Down-regulation of histone deacetylase 7 reduces biological activities of retinal microvascular endothelial cells under high glucose condition and related mechanism

AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the retinal tissue in diabetic rats.METHODS:The expression of HDAC7 in HRMECs under high glucose and the retinal tissue from normal or diabetic rats were detected with immunohistochemistry and Western blot.LV-shHDAC7 HRMECs were used to study the effect of HDAC7 on cell activities.Cell count kit-8(CCK-8),5-ethynyl2’-deoxyuridine(EdU),flow cytometry,scratch test,Transwell test and tube formation assay were used to examine the ability of cell proliferation,migration,and angiogenesis.Finally,a preliminary exploration of its mechanism was performed by Western blot.RESULTS:The expression of HDAC7 was both upregulated in retinal tissues of diabetic rats and high glucosetreated HRMECs.Down-regulation of HDAC7 expression significantly reduced the ability of proliferation,migration,and tube formation,and reversed the high glucose-induced high expression of CDK1/Cyclin B1 and vascular endothelial growth factor in high glucose-treated HRMECs.CONCLUSION:High glucose can up-regulate the expression of HDAC7 in HRMECs.Down-regulation of HDAC7 can inhibit HRMECs activities.HDAC7 is proposed to be involved in pathogenesis of diabetic retinopathy and a therapeutic target.

Study on regulating mechanisms of oxocrebanine obtained from Stephania hainanensis H.S.Lo et Y.Tsoong on microtubule sites and tubulin in human breast cancer MCF-7 cells

Objective:To determine the destructive ability of oxocrebanine,an anti-breast cancer active compound obtained from Stephania hainanensis H.S.Lo et Y.Tsoong,on microtubule network,and investigate the effect of oxocrebanine on microtubule network homeostasis at both molecular and cellular levels.Methods:the EBI site competition method and molecular docking method were used to determine the occupation of the microtubule site of oxocrebanine.Western Blot was used to detect the effect of oxocrebanine on microtubule-associated proteins including STAT3,PAK1,CAMK4,and PKA.Results:The results of EBI site competition assay showed that the binding of EBI toβ-Tubulin covalent fusions produced adducts that appeared in regions of lower molecular weight thanβ-tubulin(ctrl 2).Molecular docking results showed that oxocrebanine could occupy the colchicine site of microtubule proteins.As revealed by Western Blot,the expression of STAT3 protein was decreased after MCF-7 cells have been treated with low,medium,and high concentration of oxocrebanine or the positive drug taxol for 48 h(P<0.01).The expression levels of PAK1 and Camk4 proteins aslo showed significant reductions(P<0.05,or P<0.01).Oxocrebanine also decreased the PKA protein in MCF-7 cells compared to the control group(P<0.01).Conclusions:Oxocrebanine,a ligand that binds at the colchicine site of tubulin,perturbs tubulin polymerization and causes mitosis in MCF-7 cells,thus leading to MCF-7 cell death.Oxocrebanine may promote microtubule dynamics through stathmin by inhibiting the expression levels of STAT3,PAK1,Camk4,and PKA proteins in MCF-7 cells.Oxocrebanine interfers with spindle formation,and ultimately causes mitotic catastrophe in MCF-7 cells.

非剥脱点阵激光联合侧柏叶酊对斑秃小鼠IL-7/IL-7Rα信号通路和Tregs细胞亚群的影响

目的:研究1565 nm非剥脱点阵激光联合侧柏叶酊(Platycladus orientalis tincture,POT)对斑秃(Alopecia areata,AA)小鼠治疗作用以及对白细胞介素7(Interleukin 7,IL-7)/白细胞介素7受体α(Interleukin-7 receptorα,IL-7Rα)信号通路和调节性T细胞(Regulatory T cells,Tregs)亚群的影响。方法:将50只成年雄性C3H/HeJ小鼠随机分为对照组(C组),模型组[M组,环磷酰胺(Cyclophosphamide,CTX)诱导AA模型],M+1565 nm组(1565 nm非剥脱点阵激光治疗AA),M+POT组(POT治疗AA)、M+1565 nm+POT组(1565 nm非剥脱点阵激光联合POT治疗AA),每组10只。流式细胞术检测C组和M组皮损组织中Tregs细胞亚群的比例和所有组血液中单个核细胞中Tregs细胞亚群的比例。Western blot法检测各组小鼠皮损组织中IL-7和IL-7Rα的表达。结果:与C组比,M组皮肤组织IL-7和IL-7Rα的表达均明显增加,而且Tregs细胞比例明显减少(P<0.05)。与M组比,M+1565 nm组和M+POT组IL-7的表达均降低(P<0.05)。与M组比,M+1565 nm+POT组IL-7和IL-7Rα的表达均降低,且Tregs细胞比例都显著增加(P<0.05)。结论:1565 nm非剥脱点阵激光联合POT治疗可以抑制斑秃小鼠IL-7/IL-7Rα信号并减少Tregs细胞的比例。

