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5. Redesign techniques form nonlinear sampled-data systems

Author

Gruene, L, Pannek, J, Worthmann, K, Nesic, D, Gruene, L, Pannek, J, Worthmann, K, and Nesic, D

Abstract

In the emulation approach to controller design for networked control systems the controller is first designed in continuous time ignoring the network and then implemented as a sampled-data controller. While very attractive for its simplicity, typically sufficiently small sampling periods are needed in order to ensure satisfactory performance of the resulting sampled-data closed loop. Thus, in the presence of network bandwidth constraints performance loss up to instability may occur. In this paper we present a variety of analytical and numerical techniques for the redesign of sampled-data controllers which improve the sampled-data performance of the non-redesigned controller and aim at reducing the necessary communication bandwidth.

Published
2008

6. Continuous-time controller redesign for digital implementation: A trajectory based approach

Author

Gruene, L, Worthmann, K, Nesic, D, Gruene, L, Worthmann, K, and Nesic, D

Abstract

Given a continuous-time nonlinear closed loop system, we investigate sampled-data feedback laws for which the trajectories of the sampled-data closed loop system converge to the continuous-time trajectories with a prescribed rate of convergence as the length of the sampling interval tends to zero. We derive necessary and sufficient conditions for the existence of such sampled-data feedback laws and—in case of existence—provide explicit redesign formulas and algorithms for these controllers.

Published
2008

7. High order approximations by sampled-data feedback

Author

Grune, L, Worthmann, K, NESIC, D, Grune, L, Worthmann, K, and NESIC, D

Abstract

Given a continuous time nonlinear closed loop system, we give explicit constructions for sampled–data feedback laws for which the trajectories of the sampled– data closed loop system converge to the continuous time trajectories with a prescribed rate of convergence as the sampling interval length tends to zero. In particular, we investigate necessary and sufficient conditions under which such sampled–data feedbacks exist. We give analytic solutions to the problem for local orders of convergence _ 4 and present a MAPLE code for general orders.

