Epigenetic inactivation of the CIP/KIP cell-cycle control pathway in acute leukemias

Dysregulation of the cell cycle is important in oncogenesis. We analyzed the potential inactivation of the CIP/KIP family of the cyclin E/CDK/RB pathway by gene promoter hypermethylation in leukemias. The methylation-specific polymerase chain reaction (MSP) with primers for methylated (M-MSP) and unmethylated (U-MSP) alleles of the p21, p27, and p57 genes was used to study five leukemic cell lines, 50 acute myeloid leukemia (AML) samples, and 25 acute lymphoblastic leukemia (ALL) samples. p21 was hemizygously methylated in Raji and Jurkat but remained unmethylated in U937, HL60, and NB4. p27 was hemizygously methylated in Raji but unmethylated in the other cell lines. p57 was completely methylated in Raji and NB4, hemizygously methylated in U937, and unmethylated in HL60 and Jurkat. At diagnosis, p21 methylation was not detected in any case of AML or ALL. p27 methylation occurred in 2 (4%) AML patients and in 1 (4%) ALL patient. p57 methylation occurred in 1 (2%) AML patient and in 1 (4%) ALL patient. Therefore, methylation inactivation of the INK4/CDK/RB pathway in leukemia is infrequent. A review of the literature showed a marked variation in the frequencies of methylation of these genes, which might be attributable to difference in methodologies used to detect gene methylation.     

CK19 CK20和MMP-9mRNA外周血表达与中晚期肺癌临床相关性

目的：检测肺癌外周血中细胞角蛋白19（CK19）mRNA、细胞角蛋白20（CK20）mRNA和基质金属蛋白酶-9（MMP-9）mRNA,探讨其作为判断晚期肺癌外周血检测其转移及判断预后的指标可行性。方法采用RT-PCR技术检测肺癌患者外周血中CK19mRNA、CK20mRNA、MMP-9mRNA的表达量,按表达阴阳性作为标准对其临床资料及短期疗效进行评估。结果 CK19、MMP-9及CK20表达阳性率化疗3疗程后均下降,其中CK19、MMP-9差异有统计学意义（P值0.003、0.007）;CK19mRNA、CK20mRNA和MMP-9mRNA表达阳性率化疗前后与性别,分化等均无相关性（P＆gt;0.05）;其中化疗前CK19mRNA表达Ⅳ期阳性率高于Ⅲ期患者（P=0.01）;腺癌患者CK19mRNA与MMP-9mRNA表达,化疗前表达阳性率明显高于鳞癌差异有统计学意义（P值为0.01、0.04）;化疗后腺癌CK19mRNA、CK20mRNA、MMP-9mRNA表达较鳞癌表达下降明显（P值为0.04、0.03、0.02）;治疗前CK20mRNA和MMP-9mRNA表达与有无疗效存在相关性,差异有统计学意义（P值为0.04、0.0.02）,CK19mRNA表达无相关性。化疗后CK19mRNA、CK20mRNA表达与有无疗效存在相关性,差异有统计学意义（P值为0.01、0.02）,MMP-9mRNA表达无相关性。结论通过CK19、CK20、MMMP-9mRNA表达检测,判断晚期肺癌患者化疗疗效,指导用药,提高晚期患者的生活质量及预后的监测的重要意义。     

YC-1 inhibits proliferation of human vascular endothelial cells through a cyclic GMP-independent pathway

This study was designed to investigate the effect of YC-1, 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole, in human umbilical vein endothelial cells (HUVECs) proliferation and its underlying mechanism. YC-1 at a range of concentrations (5-50 microM) inhibited DNA synthesis and decreased cell number in cultured HUVEC in a dose- and time-dependent manner. YC-1 was not cytotoxic at these concentrations. [3H]thymidine incorporation and flow cytometry analyses revealed that YC-1 treatment decreased DNA synthesis and arrested the cells at the G0/G1 phase of the cell cycle. Western blot analysis demonstrated that YC-1 (5-50 microM) increased the levels of cyclin-dependent kinase (CDK)-inhibitory proteins (CKIs), p21 and p27, but did not induce any significant changes of cyclins and CDKs. In the YC-1-treated HUVEC, the formation of CDK2-p21 complex, but not CDK2-p27 complex, was increased and the assayable CDK2 kinase activity was decreased. These changes were in a dose-dependent manner. In contrast, the formations of CDK4-p21 and CDK4-p27 complex were slightly increased and the assayable CDK4 kinase activity was slightly decreased (if there were any changes). Pretreatment with guanylyl cyclase inhibitors, 1H-(1,2,4)oxadiazolo[4,3-a]quinozalin-1-one (ODQ) and methylene blue, inhibited the YC-1-induced increase of cyclic GMP level, but did not change significantly the magnitude of the YC-1-induced inhibition of thymidine incorporation and cell number in HUVEC. These results indicate that YC-1-induced cell cycle arrest in HUVEC occurred when the cyclin-CDK system was inhibited just as p21 and p27 protein levels were augmented. This YC-1-induced antiproliferation effect in HUVEC is via a cyclic GMP-independent pathway.     

