rasal1 polyclonal antibody  (Bioss)

Journal: Kidney international

Article Title: Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

doi: 10.1016/j.kint.2016.07.042

Figure Lengend Snippet: Figure 1 | Aberrant Rasal1 promoter methylation is an epigenetic marker of AKI-to-CKD progression and is not present in AKI with effective recovery from injury. (a) Mice were challenged with either moderate or severe ischemia-reperfusion injury (IRI), representative photomicrographs of hematoxylin and eosin (HE), periodic acid–Schiff (PAS), Masson’s trichrome–stained (MTS; bars ¼ 100 mm) kidney sections, and sections immunolabeled with primary antibodies against Collagen-1, a-smooth muscle actin (a-SMA), and Ki67 (bars ¼ 25 mm) (Continued)

Article Snippet: Sections were stained using polyclonal antibody against Rasal1 (Biorbyt, Cambridge, UK), peroxidase labeling was performed using the Vectastain Universal Elite ABC Kit (Vector Laboratories).

Techniques: Methylation, Marker, Staining, Immunolabeling

Journal: Kidney international

Article Title: Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

doi: 10.1016/j.kint.2016.07.042

Figure Lengend Snippet: Figure 2 | Transgenic RASAL1 overexpression protects from loss of endogenous Rasal1 and attenuates AKI-to-CKD progression. (a,b) As analyzed by immunostaining (bars ¼ 50 mm), administration of doxycycline (þDOX) to rtTAhCMV;hRASAL1-pTreTight mice (referred as hRASAL1oe) results in robust induction of transgenic RASAL1 in interstitial compartments (n of mice as indicated, data are presented as means SD, **P < 0.01, ****P < 0.0001, P values were calculated using 1-way analysis of variance with Bonferroni post hoc analysis (Continued)

Article Snippet: Sections were stained using polyclonal antibody against Rasal1 (Biorbyt, Cambridge, UK), peroxidase labeling was performed using the Vectastain Universal Elite ABC Kit (Vector Laboratories).

Techniques: Transgenic Assay, Over Expression, Immunostaining

Journal: Kidney international

Article Title: Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

doi: 10.1016/j.kint.2016.07.042

Figure Lengend Snippet: Figure 4 | Attenuation of AKI-to-CKD progression by low-dose hydralazine is associated with normalization of Rasal1 promoter methylation and restoration of intrarenal Rasal1 expression. (a) Analyzed by quantitative reverse-transcription polymerase chain reaction (PCR), administration of low-dose hydralazine induced intrarenal mRNA expression of ten eleven translocation 3 (Tet3, n of mice as (Continued)

Article Snippet: Sections were stained using polyclonal antibody against Rasal1 (Biorbyt, Cambridge, UK), peroxidase labeling was performed using the Vectastain Universal Elite ABC Kit (Vector Laboratories).

Techniques: Methylation, Expressing, Reverse Transcription, Polymerase Chain Reaction, Translocation Assay

Journal: Kidney international

Article Title: Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

doi: 10.1016/j.kint.2016.07.042

Figure Lengend Snippet: Figure 5 | Low-dose hydralazine rescues Rasal1 expression in the context of aberrant Rasal1 promoter methylation. (a–d) We induced Rasal1 promoter methylation in primary fibroblast cultures by exposing cells to transforming growth factor (TGF)-b1 for 5 days. After removal of TGF-b1, vehicle dimethylsulfoxide, hydralazine, or ramipril were added to culture media for an additional 1 day to elucidate their potential demethylating activities. The pictures show virtual gel images of Rasal1 polymerase chain reaction products of captured (Continued)

Article Snippet: Sections were stained using polyclonal antibody against Rasal1 (Biorbyt, Cambridge, UK), peroxidase labeling was performed using the Vectastain Universal Elite ABC Kit (Vector Laboratories).

