The Experts below are selected from a list of 783 Experts worldwide ranked by ideXlab platform
Elena Levtchenko - One of the best experts on this subject based on the ideXlab platform.
-
molecular basis of cystinosis geographic distribution functional consequences of mutations in the CTNS Gene and potential for repair
2019Co-Authors: Dries David, Sante Princiero Berlingerio, Mohamed A Elmonem, Fanny Oliveira Arcolino, Neveen A Soliman, Bert Van Den Heuvel, Rik Gijsbers, Elena LevtchenkoAbstract:Mutations in the CTNS Gene encoding the lysosomal membrane cystine transporter cystinosin are the cause of cystinosis, an autosomal recessive lysosomal storage disease. More than 140 CTNS mutations have been reported worldwide. Recent studies have discovered that cystinosin exerts other key cellular functions beyond cystine transport such as regulation of oxidative state, lysosomal dynamics and autophagy. Here, we review the different mutations described in the CTNS Gene and the geographical distribution of incidence. In addition, the characteristics of the various mutations in relation to the functions of cystinosin needs to be further elucidated. In this review, we highlight the functional consequences of the different mutations in correlation with the clinical phenotypes. Moreover, we propose how this understanding would be fundamental for the development of new technologies through targeted Gene therapy, holding promises for a possible cure of the kidney and extra-renal phenotypes of cystinosis.
-
Original Article Altered status of glutathione and its metabolites in cystinotic cells
2016Co-Authors: Elena Levtchenko, Leo A H Monnens, Lambertus Van Den Heuvel, Adriana De Graaf-hess, Martijn Wilmer, Henk BlomAbstract:Background. Cystinosis is an autosomal recessive disorder, caused by mutations of the lysosomal cystine carrier cystinosin, encoded by the CTNS Gene (17p13). The concomitant intralysosomal cystine accumula-tion leads to multi-organ damage, with kidneys being the first affected. Altered mitochondrial oxidative phosphorylation has been demonstrated in animal proximal tubules loaded with cystine dimethyl ester, mimicking cystine accumulation in cystinosis, but has not been confirmed in cells of patients with cystinosis. Furthermore, the link between cystine accumulation and mitochondrial damage is also missing. We hypothesized that cytosolic cysteine deficiency result-ing in intracellular glutathione (GSH) shortage migh
-
Stem Cell Microvesicles Transfer Cystinosin to Human Cystinotic Cells and Reduce Cystine Accumulation In
2016Co-Authors: Diana M Iglesias, Elena Levtchenko, Francesco Emma, Reyhan El Kares, Anna Taranta, Francesco Bellomo, Martine Besouw, Jaan Toelen, Lambertus Van Den Heuvel, Leelee ChuAbstract:Cystinosis is a rare disease caused by homozygous mutations of the CTNS Gene, encoding a cystine efflux channel in the lysosomal membrane. In CTNS knockout mice, the pathologic intralysosomal accumulation of cystine that drives progressive organ damage can be reversed by infusion of wildtype bone marrow-derived stem cells, but the mechanism involved is unclear since the exoGeneous stem cells are rarely integrated into renal tubules. Here we show that human mesenchymal stem cells, from amniotic fluid or bone marrow, reduce pathologic cystine accumulation in co-cultured CTNS mutant fibroblasts or proximal tubular cells from cystinosis patients. This paracrine effect is associated with release into the culture medium of stem cell microvesicles (100–400 nm diameter) containing wildtype cystinosin protein and CTNS mRNA. Isolated stem cell microvesicles reduce target cell cystine accumulation in a dose-dependent, Annexin V-sensitive manner. Microvesicles from stem cells expressing CTNSRed transfer tagged CTNS protein to the lysosome/endosome compartment of cystinotic fibroblasts. Our observations suggest that exogenous stem cells may reprogram the biology of mutant tissues b
-
Cystinosis: a review
2016Co-Authors: Mohamed A Elmonem, Koenraad Veys, Neveen A Soliman, Lambertus P Van Den Heuvel, Maria Van Dyck, Elena LevtchenkoAbstract:Cystinosis is the most common hereditary cause of renal Fanconi syndrome in children. It is an autosomal recessive lysosomal storage disorder caused by mutations in the CTNS Gene encoding for the carrier protein cystinosin, transporting cystine out of the lysosomal compartment. Defective cystinosin function leads to intra-lysosomal cystine accumulation in all body cells and organs. The kidneys are initially affected during the first year of life through proximal tubular damage followed by progressive glomerular damage and end stage renal failure during mid-childhood if not treated. Other affected organs include eyes, thyroid, pancreas, gonads, muscles and CNS. Leucocyte cystine assay is the cornerstone for both diagnosis and therapeutic monitoring of the disease. Several lines of treatment are available for cystinosis including the cystine depleting agent cysteamine, renal replacement therapy, hormonal therapy and others; however, no curative treatment is yet available. In the current review we will discuss the most important clinical features of the disease, advantages and disadvantages of the current diagnostic and therapeutic options and the main topics of future research in cystinosis.
