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Mitochondrial disease can be defined as a genetic disease that manifests due to abnormalities in mitochondrial respiratory chain (MRC) caused by pathogenic variants in genes located on mtDNA or nDNA. It has been known that measurement of MRC enzyme activity in cultured fibroblasts from skin biopsies and in affected organ tissues is useful for the diagnosis of mitochondrial diseases. There have been reports of the usefulness of measuring MRC activity in renal tissue in kidney as well. On the other hand, the use of renal tissue is risky as a kidney biopsy must be performed. In this study, we investigated whether measurement of MRC enzyme activity in cultured tubular cells from urinary droplet would be useful for the diagnosis of mitochondrial nephropathy.
Five patients with mitochondrial nephropathy with the 3243A>G mutation and 21 patients with other renal diseases were enrolled. Urinary drop cells (P0) were cultured and P3 cells were used for measurements. Among 21 patients with other renal diseases, fourteen patients with cultured cells over 0.4x104 cells were selected as controls. For measurement of the activities of the MRC complexes I, II, III, and IV, the absorbance of the increasing and decreasing substances before and after the redox reaction of each complex was measured spectrophotometrically as followed previously reported methods. Enzyme activities of each complex were presented as the percentage of normal control mean relative to citrate synthase (CS) activity.
PCR using RNA extracted from the cultured cells confirmed that these cells expressed CD10. The activity of each MRC complex in the CS ratio of the 14 controls was set to 100%. However, the SD of complex I, II, III and IV was highly variable even within normal except for complex I, which was 13.9%, 25.5%, 23.5% and 39.4%, respectively. The mean (SD) ratios of complex I, II, III and IV to normal controls in mitochondrial disease patients were 57.4 (19.4) %, 66.7 (37.2) %, 65.7 (26.5) % and 75.0 (31.5) % respectively. The p-values between controls and patients were 0.01, 0.16,0.06, and 0.23, respectively. If less than 2 SD of normal control is defined as a reduction, complex I activity is reduced in patients with mitochondrial nephropathy. In addition, complex I activity of four among five patients were less than that of mean minus 2SD (72.2%). In one of the five cases, this test was performed before the genetic diagnosis was confirmed.
In patients with mitochondrial nephropathy with m.3243A>G, measurement of MRC enzyme activity in cultured cells from urinary drop cells may be useful for diagnosis. Whether it is also useful in mitochondrial nephropathy with other mutations is a subject for future investigation.