Implementation of supportive care and best supportive care interventions in clinical trials enrolling patients with cancer†.

BACKGROUND: With the growing and evolving role of palliative care in oncology, we examined how supportive care (SC) and best supportive care (BSC) are implemented in clinical trials when used as a comparison treatment arm. METHODS: We conducted a systematic review of the literature for clinical trials published between 1980 and 2012 in which systemic anticancer therapy was compared with an SC-only arm and compared SC implementation with World Health Organization (WHO) published guidelines. RESULTS: Our search identified 189 articles, 73 of which met our inclusion criteria with the following cancer types: 29 lung, 7 colorectal, 6 pancreatic, 5 gastric and 26 others. Fifty-five studies (75%) provided some definition of SC, and 48 studies (66%) used the term BSC. Twenty-one of the 55 studies that provided a definition described the use of palliative therapies as being 'at the discretion of the treating physician' without standardization. Only two studies provided SC that incorporated routine physical, psychological and social assessments including rapid referral to SC specialists. SC interventions most commonly included analgesics (47%) and radiotherapy (44%). Trials using the term BSC versus SC were more likely to include blood transfusions (P = 0.002) and antibiotics (P = 0.033), but less likely to include steroids (P = 0.05) and palliative specialists (P = 0.047). CONCLUSIONS: The implementation of SC in clinical trials in this systematic review is highly variable. The vast majority of the studies did not meet the WHO guidelines on SC because palliative care therapies were not recommended or integrated into care. Future clinical trials utilizing a SC intervention arm should define these interventions in a standardized approach that meets current guidelines such as the WHO recommendations.

Characteristics of a widespread community cluster of H275Y oseltamivir-resistant A(H1N1)pdm09 influenza in Australia.

BACKGROUND: Oseltamivir resistance in A(H1N1)pdm09 influenza is rare, particularly in untreated community cases. Sustained community transmission has not previously been reported. METHODS: Influenza specimens from the Asia-Pacific region were collected through sentinel surveillance, hospital, and general practitioner networks. Clinical and epidemiological information was collected on patients infected with oseltamivir-resistant viruses. RESULTS: Twenty-nine (15%) of 191 A(H1N1)pdm09 viruses collected between May and September 2011 from Hunter New England (HNE), Australia, contained the H275Y neuraminidase substitution responsible for oseltamivir resistance. Only 1 patient had received oseltamivir before specimen collection. The resistant strains were genetically very closely related, suggesting the spread of a single variant. Ninety percent of cases lived within 50 kilometers. Three genetically similar oseltamivir-resistant variants were detected outside of HNE, including 1 strain from Perth, approximately 4000 kilometers away. Computational analysis predicted that neuraminidase substitutions V241I, N369K, and N386S in these viruses may offset the destabilizing effect of the H275Y substitution. CONCLUSIONS: This cluster represents the first widespread community transmission of H275Y oseltamivir-resistant A(H1N1)pdm09 influenza. These cases and data on potential permissive mutations suggest that currently circulating A(H1N1)pdm09 viruses retain viral fitness in the presence of the H275Y mutation and that widespread emergence of oseltamivir-resistant strains may now be more likely.

Increased detection in Australia and Singapore of a novel influenza A(H1N1)2009 variant with reduced oseltamivir and zanamivir sensitivity due to a S247N neuraminidase mutation.

A novel influenza A(H1N1)2009 variant with mildly reduced oseltamivir and zanamivir sensitivity has been detected in more than 10% of community specimens in Singapore and more than 30% of samples from northern Australia during the early months of 2011. The variant, which has also been detected in other regions of the Asia-Pacific, contains a S247N neuraminidase mutation. When combined with the H275Y mutation, as detected in an oseltamivir-treated patient, the dual S247N+H275Y mutant had extremely high oseltamivir resistance.

A new pandemic influenza A(H1N1) genetic variant predominated in the winter 2010 influenza season in Australia, New Zealand and Singapore.

