Antitumor effects and compatibility mechanism of Hedyotis Diffusa-Sculellaria Barbata herb pair

OBJECTIVE Hedyotis diffusa Willd and Scutellaria barbata D Don are most commonly used herb pair for cancer treatment in traditional Chinese medicine.This study aimed to evaluated the in vitro and in vivo antitumor effects and the compatibility mechanisms of Hedyotis diffusa-Sculellaria barbata herb pair.METHODS Hedyotic diffusa extract,Scutellariae Barbatae extract,and herb pair extract were prepared and the component were detected by UPLC-Q-TOF-MS.The effects of different concentrations of Hedyotic diffusa extract,Scutellariae Barbatae extract,and herb pair extract were detected using MTS assay.The in vivo antitumor effects were evaluated in Panc28 pancreatic cells bearing nude mice model.The compatibility mechanism of Hedyotis diffusa-Sculellaria barbata herb pair were evaluated based on the network pharmacology,using TCMSP,Cytoscape 3.6.1 software,and DAVID 6.8.The anticancer mechanism were further validated in vitro.RESULTS Both the MTS and in vivo results showed that herb pair extract showed more obvious inhibitory effects on cancer cells compared to each individual.A total of 37 active components were selected from Hedyotis diffusa and Sculellaria barbata,33 kinds of active ingredients are involved in their anti-tumor effects.58 cancer-related targets and 66 KEGG pathways were identified.The potential targets for the herb pair might be prostaglandin G/H synthase 2(PTGS2),HSP90,EGFR,72 ku typeⅣcollagenase,PPAR-γ,et al.In vitro validation result showed that compatibility mech⁃anisms was related with HSP90,EGFR related pathways.CONCLUSION The result of the study preliminarily verified the basic anti-tumor pharmacological effects of Hedyotis diffusa-Sculellaria barbata herb pair,and lays a solid foundation for further studies on the anti-tumor mechanism of the herbal pair.

Effect of Polysaccharides from Hedyotis diffusa on Immune Cells of Small Intestinal Mucosa of Chicks

[Objectives]This study aimed to investigate the effect of polysaccharides from Hedyotis diffusa on the immune cells of the intestinal mucosa of chicks.[Methods]Total 200 seven-day-old AA chicks were evenly and randomly divided into four groups:control group and three treatment groups.The chicks in the control group(group I)were fed basal diet,and those in the treatment groups were fed with basal diet supplemented with 0.4(group II),0.8(group III)and 1.2(group IV)g/kg H.diffusa polysaccharides(OPS),respectively.At the age of 14,28 and 42 d,10 chicks from each of the groups were sacrificed,respectively for counting of intraepithelial lymphocytes(IELs)and lamina propria secretory IgA(SIgA)positive cells in small intestinal mucosa.[Results]At the age of 14 d,the duodenal and jejunal IEL and SIgA positive cell counts in groups II,III and IV were significantly greater than those in group I(P<0.05).At the age of 28 d,the jejunal IEL count of group IV was significantly lower than those of groups I,II and III(P<0.05);and there was no significant difference in IEL or SIgA positive cell count between group I and the other groups(P>0.05).At the age of 42 d,the duodenal and jejunal IEL counts in group II were significantly greater than those in groups I,III and IV(P<0.05),while the differences in duodenal and jejunal IEL count between groups III,IV and I were insignificant(P>0.05).The duodenal SIgA positive cell counts in the treatment groups were insignificantly different from that in group I(P>0.05).The jejunal SIgA positive cell count in group II was significantly greater than those in groups I,III and IV(P<0.05).There was no significant difference in jejunal SIgA positive cell count between groups III and IV and group I(P>0.05).[Conclusions]OPS at 0.4 g/kg can promote the proliferation of immune cells in the small intestine mucosa of 14-42-day-old AA chicks,and it is recommended to be added to the feed.

Inhibition of vascular endothelial growth factor expression by Chinese medicine of Hedyotis diffusa Willd herbal compounds

The Hedyotis diffusa Willd herbal compounds(HDWHCs)are commonly used as Chinese medicine to treat cancer patients with established clinical therapeutic efficacy in China.However,the underlying mechanisms remain to be elucidated.In this study,we used freeze-dried powder of the water extracts of HDWHCs to investigate the potential mechanisms of HDWHCs in cancer treatment.HDWHCs treatment significantly inhibited vascular endothelial growth factor(VEGF)mRNA levels and VEGF transcriptional activation in cancer cells.HDWHCs also had a remarkable inhibitory effect on the expression of hypoxia-inducible factor 1alpha(HIF-1alpha).Forced expression of HIF-1αrestored VEGF transcriptional activation inhibited by HDWHCs,indicating that HDWHCs suppressed VEGF expression through decreasing HIF-1alpha expression.Moreover,HDWHCs inhibited cyclooxygenase-2(COX-2)expression,and overexpression of HIF-1alpha restored HDWHCs’inhibitory effect on COX-2 at transcriptional level.These findings may provide better understanding of HDWHCs’anti-cancer mechanism in cancer treatment.

