HSP90 limits thrombin‑stimulated IL‑6 synthesis in osteoblast‑like MC3T3‑E1 cells: Regulation of p38 MAPK

Fujita,Kazuhiko; Otsuka,Takanobu; Kawabata,Tetsu; Kainuma,Shingo; Sakai,Go; Matsushima‑Nishiwaki,Rie; Kozawa,Osamu; Tokuda,Haruhiko

doi:10.3892/ijmm.2018.3785

HSP90 limits thrombin‑stimulated IL‑6 synthesis in osteoblast‑like MC3T3‑E1 cells: Regulation of p38 MAPK

Authors:
- Kazuhiko Fujita
- Takanobu Otsuka
- Tetsu Kawabata
- Shingo Kainuma
- Go Sakai
- Rie Matsushima‑Nishiwaki
- Osamu Kozawa
- Haruhiko Tokuda
View Affiliations

Affiliations: Department of Orthopedic Surgery, Nagoya City University Graduate School of Medical Sciences, Nagoya, Aichi 467‑8601, Japan, Department of Pharmacology, Gifu University Graduate School of Medicine, Gifu 501‑1194, Japan
Published online on: July 18, 2018 https://doi.org/10.3892/ijmm.2018.3785
Pages: 2185-2192

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Heat shock protein 90 (HSP90), expressed abundantly in a variety of cell types, is a molecular chaperone, and has a central role in protein homeostasis under stress conditions. In our previous study, it was shown that thrombin stimulates interleukin‑6 (IL‑6) synthesis via p44/p42 mitogen‑activated protein kinase (MAPK) and p38 MAPK in osteoblast‑like MC3T3‑E1 cells, and that Rho‑kinase acts as a positive regulator at a point upstream of p38 MAPK, but not p44/p42 MAPK. The present study investigated whether or not HSP90 is involved in the thrombin‑stimulated synthesis of IL‑6 and examined the mechanism by which HSP90 is involved in MC3T3‑E1 cells. Cultured cells were stimulated by treatment with thrombin. IL‑6 concentrations in MC3T3‑E1 cells were determined using an ELISA assay, and levels of phosphorylated p38 MAPK, p44/p42 MAPK and myosin phosphatase targeting subunit, a substrate of Rho‑kinase; were analyzed by western blotting. The 17‑allylamino‑17demethoxy‑geldanamycin (17‑AAG) and 17‑dimethylamino‑ethylamino‑17‑demethoxy‑geldanamycin (17‑DMAG) HSP90 inhibitors significantly enhanced the thrombin‑stimulated release of IL‑6. Geldanamycin, another inhibitor of HSP90, also upregulated the release and mRNA expression of IL‑6. 17‑AAG and geldanamycin markedly potentiated the thrombin‑induced phosphorylation of p38 MAPK without affecting the phosphorylation of p44/p42 MAPK or myosin phosphatase targeting subunit, a substrate of Rho‑kinase. Additionally, the enhancement by 17‑AAG of the thrombin‑stimulated release of IL‑6 was significantly reduced by SB203580, an inhibitor of p38 MAPK. These results suggested that the thrombin‑stimulated synthesis of IL‑6 was limited by HSP90 in osteoblasts, and that the effects of HSP90 were exerted at the point between Rho‑kinase and p38 MAPK.

