The Experts below are selected from a list of 282 Experts worldwide ranked by ideXlab platform
Patrick Vinay - One of the best experts on this subject based on the ideXlab platform.
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Receptor-mediated endocytosis in kidney Proximal Tubules: recent advances and hypothesis.
Electrophoresis, 1997Co-Authors: Vladimir Marshansky, Sylvain G. Bourgoin, Irène Londono, Moise Bendayan, Bruno Maranda, Patrick VinayAbstract:Preparation of kidney Proximal Tubules in suspension allows the study of receptor-mediated endocytosis, protein reabsorption, and traffic of endosomal vesicles. The study of tubular protein transport in vitro coupled with that of the function of endosomal preparation offers a unique opportunity to investigate a receptor-mediated endocytosis pathway under physiological and pathological conditions. We assume that receptor-mediated endocytosis of albumin in kidney Proximal Tubules in situ and in vitro can be regulated, on the one hand, by the components of the acidification machinery (V-type H+-ATPase, Cl(-)-channel and Na+/H+-exchanger), giving rise to formation and dissipation of a proton gradient in endosomal vesicles, and, on the other hand, by small GTPases of the ADP-ribosylation factor (Arf)-family. In this paper we thus analyze the recent advances of the studies of cellular and molecular mechanisms underlying the identification, localization, and function of the acidification machinery (V-type H+-ATPase, Cl(-)-channel) as well as Arf-family small GTPases and phospholipase D in the endocytotic pathway of kidney Proximal Tubules. Also, we explore the possible functional interaction between the acidification machinery and Arf-family small GTPases. Finally, we propose the hypothesis of the regulation of translocation of Arf-family small GTPases by an endosomal acidification process and its role during receptor-mediated endocytosis in kidney Proximal Tubules. The results of this study will not only enhance our understanding of the receptor-mediated endocytosis pathway in kidney Proximal Tubules under physiological conditions but will also have important implications with respect to the functional consequences under some pathological circumstances. Furthermore, it may suggest novel targets and approaches in the prevention and treatment of various diseases (cystic fibrosis, Dent's disease, diabetes and autosomal dominant polycystic kidney disease).
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Identification of ADP-ribosylation factor-6 in brush-border membrane and early endosomes of human kidney Proximal Tubules
Electrophoresis, 1997Co-Authors: Vladimir Marshansky, Sylvain G. Bourgoin, Irène Londono, Moise Bendayan, Patrick VinayAbstract:The expression and distribution of ADP-ribosylation factor (ARF) small GTP-binding proteins in kidney tissue was examined. Various anti-ARF antibodies were raised against purified rec-ARF 1 and rec-ARF 6 and their specificity was determined. Using indirect immunofluorescence analysis of intact kidney, ARF proteins were found to be predominantly expressed in kidney Tubules as compared to glomeruli. This result was further supported by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis of purified human kidney glomeruli and Proximal Tubules. Both ARF 1 and ARF 6 were detected in purified human glomeruli and Proximal Tubules; however, ARF 1 was more abundant than ARF 6 in these kidney structures. Brush-border membrane vesicles (BBMV) and early endosomes (EE) derived from the receptor-mediated endocytosis pathway were isolated from purified Proximal Tubules of rat, dog and human kidney using a combination of magnesium precipitation and wheat-germ agglutinin negative selection techniques. We demonstrated that ARF 6 is associated with BBMV and with EE derived from receptor-mediated endocytosis pathway of human kidney Proximal Tubules. Using a combination of SDS-PAGE and quantitative enhanced chemiluminescence Western blot analysis, the quantification of the ARF 6 distribution in membrane and cytoplasmic fractions of Proximal Tubules was made and its predominance in membrane fractions was demonstrated. By analogy with the functional role of ARF 1 in Golgi protein transport, we suggest that ARF 6 may play an important role in the regulation of receptor-mediated endocytosis and protein reabsorption by kidney Proximal Tubules.
