Angiopoietin 1 reduces rat follicular atresia mediated by apoptosis through the PI3K/Akt pathway

Elevated fatty acid levels have been thought to contribute to insulin resistance. Repression of the glucose transporter 4 (GLUT4) gene as well as impaired GLUT4 translocation may be a mediator for fatty acid-induced insulin resistance. This study was initiated to determine whether palmitate treatment repressed GLUT4 expression, whether glucose/fatty acid metabolism influenced palmitate-induced GLUT4 gene repression (PIGR), and whether attempts to prevent PIGR restored palmitate-induced impairment of glucose uptake (PIIGU) in C2 myotubes. Not only stimulators of fatty acid oxidation, such as bezafibrate, AICAR, and TOFA, but also TCA cycle substrates, such as pyruvate, leucine/glutamine, and α-ketoisocaproate/monomethyl succinate, significantly prevented PIGR. In particular, supplementing with pyruvate through methyl pyruvate resulted in nearly complete prevention of PIIGU, whereas palmitate treatment reduced the intracellular pyruvate level. These results suggest that pyruvate depletion plays a critical role in PIGR and PIIGU; thus, pyruvate supplementation may help prevent obesity-induced insulin resistance in muscle cells.     

Melatonin induces apoptosis of colorectal cancer cells through HDAC4 nuclear import mediated by CaMKII inactivation

Melatonin induces apoptosis in many different cancer cell lines, including colorectal cancer. However, the precise mechanisms involved remain largely unresolved. In this study, we provide evidence to reveal a new mechanism by which melatonin induces apoptosis of colorectal cancer LoVo cells. Melatonin at pharmacological concentrations significantly suppressed cell proliferation and induced apoptosis in a dose‐dependent manner. The observed apoptosis was accompanied by the melatonin‐induced dephosphorylation and nuclear import of histone deacetylase 4 (HDAC4). Pretreatment with a HDAC4‐specific siRNA effectively attenuated the melatonin‐induced apoptosis, indicating that nuclear localization of HDAC4 is required for melatonin‐induced apoptosis. Moreover, constitutively active Ca²⁺/calmodulin‐dependent protein kinase II alpha (CaMKIIα) abrogated the melatonin‐induced HDAC4 nuclear import and apoptosis of LoVo cells. Furthermore, melatonin decreased H3 acetylation on bcl‐2 promoter, leading to a reduction of bcl‐2 expression, whereas constitutively active CaMKIIα(T286D) or HDAC4‐specific siRNA abrogated the effect of melatonin. In conclusion, the present study provides evidence that melatonin‐induced apoptosis in colorectal cancer LoVo cells largely depends on the nuclear import of HDAC4 and subsequent H3 deacetylation via the inactivation of CaMKIIα.     

COX-2 inhibits Fas-mediated apoptosis in cholangiocarcinoma cells

Fas expression has been shown to negatively regulate the progression of cholangiocarcinoma cells in xenografts. However, many human cholangiocarcinomas express Fas, suggesting these cancers have developed mechanisms to inhibit Fas-mediated apoptosis. Cyclooxygenase-2 (COX-2), which generates prostanoids, is expressed by many cholangiocarcinomas. Therefore, our aim was to determine whether COX-2 expression inhibits death receptor--mediated apoptosis in KMBC cells, a cholangiocarcinoma cell line. These cells express messenger RNA for the death receptors Fas, tumor necrosis factor receptor 1 (TNF-R1), death receptor 4 (DR4), and DR5. Agonists for these death receptors, CH-11, TNF-alpha, and TRAIL all induced apoptosis. However, COX-2, whether induced by proinflammatory cytokines or transient transfection, only significantly inhibited Fas-mediated apoptosis. The COX-2 inhibitor NS-398 restored Fas-mediated apoptosis in COX-2 transfected cells. Prostaglandin E2 reduced apoptosis and mitochondrial depolarization after treatment with the Fas agonist CH-11. Of a variety of antiapoptotic proteins examined, COX-2/prostaglandin E2 only increased expression of Mcl-1, an antiapoptotic member of the Bcl-2 family. In conclusion, these data suggest that prostanoid generation by COX-2 specifically inhibits Fas-mediated apoptosis, likely by up-regulating Mcl-1 expression. Pharmacologic inhibition of COX-2 may be useful in augmenting Fas-mediated apoptosis of cholangiocarcinoma cells.     

