医学实验方法与技术课程培养中西医结合研究生创新能力的实践

研究生教育的理论与实践证实,创新型研究型人才的培养是研究生教育的主要目标。从医学生本科阶段到研究生阶段的过渡面临的一个主要变化是医学实验技术的熟悉和掌握。中西医结合研究生更是面临理论与实践有机结合、中西方医学并重的科研挑战。在研究生学习的初始阶段,最先面对的就是科研实验的学习和科研思维的转变。因此,医学实验方法与技术的教学模式的优化是提高中西医结合专业研究生创新能力的一个重要手段。

黄芪多糖抗肿瘤作用的研究进展

黄芪多糖是中药黄芪的提取物,具有抗肿瘤、调节免疫、抗氧化、降血糖等多种作用。近年来,其在抗肿瘤方面的作用备受关注。本文通过阅读近几年国内外黄芪多糖抗肿瘤相关文献,主要从调节肿瘤免疫、抑制肿瘤细胞增殖、抑制肿瘤细胞迁移侵袭、诱导肿瘤细胞凋亡、抗血管和淋巴管新生以及联合化疗药物的增效减毒作用几方面展开综述,深入了解黄芪多糖在抗肿瘤方面的作用及机制、研究新进展,为黄芪多糖今后的研究开发和广泛应用提供参考。

Scutellaria barbata D.Don Inhibits Colorectal Cancer Growth via Suppression of Wnt/β-Catenin Signaling Pathway

Objective： To investigate the effect of the ethanol extract of Scutellaria barbata D. Don（EESB） on colorectal cancer（CRC） growth and Wnt/β-catenin signaling pathway in vivo and in vitro. Methods： In vivoexperiment, CRC xenograft mouse model was constructed with injection of HT-29 cells. Following xenograft implantation, twenty mice were randomly divided into EESB-treated group（n=10） and control group（n=10） by a random number table, and were given with intra-gastric administration of 2 g/kg EESB or saline, 5 days a week for 16 days, respectively. At the end of experiment, tumors were removed and weighed by electronic scales. The proliferation biomarker Ki-67 of tumor was evaluated by immunohistochemistry（IHC） assay. In vitro study, HT-29 cells were treated with 0, 0.5, 1.5, 2.5 mg/m L EESB for 24 h. At the end of the treatment, the viability and survival of HT-29 cells were determined by methylthiazolyldiphenyl-tetrazolium bromide（MTT） assay and colony formation assay, respectively. The m RNA expression of c-Myc, Survivin and adenomatous polyposis coli（APC） was examined by reverse transcription-polymerase chain reaction（RT-PCR） both in tumor tissues of CRC xenograft mice and HT-29 cells. Protein expression of c-Myc, Survivin, APC, and β-catenin as well as β-catenin phosphorylation level were evaluated by IHC assay or Western blotting. Results： EESB significantly reduced tumor weight in CRC xenografts mice, compared with the control group（P〈0.05）. IHC assay showed that EESB significantly inhibited protein expression of Ki-67 in tumor tissues（P〈0.05）. MTT assay showed that EESB significantly reduced HT-29 cell viability in a dose-dependent manner（P〈0.05）. Colony formation assay showed that EESB dose-dependently decreased the survival of HT-29 cells（P〈0.05）. In addition, RT-PCR assay showed that EESB decreased the m RNA expression of c-Myc and Survivin and increased APC expression, both in tumor tissues of CRC xenograft mice and HT-29 cells（P〈0.05）. IHC assay or Western blotting showed that EESB decreased protein expression of β-catenin, c-Myc and Survivin, as well as increased APC expression and β-catenin phosphorylation in tumor tissues or HT-29 cells（P〈0.05）. Conclusions： EESB significantly reduced tumor growth in CRC xenografts mice, and inhibited the viability and survival of HT-29 cells. EESB could suppress the activation of the Wnt/β-catenin pathway, which might be one of the mechanisms whereby Scutellaria barbata D. Don exerts its anticancer activity.

