Triptolide promotes generation of FoxP3+ T regulatory cells in rats

Ethnopharmacological relevance

Triptolide (TPT), a component of the Chinese herb Triptergium wilfordii, has potent immunosuppressive and anti-inflammatory activity and is used clinically in recipients of kidney transplantation.

Aim of the study

This work aimed to investigate the effect of TPT on the differentiation of regulatory T lymphocytes (Tregs) from CD4+ cells in rats.

Materials and methods

MACS-purified rat CD4+ cells were costimulated with anti-CD3 and anti-CD28 in the presence of TGF-β to induce the expression of FoxP3, which was detected by flow cytometry. TPT and cyclosporine A (CsA) were separately added into the cultures to observe the effect on the expression of FoxP3. Kidney transplantation was performed in rats that either received no treatment or were treated with TPT after transplantation.

Results

TPT treatment enhanced the expression of FoxP3 in CD4+ cells, whereas CsA inhibited the FoxP3 expression. In the rat kidney transplantation model, the recipient rats treated with TPT survived longer than the control rats (18–19.83 vs 6.83 days, P < 0.05). Meanwhile, the FoxP3+ T cells in the spleens of treated rats were higher than those from the untreated rats (12.4% vs 4.7%, P < 0.05).

Conclusions

These data suggest that TPT may promote the differentiation of CD4+ cells to FoxP3+ Tregs. This would be at least one of the pathways responsible for the immunosuppressive activity of TPT.     

MTT法检测雷公藤内酯醇对晶状体上皮细胞增殖的影晌

目的:研究雷公藤内酯醇对体外培养的牛晶状体上皮细胞增殖的抑制作用。方法:取第4代对数生长期的牛晶状体上皮细胞培养24h后,加入重组人表皮生长因子(终浓度50μg/L)和不同浓度的雷公藤内酯醇(40,20,10μg/L),分别作用6,12,24,48,72h后,采用MTT法观察雷公藤内酯醇对牛晶状体上皮细胞增殖的影响。结果:不同浓度的雷公藤内酯醇,于不同时间作用在处于增殖状态的LECs,其增殖抑制率升高呈现明显的时间及剂量依赖性。结论:雷公藤内酯醇对晶状体上皮细胞增殖有明显的抑制作用在防止后囊混浊方面有广阔的前景。     

Triptolide inhibits IL-12 production and T cell-stimulatory capacity of dendritic cells

Extracts of Tripterygium wilfordii Hook F.(TWHF ) are effective in traditional Chinesemedicine for treatment of autoimmune diseasessuch as rheumatoid arthritis, systemic lupus ery thematosus, nephritis and asthma [1, 2]. Trip tolide, a diterpenoid triepoxide, is derived fromTWHF and is responsible for most of the immuno suppressive and anti inflammatory effects ofTWHF. Triptolide has been shown to be effectivein the treatment of autoimmune diseases, …     

雷公藤甲素通过抑制破骨细胞生成预防骨丢失的研究

目的 探讨雷公藤甲素抗骨质疏松的作用及机制。方法 建立大鼠老年性骨质疏松模型。40只22月龄雄性SD大鼠随机分为雷公藤甲素（每天15μg/kg腹腔注射）治疗组和生理盐水对照组（每天15μg/kg腹腔注射）,连续8周。采用显微CT分析胫骨近端骨松质的骨密度(BMD)和骨显微结构。WB检测成骨相关蛋白表达水平。TRACP-5b染色法测定破骨细胞数,同时检测骨吸收标志物表达水平。结果 显微CT结果显示,雷公藤甲素治疗组大鼠骨密度、骨体积/总体积比值(Bv/Tv)、骨小梁厚度(Tb.Th)、骨小梁数目(Tb.N)、骨小梁间距(Tb.Sp)均显著高于对照组（P<0.05）。两组的成骨相关蛋白表达水平无明显差异,TRACP-5b染色显示雷公藤甲素减少了体内破骨细胞的数量（P<0.05）,同时血液骨吸收标志物水平也明显降低（P<0.05）。结论 雷公藤甲素通过抑制破骨细胞生成进而对老年性骨质疏松有保护作用。雷公藤甲素可能是治疗老年性骨质疏松症的一种可行方案。     

