Stope Matthias B, Klinkmann Gerd, Diesing Karoline, Koensgen Dominique, Burchardt Martin, Mustea Alexander
Department of Urology, University Medicine Greifswald, Ferdinand-Sauerbruch-Strasse, 17475 Greifswald, Germany.
Department of Gynaecology and Obstetrics, University Medicine Greifswald, Ferdinand-Sauerbruch-Strasse, 17475 Greifswald, Germany.
Dis Markers. 2017;2017:1575374. doi: 10.1155/2017/1575374. Epub 2017 Feb 23.
The heat shock protein HSP27 has been correlated in ovarian cancer (OC) patients with aggressiveness and chemoresistance and, therefore, represents a promising potential biomarker for OC diagnosis, prognosis, and treatment response. Notably, secretion of soluble HSP27 has been described by a few cell types and may take place as well in OC cells. Therefore, we studied HSP27 secretion mechanisms under diverse cellular conditions in an OC cell model system. Secretion of HSP27 was characterized after overexpression of HSP27 by transfected plasmids and after heat shock. Intra- and extracellular HSP27 amounts were assessed by Western blotting and ELISA. Protein secretion was blocked by brefeldin A and the impact of the HSP27 phosphorylation status was analyzed overexpressing HSP27 phosphomutants. The present study demonstrated that HSP27 secretion by OVCAR-3 and SK-OV-3 cells depends on intracellular HSP27 concentrations. Moreover, HSP27 secretion is independent of the endoplasmic reticulum secretory pathway and HSP27 phosphorylation. Notably, analysis of OC cell-born exosomes not only confirmed the concentration-dependent correlation of HSP27 expression and secretion but also demonstrated a concentration-dependent incorporation of HSP27 protein into exosomes. Thus, secreted HSP27 may become more important as an extracellular factor which controls the tumor microenvironment and might be a noninvasive biomarker.
热休克蛋白HSP27已被证实与卵巢癌（OC）患者的侵袭性和化疗耐药性相关，因此，它是一种很有前景的OC诊断、预后及治疗反应的潜在生物标志物。值得注意的是，已有少数细胞类型描述了可溶性HSP27的分泌情况，OC细胞中也可能发生这种情况。因此，我们在OC细胞模型系统中研究了不同细胞条件下HSP27的分泌机制。通过转染质粒过表达HSP27以及热休克后，对HSP27的分泌进行了表征。通过蛋白质免疫印迹法和酶联免疫吸附测定法评估细胞内和细胞外HSP27的含量。用布雷菲德菌素A阻断蛋白质分泌，并通过过表达HSP27磷酸化突变体分析HSP27磷酸化状态的影响。本研究表明，OVCAR-3和SK-OV-3细胞分泌HSP27取决于细胞内HSP27的浓度。此外，HSP27的分泌不依赖于内质网分泌途径和HSP27的磷酸化。值得注意的是，对OC细胞产生的外泌体的分析不仅证实了HSP27表达与分泌的浓度依赖性相关性，还证明了HSP27蛋白以浓度依赖性方式掺入外泌体。因此，分泌型HSP27作为一种控制肿瘤微环境的细胞外因子可能变得更加重要，并且可能是一种非侵入性生物标志物。
