12-Lipoxygenases and 12(S)-HETE: role in cancer metastasis

Arachidonic acid metabolites have been implicated in multiple steps of carcinogenesis. Their role in tumor cell metastasis, the ultimate challenge for the treatment of cancer patients, are however not well-documented. Arachidonic acid is primarily metabolized through three pathways, i.e., cyclooxygenase, lipoxygenase, and P450-dependent monooxygenase. In this review we focus our attention on one specific lipoxygenase, i.e., 12-lipoxygenase, and its potential role in modulating the metastatic process. In mammalian cells there exist three types of 12-lipoxygenases which differ in tissue distribution, preferential substrates, and profile of their metabolites. Most of these 12-lipoxygenases have been cloned and sequenced, and the molecular and biochemical determinants responsible for catalysis of specific substrates characterized. Solid tumor cells express 12-lipoxygenase mRNA, possess 12-lipoxygenase protein, and biosynthesize 12(S)-HETE [12(S)-hydroxyeicosatetraenoic acid], as revealed by numerous experimental approaches. The ability of tumor cells to generate 12(S)-HETE is positively correlated to their metastatic potential. A large collection of experimental data suggest that 12(S)-HETE is a crucial intracellular signaling molecule that activates protein kinase C and mediates the biological functions of many growth factors and cytokines such as bFGF, PDGF, EGF, and AMF. 12(S)-HETE plays a pivotal role in multiple steps of the metastatic cascade encompassing tumor cell-vasculature interactions, tumor cell motility, proteolysis, invasion, and angiogenesis. The fact that 12-lipoxygenase is expressed in a wide diversity of tumor cell lines and 12(S)-HETE is a key modulatory molecule in metastasis provides the rationale for targeting these molecules in anti-cancer and anti-metastasis therapeutic protocols.     

CacyBP/SIP protein promotes proliferation and G1/S transition of human pancreatic cancer cells

Calcyclin-binding protein or Siah-1-interacting protein (CacyBP/SIP), a component of the ubiquitin-mediated proteolysis, could participate in beta-catenin degradation, which was found to be related to the malignant phenotypes of pancreatic cancer previously. However, the role of CacyBP/SIP itself in pancreatic cancer has not been investigated. In the present study, CacyBP/SIP expression was assayed and manipulated to reveal the potential mechanism in pancreatic cancer carcinogenesis. Here, we show that CacyBP/SIP is over-expressed in pancreatic cancer cells. Down-regulation of CacyBP/SIP by small interference RNA (siRNA) severely suppresses the proliferation and tumorigenesis in pancreatic cancer. G1/S transition arrest induced by inhibition of CacyBP/SIP is at least partly mediated by down-regulation of Cyclin E and CDK2 as well as up-regulation of p27 and Rb. Collectively, CacyBP/SIP as an enhancer of pancreatic cancer malignance might develop into another possible therapeutic target.     

Nestin phosphorylation at threonines 315 and 1299 correlates with proliferation and metastasis of human pancreatic cancer

The neuroepithelial stem cell marker nestin is a cytoskeletal protein that regulates cell proliferation, invasion, and stemness in various tumors, including pancreatic tumors. In the present study, we examined the expression and roles of phosphorylated nestin in pancreatic cancer cells. Nestin phosphorylation at threonines 315 (Thr315) and 1299 (Thr1299) was observed during mitosis in human pancreatic cancer cells. Nestin phosphorylation was positively correlated with a cell proliferation marker, MIB‐1 expression in human pancreatic cancer samples. Transfection of MIA PaCa‐2 cells with nestin mutated at Thr315 and/or Thr1299 (to suppress phosphorylation) resulted in lower proliferation rates than those in control groups. Transfecting MIA PaCa‐2 cells with wild‐type nestin or with nestin mutated at Thr315 increased migration and invasion. In contrast, transfection with nestin mutated at both phosphorylation sites (Thr315 and Thr1299) did not enhance cell migration or invasion. In an intra‐splenic xenograft experiment using MIA PaCa‐2 cells, tumors expressing the nestin double mutant formed fewer liver metastases than tumors expressing wild‐type nestin. Nestin phosphorylation at these two sites was decreased upon treatment with inhibitors for cyclin dependent kinases, AKT, and Aurora in PANC‐1 cells, which express a high baseline level of phosphorylated nestin. These findings suggest that phosphorylation of nestin at Thr315 and/or Thr1299 affects cell proliferation, and inhibition of both phosphorylation sites suppresses invasion and metastasis of human pancreatic cancer. Inhibiting nestin phosphorylation at these two sites may represent a novel therapeutic strategy for pancreatic cancer.     