Published
2006

8. Experimental pathology

Author

Yi Chun, D. X., primary, Alexandre, H., additional, Edith, B., additional, Nacera, O., additional, Julie, P., additional, Chantal, J., additional, Eric, R., additional, Zhang, X., additional, Jin, Y., additional, Miravete, M., additional, Dissard, R., additional, Klein, J., additional, Gonzalez, J., additional, Caubet, C., additional, Pecher, C., additional, Pipy, B., additional, Bascands, J.-L., additional, Mercier-Bonin, M., additional, Schanstra, J., additional, Buffin-Meyer, B., additional, Claire, R., additional, Rigothier, C., additional, Richard, D., additional, Sebastien, L., additional, Moin, S., additional, Chantal, B., additional, Christian, C., additional, Jean, R., additional, Migliori, M., additional, Cantaluppi, V., additional, Mannari, C., additional, Medica, D., additional, Giovannini, L., additional, Panichi, V., additional, Goldwich, A., additional, Alexander, S., additional, Andre, G., additional, Amann, K., additional, Migliorini, A., additional, Sagrinati, C., additional, Angelotti, M. L., additional, Mulay, S. R., additional, Ronconi, E., additional, Peired, A., additional, Romagnani, P., additional, Anders, H.-J., additional, Chiang, W. C., additional, Lai, C. F., additional, Peng, W.-H., additional, Wu, C. F., additional, Chang, F.-C., additional, Chen, Y.-T., additional, Lin, S.-L., additional, Chen, Y. M., additional, Wu, K. D., additional, Lu, K.-S., additional, Tsai, T. J., additional, Virgine, O., additional, Qing Feng, F., additional, Zhang, S.-Y., additional, Dominique, D., additional, Vincent, A., additional, Marina, C., additional, Philippe, L., additional, Georges, G., additional, Pawlak, A., additional, Sahali, D., additional, Matsumoto, S., additional, Kiyomoto, H., additional, Ichimura, A., additional, Dan, T., additional, Nakamichi, T., additional, Tsujita, T., additional, Akahori, K., additional, Ito, S., additional, Miyata, T., additional, Xie, S., additional, Zhang, B., additional, Shi, W., additional, Yang, Y., additional, Nagasu, H., additional, Satoh, M., additional, Kidokoro, K., additional, Nishi, Y., additional, Ihoriya, C., additional, Kadoya, H., additional, Sasaki, T., additional, Kashihara, N., additional, Wu, C.-F., additional, Chou, Y.-H., additional, Duffield, J., additional, Rocca, C., additional, Gregorini, M., additional, Corradetti, V., additional, Valsania, T., additional, Bedino, G., additional, Bosio, F., additional, Pattonieri, E. F., additional, Esposito, P., additional, Sepe, V., additional, Libetta, C., additional, Rampino, T., additional, Dal Canton, A., additional, Omori, H., additional, Kawada, N., additional, Inoue, K., additional, Ueda, Y., additional, Yamamoto, R., additional, Matsui, I., additional, Kaimori, J., additional, Takabatake, Y., additional, Moriyama, T., additional, Isaka, Y., additional, Rakugi, H., additional, Wasilewska, A., additional, Taranta-Janusz, K., additional, Deebek, W., additional, Kuroczycka-Saniutycz, E., additional, Lee, A. S., additional, Lee, J. E., additional, Jung, Y. J., additional, Kang, K. P., additional, Lee, S., additional, Kim, W., additional, Arfian, N., additional, Emoto, N., additional, Yagi, K., additional, Nakayama, K., additional, Hartopo, A. B., additional, Nugrahaningsih, D. A., additional, Yanagisawa, M., additional, Hirata, K.-I., additional, Munoz-Felix, J. M., additional, Lopez-Novoa, J. M., additional, Martinez-Salgado, C., additional, Oujo, B., additional, Arevalo, M., additional, Bernabeu, C., additional, Perez-Barriocanal, F., additional, Jesper, K., additional, Nathalie, V., additional, Pierre, G., additional, Yi Chun, D. X., additional, Iyoda, M., additional, Shibata, T., additional, Matsumoto, K., additional, Shindo-Hirai, Y., additional, Kuno, Y., additional, Wada, Y., additional, Akizawa, T., additional, Schwartz, I., additional, Schwartz, D., additional, Prot Bertoye, C., additional, Terryn, S., additional, Claver, J., additional, Beghdadi, W. B., additional, Monteiro, R., additional, Blank, U., additional, Devuyst, O., additional, Daugas, E., additional, Van Beneden, K., additional, Geers, C., additional, Pauwels, M., additional, Mannaerts, I., additional, Van den Branden, C., additional, Van Grunsven, L. A., additional, Seckin, I., additional, Pekpak, M., additional, Uzunalan, M., additional, Uruluer, B., additional, Kokturk, S., additional, Ozturk, Z., additional, Sonmez, H., additional, Yaprak, E., additional, Furuno, Y., additional, Tsutsui, M., additional, Morishita, T., additional, Shimokawa, H., additional, Otsuji, Y., additional, Yanagihara, N., additional, Kabashima, N., additional, Ryota, S., additional, Kanegae, K., additional, Miyamoto, T., additional, Nakamata, J., additional, Ishimatsu, N., additional, Tamura, M., additional, Nakagawa, T., additional, Ichikawa, K., additional, Miyamoto, M., additional, Takabayashi, D., additional, Yamazaki, H., additional, Kakeshita, K., additional, Koike, T., additional, Kagitani, S., additional, Tomoda, F., additional, Hamashima, T., additional, Ishii, Y., additional, Inoue, H., additional, Sasahara, M., additional, El Machhour, F., additional, Kerroch, M., additional, Mesnard, L., additional, Chatziantoniou, C., additional, Dussaule, J.-C., additional, Inui, K., additional, Sasai, F., additional, Maruta, Y., additional, Nishiwaki, H., additional, Kawashima, E., additional, Inoue, Y., additional, Yoshimura, A., additional, Musacchio, E., additional, Priante, G., additional, Valvason, C., additional, Sartori, L., additional, Baggio, B., additional, Kim, J. H., additional, Gross, O., additional, Diana, R., additional, Gry, D. H., additional, Asimal, B., additional, Johanna, T., additional, Imke, S.