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M C Capogrossi

Publications and source records attributed to M C Capogrossi.

At least 37 records · Page 2Linked to original sources
PubMed2000

E1A stimulates FGF-2 release promoting differentiation of primary endothelial cells.

Basic Fibroblast Growth Factor (FGF-2) is a growth and survival factor and represents one of the most potent differentiation agents of vascular system. In the present study we describe that adenoviral oncoprotein E1A regulates FGF-2 production and determines the acquisition of a pro-angiogenic phenotype in primary bovine aortic endothelial cells (BAEC). Following their transfection, wild type E1A proteins 12S and 13S (wtE1A) stimulated BAEC to differentiate on reconstituted basement membrane matrix (Matrigel). This outcome was paralleled by invasion and migration enhancement in wtE1A-transfected cells. This stimulating effect was absent with the E1A mutant dl646N. Accordingly, zymography and RT - PCR analyses showed that matrix metalloproteinase-9 protein- and mRNA-levels increased following wtE1A transfection. Interestingly, wtE1A-transfected BAEC showed FGF-2 mRNA- and protein-levels higher than controls. Further, FGF-2 neutralization reduced the amount of MMP-9 released in the supernatant of E1A-transfected cells and strongly inhibited BAEC differentiation, thus suggesting that wtE1A activates BAEC by a mechanism, at least partially, dependent on a FGF-2 autocrine/paracrine loop.

Adenovirus E1A Proteins↗
PubMed2000

Transcriptionally active drugs improve adenovirus vector performance in vitro and in vivo.

Cytomegalovirus (CMV) promoter is often present in recombinant adenovirus vectors (AdVs) suitable for gene therapy, ensuring high levels of transgene production in a wide range of hosts. Despite this characteristic, the presence of the AdV genome in target cells and tissues typically lasts longer than transgene production that may be rapidly extincted by ill-defined silencing mechanisms. In the present article, it is reported that transcriptionally active drugs, retinoic acid (RA) and histone deacetylase inhibitor trichostatin A (TSA), enhance AdV transgene expression in infected cells and tissues. The association of RA and TSA increased more than seven-fold above control the activity of AdVs encoding for LacZ or VEGF165. This effect was, at least in part, mediated by the direct activation of retinoic acid receptors. Finally, administration of RA and TSA alone at days 0 and 5 after infection prolonged transgene production up to 21 days after infection versus 6-8 days in untreated controls. These results indicate that transcriptionally active drugs improve AdV function and may represent a novel strategy to more efficiently design AdVs for gene therapy interventions.

Adenoviridae↗
PubMed2000

Wild-type p53 gene transfer inhibits invasion and reduces matrix metalloproteinase-2 levels in p53-mutated human melanoma cells.

The tumor suppressor gene p53 has inhibitory effects on cell growth and angiogenesis and induces apoptosis when overexpressed in melanoma and in a variety of tumor cells by adenovirus-mediated gene transfer. The invasive ability of tumor cells, facilitating local infiltration and metastasis, is related to matrix metalloproteinase levels. In melanoma, matrix metalloproteinase-2 and matrix metalloproteinase-9 have a prominent role in this process. The aim of this study was to evaluate whether wild-type p53 overexpression, obtained by a recombinant adenovirus vector (AdCMV.p53), affects cell invasiveness through modulation of matrix metalloproteinase-2 and matrix metalloproteinase-9. Two human melanoma cell lines were used in this study: the SK-MEL-110, carrying a mutated p53 gene, and the SK-MEL-147, carrying the wild-type p53 gene. SK-MEL-110 cells infected with AdCMV.p53 exhibited decreased invasion capability from day 1 after infection, compared with cells not infected or infected with the control vector AdCMV.Null. This reduced invasiveness was associated with decreased matrix metalloproteinase-2 levels in conditioned media whereas no changes were detected in matrix metalloproteinase-9 secreted levels. No modulation in matrix metalloproteinase-2 mRNA levels was detectable, however, after wild-type p53 gene transfer. Furthermore, protein expression of secreted tissue inhibitor of metalloproteinase-2 was not altered by AdCMV.p53 treatment. In contrast, in SK-MEL-147 cells, AdCMV.p53 did not affect cell invasiveness and levels of secreted matrix metalloproteinase-2. Gene transfer of wild-type p53 inhibited proliferation of both cell lines, showing that also SK-MEL-147 cells respond to wild-type p53 overexpression. This novel mechanism of action of wild-type p53 gene transfer may contribute to its antitumor effect by downregulating cell invasion and matrix metalloproteinase-2 secreted levels in mutated p53 human melanoma cell lines.