-
doi:10.1093/ndt/gfh932 Original Article Altered status of glutathione and its metabolites in cystinotic cells
2015Co-Authors: Elena Levtchenko, Leo A H Monnens, Lambertus Van Den Heuvel, Adriana De Graaf-hess, Martijn Wilmer, Henk BlomAbstract:Background. Cystinosis is an autosomal recessive disorder, caused by mutations of the lysosomal cystine carrier cystinosin, encoded by the CTNS Gene (17p13). The concomitant intralysosomal cystine accumula-tion leads to multi-organ damage, with kidneys being the first affected. Altered mitochondrial oxidative phosphorylation has been demonstrated in animal proximal tubules loaded with cystine dimethyl ester, mimicking cystine accumulation in cystinosis, but has not been confirmed in cells of patients with cystinosis. Furthermore, the link between cystine accumulation and mitochondrial damage is also missing. We hypothesized that cytosolic cysteine deficiency result-ing in intracellular glutathione (GSH) shortage migh
Francesco Emma - One of the best experts on this subject based on the ideXlab platform.
-
mitochondrial dynamics of proximal tubular epithelial cells in nephropathic cystinosis
2019Co-Authors: Domenico De Rasmo, Francesco Emma, Elena V Polishchuk, Ester De Leo, Roman S Polishchuk, Anna Signorile, Anna Ferretta, Roberto Raso, Silvia Russo, Francesco BellomoAbstract:Nephropathic cystinosis is a rare lysosomal storage disorder caused by mutations in CTNS Gene leading to Fanconi syndrome. Independent studies reported defective clearance of damaged mitochondria and mitochondrial fragmentation in cystinosis. Proteins involved in the mitochondrial dynamics and the mitochondrial ultrastructure were analyzed in CTNS-/- cells treated with cysteamine, the only drug currently used in the therapy for cystinosis but ineffective to treat Fanconi syndrome. CTNS-/- cells showed an overexpression of parkin associated with deregulation of ubiquitination of mitofusin 2 and fission 1 proteins, an altered proteolytic processing of optic atrophy 1 (OPA1), and a decreased OPA1 oligomerization. According to molecular findings, the analysis of electron microscopy images showed a decrease of mitochondrial cristae number and an increase of cristae lumen and cristae junction width. Cysteamine treatment restored the fission 1 ubiquitination, the mitochondrial size, number and lumen of cristae, but had no effect on cristae junction width, making CTNS-/- tubular cells more susceptible to apoptotic stimuli.
-
impact of atypical mitochondrial cyclic amp level in nephropathic cystinosis
2018Co-Authors: Francesco Bellomo, Francesco Emma, Domenico De Rasmo, Anna Signorile, Grazia Tamma, Marianna RanieriAbstract:Nephropathic cystinosis (NC) is a rare disease caused by mutations in the CTNS Gene encoding for cystinosin, a lysosomal transmembrane cystine/H+ symporter, which promotes the efflux of cystine from lysosomes to cytosol. NC is the most frequent cause of Fanconi syndrome (FS) in young children, the molecular basis of which is not well established. Proximal tubular cells have very high metabolic rate due to the active transport of many solutes. Not surprisingly, mitochondrial disorders are often characterized by FS. A similar mechanism may also apply to NC. Because cAMP has regulatory properties on mitochondrial function, we have analyzed cAMP levels and mitochondrial targets in CTNS−/− conditionally immortalized proximal tubular epithelial cells (ciPTEC) carrying the classical homozygous 57-kb deletion (delCTNS−/−) or with compound heterozygous loss-of-function mutations (mutCTNS−/−). Compared to wild-type cells, cystinotic cells had significantly lower mitochondrial cAMP levels (delCTNS−/− ciPTEC by 56% ± 10.5, P < 0.0001; mutCTNS−/− by 26% ± 4.3, P < 0.001), complex I and V activities, mitochondrial membrane potential, and SIRT3 protein levels, which were associated with increased mitochondrial fragmentation. Reduction of complex I and V activities was associated with lower expression of part of their subunits. Treatment with the non-hydrolysable cAMP analog 8-Br-cAMP restored mitochondrial potential and corrected mitochondria morphology. Treatment with cysteamine, which reduces the intra-lysosomal cystine, was able to restore mitochondrial cAMP levels, as well as most other abnormal mitochondrial findings. These observations were validated in CTNS-silenced HK-2 cells, indicating a pivotal role of mitochondrial cAMP in the proximal tubular dysfunction observed in NC.
