Synergistic antileukemic interactions between 17-AAG and UCN-01 involve interruption of RAF/MEK- and AKT-related pathways

Interactions between the protein kinase C (PKC) and Chk1 inhibitor UCN-01 and the heat shock protein 90 (Hsp90) antagonist 17-AAG have been examined in human leukemia cells in relation to effects on signal transduction pathways and apoptosis. Simultaneous exposure (30 hours) of U937 monocytic leukemia cells to minimally toxic concentrations of 17-AAG (eg, 400 nM) and UCN-01 (eg, 75 nM) triggered a pronounced increase in mitochondrial injury (ie, loss of mitochondrial membrane potential [Deltapsim]; cytosolic release of cytochrome c), caspase activation, and apoptosis. Synergistic induction of apoptosis was also observed in other human leukemia cell types (eg, Jurkat, NB4). Coexposure of human leukemia cells to 17-AAG and the PKC inhibitor bisindolylmaleimide (GFX) did not result in enhanced lethality, arguing against the possibility that the PKC inhibitory actions of UCN-01 are responsible for synergistic interactions. The enhanced cytotoxicity of this combination was associated with diminished Akt activation and marked down-regulation of Raf-1, MEK1/2, and mitogen-activated protein kinase (MAPK). Coadministration of 17-AAG and UCN-01 did not modify expression of Hsp90, Hsp27, phospho-JNK, or phospho-p38 MAPK, but was associated with further p34cdc2 dephosphorylation and diminished expression of Bcl-2, Mcl-1, and XIAP. In addition, inducible expression of both a constitutively active MEK1/2 or myristolated Akt construct, which overcame inhibition of ERK and Akt activation, respectively, significantly attenuated 17-AAG/UCN-01-mediated lethality. Together, these findings indicate that the Hsp90 antagonist 17-AAG potentiates UCN-01 cytotoxicity in a variety of human leukemia cell types and suggest that interference with both the Akt and Raf-1/MEK/MAP kinase cytoprotective signaling pathways contribute to this phenomenon.     

项七针治疗椎动脉型颈椎病和对椎-基底动脉血流动力学的影响

目的 观察项七针治疗椎动脉型颈椎病的疗效和对椎-基底动脉血流动力学的影响。方法 将病人随机分为项七针组38例和颈夹脊组34例，进行症状评分和经颅彩色多普勒检查椎-基底动脉血流速指标。结果 经治疗2个疗程后，项七针组总有效率为89．47％，颈夹脊组为76．47％，两组差异有统计学意义（P〈0．05）；项七针组治疗后椎-基底动脉平均血流速度（MV）、椎-基底动脉搏动指数（PI）、阻力指数（RI）的改变与颈夹脊组比较均有统计学意义（P〈0．05）。项七针组疗效和改善脑供血方面均优于颈夹脊组。结论 运用针灸治疗椎动脉型颈椎病，可收到一定的临床疗效，且在改善病人症状积分及椎-基底动脉血流速度等方面亦显示出较好好的治疗作用。     

Cloning and functional characterization of GMPR2, a novel human guanosine monophosphate reductase,which promotes the monocytic differentiation of HL-60 leukemia cells

PURPOSE: To identify the biological function of a novel molecule which shows high homology with human guanosine monophosphate reductase (GMPR) and is named GMPR2. METHODS: GMPR2 cDNA was cloned from the cDNA library of human dendritic cells and was characterized by Bioinformatics. The expression pattern of GMPR2 was analyzed by Northern blotting. The enzymatic activity of the purified recombinant GMPR2 protein was determined using a spectrophotometric assay. HL-60 leukemia cells were transfected with GMPR2 and the expression of CD14 and myeloperoxidase (MPO) in HL-60 cells with and without 12- o-tetra-decanoyl-phorbol-13-acetate (TPA) induction was monitored by FACS analysis. RESULTS: The novel gene contained ten exons and nine introns and was mapped to 14q11-21. Northern blotting indicated a ubiquitous expression of GMPR2 mRNA in most of the human tissues and cancer cell lines investigated. The recombinant GMPR2 protein was able to reduce GMP. The expression of CD14 and MPO in HL-60 leukemia cells overexpressing GMPR2 clearly increased after induction by TPA. CONCLUSIONS: GMPR2 is a novel human GMP reductase, and overexpression of GMPR2 can promote the monocytic differentiation of HL-60 leukemia cells.     

