游离脂肪酸激活HaCaT细胞表面TLR对NF-ΚB信号转导通路的影响

目的:观察长链游离脂肪酸对体外培养的角质形成细胞的作用,探讨其对TLR及NF-ΚB信号转导通路的影响。方法:①体外培养Hacat细胞,实验分为N、PA、SA、LPS四组,N组为正常对照组,软脂酸(Palmitic Acid,PA)750μmol/L、硬脂酸(Stearic Acid,SA)750μmol/L、脂多糖(lipopolysaccharide,LPS)500 ng/L和Hacat细胞共培养72h组分别为N组、PA组、SA组、LPS组;②LPS、SA与角质形成细胞共培养72h检测LPS、SA对Hacat细胞NF-κB、p-IκBα、IκBα表达的影响。通过免疫印迹法(Weston blot)检测SA、LPS对Hacat细胞TLR2、TLR4、NF-ΚB、p-IκBα、IκBα表达的影响;Real Ti me PCR方法检测对Hacat细胞内NF-κB、TNF-αmRNA表达的影响。结果:SA、LPS激活细胞表面TLR后可以明显促进细胞表面TLR2、TLR4表达及细胞内NF-κB、TNF-α等促炎症因子的表达。PA促进Hacat细胞内NF-ΚB、p-IκBα、IκBα表达无明显影响(P>0.05)结论:SA可促进角质形成细胞分泌NF-κB、TNF-α等因子表达促进炎症反应;其机制可能为:SA可能通过TLR及NF-κB信号传导通路促进细胞释放NF-κB、TNF-α等促炎因子。

Gualou Xiebai Banxia Decoction(瓜蒌薤白半夏汤) Inhibits NF-kappa B-dependent Inflammation in Myocardial Ischemia-reperfusion Injury in Rats

Objective: To evaluate the myocardial protective effect of Gualou Xiebai Banxia decoction (瓜蒌薤白半夏汤GXBD) and explore the mechanisms of inhibition of NF-kappa B activation and blockade of inflammatory responses induced by ischemia-reperfusion in rats. Methods: Twenty-four Sprague Dawley (SD) rats were randomly divided into three groups. Rats in the treatment group received GXBD (13 g crude drug/kg) for three weeks, while rats in the model control and normal control groups received equal volumes of distilled water. On the 22nd day, rats in the ischemia-reperfusion (I/R) control and GXBD-treated groups underwent 30 min occlusion of the left anterior descending (LAD) coronary artery, followed by 120 min reperfusion. Electrocardiogram was recorded, and the activities of cardiac enzymes, cytokines, and NF-кB were assessed after I/R. Results: Compared with the I/R control group, GXBD treatment restored the activity of the specific myocardial-injury marker creatine kinase (CK) and lactate dehydrogenase (LDH), and inhibited the inflammatory response involving the nuclear factor-кB (NF-кB) pathway, including down-regulation of interleukin (IL)-1β and IL-6, and up-regulation of IL-10 gene expression. Conclusion: GXBD strongly reduced myocardial impairment in our I/R model, including inhibition of NF-кB activation and inflammatory cytokine responses.

Effect of warming yang and supplementing kidney moxibustion therapy on the NF-κB signaling pathway of vascular dementia rats

Objective： To observe the effect of warming yang and supplementing kidney moxibustion therapy on the behavioristics and the expression levels of hippocampal IL-1β, TNF-α and NF-κB related genes and proteins of vascular dementia（VD） rats, and to explore the mechanism of warming yang and supplementing kidney moxibustion therapy in inhibiting VD inflammatory response. Methods Seventy-eight SD rats, except 10 rats as sham operation group（group A）, were established into VD models by applying the ischemia reperfusion method in bilateral common carotid arteries. 30 models were successful and were randomly divided into model group（group B）, moxibustion group（group C） and western medication group（group D）, with 10 rats in each group. After modeling, suspended moxibustion was conducted at ＂Dàzhuī（大椎GV 14）＂,＂Mìngmén（命门GV 4）＂ and ＂Guānyuán（关元CV 4）＂ of the rats in group C for15 min/time. The intervention was performed for once a day, and intervention for 4 consecutive weeks was needed. Intragastric administration with nimodipine（2 mgkg-1d-1） was carried out in the rats in group D for 4 consecutive weeks. Morris water maze test was adopted for behavioral test in the rats in each group. HE staining was conducted in order to observe the pathological changes. RT-qPCR method and Western blot method were used for detecting the expression levels of hippocampal IL-1β, TNF-αand NF-κB related genes and proteins.Results： After modeling, the Morris water maze escape latency of rats in operation groups was obviously longer than that in group A（P〈0.05）. According to HE staining results, the arrangement of neurons in group B was incompact, the cell membranes were undefined, the cytoplasm was heterogeneous, karyopyknosis was found, the amount of necrosis increased, and the number of inflammatory cells significantly increased. Compared with group B, the number of inflammatory cells significantly reduced in group C and group D, and the amount of neuron necrosis also reduced. After intervention, compared with group B, the expression levels of TNF-a, p-IκB and NF-κBp65 mRNA in the hippocampal of the rats in group C and group D significantly decreased（all P〈0.05）, compared with group B, the expression levels of IL-1β, TNF-α and NF-κBp65 mRNA in the hippocampal of rats in group C and group D also significantly decreased（all P〈0.05）.Conclusion： Warming yang and supplementing kidney moxibustion therapy may inhibit the NF-κB signaling pathway and reduce the inflammatory response through lowering the expression levels of IL-1β,TNF-α and NF-κB mRNA in the brain, thus achieving the purpose of treating VD.