Adenoviridae↗
PubMed2000

Nonviral and viral gene transfer to the kidney in the context of transplantation.

BACKGROUND/AIMS: Local modulation of the immune response through genetic manipulation of the graft is an attractive novel approach to overcome the toxicity of immunosuppressive therapy to prevent acute graft rejection. We have previously reported that the cationic polymer polyethylenimine 25k (PEI 25k) transduced reporter genes when injected into the renal artery, but with a low transfection efficiency. Here we compare the risk/benefit profiles of such a nonviral versus a viral technique of gene transfer to the kidney in the context of renal transplantation. METHODS: Donor kidneys from Lewis rats were perfused in a closed circuit with an artificial cell-free medium containing PEI 25k complexed to an expression vector coding for the beta-galactosidase (beta-gal) gene and subsequently transplanted in a syngeneic animal. In a second set of experiments, donor kidneys were injected or perfused with a replication-deficient adenovirus encoding the beta-gal gene (AdCMV. beta-gal; 1 x 10(9) plaque-forming units) before transplantation. RESULTS: Perfusion of the kidney with PEI 25k/DNA complexes resulted in large areas of hypoperfusion characterized by injured glomeruli and tubuli, capillary thrombosis and accumulation of C3 in glomerular capillaries. Reperfusion of the kidney was achieved by lowering the PEI 25k/DNA ratio, but no detectable transfection was observed. In animals receiving adenovirus, the beta-gal activity increased with time and was localized mainly in proximal and distal tubular cells, as documented by beta-gal histochemistry and in situ hybridization. A significantly increased expression of beta-gal was achieved by perfusion of the kidney with AdCMV.beta-gal before transplantation, beta-gal staining mainly localizing in proximal and distal tubular cells. CONCLUSIONS: Unlike nonviral methods of gene delivery, adenovirus-mediated gene transfer to the kidney offers exciting perspectives for the development of molecular medicine in the field of organ transplantation.

Adenoviridae↗
PubMed2000

Adenovirus-mediated human tissue kallikrein gene delivery induces angiogenesis in normoperfused skeletal muscle.

We investigated whether local delivery of the tissue kallikrein gene induces angiogenesis in normoperfused mouse hindlimb muscles. Intramuscular injection of adenovirus containing the human tissue kallikrein gene under the control of a cytomegalovirus enhancer/promoter sequence resulted in local production and release of recombinant human tissue kallikrein, whereas transgene expression was absent in muscles of the contralateral hindlimb. Angiogenesis in infected muscles was documented by histological evidence of increased capillary density. In contrast, no angiogenic effect was seen either in the ipsilateral gastrocnemius or contralateral hindlimb muscles. Neovascularization was associated with a transient increase in muscular blood flow as determined by laser Doppler flowmetry. We also investigated the mechanisms of kallikrein-induced angiogenesis. We found that the angiogenic response to kallikrein was abolished by chronic blockade of the kinin B(1) or B(2) receptor or by inhibition of nitric oxide synthase. In addition, inhibition of cyclooxygenase-2 by nimesulide significantly reduced kallikrein-induced effects. These results indicate that (1) human tissue kallikrein acts as an angiogenic factor in normoperfused skeletal muscle and (2) nitric oxide and prostacyclin are essential mediators of kallikrein-induced angiogenesis. Our findings provide new insights into the role of the tissue kallikrein-kinin system in vascular biology.

Adenoviridae↗
PubMed2000

Adenovirus-mediated human tissue kallikrein gene delivery inhibits neointima formation induced by interruption of blood flow in mice.

