HPLC法测定冬凌草萜含片中冬凌草甲素的含量

目的：对冬凌草萜含片中的冬凌草甲素进行含量测定。方法：采用ＨＰＬＣ法,以Ｃ１８－ＯＤＳ为色谱柱,甲醇－水（５５：４５）为流动相,检测波长为２４２ｍｍ。结果：回收率为９８．２３％,ＲＳＤ为２．１２％。结论：该方法简便、可靠,适合冬凌草及其制剂的质量控制。     

HPLC法测定冬凌草片中冬凌草甲素的含量

用HPLC法对不同厂家和自制冬凌草片中冬凌草甲素的含量进行了比较,采用HypersilODS2C18色谱柱,以甲醇－水（50：50）为流动相,检测波长为238nm,结果表明含量有较大差异。建议通过控制药材的含叶量来控制冬凌草片中冬凌草甲素的含量。     

高效液相色谱法测定冬凌草糖浆中冬凌草甲素含量

目的 建立测定冬凌草糖浆的冬凌草甲素含量的高效液相色谱法.方法采用Diamonsil C18色谱柱(250mm×4.6mm,5μm),以甲醇-水(50:50)为流动相,流速为1.0 mL/min,检测波长为239 nm.结果冬凌草甲素进样量在0.093～0.746μg范围内与峰面积积分值线性关系良好,回归方程为Y=413 933.35-63 428.66X,r=0.999 7(n=5);平均加样回收率为99.0%,RSD为1.06%(n=5).结论该法简便、准确、专属性和重复性好,为冬凌草糖浆中冬凌草甲素的定量分析提供了科学有效的方法.     

HPLC测定冬凌草药材及片剂中的冬凌草甲素

目的 采用HPLC法测定冬凌草药材及片剂中的冬凌草甲素.方法 用Hypersil C_(18)(250 mm×4.6 mm,5μm)色谱柱,甲醇-水(50:50)为流动相,流速0.8 mL·min~(-1);检测波长238 nm;柱温25 ℃;进样量20μL.结果 冬凌草甲素20～200 mg·mL~(-1)与峰面积有良好的线性关系(r=0.9999),最低检出限50 ng.按<中国药典>方法对样品预处理后进样测量,在药材中可检出冬凌草甲素,但片剂中无法检出.对供试品脱脂后,片剂中仍未检出冬凌草甲素.结论 所建方法简便、重复性好、准确度高,可用于冬凌草片剂的质量控制.文中所选的冬凌草片剂中未检测出冬凌草甲素,说明其质量有待提高.     

HPLC测定微波提取冬凌草中的冬凌草甲素

目的从冬凌草中提取冬凌草甲素并测定其含量。方法采用微波萃取技术提取,HPLC法测定。结果测得冬凌草叶中冬凌草甲素含量为0.6152%,平均回收率为97.86%,RSD=2.14%。结论采用微波萃取技术提取冬凌草甲素,具有提取快速、操作简便、节省溶剂和有效成分提取率高等优点。     

冬凌草甲素上调人前列腺癌PC-3细胞PTEN基因的表达及其诱导凋亡作用

目的：研究冬凌草甲素诱导人前列腺癌PC-3细胞凋亡及与PTEN基因表达的关系。方法：MTT比色法测定冬凌草甲素诱导PC-3细胞的增长抑制;流式细胞术分析DNA倍体测定细胞凋亡;RT-PCR半定量分析冬凌草甲素诱导人前列腺癌PC-3细胞后PTENmRNA的表达情况。结果：MTT比色法测得冬凌草甲素对PC-3细胞具有明显的增殖抑制作用,并随冬凌草甲素浓度的升高及作用时间的延长而增强;流式细胞术分析PC-3细胞经冬凌草甲素诱导后,细胞凋亡比率随作用时间延长而增加。RT-PCR测得PTEN基因mRNA的表达随细胞凋亡的增加而逐渐增高。结论：冬凌草甲素能够明显抑制PC-3细胞的增殖,并诱导前列腺癌细胞凋亡,其作用途径可能与上调PTENmRNA表达有关。     