-
cystinosin lkg rescues cystine accumulation and decreases apoptosis rate in cystinotic proximal tubular epithelial cells
2017Co-Authors: Anna Taranta, Anna Pastore, Stefania Petrini, Francesco Bellomo, Elena V Polishchuk, Ester De Leo, Laura Rita Rega, Roman S Polishchuk, Maria Antonietta De Matteis, Francesco EmmaAbstract:Nephropathic cystinosis is a lysosomal storage disease that is caused by mutations in the CTNS Gene encoding a cystine/proton symporter cystinosin and an isoform cystinosin-LKG which is Generated by an alternative splicing of exon 12. We have investigated the physiological role of the cystinosin-LKG that is widely expressed in epithelial tissues. We have analyzed the intracellular localization and the function of the cystinosin-LKG conjugated with DsRed (cystinosin-LKG-RFP) in Madin-Darby canine kidney cells (MDCK II) and in proximal tubular epithelial cells carrying a deletion of the CTNS Gene (cystinotic PTEC), respectively. Cystinosin-LKG-RFP colocalized with markers of lysosomes, late endosomes and was also expressed on the apical surface of polarized MDCK II cells. Moreover, immune-electron microscopy images of MDCK II cells overexpressing cystinosin-LKG-RFP showed stacked lamellar membranes inside perinuclear lysosomal structures. To study the role of LKG-isoform, we have investigated cystine accumulation and apoptosis that have been described in cystinotic cells. Cystinosin-LKG decreased cystine levels by approximately 10-fold similarly to cystinosin-RFP. The levels of TNFα- and actinomycin D-inducted apoptosis dropped in cystinotic cells expressing LKG-isoform. This effect was also similar to the main isoform. Our results suggest that cystinosin-LKG and cystinosin move similar functional activities in cells.
-
Stem Cell Microvesicles Transfer Cystinosin to Human Cystinotic Cells and Reduce Cystine Accumulation In
2016Co-Authors: Diana M Iglesias, Elena Levtchenko, Francesco Emma, Reyhan El Kares, Anna Taranta, Francesco Bellomo, Martine Besouw, Jaan Toelen, Lambertus Van Den Heuvel, Leelee ChuAbstract:Cystinosis is a rare disease caused by homozygous mutations of the CTNS Gene, encoding a cystine efflux channel in the lysosomal membrane. In CTNS knockout mice, the pathologic intralysosomal accumulation of cystine that drives progressive organ damage can be reversed by infusion of wildtype bone marrow-derived stem cells, but the mechanism involved is unclear since the exoGeneous stem cells are rarely integrated into renal tubules. Here we show that human mesenchymal stem cells, from amniotic fluid or bone marrow, reduce pathologic cystine accumulation in co-cultured CTNS mutant fibroblasts or proximal tubular cells from cystinosis patients. This paracrine effect is associated with release into the culture medium of stem cell microvesicles (100–400 nm diameter) containing wildtype cystinosin protein and CTNS mRNA. Isolated stem cell microvesicles reduce target cell cystine accumulation in a dose-dependent, Annexin V-sensitive manner. Microvesicles from stem cells expressing CTNSRed transfer tagged CTNS protein to the lysosome/endosome compartment of cystinotic fibroblasts. Our observations suggest that exogenous stem cells may reprogram the biology of mutant tissues b
-
nephropathic cystinosis an international consensus document
2014Co-Authors: Francesco Emma, Stephanie Cherqui, Rezan Topaloglu, Galina Nesterova, Craig B Langman, Antoine Labbe, Paul Goodyer, Mirian C H Janssen, Marcella Greco, Ewa ElenbergAbstract:Cystinosis is caused by mutations in the CTNS Gene (17p13.2), which encodes for a lysosomal cystine/proton symporter termed cystinosin. It is the most common cause of inherited renal Fanconi syndrome in young children. Because of its rarity, the diagnosis and specific treatment of cystinosis are frequently delayed, which has a significant impact on the overall prognosis. In this document, we have summarized expert opinions on several aspects of the disease to improve knowledge and provide guidance for diagnosis and treatment.