雷公藤内酯醇抗人肝癌SMMC-7721细胞活性研究

目的: 评价雷公藤内酯醇抗人肝癌SMMC-7721细胞的作用.方法: 采用体外细胞培养和SD大鼠体内肿瘤接种两种衡量方法,体外细胞培养检测指标包括细胞形态观察,台盼蓝染色,DAPI染色,DNA ladder;SD大鼠体内接种肿瘤指标检测用游标卡尺测量肿瘤体积大小.结果: 与对照组相比,体外培养肿瘤细胞在用药后活细胞大幅减少,体内接种肿瘤细胞在用药后,肿瘤块体积较小且增长缓慢,而对照组肿瘤体积却大幅增长.结论: 雷公藤内酯醇对体外和体内培养的人肝癌SMMC-7721细胞有明显的生长抑制作用和促凋亡作用.     

雷公藤内酯醇诱导胰腺癌细胞凋亡及对5-脂氧合酶和凋亡基因表达的影响

目的探讨雷公藤内酯醇(TL)在胰腺癌化学预防和治疗中的应用价值。方法采用人胰腺癌细胞株SW1990作为研究对象,MTT法和吖啶橙(AO)染色分别检测SW1990细胞的增殖和凋亡,采用半定量RT-PCR法检测5-脂氧合酶(5-LOX)mRNA、bcl-2 mRNA和bax mRNA的表达。结果10μg/ml的TL对SW1990细胞生长的抑制率达30%以上,0.1μg/mlTL处理24h后,SW1990细胞凋亡指数达38.5%,显著高于对照组的14.97%。5-LOX、bcl-2、bax mRNA表达从0.7160±0.0251,0.2446±0.0311和0.0260±0.0065分别变化为0.2400±0.0316,0.0700±0.0158和0.0700±0.0158。结论TL能抑制胰腺癌细胞增殖和诱导细胞凋亡,其凋亡机制可能与下调5-LOXmRNA和bcl-2mRNA、上调bax mRNA基因表达有关。     

Apoptosis of human pancreatic cancer cells induced by Triptolide

AIM： To investigate apoptosis in human pancreatic cancer cells induced by Triptolide （TL）, and the relationship between this apoptosis and expression of caspase-3＇ bcl-2 and bax. METHODS： Human pancreatic cancer cell line SW1990 was cultured in DMEM media for this study. MTT assay was used to determine the cell growth inhibitory rate in vitro. Flow cytometry and TUNEL assay were used to detect the apoptosis of human pancreatic cancer cells before and after TL treatment. RT-PCR was used to detect the expression of apoptosis-associated gene caspase-3＇ bcl-2 and bax. RESULTS： TL inhibited the growth of human pancreatic cancer cells in a dose-and time-dependent manner. TL induced human pancreatic cancer cells to undergo apoptosis with typically apoptotic characteristics. TUNEL assay showed that after the treatment of human pancreatic cancer cells with 40 ng/mL TL for 12 h and 24 h, the apoptotic rates of human pancreatic cancer cells increased significantly. RT-PCR demonstrated that caspase-3 and bax were significantly up-regulated in SW1990 cells treated with TL while bcl-2 mRNA was not. CONCLUSION： TL is able to induce the apoptosis in human pancreatic cancer cells. This apoptosis may be mediated by up-regulating the expression of apoptosisassociated caspase-3 and bax gene.     

