目的 研究安神养血口服液对更年期综合征大鼠的治疗效果及机制.方法 大鼠随机分为正常对照组,假手术组,模型组,安神养血口服液低、中、高剂量组,戊酸雌二醇组,每组10只.各组小鼠灌胃给予相应药物28 d后,HE染色观察子宫组织病理变化,ELISA法检测血清雌二醇(E2)、促黄体生成素(LH)、促卵泡生成素(FSH)、促性腺激素释放激素(GnRH)、睾酮(T)及下丘脑多巴胺(DA)、去甲肾上腺素(NE)、五羟色胺(5-HT)、五羟吲哚乙酸(5-HIAA)、β-内啡肽(β-EP)水平,WST-1法、TBA法检测血清超氧化物歧化酶(SOD)、丙二醛(MDA)水平.结果 与模型组比较,安神养血口服液组大鼠血清E2、T、SOD水平升高(P<0.05,P<0.01),LH、FSH、Gn RH、MDA水平降低(P<0.05,P<0.01),下丘脑5-HT、5-HIAA水平降低(P<0.01),DA、NE、β-EP水平升高(P<0.05,P<0.01),子宫系数增加(P<0.05),可有效改善子宫组织病理变化.结论 安神养血口服液可以改善更年期综合征模型大鼠血清相关性激素及下丘脑单胺类神经递质水平,同时有效提高血液过氧化物酶活性,增强抗氧化作用.

目的 建立瑶药止痛散的质量标准.方法 采用薄层色谱(TLC)法对瑶药止痛散中毛两面针、洋金花、草乌进行定性鉴别;采用高效液相色谱(HPLC)法测定瑶药止痛散中氯化两面针碱的含量,色谱柱为Waters XBridge Phengl C18柱(150 mm×2.1 mm,3.5μm),流动相为乙腈-0.2％磷酸溶液-三乙胺(20:80:1,V/V/V),流速为1.0 mL/min,检测波长为271 nm,柱温为30℃,进样量为10μL.结果 瑶药止痛散中毛两面针、洋金花、草乌的TLC图斑点清晰,分离度好,阴性对照无干扰.氯化两面针碱质量浓度在1.25～12.50μg/mL范围内与峰面积线性关系良好;精密度、重复性、稳定性试验结果的RSD均小于2.0％;平均加样回收率为100.28％,RSD为1.75％(n=9).结论 该研究所建立的标准可用于瑶药止痛散的质量控制.

目的 优选复方杨桃颗粒的成型工艺.方法 以吸湿率、成型率、溶化性因素为考察指标,筛选最佳辅料与配比,确定复方杨桃颗粒的成型工艺.结果 最佳成型工艺条件为:干浸膏:可溶性淀粉:糊精=1:1:0.2,90％乙醇为润湿剂,按照该成型工艺制备3批颗粒并作评价.颗粒的成型率可达80％以上,粒度检查、水分测定和溶化性均符合规定,颗粒临界相对湿度(CRH)为66.91％.结论 该成型制备工艺方法合理可行,可为大规模生产提供科学的试验依据.

急性缺血性脑卒中具有高发病率、致残率和病死率,已成为影响人类健康和生活的主要疾病.溶栓治疗是目前国内外指南一致推荐的治疗急性缺血性脑卒中的首选方法.瑞替普酶是目前临床上广泛使用的溶栓药物,目前美国FDA已批准用于治疗心肌梗死,具有溶栓特异性高,溶栓速度快等特点.瑞替普酶在治疗急性缺血性脑卒中取得较好的疗效,但仍需大量样本临床试验来论证,本文就瑞替普酶静脉溶栓治疗急性缺血性脑卒中的研究进展做以概述和总结,为今后治疗脑梗死选择溶栓药物提供依据.