Pandemic H1N1 influenza virus is of global health concern and is currently the predominant influenza virus subtype circulating in the southern hemisphere 2010 winter. The virus has changed little since it emerged in 2009, however, in this report we describe several genetically distinct changes in the pandemic H1N1 influenza virus. These variants were first detected in Singapore in early 2010 and have subsequently spread through Australia and New Zealand. At this stage, these signature changes in the haemagglutinin and neuraminidase proteins have not resulted in significant antigenic changes which might make the current vaccine less effective, but such adaptive mutations should be carefully monitored as the northern hemisphere approaches its winter influenza season.

Longitudinal sampling is required to maximize detection of intrahost A/H3N2 virus variants.

Seasonal human influenza viruses continually change antigenically to escape from neutralizing antibodies. It remains unclear how genetic variation in the intrahost virus population and selection at the level of individual hosts translates to the fast-paced evolution observed at the global level because emerging intrahost antigenic variants are rarely detected. We tracked intrahost variants in the hemagglutinin and neuraminidase surface proteins using longitudinally collected samples from 52 patients infected by A/H3N2 influenza virus, mostly young children, who received oseltamivir treatment. We identified emerging putative antigenic variants and oseltamivir-resistant variants, most of which remained detectable in samples collected at subsequent days, and identified variants that emerged intrahost immediately prior to increases in global rates. In contrast to most putative antigenic variants, oseltamivir-resistant variants rapidly increased to high frequencies in the virus population. Importantly, the majority of putative antigenic variants and oseltamivir-resistant variants were first detectable four or more days after onset of symptoms or start of treatment, respectively. Our observations demonstrate that de novo variants emerge, and may be positively selected, during the course of infection. Additionally, based on the 4-7 days post-treatment delay in emergence of oseltamivir-resistant variants in six out of the eight individuals with such variants, we find that limiting sample collection for routine surveillance and diagnostic testing to early timepoints after onset of symptoms can potentially preclude detection of emerging, positively selected variants.

Pain experience during initial alignment with a self-ligating and a conventional fixed orthodontic appliance system. A randomized controlled clinical trial.

OBJECTIVES: To test the hypotheses that (1) there is no difference in the pain experience during the week following initial placement of two orthodontic appliances (SmartClip and Victory; 3M Unitek, Monrovia, Calif); and (2) there is no difference in the pain experience during removal and insertion of orthodontic archwires with these brackets. MATERIALS AND METHODS: Sixty-six consecutive patients were treated with a self-ligating bracket system (SmartClip) or a conventional appliance (Victory) on the basis of computer-generated random allocation. After appliance placement and engagement of a 0.016'' nickel-titanium archwire, pain experience was recorded after 4, 24, and 72 hours and after 7 days with the use of a visual analog system (VAS) questionnaire. At a subsequent visit, participants documented pain experiences during removal and insertion of 0.019 x 0.025'' archwires on an additional 100 mm VAS questionnaire. Independent t-tests and analyses of covariance were used to analyze normally distributed data; the Mann-Whitney U-test was used for skewed distributions. RESULTS: Forty-eight (72.2%) and fifty-one (77.3%) subjects completed the first and second parts of the study, respectively. Bracket type had no influence on pain experience at 4 hours (P = .958), 24 hours (P = .289), 72 hours (P = .569), and 7 days (P = .756) following appliance placement. However, bracket type significantly influenced pain experience during archwire removal (P = .001) and insertion (P = .013). CONCLUSIONS: Hypothesis 1 cannot be rejected. The bracket type had no effect on subjective pain experience during the first week after initial placement of two preadjusted orthodontic appliances. Hypothesis 2 was rejected. Significantly greater discomfort was experienced during archwire insertion and removal with the SmartClip appliance.

Plasma growth differentiation factors 8 and 11 levels in cats with congestive heart failure secondary to hypertrophic cardiomyopathy.

