Inhibition of sPLA2 enzyme activity by cell‐permeable antioxidant EUK‐8 and downregulation of p38, Akt, and p65 signals induced by sPLA2 in inflammatory mouse paw edema model. Issue 2 (31st December 2022) - Jayachandra, Krishnegowda
Abstract: The arachidonic acid (AA) metabolic pathway, plays a vital role in the production of eicosanoids by the action of pro‐inflammatory secretory phospholipase A2 (PLA2 ). Release of eicosanoids is known to be involved in many inflammatory diseases. Identification of the inhibitory molecules of this AA pathway enzyme along with the regulation of intracellular signaling cascades may be a finer choice to develop as a powerful anti‐inflammatory drug. In this regard, we have screened few cell‐permeable antioxidant molecules Tempo, Mito‐TEMPO, N, N'‐Bis(salicylideneamino)ethane‐manganese(II) (EUK)‐134, and EUK‐8 against pro‐inflammatory sPLA2 s. Among these, we found EUK‐8 is a potent inhibitor with its IC50 value ranges 0.7–2.0 µM for sPLA2 s isolated from different sources. Furthermore, docking studies confirm the strong binding of EUK‐8 towards sPLA2 . In vivo effect of EUK‐8 was studied in HSF‐sPLA2 ‐induced edema in mouse paw model. In addition to neutralizing the edema, EUK‐8 significantly reduces the phosphorylation level of inflammatory proteins such as p38 member of MAPK pathway, Akt, and p65 along with the suppression of pro‐inflammatory cytokine (interleukin‐6) and chemokine (CXCL1) in edematous tissue. This shows that EUK‐8 not only inhibits the sPLA2 activity, it also plays an important role in the regulation of sPLA2 ‐induced cell signaling cascades. Apart from the sPLA2 inhibition, we also examine the regulatory actions of EUK‐8 with other downstream enzymes of AA pathway such as 5‐LOX assay in human polymorphonuclear leukocytes (PMNs) and COX‐2 expression in carrageenan‐λ induced paw edema. Here EUK‐8 significantly inhibits 5‐LOX enzyme activity and downregulates COX‐2 expression. These data indicate that EUK‐8 found to be a promising multitargeted inhibitory molecule toward inflammatory pathway. In conclusion, mitochondrial targeted antioxidant EUK‐8 is not only the powerful antioxidant, also a potent anti‐inflammatory molecule and may be a choice of molecule for pharmacological applications.