Structured Review

Progen Biotechnik mouse anti synaptopodin
Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using <t>anti-synaptopodin</t> and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.
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1) Product Images from "Novel Microscopic Techniques for Podocyte Research"

Article Title: Novel Microscopic Techniques for Podocyte Research

Journal: Frontiers in Endocrinology

doi: 10.3389/fendo.2018.00379

Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using anti-synaptopodin and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.
Figure Legend Snippet: Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using anti-synaptopodin and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.

Techniques Used: Labeling, Staining

2) Product Images from "Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation"

Article Title: Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation

Journal: Scientific Reports

doi: 10.1038/srep17694

Effects of CsA on podocyte WAVE1 expression in PAN-induced rat nephropathy. ( A ) Western blot analysis of WAVE1 in isolated glomeruli. ( B ) WAVE1 expression was quantified and normalized to GAPDH expression. ( C ) Immunofluorescent staining of WAVE1 and synaptopodin in rats. Scale bar = 20 μm. WAVE1 is labelled in green, and synaptopodin is labelled red. The data are presented as the mean ± SD. n = 5. **P
Figure Legend Snippet: Effects of CsA on podocyte WAVE1 expression in PAN-induced rat nephropathy. ( A ) Western blot analysis of WAVE1 in isolated glomeruli. ( B ) WAVE1 expression was quantified and normalized to GAPDH expression. ( C ) Immunofluorescent staining of WAVE1 and synaptopodin in rats. Scale bar = 20 μm. WAVE1 is labelled in green, and synaptopodin is labelled red. The data are presented as the mean ± SD. n = 5. **P

Techniques Used: Expressing, Western Blot, Isolation, Staining

The interaction between WAVE1 and calcineurin and WAVE1 phosphorylation in PAN-stimulated podocytes. ( A ) Co-immunoprecipitation analysis of the interaction between WAVE1 and calcineurin. Control IgG represents normal rabbit IgG (RIgG), which replaced the anti-calcineurin antibody in the precipitation process. ( B ) WAVE1 serine phosphorylation was detected by co-immunoprecipitation. Control IgG represents normal rabbit IgG, which replaced the anti-calcineurin antibody in the precipitation process. ( C ) WAVE1 serine phosphorylation was quantified based on the expression of total WAVE1 in the sample precipitated with the anti-calcineurin antibody. ( D ) Western blot of synaptopodin expression in WAVE1-overexpressing podocytes. ( E , F ) WAVE1 and synaptopodin expression levels were quantified after normalization to GAPDH expression. ( G ) Western blot of WAVE1 expression in synaptopodin-knockdown podocytes. ( H,I ) Synaptopodin and WAVE1 expression levels were quantified after normalization to GAPDH expression. The data are presented as the mean ± SD. n = 3. *P
Figure Legend Snippet: The interaction between WAVE1 and calcineurin and WAVE1 phosphorylation in PAN-stimulated podocytes. ( A ) Co-immunoprecipitation analysis of the interaction between WAVE1 and calcineurin. Control IgG represents normal rabbit IgG (RIgG), which replaced the anti-calcineurin antibody in the precipitation process. ( B ) WAVE1 serine phosphorylation was detected by co-immunoprecipitation. Control IgG represents normal rabbit IgG, which replaced the anti-calcineurin antibody in the precipitation process. ( C ) WAVE1 serine phosphorylation was quantified based on the expression of total WAVE1 in the sample precipitated with the anti-calcineurin antibody. ( D ) Western blot of synaptopodin expression in WAVE1-overexpressing podocytes. ( E , F ) WAVE1 and synaptopodin expression levels were quantified after normalization to GAPDH expression. ( G ) Western blot of WAVE1 expression in synaptopodin-knockdown podocytes. ( H,I ) Synaptopodin and WAVE1 expression levels were quantified after normalization to GAPDH expression. The data are presented as the mean ± SD. n = 3. *P

Techniques Used: Immunoprecipitation, Expressing, Western Blot

3) Product Images from "Laminin α1 Regulates Age-Related Mesangial Cell Proliferation and Mesangial Matrix Accumulation through the TGF-β Pathway"

Article Title: Laminin α1 Regulates Age-Related Mesangial Cell Proliferation and Mesangial Matrix Accumulation through the TGF-β Pathway

Journal: The American Journal of Pathology

doi: 10.1016/j.ajpath.2014.02.006

Analysis of primary mesangial cells. Mesangial cells are elongated, arranged in multiple layers, and stained positively for smooth muscle α-actin ( A ), but not synaptopodin ( B ) or PECAM ( C ). D: Proliferation was assessed by immunofluorescence assay for BrdU-positive cells as a percentage of the total number of cells, averaged from five separate fields of view. Mesangial cells from Lama1 CKO mice exhibit significantly increased proliferation compared with control cells. This increase is inhibited by exogenous LAMA1-containing LM-111, but not by LAMA2- or LAMA5-containing isoforms. E: Mesangial cell proliferation determined in cells transfected with vector alone or with a LAMA1 expression vector. Control (dark gray bars) and Lama1 CKO (light gray bars) cells were cultured on slide chambers for 24 hours and transfected for 48 hours before proliferation analysis. Restoration of LAMA1 expression by Lama1 CKO cells suppresses their proliferation. Data represent means ± SEM. ∗ P
Figure Legend Snippet: Analysis of primary mesangial cells. Mesangial cells are elongated, arranged in multiple layers, and stained positively for smooth muscle α-actin ( A ), but not synaptopodin ( B ) or PECAM ( C ). D: Proliferation was assessed by immunofluorescence assay for BrdU-positive cells as a percentage of the total number of cells, averaged from five separate fields of view. Mesangial cells from Lama1 CKO mice exhibit significantly increased proliferation compared with control cells. This increase is inhibited by exogenous LAMA1-containing LM-111, but not by LAMA2- or LAMA5-containing isoforms. E: Mesangial cell proliferation determined in cells transfected with vector alone or with a LAMA1 expression vector. Control (dark gray bars) and Lama1 CKO (light gray bars) cells were cultured on slide chambers for 24 hours and transfected for 48 hours before proliferation analysis. Restoration of LAMA1 expression by Lama1 CKO cells suppresses their proliferation. Data represent means ± SEM. ∗ P

Techniques Used: Staining, Immunofluorescence, Mouse Assay, Transfection, Plasmid Preparation, Expressing, Cell Culture

4) Product Images from "KIBRA Modulates Directional Migration of Podocytes"