-E., additional, Lydia, W., additional, Gerhard-Anton, M., additional, Hassan, D., additional, Cano, J. L., additional, Griera, M., additional, Olmos, G., additional, Martin, P., additional, Cortes, M. A., additional, Lopez-Ongil, S., additional, Rodriguez-Puyol, D., additional, DE Frutos, S., additional, Gonzalez, M., additional, Luengo, A., additional, Rodriguez-Puyol, M., additional, Calleros, L., additional, Lupica, R., additional, Lacquaniti, A., additional, Donato, V., additional, Maggio, R., additional, Mastroeni, C., additional, Lucisano, S., additional, Cernaro, V., additional, Fazio, M. R., additional, Quartarone, A., additional, Buemi, M., additional, Kacik, M., additional, Goedicke, S., additional, Eggert, H., additional, Hoyer, J. D., additional, Wurm, S., additional, Steege, A., additional, Banas, M., additional, Kurtz, A., additional, Banas, B., additional, Lasagni, L., additional, Lazzeri, E., additional, Romoli, S., additional, Schaefer, I., additional, Teng, B., additional, Worthmann, K., additional, Haller, H., additional, Schiffer, M., additional, Prattichizzo, C., additional, Netti, G. S., additional, Rocchetti, M. T., additional, Cormio, L., additional, Carrieri, G., additional, Stallone, G., additional, Grandaliano, G., additional, Ranieri, E., additional, Gesualdo, L., additional, Kucher, A., additional, Smirnov, A., additional, Parastayeva, M., additional, Beresneva, O., additional, Kayukov, I., additional, Zubina, I., additional, Ivanova, G., additional, Abed, A., additional, Schlekenbach, L., additional, Foglia, B., additional, Kwak, B., additional, Chadjichristos, C., additional, Queisser, N., additional, Schupp, N., additional, Brand, S., additional, Himer, L., additional, Szebeni, B., additional, Sziksz, E., additional, Saijo, S., additional, Kis, E., additional, Prokai, A., additional, Banki, N. F., additional, Fekete, A., additional, Tulassay, T., additional, Vannay, A., additional, Hegner, B., additional, Schaub, T., additional, Lange, C., additional, Dragun, D., additional, Klinkhammer, B. M., additional, Rafael, K., additional, Monika, M., additional, Anna, M., additional, Van Roeyen, C., additional, Boor, P., additional, Eva Bettina, B., additional, Simon, O., additional, Esther, S., additional, Floege, J., additional, Kunter, U., additional, Janke, D., additional, Jankowski, J., additional, Hayashi, M., additional, Takamatsu, I., additional, Horimai, C., additional, Yoshida, T., additional, Seno DI Marco, G., additional, Koenig, M., additional, Stock, C., additional, Reiermann, S., additional, Amler, S., additional, Koehler, G., additional, Fobker, M., additional, Buck, F., additional, Pavenstaedt, H., additional, Lang, D., additional, Brand, M., additional, Plotnikov, E., additional, Morosanova, M., additional, Pevzner, I., additional, Zorova, L., additional, Pulkova, N., additional, Zorov, D., additional, Wornle, M., additional, Ribeiro, A., additional, Belling, F., additional, Merkle, M., additional, Nakazawa, D., additional, Nishio, S., additional, Shibasaki, S., additional, Tomaru, U., additional, Akihiro, I., additional, Kobayashi, I., additional, Imanishi, Y., additional, Kurajoh, M., additional, Nagata, Y., additional, Yamagata, M., additional, Emoto, M., additional, Michigami, T., additional, Ishimura, E., additional, Inaba, M., additional, Wu, C.-C., additional, Lu, K.-C., additional, Chen, J.-S., additional, Chu, P., additional, Lin, Y.-F., additional, Eller, K., additional, Schroll, A., additional, Kirsch, A., additional, Huber, J., additional, Weiss, G., additional, Theurl, I., additional, Rosenkranz, A. R., additional, Zawada, A., additional, Rogacev, K., additional, Achenbach, M., additional, Fliser, D., additional, Held, G., additional, Heine, G. H., additional, Miyamoto, Y., additional, Iwao, Y., additional, Watanabe, H., additional, Kadowaki, D., additional, Ishima, Y., additional, Chuang, V. T. G., additional, Sato, K., additional, Otagiri, M., additional, Maruyama, T., additional, Iwatani, H., additional, Honda, D., additional, Noguchi, T., additional, Tanaka, M., additional, Tanaka, H., additional, Fukagawa, M., additional, Pircher, J., additional, Koppel, S., additional, Mannell, H., additional, Krotz, F., additional, Virzi, G. M., additional, Bolin, C., additional, Cruz, D., additional, Scalzotto, E., additional, De Cal, M., additional, Vescovo, G., additional, Ronco, C., additional, Grobmayr, R., additional, Lech, M., additional, Ryu, M., additional, Aoshima, Y., additional, Mizobuchi, M., additional, Ogata, H., additional, Kumata, C., additional, Nakazawa, A., additional, Kondo, F., additional, Ono, N., additional, Koiwa, F., additional, Kinugasa, E., additional, Freisinger, W., additional, Lale, N., additional, Lampert, A., additional, Ditting, T., additional, Heinlein, S., additional, Schmieder, R. E., additional, Veelken, R., additional, Nave, H., additional, Perthel, R., additional, Suntharalingam, M., additional, Bode-Boger, S., additional, Beutel, G., additional, Kielstein, J., additional, Rodrigues-Diez, R., additional, Rayego-Mateos, S., additional, Lavoz, C., additional, Stark Aroeira, L. G., additional, Orejudo, M., additional, Alique, M., additional, Ortiz, A., additional, Egido, J., additional, Ruiz-Ortega, M., additional, Oskar, W., additional, Rusan, C., additional, Padberg, J.-S., additional, Wiesinger, A., additional, Reuter, S., additional, Grabner, A., additional, Kentrup, D., additional, Lukasz, A., additional, Oberleithner, H., additional, Pavenstadt, H., additional, Kumpers, P., additional, Eberhardt, H. U., additional, Skerka, C., additional, Chen, Q., additional, Hallstroem, T., additional, Hartmann, A., additional, Kemper, M. J., additional, Zipfel, P. F., additional, N'gome-Sendeyo, K., additional, Fan, Q.-F., additional, Toblli, J., additional, Cao, G., additional, Giani, J. F., additional, Dominici, F. P., additional, Kim, J. S., additional, Yang, J. W., additional, Kim, M. K., additional, Han, B. G., additional, and Choi, S. O., additional

