IL-6-accelerated calcification by induction of ROR2 in human adipose tissue-derived mesenchymal stem cells is STAT3 dependent

Shunsuke Fukuyo, Kunihiro Yamaoka, Koshiro Sonomoto, Koichi Oshita, Yosuke Okada, Kazuyoshi Saito, Yasuhiro Yoshida, Tamotsu Kanazawa, Yasuhiro Minami, Yoshiya Tanaka

Research output: Contribution to journalArticle

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Abstract

Objective: The mechanisms of ectopic calcification in inflammatory diseases are poorly understood. We investigated the effects of inflammatory cytokines on the mechanisms of calcification in human adipose tissue-derived mesenchymal stem cells (hADSCs). Methods: The effects of inflammatory cytokines were evaluated using hADSCs cultured in osteoblast induction medium. mRNA expression was measured by real-time PCR and protein levels were measured by western blotting. Cell mineralization was evaluated by Alizarin Red S staining. Results: In hADSCs, administration of IL-6/soluble IL-6 receptor (sIL-6R), TNF or IL-1b accelerated calcification through enhanced expression of an osteoblast differentiation marker, runt-related transcription factor 2 (RUNX2). IL-6/sIL-6R had the greatest effect. The transcription of mRNA for receptor tyrosine kinase-like orphan receptor 2 (ROR2), involved in the non-canonical wingless-type (WNT) MMTV integration site pathway, was increased, while b-catenin expression, an essential factor in the canonical WNT signalling pathway for osteoblast differentiation, did not change. Suppression of signal transducer and activator of transcription 3 (STAT3), but not STAT1, by small interfering RNA (siRNA) exerted a strong inhibitory effect on RUNX2 and ROR2 expression, and inhibited accelerated calcification. Conclusion: IL-6/sIL-6R stimulation accelerated the ROR2/WNT5A pathway in hADSCs in a STAT3- dependent manner, resulting in augmented calcification. These results suggest that the mechanisms of ectopic calcification accelerated by IL-6 in hADSCs may be involved in chronic inflammatory tissues and that IL-6 inhibitors may be beneficial in the treatment of ectopic calcification in inflammatory diseases.

Original languageEnglish
Article numberket496
Pages (from-to)1282-1290
Number of pages9
JournalRheumatology (United Kingdom)
Volume53
Issue number7
DOIs
Publication statusPublished - 2014
Externally publishedYes

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STAT3 Transcription Factor
Mesenchymal Stromal Cells
Adipose Tissue
Interleukin-6
Interleukin-6 Receptors
Osteoblasts
Receptor Tyrosine Kinase-like Orphan Receptors
Cytokines
Catenins
Messenger RNA
Differentiation Antigens
Small Interfering RNA
Real-Time Polymerase Chain Reaction
Transcription Factors
Western Blotting
Staining and Labeling
Proteins

Keywords

  • ADSCs
  • Ectopic calcification
  • IL-6
  • ROR2
  • STAT3
  • WNT5A

ASJC Scopus subject areas

  • Rheumatology
  • Pharmacology (medical)

Cite this

IL-6-accelerated calcification by induction of ROR2 in human adipose tissue-derived mesenchymal stem cells is STAT3 dependent. / Fukuyo, Shunsuke; Yamaoka, Kunihiro; Sonomoto, Koshiro; Oshita, Koichi; Okada, Yosuke; Saito, Kazuyoshi; Yoshida, Yasuhiro; Kanazawa, Tamotsu; Minami, Yasuhiro; Tanaka, Yoshiya.

In: Rheumatology (United Kingdom), Vol. 53, No. 7, ket496, 2014, p. 1282-1290.

Research output: Contribution to journalArticle

Fukuyo, S, Yamaoka, K, Sonomoto, K, Oshita, K, Okada, Y, Saito, K, Yoshida, Y, Kanazawa, T, Minami, Y & Tanaka, Y 2014, 'IL-6-accelerated calcification by induction of ROR2 in human adipose tissue-derived mesenchymal stem cells is STAT3 dependent', Rheumatology (United Kingdom), vol. 53, no. 7, ket496, pp. 1282-1290. https://doi.org/10.1093/rheumatology/ket496
Fukuyo, Shunsuke ; Yamaoka, Kunihiro ; Sonomoto, Koshiro ; Oshita, Koichi ; Okada, Yosuke ; Saito, Kazuyoshi ; Yoshida, Yasuhiro ; Kanazawa, Tamotsu ; Minami, Yasuhiro ; Tanaka, Yoshiya. / IL-6-accelerated calcification by induction of ROR2 in human adipose tissue-derived mesenchymal stem cells is STAT3 dependent. In: Rheumatology (United Kingdom). 2014 ; Vol. 53, No. 7. pp. 1282-1290.
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abstract = "Objective: The mechanisms of ectopic calcification in inflammatory diseases are poorly understood. We investigated the effects of inflammatory cytokines on the mechanisms of calcification in human adipose tissue-derived mesenchymal stem cells (hADSCs). Methods: The effects of inflammatory cytokines were evaluated using hADSCs cultured in osteoblast induction medium. mRNA expression was measured by real-time PCR and protein levels were measured by western blotting. Cell mineralization was evaluated by Alizarin Red S staining. Results: In hADSCs, administration of IL-6/soluble IL-6 receptor (sIL-6R), TNF or IL-1b accelerated calcification through enhanced expression of an osteoblast differentiation marker, runt-related transcription factor 2 (RUNX2). IL-6/sIL-6R had the greatest effect. The transcription of mRNA for receptor tyrosine kinase-like orphan receptor 2 (ROR2), involved in the non-canonical wingless-type (WNT) MMTV integration site pathway, was increased, while b-catenin expression, an essential factor in the canonical WNT signalling pathway for osteoblast differentiation, did not change. Suppression of signal transducer and activator of transcription 3 (STAT3), but not STAT1, by small interfering RNA (siRNA) exerted a strong inhibitory effect on RUNX2 and ROR2 expression, and inhibited accelerated calcification. Conclusion: IL-6/sIL-6R stimulation accelerated the ROR2/WNT5A pathway in hADSCs in a STAT3- dependent manner, resulting in augmented calcification. These results suggest that the mechanisms of ectopic calcification accelerated by IL-6 in hADSCs may be involved in chronic inflammatory tissues and that IL-6 inhibitors may be beneficial in the treatment of ectopic calcification in inflammatory diseases.",
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T1 - IL-6-accelerated calcification by induction of ROR2 in human adipose tissue-derived mesenchymal stem cells is STAT3 dependent