Article Title: KIBRA Modulates Directional Migration of Podocytes

Journal: Journal of the American Society of Nephrology : JASN

doi: 10.1681/ASN.2007080916

KIBRA binds directly to synaptopodin via the WW domains. (A) Synaptopodin and dendrin carry two independent PPxY motifs with an S/T at the third position. The regions between the flanking PPxY motifs contain additional identical aa (magenta). Dendrin
Figure Legend Snippet: KIBRA binds directly to synaptopodin via the WW domains. (A) Synaptopodin and dendrin carry two independent PPxY motifs with an S/T at the third position. The regions between the flanking PPxY motifs contain additional identical aa (magenta). Dendrin

Techniques Used:

Model of the cellular function of KIBRA. KIBRA directly interacts with dendrin, synaptopodin (SYNPO), and PATJ and aPKC (red arrows). Thus, KIBRA could serve as a linker molecule between polarity proteins and components of the cytoskeleton (actin, α-actinin,
Figure Legend Snippet: Model of the cellular function of KIBRA. KIBRA directly interacts with dendrin, synaptopodin (SYNPO), and PATJ and aPKC (red arrows). Thus, KIBRA could serve as a linker molecule between polarity proteins and components of the cytoskeleton (actin, α-actinin,

Techniques Used: Cell Function Assay

5) Product Images from "Origin of Parietal Podocytes in Atubular Glomeruli Mapped by Lineage Tracing"

Article Title: Origin of Parietal Podocytes in Atubular Glomeruli Mapped by Lineage Tracing

Journal: Journal of the American Society of Nephrology : JASN

doi: 10.1681/ASN.2013040376

Podocyte and PEC marker expression in humans. (A–A’’’) Immunofluorescence double stainings of an apparently normal kidney of an 18-year-old old healthy man show no colocalization of the podocyte marker synaptopodin and
Figure Legend Snippet: Podocyte and PEC marker expression in humans. (A–A’’’) Immunofluorescence double stainings of an apparently normal kidney of an 18-year-old old healthy man show no colocalization of the podocyte marker synaptopodin and

Techniques Used: Marker, Expressing, Immunofluorescence

Statistical analysis of the proportion of Bowman’s capsules covered by synaptopodin-positive cells (presumptive parietal podocytes). More than half of the glomeruli do not show parietal podocytes. In a significant portion of glomeruli (approximately
Figure Legend Snippet: Statistical analysis of the proportion of Bowman’s capsules covered by synaptopodin-positive cells (presumptive parietal podocytes). More than half of the glomeruli do not show parietal podocytes. In a significant portion of glomeruli (approximately

Techniques Used:

Cysts induced by electrocoagulation are glomerular cysts. (A) In normal glomeruli, synaptopodin (A, arrow) and SSeCKS (A’, arrow) are specifically detected in podocytes and PECs, respectively. The boundary from synaptopodin-positive podocytes
Figure Legend Snippet: Cysts induced by electrocoagulation are glomerular cysts. (A) In normal glomeruli, synaptopodin (A, arrow) and SSeCKS (A’, arrow) are specifically detected in podocytes and PECs, respectively. The boundary from synaptopodin-positive podocytes

Techniques Used:

Parietal podocytes in atubular glomeruli. (A and B) Bowman's capsules of atubular glomeruli are lined by synaptopodin-positive parietal podocytes regardless of the cyst diameter. (C–F) Influence of genetic background (38 Sv129 mice versus 36 C57BL/6
Figure Legend Snippet: Parietal podocytes in atubular glomeruli. (A and B) Bowman's capsules of atubular glomeruli are lined by synaptopodin-positive parietal podocytes regardless of the cyst diameter. (C–F) Influence of genetic background (38 Sv129 mice versus 36 C57BL/6

Techniques Used: Mouse Assay

6) Product Images from "Metformin ameliorates the severity of experimental Alport syndrome"

Article Title: Metformin ameliorates the severity of experimental Alport syndrome

Journal: Scientific Reports

doi: 10.1038/s41598-021-86109-1

Metformin protects against podocyte dysfunction in Col4a5 G5X Alport syndrome mice. ( a ) Visualization of indicated podocyte and glomerular cell proteins in renal tissue sections. Scale bars, 50 μm. ( b ) Quantification of WT1-positive cells in the glomerulus showed losartan or metformin protected against podocyte loss in late stage C57BL/6 Col4a5 G5X vAlport syndrome mice. ( c ) Quantification of nephrin- and synaptopodin-positive area in the glomerulus showed that the decrease of these podocyte proteins in glomerulus was suppressed in losartan- or metformin-treated C57BL/6 Col4a5 G5X Alport syndrome mice. ( d ) Quantification of PCNA-positive cells in the glomerulus showed the number of proliferating cells in glomerulus was decreased in losartan- or metformin-treated C57BL/6 Col4a5 G5X Alport syndrome mice. Data are expressed as the means ± S.E. (n = 4 per group). P values were assessed by Dunnett’s test ( ## P
Figure Legend Snippet: Metformin protects against podocyte dysfunction in Col4a5 G5X Alport syndrome mice. ( a ) Visualization of indicated podocyte and glomerular cell proteins in renal tissue sections. Scale bars, 50 μm. ( b ) Quantification of WT1-positive cells in the glomerulus showed losartan or metformin protected against podocyte loss in late stage C57BL/6 Col4a5 G5X vAlport syndrome mice. ( c ) Quantification of nephrin- and synaptopodin-positive area in the glomerulus showed that the decrease of these podocyte proteins in glomerulus was suppressed in losartan- or metformin-treated C57BL/6 Col4a5 G5X Alport syndrome mice. ( d ) Quantification of PCNA-positive cells in the glomerulus showed the number of proliferating cells in glomerulus was decreased in losartan- or metformin-treated C57BL/6 Col4a5 G5X Alport syndrome mice. Data are expressed as the means ± S.E. (n = 4 per group). P values were assessed by Dunnett’s test ( ## P

Techniques Used: Mouse Assay

7) Product Images from "Characterization and Culture of Fetal Rhesus Monkey Renal Cortical Cells"

Article Title: Characterization and Culture of Fetal Rhesus Monkey Renal Cortical Cells

Journal: Pediatric research

doi: 10.1203/PDR.0b013e3181b45565

Immunocytochemistry - Cultured cortical cells on fibronectin and renal ECM Cells from passage 5 cultured on fibronectin contained more synaptopodin positive cells (red) (arrow) than those cultured on other substrates. Nuclei stained with DAPI (blue) (A-D). Synaptopodin positive cells were positive for nestin (green) and showed morphology consistent with mature podocytes (A-C). (A-B, 20x, bar = 100 μm; C, 60x, bar = 50 μm; D, 100x, bar = 20 μm).
Figure Legend Snippet: Immunocytochemistry - Cultured cortical cells on fibronectin and renal ECM Cells from passage 5 cultured on fibronectin contained more synaptopodin positive cells (red) (arrow) than those cultured on other substrates. Nuclei stained with DAPI (blue) (A-D). Synaptopodin positive cells were positive for nestin (green) and showed morphology consistent with mature podocytes (A-C). (A-B, 20x, bar = 100 μm; C, 60x, bar = 50 μm; D, 100x, bar = 20 μm).

