活血解毒方对缺氧/复氧所致心肌细胞凋亡的影响＊

目的 研究活血解毒方对缺氧/复氧（H/R）所致的心肌细胞凋亡的影响。方法 体外培养H9C2心肌细胞并分成正常对照组（Control组）、缺氧复氧组（H/R组）、H/R + 活血解毒中药组、LY294002组和活血解毒中药 + LY294002组，其中正常对照组给予DMEM培养基培养，缺氧复氧组给予缺氧4 h、复氧6 h处理，缺氧复氧 + 活血解毒中药组、LY294002组和活血解毒中药 + LY294002组分别给予活血解毒中药、LY294002、活血解毒中药配合LY294002预处理24 h，然后均给予缺氧4 h、复氧6 h处理。采用CCK8检测各组心肌细胞的存活率，流式细胞术检测各组细胞凋亡水平，电镜观察各组心肌细胞中线粒体、自噬体的变化，Western Blot检测各组细胞凋亡蛋白半胱氨酸天冬氨酸蛋白酶3（Caspase3）、半胱氨酸天冬氨酸蛋白水解酶3（Cleaved caspase3）、β-连环蛋白（β-Catenin）、p-p65、B淋巴细胞瘤-2（B-cell lymphoma-2，Bcl-2）蛋白的表达。结果 活血解毒方预处理显著增加H/R诱导的H9C2心肌细胞凋亡，降低凋亡相关蛋白Cleaved caspase3、β-Catenin、p-p65表达，增加Bcl-2表达。结论 活血解毒方可通过抑制细胞凋亡，降低缺氧/复氧所致的心肌细胞损伤，其作用机制可能与PI3K信号通路相关。     

Lateral Lymph Nodes in Rectal Cancer: Do we all Think the Same? A Review of Multidisciplinary Obstacles and Treatment Recommendations

Lateral lymph nodes in low, locally advanced, rectal cancer have proven implications for local recurrence rates, which increase drastically in the presence of persistently enlarged lateral lymph nodes. These clinical implications warrant a thorough understanding of lateral nodal disease with awareness and knowledge from all three specialties involved – radiology, radiation oncology, and surgery – to ensure proper treatment. Relevant literature for each specialty, including all current guidelines and perspectives, were examined. Variations in definitions and treatment paradigms were evaluated. There is still no consensus for the standardized treatment of lateral nodal disease. Each discipline works according to their own available evidence, but relevant data are scarce. Current international guidelines and standard recommendations for the diagnostics and treatment of lateral lymph nodes are lacking. This results in differing perspectives and interpretations between the disciplines which can lead to challenging communication in an area where multidisciplinary collaboration is essential. This review addresses this by presenting the current evidence, perspectives and practices of each specialty and makes suggestions for each phase of the diagnostic and treatment process for patients with lateral nodal disease. By doing this, steps are taken toward achieving international consensus, and multidisciplinary collaboration.     

Molecular mechanisms and cellular events involved in the neuroprotective actions of estradiol. Analysis of sex differences

Estradiol, either from peripheral or central origin, activates multiple molecular neuroprotective and neuroreparative responses that, being mediated by estrogen receptors or by estrogen receptor independent mechanisms, are initiated at the membrane, the cytoplasm or the cell nucleus of neural cells. Estrogen-dependent signaling regulates a variety of cellular events, such as intracellular Ca²⁺ levels, mitochondrial respiratory capacity, ATP production, mitochondrial membrane potential, autophagy and apoptosis. In turn, these molecular and cellular actions of estradiol are integrated by neurons and non-neuronal cells to generate different tissue protective responses, decreasing blood-brain barrier permeability, oxidative stress, neuroinflammation and excitotoxicity and promoting synaptic plasticity, axonal growth, neurogenesis, remyelination and neuroregeneration. Recent findings indicate that the neuroprotective and neuroreparative actions of estradiol are different in males and females and further research is necessary to fully elucidate the causes for this sex difference.     

健脾益肾方对非小细胞肺癌细胞体外增殖凋亡作用的实验研究

目的:探讨健脾益肾方对非小细胞肺癌(NSCLC)细胞体外增殖凋亡的作用。方法:人NSCLC细胞系A549分为四组:空白对照组(仅加入细胞培养液)、阴性对照组(加入细胞,不进行中药处理)、实验组(加细胞加中药处理)。荧光定量PCR和Western blot分别检测Survivin、Bcl-2和Caspase-3的mRNA和蛋白表达。MTT检测细胞增殖;流式细胞术检测细胞凋亡。结果:与空白对照组相比,阴性对照组细胞在24、48、72 h的吸光度值明显升高,细胞凋亡率下降,Survivin和Bcl-2 mRNA和蛋白相对表达量上调,Caspase-3 mRNA和蛋白相对表达量下调,组间比较差异有统计学意义(P<0.05);而健脾益肾方处理的实验组24、48、72 h的吸光度值均显著降低,细胞凋亡率显著上升,Survivin和Bcl-2 mRNA和蛋白相对表达量下调,Caspase-3 mRNA和蛋白相对表达量上调,与空白对照组相比差异有统计学意义(P<0.05)。结论:健脾益肾方可通过下调Survivin和Bcl-2、上调Caspase-3表达诱导NSCLC细胞凋亡,并抑制肿瘤细胞的增殖,进而抑制NSCLC的发展。     