Tissue kallikrein cleaves kininogen to produce vasoactive kinin peptides. Binding of kinins to bradykinin B(2) receptors on vascular endothelial cells stimulates the release of nitric oxide and prostacyclin, thus activating the cGMP and cAMP pathways. In this study, we evaluated the effects of adenovirus-mediated human tissue kallikrein gene (Ad.CMV-cHK) delivery in a mouse model of arterial remodeling induced by permanent alteration in shear stress conditions. Mice underwent ligature of the left common carotid artery and were injected intravenously with saline or 1.8 x 10(9) plaque-forming units of Ad.CMV-cHK or control virus (Ad.CMV-LacZ). Fourteen days after surgery, morphometric analysis revealed that Ad. CMV-cHK reduced neointima formation by 52% (P<0.05) compared with Ad. CMV-LacZ. Expression of human tissue kallikrein (HK) mRNA was detected in mouse carotid artery, aorta, kidney, heart, and liver, and recombinant HK was present in the urine and plasma of mice receiving HK gene. Kallikrein gene transfer resulted in increases in urinary kinin, cGMP, and cAMP levels. The protective action of Ad. CMV-cHK on neointima formation was significantly reduced (P<0.05) in mice with knockout of the kinin B(2) receptor gene compared with wild-type control mice (J129Sv mice). In contrast, the effect of Ad. CMV-cHK was amplified (P<0.05) in transgenic mice overexpressing human B(2) receptor compared with wild-type control mice (c57/Bl6 mice). Thus, the inhibitory effect of recombinant kallikrein on structural alterations caused by the interruption of blood flow appears to be mediated by the B(2) receptor. These results provide new insight into the role of the tissue kallikrein-kinin system in vascular remodeling and suggest the application of HK gene therapy to treat restenosis and atherosclerosis.

Adenoviridae↗
PubMed2000

Angiotensin II type 1 receptor blockade prevents cardiac remodeling in bradykinin B(2) receptor knockout mice.

Knockout mice (B(2)(-/-)) lacking the bradykinin (BK) B(2) receptor gene develop mild hypertension, cardiac hypertrophy, and myocardial damage. We hypothesized that these effects are due to the hypertrophying and damaging actions of angiotensin II (Ang II) in the absence of the balancing protection of BK. To verify this hypothesis, B(2)(-/-) or wild-type mice (B(2)(+/+)) were administered a nonpeptide antagonist of Ang II type 1 (AT(1)) receptors (A81988) from conception through 180 days of age. Untreated B(2)(+/+) and B(2)(-/-) served as controls. Blood pressure (BP) and heart rate were monitored with the use of tail-cuff plethysmography at regular intervals. Ventricular weights, diameters, wall thickness, chamber volume, and myocardial fibrosis were measured at 40 and 180 days. No differences were observed in BP, heart rate, and cardiac weight and dimensions between treated and untreated B(2)(+/+). The BP of AT(1) antagonist-treated B(2)(-/-) was reduced until 70 days; then, it increased to the levels found in untreated B(2)(-/-). AT(1) receptor blockade resulted in a reduction in left ventricular mass, chamber volume, and wall thickness and abrogated myocardial fibrosis in B(2)(-/-). These results indicate that Ang II is the major factor responsible for ventricular remodeling and myocardial damage in mice with disruption of BK B(2) receptor signaling. The interaction of Ang II and BK appears to be essential for the development of a normal heart.

Angiotensin II↗
PubMed2000

The chemotactic and mitogenic effects of platelet-derived growth factor-BB on rat aorta smooth muscle cells are inhibited by basic fibroblast growth factor.

