ROS-induced oxidative damage in lymphocytes ex vivo/in vitro from healthy individuals and MGUS patients: protection by myricetin bulk and nanoforms

Shabana Akhtar,Diana Anderson,Lisa Newton,Mohammad Isreb,Mojgan Najafzadeh,Rajendran C Gopalan

doi:10.1007/s00204-020-02688-4

Abstract

We investigated the protective role of myricetin bulk and nanoforms, against reactive oxygen species (ROS)-induced oxidative stress caused by hydrogen peroxide and tertiary-butyl hydro peroxide in lymphocytes in vitro from healthy individuals and those from pre-cancerous patients suffering with monoclonal gammopathy of undetermined significance (MGUS). The change in intracellular reactive oxygen species was measured once cells were treated with myricetin bulk forms and nanoforms with and without either hydrogen peroxide or tertiary-butyl hydro peroxide co-supplementation. The direct and indirect antioxidant activity of myricetin was spectrofluometrically measured using the fluorescent dye 2′,7′-dichlorofluorescin diacetate and using the Comet assay, respectively. Hydrogen peroxide (50 µM) and tertiary-butyl hydro peroxide (300 µM) induced a higher level of reactive oxygen species-related DNA damage and strand breaks. Addition of myricetin nanoform (20 µM) and bulk (10 µM) form could, however, significantly prevent hydrogen peroxide- and tertiary-butyl hydro peroxide-induced oxidative imbalances and the nanoform was more effective. Glutathione levels were also quantified using a non-fluorescent dye. Results suggest that myricetin treatment had no significant effect on the cellular antioxidant enzyme, glutathione. The current study also investigates the effect of myricetin on the induction of double-strand breaks by staining the gamma-H2AX foci immunocytochemically. It was observed that myricetin does not induce double-strand breaks at basal levels rather demonstrated a protective effect.

Highlights

Oxidative stress is a major factor contributing towards the development of various illnesses and it is a key inducer of cellular lesions
Cytotoxicity of test chemicals was determined by culturing lymphocytes from healthy individuals and monoclonal gammopathy of undetermined significance (MGUS) patients and measuring the mean absorbance at 1 and 24 h using the MTT dye
Results from the MTT assay showed that the test chemicals used in this study exhibited non-significant level of cytotoxicity occurred only after 24 h treatment, for all the treatment groups assessed in lymphocytes from both healthy and patient group (Fig. 2a, b)

Summary

Introduction

Oxidative stress is a major factor contributing towards the development of various illnesses and it is a key inducer of cellular lesions. It is triggered by a disparity between the production of reactive oxygen species (ROS) and the capacity of biological system to cleanse the reactive intermediates. Intracellular ROS production has been associated with the mediation of the effects caused by anti-cancer drugs like taxol (Perkins et al 2000; Varbiro et al 2001). It has been suggested that cellular redox potential may be regulated through increasing glutathione (GSH) pools to the mitochondria, preventing ROS formation and cellular injury caused by oxidative stress (Slikker et al 2001)

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