Inhibitory effect and mechanism of total flavonoids from Melicope pteleifolia on transplanted tumor of colorectal cancer in nude mice / 中国药房

OBJECTIVE To study the inhibitory effect and mechanism of total flavonoids from Melicope pteleifolia （TF-MPL） on transplanted tumor of colorectal cancer in nude mice. METHODS The transplanted tumor model of colorectal cancer was induced by injecting 0.2 mL colorectal cancer cell LoVo subcutaneously via the right armpit of nude mice. After successful modeling， nude mice were randomly divided into model group， 5-fluorouracil group （positive control， 10 mg/kg）， TF-MPL high- dose and low-dose groups （25， 12.5 mg/kg）； a normal group （normal saline containing 0.3% carboxymethyl cellulose sodium） without modeling was additionally set up， with 6 mice in each group. Each group was intraperitoneally injected with the corresponding drug solution/solvent for 21 consecutive days. The inhibitory rate of the transplanted tumor， liver and spleen index， and the levels of tumor necrosis factor-α （TNF-α） and interleukin-6 （IL-6） in serum were detected after the last medication； the morphological changes of tumor tissue were observed； immunohistochemical staining was used to detect protein expressions of Toll- like receptor 4 （TLR4） and nuclear factor-κB subunit p65 （NF-κB p65） in tumor tissue of nude mice. Western blot assay was used to detect protein expressions of TLR4， myeloid differentiation factor 88 （MyD88）， TNF receptor-associated factor 6 （TRAF6）， interleukin-1 receptor-associated kinase 1 （IRAK-1）， NF-κB p65 and caspase-3 in tumor tissue of nude mice. RESULTS Compared with the model group， TF-MPL high-dose group showed a significant decrease in tumor weight （inhibitory rate of 36.91%）， liver and spleen index， serum levels of TNF-α and IL-6 and protein expressions of TLR4， MyD88， TRAF6，IRAK-1 and NF- κB p65 （P＜0.05 or P＜0.01）； the expression of caspase-3 protein was increased significantly （P＜0.05）， and more tumor cell shrinkage and deformation， nuclear pyknosis and fragmentation were observed. CONCLUSIONS TF-MPL can significantly inhibit the growth of transplanted tumor of colorectal cancer in nude mice， the mechanism of which may be associated with reducing inflammatory response， inhibiting TLR4/MyD88/NF-κB signaling pathway， and promoting apoptosis in colorectal cancer cells.

Effects of Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination on inflammatory responses in atherosclerotic mice / 中国中药杂志

The study aims to observe the effects and explore the mechanisms of Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination in the treatment of the inflammatory response of mice with atherosclerosis(AS) via the Toll-like receptor 4(TLR4)/myeloid differentiation primary response protein 88(MyD88)/nuclear factor-κB(NF-κB) signaling pathway. Male ApoE~(-/-) mice were randomly assigned into a model group, a Buyang Huanwu Decoction group, an Astragali Radix-Angelicae Sinensis Radix combination group, and an atorvastatin group, and male C57BL/6J mice of the same weeks old were used as the control group. Other groups except the control group were given high-fat diets for 12 weeks to establish the AS model, and drugs were administrated by gavage. Aortic intimal hyperplasia thickness, blood lipid level, plasma inflammatory cytokine levels, M1/M2 macrophage markers, and expression levels of proteins in TLR4/MyD88/NF-κB pathway in the vessel wall were measured to evaluate the effects of drugs on AS lesions and inflammatory responses. The results showed that the AS model was successfully established with the ApoE~(-/-) mice fed with high-fat diets. Compared with the control group, the model group showed elevated plasma total cholesterol(TC), triglyceride(TG), and low-density lipoprotein cholesterol(LDL-c) levels(P<0.05), thickened intima(P<0.01), and increased plasma tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) levels(P<0.01). Moreover, the model group showed increased expression of vascular cell adhesion molecule-1(VCAM-1) and inducible nitric oxide synthase(iNOS)(P<0.01), inhibited expression of endothelial nitric oxide synthase(eNOS) and cluster of differentiation 206(CD206)(P<0.01), and up-regulated mRNA and protein levels of TLR4, MyD88, NF-κB inhibitor alpha(IκBα), and NF-κB in the vessel wall(P<0.05). Compared with the model group, Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination lowered the plasma TC and LDL-c levels(P<0.01), alleviated the intimal hyperplasia(P<0.01), and reduced the plasma TNF-α and IL-6 levels(P<0.05). Moreover, the two interventions promoted the expression of eNOS and CD206(P<0.05), inhibited the expression of VCAM-1 and iNOS(P<0.01), and down-regulated the mRNA and protein levels of TLR4, MyD88, IκBα, and NF-κB(P<0.05) in the vessel wall. This study indicated that Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination could delay the progression of AS, inhibit the polarization of vascular wall macrophages toward M1 type, and attenuate vascular inflammatory response by inhibiting the activation of TLR4/MyD88/NF-κB signaling pathway in the vascular wall. Astragali Radix and Angelicae Sinensis Radix were the main pharmacological substances in Buyang Huanwu Decoction for alleviating the AS vascular inflammatory response.

