野生型p53基因cDNA转染对卵巢癌细胞系SKOV-3生物学行为的影响

目的：研究野生型ｐ５３基因ｃＤＮＡ转染对卵巢癌细胞系ＳＫＯＶ－３生物学行为的影响。方法：用脂质体介导的转染技术，将含有全长人野生型ｐ５３ｃＤＮＡ的真核表达重组质粒和空载体质粒，分别导入不表达ｐ５３的ＳＫＯＶ－３细胞，观察细胞生物学行为的改变。结果：（１）共获２个ｐ５３ｃＤＮＡ转染克隆（ｐＣ５３）和２个空载体转染克隆（ｐＮｅｏ）；（２）ｐＣ５３和ｐＮｅｏ细胞形态学与ＳＫＯＶ－３无显著差别；（３）ｐＣ５３细胞生长明显比ＳＫＯＶ－３慢，而ｐＮｅｏ细胞生长与ＳＫＯＶ－３相当；（４）ｐＣ５３在软琼脂中形成集落数显著少于ＳＫＯＶ－３细胞和ｐＮｅｏ细胞；（５）ｐＣ５３Ｇ１／Ｇ０期细胞百分比高于ＳＫＯＶ－３和ｐＮｅｏ。结论：野生型ｐ５３ｃＤＮＡ可作为卵巢癌基因治疗的目的基因之一。     

p53基因转染对卵巢癌MDR细胞药物敏感性及恶性表型的影响

研究 ｐ５３基因导入已知内源背景的肿瘤 ＭＤＲ细胞所致的恶性表型和ＭＤＲ表型的改变及两者的关系。方法：用磷酸钙沉淀法将含有野生型及全长反义 ｐ５３ｃＤＮＡ的逆转录病毒载体ｐＤＷｐ５３及 ｐＤＡｐ５３转染病毒包装细胞 ＰＡ３１７,测定病毒滴度。用此病毒感染卵巢癌多药耐药细胞株Ａ２７８０／ＡＤＭ,Ｓｏｕｔｈｅｒｎ Ｂｌｏｔ鉴定,检测转导基因后细胞株的恶性度、多药耐药性等情况。结果：野生型及反义全长ｐ５３ｃＤＮＡ均转入 ＰＡ３１７细胞获得效价为（１－ １．５） Ｘ１０５ＣＦＵ／ｍｌ的前病毒,以此感染卵巢癌多药耐药细胞株Ａ２７８０／ＡＤＭ,Ｓｏｕｔｈｅｍ Ｂｌｏｔ证实ｐ５３基因导入该细胞并整合到基因组ＤＮＡ中,进一步测试观察到：①导入野生型ｐ５３基因的Ａ２７８０／ＡＤＭ细胞生长被抑制、恶性度降低,细胞形态和生长曲线改变,软琼脂集落形成率及裸鼠接种成瘤率降低;②细胞多药耐药性减弱,对ＡＤＭ耐药性下降,Ｐ－ｇｐ表达降低;③反义ｐ５３的导入也对Ａ２７８０／ＡＤＭ恶性度有一定的影响;④野生型ｐ５３导致的ＭＤＲ细胞恶性表型与ＭＤＲ水平的降低似有平行关系。结论：ｐ５３基因对肿瘤细胞ｍｄｒ－ １基因的表达可能起调控作用,ｐ５３发生突变的 Ｍ?     

化疗及放疗对卵巢的影响及预防

随治疗方法不断完善及新技术应用,肿瘤患者的生存率明显提高.但如何保持放化疗后各重要器官的生理功能,尤其是年轻患者的卵巢功能这个临床新问题日益引起临床医生的重视.详细综述有关放化疗后卵巢的损害及预防的最新研究动态.     

