Chu Kataoka

New Anti–Monocyte Chemoattractant Protein-1 Gene Therapy Attenuates Atherosclerosis in Apolipoprotein E–Knockout Mice

BackgroundMonocyte recruitment into the arterial wall and its activation may be the central event in atherogenesis. Monocyte chemoattractant protein-1 (MCP-1) is an important chemokine for monocyte recruitment, and its receptor (CCR2) may mediate such in vivo response. Although the importance of the MCP-1/CCR2 pathway in atherogenesis has been clarified, it remains unanswered whether postnatal blockade of the MCP-1 signals could be a unique site-specific gene therapy. Methods and ResultsWe devised a new strategy for anti-MCP-1 gene therapy to treat atherosclerosis by transfecting an N-terminal deletion mutant of the human MCP-1 gene into a remote organ (skeletal muscle) in apolipoprotein E-knockout mice. This strategy effectively blocked MCP-1 activity and inhibited the formation of atherosclerotic lesions but had no effect on serum lipid concentrations. Furthermore, this strategy increased the lesional extracellular matrix content. ConclusionsWe conclude that this anti-MCP-1 gene therapy may serve not only to reduce atherogenesis but also to stabilize vulnerable atheromatous plaques. This strategy may be a useful and feasible form of gene therapy against atherosclerosis in humans.

Importance of Monocyte Chemoattractant Protein-1 Pathway in Neointimal Hyperplasia After Periarterial Injury in Mice and Monkeys

Neointimal hyperplasia is a major cause of restenosis after coronary intervention. Because vascular injury is now recognized to involve an inflammatory response, monocyte chemoattractant protein-1 (MCP-1) might be involved in underlying mechanisms of restenosis. In the present study, we demonstrate the important role of MCP-1 in neointimal hyperplasia after cuff-induced arterial injury. In the first set of experiments, placement of a nonconstricting cuff around the femoral artery of intact mice and monkeys resulted in inflammation in the early stages and subsequent neointimal hyperplasia at the late stages. We transfected with an N-terminal deletion mutant of the human MCP-1 gene into skeletal muscles to block MCP-1 activity in vivo. This mutant MCP-1 works as a dominant-negative inhibitor of MCP-1. This strategy inhibited early vascular inflammation (monocyte infiltration, increased expression of MCP-1, and inflammatory cytokines) and late neointimal hyperplasia. In the second set of experiments, the cuff-induced neointimal hyperplasia was found to be less in CCR2-deficient mice than in control CCR2+/+ mice. The MCP-1/CCR2 pathway plays a central role in the pathogenesis of neointimal hyperplasia in cuffed femoral artery of mice and monkeys. Therefore, the MCP-1/CCR2 pathway can be a therapeutic target for human restenosis after coronary intervention.

Essential role of monocyte chemoattractant protein-1 in development of restenotic changes (neointimal hyperplasia and constrictive remodeling) after balloon angioplasty in hypercholesterolemic rabbits

Background—Renarrowing of dilated arterial sites (restenosis) hampers the clinical benefits of coronary angioplasty. Infiltration and activation of monocytes in the arterial wall mediated by monocyte chemoattractant protein-1 (MCP-1) might be a major cause of restenosis after angioplasty. However, there is no direct evidence to support a definite role of MCP-1 in the development of restenosis. Methods and Results—We recently devised a new strategy for anti–MCP-1 gene therapy by transfecting an N-terminal deletion mutant of the MCP-1 gene into skeletal muscles. We used this strategy to investigate the role of MCP-1 in the development of restenotic changes after balloon injury in the carotid artery in hypercholesterolemic rabbits. Intramuscular transfection of the mutant MCP-1 gene suppressed monocyte infiltration/activation in the injured arterial wall and thus attenuated the development of neointimal hyperplasia and negative remodeling. Conclusions—MCP-1–mediated monocyte infiltration is necessary in the development of restenotic changes to balloon injury in hypercholesterolemic rabbits. This strategy may be a useful and practical form of gene therapy against human restenosis.

Pathogenic Role of Oxidative Stress in Vascular Angiotensin-Converting Enzyme Activation in Long-Term Blockade of Nitric Oxide Synthesis in Rats

Inhibition of nitric oxide (NO) synthesis with N(omega)-nitro-L-arginine methyl ester (L-NAME) activates vascular angiotensin-converting enzyme (ACE) and causes oxidative stress. We investigated the role of oxidative stress in the pathogenesis of ACE activation in rats. Studies involved aortas of rats receiving no treatment, L-NAME, L-NAME plus L-arginine, or L-NAME plus an antioxidant drug (N-acetylcysteine, allopurinol, or ebselen) for 7 days. L-NAME significantly increased oxidative stress (O(2)(-)) and ACE activity. The increased O(2)(-) production was normalized by removal of endothelium. Immunohistochemistry showed the increased ACE activity in the endothelial layer. Treatment with antioxidant drugs did not affect the L-NAME-induced increase in systolic arterial pressure but did prevent increases in vascular O(2)(-) production and ACE activity. These results implicate oxidative stress in the pathogenesis of vascular ACE activation in rats with long-term inhibition of NO synthesis. The observed effects of antioxidant drugs on ACE activation do not appear to involve the hypertension induced by L-NAME.

