1. A critical role for TNFα in the selective attachment of mononuclear leukocytes to angiotensin-II-stimulated arterioles
by Teresa Mateo, Yafa Naim Abu Nabah, Mercedes Losada, Rossana Estellés, Chantal Company, Begoña Bedrina, Jose Miguel Cerdá-Nicolás, Stephen Poole, Peter J. Jose, Julio Cortijo, Esteban J. Morcillo, and Maria-Jesus Sanz
Blood
Volume 110(6):
September 15, 2007
©2007 by American Society of Hematology

2. Effect of Ang-II on TNFα release and leukocyte accumulation into the rat peripheral cavity.
Effect of Ang-II on TNFα release and leukocyte accumulation into the rat peripheral cavity. Time course of Ang-II-induced TNFα release (A), neutrophil polymorphonuclear leukocytes [PMNs] (B), and mononuclear leukocyte accumulation (C). Rats were intraperitoneally injected with 5 mL PBS or 1 nM Ang-II. Results are mean (± SEM) for 5 animals per group: *P < .05 or **P < .01 relative to values in the PBS-injected group.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology

3. Effects of an antirat TNFα antiserum on Ang-II-induced leukocyte accumulation and chemokine mRNA expression and generation in the rat peritoneal cavity.
Effects of an antirat TNFα antiserum on Ang-II-induced leukocyte accumulation and chemokine mRNA expression and generation in the rat peritoneal cavity. Neutrophil accumulation (A), mononuclear leukocyte accumulation (B), chemokine mRNA expression (C), and the generation of CINC/KC (D), MIP-2 (E), MCP-1 (F), RANTES (G), and MIP-1α (H) in the rat peritoneal cavity. Cell counts and chemokine levels in the peritoneal exudate as well as chemokine mRNA expression in the mesenteric tissue were determined in the following experimental groups: untreated rats intraperitoneally injected with 5 mL PBS, untreated rats exposed to 1 nM Ang-II intraperitoneally, and Ang-II-exposed rats pretreated with the antirat TNFα antiserum (1 mL per rat intravenously) 15 minutes prior to Ang-II intraperitoneally. Results are the mean (± SEM) for 5 to 8 animals per group. *P < .05 or **P < .01 relative to values in the PBS-injected group; + P < .05 or ++ P < .01 relative to the Ang-II untreated group.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology

4. Effect of antirat TNFα and antirat IL-4 antisera on subacute (4-hour) Ang-II-induced leukocyte adhesion to rat mesenteric arterioles.
Effect of antirat TNFα and antirat IL-4 antisera on subacute (4-hour) Ang-II-induced leukocyte adhesion to rat mesenteric arterioles. Rats were treated intraperitoneally with PBS (buffer; n = 5) or 1 nM Ang-II (n = 5). Some animals were pretreated with the antirat TNFα antiserum (1 mL per rat intravenously; n = 6), or the antirat IL-4 antiserum (1 mL per rat intravenously; n = 6), or a combination of both antisera (n = 4) 15 minutes before the administration of Ang-II. Results are mean (± SEM). *P < .05 or **P < .01 relative to the PBS group; + P < .05 or ++ P < .01 relative to the Ang-II group.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology

5. Effect of antirat TNFα and antirat IL-4 antisera on subacute (4-hour) Ang-II-induced leukocyte responses within rat mesenteric postcapillary venules.
Effect of antirat TNFα and antirat IL-4 antisera on subacute (4-hour) Ang-II-induced leukocyte responses within rat mesenteric postcapillary venules. These responses were measured in the same rats as those described in the legend to Figure 3: venular responses of leukocyte rolling flux (A), leukocyte rolling velocity (B), leukocyte adhesion (C), and leukocyte emigration (D) are mean (± SEM). *P < .05 or **P < .01 relative to the PBS group.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology

6. Representative photomicrographs showing immunolocalization of TNFα and IL-4 in rat mesenteric arterioles and postcapillary venules.
Representative photomicrographs showing immunolocalization of TNFα and IL-4 in rat mesenteric arterioles and postcapillary venules. Mesentery was fixed for staining with anti-TNFα (A,C,E,G) or IL-4 (B,D,F,H) antibodies 4 hours after the intraperitoneal injection of PBS (A-D) or Ang-II (1 nM; E-H). Brown reaction product indicates positive immunoperoxidase localization on the vascular endothelium. All panels are lightly counterstained with hematoxylin. Results are representative of 4 to 5 experiments with each treatment. A 100 × 1.25 oil objective lens was used. Leica IM 1000 software capture imaging (Leica Microsystems, Wetzlar, Germany) was used to obtain the images.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology

7. Effect of Ang-II on TNFα and IL-4 in human cells.
Effect of Ang-II on TNFα and IL-4 in human cells. Effect of TNFα and/or IL-4 on HPBMC recruitment by HUAECs. Time course of Ang-II-induced TNFα mRNA increase in HUAECs (A), HPBMCs (B), TNFα release in HPBMCs, (C) IL-4 mRNA expression in HPBMs (D), and recruitment of HPBMCs by TNFα and/or IL-4-stimulated HUAECs (E). HUAECs and HPBMCs (5 × 106/mL) were stimulated with 1 μM Ang-II in the presence or absence of losartan (100 μM) for 1, 4, and 24 hours. Total mRNA was extracted. Relative quantification of the mRNA levels of TNFα, IL-4, and GAPDH was determined by using real-time quantitative RT-PCR by the comparative Ct method (ΔΔCt method). Columns show the fold increase in expression of TNFα and IL-4 mRNA relative to control GAPDH values, calculated as mean ± SEM of the 2-ΔΔ Ct values of 4 to 5 experiments. Protein content in the supernatant was determined by conventional sandwich ELISA and expressed as pM concentration of the cytokine of mean ± SEM from 4 to 5 experiments. *P < .05 or **P < .01 relative to values in the control group. + P < .05 relative to the Ang-II untreated group. HUAECs were incubated with medium, TNFα (0.2 ng/mL), IL-4 (20 ng/mL), or with a combination of both cytokines for 24 hours. Isolated HPBMCs (1 × 106/mL) were perfused over the HUAECs for 5 minutes at 1 dyn/cm2 and leukocyte accumulation quantified. Results are the mean (± SEM) for 4 experiments. **P < .01 relative to values in the control group; ++ P < .01 relative to the values in the TNFα or IL-4 groups.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology

8. Effect of a neutralizing antihuman TNFα mAb on Ang-II-induced MCP-1, RANTES, IL-8, and MCP-3 release from HUAECs, MIP-1α, and IL-8 release in human whole blood.
Effect of a neutralizing antihuman TNFα mAb on Ang-II-induced MCP-1, RANTES, IL-8, and MCP-3 release from HUAECs, MIP-1α, and IL-8 release in human whole blood. HUAECs and human whole blood were stimulated with Ang-II (1 μM) or with 1 μM Ang-II plus anti-TNFα mAb (2 μg/mL). The release of MCP-1, RANTES, and IL-8 at 4 hours (A) and MCP-1 and MCP-3 at 24 hours (B; pM in the cell supernatant) as well as the release of MIP-1α (C) and IL-8 (D; pM in the plasma) in response to Ang-II was measured by ELISA and is expressed as mean (± SEM) of 4 to 6 experiments: *P < .05 or **P < .01 relative to values in the medium control group; + P < .05 relative to the 1 μM Ang-II group.
Teresa Mateo et al. Blood 2007;110:
©2007 by American Society of Hematology