Point of care, bone marrow mononuclear cell therapy in ischemic heart failure patients personalized for cell potency: 12-month feasibility results from CardiAMP heart failure roll-in cohort.

AIM: Heart failure following myocardial infarction (MI) is a potentially lethal problem with a staggering incidence. The CardiAMP Heart Failure trial represents the first attempt to personalize marrow-derived cell-based therapy to individuals with cell characteristics associated with beneficial responses in prior trials. Before the initiation of the randomized pivotal trial, an open-label "roll-in cohort" was completed to ensure the feasibility of the protocol's procedures. METHODS: Patients with chronic post-MI heart failure (NYHA class II-III) receiving stable, guideline-directed medical therapy with a left ventricular ejection fraction between 20 and 40% were eligible. Two weeks prior to treatment, a ~ 5 mL bone marrow aspiration was performed to examine "cell potency". On treatment day, a 60 mL bone marrow aspiration, bone marrow mononuclear cell (BM MNC) enrichment and transendocardial injection of 200 million BM MNC's was performed in a single, point of care encounter. Patients were then followed to assess clinical outcomes. RESULTS: The cell potency small volume bone marrow aspirate, the 60 mL bone marrow aspirate, and transendocardial injections were well tolerated in 10 patients enrolled. There were no serious adverse events related to bone marrow aspiration or cell delivery. Improvement in 6-min walk distance was observed at 6 months (+47.8 m, P = 0.01) and trended to improvement at 12 months (+46.4, P = 0.06). Similarly, trends to improved NYHA heart failure functional class, quality of life, left ventricular ejection fraction and recruitment of previously akinetic left ventricular wall segments were observed. CONCLUSION: All CardiAMP HF protocol procedures were feasible and well tolerated. Favorable functional, echo and quality of life trends suggest this approach may offer promise for patients with post MI heart failure. The randomized CardiAMP Heart Failure pivotal trial is underway to confirm the efficacy of this approach. CLINICAL TRIAL REGISTRATION: https://clinicaltrials.gov/ct2/show/NCT02438306.

The CardiAMP Heart Failure trial: A randomized controlled pivotal trial of high-dose autologous bone marrow mononuclear cells using the CardiAMP cell therapy system in patients with post-myocardial infarction heart failure: Trial rationale and study design.

BACKGROUND: Heart failure following myocardial infarction is a common, disabling, and deadly condition. Direct injection of autologous bone marrow mononuclear cells into the myocardium may result in improved functional recovery, relieve symptoms, and improve other cardiovascular outcomes. METHODS: CardiAMP-HF is a randomized, double-blind, sham-controlled, pivotal trial designed to investigate the safety and efficacy of autologous bone marrow mononuclear cells treatment for patients with medically refractory and symptomatic ischemic cardiomyopathy. The primary end point is change in 6-minute walk distance adjusted for major adverse cardiovascular events at 12 months following treatment. Particularly novel aspects of this trial include a cell potency assay to screen subjects who have bone marrow cell characteristics that suggest a favorable response to treatment, a point-of-care treatment method, a high target dose of 200 million cells, and an efficient transcatheter intramyocardial delivery method that is associated with high cell retention. CONCLUSIONS: This novel approach may lead to a new treatment for those with ischemic heart disease suffering from medically refractory heart failure.

Intracoronary infusion of encapsulated glucagon-like peptide-1-eluting mesenchymal stem cells preserves left ventricular function in a porcine model of acute myocardial infarction.

BACKGROUND: Engraftment and survival of stem cells in the infarcted myocardium remain problematic in cell-based therapy for cardiovascular disease. To overcome these issues, encapsulated mesenchymal stem cells (eMSCs) were developed that were transfected to produce glucagon-like peptide-1, an incretin hormone with known cardioprotective effects, alongside MSC endogenous paracrine factors. This study was designed to investigate the efficacy of different doses of intracoronary infusion of eMSC in a porcine model of acute myocardial infarction (AMI). METHODS AND RESULTS: One hundred pigs were subjected to a moderate AMI (posterolateral AMI; n=50) or a severe AMI (anterior AMI; n=50), whereupon surviving animals (n=36 moderate, n=33 severe) were randomized to receive either intracoronary infusion of 3 incremental doses of eMSC or Ringers' lactate control. Cardiac function was assessed using invasive hemodynamics, echocardiography, and histological analysis. A trend was observed in the moderate AMI model, whereas in the severe AMI model, left ventricular ejection fraction improved by +9.3% (P=0.004) in the best responding eMSC group, because of a preservation of left ventricular end-systolic volume. Arteriolar density increased 3-fold in the infarct area (8.4±0.9/mm(2) in controls versus 22.2±2.6/mm(2) in eMSC group; P<0.001). Although not statistically significant, capillary density was 30% higher in the border zone (908.1±99.7/mm(2) in control versus 1209.0±64.6/mm(2) in eMSC group; P=ns). CONCLUSIONS: eMSCs enable sustained local delivery of cardioprotective proteins to the heart, thereby enhancing angiogenesis and preserving contractile function in an animal AMI model.

