A CCR2/CCR5 antagonist attenuates an increase in angiotensin II-induced CD11b+ monocytes from atherogenic ApoE-/- mice.

OBJECTIVE: Monocyte infiltration into the vessel wall, a process primarily mediated by the interaction between monocyte chemoattractant protein-1 (MCP-1) and its receptor, CCR2, is a key step in atherogenesis. Angiotensin II (Ang II) enhances this monocyte infiltration by increasing the endothelial binding integrin, CD11b. However, the modulation of the Ang II-induced CD11b expression in monocytes in not clear. The aim of this study was to determine if MCP-1/MCP-2 receptor (CCR2) interaction regulates monocyte CD11b expression after 7 days of Ang II infusion. METHODS AND RESULTS: In ApoE(-/-) mice continuous subcutaneous infusion of Ang II (0.75 mg/kg/day) for 7 days significantly increased CD11b expression in circulating monocytes as measured by flow cytometry. CD11b expression in ApoE(-/-) was increased from 135 +/- 9 to 176 +/- 12 mean fluorescent intensity (MFI), control and Ang II-treated, respectively while in C57B/J wildtype mice CD11b increased from 128 +/- 13 to 174 +/- 8 MFI, control and Ang II-treated, respectively. Interestingly, co-infusion of either MCP-1 neutralizing antibody (25 microg/kg/day) or a CCR2 antagonist (500 microg/kg/day) with Ang II for 7 days effectively inhibited monocyte CD11b expression and this inhibition was accompanied by a down-regulated vascular infiltration of Mac-2 positive monocyte-derived macrophages. CONCLUSION: Our data in the atherogenic ApoE(-/-) mouse demonstrates that the Ang II induced increase in both monocytic CD11b integrin expression and monocyte vascular infiltration occurs early in atherogenesis. These Ang II-induced monocytic changes are in part regulated through the MCP-1/CCR2 interaction.

A nonpeptide, piperidine renin inhibitor provides renal and cardiac protection in double-transgenic mice expressing human renin and angiotensinogen genes.

INTRODUCTION: Controlling hypertension by angiotensin converting enzyme inhibitors (ACEI) or angiotensin receptor blockers (ARB), mechanisms that inhibit later pathway steps in the renin-angiotensin system (RAS), have clinically afforded protection against cardiac and renal disease. MATERIALS AND METHODS: In order to determine if blocking the RAS rate-limiting step of angiotensin II generation via renin inhibition could afford similar end organ protection in a human-relevant preclinical model, this study investigated the cardiac and renal effects of a nonpeptide, piperidine renin inhibitor (RI; 100 mg/kg/day PO) in double transgenic mice (dTGM) which express both human renin and angiotensinogen genes. RI was compared to the ARB, candesartan (3 mg/kg/day PO), and to the ACEI, enalapril (60 mg/kg/day PO) in a 4-week dosing paradigm. These doses of RI, ACEI and ARB were previously found to normalize mean blood pressure (MBP) to 110 + 3, 109 + 7 and 107 + 6 mmHg, respectively, after 1 day of treatment. RESULTS AND DISCUSSION: In the dTGM, PRA, plasma aldosterone, GFR, microalbuminuria and left ventricular free wall thickness (LVH) were higher than in the wild type C57BL/6 mice. Microalbuminuria and LVH were significantly reduced by 93% and 9% for the RI, 83% and 13% for enalapril and 73% and 6% for candesartan, respectively. PRA and aldosterone were reduced by the RI 56% and 23%, respectively. These results suggest that the RI provides protection against cardiac and renal disease, similar to ARB and ACEI.

Delayed angiogenesis in aging rats and therapeutic effect of adenoviral gene transfer of VEGF.

