Exploring the cytoprotective potential of flavonoid fraction from Carica papaya L. Cultivar ‘Red Lady' Leaf: Insights into Nrf2 activation via in-vitro and in-silico approaches

Flavonoids play a significant role in mitigating the production of reactive oxygen species (ROS) and subsequent oxidative stress. In the present study we aimed to evaluate the cytoprotective and antioxidant defense mechanism of flavonoid fraction from Carica papaya L. (FFCP) leaf in human PBMC. For this investigation, the human PBMCs were lotted into three groups viz., negative control, positive control (treated with H2O2), and experiment group (pretreated with FFCP and challenged with H2O2). The cytoprotective mechanism was evaluated by assessing the intracellular ROS and Ca2+ mobilization, membrane and DNA damages, antioxidant enzymes, and Nrf2 activation. In-silico analysis was employed for assessing the interactions of flavonoids in FFCP with the Nrf2 binding pocket of Keap 1 protein. The results of the experiment unveiled that FFCP protected the human PBMCs from H2O2 induced oxidative damages such as lipid peroxidation, LDH leakage, DNA breakage, reduced the intracellular ROS and Ca2+ mobilization as compared to positive control. FFCP pretreatment effectively improved the activity of key antioxidant enzyme markers, such as SOD (1.66 ± 0.032-fold increase), CAT (2.24 ± 0.09-fold increase), GPx (1.94 ± 0.05-fold increase), and GR (1.81 ± 0.14-fold increase). Furthermore, the FFCP modulated the Nrf2 gene expression (a 2.38 ± 0.25 -fold) and its translocation into the nucleus as compared to the positive control. In-silico analysis enunciated that all flavonoids exhibited strong affinity with the Nrf2 binding pocket of Keap1 protein via stable hydrogen bonding and hydrophobic interactions. These observations suggest that FFCP is a natural source of bioactive compounds with nutraceutical benefits for combating oxidative stress-associated diseases by protecting the cells from oxidative stress induced damages as well as by instigating antioxidant defense mechanisms. [Display omitted] •Flavonoid fraction of Carica papaya L. (FFCP) protected the human PBMCs from oxidative damages- Lipid peroxidation, LDH leakage and DNA damage.•FFCP pretreatment effectively reduced the intracellular calcium mobilization and ROS production induced by oxidative stress.•FFCP pretreatment enhanced the activity of key antioxidant enzymes under oxidative stress.•FFCP upregulated the expression and translocation of Nrf2 transcription factor into nucleus.•Flavonoids in FFCP exhibited good binding affinity with the Nrf2 binding pocket of Keap 1•All flavonoids exhibited good ADME propert