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目的观察痰湿壅盛证高血压模型大鼠的证候表现、理化指标及可能的病理机制。方法 50只Wistar大鼠采用高脂饲料饲养的方法制备痰湿壅盛证大鼠模型,同时设对照组(10只),给予普通饲料饲养。两组均持续饲养25周后,选取造模大鼠中体重、血压均超过对照组平均值25%的大鼠纳入模型组(22只)。检测两组大鼠体重、血压、血脂及相关血清学指标;观察靶器官形态变化;采用实时荧光定量PCR(q-PCR)检测主动脉中瘦素受体(leptin receptor,LepR)、Janus蛋白络氨酸激酶2(Janus kinase2,Jak2)、信号转导子和转录激活子3(signal transducer and activator of transcription 3,Stat3)、细胞因子信号抑制蛋白-3(suppressor of cytokine signaling-3,Socs3)、血管紧张素Ⅱ1型受体(angiotensinⅡreceptor type 1,AT1)、AT 2、磷脂酰肌醇3-激酶(phosphatidylinositol 3 kinase,PI3K)、丝苏氨酸蛋白激酶(serine threonine kinase,Akt)、核转录因子kBp65(nuclear factor of kappa B,NF-κBp65)、核因子kB激酶抑制蛋白α(inhibitor of nuclear factor kappa-B kinaseα,IKKα)、核因子kB抑制蛋白β亚基(NF-kappa-B inhibitorβ,IKKβ)、核因子kB抑制蛋白α亚基(NF-kappa-B inhibitorα,IKBα)和磷酸腺苷活化蛋白激酶(AMP-activated protein kinase,AMPK)mRNA表达水平。根据qPCR结果,分别采用免疫组化和Western blot检测主动脉中AT1、LepR的表达。结果与对照组比较,模型组大鼠体重、血压、血脂升高,血清Lep、AngⅡ、同型半胱氨酸(Hcy)、内皮素1(ET-1)、TNF-α、IL-6、β_2微球蛋白(β_2-MG)升高,NO下降,差异均有统计学意义(P<0.05,P<0.01)。模型组大鼠主动脉内皮损伤、平滑肌细胞增生,伴发心、肾损害。与对照组比较,模型组大鼠主动脉中LepR、Jak2、Stat3、Socs3、AT1、PI3K、Akt、NF-κB p65、IKKβ、IKBα、AMPK mRNA表达上调(P<0.05),IKKα下调(P<0.05)。免疫组化显示:模型组AT1和LepR棕黄色沉积明显增多、阳性部位分布更加广泛。Western blot显示:与对照组比较,模型组大鼠主动脉中AT1和LepR蛋白表达增加(P<0.05)。结论模型大鼠呈现典型的证候特征,体重增加,呈腹形肥胖状态,毛色黯淡,纳呆嗜睡,活动度下降,饮食减少,便溏,舌黯红,苔白厚腻。瘦素可作为评价痰湿壅盛证高血压大鼠模型的客观指标之一。
Objective To observe the syndromes, physicochemical indexes and possible pathological mechanisms of phlegm-dampness-hypertensive rats. Methods Fifty Wistar rats were fed with high-fat diet to establish the rat model of phlegm-dampness syndrome. At the same time, the control group (n = 10) was given normal feed. After 25 weeks of continuous feeding in both groups, rats of model group (22 rats) with body weight and blood pressure exceeding 25% of the average value of the control group were selected. The body weight, blood pressure, blood lipid and related serological indexes of the two groups were detected; the morphological changes of target organs were observed; the expressions of leptin receptor (LepR) and Janus protein in the aorta were detected by real-time quantitative PCR (q- Janus kinase 2 (Jak2), signal transducer and activator of transcription 3 (Stat3), suppressor of cytokine signaling-3 (Socs3) Angiotensin II receptor type 1 (AT1), AT2, phosphatidylinositol 3 kinase (PI3K), serine threonine kinase (Akt), nuclear transcription factor NF-κBp65, IκBα, NF-κBpβ, IKKβ, NF-κBp65, NF-κBp65, ), NF-kappa-B inhibitorα (IKBα) and AMP-activated protein kinase (AMPK) mRNA levels were detected. According to qPCR results, immunohistochemistry and Western blot were used to detect the expression of AT1 and LepR in the aorta respectively. Results Compared with the control group, body weight, blood pressure, blood lipids, Lep, AngⅡ, Hcy, ET-1, TNF- α, IL-6, Microglobulin (β_2-MG) increased, NO decreased, the difference was statistically significant (P <0.05, P <0.01). The aortic endothelium injury, smooth muscle cell proliferation, concomitant heart and kidney damage in model group rats. Compared with the control group, the expressions of LepR, Jak2, Stat3, Socs3, AT1, PI3K, Akt, NF-κB p65, IKKβ, IKBα and AMPK mRNA in the aorta of model group were significantly increased 0.05). Immunohistochemistry showed that the model group, AT1 and LepR brown yellow deposits increased significantly, the distribution of positive parts more widely. Western blot showed that compared with the control group, AT1 and LepR protein expression in the aorta of model rats increased (P <0.05). Conclusion The model rats showed typical syndrome characteristics, weight gain, belly-shaped obesity, dull coat, sleepiness, decreased activity, decreased diet, loose stools, dark red tongue and thick white fur. Leptin can be used as one of the objective indicators to evaluate phlegm-dampness syndrome-induced hypertension rat model.