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Proton gradient formation in early endosomes from Proximal Tubules
Biochimica et biophysica acta, 1996Co-Authors: Vladimir Marshansky, Patrick VinayAbstract:Heavy endosomes were isolated from Proximal Tubules using a combination of magnesium precipitation and wheat-germ agglutinin negative selection techniques. Two small GTPases (Rab4 and Rab5) known to be specifically present in early endosomes were identified in our preparations. Endosomal acidification was followed fluorimetrically using acridine orange. In presence of chloride ions and ATP, the formation of a proton gradient (delta pH) was observed. This process is due to the activity of an electrogenic V-type ATPase present in the endosomal membrane since specific inhibitors bafilomycin and folimycin effectively prevented or eliminated endosomal acidification. In presence of chloride ions (K(m) = 30 mM) the formation of the proton gradient was optimal. Inhibitors of chloride channel activity such as DIDS and NPPB reduced acidification. The presence of sodium ions stimulated the dissipation of the proton gradient. This effect of sodium was abolished by amiloride derivative (MIA) but only when loaded into endosomes, indicating the presence of a physiologically oriented Na+/H(+)-exchanger in the endosomal membrane. Monensin restored the gradient dissipation. Thus three proteins (V-type ATPase, Cl(-)-channel, Na+/H(+)-exchanger) present in early endosomes isolated from Proximal Tubules may regulate the formation, maintenance and dissipation of the proton gradient.
Joel M Weinberg - One of the best experts on this subject based on the ideXlab platform.
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Substrate Modulation of Fatty Acid Effects on Energization and Respiration of Kidney Proximal Tubules during Hypoxia/Reoxygenation
PloS one, 2014Co-Authors: Anja Bienholz, Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ahmad Al-taweel, Joel M WeinbergAbstract:Kidney Proximal Tubules subjected to hypoxia/reoxygenation develop a nonesterified fatty acid-induced energetic deficit characterized by persistent partial mitochondrial deenergization that can be prevented and reversed by citric acid cycle substrates. To further assess the role of competition between fatty acids and substrates on inner membrane substrate carriers in the deenergization and the contribution to deenergization of fatty acid effects on respiratory function, digitonin-permeabilized rabbit and mouse Tubules were studied using either addition of exogenous oleate after control normoxic incubation or increases of endogenous fatty acids produced by hypoxia/reoxygenation. The results demonstrated major effects of matrix oxaloacetate accumulation on succinate-supported energization and respiration and their modification by fatty acids. Improvements of energization in the presence of fatty acids by glutamate were shown to result predominantly from lowering matrix oxaloacetate rather than from amelioration of transmembrane cycling of fatty acids and uncoupling. Mouse Tubules had 2.5 fold higher rates of succinate utilization, which resulted in stronger effects of oxaloacetate accumulation than rabbit Tubules. Hypoxia/reoxygenation induced respiratory inhibition that was more severe for complex I-dependent substrates. Fatty acids themselves did not acutely contribute to this respiratory inhibition, but lowering them during 60 min. reoxygenation to allow recovery of ATP during that period alleviated it. These data clarify the basis for the nonesterified fatty acid-induced mitochondrial energetic deficit in kidney Proximal Tubules that impairs structural and functional recovery and provide insight into interactions that need to be considered in the design of substrate-based interventions to improve mitochondrial function.
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substrate modulation of fatty acid effects on energization and respiration of kidney Proximal Tubules during hypoxia reoxygenation
PLOS ONE, 2014Co-Authors: Anja Bienholz, Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ahmad Altaweel, Joel M WeinbergAbstract:Kidney Proximal Tubules subjected to hypoxia/reoxygenation develop a nonesterified fatty acid-induced energetic deficit characterized by persistent partial mitochondrial deenergization that can be prevented and reversed by citric acid cycle substrates. To further assess the role of competition between fatty acids and substrates on inner membrane substrate carriers in the deenergization and the contribution to deenergization of fatty acid effects on respiratory function, digitonin-permeabilized rabbit and mouse Tubules were studied using either addition of exogenous oleate after control normoxic incubation or increases of endogenous fatty acids produced by hypoxia/reoxygenation. The results demonstrated major effects of matrix oxaloacetate accumulation on succinate-supported energization and respiration and their modification by fatty acids. Improvements of energization in the presence of fatty acids by glutamate were shown to result predominantly from lowering matrix oxaloacetate rather than from amelioration of transmembrane cycling of fatty acids and uncoupling. Mouse Tubules had 2.5 fold higher rates of succinate utilization, which resulted in stronger effects of oxaloacetate accumulation than rabbit Tubules. Hypoxia/reoxygenation induced respiratory inhibition that was more severe for complex I-dependent substrates. Fatty acids themselves did not acutely contribute to this respiratory inhibition, but lowering them during 60 min. reoxygenation to allow recovery of ATP during that period alleviated it. These data clarify the basis for the nonesterified fatty acid-induced mitochondrial energetic deficit in kidney Proximal Tubules that impairs structural and functional recovery and provide insight into interactions that need to be considered in the design of substrate-based interventions to improve mitochondrial function.
