Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: AT1 (A) and AT2 (B) were detected in kidney slices as described in the methods section. Arrows indicate localization of AT1 and AT2. Magnification = 20X. AT1 is strongly expressed in the apical and lateral membranes of the tubules (thin arrows), and in mesangial cells (thick arrow),.AT2 is expressed mostly in glomerular endothelial cells (thick arrow) and in the proximal tubules (thin arrow). The pictures are representative of kidneys from 2 individual mice. C) detection of AT1 and AT2 by western blot in kidney cortex homogenates using the same antibodies as in A and B.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Western Blot

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: A. Angiotensin (Ang I and Ang II) concentration in kidney cortex homogenates was determined by ELISA. **p<0.01 by Student's t-test. Activation of AT1 (B) and AT2 (C) was assessed on kidney cortex homogenates by immunoblot using conformation-specific antibodies. For each analysis, loading control was assessed by detection of the corresponding receptor, regardless of its activation state, using antibodies directed against the extracellular part of the receptors. The lower panel shows combined data obtained on kidneys from 5 individual mice in each group. **p<0.01, ns = not significant by ANOVA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, Activation Assay, Western Blot

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: A. VEGF mRNA translation was studied by polysome assay. The shift of VEGF mRNA toward the heaviest fractions in diabetic kidneys indicates active translation, and is reversed by the AT2 but not the AT1 antagonist. Note that the distribution of GAPDH mRNA is unaffected by the various treatments. The figure is representative of experiments performed on kidneys from 3 individual mice. B. Association of hnRNP K with VEGF mRNA was studied by immunoprecipitation with anti-hnRNP K antibody followed by extraction of associated RNA associated and RT-PCR. Note that hnRNP K is constitutively associated with both VEGF and GAPDH mRNAs and that only the association with VEGF mRNA is increased in diabetic kidneys. The figure is representative of experiments performed on kidneys from 3 individual mice. **p<0.01, ns = not significant by ANOVA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Immunoprecipitation, Reverse Transcription Polymerase Chain Reaction

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: The diagram depicts a summary of the data obtained in this study. AT1 activation by hyperglycemia activates mTOR independently of Akt and leads to inactivation of eEF2K but is not sufficient to cause the dephosphorylation and activation of eEF2. Activation of AT2 stimulates the Akt-mTOR pathway that inactivates eEF2K and activates PP2A that dephosphorylates eEF2; AT2 also mediates hyperglycemia-induced increment in eEF1A. Activation of eEF2 and up-regulation of eEF1A stimulate elongation phase of mRNA translation, leading to increased translation of VEGF mRNA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Activation Assay, De-Phosphorylation Assay

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: AT1 (A) and AT2 (B) were detected in kidney slices as described in the methods section. Arrows indicate localization of AT1 and AT2. Magnification = 20X. AT1 is strongly expressed in the apical and lateral membranes of the tubules (thin arrows), and in mesangial cells (thick arrow),.AT2 is expressed mostly in glomerular endothelial cells (thick arrow) and in the proximal tubules (thin arrow). The pictures are representative of kidneys from 2 individual mice. C) detection of AT1 and AT2 by western blot in kidney cortex homogenates using the same antibodies as in A and B.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Western Blot

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: A. Angiotensin (Ang I and Ang II) concentration in kidney cortex homogenates was determined by ELISA. **p<0.01 by Student's t-test. Activation of AT1 (B) and AT2 (C) was assessed on kidney cortex homogenates by immunoblot using conformation-specific antibodies. For each analysis, loading control was assessed by detection of the corresponding receptor, regardless of its activation state, using antibodies directed against the extracellular part of the receptors. The lower panel shows combined data obtained on kidneys from 5 individual mice in each group. **p<0.01, ns = not significant by ANOVA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, Activation Assay, Western Blot

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: A. VEGF mRNA translation was studied by polysome assay. The shift of VEGF mRNA toward the heaviest fractions in diabetic kidneys indicates active translation, and is reversed by the AT2 but not the AT1 antagonist. Note that the distribution of GAPDH mRNA is unaffected by the various treatments. The figure is representative of experiments performed on kidneys from 3 individual mice. B. Association of hnRNP K with VEGF mRNA was studied by immunoprecipitation with anti-hnRNP K antibody followed by extraction of associated RNA associated and RT-PCR. Note that hnRNP K is constitutively associated with both VEGF and GAPDH mRNAs and that only the association with VEGF mRNA is increased in diabetic kidneys. The figure is representative of experiments performed on kidneys from 3 individual mice. **p<0.01, ns = not significant by ANOVA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Immunoprecipitation, Reverse Transcription Polymerase Chain Reaction

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: A. AT2 protein level was measured by immunoblot using a specific antibody. The lower panel shows combined data obtained on kidneys from 5 individual mice for each experimental condition. B. VEGF protein (B) and mRNA (C) levels were measured by immunoblot and RT-qPCR, respectively, on the same number of kidneys as in panel A. Identical experiments were performed for the protein and mRNA levels of fibronectin (D - E) and laminin β1 (F - G). **p<0.01, ns = not significant by ANOVA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Western Blot, Quantitative RT-PCR

Journal:

Article Title: Acute hyperglycemia rapidly stimulates VEGF mRNA translation in the kidney. Role of angiotensin type 2 receptor (AT2)

doi: 10.1016/j.cellsig.2010.07.012

Figure Lengend Snippet: The diagram depicts a summary of the data obtained in this study. AT1 activation by hyperglycemia activates mTOR independently of Akt and leads to inactivation of eEF2K but is not sufficient to cause the dephosphorylation and activation of eEF2. Activation of AT2 stimulates the Akt-mTOR pathway that inactivates eEF2K and activates PP2A that dephosphorylates eEF2; AT2 also mediates hyperglycemia-induced increment in eEF1A. Activation of eEF2 and up-regulation of eEF1A stimulate elongation phase of mRNA translation, leading to increased translation of VEGF mRNA.

Article Snippet: Conformation-specific antibodies directed against activated AT1 and AT2 were from Assay Designs (Ann Arbor, MI), and antibodies directed against AT1 and AT2 were purchased from Alomone Labs (Jerusalem, Israel).

Techniques: Activation Assay, De-Phosphorylation Assay