Published
2012
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9. Mechanisms and targets of glomerular damage

Author

Rayego-Mateos, S., primary, Rodrigues-Diez, R., additional, Rodrigues-Diez, R. R., additional, Lavoz-Barria, C., additional, Alique, M., additional, Mas, S., additional, Pato, J., additional, Keri, G., additional, Egido, J., additional, Ortiz, A., additional, Ruiz-Ortega, M., additional, Ying, L., additional, Tepel, M., additional, Frank, E., additional, Florian, T., additional, Gregor, T., additional, Boye, J., additional, Maik, G., additional, Teng, B., additional, Gu, C., additional, Haller, H., additional, Sever, S., additional, Schiffer, M., additional, Worthmann, K., additional, Leitges, M., additional, Dittrich-Breiholz, O., additional, Kracht, M., additional, Peired, A., additional, Angelotti, M. L., additional, Ronconi, E., additional, Lazzeri, E., additional, Sisti, A., additional, Lasagni, L., additional, and Romagnani, P., additional

Published
2012
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10. Model Predictive Temperature Control for Retinal Laser Treatments.

Author

Kleyman V, Eggert S, Schmidt C, Schaller M, Worthmann K, Brinkmann R, and Müller MA

Subjects
Animals, Swine, Laser Coagulation methods, Temperature, Photoacoustic Techniques methods, Laser Therapy methods, Retina surgery, Retina radiation effects
Abstract

Purpose: Manual, individual adjustment of the laser power in retinal laser therapies is time-consuming, is inaccurate with respect to uniform effects, and can only prevent over- or undertreatment to a limited extent. Automatic closed-loop temperature control allows for similar temperatures at each irradiated spot despite varying absorption. This is of crucial importance for subdamaging hyperthermal treatments with no visible effects and the safety of photocoagulation with short irradiation times. The aim of this work is to perform extensive experiments on porcine eye explants to demonstrate the benefits of automatic control in retinal laser treatments., Methods: To ensure a safe and reliable temperature rise, we utilize a model predictive controller. For model predictive control, the current state and the spot-dependent absorption coefficients are estimated by an extended Kalman filter (EKF). Therein, optoacoustic measurements are used to determine the temperature rise at the irradiated areas in real time. We use fluorescence vitality stains to measure the lesion size and validate the proposed control strategy., Results: By comparing the lesion size with temperature values for cell death, we found that the EKF accurately estimates the peak temperature. Furthermore, the proposed closed-loop control scheme works reliably with regard to similar lesion sizes despite varying absorption with a smaller spread in lesion size compared to open-loop control., Conclusions: Our closed-loop control approach enables a safe subdamaging treatment and lowers the risk for over- and undertreatment for mild coagulations in retinal laser therapies., Translational Relevance: We demonstrate that modern control strategies have the potential to improve retinal laser treatments for several diseases.

