats with functional dyspepsia and increased in the gastric and hypothalamus, and increased in circulating blood. The increase of SP and CGRP protein in the stomach of the compound small platform group was more obvious, and the CGRP in the serum was obviously decreased, and the rest had no significant difference. Therefore, iodine B was not significant. Ghrelin, CCK, VIP secretion and circulation decreased, and the content of inhibitory brain gut peptide SS in the body increased, which may be the pathogenesis of functional dyspepsia spleen deficiency syndrome. The difference between the expression of SP and CGRP protein in the two spleen deficiency syndrome model in the stomach, as well as the difference in the expression of SP and CGRP protein in the stomach, and The difference in the expression content of SS may be the mechanism of the two types of spleen asthenia. The deficiency of the spleen deficiency syndrome can not be eaten and thin, the pain threshold is reduced, the sensitivity is relatively high, and the spleen deficiency syndrome can not be fed and fat, and the main manifestation is the gastrointestinal hormone disorder. At the same time, the traditional Chinese medicine can also significantly increase the percentage of the sugar water consumption rate of 75% of the rats in the group. And there is a quantitative effect relationship. In the case of gastrointestinal sensitivity, the spleen six gentleman soup of Jianpi traditional Chinese medicine can significantly reduce the high sensitivity of the stomach in the model group, and it has a quantitative effect correlation. The study shows that the prescription of Six Gentlemen of Jianpi prescription is effectively modified. Good functional dyspepsia spleen deficiency syndrome has three major main symptoms, such as less food, Na, and abdominal distention. In addition, it also improves the color of the rats and the symptoms of burnout and spleen deficiency. Therefore, the prescription of Six Gentlemen of Jianpi prescription has a therapeutic effect on the spleen deficiency syndrome of functional dyspepsia, and the model of iodine acetamide and iodine B copied in this study are also verified. Amido compound small platform model is highly consistent with the animal model of functional dyspepsia with spleen deficiency. In biological indexes, the addition and subtraction of Six Gentlemen of Jianpi prescription fragrant sand can increase the secretion of Ghrelin, CCK, VIP, and then increase circulating blood and the content of hypothalamus. Meanwhile, it can lower the content of SS protein in the gastric mucosa, serum and hypothalamus. There was no significant difference between SP and CGRP, and there was no significant difference between the drug treatment group and the IIb group. The SP and CGRP protein in the gastric mucosa of the addition and subtraction group of the Six Gentlemen of the fragrant sand was lower than that in the model group, and the changes of the SP in the gastric mucosa were significantly lower than those in the model group, but the sera and hypothalamus of the addition and subtraction of the six monarch of the fragrant sand were found in the hypothalamus. The changes of SP and CGRP were not obvious. The study found that the prescription of Jianpi Fang Decoction Six Gentlemen added and subtraction had a therapeutic effect on the spleen deficiency functional dyspepsia. Its target focused on the gastric mucosa, by regulating the formation of the brain gut peptide and then improving the symptoms of the functional dyspepsia of the spleen deficiency type. The mechanism of the dysfunction of the spleen deficiency syndrome is related to the regulation of the formation of the brain gut peptide. Conclusion the comprehensive study found that the main symptoms of functional dyspepsia in Chinese medicine are "upper abdominal distention", while "upper abdominal distention" as the main symptom of TCM is "ruffian". The loss of appetite is the criterion for determining the prosperity and decline of the pathogenic factors. According to this characteristic, we establish the animal model of functional dyspepsia and spleen deficiency syndrome, and then discuss the biological mechanism of the spleen deficiency syndrome of indigestion. The study found that the IIb and the compound small platform stand method conforms to the standard of the spleen deficiency of the dyspepsia, and the brain gut peptide. The change of hormone content is closely related to the spleen deficiency syndrome of indigestion type. Using Chinese medicine Jianpi to prove the model, it is found that the Chinese herbal medicine can adjust the formation of the peptide hormone of the brain and then improve the symptoms of the spleen deficiency syndrome. Therefore, this study proves that the possible biological mechanism of the functional dyspepsia is that the possible biological mechanism of the spleen deficiency syndrome is the change of the brain gut peptide, which is manifested as the stomach with the stomach. The gut gut peptide imbalance in intestinal promotion and inhibition has broadened the theory of "Spleen Deficiency" in traditional Chinese medicine to a certain extent.
【学位授予单位】：北京中医药大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R259
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本文编号：1984186