Effects And Mechanisms Of Chaihushugan Powder On Gastric Ulcer In Rats With Stress

Research background:Stress is that body by a variety of stressors (internal and external environment, social and psychological factors) causes systemic non-specific adaptive response, which is the result of the interaction of the central nervous system, immune system and endocrine, mainly with the LC-Noradrenergic neurons/Sympathetic-adrenal medulla axis and the Hypothalamic-Pituitary-Adrenal axis excited which causes neuroendocrine response, and general adaptation syndrome. Appropriate stress can mobilize the body’s non-specific adaptive system, conducive to the body which quickly adapts to the environment, and maintain the body’s homeostasis. while the load is too strong or too long, stress can damage body tissues or organs. Stress effects are not only manifested in the body, but also psychological harm. Continuous body stress will inevitably lead to psychological stress. Psychological stress is that the body through awareness, evaluation feels the threat of stressors, which causes the process of change of psychological and physiological function. Clinical studies have shown that stress and75%to90%of diseases are closely related. With the life pace accelerating and competitive pressures increasing, there is a rising trend in the prevalence of psychosomatic diseases. In modern psycho-social medical model, the relationship between stress and disease as well as the mechanism of stress is a current research focus of the medical profession.Stress is closely related to the development of psychosomatic diseases of the gastrointestinal. gastrointestinal is the most sensitive psychosomatic organ, which is vulnerable to the impact of internal and external environmental stimuli and emotional factors. Stress may cause disorder of gastrointestinal hormone through neuroendocrine changes, affecting the movement and function of the gastrointestinal. The incidence of gastric ulcer is related to gastrointestinal hormones. Gastrointestinal hormones which exist not only in the gastrointestinal endocrine cells and nerve cells, also in the central nervous system, are small molecule active substances or peptides, as a nerve transmitter substance of the information, also known as brain-gut peptide. The brain-gut peptide through regulation of the brain-gut axis regulate gastric acid and gastrointestinal motility. Researches on the brain-gut peptide in the central level (LC) affecting gastrointestinal function are poor. Traditional Chinese Medicine(TCM) believes that emotional abnormalities or any form of stress can cause disorder of Qi ascending, descending, exiting and entering, disorder of Qi, Blood, Yin and Yang, which led to the dysfunctional organs and a series of body homeostasis. Liver in TCM has a certain function of adjustment mechanism of neuroendocrine immunomodulation (NIM) network. Some scholars believe that the liver is the body’s regulation center of stress response, is the core of the body to regulate brain-gut peptide. Failure of liver to convey and disperse causes stagnation of liver Qi, stomach disorder due to liver Qi or liver stagnation and spleen deficiency, which led to dysfunction of gastrointestinal digestion, absorption, secretion, movement.《Su Wen·Liu Yuan Zheng Ji Da Lun》 was said,"Relieving liver depression","Joy increases while the evil liver depression","liver being the resolute viscera, liver of yin nature with yang functioning". Posterity think the method of the sparse liver to regulate the flow of vital energy as the main treatment method. Related experiments show that the method of the sparse liver to regulate the flow of vital energy has a good effect on treating psychosomatic diseases of gastrointestinal, regulating gastrointestinal hormone, improving gastrointestinal function. Chaihushugan powder is from 《Jing Yue Quan Shu》,and studies confirm the Chaihushugan powder can affect the metabolism of neurotransmitters.Traditional Chinese Medicine and modern medicine both consider that Emotional changes can lead to gastrointestinal motility and absorption change. Modern medicine from the angle of the brain-gut axis explains this process and the pathophysiological mechanisms, however Traditional Chinese Medicine explains the reason that incoordination between the liver and the spleen, stomach disorder due to liver qi and liver stagnation and spleen deficiency. We assume that emotional changes can through liver depression (neurotransmitter changes)lead to spleen deficiency(disorders of gastrointestinal motility, absorption,etc), in fact which is the the brain gut axis between the contact. The link between the brain-gut axis is one of the main lines which reflect Functional network of the liver depression and spleen deficiency syndrome. Researching neuropeptides (brain-gut peptides) which are an important factor linking the central nervous system and gastrointestinal system, helps to understand the relationship of the emotions-gastric ulcer, emotional-liver-stomach, to explore the essence of TCM syndrome.Objective:To find Effects and mechanisms of Chaihushugan Powder on gastriculcer in Rats with Acute stress, the behavior and Gastric histology in Rats with Acute Stress were observed. Gas and SS in plasma gastric antrum and LC were tested by Elisa and the expression of Gas and SS in gastric antrum and LC by immunohistochemistry.Methods:1. Influences of Chaihushugan Powder on Behavior and Gastric histology in Rats