胃萎清治疗慢性萎缩性胃炎的临床疗效及作用机制研究

发布时间：2018-01-17 16:30

本文关键词：胃萎清治疗慢性萎缩性胃炎的临床疗效及作用机制研究　出处：《广州中医药大学》2017年博士论文　论文类型：学位论文

【摘要】：背景:慢性萎缩性胃炎是慢性胃炎的一种类型,是指胃黏膜固有腺体出现萎缩,可伴有肠上皮化生及异型增生,带来严重的公共卫生及社会问题,严重影响患者社会工作能力及生存质量。慢性萎缩性胃炎是慢性浅表性胃炎和胃癌之间的"桥梁",是胃黏膜恶性转变的关键阶段,通过对慢性萎缩性胃炎机制的深入研究,不仅可以一定程度揭示炎症导致炎性肿瘤的发病机理,更能实施针对性预防措施,降低胃癌的发病率及死亡率。胃萎清是我们临床治疗慢性萎缩性胃炎的经验方,该方由北芪,白术,半枝莲,莪术,五指毛桃,枳壳组成,共奏健脾理气,化瘀解毒的功效,以促进脏腑功能恢复,改善患者的临床症状为首务。JAK2/STAT3信号通路和细胞增殖凋亡密切相关,活化的STAT3可以诱导Bcl-2、Bcl-xL基因的表达,从而抑制胃黏膜细胞的凋亡,促进胃黏膜慢性炎症相关的胃癌发生,因此抑制JAK2/STAT3信号通路提供了一种防治胃癌发生的可能。本研究旨在评估胃萎清颗粒治疗慢性萎缩性胃炎的临床有效性及安全性,同时以JAK2/STAT3信号通路为切入点,分析胃萎清对慢性萎缩性胃炎(CAG)大鼠的作用机制,进一步揭示胃萎清改善慢性萎缩性胃炎病理的分子生物学机制。目的:(1)评估胃萎清颗粒治疗慢性萎缩性胃炎患者的临床有效性及安全性。(2)本实验用MNNG配合饥饱失常建立慢性萎缩性胃炎的大鼠模型,观察胃萎清颗粒对慢性萎缩性胃炎大鼠一般情况的影响,同时胃萎清对慢性萎缩性胃炎大鼠腺体萎缩、肠上皮化生的治疗效果。(3)探讨通过观察JAK2/STAT3信号通路在慢性萎缩性胃炎中的表达情况,探讨胃萎清颗粒对JAK2/STAT3信号通路关键因子的调控作用。方法:(1)临床研究:纳入慢性萎缩性胃炎患者,随机分为胃萎清组和叶酸组,胃萎清组予胃萎清颗粒剂,1次1包,1日3次。叶酸组予叶酸片,1次5mg,1日3次。给药24周后复查胃镜及病理,评估组间胃黏膜萎缩、肠化、异型增生、慢性炎症、活动性的治疗前后改变。并且评估组间上腹(胃脘)疼痛、上腹(胃脘)胀闷、上腹(胃脘)堵闷、烧心、反酸、嗳气、食欲减退、食量减少各单项症状积分及症状总积分的治疗前后改变情况。(2)实验研究:70-80天SPF级SD雄性大鼠94只,体重170-200克,适应性喂养1周后,随机分为2组,即:空白组,慢性萎缩性胃炎模型组。空白组14只,慢性萎缩性胃炎模型组80只。空白组给予正常饮食,自由饮水。慢性萎缩性胃炎模型组予180μg/mL的MNNG饮用液自由饮用,结合饥饱失常法复制慢性萎缩性胃炎大鼠模型,连续造模至第14周,随机抽取2只慢性萎缩性胃炎模型组老鼠,进行胃黏膜病理形态组学检查,两只大鼠病理结果均显示为胃黏膜固有腺体萎缩伴肠上皮化生,表明造模成功。造模成功后将慢性萎缩性胃炎模型组随机分为5组:模型组,胃萎清高剂量组,胃萎清中剂量组,胃萎清低剂量组,叶酸组。正常对照组和模型对照组给予生理盐水灌胃。胃萎清高、中、低剂量组灌胃给药浓度分别为15.6g/kg/d、7.8g/kg/d、3.9g/kg/d。叶酸组给药浓度为1.46mg/kg/d。灌胃容积10ml/kg,每日1次,连续灌胃10周。实验结束后取小弯侧胃体—窦交界处胃黏膜,制备胃黏膜上皮组织切片。HE染色法观察各组大鼠胃黏膜上皮萎缩、肠上皮化生情况。AB-PAS染色法观察各组大鼠胃黏膜上皮肠上皮化生病变范围;刮取剩余大鼠胃黏膜,Western Blot法检测JAK2、STAT3、p-STAT3 的表达。RT-PCR 法检测 Bcl-2、Bcl-xL 的表达。结果:(1)临床研究:本试验共入组64例,治疗组34例,对照30例。其中2例治疗组患者脱落,总共62例患者参与数据分析。①病理方面,胃萎清对慢性炎症积分改善的效果优于叶酸组。②在主要症状积分改善方面,治疗组在改善上腹(胃脘)疼痛方面效果优于对照组,对照组在治疗上腹(胃脘)胀满方面效果优于治疗组,其它方面积分治疗组与对照组对比,差异无统计学意义。(2)实验研究:①HE染色显示模型组胃黏膜腺体大部分萎缩消失,粘膜肌层增厚,间质内有炎性细胞浸润,淋巴滤泡形成,可见肠上皮化生。有不同程度的变性或坏死的脱落细胞,粘膜层变薄,腺体细胞排列紊乱,腺体呈囊状扩张,上皮细胞形态大小不一,间质水肿、充血。胃黏膜全层出现肠上皮化生,可见大量杯状细胞。叶酸组大鼠胃黏膜呈粉红色,光泽度差,胃黏膜变薄、皱襞紊乱,见大量炎细胞浸润。间质水肿、充血、炎细胞浸润,区域粘膜可见大量肠上皮化生。胃萎清三个剂量组病理介于叶酸与空白组之间,其中以胃萎清高剂量组病理改善最明显,胃萎清中剂量组次之,胃萎清低剂量组病理改善效果不明显。AB-PAS染色显示正常组大鼠胃黏膜上皮无蓝色或紫色肠上皮化生灶;模型组出现弥漫性肠上皮化生,胃萎清治疗后,则主要集中在胃腔侧的细胞,其中以胃萎清高剂量组和中剂量组改善最明显,叶酸组改善不明显;②JAK2蛋白在模型组的表达最高,其中胃萎清高剂量组和胃萎清中剂量组表达较模型组降低。STAT3蛋白在模型组的表达最高,胃萎清三个剂量组表达较模型组降低,其中高剂量组降低最明显。p-STAT3蛋白在模型组的表达最高,胃萎清三个剂量组表达较模型组降低,其中胃萎清高剂量组较模型组降低最明显。和模型组比较,胃萎清高剂量组Bcl-2mRNA表达下降,差异有统计学意义(P0.05)。和模型组比较,胃萎清高剂量组Bcl-xL mRNA表达下降,差异有统计学意义(P0.05)。结论:(1)胃萎清可改善患者病理的慢性炎症积分。同时胃萎清可改善上腹(胃脘)疼痛。这为胃萎清治疗慢性萎缩性胃炎提供临床证据。(2)本次研究采用MNNG溶液自由饮用、饥饱失常法的综合造模方法,成功复制了慢性萎缩性胃炎大鼠模型。(3)胃萎清能在一定程度上缓解大鼠胃黏膜萎缩、肠上皮化生。(4)慢性萎缩性胃炎阶段JAK2/STAT3的激活可能是促进黏膜细胞过度增殖,参与胃黏膜恶性转变过程的重要步骤,胃萎清可一定程度抑制JAK2/STAT3信号通路的活化。
