本文選題：表征　切入點：葛根芩連湯　出處：《北京中醫(yī)藥大學(xué)》2014年碩士論文　論文類型：學(xué)位論文

【摘要】：目的 觀察葛根芩連湯對2型糖尿病大鼠宏觀表征、肛溫、抓力及相關(guān)指標是否具有改善作用,探索葛根芩連湯對模型動物中醫(yī)相關(guān)指標的作用及其與中醫(yī)臨床既往報道的一致性,為豐富中藥藥效評價體系、體現(xiàn)中醫(yī)藥治療特色奠定基礎(chǔ)；分析多項胰高血糖素樣肽(GLP-1)相關(guān)指標,從GLP-1相關(guān)機制的角度探索葛根芩連湯對2型糖尿病的可能作用機制,為利用GLP-1相關(guān)機制降糖中藥增添新的中藥復(fù)方 方法 1.高脂喂養(yǎng)8周后,腹腔注射30mg/kg鏈脲佐菌素(STZ)誘導(dǎo)構(gòu)建2型糖尿病大鼠模型,造模成功后,按大鼠按體重和血糖進行隨機分組,分為4組：2型糖尿病模型組、葛根芩連湯劑量1組(9.10g/kg, GGQL1組)、葛根芩連湯劑量2組(18.20g/kg, GGQL2組)、西格列汀組(0.01g/kg),另設(shè)正常對照組。 2.分組后對各組大鼠肛溫、抓力、飲食量、胰島素耐量、葡萄糖耐量進行檢測,其后進行上述藥物的干預(yù),灌胃6周,期間,模型組及給藥各組仍以高脂喂養(yǎng),正常組普通飼料喂養(yǎng)。每天給藥一次,進行一般情況觀察。 3.對大鼠體重、飲食量、空腹血糖值、抓力、肛溫進行動態(tài)檢測；給藥3周、6周,對各組大鼠進行胰島素耐量、葡萄糖耐量檢測；給藥6周,對各組大鼠進行表征量表采集。 4.給藥6周后,腹腔麻醉大鼠,腹主動脈取血并分離血清,應(yīng)用終點法測定血清中TG和TC值；取大鼠胰腺、回腸,甲醛固定,備用。 5.取甲醛固定組織胰腺、回腸,采用石蠟包埋技術(shù),制成石蠟切片備用；采用石蠟切片HE染色技術(shù)觀察胰腺、回腸組織病理結(jié)構(gòu)改變。 6.取大鼠胰腺石蠟包埋切片,應(yīng)用免疫組織化學(xué)檢測方法,檢測各組大鼠胰島內(nèi)胰島素的表達；采用TUNEL法檢測胰島細胞,觀察胰島B細胞凋亡情況。 7.取大鼠回腸石蠟包埋切片,應(yīng)用免疫組織化學(xué)方法檢測各組大鼠GLP-1、GIP的表達。 結(jié)果 1.體重：模型組體重顯著降低,GGQL1組體重值略有降低,GGQL2組基本保持穩(wěn)定； 2.空腹血糖：GGQL1組、GGQL2組于給藥4周、6周顯著降低,血糖值低于模型組(P0.05)； 3.飲食量：GGQL1組給藥后1-3周內(nèi)的飲食量低于模型組(P0.05)；GGQL2組于給藥1周后的飲食量均低于模型組(P0.05)； 4.表征觀察：與模型組相比,GGQL1組大鼠耳血管可見度適中、耳淺粉色、尾淡白、大便黑黃(P0.01/6),GGQL2組大鼠舌體適中、耳血管可見度適中、大便黑黃(P0.01/6),活動度增加、舌潤滑、耳淺粉色、鼻淡紅、爪粉紅不潤、尾黃中有白(P0.05/6)； 5.抓力：GGQL1組抓力值升高,給藥1周、4周抓力值高于模型組(P0.05),GGQL2組抓力值升高,給藥后4周、5周抓力值高于模型組(P0.05),且于給藥后5周高于GGQL1組； 6.肛溫：GGQL1組、GGQL2組肛溫值呈降低趨勢,于給藥5周時,GGQL1組肛溫值低于模型組(P0.05)； 7.胰島素耐量：給藥各組其血糖下降速率較高,血糖變化AUC值明顯低于模型組(P0.05),而與正常組無顯著性差異； 8.葡萄糖耐量：給藥各組血糖變化速率高于模型組,血糖變化AUC值高于模型組,其中GGQL2組明顯高于模型組(P0.05)； 9. TG、TC:GGQL1組TG、TC值均低于模型組但差異無統(tǒng)計學(xué)意義GGQL2組TG值(P0.05)低于模型組,TC值差異無統(tǒng)計學(xué)差異； 10.HE染色：與模型組相比,GGQL1組、GGQL2組胰島細胞均有所改善胰島較有規(guī)則,皺縮胰島少見,胰島細胞中空泡變性細胞有所減少,胰島細胞有所增加；回腸染色略有增加,由環(huán)狀黏膜肌層、黏膜下層、肌層組成的環(huán)層有所增厚,環(huán)狀較規(guī)則,皺襞分布緊密,內(nèi)部空腔相對減小 11.免疫組化、凋亡結(jié)果：GGQL1組、GGQL2組胰腺內(nèi)細胞凋亡率明顯低于模型組(P0.05),胰島素IOD/Area則高于模型組(P0.05),腸內(nèi)GLP-1平均光密度值高于模型組(P0.05),GIP平均光密度值低于模型組(P0.05)。 結(jié)論 1葛根芩連湯對2型糖尿病大鼠表征作用 葛根芩連湯可維持2型糖尿病大鼠的體重,減輕體重減輕癥狀；降低2型糖尿病大鼠的空腹血糖值,減輕高血糖癥；減少2型糖尿病大鼠的飲食量,減輕多食癥狀；增加2型糖尿病大鼠輕響抬頭率,改善眼睛黯淡無神、唇少光澤不潤、毛干燥、爪干澀不潤,改善大鼠活動減少、舌少津液、抓取反抗、鼻少光澤不潤,改善倦怠乏力、津液虧虛癥狀；增加2型糖尿病大鼠抓力,改善乏力癥狀；改善2型糖尿病病程中肛溫升溫階段,改善煩熱癥狀。由此可知,葛根芩連湯可以從不同程度改善2型糖尿病的癥狀。 2葛根芩連湯與GLP-1相關(guān)的生物學(xué)基礎(chǔ) 葛根芩連湯可改善胰島素抵抗,增加2型糖尿病大鼠注射胰島素30min后的血糖下降速率,增加2型糖尿病大鼠口服葡萄糖后血糖變化趨勢；調(diào)節(jié)血脂,有降低TG、TC值的趨勢作用；改善胰島、回腸形態(tài)；增加胰島內(nèi)分泌胰島素的胰島B細胞面積和胰島素分泌量；增加回腸內(nèi)GLP-1水平、減少GIP水平；減少胰島細胞內(nèi)的細胞凋亡率。葛根芩連湯除增加GLP-1水平、減少GIP水平、調(diào)節(jié)血脂的作用外,其他作用均與GLP-1的生物學(xué)機制相一致,故而認為,葛根芩連湯可能通過GLP-1的相關(guān)機制起到治療2型糖尿病的作用。
