苦参碱对视网膜母细胞瘤细胞survivin和端粒酶活性表达的影响

目的 研究苦参碱对视网膜母细胞瘤(RB)细胞凋亡抑制蛋白survivin和端粒酶活性表达的影响,探讨苦参碱对Rb的作用机制.方法 体外培养视网膜母细胞瘤患者HXO-Rb细胞经不同浓度的苦参碱作用后,采用噻唑蓝比色法测定细胞生长情况,RT-PCR检测survivin RNA水平,聚合酶链法(PCR)检测端粒酶活性.结果 苦参碱在一定范围内可抑制HXO-Rb细胞的生长,使survivin mRNA表达下调.可明显观察到细胞凋亡的形态学改变,端粒酶活性明显下降,细胞凋亡率增加.结论 苦参碱可以增加HXO-RB细胞凋亡,可以减少RB细胞mRNA表达,减弱RB细胞端粒酶活性,影响其周期变化,抑制RB细胞系增殖.其对survivin的抑制,及端粒酶活性的下降可能是诱发RB细胞凋亡的原因.     

New RB1 oncogenic mutations and intronic polymorphisms in Serbian retinoblastoma patients: genetic counseling implications

The purpose of this work was to identify germ line RB1 mutations in 16 Serbian retinoblastoma patients for genetic counselling. Mutation analysis was carried out by PCR directed sequencing of the 27 exons. Loss of heterozygosity for two RB1 intragenic markers was also analyzed in 14 tumour samples. Five new RB1 oncogenic mutations (g.2078 del C, g.77047_48 del GC, g.78117_8 del TT, g.160797 del T, and g.64439+2 T>C) and two recurrences (R445X and Q383X) have been found in this study. In addition, four intronic variants were observed germ line in some unilateral patients. Two of these variants (g.44668-15T/G, and g.166204-8T/A) are discussed as potential oncogenic mutation candidates. The results show the relevance of studies aimed to investigate the role of intronic variants in exon splicing regulation. Such studies will help to disclose hidden retinoblastoma susceptibilities, important for accurate genetic counselling.     

Correlation of cyclin D1 and E1 expression with bladder cancer presence, invasion, progression, and metastasis

The objective of this study was to determine the correlation of the expression of cyclin D1 and E1 with the expression of commonly altered cell cycle regulators and bladder cancer presence, staging, and clinical outcomes. We performed immunohistochemical staining for cyclin D1, cyclin E1, p53, p21, p27, and retinoblastoma protein (pRB) on serial cuts from normal urothelium from 9 controls, radical cystectomy specimens from 226 consecutive patients with advanced transitional cell carcinoma, and lymph nodes with metastasis from 50 of the 226 cystectomy patients. Cyclin D1 and E1 immunoreactivity were considered low when samples demonstrated less than 10% and 30% nuclear reactivity, respectively. Normal bladder urothelium from all 9 control patients showed uniformly intense expression of cyclin D1 and E1. Cyclin D1 expression was low in 99 (43.8%) of 226 cystectomy specimens and 25 (50.0%) of 50 metastatic lymph node specimens. Cyclin D1 immunoreactivity was not associated with any pathologic characteristics or clinical outcomes. Cyclin E1 expression was low in 125 (55.3%) of 226 cystectomy specimens and 22 (44.0%) of 50 metastatic lymph node specimens. Low cyclin E1 expression was significantly associated with advanced pathologic stage, lymphovascular invasion, and lymph node metastases. In multivariate analyses, low cyclin E1 expression was significantly associated with bladder cancer-specific mortality (P = .048), but not disease recurrence (P = .056). Low cyclin E1 expression was significantly associated with altered expression of pRB, p27, and cyclin D1. Low cyclin D1 expression was significantly associated with altered expression of pRB, p21, and cyclin E1. Cyclin E1 expression stratifies patients with bladder transitional cell carcinoma into those with more "indolent" behavior and those with features of biologically and clinically aggressive disease.     

CT CDFI在视网膜母细胞瘤诊断中的价值

目的评价CT、彩色多普勒血流显像（CDFI）在视网膜母细胞瘤诊断中的价值。方法回顾性分析20例经临床和手术病理证实为视网膜母细胞瘤患者的CT、CDFI表现特点。结果20例患者均发现球后壁肿块,并伴有钙化。CT、CDFI检查分别显示不规则型软组织肿块9眼（42．85％）、7眼（33．33％）,圆形或半圆形软组织肿块11眼（57．15％）、14眼（66．67％）,显示有钙化斑的21眼（100％）、19眼（90．50％）,显示视神经增粗的5眼（23．80％）、5眼（23．80％）,显示有向眶内蔓延的3眼（9．50％）、4眼（19．47％）。结论CT及CDFI检查在视网膜母细胞瘤诊断中各有所长,具有重要的临床应用价值。     

Retinoblastoma trilateral. Correlación de las alteraciones genéticas del gen RB1 y la presencia de quistes de la glándula pineal

ObjetiveTo determine the correlation between the presence of genetic anomalies identified in the RB1 gene and the development of trilateral retinoblastoma.MethodNo patients with primitive neuroectodermal tumour (PNET) were identified out of a total of 206 patients, but there were 17 cases of pineal cysts, of which 11 had a genetic study.ResultsOf the 11 patients who had a genetic study performed, the anomaly in the germinal line was identified in 8 cases, which was equivalent to 100% of the bilateral retinoblastomas, and 25% of the unilateral ones. It is more common to find a germinal mutation in patients with bilateral disease (P=.024). There are no significant differences in the type of anomaly identified, although the nonsense-frameshift type is more frequent in cases with bilateral involvement. Identification of the genetic anomaly is more frequent in patients who have pineal cysts (Fisher test; P=.490). Nine of the 17 patients received systemic chemotherapy (52.29% of the cases), which could be able to prevent the development of PNET. Although a certain trend was observed in all the mentioned parameters, there was a relationship between, the presence of pineal cysts and bilateral disease (Pearson Chi X2: P=.191), a known family history (Fisher test; P=.114) and age of early diagnosis (Fisher test; P=.114). There were no significant differences in the mutation type identified.ConclusionsConsidering pineal cysts as a pre-malignant form of pinealoblastoma, we found a relationship between the germinal line mutation of the RB1 gene and the cases with bilateral or unilateral retinoblastoma.     

