Efficacy of stem cell-based therapies for colistin-induced nephrotoxicity

The increase in infections with multidrug resistant bacteria has forced to return to the use of colistin, antibiotic with known nephrotoxicity. Mesenchymal stem cells (MSCs) are being extensively investigated for their potential in regenerative medicine. This study aimed to investigate the possible protective mechanisms of the MSCs against kidney injury induced by colistin. Forty adult female albino rats were randomly classified into 4 equal groups; the control group, the MSC-treated group (a single dose of 1 ×10⁶ /ml MSCs through the tail vein), the colistin-treated group (36 mg/kg/day colistin was given for 7 days), and the both colistin and MSC group (36 mg/kg/day colistin and 1 ×10⁶ /ml MSCs). Main outcome measures were histopathological alterations, kidney malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and immunohistological autophagy evaluation. MSC repressed the progression of colistin-induced kidney injury as evidenced by the improvement of histopathological alterations and the substantial increase MDA, and decrease SOD and CAT in serum levels. Moreover, MSC resulted in a profound reduction in oxidative stress as manifested by decreased MDA and increased SOD in serum. Notably, MSC suppressed colistin-induced autophagy; it reduced renal levels of Beclin-1, P62 and LC3A/B. Furthermore, MSC decreased renal levels of eNOS. Lastly, MSC efficiently decreased expression of the TUNEL positive cell number. MSC confers protection against colistin-induced kidney injury by alleviating oxidative stress, nitric oxide synthase besides modulating reducing autophagy and apoptosis.     

Evaluation of Ultra-Microscopic Changes and Proliferation of Apoptotic Glioblastoma Multiforme Cells Induced by Velogenic Strain of Newcastle Disease Virus AF2240

Aim: Newcastle disease virus (NDV) is a member of genus Avulavirus within the family Paramyxoviridae. Interestof using NDV as an anticancer agent has arisen from its ability to kill tumor cells with limited toxicity to normal cells.Methods: In this investigation, the proliferation of brain tumor cell line, glioblastoma multiform (DBTRG.05MG)induced by NDV strain AF2240 was evaluated in-vitro, by using MTT proliferation assay. Furthermore, Cytologicalobservations were studied using fluorescence microscopy and transmission electron microscopy, DNA laddering inagarose gel electrophoresis assay used to detect the mode of cell death and analysis of the cellular DNA content byflowcytometery. Results: MTT proliferation assay, Cytological observations using fluorescence microscopy andtransmission electron microscopy show the anti-proliferation effect and apoptogenic features of NDV on DBTRG.05MG.Furthermore, analysis of the cellular DNA content showed that there was a loss of treated cells in all cell cycle phases(G1, S and G2/M) accompanied with increasing in sub-G1 region (apoptosis peak). Conclusion: It could be concludedthat NDV strain AF2240 is a potent antitumor agent that induce apoptosis and its cytotoxicity increasing while increasingof time and virus titer.     

Anti-Oxidant,Anti-Hemolytic Effects of Crataegus aronia Leaves and Its Anti- Proliferative Effect Enhance Cisplatin Cytotoxicity in A549 Human Lung Cancer Cell Line

Objective: For Arabian traditional medicine, Crataegus aronia syn. Azarolus (L) Bosc. ex DC (Rosaceae) is widely used to treat diabetes, sexual weakness, cardiovascular diseases and cancer. The anti-cancerous and anti-hemolysis effects of the hydroalcoholic extract of this plant have never been investigated before. The present study aims to evaluate the biological activities of the hydroalcoholic extract of Crataegus aronia leaves in combination with cisplatin, one of the most widely employed chemotherapeutics, on A549 human lung cancer cell line. Methods: The anti-oxidant and anti-proliferative activities of leaves, fruits, seeds of C. aronia were investigated by DPPH method and MTT assay; respectively. Cell migration activity was investigated by wound healing and by cell aggregation assays. The effect of C. aronia in inducing cell cycle arrest along with activating cell apoptosis was evaluated by flow cytometry and Western blot assays, respectively. Results: Our results showed that C. aronia leaves (C. aronia L.) had the highest anti-oxidant and anti-proliferative activities. The leaves extract was potent against hemolysis of the human erythrocytes and showed elevated decrease in migration by reducing wound healing migration and by increasing cell aggregation. Finally, C. aronia L. treatment exhibited apoptotic activity on A549 cells by the down-regulation of PARP-1, caspase-3 and Bcl-2 proteins and by increasing the percentage of A549 cells in sub G0 cell cycle. Moreover, the co-treatment of C. aronia L. and cisplatin remarkably sensitised A549 cells to cisplatin. Conclusion: The results suggested that C. aronia L. could be used as a potential treatment against human lung cancer exhibiting minimal side effects on human health.     