HPLC法测定冬凌草萜含片中冬凌草甲素的含量

目的:对冬凌草萜含片中的冬凌草甲素进行含量测定。方法:采用HPLC 法,以C₁₈-ODS 为色谱柱,甲醇-水(55∶45) 为流动相,检测波长为242nm 。结果:回收率为98 .23 % ,RSD为2 .12 % 。结论:该方法简便、可靠,适合冬凌草及其制剂的质量控制     

冬凌草甲素诱导HL-60细胞凋亡

目的　研究冬凌草甲素诱导人白血病HL 6 0细胞凋亡的作用。方法　形态学观察 ,DNA凝胶电泳及流式细胞术。结果　冬凌草甲素能显著地诱导HL 6 0细胞发生凋亡 ,其作用呈明显的浓度效应关系和时间依赖性。形态学观察可见凋亡小体的形成 ,琼脂糖凝胶电泳可见明显的DNA梯带 ;流式细胞仪检测到G1亚峰。结论　冬凌草甲素能诱导HL 6 0细胞凋亡 ,并与其细胞杀伤活性相互平行 ,提示冬凌草甲素的抗癌活性与诱导肿瘤细胞凋亡相关     

高效液相色谱法同时测定冬凌草中冬凌草甲、乙素的含量

目的研究冬凌草中冬凌草甲、乙素的含量测定.方法采用高效液相色谱法进行测定.结果冬凌草甲素进样量在0.196～0.980μg之间线性关系良好,相关系数r=0.9999,回收率为99.29%,测定的RSD=2.1%.冬凌草乙素进样量在0.1712～0.8560μg之间线性关系良好,相关系数r=0.9999,回收率为98.63%,测定的RSD=1.9%.结论本法简便、准确、灵敏,重现性好,可用于冬凌草及其制剂的质量控制.     

高效毛细管电泳法测定冬凌草片中冬凌草甲素的含量

目的:建立用高效毛细管电泳测定冬凌草片中冬凌草甲素的含量测定方法.方法:样品用乙醚超声提取,缓冲液为20 mmol·L-1磷酸盐缓冲液,pH=7.0,分离电压15 kM,石英毛细管33 cm×75 μm,操作温度25℃,紫外检测波长238 nm,压力进样6.0 kV×10 s,内标法定量.结果:线性范围为50～400 mg·L-1,平均回收率为99.5%,RSD为3.9%.结论:该方法简便、快速且消耗试剂少,适合于冬凌草片剂的质量控制.     

Cellular Responses to Cancer Chemopreventive Agent D,L-Sulforaphane in Human Prostate Cancer Cells Are Initiated by Mitochondrial Reactive Oxygen Species

Purpose  Present study was undertaken to elucidate the mechanism of cellular responses to D,L-sulforaphane (SFN), a highly promising cancer chemopreventive agent. Methods  Mitochondrial DNA deficient Rho-0 variants of LNCaP and PC-3 cells were generated by culture in the presence of ethidium bromide. Apoptosis was assessed by analysis of cytoplasmic histone-associated DNA fragmentation and activation of caspase-3. Immunoblotting was performed to determine the expression of apoptosis- and cell cycle-regulating proteins. Generation of reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and cell cycle distribution were measured by flow cytometry. Results  The Rho-0 variants of LNCaP and PC-3 cells were significantly more resistant to SFN-induced ROS generation, apoptotic DNA fragmentation, disruption of MMP, cytosolic release of cytochrome c, and G2/M phase cell cycle arrest compared with corresponding wild-type cells. SFN-induced autophagy, which serves to protect against apoptotic cell death in PC-3 and LNCaP cells, was also partially but markedly suppressed in Rho-0 variants compared with wild-type cells. SFN statistically significantly inhibited activities of mitochondrial respiratory chain enzymes in LNCaP and PC-3 cells. Conclusion  These results indicate, for the first time, that mitochondria-derived ROS serve to initiate diverse cellular responses to SFN exposure in human prostate cancer cells. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. This investigation was supported in part by the USPHS grants CA115498 and CA101753 (to S.V.S.), awarded by the National Cancer Institute, and grant 2718/P01/2007/32 (to A.H-A.), awarded by the Polish Ministry of Science and Higher Education. Dong Xiao and Anna A. Powolny contributed equally.     