磷酸化的细胞周期相关蛋白在大鼠海马神经元发育和老化过程中的表达

目的： 观察正常Wistar大鼠发育及老化过程中海马神经元磷酸化的细胞周期相关蛋白的表达,探讨这一过程中海马神经元的细胞周期的变化以及磷酸化细胞周期相关蛋白在其中的调控作用。方法：采用免疫荧光方法观察不同发育时期(1天、11天、1月、3月、15月) 磷酸化周期素依赖激酶2（CDK2）、磷酸化细胞分裂周期激酶2（CDC2）、磷酸化视网膜母细胞瘤（Rb）蛋白表达的规律,并应用Western blotting方法测定不同阶段大鼠海马内磷酸化CDK2、磷酸化CDC2、磷酸化Rb的含量。结果：在各年龄组中神经元特异性核蛋白（NeuN）阳性细胞数量随着年龄增加而逐渐减少,提示神经元数量随年龄的增加而逐渐减少。磷酸化CDK2、磷酸化Rb阳性细胞的数量随着年龄增加而逐渐增多,老年组增加明显,与其它各组间有显著差异,磷酸化CDC2阳性细胞在各年龄组神经元中表达量均较低;蛋白定量亦提示老年组磷酸化CDK2、磷酸化Rb的含量较其它组明显增高。结论：海马神经元数量随年龄增加逐渐减少,而其中磷酸化CDK2和磷酸化Rb却随年龄的增长逐渐增多,提示老化过程中部分神经元再次进入细胞周期,说明磷酸化细胞周期相关蛋白可能参与了这一过程中海马神经元的凋亡。     

Human papillomavirus type 45 E7 is a transforming protein inducing retinoblastoma protein degradation and anchorage-independent cell cycle progression

High-risk human papillomaviruses (HPV) cause cervical cancer. The biological properties of HPV-45, the third most prevalent high-risk HPV-genotype, are unknown. We demonstrate here that the HPV-45 E7 protein transforms immortalized NIH3T3 fibroblasts, while mutations in either the conserved LXCXE sequence (C28G) or the carboxyl-terminus (Δ87LQQLF91) significantly abolish this activity. To address the mechanisms underlying cell transformation by HPV-45 E7, we investigated its impact on the cell cycle. We show that HPV-45 E7 associates with the hypophosphorylated form of the retinoblastoma protein (pRb) and induces a significant reduction in the pRb half-life which can be blocked by epoxomicin. Moreover, HPV-45 E7 induces anchorage-independent cell cycle progression of NIH3T3 cells and extends the lifespan of primary human keratinocytes. HPV-45 E7C28G did not bind pRb and could neither induce pRb-proteolysis nor promote cell cycle progression. HPV-45 E7Δ87LQQLF91 had intermediate pRb-binding affinity and retained a residual activity to induce the degradation of pRb but lost the capability to promote cell cycle progression in suspension. Another carboxyl-terminal mutant, HPV-45 E7Δ81AEDL84, showed a trend to reduced transforming activity, had reduced pRb-binding activity and lost the capability to induce pRb-degradation; however, this mutant could induce anchorage-independent cell cycle progression with the same efficiency as HPV-45 E7 wild type. In summary, these data suggest that HPV-45 E7 is a transforming protein and that abrogation of cell cycle control contributes to its oncogenic potential.     

Direct association of the HPV16 E7 oncoprotein with cyclin A/CDK2 and cyclin E/CDK2 complexes

The human papillomavirus (HPV) E7 oncoprotein has been shown to associate with cyclin/CDK2 complexes. Here we present evidence that HPV E7 proteins can associate with cyclin A/CDK2 and cyclin E/CDK2 complexes in cells that lack retinoblastoma tumor suppressor family members through sequences outside of the core retinoblastoma tumor suppressor binding site. Moreover, we show that HPV16 E7 can directly associate with cyclin A/CDK2 and cyclin E/CDK2 complexes. These results suggest that cyclin/CDK2 complexes may be components of HPV E7-associated cellular complexes that do not contain retinoblastoma tumor suppressor family members.     

视网膜母细胞瘤的CT与MRI诊断

目的:探讨CT与MRI对视网膜母细胞瘤的诊断价值。方法:16例视网膜母细胞瘤,11例行CT检查,4例行MRI检查,1例同时行CT和MRI检查。回顾性分析其影像学征像。结果:视网膜母细胞瘤的CT征象主要表现为眼环后份或以后份为主的软组织肿块,伴有斑点及斑块状钙化,钙化显示率达92.9%(13/14)。MRI主要表现为眼环内软组织信号肿块,部分病例肿块内可见更低信号影,钙化显示率仅40%(2/5),但MRI对肿瘤侵犯周围结构较敏感。结论:CT与MRI对视网膜母细胞瘤的诊断价值,以CT较具优势,肿块内钙化可以认为是视网膜母细胞瘤定性诊断依据。影像学检查对于临床确定治疗方案和判断预后具有重要临床价值。