DHCR7基因在胃癌中的表达及其与免疫相关基因的关系

目的探究7-脱氢胆固醇还原酶(DHCR7)基因在胃癌中的表达及其与免疫相关基因的关系。方法使用UALCAN、TIMER数据库分析DHCR7基因在不同类型肿瘤中的表达情况。使用GEPIA、TCGA、GEO数据库分析DHCR7基因在胃癌中的表达。Kaplan-Meier Plotter数据库分析DHCR7基因表达与胃癌患者预后的相关性。通过cBioPortal数据库找出与DHCR7共表达的基因,并展示这些基因中相关系数比较高的基因。对胃癌中与DHCR7表达正负相关的基因进行基因本体分析(GO)和京都基因与基因组百科全书(KEGG)分析,首先分析DHCR7基因与CD8^(+)T细胞以及招募CD8^(+)T细胞相关趋化因子的相关性;其次分析DHCR7基因与免疫激活相关基因的相关性;最后分析DHCR7表达与胃癌患者临床病理学特征的关联。结果DHCR7基因在大多数肿瘤类型中均有高表达趋势。DHCR7基因在胃癌中的表达高于正常胃黏膜。Kaplan-Meier Plotter数据库中201790-s-at芯片结果显示高表达DHCR7组患者生存期较短。与DHCR7负相关基因的GO、KEGG通路富集分析主要富集在免疫相关功能和信号通路上,DHCR7与CD8^(+)T细胞、招募CD8^(+)T细胞相关的趋化因子以及免疫激活基因都具有负相关性。DHCR7高表达与胃癌患者年龄呈正相关。结论DHCR7在胃癌组织中呈高表达,高表达DHCR7患者预后不良,DHCR7基因能够影响胃癌免疫微环境,有望成为胃癌免疫治疗的新靶点。

EGFR突变NSCLC组织LncRNA TCF7L2表达及临床病理特征和预后分析

目的 探究长链非编码RNA(LncRNA)转录因子7类似物2(TCF7L2)在表皮生长因子受体基因(EGFR)突变的非小细胞肺癌(NSCLC)组织表达及其与病人临床病理特征和预后相关性。方法 选取2019年3月—2021年6月河北北方学院附属第一医院治疗的EGFR突变NSCLC病人104例为研究对象,分别取其癌组织和癌旁组织应用逆转录PCR(RT-PCR)检测LncRNA TCF7L2的表达,比较不同组织TCF7L2表达及其与临床病理特征和预后的相关性。结果 RT-PCR检测结果显示,癌组织中LncRNA TCF7L2表达量显著高于癌旁组织(t=12.410,P<0.05)。与LncRNA TCF7L2低表达病人比较,高表达者发生淋巴结转移更多、肿瘤体积更大(χ^(2)=4.579、7.762,P<0.05);LncRNA TCF7L2高表达病人6、9和12个月的生存率较低,但差异均无统计学意义(P>0.05)。结论 LncRNA TCF7L2在EGFR突变NSCLC病人癌组织表达高于癌旁组织,且TCF7L2高表达病人的淋巴结转移风险较高、肿瘤体积较大,其生存率可能较低。