Radiosensitivity of human prostate cancer cells can be modulated by inhibition of 12-lipoxygenase

Nearly 30% of prostate cancer (PCa) patients treated with potentially curative doses relapse at the sites of irradiation. How some tumor cells acquire radioresistance is poorly understood. The platelet-type 12-lipoxygenases (12-LOX)-mediated arachidonic acid metabolism is important in PCa progression. Here we show that 12-LOX confers radioresistance upon PCa cells. Treatment with 12-LOX inhibitors baicalein or BMD122 sensitizes PCa cells to radiation, without radiosensitizing normal cells. 12-LOX inhibitors and radiation, when combined, have super additive or synergistic inhibitory effects on the colony formation of both androgen-dependent LNCaP and androgen-independent PC-3 PCa cells. In vivo, the combination therapy significantly reduced tumor growth.     

The MEK/ERK pathway mediates COX-2-selective NSAID-induced apoptosis and induced COX-2 protein expression in colorectal carcinoma cells

Nonsteroidal antiinflammatory drugs (NSAIDs) can prevent colorectal tumorigenesis in humans and in rodents. In vitro and in vivo studies indicate that one of their principal antineoplastic avenues is the induction of apoptosis. We have shown previously that NS-398, which selectively inhibits cyclooxygenase-2 (COX-2) over cyclooxygenase-1, induces apoptosis of colorectal tumour cells and elevates COX-2 protein expression. Here, we have determined that the extracellular signal-regulated kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway mediates these effects of NS-398. Treatment of HT29 colorectal carcinoma cells with 75 microM NS-398 caused activation of ERK-1/-2 but not of the p38 and c-Jun N-terminal kinase (JNK) mitogen-activated protein kinases. This was apparent at 24 hr and maintained at 72 hr. U0126, a specific inhibitor of the ERK-activating kinases MEK-1/-2, prevented the activation of ERK induced by NS-398 and blocked the increase in COX-2 protein expression seen when HT29 cells were treated with NS-398 alone. The activation of ERK by NS-398 preceded and accompanied a decrease in attached cell yield and an increase in apoptosis. U0126 dose-dependently protected HT29 cells from these antiproliferative effects of NS-398, indicating an antiproliferative role for sustained ERK-1/-2 activation in response to this NSAID. These results point to a key role for the MEK/ERK signalling pathway in mediating the effects of a COX-2-selective NSAID on colorectal carcinoma cells.     

Decreased mitogenic response to epidermal growth factor in human squamous cell carcinoma lines overexpressing epidermal growth factor receptor owing to limiting amounts of the adaptor protein Grb2: Rescue by retinoic acid treatment

