Hombach-Klonisch S, Buchmann J, Sarun S, Fischer B, Klonisch T
Department of Anatomy and Cell Biology, Martin Luther University Faculty of Medicine, Halle (Saale), Germany.
Cancer. 2000 Dec 1;89(11):2161-8.
Expression of relaxin-like factor (RLF), a member of the relaxin family, was studied in normal, benign, and malignant neoplastic human breast tissue.
Reverse transcription polymerase chain reaction (RT-PCR) and nonradioactive in situ hybridization were employed to detect RLF transcripts. RLF epitopes were detected with a rabbit polyclonal antiserum generated against the putative receptor binding domain of human RLF. The RLF antiserum was characterized by Western blot analysis on human testicular and placental tissues, recombinant glutathione S-transferase-RLF fusion protein, and baculovirus-derived recombinant marmoset-RLF and marmoset-relaxin.
RT-PCR analysis revealed RLF amplicons in a cDNA library of normal human breast tissue and in malignant neoplastic breast tissue. RLF hybridization signals were localized exclusively in the tubuloalveolar and ductal breast epithelium but were absent in stromal cells. Benign breast disease displayed weaker RLF hybridization signals compared with normal tubuloalveolar breast tissue. Malignant transformation of breast epithelial tissues resulted in down-regulation of RLF gene expression. The weakest expression of RLF mRNA was observed in lymph node metastases of corresponding primary ductal carcinomas. Immunoreactive RLF was exclusively expressed in breast epithelial cells. Despite strong RLF hybridization signals, the tubuloalveolar epithelial cells of normal breast tissue displayed only very weak immunoreactive RLF. Benign breast disease showed clearly detectable levels of both RLF mRNA and immunoreactive protein. In contrast, epithelial cells in breast carcinoma and lymph node metastases displayed strong expression of immunoreactive RLF, although expression of RLF transcripts was weak.
Results demonstrated that transcriptional and posttranscriptional mechanisms affected human RLF gene expression in normal and neoplastic epithelial breast cells.
研究了松弛素家族成员松弛素样因子（RLF）在人正常乳腺组织、良性和恶性肿瘤组织中的表达情况。
采用逆转录聚合酶链反应（RT-PCR）和非放射性原位杂交检测RLF转录本。用针对人RLF假定受体结合域产生的兔多克隆抗血清检测RLF表位。通过对人睾丸和胎盘组织、重组谷胱甘肽S-转移酶-RLF融合蛋白以及杆状病毒衍生的重组狨猴-RLF和狨猴松弛素进行蛋白质印迹分析，对RLF抗血清进行了鉴定。
RT-PCR分析显示，在人正常乳腺组织和恶性肿瘤乳腺组织的cDNA文库中存在RLF扩增子。RLF杂交信号仅定位在乳腺的小管泡状和导管上皮细胞中，而基质细胞中没有。与正常小管泡状乳腺组织相比，良性乳腺疾病显示出较弱的RLF杂交信号。乳腺上皮组织的恶性转化导致RLF基因表达下调。在相应原发性导管癌的淋巴结转移中观察到RLF mRNA的表达最弱。免疫反应性RLF仅在乳腺上皮细胞中表达。尽管RLF杂交信号很强，但正常乳腺组织的小管泡状上皮细胞仅显示出非常弱的免疫反应性RLF。良性乳腺疾病显示出RLF mRNA和免疫反应性蛋白的可检测水平。相比之下，乳腺癌和淋巴结转移中的上皮细胞显示出强烈的免疫反应性RLF表达，尽管RLF转录本的表达较弱。
结果表明，转录和转录后机制影响人正常和肿瘤性乳腺上皮细胞中RLF基因的表达。