SOX7靶向ERK1/2/PD-L1通路抑制结直肠癌血管生成

目的探讨性别决定区Y框蛋白7(SOX7)对结直肠癌血管生成的影响及潜在作用机制。方法应用免疫荧光检测结直肠癌患者组织样本中SOX7表达水平,之后通过裸鼠、转染SOX7 mimic的人结直肠癌细胞系SW480细胞和人脐静脉内皮细胞(HUVEC)共培养进一步研究。用Western-blot验证SOX7与ERK1/2/PD-L1对结直肠癌细胞的相关蛋白表达的影响。用CCK8检测SOX7与ERK1/2/PD-L1对HUVEC增殖的影响。通过体外内皮细胞成管实验测定SOX7与ERK1/2/PD-L1对肿瘤血管生成的影响。结果SOX7在人结直肠癌组织中表达被抑制(P<0.01),同时SOX7的过表达抑制了小鼠体内肿瘤生长(P<0.01)。SW480细胞中SOX7的过表达抑制了ERK1/2、c-Jun的表达,并在ERK1/2的激动剂Senkyunolide I的作用下上调了SW480细胞的ERK1/2、c-Jun蛋白表达(P<0.01),逆转了SOX7对SW480细胞中ERK1/2、c-Jun蛋白表达的影响(P<0.01)。HUVEC中SOX7抑制了PD-L1、V-EGFR2、p-PI3K、HIF-1α的蛋白表达,Senkyunolide I上调了HUVEC的PD-L1、V-EGFR2、p-PI3K、HIF-1α的蛋白表达,并逆转了SOX7对HUVEC中上述相关蛋白表达的影响(P<0.01)。PD-1/PD-L1 Inhibitor 3抑制了PD-L1、V-EGFR2、p-PI3K、HIF-1α的蛋白表达,SOX7过表达在PD-1/PD-L1 Inhibitor 3的影响下并没有表现出抑制作用。CCK8实验结果显示SOX7过表达显著抑制了HUVEC的增殖能力,Senkyunolide I作用下的两组HUVEC增殖能力较SOX7 NC组与SOX7 mimic组明显上升,PD-1/PD-L1 Inhibitor 3作用下的两组HUVEC增殖能力较SOX7 NC组与SOX7 mimic组明显下降,以上均有明显统计学差异(P<0.01)。成管实验结果显示SOX7过表达抑制了HUVEC的血管生成,Senkyunolide I强烈加速了血管生成,而PD-1/PD-L1 Inhibitor 3血管生成则被显著抑制,以上均有明显统计学差异(P<0.01)。结论SOX7通过ERK1/2/PD-L1通路抑制结直肠肿瘤的增殖和血管生成,SOX7可能是晚期CRC患者临床治疗中潜在的抗血管生成靶点。

雷公藤内酯醇通过调控miR-142-3p/HSP70通路抑制人乳腺癌MCF-7细胞增殖、侵袭和迁移

目的:探究雷公藤内酯醇(TP)通过miR-142-3p/HSP70信号通路对人乳腺癌MCF-7细胞恶性生物学行为的影响。方法:常规培养MCF-7细胞,将其分为6组:对照组、TP组、miR-142-3p inhibitor组、TP+inhibitor组、miR-142-3p mimic组和TP+mimic组,用转染试剂将相应的核酸或质粒转染MCF-7细胞。qPCR法、EdU细胞增殖实验、Transwell小室实验、细胞划痕实验、WB法分别检测转染后各组MCF-7细胞中miR-142-3p和HSP70 mRNA的表达,MCF-7细胞的增殖、侵袭、迁移能力和HSP70蛋白表达水平。结果:TP或miR-142-3p过表达能显著促进MCF-7细胞中miR-142-3p和HSP70的表达,敲减miR-142-3p则可明显抑制MCF-7细胞中miR-142-3p和HSP70的表达,TP可逆转由敲减miR-142-3p对MCF-7细胞中miR-142-3p和HSP70表达的影响;TP、过表达miR-142-3p均可明显抑制MCF-7细胞的增殖、迁移和侵袭能力(均P<0.05),敲减miR-142-3p则均可促进MCF-7细胞的增殖、迁移和侵袭能力(均P<0.05),TP可逆转由敲减miR-142-3p对MCF-7细胞恶性生物学行为的影响(均P<0.05)。结论:TP可通过调控miR-142-3p/HSP70信号通路,进而抑制MCF-7细胞的增殖、侵袭和迁移能力。