View Figures

View References

1	Kular J, Tickner J, Chim SM and Xu J: An overview of the regulation of bone remodeling at the cellular level. Clin Biochem. 45:863–873. 2012. View Article : Google Scholar : PubMed/NCBI
2	Chim SM, Tickner J, Chow ST, Kuek V, Guo B, Zhang G, Rosen V, Erber W and Xu J: Angiogenic factors in bone local environment. Cytokine Growth Factor Rev. 24:297–310. 2013. View Article : Google Scholar : PubMed/NCBI
3	Parfitt AM: Targeted and nontargeted bone remodeling: Relationship to basic multicellular unit origination and progression. Bone. 30:5–7. 2002. View Article : Google Scholar : PubMed/NCBI
4	Hirano T, Yasukawa K, Harada H, Taga T, Watanabe Y, Matsuda T, Kashiwamura S, Nakajima K, Koyama K, Iwamatsu A, et al: Complementary DNA for a novel human interleukin (BSF-2) that induces B lymphocytes to produce immunoglobulin. Nature. 324:73–76. 1986. View Article : Google Scholar : PubMed/NCBI
5	Kwan TS, Padrines M, Théoleyre S, Heymann D and Fortun Y: IL-6, RANKL, TNF-alpha/IL-1: Interrelations in bone resorption pathophysiology. Cytokine Growth Factor Rev. 15:49–60. 2004. View Article : Google Scholar
6	Prystaz K, Kaiser K, Kovtun A, Haffner-Luntzer M, Fischer V, Rapp AE, Liedert A, Strauss G, Waetzig GH, Rose-John S and Ignatius A: Distinct effects of IL-6 classic and trans-signaling in bone fracture healing. Am J Pathol. 188:474–490. 2018. View Article : Google Scholar
7	Sims NA: Cell-specific paracrine actions of IL-6 family cytokines from bone, marrow and muscle that control bone formation and resorption. Int J Biochem Cell Biol. 79:14–23. 2006. View Article : Google Scholar
8	Schopf FH, Biebl MM and Buchner J: The HSP90 chaperone machinery. Nat Rev Mol Cell Biol. 18:345–360. 2017. View Article : Google Scholar : PubMed/NCBI
9	Chiosis G: Targeting chaperones in transformed systems-a focus on Hsp90 and cancer. Expert Opin Ther Targets. 10:37–50. 2006. View Article : Google Scholar : PubMed/NCBI
10	Prodromou C: Mechanisms of Hsp90 regulation. Biochem J. 473:2439–2452. 2016. View Article : Google Scholar : PubMed/NCBI
11	Xu W and Neckers L: Targeting the molecular chaperone heat shock protein 90 provides a multifaceted effect on diverse cell signaling pathways of cancer cells. Clin Cancer Res. 13:1625–1629. 2007. View Article : Google Scholar : PubMed/NCBI
12	Whitesell L and Lindquist SL: HSP90 and the chaperoning of cancer. Nat Rev Cancer. 5:761–772. 2005. View Article : Google Scholar : PubMed/NCBI
13	Schulte TW and Neckers LM: The benzoquinone ansamycin 17-allylamino-17-demethoxygeldanamycin binds to HSP90 and shares important biologic activities with geldanamycin. Cancer Chemother Pharmacol. 42:273–279. 1998. View Article : Google Scholar : PubMed/NCBI
14	Jez JM, Chen JC, Rastelli G, Stroud RM and Santi DV: Crystal structure and molecular modeling of 17-DMAG in complex with human Hsp90. Chem Biol. 10:361–368. 2003. View Article : Google Scholar : PubMed/NCBI
15	Ochel HJ, Eichhorn K and Gademann G: Geldanamycin: The prototype of a class of antitumor drugs targeting the heat shock protein 90 family of molecular chaperones. Cell Stress Chaperones. 6:105–112. 2001. View Article : Google Scholar : PubMed/NCBI
16	Price JT, Quinn JM, Sims NA, Vieusseux J, Waldeck K, Docherty SE, Myers D, Nakamura A, Waltham MC, Gillespie MT and Thompson EW: The heat shock protein 90 inhibitor, 17-allylamino-17-demethoxygeldanamycin, enhances osteoclast formation and potentiates bone metastasis of a human breast cancer cell line. Cancer Res. 65:4929–4938. 2005. View Article : Google Scholar : PubMed/NCBI
17	Mori M, Hitora T, Nakamura O, Yamagami Y, Horie R, Nishimura H and Yamamoto T: Hsp90 inhibitor induces autophagy and apoptosis in osteosarcoma cells. Int J Oncol. 46:47–54. 2015. View Article : Google Scholar
