Dual drug nanoparticle synergistically induced apoptosis, suppressed inflammation, and protected autophagic response in rheumatoid arthritis fibroblast-like synoviocytes.

Rheumatoid arthritis (RA) is a chronic immune-mediated joint inflammatory disorder associated with aberrant activation of fibroblast-like synoviocytes (FLS). Recently, FLS gained importance due to its crucial role in RA pathogenesis, and thus, targeting FLS is suggested as an attractive treatment strategy for RA. FLS-targeted approaches may be combined with disease-modifying antirheumatic drugs (DMARDs) and natural phytochemicals to improve efficacy in RA control and negate immunosuppression. In this study, we assessed the therapeutic effectiveness of DD NP HG in primary RA-FLS cells isolated from the synovial tissue of FCA-induced RA rats. We observed that DD NP HG had good biosafety for healthy FLS cells and, at higher concentrations, a mild inhibitory effect on RA-FLS. The combination therapy (DD NP HG) of MTX NP and PEITC NE in RA-FLS showed a higher rate of apoptosis with significantly reduced LPS-induced expression of pro-inflammatory cytokines (TNF-α, IL-17A, and IL-6) in arthritic FLS. Further, the gene expression studies showed that DD NP HG significantly down-regulated the mRNA expression of IL-1β, RANKL, NFATc1, DKK1, Bcl-xl, Mcl-1, Atg12, and ULK1, and up-regulated the mRNA expression of OPG, PUMA, NOXA and SQSTM1 in LPS-stimulated RA-FLS cells. Collectively, our results demonstrated that DD NP HG significantly inhibited the RA-FLS proliferation via inducing apoptosis, down-regulating pro-inflammatory cytokines, and further enhancing the expression of genes associated with bone destruction in RA pathogenesis. A nanotechnology approach is a promising strategy for the co-delivery of dual drugs to regulate the RA-FLS function and achieve synergistic treatment of RA.

Haloi P ，Choudhary R ，Lokesh BS ，Konkimalla VB ... -  《-》

Silencing aquaporin 1 inhibits autophagy to exert anti-rheumatoid arthritis effects in TNF-α-induced fibroblast-like synoviocytes and adjuvant-induced arthritis rats.

Fibroblast-like synoviocytes (FLS) are key players in rheumatoid arthritis (RA) by resisting apoptosis via increased autophagy. Elevated synovial aquaporin 1 (AQP1) affects RA FLS behaviors, but its relationship with FLS autophagy is unclear. We aim to clarify that silencing AQP1 inhibits autophagy to exert its anti-RA effects. We studied the effects and mechanisms of AQP1 silencing on autophagy in TNF-α-induced RA FLS and examined the crucial role of autophagy inhibition in its impacts on RA FLS pathogenic behaviors. We explored whether silencing synovial AQP1 relieved rat adjuvant-induced arthritis (AIA) by reducing synovial autophagy. TNF-α stimulation increased AQP1 expression and autophagy levels in RA FLS, with a positive correlation between them. AQP1 silencing inhibited autophagy in TNF-α-stimulated RA FLS, along with suppressing proliferation, promoting apoptosis, and mitigating inflammation. Notably, the inhibitory effects of AQP1 silencing on RA FLS pathogenic behaviors were cancelled by autophagy activation with rapamycin (Rapa) but enhanced by autophagy inhibition using 3-Methyladenine. Mechanistically, silencing AQP1 enhanced the binding of Bcl-2 to Beclin1 by decreasing Beclin1-K63 ubiquitination, thus inhibiting RA FLS autophagy. In vivo, silencing synovial AQP1 relieved the severity and development of rat AIA, alongside reducing Ki67 expression, promoting apoptosis, and decreasing autophagy within AIA rat synovium. Expectedly, the Rapa co-administration nullified the anti-AIA effects of silencing synovial AQP1. These findings reveal that silencing AQP1 inhibits RA FLS pathogenic behaviors and attenuates rat AIA through autophagy inhibition. This study may help clarify the pathogenic role of AQP1 in enhancing autophagy during RA development.