Luteolin Suppresses Teratoma Cell Growth and Induces Cell Apoptosis via Inhibiting Bcl-2

Luteolin, which is found in plant foods, has a range of therapeutic applications. In order to examine the potential roles of luteolin in ovarian teratocarcinoma, the human ovarian teratocarcinoma cell line PA-1 was selected for functional experiments in vitro and in vivo. We demonstrated that luteolin inhibited the proliferation and colony formation of PA-1 cells in vitro. The flow cytometry results suggested that luteolin induced apoptosis of PA-1 cells in a dose-dependent manner. Immunofluorescence and qRT-PCR results showed that the expression of B-cell lymphoma-2 (Bcl-2) was decreased in luteolin-treated cells, whereas the expression of Bcl-2- associated X (Bax) was increased compared with that in the control group. In addition, luteolin inhibited the tumor growth of ovarian teratocarcinoma cells in a xenograft model. All the results suggested that luteolin induced cell apoptosis and inhibited tumor growth of PA-1 cells.     

Combined Hyperthermia and Radiation Therapy for Treatment of Hepatocellular Carcinoma

Background: There is no doubt that hyperthermia is one of the powerful radiosensitizers. Finding a proper mechanismworking in hyperthermia/radiation combination is still pronounced challenge. Objectives: This study is focusing onthe anti-cancer activities (anti-proliferative, anti-angiogenic and antiapoptotic) of thermoradiotherapy. Materials andMethods: Liver cancer cell line (HepG2) was treated by 37oC, 40oC and 43oC hyperthermia degrees combined withthree radiation doses (2 Gy, 4 Gy and 8 Gy) for 24, 48 and 72 hrs. Cell viability, apoptotic/necrotic cell screening,apoptotic (BAX and FasL) and antiapoptotic (BCL-2 and GRP78) genes, and pro-angiogenic mediators [vascularendothelial- (VEGF) and Platelet derived-growth factors (PDGF) ware investigated. Results: Our data showed that 40oCtemperature combined with 4 Gy radiation gives a significant decrease (p<0.05) in cell viability. Maximum cytotoxicitywas reported 48 hr post-treatment followed by slight restoration of cell viability after 72 hr. Compared with untreatedcells, only 5% of viable cells with a high percentage of apoptotic (31%) and necrotic (63%) cells were demonstratedin 40oC/4 Gy/48 hr group. Expression of pro-apoptotic genes (BAX and FasL) were increased after hyperthermia withapparent elevation in 40oC/4 Gy/48 hr group coincides with moderate expression of antiapoptotic BCL-2 and GRP78genes. A significant reduction (p<0.001; p<0.05) in VEGF and PDGF levels; respectively was shown at 40oC/4 Gy/48hr group. Conclusions: This pilot study proposed 40oC mild temperature hyperthermia as a favorable hyperthermalcondition with 4 Gy radiotherapy in HCC treatment. A further research has to be performed considering an applicationof more than one session of radiothermal therapy at 40oC/4 Gy for total abrogation of cancer cells.     