OBJECTIVES: Growth differentiation factor (GDF) 11 has been shown to reduce cardiac hypertrophy in mice. Low levels of GDF-11 are associated with cardiac hypertrophy in humans. The authors hypothesized that plasma GDF-11 level is decreased in cats with hypertrophic cardiomyopathy (HCM). Given the close homology between GDF-11 and myostatin/GDF-8, GDF-8 levels were also assessed. ANIMALS: Thirty-seven client-owned cats were enrolled, including cats with normal cardiac structure (n = 16), cats with HCM or hypertrophic obstructive cardiomyopathy (HOCM; n = 14), and cats with HCM and congestive heart failure (CHF; n = 7). METHODS: Plasma samples were analyzed for GDF-8 and GDF-11 using liquid chromatography tandem-mass spectrometry. Levels of GDF-8 and GDF-11 were compared between cats with normal cardiac structure, HCM or HOCM, and CHF. RESULTS: No differences in GDF-11 concentrations were found between cats with normal cardiac structure and cats with HCM/HOCM, with or without history of CHF. Decreased GDF-8 concentrations were detected in cats with CHF compared to cats with HCM/HOCM without history of CHF (p=0.031) and cats with normal cardiac structure (p=0.027). Growth differentiation factor 8 was higher in cats with HOCM compared to those with CHF (p=0.002). No statistical difference was noted in GDF-8 level as a function of age, weight, or body condition score. CONCLUSIONS: Plasma GDF-11 was not different between cats with HCM/HOCM and cats with normal cardiac structure regardless of age. Plasma GDF-8 was decreased in cats with CHF compared to cats with normal cardiac structure and cats with asymptomatic HCM/HOCM, suggesting a possible role in CHF development.

Toll4 (TLR4) expression in cardiac myocytes in normal and failing myocardium.

Expression of innate immune response proteins, including IL-1beta, TNF, and the cytokine-inducible isoform of nitric oxide synthase (iNOS), have been documented in the hearts of humans and experimental animals with heart failure regardless of etiology, although the proximal events leading to their expression are unknown. Noting that expression of a human homologue of Drosophila Toll, a proximal innate immunity transmembrane signaling protein in the fly, now termed human Toll-like receptor 4 (hTLR4), appeared to be relatively high in the heart, we examined TLR4 mRNA and protein abundance in isolated cellular constituents of cardiac muscle and in normal and abnormal murine, rat, and human myocardium. TLR4 expression levels in cardiac myocytes and in coronary microvascular endothelial cells could be enhanced by either LPS or IL-1beta, an effect inhibited by the oxygen radical scavenger PDTC. Transfection of a constitutively active TLR4 construct, CD4/hTLR4, resulted in activation of a nuclear factor-kappaB reporter construct, but not of an AP-1 or an iNOS reporter construct, in cardiac myocytes. In normal murine, rat, and human myocardium, TLR4 expression was diffuse, and presumably cytoplasmic, in cardiac myocytes. However, in remodeling murine myocardium remote from sites of ischemic injury and in heart tissue from patients with idiopathic dilated cardiomyopathy, focal areas of intense TLR4 staining were observed in juxtaposed regions of 2 or more adjacent myocytes; this staining was not observed in control myocardium. Increased expression and signaling by TLR4, and perhaps other Toll homologues, may contribute to the activation of innate immunity in injured myocardium.

Atrial natriuretic factor gene expression in ventricles of rats with spontaneous biventricular hypertrophy.

A subset of Wistar-Kyoto (WKY) rats that spontaneously develops biventricular hypertrophy (BVH) in response to increased cardiac output was evaluated for ventricular expression of the atrial natriuretic factor (ANF) gene. Normal WKY rats had low levels of left ventricular ANF mRNA and minimally detectable ANF transcripts in the right ventricle. In contrast, BVH rats showed a sixfold greater ANF mRNA concentration in the left ventricle than age-matched WKY controls. BVH right ventricular ANF mRNA levels equaled those found in BVH left ventricles and were dramatically greater than WKY right ventricular controls. Unlike experimental models of hypertrophy, both left and right ventricles significantly increase ANF gene transcripts in the natural development of BVH. The left and right ventricles can concordantly respond to hypertrophy and increase ANF gene transcription.

Cystatin C deficiency in human atherosclerosis and aortic aneurysms.