Sphingosine 1-phosphate, a specific endogenous signaling molecule controlling cell motility and tumor cell invasiveness.

Sphingosine 1-phosphate (Sph-1-P), the initial product of Sph degradation by Sph kinase, was shown to be a strong inhibitor of cell motility and phagokinesis of B16 melanoma and other types of cells at 10-100 nM concentration. It also inhibited "chemoinvasion" of tumor cells through a thick layer of Matrigel on a filter membrane. Such inhibitory effects were produced minimally or not at all by Sph, N-methyl derivatives of Sph, or other related sphingolipids and phospholipids. Sph-1-P did not inhibit cell proliferation or protein kinase C (PKC) activity, in contrast to Sph and N-methyl-Sph, which inhibit PKC activity and cell growth in general. Radiolabeled [3H]Sph and [14C]N-methyl-Sph were rapidly incorporated into B16 melanoma cells. However, [14C]N-methyl-Sph was not metabolically converted into other compounds, whereas [3H]Sph was efficiently converted within 10 min to Sph-1-P, followed by conversion to other sphingolipids and phospholipids. The inhibitory effect of Sph-1-P on cell motility and tumor cell invasiveness could be a specific phenomenon independent of PKC and other known transmembrane signaling mechanisms, based on an unknown mechanism. It may directly affect organizational assembly of actin filaments. Since exogenous Sph is rapidly converted into Sph-1-P, some reported effects of Sph may be ascribable to such conversion.     

腹股沟斜疝少见和特殊征象的多层螺旋CT表现

目的探讨腹股沟斜疝少见和特殊征象的多层螺旋CT表现。方法收集19例经手术证实的少见腹股沟斜疝病例,均行多层螺旋CT扫描,其中平扫4例,增强扫描15例,均将原始图像传至工作站行图像后处理,分析影像特点及特殊征像。结果 19例中单侧疝12例,双侧7例。巨大疝6例,1例同时见膀胱、肠管、脂肪及腹膜等疝入,另5例疝内容物为肠管、网膜等;少见疝内容物6例,包括膀胱疝入5例,1例为大部分膀胱疝入,表现为肠管及腹膜伴随膀胱一起疝入阴囊;其余为部分膀胱疝入,见膀胱呈"哑铃"状改变;输尿管疝入阴囊1例,表现为迂曲扩张的巨输尿管疝入阴囊,并伴双肾积水。嵌顿疝合并肠梗阻5例,表现为近端肠管扩张积气并出现液-气平面;绞窄性疝2例,增强扫描肠管无强化。结论多层螺旋CT能够正确判断少见腹股沟斜疝的类型,明确疝内容物的种类、大小及周围解剖关系,为少见和特殊腹股沟斜疝的诊断积累影像经验。     

心理干预对妇科宫颈癌术后患者化疗依从性的影响

目的:探讨心理干预对妇科宫颈癌术后患者化疗依从性的影响。方法:将我院2013年1月~2014年1月收治的120例宫颈癌患者随机分为观察组和对照组各60例,两组均给予常规联合化疗和护理,观察组在此基础上对患者进行心理干预,比较两组患者化疗依从性。结果:观察组患者抑郁自评量表(SDS)和焦虑自评量表(SAS)得分均低于对照组(P<0.05);观察组患者依从性明显高于对照组(P<0.05);观察组患者心理干预前后掌握相关知识、自我防护不合理、不愿继续化疗比例差异比较有统计学意义(P<0.05)。结论:对妇科宫颈癌患者进行心理干预,有助于患者预后恢复,显著提高了妇科宫颈癌术后患者化疗依从性。     

Inhibition of the NF-κB pathway by R65 ribozyme gene via adeno-associatedvirus serotype 9 ameliorated oxidized LDL induced human umbilical vein endothelial cell injury