D-半乳糖联合对氯苯丙氨酸致肾不藏志不寐大鼠下丘脑神经递质和凋亡基因的实验研究

目的:观察D-半乳糖联合对氯苯丙氨酸(PCPA)致肾不藏志不寐大鼠下丘脑神经递质和凋亡基因的变化。方法:40只SD大鼠随机分为对照组、老年组、失眠组、肾不藏志不寐组(模型组),每组10只。老年组大鼠给予D-半乳糖皮下注射,失眠组给予对PCPA腹腔注射,肾不藏志不寐组给予D-半乳糖皮下注射联合PCPA腹腔注射建立模型,对照组给予等剂量生理盐水注射。造模结束后测量各组大鼠体质量、采用水迷宫观察大鼠定位航行探索运动轨迹、采用RT-PCR法检测各组大鼠下丘脑GABAARα1、GABAARβ2、GABAARγ2、Bax、Bcl-2 mRNA的相对表达量。结果:与空白组比较,模型组体质量显著降低(P<0.01);定位航行探索运动轨迹显著延长;模型组下丘脑GABAARα1、GABAARβ2、GABAARγ2、Bcl-2mRNA相对表达量均显著性降低(P<0.05,P<0.05,P<0.01,P<0.05);下丘脑Bax mRNA相对表达量显著性增加(P<0.05)。结论:D-半乳糖联合PCPA致肾不藏志不寐大鼠下丘脑睡眠相关的神经递质和凋亡基因表达发生改变。     

Anticancer Activity of Solvent Extracts of Hexogonia glabra against Cervical Cancer Cell Lines

Objective: In this study, we aimed to harness some solvent extracts of one wild mushroom Hexagonia glabra and test their anti-cancer activity against cervical human cell lines, namelyHeLa, SiHa, and CaSki. Methods: It includes cell morphological study by microscope, nuclear morphology by DAPI staining under fluorescence microscopy, apoptosis assay by fluorescence technique, anti-proliferation by MTT assay and expression of apoptotic and anti-apoptotic genes by Western blotting and cell cycle analysis was done. Results: The selected cervical cancer cells were treated separately with 150 µg/mL of three extracts, namely of ethanolic (EE), ethyl acetate (EAE), and water extract (WE) and exhibited features like round, shrink and dead. All extracts caused apoptosis in cell lines and EE had the highest effect in this regard. The percentages of apoptotic cells in HeLa, SiHa and CaSki, at the same concentration of EE were 79.23, 75.42, and 76.36% respectively. Cytotoxicity assay showed that all three extracts (50 – 250 μg/mL) were potent for inhibition of cell growth of three cell lines and again EE had the highest effect. The percentages of cell growth inhibition in HeLa, SiHa, and CaSki cells treated with EE at 24 h at 50 µg/mL were 45.79±4.11, 41.66±4.03, and 36.72±2.67, while they were 74.23±7.45, 62.31±5.97, and 54.23±5.04 at 150 µg/mL concentration. At 250 µg/mL concentration, the percentages of cell growth inhibition were 94.25 ±8.11, 90.02 ±8.67, and 85.43±6.21, respectively. The expression of apoptotic gene (Caspase 3, 9) and tumor guard gene (p53), as their proteins in Western blotting increased . However, anti-apoptotic BcL2 gene of all cell lines was decreased following treatment with extracts. In addition, the cell cycle analysis (CaSki cell) showed that treatment (EE) arrested at G2/M check point cell cycle. Conclusion: All extracts of this mushroom were active in arresting growth of three cell lines and EE had the highest effect, indicating that this mushroom can be a valuable source of anticancer agents.     

