葡萄膜黑色素瘤转移相关基因的表达及意义

目的：探讨肿瘤转移相关基因在葡萄膜黑色瘤中的表达及与浸润能力和病理分型的关系。方法：采用免疫组化方法定理检测96例葡萄膜黑色素瘤体标本中EGFR和nm23蛋白的表达,结果：肿瘤的浸润能力与EGFR的表达呈正相关,与nm23的表达则呈负相关,类上皮细胞型,混合细胞型,梭形细胞型EGFR的表达率依次降低,nm23的表达率依次增高,结论：EGFR和nm23是评价葡萄膜黑色素瘤转移潜能的有效指标。     

端粒酶在葡萄膜恶性黑色素瘤中的表达及意义

目的 检测端粒酶在人眼葡萄膜黑色素瘤中的表达,探讨其在葡萄膜黑色素瘤发病机制中的作用,为其临床治疗提供新的思路.方法 选取16例因患葡萄膜黑色素瘤而行眼球摘除术的患者,取其葡萄膜黑色素瘤组织,采用PCR-ELISA及PCR-PAGE检测其端粒酶的含量及表达.结果 16例患者中15例的端粒酶检测呈阳性表达,占93.8%.PCR-ELISA结果显示其活性绝大多数呈中度、高度表达,仅1例△A值＜0.15.电泳结果显示为多少不等的梯形条带,其中15例显示为4条以上.结论 葡萄膜黑色素瘤患者端粒酶活性明显增高,可能是肿瘤发生、发展的重要原因之一,抑制端粒酶活性有可能成为葡萄膜黑色素瘤的新型疗法之一.     

重视对葡萄膜黑色素瘤侵袭转移机制的研究

葡萄膜黑色素瘤（UM）是成人眼内最常见的恶性肿瘤。肿瘤组织浸润侵袭性强,易经血行途径发生转移,转移后临床预后较差。UM转移存在多种途径,目前认为与染色体异常、基因表达异常、细胞因子及信号转导通路调控异常等有关。通过对UM临床病理学特征及其侵袭转移机制进行研究,寻求检测UM早期转移的标志物及可能的治疗靶点,进行有效地监测和干预,对于提高患者生存率以及UM的防治水平具有重要意义。     

蛋白酶活化受体2在葡萄膜黑色素瘤中的表达及对细胞增殖和侵袭的影响

目的：探讨蛋白酶活化受体2(protease-activated receptor 2,PAR2)在葡萄膜黑色素瘤(uveal melanoma,UM)中的表达,以及沉默人UM 细胞系M23中PAR2基因对细胞增殖和侵袭的影响。

方法：选取2012-02/2017-12在我院行手术治疗且资料完整的UM患者45例45眼,选取同期因眼部外伤行眼球摘除且葡萄膜正常的患者30例30眼,实时荧光定量PCR术检测UM和正常脉络膜组织中PAR2基因表达,培养M23细胞并分为PAR2干扰组、阴性对照序列组和空白组,实时荧光定量PCR技术检测细胞中PAR2基因表达,MTT法检测细胞增殖能力,Transwell法检测细胞迁移和侵袭能力。

结果：UM组织中PAR2 mRNA相对表达量为1.73±0.13,正常脉络膜组织中PAR2 mRNA相对表达量为1.06±0.10,差异有统计学意义(t=23.732,P<0.01); UM组织中PAR2 mRNA相对表达量与病理学类型、巩膜浸润、视盘受累和眼外生长有关,差异有统计学意义(P<0.05); PAR2干扰组细胞中PAR2 mRNA相对表达量低于阴性对照序列和空白组,差异有统计学意义(P<0.05); PAR2干扰组细胞24、48、72和96h时吸光度A值低于阴性对照组和空白组,差异有统计学意义(P<0.05); PAR2干扰组迁移细胞数和侵袭细胞数低于阴性对照序列组和空白组(P<0.05)。

结论：PAR2在UM组织中呈高表达,且与肿瘤高转移风险有关,特异性沉默M23细胞中PAR2基因表达可有效抑制细胞增殖、迁移及侵袭。     

HMGA1在葡萄膜黑色素瘤中的表达及对细胞增殖和侵袭的影响

目的：探讨高迁移率族蛋白A1(HMGA1)蛋白在葡萄膜黑色素瘤(UM)组织中的表达,以及抑制该基因表达对细胞增殖和侵袭的影响。

方法：选取2014-02/2019-08在我院接受手术治疗的UM患者53例53眼,同期留取因外伤摘除眼球的正常葡萄膜组织34例34眼。采用免疫组织化学法检测组织中HMGA1蛋白表达。将体外培养的人UM细胞系M23分为HMGA1下调组、阴性对照组和空白组,分别转染HMGA1干扰序列、阴性对照序列和不作任何处理,采用实时荧光定量PCR术检测HMGA1 mRNA表达,CCK-8法检测细胞增殖能力,Transwell法检测细胞迁移和侵袭能力。