USP7-MDM2-p53信号轴对子宫内膜癌细胞增殖、凋亡和细胞周期的影响

目的:探讨泛素特异性蛋白酶7(USP7)调节Mdm2 p53结合蛋白同源物(MDM2)-p53轴对子宫内膜癌细胞增殖、凋亡和细胞周期的影响。方法:Western blot检测人子宫内膜癌组织、癌旁组织、人子宫内膜上皮细胞hEEC及人子宫内膜癌细胞系Ishikawa、HEC-1-A、KLE中USP7蛋白表达。将Ishikawa细胞分为NC组、P22077(USP7抑制剂)组、pcDNA组、pcDNA-MDM2组、P22077+pcDNA组、P22077+pcDNA-MDM2组,CCK-8法和克隆形成实验检测Ishikawa细胞增殖;流式细胞术检测Ishikawa细胞凋亡与细胞周期变化;Western blot检测Ishikawa细胞中USP7、细胞周期蛋白D1(CyclinD1)、周期素依赖性激酶2(CDK2)、Bcl-2相关X蛋白(Bax)、MDM2、p53蛋白表达。以RG7388(MDM2抑制剂)或PFT-α(p53抑制剂)与20μmol/L P22077共处理Ishikawa细胞48 h以验证USP7-MDM2-p53信号轴上下游关系。结果:USP7蛋白在子宫内膜癌组织和细胞中高表达,且Ishikawa细胞中USP7蛋白表达量最高,因此,选择Ishikawa细胞为研究对象。与NC组比较,P22077组Ishikawa细胞OD 450值、克隆形成率、S期和G 2/M期细胞数、USP7、CyclinD1、CDK2、MDM2蛋白表达降低,细胞凋亡率、G_(0)/G_(1)期细胞数、p53、Bax蛋白表达升高(P<0.05);与NC组、pcDNA组比较,pcDNA-MDM2组Ishikawa细胞OD 450值、克隆形成率、S期和G 2/M期细胞数、USP7、CyclinD1、CDK2、MDM2蛋白表达升高,细胞凋亡率、G_(0)/G_(1)期细胞数、p53、Bax蛋白表达降低(P<0.05);与P22077组、P22077+pcDNA组比较,P22077+pcDNA-MDM2组Ishikawa细胞OD 450值、克隆形成率、S期和G 2/M期细胞数、USP7、CyclinD1、CDK2、MDM2蛋白表达升高,细胞凋亡率、G_(0)/G_(1)期细胞数、p53、Bax蛋白表达降低(P<0.05)。p53为USP7-MDM2通路下游分子。结论:抑制USP7表达可能通过下调MDM2来激活p53进而抑制Ishikawa细胞增殖、促进细胞凋亡及周期停滞。

Neuroprotective effects of neural stem cells pretreated with neuregulin1β on PC12 cells exposed to oxygen-glucose deprivation/reoxygenation

Studies on ischemia/reperfusion(I/R)injury suggest that exogenous neural stem cells(NSCs)are ideal candidates for stem cell therapy reperfusion injury.However,NSCs are difficult to obtain owing to ethical limitations.In addition,the survival,differentiation,and proliferation rates of transplanted exogenous NSCs are low,which limit their clinical application.Our previous study showed that neuregulin1β(NRG1β)alleviated cerebral I/R injury in rats.In this study,we aimed to induce human umbilical cord mesenchymal stem cells into NSCs and investigate the improvement effect and mechanism of NSCs pretreated with 10 nM NRG1βon PC12 cells injured by oxygen-glucose deprivation/reoxygenation(OGD/R).Our results found that 5 and 10 nM NRG1βpromoted the generation and proliferation of NSCs.Co-culture of NSCs and PC12 cells under condition of OGD/R showed that pretreatment of NSCs with NRG1βimproved the level of reactive oxygen species,malondialdehyde,glutathione,superoxide dismutase,nicotinamide adenine dinucleotide phosphate,and nuclear factor erythroid 2-related factor 2(Nrf2)and mitochondrial damage in injured PC12 cells;these indexes are related to ferroptosis.Research has reported that p53 and solute carrier family 7 member 11(SLC7A11)play vital roles in ferroptosis caused by cerebral I/R injury.Our data show that the expression of p53 was increased and the level of glutathione peroxidase 4(GPX4)was decreased after RNA interference-mediated knockdown of SLC7A11 in PC12 cells,but this change was alleviated after co-culturing NSCs with damaged PC12 cells.These findings suggest that NSCs pretreated with NRG1βexhibited neuroprotective effects on PC12 cells subjected to OGD/R through influencing the level of ferroptosis regulated by p53/SLC7A11/GPX4 pathway.