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Accumulation of nonesterified fatty acids causes the sustained energetic deficit in kidney Proximal Tubules after hypoxia-reoxygenation.
American journal of physiology. Renal physiology, 2005Co-Authors: Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ruth A Senter, Joel M WeinbergAbstract:Kidney Proximal Tubules exhibit decreased ATP and reduced, but not absent, mitochondrial membrane potential (Δψm) during reoxygenation after severe hypoxia. This energetic deficit, which plays a pi...
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assessment of mitochondrial membrane potential in Proximal Tubules after hypoxia reoxygenation
American Journal of Physiology-renal Physiology, 2005Co-Authors: Thorsten Feldkamp, Andreas Kribben, Joel M WeinbergAbstract:Proximal Tubules develop a severe energetic deficit during hypoxia-reoxygenation (H/R) that previous studies using fluorescent potentiometric probes have suggested is characterized by sustained, pa...
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anaerobic and aerobic pathways for salvage of Proximal Tubules from hypoxia induced mitochondrial injury
American Journal of Physiology-renal Physiology, 2000Co-Authors: Joel M Weinberg, Manjeri A Venkatachalam, Nancy F Roeser, Pothana Saikumar, Zheng Dong, Ruth A Senter, Itzhak NissimAbstract:We have further examined the mechanisms for a severe mitochondrial energetic deficit, deenergization, and impaired respiration in complex I that develop in kidney Proximal Tubules during hypoxia-re...
Vladimir Marshansky - One of the best experts on this subject based on the ideXlab platform.
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Receptor-mediated endocytosis in kidney Proximal Tubules: recent advances and hypothesis.
Electrophoresis, 1997Co-Authors: Vladimir Marshansky, Sylvain G. Bourgoin, Irène Londono, Moise Bendayan, Bruno Maranda, Patrick VinayAbstract:Preparation of kidney Proximal Tubules in suspension allows the study of receptor-mediated endocytosis, protein reabsorption, and traffic of endosomal vesicles. The study of tubular protein transport in vitro coupled with that of the function of endosomal preparation offers a unique opportunity to investigate a receptor-mediated endocytosis pathway under physiological and pathological conditions. We assume that receptor-mediated endocytosis of albumin in kidney Proximal Tubules in situ and in vitro can be regulated, on the one hand, by the components of the acidification machinery (V-type H+-ATPase, Cl(-)-channel and Na+/H+-exchanger), giving rise to formation and dissipation of a proton gradient in endosomal vesicles, and, on the other hand, by small GTPases of the ADP-ribosylation factor (Arf)-family. In this paper we thus analyze the recent advances of the studies of cellular and molecular mechanisms underlying the identification, localization, and function of the acidification machinery (V-type H+-ATPase, Cl(-)-channel) as well as Arf-family small GTPases and phospholipase D in the endocytotic pathway of kidney Proximal Tubules. Also, we explore the possible functional interaction between the acidification machinery and Arf-family small GTPases. Finally, we propose the hypothesis of the regulation of translocation of Arf-family small GTPases by an endosomal acidification process and its role during receptor-mediated endocytosis in kidney Proximal Tubules. The results of this study will not only enhance our understanding of the receptor-mediated endocytosis pathway in kidney Proximal Tubules under physiological conditions but will also have important implications with respect to the functional consequences under some pathological circumstances. Furthermore, it may suggest novel targets and approaches in the prevention and treatment of various diseases (cystic fibrosis, Dent's disease, diabetes and autosomal dominant polycystic kidney disease).
