scholarly journals Knockdown of Tmem234 in zebrafish results in proteinuria

2015 ◽  
Vol 309 (11) ◽  
pp. F955-F966 ◽  
Author(s):  
Patricia Q. Rodriguez ◽  
Asmundur Oddsson ◽  
Lwaki Ebarasi ◽  
Bing He ◽  
Kjell Hultenby ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
The Body ◽  
Zebrafish Larvae ◽  
Functional Integrity ◽  
Glomerular Capillary ◽  
Filtration Barrier ◽  
Foot Process Effacement ◽  
Foot Process ◽  
Critical Components ◽  
Molecular Components

Podocytes are highly specialized epithelial cells located at the outer aspects of the glomerular capillary tuft and critical components of the kidney filtration barrier. To maintain their unique features, podocytes express a number of proteins that are only sparsely found elsewhere in the body. In this study, we have identified four (Tmem234, Znf185, Lrrc49, and Slfn5) new highly podocyte-enriched proteins. The proteins are strongly expressed by podocytes, while other parts of the kidney show only weak or no expression. Tmem234, Slfn5, and Lrrc49 are located in foot processes, whereas Znf185 is found in both foot and major processes. Expressional studies in developing kidneys show that these proteins are first expressed at the capillary stage glomerulus, the same stage when the formation of major and foot processes begins. We identified zebrafish orthologs for Tmem234 and Znf185 genes and knocked down their expression using morpholino technology. Studies in zebrafish larvae indicate that Tmem234 is essential for the organization and functional integrity of the pronephric glomerulus filtration barrier, as inactivation of Tmem234 expression results in foot process effacement and proteinuria. In summary, we have identified four novel highly podocyte-enriched proteins and show that one of them, Tmem234, is essential for the normal filtration barrier in the zebrafish pronephric glomerulus. Identification of new molecular components of the kidney filtration barrier opens up possibilities to study their role in glomerulus biology and diseases.

PAR-3 Activation by Activated Protein C: a Novel Podocyte Protective Signalling Pathway

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 329-329
Author(s):  
Thati Madhusudhan ◽  
Hongjie Wang ◽  
Sandra Müller-Krebs ◽  
Vedat Schwenger ◽  
Martin G. Zeier ◽  
...  
Keyword(s):  
Protein C ◽  
Limited Proteolysis ◽  
Activated Protein C ◽  
Signalling Pathway ◽  
Diabetic Patients ◽  
Cytoprotective Effect ◽  
Filtration Barrier ◽  
Foot Process Effacement ◽  
Foot Process

Abstract Abstract 329 Activated protein C (APC) has anti-coagulant and cytoprotective effects. Recently we have shown that APC protects against diabetic nephropathy by inhibiting endothelial and podocyte apoptosis. The cytoprotecive effects of APC in endothelial cells require endothelial protein C receptor (EPCR) and protease activated recetor-1 (PAR-1). However, the signalling mechanism through which APC exerts its cytoprotective effects in podocytes is not known. Here we have used a mouse model of LPS-mediated podocyte injury and show that APC protects against LPS induced podocyte apoptosis and proteinuria. These cytoprotective effects of APC require activation of PAR-3. Supplementary in vitro experiments were performed using immortalized differentiated human and mouse podocytes to delineate the receptor mechanism. LPS administration (200 μg/mice, i.p.) induced podocyte apoptosis, foot process effacement and albuminuria by 24h. Administration of APC (20 μg/ mice, i.v, 6 h after LPS injection) protected against podocyte apoptosis, foot process effacement and albuminuria. Pre-incubation of APC with HAAPC antibody which blocks the anticoagulant, but not the cytoprotective properties of APC did not abolish the cytoprotective effect, establishing that the protective effect of APC is independent of its anticoagulant property. To demonstrate the direct cytoprotective effect of APC on podocytes, FITC labelled APC was administered i.v. Within 3 min FITC-APC accumulated in the pericapillary space of mouse glomeruli. In addition significantly elevated levels of PC were detected in urine samples of LPS treated mice as well as of diabetic patients or mice. These results suggest that APC transverses the glomerular filtration barrier, thus being able to directly modulate podocyte signalling in vivo. Furthermore experiments with PAR-3-/- mice, EPCR deficient (EPCRd/d) mice or PAR-1 antagonist (pepducin PI pal12S) showed that the cytoprotective effect of APC in podocytes is independent of EPCR but requires PAR-3 and PAR-1. In vitro APC (2 nM) inhibited puromycin aminonucleoside (PAN) induced podocyte apoptosis. These anti-apoptotic effect of APC require limited proteolysis of PAR-3 and cross-activation of either PAR-1 (mouse, rat podocytes) or PAR-2 (human podocytes). In addition, ectopic expression of PAR-3 in mesangial cells which lack PAR-3 is sufficient to render these cells APC sensitive and inhibit staurosporine induced apoptosis. In conclusion, we demonstrate a novel cyto-protective mechamism of APC in an acute model of nephropathy, which depends on APC mediated limited proteolysis of PAR-3. This novel podocyte protective signalling pathway may lay ground to the delineation of new pathophysiological concepts and therapeutic approaches. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Dosage-dependent role of Rac1 in podocyte injury