Published
2024
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11. Tyrosine Phosphorylation of CD2AP Affects Stability of the Slit Diaphragm Complex.

Author

Tossidou I, Teng B, Worthmann K, Müller-Deile J, Jobst-Schwan T, Kardinal C, Schroder P, Bolanos-Palmieri P, Haller H, Willerding J, Drost DM, de Jonge L, Reubold T, Eschenburg S, Johnson RI, and Schiffer M

Subjects
Animals, Drosophila melanogaster, HEK293 Cells, Humans, Membrane Proteins metabolism, Phosphorylation, Podocytes metabolism, Protein Stability, Vascular Endothelial Growth Factor A pharmacology, Zebrafish, Adaptor Proteins, Signal Transducing metabolism, Cytoskeletal Proteins metabolism, Membrane Proteins chemistry, Tyrosine metabolism
Abstract

Background: CD2-associated protein (CD2AP), a slit diaphragm-associated scaffolding protein involved in survival and regulation of the cytoskeleton in podocytes, is considered a "stabilizer" of the slit diaphragm complex that connects the slit diaphragm protein nephrin to the cytoskeleton of the cell. Tyrosine phosphorylation of slit diaphragm molecules can influence their surface expression, but it is unknown whether tyrosine phosphorylation events of CD2AP are also physiologically relevant to slit diaphragm stability., Methods: We used isoelectric focusing, western blot analysis, and immunofluorescence to investigate phosphorylation of CD2AP, and phospho-CD2AP antibodies and site-directed mutagenesis to define the specific phosphorylated tyrosine residues. We used cross-species rescue experiments in Cd2ap KD zebrafish and in Drosophila cindr RNAi mutants to define the physiologic relevance of CD2AP phosphorylation of the tyrosine residues., Results: We found that VEGF-A stimulation can induce a tyrosine phosphorylation response in CD2AP in podocytes, and that these phosphorylation events have an important effect on slit diaphragm protein localization and functionality in vivo . We demonstrated that tyrosine in position Y10 of the SH3-1 domain of CD2AP is indispensable for CD2AP function in vivo . We found that the binding affinity of nephrin to CD2AP is significantly enhanced in the absence of Y10; however, unexpectedly, this increased affinity leads not to stabilization but to functional impairment of the glomerular filtration barrier., Conclusions: Our findings provide insight into CD2AP and its phosphorylation in the context of slit diaphragm functionality, and indicate a fine-tuned affinity balance of CD2AP and nephrin that is influenced by receptor tyrosine kinase stimulation., (Copyright © 2019 by the American Society of Nephrology.)

Published
2019
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12. Def-6, a novel regulator of small GTPases in podocytes, acts downstream of atypical protein kinase C (aPKC) λ/ι.

Author

Worthmann K, Leitges M, Teng B, Sestu M, Tossidou I, Samson T, Haller H, Huber TB, and Schiffer M

Subjects
Actin Cytoskeleton genetics, Animals, Cell Membrane genetics, Cells, Cultured, DNA-Binding Proteins genetics, GTP Phosphohydrolases genetics, GTP Phosphohydrolases metabolism, Guanine Nucleotide Exchange Factors, Isoenzymes genetics, Mice, Mice, Knockout, Nuclear Proteins genetics, Podocytes cytology, Protein Kinase C genetics, Protein Kinase C-epsilon genetics, Protein Kinase C-epsilon metabolism, Actin Cytoskeleton metabolism, Cell Membrane metabolism, DNA-Binding Proteins metabolism, Gene Expression Regulation physiology, Isoenzymes metabolism, Nuclear Proteins metabolism, Podocytes metabolism, Protein Kinase C metabolism
Abstract