Francesco Bellomo - One of the best experts on this subject based on the ideXlab platform.
-
mitochondrial dynamics of proximal tubular epithelial cells in nephropathic cystinosis
2019Co-Authors: Domenico De Rasmo, Francesco Emma, Elena V Polishchuk, Ester De Leo, Roman S Polishchuk, Anna Signorile, Anna Ferretta, Roberto Raso, Silvia Russo, Francesco BellomoAbstract:Nephropathic cystinosis is a rare lysosomal storage disorder caused by mutations in CTNS Gene leading to Fanconi syndrome. Independent studies reported defective clearance of damaged mitochondria and mitochondrial fragmentation in cystinosis. Proteins involved in the mitochondrial dynamics and the mitochondrial ultrastructure were analyzed in CTNS-/- cells treated with cysteamine, the only drug currently used in the therapy for cystinosis but ineffective to treat Fanconi syndrome. CTNS-/- cells showed an overexpression of parkin associated with deregulation of ubiquitination of mitofusin 2 and fission 1 proteins, an altered proteolytic processing of optic atrophy 1 (OPA1), and a decreased OPA1 oligomerization. According to molecular findings, the analysis of electron microscopy images showed a decrease of mitochondrial cristae number and an increase of cristae lumen and cristae junction width. Cysteamine treatment restored the fission 1 ubiquitination, the mitochondrial size, number and lumen of cristae, but had no effect on cristae junction width, making CTNS-/- tubular cells more susceptible to apoptotic stimuli.
-
impact of atypical mitochondrial cyclic amp level in nephropathic cystinosis
2018Co-Authors: Francesco Bellomo, Francesco Emma, Domenico De Rasmo, Anna Signorile, Grazia Tamma, Marianna RanieriAbstract:Nephropathic cystinosis (NC) is a rare disease caused by mutations in the CTNS Gene encoding for cystinosin, a lysosomal transmembrane cystine/H+ symporter, which promotes the efflux of cystine from lysosomes to cytosol. NC is the most frequent cause of Fanconi syndrome (FS) in young children, the molecular basis of which is not well established. Proximal tubular cells have very high metabolic rate due to the active transport of many solutes. Not surprisingly, mitochondrial disorders are often characterized by FS. A similar mechanism may also apply to NC. Because cAMP has regulatory properties on mitochondrial function, we have analyzed cAMP levels and mitochondrial targets in CTNS−/− conditionally immortalized proximal tubular epithelial cells (ciPTEC) carrying the classical homozygous 57-kb deletion (delCTNS−/−) or with compound heterozygous loss-of-function mutations (mutCTNS−/−). Compared to wild-type cells, cystinotic cells had significantly lower mitochondrial cAMP levels (delCTNS−/− ciPTEC by 56% ± 10.5, P < 0.0001; mutCTNS−/− by 26% ± 4.3, P < 0.001), complex I and V activities, mitochondrial membrane potential, and SIRT3 protein levels, which were associated with increased mitochondrial fragmentation. Reduction of complex I and V activities was associated with lower expression of part of their subunits. Treatment with the non-hydrolysable cAMP analog 8-Br-cAMP restored mitochondrial potential and corrected mitochondria morphology. Treatment with cysteamine, which reduces the intra-lysosomal cystine, was able to restore mitochondrial cAMP levels, as well as most other abnormal mitochondrial findings. These observations were validated in CTNS-silenced HK-2 cells, indicating a pivotal role of mitochondrial cAMP in the proximal tubular dysfunction observed in NC.
-
cystinosin lkg rescues cystine accumulation and decreases apoptosis rate in cystinotic proximal tubular epithelial cells
2017Co-Authors: Anna Taranta, Anna Pastore, Stefania Petrini, Francesco Bellomo, Elena V Polishchuk, Ester De Leo, Laura Rita Rega, Roman S Polishchuk, Maria Antonietta De Matteis, Francesco EmmaAbstract:Nephropathic cystinosis is a lysosomal storage disease that is caused by mutations in the CTNS Gene encoding a cystine/proton symporter cystinosin and an isoform cystinosin-LKG which is Generated by an alternative splicing of exon 12. We have investigated the physiological role of the cystinosin-LKG that is widely expressed in epithelial tissues. We have analyzed the intracellular localization and the function of the cystinosin-LKG conjugated with DsRed (cystinosin-LKG-RFP) in Madin-Darby canine kidney cells (MDCK II) and in proximal tubular epithelial cells carrying a deletion of the CTNS Gene (cystinotic PTEC), respectively. Cystinosin-LKG-RFP colocalized with markers of lysosomes, late endosomes and was also expressed on the apical surface of polarized MDCK II cells. Moreover, immune-electron microscopy images of MDCK II cells overexpressing cystinosin-LKG-RFP showed stacked lamellar membranes inside perinuclear lysosomal structures. To study the role of LKG-isoform, we have investigated cystine accumulation and apoptosis that have been described in cystinotic cells. Cystinosin-LKG decreased cystine levels by approximately 10-fold similarly to cystinosin-RFP. The levels of TNFα- and actinomycin D-inducted apoptosis dropped in cystinotic cells expressing LKG-isoform. This effect was also similar to the main isoform. Our results suggest that cystinosin-LKG and cystinosin move similar functional activities in cells.