Zhang MY ，Wang MQ ，Huang Y ，Gu SL ，Zhou MY ，Xu ZS ，Li LL ，Lv M ，Cai L ，Li R ... -  《-》

Aryl hydrocarbon receptor antagonism attenuates growth factor expression, proliferation, and migration in fibroblast-like synoviocytes from patients with rheumatoid arthritis.

Lahoti TS ，Hughes JM ，Kusnadi A ，John K ，Zhu B ，Murray IA ，Gowda K ，Peters JM ，Amin SG ，Perdew GH ... -  《-》

Ferroportin inhibits the proliferation and migration of fibroblast-like synoviocytes in rheumatoid arthritis via regulating ROS/PI3K/AKT signaling pathway.

The aberrant proliferation of fibroblast-like synoviocytes (FLS) significantly contributes to excessive synovial hyperplasia and joint deformity in rheumatoid arthritis (RA). It has been observed that the membrane iron transporter protein, ferroportin (FPN), is commonly downregulated in tumor cells, while its overexpression can inhibit tumor cell proliferation. However, limited studies have investigated the role of iron in the pathogenesis of RA. In this study, we examined the functional relevance of FPN in RA. The expression of FPN in RA tissue specimens and primary cells was assessed using western blotting and RT-PCR. An adjuvant-induced arthritis (AIA) rat model was established to further validate the expression level of FPN. Phenotypic analysis of FLS cell proliferation was performed via CCK-8, clonogenic formation, and cell scratch assays. The involvement of membrane iron transporter proteins was analyzed through RNAseq and reactive oxygen species (ROS) detection. The results demonstrated decreased expression of FPN in the synovial tissue of RA patients compared to the normal group. Overexpression of FPN can inhibit RA-FLS proliferation and migration by suppressing the PI3K/AKT pathway, and this effect is associated with the elevation of ROS levels. Our findings suggest that the downregulation of FPN may contribute to the pathogenesis of RA, indicating a potential role of iron dysregulation in this disease, and FPN regulates the proliferation and migration of FLS by promoting the levels of ROS in FLS as well as suppressing the PI3K/AKT signaling pathway. These results suggest that FPN could be a potential target for alleviating joint damage in RA.

Shao W ，Liu F ，Zhu L ，Qian W ，Meng Q ，Zhang A ，Jin S ，Lu J ，Yan SG ... -  《-》

Effect of Aconitum diphtheria on the Proliferation, Apoptosis, and Inflammatory Response of Rheumatoid Arthritis Fibroblast-Like Synoviocytes.

Rheumatoid arthritis (RA), a highly disabling autoimmune disease, is characterized by joint damage and synovial hyperplasia. This paper was designed to explore the effect and mechanisms of Aconitum diphtheria on the proliferation and apoptosis of RA fibroblast-like synoviocytes (RA-FLS). First, RA-FLS was treated with different doses of A. diphtheria extracts. Then, an inverted biological microscope was adopted to observe the RA-FLS morphology. Subsequently, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining, Cell Counting Kit-8 and western blot were utilized to analyze the apoptosis, proliferation, and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway-related proteins in RA-FLS, respectively. The experimental results disclosed that, after treatment of RA-FLS with A. diphtheria extracts, the cells became round and the intercellular space was increased. Besides, the RA-FLS proliferation was effectively inhibited by A. diphtheria extracts, while the apoptosis was promoted. Additionally, A. diphtheria extracts could effectively downregulate the expression levels of p-NF-κB (p50), NF-κB (p50), p-NF-κB (p65), and NF-κB (p65). Collectively, A. diphtheria can effectively inhibit RA-FLS proliferation and promote apoptosis in a dose-dependent manner. Similarly, A. diphtheria is able to downregulate p-NF-κB and NF-κB protein expression, but there is no dose-response relationship.

Meng Y ，Cai XL ，Cong S ，Sun J ，Hu YW ，Luo L ... -  《-》