Mitochondrial Transplantation Therapy against Ifosfamide Induced Toxicity on Rat Renal Proximal Tubular Cells

Abstract

Mitochondrial dysfunction is a basic mechanism leading to drug nephrotoxicity. Replacement of defective mitochondria with freshly isolated mitochondria is potentially a comprehensive tool to inhibit cytotoxicity induced by ifosfamide on renal proximal tubular cells (RPTCs). We hypothesize that the direct exposure of freshly isolated mitochondria into RPTCs affected by ifosfamide might restore mitochondrial function and reduce cytotoxicity. So, the aim of this study was to assess the protective effect of freshly isolated mitochondrial transplantation against ifosfamide-induced cytotoxicity in RPTCs. Therefore, the suspension of rat RPTCs (106 cells/ml) in Earle’s solution with the pH of 7.4 at 37°C was incubated for 2 h after ifosfamide (4 mM) addition. Fresh mitochondria were isolated from the rat kidney and diluted to the needed concentrations at 4°C. The media containing suspended RPTCs was replaced with mitochondrial-supplemented media, which was exposed to cells for 4 hours in flasks-rotating in a water bath at 37°C. Statistical analysis demonstrated that mitochondrial administration reduced cytotoxicity, lipid peroxidation (LPO), reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP) collapse, lysosomal membrane damage, extracellular oxidized glutathione (GSSG) level, and caspase-3 activity induced by ifosfamide in rat RPTCs. Moreover, mitochondrial transplantation increased the intracellular reduced glutathione (GSH) level in RPTCs affected by ifosfamide. According to the current study, mitochondrial transplantation is a promising therapeutic method in xenobiotic-caused nephrotoxicity pending successful complementary in vivo and clinical studies.

Role of Mitochondria and Mitochondrial Transplantation in Drug-induced Toxic Organ Injury

“…78 Ifosfamide, a chemotherapeutic, was reported as a cause of tubular dysfunction. 79 Activation of complex I of rat renal cortex mitochondria was found to decrease in a study after ifosfamide treatment. 80 Antibiotic gentamicin can directly interact with complex II, and it can inhibit mitochondrial protein synthesis.…”

Renal-Protective Roles of Lipoic Acid in Kidney Disease

“…The authors found that ALA is equivalent to the Huangkui capsule in renoprotection against diabetic kidney injury, and both agents improve kidney function by attenuating oxidative stress and downregulating the activation of the p38MAPK and Akt pathways. ALA has also been compared with N -acetylcysteine (NAC) in one study, whereby the authors found that NAC is better than ALA in protecting oxidative kidney injury induced by the chemotherapeutic drug ifosfamide, which is highly toxic to the kidney [ 173 , 174 , 175 , 176 ]. However, the authors used an NAC concentration (200 mg/kg) that was twice that of ALA (100 mg/kg) [ 176 ].…”

Transplantation of Healthy Mitochondria into Rat Renal Proximal Tubular Cells Reduces Colistin- Induced Mitochondrial Dysfunction and Cellular Damage: An In Vitro Study

“…Cytotoxicity in the RPTCs was measured by lactate dehydrogenase (LDH) release. LDH activity was measured by using the LDH kit (Sigma-Aldrich, USA) (21). Brie y, after 4 hours of incubation, the RPTCs were washed with PBS and 10µl of the sample with 1ml of the indicator was mixed at 37°C for 4min.…”

“…Caspase 3 activation in RPTCs was measured by a caspase3 assay kit (Sigma-Aldrich, USA). The base for this assessment is hydrolysis peptide substrates (ACDEVEpNA) by caspase 3, releasing the pNA chromophore which has a λmax of 405 nm (21).…”

“…MMP collapse in RPTCs was measured by detection of the uorescence intensity of rhodamine 123 using a spectro uorometer (21). Brie y, after 4 hours of incubation, the RPTCs were washed with PBS and incubated with rhodamine 123 (5 µM) at 37°C for 20 min.…”