目的:探讨杨桃根中2-十二烷基-6-甲氧基-2,5-二烯-1,4-环己二酮(DMDD)对裸鼠肝癌的抑瘤作用,并研究其机制.方法:建立人肝癌裸鼠皮下移植瘤模型,48只裸鼠随机分为6个组:DMDD高剂量组,DMDD中剂量组,DMDD低剂量组,多柔比星组,模型组,空白组,每组8只.记录裸鼠体质量和肿瘤大小变化,计算瘤重,抑瘤率,肿瘤指数及脏器指数;检测血常规指标:白细胞(WBC)、红细胞(RBC)、血红蛋白(HGB)和血小板(PLT);采用酶联免疫法检测裸鼠血清中白介素2(IL-2)和白介素10(IL-10)的含量;免疫组化法检测TLR4,MyD88和NF-κB的蛋白表达水平.结果:DMDD各剂量组及多柔比星组的瘤体质量均小于模型组(P＜0.01),DMDD高、中、低剂量组及多柔比星组的抑瘤率分别:66.39％,63.11％,47.33％,49.38％,DMDD各剂量组和多柔比星组的肿瘤指数低于模型组(P＜0.01),DMDD各剂量组的脾脏指数低于模型组(P＜0.01);DM-DD各剂量组均能升高WBC、HGB和PLT的数量(P＜0.01或P＜0.05);DMDD各剂量组能降低的血清中IL-2和IL-10的表达水平(P＜0.01或P＜0.05);免疫组化结果显示,DMDD各剂量组均能下调TLR4,MyD88和NF-κB的蛋白表达水平.结论:DMDD具有良好的抗肝癌作用,该作用可能与其能改善裸鼠免疫器官的功能和血常规指标,下调IL-2和IL-10的水平,调控TLR4/MyD88/NF-κB信号通路有关.

Background: 2-Dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione (DMDD) has been reported to inhibit a variety of cancer cell lines.The purpose of this study was to investigate the effects of DMDD on 4T1 breast cancer cells and the effects of DMDD on 4T1 breast cancer in mice and its molecular mechanisms.Methods: 4T1 breast cancer cells were treated with different concentrations of DMDD, and their proliferation, apoptosis, cell-cycle distribution, migration, and invasion were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT, Acridine orange and ethidium bromide dual staining analysis (AO/EB) dual staining, flow cytometry, scratch test, and the Transwell assay.Relative quantitative real-time qPCR analysis and Western blot were applied to examine the expression levels of related genes and proteins.In animal experiments, we established a xenograft model to assess the anti-breast cancer effects of DMDD by evaluating the inhibition rate.The apoptotic activity of DMDD was evaluated by hematoxylin-eosin (HE) staining, transmission electron microscope (TEM) analysis and TdT-mediated dUTP nick end labeling (TUNEL) assays.The mRNA expression levels of MAPK pathway components were detected by relative quantitative real-time qPCR.In addition, the protein expression levels of MAPK pathway components were assessed through immunohistochemical assays and Western blotting.Results: Experiments showed that DMDD could inhibit the proliferation, migration, invasion of 4T1 cells and induce cellular apoptosis and G1 cell cycle arrest.Moreover, DMDD down-regulated the mRNA expressions of raf1, mek1, mek2, erk1, erk2, bcl2, and upregulated the mRNA expression of bax.DMDD reduced the protein expressions of p-raf1, p-mek, p-erk, p-p38, Bcl2, MMP2, MMP9 and increased the protein expressions of Bax and p-JNK.The results showed that DMDD can effectively reduce the tumor volume and weight of breast cancer in vivo, up-regulate the expression of IL-2, down-regulate the expression of IL-4 and IL-10, induce the apoptosis of breast cancer cells in mice, and regulate the expression of genes and proteins of the MAPK pathway.Conclusion: Our study indicates that DMDD can inhibit proliferation, migration, and invasion and induces apoptosis and cell-cycle arrest of 4T1 breast cancer cells.Also, our findings indicate that DMDD induces the apoptosis of breast cancer cells and inhibits the growth in mice.Its mechanism may be related to the MAPK pathway.