Techniques Used: Immunocytochemistry, Cell Culture, Staining

Quantitative PCR - Synaptopodin and nestin While synaptopodin expression increased (A), nestin expression decreased (B) in cells grown on all substrates studied (black column - P1, white column - P5). Furthermore, podocyte specific expression was present in cells at P5 (C) (black - plastic, white - renal ECM) (D) (black - collagen I, white - collagen IV, gray - laminin, hatched - fibronectin). Passage 1 (P1) compared to passage 5 (P5) (*p
Figure Legend Snippet: Quantitative PCR - Synaptopodin and nestin While synaptopodin expression increased (A), nestin expression decreased (B) in cells grown on all substrates studied (black column - P1, white column - P5). Furthermore, podocyte specific expression was present in cells at P5 (C) (black - plastic, white - renal ECM) (D) (black - collagen I, white - collagen IV, gray - laminin, hatched - fibronectin). Passage 1 (P1) compared to passage 5 (P5) (*p

Techniques Used: Real-time Polymerase Chain Reaction, Expressing

Renal morphology and immunohistochemistry - Early third trimester fetal monkey kidney and nephrogenic zone (A) The fetal monkey kidney in the early third trimester shows a clear nephrogenic zone in the cortical region with the ureteric bud (arrowhead) and the metenephric mesenchyme (arrow) evident. The four stages of developing glomeruli are also present and represent the vesicle (a), S-shape (b), capillary (c), and mature (d) glomerulus. Hematoxylin and eosin (H E). (B-C) The nephrogenic zone is shown with the nuclei stained with DAPI (blue), metanephric mesenchyme (arrowheads), developing podocytes, and mature podocytes (arrows) positive for synaptopodin (green) and nestin (red). (D-I) Mature podocytes in terminally differentiated glomeruli are positive for nestin (red) and synaptopodin (green). (J-O) The endothelial marker CD31 (green) is not evident with the nestin positive cells (red) in these specimens (A-C, 20x, bar = 100 μm; D-F, 40x, bar = 50 μm; G-I, 100x, bar = 20 μm; J-L, 40x, bar = 50 μm; M-O, 100x, bar = 20 μm).
Figure Legend Snippet: Renal morphology and immunohistochemistry - Early third trimester fetal monkey kidney and nephrogenic zone (A) The fetal monkey kidney in the early third trimester shows a clear nephrogenic zone in the cortical region with the ureteric bud (arrowhead) and the metenephric mesenchyme (arrow) evident. The four stages of developing glomeruli are also present and represent the vesicle (a), S-shape (b), capillary (c), and mature (d) glomerulus. Hematoxylin and eosin (H E). (B-C) The nephrogenic zone is shown with the nuclei stained with DAPI (blue), metanephric mesenchyme (arrowheads), developing podocytes, and mature podocytes (arrows) positive for synaptopodin (green) and nestin (red). (D-I) Mature podocytes in terminally differentiated glomeruli are positive for nestin (red) and synaptopodin (green). (J-O) The endothelial marker CD31 (green) is not evident with the nestin positive cells (red) in these specimens (A-C, 20x, bar = 100 μm; D-F, 40x, bar = 50 μm; G-I, 100x, bar = 20 μm; J-L, 40x, bar = 50 μm; M-O, 100x, bar = 20 μm).

Techniques Used: Immunohistochemistry, Staining, Marker

8) Product Images from "The Expression Profile of Complement Components in Podocytes"

Article Title: The Expression Profile of Complement Components in Podocytes

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms17040471

Representative complement immunofluorescent staining in the PAN rat nephropathy model. Immunofluorescent staining of C3 ( A ), DAF ( C ), C3aR ( E ), C1q ( G ), and C3d ( I ), with synaptopodin in a PAN nephropathy rat model. Complement factors are labeled in green, and synaptopodin is labeled in red. Scale bar for overview: 20 μm; higher magnification: 30 μm. ( B , D , F , H ) Immunofluorescence staining of C3, C3aR, DAF, and C1q in glomeruli, the positive signals were quantified as the relative fluorescence intensity. Each column represents the mean value that was derived from four glomeruli in a single experiment. The results are presented as the mean value of three independent experiments; ( J ) Images J 1 – 6 are the negative controls by without primary antibodies; images J 1 , 2 , 4 , 5 were incubated with Alexa Fluor ®596 goat anti-mouse IgG, Alexa Fluor ®488 goat anti-rabbit IgG, Alexa Fluor ®596 goat anti-rabbit IgG, and Alexa Fluor ®488 goat anti-mouse IgG, respectively. Data are presented as means ± SD. * p
Figure Legend Snippet: Representative complement immunofluorescent staining in the PAN rat nephropathy model. Immunofluorescent staining of C3 ( A ), DAF ( C ), C3aR ( E ), C1q ( G ), and C3d ( I ), with synaptopodin in a PAN nephropathy rat model. Complement factors are labeled in green, and synaptopodin is labeled in red. Scale bar for overview: 20 μm; higher magnification: 30 μm. ( B , D , F , H ) Immunofluorescence staining of C3, C3aR, DAF, and C1q in glomeruli, the positive signals were quantified as the relative fluorescence intensity. Each column represents the mean value that was derived from four glomeruli in a single experiment. The results are presented as the mean value of three independent experiments; ( J ) Images J 1 – 6 are the negative controls by without primary antibodies; images J 1 , 2 , 4 , 5 were incubated with Alexa Fluor ®596 goat anti-mouse IgG, Alexa Fluor ®488 goat anti-rabbit IgG, Alexa Fluor ®596 goat anti-rabbit IgG, and Alexa Fluor ®488 goat anti-mouse IgG, respectively. Data are presented as means ± SD. * p