Growth factor receptors of the tyrosine kinase family regulate proliferation of a variety of cell types. In some human cancers, the epidermal growth factor receptor (EGFR) and its ligands often are overexpressed, leading to both constitutive and autocrine activation. Intracellular signaling via this receptor takes place through several mechanisms of action, including activation of ras and the mitogen‐activated protein kinase (MAPK) pathway. Our previous studies have shown that human squamous cell carcinoma (SCC) lines overexpress EGFR and do not increase proliferation in response to exogenous epidermal growth factor (EGF). The vitamin A metabolite retinoic acid (RA) has been used as a chemotherapeutic drug in the treatment of SCC. RA decreases proliferation of SCC lines, in part owing to inhibition of EGFR expression. However, we previously found that treatment of SCC lines with inhibitory doses of RA sensitized cells to the proliferative effects of EGF. We now present a mechanism of action for this effect. RA inhibited expression of EGFR and proteins in the MAPK signaling pathway. Expression of these molecules returned to basal levels within 24 h after RA withdrawal. RA also inhibited autocrine secretion of EGF, which returned to basal levels with slower kinetics. During this time, addition of exogenous EGF stimulated mitosis in SCC lines. These data suggested that signaling proteins downstream of overexpressed EGFR may have limited the mitotic response in SCC lines. In support of this hypothesis, overexpression of the EGFR adaptor protein Grb2 increased cell proliferation and restored EGF‐induced mitosis. © 2001 Wiley‐Liss, Inc.     

Influence of epidermal growth factor receptor (EGFR) gene mutations on the expression of EGFR, phosphoryl-Akt, and phosphoryl-MAPK, and on the prognosis of patients with non-small cell lung cancer

BACKGROUND AND OBJECTIVES: In this paper we examined the influence of epidermal growth factor receptor (EGFR) gene mutations on EGFR expression, downstream mediators, and survival in patients with non-small cell lung cancer (NSCLC). METHODS: We retrospectively analyzed the tumors of 53 patients with completely resected pathological stage I-IIIA NSCLC for the presence of EGFR gene mutations, the expression of EGFR mRNA and protein, phosphoryl-Akt, and phosphoryl-mitogen-activated protein kinase (MAPK) using immunostaining, and patients' prognosis. RESULTS: EGFR mutations were associated with elevations in EGFR mRNA (P = 0.004) and protein (P = 0.029) expression, but not with the expression of phosphoryl-Akt or phosphoryl-MAPK. The 5-year survival rate for all patients who exhibited an EGFR mutation was similar to those who were free of such mutations (71% vs. 56%, P = 0.252). However, the 5-year survival rate of patients with either a stage I adenocarcinoma or large cell carcinoma who had an EGFR mutation was significantly greater than for those who did not have such a mutation (92% vs. 57%, P = 0.037). CONCLUSIONS: EGFR gene mutations were significantly associated with higher EGFR expression, but not with p-Akt or p-MAPK status. In early stage NSCLC, the presence of an EGFR gene mutation bode well for the patient's prognosis.     

Radiation stimulates HGF receptor/c-Met expression that leads to amplifying cellular response to HGF stimulation via upregulated receptor tyrosine phosphorylation and MAP kinase activity in pancreatic cancer cells

Hepatocyte growth factor (HGF) is a stromal-derived cytokine that plays a crucial role in invasion and metastasis of tumor cells through the interaction with HGF receptor, c-Met, which is frequently overexpressed in pancreatic cancer. The present study was designed to investigate the change in HGF receptor and HGF-mediated signaling after irradiation in pancreatic cancer cells. Six cell lines from human pancreatic cancer were included in the study. Gamma-radiation was used for irradiation treatment. The changes in expression levels of c-Met were evaluated by immunoblot and confirmed morphologically by indirect immunofluorescence staining. Whether the resultant alteration in c-Met would cascade as biologically usable signals upon HGF ligation was traced by receptor tyrosine phosphorylation analysis and mitogen activated protein kinase (MAP kinase or MAPK) activity assay. The various biological responses to HGF (including cell proliferation, cell scattering, migration and invasion) were evaluated as well. We also used a 4-kringle antagonist of HGF, NK4, to block the HGF/c-Met signaling pathway. Both immunoblot and immunofluorescent analysis showed moderate increased expression of c-Met in 3 of 6 pancreatic cancer cell lines after irradiation. The actions seemed to be dose-responsible, which began at 3 hr and reached its peak value at 24 hr following irradiation. The radiation-increased expression of c-Met could transform into magnifying receptor tyrosine phosphorylation reaction and MAP kinase activity once the ligand was added, fairly corresponding with alteration in the receptor. Sequentially, the cellular responses to HGF, including scattering and invasion but not proliferation, were enhanced. Also, in the presence of HGF, the elevated receptor could help to recover the radiation-compromised cell migration. A recombinant HGF antagonist, NK4 could effectively block these aberrant effects activated by irradiation both in molecular and cellular levels, thus suggesting the deep involvement of the c-Met/HGF pathway in the enhanced malignant potential after irradiation. These results suggest that radiation may promote HGF-induced malignant biological behaviors of certain pancreatic cancer cells through the up-regulated HGF/c-Met signal pathway. Selectively targeted blockade of the HGF/c-Met pathway could help to abolish the enforced malignant behavior of tumor cells by irradiation and therefore may improve the efficacy of radiotherapy for pancreatic cancer.     

