Systemic delivery of IL-10 by an AAV vector prevents vascular remodeling and end-organ damage in stroke-prone spontaneously hypertensive rat.

Interleukin-10 (IL-10) ameliorates various T-helper type 1 cell-mediated chronic inflammatory diseases. Although the therapeutic benefits of IL-10 include antiatherosclerotic effects, pathophysiological effects of IL-10 on vascular remodeling in hypertension have not yet been elucidated. These studies were designed to determine whether sustained IL-10 expression, mediated by an adeno-associated virus (AAV) vector, prevents vascular remodeling and target-organ damage in the stroke-prone spontaneously hypertensive rat (SHR-SP)-an animal model of malignant hypertension. A single intramuscular injection of an AAV1 vector encoding rat IL-10 introduced long-term IL-10 expression. These IL-10-transduced rats had decreased stroke episodes and proteinuria, resulting in improved survival. Histological examination revealed a reduced level of deleterious vascular remodeling of resistance vessels in the brain and kidney of these rats. Immunohistochemical analysis indicated that IL-10 inhibited the enhanced renal transforming growth factor-beta expression and perivascular infiltration of monocytes/macrophages and nuclear factor-kappaB-positive cells normally observed in the SHR-SP. Four weeks after IL-10 vector injection, systolic blood pressure significantly decreased and this effect persisted for several months. Overall, AAV vector-mediated systemic IL-10 expression prevented vascular remodeling and inflammatory lesions of target organs in the SHR-SP. This approach provides significant insights into the prevention strategy of disease onset with unknown genetic predisposition or intractable polygenic disorders.

Adeno-associated virus vector-mediated interleukin-10 gene transfer inhibits atherosclerosis in apolipoprotein E-deficient mice.

Inflammation is a major contributor to atherosclerosis by its effects on arterial wall biology and lipoprotein metabolism. Interleukin-10 (IL-10) is an anti-inflammatory cytokine that may modulate the atherosclerotic disease process. We investigated the effects of adeno-associated virus (AAV) vector-mediated gene transfer of IL-10 on atherogenesis in apolipoprotein E (ApoE)-deficient mice. A murine myoblast cell line, C2C12, transduced with AAV encoding murine IL-10 (AAV2-mIL10) secreted substantial amounts of IL-10 into conditioned medium. The production of monocyte chemoattractant protein-1 (MCP-1) by the murine macrophage cell line, J774, was significantly inhibited by conditioned medium from AAV2-mIL10-transduced C2C12 cells. ApoE-deficient mice were injected with AAV5-mIL10 into their anterior tibial muscle at 8 weeks of age. The expression of MCP-1 in the vascular wall of the ascending aorta and serum MCP-1 concentration were decreased in AAV5-mIL10-transduced mice compared with AAV5-LacZ-transduced mice. Oil red-O staining of the ascending aorta revealed that IL-10 gene transfer resulted in a 31% reduction in plaque surface area. Serum cholesterol concentrations were also significantly reduced in AAV5-mIL10-transduced mice. To understand the cholesterol-lowering mechanism of IL-10, we measured the cellular cholesterol level in HepG2 cells, resulting in its significant decrease by the addition of IL-10 in a dose-dependent manner. Furthermore, IL-10 suppressed HMG-CoA reductase expression in the HepG2 cells. These observations suggest that intramuscular injection of AAV5-mIL10 into ApoE-deficient mice inhibits atherogenesis through anti-inflammatory and cholesterol-lowering effects.

Increased urinary 15-F2t-isoprostane concentrations in patients with non-ischaemic congestive heart failure: a marker of oxidative stress.

OBJECTIVE: To investigate a novel marker of oxidative stress in patients with congestive heart failure (CHF). PATIENTS: 15 patients with mild CHF, 15 patients with severe CHF with acute exacerbation, and 15 control subjects. MAIN OUTCOME MEASURES: Measurement of urinary 15-F2t-isoprostane, plasma brain natriuretic peptide (BNP), serum interleukin 6 (IL-6), and serum thrombomodulin concentrations. In patients with severe CHF, samples were taken at admission and 4, 7, and 14 days after admission. RESULTS: Urinary 15-F2t-isoprostane, plasma BNP, and serum IL-6 concentrations in patients with severe CHF were significantly higher than those in control subjects or in patients with mild CHF. However, concentrations of serum thrombomodulin, a marker of endothelial damage, were not different between patients with CHF and control subjects. In addition, urinary 15-F2t-isoprostane, plasma BNP, and serum IL-6 concentrations in patients with severe CHF gradually decreased in proportion to the severity of CHF during hospitalisation. Interestingly, urinary 15-F2t-isoprostane concentrations significantly correlated with plasma BNP concentrations and serum IL-6 concentrations, but not with serum thrombomodulin concentrations. CONCLUSIONS: Urinary 15-F2t-isoprostane concentrations increased in proportion to the severity of CHF in patients. This may be caused by increased 15-F2t-isoprostane production. These findings suggest that urinary 15-F2t-isoprostane may be a marker of morbidity as well as oxidative stress in patients with CHF.

Augmentation of monocyte interleukin-8 production by psoralen/UVA-treated CD4+ T cells.

Treatment of cells with psoralen and ultraviolet A light (UVA) modulates their cytokine production. As extracorporeal photochemotherapy has been reported to induce cytokine production by monocytes, we quantified interleukin-8 (IL-8), a representative chemokine produced by monocytes, in culture supernatants from human peripheral blood mononuclear cells (PBMC) treated with 8-methoxypsoralen (8-MOP) and UVA. Lipopolysaccharide stimulated IL-8 production in 8-MOP-phototreated PBMC more efficiently than those untreated or treated with 8-MOP or UVA. More interestingly, when cultured with T-cell-stimulating anti-CD3 and anti-CD28 antibodies, 8-MOP/UVA-treated PBMC produced enhanced amounts of IL-8 with an increased level of IL-8 mRNA expression. Depletion of CD4 but not CD8 T cells from PBMC abrogated this augmented IL-8 elaboration, and CD4 T cells per se secreted no substantial amount of IL-8 even upon CD3/CD28 stimulation. Thus, 8-MOP/UVA-treated CD4 T cells stimulated monocytes to secrete IL-8. The IL-8 overproduction was induced by direct contact of monocytes with 8-MOP/UVA-treated CD4 T cells but not by cytokines from the treated CD4 T cells. These findings imply that in extracorporeal photochemotherapy, monocytes effectively produce IL-8 by cell-to-cell contact with 8-MOP/UVA-treated malignant CD4 T cells. The augmentation of monocyte cytokine/chemokine production by 8-MOP/UVA may be one of the mechanisms underlying the therapeutic efficacy of extracorporeal photochemotherapy.