结果：UM组织中HMGA1蛋白阳性表达率为77%,高于正常葡萄膜组织中的29%(P<0.001); 与未发生巩膜浸润、未累及睫状体和未发生眼外生长相比,HMGA1蛋白在发生巩膜浸润、累及睫状体和发生眼外生长的组织中阳性表达率升高(均P<0.05)。与阴性对照组和空白组相比,HMGA1 mRNA在HMGA1下调组细胞中相对表达量降低,且HMGA1下调组细胞培养24、48、72、96h时吸光度OD值降低,迁移细胞数和侵袭细胞数均明显减少(均P<0.05)。

结论：UM组织中HMGA1蛋白阳性表达率升高,下调M23细胞中HMGA1表达可减少细胞增殖,抑制细胞迁移和侵袭。     

E26转录因子-1在不同类型葡萄膜黑色素瘤中的表达及意义

目的　探讨E2 6转录因子 1(E2 6transformation specific 1,Ets 1)在不同类型的葡萄膜黑色素瘤中的表达及其与预后的相关关系。方法　以原位杂交和免疫组织化学法检测 78例葡萄膜黑色素瘤中Ets 1mRNA和蛋白的表达 ,并按WHO 1980年的标准进行分型 :梭型细胞型、类上皮细胞型和混合细胞型。结果　 78例中 ,梭型细胞型占 2 1例 ,类上皮细胞型占 34例 ,混合细胞型占 2 3例。Ets 1mRNA和蛋白在 3种类型的葡萄膜黑色素瘤中均有表达 ,但表达强度随梭型细胞型 ,混合细胞型和类上皮细胞型依次递增。回访37例患者 ,其中梭型细胞型 18例 ,平均生存时间为 (78.33± 2 4 .6 9)月 ;混合细胞型 10例 ,平均生存时间 (6 1.4 4± 2 0 .4 6 )月 ;类上皮细胞型 9例 ,平均生存时间 (36 .76± 12 .19)月。患者生存时间与Ets 1表达强度呈负相关。结论　Ets 1可能在葡萄膜黑色素瘤的转移、浸润中发挥重要作用 ,Ets 1的检测可作为葡萄膜黑色素瘤恶性程度的参考指标。     

MAPK通路中肿瘤转移抑制基因对葡萄膜黑色素瘤侵袭能力的影响

丝裂原活化蛋白激酶(MAPK)通路是进化上高度保守的信号通路,它将细胞外信号转化为细胞增生信号传递到细胞核,具有调控细胞的生长、分化、凋亡、基因表达的功能。MAPK信号通路的正常活化与某些肿瘤转移抑制基因的调控有关。在葡萄膜黑色素瘤中存在着nm23、RKIP、MKK4等肿瘤转移抑制基因异常表达所导致的MAPK通路信号传导异常,这种分子信号通路的异常变化对葡萄膜黑色素瘤的侵袭和转移能力有着重要的影响。     

转化生长因子-β1在葡萄膜黑色素瘤中的表达及意义

目的:探讨转化生长因子-β(1TGF-β1)在葡萄膜黑色素瘤中的表达及意义。方法:以原位杂交法检测78例葡萄膜黑色素瘤中TGF-β1mRNA的表达,并按WHO1980年的标准进行分型:梭型细胞型,类上皮细胞型和混合细胞型。进行临床随访。结果:78例中,梭型细胞型21例,类上皮细胞型34例,混合细胞型23例。TGF-β1mRNA在3种类型的葡萄膜黑色素瘤中均有不同程度的表达,表达强度随梭型细胞型,混合细胞型和类上皮细胞型依次递增。回访37例患者,其中梭型细胞型18例,平均生存时间为(78.3±14.2)mo,混合细胞型10例,平均生存时间(69.1±17.4)mo,类上皮细胞型9例,平均生存时间(36.8±12.2)mo。患者生存时间与TGF-β1表达强度呈负相关。结论:TGF-β1可能在葡萄膜黑色素瘤的转移,浸润中发挥重要作用。     