Bax translocation and mitochondrial fragmentation induced by Helicobacter pylori

BACKGROUND AND AIMS: Previous in vitro and in vivo studies have revealed an association between Helicobacter pylori infection and apoptosis in gastric epithelial cells. Although involvement of the Bcl-2 family of proteins as well as cytochrome c release has been demonstrated in H pylori induced cell death, the exact role of the mitochondria during this type of programmed cell death has not been fully elucidated. Therefore, we sought to determine whether or not Bax translocation and mitochondrial fragmentation occur on exposure of gastric epithelial cells to H pylori, resulting in cell death. METHODS: Experiments were performed with human gastric adenocarcinoma (AGS) cells, AGS cells transfected with the HPV-E6 gene (which inactivates p53 function), AGS-neo cells (transfected with the backbone construct), mouse embryonic fibroblasts (MEFs), and p19(ARF) null (ARF(-/-)) MEFs. Cells were incubated with a cag positive H pylori strain for up to 24 hours, lysed, and cytoplasmic and mitochondrial membrane fractions were analysed by western blot for Bax translocation. RESULTS: Bax translocation was detected in AGS, AGS-neo, and normal MEF cells after exposure to H pylori for three hours, but not in ARF(-/-) MEFs cells. Translocation of Bax after H pylori incubation was also detected in AGS-E6 cells (inactive p53 gene) but to a lesser degree than in AGS-neo cells. In parallel studies, the mitochondrial morphology of living cells infected with H pylori was assessed by confocal microscopy. Mitochondrial fragmentation was detectable after 10 hours of H pylori incubation with AGS cells and after seven hours with MEF cells. In wild-type MEFs, mitochondrial fragmentation was significantly increased in comparison with ARF null MEFs (43% v 10.4%, respectively). Furthermore, mitochondrial depolarisation and caspase-3 activity were initiated within four hours in cells incubated with H pylori, and these events were inhibited by forced expression of Bcl-2. CONCLUSIONS: These data suggest that during H pylori induced apoptosis, Bax translocates to the mitochondria which subsequently undergo depolarisation and profound fragmentation. Functional ARF and p53 proteins may play an important role in H pylori induced mitochondrial modification.     

Spontaneous and drug-induced apoptosis is mediated by conformational changes of Bax and Bak in B-cell chronic lymphocytic leukemia

The role of Bax and Bak, 2 proapoptotic proteins of the Bcl-2 family, was analyzed in primary B-cell chronic lymphocytic leukemia (CLL) cells following in vitro treatment with fludarabine, dexamethasone, or the combination of fludarabine with cyclophosphamide and mitoxantrone (FCM). A strong correlation was found between the number of apoptotic cells and the percentage of cells stained with antibodies recognizing conformational changes of Bax (n = 33; r = 0.836; P <.001) or Bak (n = 10; r = 0.948; P <.001). Preincubation of CLL cells with Z-VAD.fmk (N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone), a broad caspase inhibitor, abolished caspase-3 activation, exposure of phosphatidylserine residues, and reactive oxygen species generation; partially reversed the loss of transmembrane mitochondrial potential (DeltaPsim); but did not affect Bax or Bak conformational changes. These results indicate that the conformational changes of Bax and Bak occur upstream of caspase activation or are caspase independent. Following drug-induced apoptosis, Bax integrates into mitochondria, as demonstrated by fluorescence microscopy and Western blot, without changes in the total amount of Bax or Bak protein. Fludarabine and FCM induce p53 stabilization, but do not seem to be essential in inducing Bax and Bak conformational changes, as they are also observed in dexamethasone-treated CLL cells. These results demonstrate that, in CLL cells, the change in the intracellular localization of Bax from cytosol to mitochondria and the conformational changes of Bax and Bak are among the early steps in the induction of cell death.     