In response to endovascular injury, platelet-derived growth factor-BB (PDGF-BB) and basic fibroblast growth factor (bFGF) are released locally and modulate vascular smooth muscle cells (SMC) proliferation and migration within the vascular wall. The aim of the present in vitro study was to determine how rat aorta SMC respond to the simultaneous exposure to PDGF-BB and bFGF. In a modified Boyden chamber assay bFGF exhibited a dose-dependent effect to inhibit the chemotactic action of PDGF-BB. A comparable result was observed in proliferation assays. In contrast, MIP-1 beta, epidermal growth factor (EGF), fibronectin and acidic FGF (aFGF) did not inhibit the chemotactic effect of PDGF-BB. Denatured bFGF did not exert an inhibitory effect and neutralizing antibodies either to bFGF or to bFGF-receptor abolished the inhibition observed in the presence of bFGF. The role played by PDGF receptor alpha (PDGF-Ralpha) was investigated in PDGF-Ralpha-dominant negative-transfected SMC, by selectively blocking PDGF-BB-binding to PDGF-Ralpha with neomycin, by neutralizing PDGF-Ralpha with a monoclonal antibody and by selectively stimulating PDGF-Ralpha with PDGF-AA; in all cases the effect of bFGF to inhibit PDGF-BB-directed SMC migration was abolished. These in vitro studies show that bFGF significantly inhibits PDGF-BB-induced SMC migration and proliferation and that this effect is mediated by both PDGF-Ralpha and bFGF receptor.

Animals↗
PubMed2000

Induction of interleukin-1beta and interleukin-6 gene expression in hypoperfused skeletal muscle of patients with peripheral arterial disease.

BACKGROUND: A growing amount of data supports the role of inflammation in the pathophysiology of atherosclerotic diseases but the cellular source of cytokines has not been clearly identified. Cytokines could be produced by inflammatory cells, activated endothelial and smooth muscle cells, and by the tissue exposed to recurrent ischemia. Accordingly, we evaluated whether hypoperfusion induces gene expression of interleukin (IL)-1beta and IL-6 in the skeletal muscle of patients with peripheral arterial disease and critical limb ischemia. METHODS: Skeletal muscle biopsies were obtained, during a femoral-distal bypass, from normoperfused (control) and hypoperfused skeletal muscles in 8 patients. Gene expression was assessed by semiquantitative reverse transcriptase-polymerase chain reaction, using glyceraldehyde-phosphate-deydrogenase mRNA levels as a normalization factor. RESULTS: In the hypoperfused biopsies, the level of IL-1beta gene expression was significantly higher in all but 2 patients (mean upregulation > 8.8 fold, p = 0.043), and the level of IL-6 gene expression was significantly higher in all but 1 patient (mean upregulation > 23.7 fold, p = 0.031). CONCLUSIONS: We report that IL-1beta and IL-6 gene expression is markedly upregulated in hypoperfused skeletal muscle of patients with critical lower limb ischemia. To our knowledge this is the first report of a local activation of the inflammatory cascade at the level of hypoperfused skeletal muscle. This activation, which could worsen symptoms and tissue viability and be involved in the pathophysiology of reperfusion injury, might be considered as a therapeutic target. It remains to be investigated whether our results may also apply to coronary artery disease.

Aged↗
PubMed1999

Dilated and failing cardiomyopathy in bradykinin B(2) receptor knockout mice.

BACKGROUND: The activation of B(2) receptors by kinins could exert cardioprotective effects in myocardial ischemia and heart failure. METHODS AND RESULTS: To test whether the absence of bradykinin B(2) receptors may affect cardiac structure and function, we examined the developmental changes in blood pressure (BP), heart rate, and heart morphology of bradykinin B(2) receptor gene knockout (B(2)(-/-)), heterozygous (B(2)(+/-)), and wild-type (B(2)(+/+)) mice. The BP of B(2)(-/-) mice, which was still normal at 50 days of age, gradually increased, reaching a plateau at 6 months (136+/-3 versus 109+/-1 mm Hg in B(2)(+/+), P<0.01). In B(2)(+/-) mice, BP elevation was delayed. At 40 days, the heart rate was higher (P<0.01) in B(2)(-/-) and B(2)(+/-) than in B(2)(+/+) mice, whereas the left ventricular (LV) weight and chamber volume were similar among groups. Thereafter, the LV growth rate of B(2)(-/-) and B(2)(+/-) mice was accelerated, leading at 360 days to a LV weight-to-body weight ratio that was 9% and 17% higher, respectively, than that of B(2)(+/+) mice. In B(2)(-/-) mice, hypertrophy was associated with a marked chamber dilatation (42% larger than that of B(2)(+/+) mice), an elevation in LV end-diastolic pressure (25+/-3 versus 5+/-1 mm Hg in B(2)(+/+) mice, P<0.01), and reparative fibrosis. CONCLUSIONS: The disruption of the bradykinin B(2) receptor leads to hypertension, LV remodeling, and functional impairment, implying that kinins are essential for the functional and structural preservation of the heart.

