米诺环素通过影响下丘脑室旁核小胶质细胞活化水平降低两肾一夹所致高血压的作用机制

Mechanism of Minocycline in Reducing Hypertension Induced by Two Kidneys and One Clip by Affecting the Activation Level of Microglia in the Paraventricular Nucleus of Hypothalamus

目的:观察米诺环素对两肾一夹(2k1c)型高血压大鼠血浆炎性因子、神经递质浓度及血压的影响,并探讨其是否通过影响下丘脑室旁核(PVN)发挥调控作用。方法:将雄性无特定病原体(SPF)级SD大鼠28只随机分为假手术(SHAM)+人工脑脊液(aCSF)组、SHAM+minocycline组、2k1c+aCSF组、2k1c+minocycline组。通过肾动脉缩窄术制作模型并进行室旁核置管,利用无创血压测量仪监测大鼠血压;10周后进行肾脏交感神经放电(RSNA)检测;通过小胶质细胞染色统计激活小胶质细胞的数量及细胞突起的长度;采用高效液相色谱法(HPLC)检测下丘脑室旁核(PVN)中白细胞介素1β(IL-1β)、白细胞介素6(IL-6)、去甲肾上腺素(NE)、谷氨酸(GLU)、γ-氨基丁酸(GABA)含量。实时定量聚合酶链式反应(PCR)检测gp^(91phox)mRNA在PVN中的表达。采用酶联免疫吸附法(ELISA)检测大鼠血浆IL-1β、IL-6和NE浓度。结果:中枢缓慢输注米诺环素可减少激活小胶质细胞数量,增加细胞突起长度(P<0.05)。2k1c+aCSF组和2k1c+minocycline组大鼠PVN和血浆IL-1β、IL-6、NE水平及PVN中GLU含量高于SHAM+aCSF组和SHAM+minocycline组(P<0.05),米诺环素可逆转上述表达(P<0.05)。2k1c+aCSF组和2k1c+minocycline组大鼠PVN中GABA含量低于SHAM+aCSF组和SHAM+minocycline组(P<0.05),经米诺环素治疗可有效提高2k1c+minocycline组大鼠GABA表达量(P<0.05)。与SHAM+aCSF组和SHAM+minocycline组大鼠比较,2k1c+aCSF组和2k1c+minocycline组大鼠PVN中gp^(91phox)mRNA表达升高(P<0.05),米诺环素可降低PVN中gp^(91phox)mRNA表达(P<0.05)。2k1c+aCSF组和2k1c+minocycline组大鼠肾交感放电高于SHAM+aCSF组和SHAM+minocycline组(P<0.05),经米诺环素治疗后肾交感放电有效减弱(P<0.05)。结论:缓慢输注米诺环素通过影响下丘脑室旁核小胶质细胞活化水平从而降低2k1c所致的高血压。

Objective:To observe the effects of minocycline on plasma inflammatory factor,neurotransmitter concentration and blood pressure in 2 kidney and 1 clip(2k1c)hypertensive rats,and to explore a regulatory role by hypothalamic paraventricular nucleus(PVN).Methods:Twenty-eight male specific pathogen free(SPF)grade SD rats were randomly divided into SHAM+artificial cerebrospinal fluid(aCSF)group,SHAM+minocycline group,2k1c+aCSF group,and 2k1c+minocycline group.The model was established by renal artery narrowing and paraventricular nuclear cannulation was performed,and the blood pressure of rats was monitored using a non-invasive blood pressure meter.Renal sympathetic nerve discharge(RSNA)detection was performed after 10 weeks.The number of activated microglia and the length of the cell process were measured by microglia staining.The contents of interleukin-1β(IL-1β),interleukin-6(IL-6),noradrenaline(NE),glutamic acid(GLU)and gamma-aminobutyric acid(GABA)in PVN were detected by high performance liquid chromatography(HPLC).Rea-l time quantitative polymerase chain reaction(PCR)was used to detect gp^(91phox)mRNA expression in PVN.Plasma levels of IL-1β,IL-6,and NE were detected by enzyme-linked immunosorbent assay(ELISA).Results:Central chronic infusion of minocycline could reduce the number of activated microglia and increase the length of cell processes(P<0.05).The levels of IL-1β,IL-6 and NE in plasma and the GLU in PVN of the 2k1c+aCSF group and 2k1c+minocycline group were higher than those of the SHAM+aCSF group and SHAM+minocycline group(P<0.05),and minocycline could reverse the above expression(P<0.05).The GABA content of PVN in the 2k1c+aCSF group and 2k1c+minocycline group was lower than that in the SHAM+aCSF group and SHAM+minocycline group(P<0.05).GABA expression in the 2k1c+minocycline group effectively increased by minocycline treatment(P<0.05).Compared with SHAM+aCSF group and SHAM+minocycline group,gp^(91phox) mRNA expression in PVN of 2k1c+aCSF group and 2k1c+minocycline group was increased(P<0.05).Minocycline decreased gp^(91phox)mRNA expression in PVN(P<0.05).The renal sympathetic discharge in the 2k1c+aCSF group and 2k1c+minocycline group was higher than that in the SHAM+aCSF group and SHAM+minocycline group(P<0.05),and the renal sympathetic discharge was effectively weakened after minocycline treatment(P<0.05).Conclusion:Chronic infusion of minocycline could reduce hypertension by affecting the activation level of microglia in the paraventricular nucleus of hypothalamus.