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Identification of ADP-ribosylation factor-6 in brush-border membrane and early endosomes of human kidney Proximal Tubules
Electrophoresis, 1997Co-Authors: Vladimir Marshansky, Sylvain G. Bourgoin, Irène Londono, Moise Bendayan, Patrick VinayAbstract:The expression and distribution of ADP-ribosylation factor (ARF) small GTP-binding proteins in kidney tissue was examined. Various anti-ARF antibodies were raised against purified rec-ARF 1 and rec-ARF 6 and their specificity was determined. Using indirect immunofluorescence analysis of intact kidney, ARF proteins were found to be predominantly expressed in kidney Tubules as compared to glomeruli. This result was further supported by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis of purified human kidney glomeruli and Proximal Tubules. Both ARF 1 and ARF 6 were detected in purified human glomeruli and Proximal Tubules; however, ARF 1 was more abundant than ARF 6 in these kidney structures. Brush-border membrane vesicles (BBMV) and early endosomes (EE) derived from the receptor-mediated endocytosis pathway were isolated from purified Proximal Tubules of rat, dog and human kidney using a combination of magnesium precipitation and wheat-germ agglutinin negative selection techniques. We demonstrated that ARF 6 is associated with BBMV and with EE derived from receptor-mediated endocytosis pathway of human kidney Proximal Tubules. Using a combination of SDS-PAGE and quantitative enhanced chemiluminescence Western blot analysis, the quantification of the ARF 6 distribution in membrane and cytoplasmic fractions of Proximal Tubules was made and its predominance in membrane fractions was demonstrated. By analogy with the functional role of ARF 1 in Golgi protein transport, we suggest that ARF 6 may play an important role in the regulation of receptor-mediated endocytosis and protein reabsorption by kidney Proximal Tubules.
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Proton gradient formation in early endosomes from Proximal Tubules
Biochimica et biophysica acta, 1996Co-Authors: Vladimir Marshansky, Patrick VinayAbstract:Heavy endosomes were isolated from Proximal Tubules using a combination of magnesium precipitation and wheat-germ agglutinin negative selection techniques. Two small GTPases (Rab4 and Rab5) known to be specifically present in early endosomes were identified in our preparations. Endosomal acidification was followed fluorimetrically using acridine orange. In presence of chloride ions and ATP, the formation of a proton gradient (delta pH) was observed. This process is due to the activity of an electrogenic V-type ATPase present in the endosomal membrane since specific inhibitors bafilomycin and folimycin effectively prevented or eliminated endosomal acidification. In presence of chloride ions (K(m) = 30 mM) the formation of the proton gradient was optimal. Inhibitors of chloride channel activity such as DIDS and NPPB reduced acidification. The presence of sodium ions stimulated the dissipation of the proton gradient. This effect of sodium was abolished by amiloride derivative (MIA) but only when loaded into endosomes, indicating the presence of a physiologically oriented Na+/H(+)-exchanger in the endosomal membrane. Monensin restored the gradient dissipation. Thus three proteins (V-type ATPase, Cl(-)-channel, Na+/H(+)-exchanger) present in early endosomes isolated from Proximal Tubules may regulate the formation, maintenance and dissipation of the proton gradient.
Andreas Kribben - One of the best experts on this subject based on the ideXlab platform.
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Substrate Modulation of Fatty Acid Effects on Energization and Respiration of Kidney Proximal Tubules during Hypoxia/Reoxygenation
PloS one, 2014Co-Authors: Anja Bienholz, Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ahmad Al-taweel, Joel M WeinbergAbstract:Kidney Proximal Tubules subjected to hypoxia/reoxygenation develop a nonesterified fatty acid-induced energetic deficit characterized by persistent partial mitochondrial deenergization that can be prevented and reversed by citric acid cycle substrates. To further assess the role of competition between fatty acids and substrates on inner membrane substrate carriers in the deenergization and the contribution to deenergization of fatty acid effects on respiratory function, digitonin-permeabilized rabbit and mouse Tubules were studied using either addition of exogenous oleate after control normoxic incubation or increases of endogenous fatty acids produced by hypoxia/reoxygenation. The results demonstrated major effects of matrix oxaloacetate accumulation on succinate-supported energization and respiration and their modification by fatty acids. Improvements of energization in the presence of fatty acids by glutamate were shown to result predominantly from lowering matrix oxaloacetate rather than from amelioration of transmembrane cycling of fatty acids and uncoupling. Mouse Tubules had 2.5 fold higher rates of succinate utilization, which resulted in stronger effects of oxaloacetate accumulation than rabbit Tubules. Hypoxia/reoxygenation induced respiratory inhibition that was more severe for complex I-dependent substrates. Fatty acids themselves did not acutely contribute to this respiratory inhibition, but lowering them during 60 min. reoxygenation to allow recovery of ATP during that period alleviated it. These data clarify the basis for the nonesterified fatty acid-induced mitochondrial energetic deficit in kidney Proximal Tubules that impairs structural and functional recovery and provide insight into interactions that need to be considered in the design of substrate-based interventions to improve mitochondrial function.
