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In vitro study of putative genomic biomarkers of nephrotoxicity through differential gene expression using gentamicin
Sarah Cristina Teixeira Silva1, Leonardo Augusto de Almeida2, Stellamaris Soares1
1a Departamento de Análises Clínicas e Toxicológicas, Faculdade de Farmácia, Laboratório de Toxicologia Experimental in vitro , Universidade Federal de Minas Gerais (UFMG) , Belo Horizonte , Brazil.
Abstract:
Drug-induced nephrotoxicity is one of the most frequently observed effects in long-term pharmacotherapy. The effects of nephrotoxicity are commonly discovered later due to a lack of sensitivity in in vivo methods. Therefore, researchers have tried to develop in vitro alternative methods for early identification of toxicity. In this study, LLC-PK1 cells were exposed to gentamicin through MTT and trypan blue assay. Concentrations of 4 (low), 8 (medium) and 12 (high) mM, were used to evaluate differential gene expression. A panel of genes was selected based on gene expression changes. The search for sequences of mRNA encoding proteins previously associated with kidney damage was conducted in the databases of the National Center for Biotechnology Information (USA). RNA was extracted from the cells, and RT-qPCR was performed to evaluate differential expression profiles of the selected genes. Among the 11 analyzed genes, four proved to be differentially up-regulated in cells exposed to gentamicin: HAVcr1, caspase 3, ICAM-1 and EXOC6. According to this study's results, we suggest that these genes play an important role in the mechanism of in vitro nephrotoxicity caused by gentamicin and can be used as early in vitro biomarkers to identify nephrotoxicity when developing safer drugs.
Insights
This study identifies four key genes (HAVcr1, caspase 3, ICAM-1, EXOC6) as early biomarkers for drug-induced nephrotoxicity. These findings aid in developing safer drugs by enabling early in vitro toxicity detection.
Area of Science:
- Biomedical Science
- Toxicology
- Molecular Biology
Background:
- Drug-induced nephrotoxicity is a common complication of long-term pharmacotherapy.
- Current in vivo methods for detecting nephrotoxicity lack sensitivity, leading to late diagnosis.
- There is a critical need for reliable in vitro methods for early toxicity identification.
Purpose of the Study:
- To investigate the potential of specific genes as early biomarkers for gentamicin-induced nephrotoxicity.
- To establish an in vitro model for assessing drug-induced kidney damage.
- To identify novel molecular targets for predicting nephrotoxicity during drug development.
Main Methods:
- LLC-PK1 cells were exposed to varying concentrations of gentamicin.
- Cell viability was assessed using MTT and trypan blue assays.
- Differential gene expression was analyzed using RT-qPCR on a selected panel of 11 genes.
Main Results:
- Four genes, HAVcr1, caspase 3, ICAM-1, and EXOC6, were significantly up-regulated in gentamicin-exposed cells.
- These genes showed differential expression patterns indicative of cellular stress and damage.
- The identified genes are associated with kidney damage mechanisms.
Conclusions:
- HAVcr1, caspase 3, ICAM-1, and EXOC6 can serve as early in vitro biomarkers for gentamicin-induced nephrotoxicity.
- These biomarkers facilitate the early detection of kidney toxicity during drug development.
- This study contributes to the development of safer pharmaceutical agents through improved toxicity screening.