Ethyl Acetate Fraction in Hedyotis Diffusa Willd Inhibits T Cell Proliferation to Improve the Pathogenesis of Systemic Lupus Erythematosus

Background:Abnormal proliferation of T cells plays an essential role in the pathogenesis of Systemic lupus erythematosus(SLE).The pharmaceutical effect of Hedyotis Diffusa Willd(HDW)on SLE has been investigated previously.Nevertheless,the biomedical mechanism is still left unclear.Objective:This study has been arranged to evaluate the therapeutic effect of the ethyl acetate fraction of HDW(EAHDW)on lupus mice and explore the potential therapeutic mechanism.Methods:EAHDW was prepared with 80%ethanol reflex extraction followed by successive extraction,and ana-lyzed with HPLC and UPLC-Q/TOP-MS.The potential targets and STAT3 affinity regulators were predicted with network pharmacology.The pharmaceutic effect of EAHDW was studied with MRL/lpr mice.Cytokines and au-toantibodies were quantified with ELISA assays.The pathological damage of glomerulus and STAT3 expression in the kidney was detected with histochemical and immunohistochemical techniques.The cell cycle properties in cell proliferation were identified with the flow cytometry.The western blot and dual-Luciferase reporter assay were applied to evaluate translational and transcriptional activity of STAT3,respectively.Results:In this study,the extraction ratio of EAHDW was 2.7±1%,in which 19 ingredients were identified.Network pharmacological analysis showed that the target genes of EAHDW were highly focused on influencing the abnormal T cell proliferation in SLE.EAHDW showed the beneficial effects on pathological changes and STAT3 expression in the glomerulus of lupus mice,and the levels of cytokines and autoantibodies in serum.In cytological study,EAHDW treatment attenuated the transcription and phosphorylation of STAT3,which inhibited T cell proliferation by prolonged S-phase of the cell cycle.A total of 5 compounds in EAHDW exhibited high docking affinity to the DNA-binding site of STAT3.Conclusion:EAHDW could reduce the inflammatory response and inhibit the proliferation of T cells by interfering with the STAT3 signaling pathway,thereby playing a therapeutic effect on SLE.

乙醇-磷酸氢二钾双水相体系萃取白花蛇舌草总黄酮的工艺研究

利用乙醇-磷酸氢二钾双水相体系对白花蛇舌草(Hedyotis diffusa)总黄酮进行萃取分离。通过单因素实验分别考察了m(K2HPO4)、超声时间及超声温度对白花蛇舌草总黄酮萃取效果的影响,并用响应面法优化了萃取工艺。经优化后,白花蛇舌草总黄酮的最佳萃取条件为:m(K2HPO4)2.0g,超声时间37min,超声温度57℃。在此条件下,白花蛇舌草总黄酮的萃取率可高达99.45%。本方法精密度良好,准确度较高,可为白花蛇舌草总黄酮的高效萃取提供参考。

白花蛇舌草提取物诱导人胃癌MKN-45细胞凋亡

为探究白花蛇舌草提取物对人胃癌MKN-45细胞凋亡的影响。用不同浓度的白花蛇舌草提取物处理MKN-45细胞,然后在光学显微镜和激光共聚焦显微镜下观察数量和形态。流式细胞仪检测细胞凋亡率,RT-qPCR和Western Blot检测凋亡相关基因Bax和Bcl-2的m RNA和蛋白表达水平。结果表明,不同浓度的白花蛇舌草提取物处理48 h后,在显微镜下观察到细胞体积缩小、细胞核裂解和染色质形态变化。细胞凋亡率随着药物浓度的增加而增加,用30μg/mL白花蛇舌草提取物处理后细胞凋亡率达到23.6%,Bax基因表达水平显著增加,Bcl-2基因表达水平显著降低。综上所述,白花蛇舌草提取物可诱导人胃癌MKN-45细胞凋亡,具有潜在的医学和药用价值。

白花蛇舌草对人胃癌SGC-7901细胞增殖的影响

本研究以不同浓度的白花蛇舌草注射液处理人胃癌SGC-7901细胞,探究其对细胞增殖的影响。利用台盼蓝拒染法和吉姆萨染色法分别检测不同浓度的白花蛇舌草注射液对人胃癌SGC-7901细胞存活率和细胞克隆形成率的影响;采用EdU法检测药物处理后SGC-7901的细胞增殖率;最后应用RT-qPCR和Western blot分别检测药物处理后对细胞周期相关蛋白CCNB1和CCND1在mRNA水平和蛋白水平表达的影响。实验结果表明:白花蛇舌草注射液可抑制人胃癌细胞SGC-7901的细胞活力,其细胞存活率随着药物浓度的升高而降低;不同浓度的白花蛇舌草注射液处理人胃癌SGC-7901细胞48 h后,其细胞克隆形成率和细胞增殖率显著降低;RT-qPCR和Western blot结果显示,CCNB1和CCND1基因的mRNA及蛋白表达水平随着药物浓度的升高而显著降低。综上所述,白花蛇舌草通过下调CCNB1和CCND1基因的表达抑制人胃癌SGC-7901细胞增殖。