MAP3K9在肺癌组织中的表达及其对肺癌细胞增殖、迁移、侵袭的影响

目的:检测MAP3K9在临床肺癌样本及肺癌细胞系中的表达,研究MAP3K9对肺癌细胞增殖、迁移、侵袭能力的影响,并结合受试者工作特征曲线（ROC曲线）评估其诊断价值。方法:从云南省肿瘤医院收集2018年09月至2019年12月经病理诊断确诊为肺癌患者的癌组织及其对应的癌旁组织。MAP3K9在肺癌组织及癌旁组织、肺癌细胞系A549和H1299、人正常肺上皮细胞系BEAS-2B中的mRNA表达量采用实时荧光定量PCR技术检测。MAP3K9在细胞系中的蛋白表达量采用蛋白质印迹技术检测。分别用CCK8、划痕和Transwell实验检测细胞的增殖、迁移、侵袭能力。利用在线生存数据库分析表达MAP3K9患者的生存率。使用SPSS软件绘制ROC曲线。结果:MAP3K9在肺癌组织中的表达量显著高于癌旁组织（P＜0.05）。MAP3K9在A549和H1299中的表达量均高于BEAS-2B（P均<0.05）。体外细胞实验表明敲低MAP3K9能够明显抑制A549、H1299细胞的增殖、迁移和侵袭（P均<0.05）。生存曲线显示高表达MAP3K9的肺癌患者生存率低（P＜0.05）。ROC曲线下面积为0.894（P＜0.05）,诊断临界值为1.36×10-3,诊断敏感度为71.2%,特异度为98.1%。结论:MAP3K9在临床肺癌样本及肺癌细胞系中高表达,该基因高表达能够促进肺癌细胞的增殖、迁移和侵袭,与患者预后不良有关,MAP3K9作为一个致癌基因有潜力成为临床诊断和治疗肺癌的有效靶点。     

Protease-activated receptor-2 regulates cell proliferation and enhances cyclooxygenase-2 mRNA expression in human pancreatic cancer cells

BACKGROUND AND OBJECTIVES: Protease-activated receptor-2 (PAR-2) is a G protein-coupled receptor that is activated by trypsin. Recent studies have suggested that PAR-2 activity correlates with inflammatory processes and cell proliferation and that PAR-2 activation in non-neoplastic cells induces expression of cyclooxygenase-2 (COX-2). In the present study, we examined whether PAR-2 activation regulates cell proliferation and COX-2 expression by pancreatic cancer cells. METHODS: We analyzed PAR-2 expression immunohistochemically in 40 intraductal papillary-mucinous neoplasms (IPMNs) and 73 invasive ductal carcinomas (IDCs) of the pancreas. We used four pancreatic cancer cell lines (Panc1, T3M4, BxPC3, and MIApaca2) to measure cell proliferation and COX-2 mRNA expression after PAR-2 activation. RESULTS: PAR-2 protein was detected immunohistochemically in 85.0% of IPMNs and 65.8% of IDCs. Trypsin and a PAR-2 agonist peptide, SLIGKV, stimulated proliferation of each cell line in a dose-dependent manner. Exposure of cells to anti-PAR-2 neutralizing antibody prior to PAR-2 activation suppressed cell proliferation. In COX-2-positive cell lines (T3M4 and BxPC3), PAR-2 activation significantly increased COX-2 mRNA expression. CONCLUSIONS: Our results suggest that PAR-2 activation is associated with cell proliferation and COX-2 expression in pancreatic cancer cells. Blockade of the PAR-2 signaling pathway may be a novel strategy for suppressing pancreatic tumor growth.     

