受体的表达及临床意义
杨晓宇
中文摘要
目的 研究恶性黑色素瘤瘤组织中IGF-1R的表达及其临床意义.方法 选取26例经病理诊断明确的恶性黑色素瘤的石蜡切片（其中有远处转移的病例18例，无远处转移的病例8例）及20例正常色素痣的石蜡切片.采用EnVision两步法，用兔抗人IGF-1R多克隆抗体检测26例恶性黑色素瘤组织及20例色素痣组织中的IGF-1R 的表达情况，并分析恶性黑色素瘤中的IGF-1R表达与年龄、性别及有无远处转移的关系.应用χ2检验及Fisher确切概率法进行统计，以P<0.05为有统计学意义.结果 应用免疫组化EnVision方法，IGF-1R在26例恶性黑色素瘤组织中阳性表达率为57.7%（15/26）；20例色素痣组织中IGF-1R阳性表达率为25.0%（5/20）.统计学分析表明两种组织中IGF-1R阳性表达差异显著（χ2=4.912，P=0.023,P<0.05）,恶性黑色素瘤组织中IGF-1R的高表达与患者有无远处转移相关，而与性别、年龄无关.结论IGF-1R在恶性黑色素瘤伴有转移的病例中呈高表达，且伴有转移的晚期黑色素瘤存活率较差，故IGF-1R的表达量可以预测原发性黑色素瘤的预后及转移，对恶性黑色素瘤的临床治疗及改善生存率可能具有一定的意义.
[关键词] IGF-1R；免疫组化；恶性黑色素瘤；远处转移
Expression and Clinical significance of Insulin-like Growth Factor-1 Receptor in Malignant Melanoma
ABSTRACT
OBJECTIVE：To investigate the expression and clinical significance of the
insulin-like growth factor-1 receptor (IGF-1R) in malignant melanoma tumor.
METHODS: Twenty-six paraffin-embedded samples of malignant melanoma were obtained and had complete clinical and pathologic data (18 cases have distant metastasis,8 cases have no sign of distant metastasis).Twenty samples of pigmented nevus were also obtained.Envision method was used to dyeing the sheets.PBS was used as a substitute for the primary antibody on the megative control.The expression intensity and distribution of IGF-1R in the above 2 kinds of different tissues of 46 cases under 400 times light microscope were observed.
Six high power fields were taken in each sheet.Statistics was used to analyze the data,and analyzed clinical revelance.RESULTS:EnVision immunohistochemical analysis and immunohistochemistry confirmed that IGF-1R is positive expressed in the cell membrane.The IGF-1R expression positive rate in malignant melanom
-a was 57.7%(15/26),significantly higher than that in pigmented nevus(25.0%,5/
20,p<0.05).Statistical analysis showed that significant difference of IGF-1R posit-
ive expression（χ2=4.912，P=0.023,P<0.05）between the two types of tissues,the high IGF-1R expression in malignant are only related to metastasis.CONCLUSI
-ONS:IGF-1R is positive expressed in the malignant melanoma cell membrane expression. Cases of primary melanoma with distant metastasis showed high expression and low survival rate.the IGF-1R expression can be prediction of primary melanoma metastasis and prognosis of malignant melanoma in clinical treatment.
[Keywords]:IGF-1R;immunohistochemistry;malignant melanoma;distant metas-
tasis
前言
大约有1/3诊断“恶黑”的患者伴有远处转移，其中肝转移占20%[1-3]. IGF-1是生长激素（GH）依赖性生长因子之一,研究表明，IGF-1R水平升高与前列腺癌、乳腺癌、结肠癌等发病率呈明显正相关.而关于胰岛素样生长因子-1R与原发性恶性黑色素瘤的之间的关系尚无相关研究.故本研究旨在探讨胰岛素样生长因子-1R与原发性恶性黑素瘤的发病、转移及预后等之间的关系，以预测恶性黑色素瘤的预后，对其临床治疗有一定的帮助.
1资料与方法
1.1 标本来源:26例病理诊断明确的恶性黑色素瘤标本来自上海第二军医大学附属长海医院病理科存档蜡块，均经10%福尔马林固定，石蜡包埋存档，具备完整的临床和病理资料(其中伴有远处转移的为18例，无远处转移的为8例).20例色素痣标本取自长海医院门诊行色素痣切除术的患者.其H E切片经两位病理医生独立阅片，无或少色素性的疑难病例经HMB45免疫组化染色确诊.
1.2 临床资料:26例恶性黑色素瘤中，男性14例，女性12例;最小年龄32岁，最大年龄76岁，平均年龄为54岁;伴有远处转移者18例，无远处转移者8例.
1.3 方法采用EnVision两步法，兔抗人IGF-1R多克隆抗体稀释度为1 :100，检测IGF-1R在恶性黑色素瘤中的定位和表达水平，分别以PBS代替一抗作为阴性对照，以已知阳性的肝癌组织切片作阳性对照.
2结果判定阳性结果为胞膜内出现深褐色或棕黄色颗粒.(l)染色强度:阴性（-):不着色或显色强度与背景无差别者为阴性;阳性(+): 淡黄、黄或深黄色;中等阳性（++):浅棕黄色;强阳性(+++):深棕黄色.(2)染色频度:每张切片在高倍显微镜(x400)下取6个不同视野各计数200个细胞，按阳性细胞所占的百分率分为:阴性(-)：阳性细胞计数<10%;阳性(+):阳性细胞计数10%-30%;中等阳性(++):阳性细胞计数30%-50%;强阳性(+++):阳性细胞计数>50%.每张切片由两位病理专家独立阅片，若有分歧，则再次阅片，讨论后决定最后结果.