We have studied an age-related impairment in angiogenesis and evaluated the effect of overexpressing VEGF in this situation. Polyvinyl alcohol sponges were implanted subcutaneously into aged (24-month), adult (12-month), and young (2-month) rats. Blood vessel ingrowth and proliferative activity in the sponges were assessed by histology with immunostaining for von Willebrand's factor and proliferating cell nuclear antigen (PCNA), respectively. The percentage of total sponge area filled with ingrowing fibrovascular tissue was minimal in aged rats, intermediate in adult rats and highest in young rats. A similar pattern was observed for the total blood vessel numbers in the sponges from old to young animals. The percentage of total sponge endothelial cells (ECs) showing proliferative activity (PCNA positive) was lowest in the aged animals, intermediate in the adult rats and highest in the young rats. To further explore the mechanism of impaired angiogenesis in aged animals, we investigated and found a reduced level of endogenous VEGF protein expression in 12-month-old rats compared to that in 2-month-old rats. VEGF121 gene transfer significantly enhanced blood vessel and fibrovascular tissue ingrowth in adult/aged rats. Adenoviral-VEGF gene transfer also significantly stimulated EC proliferation in aged and adult rats. However, identical treatment failed to further stimulate the already more robust angiogenesis in young animals. The different angiogenic response in adult vs. young rats was not due to differences in gene transfer efficiency, since similar levels of human VEGF121 protein was detected in adult and young rats. Our results indicate that the decreased angiogenic response with aging is associated with reduced EC proliferation and reduced endogenous VEGF production. Adenoviral-VEGF121 gene transfer is effective in augmenting angiogenesis, particularly in older animals.

Dose-dependent neovascularization-promoting effect of adenovirus vector CI-1023 in a rat hindlimb ischaemic model.

CI-1023 (AdGVVEGF121.10) is a replication-deficient adenovirus vector (complete E1a-, partial E1b-, partial E3-) delivering human vascular endothelial growth factor-121 gene. Previous studies from this group have established that CI-1023 can successfully transfer human vascular endothelial growth factor-121 gene resulting in local tissue expression of vascular endothelial growth factor protein. The purpose of this study was to evaluate neovascularization-promoting potency and efficacy of CI-1023 in a wide dose range. In a rat hindlimb ischaemic model, we measured neovascularization-promoting effect of CI-1023 using three end-points: post mortem angiography, immuno-histochemistry and Laser Doppler scanning of tissue blood perfusion. Neovascularization-promoting activity of CI-1023 over the dose range of 4 x 10(6) pu-4 x 10(10) pu was evaluated. Our data demonstrated an obvious dose-dependent effect between 4 x 10(6) pu-4 x 10(8) pu. The neovascularizing effect is somewhat plateaued at the levels between 4 x 10(8) pu and 4 x 10(10) pu. We conclude CI-1023 is a potent neovascularization-promoting compound, with a dose-dependent effect between 4 x 10(6) pu-4 x 10(8) pu in the rat hindlimb ischaemic model.

Impaired angiogenesis in SHR is associated with decreased KDR and MT1-MMP expression.

This study examined whether retarded angiogenesis in a hypertension animal model was associated with impaired VEGF signaling. Furthermore, we sought to determine whether this impairment could be overcome by VEGF addition. Using a rat sponge implantation model, we confirmed impaired angiogenesis in spontaneous hypertensive rats (SHRs). Fourteen days after sponge implantation, the level of angiogenesis in SHRs was approximately half of those in age-matched normotensive Wistar-Kyoto or Sprague-Dawley rats. Significantly, expression of kinase-insert domain-containing receptor (KDR) and membrane type 1 matrix metalloproteinase (MT1-MMP) was reduced in SHRs compared to controls. Immunohistological analysis indicated endothelial proliferation was decreased in SHRs. Gene transfer of human VEGF(121) increased KDR and MT1-MMP expression in SHRs. VEGF(121) also up-regulated endothelial proliferation and angiogenesis. Our results indicate down-regulated KDR and MT1-MMP expression is associated with an impaired angiogenesis in SHRs. VEGF gene transfer is effective in ameliorating the impaired angiogenesis in SHRs.