电针“足三里”减轻大肠癌小鼠化疗后肠黏膜损伤及其对氧化应激和细胞凋亡的影响

目的:观察电针“足三里”对大肠癌荷瘤小鼠5-氟尿嘧啶(5-FU)化疗后肠黏膜损伤的减轻作用及对结肠组织氧化应激及细胞凋亡的影响,探讨电针缓解化疗后肠黏膜损伤的部分机制。方法:BALB/c小鼠随机分为正常组、CT26组、5-FU组、非穴组和足三里组,每组6只。除正常组外,其余4组采用大肠癌CT26细胞皮下注射种植移植瘤;5-FU组、非穴组和足三里组腹腔注射5-FU(5 mg/mL)溶液,每3 d注射1次,共7次。足三里组和非穴组小鼠在每次给予5-FU腹腔注射后进行电针,足三里组电针双侧“足三里”,非穴组电针双侧非穴,每次5 min,3 d治疗1次,共治疗21 d。治疗结束后评估小鼠腹泻指数,测量小鼠结肠长度;HE染色法观察小鼠结肠黏膜病理形态,测量结肠绒毛长度;生化分析法检测小鼠结肠组织丙二醛(MDA)含量、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)活性;TUNEL法检测小鼠结肠组织细胞凋亡水平;免疫组织化学法检测小鼠结肠组织B淋巴细胞瘤-2蛋白(Bcl-2)、B淋巴细胞瘤-2基因相关X蛋白(Bax)阳性表达水平。结果:与正常组比较,CT26组小鼠腹泻指数、结肠长度、结肠绒毛长度、MDA含量、SOD活性、GSH-Px活性、细胞凋亡率、Bax、Bcl-2阳性表达率差异均无统计学意义。与CT26组比较,5-FU组小鼠腹泻指数升高(P<0.01),结肠长度及结肠绒毛均缩短(P<0.01),结肠组织MDA含量升高(P<0.05),SOD和GSH-Px活性降低(P<0.01),细胞凋亡率升高(P<0.01),Bax阳性表达升高(P<0.05),Bcl-2阳性表达率降低(P<0.01)。与5-FU组比较,足三里组小鼠腹泻指数降低(P<0.01),结肠长度及结肠绒毛均增长(P<0.01),结肠组织MDA含量降低(P<0.05),SOD和GSH-Px活性升高(P<0.01),细胞凋亡率降低(P<0.01),Bax表达降低(P<0.05),Bcl-2表达升高(P<0.01);非穴组小鼠结肠绒毛长度增长(P<0.01),结肠组织SOD、GSH-Px活性升高(P<0.05,P<0.01),结肠组织细胞凋亡率降低(P<0.01)。结论:电针“足三里”可减轻大肠癌荷瘤小鼠5-FU化疗后肠黏膜损伤,其作用机制可能与调节结肠组织氧化应激、抑制细胞凋亡有关。

片仔癀胶囊治疗转移性结肠癌随机双盲对照试验研究

目的评价片仔癀胶囊对转移性结肠癌患者的临床疗效与安全性。方法采用随机双盲安慰剂对照研究方法,将患者按2:1的比例分配至试验组(79例)和对照组(38例),分别给予片仔癀胶囊和安慰剂,每次2粒,每日3次,连续治疗28天为1个疗程,以疾病出现进展为干预终点。检测患者肿瘤标志物(CEA、CA199)、血常规(RBC、WBC、NEU、PLT)及肝肾功能(ALT、AST、γ-GGT、TBIL、BUN、SCr),计算两组无进展生存期(PFS),观察两组患者不良事件(AE)。结果与本组治疗前比较,对照组治疗2个疗程后CA199降低(P<0.01),试验组治疗2个疗程后CEA和CA199降低(P<0.01)。与对照组同期比较,试验组治疗2个疗程后CA199升高(P<0.05)。使用片仔癀胶囊和用药周期≥3个疗程为保护因素(P<0.01,P<0.05)。片仔癀胶囊可提高患者1年无进展生存率(P=0.011)。结论服用片仔癀胶囊3个疗程以上,可降低患者肿瘤标志物CA199,提高PFS,且不增加AE。

Shexiang Tongxin Dropping Pill（麝香通心滴丸）Protects against Na2S2O4-Induced Hypoxia-Reoxygenation Injury in H9c2 Cells

Objectives: To investigate the protective effects of Shexiang Tongxin Dropping Pill(麝香通心滴丸,STP) on Na2S2O4-induced hypoxia-reoxygenation injury in cardiomyoblast H9c2 cells. Methods: The cell viability and levels of mRNA and protein expression in H9c2 cells were determined following Na2S2O4-induced hypoxia using Hoechst staining, annexin V/propidium iodide(PI) flow cytometry, real-time polymerase chain reaction and Western blot analysis. Results: STP pretreatment signi?cantly increased the viability and inhibited aberrant morphological changes in H9c2 cardiomyoblast cells induced by Na2S2O4 treatment(P<0.05). In addition, STP pretreatment attenuated Na2S2O4-induced hypoxic damage, down-regulated the expression of pro-apoptotic Bax,and up-regulated the expression of anti-apoptotic Bcl-2 in H9c2 cells(P<0.05). Conclusions: STP was strongly cardioprotective in hypoxia-reoxygenation injury by preventing hypoxic damage and inhibiting cellular apoptosis.These results further support the use of STP as an effective drug for the treatment of ischemic heart disease.