Hesperidin as a preventive resistance agent in MCF-7 breast cancer cells line resistance to doxorubicin

Objective:To evaluate of hesperidin to overcome resistance of doxorubicin in MCF-7 resistant doxorubicin cells(MCF-7/Dox)in cytotoxicity apoptosis and P-glycoprotein(Pgp)expression in combination with doxorubicin.Methods:The cytotoxic properties.50%inhibition concentration(IC_(50))and its combination with doxorubicin in MCF-7 cell lines resistant to doxorubicin(MCF-7/Dox)cells were determined using MTT assay.Apoptosis induction was examined by double staining assay using ethidium bromide-acridine orange.Immunocytochemistry assay was performed to determine the level and localization of Pgp.Results:Single treatment of hesperidin showed cytotoxic activity on MCF-7/Dox cells with IC_(50)value of 11μmol/L.Thus,combination treatment from hesperidin and doxorubicin showed addictive and antagonist effect(CI>1.0).Hesperidin did not increase the apoptotic induction,but decreased the Pgp expressions level when combined with doxorubicin in low concentration.Conclusions:Hesperidin has cytotoxic effect on MCF-7/Dox cells with IC_(50)of 11μmol/L.Hesperidin did not increased the apoptotic induction combined with doxorubicin.Cochemotherapy application of doxorubicin and hesperidin on MCF-7/Dox cells showed synergism effect through inhibition of Pgp expression.

Triterpenoid of avocado (Persea americana) seed and its cytotoxic activity toward breast MCF-7 and liver HepG2 cancer cells

Objective:To determine the structure of triterpenoid isolated from avocado seeds and the cytotoxic effect on MCF-7 and Hep G2 cells.Methods:The powder sample was macerated with ethanol,followed with separation of the extract by column chromatography.The target compound was monitored on thin layer chromatography plate and reagent Lieberman–Buchard.The isolated compound was characterized by spectral analysis,mainly ultraviolet,infrared,and liquid chromatographymass spectroscopy and their spectroscopic data with those reported in literature were compared.In vitro cytotoxic activity was investigated against Vero,MCF-7,and Hep G2 cell lines using MTT assay.Results:A triterpenoid compound was isolated from ethanol extract.The extracts,fraction(F3),and the isolated compound showed a significant cytotoxic activity against all investigated cell lines.MTT assay showed that the triterpenoid isolate inhibited cell proliferation of MCF-7 and Hep G2 cell line with the IC50 values of 62 mg/m L and 12 mg/m L,respectively,and was safe to normal cells.Conclusions:The results of the present study reveal that triterpenoid from avocado seeds have the potential for further development as anticancer agents.

Fucoidan Induces G_1 Phase Arrest and Apoptosis through Caspases-dependent Pathway and ROS Induction in Human Breast Cancer MCF-7 Cells

Fucoidan is an active component of seaweed, which inhibits proliferation and induces apoptosis of several tumor cells while the detailed mechanisms underlying this process are still not clear. In this study, the effect of Fucoidan on the proliferation and apoptosis of human breast cancer MCF-7 cells and the molecular mechanism of Fucoidan action were investigated. Viable cell number of MCF-7 cells was decreased by Fucoidan treatment in a dose-dependent manner as measured by MTT assay. Fucoidan treatment resulted in G1 phase arrest of MCF-7 cells as revealed by flow cytometry, which was associated with the decrease in the gene expression of cyclin D 1 and CDK-4. Annexin V/PI staining results showed that the number of apoptotic cells was associated with regulation of cytochrome C, cas- pase-8, Bax and Bcl-2 at transcriptional and translational levels. Both morphologic observation and Hoechst 33258 assay results confirmed the pro-apoptotic effect of Fucoidan. Meanwhile, the ROS pro- duction was also increased by Fucoidan treatment, which suggested that Fucoidan induced oxidative damage in MCF-7 cells. The results of present study demonstrated that Fucoidan could induce GI phase arrest and apoptosis in MCF-7 cells through regulating the cell cycle and apoptosis-related genes or proteins expression, and ROS generation is also involved in these processes.

