Jun 2023
S. Prakasha Gowda A.Corresponding author
Insulin is a frequent peptide hormone addition in serum-free mammalian cell culture media. It contributes in a variety of biological functions, including as promoting cell proliferation, cell cycle progression, and glucose uptake. However, it is unknown how stable insulin is under in vitro cell culture media treatment conditions. The instability of insulin in aqueous solutions has caused a number of issues, necessitating the development of new therapeutic strategies that can keep insulin stable and functioning. Such choices are required to accommodate updated insulin delivery guidelines as well as the storage and transportation of insulin. To preserve structural and functional integrity, protein medicines are frequently stabilized with antioxidants in aqueous solutions. In the present study, the effects of the antioxidants disodium ethylenediaminetetraacetic acid dihydrate (EDTA) and sodium selenite (Se) and their ability to scavenge free radicals on insulin stability in the medium Dulbecco's Modified Eagle Medium (DMEM) and Roswell Park Memorial Institute (RPMI) were examined. To investigate the stability of human recombinant insulin, in vitro serum-free DMEM and RPMI media were utilized for 5 days at 37˚C containing different EDTA and Se concentrations. Reversed phase high performance liquid chromatography (RP-HPLC) was used to detect and quantify insulin. Sodium dodecyl sulfate polyacrylamide gel (SDS-PAGE) electrophoresis was used to assess conformational stability. The results demonstrated that, when EDTA and Se were added separately to DMEM and RPMI media, insulin stability was improved compared to when neither compound was added.
Apr 2020 DOI 10.14302/issn.2470-0436.jos-20-3303
Manikandan R.Corresponding author
Department of Zoology, University of Madras, Guindy Campus, Chennai-600025
The effect of resveratrol, a free radical scavenger, during cataract development was evaluated in the Wistar rat pup model. This study investigated the possible free radical scavenging potential of resveratrol at 40 mg/ kg body wt dose in selenite-induced cataract in rat pups. Intraperitoneal injection of sodium selenite (15 µm mol/ kg body wt) in 8 to 10 day old rat pups lead to severe oxidative stress in the tissues evidenced by decreased antioxidants and increased lipid peroxidase, nitric oxide, superoxide anion, hydroxyl radical generation, inducible nitric oxide synthase (iNOS) as well as nuclear factor kappa B (NF-kB) expression levels that probably led to cataract formation. Selenite exposure also caused an increase in total calcium in the eye lens and significantly inhibited the activity of Ca2+ ATPase but not Na+/ K+ ATPase or Mg2+ ATPase. However, both pre- and co-treatments with resveratrol, but not post-treatment, led to an increase in antioxidant levels with a concomitant reduction in oxidative stress and also rescued the selenite-mediated increase in lens Ca2+ and inhibition of Ca2+ ATPase activity in the eye lens. The results of this study demonstrate antioxidants decrease and increase in free radical generation triggered by selenite causes the inactivation of lens Ca2+ ATPase leading to a rise in intracellular Ca2+ level. Resveratol treatment was able to prevent selenite-induced oxidative stress and in turn the inhibition of lens opacification. Thus, resveratrol has the potential to function as an anti-cataractogenic agent, possibly by preventing free radical-mediated accumulation of Ca2+ in the eye lens.