Decreased Intracellular ATP Content and Intact Mitochondrial Energy Generating Capacity in Human Cystinotic Fibroblasts

Levtchenko, Elena; Wilmer, Martijn J.; Janssen, A.J.M.; Koenderink, Jan B.; Visch, Henk-Jan; Willems, Peter H.G.M.; Graaf-Hess, Adriana de; Blom, Henk J.; Heuvel, Lambertus P. van den; Monnens, L.A.H.

doi:10.1203/01.pdr.0000196334.46940.54

Cited by 54 publications

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“…ATP generation in these cells appeared to be normal under basal and stimulated conditions. A moderate decrease in intracellular ATP content did not cause alterations of Na,K-ATPase activity (13). Furthermore, we found normal intracellular ATP levels in human immortalized PTEC, derived from urine of cystinotic patients compared with healthy controls (14).…”

mentioning

confidence: 50%

“…Fibroblasts were cultured from skin biopsies after obtaining informed consent of healthy controls (ct1 and ct2) and cystinotic patients (pt1 and pt2), as described previously (13). Cystinosis was diagnosed in all patients by measuring elevated cystine content in polymorphonuclear cells (Ͼ0.5 nmol cystine/mg protein) and was confirmed by molecular analysis of the CTNS gene.…”

Section: Cell Culturesmentioning

confidence: 99%

“…After culturing fibroblasts to confluence, cells were incubated with CDME as indicated above (1.0 mM CDME for 30 min) and harvested using trypsin-EDTA to determine intracellular ATP levels in three independent experiments using an ATP luminescence kit (Roche) as described previously. Data are presented as nmol ATP/mg protein (13) and expressed as mean Ϯ SEM.…”

Section: Cell Culturesmentioning

confidence: 99%

“…Mitochondrial ATP production was monitored by transfecting fibroblasts cultured on a coverslip with a baculovirus containing cDNA for mitochondria-targeted luciferase (13,18). Perfusing the transfected cells with N-2-hydroxyethylpiperazine-N 1 -2-ethanesulfonic acid (HEPES)-Tris medium (132 mM NaCl, 4.2 mM KCl, 1 mM MgCl 2 , 5.5 mM D-glucose, 10 mM HEPES, 1 mM CaCl 2 , pH 7.4) supplemented with 5 M beetle luciferin allowed us to monitor mitochondrial ATP production in intact cells at 37°C using a photomultiplier tube.…”

Section: Cell Culturesmentioning

confidence: 99%

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Cystine Dimethylester Model of Cystinosis: Still Reliable?

et al. 2007

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ABSTRACT:The ability of cystine dimethylester (CDME) to load lysosomes with cystine has been used to establish the basic defect in cystinosis: defective cystine exodus from lysosomes. Using CDME loading, it has been postulated that cystine accumulation in cystinosis affects mitochondrial ATP production, resulting in defective renal tubular reabsorption. Recent studies in cystinotic fibroblasts, however, show normal adenosine triphosphate (ATP) generation capacity. To investigate the effect of CDME in more detail, mitochondrial ATP generation, reactive oxygen species production, and viability are compared in fibroblasts loaded with CDME with those of cystinotic cells with a defective cystine transporter. Intracellular cystine levels were comparable in fibroblasts loaded with CDME (1 mM, 30 min) and cystinotic fibroblasts. Intracellular ATP levels and mitochondrial ATP production were decreased in fibroblasts loaded with CDME, but normal in cystinotic fibroblasts. Superoxide production was increased with 300% after CDME loading, whereas no changes were observed in cystinotic fibroblasts. Exposure to CDME led to cell death in a time-and concentration-dependent manner. Our data demonstrate that CDME has a toxic effect on mitochondrial ATP production and cell viability. These effects are not observed in cystinotic cells, indicating that a more appropriate model is required for studying the pathogenesis of cystinosis.

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confidence: 50%

Section: Cell Culturesmentioning

confidence: 99%

Section: Cell Culturesmentioning

confidence: 99%

Section: Cell Culturesmentioning

confidence: 99%

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Cystine Dimethylester Model of Cystinosis: Still Reliable?

et al. 2007

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“…The decrease of cystinosin activity causes accumulation of cystine in lysosomes, causing cystinosis, a lethal disease affecting mainly the kidneys (Bellomo et al 2010). Data obtained using in vitro models of cystinosis show to abnormal cysteinylation of proapoptotic kinases, altered cell redox state and decreased the reduced glutathione synthesis, which can impair mitochondrial activity, although mitochondria do not seem to be compromised in cystinosis (Laube et al 2006, Levtchenko et al 2006, Park et al 2006. Although CDME load was used as a model of cystinosis, the elicited effects are considered toxic, and this load is no longer used as a reliable model of cystinosis (Wilmer et al 2007).…”

Section: Discussionmentioning

confidence: 99%

Thiol/disulfide status regulates the activity of thiol-containing kinases related to energy homeostasis in rat kidney

Rech

Mezzomo

Athaydes

et al. 2017

An. Acad. Bras. Ciênc.