AU - Fukuyo, Shunsuke

AU - Yamaoka, Kunihiro

AU - Sonomoto, Koshiro

AU - Oshita, Koichi

AU - Okada, Yosuke

AU - Saito, Kazuyoshi

AU - Yoshida, Yasuhiro

AU - Kanazawa, Tamotsu

AU - Minami, Yasuhiro

AU - Tanaka, Yoshiya

PY - 2014

Y1 - 2014

N2 - Objective: The mechanisms of ectopic calcification in inflammatory diseases are poorly understood. We investigated the effects of inflammatory cytokines on the mechanisms of calcification in human adipose tissue-derived mesenchymal stem cells (hADSCs). Methods: The effects of inflammatory cytokines were evaluated using hADSCs cultured in osteoblast induction medium. mRNA expression was measured by real-time PCR and protein levels were measured by western blotting. Cell mineralization was evaluated by Alizarin Red S staining. Results: In hADSCs, administration of IL-6/soluble IL-6 receptor (sIL-6R), TNF or IL-1b accelerated calcification through enhanced expression of an osteoblast differentiation marker, runt-related transcription factor 2 (RUNX2). IL-6/sIL-6R had the greatest effect. The transcription of mRNA for receptor tyrosine kinase-like orphan receptor 2 (ROR2), involved in the non-canonical wingless-type (WNT) MMTV integration site pathway, was increased, while b-catenin expression, an essential factor in the canonical WNT signalling pathway for osteoblast differentiation, did not change. Suppression of signal transducer and activator of transcription 3 (STAT3), but not STAT1, by small interfering RNA (siRNA) exerted a strong inhibitory effect on RUNX2 and ROR2 expression, and inhibited accelerated calcification. Conclusion: IL-6/sIL-6R stimulation accelerated the ROR2/WNT5A pathway in hADSCs in a STAT3- dependent manner, resulting in augmented calcification. These results suggest that the mechanisms of ectopic calcification accelerated by IL-6 in hADSCs may be involved in chronic inflammatory tissues and that IL-6 inhibitors may be beneficial in the treatment of ectopic calcification in inflammatory diseases.

AB - Objective: The mechanisms of ectopic calcification in inflammatory diseases are poorly understood. We investigated the effects of inflammatory cytokines on the mechanisms of calcification in human adipose tissue-derived mesenchymal stem cells (hADSCs). Methods: The effects of inflammatory cytokines were evaluated using hADSCs cultured in osteoblast induction medium. mRNA expression was measured by real-time PCR and protein levels were measured by western blotting. Cell mineralization was evaluated by Alizarin Red S staining. Results: In hADSCs, administration of IL-6/soluble IL-6 receptor (sIL-6R), TNF or IL-1b accelerated calcification through enhanced expression of an osteoblast differentiation marker, runt-related transcription factor 2 (RUNX2). IL-6/sIL-6R had the greatest effect. The transcription of mRNA for receptor tyrosine kinase-like orphan receptor 2 (ROR2), involved in the non-canonical wingless-type (WNT) MMTV integration site pathway, was increased, while b-catenin expression, an essential factor in the canonical WNT signalling pathway for osteoblast differentiation, did not change. Suppression of signal transducer and activator of transcription 3 (STAT3), but not STAT1, by small interfering RNA (siRNA) exerted a strong inhibitory effect on RUNX2 and ROR2 expression, and inhibited accelerated calcification. Conclusion: IL-6/sIL-6R stimulation accelerated the ROR2/WNT5A pathway in hADSCs in a STAT3- dependent manner, resulting in augmented calcification. These results suggest that the mechanisms of ectopic calcification accelerated by IL-6 in hADSCs may be involved in chronic inflammatory tissues and that IL-6 inhibitors may be beneficial in the treatment of ectopic calcification in inflammatory diseases.

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