Celastrol protects kidney against ischemia–reperfusion-induced injury in rats

Background

Ischemia–reperfusion (IR) causes various damages in renal tissues, which is exacerbated by hypoxia-induced excessive inflammation and deteriorates the prognosis of patients after kidney surgery. Celastrol is a potent inflammation inhibitor that has little toxicity. In this report, we investigated whether celastrol protects against IR-induced renal injury in rats.

Materials and methods

Renal IR injury was induced by occlusion of the bilateral renal pedicles for 45 min followed by reperfusion for 6 h. Celastrol or vehicle solution was intraperitoneally injected 30 min before renal ischemia, respectively. Renal histology, function, and pro-inflammatory cytokines and mediators were assessed. The effect of celastrol on nuclear translocation of nuclear factor kappa B (NF-κB) was also measured.

Results

Celastrol significantly suppressed elevation of the renal function markers and the lipid peroxidation level, alleviated renal tubular damage, and decreased the levels of tumor necrosis factor-α, interleukin-1β, and monocyte chemotactic protein-1 (MCP-1) messenger RNA in kidney caused by IR. Moreover, celastrol prevented IR-induced expression of pro-inflammatory mediators, which was associated with suppression of nuclear translocation of NF-κB subunit p65.

Conclusions

Celastrol ameliorated the acute kidney injury caused by IR, which was associated with inhibiting local NF-κB activation and inflammation. Our findings suggest that celastrol could be useful for preventing IR-induced renal injury.     

南蛇藤醇抑制宫颈癌细胞生物学行为的机制研究

目的　探讨南蛇藤醇抑制宫颈癌细胞增殖和侵袭的作用机制。 方法　将正常培养HeLa细胞分为对照组、南蛇藤醇低、中、高剂量组（终浓度分别为4 μmol/L、8 μmol/L和12 μmol/L）、阳性对照组（顺铂,终浓度为10 μmol/L）、LY294002组（LY294002,终浓度为20 μmol/L）和（南蛇藤醇+LY294002）组（南蛇藤醇和LY294002,终浓度分别为12 μmol/L和20 μmol/L）。MTT法检测各组细胞存活情况;流式细胞仪检测细胞凋亡情况;Transwell检测细胞的侵袭;蛋白质印迹法检测PI3K/AKT/NF-κB信号通路的表达;采用皮下注射HeLa细胞建立裸鼠模型,观察南蛇藤醇（120 mg/kg）对裸鼠移植瘤体积和重量的影响。 结果　与对照组相比,南蛇藤醇组、阳性对照组和LY294002组HeLa细胞的相对增殖率、侵袭细胞数均明显下降（P<0.001）,细胞凋亡率明显升高（P<0.001）,p-PI3K、p-AKT、NF-κB p65蛋白表达明显下调（P<0.001）,且随着南蛇藤醇浓度升高,其作用增强;与LY294002组相比,（南蛇藤醇+LY294002）组HeLa细胞的相对增殖率和侵袭细胞数明显下降（q=10.182,q=10.217,P均<0.001）,细胞凋亡率明显升高（q=23.636,P<0.001）;与对照组相比,南蛇藤醇组裸鼠体重、移植瘤的体积和重量明显下降（q=4.100,P<0.020;q=13.501,P<0.001;q=5.078,P=0.005）。 结论　南蛇藤醇可能通过抑制HeLa细胞的PI3K/Akt/NF-κB信号通路,抑制其恶性生物学行为。     