8-bromo-7-methoxychrysin inhibits properties of liver cancer stem cells via downregulation of β-catenin

AIM:To evaluate whether 8-bromo-7-methoxychrysin(BrMC),a synthetic analogue of chrysin,inhibits the properties of cancer stem cells derived from the human liver cancer MHCC97 cell line and to determine the potential mechanisms.METHODS:CD133+cells were sorted from the MHCC97 cell line by magnetic activated cell sorting,and amplified in stem cell-conditioned medium to obtain the enriched CD133+sphere forming cells(SFCs).The stem cell properties of CD133+SFCs were validated by the tumorsphere formation assay in vitro and the xenograft nude mouse model in vivo,and termed liver cancer stem cells(LCSCs).The effects of BrMC on LCSCs in vitro were evaluated by MTT assay,tumorsphere formation assay and transwell chamber assay.The effects of BrMC on LCSCs in vivo were determined using a primary and secondary xenograft model in Balb/c-nu mice.Expressions of the stem cell markers,epithelialmesenchymal transition(EMT)markers andβ-catenin protein were analyzed by western blotting or immunohistochemical analysis.RESULTS:CD133+SFCs exhibited stem-like cell properties of tumorsphere formation and tumorigenesis capacity in contrast to the parental MHCC97 cells.We found that BrMC preferentially inhibited proliferation and self-renewal of LCSCs(P<0.05).Furthermore,BrMC significantly suppressed EMT and invasion of LCSCs.Moreover,BrMC could efficaciously eliminate LCSCs in vivo.Interestingly,we showed that BrMC decreased the expression ofβ-catenin in LCSCs.Silencing ofβ-catenin by small interfering RNA could synergize the inhibition of self-renewal of LCSCs induced by BrMC,while Wnt3a treatment antagonized the inhibitory effects of BrMC.CONCLUSION:BrMC can inhibit the functions and characteristics of LCSCs derived from the liver cancer MHCC97 cell line through downregulation ofβ-catenin expression.

B7-H1 expression is associated with expansion of regulatory T cells in colorectal carcinoma

AIM： To investigate the expression of B7-H1 in human colorectal carcinoma （CRC） to define its regulating ef- fects on T cells in tumor microenvironment.

基质金属蛋白酶7在肝癌细胞迁移及免疫细胞浸润中的作用与预后价值

目的评估基质金属蛋白酶7(MMP7)在肝癌细胞迁移及免疫细胞浸润中的作用及预后价值。方法分别构建下调或上调靶基因MMP7的MMP7_siRNA、pMMP7转染肝癌细胞株(MHCC97H)。采用RT-qPCR、Western Blot分别检测细胞中靶基因mRNA及蛋白的表达水平。扫描电镜和Transwell小室实验分别观察细胞伪足和迁移能力的变化,并采用生物信息学的方法,在TCGA、TIMER数据库分析MMP7与免疫细胞及肝癌患者免疫浸润评分之间的相关性,并进一步研究MMP7与肝癌患者预后的相关性。相关性分析采用Spearman方法。Sanger Box在线工具评估MMP7在肝癌总体生存期、疾病特异性生存期方面的意义。Kaplan-Meier法绘制生存曲线,Log-rank检验评估不同组样本之间的预后差异。结果构建的MMP7_siRNA、pMMP7转染MHCC97H细胞后均能有效下调或上调靶基因MMP7的表达,在MHCC97H细胞中MMP7被干扰后细胞伪足明显减少,并且变短,但是过表达MMP7之后细胞表面丝状伪足数量明显增多,并且伪足长度变长,放射状排列。Transwell小室结果发现,MMP7_siRNA2能够显著降低细胞迁移能力(P<0.05),而转染pMMP7后,细胞迁移能力显著增加(P<0.05)。MMP7的表达与B淋巴细胞(r=0.37)、CD4+T淋巴细胞(r=0.40)、中性粒细胞(r=0.49)、巨噬细胞(r=0.49)、树突状细胞(r=0.47)显著相关(P值均<0.05)。在TCGA数据库中,基于总体生存期最佳截断值将肝癌患者分成MMP7高表达组(n=267)和MMP7低表达组(n=146),结果发现,MMP7高表达组的总体生存期明显低于MMP7低表达组(P<0.05);基于疾病特异性生存期最佳截断值将肝癌患者分成MMP7高表达组(n=257)和MMP7低表达组(n=145),结果发现,MMP7高表达组的疾病特异性生存期也低于MMP7低表达组(P<0.05)。结论MMP7促进了肝癌细胞的迁移,并在免疫细胞浸润中发挥主要作用,MMP7的表达也与肝癌的预后具有明显相关性。