Intracoronary stem cell infusion after acute myocardial infarction: a meta-analysis and update on clinical trials.

BACKGROUND: Several cell-based therapies for adjunctive treatment of acute myocardial infarction have been investigated in multiple clinical trials, but the benefits still remain controversial. This meta-analysis aims to evaluate the efficacy of bone marrow-derived mononuclear cell (BMMNC) therapy in patients with acute myocardial infarction, but also explores the effect of newer generations of stem cells. METHODS AND RESULTS: A random-effects meta-analysis was performed on randomized controlled trials investigating the effects of stem cell therapy in patients with acute myocardial infarction that were published between January 2002 and September 2013. The defined end points were left ventricular (LV) ejection fraction, LV end-systolic and end-diastolic volumes, infarct size, and major adverse cardiac and cerebrovascular event rates. Also, several subgroup analyses were performed on BMMNC trials. Overall, combining the results of 22 randomized controlled trials (RCTs), LV ejection fraction increased by +2.10% (95% confidence interval [CI], 0.68-3.52; P=0.004) in the BMMNC group as compared with controls, evoked by a preservation of LV end-systolic volume (-4.05 mL; 95% CI, -6.91 to -1.18; P=0.006) and a reduction in infarct size (-2.69%; 95% CI, -4.83 to -0.56; P=0.01). However, there is no effect on cardiac function, volumes, or infarct size, when only RCTs (n=9) that used MRI-derived end points were analyzed. Moreover, no beneficial effect could be detected on major adverse cardiac and cerebrovascular event rates after BMMNC infusion after a median follow-up duration of 6 months. CONCLUSIONS: Intracoronary infusion of BMMNC is safe, but does not enhance cardiac function on MRI-derived parameters, nor does it improve clinical outcome. New and possibly more potent stem cells are emerging in the field, but their clinical efficacy still needs to be defined in future trials.

Absence of chemokine (C-x-C motif) ligand 10 diminishes perfusion recovery after local arterial occlusion in mice.

OBJECTIVE: In arteriogenesis, pre-existing anastomoses undergo enlargement to restore blood flow in ischemic tissues. Chemokine (C-X-C motif) ligand 10 (CXCL10) is secreted after Toll-like receptor activation. Toll-like receptors are involved in arteriogenesis; however, the role of CXCL10 is still unclear. In this study, we investigated the role for CXCL10 in a murine hindlimb ischemia model. APPROACH AND RESULTS: Unilateral femoral artery ligation was performed in wild-type (WT) and CXCL10(-/-) knockout (KO) mice and perfusion recovery was measured using laser-Doppler perfusion analysis. Perfusion recovery was significantly lower in KO mice compared with WT at days 4 and 7 after surgery (KO versus WT: 28±5% versus 81±13% at day 4; P=0.003 and 57±12% versus 107±8% at day 7; P=0.003). Vessel measurements of α-smooth muscle actin-positive vessels revealed increasing numbers in time after surgery, which was significantly higher in WT when compared with that in KO. Furthermore, α-smooth muscle actin-positive vessels were significantly larger in WT when compared with those in KO at day 7 (wall thickness, P<0.001; lumen area, P=0.003). Local inflammation was assessed in hindlimb muscles, but this did not differ between WT and KO. Chimerization experiments analyzing perfusion recovery and histology revealed an equal contribution for bone marrow-derived and circulating CXCL10. Migration assays showed a stimulating role for both intrinsic and extrinsic CXCL10 in vascular smooth muscle cell migration. CONCLUSIONS: CXCL10 plays a causal role in arteriogenesis. Bone marrow-derived CXCL10 and tissue-derived CXCL10 play a critical role in accelerating perfusion recovery after arterial occlusion in mice probably by promoting vascular smooth muscle cell recruitment and maturation of pre-existing anastomoses.

