Journal: Molecular Metabolism

Article Title: P2Y2R contributes to the development of diabetic nephropathy by inhibiting autophagy response

doi: 10.1016/j.molmet.2020.101089

Figure Lengend Snippet: Antibodies used in this study.

Article Snippet: Wilms Tumor-1 (WT1) , Boster Biological Technology , #M00199-1 , Rabbit , 1 μg/ml (IHC).

Techniques:

Journal: Molecular Metabolism

Article Title: P2Y2R contributes to the development of diabetic nephropathy by inhibiting autophagy response

doi: 10.1016/j.molmet.2020.101089

Figure Lengend Snippet: P2Y2R deficiency protects against podocyte loss and glomerular injury in DN. (A) Relative mRNA levels of Nphs1 (nephrin) and Nphs2 (podocin) were determined by real-time PCR analysis (n = 3–5). (B) Localisation of WT1, a podocyte marker, was analysed by immunohistochemistry, and representative images are shown. The number of stained podocytes per glomeruli was counted by using ImageJ software (n = 3). (C) Kidney sections were stained with PAS staining and glomerular morphological changes were scored as described in the method section (n = 3). Data are presented as mean ± SEM. One-way ANOVA was used for statistical analysis followed by Bonferroni's multiple comparisons test. ∗∗∗p < 0.001 vs WT control mice; and ### p < 0.001 vs WT DN mice. Scale bar, 50 μm.

Article Snippet: Wilms Tumor-1 (WT1) , Boster Biological Technology , #M00199-1 , Rabbit , 1 μg/ml (IHC).

Techniques: Real-time Polymerase Chain Reaction, Marker, Immunohistochemistry, Staining, Software, Control

Journal: Journal of Biomedical Science

Article Title: ADAM10 is expressed in human podocytes and found in urinary vesicles of patients with glomerular kidney diseases

doi: 10.1186/1423-0127-17-3

Figure Lengend Snippet: Podocytes isolated out of the urine of patients with nephrotic syndrome express ADAM10 . Cells isolated out of the urine of a patient with nephrotic syndrome were analyzed by flow cytometry (A+E) , Western blot (B+C) , RT-PCR (C lower panel) , and immunofluorescence (D+F) . (A) Cells isolated from the urine were stained with ADAM10 or L1 adhesion molecule and analyzed with Cellquest software from Becton Dickinson (Heidelberg, Germany). (B) Urinary cells were lysed and western blots (WB) with ADAM10 and L1 (L1 11A) specific antibodies were performed. (C) Lower panel: RT-PCR with α3, β1, and podocin specific primers on cDNA of cells isolated from the urine. Upper panel: Western blot analysis with α3, β1 and podocin specific antibodies in lysats of cells isolated from the urine. (D) Immunofluorescence double staining of cells isolated from the urine with podocyte specific marker proteins α3, nephrin, podocyin antibodies. Images were documented with a Zeiss camera. (E) Urinary cells were investigated by intracellular FACS staining using WT1 (podocyte specific marker protein) and ADAM10 antibodies. Stained cells were analyzed with Cellquest software from Becton Dickinson (Heidelberg, Germany). (F) Immunofluorescence staining of untreated (control) and IFN-γ treated urinary podocytes with L1 specific primary antibodies followed by Alexa 488 coupled secondary antibodies. Nuclei of urinary podocytes were stained and visualized with DAPI. Images were documented with a Zeiss camera. (G) Glomeruli from human kidney were isolated and glomerular lysats were prepared, proteins were loaded on a SDS gel and western blot analysis were performed using a polyclonal antibody against the cytoplasmic tail of L1.

Article Snippet: WT1 antibody for immunofluorescence analysis was purchased from Santa Cruz (Heidelberg, Germany).