MicroRNA在葡萄膜恶性黑色素瘤中的研究进展

恶性黑色素瘤恶性程度高,易经血流转移,是成年人中最多见的一种恶性眼内肿瘤,其发生发展涉及到多步骤的分子生物学机制,包括原癌基因和抑癌基因的遗传和表观遗传学改变。因此研究参与此过程的分子机制能为肿瘤的有效治疗提供有益的见解。微小核糖核酸（ｍｉｃｒｏＲＮＡ,ｍｉＲＮＡ）是一类长约２２个核苷酸的非编码单链小分子ＲＮＡ,在各种生理病理过程中发挥了重要作用,最新的研究发现,异常表达的ｍｉｃｒｏＲＮＡ参与了葡萄膜恶性黑色素瘤众多的病理过程。本文综述了ｍｉｃｒｏＲＮＡ的发现、形成及作用机制,葡萄膜恶性黑色素瘤中ｍｉｃｒｏＲＮＡ的异常表达及其可能机制,ｍｉｃｒｏＲＮＡ与葡萄膜恶性黑色素瘤的发生与增殖、侵袭和转移以及ｍｉｃｒｏＲＮＡ在葡萄膜恶性黑色素瘤中的临床应用价值。     

胰岛素样生长因子-1受体在原发性葡萄膜恶性黑色素瘤中的表达及临床意义

目的 观察胰岛素样生长因子-1受体(insulin-like growth factor-1-receptor,IGF-1R)对人葡萄膜恶性黑色素瘤预后评估的作用.方法 用免疫组织化学SABC法检测IGF-1R在41例葡萄膜恶性黑色素瘤组织的表达.用x2检验分析其表达与临床病理参数之间的关系.采用乘积极限法分析生存率,并绘制Kaplan-Meier曲线.建立Cox比例风险模型,探讨影响预后的相关危险因素.结果 41例葡萄膜恶性黑色素瘤标本中,IGF-1R在28例肿瘤的细胞胞浆中表达,阳性率68.3%.x2检验分析显示:IGF-1R的表达水平与肿瘤的细胞类型、最大基底直径、肿瘤的高度显著相关(P<0.05).单因素及多因素分析提示:IGF-1R的表达为影响预后的因素.结论 IGF-1R在葡萄膜恶性黑色素瘤中的高表达是影响预后的因素,可能成为临床判断预后和指导治疗的重要指标.     

Prognostic factors of liver metastases from uveal melanoma

Objectives This study was designed to assess survival and identify prognostic factors for liver metastases diagnosed by systematic screening in uveal melanoma patients. Methods Among 602 consecutive patients treated over 10 years for uveal melanoma and followed by systematic semi-annual hepatic screening (abdominal ultrasonography), 63 (10.5%) developed liver metastases; these patients form the basis of this study. Factors including patient demographics, characteristics of the uveal tumor, metastasis-free interval, severity of liver metastatic involvement, and treatments of metastases were studied retrospectively regarding their prognostic value, using univariate (Kaplan-Meier method) and multivariate (Cox model) analyses. Results Thirty-five patients (55.6% of the metastatic population) received systemic chemotherapy or best supportive care only; 14 patients (22.2% of the metastatic population) diagnosed with diffuse liver involvement had cytoreductive surgery and intra-arterial chemotherapy; 14 (22.2% of the metastatic population) had complete surgical removal of liver metastases followed by postoperative intra-arterial chemotherapy. No significant surgical complications were experienced. The median overall survival after diagnosis of liver metastases was 15 months. It reached 25 months for selected patients with complete resection (P=0.0002). In this cohort of 63 patients, ten or fewer preoperatively diagnosed metastases and primary uveal melanoma not involving the ciliary body were independently associated with better prognosis. Conclusions This study suggests that selected patients with screened liver metastases from uveal melanoma may benefit from aggressive treatment, including surgery. The two independent favorable prognostic factors are fewer than ten metastases at screening and the absence of ciliary body involvement. Laurent Kodjikian had full access to all the data and takes responsibility for the integrity of the data in the study and the accuracy of the data analysis A summary of this work was presented at the 27th congress of the “Societa Italiana di Chirurgia Oncologica” (May 29–31, 2003) and published as an abstract This work was presented at the 24th Meeting of the Club Jules Gonin, September 2004, Athens (Greece)     

The electro-oculogram in uveal melanoma

The electro-oculogram in 52 patients with a suspected malignant melanoma of the choroid or ciliary body was plotted in a diagram constructed for the differential diagnosis of malignant melanoma, metastasis, naevus and retinal detachment. Thirty-one patients were diagnosed as suffering from malignant melanoma on clinical grounds (19 histologically confirmed). Twenty-six were classified correctly as a melanoma using our EOG probability score. Rupture of Bruch's membrane and tumor localization were of no influence on the EOG classification. Accompanying retinal detachment lowered the Lp/Dt-ratio significantly without affecting the Dt, as was also the case in tumors with a prominence equal to or greater than 6mm when compared with smaller tumors. However melanomas were still classified correctly in the majority of the patients by means of EOG. We conclude that an acceptable differentiation can be made between melanomas, retinal detachments and naevi. Melanomas cannot be differentiated from choroidal metastases.     