Aβ1–42-induced neurotoxicity has been considered as a possible mechanism to aggravate the onset and progression of Alzheimer’s disease (AD). In this study, we aim to determine the protective effect of DMDD on the apoptosis of SH-SY5Y cells induced by Aβ1–42 and elucidate potential mechanism of DMDD’s protective function in apoptosis. CCK-8, AnnexinV-FITC/PI flow cytometry, and transmission electron microscopy analysis were used to determine the protection of DMDD on Aβ1–42-evoked apoptosis of SH-SY5Y cells. Cytochrome c release, JC-1 staining, and measuring the protein of Bcl-2 family by Western blot were applied to elucidate the mechanism of DMDD’s protective function in apoptosis. Three concentration of DMDD (5 μmol/L, 10 μmol/L, and 20 μmol/L) rescues the cell viability loss and apoptosis of SH-SY5Y cells cultivated in Aβ1–42. The expressions of cleaved Caspase-3, -8, -9, the cytochrome c release, and mitochondrial membrane potential loss were inhibited by DMDD in Aβ1–42-insulted SH-SY5Y cells. The Western blot analysis showed that DMDD pretreatment clearly downregulated the protein of Bax and upregulated Bcl-2. Moreover, the Bcl-2/Bax ratio was obviously decreased in cells only exposed to Aβ1–42, but, which was suppressed by treated with DMDD. DMDD attenuated the apoptosis of SH-SY5Y cells induced by Aβ1–42 through reversing the Bcl-2/Bax ratio.

[目的]研究瑶药止痛散的最佳制备工艺条件.[方法]采用L9(34)正交设计,以氯化两面针碱含量为指标,考察烘烤温度、粉碎细度和烘烤时间因素的影响,优选最佳制备工艺参数.[结果]最佳制备工艺为:烘干温度80℃,烘干3 h,粉碎成100目.[结论]瑶药止痛散的制备工艺稳定可靠,可为产品实际生产提供可靠保证.

[This corrects the article DOI: 10.2147/DMSO.S209436.].

ObjectiveDiabetic kidney disease (DKD) is the leading cause of death and disability of diabetes mellitus. However, there is still a lack of specific drugs for the treatment of DKD. The chief aim of this research is to investigate the role and mechanism of 2-Dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione (DMDD) for DKD.MethodsWild type and TLR4 knockout mice were induced to diabetes. After 4-week treatment with DMDD, blood sugar, renal function, blood lipid and pathological changes were assessed. Real-time PCR, western blotting, and immunohistochemistry were employed to detect the expressions of TLR4, TGFβ1 and Smad2/3 in the renal tissue.ResultsDMDD improved the serum lipid and decreased fasting blood glucose levels in diabetic mice. CysC and urinary albumin levels increased markedly in the diabetic group, and they were obviously decreased after 4 weeks of DMDD treatment. Compared with the WT diabetic mice, the urinary albumin and CysC in the TLR4-/- mice were expressed at lower levels. HE and Masson’s staining revealed that DMDD clearly ameliorated pathological changes and renal fibrosis. When TLR4 gene was knock out, the pathological was improved. Mechanistically, TLR4, TGF-β1 and Smad2/3 were obvious up-regulation in the renal tissues of diabetic mice. The expressions of these proteins were significantly down-regulated after DMDD treatment (p < 0.05). In the TLR4-/- mice, mRNA and protein levels of TGF-β1 and Smad2/3 were obviously lower than those in the WT mice. In addition, IHC revealed that a strong in situ expressions of TLR4, TGF-β1 and Smad2/3 were seen in the kidney tissues of diabetic mice, which were distinctly weakened in the DMDD-treated mice. In the TLR4-/- mice, however, expressions of TGF-β1 and Smad2/3 were not remarkable increase in the diabetic mice compared with normal mice.ConclusionsThese results strongly indicate that TLR4 is essential for DMDD protection against renal dysfunction in diabetic mice. Its hypoglycemic and anti-fibrosis effects were likely mediated by the TLR4/TGFβ signaling pathway.