2016 ◽  
Vol 310 (8) ◽  
pp. F777-F784 ◽  
Author(s):  
Xiaoyang Wan ◽  
Mi-Sun Lee ◽  
Weibin Zhou
Keyword(s):  
Kidney Injury ◽  
Small Gtpase ◽  
Glomerular Sclerosis ◽  
Transgenic Zebrafish ◽  
Dependent Manner ◽  
Podocyte Injury ◽  
Filtration Barrier ◽  
Foot Process Effacement ◽  
Rac1 Activity ◽  
Foot Process

Activation of small GTPase Rac1 in podocytes is associated with rodent models of kidney injury and familial nephrotic syndrome. Induced Rac1 activation in podocytes in transgenic mice results in rapid transient proteinuria and foot process effacement, but not glomerular sclerosis. Thus it remains an open question whether abnormal activation of Rac1 in podocytes is sufficient to cause permanent podocyte damage. Using a number of transgenic zebrafish models, we showed that moderate elevation of Rac1 activity in podocytes did not impair the glomerular filtration barrier but aggravated metronidazole-induced podocyte injury, while inhibition of Rac1 activity ameliorated metronidazole-induced podocyte injury. Furthermore, a further increase in Rac1 activity in podocytes was sufficient to cause proteinuria and foot process effacement, which resulted in edema and lethality in juvenile zebrafish. We also found that activation of Rac1 in podocytes significantly downregulated the expression of nephrin and podocin, suggesting an adverse effect of Rac1 on slit diaphragm protein expression. Taken together, our data have demonstrated a causal link between excessive Rac1 activity and podocyte injury in a dosage-dependent manner, and transgenic zebrafish of variable Rac1 activities in podocytes may serve as useful animal models for the study of Rac1-related podocytopathy.

scholarly journals Sulfatases, in Particular Sulf1, Are Important for the Integrity of the Glomerular Filtration Barrier in Zebrafish

2019 ◽  
Vol 2019 ◽  
pp. 1-11
Author(s):  
Heiko Schenk ◽  
Anna Masseli ◽  
Patricia Schroder ◽  
Patricia Bolaños-Palmieri ◽  
Michaela Beese ◽  
...  
Keyword(s):  
Glomerular Filtration ◽  
Arteriovenous Malformations ◽  
Structural Changes ◽  
Protein Loss ◽  
Tail Region ◽  
Glomerular Filtration Barrier ◽  
Zebrafish Larvae ◽  
Filtration Barrier ◽  
Foot Process ◽  
Cell Crosstalk