[Abstract]:Background: chronic atrophic gastritis is a type of chronic gastritis, refers to gastric glands atrophy, intestinal metaplasia and dysplasia, bring social and public health problem is serious, serious impact on social work ability and life quality of patients with chronic atrophic gastritis, chronic superficial gastritis and gastric cancer between the "bridge", is the key stage of gastric malignant transformation, through in-depth study of the mechanism of chronic atrophic gastritis, can not only reveal the degree of inflammation pathogenesis of inflammatory tumor, it can implement the preventive measures to reduce the incidence and mortality of gastric cancer. Gastric atrophy is the experience of chronic atrophy gastritis treated in our clinic, the side from Beiqi, Atractylodes, Scutellaria barbata, rhizoma curcumae, Radix Fici Hirtae, Fructus aurantii, played a total of Jianpiliqi, removing blood stasis detoxification, to promote viscera function recovery, improve patients The clinical symptoms of the first.JAK2/STAT3 signal pathway and proliferation and apoptosis is closely related to the activation of STAT3 can induce Bcl-2, Bcl-xL gene expression, thereby inhibiting the apoptosis of gastric mucosal cells, promote gastric mucosa of chronic inflammation related, the inhibition of JAK2/ STAT3 signaling pathway provides a gastric cancer prevention. This research to evaluate the clinical efficacy and safety of Wei Wei Qing granule in the treatment of chronic atrophic gastritis, and the JAK2/STAT3 pathway as a starting point, analysis of Wei Wei Qing on chronic atrophic gastritis (CAG) rats, to further reveal the molecular mechanism of Wei Wei Qing to improve the pathology of chronic atrophic gastritis. Objective: (1) to assess the clinical efficacy and safety of Wei Wei Qing granule in the treatment of patients with chronic atrophic gastritis (2). The experiment used MNNG with the establishment of chronic atrophic gastritis hunger The rat model, to observe the effect of Weiwei Granule on chronic atrophic gastritis rats in general, at the same time, Wei Wei Qing on chronic atrophic gastritis rats glandular atrophy, intestinal metaplasia (3) to investigate the effect of treatment. By observing the expression of JAK2/STAT3 signaling pathway in chronic atrophic gastritis, to explore the regulation effect of Wei Wei Qing Granule on the key factors of the JAK2/STAT3 signaling pathway. Methods: (1) clinical research: in patients with chronic atrophic gastritis, gastric atrophy were randomly divided into clear group and folic acid group, Weiwei group, Qing Wei Wei Qing granules, 1 Pack 1 times 1, 3 times a day. The folic acid group to Folic Acid Tablets. The 1 time 5mg, 1 3 times a day. Review of gastroscope and pathology after treatment for 24 weeks, the evaluation group of gastric atrophy, intestinal metaplasia, dysplasia, chronic inflammation, activity changes before and after treatment and between groups were evaluated. The abdomen (stomach pain), abdomen (stomach fullness), abdomen (stomach) plugging heartburn, acid regurgitation and belching. , loss of appetite, reduce food intake changes before and after the treatment of each single symptom score and