The pathogenesis of atherosclerosis and abdominal aortic aneurysm involves breakdown of the elastic laminae. Elastolytic cysteine proteases, including cathepsins S and K, are overexpressed at sites of arterial elastin damage, but whether endogenous local inhibitors counterbalance these proteases is unknown. We show here that, whereas cystatin C is normally expressed in vascular wall smooth muscle cells (SMCs), this cysteine protease inhibitor is severely reduced in both atherosclerotic and aneurysmal aortic lesions. Furthermore, increased abdominal aortic diameter among 122 patients screened by ultrasonography correlated inversely with serum cystatin C levels. In vitro, cytokine-stimulated vascular SMCs secrete cathepsins, whose elastolytic activity could be blocked when cystatin C secretion was induced by treatment with TGF-beta(1). The findings highlight a potentially important role for imbalance between cysteine proteases and cystatin C in arterial wall remodeling and establish that cystatin C deficiency occurs in vascular disease.

Targeted deletion of matrix metalloproteinase-9 attenuates left ventricular enlargement and collagen accumulation after experimental myocardial infarction.

Matrix metalloproteinase-9 (MMP-9) is prominently overexpressed after myocardial infarction (MI). We tested the hypothesis that mice with targeted deletion of MMP9 have less left ventricular (LV) dilation after experimental MI than do sibling wild-type (WT) mice. Animals that survived ligation of the left coronary artery underwent echocardiographic studies after MI; all analyses were performed without knowledge of mouse genotype. By day 8, MMP9 knockout (KO) mice had significantly smaller increases in end-diastolic and end-systolic ventricular dimensions at both midpapillary and apical levels, compared with infarcted WT mice; these differences persisted at 15 days after MI. MMP-9 KO mice had less collagen accumulation in the infarcted area than did WT mice, and they showed enhanced expression of MMP-2, MMP-13, and TIMP-1 and a reduced number of macrophages. We conclude that targeted deletion of the MMP9 gene attenuates LV dilation after experimental MI in mice. The decrease in collagen accumulation and the enhanced expression of other MMPs suggest that MMP-9 plays a prominent role in extracellular matrix remodeling after MI.

Dilated cardiomyopathy in homozygous myosin-binding protein-C mutant mice.

To elucidate the role of cardiac myosin-binding protein-C (MyBP-C) in myocardial structure and function, we have produced mice expressing altered forms of this sarcomere protein. The engineered mutations encode truncated forms of MyBP-C in which the cardiac myosin heavy chain-binding and titin-binding domain has been replaced with novel amino acid residues. Analogous heterozygous defects in humans cause hypertrophic cardiomyopathy. Mice that are homozygous for the mutated MyBP-C alleles express less than 10% of truncated protein in M-bands of otherwise normal sarcomeres. Homozygous mice bearing mutated MyBP-C alleles are viable but exhibit neonatal onset of a progressive dilated cardiomyopathy with prominent histopathology of myocyte hypertrophy, myofibrillar disarray, fibrosis, and dystrophic calcification. Echocardiography of homozygous mutant mice showed left ventricular dilation and reduced contractile function at birth; myocardial hypertrophy increased as the animals matured. Left-ventricular pressure-volume analyses in adult homozygous mutant mice demonstrated depressed systolic contractility with diastolic dysfunction. These data revise our understanding of the role that MyBP-C plays in myofibrillogenesis during cardiac development and indicate the importance of this protein for long-term sarcomere function and normal cardiac morphology. We also propose that mice bearing homozygous familial hypertrophic cardiomyopathy-causing mutations may provide useful tools for predicting the severity of disease that these mutations will cause in humans.

Peroxisome proliferator-activated receptor gamma activators inhibit cardiac hypertrophy in cardiac myocytes.