[Abstract]:objective
Observation of Gegenqinlian Decoction on rats with type 2 diabetes mellitus macro characterization, rectal temperature, grip and related indicators are improved, the role of Gegenqinlian Decoction on the related indexes of TCM animal model to explore the consistency and clinical reports, to enrich the efficacy of traditional Chinese medicine evaluation system, lay the foundation of TCM treatment characteristics a number of analysis; glucagon like peptide (GLP-1) related indicators, from the GLP-1 point of view to explore the related mechanism of Gegenqinlian Decoction on the possible mechanism of type 2 diabetes, add in medicine is a new mechanism of traditional Chinese medicine by GLP-1
Method
1. high fat diet for 8 weeks after intraperitoneal injection of 30mg/kg streptozotocin (STZ) induced rat model of type 2 diabetes, after the success of the model, according to the rats with the body weight and blood glucose were randomly divided into 4 groups: type 2 diabetes model group, Gegen Qinlian Decoction group (1 9.10g/kg GGQL1, group), Gegenqinlian Decoction group (2 18.20g/kg, GGQL2 group), Sig Leo Dean group (0.01g/kg), and normal control group.
2. groups of rats after rectal temperature, grip strength, diet, insulin tolerance, glucose tolerance test, followed by the drug intervention period by gavage for 6 weeks, the model group and the treatment groups with high fat diet, normal group fed with normal diet. Administered once a day, the general situation of the observation.
3., weight, diet, fasting blood glucose, grasping force and Anal temperature were dynamically detected in rats. After 3 weeks, 6 weeks, the rats were tested for insulin tolerance and glucose tolerance. After 6 weeks, the rats were collected for each group.
4. after 6 weeks of administration, the abdominal aorta blood was taken out and the serum was separated from the abdominal anesthetized rats. The TG and TC values in serum were determined by endpoint method, and the pancreas, ileum and formaldehyde were fixed in rats.
5. formaldehyde was used to immobilate tissue pancreas and ileum. Paraffin sections were made by paraffin embedding technology. Paraffin sections and HE staining were used to observe the pathological changes of pancreas and ileum.
6. the paraffin embedded sections of rat pancreas were taken. Immunohistochemistry was used to detect the expression of insulin in the islets of rats. The islet cells were detected by TUNEL and the apoptosis of islet B cells was observed.