Imatinib mesylate induces apoptosis in human cholangiocarcinoma cells.

BACKGROUND: Cholangiocarcinoma is a highly malignant, usually fatal cancer with limited therapeutic options. Receptor tyrosine kinases contribute to the development and progression of this cancer. The relatively selective tyrosine kinase inhibitor imatinib mesylate (STI-571 or Gleevec(R)) has recently been licensed. However, the ability of this drug to inhibit signal transduction and induce apoptosis in human cholangiocarcinoma cells is incompletely studied. Thus, our goal was to examine the ability of STI-571 to induce apoptosis in KMCH-1 cells, a human cholangiocarcinoma cell line. METHODS: Apoptosis was assessed morphologically and also biochemically by measuring caspase activity and the mitochondrial membrane potential. STI-571 induced apoptosis and inhibited growth of KMCH-1 cells in a time- and concentration-dependent manner. The induction of apoptosis was accompanied by mitochondrial depolarization followed by a 4.5-fold increase in caspase activation and was abrogated by the pancaspase inhibitor z-VAD(OMe)-fmk. Interestingly, cholangiocarcinoma cells do not express detectable PDGFR, c-Abl or c-Kit, which are protein kinases known to be directly inhibited by STI-571. However, a significant decrease in epidermal growth factor receptor (EGFR) and focal adhesion kinase (FAK) phosphorylation was observed following treatment with STI-571. This decrease in EGFR and FAK phosphorylation was associated with a reduction in Akt activity resulting in loss of Mcl-1, a potent anti-apoptotic Bcl-2 family protein. CONCLUSIONS: These results indicate that STI-571 induces caspase-dependent apoptosis in a human cholangiocarcinoma cell line and suggest that STI-571 might warrant further investigation as a possible agent for treatment of human cholangiocarcinoma.     

心力衰竭大鼠心肌细胞凋亡及Bax蛋白表达的探讨

为了探讨心肌细胞凋亡在心力衰竭中的地位和作用,用结扎冠状动脉方法建立大鼠心力衰竭模型.采用末端脱氧核糖酸转移酶介导的dUTP缺口末端标记(TUNEL)染色法对心肌组织进行原位细胞凋亡检测,发现心衰组心肌细胞核有黄褐颗粒状改变.用SP免疫组化染色,心衰组Bax蛋白在膜及胞浆有阳性表达;而血管紧张素转换酶抑制剂(ACEI)治疗组,心肌凋亡细胞数和Bax蛋白表达明显减少,表明心肌细胞凋亡是心力衰竭发生原因之一,在心衰的发生中起着一定的作用,而ACEI能减轻凋亡的发生.     

Attenuation of apoptosis by chromogranin A-induced Akt and survivin pathways in prostate cancer cells

Multiple studies indicate that neuroendocrine (NE) differentiation in prostate cancer (PC) contributes to androgen-independent progression. Levels of chromogranin A (CgA), which is produced by NE cells, are increased in advanced PC. However, the mechanism by which high levels of circulating CgA contribute to PC progression is unknown. To examine the effects of CgA on PC cells, we first performed proliferation assays in the presence of recombinant CgA (rCgA) in LNCaP and C4-2 PC cells, and found that rCgA increased cell proliferation in a dose and time-dependent manner. NE differentiated PC cells, also overexpress the antiapoptosis protein survivin. Therefore, we examined survivin expression in the presence of CgA in PC cells. Western blot analysis showed that survivin was significantly increased by rCgA and inhibited by an anti-CgA antibody in both LNCaP and C4-2 cells. Survivin expression is believed to be regulated by PI3K/Akt pathway. We next assessed the phosphorylation status of Akt and found that Akt phosphorylation was increased by treatment with rCgA. To determine whether Akt phosphorylation is necessary for rCgA-induced survivin expression, we examined the effects of Akt, MAPK kinase, and protein kinase C inhibitors on CgA-induced survivin expression, and found that survivin expression was reduced in the presence of Akt inhibitors, but not MAPK kinase or protein kinase C inhibitors. Furthermore, in the presence of an Akt inhibitor or small interfering RNA of survivin, CgA-enhanced proliferation of C4-2 and LNCaP cells significantly decreased. Together, our results demonstrate that CgA can increase PC cell survival through Akt-mediated survivin up-regulation.     