BACKGROUND: Peroxisome proliferator-activated receptors (PPARs) are transcription factors belonging to the nuclear receptor superfamily. PPARgamma mRNA is present in cardiac myocytes; however, whether PPARgamma affects cardiac hypertrophy remains unknown. METHODS AND RESULTS: We investigated the effects of PPARgamma activators on cardiac hypertrophy in neonatal rat cardiac myocytes. Cyclic 4% biaxial mechanical strain caused enlargement of cardiac myocytes (1.3-fold versus control, P<0.0001), but the PPARgamma activators troglitazone and 15-deoxy-Delta(12-14)-prostaglandin J(2) (15d-PGJ(2)) (10 micromol/L) inhibited this effect (troglitazone, -72%, P<0.0005; 15d-PGJ(2), -88%, P<0.0002). Total cell protein was increased by mechanical strain (control, 164.3 microgram/dish; strain, 265.5, P<0.0002), and this effect was inhibited by troglitazone and 15d-PGJ(2) (troglitazone, -61%, P<0.005; 15d-PGJ(2), -72%, P<0.001). [(3)H]Leucine uptake was also increased by mechanical strain (1.9-fold versus control, P<0.002), and this increase was inhibited by troglitazone and 15d-PGJ(2) (troglitazone, -52% at 10 micromol/L, P<0.01; 15d-PGJ(2), -70% at 10 micromol/L, P<0.005). An increase in [(3)H]leucine uptake induced by angiotensin II or phenylephrine was significantly inhibited by troglitazone and 15d-PGJ(2). Mechanical strain induced mRNA expression for brain natriuretic peptide, but PPARgamma activators inhibited this induction. Furthermore, PPARgamma activators inhibited mechanically induced activation of nuclear factor (NF)-kappaB. Pyrrolidine dithiocarbamate, an inhibitor of NF-kappaB activation, inhibited strain-induced [(3)H]leucine uptake (-50% at 100 micromol/L, P<0.05). CONCLUSIONS: These results demonstrate that PPARgamma activators inhibit cardiac hypertrophy in cardiac myocytes and suggest that PPARgamma activators may regulate cardiomyocyte hypertrophy at least partially through the NF-kappaB pathway.

Regulation of cardiomyocyte mechanotransduction by the cardiac cycle.

BACKGROUND: Overloading the left ventricle in systole (pressure overload) is associated with a distinct morphological response compared with overload in diastole (volume overload). METHODS AND RESULTS: We designed a novel computer-controlled experimental system that interfaces biaxially uniform strain with electrical pacing, so that cellular deformation can be imposed during a specified phase of the cardiac cycle. Cardiomyocytes were exposed to strain (4%) during either the first third (systolic phase) or last third (diastolic phase) of the cardiac cycle. Strain imposed during the systolic phase selectively activated p44/42 mitogen-activated protein kinase (MAPK) and MAPK/extracellular signal-regulated protein kinase kinase (MEK1/2, an activator of p44/42 MAPK) compared with strain imposed during the diastolic phase. In contrast, there was no difference in activation of p38 and c-Jun NH(2)-terminal kinases induced by strain imposed during the systolic phase (5.8- and 3.3-fold versus control, n=4) compared with the diastolic phase (5.5- and 3.1-fold). Induction of both brain natriuretic peptide (5.8-fold versus control, P:<0.05, n=3) and tenascin-C (7.0-fold, P:<0.02) mRNA expression by strain imposed during the systolic phase was greater than during the diastolic phase (3.9- and 3.6-fold, respectively). [(3)H]leucine incorporation induced by strain imposed during the systolic phase (4.0-fold versus control) was greater than during the diastolic phase (2.7-fold, P:<0.02, n=4); a selective inhibitor of MEK1/2 inhibited this difference. CONCLUSIONS: Mechanical activation of p44/42 MAPK and MEK1/2, gene expression, and protein synthesis is regulated by the cardiac cycle, suggesting that mechanotransduction at the cellular level may underlie differences between pressure and volume overload of the heart.

The impact of calcification on the biomechanical stability of atherosclerotic plaques.

