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Updated: Jul 26, 2025
An Adoptive Transfer Model of Rheumatoid Arthritis in Mice
Published on: June 6, 2025
HIF-1α dependent RhoA as a novel therapeutic target to regulate rheumatoid arthritis fibroblast-like synoviocytes
Jianhai Chen1,2,3, Jingqin Chen3, Jianwei Tan3
1Rehabilitation Department of The Second Affiliated Hospital, School of Medicine, The Chinese University of Hong Kong, Shenzhen & Longgang District People's Hospital of Shenzhen, Shenzhen, Guangdong, 518172, China.
Objective:
The purpose of this work is to investigate how the Rho family of GTPases A (RhoA) mediates the pathogenesis of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS).
Methods:
The expression of RhoA in the synovial tissues of RA and Healthy people (Control) was detected using immunohistochemistry methods. The expression of RhoA and hypoxia-inducible factor-1α (HIF-1α) is inhibited by small interfering RNAs (siRNAs). The inhibition effect on RA-FLS migration was further investigated. The protein expression level of HIF-1α, RhoA, focal adhesion kinase (FAK), and myosin light chain (MLC) was also analysed using western blotting (WB). DBA1 mice were immunised with the mixture of bovine type II collagen and Freund's adjuvant to establish collagen induced arthritis (CIA) mouse model. Lip-siRhoA is administered through joint injection every two days. Micro-computed tomography (micro-CT) was used to detect mouse ankle joint destruction and evaluate the bone loss of the periarticular side. Destruction of the ankle articular cartilage was tested by histology. Expressions of P-RhoA, P-FAK and P-MLC in the ankle joint was detected by immunohistochemistry assay.
Results:
The expression level of RhoA in the synovial tissues of RA patients was significantly higher than that in control group. Hypoxia was able to up-regulate the expression of RhoA. Whereas, HIF-1α siRNA (siHIF-1α) could down-regulate the expression of RhoA. Additionally, both of siHIF-1α and RhoA siRNA (siRhoA) delivered by liposome (Lip-siHIF-1α and Lip-siRhoA) were found to suppress FAK and MLC phosphorylation in vitro. In CIA mouse model, Lip-siRhoA was demonstrated to ameliorate the destruction of ankle joint and reduce the severity of ankle joint cartilage damage by micro-CT and histological staining, respectively. Therefore, inhibition of FLS cell migration can protect articular bone from destruction. Furthermore, the expression of P-RhoA, P-FAK and P-MLC was evaluated and found to be down-regulated by Lip-siRhoA in vivo.
Conclusion:
The results demonstrated that under hypoxic environment, HIF-1α dependent RhoA pathway played an important role on cytoskeleton remodelling and RA-FLS migration. Through down-regulating RhoA expression, it could effectively treat RA in vitro and in vivo.
The Translational Potential Of This Article:
Our study provides new evidence for the potential clinical application of RhoA as a candidate for the treatment of RA.
Insights
This study reveals that RhoA, regulated by HIF-1α under hypoxia, drives rheumatoid arthritis fibroblast-like synoviocyte migration. Inhibiting RhoA effectively treats rheumatoid arthritis in vitro and in vivo.
Area of Science:
- Rheumatology
- Molecular Biology
- Cell Biology
Background:
- Rheumatoid arthritis (RA) pathogenesis involves fibroblast-like synoviocytes (RA-FLS).
- The Rho family of GTPases, specifically RhoA, is implicated in cellular processes relevant to RA.
- Understanding the role of RhoA in RA-FLS is crucial for developing targeted therapies.
Purpose of the Study:
- To investigate the role of RhoA in the pathogenesis of RA-FLS.
- To elucidate the mechanism by which RhoA mediates RA-FLS activity, particularly in relation to hypoxia and cytoskeleton remodeling.
- To evaluate the therapeutic potential of inhibiting RhoA in RA models.
Main Methods:
- Immunohistochemistry to detect RhoA expression in synovial tissues from RA patients and controls.
- siRNA-mediated inhibition of RhoA and hypoxia-inducible factor-1α (HIF-1α) to assess their effects on RA-FLS migration and protein expression (HIF-1α, RhoA, FAK, MLC).
- Collagen-induced arthritis (CIA) mouse model treated with liposomal RhoA siRNA (Lip-siRhoA) to evaluate therapeutic efficacy using micro-CT, histology, and immunohistochemistry.
Main Results:
- RhoA expression was significantly elevated in RA synovial tissues compared to controls and was upregulated by hypoxia.
- HIF-1α inhibition reduced RhoA expression, while both HIF-1α and RhoA siRNA suppressed RA-FLS migration, FAK, and MLC phosphorylation in vitro.
- In CIA mice, Lip-siRhoA treatment ameliorated joint destruction and cartilage damage, reducing phosphorylated RhoA, FAK, and MLC levels in vivo.
Conclusions:
- The HIF-1α-dependent RhoA pathway is critical for cytoskeleton remodeling and RA-FLS migration in hypoxic environments.
- Down-regulating RhoA expression offers a promising therapeutic strategy for treating RA, both in vitro and in vivo.
- This study highlights RhoA as a potential therapeutic target for RA treatment.
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