Staurosporine诱导乳鼠心肌细胞凋亡与Bcl-2/Bax的关系

目的 探讨staurosporine（STS）诱导心肌细胞凋亡过程中,凋亡调节蛋白Bcl-2和Bax的变化及意义。方法 以STS诱导原代培养的SD乳鼠心肌细胞,检测半胱天冬蛋白酶(caspase)-3的活性及用Hoechst-33258荧光染色观察细胞凋亡。用免疫印迹法（Western blot）检测Bcl-2和Bax表达的变化。用免疫荧光共聚焦显微镜和免疫印迹法观察Bax在细胞内的分布。结果 以4 μmol/L STS处理心肌细胞,随着处理时间的延长,细胞中caspase-3的活性逐渐增加,与对照组比较,8 h达到峰值（P<0.01）。Hoechst-33258荧光染色显示,多数细胞核呈现核分叶。STS诱导心肌细胞凋亡过程中,Bcl-2和Bax的水平与对照组比较无显著变化。正常细胞内的Bax均匀分布于细胞质中,在STS处理的细胞中,Bax明显聚集于线粒体上,呈现明显的线粒体的转位。STS诱导心肌细胞凋亡过程中,胞质片段Bax的水平明显下降,而线粒体片段Bax的水平明显升高,进一步证实STS诱导细胞凋亡过程中伴有明显的Bax线粒体转位。结论 STS诱导细胞凋亡中,伴有明显的Bax由胞质向线粒体的转位。     

阻塞性黄疸大鼠肝组织Bcl-2及BaX的表达与细胞凋亡

目的探讨在阻塞性黄疸肝损害中Bcl-2,Bax基因的表达.方法结扎Wistar大鼠胆总管建立阻塞性黄疸大鼠动物模型.分为结扎后3,7,14,21d及阴性对照各组,每组8只.结扎后3,7,14,21d处死大鼠,门静脉抽血,离心后取上层血清行胆红素测定.HE染色,光镜下观察阻塞性黄疸大鼠肝脏组织的病理改变.用末端脱氧核苷酸转移酶(TdT)介导的dUTP缺口末端标记技术(TUNEL)和免疫组织化学方法检测阻塞性黄疸大鼠肝脏组织细胞凋亡状态及凋亡相关基因Bcl-2,Bax蛋白的表达.结果胆总管结扎后,随结扎时间的延长胆红素值逐渐增高.光镜下,胆总管结扎3d即见肝内胆管扩张,结扎7d,肝细胞内或细胞间瘀胆,纤维组织增生;结扎14 d,纤维组织增生向肝小叶内扩展;结扎21d,纤维组织带增宽,肝脏结构广泛改变.TUNEL检测表明,正常大鼠肝组织偶见细胞凋亡,结扎3d即见细胞凋亡增加,结扎7d明显增加,结扎14 d后细胞凋亡达高峰,凋亡指数(AI)达58.2%±1.6%,各组AI有显著性差异(P＜0.05),21d细胞凋亡数目明显减少.SABC免疫组化检测,Bcl-2蛋白的表达,对照组、胆总管结扎3d,7 d均为阴性表达,14 d弱阳性表达,21 d强阳性表达;结扎14 d,Bcl-2蛋白弱阳性表达,AI达高峰,结扎21d Bcl-2强阳性表达,AI下降.Bax蛋白的表达,对照组、胆总管结扎3 d均可见表达,7d表达增多,14 d明显增多;21 d表达减少;结扎7 d,Bax蛋白中阳性表达,AI增高;结扎14d,Bax蛋白强阳性表达,AI达高峰.表明在阻塞性黄疸过程中,Bcl-2蛋白表达越强,凋亡指数越少;Pax蛋白表达越强,凋亡指数越强.结论Bcl-2和Bax蛋白均参与了阻塞性黄疸肝组织中细胞凋亡的调节,并在阻塞性黄疸肝损害的发生和发展中起重要作用.     