The atypical protein kinase C (aPKC) isotypes PKCλ/ι and PKCζ are both expressed in podocytes; however, little is known about differences in their function. Previous studies in mice have demonstrated that podocyte-specific loss of PKCλ/ι leads to a severe glomerular phenotype, whereas mice deficient in PKCζ develop no renal phenotype. We analyzed various effects caused by PKCλ/ι and PKCζ deficiency in cultured murine podocytes. In contrast to PKCζ-deficient podocytes, PKCλ/ι-deficient podocytes exhibited a severe actin cytoskeletal phenotype, reduced cell size, decreased number of focal adhesions, and increased activation of small GTPases. Comparative microarray analysis revealed that the guanine nucleotide exchange factor Def-6 was specifically up-regulated in PKCλ/ι-deficient podocytes. In vivo Def-6 expression is significantly increased in podocytes of PKCλ/ι-deficient mice. Cultured PKCλ/ι-deficient podocytes exhibited an enhanced membrane association of Def-6, indicating enhanced activation. Overexpression of aPKCλ/ι in PKCλ/ι-deficient podocytes could reduce the membrane-associated expression of Def-6 and rescue the actin phenotype. In the present study, PKCλ/ι was identified as an important factor for actin cytoskeletal regulation in podocytes and Def-6 as a specific downstream target of PKCλ/ι that regulates the activity of small GTPases and subsequently the actin cytoskeleton of podocytes., (Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Published
2013
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13. aPKCλ/ι and aPKCζ contribute to podocyte differentiation and glomerular maturation.

Author

Hartleben B, Widmeier E, Suhm M, Worthmann K, Schell C, Helmstädter M, Wiech T, Walz G, Leitges M, Schiffer M, and Huber TB

Subjects
Animals, Capillaries metabolism, Capillaries pathology, Capillaries ultrastructure, Cell Differentiation physiology, Centrosome enzymology, Centrosome pathology, Centrosome ultrastructure, Female, Golgi Apparatus enzymology, Golgi Apparatus pathology, Golgi Apparatus ultrastructure, Isoenzymes genetics, Kidney Glomerulus blood supply, Kidney Glomerulus cytology, Kidney Glomerulus enzymology, Male, Mice, Mice, Knockout, Microscopy, Electron, Podocytes ultrastructure, Protein Kinase C genetics, Proteinuria metabolism, Proteinuria pathology, Signal Transduction physiology, Tight Junctions enzymology, Tight Junctions pathology, Tight Junctions ultrastructure, Isoenzymes metabolism, Podocytes enzymology, Podocytes pathology, Protein Kinase C metabolism, Proteinuria physiopathology
Abstract

Precise positioning of the highly complex interdigitating podocyte foot processes is critical to form the normal glomerular filtration barrier, but the molecular programs driving this process are unknown. The protein atypical protein kinase C (aPKC)--a component of the Par complex, which localizes to tight junctions and interacts with slit diaphragm proteins--may play a role. Here, we found that the combined deletion of the aPKCλ/ι and aPKCζ isoforms in podocytes associated with incorrectly positioned centrosomes and Golgi apparatus and mislocalized molecules of the slit diaphragm. Furthermore, aPKC-deficient podocytes failed to form the normal network of foot processes, leading to defective glomerular maturation with incomplete capillary formation and mesangiolysis. Our results suggest that aPKC isoforms orchestrate the formation of the podocyte processes essential for normal glomerular development and kidney function. Defective aPKC signaling results in a dramatically simplified glomerular architecture, causing severe proteinuria and perinatal death.

Published
2013
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14. Deficits in sialylation impair podocyte maturation.

Author

Weinhold B, Sellmeier M, Schaper W, Blume L, Philippens B, Kats E, Bernard U, Galuska SP, Geyer H, Geyer R, Worthmann K, Schiffer M, Groos S, Gerardy-Schahn R, and Münster-Kühnel AK

Subjects
Animals, Cell Nucleus metabolism, Gene Knock-In Techniques, Mice, Mice, Inbred C57BL, N-Acylneuraminate Cytidylyltransferase genetics, Phenotype, Podocytes ultrastructure, Sialoglycoproteins metabolism, Glomerular Filtration Barrier embryology, Membrane Proteins metabolism, N-Acetylneuraminic Acid metabolism, N-Acylneuraminate Cytidylyltransferase metabolism, Podocytes physiology
Abstract