Evaluation of anticancer effects of a pharmaceutically viable extract of a traditional polyherbal mixture against non-small-cell lung cancer cells

ObjectiveThe present work tested organic solvents to prepare an extract with anticancer properties from a polyherbal mixture containing Nigella sativa (seeds), Hemidesmus indicus (roots) and Smilax glabra (rhizomes). We evaluate anticancer effects in non-small-cell lung cancer cells (NCI-H292), and discuss optimization for pharmaceutical use in the context of efficacy, yield and toxicity.MethodsUsing different organic solvents, six extracts were prepared from the polyherbal mixture. Based on the cytotoxic effects of these extracts on NCI-H292 cells and normal lung cells (MRC-5), as evaluated by the sulphorhodamine B assay, the total ethyl acetate (T-EA) extract was selected for further analysis. The possible anticancer mechanisms were assessed by evaluating the extract’s effects on apoptosis (through fluorescent microscopic analysis, DNA fragmentation analysis, caspase 3/7 assay and analysis of expression levels of apoptosis-related genes p53, Bax, survivin, Hsp70 and Hsp90), colony formation and antioxidant activity.ResultsThe extract had cytotoxic effects against NCI-H292 cells in a time- and dose-dependent manner. Significant antioxidant activity and inhibition of colony formation were also observed. The expression level of caspase 3/7 significantly (P < 0.001) increased in NCI-H292 cells treated with 50 μg/mL of the extract. The same dosage led to a significant increase in expression levels of Bax and p53 (P < 0.05 and P < 0.01 respectively), accompanied by a significant decrease (P < 0.0001) in survivin, Hsp70 and Hsp90.ConclusionT-EA extract of the above polyherbal mixture has cytotoxicity against NCI-H292 cells via induction of apoptosis, antioxidant effects and inhibition of colony formation.     

沉默信息调节因子过表达或谷氨酰胺剥夺对肾透明细胞癌细胞凋亡、增殖的影响

背景与目的：肾透明细胞癌（clear cell renal cell carcinoma，ccRCC）是最常见的肾癌类型，它与代谢密切相关。探讨沉默信息调节因子4（silent information regulator 4，SIRT4）过表达或谷氨酰胺（glutamine，Gln）剥夺对ccRCC细胞增殖、凋亡的影响。方法：慢病毒构建SIRT4和突变体H161Y过表达的Caki-2细胞株，利用无Gln的培养基来构建Gln剥夺模型，并通过体外增殖活力实验［细胞计数试剂盒-8（cell counting kit-8，CCK-8）］和克隆形成实验来分析两者对Caki-2细胞增殖和生长能力的影响；利用DCFH-DA荧光探针检测细胞内活性氧自由基（reactive oxygen species，ROS）水平进而评估Gln代谢对细胞ROS含量的影响；进一步通过线粒体膜电位检测、凋亡检测和蛋白质印迹法（Western blot）检测凋亡相关分子，分析SIRT4过表达以及Gln剥夺对Caki-2细胞凋亡的影响。结果：过表达SIRT4可抑制Gln代谢从而抑制Caki-2细胞增殖，另外还原性物质还原型烟酰胺腺嘌呤二核苷酸磷酸（reduced nicotinamide adenine dinucleotide phostate，NADPH）的生成减少能够增加细胞内ROS含量，促进细胞凋亡。而Gln剥夺抑制细胞增殖和促进细胞凋亡的效果均比过表达SIRT4明显，但长期缺乏Gln将导致细胞无法生长。结论：无论是过表达SIRT4还是Gln剥夺均能抑制ccRCC细胞增殖，促进凋亡。     