过表达RSK4m1对人乳腺癌细胞MDA-MB-231生长和侵袭的影响

目的 探讨过表达p90核糖体S6蛋白激酶4变异体1（ribosomal protein S6 kinase 4 variant 1,RSK4m1）对人乳腺癌MDA-MB-231细胞生长和侵袭的影响。方法 通过负载RSK4m1慢病毒在人乳腺癌MDA-MB-231细胞中稳定过表达RSK4m1。以MDA-MB-231亲本细胞为空白对照组（Con组）、转染空载病毒的MDA-MB-231细胞为阴性对照组（Mock组）、转染过表达RSK4m1的MDA-MB-231细胞为实验组（OE组）。采用qRT-PCR和Westen Blot法检测各组RSK4m1 mRNA及蛋白的表达;CCK-8法检测细胞增殖情况;细胞划痕实验检测细胞的迁移能力;Transwell小室侵袭实验检测细胞的侵袭能力。结果 OE组中RSK4m1 mRNA及蛋白的表达量均明显高于Con组及Mock组（P<0.05）。CCK-8实验显示,24 h、48 h、72 h和96 h时OE组细胞增殖均低于Mock组和Con组（P<0.05）;细胞划痕实验显示,细胞培养24 h后,OE组细胞迁移率显著低于Con组和Mock组［（14.53±0.64）% vs （25.67±2.44）%、（24.47±2.25）%,P<0.05］。Transwell小室侵袭实验显示,OE组细胞侵袭能力显著低于Con组和Mock组［（35.00±5.57）个 vs （66.70±5.86）个、（58.67±4.16）个,P<0.05］。结论 过表达RSK4m1可显著抑制人乳腺癌MDA-MB-231细胞增殖、迁移和侵袭能力,为RSK4m1可能成为乳腺癌的治疗新靶点提供了实验依据。     

Demethylzeylasteral (ZST93) inhibits cell growth and enhances cell chemosensitivity to gemcitabine in human pancreatic cancer cells via apoptotic and autophagic pathways

The overall 5‐year survival rate of patients with human pancreatic cancer remains less than 8% because of its aggressive growth, early metastasis and resistance to conventional chemoradiotherapy. It is essential to develop innovative and effective therapeutic agents to improve its prognosis. Demethylzeylasteral (ZST93) is a novel triterpenoid monomer extracted from the xylem of Tripterygium roots. Our study aimed to assess the effects of ZST93 on cell proliferation and its role in the chemosensitivity to gemcitabine in human pancreatic cancer cells. The effects of ZST93 on cancer cell proliferation, cell cycle distribution, apoptosis and autophagy were evaluated in various human pancreatic cancer cell lines, and the antitumor effects of ZST93 alone and in combination with gemcitabine were identified in a xenograft mouse model. The results showed that ZST93 could inhibit the proliferation of pancreatic cancer cells and arrest cell cycle at G0/G1 phase by regulating the expression of Cyclin D1 and Cyclin A2. Moreover, ZST93 killed pancreatic cancer cells through two different mechanisms: inducing autophagic cell death at low concentrations and apoptotic cell death at high concentrations. Furthermore, ZST93 could enhance the chemosensitivity of pancreatic cancer cells to gemcitabine both in vitro and in vivo through modulation of the cross talk between autophagy and apoptosis. ZST93 is a potential therapeutic agent for developing novel therapeutic strategies in human pancreatic cancer.     

