scholarly journals Laminin-521 Protein Therapy for Glomerular Basement Membrane and Podocyte Abnormalities in a Model of Pierson Syndrome

2018 ◽  
Vol 29 (5) ◽  
pp. 1426-1436 ◽  
Author(s):  
Meei-Hua Lin ◽  
Joseph B. Miller ◽  
Yamato Kikkawa ◽  
Hani Y. Suleiman ◽  
Karl Tryggvason ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Congenital Nephrotic Syndrome ◽  
Super Resolution ◽  
Diffuse Mesangial Sclerosis ◽  
Protein Therapy ◽  
Correct Orientation ◽  
Pierson Syndrome ◽  
Foot Process ◽  
And Function

Background Laminin α5β2γ1 (LM-521) is a major component of the GBM. Mutations in LAMB2 that prevent LM-521 synthesis and/or secretion cause Pierson syndrome, a rare congenital nephrotic syndrome with diffuse mesangial sclerosis and ocular and neurologic defects. Because the GBM is uniquely accessible to plasma, which permeates endothelial cell fenestrae, we hypothesized that intravenous delivery of LM-521 could replace the missing LM-521 in the GBM of Lamb2 mutant mice and restore glomerular permselectivity.Methods We injected human LM-521 (hLM-521), a macromolecule of approximately 800 kD, into the retro-orbital sinus of Lamb2−/− pups daily. Deposition of hLM-521 into the GBM was investigated by fluorescence microscopy. We assayed the effects of hLM-521 on glomerular permselectivity by urinalysis and the effects on podocytes by desmin immunostaining and ultrastructural analysis of podocyte architecture.Results Injected hLM-521 rapidly and stably accumulated in the GBM of all glomeruli. Super-resolution imaging showed that hLM-521 accumulated in the correct orientation in the GBM, primarily on the endothelial aspect. Treatment with hLM-521 greatly reduced the expression of the podocyte injury marker desmin and attenuated the foot process effacement observed in untreated pups. Moreover, treatment with hLM-521 delayed the onset of proteinuria but did not prevent nephrotic syndrome, perhaps due to its absence from the podocyte aspect of the GBM.Conclusions These studies show that GBM composition and function can be altered in vivovia vascular delivery of even very large proteins, which may advance therapeutic options for patients with abnormal GBM composition, whether genetic or acquired.

scholarly journals Pierson Syndrome Associated with Hypothyroidism and Septic Shock

2020 ◽  
Vol 20 (4) ◽  
pp. e385-389
Author(s):  
Areeba Ejaz ◽  
Meher B. Ali ◽  
Fatima Siddiqui ◽  
Mashal B. Ali ◽  
Ammarah Jamal
Keyword(s):  
Septic Shock ◽  
Nephrotic Syndrome ◽  
Neuromuscular Junctions ◽  
Normal Component ◽  
Congenital Nephrotic Syndrome ◽  
Psychomotor Retardation ◽  
Basement Membranes ◽  
High Dose ◽  
Converting Enzyme Inhibitors ◽  
Pierson Syndrome

Pierson syndrome is caused by mutations in the laminin β2 gene causing absent β2 laminin, which is a normal component of the basement membranes of the mature glomerulus, structures in the anterior eye and neuromuscular junctions. The mutations manifest as congenital nephrotic syndrome and microcoria which are characteristic ocular features of this disease. These mutations may also result in neurological abnormalities such as hypotonia and psychomotor retardation. We report a two-month old boy who presented to the Pediatrics Department of Dr. Ruth K. M. Pfau Civil Hospital, Karachi, Pakistan, in 2015, with the typical features of microcoria and congenital nephrotic syndrome. The hypocalcaemia, hypoproteinaemia and probable immunocompromised state consequent to nephrotic syndrome resulted in seizures, hypothyroidism and urosepsis. Despite being treated aggressively with high dose antibiotics, ionotropic support, angiotensin-converting enzyme inhibitors, thyroxine replacement and nutritional support, the infant died due to significant multiorgan disease including renal failure and septic shock. Keywords: Pierson Syndrome; Microcoria and Congenital Nephrotic Syndrome; Congenital Microcoria; Hypothyroidism; Septic Shock; Case Report; Pakistan.