Techniques: Expressing, Methylation, Polymerase Chain Reaction

Journal: Kidney international

Article Title: Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

doi: 10.1016/j.kint.2016.07.042

Figure Lengend Snippet: Figure 6 | Low-dose hydralazine attenuates AKI-to-CKD progression in already established kidney injury. (a) Mice underwent severe ischemia-reperfusion injury (IRI) and received either daily intraperitoneal injections of vehicle dimethylsulfoxide (DMSO) or low-dose hydralazine (5 mg/kg/d) starting 3 days after surgery in established kidney injury, contralateral kidneys were removed 35 days after severe IRI. (b,c) The pictures show virtual gel images of Rasal1 polymerase chain reaction products of captured methylation by DNA (Continued)

Article Snippet: Sections were stained using polyclonal antibody against Rasal1 (Biorbyt, Cambridge, UK), peroxidase labeling was performed using the Vectastain Universal Elite ABC Kit (Vector Laboratories).

Techniques: Polymerase Chain Reaction, Methylation

Journal: Kidney international

Article Title: Low-dose hydralazine prevents fibrosis in a murine model of acute kidney injury-to-chronic kidney disease progression.

doi: 10.1016/j.kint.2016.07.042

Figure Lengend Snippet: Figure 7 | Aberrant RASAL1 promoter methylation can be targeted therapeutically by low-dose hydralazine in human fibroblast cultures. (a) We induced activation of human fibroblast cultures by prolonged exposure to transforming growth factor (TGF)-b1 for 5 days. As analyzed by quantitative reverse-transcription polymerase chain reaction (PCR), low-dose hydralazine induced TET3 mRNA expression levels (n of independent experiments as indicated, measurements were performed in technical triplicates, data are presented as means SD, ****P < 0.0001, P values were calculated using 1-way analysis of variance with Bonferroni post hoc analysis comparing the indicated pairs of columns). (b,c) Representative photomicrographs of TET3 immunostainings overlayed with differential interference contrast (DIC; bars ¼ 25 mm) and 40,6-diamidino-2-phenylindole (DAPI; bars ¼ 5 mm) are shown. Low-dose hydralazine robustly induced Tet3 expression levels in primary fibroblast cultures (n of independent replicates as indicated, data are presented as means SD, ****P < 0.0001, P values were calculated using 1-way analysis of variance with Bonferroni post hoc analysis comparing the indicated pairs of columns). (d–g) The pictures show virtual gel images of RASAL1 PCR products of captured methylation (MeDIP), hydroxymethylation (hMeDIP), and carboxylation (CaDIP) by DNA immu- noprecipitation and corresponding input DNA (dilution 1:20, as controls for equal loading in immunoprecipitation). The bar graphs summarize the average RASAL1 DNA concentrations relative to input DNA. Although exposure to TGF-b1 alone was associated with robust RASAL1 pro- moter methylation, the addition of low-dose hydralazine normalized RASAL1 promoter methylation involving RASAL1 promoter hydrox- ymethylation and carboxylation (n of independent experiments as indicated, data are presented as means SD, *P < 0.05, **P < 0.01, #no significance, P values were calculated using 1-way analysis of variance with Bonferroni post hoc analysis comparing the indicated pairs of columns). (h) Analyzed by quantitative reverse-transcription PCR, RASAL1 mRNA expression levels observed as repressed in response to TGF-b1 were normalized by low-dose hydralazine (n of independent experiments as indicated, measurements were performed in technical triplicates, data are presented as means SD, ****P < 0.0001, #no significance, P values were calculated using 1-way analysis of variance with Bonferroni post hoc analysis comparing the indicated pairs of columns). Boxed portions correspond to a higher magnification view.

Article Snippet: Sections were stained using polyclonal antibody against Rasal1 (Biorbyt, Cambridge, UK), peroxidase labeling was performed using the Vectastain Universal Elite ABC Kit (Vector Laboratories).

Techniques: Methylation, Activation Assay, Reverse Transcription, Polymerase Chain Reaction, Expressing, Methylated DNA Immunoprecipitation, Immunoprecipitation