Objective: NF-κB signaling plays a central role in the regulation of inflammatory responses in atherosclerosis. R65 ribozyme gene suppresses activation of NF-κB pathway, therefore we studied whether R65 gene therapy can ameliorate oxidized low-density lipoprotein (ox-LDL) induced human umbilical vein endothelial cells (HUVECs) injury. Methods and results: Recombinant adeno-associated virus serotype 9 (rAVV9) vector was used to transfect the R65 ribozyme gene (rAVV9-R65) into HUVECs then following ox-LDL stimulation, expression of NF-κB p65 and p50 subunits, inflammatory mediators and cell apoptosis were examined. First, rAVV9-enhanced green fluorescent protein (eGFP)-R65 at 1×10⁷ v.g./cell multiplicity of infection reached a long-lasting and significant increase in R65 gene expression. Second, ox-LDL treatment led to time- and dose-dependent activation of NF-κB pathway, and enhanced inflammatory response and cell death evidenced by increased expression of nuclear NF-κB p65 and p50 subunits, greater production of tumor necrosis factor α, interleukin-6 and von willebrand factor and 20.57% increasedapoptotic HUVECs. Third, over-expression ofR65 gene was 2-fold increased in HUVECs attenuated ox-LDL induced unclear accumulation and expression of p65 subunit and ameliorated inflammation and cell death (all P < 0.05). Conclusion: rAAV9-mediated R65 ribozyme gene transfection in cultured HUVECs effectively inhibits ox-LDL induced activation of NF-κB and production of inflammatory cytokines and prevents cell apoptosis.     

MicroRNA-200a inhibits epithelial-mesenchymal transition in human hepatocellular carcinoma cell line

Objective: Our study investigated the role of microRNA (miR)-200a and its molecular targets in hepatocellular carcinoma (HCC) cells. Methods: An inhibitor of miR-200a was transiently transfected into the hepatocellular carcinoma cell line, MHCC-97L. The effect of this transfection on mRNA levels of epithelial-mesenchymal transition (EMT)-related genes was measured by fluorescence-based quantitative real-time polymerase chain reaction (qRT-PCR). Further, protein levels of EMT-related genes, cell proliferation and apoptosis-related markers were assessed by Western blot analysis in these transfected cells. MTT and wound-healing assay were used to evaluate the proliferation and migration of MHCC-97L cells in presence and in absence of miR-200a inhibitor. Results: Compared with miR-NC control group, qRT-PCR results in anti-miR-200a group revealed a significant reduction in the mRNA levels of E-cadherin, with a concomitant increasing in vimentin mRNA level (all P < 0.05). Western blot results showed higher E-cadherin and Caspase-3 protein expressions in anti-miR-200a group compared to miR-NC group (P < 0.05). In addition, vimentin and Ki-67 protein expression was found sharply decreased in anti-miR-200a group compared to miR-NC group (P < 0.05). Consistent with this, wound-healing and MTT assay showed that migration and proliferation capacity of MHCC-97L cells in anti-miR-200a group is significantly increased compared with miR-NC group (both P < 0.05). Conclusion: Our study reveals an important role of miR-200a in inhibiting EMT, proliferation and migration in HCC cells, suggesting the possibility of miR-200a-based therapeutics in HCC.     

Hypermethylation of TFPI2 correlates with cervical cancer incidence in the Uygur and Han populations of Xinjiang,China

Tissue factor pathway inhibitor 2 (TFPI2) is a Kunitz-type serine proteinase inhibitor, which plays an important role in the etiology of human malignancies. DNA methylation is a common epigenetic modification of the genome that is involved in regulating many cellular processes. In addition to human papilloma virus (HPV) infection, DNA methylation may play a role in the carcinogenesis of cervical cancer. Methylation of 22 CpG sites in the promoter region of the TFPI2 gene was detected by MassARRAY spectrometry and a gene mass spectrogram was drawn using MALDI-TOF MS. HPV16 was detected by PCR. We show that aberrant methylation of TFPI2 is present in a higher proportion of invasive cervical carcinoma (ICC) clinical samples as compared to normal cervical samples in Uygur and Han. Across the four pathologic lesions of the progression of cervical cancer, ICC showed the highest level of aberrant methylation, and with a stronger correlation between CpG site and lesion grade in Uygur than in Han. Moreover, a difference in TFPI2 methylation between Uygur patients positive and negative for HPV16 infection was observed at CpG_6 (P = 0.028) and CpG_15 (P = 0.007). Altogether, these results indicate that DNA methylation of TFPI2 may play an important role in the carcinogenesis of cervical cancer and that the differential methylation of TFPI2 may at least partially explain the disparity in cervical cancer incidence between Uygur and Han women.