rabbit polyclonal antibody anti vegfa  (St Johns Laboratory)

Journal: Renal Failure

Article Title: miR-29a-3p/ Vegfa axis modulates high phosphate-induced vascular smooth muscle cell calcification

doi: 10.1080/0886022x.2025.2489712

Figure Lengend Snippet: Figure 1. Identification of Vegfa as a direct target of miR-29a-3p in VSMCs using bioinformatics and dual-luciferase assay. (A) Venn diagram depicting the overlap of miR-29a-3p target genes predicted by four miRNA target prediction tools: miRTarBase, miRWalk, TargetScan, and miRDB. The box highlights the seven genes identified as common targets across all tools. (B) Sequence alignment of miR-29a-3p with the 3′-UTR region of wild-type and mutated Vegfa mRNA. The predicted binding site and seed region are marked in red. (C) Dual-luciferase reporter assay results for VSMCs co-transfected with a control vector, a vector containing the wild type (WT) Vegfa 3′-UTR sequence, or the MUT Vegfa sequence, along with miR-29a-3p mimics or a negative control miRNA (NT). Relative luciferase activity was measured, and data are presented as mean ± SD from three independent experiments.

Article Snippet: VSMCs recovered by centrifugation were homogenized and separated using 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (China National Pharmaceutical Group Corporation Chemical reagent Co., Ltd., China; Cat. #30166428). the proteins were then transferred to polyvinylidene fluoride membranes (Millipore Co., uSa; Cat. #ISEQ00010). the blots were stained with rabbit polyclonal antibodies against VEGFa (1:10,000; Proteintech, uSa; Cat. #19003-1-aP), and mouse monoclonal antibodies against β-actin (1:10,000, Servicebio, China; Cat. #Gb15001), followed by anti-rabbit IgG (1:10,000, SIMubIotECH, China; Cat. #S2001), and anti-mouse IgG horseradish peroxidase conjugates (1:10,000, abclonal, China; Cat. #aS003).

Techniques: Luciferase, Sequencing, Binding Assay, Reporter Assay, Transfection, Control, Plasmid Preparation, Negative Control, Activity Assay

Journal: Renal Failure

Article Title: miR-29a-3p/ Vegfa axis modulates high phosphate-induced vascular smooth muscle cell calcification

doi: 10.1080/0886022x.2025.2489712

Figure Lengend Snippet: Figure 2. miR-29a-3p overexpression suppresses Vegfa expression in VSMCs. (A) Representative Western blot analysis of VEGFA protein levels in VSMCs under standard culture conditions (control), high phosphate conditions (Pi), and high phosphate conditions following miR-29a-3p overexpression (Pi + miR-29a-3p). β-actin served as a loading control. Quantitative analysis of VEGFA protein levels is shown as relative fold change normalized to β-actin. (B) Vegfa mRNA levels quantified using RT-qPCR in the same experimental groups. GAPDH was used as an internal control for normalization. Data represent the mean ± SD of five independent biological replicates.

Article Snippet: VSMCs recovered by centrifugation were homogenized and separated using 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (China National Pharmaceutical Group Corporation Chemical reagent Co., Ltd., China; Cat. #30166428). the proteins were then transferred to polyvinylidene fluoride membranes (Millipore Co., uSa; Cat. #ISEQ00010). the blots were stained with rabbit polyclonal antibodies against VEGFa (1:10,000; Proteintech, uSa; Cat. #19003-1-aP), and mouse monoclonal antibodies against β-actin (1:10,000, Servicebio, China; Cat. #Gb15001), followed by anti-rabbit IgG (1:10,000, SIMubIotECH, China; Cat. #S2001), and anti-mouse IgG horseradish peroxidase conjugates (1:10,000, abclonal, China; Cat. #aS003).

Techniques: Over Expression, Expressing, Western Blot, Control, Quantitative RT-PCR

Journal: Renal Failure

Article Title: miR-29a-3p/ Vegfa axis modulates high phosphate-induced vascular smooth muscle cell calcification

doi: 10.1080/0886022x.2025.2489712

Figure Lengend Snippet: Figure 3. Vegfa knockdown and miR-29a-3p overexpression mitigate high phosphate-induced VSMCs calcification. (A–C) Analysis of Vegfa knockdown effects in VSMCs cultured in normal media (control) or high phosphate conditions (Pi). (A) Vegfa mRNA levels were quantified via RT-qPCR. (B) ARS with corresponding spectrophotometric quantification to assess cellular mineralization. (C) Quantification of intracellular Ca2+ content. Cells were treated with small interfering RNA targeting Vegfa (siVegfa) or a NT control. (D–F) Investigation of the role of miR-29a-3p in VSMCs cultured under high phosphate conditions. (D) Vegfa mRNA expression measured using RT-qPCR, (E) mineralization assessed using ARS and spectrophotometric quantification, and (F) intracellular Ca2+ levels were quantified. Cells were transfected with either an empty vector (VE) or a Vegfa-overexpressing construct (OE), combined with a NT miRNA mimic or a miR-29a-3p mimic. All experiments were performed in five replicates, and error bars represent the SD. GAPDH was used as an internal control in RT-qPCR experiments. Scale bar = 100 µm.

Article Snippet: VSMCs recovered by centrifugation were homogenized and separated using 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (China National Pharmaceutical Group Corporation Chemical reagent Co., Ltd., China; Cat. #30166428). the proteins were then transferred to polyvinylidene fluoride membranes (Millipore Co., uSa; Cat. #ISEQ00010). the blots were stained with rabbit polyclonal antibodies against VEGFa (1:10,000; Proteintech, uSa; Cat. #19003-1-aP), and mouse monoclonal antibodies against β-actin (1:10,000, Servicebio, China; Cat. #Gb15001), followed by anti-rabbit IgG (1:10,000, SIMubIotECH, China; Cat. #S2001), and anti-mouse IgG horseradish peroxidase conjugates (1:10,000, abclonal, China; Cat. #aS003).

Techniques: Knockdown, Over Expression, Cell Culture, Control, Quantitative RT-PCR, Small Interfering RNA, Expressing, Transfection, Plasmid Preparation, Construct