The role of sialylation in kidney biology is not fully understood. The synthesis of sialoglycoconjugates, which form the outermost structures of animal cells, requires CMP-sialic acid, which is a product of the nuclear enzyme CMAS. We used a knock-in strategy to create a mouse with point mutations in the canonical nuclear localization signal of CMAS, which relocated the enzyme to the cytoplasm of transfected cells without affecting its activity. Although insufficient to prevent nuclear entry in mice, the mutation led to a drastically reduced concentration of nuclear-expressed enzyme. Mice homozygous for the mutation died from kidney failure within 72 hours after birth. The Cmas(nls) mouse exhibited podocyte foot process effacement, absence of slit diaphragms, and massive proteinuria, recapitulating features of nephrin-knockout mice and of patients with Finnish-type congenital nephrotic syndrome. Although the Cmas(nls) mouse displayed normal sialylation in all organs including kidney, a critical shortage of CMP-sialic acid prevented sialylation of nephrin and podocalyxin in the maturing podocyte where it is required during the formation of foot processes. Accordingly, the sialylation defects progressed with time and paralleled the morphologic changes. In summary, sialylation is critical during the development of the glomerular filtration barrier and required for the proper function of nephrin. Whether altered sialylation impairs nephrin function in human disease requires further study.

Published
2012
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15. CD2AP regulates SUMOylation of CIN85 in podocytes.

Author

Tossidou I, Niedenthal R, Klaus M, Teng B, Worthmann K, King BL, Peterson KJ, Haller H, and Schiffer M

Subjects
Adaptor Proteins, Signal Transducing genetics, Animals, Cell Line, Cytoskeletal Proteins genetics, Gene Deletion, Gene Expression Regulation, Humans, Mice, Neoplasm Proteins genetics, Nerve Tissue Proteins genetics, Phylogeny, Adaptor Proteins, Signal Transducing metabolism, Cytoskeletal Proteins metabolism, Neoplasm Proteins metabolism, Nerve Tissue Proteins metabolism, Podocytes metabolism, Small Ubiquitin-Related Modifier Proteins metabolism, Sumoylation
Abstract

Podocytes are highly differentiated and polarized epithelial cells located on the visceral side of the glomerulus. They form an indispensable component of the glomerular filter, the slit diaphragm, formed by several transmembrane proteins and adaptor molecules. Disruption of the slit diaphragm can lead to massive proteinuria and nephrotic syndrome in mice and humans. CD2AP is an adaptor protein that is important for the maintenance of the slit diaphragm. Together with its paralogue, CIN85, CD2AP belongs to a family of adaptor proteins that are primarily described as being involved in endocytosis and downregulation of receptor tyrosine kinase activity. We have shown that full-length CIN85 is upregulated in podocytes in the absence of CD2AP, whereas in wild-type cells, full-length CIN85 is not detectable. In this study, we show that full-length CIN85 is postranslationally modified by SUMOylation in wild-type podocytes. We can demonstrate that CIN85 is SUMOylated by SUMO-1, -2, and -3 and that SUMOylation is enhanced in the presence of CD2AP. Conversion of lysine 598 to arginine completely abolishes SUMOylation and leads to increased binding of CIN85 to nephrin. Our results indicate a novel role for CD2AP in regulating posttranslational modification of CIN85.

Published
2012
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16. CIN85/RukL is a novel binding partner of nephrin and podocin and mediates slit diaphragm turnover in podocytes.

Author

Tossidou I, Teng B, Drobot L, Meyer-Schwesinger C, Worthmann K, Haller H, and Schiffer M

Subjects
Adaptor Proteins, Signal Transducing chemistry, Adaptor Proteins, Signal Transducing genetics, Adaptor Proteins, Signal Transducing metabolism, Animals, Blotting, Western, Cell Line, Cells, Cultured, Cytoskeletal Proteins chemistry, Cytoskeletal Proteins genetics, Cytoskeletal Proteins metabolism, Endocytosis physiology, Enzyme-Linked Immunosorbent Assay, Fibroblast Growth Factors pharmacology, Fluorescent Antibody Technique, Humans, Immunohistochemistry, Immunoprecipitation, Intracellular Signaling Peptides and Proteins genetics, Membrane Proteins genetics, Mice, Neoplasm Proteins chemistry, Neoplasm Proteins genetics, Nerve Tissue Proteins chemistry, Nerve Tissue Proteins genetics, Podocytes drug effects, Protein Binding genetics, Protein Binding physiology, Proteinuria metabolism, RNA, Small Interfering, Ubiquitination drug effects, Intracellular Signaling Peptides and Proteins metabolism, Membrane Proteins metabolism, Neoplasm Proteins metabolism, Nerve Tissue Proteins metabolism, Podocytes metabolism
Abstract

