The kidney is especially vulnerable to toxic insult by various drugs and xenobiotics because it receives nearly one quarter of the cardiac output, and transports, metabolizes and concentrates a variety of potentially toxic substances within its parenchyma As a consequence of its primary functions, the kidney is especially vulnerable to toxic insults by various drugs or xenobiotics, and thus nephrotoxicity is one of the major concerns in safety evaluation. Thus, despite the morphological complexity of the kidney, the epithelial cells of proximal tubule stand out as one of the most sensitive components in the kidney and are thus highly susceptible to damage. Traditional markers of kidney toxicity, blood creatinine and urea, have some limitations. First, significant changes may not occur until frank renal damage has occurred (determined by histopathology assessment and/or functional tests) and second they are not region specific. As a consequence of the difficulties in assessing renal damage by currently available/traditional biomarkers, there is an ongoing interest in developing new, early biomarkers of nephrotoxicity. Integratation of emerging transcription profiling technologies into traditional drug safety assessment evaluations offers the possibility to take new steps toward understanding mechanism of target organ toxicity and elucidating putative new biomarkers of exposure and safety. The renal toxicity process is typically initiated by a toxic injury to tubular epithelial cells in various nephron segments and this initial injury is often followed by cellular proliferation and repair that attempts to restore normal renal function. Changes in the expression of mRNA specifically expressed in the injured kidney cells are some of the earliest events that accompany the renal injury. This is accompanied by changes in the expression of other genes that contribute either to cellular repair or recovery of renal function. In many of published literature employed microarray technology has been used to identify some patterns of gene expression that reflect different types of nephrotoxicity. Signature of genes designed to indicate nephrotoxicity obtained need to be validated with an independent gene expression detection technology such as quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). The objective of the present work was to evaluate potential molecular markers of proximal tubular damage in rat kidney caused by segment-specific toxic agents . A molecular biology approach has been applied to evaluate the expression of a selected panel of four genes that are involved in different mechanisms of toxicity and can reflect different types of nephrotoxicity. The selected genes, Kidney injury molecule-1 (Kim-1), Clusterin (Cln), Osteopontin (Spp1), and Regucalcin (Rgn) are involved in different pathways, such as tissue damage, remodeling and regeneration, disruption of calcium homeostasis, oxidative stress. Gene expression was quantified by Real-Time quantitative Polymerase Chain Reaction (qRT-PCR) in kidney samples from male rats treated with HCBD, K2Cr2O7, and Cephaloridine, three nephrotoxicants that primarily injure specific regions of the proximal tubule via different mechanisms of action. Conventional clinical chemistry and histopathological analysis were performed to confirm the induction of segment-specific proximal tubule damage caused by chemicals. Results showed a correlation of gene changes with microscopic modification induced by nephrotoxicants. Moreover, the severity in proximal tubular damage evidenced in the dose-response experiments with the three compounds, were correlated with the magnitude of the transcriptional response. On the contrary, traditional clinical markers changes were observed for severe damage only, confirming the low sensitivity. Interestingly, even low severity microscopic observations were evidenced by gene expression quantification. Moreover, since KIM-1 mRNA levels modifications were evidenced also for low dose of nephrotoxicants where no morphologic modifications were observed, it is likely to represent a potential predictive biomarker of nephrotoxicity. In conclusion, this study confirmed that gene expression quantification may be used to detect renal tubular damage induced by nephrotoxicants. Genomic responses could represent more sensitive tool to monitor renal damage in comparison with traditional morphological and biochemical end points.
|Titolo:||Drug-inducted nephrotoxicity: a molecular biology approach to identify early markers of segment-specific proximal tubule injury in rat|
|Data di pubblicazione:||2008|
|Appare nelle tipologie:||07.13 Doctoral Thesis|