Marta Leon-Monzon

Mitochondrial Alterations with Mitochondrial DNA Depletion in the Nerves of AIDS Patients with Peripheral Neuropathy Induced by 2′3′-Dideoxycytidine (ddC)

The 2′3′-dideoxycytidine (ddC), a nonazylated dideoxynucleoside analog used for the treatment of AIDS, causes a dose-dependent, painful, sensorimotor axonal peripheral neuropathy in up to 30% of the patients. To investigate the cause of the neuropathy, we performed morphological and molecular studies on nerve biopsy specimens from well-selected patients with ddC-neuropathy and from control subjects with disease, including patients with AIDS-related neuropathy never treated with ddC. Because ddC, in vitro, inhibits the replication of mitochondrial DNA (mtDNA), we counted the number of normal and abnormal mitochondria in a 0.04 mm2 cross-sectional area of the nerves and quantified the copy numbers of mtDNA by competitive PCR in all specimens. A varying degree of axonal degeneration was present in all nerves. Abnormal mitochondria with enlarged size, excessive vacuolization, electron-dense concentric inclusions and degenerative myelin structures were prominent in the ddC-neuropathy and accounted for 55% ± 2.5% of all counted mitochondria in the axon and Schwann cells, compared with 9% ± 0.7% of the controls (p < 0.001). Significantly (p < 0.005) reduced copy numbers, with as high as 80% depletion, of the mtDNA was demonstrated in the nerves of the ddC-treated patients compared with the controls. We conclude that ddC induces a mitochondrial neuropathy with depletion of the nerves mtDNA. The findings are consistent with the ability of ddC to selectively inhibit the γ-DNA polymerase in neuronal cell lines. Toxicity to mitochondria of the peripheral nerve is a new cause of acquired neuropathy induced by exogenous toxins and may be the cause of neuropathy associated with the other neurotoxic antiretroviral drugs or toxic-metabolic conditions.

Detection of Poliovirus Antibodies and Poliovirus Genome in Patients with the Post-Polio Syndrome

To investigate the role of poliovirus (PV) infection in the development of the post-polio syndrome (PPS), we studied the serum, spinal fluid, peripheral blood lymphocytes, and muscle from 47 patients with PPS. We found high titers of IgM PV antibodies (up to 1:250) in the serum of 6 patients, compared to very low titers (less than 1:50) in normal subjects or disease controls. By polymerase chain reaction, using primers of the replicase PV gene, we amplified PV sequences in the peripheral blood lymphocytes in 7 of 37 patients and in the CSF in 4 of 40 patients, but in none of the controls. Sequencing of the amplified product confirmed that it belonged to PV type 1 with a 99.3% homology. We conclude that some patients with PPS have in the serum high titers of IgM anti-PV antibodies, implying an ongoing antibody response to a viral antigen. The presence of PV-RNA in the CSF or lymphocytes suggests possible persistence of mutated virus or defective PV particles. The significance of these findings in the pathogenesis of PPS remains to be determined.

Expression of Poliovirus Receptor in Human Spinal Cord and Muscle

Because a prerequisite for infection of a cell with the poliovirus is the presence of poliovirus receptor (PVR), we examined its tissue localization in the human muscle, spinal cord, and muscle cultures using a specific monoclonal antibody against PVR in immunocytochemical studies on serial sections. We found weak expression of PVR in the motor neurons but not the axons. In normal muscle, PVR was expressed at the end plate as confirmed by immunolocalization in serial sections with alpha-bungarotoxin. In neurogenic conditions and in myopathies, PVR was found in occasional denervated muscle fibers and in several regenerating ones. Human myotubes expressed PVR and were susceptible to the poliovirus infection. We conclude that PVR is present at the motor end-plate that can serve as one of the routes of entry of the virus to the motor neurons. The presence of PVR in the regenerating muscle fibers is in accord with clinical observations that muscle injuries can predispose patients to paralytic poliomyelitis.

Effect of L-Carnitine on AZT-Induced Mitochondrial Toxicity: Studies on Human Muscle Cultures

M itochondrial myopathy in zidovudine-treated patients was initially reported in 1990 by Dalakas.1 Zidovudine (AZT) is one of the nucleotide analogs, the antiretroviral drugs, provided to patients infected with human immunodeficiency virus (HIV) to control the amount of virus present in those individuals and avoid the full development of the disease (AIDS).2 The first treatment consisted of a high dose of AZT for prolonged periods; in some of the HIV-patients treated with this regimen, it was observed that AZT caused a myopathic disease that was called “AZT myopathy.”1,3,4,5 Muscle biopsy studies from those patients revealed that this myopathy was histo- logically characterized by an increased number of a unique type of muscle fiber described as ragged red fibers.1,6 When the muscle biopsies were studied by electron microscopy, a proliferation of mitochondria with enlarged size and abnormal cristae was observed.1,6,7 Molecular analysis has also determined a severe reduction of the mitochondrial DNA in the muscle of HIV patients treated with AZT.6 We and others have reported on the mechanism by which AZT damages the muscle.8–11 Our studies were performed in vitro, using human muscle cultures,8,12,13 and in vivo, using rats treated with daily intraperitoneal injections of AZT.8 In an effort toward preventing this disease, our laboratory has also found that L-carnitine (3-hydroxy-4-me- thyl-ammoniobutanoate) in our model systems prevents and improves the pathological alterations caused by AZT.12,13