Background/Purpose: Angiogenesis is an important contributor to the development of rheumatoid arthritis (RA). Tocilizumab (TCZ), an anti-IL-6 receptor antibody used in the treatment of RA patients, has been shown to exert anti-inflammatory effects. However, its effects on angiogenesis are not fully elucidated, and the molecular mechanisms regulating this effect are unknown.
We aimed to evaluate the expression levels of several microRNA molecules and the concentrations of pro- and anti-angiogenic factors in sera of RA patients before and after the initiation of TCZ treatment and to explore the mechanisms of TCZ action in an in vitro co-culture system.

Methods: Sera from 40 RA patients before and 4 months after the initiation of TCZ treatment were collected. Using commercial ELISA kits, the concentrations of the pro-angiogenic factors EMMPRIN, VEGF, MMP-9, IL-6, NGAL, and of the anti-angiogenic factors endostatin and thrombospondin-1 (Tsp-1) were measured, as were the expression levels of microRNA molecules miR-16-5p, miR-21-5p, miR-132-3p, miR-146a-5p, miR-150-5p, miR-155-5p, miR-203a-3p, miR-221-3p, and miR-323a-3p. In vitro, the levels of secreted EMMPRIN, VEGF MMP-9 and Tsp-1 were measured in a co-culture system of HT1080 fibroblasts and U937 monocytes with and without addition of anti-EMMPRIN blocking antibody.

Results: The mean age of RA patients was 57.5±11.1 years, 33 (82.5%) females, mean disease duration 7.7±5.6 years. Of these patients, 25/40 (62.5%) were classified as “responders” based on EULAR criteria.

Statistically significant reduction in the level of EMMPRIN/CD147 (p=0.035), without significant changes in serum levels of MMP-9, VEGF, MMP-3, MMP-7 and of the anti-angiogenic factors Tsp-1 and endostatin were found 4 months after TCZ treatment. The ratio between EMMPRIN and Tsp-1 calculated for each patient 4 months after initiating TCZ decreased significantly (p=0.031), most notably in responding patients versus non-responders (p=0.033), while the levels of VEGF, MMP-9, Tsp-1, and EMMPRIN were unchanged. Of microRNA, only miR-146a-5p and miR-150-5p levels were significantly increased after 4 months of TCZ treatment relative to treatment initiation (p=0.0178, p=0.0028 respectively), with no difference noted in microRNA levels between patients considered responders and non-responders to TCZ.

In vitro, the accumulation of EMMRPIN, VEGF and MMP-9 in the supernatants was increased by co-culturing the HT1080 fibroblasts and the U937 monocytes (p< 0.05, p< 0.05, P< 0.001 respectively), while the accumulation of the anti-angiogenic factor thrombospondin-1 (Tsp-1) (p< 0.001) and the expression levels of miR-146a-5p were reduced (p< 0.001). Transfection of the miR-146-5p mimic reduced EMMPRIN, VEGF and MMP-9 levels by 1.3, 2.3 and 2.2-fold, respectively (p< 0.05).

Adding anti-EMMPRIN antibody to the co-culture reduced the accumulation of VEGF and MMP-9 in the supernatants by 1.8 and 1.4, respectively (p< 0.05).

Conclusion: Our findings implicate miR-146a-5p in the regulation of EMMPRIN and suggest that TCZ affects angiogenesis through its effects on EMMRPIN and miR-146a-5p.

« Back to ACR Convergence 2021

ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-effect-of-tocilizumab-on-mir-146a-5p-and-emmprin-cd147-in-rheumatoid-arthritis-patients/