Techniques Used: Staining, Labeling, Immunofluorescence, Fluorescence, Derivative Assay, Incubation

9) Product Images from "Podocyte-Specific Overexpression of Wild Type or Mutant Trpc6 in Mice Is Sufficient to Cause Glomerular Disease"

Article Title: Podocyte-Specific Overexpression of Wild Type or Mutant Trpc6 in Mice Is Sufficient to Cause Glomerular Disease

Journal: PLoS ONE

doi: 10.1371/journal.pone.0012859

Protein expression in transgenic mice. (A) The in vivo Trpc6-HA expression was confirmed by Immunoprecipitation (IP). The IP was performed from isolated mouse glomeruli (n = 8 kidney from each genotype) as described in material and methods . The glomeruli fraction was loaded into an HA affinity column, and a 30 µl aliquot of the sample which did not bind into the column was loaded in a gel and stained with Coomassie brilliant blue as a loading control (effluent control) for transgenic and non transgenic samples. (B) 30 µl of each IP sample was run in a SDS PAGE and a Western blot analysis against HA epitope was performed. (C) Double immunofluorescence of kidney cryosections to detect synaptopodin (green) and HA (red) for every transgenic line utilized in this study (400x).
Figure Legend Snippet: Protein expression in transgenic mice. (A) The in vivo Trpc6-HA expression was confirmed by Immunoprecipitation (IP). The IP was performed from isolated mouse glomeruli (n = 8 kidney from each genotype) as described in material and methods . The glomeruli fraction was loaded into an HA affinity column, and a 30 µl aliquot of the sample which did not bind into the column was loaded in a gel and stained with Coomassie brilliant blue as a loading control (effluent control) for transgenic and non transgenic samples. (B) 30 µl of each IP sample was run in a SDS PAGE and a Western blot analysis against HA epitope was performed. (C) Double immunofluorescence of kidney cryosections to detect synaptopodin (green) and HA (red) for every transgenic line utilized in this study (400x).

Techniques Used: Expressing, Transgenic Assay, Mouse Assay, In Vivo, Immunoprecipitation, Isolation, Affinity Column, Staining, SDS Page, Western Blot, Immunofluorescence

10) Product Images from "Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation"

Article Title: Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation

Journal: Scientific Reports

doi: 10.1038/srep17694

Effects of CsA on podocyte WAVE1 expression in PAN-induced rat nephropathy. ( A ) Western blot analysis of WAVE1 in isolated glomeruli. ( B ) WAVE1 expression was quantified and normalized to GAPDH expression. ( C ) Immunofluorescent staining of WAVE1 and synaptopodin in rats. Scale bar = 20 μm. WAVE1 is labelled in green, and synaptopodin is labelled red. The data are presented as the mean ± SD. n = 5. **P
Figure Legend Snippet: Effects of CsA on podocyte WAVE1 expression in PAN-induced rat nephropathy. ( A ) Western blot analysis of WAVE1 in isolated glomeruli. ( B ) WAVE1 expression was quantified and normalized to GAPDH expression. ( C ) Immunofluorescent staining of WAVE1 and synaptopodin in rats. Scale bar = 20 μm. WAVE1 is labelled in green, and synaptopodin is labelled red. The data are presented as the mean ± SD. n = 5. **P

Techniques Used: Expressing, Western Blot, Isolation, Staining

The interaction between WAVE1 and calcineurin and WAVE1 phosphorylation in PAN-stimulated podocytes. ( A ) Co-immunoprecipitation analysis of the interaction between WAVE1 and calcineurin. Control IgG represents normal rabbit IgG (RIgG), which replaced the anti-calcineurin antibody in the precipitation process. ( B ) WAVE1 serine phosphorylation was detected by co-immunoprecipitation. Control IgG represents normal rabbit IgG, which replaced the anti-calcineurin antibody in the precipitation process. ( C ) WAVE1 serine phosphorylation was quantified based on the expression of total WAVE1 in the sample precipitated with the anti-calcineurin antibody. ( D ) Western blot of synaptopodin expression in WAVE1-overexpressing podocytes. ( E , F ) WAVE1 and synaptopodin expression levels were quantified after normalization to GAPDH expression. ( G ) Western blot of WAVE1 expression in synaptopodin-knockdown podocytes. ( H,I ) Synaptopodin and WAVE1 expression levels were quantified after normalization to GAPDH expression. The data are presented as the mean ± SD. n = 3. *P
Figure Legend Snippet: The interaction between WAVE1 and calcineurin and WAVE1 phosphorylation in PAN-stimulated podocytes. ( A ) Co-immunoprecipitation analysis of the interaction between WAVE1 and calcineurin. Control IgG represents normal rabbit IgG (RIgG), which replaced the anti-calcineurin antibody in the precipitation process. ( B ) WAVE1 serine phosphorylation was detected by co-immunoprecipitation. Control IgG represents normal rabbit IgG, which replaced the anti-calcineurin antibody in the precipitation process. ( C ) WAVE1 serine phosphorylation was quantified based on the expression of total WAVE1 in the sample precipitated with the anti-calcineurin antibody. ( D ) Western blot of synaptopodin expression in WAVE1-overexpressing podocytes. ( E , F ) WAVE1 and synaptopodin expression levels were quantified after normalization to GAPDH expression. ( G ) Western blot of WAVE1 expression in synaptopodin-knockdown podocytes. ( H,I ) Synaptopodin and WAVE1 expression levels were quantified after normalization to GAPDH expression. The data are presented as the mean ± SD. n = 3. *P

Techniques Used: Immunoprecipitation, Expressing, Western Blot

11) Product Images from "Schip1 Is a Novel Podocyte Foot Process Protein that Mediates Actin Cytoskeleton Rearrangements and Forms a Complex with Nherf2 and Ezrin"

Article Title: Schip1 Is a Novel Podocyte Foot Process Protein that Mediates Actin Cytoskeleton Rearrangements and Forms a Complex with Nherf2 and Ezrin