RNA干扰技术沉默survivin基因对卵巢上皮性癌细胞株SKOV3生物学行为的影响

survivin基因是凋亡抑制蛋白家族（IAP）成员之一,具有肿瘤组织表达特异性,与肿瘤化疗耐药相关,已有研究证明,survivin基因在卵巢肿瘤组织高表达,与肿瘤的恶性程度及预后密切相关.RNA干扰（RNA interference,RNAi）技术具有很强的转录后基因沉默作用,已广泛用于抗肿瘤的研究中.     

卵巢克鲁根勃瘤p53蛋白表达的研究

分析胃肠道粘液腺癌转移至卵巢后Ｐ５３蛋白的表达。方法应用免疫组织化学ＢＳＡ法检测３２例卵的克鲁根勃瘤。结果阳性表达率４０．９％,且以３０岁以下,双侧肿瘤术后存活期短患者的高表达,有统计学意义。结论Ｐ５３蛋白作为独立因素影响卵巢克鲁根勃瘤的生物学行为患者的预后。     

p53基因与白细胞介素6对卵巢癌生物学行为的影响

卵巢癌死亡率居女性生殖系统恶性肿瘤之首，恶性肿瘤的生长与转移，往往是患者死亡的直接原因，也是恶性肿瘤研究中最重要的问题。基础研究证明，癌基因的过度表达、抑癌基因的失活及细胞生长因子调节的异常，是患者体内正常组织癌变并导致其生物学行为一系列改变的基本原...     

p53基因多态性与卵巢上皮性癌发病风险的关系

目的　探讨中国北方汉族妇女卵巢上皮性癌 (卵巢癌 )易感性与 p5 3基因第 4外显子的第 72密码子和第 3内含子多态性的关系。方法　应用序列特异性引物 ,以PCR技术检测 12 4例卵巢癌患者 (卵巢癌组 )和 12 8例健康妇女 (对照组 )的p5 3基因第 4外显子的第 72密码子和第 3内含子的基因型。结果　卵巢癌组和对照组脯氨酸 (Pro)、精氨酸 (Arg)等位基因频率分别为 5 3 2 %、4 6 8%和 4 6 1%、5 3 9%,两组比较 ,差异无显著性 (χ2 =2 5 6 3,P =0 10 9) ;卵巢癌组Pro/Pro、Pro /Arg、Arg/Arg 3种基因型频率分别为 2 9 0 %、4 8 4 %、2 2 6 %,与对照组 (2 1 1%、5 0 0 %、2 8 9%)相比 ,差异也无显著性 (χ2 =2 5 98,P =0 2 73) ;按病理类型分类 ,浆液性癌和宫内膜样癌两者间或分别与对照组间 ,其基因型频率与等位基因频率比较 ,差异均无显著性 (P >0 0 5 ) ;按手术病理分期分类 ,Ⅲ～Ⅳ期卵巢癌患者Arg等位基因及Arg/Arg基因型频率明显高于Ⅰ～Ⅱ期卵巢癌患者 (χ2 =7 4 94 ,P =0 0 0 6和 χ2 =8 318,P =0 0 0 4 )。卵巢癌组及对照组p5 3基因第 3内含子 16bp插入或缺序列 (PIN3)的A、A′等位基因频率分别为 94 8%、5 2 %及 94 5 %、5 5 %,两组比较 ,差异无显著性(χ2 =0 0 13,P =0 910 ) ;两组     