PTPN12与非小细胞肺癌放射敏感性的关系研究

目的:探讨非小细胞肺癌（non-small cell lung cancer ,NSCLC ）组织中非受体型蛋白酪氨酸磷酸酶12（protein tyrosine phosphatase non-receptor type 12,PTPN12）表达与放疗疗效的关系,评估PTPN12基因下调与H 1299细胞放射敏感性的关系。方法:回顾性分析2013年9 月至2014年10月河北医科大学第四医院92例接受根治性放疗的NSCLC 患者的临床资料,通过免疫组织化学方法检测肿瘤组织中PTPN12的表达,并分析其与患者放疗疗效的关系。通过shRNA-PTPN 12干扰质粒转染H 1299细胞,利用实时定量PCR 技术及Westernblot技术检测PTPN12在mRNA 及蛋白水平的表达。通过克隆形成实验构建细胞生存曲线,评估PTPN12基因下调对H 1299细胞放射敏感性的作用。结果:PTPN12低表达患者的放疗疗效明显优于高表达者（80.0% vs . 57.1% ,P = 0.018）。 多因素分析显示PTPN12表达是影响放疗疗效的唯一因素。构建shRNA-GFP 及shRNA-PTPN 12干扰质粒转染H 1299细胞,获得稳定转染细胞株H 1299-shGFP及H 1299-shPTPN 12。细胞生存曲线显示H 1299-shGFP及H 1299-shPTPN 12细胞的D 0、Dq、SF2 值分别为3.02Gy、3.41Gy、0.94及1.81Gy、2.40Gy、0.72,具有显著性差异（P < 0.05）。 结论:PTPN12低表达与NSCLC 患者的放疗疗效密切相关,而PTPN12基因下调能明显增加H 1299细胞的放射敏感性。      

ERK通路可通过调节mdr-1和RRM1基因的表达参与诱导胰腺癌细胞株SW1990的吉西他滨化疗耐药

Objective: To investigate the relationship between extracellular signal-regulated kinase (ERK) pathway, multidrug resistance gene (mdr-1), ribonucleotide reductase M1 (RRM1) gene and their roles in gemcitabine (GEM) chemoresistance in pancreatic cancer cell line SW1990. Methods: The GEM-resistance cell model was constructed by a stepwise method. Immunohistochemistry was used to measure the expression of ERK protein (ERK1/2) in the established cell strains in a semiquantitative way. The mRNA expression of mdr-1 and RRM1 were detected by RT-PCR. MTT assay was performed to determine the IC50 value. Results: The established GEM-resistant cell strains were able to gain stable growth and passage ability in the medium contained different concentration levels of GEM (0, 30, 60, 100, 150 and 200 nmol/L). The expression of ERK protein, mdr-1 and RRM1 gene were elevated accompanied by the increase of GEM concentration. There was a highly positive correlation between mdr-1, RRM1 expression and GEM-resistance level (r = 0.960, P = 0.002 and r = 0.966, P = 0.002). The expression of ERK protein also correlated with the mdr-1 and RRM1 level (r = -0.943, P = 0.005 and r = -0.883, P = 0.02). At the GEM-resistance level of 200 nmol/L, the grey scale value of ERK1/2 was 164.22 ± 13.17, mdr-1/βactin and RRM1/β-actin were 1.41 ± 0.04 and 1.45 ± 0.18, respectively; after treated with ERK pathway inhibitor U0126, these values synchronously decreased to 186.85 ± 13.14, 0.2 3 ± 0.02 and 0.21 ± 0.03, respectively; inversely, the ERK1/2 grey scale value was 106.55 ± 16.45, mdr-1/βactin and RRM1/β-actin were 1.50 ± 0.07 and 1.52 ± 0.12, respectively, which presented a tendency of synchronously increase after treated with ERK pathway activator EGF. The IC50 values in GEM-resistant cells of 0 nmol/L and 200 nmol/L levels were 4.104 and 10.20, respectively. After treated with U0126, these values decreased to 3.26 and 4.50, respectively; while treated with EGF, the IC50 values increased to 8.89 and 17.17, respectively. Conclusion: The ERK pathway may induce the GEM-chemoresistance in pancreatic cell line SW1990 through the participation in the regulation of the mdr-1 and RRM1 gene expression.     