Journal: PLoS ONE

doi: 10.1371/journal.pone.0122067

Schip1 is expressed by glomerular podocytes. (A) RT-PCR shows SCHIP1 expression in both glomerulus and the kidney fraction lacking glomeruli. In the glomerulus, expression is mostly detected in FACS-sorted podocytes. As an internal control, expression levels of GAPDH were measured. Controls for markers of various fractions are presented in S3 Fig. (Pod-podocytes, ROG-rest of glomerulus, GLOM-glomerulus, ROK-rest of kidney). (B) Northern blotting on a mouse multiple tissue panel shows the presence of two SCHIP1 mRNA transcripts enriched in the brain, heart, testes and kidney tissues. (C) Northern blotting on mouse glomerulus and ROK tissue confirms stronger SCHIP1 expression in the glomerulus, and presence of two transcripts. (D) By radioactive in situ hybridization on newborn mouse kidney sections SCHIP1 mRNA is localized to developing podocytes of the capillary loop stage glomerulus. (E) By Western blotting, the mouse 55kDa Schip1 protein is detected mostly in the glomerulus. Podocin was used as a positive control for the glomerular fraction, β-actin as a loading control. (F) Immunofluorescence on mouse and human kidney sections shows Schip1 glomerulus signal that partially overlaps with a podocyte foot process marker synaptopodin (Synpo). (G) By immunoelectron microscopy, Schip1 localizes to the glomerular podocyte foot processes (FP) in human kidney sections (GBM-glomerular basement membrane).
Figure Legend Snippet: Schip1 is expressed by glomerular podocytes. (A) RT-PCR shows SCHIP1 expression in both glomerulus and the kidney fraction lacking glomeruli. In the glomerulus, expression is mostly detected in FACS-sorted podocytes. As an internal control, expression levels of GAPDH were measured. Controls for markers of various fractions are presented in S3 Fig. (Pod-podocytes, ROG-rest of glomerulus, GLOM-glomerulus, ROK-rest of kidney). (B) Northern blotting on a mouse multiple tissue panel shows the presence of two SCHIP1 mRNA transcripts enriched in the brain, heart, testes and kidney tissues. (C) Northern blotting on mouse glomerulus and ROK tissue confirms stronger SCHIP1 expression in the glomerulus, and presence of two transcripts. (D) By radioactive in situ hybridization on newborn mouse kidney sections SCHIP1 mRNA is localized to developing podocytes of the capillary loop stage glomerulus. (E) By Western blotting, the mouse 55kDa Schip1 protein is detected mostly in the glomerulus. Podocin was used as a positive control for the glomerular fraction, β-actin as a loading control. (F) Immunofluorescence on mouse and human kidney sections shows Schip1 glomerulus signal that partially overlaps with a podocyte foot process marker synaptopodin (Synpo). (G) By immunoelectron microscopy, Schip1 localizes to the glomerular podocyte foot processes (FP) in human kidney sections (GBM-glomerular basement membrane).

Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing, FACS, Northern Blot, In Situ Hybridization, Western Blot, Positive Control, Immunofluorescence, Marker, Immuno-Electron Microscopy

12) Product Images from "An Essential Role of the Universal Polarity Protein, aPKC?, on the Maintenance of Podocyte Slit Diaphragms"

Article Title: An Essential Role of the Universal Polarity Protein, aPKC?, on the Maintenance of Podocyte Slit Diaphragms

Journal: PLoS ONE

doi: 10.1371/journal.pone.0004194

Selective depletion of aPKCλ from mouse podocytes causes FSGS. Periodic acid-Schiff staining (PAS) and immunohistochemistry for nephrin and synaptopodin in mutant (cKO) and control kidneys at P0, P10 and P21 show that mutant mice develop segmental to global glomerulosclerosis. Boxed regions are enlarged to show an irregular pattern of nephrin staining in mutant podocytes. Loss of podocytes (arrows) and occasional adhesion of glomeruli to Bowman's capsules (arrowheads) reveal the development of focal segmental glomerulosclerosis. Consistent with massive proteinuria in mutant mice at birth, PAS staining reveals occasional hyaline droplets, representing reabsorbed urinary protein, in the proximal renal tubules at P0. Bars, 50 µm.
Figure Legend Snippet: Selective depletion of aPKCλ from mouse podocytes causes FSGS. Periodic acid-Schiff staining (PAS) and immunohistochemistry for nephrin and synaptopodin in mutant (cKO) and control kidneys at P0, P10 and P21 show that mutant mice develop segmental to global glomerulosclerosis. Boxed regions are enlarged to show an irregular pattern of nephrin staining in mutant podocytes. Loss of podocytes (arrows) and occasional adhesion of glomeruli to Bowman's capsules (arrowheads) reveal the development of focal segmental glomerulosclerosis. Consistent with massive proteinuria in mutant mice at birth, PAS staining reveals occasional hyaline droplets, representing reabsorbed urinary protein, in the proximal renal tubules at P0. Bars, 50 µm.

Techniques Used: Staining, Immunohistochemistry, Mutagenesis, Mouse Assay

13) Product Images from "Characterization of a Trpc6 Transgenic Mouse Associated with Early Onset FSGS"

Article Title: Characterization of a Trpc6 Transgenic Mouse Associated with Early Onset FSGS

Journal: British journal of medicine and medical research

doi: 10.9734/bjmmr/2015/12493

Molecular characterization of the Trpc6-M131T transgenic line (A)Scheme of the microinjected transgene Trpc6- M131T and its comparison with the wild type allele. The complete Trpc6-M131T cDNA were subcloned downstream the pNPHS2 podocin promoter as previously described [ 21 ]. (B) Relative mRNA Trpc6/Gapdh expression levels in glomeruli were determined by real-time PCR. Values represent mean +/− SEM; n=5. (C) Immunofluorescence in kidney cryosections showing podocyte-specific Trpc6-M131T transgen expression. A podocyte specific marker, Synaptopodin (green) was used as control to determine HA (red) HA (400x)
Figure Legend Snippet: Molecular characterization of the Trpc6-M131T transgenic line (A)Scheme of the microinjected transgene Trpc6- M131T and its comparison with the wild type allele. The complete Trpc6-M131T cDNA were subcloned downstream the pNPHS2 podocin promoter as previously described [ 21 ]. (B) Relative mRNA Trpc6/Gapdh expression levels in glomeruli were determined by real-time PCR. Values represent mean +/− SEM; n=5. (C) Immunofluorescence in kidney cryosections showing podocyte-specific Trpc6-M131T transgen expression. A podocyte specific marker, Synaptopodin (green) was used as control to determine HA (red) HA (400x)

Techniques Used: Transgenic Assay, Expressing, Real-time Polymerase Chain Reaction, Immunofluorescence, Marker

14) Product Images from "Novel Microscopic Techniques for Podocyte Research"

Article Title: Novel Microscopic Techniques for Podocyte Research

Journal: Frontiers in Endocrinology

doi: 10.3389/fendo.2018.00379

Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using anti-synaptopodin and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.
Figure Legend Snippet: Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using anti-synaptopodin and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.