The 6-O-endosulfatases (sulfs) are important enzymatic components involved in the regulation of heparan sulfate by altering the sulfatation pattern. Specifically in the kidney, sulfs have been implicated in the glomerular podocyte-endothelial cell crosstalk and in the preservation of the glomerular filtration barrier (GFB) in different mouse models. Since it has been shown that in zebrafish larvae, Sulf1, Sulf2a, and Sulf2b are expressed in the pronephric kidney we set out to establish if a reduction in sulf expression leads to GFB dysfunction. Here, we show that a reduced sulf expression following morpholino (MO) induced knockdown in zebrafish larvae promotes damage to the GFB leading to renal plasma protein loss from the circulation. Moreover, a combined knockdown of Sulf1, Sulf2a, and Sulf2b is associated with severe morphologic changes including narrowing of the fenestration between glomerular endothelial cells as well as thickening of the glomerular basement membrane and podocyte foot process effacement, suggesting that glomerular damage is an underlying cause of the circulatory protein loss observed after MO injection. Additionally, we show that a decrease in sulf expression reduces the bioavailability of VegfA in the glomerulus of the pronephros, which may contribute to the structural changes observed in the glomeruli of morphant fish. Furthermore, consistent with previous results, knockdown of the sulfs is associated with arteriovenous malformations in particular in the tail region of the larvae. Overall, taken together our results suggest that 6-O-endosulfatases are important in the preservation of GFB integrity and a reduction in their expression levels induces phenotypic changes that are indicative of renal protein loss.

scholarly journals Tangeretin Ameliorates Podocyte Injury Through Blocking Epithelial to Mesenchymal Transition in High Glucose-exposed Podocytes and Diabetic Kidneys (P06-009-19)

2019 ◽  
Vol 3 (Supplement_1) ◽  
Author(s):  
Min-Kyung Kang ◽  
Young-Hee Kang
Keyword(s):  
Epithelial Cells ◽  
High Glucose ◽  
Epithelial To Mesenchymal Transition ◽  
Kidney Tissue ◽  
Slit Diaphragm ◽  
Podocyte Injury ◽  
Mesenchymal Transition ◽  
Tissue Sections ◽  
Foot Process Effacement ◽  
Foot Process

Abstract Objectives Epithelial to mesenchymal transition (EMT) is a process by which epithelial cells acquire mesenchymal properties. This process contributes to the accumulation of matrix proteins in kidney and leads to renal glomeruli fibrosis. Tangeretin is an O-polymethoxylated flavone with anti-inflammatory and antioxidant properties that is found in citrus peels. This study investigated the renoprotective effects of tangeretin on EMT-mediated podocyte injury and fibrosis caused by hyperglycemia. Methods Mouse glomerular epithelial cells (podocyte) were incubated in media containing 5.5 mM glucose, 27.5 mM mannitol as an osmotic control or 33 mM glucose (high glucose) in the absence and presence of 1–20 μM tangeretin for up to 4 days. Antibodies of E-cadherin, N-cadherin, α-SMA, nephrin, podocin, P-cadherin and collagen1 were used for Western blotting. The in vivo animal model employed db/db mice orally administrated with 10 mg/kg tangeretin for 8 weeks. Kidney tissue extracts of were Western-blotted, and tissue sections cut down in 5 µM thickness were immunohistochemically stained. In addition, kidney tissue sections (ultrathin sections, 70 nm) were observed with transmission electron microscopy (TEM). Results Non-toxic tangeretin enhanced expression of the podocyte slit diaphragm proteins of nephrin, podocin and P-cadherin down-regulated by glucose stimulation. Also, tangeretin inhibited high glucose-induced expression of the mesenchymal markers of N-cadherin and α-smooth muscle actin, whereas the induction of the epithelial marker E-cadherin was enhanced. Furthermore tangeretin attenuated the fibronectin induction and collagen production elevated by the presence of high glucose. The TEM images revealed that podocyte foot process effacement occurred in diabetic mouse glomeruli. However, oral administration of 10 mg/kg tangeretin reduced urine albumin excretion and improved foot process effacement of diabetic podocytes through inhibiting loss of glomerular slit junction proteins. Conclusions These results demonstrated that tangeretin maintained the structures of podocyte slit diaphragm in a robust form, and inhibited podocyte injury through blocking epithelial to mesenchymal transition of podocytes. Therefore, tangeretin may be a potent renoprotective agent counteracting diabetes-associated loss of podocyte slit diaphragm and maintaining glomerular filtration barrier. Funding Sources This work was supported by the National Research Foundation of Korea (NRF) grants funded by the Korea government (MEST) (NRF-2017R1A6A3A04011473).