BACKGROUND: Increased biomechanical stresses in the fibrous cap of atherosclerotic plaques contribute to plaque rupture and, consequently, to thrombosis and myocardial infarction. Thin fibrous caps and large lipid pools are important determinants of increased plaque stresses. Although coronary calcification is associated with worse cardiovascular prognosis, the relationship between atheroma calcification and stresses is incompletely described. METHODS AND RESULTS: To test the hypothesis that calcification impacts biomechanical stresses in human atherosclerotic lesions, we studied 20 human coronary lesions with techniques that have previously been shown to predict plaque rupture locations accurately. Ten ruptured and 10 stable lesions derived from post mortem coronary arteries were studied using large-strain finite element analysis. Maximum stress was not correlated with percentage of calcification, but it was positively correlated with the percentage of lipid (P:=0.024). When calcification was eliminated and replaced with fibrous plaque, stress changed insignificantly; the median increase in stress for all specimens was 0.1% (range, 0% to 8%; P:=0.85). In contrast, stress decreased by a median of 26% (range, 1% to 78%; P:=0.02) when lipid was replaced with fibrous plaque. CONCLUSIONS: Calcification does not increase fibrous cap stress in typical ruptured or stable human coronary atherosclerotic lesions. In contrast to lipid pools, which dramatically increase stresses, calcification does not seem to decrease the mechanical stability of the coronary atheroma.

Biomechanical strain induces class a scavenger receptor expression in human monocyte/macrophages and THP-1 cells: a potential mechanism of increased atherosclerosis in hypertension.

BACKGROUND: Although hypertension is an important risk factor for the development of atherosclerosis, the mechanisms for this interaction are incompletely described. Previous studies have suggested that biomechanical strain regulates macrophage phenotype. We tested the hypothesis that biomechanical strain can induce expression of the class A scavenger receptor (SRA), an important lipoprotein receptor in atherogenesis. METHODS AND RESULTS: Human monocyte/macrophages or THP-1 cells were cultured in a device that imposes uniform biaxial cyclic 1-Hz strains of 0%, 1%, 2%, or 3%, and SRA expression was analyzed. Mechanical strains induced SRA mRNA (3.5+/-0.6-fold at 3% strain for 48 hours, P<0.01) and SRA protein in THP-1 cells in an amplitude-dependent manner. This induction was accompanied by augmented expression of the class B scavenger receptor CD36 (2.8+/-0.3-fold, P<0.001) but not by increased peroxisome proliferator-activated receptor-gamma expression. To evaluate this effect in vivo, apolipoprotein E(-/-) mice were randomly assigned to receive standard chow, a high-cholesterol diet, or a high-cholesterol diet with hypertension induced by angiotensin II infusion for 8 weeks. Immunohistochemistry revealed that among macrophages in atherosclerotic lesions of the aorta, the proportion of macrophages with SRA expression was highest in hypertensive animals on a high-cholesterol diet (43.9+/-0.7%, versus 12.0+/-2.0% for normotensive animals on a high-cholesterol diet and 4.7+/-4.7% for animals on standard chow; P<0.001). CONCLUSIONS: Biomechanical strain induces SRA expression by monocyte/macrophages, suggesting a novel mechanism for promotion of atherosclerosis in hypertensive patients.

Matrix metalloproteinase inhibition attenuates early left ventricular enlargement after experimental myocardial infarction in mice.

BACKGROUND: Extracellular matrix synthesis and degradation contribute to the morphological changes that occur after myocardial infarction (MI). METHODS AND RESULTS: We tested the hypothesis that inhibition of matrix metalloproteinases (MMPs) attenuates left ventricular remodeling in experimental MI. Seventy-one male FVB mice that survived ligation of the left anterior coronary artery were randomized to a broad-spectrum MMP inhibitor (CP-471,474) or placebo by gavage. Echocardiographic studies were performed before randomization (within 24 hours of surgery) and 4 days later and included short-axis imaging at the midpapillary and apical levels. Infarction as defined by wall motion abnormality was achieved in 79% of the procedures (n=56), and mortality rate during the 4-day protocol was 23% (9 of 36 on treatment vs 7 of 35 on placebo; P=NS). Baseline end-diastolic and end-systolic dimensions and areas were similar (P=NS) between treated and placebo groups. At follow-up, infarcted mice allocated to MMP inhibitor had significantly smaller increases in end-systolic and end-diastolic dimensions and areas at both midpapillary and apical levels compared with infarcted mice allocated to placebo (all P<0.05). In addition, infarcted animals that received MMP inhibitor had no change in fractional shortening (-3+/-13%), whereas animals that received placebo had a decrease in fractional shortening (-12+/-12%) (P<0.05). In an analysis stratified by baseline end-diastolic area, the effects of MMP inhibition on the changes in end-systolic area and end-diastolic area were most prominent in animals that had more initial left ventricular dilatation (both P<0.05). CONCLUSIONS: -Administration of an MMP inhibitor attenuates early left ventricular dilation after experimental MI in mice. Further studies in genetically altered mice and other models will improve understanding of the role of MMPs in left ventricular remodeling.