Objective: Diabetic kidney disease (DKD) is the leading cause of death and disability of diabetes mellitus. However, there is still a lack of specific drugs for the treatment of DKD. Objective: The chief aim of this research is to investigate the role and mechanism of DMDD for DKD.Methods: Wild type and TLR4 knockout mice were induced to diabetes. After 4-week treatment with DMDD, blood sugar, renal function, blood lipid and pathological changes were assessed. Real-time PCR, western blotting, and immunohistochemistry were employed to detect the expressions of TLR4, TGFβ1 and Smad2/3 in the renal tissue.Results: DMDD improved the serum lipid and decreased fasting blood glucose levels in diabetic mice. CysC and urinary albumin levels increased markedly in the diabetic group, and they were obviously decreased after 4 weeks of DMDD treatment. Compared with the WT diabetic mice, the urinary albumin and CysC in the TLR4-/- mice were expressed at lower levels. HE and Masson's staining revealed that DMDD clearly ameliorated pathological changes and renal fibrosis. When TLR4 gene was knock out, the pathological was improved. Mechanistically, TLR4, TGF-β1 and Smad2/3 were obvious up-regulation in the renal tissues of diabetic mice. The expressions of these proteins were significantly down-regulated after DMDD treatment (p< 0.05). In the TLR4-/- mice, mRNA and protein levels of TGF-β1 and Smad2/3 were obviously lower than those in the WT mice. In addition, IHC revealed that a strong in situ expressions of TLR4, TGF-β1 and Smad2/3 were seen in the kidney tissues of diabetic mice, which were distinctly weakened in the DMDD-treated mice. In the TLR4-/- mice, however, expressions of TGF-β1 and Smad2/3 were not remarkable increase in the diabetic mice compared with normal mice.Conclusions: These results strongly indicate that TLR4 is essential for DMDD protection against renal dysfunction in diabetic mice. Its hypoglycemic and anti-fibrosis effects were likely mediated by inhibiting of the TLR4/TGFβ signaling pathway, and the downregulation of Smads in the renal interstitium.Funding Statement: This research was supported by the National Natural Science Foundation of China (No. 81760665, 81460205), and Guangxi Natural Science Foundation under Grant No. 2017GXNSFBA198177. In addition, this project was financially supported by the project of improving the basic ability of young teachers in colleges and universities in Guangxi (KY2016YB098).Declaration of Interests: The authors declare no conflicts of interest in this work.Ethics Approval Statement: All experimental procedures were approved by the Institutional Animal Care and Use Committee of Guangxi Medical University.

目的:探讨玉郎伞多糖(YLSP)对大鼠放射性肺损伤的作用及机制.方法:将120只SD大鼠随机分为5组:正常对照组、单纯照射组及玉郎伞多糖低剂量(YLSPL)组、中剂量(YLSPM)组和高剂量(YLSPH)组,每组24只.YLSPL组、YLSPM组、YLSPH组分别按150 mg/kg、300 mg/kg、600 mg/kg于照射前1周开始灌胃给予YLSP,正常对照组和单纯照射组给予等量生理盐水.单纯照射组和YLSP各给药组大鼠麻醉后用DT γ射线单次20 Gy照射大鼠右肺,正常对照组仅麻醉不予照射.分别于照射后第10、第40、第70天,每组取8只大鼠行腹主动脉取血并处死,采用酶联免疫吸附试验(ELISA)法检测血清转化生长因子β1(TGF-β1)和白细胞介素-6(IL-6)含量;取出双肺,计算湿肺系数;采用苏木精—伊红(HE)染色法观察肺组织病理学改变;检测肺组织超氧化物歧化酶(SOD)活性及丙二醛(MDA)含量;荧光定量PCR (qPCR)法检测Smad2 mRNA相对表达量.结果:正常对照组肺组织及肺泡结构完好,无充血、渗出、水肿等;单纯照射组照射后早期出现充血、水肿、炎症细胞浸润、肺泡间隔增宽,之后肺组织纤维化加重、肺泡腔变小、消失;YLSP各给药组肺组织病理明显改善.照射后,YLSP各给药组大鼠血清TGF-β1和IL-6含量、Smad2 mRNA相对表达量及肺组织MDA含量均下降,肺组织SOD活性升高,以YLSPH组最为显著(P＜0.05).YLSP各给药组湿肺系数照射后第10、第40、第70天均显著下降(均P＜0.05).结论:YLSP对放射性肺损伤大鼠有保护作用,其机制可能与降低血清TGF-β1及IL-6含量、下调Smad2 mRNA表达、抗自由基和抑制脂质过氧化损伤作用有关.