人葡萄膜黑色素瘤的细胞培养经验及鉴定

目的：探讨人葡萄膜黑色素瘤细胞体外培养的方法及生物学鉴定的研究。方法：分别用机械取材法、化学取材法及组织块培养法,体外培养人葡萄膜黑色素瘤细胞。通过倒置显微镜下观察细胞形态及生长特性,收集第三代融合的细胞经原位固定后作HE染色、HMB45,S-100,神经元特异性烯醇化酶（NSE）免疫组化染色,电子显微镜鉴定培养细胞。结果：三种取材方法中,机械分离的肿瘤细胞活性好、繁殖快、细胞呈梭型或类上皮型,胞核透明呈梭形或椭圆形,胞浆富含黑色素颗粒。HMB45,S-100,NSE呈阳性反应。透射电镜发现肿瘤细胞内不同发育阶段的黑色素小体及大小不等的黑色素颗粒。结论：机械取材法是获取人葡萄膜黑色素瘤细胞体外培养的最佳方法。用免疫组化及电镜可以鉴定细胞来源及纯度。     

Expression of the SST receptor 2 in uveal melanoma is not a prognostic marker

Introduction  Uveal melanoma (UM) cells and neurohormone-producing cells both originate from the neural crest. Somatostatin receptors subtype 2 (SSTR2) are over-expressed in several tumors, often from neuroendocrine origin, and synthetic antagonists like octreotide and octreotate are being used as diagnostic or therapeutic agents. We investigated the SSTR2 expression in UM, and determined whether this expression was related to prognosis of the disease. Materials and methods  UM cell lines and fresh primary UM samples were tested for SSTR2 expression by autoradiography (AR) using 125I-Tyr3-octreotate. Furthermore, UM cell lines were analyzed for SSTR2 mRNA expression with quantitative real-time RT-PCR. Results  Using AR, cell-surface SSTR2 expression was demonstrated in two UM metastatic cell lines, but no expression was detected in three cell lines derived from primary UM. However, all primary and metastatic UM cell lines showed mRNA expression levels for SSTR2 using quantitative real-time RT-PCR. Only three of 14 primary UM demonstrated moderate SSTR2 expression, and this expression was not significantly associated with tumor-free survival or any tested prognostic factor. Conclusions  Based on the rare and low expression of SSTR2 found in primary UM specimens and in UM cell lines, we conclude that SSTR2 is not widely expressed in UM. Furthermore, SSTR2 expression was not associated with tumor-free survival and prognostic factors. Therefore SSTR2 is not suited as prognostic marker or therapeutic target in UM.     

Electro-oculographic changes after local excision of uveal melanoma

Standard electro-oculography was performed in 10 patients with malignant unilateral uveal melanoma All eyes were treated by means of surgical local excision. After a mean ± standard deviation follow-up of 10.5 ± 5.5 months (range, 6 to 20 months) in the five patients with choroidal melanoma, the electro-oculographic recordings, which had appeared reduced or flat before the surgery, increased significantly just after the local excision. Thereafter the amplitudes of the electro-oculographic light peaks were reduced in all cases up to extinction, possibly because of a retinal detachment. On the contrary, the electro-oculographic value in the five melanomas localized at the iris and ciliary body, normal before the surgery, showed a marked and persistent increase after surgery, probably related to an electric hole in the globe.Abbreviations AI Arden index - EOG electro-oculogram     