解毒消癥饮对血管生成因子A诱导人脐静脉内皮细胞的体外血管生成抑制作用

目的:探讨解毒消癥饮(JXY)对血管生成因子A(VEGFA)诱导的人脐静脉内皮细胞(HUVECs)凋亡、迁移以及血管生成的体外影响。方法:HUVECs分为对照组、诱导组(VEGFA 10ng/mL)、JXY低剂量组(VEGFA10ng/L+JXY0.05mg/mL)、JXY中剂量组(VEGFA10ng/mL+JXY0.1mg/mL)、JXY高剂量组(VEGFA10ng/mL+JXY 0.2mg/mL),通过MTT、Hoechst、划痕实验,管腔形成实验、Transwell以及Western blot分别检测细胞活力、凋亡、修复能力、血管生成、细胞迁移和VEGFR2、MMP2、MMP9蛋白表达。结果:与诱导组比较,JXY以剂量依赖性地抑制VEGFA诱导的细胞增殖、修复能力,迁移以及体外管腔的形成能力(P<0.01),促进凋亡(P<0.05,P<0.01)。进一步研究表明JXY剂量依赖性降低VEGFR2、MMP2、MMP9蛋白表达量。结论:JXY显著抑制VEGFA诱导的HUVECs细胞体外血管生成,与其降低VEGFR2、MMP2、MMP9表达直接相关。

Babao Dan Alleviates 5-Fluorouracil-Induced Intestinal Damage via Wnt/β-Catenin Pathway

Objective:To evaluate the protective function of Babao Dan(BBD)on 5-flurouracil(5-FU)-induced intestinal mucositis(IM)and uncover the underlying mechanism.Methods:A total of 18 male mice were randomly divided into 3 groups by a random number table,including control,5-FU and 5-FU combined BBD groups,6 mice in each group.A single intraperitoneal injection of 5-FU(150 mg/kg)was performed in 5-FU and 5-FU combined BBD groups on day 0.Mice in 5-FU combined BBD group were gavaged with BBD(250 mg/kg)daily from day 1 to 6.Mice in the control group were gavaged with saline solution for 6 days.The body weight and diarrhea index of mice were recorded daily.On the 7th day,the blood from the heart of mice was collected to analyze the proportional changes of immunological cells,and the mice were subsequently euthanized by mild anesthesia with 2% pentobarbital sodium.Colorectal lengths and villus heights were measured.Intestinal-cellular apoptosis and proliferation were evaluated by Tunel assay and immunohistochemical staining of proliferating cell nuclear antigen,respectively.Immunohistochemistry and Western blot were performed to investigate the expressions of components in Wnt/β-catenin pathway(Wnt3,LRP5,β-catenin,c-Myc,LRG5 and CD44).Results:BBD obviously alleviated 5-FU-induced body weight loss and diarrhea,and reversed the decrease in the number of white blood cel s,including monocyte,granulocyte and lymphocyte,and platelet(P<0.01).The shortening of colon caused by 5-FU was also reversed by BBD(P<0.01).Moreover,BBD inhibited apoptosis and promoted proliferation in jejunum tissues so as to reduce the intestinal mucosal damage and improve the integrity of vil us and crypts.Mechanical y,the expression levels of Wnt/β-catenin mediators such as Wnt3,LRP5,β-catenin were upregulated by BBD,activating the transcription of c-Myc,LRG5 and CD44(P<0.01).Conclusions:BBD attenuates the adverse effects induced by 5-FU via Wnt/β-catenin pathway,suggesting it may act as a potential agent against chemotherapy-induced intestinal mucositis.

Chemical Composition,Anticancer,Anti-neuroinflammatory,and Antioxidant Activities of the Essential Oil of Patrinia scabiosaefolia

Objective：To study the chemical composition,anticancer,anti-neuroinflammatory,and antioxidant activities of the essential oil of Patrinia scabiosaefolia（EO-PS）.Methods：Patrinia scabiosaefolia was analyzed by gas chromatography-mass spectrometry.Eight human carcinoma cell lines,including SGC-7901,AGS,Hep G2,HT-29,HCT-8,5-FU/HCT-8,He La,and MDA-MB-231,were assessed by methylthiazolyldiphenyltetrazolium bromide（MTT） assay.Anti-neuroinflammatory activity was assessed by production of interleukin（IL）-1β and IL-6 induced by lipopolysaccharide in BV-2 cells（microglia from mice）.The antioxidant activity was evaluated with a 2,2-diphenyl-1-picrylhydrazyl（DPPH） scavenging assay.Results：Forty-four components,representing 83.919% of the total oil,were identified in the EO-PS.The major constituents were caryophyllene oxide（12.802%）,caryophyllene（6.909%）,α-caryophyllene（2.927%）,β-damascenone（3.435%）,calarene（5.621%）,and phenol（3.044%）.The MTT assay showed that the EO-PS exhibited significant dose-dependent growth inhibition in the 50–200 μg/m L dilution range.The EO-PS exhibited a dose-dependent scavenging activity against the DPPH radical,with an half of maximal inhibitory concentration 1.455 mg/m L.Conclusion：The EO-PS possesses a wide range of antitumor,anti-neuroinflammatory and antioxidant activities,suggesting that it may be a good candidate for further investigations of new bioactive substances.

以环状RNA为靶点的中药抗肿瘤治疗研究进展

环状RNA(circular RNA,circRNA)是一组分子呈封闭环状结构,且不具有编码功能的转录本。由于其共价闭合的环结构,没有3’和5’末端,因此不易被外切酶降解[1]。与线性RNA比较,它更为保守和稳定,广泛存在于真核细胞的细胞质中。1976年,Sanger HL等[2]首次在植物病毒中发现了单链共价闭合circRNA分子。