布鲁氏菌病患者外周血中Siglec-7^(+)单核细胞与Th1/Th2细胞的相关性研究

目的:探讨布鲁氏菌病患者外周血中单核细胞的Siglec-7表达以及Siglec-7^(+)单核细胞与Th1/Th2细胞的相关性,分析Siglec-7分子在布鲁氏菌病患者中的临床意义。方法:纳入初诊为布鲁氏菌感染的患者50例(BI组),健康对照46例(正常对照组)。流式细胞术检测外周血单核细胞Siglec-7表达,分析Siglec-7^(+)单核细胞与布鲁氏菌病患者临床指标的相关性;流式细胞术检测Th1/Th2细胞水平,CBA法检测外周血清IFN-γ和IL-4含量,分析Siglec-7^(+)单核细胞与Th1/Th2水平、IFN-γ/IL-4含量的相关性。结果:BI组Siglec-7^(+)单核细胞水平较对照组明显升高(P<0.001);肝功能和肺X线异常的布鲁氏菌病患者体内Siglec-7^(+)单核细胞水平较两者正常的患者升高(P<0.001,P<0.05);Th2细胞水平和IL-4含量较对照组升高(P<0.05),而Th1细胞水平、IFN-γ含量与Th1/Th2、IFN-γ/IL-4较对照组明显降低(P<0.001);Siglec-7^(+)单核细胞水平与Th1细胞水平、IFN-γ含量呈负相关(r=-0.651,r=-0.407);与Th2细胞水平、IL-4含量呈正相关(r=0.706,r=0.530)。结论:在布鲁氏菌感染过程中,Siglec-7^(+)单核细胞增多并可能参与Th1/Th2细胞失衡。

Dietary Daidzein Enhances Antiapoptotic Effect of 17β-Estradiol (E_2) on Breast Cancer MCF-7 Cells

Objective： To investigate whether dietary daidzein interact with endogenous 17β-Estradiol （E2） to give rise to additive or inhibitory effects on proliferation and apoptosis in breast cancer cells. Methods： Cell cycle distribution and apoptosis induction were analyzed by using flow cytometry when breast cancer cell lines MCF-7 were cotreated with daidzein （1, 5 μmol/L） and E2 （0.1-10 nmol/L） for 5 days. Whether daidzein could alter E2-modulated mRNA expression of estrogen receptor alpha （ERα）, estrogen receptor beta （ERI3） and ERβ-estrogen response element （ERE） dependent transcription was investigated by RT-PCR and luciferase induction assays. The effects of daidzein on E2-modulated expression of proapoptotic p53, bax and antiapoptotic bcl-2 at both mRNA and protein levels were also investigated by RT-PCR and Western blot. Results： Daidzein enhanced the antiapoptotic effect in an Ea dose-dependent manner, but had no effect on E2-induced proliferation. Daidzein antagonized E2-induced ERβ mRNA expression and ERβ-ERE dependent transcription. In addition, daidzein only antagonized E2-upregulated expression of p53 and bax, but had no effect on E2-upregulated expression of bcl-2. Conclusion： Daidzein enhances the antiapoptotic effect of E2 on breast cancer cells by inhibiting E2-mediated p53-bax proapoptotic pathway. These results suggest that dietary daidzein may enhance deleterious effect of endogenous E2 in hormone-dependent breast cancer.