Techniques Used: Labeling, Staining

15) Product Images from "Albuminuria and Glomerular Damage in Mice Lacking the Metabotropic Glutamate Receptor 1"

Article Title: Albuminuria and Glomerular Damage in Mice Lacking the Metabotropic Glutamate Receptor 1

Journal: The American Journal of Pathology

doi: 10.1016/j.ajpath.2010.11.050

Podocyte proteins in 8-month-old animals. Immunofluorescence displays diffuse progressive loss of the podocyte markers nephrin ( B and C ) and synaptopodin ( E and F ) and segmental loss of podocin ( H and I ) and ZO-1 ( K and L ) in glomeruli of 8 month-old
Figure Legend Snippet: Podocyte proteins in 8-month-old animals. Immunofluorescence displays diffuse progressive loss of the podocyte markers nephrin ( B and C ) and synaptopodin ( E and F ) and segmental loss of podocin ( H and I ) and ZO-1 ( K and L ) in glomeruli of 8 month-old

Techniques Used: Immunofluorescence

16) Product Images from "A Homozygous Missense Mutation in the Ciliary Gene TTC21B Causes Familial FSGS"

Article Title: A Homozygous Missense Mutation in the Ciliary Gene TTC21B Causes Familial FSGS

Journal: Journal of the American Society of Nephrology : JASN

doi: 10.1681/ASN.2013101126

IFT139, expressed at the base of primary cilium in tubules, fetal and undifferentiated podocytes, relocalizes along the microtubule network in mature and differentiated podocytes. (A) Immunolocalization of IFT139 in adult control human kidney using lotus tetragonolobus (LTA) and peanut agglutinin lectin (PNA) to identify proximal and distal tubules, respectively, revealed that IFT139 is expressed predominantly in distal tubules and glomeruli. Scale bar: 50 µ m. (B) Western blot showing expression of IFT139 in total kidney and isolated glomeruli or tubules from adult wild-type mice of mixed background. (C) Immunolabeling of IFT139; acetylated α -tubulin, a ciliary component; and nephrin, a podocyte marker, in fetal (14 weeks) and mature kidneys. IFT139 is expressed in developing glomeruli, notably at the base of primary cilia (arrow). In contrast, we observed a rearrangement of the acetylated α -tubulin network in mature glomeruli, and IFT139 colocalizes with this network. Scale bar: 50 µ m. (D) Immunolocalization of IFT139 in podocytes after 3 and 14 days of differentiation (D-3 and D-14). IFT139 localizes at the base of the cilium in undifferentiated podocytes and is redistributed along the acetylated α -tubulin network in differentiated podocytes, labeled by synaptopodin. Scale bar: 20 µ m. (E) Percentage of ciliated cells in cultured podocytes demonstrated that differentiated podocytes lose their primary cilium. For statistical analysis, four independent experimentations were performed. A t test was conducted; the mean±SEM is shown (* P
Figure Legend Snippet: IFT139, expressed at the base of primary cilium in tubules, fetal and undifferentiated podocytes, relocalizes along the microtubule network in mature and differentiated podocytes. (A) Immunolocalization of IFT139 in adult control human kidney using lotus tetragonolobus (LTA) and peanut agglutinin lectin (PNA) to identify proximal and distal tubules, respectively, revealed that IFT139 is expressed predominantly in distal tubules and glomeruli. Scale bar: 50 µ m. (B) Western blot showing expression of IFT139 in total kidney and isolated glomeruli or tubules from adult wild-type mice of mixed background. (C) Immunolabeling of IFT139; acetylated α -tubulin, a ciliary component; and nephrin, a podocyte marker, in fetal (14 weeks) and mature kidneys. IFT139 is expressed in developing glomeruli, notably at the base of primary cilia (arrow). In contrast, we observed a rearrangement of the acetylated α -tubulin network in mature glomeruli, and IFT139 colocalizes with this network. Scale bar: 50 µ m. (D) Immunolocalization of IFT139 in podocytes after 3 and 14 days of differentiation (D-3 and D-14). IFT139 localizes at the base of the cilium in undifferentiated podocytes and is redistributed along the acetylated α -tubulin network in differentiated podocytes, labeled by synaptopodin. Scale bar: 20 µ m. (E) Percentage of ciliated cells in cultured podocytes demonstrated that differentiated podocytes lose their primary cilium. For statistical analysis, four independent experimentations were performed. A t test was conducted; the mean±SEM is shown (* P

Techniques Used: Western Blot, Expressing, Isolation, Mouse Assay, Immunolabeling, Marker, Labeling, Cell Culture

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    Progen Biotechnik primary mouse monoclonal anti synaptopodin
    Double immunofluorescence staining of kidney sections from Wistar rats. Immunofluorescence staining of glomeruli: (A) anti-meprinβ antibody, (B) <t>anti-synaptopodin</t> antibody, (C) merged image meprin/synaptopodin, (D) anti-meprinβ antibody, (E) anti-Thy 1.1 antibody, (F) merged image meprin/Thy 1.1. Nuclei are marked by staining with 4,6-diamidino-2-phenylindole (DAPI).
    Primary Mouse Monoclonal Anti Synaptopodin, supplied by Progen Biotechnik, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Progen Biotechnik mouse anti synaptopodin
    Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using <t>anti-synaptopodin</t> and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.
    Mouse Anti Synaptopodin, supplied by Progen Biotechnik, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti synaptopodin/product/Progen Biotechnik
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti synaptopodin - by Bioz Stars, 2022-09
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    Progen Biotechnik synaptopodin antibody
    MES+HS activates Akt and induces Hsp72 expression in podocytes of Alport mice in vivo. ( a , c e ) One hour after the final treatment, whole kidney protein lysates or glomeruli protein lysates were isolated from 8 weeks-MES+HS-treated and control Alport mice, and immunoblotted to detect p-Akt and total Akt proteins. Actin was used as loading control ( a, c ). Renal cryosections were subjected to immunohistochemical analysis to assess phosphorylated Akt. Arrowheads indicate phosphorylated Akt in podocytes. Cryosections were counterstained with DAPI ( e ). ( b, d f ) Eight hours ( b f ) or five hours ( d ) after the final treatment, whole kidney or glomeruli protein lysates were isolated from MES+HS-treated (8 weeks treated for b f , or 4 weeks treated for d ) and control Alport mice, and immunoblotted to assess Hsp72 and HSC70 expression. α-Tubulin or actin was used as loading control ( b, d ). Renal cryosections were subjected to immunohistochemical analysis to assess Hsp72. Arrowheads indicate up-regulated Hsp72 in podocytes. Cryosections were counterstained with DAPI ( f ). ( e f ) Representative glomeruli images of three independent experiments are shown. Renal sections were immunostained with <t>anti-synaptopodin</t> antibody to assess podocyte localization ( lower panels ). Sections were counterstained with DAPI. White bars; 10 µm.
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    Double immunofluorescence staining of kidney sections from Wistar rats. Immunofluorescence staining of glomeruli: (A) anti-meprinβ antibody, (B) anti-synaptopodin antibody, (C) merged image meprin/synaptopodin, (D) anti-meprinβ antibody, (E) anti-Thy 1.1 antibody, (F) merged image meprin/Thy 1.1. Nuclei are marked by staining with 4,6-diamidino-2-phenylindole (DAPI).