PARP14 promotes the proliferation and gemcitabine chemoresistance of pancreatic cancer cells through activation of NF‐κB pathway

Pancreatic cancer (PC) is the most fatal gastrointestinal malignancy in the world, with a 5‐year relative survival of only 8%. Poly(ADP‐ribose) polymerase (PARP)14, a member of the macro‐PARP subfamily proteins, has been reported to participate in various biologic and pathologic processes in multiple cancers. The roles and underlying molecular mechanisms of PARP14 in PC carcinogenesis, however, remain to be elucidated. In this study, we for the first time discovered that PARP14 was highly expressed in human primary PC specimens and significantly correlated with poor patient prognosis. Using loss‐of‐function studies in vitro and in vivo, we showed that the knockdown of PARP14 led to enhanced apoptosis, repressed proliferation, and gemcitabine (GEM) resistance of PC cells. Further investigations revealed that PARP14 was significantly overexpressed in GEM‐resistant PC cells (SW1990/GZ). And silencing of PARP14 significantly reversed the GEM resistance of SW1990/GZ cells. To the mechanism, PARP14 could stimulate PC progression by the activation of nuclear factor‐κB (NF‐κB) signaling pathway. And inhibition of NF‐κB signal could significantly reverse PARP14–overexpression triggered PC carcinogenesis. In conclusion, PARP14 could promote PC cell proliferation, antiapoptosis, and GEM resistance via NF‐κB signaling pathway, highlighting its potential role as a therapeutic target for PC.     

蛋白激酶SGK3过表达对乳腺癌细胞株MDA-MB-231影响的研究

目的 血清和糖皮质激素诱导的蛋白激酶3 (serum and glucocorticoid-inducible protein kinase 3,SGK3)参与细胞增殖、分化和物质转运等生命活动,在某些肿瘤形成中担任重要角色.本研究设计观察SGK3过表达对乳腺癌细胞株MDA-MB-231细胞增殖和存活的影响,并初步探讨其作用的分子机制.方法 免疫组化方法检测乳腺癌组织中SGK3蛋白的表达;通过GenEscortTMⅠ介导用重组质粒pEGFPN1-SGK3瞬时转染乳腺癌MDA-MB-231细胞,荧光显微镜和蛋白质印迹法鉴定转染细胞中融合蛋白SGK3-GFP的表达;细胞增殖实验观察转染细胞的生长增殖情况;流式细胞术分析转染细胞的细胞周期时相变化;蛋白质印迹方法检测细胞增殖与存活相关基因的表达.结果 SGK3在乳腺癌组织中的表达量明显高于癌旁组织和正常乳腺组织(F=29.672,P＜0.001),与二者组间比较,均P＜0.01.荧光显微镜下可见转染细胞因外源基因的表达而发出绿色荧光,且蛋白质印迹法检测可见融合蛋白SGK3-GFP的表达;过表达SGK3的MDA-MB-231细胞,其细胞生长速度加快,细胞倍增时间缩短(F=19.557,P-0.001),与亲本细胞及转染空载体细胞组间比较均P＜0.01.与亲本细胞和转染空载体组比较,SGK3过表达使细胞凋亡比例明显下降(F=12.156,P=0.008),与前两者组间比较P值分别为0.019和0.030;细胞内源性Bad的表达量降低(F=11.152,P=0.010),与前两者组间比较P值分别为0.008和0.005;而Bcl-xL(F=8.810,P=0.016,与前两者组间比较P值分别为0.014和0.009)以及p-GSK3β(F=42.253,P＜0.001,与前两者组间比较P值分别为＜0.001和0.009)的表达量均升高.结论 乳腺癌组织高表达SGK3,SGK3过表达可促进MDA-MB-231细胞的增殖与存活,影响细胞增殖与存活相关基因的表达可能是其发挥作用的主要分子机制.