18	Fujita K, Tokuda H, Kuroyanagi G, Yamamoto N, Kainuma S, Kawabata T, Sakai G, Matsushima-Nishiwaki R, Kozawa O and Otsuka T: HSP90 inhibitors potentiate PGF2α-induced IL-6 synthesis via p38 MAPK in osteoblasts. PLoS One. 12:e01778782017. View Article : Google Scholar
19	Kyriakis JM and Avruch J: Mammalian mitogen-activated protein kinase signal transduction pathways activated by stress and inflammation. Physiol Rev. 81:807–869. 2001. View Article : Google Scholar : PubMed/NCBI
20	Lane DA, Philippou H and Huntington JA: Directing thrombin. Blood. 106:2605–2612. 2005. View Article : Google Scholar : PubMed/NCBI
21	Mackie EJ, Loh LH, Sivagurunathan LS, Uaesoontrachoon K, Yoo HJ, Wong D, Georgy SR and Pagel CN: Protease-activated receptors in the musculoskeletal system. Int J Biochem Cell Biol. 40:1169–1184. 2008. View Article : Google Scholar : PubMed/NCBI
22	Kozawa O, Tokuda H, Kaida T, Matsuno H and Uematsu T: Thrombin regulates interleukin-6 synthesis through phosphate-dylcholine hydrolysis by phospholipase D in osteoblasts. Arch Biochem Biophys. 345:10–15. 1997. View Article : Google Scholar : PubMed/NCBI
23	Kato K, Otsuka T, Matsushima-Nishiwaki R, Natsume H, Kozawa O and Tokuda H: Rho-kinase regulates thrombin-stimulated interleukin-6 synthesis via p38 mitogen-activated protein kinase in osteoblasts. Int J Mol Med. 28:653–658. 2011.PubMed/NCBI
24	Sudo H, Kodama HA, Amagai Y, Yamamoto S and Kasai S: In vitro differentiation and calcification in a new clonal osteogenic cell line derived from newborn mouse calvaria. J Cell Biol. 96:191–198. 1983. View Article : Google Scholar : PubMed/NCBI
25	Kozawa O, Tokuda H, Miwa M, Kotoyori J and Oiso Y: Cross-talk regulation between cyclic AMP production and phosphoinositide hydrolysis induced by prostaglandin E2 in osteoblast-like cells. Exp Cell Res. 198:130–134. 1992. View Article : Google Scholar : PubMed/NCBI
26	Laemmli UK: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 227:680–685. 1970. View Article : Google Scholar : PubMed/NCBI
27	Kato K, Ito H, Hasegawa K, Inaguma Y, Kozawa O and Asano T: Modulation of the stress-induced synthesis of hsp27 and alpha B-crystallin by cyclic AMP in C6 rat glioma cells. J Neurochem. 66:946–950. 1996. View Article : Google Scholar : PubMed/NCBI
28	Chen H, Xing J, Hu X, Chen L, Lv H, Xu C, Hong D and Wu X: Inhibition of heat shock protein 90 rescues glucocorticoid-induced bone loss through enhancing bone formation. J Steroid Biochem Mol Biol. 171:236–246. 2017. View Article : Google Scholar : PubMed/NCBI
29	Fukata Y, Amano M and Kaibuchi K: Rho-Rho-kinase pathway in smooth muscle contraction and cytoskeletal reorganization of non-muscle cells. Trends Pharmacol Sci. 22:32–39. 2001. View Article : Google Scholar : PubMed/NCBI
30	Ito M, Nakano T, Erdori F and Hartshorne DJ: Myosin phosphatase: Structure, regulation and function. Mol Cell Biochem. 259:197–209. 2004. View Article : Google Scholar : PubMed/NCBI
31	Cuenda A, Rouse J, Doza YN, Meier R, Cohen P, Gallagher TF, Young PR and Lee JC: SB 203580 is a specific inhibitor of a MAPK homologue which is stimulated by cellular stresses and interleukin-1. FEBS Lett. 364:229–233. 1995. View Article : Google Scholar : PubMed/NCBI
32	Kozawa O, Niwa M, Hatakeyama D, Tokuda H, Oiso Y, Matsuno H, Kato K and Uematsu T: Specific induction of heat shock protein 27 by glucocorticoid in osteoblasts. J Cell Biochem. 86:357–364. 2002. View Article : Google Scholar : PubMed/NCBI
33	Verma S, Goyal S, Jamal S, Singh A and Grover A: Hsp90: Friends, clients and natural foes. Biochimie. 127:227–240. 2016. View Article : Google Scholar : PubMed/NCBI