Abstract
Background: Pulmonary Fibrosis is a chronic lung disease characterized by scar formation and respiratory insufficiency. Metformin is an antidiabetic drug that has recently been shown to prevent lung fibrosis via NADPH oxidase-4 suppression. Flaxseed oil has been shown to be a potent antioxidant that attenuates oxidative damage by inhibition of lipid peroxidation and the proinflammatory cytokine production of IL-6 and TNF-alpha. Aim: To investigate whether metformin and/or flaxseed oil could protect against bleomycin-induced pulmonary fibrosis and if this protection is mediated by inhibiting NOX-4 signaling pathway and the synthesis and release of inflammatory and oxidative stress markers. Material and Methods: Fifty adult male rats were randomized into 5 groups/10 rats each. Pulmonary fibrosis was induced by Bleomycin, delivered intratracheally at a concentration of 1.5mg/kg body weight once in group PF, MET, Flax and combined MET& Flax. Both metformin and Flaxseed oil were administered orally for 21 day after induction of PF. Withdrawal of blood samples for chemical and spectral assay of antioxidant markers was done. Histopathology and assessment of inflammatory and fibrotic markers and PCR expression of NOX4 gene were done. Results: There was a significant reduction in inflammatory cell count, histopathological score of inflammation and fibrosis in the three treatment groups. Also, there was marked improvement in inflammatory (IL-6, TNF-a) and fibrotic markers (TGFB and Col-1) in metformin group rather than flaxseed and combined group. Nox4 gene expression was significantly reduced in metformin treated group rather than other 2 groups.
Key Points
Flaxseed oil has been shown to be a potent antioxidant, it attenuates oxidative damage effectively due to their protective antioxidant properties. This can protect against lung pulmonary fibrosis in bleomycin-induced lung fibrosis. Flaxseed oil also inhibits lipid peroxidation by inhibiting the proinflammatory cytokine production of IL-6 and TNF-α. This study compared the therapeutic effect of metformin, flaxseed oil or combined treatment, in resolution of bleomycin induced lung fibrosis model, and to investigate if this therapeutic effect is due to NOX4 suppression.
In the present study, it was demonstrated that metformin, flaxseed oil treatment significantly improved lung injury in BLM-treated rats. Indeed, both individually was equally beneficial in improving lung injury, including improvement of functional and structural changes and inhibition of fibrosis, inflammation, lipid accumulation in the lung. Dual therapy with metformin and flaxseed oil, in some, but not all aspects, produced additive and synergistic therapeutic effects in the treatment of lung fibrosis. Pulmonary fibrosis pathogenesis involves variety of factors such as inflammation, ROS, activation of profibrotic cytokines. Among a variety of profibrotic cytokines is trans forming growth factor (TGF)-β that play a crucial role in orchestrating fibrosis development during IPF pathogenesis through regulating myofibroblast differentiation and proliferation result in accumulation of excessive deposition of ECM proteins. Increased expression levels of NOX4 have been reported in IPF lung not only in fibroblast of actively fibrosing areas but also injured epithelial cells , NOX4 is essential for a variety of issues include myofibroblast differentiation through TGF-β, apoptosis resistance by accelerating cellular senescence, which is associated with prolonged ECM production.
This study found that level of TGF- β and collagen type I was elevated in the BLM induced fibrosis group compared to normal control, associated with increased expression of NOX4 and elevated MDA level in the BLM induced fibrosis group compared to normal group with associated fibrosis appeared in histological sections while in the treatment groups it was found that TGF-β and collagen type I level was reduced and NOX4 expression was decreased with subsequent decrease in MDA level and associated with reduced fibrosis in histological sections, with the metformin reducing TGF-β , collagen type I ,and NOX4 expression a little more than flaxseed oil although the difference was nonsignificant the combined treatment group show a little bit more reduction than each individual drug regarding TGF-β , collagen.
The anti-fibrotic mechanism of metformin and flaxseed oil that is assigned to inhibition of TGF-β mediated fibrosis and to it is effect in regulation of NOX4 expression and to reduction of NOX4 mediated ROS production with the anti-inflammatory effect that was obvious in the study may provide a treatment modality for lung fibrosis.