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substrate modulation of fatty acid effects on energization and respiration of kidney Proximal Tubules during hypoxia reoxygenation
PLOS ONE, 2014Co-Authors: Anja Bienholz, Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ahmad Altaweel, Joel M WeinbergAbstract:Kidney Proximal Tubules subjected to hypoxia/reoxygenation develop a nonesterified fatty acid-induced energetic deficit characterized by persistent partial mitochondrial deenergization that can be prevented and reversed by citric acid cycle substrates. To further assess the role of competition between fatty acids and substrates on inner membrane substrate carriers in the deenergization and the contribution to deenergization of fatty acid effects on respiratory function, digitonin-permeabilized rabbit and mouse Tubules were studied using either addition of exogenous oleate after control normoxic incubation or increases of endogenous fatty acids produced by hypoxia/reoxygenation. The results demonstrated major effects of matrix oxaloacetate accumulation on succinate-supported energization and respiration and their modification by fatty acids. Improvements of energization in the presence of fatty acids by glutamate were shown to result predominantly from lowering matrix oxaloacetate rather than from amelioration of transmembrane cycling of fatty acids and uncoupling. Mouse Tubules had 2.5 fold higher rates of succinate utilization, which resulted in stronger effects of oxaloacetate accumulation than rabbit Tubules. Hypoxia/reoxygenation induced respiratory inhibition that was more severe for complex I-dependent substrates. Fatty acids themselves did not acutely contribute to this respiratory inhibition, but lowering them during 60 min. reoxygenation to allow recovery of ATP during that period alleviated it. These data clarify the basis for the nonesterified fatty acid-induced mitochondrial energetic deficit in kidney Proximal Tubules that impairs structural and functional recovery and provide insight into interactions that need to be considered in the design of substrate-based interventions to improve mitochondrial function.
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N-cadherin is depleted from Proximal Tubules in experimental and human acute kidney injury
Histochemistry and Cell Biology, 2010Co-Authors: Jens Nürnberger, Markus Hörbelt, Thorsten Feldkamp, Rosmaria Kavapurackal, Anabelle Opazo Saez, Jan Becker, Andreas KribbenAbstract:Ischemia remains the most common cause of acute kidney injury (AKI). Decreased intercellular adhesion and alterations in adhesion molecules may contribute to the loss of renal function observed in AKI. In the present study, we evaluated the distribution of adhesion molecules in the human kidney and analyzed their expression in human and experimental AKI. Specimens of human kidneys obtained from patients with and without AKI were stained for the cell adhesion molecules E-cadherin, N-cadherin and β-catenin. Experimental AKI in rats was induced by renal artery clamping. Immunostaining and immunoblotting were carried out for E-cadherin, N-cadherin and β-catenin. Proximal tubule cells from opossum kidneys (OKs) were used to analyze the effect of chemical hypoxia (ATP depletion) in vitro. In the adult human kidney, N-cadherin was expressed in Proximal Tubules, while E-cadherin was expressed in other nephron segments. β-Catenin was expressed in both Proximal and distal Tubules. In human AKI and in ischemic rat kidneys, N-cadherin immunostaining was depleted from Proximal Tubules. There was no change in E-cadherin or β-catenin. In vitro, OK cells expressed N-cadherin only in the presence of collagen, and ATP depletion led to a depletion of N-cadherin. Collagen IV staining was reduced in ischemic rat kidneys compared to controls. The results of the study suggest that N-cadherin may play a significant role in human and experimental AKI.
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Digital fluorescence imaging of organic cation transport in freshly isolated rat Proximal Tubules.
