Naringenin Against Cadmium Toxicity in Fibroblast Cells: An Integrated Network Pharmacology and In Vitro Metabolomics Approach.

Cadmium, a heavy metal, disrupts cellular homeostasis and is highly toxic, with no effective treatments currently available against its toxicity. According to studies, phytochemicals provide a promising strategy for mitigating cadmium toxicity. Naringenin (NG), a potent antioxidant found primarily in citrus fruits, showed protective properties against cadmium toxicity in rats. Nonetheless, the precise mechanism of cadmium cytotoxicity in fibroblasts remains unknown. This study evaluated NG against cadmium (CdCl2) toxicity utilizing network pharmacology and in silico molecular docking, and was further validated experimentally in rat fibroblast F111 cells. Using network pharmacology, 25 possible targets, including the top 10 targets of NG against cadmium, were identified. Molecular docking of interleukin 6 (IL6), the top potential target with NG, showed robust binding with an inhibition constant (Ki) of 58.76 µM, supporting its potential therapeutic potential. Pathway enrichment analysis suggested that "response to reactive oxygen species" and "negative regulation of small molecules metabolic process" were the topmost pathways targeted by NG against cadmium. In vitro analysis showed that NG (10 µM) attenuated CdCl2-induced oxidative stress by reducing altered intracellular ROS, mitochondrial mass, and membrane potential. Also, NG reversed CdCl2-mediated nuclear damage, G2/M phase arrest, and apoptosis. GC/MS-based metabolomics of F111 cells revealed CdCl2 reduced cholesterol levels, which led to alterations in primary bile acid, steroid and steroid hormone biosynthesis pathways, whereas, NG restored these alterations. In summary, combined in silico and in vitro analysis suggested that NG protected cells from CdCl2 toxicity by mitigating oxidative stress and metabolic pathway alterations, providing a comprehensive understanding of its protective mechanisms against cadmium-induced toxicity.

Phytochemical Profiling of Tupistra nutans Wall. ex Lindl. Inflorescence Extract and Evaluation of Its Antioxidant Activity and Toxicity in Hepatocarcinoma (HepG2) and Fibroblast (F111) Cells.

Tupistra nutans Wall. ex Lindl. is a medicinal plant found in the Eastern Himalayan region. Besides being used as a folk medicine for pain and high blood sugar, its inflorescence is consumed as a vegetable. However, its medicinal properties have not been proven in vitro and in vivo till now. Therefore, in this study, we reported the phytochemicals present in the methanolic extract of Tupistra nutans Wall. ex Lindl. inflorescence (METNI) and its comparative effect in liver carcinoma HepG2 cells against non-cancerous murine fibroblast F111 cells. Phytochemical profiling by gas chromatography-mass spectrometry (GC-MS) analysis showed that METNI was rich in unsaturated fatty acids, vitamin E, and anticancer compounds like diosgenin, linoleic acid, and palmitoleic acid. METNI was found to have in vitro antioxidant property as determined by DPPH and pyrogallol methods, and UV protection property as investigated by fluorescence-based and spectrophotometric methods. MTT assay revealed METNI caused significantly more cell proliferation inhibition in HepG2 (IC50 = 138 µg/ml) compared to F111 (IC50 = 347 µg/ml) cells. Although in both HepG2 and F111 cells METNI showed significant antioxidant activity, it led to intracellular ROS generation and cell cycle alteration at higher exposure. The obtained results suggest that Tupistra nutans can be a promising application for anticancer drug and skin care product development, but can be harmful if overconsumed.

Effect of differential deprivation of nutrients on cellular proliferation, oxidative stress, mitochondrial function, and cell migration in MDA-MB-231, HepG2, and HeLa cells.

Cancerous cells display metabolic engineering through enhanced utilization of nutrients to support their increased requirements for proliferation, bioenergetics, biosynthesis, redox homeostasis, and cell signaling. To investigate the extent to which malignant cells rely on glycolysis and glutaminolysis, the effects of differential deprivation of nutrients such as d-glucose, l-glutamine, and pyruvate on proliferation, morphology, cell cycle, oxidative stress, mitochondrial function, autophagic vacuole formation, and migration in MDA-MB-231, HepG2, and HeLa cells were investigated in this study. Cell viability assay,  cell morphology, and ATP assay showed higher dependence of MDA-MB-231 and HepG2 cells on glucose and glutamine, respectively, for cell survival, growth, ATP production, and proliferation, while HeLa cells were equally dependent on both. However, the combination of all three nutrients displayed maximum proliferation. Differential deprivation of glucose in the absence of glutamine resulted in G0/G1 plus G2/M arrest in MDA-MB-231, whereas G0/G1 arrest in HepG2 and S-phase arrest in HeLa cells occurred at 48 h. Although the differential withdrawal of nutrients revealed a varying degree of effect dependent on cell type, nutrient type, nutrient concentrations, and deprivation time, a general trend of increased oxidative stress, loss of mitochondrial membrane potential, and ATP and antioxidant (GSH) depletion led to mitochondrial dysfunction in all three cell lines and inhibition of cell migration in MDA-MB-231 and HeLa cells at 48 h. Extreme deprivation of nutrients formed autophagic vacuoles. Importantly, normal cells (HEK293) remained unaffected under most of the nutrient-deprived conditions examined. This study enhances our understanding of the impact of differential nutrient deprivation on critical characteristics of cancer cells, contributing to the development of metabolism-based effective anticancer strategies. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03759-w.