Clinical study using adipose-derived mesenchymal-like stem cells in acute myocardial infarction and heart failure.

Adipose tissue represents an abundant, accessible source of regenerative cells that can be easily obtained in sufficient amount for therapy. Adipose-derived regenerative cells (ADRC) are comprised of leukocytes, smooth muscles, endothelial cells, and mesenchymal stem cells. In contrast to bone-marrow-derived MSC, the abundance of adipose tissue in patients and the higher frequency per unit mass of regenerative cells allow for the isolation of cells in therapeutic meaningful amounts in less than 2h after donor tissue acquisition.Harvest of adipose tissue can thus follow primary PCI, allowing efficient treatment within 24h. This obviates the need for extensive cell culturing in GMP clean room facilities and makes ADSCs a promising and practical autologous cell source. In the following chapter, we will describe the liposuction procedure for stem cell harvest, two cell delivery techniques, and pressure/volume loop analysis for the follow-up of our patients enrolled in the clinical studies.

Intracoronary infusion of allogeneic mesenchymal precursor cells directly after experimental acute myocardial infarction reduces infarct size, abrogates adverse remodeling, and improves cardiac function.

RATIONALE: Mesenchymal precursor cells (MPCs) are a specific Stro-3+ subpopulation of mesenchymal stem cells isolated from bone marrow. MPCs exert extensive cardioprotective effects, and are considered to be immune privileged. OBJECTIVE: This study assessed the safety, feasibility, and efficacy of intracoronary delivery of allogeneic MPCs directly after acute myocardial infarction in sheep. METHODS AND RESULTS: Initially, intracoronary delivery conditions were optimized in 20 sheep. These conditions were applied in a randomized study of 68 sheep with an anterior acute myocardial infarction. Coronary flow was monitored during MPC infusion, and cardiac function was assessed using invasive hemodynamics and echocardiography at baseline and during 8 weeks follow-up. Coronary flow remained within thrombolysis in myocardial infarction III definitions in all sheep during MPC infusion. Global left ventricular ejection fraction as measured by pressure-volume loop analysis deteriorated in controls to 40.7±2.6% after 8 weeks. In contrast, MPC treatment improved cardiac function to 52.8±0.7%. Echocardiography revealed significant improvement of both global and regional cardiac functions. Infarct size decreased by 40% in treated sheep, whereas infarct and border zone thickness were enhanced. Left ventricular adverse remodeling was abrogated by MPC therapy, resulting in a marked reduction of left ventricular volumes. Blood vessel density increased by >50% in the infarct and border areas. Compensatory cardiomyocyte hypertrophy was reduced in border and remote segments, accompanied by reduced collagen deposition and apoptosis. No microinfarctions in remote myocardial segments or histological abnormalities in unrelated organs were found. CONCLUSIONS: Intracoronary infusion of allogeneic MPCs is safe, feasible, and markedly effective in a large animal model of acute myocardial infarction.

Feasibility of intracoronary GLP-1 eluting CellBead™ infusion in acute myocardial infarction.

Cell therapy is a field of growing interest in the prevention of post acute myocardial infarction (AMI) heart failure. Stem cell retention upon local delivery to the heart, however, is still unsatisfactory. CellBeads were recently developed as a potential solution to this problem. CellBeads are 170-µm alginate microspheres that contain mesenchymal stem cells (MSCs) genetically modified to express glucagon-like peptide-1 (GLP-1) supplementary to inherent paracrine factors. GLP-1 is an incretin hormone that has both antiapoptotic and cardioprotective effects. Transplanting CellBeads in the post-AMI heart might induce cardiomyocyte salvage and ultimately abrogate adverse cardiac remodeling. We aimed to investigate the feasibility of intracoronary infusion of CellBeads in a large animal model of AMI. Four pigs were used in a pilot study to assess the maximal safe dose of CellBeads. In the remaining 21 animals, an AMI was induced by balloon occlusion of the left circumflex coronary artery for 90 min. During reperfusion, 60,000 CellBeads (n = 11), control beads (n = 4), or lactated Ringers' (n = 6) were infused. Animals were sacrificed after 2 or 7 days, and the hearts were excised for histological analyses. Intracoronary infusion did not permanently affect coronary flow in any of the groups. Histological analysis revealed CellBeads containing viable MSCs up to 7 days. Viability and activity of the MSCs was confirmed by qPCR analysis that showed expression of recombinant GLP-1 and human genes after 2 and 7 days. CellBeads reduced inflammatory infiltration by 29% (p = 0.001). In addition, they decreased the extent of apoptosis by 25% (p = 0.001) after 2 days. We show that intracoronary infusion of 5 million encapsulated MSCs is safe and feasible. Also, several parameters indicate that the cells have paracrine effects, suggesting a potential therapeutic benefit of this new approach.