Regulatory effect of Di'ao Xinxuekang on TLR4/MyD88/NF-κB signaling pathway in atherosclerotic rats / 中国中药杂志

The aim of this paper was to observe the effect of Di'ao Xinxuekang(DXXK) on TLR4/MyD88/NF-κB signaling pathway in atherosclerotic rats, and to explore its anti-atherosclerotic mechanism. Sixty SD rats were randomly divided into normal group, model group, atorvastatin group(4.0 mg·kg~(-1)), and DXXK groups(100, 30, 10 mg·kg~(-1)), with 10 rats in each group. The atherosclerosis model was induced by high fat diet plus vitamin D_2. Experimental drugs were administered intragastrically once daily for 8 weeks starting from the 9 th week. Biochemical analyzers were used to detect levels of triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C) and high-density lipoprotein cholesterol(HDL-C) in blood lipid. The levels of serum tumor necrosis factor(TNF)-α, interleukin(IL)-6 and IL-1β were detected by ELISA. Pathological changes of aortic tissues were observed by using Sudan Ⅳ and HE staining. The mRNA and protein expressions of TLR4, MyD88 and NF-κB p65 in aortic tissues were detected by RT-PCR and Western blot, respectively. As compared with the model group, TC, TG, and LDL-C levels in serum were significantly decreased, HDL-C content was significantly increased, and levels of TNF-α, IL-6, and IL-1β in serum were significantly decreased in atorvastatin group and DXXK high and middle dose groups. Aortic lesions in atorvastatin group and DXXK group were significantly improved, and the mRNA and protein expressions of TLR4, MyD88, NF-κB p65 in the aorta were decreased. DXXK has a preventive and therapeutic effect on atherosclerosis in rats, and its mechanism may be related to inhibiting inflammatory reaction by regulating TLR4/MyD88/NF-κB signal transduction, thereby inhibiting the progression of atherosclerosis.

Intervention Effects of Berberine on Mice Macrophage Polarization Based on TLR 4/MyD88/NF-κB Signaling Pathway / 中国药房

OBJECTIVE：To study the effects of berberine on mic e macrophage polarization based on TLR 4-MyD88-NF-κB signaling pathway. METHODS ：Using mice RAW 264.7 macrophage as the object ，atorvastatin calcium as positive control ， inflammatory cell model was induced by lipopolysaccharide （LPS）；ELISA method was used to detect the contents of TNF-α，IL-6 and NF-κB in cell culture medium after treated with low，medium and high doses of berberine （5，10，20 μmol/L）for 24 h. The real-time fluorescence quantitative PCR was conducted to determine the mRNA expression of TLR 4 and MyD 88 in cells. Western blotting assay was used to detect the protein expression of TLR 4，MyD88，iNOS and CD 206 in cells. RESULTS ：Compared with blank control group ，the contents of TNF-α，IL-6 and NF-κB in cell culture medium，mRNA expression of TLR 4 and MyD 88， protein expression of TLR 4，MyD88 and iNOS in cells were increased significantly in LPS induction group （P＜0.05）. Compared with LPS induction group ，the contents of TNF-α and IL-6，mRNA and protein expression of TLR 4 and MyD 88 in atorvastatin calcium group ，berberine medium-dose and high-dose groupsas well as the content of NF-κ B and protein expression of iNOS in administration groups were decreased significantly ， while the content of NF-κB in berberine high-dose group was significantly lower than atorvastatin calcium group （P＜0.05）. The protein expressions of CD206 in atorvastatin calcium group and berberine high-dose group were increased significantly ，while the protein expression of CD 206 in berberine high-dose group was significantly higher than atorvastatin calcium group （P＜0.05）. CONCLUSIONS ：Different doses of berberine can intervene in mice macrophage polarization to different extents ，the mechanism of which may be associated with the regulation of TLR4/MyD88/NF-κB signaling pathway.