-
ORIGINAL ARTICLE Analysis of CTNS Gene transcripts in nephropathic cystinosis
2016Co-Authors: Francesco BellomoAbstract:Abstract Nephropathic cystinosis (NC) is an autosomal recessive disorder caused by mutations of the CTNS Gene that encodes for a cystine transmembrane transporter. Several mutations have been described in the coding and promoter regions of the CTNS Gene in affected individuals. We selected three patients with NC from two unrelated families, in whom sequence analysis of the CTNS Gene detected only one or no mutations. Total RNAwas isolated from peripheral blood mononuclear cells or fibroblasts and CTNS transcripts were analyzed. We observed a skipping of exon 5 (85 bp) in two siblings and an intron 9 retention of 75 bp associated with partial replication of exon 9 in the third patient. Genomic DNA analysis of intron regions surrounding exon 5 showed a point mutation in the hypothetical lariat branch site of intron 4 at position –24 (c.141–24 T>C) in the first two patients and a duplication of 266 bp including a part of exon and intron 9 in the third patient. Analysis of CTNS Gene transcripts allowed identification of mutations in patients in whom CTNS mutations could not be detected by traditional DNA sequencing. These results support the hypothesis that cystinosis is a monogenic disorder
-
Transcriptional and Posttranscriptional Regulation of the
2016Co-Authors: Ctns Gene, Anna Taranta, Francesco Bellomo, Serena Corallini, Alessia Palma, Anna PastoreAbstract:ABSTRACT: Cell cysteine (Cys) levels and/or the [Cys/CySS] redox potential have been shown to regulate mRNA levels of the CTNS Gene, which encodes for a lysosomal cystine (CySS) carrier that is defective in cystinosis. To investigate the mechanisms in-volved CTNS mRNA regulation, different portions of the CTNS promotor were cloned into a luciferase vector and transfected in HK2 cells. A 1.5–2.4-fold increase in luciferase activity was observed when cells were incubated in culture medium containing low CySS concentrations. Conversely, CTNS mRNA levels decreased by 47– 56 % in the presence of N-acetyl-L-cysteine (NAC). Chase experi-ments with actinomycin D (ActD) demonstrated a 3-fold stabilization of the CTNS mRNA when cells were cultured in low CySS medium for 48 h. Treatment of control cells with cyclohexamide (CHX) increased CTNS mRNA levels, suggesting that CHX blocked the synthesis of proteins involved in mRNA degradation or in repressio
Anna Taranta - One of the best experts on this subject based on the ideXlab platform.
-
cystinosin lkg rescues cystine accumulation and decreases apoptosis rate in cystinotic proximal tubular epithelial cells
2017Co-Authors: Anna Taranta, Anna Pastore, Stefania Petrini, Francesco Bellomo, Elena V Polishchuk, Ester De Leo, Laura Rita Rega, Roman S Polishchuk, Maria Antonietta De Matteis, Francesco EmmaAbstract:Nephropathic cystinosis is a lysosomal storage disease that is caused by mutations in the CTNS Gene encoding a cystine/proton symporter cystinosin and an isoform cystinosin-LKG which is Generated by an alternative splicing of exon 12. We have investigated the physiological role of the cystinosin-LKG that is widely expressed in epithelial tissues. We have analyzed the intracellular localization and the function of the cystinosin-LKG conjugated with DsRed (cystinosin-LKG-RFP) in Madin-Darby canine kidney cells (MDCK II) and in proximal tubular epithelial cells carrying a deletion of the CTNS Gene (cystinotic PTEC), respectively. Cystinosin-LKG-RFP colocalized with markers of lysosomes, late endosomes and was also expressed on the apical surface of polarized MDCK II cells. Moreover, immune-electron microscopy images of MDCK II cells overexpressing cystinosin-LKG-RFP showed stacked lamellar membranes inside perinuclear lysosomal structures. To study the role of LKG-isoform, we have investigated cystine accumulation and apoptosis that have been described in cystinotic cells. Cystinosin-LKG decreased cystine levels by approximately 10-fold similarly to cystinosin-RFP. The levels of TNFα- and actinomycin D-inducted apoptosis dropped in cystinotic cells expressing LKG-isoform. This effect was also similar to the main isoform. Our results suggest that cystinosin-LKG and cystinosin move similar functional activities in cells.
