Uridine Abrogates the Adverse Effects of Antiretroviral Pyrimidine Analogues on Adipose Cell Functions

Abstract

Objectives Side effects of antiretroviral treatment such as lipoatrophy have been mainly attributed to mitochondrial toxicity of nucleoside reverse transcriptase inhibitors (NRTIs). We assessed whether uridine can abrogate the adverse effects of NRTIs on adipocyte functions. Methods 3T3-F442A preadipocytes were exposed to stavudine (d4T; 10 μM), zidovudine (ZDV; 1 μM), zalcitabine (ddC; 0.2 μM) or didanosine (ddI; 10 μM) in the absence or presence of uridine 21 days prior to and 7 days after induction of differentiation. Then, lipid accumulation (oil red staining), apoptosis (flow cytometry, PARP-cleavage), mitochondrial mass (Mitotracker) and DNA (mtDNA), cytochrome c oxidase (COX) subunits and mitochondrial membrane potential (JC-1) were quantified. Results Whereas ddI had no effects, d4T, ZDV and ddC significantly decreased cellular lipid accumulation (by 32%, 46% and 24%, respectively), increased apoptosis and induced mitochondrial depolarization. d4T, ZDV and ddC decreased adipocyte mtDNA (by 64%, 53% and 46%, respectively) and reduced the mtDNA encoded COX II subunit. Uridine (200 μM) had no intrinsic effect, but prevented all adverse effects of d4T, ZDV and ddC on adipocyte morphology, lipid staining, apoptosis, mtDNA depletion (partial prevention with ZDV), mitochondrial mass and membrane potential. The effects of uridine were concentration-dependent. Uridine also fully reverted established d4T toxicities despite continued d4T exposure. Conclusions Uridine supplementation protects adipocytes from the adverse effects of d4T, ZDV and ddC on lipid accumulation, cell survival and mitochondrial functions, suggesting that the toxic effects could be linked to intracellular depletion of uridine or its metabolites. Uridine is an interesting candidate in the prevention of NRTI-induced lipoatrophy in vivo.

3D Spheroids of Human Primary Urine-Derived Stem Cells in the Assessment of Drug-Induced Mitochondrial Toxicity

“…Each drug with 3 doses was added into a culture medium and incubated, Mito-Tox was evaluated in 3D spheroids, as compared to 2D cultures a three-time points (3 day, 2, and 4 weeks) Three FDA approved anti-HIV drugs were tested compared to two controls, including (1) ddC (0.1, 2, 10 µM) [ 39 , 40 ] known for significantly inducing Mito-Tox, 0.1 µM ddC is a typical single dose maximal plasma concentration (Cmax) [ 40 ]; (2) TFV (3, 30, 300 µM) [ 41 , 42 , 43 ] known for inducing only minimal Mito-Tox, 2.12µM TFV is a typical single dose maximal plasma concentration [ 40 , 41 , 42 , 43 , 44 ] (3) RAL (2, 20, 200 µM) [ 45 , 46 , 47 ] that is controversial in its ability to cause Mito-Tox, 2.25 µM is a typical RAL maximal plasma concentration [ 48 ], (4) rotenone (10 µM) [ 49 ] as a positive control; and (5) 0.1% DMSO as a negative control, see Table 1 .…”