冬凌草叶和茎中冬凌草甲素含量的对比研究

目的：建立了冬凌草叶及茎中冬凌草甲素的高效液相色谱测定法。方法：采用Ｈｙｐｅｒｓｉｌ ＯＤＳ２Ｃ１８色谱柱,以甲醇－水（５０：５０）为流动相,在２３８ｎｍ波长处检测。结果：叶和茎中冬凌草甲素的含量分别约为０．５５％和０．０２５％,平均回收率为９７．４５％。结论：该方法准确、灵敏、简便。     

shRNA 沉默CtBP2 基因对前列腺癌细胞的增殖作用

【摘要】目的 探讨RNA 干扰技术沉默羧基末端结合蛋白（CtBP）2 基因表达对前列腺癌PC3 细胞增殖能力的影响。方法 实验分为空白对照组、转染空质粒组、转染shRNA 组。设计并合成针对CtBP2 的3 条特异性短发卡 RNA（shRNA）模板,构建CtBP2-shRNA 重组质粒,转染PC3 细胞。通过RT-PCR 检测CtBP2 mRNA 的表达水平;采用Western blot 法检测CtBP2 蛋白表达的水平,运用MTT 法检测沉默CtBP2 对PC3 细胞体外增殖的抑制作用。结果 将CtBP2-shRNA 转染前列腺癌PC3 细胞后,CtBP2 mRNA 以及蛋白的表达水平均明显下降。沉默CtBP2 表达后,MTT 结果显示转染shRNA 组的PC3 细胞较空白对照组、转染空质粒组细胞增殖能力明显减弱,差异有统计学意义（P < 0.01）。结论 CtBP2-shRNA 可抑制CtBP2 在前列腺癌细胞中的表达并抑制肿瘤细胞生长,提示CtBP2 可作为前列腺癌基因治疗的一个新靶点。     

Polyazamacrocycles as Potential Antitumor Agents for Human Prostate Cancer Cells

Polyazamacrocycles are currently being studied and used in a variety of applications beyond their traditional place in supramolecular and co‐ordination chemistry. This study suggests additional applications of these compounds with particular emphasis on their use as antiproliferative agents that could be potentially used to treat cancer. Four polyazamacrocycles were tested in human prostate cancer LNCaP and prostate epithelial PNTA1 cells to analyze changes in cell proliferation and cell death capabilities. Their intracellular localization was also evaluated by confocal microscopy. The results show a decrease in proliferation rate and cell viability of LNCaP and PNTA1, after treatment with these compounds. The decrease in the number of viable cells is similar for the majority of the compounds studied, and at higher concentration, the proliferation efficiency decreased significantly in the cell lines studied. Also, our results suggest that L and L₂ induce early apoptosis in PNTA1 cells and late apoptosis/necrosis in LNCaP cells. The compounds did not induce a significant increase in necrosis of both cell types. Although the compounds did not localize in a unique organelle, all of them have as main target the Golgi apparatus and other localization profiles differed depending on the cell line.     

Activation of RhoA,B,C by Yersinia Cytotoxic Necrotizing Factor (CNFy) Induces Apoptosis in LNCaP Prostate Cancer Cells

Prostate cancer is the most common malignancy, accounting for about 25% of all incident cases among men in industrialized countries. The human androgen-dependent prostate cancer cell line LNCaP, which is derived from a metastatic lesion of human prostatic adenocarcinoma, is frequently used to study prostate cancer associated signaling pathways in vitro. Recently it was described that Rho GTPase activation in these cells leads to apoptotic responses. We used the bacterial toxins CNFy and CNF1, which specifically and directly activate Rho GTPases by deamidation of a single glutamine. We asked whether these Rho activators could induce apoptosis in LNCaP cells. Our results indicate that RhoA activation, induced by CNFy, does lead to intrinsic apoptosis of the cells. Analysis of the underlying signaling pathway reveals that apoptosis induction requires the activity of Rho kinase (ROCK) and myosin activation, an apoptotic pathway previously identified in cancer stem cells.     