Drug Metabolism and Disposition, 2005Co-Authors: Frank Pietruck, Katrin Engeln, Markus Hörbelt, Stefan Herget-rosenthal, Thomas Philipp, Thorsten Feldkamp, Andreas KribbenAbstract:The secretion of cationic drugs and endogenous metabolites is a major function of the kidney. This is accomplished by organic cation transport systems, mainly located in the Proximal Tubules. Here, we describe a model for continuous measurement of organic cation (OC) transport. In this model, organic cation transport in individual freshly isolated rat Proximal Tubules is investigated by use of digital fluorescence imaging. To directly measure organic cation transport across the basolateral membrane, the fluorescent organic cation 4-(4-dimethylaminostyryl)- N -methylpyridinium (ASP + ) is used with a customized perfusion chamber. ASP + uptake in this model displayed the characteristics of organic cation transport. Over the tested range of 1 to 50 μM, it showed a concentration-dependent uptake across the basolateral membrane. In the presence of competitive inhibitors of OC transport such as N 1 -methylnicotinamide + , tetraethylammonium + , and choline + , a concentration-dependent and reversible inhibition of ASP + uptake could be documented. In conclusion, continuous measurement of organic cation transport in freshly isolated rat Proximal Tubules by digital fluorescence imaging using ASP + is a useful tool for investigation of drug transport and interactions and, furthermore, may be helpful for investigation of organic cation transport under pathophysiological conditions.
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Accumulation of nonesterified fatty acids causes the sustained energetic deficit in kidney Proximal Tubules after hypoxia-reoxygenation.
American journal of physiology. Renal physiology, 2005Co-Authors: Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ruth A Senter, Joel M WeinbergAbstract:Kidney Proximal Tubules exhibit decreased ATP and reduced, but not absent, mitochondrial membrane potential (Δψm) during reoxygenation after severe hypoxia. This energetic deficit, which plays a pi...
Thorsten Feldkamp - One of the best experts on this subject based on the ideXlab platform.
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Substrate Modulation of Fatty Acid Effects on Energization and Respiration of Kidney Proximal Tubules during Hypoxia/Reoxygenation
PloS one, 2014Co-Authors: Anja Bienholz, Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ahmad Al-taweel, Joel M WeinbergAbstract:Kidney Proximal Tubules subjected to hypoxia/reoxygenation develop a nonesterified fatty acid-induced energetic deficit characterized by persistent partial mitochondrial deenergization that can be prevented and reversed by citric acid cycle substrates. To further assess the role of competition between fatty acids and substrates on inner membrane substrate carriers in the deenergization and the contribution to deenergization of fatty acid effects on respiratory function, digitonin-permeabilized rabbit and mouse Tubules were studied using either addition of exogenous oleate after control normoxic incubation or increases of endogenous fatty acids produced by hypoxia/reoxygenation. The results demonstrated major effects of matrix oxaloacetate accumulation on succinate-supported energization and respiration and their modification by fatty acids. Improvements of energization in the presence of fatty acids by glutamate were shown to result predominantly from lowering matrix oxaloacetate rather than from amelioration of transmembrane cycling of fatty acids and uncoupling. Mouse Tubules had 2.5 fold higher rates of succinate utilization, which resulted in stronger effects of oxaloacetate accumulation than rabbit Tubules. Hypoxia/reoxygenation induced respiratory inhibition that was more severe for complex I-dependent substrates. Fatty acids themselves did not acutely contribute to this respiratory inhibition, but lowering them during 60 min. reoxygenation to allow recovery of ATP during that period alleviated it. These data clarify the basis for the nonesterified fatty acid-induced mitochondrial energetic deficit in kidney Proximal Tubules that impairs structural and functional recovery and provide insight into interactions that need to be considered in the design of substrate-based interventions to improve mitochondrial function.
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substrate modulation of fatty acid effects on energization and respiration of kidney Proximal Tubules during hypoxia reoxygenation
PLOS ONE, 2014Co-Authors: Anja Bienholz, Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ahmad Altaweel, Joel M WeinbergAbstract:Kidney Proximal Tubules subjected to hypoxia/reoxygenation develop a nonesterified fatty acid-induced energetic deficit characterized by persistent partial mitochondrial deenergization that can be prevented and reversed by citric acid cycle substrates. To further assess the role of competition between fatty acids and substrates on inner membrane substrate carriers in the deenergization and the contribution to deenergization of fatty acid effects on respiratory function, digitonin-permeabilized rabbit and mouse Tubules were studied using either addition of exogenous oleate after control normoxic incubation or increases of endogenous fatty acids produced by hypoxia/reoxygenation. The results demonstrated major effects of matrix oxaloacetate accumulation on succinate-supported energization and respiration and their modification by fatty acids. Improvements of energization in the presence of fatty acids by glutamate were shown to result predominantly from lowering matrix oxaloacetate rather than from amelioration of transmembrane cycling of fatty acids and uncoupling. Mouse Tubules had 2.5 fold higher rates of succinate utilization, which resulted in stronger effects of oxaloacetate accumulation than rabbit Tubules. Hypoxia/reoxygenation induced respiratory inhibition that was more severe for complex I-dependent substrates. Fatty acids themselves did not acutely contribute to this respiratory inhibition, but lowering them during 60 min. reoxygenation to allow recovery of ATP during that period alleviated it. These data clarify the basis for the nonesterified fatty acid-induced mitochondrial energetic deficit in kidney Proximal Tubules that impairs structural and functional recovery and provide insight into interactions that need to be considered in the design of substrate-based interventions to improve mitochondrial function.