二甲双胍对甲状腺乳头状癌TPC-1细胞凋亡的影响及其机制研究

目的 探讨二甲双胍对甲状腺乳头状癌TPC-1细胞凋亡的影响及其机制。方法 用不同浓度(0mmol/L、1mmol/L、5mmol/L、10mmol/L、20mmol/L、40mmol/L)二甲双胍分别处理TPC-1细胞24h。采用四甲基偶氮唑蓝（MTT）法检测细胞增殖能力；采用Annexin Ⅴ-FITC/PI流式细胞术检测不同浓度二甲双胍对TPC-1细胞凋亡的影响，同时在40mmol/L组加入腺苷酸活化蛋白激酶(AMPK)抑制剂Compound C，观察其对TPC-1细胞凋亡的影响；采用Western blot法检测磷酸化AMPK（p-AMPK）及Caspase-9蛋白的表达。结果 与0mmol/L组比较，1mmol/L及5mmol/L组的TPC-1细胞增殖能力无明显改变（P＞0.05）；而10mmol/L、20mmol/L、40mmol/L二甲双胍均可明显抑制TPC-1细胞的增殖能力，且呈剂量依赖性（P＜0.05）。与0mmol/L组比较，不同浓度二甲双胍均可明显增加TPC-1细胞凋亡率（P＜0.001）；而加入Compound C后，40mmol/L组TPC-1细胞凋亡率较前明显下降（P＜0.001）。与0mmol/L组比较，1mmol/L、5mmol/L、10mmol/L、20mmol/L、40mmol/L组p-AMPK、Caspase-9表达量增加。结论 二甲双胍可诱导甲状腺乳头状癌TPC-1细胞凋亡，且二甲双胍诱导TPC-1细胞凋亡的过程可能与激活AMPK通路有关，可能由Caspase-9介导。     

灯盏乙素介入IP3R-Ca2+途径抑制β淀粉样蛋白诱导的神经细胞凋亡

目的：观察灯盏乙素（Scu）对β淀粉样蛋白（Aβ）诱导的细胞模型中1，4，5-三磷酸肌醇受体（IP₃R）-Ca²⁺途径的影响，探讨其在阿尔茨海默病（AD）病程中可能发挥的积极作用。方法：选用神经母细胞瘤细胞分为对照组、Scu处理组、Aβ处理组、Aβ+Scu （高、中、低）处理组及Aβ+IP₃R拮抗剂组，用CCK-8法筛选药物浓度并检测各组细胞生存率；用酶联免疫吸附法检测各组细胞中1，4，5-三磷酸肌醇（IP₃）的含量；用蛋白印迹和实时荧光定量聚合酶链反应方法检测各组细胞IP₃R和凋亡相关因子Caspase-3、Bcl-2、Bax的蛋白及mRNA的表达水平；用激光共聚焦显微镜观察各组细胞内Ca²⁺浓度的变化；用AnnexinV/PI双染法测定各组细胞的凋亡率。结果：与对照组和Scu处理组相比，Aβ处理组细胞存活率下降，IP₃含量升高，IP₃R、Bax和Caspase-3的蛋白及mRNA表达上调，Bcl-2蛋白及mRNA的表达下调，细胞胞浆内Ca²⁺浓度及细胞凋亡率升高；Aβ+Scu处理组细胞中各检测指标的变化与Aβ处理组的结果正好相反，IP₃R通道下游指标的变化与Aβ+IP₃R拮抗剂组基本一致。结论：Scu能够下调通路蛋白IP₃、IP₃R的表达，抑制Aβ介导的Ca²⁺内流所致的细胞凋亡，可能通过对IP₃R-Ca²⁺途径的调控来影响AD病程。     

益气逐瘀方对缺氧诱导乳鼠心肌细胞凋亡及miR-24/Bim通路的影响

目的研究益气逐瘀方(参元丹)对缺氧诱导乳鼠心肌细胞凋亡及miR-24/Bim通路的影响。方法分离并培养乳鼠心肌细胞,建立心肌细胞缺氧模型,Lipofectamine2000转染miR-24 mimic/inhibitor至心肌细胞,实验分为正常组、缺氧组、参元丹组、参元丹+miR-24 mimic组、参元丹+miR-24 inhibitor组,治疗组予参元丹含药血清干预。比色法检测心肌细胞培养基上清液LDH、CPK活性,MTT法检测心肌细胞存活率,Annexin V/PI双染流式细胞术检测心肌细胞凋亡,qRT-PCR检测心肌细胞miR-24、Bim mRNA表达。结果与低氧组相比,参元丹组、参元丹+miR-24 mimic组、参元丹+miR-24 inhibitor组均能显著降低细胞培养基上清液LDH、CPK活性(P<0.05),提高心肌细胞存活率(P<0.05),降低心肌细胞凋亡(P<0.05),升高心肌细胞miR-24 mRNA表达(P<0.05),降低Bim mRNA表达(P<0.05);治疗组之间比较,参元丹+miR-24 mimic组作用更显著(P<0.05)。结论益气逐瘀方参元丹能够减轻缺氧诱导乳鼠心肌细胞损伤及细胞凋亡,其作用机制可能与其上调miR-24表达,同时抑制其靶基因Bim mRNA表达有关。