目的:研究柿寄生提取物对对乙酰氨基酚(APAP)诱导小鼠肝损伤的保护作用及作用机制.方法:将60只健康雄性昆明种小鼠随机分成正常组、模型组、联苯双酯阳性组(150 mg·kg-1)、柿寄生提取物低、中、高剂量组(11.67,23.34,46.67mg·kg-1),灌胃给药8d后,除正常组外其他各组腹腔注射265 mg· kg-1对乙酰氨基酚,建立急性肝损伤模型,眼球取血后常规检测血清谷丙转氨酶(ALT)、谷草转氨酶(AST)活性水平、肝匀浆生化指标和肝脏病理.结果:与模型组相比,柿寄生提取物中剂量能明显降低AST、ALT活性水平,降低肿瘤坏死因子-α、白细胞介素-6、白细胞介素-1β表达水平(P＜0.05),降低丙二醛活力水平(P＜0.05),显著提高超氧化物歧化酶和谷胱甘肽过氧化物酶活性、提高微量还原型谷胱甘肽的水平(P＜0.05),改善肝组织病理学变化.结论:柿寄生萃取物能明显减轻APAP所致的肝损伤,其机制与降低炎症因子水平,提高机体抗氧化能力,降低氧化应激有关.

目的:探讨山栀子根提取物对四氯化碳诱导肝纤维化大鼠的影响.方法:采用连续灌胃法(8 w,2次/w)给予50％四氯化碳花生油(1∶1混合)复制肝纤维化模型.检测血清中门冬酸氨基转移酶(AST)、丙氨酸氨基转移酶(ALT),肝脏组织中超氧化物歧化酶(SOD)、丙二醛(MDA)、还原型谷胱甘肽(GSH)、谷胱甘肽过氧化物酶(GSH-Px)的活性.HE染色观察大鼠肝脏组织病变情况.免疫组织化学法检测肝脏组织中α平滑肌肌动蛋白(α-SMA)及Ⅳ型胶原蛋白(ColⅣ)的蛋白表达情况.结果:与模型组比较,山栀子根提取物给药组血清ALT、AST及肝脏组织中MDA显著降低,肝脏组织中的SOD、GSH、GSH-Px显著升高.HE染色结果可见肝脏组织病变减轻.α-SMA及ColⅣ阳性表达率明显减少.结论:山栀子根提取物对四氯化碳诱导的肝纤维化大鼠具有保护作用,其机制可能与降解细胞外基质及抗脂质过氧化作用有关.

BACKGROUND:Hyperglycemia stimulated epithelial-mesenchymal transition (EMT) plays a critical role in initiating and progressing renal fibrosis in diabetic kidney disease (DKD). It is crucial to explore novel renal protective drugs for the treatment of DKD.OBJECTIVE:The present study is to confirm our hypothesis and to accumulate the information for the application of DMDD (2-Dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione) as a novel therapeutic agent to potentially inhibit renal fibrogenesis and EMT in the DKD.METHODS:High glucose induced renal proximal tubular epithelial cell line (HK-2 cells) was cultured and treated with DMDD. The cell viability and DMDD cytotoxicity were assessed by CCK8. Immunofluorescence was used for detection of TLR4 and downstream protein in normal and high glucose induced HK-2 cells. HK-2 cells were transfected with lentivirus codifying for BAMBI (BMP and activin membrane bound inhibitor) and interfering RNA for determination of the effect of BAMBI over-expression and silencing, respectively. TLR4-BAMBI-Smad2/3 pathway was analyzed by means of RT-PCR and western blot.RESULTS:A high concentration (60mM) of glucose induced significant EMT process and TLR4 expression was increased obviously in this circumstance. DMDD inhibited high expressions of TLR4 and Smad2/3 in HG induced cells and decreased the expression of BAMBI. In addition, the effects of decreased BAMBI expression and increased Smad2/3 expression in HG cultured cells were reversed in the cells of TAK-242 (TLR4 signaling inhibitor) intervention. BAMBI gene silencing dramatically increased EMT process and the over-expression of BAMBI was opposite in HK-2 cells with HG condition. These observations of EMT were ameliorated when the HK-2 cells were pre-treated with DMDD.CONCLUSIONS:Our study demonstrates that DMDD treatment improves EMT in the HG induced HK-2 cells. In addition, DMDD significantly inhibits EMT by TLR4-BAMBI-Smad2/3 pathway, which hints that DMDD may be an alternative approach in diabetic renal injury.