with Acute Stress1.1Animal grouping and preparation of GU model Thirty-two female Wistar rats with SPF level and at the weight of180-220g were into four groups according to a completely random method.those were normal control group, gastric ulcer model group, Chaihushugan powder group and ranitidine group. In each group, there were eight rats. Except normal group, rats of other groups were induced three times12hours by using unpredictable stress of cool-water swim, clipping tail, shock, etc. Before preparing GU model, Chaihushugan powder group and the ranitidine group were administrated Chaihushugan powder and ranitidine suspension (1ml per100g body weight)for a week (once everyday). At the same time, the model group and the normal group were given equal volume of physiological saline.1.2Indicators detection The changes of behavior (Scores of horizontal and vertical movements, consumption of sucrose solutions and the forced swim test)in Rats with Acute Stress were observed. After the last stress two hours, all rats were killed, removed stomach, then injected10ml10%neutral formalin solution to the stomach from the pylorus, cut the stomach along the greater curvature after10minutes and flatted it. Tested the tissue indexes, and then measured the maximum transverse diameter and longitudinal diameter of ulcers with a caliper, and denote them by d1、 d2. Both the product were the ulcer area. Depth of ulcer was calculated more than or less than2mm. Ulcer size was expressed with ulcer index. Then embedded stomach tissues with olefin and sliced up. At last, the tissue indexes were tested, pathology examination by HE dyeing.2. The Effect of Chaihushugan powder on Gastrin(Gas) and Somatostatin(SS) in plasma gastric antrum and locus coeruleus (LC)of gastric ulcer model rats2.1Animal grouping and preparation of GU model (as1.1)2.2Indicators detection Anaesthetized rats, prepared for abdominal skin, opened abdomen, and collected about3ml abdominal aortic blood into ordinary vacuum mining vessels, after natural solidification, centrifugated them for10min at the speed of2000r/min, separated the plasma and saved them at-20℃refrigerator. Isolated and cut the stomach along the greater curvature side, scraped about0.1g of the gastric mucosa with slide, weighing them by electronic analytical balance, heating to boiling and then putting it into homogenizer which filled with lml lmol/L CH3COOH homogenized, following by lml lmol/L NaOH. At last, put them into glass test tube,3500r/min centrifugal15min, and the supernatant fluid was packed in the refrigerator at-70℃refrigerator. Opened head and removed LC, prepared homogenate by routine, then put them into glass test tube,5000r/min centrifugal10min,the supernatant fluid reserved at-20℃, respectively in the determination of content Of Gas and SS in plasma gastric antrum and LC of gastric ulcer model rats by Elisa.3. Statistical analysisThe experimental results are expressed by x±s. SPSS13.0software was applied into One-way ANOVA for mean comparison based on multiple samples. When the variance was homogeneous, Least-significant difference (Bonferroni)was applied for test and the value of P<0.05would mean significant difference.Results:1. Influences of Chaihushugan Powder on Behavior and Gastric histology in Rats with Acute StressIn comparison with normal control group, body quality, scores of horizontal and vertical movements in model group was significantly reduced (P<0.05),whereas in the forced swim test the static time increased remarkably (P<0.05). While compared with model group, Chaihushugan Powder not only increased scores of horizontal and vertical movements, induced static time and reduce ulcer index (all P<0.05) increased consumption of sucrose solutions,(P>0.05),but also alleviated Gastric residues, the quantities and extents of gastric mucosal lesion.2. The Effect of Chaihushugan powder on Gastrin(Gas) and Somatostatin(SS) in plasma gastric antrum and locus coeruleus (LC)of gastric ulcer model ratsIn comparison with gastric ulcer model group, Gas in gastric antrum and in plasma, SS in gastric antrum of Chaihushugansan group were significantly lower (P<0.05),while SS in plasma,GAS and SS in LC were not evidently different (P>0.05); Ranitidine reduced obviously GAS in LC and SS in plasma (both P<0.05),no effect on SS in LC.Conclusions:1.The method of replicating gastric ulcer model by using unpredictable stress of cool-water swim, clipping tail, shock, etc, not only had a high stability and good feasibility,but also simulated that abnormal emotions triggered the process of hepatic depression syndrome, and a more reasonable observation of emotional change caused gastric ulcers.2.Chaihushugan Powder can alleviate behavior of rats with acute stress,increased scores of horizontal and vertical movements, consumption of sucrose solutions, induced static time. In a word, Chaihushugan Powder can improve Depression state.3.Chaihushugan Powder not only reduced ulcer index(P<0.05), alleviated Gastric residues,the quantities and extents of gastric mucosal lesion, but also promoted mucosal repair, proliferation, reduced inflammatory cells infiltration. It may play certain effects against psychological stress.4.Chaihushugan powder has the function of alleviating the content of peripheric or central GAS and SS, which may be one of its therapeutic mechanisms on gastric ulcer with acute stress, which suggested traditional Chinese formulations had some targets in overall modulating role.