scholarly journals Human pluripotent stem cell-derived kidney organoids for personalized congenital and idiopathic nephrotic syndrome modeling

2021 ◽  
Author(s):  
Jitske Jansen ◽  
Bartholomeus T van den Berge ◽  
Martijn van den Broek ◽  
Rutger J Maas ◽  
Brigith Willemsen ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Idiopathic Nephrotic Syndrome ◽  
Super Resolution ◽  
Induced Pluripotent Stem Cell ◽  
Glomerular Injury ◽  
Kidney Organoids

Nephrotic syndrome (NS) is characterized by severe proteinuria as a consequence of kidney glomerular injury due to podocyte damage. In vitro models mimicking in vivo podocyte characteristics are a prerequisite to resolve NS pathogenesis. Here, we report human induced pluripotent stem cell derived kidney organoids containing a podocyte population that heads towards adult podocytes and were superior compared to 2D counterparts, based on scRNA sequencing, super-resolution imaging and electron microscopy. In this study, these next-generation podocytes in kidney organoids enabled personalized idiopathic nephrotic syndrome modeling as shown by activated slit diaphragm signaling and podocyte injury following protamine sulfate treatment and exposure to NS plasma containing pathogenic permeability factors. Organoids cultured from cells of a patient with heterozygous NPHS2 mutations showed poor NPHS2 expression and aberrant NPHS1 localization, which was reversible after genetic correction. Repaired organoids displayed increased VEGFA pathway activity and transcription factor activity known to be essential for podocyte physiology, as shown by RNA sequencing. This study shows that organoids are the preferred model of choice to study idiopathic and congenital podocytopathies.

scholarly journals Mitochondrial division, fusion and degradation

2019 ◽  
Vol 167 (3) ◽  
pp. 233-241 ◽  
Author(s):  
Daisuke Murata ◽  
Kenta Arai ◽  
Miho Iijima ◽  
Hiromi Sesaki
Keyword(s):  
Super Resolution ◽  
Healthy Population ◽  
Lipid Biochemistry ◽  
Mitochondrial Division ◽  
Cellular Processes ◽  
Size Number ◽  
Wide Range ◽  
Autophagic Degradation ◽  
And Function

Abstract The mitochondrion is an essential organelle for a wide range of cellular processes, including energy production, metabolism, signal transduction and cell death. To execute these functions, mitochondria regulate their size, number, morphology and distribution in cells via mitochondrial division and fusion. In addition, mitochondrial division and fusion control the autophagic degradation of dysfunctional mitochondria to maintain a healthy population. Defects in these dynamic membrane processes are linked to many human diseases that include metabolic syndrome, myopathy and neurodegenerative disorders. In the last several years, our fundamental understanding of mitochondrial fusion, division and degradation has been significantly advanced by high resolution structural analyses, protein-lipid biochemistry, super resolution microscopy and in vivo analyses using animal models. Here, we summarize and discuss this exciting recent progress in the mechanism and function of mitochondrial division and fusion.

Pierson Syndrome: A Novel Cause of Congenital Nephrotic Syndrome

PEDIATRICS ◽  
2006 ◽  
Vol 118 (2) ◽  
pp. e501-e505 ◽  
Author(s):  
R. VanDeVoorde ◽  
D. Witte ◽  
J. Kogan ◽  
J. Goebel
Keyword(s):  
Nephrotic Syndrome ◽  
Congenital Nephrotic Syndrome ◽  
Pierson Syndrome

High spatiotemporal resolution and low photo-toxicity fluorescence imaging in live cells and in vivo

2019 ◽  
Vol 47 (6) ◽  
pp. 1635-1650 ◽  
Author(s):  
Xiaohong Peng ◽  
Xiaoshuai Huang ◽  
Ke Du ◽  
Huisheng Liu ◽  
Liangyi Chen
Keyword(s):  
Fluorescence Microscopy ◽  
Spatial Resolution ◽  
Cell Biology ◽  
Critical Role ◽  
Super Resolution ◽  
Light Sheet ◽  
Live Cells ◽  
Spatiotemporal Resolution ◽  
And Function

Taking advantage of high contrast and molecular specificity, fluorescence microscopy has played a critical role in the visualization of subcellular structures and function, enabling unprecedented exploration from cell biology to neuroscience in living animals. To record and quantitatively analyse complex and dynamic biological processes in real time, fluorescence microscopes must be capable of rapid, targeted access deep within samples at high spatial resolutions, using techniques including super-resolution fluorescence microscopy, light sheet fluorescence microscopy, and multiple photon microscopy. In recent years, tremendous breakthroughs have improved the performance of these fluorescence microscopies in spatial resolution, imaging speed, and penetration. Here, we will review recent advancements of these microscopies in terms of the trade-off among spatial resolution, sampling speed and penetration depth and provide a view of their possible applications.