Cyclin E与恶性肿瘤

近年研究发现 ,细胞周期调控机制紊乱是细胞增生失控从而导致癌变的重要原因。哺乳动物中 ,一个典型的细胞周期由有严格顺序并循精确时相的 4个期即 G1 - S- G2 - M构成 ,并受到胞内外信号传导途径及反馈环路的调控 [1 ] 。其中 ,G1期是细胞增生周期的关键。目前已发现A- J10种细胞周期素 (cyclins) ,有些含亚型如 cyclin D1 、cyclin D2 、cyclin D3以及 cyclin E1 、cyclin E2 等共 15种 cy-clins。另据其峰值和主导作用通常又划为 G1 期和 M期 cyclins,前者包括 cy-clin C、D、E;后者包括 cyclin A、B。细胞周期素在结构上的共性…     

The cyclin-dependent kinase inhibitors p18INK4c and p19INK4d are highly expressed in CD34+ progenitor and acute myeloid leukaemic cells but not in normal differentiated myeloid cells

Cyclin-dependent kinase inhibitors (CKIs) are important for the differentiation of cells in various tissues. In acute myeloid leukaemia (AML) the cells accumulate at particular stages of myeloid maturation. We therefore analysed the expression pattern of different CKIs in fresh samples of AML patients and compared it with that in CD34+ progenitor and normal differentiated myeloid cells. Competitive RT-PCR and Western analysis revealed a significantly higher expression of p18INK4c and p19INK4d in leukaemic and CD34+ progenitor cells than in granulocytes and monocytes. A different pattern was seen for p27Kip1 and p57Kip2 expression being low in leukaemic cells but high in normal immature and differentiated cells. No marked differences were found in p15INK4b and p21Cip1 mRNA expression between leukaemic and CD34+ progenitor or mature myeloid cells. Our findings therefore indicate that high expression of p18INK4c and p19INK4d in haemopoietic progenitor and leukaemic blast cells may contribute to the premature differentiation block seen in AML.     

Calcium, calcium-sensing receptor and colon cancer

There is much evidence that dietary Ca(2+) loading reduces colon cell proliferation and carcinogenesis in humans and rodents, but during carcinogenesis it becomes ineffective or even tumor-promoting. We are beginning to see how Ca(2+) balances the continuous massive cell production in colon crypts by driving the terminal differentiation and eventually the apoptosis of the cells mainly on the mucosal surface, and how this Ca(2+) control is lost during colon carcinogenesis. The rapid proliferation of the transit-amplifying (TA) progeny of the colon stem cells is driven by the so-called "Wnt" signaling mechanism, which involves the stimulation of proliferogenic genes such as those for c-Myc and cyclin D1 and the silencing of the gene for the cell cycle-stopping p21(Cip1/WAF1) protein by nuclear beta-catenin*Tcf-4 complexes. TA cells avoid mitotic damage and premature apoptosis by expressing the protein survivin. It appears that TA cell cycling stops and terminal differentiation starts when the cells reach a higher level in the crypt where there is enough lumenal Ca(2+) to stimulate the expression and activation of CaSRs (Ca(2+)-sensing receptors), the signals from which stimulate the expression of E-cadherin. Along with this, the APC (adenomatous polyposis coli) protein appears and some of it enters the nucleus. There it makes the TA cells susceptible to the eventual apoptotic balancing by stopping survivin expression and the beta-catenin*Tcf-4 complex from driving further cell cycling by releasing beta-catenin from the nucleus, and delivering it to cytoplasmic APC*axin*GSK-3beta complexes for ultimate proteasomal destruction. Cytoplasmic beta-catenin is then prevented from returning to the nucleus by either being intercepted and destroyed by APC*axin*GSK-3beta complexes or locked by the emerging E-cadherin into membrane adherens junctions which tie the cell into the sheet of proliferatively shut-down cells with APC-dependent cytoskeletons moving to the mouth of the crypt and onto the flat mucosal surface. A common first step in sporadic colon carcinogenesis is the loss of functional APC which disorients upwardly directed migration and causes the retention of nuclear beta-catenin and proliferogenic beta-catenin*Tcf-4 complexes as well as genomic instability. Eventually the balance between cell proliferation and terminal differentiation and death is radically tipped in favour of proliferation by the appearance of apoptosis-resistant, survivin-expressing clones of Ca(2+)-insensitive cells which are locked into the proliferative, mutation-prone mode because of CaSR-disabling gene mutations which prevent the stimulation of E-cadherin expression and terminal differentiation.     

CD56、TPO、p63及CK19联合检测与甲状腺乳头状癌临床病理的相关性

目的：探讨CD56、TPO、p63及CK19联合检测与甲状腺乳头状癌（PTC）临床病理因素问的相互关系。方法：应用免疫组织化学sP法检测100例PTC中CD56、TPO、p63及CK19的表达，分析四种标志物与性别、年龄、病灶数目、病灶大小、病灶部位、被膜侵犯、淋巴结转移和组织学亚型等临床病理因素间的关系。结果：CD56、TPO、p63及CK19在100例PTC中的表达阳性率分别为11％、5％、17％、87％。四种标志物的表达与性别、年龄、病灶数目、病灶大小、病灶部位、有否林巴结转移、组织学亚型无关，差异无统计学意义（P〉0．05），其中有被膜侵犯时CD56的表达阳性率显著低于无被膜侵犯时，差异有统计学意义（P〈0．05），其他三种抗体的表达与是否有被膜侵犯无关，差异无统计学意义（JD〉0．05）。结论：CD56的表达与是否有被膜侵犯相关，有被膜侵犯时CD56的表达显著低于无被膜侵犯时，CD56低表达可预示PTC预后较差。