Hyperhalophilic Diatom Extract Protects against Lead-Induced Oxidative Stress in Rats and Human HepG2 and HEK293 Cells

Abstract

This work investigated the protective effects of microalga Halamphora sp. extract (HExt), a nutraceutical and pharmacological natural product, on human lead-intoxicated liver and kidney cells in vitro and in vivo in Wistar rats. The human hepatocellular carcinoma cell line HepG2 and the human embryonic kidney cell line HEK293 were used for the in vitro study. The analysis of the fatty acid methyl esters in the extract was performed via GC/MS. The cells were pretreated with HExt at 100 µg mL^-1, followed by treatment with different concentrations of lead acetate, ranging from 25 to 200 µM for 24 h. The cultures were incubated (5% CO, 37 °C) for 24 h. Four groups, each containing six rats, were used for the in vivo experiment. The rats were exposed to subchronic treatment with a low dose of lead acetate (5 mg kg^-1 b.w. per day). Pretreating HepG2 and HEK293 cells with the extract (100 µg mL^-1) significantly (p < 0.05) protected against the cytotoxicity induced by lead exposure. For the in vivo experiment, the biochemical parameters in serum-namely, the level of malondialdehyde (MDA), and the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx)-were measured in the organ homogenate supernatants. HExt was found to be rich in fatty acids, mainly palmitic and palmitoleic acids (29.464% and 42.066%, respectively). In both the in vitro and in vivo experiments, cotreatment with HExt protected the liver and kidney cell structures and significantly preserved the normal antioxidant and biochemical parameters in rats. This study discovered the possible protective effect of HExt, which could be beneficial for Pb-intoxicated cells.

Keywords: fatty acid; human cell line; lead exposure; microalgal extract; oxidative stress; rats.

Figure 1

Effect of Halamphora extract on the viability (%) of HepG2 and HEK293 cells. Data are expressed as the mean ± SD. R²: coefficient of determination. Dashed line corresponds to the curve of regression.

Figure 2

Cytotoxicity assay using HepG2 and HEK293 cells. (A) Cytotoxicity of different concentrations of lead acetate. Data are expressed as the mean ± standard deviation of triplicate measurements. (B) Morphologic changes in HepG2 cells after treatment with different concentrations of lead acetate. (C) Morphological changes in HEK293 cells after treatment with different concentrations of lead acetate.

Figure 3

Protective effect of Halamphora extract (HExt) against lead-acetate-induced toxicity to HepG2 and HEK293 cells. (A) Viability of cells treated with different concentrations of lead acetate, 50 µM, 100 µM, 150 µM, and 200 µM (C1, C2, C3, and C4, respectively), and HExt (100 µg mL⁻¹). (B) Morphological changes in HepG2 cells pretreated with HExt before the administration of different concentrations of lead. (C) Morphological changes in HEK293 cells pretreated with HExt before the administration of different concentrations of lead. All data are expressed as the mean ± SD. Different letters represent significant differences (p < 0.05) between each treatment and the control (cells treated with lead acetate).

Figure 4

Histopathology of the rat liver (H & E × 400). Photomicrographs of the liver sections from (A) Group I, (B) Group II, (C1,C2) Group III, and (D) Group IV. Necrotic focus (circles), cytolysis (arrows), inflammatory cells (hyphen), and the congested central veins (large arrows). Optic microscopy: H & E (×400). H & E: hematoxylin and eosin.

Figure 5

Histopathology of the rat kidney (H & E × 400). Photomicrographs of the kidney sections from (A) Group I, (B) Group II, (C1,C2) Group III, and (D) Group IV. Intraluminal cellular desquamation (arrows), a fibronecrotic small focus that has Bowman’s space (hashtag), and the vacuolar degeneration lesion of tubular cells (apoptosis) (hyphen) and tubular necrosis (stars). Optic microscopy: H & E (×400). H & E: hematoxylin and eosin.