野生型p53基因对人卵巢癌细胞株的生长抑制作用

目的：探讨野生型ｐ５３(ｗｔ ｐ５３)基因对人卵巢癌细胞株的生长抑制作用。方法：利用脂质体介导,将含有人全长ｗｔ－ｐ５３基因ｃＤＮＡ的真核表达载体质粒ｐＣＥＰ４／ｐ５３和空载体质粒ｐＣＥＰ４分别转染卵巢癌ＳＫＯＶ３细胞株,分别命名为ＳＫＯＶ３－ｐＣＥＰ４／ｐ５３细胞(C组)、ＳＫＯＶ３－ｐＣＥＰ４细胞(B组),另设ＳＫＯＶ３细胞为正常对照组（Ａ组）。观察细胞体外生长情况和凋亡变化。结果：外源性ｐ５３基因在Ｃ组细胞中获表达,细胞生长曲线显示ｐ５３的导入使ＳＫＯＶ３细胞生长受到明显抑制,Ｃ组平均细胞集落数（１８．６±１．８个）少于Ａ组（２４．３±２．２个）和Ｂ组（２２．７±２．６）,差异有显著性（Ｐ（005）。ＭＴＴ法检测C组细胞活力明显低于Ａ、Ｂ组。Ｃ组细胞Ｇ０～Ｇ１期百分比（５７．７９％）及凋亡细胞百分比（13．９1％）均高于Ｂ组（４６．０２％、２．０８％）和Ａ组（４３．６２％、０）。结论：ｗｔ－ｐ５３基因可介导ＳＫＯＶ３细胞Ｇ１期停滞和细胞凋亡,抑制细胞体外生长。     

野生型p53基因对卵巢癌细胞株生长及凋亡的影响

目的 探讨野生型p53基因对卵巢癌细胞生长抑制及凋亡的作用，为卵巢癌的基因治疗提供实验依据。方法 构建野生型p53基因重组腺病毒载体、体外转染卵巢癌细胞株CaOV3；应用生化染色、免疫组化、聚合酶链技术，检测外源基因的转染率及表达效果；应用细胞计数、MTT法（四甲基偶氮唑盐微量酶反应比色法）、流式细胞术和TUNEL技术（TDL-mediated dUTP-hiotin end labeling)，检测细胞生长及凋亡的情况。结果 野生型p53基因重组腺病毒载体可以有效地转染卵巢癌细胞株CaOV3，转染后的CaOV3细胞内可以检测到p53基因的cDNA及p53蛋白的表达；转染后的CaOV3细胞生长受到明显抑制，63％的细胞生长停滞于G0／G1期，40％－50％的细胞TUNEL呈阳性。结论 野生型p53基因的导入，可抑制卵巢癌细胞的生长并诱导细胞凋亡。野生型p53基因导入可能成为今后临床基因治疗卵巢癌的可行性方法之一。     

Expression of p53 in epithelial carcinoma of the ovary after chemotherapy.

OBJECTIVE: To compare the expression of p53 protein before and after chemotherapy in epithelial carcinoma of the ovary. STUDY DESIGN: Thirty patients were identified during a 10-year period with both their primary surgery and laparotomy after chemotherapy at the same institution. Tissue was processed from paraffin blocks, exposed to an antihuman p53 monoclonal antibody, then stained. Patient records were reviewed for survival data. RESULTS: Twelve of 30 (40%) patients' tissue initially stained positive for p53. Seventeen of 30 (56.7%) were negative, and one (3.3%) was mixed. Eight of 12 (66.7%) initially positive remained so after chemotherapy, 2 converted to negative, and 2 converted to mixed expression. One patient with mixed expression converted to positive after chemotherapy. Seventeen tissues initially negative remained unchanged. CONCLUSION: Chemotherapy did not permanently alter the expression of p53 detected by immunohistochemical staining in the majority (83.3%) of patients with epithelial ovarian cancer; 16.7% (all initially over-expressing p53) did show some decrease. This may have been due to deactivation of the mutated p53 gene, spontaneous mutation, selective growth within a heterogeneous population or alteration of p53 protein.     

The effects of chemotherapy and radiotherapy on fertility in premenopausal women

With the improved survival rate of childhood and young adult cancer patients, the long-term sequelae of the treatments used are increasingly important. In this review, current knowledge of the gonadotoxicity of commonly employed chemotherapeutic agents and radiotherapy regimens is examined. Differences between the effect of "high-risk" and "low-risk" agents are discussed. Tailoring treatment to suit the individual and counseling patients regarding reduced fertility have resulted in the best practice. Target Audience: Obstetricians & Gynecologists, Family Physicians. Learning Objectives: After completing this CME activity, physicians should be better able to evaluate and use appropriate methods to estimate ovarian reserve, assess the risk of infertility caused by commonly used cytotoxic chemotherapy regimens and radiation, and counsel patients regarding the gonadotoxic effects of cancer treatment.     