total symptom score. (2) experimental study: 70-80 days SPF 94 male SD rats, weighing 170-200 grams, after 1 weeks of feeding, were randomly divided into 2 groups: control group, chronic atrophic gastritis model group. The blank group 14, model of chronic atrophic gastritis group 80. Control group were given normal diet, free drinking drinking drinking liquid. The MNNG model of chronic atrophic gastritis group received 180 g/mL, combined with the hunger to copy the chronic atrophic gastritis rat model, continuous modeling to fourteenth weeks, random selected 2 rats model of chronic atrophic gastritis rats, gastric mucosa were pathological examination group, two rats pathological results showed gastric glands atrophy with intestinal metaplasia, showed that the model was successful. After a successful modeling of chronic atrophic gastritis were randomly divided into model group 5 Group: model group, high dose group, Wei Wei, Wei Wei Qing Wei Wei Qing middle dose group, low dose group, folic acid group and normal control group and model control group were given normal saline. Wei Wei Gao, in the low dose group, gavage concentration were 15.6g/kg/d, 7.8g/kg/d, 3.9g/kg/d. folic acid group. The drug dosage was 1.46mg/kg/d. intragastric volume 10ml/kg, 1 times daily, 10 weeks after the end of the experiment. The lesser curvature of gastric body and antrum junction of gastric mucosa, preparation of gastric epithelial tissue sections were observed by.HE in rat gastric mucosa atrophy and intestinal metaplasia staining, observe condition.AB-PAS pathological changes of gastric mucosa epithelial range of intestinal epithelial rats staining; gastric mucosal scraping the remaining rats, detection of JAK2, Western Blot STAT3, Bcl-2 p-STAT3 method to detect the expression of.RT-PCR, the expression of Bcl-xL. Results: (1) clinical research: this experiment into the group of 64 cases, 34 cases in the treatment group and the control 30 Cases. The treatment group of 2 cases of patients off a total of 62 patients participated in the data analysis. The pathology of gastric atrophy, cleaning effect is better than that of the folic acid group of chronic inflammation. The main points to improve the improvement of symptoms, the treatment group in improving abdominal pain (stomach) is better than the effect of the control group, the control group in the treatment of the abdomen (stomach fullness) is better than the treatment group, the other scores of the treatment group compared with the control group, the difference was not statistically significant. (2) experimental study: 1. HE staining showed that the model group of gastric mucosa gland atrophy mostly disappeared, muscularis mucosa thickening, interstitial inflammatory cell infiltration, lymph follicle formation, visible intestinal metaplasia. There are different degrees of degeneration or necrosis of exfoliated cells, mucous