scholarly journals CHANGES IN FINE STRUCTURE DURING SILK PROTEIN PRODUCTION IN THE AMPULLATE GLAND OF THE SPIDER ARANEUS SERICATUS

1969 ◽  
Vol 42 (1) ◽  
pp. 284-295 ◽  
Author(s):  
Allen L. Bell ◽  
David B. Peakall
Keyword(s):  
Fine Structure ◽  
Epithelial Cells ◽  
Protein Production ◽  
Distal Portion ◽  
Silk Gland ◽  
The Body ◽  
Single Type ◽  
Secretory Cells ◽  
The Web ◽  
Tunica Intima

The ampullate silk gland of the spider, Araneus sericatus, produces the silk fiber for the scaffolding of the web. The fine structure of the various parts of the gland is described. The distal portion of the duct consist of a tube of epithelial cells which appear to secrete a substance which forms the tunica intima of the duct wall. At the proximal end of the duct there is a region of secretory cells. The epithelium of the sac portion contains five morphologically distinct types of granules. The bulk of the synthesis of silk occurs in the tail of the gland, and in this region only a single type of secretory droplet is seen in the epithelium. Protein synthesis can be stimulated by the injection of 1 mg/kg acetylcholine into the body fluids. 10 min after injection, much of the protein stored in the cytoplasm of the epithelial cells has been secreted into the lumen. 20 min after stimulation, the ergastoplasmic sacs form large whorls in the cytoplasm. Protein, similar in electron-opacity to protein found in the lumen, begins to form in that portion of the cytoplasm which is enclosed by the whorls. The limiting membrane of these droplets is formed by ergastoplasmic membranes which lose their ribosomes. No Golgi material has been found in these cells. Protein appears to be manufactured in the cytoplasm of the tail cells in a form which is ready for secretion.

scholarly journals A faster escape does not enhance survival in zebrafish larvae

2017 ◽  
Vol 284 (1852) ◽  
pp. 20170359 ◽  
Author(s):  
Arjun Nair ◽  
Christy Nguyen ◽  
Matthew J. McHenry
Keyword(s):  
High Speed ◽  
Larval Fish ◽  
Body Position ◽  
Three Dimensional ◽  
Limited Range ◽  
The Body ◽  
Model Parameters ◽  
Fish Prey ◽  
Zebrafish Larvae ◽  
Fish Predators

An escape response is a rapid manoeuvre used by prey to evade predators. Performing this manoeuvre at greater speed, in a favourable direction, or from a longer distance have been hypothesized to enhance the survival of prey, but these ideas are difficult to test experimentally. We examined how prey survival depends on escape kinematics through a novel combination of experimentation and mathematical modelling. This approach focused on zebrafish ( Danio rerio ) larvae under predation by adults and juveniles of the same species. High-speed three-dimensional kinematics were used to track the body position of prey and predator and to determine the probability of behavioural actions by both fish. These measurements provided the basis for an agent-based probabilistic model that simulated the trajectories of the animals. Predictions of survivorship by this model were found by Monte Carlo simulations to agree with our observations and we examined how these predictions varied by changing individual model parameters. Contrary to expectation, we found that survival may not be improved by increasing the speed or altering the direction of the escape. Rather, zebrafish larvae operate with sufficiently high locomotor performance due to the relatively slow approach and limited range of suction feeding by fish predators. We did find that survival was enhanced when prey responded from a greater distance. This is an ability that depends on the capacity of the visual and lateral line systems to detect a looming threat. Therefore, performance in sensing, and not locomotion, is decisive for improving the survival of larval fish prey. These results offer a framework for understanding the evolution of predator–prey strategy that may inform prey survival in a broad diversity of animals.