Transduction of adenovirus-mediated wild-type p53 after radiotherapy in human cervical cancer cells

OBJECTIVE: Radiotherapy is the mainstay of treatment for advanced cervical cancer and most cervical cancers have evidence of HPV (human papilloma virus) infection, which inactivates the p53 gene. The goal of this study was to determine the effect of combining radiotherapy and Adp53 infection on the growth of cervical carcinoma cells. METHODS: The silta cervical carcinoma cell line contains wild-type p53 and HPV 16 infection. The C33A cell line has a p53 mutation. The Adp53 recombinant adenovirus contains the cytomegalovirus promoter, wild-type p53 cDNA, and a polyadenylation signal in a minigene cassette inserted into the El-deleted region of a modified adenovirus 5. Transduction efficiency was assessed by using an adenovirus containing the Escherichia coli beta-galactosidase gene and expression of wild-type p53 in infected cells was evaluated by Western blot analysis. One group of cells was irradiated prior to infection, the other group received no irradiation, but were either infected with virus or mock-infected. Cells were analyzed for viability 1 to 7 days after infection by using the sulforhodamine B assay. The percentage of cells undergoing apoptosis was determined by using a TUNEL assay. RESULTS: Fifty percent of C33A cells were transduced with 5 muthplicities of infection of virus whereas SiHa cells required 25 multiplicities of infection. Adp53 expression was found 48 h after infection. In the cells treated with both radiation and Adp53 infection growth inhibition was increased compared with inhibition resulting with either treatment alone. The combination treatment also increased the percentage of cells undergoing apoptosis. CONCLUSION: Combining radiotherapy with Adp53 infection increases the inhibition of growth of cervical cancer cells in vitro. This combination treatment has the potential of increasing efficacy and of therapy.     

肿瘤抑制基因p53在卵巢移行细胞癌中的过度表达

采用单克隆本免疫组化技术，对３２例卵巢称行细胞癌组织进行肿瘤抑制基因Ｐ５３表达的检测，分析Ｐ５３过度表达与卵巢移行细胞癌预后诸因素间的相关性。结果：卵巢移行细胞癌Ｐ５３表达率为４１％。     

野生型p53抑制宫颈癌HeLa细胞的研究

目的:研究外源性野生型p53(wtp53)基因对宫颈癌HeLa细胞中的致癌基因环氧合酶-2(COX-2)的影响,以及抑制HeLa细胞生长的作用。方法:用脂质体介导的转染技术将野生型p53基因转染到HeLa细胞。实验分为2组,p53转染组(p53-HeLa组)和空白对照组(HeLa组),wtp53表达用逆转录-多聚酶链反应鉴定,并检测p53mR-NA和COX-2mRNA瞬时转染及稳定转染前后表达的变化。CCK-8染色法绘制细胞生长曲线。结果:野生型p53基因转染后HeLa细胞中的p53mRNA表达增加,COX-2mRNA表达减少。通过对细胞生长曲线的分析,转染后与转染前相比,生长明显减慢(P<0.01),wtp53对HeLa细胞有抑制作用。结论:外源性wtp53基因表达可抑制宫颈癌He-La细胞中COX-2mRNA的表达;wtp53可能通过抑制COX-2对宫颈癌HeLa细胞产生抑制作用,进而阻碍肿瘤的生长。     

卵巢囊腺肿瘤中增生细胞核抗原及p53蛋白表达的意义

用免疫组化ＡＢＣ方法检测了６０例卵巢囊腺肿瘤组织中增生细胞核抗原（ＰＣＮＡ）和ｐ５３蛋白的表达。结果表明：ＰＣＮＡ和ｐ５３蛋白在良性囊腺瘤、交界性囊腺瘤和囊腺癌三类不同性质的病变中表达存在显著的差别（Ｐ〈０．００１，Ｐ〈０．０５）。ｐ５３蛋白与ＰＣＮＡ表达关系密切，高增生指数（ＰＣＮＡ＋＋＋）肿瘤ｐ５３蛋白阳性率高，低增生指数肿瘤（ＰＣＮＡ＋，－）ｐ５３表达率低。认为在卵巢囊腺肿瘤中检测ＰＣＮＡ和     

