Fernandez P M, Tabbara S O, Jacobs L K, Manning F C, Tsangaris T N, Schwartz A M, Kennedy K A, Patierno S R
Department of Pharmacology, The George Washington University Medical Center, Washington, DC 20037, USA.
Breast Cancer Res Treat. 2000 Jan;59(1):15-26. doi: 10.1023/a:1006332011207.
The 78 kDa glucose-regulated stress protein GRP78 is induced by physiological stress conditions such as hypoxia, low pH, and glucose deprivation which often exist in the microenvironments of solid tumors. Activation of this stress pathway occurs in response to several pro-apoptotic stimuli. In vitro studies have demonstrated a correlation between induced expression of GRP78 and resistance to apoptotic death induced by topoisomerase II-directed drugs. We were interested in characterizing this protein in human breast lesions for potential implications in chemotherapeutic intervention. Surgical specimens of human breast lesions and paired normal tissues from the same patients were flash frozen for these studies. Total RNA and/or protein were extracted from these tissues and used in northern and/or western blot analyses, respectively, to quantify the relative expression of GRP78. Northern blot analysis indicated that 0/5 benign breast lesions, 3/5 estrogen receptor positive (ER+) breast tumors, and 6/9 estrogen receptor negative (ER-) breast tumors exhibited overexpression of GRP78 mRNA compared to paired normal tissues, with fold overexpressions ranging from 1.8 to 20. Western blot analyses correlated with these findings since 0/5 benign breast lesions, 4/6 ER+ breast tumors, and 3/3 ER- breast tumors overexpressed GRP78 protein with fold overexpressions ranging from 1.8 to 19. Immunohistochemical analysis of these tissues demonstrated that the expression of GRP78 was heterogeneous among the cells comprising different normal and malignant glands, but confirmed the overexpression of GRP78 in most of the more aggressive ER- tumors. These results suggest that some breast tumors exhibit adverse microenvironment conditions that induce the overexpression of specific stress genes that may play a role in resistance to apoptosis and decreased chemotherapeutic efficacy.
78 kDa的葡萄糖调节应激蛋白GRP78是由生理应激条件诱导产生的，如缺氧、低pH值和葡萄糖剥夺，这些情况在实体瘤的微环境中经常存在。该应激途径的激活是对几种促凋亡刺激的反应。体外研究表明，GRP78的诱导表达与拓扑异构酶II导向药物诱导的凋亡死亡抗性之间存在相关性。我们有兴趣在人类乳腺病变中对这种蛋白质进行表征，以探讨其在化疗干预中的潜在意义。为了这些研究，将人类乳腺病变的手术标本和来自同一患者的配对正常组织进行速冻。分别从这些组织中提取总RNA和/或蛋白质，并分别用于Northern印迹和/或Western印迹分析，以量化GRP78的相对表达。Northern印迹分析表明，与配对的正常组织相比，0/5例良性乳腺病变、3/5例雌激素受体阳性（ER+）乳腺肿瘤和6/9例雌激素受体阴性（ER-）乳腺肿瘤表现出GRP78 mRNA的过表达，过表达倍数范围为1.8至20。Western印迹分析与这些结果相关，因为0/5例良性乳腺病变、4/6例ER+乳腺肿瘤和3/3例ER-乳腺肿瘤过表达GRP78蛋白，过表达倍数范围为1.8至19。对这些组织的免疫组织化学分析表明，GRP78的表达在构成不同正常和恶性腺体的细胞中是异质的，但证实了GRP78在大多数侵袭性更强的ER-肿瘤中过表达。这些结果表明，一些乳腺肿瘤表现出不利的微环境条件，可诱导特定应激基因的过表达，这些基因可能在抗凋亡和降低化疗疗效中发挥作用。