Renal targeted delivery of triptolide by conjugation to the fragment peptide of human serum albumin

We have previously demonstrated that peptide fragments (PFs) of the human serum albumin could be developed as potential renal targeting carriers, in particular, the peptide fragment, PF-A_299–585 (A_299–585 representing the amino acid sequence of the human serum albumin). In this paper, we conjugated triptolide (TP), the anti-inflammatory Chinese traditional medicine, to PF-A_299–585 via a succinic acid spacer to give TPS-PF-A_299–585 (TP loading 2.2% w/w). Compared with the free TP, TPS-PF-A_299–585 exhibited comparable anti-inflammatory activity in the lipopolysaccharide stimulated MDCK cells, but was significantly less cytotoxic than the free drug. Accumulation of TPS-PF-A_299–585 in the MDCK cells in vitro and in rodent kidneys in vivo was demonstrated using FITC-labeled TPS-PF-A_299–585. Renal targeting was confirmed in vivo in a membranous nephropathic (MN) rodent model, where optical imaging and analyses of biochemical markers were combined to show that TPS-PF-A_299–585 was capable of alleviating the characteristic symptoms of MN. The collective data affirm PF-A_299–585 to be a useful carrier for targeting TP to the kidney.     

雷公藤内酯醇对IgA肾病大鼠蛋白尿和nephrin、podocin蛋白及mRNA表达的影响

目的:观察雷公藤内酯醇(TP)对IgA肾病(IgAN)大鼠尿蛋白和足细胞nephrin、podocin蛋白及mRNA表达的影响。方法:采用牛血清白蛋白+四氯化碳+脂多糖的方法建立IgAN大鼠模型。将60只大鼠随机分为正常组、模型组、洛丁新组、TP剂量组、TP中剂量组、TP高剂量组,每组10只。于0,10,14周收集血尿标本,并处死大鼠留取肾组织标本。检测24 h尿蛋白定量,血生化指标,分别采用实时定量PCR及ELISA法检测肾组织nephrin、podocin mRNA和蛋白表达。结果:(1)实验前各组大鼠尿蛋白差异无统计学意义(P>0.05),第10周末各造模组蛋白尿均明显高于正常组(P<0.01),第14周末TP各剂量组及洛丁新组蛋白尿均较模型组明显减少(P<0.01),TP高剂量组Alt值高于其余各组(P<0.05);(2)与正常组相比,模型组nephrin、podocin蛋白、mRNA表达明显降低(P<0.01);TP各剂量组及洛丁新组nephrin、podocin蛋白、mRNA表达均较模型组明显增高(P<0.05);TP中、高剂量组较洛丁新组nephrin、podocin蛋白浓度显著增高(P<0.05);TP各剂量组与洛丁新组及TP各剂量组间nephrin、podocin mRNA表达均差异无统计学意义(P>0.05)。结论:TP能够明显减轻IgAN大鼠的蛋白尿,且能明显提高肾组织nephrin、podocin mRNA及蛋白的表达。提示TP能调节IgAN足细胞相关分子nephrin、podocin基因及蛋白的表达,减少足细胞损害,减少尿蛋白。     

雷公藤内酯醇对阿尔茨海默病模型大鼠海马iNOS表达和突触超微结构的影响

目的观察应用β淀粉样蛋白(Aβ)后大鼠海马诱导型一氧化氮合酶(i NOS)的表达和突触超微结构的变化及雷公藤内酯醇对其的影响。方法 21只SD雄性大鼠等分成对照组、模型组、用药组。在大鼠左侧海马定向注射凝聚态Aβ1⁴0作为模型组,注射等量生理盐水设为对照组,大鼠在海马注射凝聚态Aβ140作为模型组,注射等量生理盐水设为对照组,大鼠在海马注射凝聚态Aβ1⁴0后腹腔注射雷公藤内酯醇为用药组。应用免疫组织化学方法检测各组大鼠海马i NOS的表达;应用透射电镜观察各组大鼠海马突触超微结构的变化。结果免疫组织化学染色显示,模型组大鼠海马i NOS阳性细胞数和平均光密度明显高于对照组(P<0.01),用药组大鼠海马i NOS阳性细胞平均光密度较模型组明显降低(P<0.05)。透射电镜结果显示,模型组大鼠海马神经毡内突触和突触小泡数量较对照组减少,突触后致密物厚度变薄;用药组大鼠海马神经毡内突触和突触小泡数量较模型组增多,突触后致密物增厚。结论雷公藤内酯醇可以抑制Aβ诱导的大鼠海马i NOS的表达,对突触超微结构的损伤有一定的改善作用。