Analysis of ACTH receptor in adrenal tumor and tumor-like lesion.

我们从ACTH受体蛋白中选择了15个氨基酸序列。将合成肽偶联到KLH上作为载体蛋白，免疫家兔。酶联免疫吸附试验（ELISA）结果显示，在1：25，600稀释度下呈阳性反应。我们评估了人肾上腺组织石蜡切片的免疫组织化学反应性。其中一种抗体与肾上腺皮质细胞的细胞膜反应，Western印迹分析显示在约60 kDa处有一条带。用免疫肽预吸收可消除两种阳性反应。在肾上腺皮质癌、腺瘤和增生的石蜡切片上进行免疫组化反应。PCNA和Ki-67在腺癌中的反应性分别为3.43-20.08和0 ~ 20.9，在腺瘤中的反应性分别为0.14-1.33和0 ~ 1.03，在增生中的反应性分别为0.19-0.7和0 ~ 0.05。肿瘤生物学恶性程度与增殖活性有显著相关性。ACTHR在癌组织中的表达低于对照组， ...更多信息 腺瘤和增生。p53在3例癌组织中表达阳性，在腺瘤和增生组织中均为阴性。提示p53与胃癌的发生有关，但其他因素尚需进一步研究。Ras在所有病例中均为阴性。myc抗体（9 E10）可与正常肾上腺皮质细胞核、增生和腺瘤细胞核及部分胞浆反应，癌细胞主要与胞浆反应。斑点杂交检测10 pg。我们用TT二聚体探针对经甲基绿-派洛宁染色和28 SrRNA原位杂交证实为RNA保存的人肾上腺组织石蜡切片进行了原位杂交（ISH）研究。为了获得更高的反应活性，我们用地高辛探针通过ISH尝试了2种不同的寡聚DNA。结果与TT二聚体探针相似。肿瘤性病变尚未显示出特异性反应。少

We selected 15 amino acid sequences from the ACTH receptor protein. The symtthesized peptides were coupled to KLH as carrier protein and was immunized to rabbits. ELISA showed positive reaction to the peptide at the dilution of 1 : 25,600. We evaluated the immunohistochemical reactivity on the paraffin sections of human adrenal tissue. One of the antibody reacted with the cell membrane of adrenocortical cells and the Western blot analysis revealed one band at approximately 60 kDa. Both positive reactions were abolished by preabsorption with the immunizing peptide.Immunohistochemical reactivity was evaluated on the paraffin sections of the adrenocortical carcinoma, adenoma and hyperplasia. The reactivity for the PCNA and Ki-67 were 3.43-20.08 and 0-20.9 in carcinoma, 0.14-1.33 and 0-1.03 in adenoma, 0.19-0.7 and 0-0.05 in hyperplasia, respectively. The correlation of the biological malignant grading and proliferative activity was significant. ACTHR expression in carcinoma was lower than … More those in adenoma and hyperplasia. p53 was positive in three of carcinoma and negative in all of adenoma and hyperplasia. The relation between the carcinogenesis and p53 was suggested, however, the other factors are still to be considered. Ras was negative in all cases. The myc antibody (9E10) reacted with nuclei of normal adrenal cortical cells, nuclei and some cytoplasm in hyperplasia and adenoma, and predominantly cytoplasm in carcinoma.We synthesized the oligo-DNA of 60mer with a selected sequence from the ACTHR DNA with additional 5 repeater of ATT sequence. Dot blot hybridization detected l0pg. We examined the in situ hybridization (ISH) using TT dimerized probe, on the paraffin section of the human adrenal tissue that was confirmed RNA preservation by methylgreen-pyronin stain and ISH of 28s rRNA.Compact cells in zona glomerulosa and zona fasciculata showed weak positivity. For more reactivity, we tried 2 different oligo-DNAs by ISH with digoxygenin-probe. They showed similar result to TT dimerized probe. Neoplastic lesion have not showed specific reactivity yet. Less