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Considering that thiol-containing enzymes like kinases are critical for several metabolic pathways and energy homeostasis, we investigated the effects of cystine dimethyl ester and/or cysteamine administration on kinases crucial for energy metabolism in the kidney of Wistar rats. Animals were injected twice a day with 1.6 µmol/g body weight cystine dimethyl ester and/or 0.26 µmol/g body weight cysteamine from the 16 th to the 20 th postpartum day and euthanized after 12 hours. Pyruvate kinase, adenylate kinase, creatine kinase activities and thiol/disulfide ratio were determined. Cystine dimethyl ester administration reduced thiol/disulfide ratio and inhibited the kinases activities. Cysteamine administration increased the thiol/ disulfide ratio and co-administration with cystine dimethyl ester prevented the inhibition of the enzymes. Regression between the thiol/disulfide ratio, and the kinases activities were significant. These results suggest that redox status may regulate energy metabolism in the rat kidney. If thiol-containing enzymes inhibition and oxidative stress occur in patients with cystinosis, it is possible that lysosomal cystine depletion may not be the only beneficial effect of cysteamine administration, but also its antioxidant and thiol-protector effect.

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Muscle wasting and adipose tissue browning in infantile nephropathic cystinosis

Cheung

Cherqui

Ding

et al. 2015

Journal of Cachexia, Sarcopenia and Muscle

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BackgroundMuscle wasting is a common complication in patients with infantile nephropathic cystinosis, but its mechanism and association with energy metabolism is not known. We define the metabolic phenotype in Ctns−/− mice, an established murine model of infantile nephropathic cystinosis, with focus on muscle wasting and energy homeostasis.MethodsMale Ctns−/− mice and wild‐type (WT) controls were studied at 1, 4, 9, and 12 months of age. As Ctns−/− mice started to develop chronic kidney disease (CKD) at 9 months of age, 9‐ and 12‐month‐old Ctns−/− mice were also compared with age‐matched WT mice with CKD. Serum and urine chemistry and energy homeostasis parameters were measured. Skeletal muscle histomorphometry and in vivo muscle function were measured. We studied expression of genes involved in muscle mass regulation, thermogenesis, energy metabolism, adipogenesis, and adipose tissue browning in Ctns−/− mice.ResultsCtns−/− mice showed loss of weight and lean mass and increased energy expenditure. Ctns−/− mice exhibited abnormal energy homeostasis before the onset of their CKD. Food intake in Ctns−/− mice was comparable with age‐matched WT controls. However, significantly lower total body mass starting at 1 month of age and increased energy expenditure at 4 months of age preceded the onset of CKD at 9 months of age in Ctns−/− mice. Muscle accept content in 1‐ and 4‐month‐old Ctns−/− mice was significantly lower than that in age‐matched WT controls. At 12 months of age, muscle fibre area and in vivo muscle strength was reduced in Ctns−/− mice than that in WT or CKD controls. Muscle wasting in Ctns−/− mice was associated with inhibition of myogenesis, activation of muscle proteolysis pathways, and overexpression of pro‐inflammatory cytokines. Increased energy expenditure was associated with elevation of thermogenesis in skeletal muscle and adipose tissues. The development of beige adipocytes in Ctns−/− mice is a novel finding. Expression of beige adipose cell surface markers (CD137, Tmem26, and Tbx1) and uncoupling protein‐1, which is a brown adipose tissue marker, was observed in inguinal white adipose tissue of Ctns−/− mice. Expression of key molecules implicated in the pathogenesis of adipose tissue browning (Cox2, cytochrome c oxidase subunit II; PGF2α, prostaglandin F2α; IL‐1α, interleukin 1α; IL‐6, interleukin 6; TNF‐α, tumor necrosis factor α) was significantly increased in inguinal white adipose tissue of Ctns−/− mice than that in WT controls.ConclusionThis study describes a mouse model of nephropathic cystinosis presenting with profound muscle wasting. The mechanism for hypermetabolism in Ctns−/− mice may involve up‐regulation of thermogenesis pathways in skeletal muscle and adipose tissues. This study demonstrates, for the first time, the development of beige adipocytes in Ctns−/− mice. Understanding the underlying mechanisms of adipose tissue browning in cystinosis may lead to novel therapy.

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Decreased Intracellular ATP Content and Intact Mitochondrial Energy Generating Capacity in Human Cystinotic Fibroblasts

Cited by 54 publications

References 38 publications

Cystine Dimethylester Model of Cystinosis: Still Reliable?

Cystine Dimethylester Model of Cystinosis: Still Reliable?

Thiol/disulfide status regulates the activity of thiol-containing kinases related to energy homeostasis in rat kidney

Muscle wasting and adipose tissue browning in infantile nephropathic cystinosis

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