Endothelial nitric oxide synthase limits left ventricular remodeling after myocardial infarction in mice.

Background- To investigate the role of endothelial nitric oxide synthase (NOS3) in left ventricular (LV) remodeling after myocardial infarction (MI), the impact of left anterior descending coronary artery ligation on LV size and function was compared in 2- to 4-month-old wild-type (WT) and NOS3-deficient mice (NOS3(-/-)). Methods and Results- Two days after MI, both strains of mice had a similar LV size, fractional shortening, and ejection fraction by echocardiography. Twenty-eight days after MI, both strains had dilated LVs with decreased fractional shortening and lower ejection fractions. Although the infarcted fraction of the LV was similar in both strains, LV end-diastolic internal diameter, end-diastolic volume, and mass were greater, but fractional shortening, ejection fraction, and the maximum rate of developed LV pressure (dP/dt(max)) were lower in NOS3(-/-) than in WT mice. Impairment of diastolic function, as measured by the time constant of isovolumic relaxation (tau) and the maximum rate of LV pressure decay (dP/dt(min)), was more marked in NOS3(-/-) than in WT mice. Mortality after MI was greater in NOS3(-/-) than in WT mice. Long-term administration of hydralazine normalized blood pressure in NOS3(-/-) mice, but it did not prevent the LV dilatation, impaired systolic and diastolic function, and increased LV mass that followed MI. In WT mice, capillary density and myocyte width in the nonischemic portion of the LV did not differ before and 28 days after MI, whereas in NOS3(-/-) mice, capillary density decreased and myocyte width increased after MI, whether or not hydralazine was administered. Conclusions- These results suggest that the presence of NOS3 limits LV dysfunction and remodeling in a murine model of MI by an afterload-independent mechanism, in part by decreasing myocyte hypertrophy in the remote myocardium.

Overexpression of eotaxin and the CCR3 receptor in human atherosclerosis: using genomic technology to identify a potential novel pathway of vascular inflammation.

BACKGROUND: Unstable atherosclerotic lesions typically have an abundant inflammatory cell infiltrate, including activated T cells, macrophages, and mast cells, which may decrease plaque stability. The pathophysiology of inflammatory cell recruitment and activation in the human atheroma is incompletely described. METHODS AND RESULTS: We hypothesized that differential gene expression with DNA microarray technology would identify new genes that may participate in vascular inflammation. RNA isolated from cultured human aortic smooth muscle cells treated with tumor necrosis factor-alpha (TNF-alpha) was examined with a DNA microarray with 8600 genes. This experiment and subsequent Northern analyses demonstrated marked increases in steady-state eotaxin mRNA (>20 fold), a chemokine initially described as a chemotactic factor for eosinophils. Because eosinophils are rarely present in human atherosclerosis, we then studied tissue samples from 7 normal and 14 atherosclerotic arteries. Immunohistochemical analysis demonstrated overexpression of eotaxin protein and its receptor, CCR3, in the human atheroma, with negligible expression in normal vessels. Eotaxin was predominantly located in smooth muscle cells. The CCR3 receptor was localized primarily to macrophage-rich regions as defined by immunopositivity for CD 68; a minority of mast cells also demonstrated immunopositivity for the CCR3 receptor. CONCLUSIONS: Eotaxin and its receptor, CCR3, are overexpressed in human atherosclerosis, suggesting that eotaxin participates in vascular inflammation. These data demonstrate how genomic differential expression technology can identify novel genes that may participate in the stability of atherosclerotic lesions.