Effect of lycopene on glycolipid metabolism and pancreatic tissue through regulation of TLR4/MyD88/NF-κB signaling pathway in high-fat diet induced obese mice / 中草药

Objective: To investigate the effects of lycopene on glycolipid metabolism and pancreatic tissue inflammation in obese mice and its underlying mechanism. Methods: The obese mouse model was induced by a high-fat diet (HFD). The effects of lycopene on body weight, blood glucose, blood lipid and body fat were observed after 8 weeks of administration. Pathological changes of pancreas were observed by HE staining. Protein expressions involved in TLR4/MyD88/NF-κB signaling pathway were detected by Western blotting, and the degree of macrophage infiltration in the pancreatic tissues of obese mice were detected by IHC. Results: Lycopene significantly inhibited body weight gain and reduced fasting blood glucose, as well as improved glucose tolerance and blood lipid level in obese mice. In addition, lycopene also reduced vacuolization, edema degeneration, islet hypertrophy and other inflammatory lesions in pancreatic tissues. Moreover, the protein expression levels of TLR4, MyD88 and NF-κB in pancreatic tissue were decreased and the inflammatory infiltration was reduced. Conclusion: Lycopene can improve blood glucose, lipid metabolism and pancreatic inflammation in obese mice, and its mechanism may be related to the regulation of TLR4/MyD88/NF-κB signaling pathway.

Experimental study on effect and mechanism of Danzhi Jiangtang Capsules on diabetic myocardial injury / 中国中药杂志

Diabetic cardiomyopathy( DCM) is one of the major cardiovascular complications of diabetes mellitus. Based on the clinical efficacy of Danzhi Jiangtang Capsules( DJC) in the prevention and treatment of diabetes and its cardiovascular complications,both in vivo and in vitro methods were adopted to investigate its effect and underlying mechanism of protecting myocardial injury induced by diabetes. The type 2 diabetic rats were prepared by feeding high-energy food combined with streptozotin( STZ) injection,and the effects of DJC were observed by blood sugar,blood lipid,hemodynamic index,cardiac weight index and the change of cardiac pathological morphology. The protein expressions of TLR4,MyD88 and NF-κB p65 in myocardial tissue were detected and the possible mechanism was preliminarily analyzed. Besides this,DJC containing serum was prepared,H9 c2 cardiomyocyte induced by high sugar were studied to investigate the mechanism of TLR4/MyD88/NF-κB signaling pathway regulating cardiomyocyte injury and the therapeutic effect of DJC. The results demonstrated that fasting blood sugar,glycosylated hemoglobin,total cholesterol and glycerol triglyceride were significantly reduced( P<0. 01,P<0. 05). Cardiac weight index,left ventricle weight index,LVEDP and the protein expressions of TLR4,MyD88 and NF-κB p65 were significantly reduced( P<0. 01,P<0. 05). LVSP,+dp/dtmaxand-dp/dtmaxincreased significantly( P<0. 01,P< 0. 05). Moreover,the pathological damage of myocardial tissue in rats improved significantly. Meanwhile,the protein expressions of TLR4,MyD88 and NF-κB p65 in cardiomyocytes induced by high sugar were significantly inhibited( P<0. 01).It showed that DJC were effective in preventing and treating myocardial injury induced by diabetes and its mechanism may be related to the over-expression of TLR4/MyD88/NF-κB signaling pathway induced by high sugar.

Effects of Anzi Mixture on TLR4/MyD88/NF-κB Signaling Pathway of Antiphospholipid Antibodies Posi-tive Abortive Mice / 中国药房

OBJECTIVE:To study the effects of Anzi mixture on Toll like receptor 4(TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear facter-κB(NF-κB)signaling pathway of antiphospholipid antibodies(APA)positive abortive mice,and to inves-tigate the mechanism of anti-APA positive abortion. METHODS:BALB/c mice(female)were randomly divided into blank control group,model group,aspirin group (positive control,0.0195 g/kg) and Anzi mixture low-dose,medium-dose and high-dose groups (37.7,75.4,150.8 g/kg,calculated by crude drug),with 10 mice in each group. Except for blank control group,other groups were given human β2-glycoprotein Ⅰ as derivant to establish APA positive abortion model. From the first day of pregnancy, treatment groups were given relevant medicine intragastrically,and blank control group and model group were given constant vol-ume of normal saline intragastrically,once a day,for consecutive 9 d. mRNA and protein levels of TLR4,myeloid differentiation 2 (MD2),MyD88 and NF-κB in placental tissue of mice were determined by RT-PCR and immunohistochemical method. RE-SULTS:Compared with blank control group,mRNA and protein expression of TLR4,MD2,MyD88 and NF-κB in placental tis-sue were increased markedly in the model group(P<0.01). Compared with model group,mRNA and protein expression of TLR4, MD2 and MyD88 in aspirin group and Anzi mixture low-dose and medium-dose groups were decreased significantly as well as the protein expression of TLR4 in Anzi mixture high-dose group and the protein expression of NF-κB in all medicine groups(P<0.05 or P<0.01). mRNA expression of TLR4 and MD2 and the protein expression of MD2 and MyD88 in Anzi mixture low-dose groups were lower than those in aspirin group (P<0.05 or P<0.01). CONCLUSIONS:Anzi mixture can inhibit TLR4/MyD88/NF-κB signaling pathway of APA positive abortive mice,which may be one of anti-APA positive abortion mechanisms.