Abstract 19049: CASK Regulates Sodium Channels Distribution and Function in Costameric Microdomain

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Catherine A Eichel ◽  
Lin Xianming ◽  
Florent Louault ◽  
Gilles Dilanian ◽  
Mario Delmar ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Sodium Current ◽  
Systolic Dysfunction ◽  
Super Resolution ◽  
Adult Rats ◽  
Lateral Membrane ◽  
T Tubules ◽  
Conventional Cell ◽  
And Function

Introduction: Proteins of the MAGUK family have emerged as key components in ion channels organization and regulation within specialized submembrane domains of cardiomyocytes. In this context, we investigated for the first time the expression, localization and function in the heart of the MAGUK protein CASK. Methods and Results: Surprisingly, no signal was detectable for CASK at intercalated discs, making CASK the first MAGUK to be excluded from these structures. On the contrary, CASK was located at the lateral membrane where it belonged specifically to the costameric dystrophin/glycoproteins complex (DGC). Since a lateral sub-population of Nav1.5 channels have been reported to interact with syntrophin, a member of DGC, we hypothesized that CASK could modulate Nav1.5 channels. Using high resolution 3-D deconvolution microscopy, we observed a co-localization of CASK and Nav1.5 in cardiomyocytes and co-IP experiments confirmed that the two proteins are in the same complex. Whole-cell patch-clamp recordings revealed a negative regulation by CASK of the sodium current INa carried by Nav1.5 channels in cardiomyocytes. Using super resolution scanning coupled to conventional cell-attached patch-clamp, we investigated the involvement of CASK on Nav1.5 function and localization in highly confined microdomains of the lateral membrane. INa was recorded in crests and T-tubules on freshly isolated cardiomyocytes obtained from adult rats injected with Adeno Associated Virus (eGFP-AAV-ShCASK or eGFP-AAV-ShScr). CASK silencing caused a drastic reduction of INa at the crest whereas it increased the current at the T-tubules suggesting that CASK retains Nav1.5 channels at the crest and prevents their clustering in T-tubules. In vivo CASK silencing in rat using an eGFP-AAV-ShCASK-injected was associated with a conduction slowing as indicated by the prolongation of the QRS duration, cardiac dilation and left ventricle systolic dysfunction. Conclusion: Taken together these results indicate that CASK is a major determinant of the organization of a subpopulation of Nav1.5 at the costamere of cardiomyocytes and that CASK-silencing induces electrical and morphological remodeling.

scholarly journals Nanobodies as Versatile Tool for Multiscale Imaging Modalities

Biomolecules ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 1695
Author(s):  
Marco Erreni ◽  
Tilo Schorn ◽  
Francesca D’Autilia ◽  
Andrea Doni
Keyword(s):  
Molecular Imaging ◽  
Super Resolution ◽  
Imaging Modalities ◽  
Molecular Imaging Probes ◽  
Imaging Probes ◽  
Versatile Tool ◽  
And Function ◽  
Preclinical And Clinical Studies

Molecular imaging is constantly growing in different areas of preclinical biomedical research. Several imaging methods have been developed and are continuously updated for both in vivo and in vitro applications, in order to increase the information about the structure, localization and function of molecules involved in physiology and disease. Along with these progresses, there is a continuous need for improving labeling strategies. In the last decades, the single domain antigen-binding fragments nanobodies (Nbs) emerged as important molecular imaging probes. Indeed, their small size (~15 kDa), high stability, affinity and modularity represent desirable features for imaging applications, providing higher tissue penetration, rapid targeting, increased spatial resolution and fast clearance. Accordingly, several Nb-based probes have been generated and applied to a variety of imaging modalities, ranging from in vivo and in vitro preclinical imaging to super-resolution microscopy. In this review, we will provide an overview of the state-of-the-art regarding the use of Nbs in several imaging modalities, underlining their extreme versatility and their enormous potential in targeting molecules and cells of interest in both preclinical and clinical studies.

