Aging is a major risk factor for drug-induced kidney injury, yet the specific impact of age on the susceptibility of human primary kidney stem cells to mitochondrial toxicity (MtT) remains largely uncharacterized.
To address this gap, this study in Genes & Diseases by researchers from Capital Medical University and Wake Forest University Health Sciences employed an in vitro 3D silk fiber matrix model to culture human primary urine-derived stem cells (USCs) obtained from both young (yUSCs) and older (oUSCs) donors. The authors used tenofovir (TFV), a widely prescribed antiretroviral medication associated with renal adverse effects, to evaluate age-related, drug-induced mitochondrial toxicity in comparison with other antiretroviral agents such as emtricitabine (FTC) and raltegravir (RAL).
Compared with yUSCs, oUSCs exhibited greater baseline mitochondrial vulnerability, including lower mitochondrial mass and mitochondrial DNA (mtDNA) content, increased oxidative stress, enhanced apoptotic signaling, greater cellular senescence, and reduced telomerase activity. The authors observed that these age-related differences became more pronounced upon antiretroviral exposure. TFV reduced mitochondrial mass and mtDNA content even at clinically relevant concentrations during prolonged 3D culture, highlighting the sensitivity of primary USCs to chronic mitochondrial injury.
TFV also impaired mitochondrial respiratory function, causing dose-dependent reductions in electron transport chain (ETC) complexes I–V in oUSCs, with mitochondrial damage being more pronounced in older cells than in younger cells. Furthermore, increased oxidative stress was evident from elevated SOD2 expression, while mitochondrial dysfunction was accompanied by reduced ATP production and compromised renal cell function. The greater TFV-associated impairment of mitochondrial parameters in oUSCs is consistent with the authors’ conclusion that aging increases susceptibility to drug-induced MtT.
Importantly, TFV promoted apoptosis in renal stem cells. The higher baseline cleaved caspase-3/caspase-3 ratio in oUSCs than in yUSCs indicated greater age-associated apoptotic susceptibility, while exposure to TFV further increased this ratio in a dose-dependent manner. Thus, chronic TFV-induced mitochondrial injury was accompanied not only by functional mitochondrial impairment but also by increased apoptotic stress, particularly in older cells.
TFV additionally accelerated cellular senescence. Older USCs exhibited higher β-galactosidase activity and increased expression of the senescence-associated genes p53, p21, and Rb, while TFV further increased these markers and reduced telomerase activity, with stronger effects in oUSCs. Renal injury was also reflected by increased KIM-1 expression, while alterations in CYP2E1 reflected changes in renal drug-metabolic function. Other tested antiretroviral agents, FTC and RAL, also induced oxidative stress, ETC dysfunction, apoptosis, and renal cellular injury, although they did not reproduce the broader effects of TFV on mitochondrial mass, mtDNA, senescence, and stem-cell properties.
Mechanistically, the findings implicated disruption of the telomere–mitochondrial axis involving PGC-1α. Older USCs exhibited reduced TRF1, telomerase activity, and PGC-1α expression, while TFV exposure was associated with decreased mtTFA and Nrf2 expression and reduced Nrf1 expression in oUSCs relative to baseline. The authors propose that telomere dysfunction and increased p53 signaling suppress PGC-1α, thereby impairing mitochondrial homeostasis and contributing to oxidative stress, senescence, apoptosis, and renal progenitor-cell dysfunction.
Importantly, the 3D USC–SFM system detected chronic mitochondrial toxicity more sensitively than conventional HEK293 and HepG2 cell models and maintained primary USCs over prolonged exposure, supporting its potential as an age-specific preclinical platform for evaluating renal drug toxicity.
In conclusion, the study demonstrates that aging markedly increases renal stem-cell susceptibility to TFV-induced telomere–mitochondrial dysfunction, oxidative stress, senescence, apoptosis, and functional injury, while highlighting the 3D silk matrix model as a promising platform for evaluating age-specific nephrotoxicity.