Age Factors↗
PubMed1999

Adenovirus-mediated acidic fibroblast growth factor gene transfer induces angiogenesis in the nonischemic rabbit heart.

Most patients with severe coronary artery disease have normal baseline myocardial blood flow. Therefore, interventions aimed at inducing therapeutic angiogenesis in these patients should cause new blood vessel growth in the heart in the absence of chronic ischemia. It was examined whether adenovirus-mediated gene transfer of recombinant, secreted acidic fibroblast growth factor (sp+aFGF(1-154)), next to a major epicardial artery, may induce neovascularization and reduce the risk region for myocardial infarction upon coronary ligation near the injection site. Fifteen days prior to coronary artery occlusion, rabbits were treated with intramyocardial injections of AdCMV.sp+aFGF(1-154), the control vector AdCMV.NLSbetagal (1 x 10(9) plaque-forming units), or saline. Messenger RNA transcripts for aFGF(1-154) were present up to 12 days after injection in the tissues exposed to AdCMV.aFGF(1-154). Following coronary artery occlusion rabbits treated with AdCMV. sp+aFGF(1-154) showed a 50% reduction of the risk region for myocardial infarction (P < 0.01 vs control). Histologic analysis showed a twofold increase in length density of intramural coronary arterioles (P < 0.01 vs control) and a 17% increase in length density of the capillary network (P < 0.001) in the myocardium exposed to AdCMV.sp+aFGF(1-154). Thus, gene therapy with AdCMV. sp+aFGF(1-154) can induce angiogenesis in the absence of chronic ischemia. The newly formed collateral blood vessels provide an anatomical basis for the reduction in the risk region for myocardial infarction upon subsequent occlusion of the coronary artery in proximity of the site where angiogenesis was induced.

Adenoviridae↗
PubMed1999

Vascular permeability effect of adenovirus-mediated vascular endothelial growth factor gene transfer to the rabbit and rat skeletal muscle.

OBJECTIVE: Vascular endothelial growth factor has been used in preclinical studies and phase 1 and 2 clinical trials as a potent mediator of therapeutic angiogenesis; however, its ability to enhance the vascular permeability may be a source of potential complications. The objective of this work was to evaluate the effects of the intramuscular injection of an adenovirus vector coding for the 121-amino acid form of vascular endothelial growth factor (Ad.VEGF(121 )) on vascular permeability and edema development in rabbits and rats. METHODS: Different concentrations of Ad.VEGF(121 ) ranging from 10(5) to 10(10) plaque-forming units/mL (3 x 10(6)-3 x 10(11) particles/mL) were injected into hind limb or forelimb muscles of Wistar rats or rabbits. The size of the scrotum, the circumferences of limbs, and the concentration of vascular endothelial growth factor in the serum were measured daily after injection. RESULTS: The injection of different concentrations of Ad.VEGF(121 ) into the hind limb muscles of rabbits led to a dose-dependent scrotal edema in rabbits at concentrations higher than 10(7) plaque-forming units/mL (P =.002). The edema developed slowly, reached its maximum level 6 days after the injection, and spontaneously resolved thereafter. At concentrations higher than 10(9) plaque-forming units/mL the scrotal edema was accompanied by skin necrosis (P =.0001). No scrotal edema was observed in rats. CONCLUSIONS: The massive species-specific scrotal edema accompanied by skin ulceration and necrosis was observed only in rabbits treated with Ad.VEGF(121 ) in concentrations exceeding therapeutic doses. The therapeutic doses of Ad.VEGF(121 ) resulted in only moderate transient scrotal edema in rabbits, suggesting that the potential for side effects of vascular endothelial growth factor therapy as a result of increased vascular permeability should not be very alarming for generally healthy patients and may not cause a significant clinical problem in the treatment of peripheral vascular diseases.

