ABSTRACT
Stress-induced premature cell senescence is well recognized to be accompanied by emerging the senescence-associated secretory phenotype (SASP). Secreted SASP factors can promote the senescence of normal neighboring cells through autocrine/paracrine pathways and regulate the senescence response, as well. Regarding human endometrium-derived mesenchymal stem cells (MESCs), the SASP regulation mechanisms as well as paracrine activity of senescent cells have not been studied yet. Here, we examined the role of insulin-like growth factor binding protein 3 (IGFBP3) in the paracrine senescence induction in young MESCs. The H2O2-induced premature senescence of MESCs led to increased IGFBP3 in conditioned media (CM). The inhibitory analysis of both MAPK and PI3K signaling pathways showed that IGFBP3 releasing from senescent cells is mainly regulated by PI3K/Akt pathway activity. IGFBP3 appears to be an important senescence-mediating factor as its immunodepletion from the senescent CM weakened the pro-senescent effect of CM on young MESCs and promoted their growth. In contrast, young MESCs acquired the senescence phenotype in response to simultaneous addition of recombinant IGFBP3 (rIGFBP3). The mechanism of extracellular IGFBP3 internalization was also revealed. The present study is the first to demonstrate a significant role of extracellular IGFBP3 in paracrine senescence induction of young MESCs.
Subject(s)
Endometrium/cytology , Endometrium/metabolism , Mesenchymal Stem Cells/metabolism , Paracrine Communication , Cellular Senescence , Endocytosis , Female , Humans , Hydrogen Peroxide/metabolism , Hydrogen Peroxide/pharmacology , Insulin-Like Growth Factor Binding Protein 3/genetics , Insulin-Like Growth Factor Binding Protein 3/metabolism , MAP Kinase Signaling System , Phosphatidylinositol 3-Kinases , Protein Transport , Proto-Oncogene Proteins c-akt/metabolism , Signal TransductionABSTRACT
BACKGROUND INFORMATION: Heat-inducible Hsp72 is the founding member of the Hsp70 (heat shock proteins of 70 kDa) family of molecular chaperones. It is localized primarily in cytoplasm and nucleus but is also found extracellularly. The source of e-Hsp72 (extracellular Hsp72) is not precisely identified and may not be the same in every situation. A number of studies demonstrated that e-Hsp72 plays an important role in cell survival, tumour rejection and immune response. However, currently little is known about regulation of e-Hsp72 function. In cells, Hsp72 is controlled by co-chaperones. An abundant co-chaperone, HspBP1 (Hsp72-binding protein 1) was found extracellularly in the serum. In the present study we analysed the secretion and function of e-HspBP1 (extracellular HspBP1). RESULTS: A431 human squamous carcinoma cells accumulated Hsp72 and HspBP1 in chromogranin A-positive granules following heat stress or in the presence of U73122, an inhibitor of phospholipase C. Following these treatments, A431 cells also increased the secretion of both proteins into the culture medium. The secreted e-Hsp72 and e-HspBP1 were co-immunoprecipitated from the conditioned medium. Purified recombinant HspBP1 augmented e-Hsp72-mediated phosphorylation of EGFR (epidermal growth factor receptor) and its down-stream targets, ERK1 (extracellular signal-regulated kinase 1) and ERK2 in a concentration-dependent manner. Finally, a HspBP1 N-terminal domain deletion mutant and boiled recombinant HspBP1 did not affect the e-Hsp72-mediated activity. CONCLUSIONS: Heat stress and PLC (phospholipase C) inhibition result in the enhanced secretion of both Hsp72 and HspBP1. In an extracellular environment, the two chaperones interact both physically and functionally, leading to the activation of th EGFR-ERK1/2 signalling pathway. However, the magnitude of EGFR activation depends on the e-HspBP1/e-Hsp72 ratio in the medium. Extracellular chaperone-mediated activation of EGFR can provide a survival advantage to cells under heat shock and other stresses.