Decreased Bax expression by mucosal T cells favours resistance to apoptosis in Crohn's disease

BACKGROUND: Activated T cells are more susceptible to apoptosis than resting T cells. As intestinal T cells normally exhibit a higher state of activation, increased apoptosis may be necessary to maintain immune homeostasis in the specialised microenvironment of the mucosa. On the other hand, in Crohn's disease (CD) mucosal T cells are resistant to apoptosis, suggesting abnormal regulation of cell death mechanisms. AIMS: To investigate differences in expression of anti- and proapoptotic Bcl-2 family proteins, key regulators of apoptosis, between circulating and mucosal T cells, and possible alterations in CD. PATIENTS AND METHODS: Lamina propria T cells (LPT) were isolated from 10 control, seven CD, and eight ulcerative colitis (UC) patients, and peripheral blood T cells (PBT) from healthy volunteers. Purified T cells were stained intracellularly for Bcl-2, Bcl-x(L), and Bax, and mean fluorescence intensity measured by flow cytometry. RESULTS: Compared with PBT, the expression level of Bcl-2 and Bax, but not Bcl-x(L), was significantly greater in LPT, resulting in lower Bcl-x(L)/Bax ratios. In PBT, Bax expression was highly and significantly correlated with both Bcl-2 and Bcl-x(L), but correlation with Bcl-2 was absent in LPT. Bax expression in CD, but not UC, LPT was significantly lower than in control LPT, resulting in a significantly higher Bcl-x(L)/Bax ratio. The significant correlation of Bcl-x(L) to Bax was preserved in CD, but not UC, LPT. CONCLUSIONS: Regulation of Bcl-2 family protein expression differs between circulating and mucosal T cells, probably underlying diverse survival potentials. In CD LPT, a low Bax expression and a high Bcl-x(L)/Bax ratio favour resistance to apoptosis and may contribute to the chronicity of inflammation.     

Peg3/Pw1 promotes p53-mediated apoptosis by inducing Bax translocation from cytosol to mitochondria

Mitochondria is believed to play a central role in p53-mediated apoptosis. However, the signal transduction pathways leading to mitochondria remain unclear. Here, we report that translocation of Bax protein from cytosol to mitochondria is required for p53-induced apoptosis. Cytosolic Bax is unable to induce apoptosis, and blocking Bax translocation inhibits cell death. Expression of Bcl-2 blocks cytochrome c release and apoptosis but has no effect on Bax translocation, suggesting that Bax translocation acts upstream of Bcl-2. We further demonstrate that Peg3/Pw1, a protein up-regulated in p53-mediated cell death process, induces Bax translocation independent of apoptosis. The results suggest that Bax translocation represents an important regulatory step in p53-mediated apoptosis, and Peg3/Pw1 functions as a modulator downstream of p53 to regulate Bax redistribution in the cells, thus favoring the cellular decision toward apoptosis over growth arrest following p53 induction.     

Raddeanin A promotes apoptosis and ameliorates 5-fluorouracil resistance in cholangiocarcinoma cells