Adenoviridae↗
PubMed1999

Adenovirus-mediated wild-type p53 expression induces apoptosis and suppresses tumorigenesis of experimental intracranial human malignant glioma.

Adenoviral-mediated gene transfer for the treatment of experimental intrinsic malignant brain neoplasms holds promise. The role, however, of intracellular, adenoviral-mediated p53 expression to inhibit growth of experimental human intracranial malignant gliomas remains largely unexplored. Using the AdCMV.p53 vector we measured the in vitro expression of p53 and the resultant effect upon U251 human malignant glioma cellular proliferation. We further measured the survival of nude mice after intracranial injection of the infected vs. control U251 cells. The growth of the infected U251 cells was inhibited when compared to both the uninfected cells and cells infected with the control vector (AdCMV.Null). Agarose gel electrophoresis confirmed the AdCMV.p53-dependent cellular apoptosis. Nude mice having intracranial injections of the U251 cells infected with the control (AdCMV.Null) vector showed diminished survival. In contrast, mice having intracranial injections of the cells infected with the AdCMV.p53 vector showed 100% survivorship measured 100 days after treatment. Gene therapy via the AdCMV.p53 viral vector holds promise for the clinical treatment of human malignant gliomas.

Adenoviridae↗
PubMed1999

Vascular endothelial growth factor (VEGF) modulates vascular permeability and inflammation in rat brain.

Vascular endothelial growth factor (VEGF) is an angiogenic growth factor that also induces vascular permeability and macrophage migration. VEGF expression is weak in normal adult brain, but is strongly upregulated in glioma cells and reactive astrocytes, suggesting that chronic overexpression of VEGF in the brain contributes to blood-brain barrier (BBB) breakdown. We examined the effects of chronic VEGF overexposure on the integrity of the BBB using the following approaches: 1) continuous intracerebral infusion of VEGF via miniosmotic pump; and 2) intracerebral injection of an adenoviral vector encoding the VEGF165 gene (AdCMV.VEGF). After 6 days both treatments produced approximately 10-fold breakdown of the BBB (as measured by transport of 14C-aminoisobutyric acid (AIB) from blood into brain) compared with the respective controls (albumin infusion or AdCMV.beta gal virus). BBB disruption in AdCMV.VEGF-treated brains was accompanied by a severe inflammatory response not observed in brains receiving AdCMV.beta gal or VEGF protein infusion, indicating that neither VEGF nor viral particles alone were responsible for the inflammatory response. However, injection of AdCMV.beta gal followed by VEGF infusion to the same site also elicited inflammation. Chronic overexposure of normal brain to VEGF also increased intercellular adhesion molecule-1 (ICAM-1) and major histocompatibility complex (MHC) class I and II expression. Although VEGF itself is not inflammatory, VEGF may modulate immune responses in the central nervous system (CNS) by opening the BBB, altering the immunoprivileged status of the brain, and allowing contact between normally sequestered CNS antigens and blood-borne immune mediators.

Animals↗
PubMed1999

The equine herpesvirus 2 E1 open reading frame encodes a functional chemokine receptor.

Several herpesviruses contain open reading frames (ORFs) that encode potential homologs of eucaryotic genes. Equine herpesvirus 2 (EHV-2) is a gammaherpesvirus related to other lymphotropic herpesviruses such as herpesvirus saimiri and Epstein-Barr virus. The E1 ORF of EHV-2, a G protein-coupled receptor homolog, shows 31 to 47% amino acid identity with known CC chemokine receptors. To investigate whether E1 may encode a functional receptor, we cloned the E1 ORF and expressed it in stably transfected cell lines. We report here the identification of the CC chemokine eotaxin as a functional ligand for the EHV-2 E1 receptor. Chemokines are likely to play a role in the regulation of immune functions in equine hosts during EHV-2 infection and, via interaction with E1, may affect viral replication and/or escape from immune responses.