    Journal: PLoS ONE

    Article Title: Metalloprotease Meprin? in Rat Kidney: Glomerular Localization and Differential Expression in Glomerulonephritis

    doi: 10.1371/journal.pone.0002278

    Figure Lengend Snippet: Double immunofluorescence staining of kidney sections from Wistar rats. Immunofluorescence staining of glomeruli: (A) anti-meprinβ antibody, (B) anti-synaptopodin antibody, (C) merged image meprin/synaptopodin, (D) anti-meprinβ antibody, (E) anti-Thy 1.1 antibody, (F) merged image meprin/Thy 1.1. Nuclei are marked by staining with 4,6-diamidino-2-phenylindole (DAPI).

    Article Snippet: We used the following antibodies: primary rabbit polyclonal anti-meprinβ, targeting amino acids 468–612 of meprinβ, as previously described , primary mouse monoclonal anti-synaptopodin (Progen, Heidelberg, Germany), primary monoclonal mouse anti-Thy 1.1 (Oxford Biotechology, UK), secondary goat anti-rabbit and goat anti-mouse (Dako Cytomation, Denmark), diluted 1:500, 1:4, 1:20, 1:1000 and 1:400 respectively.

    Techniques: Double Immunofluorescence Staining, Immunofluorescence, Staining

    Meprinβ expression in rat glomeruli after induction of Passive Heyman nephritis (PHN). Immunofluorescence staining (A–C) of meprinβ and (D–F) synaptopodin in kidney sections from Sprague-Dawley rats. (G–I) Merged images of meprin/synaptopodin. (J–L) Immunohistochemical staining of meprinβ in glomeruli and in proximal tubules. Staining (A, D, G and J) in control rats, (B, E, H and K) in rats at day 3 and (C, F, I and L) in rats at day 6 after induction of PHN. A change in distribution of the meprinβ in PHN kidneys from a linear to a granular appearance was observed concomitantly associated with an overall reduction in signal intensity. The distribution of meprinβ and synaptopodin in glomeruli appeared increasingly divergent following progression of PHN.

    Journal: PLoS ONE

    Article Title: Metalloprotease Meprin? in Rat Kidney: Glomerular Localization and Differential Expression in Glomerulonephritis

    doi: 10.1371/journal.pone.0002278

    Figure Lengend Snippet: Meprinβ expression in rat glomeruli after induction of Passive Heyman nephritis (PHN). Immunofluorescence staining (A–C) of meprinβ and (D–F) synaptopodin in kidney sections from Sprague-Dawley rats. (G–I) Merged images of meprin/synaptopodin. (J–L) Immunohistochemical staining of meprinβ in glomeruli and in proximal tubules. Staining (A, D, G and J) in control rats, (B, E, H and K) in rats at day 3 and (C, F, I and L) in rats at day 6 after induction of PHN. A change in distribution of the meprinβ in PHN kidneys from a linear to a granular appearance was observed concomitantly associated with an overall reduction in signal intensity. The distribution of meprinβ and synaptopodin in glomeruli appeared increasingly divergent following progression of PHN.

    Article Snippet: We used the following antibodies: primary rabbit polyclonal anti-meprinβ, targeting amino acids 468–612 of meprinβ, as previously described , primary mouse monoclonal anti-synaptopodin (Progen, Heidelberg, Germany), primary monoclonal mouse anti-Thy 1.1 (Oxford Biotechology, UK), secondary goat anti-rabbit and goat anti-mouse (Dako Cytomation, Denmark), diluted 1:500, 1:4, 1:20, 1:1000 and 1:400 respectively.

    Techniques: Expressing, Immunofluorescence, Staining, Immunohistochemistry

    Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using anti-synaptopodin and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.

    Journal: Frontiers in Endocrinology

    Article Title: Novel Microscopic Techniques for Podocyte Research

    doi: 10.3389/fendo.2018.00379

    Figure Lengend Snippet: Dual color 3D-SIM of rodent (A) in mouse and (B) in rat) paraffin kidney sections shows the interdigitating foot process pattern of podocytes. Foot processes and the slit diaphragm were labeled using anti-synaptopodin and anti-nephrin antibodies, respectively. Although both proteins are in close structural vicinity the individual staining patterns do not overlap and therefore demonstrates the high resolution accomplished by 3D-SIM. All scale bars represent 1 μm.

    Article Snippet: Sections where dehydrated, antigen retrieved by pressure-cooking in citrate buffer, blocked in 1% FBS, 1% BSA, 1% NGS, 0.1% cold fish gelatin in PBS and co-labeled with guinea pig anti-nephrin (GP-N2, 1:300) and mouse anti-synaptopodin (GP94-C, 1:50, both Progen, Heidelberg, Germany) overnight at 4°C.