Podocyte damage is the basis of many glomerular diseases with ultrastructural changes and decreased expression of components of the slit diaphragm such as nephrin and podocin. Under physiological conditions it is likely that the slit diaphragm underlies permanent renewal processes to indemnify its stability in response to changes in filtration pressure. This would require constant reorganization of the podocyte foot process and the renewal of slit diaphragm components. Thus far, the mechanisms underlying the turnover of slit diaphragm proteins are largely unknown. In this manuscript we examined a mechanism of nephrin endocytosis via CIN85/Ruk(L)-mediated ubiquitination. We can demonstrate that the loss of nephrin expression and onset of the proteinuria in CD2AP(-/-) mice correlates with an increased accumulation of ubiquitinated proteins and expression of CIN85/Ruk(L) in podocytes. In cultured murine podocytes CD2AP deficiency leads to an early ubiquitination of nephrin and podocin after stimulation with fibroblast growth factor-4. Binding assays with different CIN85/Ruk isoforms and mutants showed that nephrin and podocin are binding to the coiled-coil domain of CIN85/Ruk(L). We found that in the presence of CIN85/Ruk(L), which is involved in down-regulation of receptor-tyrosine kinases, nephrin is internalized after stimulation with fibroblast growth factor-4. Interestingly, coexpression of CIN85/Ruk(L) with CD2AP led to a decreased binding of CIN85/Ruk(L) to nephrin and podocin, which indicates a functional competition between CD2AP and CIN85/Ruk(L). Our results support a novel role for CIN85/Ruk(L) in slit diaphragm turnover and proteinuria.

Published
2010
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17. The balance of autocrine VEGF-A and VEGF-C determines podocyte survival.

Author

Müller-Deile J, Worthmann K, Saleem M, Tossidou I, Haller H, and Schiffer M

Subjects
Animals, Antibodies, Monoclonal pharmacology, Antibodies, Monoclonal, Humanized, Apoptosis drug effects, Apoptosis physiology, Bevacizumab, Cell Differentiation, Cell Line, Transformed, Cell Survival drug effects, Cell Survival physiology, Enzyme Activation drug effects, Humans, Mice, Phosphatidylinositol 3-Kinases metabolism, Podocytes cytology, Protein Kinase Inhibitors pharmacology, Proto-Oncogene Proteins c-akt metabolism, Vascular Endothelial Growth Factor A antagonists & inhibitors, Vascular Endothelial Growth Factor A pharmacology, Vascular Endothelial Growth Factor B pharmacology, Vascular Endothelial Growth Factor C antagonists & inhibitors, Vascular Endothelial Growth Factor C pharmacology, Vascular Endothelial Growth Factor Receptor-2 antagonists & inhibitors, Vascular Endothelial Growth Factor Receptor-3 antagonists & inhibitors, Autocrine Communication physiology, Podocytes physiology, Vascular Endothelial Growth Factor A metabolism, Vascular Endothelial Growth Factor C metabolism
Abstract

Podocytes are an important component of the glomerular filtration barrier and are the major source of vascular endothelial growth factor (VEGF) in the glomerulus. The role of VEGF for the phenotype of the glomerular endothelium has been intensely studied; however, the direct effects of autocrine VEGF on the podocyte are largely unknown. In this study we characterized the expression of VEGF isoforms and VEGF receptors in cultured human podocytes and examined direct effects on cell signaling and apoptosis after stimulation with exogenous VEGF or ablation of autocrine VEGF. We identified VEGF-A and VEGF-C as the dominant isoforms in human podocytes and showed that autocrine levels of both are important for the intracellular activation of antiapoptotic phosphoinositol 3-kinase/AKT and suppression of the proapoptotic p38MAPK via VEGFR-2. We demonstrated that ablation of VEGF-A or VEGF-C as well as treatment with bevacizumab or a VEGFR-2/-3 tyrosine kinase inhibitor led to reduced podocyte survival. In contrast, ablation of VEGF-B had no effect on podocyte survival. Treatment with exogenous VEGF-C reversed the effect of VEGF-A neutralization, and exogenous VEGF-A abrogated the effect of VEGF-C ablation in human podocytes. Our results underline the importance of autocrine VEGF for podocyte survival and indicate the delicate balance of VEGF-A and VEGF-C to influence progression of glomerular diseases.

Published
2009
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