scholarly journals CD2AP is expressed with nephrin in developing podocytes and is found widely in mature kidney and elsewhere

2000 ◽  
Vol 279 (4) ◽  
pp. F785-F792 ◽  
Author(s):  
Cong Li ◽  
Vesa Ruotsalainen ◽  
Karl Tryggvason ◽  
Andrey S. Shaw ◽  
Jeffrey H. Miner
Keyword(s):  
Nephrotic Syndrome ◽  
Collecting Duct ◽  
Congenital Nephrotic Syndrome ◽  
Human Kidney ◽  
Distal Tubule ◽  
Slit Diaphragm ◽  
Mutant Mice ◽  
General Role ◽  
Foot Process ◽  
Punctate Pattern

CD2-associated protein (CD2AP) is an adapter molecule that can bind to the cytoplasmic domain of nephrin, a component of the glomerular slit diaphragm. Mice lacking CD2AP exhibit a congenital nephrotic syndrome characterized by extensive foot process effacement, suggesting that CD2AP-nephrin interactions are critical to maintaining slit diaphragm function. We have examined the patterns of expression of both CD2AP and nephrin in developing mouse and human kidney. Both proteins were first detected in developing podocytes at the capillary loop stage of glomerulogenesis and eventually became concentrated near the glomerular basement membrane. CD2AP was also observed diffusely in collecting duct and apically in many cells of proximal and distal tubule. Kidneys from Cd2ap −/− mice initially exhibited normal nephrin localization, but as the mice aged and foot processes became effaced, nephrin disappeared. In laminin-β2 mutant mice exhibiting nephrotic syndrome, CD2AP in glomeruli was aberrantly localized in a primarily punctate pattern. Extensive extrarenal expression of CD2AP was observed in endothelial and epithelial cells, in many cases with a specific subcellular localization. Together, these results suggest that CD2AP is not only involved in maintaining the slit diaphragm but may also have a general role in maintaining specialized subcellular architecture. The severity of kidney disease in Cd2ap mutant mice may have eclipsed manifestation of defects in other tissues.

scholarly journals Ste20-like kinase, SLK, a novel mediator of podocyte integrity

2018 ◽  
Vol 315 (1) ◽  
pp. F186-F198 ◽  
Author(s):  
Andrey V. Cybulsky ◽  
Joan Papillon ◽  
Julie Guillemette ◽  
Natalya Belkina ◽  
Genaro Patino-Lopez ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Plasma Membranes ◽  
Kidney Development ◽  
Kidney Injury ◽  
Ultrastructural Changes ◽  
Morphological Evidence ◽  
Foot Process ◽  
And Function ◽  
Specific Deletion

SLK is essential for embryonic development and may play a key role in wound healing, tumor growth, and metastasis. Expression and activation of SLK are increased in kidney development and during recovery from ischemic acute kidney injury. Overexpression of SLK in glomerular epithelial cells/podocytes in vivo induces injury and proteinuria. Conversely, reduced SLK expression leads to abnormalities in cell adhesion, spreading, and motility. Tight regulation of SLK expression thus may be critical for normal renal structure and function. We produced podocyte-specific SLK-knockout mice to address the functional role of SLK in podocytes. Mice with podocyte-specific deletion of SLK showed reduced glomerular SLK expression and activity compared with control. Podocyte-specific deletion of SLK resulted in albuminuria at 4–5 mo of age in male mice and 8–9 mo in female mice, which persisted for up to 13 mo. At 11–12 mo, knockout mice showed ultrastructural changes, including focal foot process effacement and microvillous transformation of podocyte plasma membranes. Mean foot process width was approximately twofold greater in knockout mice compared with control. Podocyte number was reduced by 35% in knockout mice compared with control, and expression of nephrin, synaptopodin, and podocalyxin was reduced in knockout mice by 20–30%. In summary, podocyte-specific deletion of SLK leads to albuminuria, loss of podocytes, and morphological evidence of podocyte injury. Thus, SLK is essential to the maintenance of podocyte integrity as mice age.

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