BACKGROUND Bile duct cancer is characterized by fast metastasis and invasion and has been regarded as one of the most aggressive tumors due to the absence of effective diagnosis at an early stage.Therefore,it is in the urgent demand to explore novel diagnostic approaches and therapeutic strategies for bile duct cancer to improve patient survival.Raddeanin A(RA)is extracted from the anemone raddeana regel and has been demonstrated to play antitumor roles in various cancers.AIM To investigate the effects of RA treatment on bile duct cancer cells.METHODS In this study,four cholangiocarcinoma cell lines(RBE,LIPF155C,LIPF178C,and LICCF)treated with RA were used to test the cell viability.The RA-associated cell functional analysis,5-fluorouracil(5-Fu)effectiveness as well as cell cycle-and apoptosis-related protein expression were investigated.RESULTS RA reduced cell viability in a dose-dependent pattern in four cell lines,and the migration and colony formation abilities were also impaired by RA in RBE and LIPF155C cell lines.RA sensitized cell lines to 5-Fu treatment and enhanced the effects of 5-Fu in cholangiocarcinoma.Also,RA decreased protein expression of Wee1,while the combinational effect of RA and 5-Fu decreased protein expressions of cyclooxygenase-2,B cell lymphoma 2,and Wee1 but increased protein levels of Bax,cyclin D1,and cyclin E.CONCLUSION Taken together,the results suggest that RA acts as an anti-cancer agent and enhancer of 5-Fu in bile duct cancer cells via regulating multiple cell cycle and apoptosis-related proteins.This finding provides novel clues to exploring a novel antitumor drug for bile duct cancer.     

The nuclear translocation of endostatin is mediated by its receptor nucleolin in endothelial cells

Endostatin, the C-terminal fragment of collagen XVIII, is a potent anti-angiogenic factor that significantly modulates the gene expression pattern in endothelial cells. Upon cell surface binding, endostatin can not only function extracellularly, but also translocate to the nucleus within minutes. However, the mechanism by which this occurs is partially understood. Here we systematically investigated the nuclear translocation mechanism of endostatin. By chemical inhibition and RNA interference, we firstly observed that clathrin-mediated endocytosis, but not caveolae-dependent endocytosis or macropinocytosis, is essential for the nuclear translocation of endostatin. We then indentified that nucleolin and integrin α5β1, two widely accepted endostatin receptors, mediate this clathrin-dependent uptake process, which also involves urokinase plasminogen activator receptor (uPAR). Either mutagenesis study, fluorescence resonance energy transfer assay, or fluorescence cell imaging demonstrates that nucleolin and integrin α5β1 interact with uPAR simultaneously upon endostatin stimulation. Blockade of uPAR decreases not only the interaction between nucleolin and integrin α5β1, but also the uptake process, suggesting that the nucleolin/uPAR/integrin α5β1 complex facilitates the internalization of endostatin. After endocytosis, nucleolin further regulates the nuclear transport of endostatin. RNA interference and mutational analysis revealed that the nuclear translocation of endostatin involves the association of nucleolin with importin α1β1 via the nuclear localization sequence. Taken together, this study reveals the pathway by which endostatin translocates to the nucleus and the importance of nucleolin in this process, providing a new perspective for the functional investigation of the nuclear-translocated endostatin in endothelial cells.     

Preconditioning decreases Bax expression, PMN accumulation and apoptosis in reperfused rat heart

OBJECTIVE: Recent studies suggest that ischemic preconditioning (IPC) inhibits myocardial apoptosis after ischemia and reperfusion. This study tested the hypothesis that IPC reduces ischemia/reperfusion-induced myocardial apoptosis by inhibiting neutrophil (PMN) accumulation and altering expression of Bcl-2 and Bax proteins. METHODS: Eighteen rats were subjected to 30 min of left coronary artery occlusion followed by 180 min of reperfusion with IPC (5 min ischemia and 10 min of reperfusion, n = 10) or without IPC (n = 8). Myocardial apoptosis was detected histologically using the terminal transferase UTP nick end labeling (TUNEL) assay and confirmed by DNA ladder on agarose gel electrophoresis. PMN accumulation was detected immunohistochemically with anti-rat CD18 antibody (WT3) and expression of Bcl-2 and Bax proteins was analyzed using Western blot assay. RESULTS: IPC significantly decreased TUNEL positive cells (% total nuclei) in the ischemic zone from 28.6 +/- 2.8 to 3.4 +/- 0.9 (P < 0.05), consistent with the absence of DNA ladders in the IPC group. IPC significantly attenuated PMN accumulation (cells/mm2 myocardium) in the ischemic zone from 243 +/- 19 to 118 +/- 19 (P < 0.05). By regression analysis, there was a significant correlation between TUNEL positive cells and accumulated CD18 positive PMNs in the ischemic zone (r = 0.8, P < 0.001), which was shifted downward by IPC. Densitometrically, IPC significantly attenuated the ischemia/reperfusion-upregulated expression of Bax protein in the ischemic zone from 204 +/- 57% in the control group to 76 +/- 7% (P < 0.05), while the expression of Bcl-2 was not different from the non-ischemic zone in either group. CONCLUSION: These data suggest that ischemic preconditioning may reduce myocardial apoptosis by inhibiting PMN accumulation and down-regulating expression of Bax.     