微小RNA-148a-3p靶向调控MAP3K9表达及对胃癌细胞增殖和凋亡的影响

目的 探讨微小RNA-148a-3p（miR-148a-3p）对丝裂原活化蛋白激酶激酶激酶9（MAP3K9）的靶向调控作用及对胃癌细胞增殖和凋亡的影响。方法 向对数生长期胃癌细胞株MGC-803转染miR-148a-3p模拟物（mimics组）和阴性对照（NC组），以未转染的MGC-803细胞为对照组；采用实时定量PCR（QPCR）检测各组miR-148a-3p水平以评价转染效率，MTT法检测各组细胞增殖能力，流式细胞术检测各组细胞凋亡情况，分别采用QPCR和Western blotting检测Bcl-2、Bax、caspase-3及MAP3K9 mRNA和蛋白水平，同时采用双荧光素酶报告实验验证miR-148a-3p与MAP3K9的靶向作用关系。结果 QPCR结果显示，对照组、NC组和mimics组的miR-148a-3p水平分别为1.021±0.123、1.087±0.196和2.854±0.368，与对照组和NC组比较，mimics组的miR-148a-3p水平升高（P＜0.05）。mimics组MGC-803细胞的增殖活力较其余两组减弱（P＜0.05）。mimics组MGC-803细胞凋亡率为（15.2±1.6）％，高于对照组的（3.5±0.9％）％和NC组的（4.5±1.1）％，差异具有统计学意义（P＜0.05）。与对照组和NC组比较，mimics组的MAP3K9和Bcl-2的mRNA和蛋白水平均下调，而Bax和caspase-3的mRNA和蛋白水平均上调（P＜0.05）；双荧光素酶报告实验证实MAP3K9是miR-148a-3p的直接作用靶点。结论 MiR-148a-3p可抑制胃癌细胞MGC-803的增殖并诱导其凋亡，可能通过靶向MAP3K9来发挥抑癌作用，调控miR-148a-3p／MAP3K9轴在胃癌防治中有一定应用前景。     

阿帕替尼联合曲妥珠单抗对胃癌细胞的协同增敏作用

目的观察阿帕替尼联合曲妥珠单抗杀伤胃癌NCI-N87细胞的协同增敏作用并探讨可能作用机制。方法CCK-8法检测空白对照组、曲妥珠单抗组(0.1、1、10μg/ml)、阿帕替尼组(1μmol/L)及曲妥珠单抗(0.1、1、10μg/ml)+阿帕替尼(1μmol/L)组对NCI-N87细胞的增殖抑制作用,流式细胞术检测NCI-N87细胞凋亡,Western blotting检测HER-2、VEGFR2、Bax、Bcl-2蛋白表达。结果CCK-8检测提示曲妥珠单抗、阿帕替尼能够抑制NCI-N87细胞增殖,在一定浓度范围内作用呈浓度依赖性和时间依赖性(P<0.01);q值计算提示曲妥珠单抗与阿帕替尼具有协同抑制NCI-N87细胞增殖的作用。流式细胞术检测显示联合组NCI-N87胃癌细胞凋亡较单药组明显升高(P<0.05),其中空白对照组、阿帕替尼组(1μmol/L)、曲妥珠单抗(0.1μg/ml)组及曲妥珠单抗(0.1μg/ml)+阿帕替尼(1μmol/L)组的凋亡率分别为(3.0±1.28)%、(5.8±1.63)%、(8.0±3.92)%和(21.6±6.85)%。曲妥珠单抗+阿帕替尼组与空白对照组、曲妥珠单抗及阿帕替尼单药组比较,HER-2蛋白表达显著下调(P<0.05);曲妥珠单抗+阿帕替尼组与空白对照组比较,Bcl-2蛋白表达、Bcl-2/Bax比值明显降低(P<0.01)。结论阿帕替尼联合曲妥珠单抗可能通过下调HER-2蛋白及调控凋亡相关蛋白Bcl-2、Bax的表达,协同抑制NCI-N87细胞增殖和促进细胞凋亡。     

乙酰紫草素通过PI3K/Akt信号通路诱导前列腺癌PC-3细胞凋亡的研究

目的：研究乙酰紫草素（ASK）对人前列腺癌PC-3细胞的诱导凋亡作用及相关的分子机制。方法：利用CCK-8实验检测ASK对体外培养PC-3细胞的杀伤作用；利用流式细胞术实验检测ASK处理后PC-3细胞凋亡情况；利用Western blotting检测凋亡蛋白和AKT信号通路蛋白的表达。结果：CCK-8实验证明ASK能够以时间和浓度依赖性抑制前列腺癌PC-3细胞的增殖；流式细胞术实验证明ASK能够诱导人前列腺癌PC-3细胞线粒体依赖性凋亡；Western blotting证明ASK能够有效抑制前列腺癌PC-3细胞中的AKT信号通路。结论：ASK能够有效抑制人前列腺癌PC-3细胞增殖，并能够有效诱导PC-3细胞发生线粒体依赖性凋亡，其机制可能是通过调控PI3K/Akt信号通路来实现，说明ASK具有一定的抗前列腺癌的潜力。