Bugide Suresh, Gonugunta Vijay Kumar, Penugurti Vasudevarao, Malisetty Vijaya Lakshmi, Vadlamudi Ratna K, Manavathi Bramanandam
Department of Biochemistry, School of Life Sciences, University of Hyderabad, Hyderabad, 500046, India.
Department of Obstetrics and Gynecology, UT Health Science Center, San Antonio, USA.
Cell Oncol (Dordr). 2017 Apr;40(2):133-144. doi: 10.1007/s13402-016-0308-2. Epub 2016 Dec 30.
Hematopoietic PBX interacting protein (HPIP), a scaffold protein, is known to regulate the proliferation, migration and invasion in different cancer cell types. The aim of this study was to assess the role of HPIP in ovarian cancer cell migration, invasion and epithelial-mesenchymal transition (EMT), and to unravel the mechanism by which it regulates these processes.
HPIP expression was assessed by immunohistochemistry of tissue microarrays containing primary ovarian tumor samples of different grades. OAW42, an ovarian carcinoma-derived cell line exhibiting a high HPIP expression, was used to study the role of HPIP in cell migration, invasion and EMT. HPIP knockdown in these cells was achieved using a small hairpin RNA (shRNA) approach. Cell migration and invasion were assessed using scratch wound and transwell invasion assays, respectively. The extent of EMT was assessed by determining the expression levels of Snail, Vimentin and E-cadherin using Western blotting. The effect of HPIP expression on AKT and MAPK activation was also investigated by Western blotting. Cell viabilities in response to cisplatin treatment were assessed using a MTT assay, whereas apoptosis was assessed by determining caspase-3 and PARP cleavage in ovarian carcinoma-derived SKOV3 cells.
We found that HPIP is highly expressed in high-grade primary ovarian tumors. In addition, we found that HPIP promotes the migration, invasion and EMT in OAW42 cells and induces EMT in these cells via activation of the PI3K/AKT pathway. The latter was found to lead to stabilization of the Snail protein and to repression of E-cadherin expression through inactivation of GSK-3β. We also found that HPIP expression confers cisplatin resistance to SKOV3 cells after prolonged exposure and that its subsequent knockdown decreases the viability of these cells and increases caspase-3 activation and PARP proteolysis in these cells following cisplatin treatment.
From these results we conclude that HPIP expression is associated with high-grade ovarian tumors and may promote their migration, invasion and EMT, a process that is associated with metastasis. In addition, we conclude that HPIP may serve as a potential therapeutic target for cisplatin resistant ovarian tumors.
造血 PBX 相互作用蛋白（HPIP）是一种支架蛋白，已知其可调节不同癌细胞类型的增殖、迁移和侵袭。本研究旨在评估 HPIP 在卵巢癌细胞迁移、侵袭和上皮-间质转化（EMT）中的作用，并阐明其调节这些过程的机制。
通过对包含不同分级原发性卵巢肿瘤样本的组织微阵列进行免疫组织化学来评估 HPIP 的表达。使用 OAW42，一种高表达 HPIP 的卵巢癌衍生细胞系，来研究 HPIP 在细胞迁移、侵袭和 EMT 中的作用。通过小发夹 RNA（shRNA）方法在这些细胞中敲低 HPIP。分别使用划痕伤口试验和 Transwell 侵袭试验评估细胞迁移和侵袭。通过蛋白质印迹法测定 Snail、波形蛋白和 E-钙黏蛋白的表达水平来评估 EMT 的程度。还通过蛋白质印迹法研究了 HPIP 表达对 AKT 和 MAPK 激活的影响。使用 MTT 试验评估顺铂处理后细胞的活力，而通过测定卵巢癌衍生的 SKOV3 细胞中的半胱天冬酶-3 和 PARP 裂解来评估细胞凋亡。
我们发现 HPIP 在高级别原发性卵巢肿瘤中高表达。此外，我们发现 HPIP 促进 OAW42 细胞的迁移、侵袭和 EMT，并通过激活 PI3K/AKT 途径在这些细胞中诱导 EMT。发现后者导致 Snail 蛋白稳定，并通过使 GSK-3β失活来抑制 E-钙黏蛋白的表达。我们还发现，长时间暴露后，HPIP 表达使 SKOV3 细胞对顺铂产生抗性，其随后的敲低降低了这些细胞的活力，并增加了顺铂处理后这些细胞中的半胱天冬酶-3 激活和 PARP 蛋白水解。
从这些结果我们得出结论，HPIP 表达与高级别卵巢肿瘤相关，可能促进其迁移、侵袭和 EMT，这一过程与转移相关。此外，我们得出结论，HPIP 可能作为顺铂耐药卵巢肿瘤的潜在治疗靶点。
