Drosophila Filamin exhibits a mechano-protective role during nephrocyte injury via induction of hypertrophic growth

Podocytes are highly specialized epithelial cells of the kidney glomerulus and are an essential part of the filtration barrier. Due to their position and function in the kidney, they are exposed to constant biomechanical forces such as shear stress and hydrostatic pressure. These forces increase during disease, resulting in podocyte injury and loss. The mechanism by which biomechanical forces are sensed and transduced to elicit an adaptive and protective response remains largely unknown. Here we show, using the Drosophila nephrocyte model, that the mechanosensor Cheerio (dFilamin) is central to this mechano-protective mechanism. We found expression of an activated mechanosensitive variant of Cheerio induced hypertrophy and rescued filtration function in injured nephrocytes. Additional analysis with human Filamin B confirmed this mechano-protective role. We delineated the mechano-protective pathway downstream of Cheerio and found activation of TOR and Yorkie induce nephrocyte hypertrophy, whereas their repression reversed the Cheerio-mediated hypertrophy. Although Cheerio/Filamin B pathway mediates a mechano-protective role in the face of injury, we found excessive activity resulted in a pathological phenotype, indicating activity levels must be tightly controlled. Taken together, our data suggest that Cheerio acts via the TOR and YAP pathway to induce hypertrophic growth, as a mechano-protective response to nephrocyte injury.

Filaminopathy type C

Filamin is an actin-binding protein that, by forming flexible molecular cross-links, stabilizes the three-dimensional F-actin networks and gives them the mechanical properties of a gel. It is represented by three isoforms: filamine A (FLNA), filamin B (FLNB), and filamin C (FLNC), derived from 3 homologous genes. Laminopathies caused by mutations in the FLNA, FLNB, and FLNC genes represent an extensive allelic series of diseases. The review discusses in detail the genotype-phenotypic correlation of all types of phylaminopathies. The neuromuscular and cardiac clinic of C-type phylaminopathy is described in detail. Three variants of C phylaminopathy known at the moment are analyzed.

Proteomic Analysis of Differentially Expressed Proteins in Endothelial Cells Under Low Shear Stress

Background: Shear stress (SS) affects the morphology, proliferation, differentiation and migration of endothelial cells, and regulates protein expression.Methods: This study aims to conduct a proteomic analysis of human umbilical vein endothelial cells (HUVECs) under low SS. HUVECs were cultured on glass slides (test group n=30; control group n=30) and transferred to parallel-plate flow chamber. The test group was subjected to low SS at 4.58 dyne/cm2 for 2 hours, and the control group was maintained under static conditions. Approximately 2×107 cells were collected from each group and analyzed by two-dimensional electrophoresis. The protein from each spot were identified by mass spectrometry. The biological functions and processes associated with the identified proteins were evaluated using the NCBInr, SwissProt, and DAVID databases. The protein expression of 14-3-3 proteins, annexin I, annexin II, zyxin, filamin A, filamin B, Ras-related nuclear protein (Ran), and talin were assessed by Western blotting.Results: A total of 234 proteins were analyzed; 14 proteins were upregulated and 78 were downregulated in the test group compared with the control group. The expression of 14-3-3 proteins, annexin I, annexin II, zyxin, filamin A, and filamin B was significantly decreased, whereas the expression of Ran and talin was significantly increased in the test group.Conclusion: The current results provide evidence that SS changes the protein profile of HUVECs.

Proteomic analysis of differentially expressed proteins in endothelial cells under low shear stress

Background: Shear stress (SS) affects the morphology, proliferation, differentiation and migration of endothelial cells, and regulates protein expression.Methods: This study aims to conduct a proteomic analysis of human umbilical vein endothelial cells (HUVECs) under low SS. HUVECs were cultured on glass slides (test group n=30; control group n=30) and transferred to parallel-plate flow chamber. The test group was subjected to low SS at 4.58 dyne/cm2 for 2 hours, and the control group was maintained under static conditions. Approximately 2´107 cells were collected from each group and analyzed by two-dimensional electrophoresis. The protein from each spot were identified by mass spectrometry. The biological functions and processes associated with the identified proteins were evaluated using the NCBInr, SwissProt, and DAVID databases. The protein expression of 14-3-3 proteins, annexin I, annexin II, zyxin, filamin A, filamin B, Ras-related nuclear protein (Ran), and talin were assessed by Western blotting.Results: A total of 234 proteins were analyzed; 14 proteins were upregulated and 78 were downregulated in the test group compared with the control group. The expression of 14-3-3 proteins, annexin I, annexin II, zyxin, filamin A, and filamin B was significantly decreased, whereas the expression of Ran and talin was significantly increased in the test group.Conclusion: The current results provide evidence that SS changes the protein profile of HUVECs.