雷公藤红素通过抑制MMP-9/sFlt-1/TNF-α减轻子痫前期模型大鼠症状的作用研究

目的:探讨雷公藤红素通过抑制金属基质酶-9/可溶性FMS样酪氨酸激酶-1/肿瘤坏死因子α(MMP-9/sFlt-1/TNF-α)减轻子痫前期模型大鼠症状的作用机制。方法:72只怀孕未经产的雌性SD大鼠随机分为6组:正常对照组、模型组、硫酸镁组(30mg·kg^-1)、雷公藤红素低、中、高剂量组(10,20,40 mg·kg^-1),每组12只。除正常对照组外,其余各组采用N-硝基-L-精氨酸甲酯盐酸盐诱导子痫前期模型,建模成功后各给药组灌胃给予相应药物,正常对照组和模型组给予等体积生理盐水。于妊娠期(第9天)测定大鼠24 h尿蛋白、血压、尿素氮(BUN),实时荧光逆转录法(RT-PCR)及蛋白质免疫印迹法(West blot)测定胎盘组织MMP-9 mRNA和蛋白水平,酶联免疫吸附法(ELISA)法测定sFlt-1、白细胞介素-4(IL-4)、TNF-α蛋白表达水平。结果:与模型组比较,硫酸镁组、雷公藤红素各剂量组收缩压、舒张压、24 h尿蛋白、BUN水平、MMP-9 mRNA和蛋白、sFlt-1、IL-4、TNF-α蛋白表达水平明显降低(P<0.05),且雷公藤红素各组呈剂量相关性(P<0.05);与硫酸镁组相比,雷公藤红素低、中剂量组以上各指标均明显升高(P<0.05)。结论:雷公藤红素能明显降低子痫前期模型大鼠血压、尿蛋白、BUN水平,缓解子痫前期症状,其机制可能与雷公藤红素通过抑制胎盘组织中MMP-9 mRNA和蛋白表达水平进而降低sFlt-1、IL-4、TNF-α蛋白表达有关。     

雷公藤单体体外抑制胶质瘤细胞的实验研究

背景与目的：已有研究发现雷公藤具有抗肿瘤作用,本研究旨在探讨雷公藤单体对胶质瘤细胞的体外抑制作用。方法：通过MTT法测定3种雷公藤单体（甲素,红素和WilforolA)对胶质瘤细胞株SHG44,C6,U251的体外抑制作用;应用免疫组化法观察雷公藤甲素与雷公藤红素对SHG44胶质瘤细胞中Bax,Bcl-2蛋白表达的影响。结果：雷公藤二萜类单体雷公藤甲素对胶质瘤细胞有极显著的抑制作用;雷公藤三萜类单体中红素的抑制作用次之,两者均使SHG44细胞Bax表达增加,Bcl-2表达下降。结论：雷公藤甲素与雷公藤红素对胶质瘤细胞有明显的抗肿瘤作用,其作用与促进Bax表达,抑制Bcl-2表达,导致细胞凋亡有关。     

雷公藤红素抑制MGC - 823细胞增殖和迁移及诱导凋亡

目的 探索雷公藤红素对MGC - 823细胞的增殖,迁移及诱导凋亡的作用,并探讨其作用机制。方法 0、1、2、4、6、8、12 μM雷公藤红素作用于MGC - 823细胞,采用MTT法检测增殖比率。0、1、2和4 μM雷公藤红素作用于MGC - 823细胞0、24、48和72 h,显微镜下观察细胞形态,采用细胞划痕实验检测细胞迁移能力,流式细胞术检测细胞凋亡,western blot法检测细胞中凋亡相关蛋白Bcl - 2、Bax、Akt、p - Akt等的表达水平。结果 经雷公藤红素作用后,MGC - 823细胞的增殖率均下降（P＜0.05）;细胞数量减少,体积缩小,皱缩变圆,部分不再贴壁,细胞核缩小;细胞迁移率减小（P＜0.05）;早期凋亡率增加（P＜0.05）;MGC - 823细胞中Bax表达升高,Bcl - 2和p - Akt表达降低（P＜0.05）。结论 雷公藤红素对MGC - 823细胞的增殖和迁移有抑制作用,并诱导其凋亡。     

雷公藤红素对裸鼠肝癌皮下移植瘤抑制作用的研究

目的 研究天然活性化合物雷公藤红素（celastrol）对裸鼠移植肿瘤的抑制作用。方法 建立人源肝癌HepG2细胞的裸鼠皮下移植肿瘤模型。20只裸鼠随机分为4组,即空白对照组和雷公藤红素高剂量组、中剂量组和低剂量组,每组分别按4.0 mg/kg、2.0 mg/kg、1.0 mg/kg雷公藤红素腹腔给药,每日一次,每周5 d,共给药30 d,实验中每5 d测量一次肿瘤体积的变化。结果 雷公藤红素高剂量组、中剂量组和低剂量组的体积抑瘤率分别是89.52%、65.60%和12.21%,与空白对照组相比,高剂量组、中剂量组的差异具有统计学意义（P〈0.05）。高剂量雷公藤红素给药后对裸鼠肝脏和肾脏等产生严重的毒性反应。结论 雷公藤红素对裸鼠肝癌皮下移植瘤具有良好的体内抗肿瘤效果,应进一步开展药物新剂型及分子结构优化方面的研究,减少其毒副反应。     

