Harmine and Kaempferol treatment enhances NFATC1 and FOXP3 mediated regulatory T-cells' suppressive capacity in generalized vitiligo

Background: Generalized vitiligo (GV) is an autoimmune disease characterized by the progressive loss of melanocytes.Objectives: Current study was undertaken to assess in-vitro therapeutic potential of Harmine and Kaempferol for GV.Methods: Calcium, calcineurin, NFATC1 levels, cell proliferation were assessed by various kits and ORAI1, PEIZO1, Calcineurin, GSK3B, DYRK1A transcripts and IFN-gamma,IL-10,TGF-beta protein levels were assessed by qPCR and ELISA in blood and skin biopsy samples from Tregs of 52 patients and 50 controls.Results: Harmine and Kaempferol treatment enhances Treg suppressive capacity, NFATs and FOXP3 expression in blood and skin Tregs of GV patients (p < 0.05). Furthermore, Harmine and Kaempferol treatment in Tregs increased calcineurin and NFATC1 activity and decreased DYRK1A transcripts in blood and skin Tregs of GV patients(p < 0.05). In-silico analysis revealed that Harmine and Kaempferol might boost Treg suppressive ca-pacity by increasing calcineurin dephosphorylation activity leading to increase NFATs activation and also in-crease nuclear retention of NFATs by inhibiting DYRK1a phosphorylation activity. Moreover, calcineurin and NFATC1 activity in Tregs were positively correlated with Treg suppressive capacity, NFATC1 and FOXP3 expression (p < 0.05), whereas, DYRK1A transcripts were negatively correlated with Treg suppressive capacity, NFATC1 and FOXP3 expression (p < 0.05). These compounds significantly increased melanocytes' survival and proliferation in Treg:CD4(+)/CD8(+):SK-Mel-28 cell line co-culture system from GV patients (p < 0.0001).Conclusions: For the first time the study suggests that Harmine and Kaempferol treated Tregs could control the CD8+ and CD4(+)T-cells' proliferation and IFN-gamma production, leading to melanocytes' survival and proliferation. These compounds may serve as novel Treg-based therapeutics for GV; however, in vivo studies are warranted to assess the safety and efficacy of these compounds.