Anti-monocyte chemoattractant protein-1 gene therapy inhibits vascular remodeling in rats: blockade of MCP-1 activity after intramuscular transfer of a mutant gene inhibits vascular remodeling induced by chronic blockade of NO synthesis

Monocyte chemoattractant protein‐1 (MCP‐1) may play an essential part in the formation of arteriosclerosis by recruiting monocytes into the arterial wall. Thus, we devised a new strategy for anti‐MCP‐1 gene therapy against arteriosclerosis by transfecting an amino‐terminal deletion mutant (missing the amino‐terminal amino acids 2 to 8) of the human MCP‐1 gene into a remote organ (skeletal muscles). Intramuscular transduction with the mutant MCP‐1 gene blocked monocyte recruitment induced by a subcutaneous injection of recombinant MCP‐1. In a rat model in which the chronic inhibition of endothelial nitric oxide synthesis induces early vascular inflammation as well as subsequent coronary vascular remodeling, this strategy suppressed monocyte recruitment into the coronary vessels and the development of vascular medial thickening, but did not reduce perivascular fibrosis. Thus, MCP‐1 is necessary for the development of medial thickening but not for fibrosis in this model. This new strategy may be a useful and feasible gene therapy against arteriosclerosis.—Egashira, K., Koyanagi, M., Kitamoto, S., Ni, W., Kataoka, C., Morishita, R., Kaneda, Y., Akiyama, C., Nishida, K.‐i., Sueishi, K., Takeshita, A. Anti‐monocyte chemoattractant protein‐1 gene therapy inhibits vascular remodeling in rats: blockade of MCP‐1 activity after intramuscular transfer of a mutant gene inhibits vascular remodeling induced by chronic blockade of NO synthesis. FASEB J. 14, 1974–1978 (2000)

Anti-monocyte chemoattractant protein-1 gene therapy inhibits restenotic changes (neointimal hyperplasia) after balloon injury in rats and monkeys

Prevention of restenosis after coronary intervention is a major clinical challenge, which highlights the need of new therapeutic options. Vascular injury may involve inflammatory responses that accelerate the recruitment and activation of monocytes through the activation of chemotactic factors, including monocyte chemoattractant protein‐1 (MCP‐1). However, there is no definitive evidence supporting the role of MCP‐1 in restenosis. We recently devised a new strategy for anti‐MCP‐1 gene therapy by transfecting an N‐terminal deletion mutant of the MCP‐1 gene into skeletal muscles. We demonstrate here that this strategy suppressed monocyte infiltration/activation in the injured site and markedly inhibited restenotic changes (neointimal hyperplasia) after balloon injury of the carotid artery in rats and monkeys. This strategy also suppressed the local production of MCP‐1 and inflammatory cytokines. Therefore, monocyte infiltration and activation mediated by MCP‐1 are essential in the development of restenotic changes after balloon injury. This strategy may be a useful form of gene therapy against human restenosis.

Increased Activity of Nuclear Factor-κB Participates in Cardiovascular Remodeling Induced by Chronic Inhibition of Nitric Oxide Synthesis in Rats

BACKGROUND Chronic inhibition of endothelial nitric oxide (NO) synthesis by the administration of N(omega)-nitro-L-arginine methyl ester (L-NAME) to rats induces early vascular inflammatory changes [monocyte infiltration into coronary vessels, nuclear factor-kappaB (NF-kappaB) activation, and monocyte chemoattractant protein-1 expression] as well as subsequent arteriosclerosis (medial thickening and perivascular fibrosis) and cardiac fibrosis. However, no direct evidence for the importance of NF-kappaB in this process is known. METHODS AND RESULTS We examined the effect of a cis element decoy strategy to address the functional importance of NF-kappaB in the pathogenesis of cardiovascular remodeling. We found here that in vivo transfection of cis element decoy oligodeoxynucleotides against NF-kappaB to hearts prevented the L-NAME-induced early inflammation and subsequent coronary vascular medial thickening. In contrast, NF-kappaB decoy oligodeoxynucleotide transfection did not decrease the development of fibrosis, the expression of transforming growth factor-beta(1) mRNA, or systolic pressure overload induced by L-NAME administration. CONCLUSIONS The NF-kappaB system participates importantly in the development of early vascular inflammation and subsequent medial thickening but not in fibrogenesis in this model. The present study may provide a new aspect of how endothelium-derived NO contributes to anti-inflammatory and/or antiarteriosclerotic properties of the vascular endothelium in vivo.

Chronic inhibition of nitric oxide synthesis in rats increases aortic superoxide anion production via the action of angiotensin II

Objective Chronic inhibition of nitric oxide (NO) synthesis by Nω-nitro-L-arginine methyl ester (L-NAME) increases vascular tissue angiotensin II activity and oxidative stress in animals by incompletely understood mechanisms. In a rat model, we investigated the role of local angiotensin II activity in the pathogenesis of increased oxidative stress. Design We studied the aortas of control rats and others receiving L-NAME or L-NAME plus an angiotensin II type 1 receptor antagonist (CS-866). Results Administration of L-NAME for 7 days significantly increased superoxide anion (O2−) and both immunoreactivity and electrophoretically demonstrable activity of redox-sensitive transcription factors (NF-κB and AP-1). Treatment with the angiotensin II type 1 receptor antagonist prevented all of the above changes. The observed effects of the type 1 receptor antagonist was independent of the L-NAME-induced arterial hypertension. Conclusions These findings suggest that chronic inhibition of NO synthesis may increase vascular oxidative stress and oxidative stress-sensitive signals via the action of angiotensin II mediated via type 1 receptors.