雷公藤红素对C6胶质瘤细胞凋亡及细胞周期阻滞的影响

目的 探讨雷公藤红素对体外C6胶质瘤细胞凋亡及细胞周期阻滞的影响及机制.方法 雷公藤红素处理体外培养的C6胶质瘤细胞,MTT法检测细胞增殖,Hochest33342染色荧光显微镜观察以及锇铀铅染色透射电镜观察胶质瘤细胞形态变化,流式细胞术分析细胞凋亡率及细胞周期改变.蛋白印迹分析检测凋亡相关蛋白及细胞周期调控蛋白的表达变化.结果 雷公藤红素对C6胶质瘤细胞的增殖抑制作用呈浓度和时间依赖性;2.56 μmol/L雷公藤红素处理C6胶质瘤细胞24h后,荧光显微镜、透射电镜下均可见凋亡形态学改变.流式细胞术检查显示,凋亡细胞比例由1.32％升高到19.34％;并且,处于G0/G1期的细胞比例由76.42％降低到54.52％,G2/M期细胞比例由9.29％升高到30.50％.蛋白印迹分析显示,雷公藤红素降低了Bcl -2及XIAP蛋白表达,促进了Bax及Caspase -3的表达以及PARP的剪切;雷公藤红素在诱导细胞周期调控蛋白p21、p27及cyclin B1蛋白表达增加的同时抑制了cdk2蛋白的表达.结论 雷公藤红素能够通过调节凋亡相关蛋白及细胞周期相关因子的表达影响C6胶质瘤细胞的凋亡及细胞周期阻滞.     

The NF-κB inhibitor celastrol attenuates acute hepatic dysfunction induced by cecal ligation and puncture in rats

Acute hepatic dysfunction associating sepsis is mediated mainly by toll-like receptor-4 (TLR-4)/nuclear factor kappa‐B (NF-κB) inflammatory pathway. This study explores potential hepatoprotective effect of the NF-κB inhibitor celastrol in cecal ligation and puncture (CLP) model in rats.Protective effect of celastrol (1 mg/kg, i.p., 1 h before CLP) was illustrated after 24 h by preventing CLP‐induced hepatic histopathological changes and elevation in serum hepatic biomarkers [alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TB) and gamma aminotransferase (γ-GT)] without affecting mortality. Celastrol anti‐inflammatory effect was illustrated by inhibiting increased serum and hepatic mRNA expression of interleukin-6 (IL-6) without affecting IL-10 elevation. Furthermore, celastrol inhibited CLP-induced elevations in hepatic mRNA expression of nuclear factor inhibitory protein kappa-B alpha (NFκBia), TLR-4, 5‐lipoxygenase (5-LOX) and prevented NF‐κB/p65 nuclear translocation and activation.In conclusion, celastrol prevented CLP-induced acute hepatic dysfunction through its anti-inflammatory effect by attenuating NF-κB activation, TLR-4 and 5-LOX expression with subsequent reduction in pro-inflammatory IL-6.     

The MVA pathway genes expressions and accumulation of celastrol in Tripterygium wilfordii suspension cells in response to methyl jasmonate treatment

Celastrol is an important bioactive triterpenoid in traditional Chinese medicinal plant, Tripterygium wilfordii. Methyl Jasmonate (MJ) is a common plant hormone which can regulate the secondary metabolism in higher plants. In this study, the mevalonate (MVA) pathway genes in T. wilfordii were firstly cloned. The suspension cells of T. wilfordii were elicited by MJ, and the expressions of MVA pathway genes were all enhanced in different levels ranging from 2.13 to 22.33 times of that at 0 h. The expressions were also enhanced compared with the CK group separately. The accumulation of celastrol in the suspension cells after the treatment was quantified and co-analyzed with the genes expression levels. The production of celastrol was significantly increased to 0.742 mg g^?1 after MJ treatment in 288 h which is consistent with the genes expressions. The results provide plenty of gene information for the biosynthesis of terpenoids in T. wilfordii and a viable way to improve the accumulation of celastrol in T. wilfordii suspension cells.