-
Transcriptional and Posttranscriptional Regulation of the
2016Co-Authors: Ctns Gene, Anna Taranta, Francesco Bellomo, Serena Corallini, Alessia Palma, Anna PastoreAbstract:ABSTRACT: Cell cysteine (Cys) levels and/or the [Cys/CySS] redox potential have been shown to regulate mRNA levels of the CTNS Gene, which encodes for a lysosomal cystine (CySS) carrier that is defective in cystinosis. To investigate the mechanisms in-volved CTNS mRNA regulation, different portions of the CTNS promotor were cloned into a luciferase vector and transfected in HK2 cells. A 1.5–2.4-fold increase in luciferase activity was observed when cells were incubated in culture medium containing low CySS concentrations. Conversely, CTNS mRNA levels decreased by 47– 56 % in the presence of N-acetyl-L-cysteine (NAC). Chase experi-ments with actinomycin D (ActD) demonstrated a 3-fold stabilization of the CTNS mRNA when cells were cultured in low CySS medium for 48 h. Treatment of control cells with cyclohexamide (CHX) increased CTNS mRNA levels, suggesting that CHX blocked the synthesis of proteins involved in mRNA degradation or in repressio
-
Stem Cell Microvesicles Transfer Cystinosin to Human Cystinotic Cells and Reduce Cystine Accumulation In
2016Co-Authors: Diana M Iglesias, Elena Levtchenko, Francesco Emma, Reyhan El Kares, Anna Taranta, Francesco Bellomo, Martine Besouw, Jaan Toelen, Lambertus Van Den Heuvel, Leelee ChuAbstract:Cystinosis is a rare disease caused by homozygous mutations of the CTNS Gene, encoding a cystine efflux channel in the lysosomal membrane. In CTNS knockout mice, the pathologic intralysosomal accumulation of cystine that drives progressive organ damage can be reversed by infusion of wildtype bone marrow-derived stem cells, but the mechanism involved is unclear since the exoGeneous stem cells are rarely integrated into renal tubules. Here we show that human mesenchymal stem cells, from amniotic fluid or bone marrow, reduce pathologic cystine accumulation in co-cultured CTNS mutant fibroblasts or proximal tubular cells from cystinosis patients. This paracrine effect is associated with release into the culture medium of stem cell microvesicles (100–400 nm diameter) containing wildtype cystinosin protein and CTNS mRNA. Isolated stem cell microvesicles reduce target cell cystine accumulation in a dose-dependent, Annexin V-sensitive manner. Microvesicles from stem cells expressing CTNSRed transfer tagged CTNS protein to the lysosome/endosome compartment of cystinotic fibroblasts. Our observations suggest that exogenous stem cells may reprogram the biology of mutant tissues b
-
Characterization of mesenchymal stem cells and CTNS(−/−) mutant target cells.
2013Co-Authors: Diana M Iglesias, Elena Levtchenko, Francesco Emma, Reyhan El Kares, Anna Taranta, Francesco Bellomo, Martine Besouw, Jaan Toelen, Lambertus Van Den Heuvel, Leelee ChuAbstract:(a) FACS analysis (left) of bmMSC surface markers (blue) and isotype controls (red) and bmMSC differentiation from baseline (upper right) to Alizarin Red S-stained calcium-containing osteogenic (middle right) and Oil-Red O-stained adipocyte (bottom right) phenotypes. (b) same FACS and differentiation analysis of amMSC cells. (c) schematic diagram depicting the common 57 Kb deletion removing the 5′ portion of the CTNS Gene and PCR-proven genotype of control and mutant fibroblasts from a cystinosis patient. Intracellular cystine content of control and homozygous CTNS mutant fibroblasts is shown on the right bar-graph.
