Protective Role of Magnesium against Oxidative Stress on SO4= Uptake through Band 3 Protein in Human Erythrocytes

Background/Aims: Magnesium, whose supplementation provides beneficial effects against oxidative stress-related conditions, has been here used to possibly protect Band 3 protein anion exchange capability and underlying signaling in an in vitro model of oxidative stress. Methods: Whole blood samples pre-exposed to 10 mM MgCl 2 , were treated for 30 min with H 2 O 2 (300 M, 600 M and 1 mM) chosen as oxidant molecule. In a separate protocol, NEM (0.5,1 and 2 mM), a phosphatase inhibitor and thiol-alkilant agent, has been also applied. The rate constant for SO 4 = uptake, accounting for Band 3 protein anion exchange capability, has been measured by a turbidimetric method, while intracellular reduced glutathione (GSH) levels and membrane -SH groups mostly belonging to Band 3 protein were spectrophotometrically quantified after reaction with DTNB (5,5'-dithiobis-(2-nitrobenzoic acid). Expression levels of Band 3 protein, phosporylated Tyrosine (P-Tyr) and tyrosine kinase (Syk) involved in signaling have been also measured. Results: Our results show that Mg 2+ prevented the reduction in the rate constant for SO 4 = uptake on H 2 O 2 -treated erythrocytes, not involving GSH levels and membrane -SH groups, unlike NEM, remaining both P-Tyr and Syk expression levels high. Conclusion: Hence, i) the measurement of the rate constant for SO 4 = uptake is a useful tool to evaluate Mg 2+ protective effect; ii) the use of two different oxidant molecules shed light on Mg 2+ effect which seems not to modulate phosphorylative pathways but would putatively stabilize membrane organization; iii) the use of Mg 2+ in food supplementation can be reasonably supported to protect erythrocytes homeostasis in case of oxidative stress-related diseases.