不同病理类型的葡萄膜黑色素瘤中微小RNA差异表达谱分析

背景 目前研究证实微小RNA(miRNA)参与大多数人类肿瘤疾病的发生和发展,其作用类似于抑癌基因或癌基因.葡萄膜黑色素瘤(UM)是成人常见的眼部恶性肿瘤,其发生和转移机制仍未完全阐明.探讨UM组织中miRNA的差异表达情况有望为UM的靶向治疗提供依据. 目的 筛选不同病理类型的UM组织中特异性miRNA表达谱. 方法 收集于2013年3月至2015年10月在北京同仁医院手术局部切除并经常规组织病理学和免疫组织化学检测证实为梭型细胞型UM的标本4例和上皮细胞型UM标本4例,采用miRNA芯片分别检测2种UM组织中miRNA的表达,收集同期死于非肿瘤疾病的8个供体眼的正常葡萄膜组织作为对照,利用组间差异倍数筛选出差异≥2倍差异表达的miRNA;用在线软件预测差异表达miRNA的靶基因,采用生物信息学方法分析靶基因参与的信号功能通路.采用实时定量PCR法验证芯片检测结果.结果 收集的梭形细胞型和上皮细胞型UM标本经组织病理学检查均得到确诊,免疫组织化学检测梭形细胞型及上皮细胞型UM组织中HMB45、黑色素-A和S-100均呈阳性反应.与正常葡萄膜组织比较,在梭形细胞型UM组织中差异表达的miRNA有109个,其中29个上调,80个下调,上调的miRNA包括miR-146a-5p、miR-25-3p和miR-29b-1-5p,下调的miRNA包括miR-126-5 p、miR-183-5p和miR-96-5p;上皮细胞型UM中差异表达的miRNA有50个,其中23个上调,27个下调,上调的miRNA包括miR-155-5p、miR-210和miR-378 a-5p;下调的miRNA包括miR-199a-5p、miR-143-3p和miR-143-5p.在梭形细胞型和上皮细胞型UM组织中共同上调的miRNA为miR-132-3p、miR-21-5p、miR-34a-5p和miR-34b-5p,共同下调的miRNA为miR-125b-2-3p、miR-126-3p、miR-199a-3p和miR-214-3p.梭形细胞型和上皮细胞型UM组织中差异表达的miRNA所预测的靶基因分别参与癌症通路、丝裂原活化蛋白激酶(MAPK)信号通路、Wnt信号通路、细胞间黏附、胞吞作用、前列腺癌通路、结直肠癌通路和细胞黏附通路.结论 与正常葡萄膜组织相比,梭形细胞型UM和上皮细胞型UM组织中存在多种miRNA的差异表达,梭形细胞型UM和上皮细胞型UM组织之间也存在明显的miRNA差异表达,这些差异表达的miRNA可通过不同的信号转导通路参与调控UM的生物学行为.     

Comparison of primary and secondary enucleation for uveal melanoma

We investigated operative course and post-operative findings of patients undergoing primary enucleation for uveal melanoma versus those requiring secondary enucleation after brachytherapy. A retrospective chart review was performed with IRB approval on patients receiving treatment for uveal melanoma. Patients with enucleation as initial treatment and patients enucleated after plaque brachytherapy were analyzed for demographic data, operative course, and post-enucleation outcome. Further cause analysis for secondary enucleations was investigated. No significant difference was seen in age, laterality, or gender between the primarily enucleated (n = 54) and secondarily enucleated (n = 34) groups. Greater difficulty with surgery was noted in 28/32 (87.5%) of secondary enucleations compared to 1/54 (1.8%) of primary enucleations (p < 0.0001). Operative time was >2 hours in 3/51 (6%) of primary enucleations (vs. 8 of 32, 25%, p = 0.02). Average implant size was similar in the 2 groups (20.6 mm), however 2/34 (6%) of secondary enucleations required dermis fat grafting. Post-enucleation anophthalmic ptosis occurred after 8/49 (16%) of primary cases (vs. 13/30, 43%, p = 0.02) and prosthetic enophthalmos after none (0%) of primary cases (vs. 5/30, 17%, p = 0.006). Class 2 gene expression profile was found in 6/8 (60%) of eyes enucleated for treatment failure. Secondary enucleation performed after plaque brachytherapy was technically more difficult, and had more anophthalmic socket and eyelid complications compared to primary enucleation for uveal melanoma. Primary enucleation may avoid additional surgery and morbidity in a subset of patients with contraindications to plaque brachytherapy.     

Treatment of metastatic uveal melanoma: Review and recommendations

This article reviews the published clinical responses of metastatic uveal melanoma and metastatic cutaneous melanoma with visceral involvement to current therapeutic protocols. Despite isolated patient responses to systemic treatment, no effective treatment currently exists for metastatic uveal melanoma. However, several new approaches involving interferons and interleukin and combination chemotherapy have shown some activity against metastatic cutaneous melanoma. The effectiveness against metastatic uveal melanomas has not been determined. A new approach to intrahepatic administration of chemotherapy also warrants further evaluation because of the high incidence of hepatic involvement with metastatic uveal melanoma. When an effective systemic treatment is found, early administration as an adjuvant to primary treatment may provide the best strategy for control of systemic spread.