-
transcriptional and posttranscriptional regulation of the CTNS Gene
2011Co-Authors: Serena Corallini, Anna Pastore, Anna Taranta, Francesco Bellomo, Alessia Palma, Francesco EmmaAbstract:Cell cysteine (Cys) levels and/or the [Cys/CySS] redox potential have been shown to regulate mRNA levels of the CTNS Gene, which encodes for a lysosomal cystine (CySS) carrier that is defective in cystinosis. To investigate the mechanisms involved CTNS mRNA regulation, different portions of the CTNS promotor were cloned into a luciferase vector and transfected in HK2 cells. A 1.5–2.4-fold increase in luciferase activity was observed when cells were incubated in culture medium containing low CySS concentrations. Conversely, CTNS mRNA levels decreased by 47–56% in the presence of N-acetyl-l-cysteine (NAC). Chase experiments with actinomycin D (ActD) demonstrated a 3-fold stabilization of the CTNS mRNA when cells were cultured in low CySS medium for 48 h. Treatment of control cells with cyclohexamide (CHX) increased CTNS mRNA levels, suggesting that CHX blocked the synthesis of proteins involved in mRNA degradation or in repression of the CTNS Gene. Finally, in vitro binding assays showed increased binding (30–110%) of the Sp-1 transcription factor to two regions of the CTNS promotor when cells were incubated in low CySS medium. These results indicate that the CTNS Gene is actively regulated at the transcriptional and posttranscriptional levels and suggest that CTNS plays a pivotal role in regulating cell thiol concentrations.
Levtchenko E.n. - One of the best experts on this subject based on the ideXlab platform.
-
Ca(2+) signalling in human proximal tubular epithelial cells deficient for cystinosin
2016Co-Authors: Ivanova E.a., Heuvel, L.p.w.j. Van Den, Levtchenko E.n., Luyten T., Missiaen L., Bultynck G.Abstract:Item does not contain fulltextNephropathic cystinosis is an autosomal recessive lysosomal storage disorder caused by loss-of-function mutations in the CTNS Gene coding for the lysosomal cystine transporter, cystinosin. Recent studies have demonstrated that, apart from cystine accumulation in the lysosomes, cystinosin-deficient cells, especially renal proximal tubular epithelial cells are characterized by abnormal vesicle trafficking and endocytosis, possible lysosomal dysfunction and perturbed intracellular signalling cascades. It is therefore possible that Ca(2+) signalling is disturbed in cystinosis, as it has been demonstrated for other disorders associated with lysosomal dysfunction, such as Gaucher, Niemann-Pick type C and Alzheimer's diseases. In this study we investigated ATP-induced, IP3-induced and lysosomal Ca(2+) release in human proximal tubular epithelial cells derived from control and cystinotic patients. No major dysregulation of intracellular Ca(2+) dynamics was found, although ATP-induced Ca(2+) release appeared slightly sensitized in cystinotic cells compared to control cells. Hence, these subtle changes in Ca(2+) signals elicited by agonists may contribute to the pathoGenesis of the disease
-
Ca(2+) signalling in human proximal tubular epithelial cells deficient for cystinosin
2016Co-Authors: Ivanova E.a., Heuvel, L.p.w.j. Van Den, Levtchenko E.n., Luyten T., Missiaen L., Bultynck G.Abstract:Nephropathic cystinosis is an autosomal recessive lysosomal storage disorder caused by loss-of-function mutations in the CTNS Gene coding for the lysosomal cystine transporter, cystinosin. Recent studies have demonstrated that, apart from cystine accumulation in the lysosomes, cystinosin-deficient cells, especially renal proximal tubular epithelial cells are characterized by abnormal vesicle trafficking and endocytosis, possible lysosomal dysfunction and perturbed intracellular signalling cascades. It is therefore possible that Ca(2+) signalling is disturbed in cystinosis, as it has been demonstrated for other disorders associated with lysosomal dysfunction, such as Gaucher, Niemann-Pick type C and Alzheimer's diseases. In this study we investigated ATP-induced, IP3-induced and lysosomal Ca(2+) release in human proximal tubular epithelial cells derived from control and cystinotic patients. No major dysregulation of intracellular Ca(2+) dynamics was found, although ATP-induced Ca(2+) release appeared slightly sensitized in cystinotic cells compared to control cells. Hence, these subtle changes in Ca(2+) signals elicited by agonists may contribute to the pathoGenesis of the disease
-
TRPV1 dysfunction in cystinosis patients harboring the homozygous 57 kb deletion