Assessment of inhibin and p53 in granulosa cell tumors of the ovary

OBJECTIVE: The goal of this work was to determine the cellular content of inhibin and p53 in granulosa cell tumors (GCTs). METHODS: Clinical records of 47 patients (mean age, 54 years; range, 20-85 years) presenting with GCT surgically managed at our institution were abstracted. International Federation of Gynecology stage I was assigned in 39 patients, stage II in 2, and stage III in 6. Concomitant endometrial carcinoma was identified in 6 patients. Mean follow-up was 13.6 years (range, 1 day to 37.6 years). Sections from paraffin-embedded tissue blocks were analyzed immunohistochemically for expression of tissue inhibin and p53 levels. Inhibin expression was graded by intensity and reactivity, and p53, by its presence or absence. RESULTS: The tumors of 27 patients (57%) stained strongly for inhibin intensity and showed >60% reactivity. Decreased intensity and reactivity of inhibin expression were associated with advanced-stage disease (P = 0.05 and P < 0.01, respectively, by Fisher exact test). Expression of p53 was detected in tumors from 27 patients (57%), and immunoreactivity was associated with compromised progression-free survival (P = 0.016, log-rank test). However, the association between p53 immunoreactivity and disease stage was not significant. Absence of p53 expression was significantly associated with concurrent endometrial carcinoma (P = 0.022), suggesting more molecularly intact tumors that retain functional activity. CONCLUSIONS: Although the majority of GCTs show strong expression of inhibin with regard to intensity and reactivity, weak expression is associated with advanced disease but not with decreased progression-free survival. By contrast, expression of p53 is not significantly associated with stage, but increased expression is associated with decreased disease-free survival. Absence of p53 expression appears to be associated with concurrent endometrial carcinoma.     

Effect of p53 gene transfer and cisplatin in a peritonitis carcinomatosa model with p53-deficient ovarian cancer cells.

OBJECTIVE: To determine whether combination treatment consisting of p53 gene transfer and cisplatin (CDDP) improves prognosis of ovarian cancer patients with peritonitis carcinomatosa, we tried this therapy in a peritonitis carcinomatosa model that we developed. METHODS: A human ovarian adenocarcinoma cell line, HRA, which has homozygous deletion of the p53 gene, was used. For p53 gene transfection, we used a recombinant adenovirus carrying a wild-type p53 gene (AxCAp53). To determine the efficiency of the recombinant adenovirus to transduce HRA cells, the cells were infected with AxCALacZ, and the transduced cells were detected by beta-galactosidase staining. The expression of the p53 protein was monitored by Western blot analysis up to 15 days after infection of 50 MOI AxCAp53. The combination effect of AxCAp53 and CDDP was evaluated by 3-(4, 5-dimethelthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. Apoptotic cells were assessed morphologically by staining with Hoechst 33258. For the peritonitis carcinomatosa model in this study, we used severe combined immunodeficiency mice with an intraperitoneal injection of HRA cells. RESULTS: The p53 protein was expressed at 24 h after infection with AxCAp53 and disappeared on the 14th day. The present in vitro study showed that wild-type p53 gene transduction significantly enhanced sensitivity to CDDP and the apoptotic index in HRA cells. A significant survival advantage was observed in the combination treatment of AxCAp53 and CDDP compared with single treatments. However, the repetitious treatment did not show significant survival advantage in the long term. CONCLUSION: The present study suggests that intraperitoneal treatment with AxCAp53 and CDDP is potentially useful as an adjuvant therapeutic modality for peritonitis carcinomatosa, although further study is needed to improve the long-term survival for those patients.