Techniques: Isolation, Flow Cytometry, Western Blot, Reverse Transcription Polymerase Chain Reaction, Immunofluorescence, Staining, Software, Double Staining, Marker, Control, SDS-Gel

Journal: Journal of Biomedical Science

Article Title: ADAM10 is expressed in human podocytes and found in urinary vesicles of patients with glomerular kidney diseases

doi: 10.1186/1423-0127-17-3

Figure Lengend Snippet: ADAM10 is expressed in podocytes in human renal tissue . (A) Glomeruli from human kidney were isolated and lysed and investigated by an ADAM10 specific westernblot (left panel). Right panel, double immunofluorescence analysis on a human kidney section with WT1 (red) and ADAM10 (green) antibodies, demonstrating ADAM10 expression in WT1 positive podocytes. (B) Increased ADAM10 levels are found in the urine of patients with glomerular kidney diseases . Western Blot analysis of ADAM10 expression in urine and urinary vesicles of healthy volunteers (HV 1-2) and patients with glomerular kidney diseases (number of patients P1-5, ADAM10 expression in supernatants (SN) and vesicles (VES) from untreated (HPC C) or treated with 1 μM ionomycin (HPC IONO) for 24 h. Membranes were reprobed with CD9 an specific marker protein of exosomes.

Article Snippet: WT1 antibody for immunofluorescence analysis was purchased from Santa Cruz (Heidelberg, Germany).

Techniques: Isolation, Immunofluorescence, Expressing, Western Blot, Marker

Journal: Scientific Reports

Article Title: Identification of m6A methyltransferase-related WTAP and ZC3H13 predicts immune infiltrates in glioblastoma

doi: 10.1038/s41598-025-88671-4

Figure Lengend Snippet: Relative expression of genes expression in GBM tissues. Expression levels of WTAP and ZC3H13 genes were compared with those of the control group on GBM tissues, 10 GBM and adjacent tissues, respectively ( A ). The expression levels of siRNA WTAP and overexpression ZC3H13 genes ( B and C ). The expression levels of CD27, CD70, CD80, CD86, ICOS, CTLA4, and LAG3 in siRNA WTAP and overexpression ZC3H13 were demonstrated in Fig. 9D and F, respectively. The result were determined by qRT-PCR. The data are mean ± standard deviation of six replicate experiments. * P < 0.05, **P < 0.01 and ***P < 0.001 compared with the corresponding control values.

Article Snippet: After blocking with 5% nonfat milk, the membranes were immunoblotted with the primary antibodies: WTAP (A04296-2, Boster Biotech, Wuhan, China), CD27 (A01148-2, Boster Biotech), CD70 (A02853-2, Boster Biotech), CD80 (A00196-3, Boster Biotech), CD86(BM4121, Boster Biotech), ICOS (A00291-2, Boster Biotech), CTLA4 (A00020-1, Boster Biotech), LAG3 (M02869-2, Boster Biotech), Beta-actin (BM3873, Boster Biotech).

Techniques: Expressing, Control, Over Expression, Quantitative RT-PCR, Standard Deviation

Journal: International Journal of Biological Sciences

Article Title: APT1-derived depalmitoylation of CD36 alleviates diabetes-induced lipotoxicity in podocytes

doi: 10.7150/ijbs.109220

Figure Lengend Snippet: Construction of db/db model to evaluate changes in renal pathology lipid metabolism. Two groups of twenty-week-old mice were analyzed: db/m (n=6) and db/db (n=6). A-B. Representative PAS staining of glomeruli in each group, and the semiquantitative analysis of mesangial area ratio. (**<0.01, scale bar: 50 μm). C-G. Biochemical examination: (C) UACR, (D) serum creatinine, (E) blood glucose, (F) serum cholesterol, (G) serum triglycerides. (**<0.01). H. Representative transmission electron microscopy (TEM) images of podocyte foot processes and glomerular basement membrane (GBM) in each group. (**<0.01, scale bar: 10 μm). I-J. Representative immunofluorescence double staining of BODIPY and WT1 in the glomeruli of db/m and db/db mice, and semiquantitative analysis. (**<0.01, scale bar:50μm). K-L. Oil Red O staining of lipid droplets (LDs) in the glomeruli of db/m and db/db mice, and semiquantitative analysis. (**<0.01, scale bar: 40 μm).

Article Snippet: Frozen kidney sections were fixed, blocked, and then stained with WT1 antibody (BM4216, Boster, China) for 1 h at 37°C, subsequently incubated in the presence of secondary antibody and BODIPY dye, and finally covered with DAPI.

Techniques: Staining, Transmission Assay, Electron Microscopy, Membrane, Immunofluorescence, Double Staining