3统计分析
应用SPSS11.0统计软件，采用四格表资料χ2检验及Fisher确切概率法进 行统计，以P<0.05为有统计学意义.
4结果
1、恶性黑色素瘤与色素痣组织中IGF-1R表达的比较
表1 恶性黑色素瘤与色素痣组织中IGF-1R表达的比较
|
组 别 |
例 数 |
IGF-1R |
阳性率 |
|||
|
- |
+ |
++ |
+++ |
|||
|
恶性黑色素瘤 |
26 |
11 8 5 2 |
57.69% |
|||
|
正常色素痣 |
20 |
15 3 1 1 |
25.0%* |
|||
(χ2=4.92，*P=0.O27，*P<0.05）
2、IGF-1R在恶性黑色素瘤中的表达及与临床因素的关系（见表2）
表2 恶性黑色素瘤中的IGF-1R免疫组化染色结果
|
组 别 |
例 数 |
IGF-1R |
阳性例数（%） |
|
- + ++ +++ |
|||
|
性别 男 女 |
14 |
7 3 3 1 |
7（50%） |
|
12 |
4 5 2 1 |
8(66.7%) |
|
|
年龄 ≦50 >50 |
11 15 |
4 5 2 0 7 3 3 2 |
7（63.6%） 8(53.3%) |
|
有转移 |
18 |
5 7 4 2 |
13（72.2%） |
|
无转移 |
8 |
6 1 1 0 |
2(25.0%) |
注：恶性黑色素瘤中IGF-1R的表达与性别无关,P=0.4527，P>0.05.
恶性黑色素瘤中IGF-1R表达与年龄无关,P＝0.7007，P>0.05.
恶性黑色素瘤中IGF-1R与有无转移相关，P=0.0384，P<0.05,有转移的患者中的阳性表达率明显高于无转移的患者.
附图1（色素痣中IGF-1R表达,HE×400）：
色素痣中IGF-1R弱阳性表达 色素痣中IGF-1R强阳性表达
附图2（黑色素瘤中IGF-1R表达,HE×400）
黑色素瘤中IGF-1R弱阳性表达 黑色素瘤中IGF-1R强阳性表达
5、讨 论
胰岛素样生长因子-1（IGF-1）主要在肝脏中产生，其对应的受体即为胰岛素样生长因子-1受体（IGF-1R），胰岛素样生长因子-1R是由a和β两个亚单位通过硫化键连接构成的细胞膜糖蛋白的异四倍体[4，5].IGF-1R一旦与其配体相结合将会激活β亚基酪氨酸激酶，从而导致IGF-1R及一些细胞内蛋白的磷酸化[6].最终将会导致Ras、细胞分裂素(丝裂原)活化蛋白激酶、磷酸肌醇-3-激酶的激活[4].正常细胞的增殖分化和机体的正常发育均受到IGF-1R介导的各信号传导途径的调节，当其介导的信号传导失常时，机体就有可能形成肿瘤.目前已发现肿瘤细胞的凋亡、转化、浸润性生长及转移与IGF-IR有关.细胞表面IGF-R数量减少可诱导细胞的凋亡及生长的停滞[6～8]. 有关文献表明IGF-1R是某些类型的细胞转化表型的形成和保持的前提及肿瘤发生的必要条件[4，9].
目前关于胰岛素样生长因子-1受体在原发性黑色素瘤的发生和发展中所起的作用尚未明确.研究表明IGF-1R在细胞周期的调控、细胞分化、细胞调亡及细胞转化过程中起重要作用[9，10～17].而且大多数研究表明，IGF-1R的高表达往往患者预后较差.xie等在对滑膜肉瘤远处转移的研究中得出IGF-1R在原发病灶中呈高度表达[18].Baserga等提出虽然IGF-1或IGF-2可以刺激肿瘤的生长，但IGF-R在患者的预后及肿瘤的转移中起到了很大的作用[19].结合国内外相关报道，本实验旨在探讨胰岛素样生长因子-1R在恶性黑色素瘤组织中的表达、分布及预后的相关性.
从本实验中我们可以得出IGF-1R在原发性恶性黑色素瘤中存在不同的表达，而高表达与恶性黑色素瘤的预后存在统计学意义.我们的实验主要探讨IGF-1R在黑色素瘤组织中的表达及意义，所以我们以原发性恶性黑色素瘤伴远处转移18例和原发性恶性黑色素瘤无远处转移8例为研究对象，从这方面来讲，我们的实验结果存在偏差，故需要大量的连续样本进行多元素的分析.
Charlotta All-Ericsson等在对原发性恶性黑色素瘤的细胞系（OCM-1、OCM-3和92-1）的体外实验中得出IGF-1R在黑色素瘤细胞系中的表达水平受到糖基化作用的抑制体调控,而IGF-1R活力的抑制与恶性黑色素瘤细胞的活力呈显著正比[21].结合国内外相关文献，已证明抗IGF一1R单克隆抗体封闭IGF一1R，能抑制横纹肌肉瘤Rh30、胰腺癌ASPC-1和COLO-357细胞、乳腺癌细胞MCF-7、结肠癌细胞系CaCo-2和结肠癌细胞生长[22～25].由此可以推测，IGF-1R的表达较高的黑色素瘤患者，有可能采用IGF一1R阻断剂作为治疗预后较差的晚期黑色素瘤患者，为恶性黑色素患者提供多渠道治疗手段.
总之，通过本实验可以发现，IGF-1R在原发性黑色素瘤伴有远处转移的病例中呈高表达，且伴有远处转移的晚期黑色素瘤治疗效果不佳，存活率较差，故测定IGF-1R的表达量可以预测原发性黑色素瘤的预后及转移，对临床治疗可能具有一定的指导意义.
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