    Techniques: Labeling, Staining

    Effects of CsA on podocyte WAVE1 expression in PAN-induced rat nephropathy. ( A ) Western blot analysis of WAVE1 in isolated glomeruli. ( B ) WAVE1 expression was quantified and normalized to GAPDH expression. ( C ) Immunofluorescent staining of WAVE1 and synaptopodin in rats. Scale bar = 20 μm. WAVE1 is labelled in green, and synaptopodin is labelled red. The data are presented as the mean ± SD. n = 5. **P

    Journal: Scientific Reports

    Article Title: Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation

    doi: 10.1038/srep17694

    Figure Lengend Snippet: Effects of CsA on podocyte WAVE1 expression in PAN-induced rat nephropathy. ( A ) Western blot analysis of WAVE1 in isolated glomeruli. ( B ) WAVE1 expression was quantified and normalized to GAPDH expression. ( C ) Immunofluorescent staining of WAVE1 and synaptopodin in rats. Scale bar = 20 μm. WAVE1 is labelled in green, and synaptopodin is labelled red. The data are presented as the mean ± SD. n = 5. **P

    Article Snippet: The following primary antibodies were used: rabbit anti-WAVE1 (1:200; Abcam, UK) and mouse anti-synaptopodin (1:100; PROGEN, USA).

    Techniques: Expressing, Western Blot, Isolation, Staining

    The interaction between WAVE1 and calcineurin and WAVE1 phosphorylation in PAN-stimulated podocytes. ( A ) Co-immunoprecipitation analysis of the interaction between WAVE1 and calcineurin. Control IgG represents normal rabbit IgG (RIgG), which replaced the anti-calcineurin antibody in the precipitation process. ( B ) WAVE1 serine phosphorylation was detected by co-immunoprecipitation. Control IgG represents normal rabbit IgG, which replaced the anti-calcineurin antibody in the precipitation process. ( C ) WAVE1 serine phosphorylation was quantified based on the expression of total WAVE1 in the sample precipitated with the anti-calcineurin antibody. ( D ) Western blot of synaptopodin expression in WAVE1-overexpressing podocytes. ( E , F ) WAVE1 and synaptopodin expression levels were quantified after normalization to GAPDH expression. ( G ) Western blot of WAVE1 expression in synaptopodin-knockdown podocytes. ( H,I ) Synaptopodin and WAVE1 expression levels were quantified after normalization to GAPDH expression. The data are presented as the mean ± SD. n = 3. *P

    Journal: Scientific Reports

    Article Title: Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation

    doi: 10.1038/srep17694

    Figure Lengend Snippet: The interaction between WAVE1 and calcineurin and WAVE1 phosphorylation in PAN-stimulated podocytes. ( A ) Co-immunoprecipitation analysis of the interaction between WAVE1 and calcineurin. Control IgG represents normal rabbit IgG (RIgG), which replaced the anti-calcineurin antibody in the precipitation process. ( B ) WAVE1 serine phosphorylation was detected by co-immunoprecipitation. Control IgG represents normal rabbit IgG, which replaced the anti-calcineurin antibody in the precipitation process. ( C ) WAVE1 serine phosphorylation was quantified based on the expression of total WAVE1 in the sample precipitated with the anti-calcineurin antibody. ( D ) Western blot of synaptopodin expression in WAVE1-overexpressing podocytes. ( E , F ) WAVE1 and synaptopodin expression levels were quantified after normalization to GAPDH expression. ( G ) Western blot of WAVE1 expression in synaptopodin-knockdown podocytes. ( H,I ) Synaptopodin and WAVE1 expression levels were quantified after normalization to GAPDH expression. The data are presented as the mean ± SD. n = 3. *P

    Article Snippet: The following primary antibodies were used: rabbit anti-WAVE1 (1:200; Abcam, UK) and mouse anti-synaptopodin (1:100; PROGEN, USA).

    Techniques: Immunoprecipitation, Expressing, Western Blot

    MES+HS activates Akt and induces Hsp72 expression in podocytes of Alport mice in vivo. ( a , c e ) One hour after the final treatment, whole kidney protein lysates or glomeruli protein lysates were isolated from 8 weeks-MES+HS-treated and control Alport mice, and immunoblotted to detect p-Akt and total Akt proteins. Actin was used as loading control ( a, c ). Renal cryosections were subjected to immunohistochemical analysis to assess phosphorylated Akt. Arrowheads indicate phosphorylated Akt in podocytes. Cryosections were counterstained with DAPI ( e ). ( b, d f ) Eight hours ( b f ) or five hours ( d ) after the final treatment, whole kidney or glomeruli protein lysates were isolated from MES+HS-treated (8 weeks treated for b f , or 4 weeks treated for d ) and control Alport mice, and immunoblotted to assess Hsp72 and HSC70 expression. α-Tubulin or actin was used as loading control ( b, d ). Renal cryosections were subjected to immunohistochemical analysis to assess Hsp72. Arrowheads indicate up-regulated Hsp72 in podocytes. Cryosections were counterstained with DAPI ( f ). ( e f ) Representative glomeruli images of three independent experiments are shown. Renal sections were immunostained with anti-synaptopodin antibody to assess podocyte localization ( lower panels ). Sections were counterstained with DAPI. White bars; 10 µm.

    Journal: PLoS ONE

    Article Title: Mild Electrical Stimulation and Heat Shock Ameliorates Progressive Proteinuria and Renal Inflammation in Mouse Model of Alport Syndrome

    doi: 10.1371/journal.pone.0043852

    Figure Lengend Snippet: MES+HS activates Akt and induces Hsp72 expression in podocytes of Alport mice in vivo. ( a , c e ) One hour after the final treatment, whole kidney protein lysates or glomeruli protein lysates were isolated from 8 weeks-MES+HS-treated and control Alport mice, and immunoblotted to detect p-Akt and total Akt proteins. Actin was used as loading control ( a, c ). Renal cryosections were subjected to immunohistochemical analysis to assess phosphorylated Akt. Arrowheads indicate phosphorylated Akt in podocytes. Cryosections were counterstained with DAPI ( e ). ( b, d f ) Eight hours ( b f ) or five hours ( d ) after the final treatment, whole kidney or glomeruli protein lysates were isolated from MES+HS-treated (8 weeks treated for b f , or 4 weeks treated for d ) and control Alport mice, and immunoblotted to assess Hsp72 and HSC70 expression. α-Tubulin or actin was used as loading control ( b, d ). Renal cryosections were subjected to immunohistochemical analysis to assess Hsp72. Arrowheads indicate up-regulated Hsp72 in podocytes. Cryosections were counterstained with DAPI ( f ). ( e f ) Representative glomeruli images of three independent experiments are shown. Renal sections were immunostained with anti-synaptopodin antibody to assess podocyte localization ( lower panels ). Sections were counterstained with DAPI. White bars; 10 µm.

    Article Snippet: Synaptopodin antibody was from PROGEN.

    Techniques: Expressing, Mouse Assay, In Vivo, Isolation, Immunohistochemistry