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N-cadherin is depleted from Proximal Tubules in experimental and human acute kidney injury
Histochemistry and Cell Biology, 2010Co-Authors: Jens Nürnberger, Markus Hörbelt, Thorsten Feldkamp, Rosmaria Kavapurackal, Anabelle Opazo Saez, Jan Becker, Andreas KribbenAbstract:Ischemia remains the most common cause of acute kidney injury (AKI). Decreased intercellular adhesion and alterations in adhesion molecules may contribute to the loss of renal function observed in AKI. In the present study, we evaluated the distribution of adhesion molecules in the human kidney and analyzed their expression in human and experimental AKI. Specimens of human kidneys obtained from patients with and without AKI were stained for the cell adhesion molecules E-cadherin, N-cadherin and β-catenin. Experimental AKI in rats was induced by renal artery clamping. Immunostaining and immunoblotting were carried out for E-cadherin, N-cadherin and β-catenin. Proximal tubule cells from opossum kidneys (OKs) were used to analyze the effect of chemical hypoxia (ATP depletion) in vitro. In the adult human kidney, N-cadherin was expressed in Proximal Tubules, while E-cadherin was expressed in other nephron segments. β-Catenin was expressed in both Proximal and distal Tubules. In human AKI and in ischemic rat kidneys, N-cadherin immunostaining was depleted from Proximal Tubules. There was no change in E-cadherin or β-catenin. In vitro, OK cells expressed N-cadherin only in the presence of collagen, and ATP depletion led to a depletion of N-cadherin. Collagen IV staining was reduced in ischemic rat kidneys compared to controls. The results of the study suggest that N-cadherin may play a significant role in human and experimental AKI.
-
Digital fluorescence imaging of organic cation transport in freshly isolated rat Proximal Tubules.
Drug Metabolism and Disposition, 2005Co-Authors: Frank Pietruck, Katrin Engeln, Markus Hörbelt, Stefan Herget-rosenthal, Thomas Philipp, Thorsten Feldkamp, Andreas KribbenAbstract:The secretion of cationic drugs and endogenous metabolites is a major function of the kidney. This is accomplished by organic cation transport systems, mainly located in the Proximal Tubules. Here, we describe a model for continuous measurement of organic cation (OC) transport. In this model, organic cation transport in individual freshly isolated rat Proximal Tubules is investigated by use of digital fluorescence imaging. To directly measure organic cation transport across the basolateral membrane, the fluorescent organic cation 4-(4-dimethylaminostyryl)- N -methylpyridinium (ASP + ) is used with a customized perfusion chamber. ASP + uptake in this model displayed the characteristics of organic cation transport. Over the tested range of 1 to 50 μM, it showed a concentration-dependent uptake across the basolateral membrane. In the presence of competitive inhibitors of OC transport such as N 1 -methylnicotinamide + , tetraethylammonium + , and choline + , a concentration-dependent and reversible inhibition of ASP + uptake could be documented. In conclusion, continuous measurement of organic cation transport in freshly isolated rat Proximal Tubules by digital fluorescence imaging using ASP + is a useful tool for investigation of drug transport and interactions and, furthermore, may be helpful for investigation of organic cation transport under pathophysiological conditions.
-
Accumulation of nonesterified fatty acids causes the sustained energetic deficit in kidney Proximal Tubules after hypoxia-reoxygenation.
American journal of physiology. Renal physiology, 2005Co-Authors: Thorsten Feldkamp, Andreas Kribben, Nancy F Roeser, Ruth A Senter, Joel M WeinbergAbstract:Kidney Proximal Tubules exhibit decreased ATP and reduced, but not absent, mitochondrial membrane potential (Δψm) during reoxygenation after severe hypoxia. This energetic deficit, which plays a pi...