Malignancy Grade–Dependent Expression of K+-Channel Subtypes in Human Prostate Cancer

K⁺ channels are key molecules in the progression of several cancer types and considered to be potential targets for cancer therapy. We examined the gene expressions of voltage-gated (K_v), Ca²⁺-activated (K_Ca), and two-pore domain (K_2P) K⁺-channel subtypes in needle-biopsy samples of human prostate cancer (PCa) by real-time PCR and compared them with those in PCa epithelial cell lines. The expression of K_v1.3, K_Ca1.1, K_Ca3.1, and K_2P1 markedly increased in the PCa group with Gleason score of 5 – 6 (GS5–6) but significantly decreased in the GS8–9 group. This malignancy grade–dependent K⁺-channel expression pattern may provide a convenient marker to understand PCa progression level.     

Bee Venom Components as Therapeutic Tools against Prostate Cancer

Prostate cancer is one of the most common cancers in men. Despite the development of a variety of therapeutic agents to treat either metastatic hormone-sensitive prostate cancer, advanced prostate cancer, or nonmetastatic/metastatic castration-resistant prostate cancer, the progression or spread of the disease often cannot be avoided. Additionally, the development of resistance of prostate cancer cells to available therapeutic agents is a well-known problem. Despite extensive and cost-intensive research over decades, curative therapy for metastatic prostate cancer is still not available. Therefore, additional therapeutic agents are still needed. The animal kingdom offers a valuable source of natural substances used for the treatment of a variety of diseases. Bee venom of the honeybee is a mixture of many components. It contains proteins acting as enzymes such as phospholipase A2, smaller proteins and peptides such as melittin and apamin, phospholipids, and physiologically active amines such as histamine, dopamine, and noradrenaline. Melittin has been shown to induce apoptosis in different cancer cell lines, including prostate cancer cell lines. It also influences cell proliferation, angiogenesis, and necrosis as well as motility, migration, metastasis, and invasion of tumour cells. Hence, it represents an interesting anticancer agent. In this review article, studies about the effect of bee venom components on prostate cancer cells are discussed. An electronic literature research was performed utilising PubMed in February 2021. All scientific publications, which examine this interesting subject, are discussed. Furthermore, the different types of application of these promising substances are outlined. The studies clearly indicate that bee venom or melittin exhibited anticancer effects in various prostate cancer cell lines and in xenografts. In most of the studies, a combination of bee venom or the modified melittin with another molecule was utilised in order to avoid side effects and, additionally, to target selectively the prostate cancer cells or the surrounding tissue. The studies showed that systemic side effects and unwanted damage to healthy tissue and organs could be minimised when the anticancer drug was not activated until binding to the cancer cells or the surrounding tissue. Different targets were used, such as the matrix metalloproteinase 2, hormone receptors expressed by prostate cancer cells, the extracellular domain of PSMA, and the fibroblast activation protein occurring in the stroma of prostate cancer cells. Another approach used loaded phosphate micelles, which were cleaved by the enzyme secretory phospholipase A2 produced by prostate cancer cells. In a totally different approach, targeted nanoparticles containing the melittin gene were used for prostate cancer gene therapy. By the targeted nonviral gene delivery, the gene encoding melittin was delivered to the prostate cancer cells without systemic side effects. This review of the scientific literature reveals totally different approaches using bee venom, melittin, modified melittin, or protoxin as anticancer agents. The toxic agents acted through several different mechanisms to produce their anti-prostate cancer effects. These mechanisms are not fully understood yet and more experimental studies are necessary to reveal the complete mode of action. Nevertheless, the researchers have conducted pioneering work. Based on these results, further experimental and clinical studies about melittin and modifications of this interesting agent deriving from nature are necessary and could possibly lead to a complementary treatment option for prostate cancer.