Amino Acid Sequence↗
PubMed1998

Adenovirus-mediated gene transfer of the human tissue inhibitor of metalloproteinase-2 blocks vascular smooth muscle cell invasiveness in vitro and modulates neointimal development in vivo.

BACKGROUND: Endovascular injury induced by balloon withdrawal leads to the increased activation of matrix metalloproteinases (MMPs) in the vascular wall, allowing smooth muscle cells (SMCs) to digest the surrounding extracellular matrix (ECM) and migrate from the media into the intima. The objective of this study was to examine the effects of a replication-deficient adenovirus carrying the cDNA for human tissue inhibitor of metalloproteinase-2 (AdCMV.hTIMP-2) on SMC function in vitro and neointimal development in the injured rat carotid artery. METHODS AND RESULTS: Infection of cultured rat aortic SMCs at a multiplicity of infection of 100 with AdCMV.hTIMP-2 resulted in high-level expression of hTIMP-2 mRNA and protein secretion into the medium. Conditioned media (CM) from AdCMV. hTIMP-2-infected but not control virus (AdCMV.null or AdCMV. betagal)-infected SMCs inhibited MMP-2 activity on gelatin zymograms as well as the chemoattractant-directed migration of SMCs across reconstituted basement membrane proteins in the Boyden chamber assay. In contrast, AdCMV.hTIMP-2 CM had no effect on chemoattractant-directed migration of SMCs occurring in the absence of an ECM barrier or on the proliferation of cultured neointimal SMCs. Delivery of AdCMV.hTIMP-2 (2.5x10(9) pfu) to the carotid artery wall at the time of balloon withdrawal injury inhibited SMC migration into the intima by 36% (P<0.05) at 4 days and neointimal area by 53% (P<0.01) at 8 days and by 12% (P=NS) at 21 days after injury. AdCMV.hTIMP-2 had no effect on medial area. CONCLUSIONS: Adenovirus-mediated hTIMP-2 gene transfer inhibits SMC invasiveness in vitro and in vivo and delays neointimal development after carotid injury.

Adenoviridae↗
PubMed1998

Cytosolic alkalinization of vascular endothelial cells produced by an abrupt reduction in fluid shear stress.

Reductions in fluid shear stress produce endothelium-dependent vasoconstriction and promote neointimal hyperplasia, but the intracellular signaling mechanisms involved in these processes are poorly understood. To examine whether decreases in fluid shear stress affect endothelial cytosolic pH, carboxy-seminaphthorhodafluor-1-loaded rat aortic endothelial cells were cultured in glass microcapillary tubes and examined during abrupt reductions in laminar flow. After a 30-minute exposure to a shear stress of 2.7 dyne/cm2 in bicarbonate buffer, the acute reduction of fluid shear stress from 2.7 to 0.3 dyne/cm2 transiently increased cytosolic pH from 7.20+/-0.02 to 7.47+/-0.07 (mean+/-SEM, P<.05 versus control). This was not affected by prior inhibition of the Na+-H+ exchanger with 10 micromol/L ethylisopropylamiloride but was abolished in bicarbonate-free buffer. Recovery from an ammonium chloride prepulse-induced acid load occurred more rapidly when fluid shear stress was abruptly reduced from 2.7 to 0.3 dyne/cm2 after maximal acidification (+0.04+/-0.02 pH unit at 2 minutes) than when shear stress was maintained at 2.7 dyne/cm2 continuously (0.00+/-0.00 pH unit at 2 minutes, P<.05). This accelerated cytosolic pH recovery was dependent on the presence of bicarbonate ion and was blocked by the addition of the exchange inhibitors DIDS (100 micromol/L) and ethylisopropylamiloride or by removal of buffer Na+, indicating that the acute reduction in fluid shear stress activates the extracellular Na+-dependent Cl(-)-HCO3- exchanger and the Na+-H+ exchanger and increases cytosolic pH in vascular endothelial cells.

Alkalies↗