Histone deacetylase inhibitor depsipeptide (FK228) induces apoptosis in leukemic cells by facilitating mitochondrial translocation of Bax, which is enhanced by the proteasome inhibitor bortezomib

Histone deacetylase (HDAC) inhibitors are promising candidates for molecular-targeted therapy for leukemia. In this study, we investigated the mechanisms of cytotoxic effects of depsipeptide (FK228), one of the most effective HDAC inhibitors against leukemia, using human myeloid leukemic cell lines HL-60 and K562. We found that FK228 activated caspase-9 and a subsequent caspase cascade by perturbing the mitochondrial membrane to release cytochrome c, which was almost completely blocked by overexpression of Bcl-2. The mitochondrial damage was caused by the translocation of Bax but not other pro-apoptotic Bcl-2 family proteins to the mitochondria. FK228 did not affect the interaction between Bax and Bax adaptor proteins such as 14-3-3theta and Ku70. FK228-induced apoptosis and mitochondrial translocation of Bax were markedly enhanced by the proteasome inhibitor bortezomib. The synergistic action of FK228 and bortezomib was at least partly mediated through conformational changes in Bax, which facilitate its translocation to the mitochondria. These results suggest that the combination of HDAC inhibitors and proteasome inhibitors is useful in the treatment of leukemia especially in the context of molecular-targeted therapy. The status of Bcl-2 and Bax may influence the sensitivity of tumors to this combination and thus can be a target of further therapeutic intervention.     

Akt expression may predict favorable prognosis in cholangiocarcinoma

BACKGROUND: Overexpression of signaling proteins including epidermal growth factor receptor (EGFR), Akt, mitogen activated protein kinase (MAPK) and cyclooxygenase-2 (COX-2) occurs in cholangiocarcinoma cell lines. However, the prognostic value of these markers is unknown. No prior study correlated the expression of these signaling proteins with clinical outcome. Further, co-expression of these proteins has not been reported. Co-expression may reflect cross-talk between signaling pathways. The aim of this clinicopathological study was to investigate the overexpression and co-expression of EGFR and related signaling proteins in cholangiocarcinoma and explore their relationship to clinical outcome. METHODS: Twenty-four consecutive cases of cholangiocarcinoma treated from 1996 to 2002 at Roswell Park Cancer Institute were included. Immunohistochemical staining of paraffin-embedded tissue sections was performed using antibodies against Akt, p-Akt, MAPK, p-MAPK, COX-2, EGFR and p-EGFR. Two pathologists independently scored the protein expression. RESULTS: Cyclooxygenase-2, Akt, and p-MAPK were commonly expressed in biliary cancers (100%, 96% and 87% of malignant cells, respectively). EGFR (60%) and p-EGFR (22%) overexpression was also detected. There was a significant association between EGFR and p-EGFR (P = 0.027) and between Akt and p-Akt (P = 0.017) expression in tumor tissue. A noteworthy association was shown between MAPK and p-Akt (P = 0.054). Multivariate analysis using the Cox proportional hazard model identified the use of chemotherapy (hazard ratio [HR] = 0.039, P = 0.0002), radiation (HR = 0.176, P = 0.0441) and Akt expression (HR = 0.139, P = 0.006) as the best predictors of overall prognosis. CONCLUSION: Epidermal growth factor receptor signaling intermediates are commonly expressed in cholangiocarcinoma. Expression of Akt and use of systemic chemotherapy or radiation may correlate with improved survival.