7. the paraffin paraffin embedded section of rat ileum was sectioned and the expression of GLP-1 and GIP was detected by immunohistochemistry.
Result
1. weight: the weight of the model group decreased significantly, the weight value of the GGQL1 group decreased slightly, and the GGQL2 group remained stable.
2. fasting blood glucose: group GGQL1, group GGQL2 was given 4 weeks, 6 weeks decreased significantly, and blood sugar was lower than that in model group (P0.05).
3. diet: the diet in group GGQL1 was lower than that in model group (P0.05) in 1-3 weeks after administration (group GGQL2), and in group GGQL2 after administration, the diet was lower than that of model group (P0.05).
4. characterization observation: compared with the model group, GGQL1 group of rats with moderate ear vascular visibility, ear pale pink, pale yellow tail, black stool (P0.01/6), GGQL2 group rats tongue moderate, ear vascular visibility is moderate, black and yellow stool (P0.01/6), increased mobility, tongue lubrication, ear pale pink, the nose is reddish, pink claw does not run, yellow tail in white (P0.05/6);
5., grasping power: the grasping force value of group GGQL1 increased, and it was given for 1 weeks. The grasping force value of 4 weeks was higher than that of model group (P0.05), and the grasping force value of GGQL2 group increased. After 4 weeks and 5 weeks, the grasping force value of group GGQL2 was higher than that of model group (P0.05), and it was higher than that of GGQL1 group at 5 weeks after administration.
6. anus temperature: in group GGQL1, the value of anus temperature in group GGQL2 decreased. At 5 weeks of administration, the value of Anal temperature in group GGQL1 was lower than that of model group (P0.05).
7. insulin tolerance: the rate of blood sugar decline was higher in each group, and the AUC value of blood sugar was significantly lower than that of the model group (P0.05), but there was no significant difference between the group and the normal group.
8. glucose tolerance: the rate of blood glucose change in each group was higher than that in the model group, and the AUC value of blood glucose was higher than that in the model group, and the GGQL2 group was significantly higher than that in the model group (P0.05).
9. TG, TC:GGQL1 group TG, TC values were lower than the model group, but there was no statistically significant difference in GGQL2 group TG value (P0.05) was lower than the model group, TC value difference was not statistically significant difference.
10.HE staining: compared with the model group, GGQL1 group, GGQL2 group of islet cells were improved islet more rules, shrinkage islet rare, vacuolar degeneration of cells decreased islet cells, increased islet cell; ileum staining increased slightly by the circular muscularis mucosa, submucosa, muscular layer ring layer increases. The ring is regular, fold distribution closely, the internal cavity is relatively reduced
11. immunohistochemistry and apoptotic results: in group GGQL1, the apoptosis rate in pancreas of group GGQL2 was significantly lower than that of model group (P0.05), and insulin IOD/Area was higher than that of model group (P0.05). The mean GLP-1 optical density of intestinal GLP-1 was higher than that of model group (P0.05), and GIP average optical density was lower than that of model group (P0.05).
conclusion
The characterization of 1 Gegen Qin Lian Decoction on type 2 diabetic rats
Gegenqinlian decoction can be maintained in type 2 diabetic rats weight, reduce weight reduce symptoms; reduce in type 2 diabetic rats fasting blood glucose, reduce hyperglycemia in type 2 diabetic rats; reduce food intake, reduce the polyphagia; increase in type 2 diabetic rats light sound rise rate, improve dark eyes dull, dry lips, dry hair, claw dry run, improve the activity of rats decreased, tongue less fluid, less shiny nose grab resistance, not run, improve fatigue, body fluid deficiency symptoms; increased in type 2 diabetic rats grip, improve the fatigue symptoms; improve type 2 diabetes in anus the temperature rise stage, improve Fanre symptoms. Therefore, Gegenqinlian decoction can improve from the different degree of symptoms of type 2 diabetes.
Biological basis of 2 Gegen Qin Lian Decoction related to GLP-1
Gegenqinlian decoction can improve insulin resistance, increase insulin 30min in type 2 diabetic rats after glucose decreased rate of blood glucose change tendency of increased oral glucose in rats with type 2 diabetes mellitus; regulate blood lipids, reduce TG, TC value trend; improve islet, ileum form; increased islet insulin secreting islet B cell area and insulin secretion; increase the level of GLP-1 in the ileum, reduce the level of GIP; decrease apoptosis of pancreatic islet cells. The rate of Gegenqinlian Decoction in addition to increase the level of GLP-1, reduce the level of GIP, the function of regulating blood fat, which he was consistent with the biological mechanism of GLP-1 and that of Gegenqinlian decoction may through the relevant mechanism of GLP-1 to treat type 2 diabetes.

【學(xué)位授予單位】：北京中醫(yī)藥大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R285.5

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