Cox7a2 mediates steroidogenesis in TM3 mouse Leydig cells

Aim： To investigate the regulatory function of Cox7a2 on steroidogenesis and the mechanism involved in TM3 mouse Leydig cells. Methods： The cDNA of Cox7a2 was cloned from TM3 mouse Leydig cells. It was subcloned to pDsRed- Express-N 1 and transfected back into TM3 mouse Leydig cells for Cox7a2 overexpression by transient gene transfection. Steroidogenesis affected by overexpressed Cox7a2 was studied by ELISA. To elicit the mechanism of this effect, expression of steroidogenic acute regulatory （STAR） protein and reactive oxygen species （ROS） were examined by Western blot and fluorometer, respectively. Results： The cDNA of Cox7a2 （249 bp） was cloned from Leydig cells and confirmed by DNA sequencing. After constructed pDsRed-Express-N1-Cox7a2 was transfected back into TM3 mouse Leydig cells, Cox7a2 inhibited not only luteinizing hormone （LH）-induced secretion of testosterone but also the expression of StAR protein. At the same time, Cox7a2 increased the activity of ROS in TM3 mouse Leydig cells. Conclusion： Cox7a2 inhibited LH-induced StAR protein expression, and consequent testosterone production, at least in part, by increasing ROS activity in TM3 mouse Leydig cells.

Roles of Smad3 and Smad7 in rat pancreatic stellate cells activated by transforming growth factor-beta 1

BACKGROUND: Pancreatic stellate cells (PSCs) play a major role in promoting pancreatic fibrosis. Transforming growth factor beta 1 (TGF-beta 1) is a critical mediator of this process. This study aimed to determine the expression of the Smad3 and Smad7 genes in the process of PSC activation, and explore the mechanisms of chronic pancreatitis. METHODS: The expressions of Smad3 and Smad7 in PSCs before and after TGF-beta 1 treatment were detected by reverse transcription-polymerase chain reaction and Western blotting analysis. Smad3 expression was detected in PSCs after treatment with 5 ng/ml of TGF-beta 1 for 24 hours. RESULTS: Smad7 expression was decreased in TGF-beta 1 -activated PSCs (P<0.05) in a dose-dependent manner. When TGF-beta 1 concentration reached 10 ng/ml, the expression of p-Smad3, Smad3, and Smad7 was inhibited (P<0.05). CONCLUSIONS: TGF-beta 1 promotes the expression of Smad3 and inhibits the expression of Smad7 during the activation of PSCs. In contrast, high-dose TGF-beta 1 downregulates the expression of Smad3 in completely activated PSCs.

Insulin-like growth factor binding protein-7 induces activation and transdifferentiation of hepatic stellate cells in vitro

AIM:To investigate the role of insulin-like growth factor binding protein-7 (IGFBP-7) in the activation and transdifferentiation of hepatic stellate cells (HSC) in vitro.METHODS:Rat HSC-T6 cells were cultured in separate dishes and treated with various concentration of transforming growth factor (TGF)-β1,IGFBP-7 or antiIGFBP-7 antibody for 24 h.The supernatant or a cytoplasm suspension was obtained from cultured HSC,followed by transfer of cells to form cell-coated dishes.Immunocytochemistry and Western blotting were used to analyze the expression of IGFBP-7 induced by TGF-β1 and the level of fibronectin,collagen and α-smooth muscle actin (SMA).The pro-apoptotic effect of antiIGFBP-7 antibody was determined by flow cytometry.RESULTS:Immunocytochemistry and Western blotting revealed that the expression of IGFBP-7 in TGF-β1 treated HSC was significantly up-regulated compared to that in the control group.In addition,fibronectin,collagen and α-SMA also showed enhanced expression in accordance with the transdifferentiation process in a dose-dependent manner to some extent.Moreover,flow cytometry suggested that anti-IGFBP-7 antibody induced apoptosis of activated HSC,which is responsible for the development of liver fibrosis,and may represent a novel pathway and target for therapeutic intervention.CONCLUSION:IGFBP-7 showed increased expression in activated HSC and played an important role in the activation and transdifferentiation process of HSC.AntiIGFBP-7 antibody may ameliorate liver fibrogenesis.