2016Co-Authors: Buntinx L., Voets T., Morlion B., Vangeel L., Janssen M., Cornelissen E.a., Vriens J., J. De ,hoon, Levtchenko E.n.Abstract:Cystinosis is a rare autosomal recessive disorder characterized by lysosomal cystine accumulation due to loss of function of the lysosomal cystine transporter (CTNS). The most common mutation in cystinosis patients of Northern Europe consists of a 57-kb deletion. This deletion not only inactivates the CTNS Gene but also extends into the non-coding region upstream of the start codon of the TRPV1 Gene, encoding the capsaicin- and heat-sensitive ion channel TRPV1. To evaluate the consequences of the 57-kb deletion on functional TRPV1 expression, we compared thermal, mechanical and chemical sensitivity of cystinosis patients with matched healthy controls. Whereas patients heterozygous for the 57-kb deletion showed normal sensory responses, homozygous subjects exhibited a 60% reduction in vasodilation and pain evoked by capsaicin, as well as an increase in heat detection threshold. Responses to cold, mechanical stimuli or cinnamaldehyde, an agonist of the related nociceptor channel TRPA1, were unaltered. We conclude that cystinosis patients homozygous for the 57-kb deletion exhibit a strong reduction of TRPV1 function, leading to sensory deficiencies akin to the phenotype of TRPV1-deficient mice. These deficits may account for the reported sensory alterations and thermoregulatory deficits in these patients, and provide a paradigm for life-long TRPV1 deficiency in humans
-
TRPV1 dysfunction in cystinosis patients harboring the homozygous 57 kb deletion
2016Co-Authors: Buntinx L., Voets T., Morlion B., Vangeel L., Janssen M., Cornelissen E.a., Vriens J., J. De ,hoon, Levtchenko E.n.Abstract:Contains fulltext : 171607.pdf (publisher's version ) (Open Access)Cystinosis is a rare autosomal recessive disorder characterized by lysosomal cystine accumulation due to loss of function of the lysosomal cystine transporter (CTNS). The most common mutation in cystinosis patients of Northern Europe consists of a 57-kb deletion. This deletion not only inactivates the CTNS Gene but also extends into the non-coding region upstream of the start codon of the TRPV1 Gene, encoding the capsaicin- and heat-sensitive ion channel TRPV1. To evaluate the consequences of the 57-kb deletion on functional TRPV1 expression, we compared thermal, mechanical and chemical sensitivity of cystinosis patients with matched healthy controls. Whereas patients heterozygous for the 57-kb deletion showed normal sensory responses, homozygous subjects exhibited a 60% reduction in vasodilation and pain evoked by capsaicin, as well as an increase in heat detection threshold. Responses to cold, mechanical stimuli or cinnamaldehyde, an agonist of the related nociceptor channel TRPA1, were unaltered. We conclude that cystinosis patients homozygous for the 57-kb deletion exhibit a strong reduction of TRPV1 function, leading to sensory deficiencies akin to the phenotype of TRPV1-deficient mice. These deficits may account for the reported sensory alterations and thermoregulatory deficits in these patients, and provide a paradigm for life-long TRPV1 deficiency in humans
-
Endo-lysosomal dysfunction in human proximal tubular epithelial cells deficient for lysosomal cystine transporter cystinosin
2015Co-Authors: Ivanova E.a., Maria Antonietta De Matteis, Heuvel, L.p.w.j. Van Den, Maria Giovanna De Leo, Pastore A., Dijkman H.b.p.m., Levtchenko E.n.Abstract:Contains fulltext : 155173.PDF (publisher's version ) (Open Access)Nephropathic cystinosis is a lysosomal storage disorder caused by mutations in the CTNS Gene encoding cystine transporter cystinosin that results in accumulation of amino acid cystine in the lysosomes throughout the body and especially affects kidneys. Early manifestations of the disease include renal Fanconi syndrome, a Generalized proximal tubular dysfunction. Current therapy of cystinosis is based on cystine-lowering drug cysteamine that postpones the disease progression but offers no cure for the Fanconi syndrome. We studied the mechanisms of impaired reabsorption in human proximal tubular epithelial cells (PTEC) deficient for cystinosin and investigated the endo-lysosomal compartments of cystinosin-deficient PTEC by means of light and electron microscopy. We demonstrate that cystinosin-deficient cells had abnormal shape and distribution of the endo-lysosomal compartments and impaired endocytosis, with decreased surface expression of multiligand receptors and delayed lysosomal cargo processing. Treatment with cysteamine improved surface expression and lysosomal cargo processing but did not lead to a complete restoration and had no effect on the abnormal morphology of endo-lysosomal compartments. The obtained results improve our understanding of the mechanism of proximal tubular dysfunction in cystinosis and indicate that impaired protein reabsorption can, at least partially, be explained by abnormal trafficking of endosomal vesicles