Metformin and omega-3 fish oil elicit anti-inflammatory effects via modulation of some dysregulated micro RNAs expression and signaling pathways in experimental induced arthritis

•Combination of Metformin and omega-3 amended the arthrogram score, hind paw swelling, serum TNF-α and interleukin (IL)-1β levels in adjuvant induced arthritis model.•On the molecular level, it activated p-AMPK and protein phosphatase 2A (PP2A) as well as p-mTOR, inhibited p-STAT3, NF-κB p65 subunit, p38 MAPK and p-JNK.•Also, it ameliorated miRNA-155, 146a and 34 levels and the imbalance between RORγT and FOXP3, correcting Th17/Treg cells balance.•The aforementioned combination had a comparable anti-inflammatory to that of Methotrexate, with less side effects so there is a probability for its use as a replacement therapy to that of methotrexate. Rheumatoid arthritis is a progressive inflammatory disease with multiple dysfunctional intracellular signaling pathways that necessitate new approaches for its management. Hence, the study aimed to inspect the ability of the combination therapy of metformin and omega-3 to modulate different signaling pathways and micro RNAs such as (miR-155, miR-146a and miR-34) as new targets in order to mitigate adjuvant-induced arthritis and compare their effect to that of methotrexate. Fourteen days post adjuvant injection, Sprague-Dawley rats were treated orally with metformin (200 mg/kg/day) and/or omega-3 (300 mg/kg/day) or intraperitoneally with methotrexate (2 mg/kg/week) for 4 weeks. All drug treatments amended the arthrogram score and hind paw swelling as well as decreased serum tumor necrosis factor (TNF)-α and interleukin (IL)-1β levels. On the molecular level, all therapies activated phospho-5′adenosine monophosphate-activated protein kinase (p-AMPK) and protein phosphatase 2A (PP2A), while they inhibited phospho-mammalian target of rapamycin (p-mTOR), phospho-signal transducers and activators of transcription (p-STAT3), nuclear factor (NF)-κB p65 subunit, phosho38 mitogen-activated protein kinase (p38 MAPK) and phospho- c-Jun N-terminal kinase (p-JNK). In addition, they decreased the elevated expression level of miRNA-155, 146a and increased the expression level of miRNA-34 and they decreased the expression level of retinoic acid receptor related orphan receptor γT (RORγT) and increased that of fork head box P3 (FOXP3), correcting Th17/Treg cells balance. On most of the aforementioned parameters, the effect of the combination therapy was comparable to that of methotrexate, emphasizing that this combination possesses better additive anti-inflammatory effect than either drug when used alone. In addition, the combinatio