Interleukin-5 is a potential mediator of angiotensin II-induced aneurysm formation in apolipoprotein E knockout mice.

BACKGROUND: The aim of this study was to evaluate alterations in Th1 and Th2 cytokines during experimental abdominal aortic aneurysm (AAA) formation. METHODS: AAAs were induced in apolipoprotein E null mice by infusing angiotensin II (Ang II, 1000 ng/kg/min). Aortic homogenates were assessed at 0, 7, 14, and 28 d (n = 11/time point) for select Th1 and Th2 cytokines by ELISA. Additional mice had co-administration of anti-IgG (n = 20) or anti-IL-5 (n = 20) and were assessed at 28 d for AAA. Aortic homogenates were assessed for MMP-2 and MMP-9 expression. Mouse aortic SMC (MASMC) and peritoneal-derived macrophages were treated with IL-5 (0-40 ng/mL), and cell extracts and media (0-48 h) were assessed for MMP-2 and MMP-9 expression. RESULTS: Ang II infusion was associated with a 3.4-fold (P < 0.01) and 3.6-fold (P < 0.01) increase in IL-5 and IL-10 (respectively), and a 0.6-fold reduction in IL-6, by 7 d. Anti-IL-5, but not anti-IgG, ameliorated Ang II-induced AAA formation. Up-regulation of MMP-2 and MMP-9 was observed in aneurysmal aortas, but not in the aortas obtained from mice treated with anti-IL-5. IL-5 stimulation of MASMC increased MMP-2 and MMP-9 mRNA (2.1-fold and 2.7-fold, respectively, P < 0.01) and protein (1.6-fold and 1.9-fold, respectively, P < 0.01) by 24 h. IL-5 stimulation of macrophages did not alter MMP expression. CONCLUSIONS: Ang II induces increased Th2 cytokines IL-5 and IL-10 early in the course of experimental AAA formation, and inhibition of IL-5 prevents AAA formation suggesting an important role. While IL-5 is capable of up-regulating MMP-2 and MMP-9 expression in MASMC, investigations into alternate roles in AAA formation is warranted.

Local and systemic alterations in signal transducers and activators of transcription (STAT) associated with human abdominal aortic aneurysms.

BACKGROUND: Signal transducers and activators of transcription (STAT) proteins are transcription factors that, when activated by phosphorylation, regulate gene expression and cellular activity. The aim of this study was to evaluate the local and systemic expression and activation of STAT proteins associated with abdominal aortic aneurysms (AAA). METHODS: Expression and activation of STAT proteins were assessed in aortic wall samples obtained from patients undergoing repair of AAA (n = 9) and from non-aneurysmal (NA) donors (n = 17). Aortic samples were evaluated for mRNA and protein expression for STAT1, 2, 3, 4, 5a, and 5b using RT-PCR and immunoblot (WB) assays and normalized to ß-actin (expressed as arbitrary units). STAT activation was assessed with WB assays using phosphorylated (p)-STAT-specific antibodies. Alterations in STAT activation were calculated by normalizing pSTAT proteins to corresponding total STAT levels. Immunohistochemistry was performed on AAA and NA samples using the total and pSTAT antibodies. Systemic alterations in STAT activation were assessed by evaluating circulating leukocytes for the presence of pSTAT from patients with AAA (AAA, n = 8), repaired aneurysm (RA, n = 8), or age/gender matched controls with no AAA (CT, n = 8). Flow cytometry was performed to assess for circulating levels of STAT1 (pY701), STAT3 (pY705), and STAT5a (pY694) in monocytes, granulocytes, and lymphocytes. Assessments were made at baseline and in response to in vitro stimulation with IFN-γ (50 ng/mL) or IL-6 (100 ng/mL). Results were analyzed using Student's t-test and are expressed as mean ± SEM. RESULTS: In AAA tissue compared with NA, STAT-1 (1.08 ± 0.09 versus 0.62 ± 0.07), -2 (0.98 ± 0.07 versus 0.55 ± 0.08), and -4 (0.89 ± 0.12 versus 0.35 ± 0.11) mRNA levels were elevated (P < 0.01, all). Corresponding increases in STAT protein were only observed for STAT1 (2.77 ± 0.93 versus 0.93 ± 0.08, P < 0.05). Increases in activation were observed in AAA compared with NA in pSTAT2 (0.77 ± 0.1 versus 0.1 ± 0.02, P < 0.01), pSTAT3 (1.6 ± 0.3 versus 0.2 ± 0.06, P < 0.02) and pSTAT5 (0.57 ± 0.03 versus 0.2 ± 0.03, P < 0.05) levels. Phosphorylated STAT1, 2, 3, and 5 were observed in inflammatory cells invading the AAA adventitia. In addition, STAT3 was observed in the media of AAA and NA, but pSTAT3 was only observed in the media of AAA. There were no differences in baseline levels of pSTAT-positive circulating leukocytes. IFN-γ stimulation decreased STAT-5a (pY694)-positive CT lymphocytes to 40% ± 13% of baseline, but had no effect on AAA or RA lymphocytes (116% ± 35%, 102% ± 19%, respectively; P = 0.01). STAT-5a (pY694)-positive CT granulocytes also decreased to 62% ± 18% of baseline compared with AAA or RA granulocytes (122% ± 25%, 126% ± 17%, respectively; P = 0.01). Alterations in STAT1 (pY701) and STAT3 (pY705) were not observed in leukocytes following cytokine stimulation. CONCLUSIONS: STAT proteins are important regulators of transcriptional activity and have been linked to cardiovascular disease. The present data suggest that altered levels of phosphorylated STATs are associated with AAA. Understanding their role may provide further insight into the mechanisms of AAA formation and allow for the development of medical treatment options.