目的:通过建立四氯化碳(CCl4)急性肝损伤模型筛选出柿树寄生植物具有保肝作用的活性部分,并探讨其对肝损伤小鼠肝细胞的保护作用.方法:(1)制备柿树寄生植物石油醚萃取物、乙酸乙酯萃取物、正丁醇萃取物,建立CCl4急性肝损伤模型,通过检测各萃取物处理组小鼠血清谷丙转氨酶(ALT)、谷草转氨酶(AST)水平及肝组织丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性,筛选柿树寄生植物的活性部分.(2)将60只昆明小鼠随机分为正常组、模型组、联苯双酯(150 mg/kg)组及正丁醇萃取物低剂量(11.67 mg/kg)组、中剂量(23.34 mg/kg)组、高剂量组(46.67 mg/kg)组,灌胃给药8d后,除正常组外,其他各组均建立急性肝损伤模型,检测各组小鼠血清白细胞介素(IL)-6和IL-1β水平,并采用免疫组织化学染色法检测肝组织IL-6蛋白表达量.结果:柿树寄生植物各萃取物处理组小鼠肝脏、脾脏、胸腺系数比较,差异均无统计学意义(均P＞0.05);与模型组比较,正丁醇萃取物组肝组织MDA含量及血清AST、ALT水平明显降低,肝组织SOD活性明显提高(P＜0.05),而石油醚萃取物组和乙酸乙酯萃取物组各指标比较,差异均无统计学意义(均P＞0.05).正丁醇萃取物中、高剂量组小鼠血清IL6水平及肝组织IL-6蛋白表达量均较模型组显著降低,正丁醇萃取物各剂量组血清IL-1β水平均显著降低(P＜0.05),且中、高剂量组IL-6水平低于低剂量组(P＜0.05),而IL-1β水平在各剂量组之间无明显差异(P＞0.05).结论:柿树寄生植物正丁醇萃取物是柿树寄生植物保肝作用的活性部分,能明显减轻CCl4所致的肝损伤,其机制与抑制炎症因子表达,提高机体抗氧化能力,降低氧化应激有关.

BACKGROUND:Averrhoa carambola L. is a traditional medicinal herb that has long been used to treat diabetes. Our previous studies found that 2-dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione (DMDD) isolated from A. carambola L. roots could ameliorate diabetic nephropathy (DN), but its exact mechanism remains unclear.METHODS:A DN model was established by streptozotocin (STZ, 100 mg/kg body weight) in TLR4 knockout (TLR4-/-, KO) mice and wild-type (WT) mice. Body weight and blood glucose were evaluated after oral administration of DMDD (12.5, 25, 50 mg/kg body weight/d) in diabetic mice. The levels of serum lipids, including TC, TG, HDL, and LDL and kidney function indexes Scr and BUN, were detected by biochemical equipment. The levels of inflammatory cytokines including IL-6 and TNF-α, were determined by ELISA kits. Furthermore, changes in renal ultrastructure were observed by electron microscopy. Western blot analysis and RT-PCR were used to assess the protein expression and mRNA levels of TLR4, MyD88 and NF-κB.RESULTS:DMDD treatment attenuated diabetic nephropathy, as a result of a decline in blood glucose, serum creatinine, and blood urine nitrogen levels and an increase in the quantity and density of podocytes, combined with improved dyslipidaemia. DMDD treatment inhibited the inflammatory response and downregulated the expression of the TLR4/MyD88/NF-κB pathway in diabetic mice, and these changes were significantly different in TLR4-/- mice.CONCLUSION:DMDD alleviates diabetic nephropathy by mitigating kidney damage and inflammation via the inhibition of the TLR4/MyD88/NF-κB signalling pathway.