Proteome Analysis of Isolated Podocytes Reveals Stress Responses in Glomerular Sclerosis

BackgroundUnderstanding podocyte-specific responses to injury at a systems level is difficult because injury leads to podocyte loss or an increase of extracellular matrix, altering glomerular cellular composition. Finding a window into early podocyte injury might help identify molecular pathways involved in the podocyte stress response.MethodsWe developed an approach to apply proteome analysis to very small samples of purified podocyte fractions. To examine podocytes in early disease states in FSGS mouse models, we used podocyte fractions isolated from individual mice after chemical induction of glomerular disease (with Doxorubicin or LPS). We also applied single-glomerular proteome analysis to tissue from patients with FSGS.ResultsTranscriptome and proteome analysis of glomeruli from patients with FSGS revealed an underrepresentation of podocyte-specific genes and proteins in late-stage disease. Proteome analysis of purified podocyte fractions from FSGS mouse models showed an early stress response that includes perturbations of metabolic, mechanical, and proteostasis proteins. Additional analysis revealed a high correlation between the amount of proteinuria and expression levels of the mechanosensor protein Filamin-B. Increased expression of Filamin-B in podocytes in biopsy samples from patients with FSGS, in single glomeruli from proteinuric rats, and in podocytes undergoing mechanical stress suggests that this protein has a role in detrimental stress responses. In Drosophila, nephrocytes with reduced filamin homolog Cher displayed altered filtration capacity, but exhibited no change in slit diaphragm structure.ConclusionsWe identified conserved mechanisms of the podocyte stress response through ultrasensitive proteome analysis of human glomerular FSGS tissue and purified native mouse podocytes during early disease stages. This approach enables systematic comparisons of large-scale proteomics data and phenotype-to-protein correlation.

Implicaciones anestésicas en el Síndrome de Larsen: A propósito de un caso

El síndrome de Larsen (SL) es una enfermedad hereditaria rara caracterizada por un defecto en la formación de colágeno debido a mutaciones en los genes que codifican la proteína citoesquelética filamina B. Su prevalencia en Europa es aproximadamente de 1/250.000 nacidos vivos. Esto implica una serie de rasgos y particularidades anatómicas de la vía aérea que debemos valorar en niños que van a ser sometidos a un acto anestésico. Se presenta el caso de un niño de 11 años diagnosticado de síndrome de Larsen que se interviene de aticotomía oído izquierdo. A este propósito, realizamos revisión bibliográfica sobre las peculiaridades del manejo anestésico de estos pacientes. ABSTRACT Anaesthetic implications in Larsen Syndrome: about a case Larsen Syndrome (SL) is a rare hereditary disease characterized by a defect in the formation of collagen due to mutations in the genes encoding the cytoskeletal protein filamin B. Its prevalence in Europe is approximately 1 to 250,000 live births. This implies a number of anatomical features of the airway that we must assess in children who are going under anesthesia. We present the case of an 11-year-old boy diagnosed with Larsen syndrome who underwent left ear aticotomy. In this regard, we conducted a literature review on the peculiarities of anesthetic management of these patients.

Proteomic analysis of differentially expressed proteins in endothelial cells under low shear stress

Shear stress (SS) affects the morphology, migration, differentiation, and proliferation of endothelial cells, and regulates protein expression. The objective of this study was to perform a proteomic analysis of human umbilical vein endothelial cells (HUVECs) under low SS. HUVEC lines were cultured on glass slides (test group n=30; control group n=30) and transferred to a parallel-plate flow chamber. The test group was subjected to low SS at 4.58 dyne/cm2 for 2 hours, and the control group was maintained under static conditions. Approximately 2×107 cells were collected from each group and analyzed by two-dimensional electrophoresis. The proteins from each spot were identified by mass spectrometry. The biological functions and processes associated with the identified proteins were evaluated using the NCBInr, SwissProt, and DAVID databases. The protein expression of 14-3-3 proteins, annexin I, annexin II, zyxin, filamin A, filamin B, Ras-related nuclear protein (Ran), and talin were assessed by Western blotting. A total of 234 proteins were analyzed; 14 proteins were upregulated and 78 were downregulated in the test group compared with the control group. The expression of 14-3-3 proteins, annexin I, annexin II, zyxin, filamin A, and filamin B was significantly decreased, whereas the expression of Ran and talin was significantly increased in the test group. It is known that SS induces the remodeling of endothelial cells by modulating protein expression. However, the mechanisms underlying this modulation are unknown. The present results provide evidence that SS changes the protein profile of HUVECs.