Roles of Histone Acetyltransferases CBP/p300 and Transcriptional Factor RORα/REV-ERBα Against TNFα-induced CCL2 Expression in RA-FLSs

Ikumi Okumura¹, Kohsuke Yoshida ¹, Kenta Kaneshiro ², Koto Uchida ³, Arisa Yaekura ⁴, Yuto Oketani ⁵, Kanta Morii ¹, Koji Tateishi ⁶, Yasuhiro Terashima ⁶, Yoshiko Kawasaki ⁷, Nao Shibanuma ⁷, Yoshitada Sakai ⁸ and Akira Hashiramoto ¹, ¹Kobe University Graduate School of Health Sciences, Kobe, Hyogo, Japan, ²Kobe University Graduate School of Health Sciences, Osaka, Osaka, Japan, ³Kobe University Graduate School of Health Sciences, Okayama, Okayama, Japan, ⁴Kobe University Graduate School of Health Sciences, Kawanishi, Hyogo, Japan, ⁵Kobe University Graduate School of Health Sciences, Himeji, Hyogo, Japan, ⁶Kohnan Kakogawa Hospital, Kakogawa, Hyogo, Japan, ⁷Kobe Kaisei Hospital, Kobe, Hyogo, Japan, ⁸Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan

Meeting: 2019 ACR/ARP Annual Meeting

Keywords: chemokines and Cell Migration

Session Information

Date: Sunday, November 10, 2019

Title: Cytokines & Cell Trafficking Poster

Session Type: Poster Session (Sunday)

Session Time: 9:00AM-11:00AM

Background/Purpose: We have reported that TNFα induced expressions of clock gene Bmal1 by up-regulating transcriptional activator RORα and down-regulating repressor REV-ERBα in RA fibroblast-like synoviocytes (RA-FLSs), and that histone acetyltransferases CREB binding protein (CBP) and p300 were involved in this process. A chemokine CCL2, associated with cell proliferation and migration of RA-FLSs, is regulated by RORα and REV-ERBα as well as Bmal1. In this study, we investigate the relation between RORα/REV-ERBα, histone acetyltransferases and TNFα-induced CCL2 expression in RA-FLSs.

Methods: To examine the effect of RORα antagonist SR1001 and REV-ERBα agonist GSK4112 on TNFα-induced CCL2 expression, primary cultured RA-FLSs were treated with SR1001 (20μM) and/or GSK4112 (20μM) in the presence of TNFα (10ng/ml) for 24h.

Next, to clarify the role of histone acetyltransferases on TNFα-induced CCL2, cells were incubated with p300/CBP inhibitor C646 or transfected with both p300 and CBP small interfering RNA (siRNA) before stimulation with TNF-α. Thereafter, intracellular CCL2 mRNA and culture supernatant CCL2 were analyzed by qPCR and ELISA, respectively.

To examine the effect of C646 on TNFα-induced cell migration of RA-FLSs, wound healing assay and immunofluorescence observation were performed by using two different culture supernatant of TNFα-stimulation; DMSO pre-treated control and C646 pre-treated one.

Results: TNFα-induced CCL2 expression was strongly inhibited by simultaneous treatment with both SR1001 and GSK4112, as compared with solo treatment of each agents. Further, CCL2 expressions were also suppressed by pretreatment with C646 and silencing of both Cbp and p300 genes. C646 significantly inhibited both TNFα-induced cell migration and reorganization of the actin cytoskeleton as compared to those with DMSO control (Figure 1).

Conclusion: We newly found that TNFα induced expression of CCL2 through RORα and REV-ERBα, which was associated with CBP/p300 in RA-FLSs, in the same manner as Bmal1.

Results propose these molecules as novel therapeutic targets of RA.

Figure 1

Figure 1. TNFα induced reorganization of the actin cytoskeleton in RA-FLSs -A and B-, while C646 significantly inhibited that -C-.

Disclosure: I. Okumura, None; K. Yoshida, None; K. Kaneshiro, None; K. Uchida, None; A. Yaekura, None; Y. Oketani, None; K. Morii, None; K. Tateishi, None; Y. Terashima, None; Y. Kawasaki, None; N. Shibanuma, None; Y. Sakai, None; A. Hashiramoto, None.

To cite this abstract in AMA style:

Okumura I, Yoshida K, Kaneshiro K, Uchida K, Yaekura A, Oketani Y, Morii K, Tateishi K, Terashima Y, Kawasaki Y, Shibanuma N, Sakai Y, Hashiramoto A. Roles of Histone Acetyltransferases CBP/p300 and Transcriptional Factor RORα/REV-ERBα Against TNFα-induced CCL2 Expression in RA-FLSs [abstract]. Arthritis Rheumatol. 2019; 71 (suppl 10). https://acrabstracts.org/abstract/roles-of-histone-acetyltransferases-cbp-p300-and-transcriptional-factor-ror%ce%b1-rev-erb%ce%b1-against-tnf%ce%b1-induced-ccl2-expression-in-ra-flss/. Accessed .

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