layer thinning, glandular cell disorder, gland cystic dilatation, epithelial cell size, interstitial edema and hyperemia. Gastric mucosal full-thickness intestinal on Metaplasia, showing a large number of goblet cells. The folic acid group in rat gastric mucosa was pink, gloss, gastric mucosa thinning, fold disorder, see a large number of inflammatory cells infiltration. Interstitial edema, hyperemia, inflammatory cell infiltration, mucous membrane area shows a large number of intestinal metaplasia. Between the Wei Wei Qing three dose group between pathology folic acid with the control group, the high dose of Weiwei group improved the most obvious pathological, Wei Wei Qing middle dose group, low dose group of Wei Wei Qing improved the pathological.AB-PAS staining showed that the effect is not obvious in normal group in rat gastric mucosa without blue or purple intestinal metaplasia lesions; diffuse intestinal metaplasia model Wei Wei Qing group, after treatment, mainly concentrated in the stomach cavity side of the cell, the Wei Wei Gao dose group and medium dose group improved the most obvious improvement is not obvious, the folic acid group; the JAK2 protein expression in the highest model group, high dose group, and the Wei Wei Wei Wei In the dose group was lower than model group.STAT3 protein expression in the model group the highest, Wei Wei Qing three dose group was lower than the model group, the high dose group decreased most significantly in the model group, the highest.P-STAT3 protein expression, Wei Wei Qing three dose group was lower than that of model group, the gastric atrophy high dose group than in the model group decreased most significantly. Compared with the model group, decreased expression of Wei Wei high dose group Bcl-2mRNA, the difference was statistically significant (P0.05). Compared with the model group, high dose group, Bcl-xL Wei Wei the expression of mRNA decreased, the difference was statistically significant (P0.05). Conclusion: (1) chronic stomach inflammation score Wei Qing can improve the patient pathology. At the same time, Wei Wei Qing (stomach) can improve abdominal pain. This is Wei Wei Qing to provide clinical evidence for the treatment of chronic atrophic gastritis (2). This study uses MNNG solution free drinking, the comprehensive modelling method into hunger method. Work to build the model of chronic atrophic gastritis rats. (3) Weiwei cleanergy to some extent alleviate rat gastric mucosal atrophy, intestinal metaplasia, chronic atrophic gastritis (4) stage of the activation of JAK2/STAT3 may promote proliferation of mucosal cells in gastric mucosa malignant transformation is an important step of the stomach Wei Qing can effectively inhibit the activation of JAK2/STAT3 signaling pathway.

【学位授予单位】：广州中医药大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R259

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