scholarly journals Mechanical challenges and cytoskeletal impairments in focal segmental glomerulosclerosis

2018 ◽  
Vol 314 (5) ◽  
pp. F921-F925 ◽  
Author(s):  
Di Feng ◽  
Clark DuMontier ◽  
Martin R. Pollak
Keyword(s):  
Blood Flow ◽  
Focal Segmental Glomerulosclerosis ◽  
Kidney Injury ◽  
Future Research ◽  
Disease States ◽  
Filtration Barrier ◽  
Segmental Glomerulosclerosis ◽  
The Face ◽  
Foot Process ◽  
Future Research Directions

Focal segmental glomerulosclerosis (FSGS) is a histologically defined form of kidney injury typically mediated by podocyte dysfunction. Podocytes rely on their intricate actin-based cytoskeleton to maintain the glomerular filtration barrier in the face of mechanical challenges resulting from pulsatile blood flow and filtration of this blood flow. This review summarizes the mechanical challenges faced by podocytes in the form of stretch and shear stress, both of which may play a role in the progression of podocyte dysfunction and detachment. It also reviews how podocytes respond to these mechanical challenges in dynamic fashion through rearranging their cytoskeleton, triggering various biochemical pathways, and, in some disease states, altering their morphology in the form of foot process effacement. Furthermore, this review highlights the growing body of evidence identifying several mutations of important cytoskeleton proteins as causes of FSGS. Lastly, it synthesizes the above evidence to show that a better understanding of how these mutations leave podocytes vulnerable to the mechanical challenges they face is essential to better understanding the mechanisms by which they lead to disease. The review concludes with future research directions to fill this gap and some novel techniques with which to pursue these directions.

scholarly journals Bordetella pertussisAdenylate Cyclase Toxin Disrupts Functional Integrity of Bronchial Epithelial Layers

2017 ◽  
Vol 86 (3) ◽  
Author(s):  
Shakir Hasan ◽  
Nikhil Nitin Kulkarni ◽  
Arni Asbjarnarson ◽  
Irena Linhartova ◽  
Radim Osicka ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Airway Epithelium ◽  
Immune Defense ◽  
Innate Immune ◽  
Camp Signaling ◽  
Apical Surface ◽  
Bronchial Epithelial ◽  
Content Type ◽  
Functional Integrity ◽  
Cell Layers

ABSTRACTThe airway epithelium restricts the penetration of inhaled pathogens into the underlying tissue and plays a crucial role in the innate immune defense against respiratory infections. The whooping cough agent,Bordetella pertussis, adheres to ciliated cells of the human airway epithelium and subverts its defense functions through the action of secreted toxins and other virulence factors. We examined the impact ofB. pertussisinfection and of adenylate cyclase toxin-hemolysin (CyaA) action on the functional integrity of human bronchial epithelial cells cultured at the air-liquid interface (ALI).B. pertussisadhesion to the apical surface of polarized pseudostratified VA10 cell layers provoked a disruption of tight junctions and caused a drop in transepithelial electrical resistance (TEER). The reduction of TEER depended on the capacity of the secreted CyaA toxin to elicit cAMP signaling in epithelial cells through its adenylyl cyclase enzyme activity. Both purified CyaA and cAMP-signaling drugs triggered a decrease in the TEER of VA10 cell layers. Toxin-produced cAMP signaling caused actin cytoskeleton rearrangement and induced mucin 5AC production and interleukin-6 (IL-6) secretion, while it inhibited the IL-17A-induced secretion of the IL-8 chemokine and of the antimicrobial peptide beta-defensin 2. These results indicate that CyaA toxin activity compromises the barrier and innate immune functions ofBordetella-infected airway epithelia.

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