Uridine adenosine tetraphosphate is a novel vasodilator in the coronary microcirculation which acts through purinergic P1 but not P2 receptors.

Uridine adenosine tetraphosphate (Up4A) has been identified as an endothelium-derived contracting factor, which acts through purinergic P2X and P2Y receptors. Since the coronary vascular actions of Up4A are unknown, we investigated the vasoactive profile of Up4A in coronary microvessels, and studied the involvement of purinergic receptor subtypes. Studies were performed in isolated porcine coronary small arteries (diameter∼250 µm), with and without endothelial denudation, mounted on a Mulvany wire myograph. Purinergic receptor expression was assessed by real-time PCR. Up4A (10(-9)-10(-5) M) failed to induce contraction at basal tone, but produced concentration-dependent vasorelaxation in precontracted microvessels. Up4A was slightly less potent than adenosine, ATP, and ADP in producing vasorelaxation, but significantly more potent than UTP and UDP. mRNA expression of P2X(4), P2Y(1), P2Y(2), P2Y(4), P2Y(6) and A(2A), but not P2X(1), receptors was observed. Up4A-induced vasodilation was unaffected by non-selective P2 receptor antagonist PPADS, P2X(1) antagonist MRS2159, P2Y(1) antagonist MRS2179 and P2Y(6) antagonist MRS2578, but was markedly attenuated by non-selective P1 receptor antagonist 8PT and A(2A) antagonist SCH58261. Up4A-induced vasodilation was not affected by ectonucleotidase inhibitor ARL67156, suggesting that A(2A) stimulation was not the result of Up4A breakdown to adenosine. Up4A-induced vasodilation was blunted in denuded vessels; additional A(2A) receptor blockade further attenuated Up4A-induced vasodilation, suggesting that A(2A) receptor-mediated vasodilation is only partly endothelium-dependent. In conclusion, Up4A exerts a vasodilator rather than a vasoconstrictor influence in coronary microvessels, which is mediated via A(2A) receptors and is partly endothelium-dependent.

Do we have a future with transcatheter adventitial delivery of stem cells?

Critically evaluating the methodology of the adventitial delivery of stem cells, some specific options should be underlined. Adventitia as the most superficial layer, consisting of connective tissue has to be distinguished of perivascular tissues. By strict definition, an adventitia is the outermost connective tissue covering any organ, or vessel. The "adventitial" delivery of stem cells with a 1mm micro-needle means a delivery to superficial so called pericardial myocardium, perivascular fat tissues, including a risk of perforation and injury of soft tissues. In fact, the mapping of the artery with visualization of the three-layer vessel structure and perivascular tissues as well as pericardial space with the state-of-the-art imaging approaches including IVUS (intravascular ultrasound) or OCT (optical coherence tomography) allows to find an optimal site for injection, prevents any technical complications and improves efficacy. NOGA magnetic navigation system still remains the optimal tool for the stem cell delivery to myocardium with appropriate visualization of necrosis and peri-infarct tissues. Potentially, more advanced imaging provides a chance to deliver infusate to the adventitial layer, which is a gate to the vessel wall for inflammation as well as a source of stem and progenitor cells, and myofibroblasts.

Apelin enhances cardiac neovascularization after myocardial infarction by recruiting aplnr+ circulating cells.

RATIONALE: Neovascularization stimulated by local or recruited stem cells after ischemia is a key process that salvages damaged tissue and shows similarities with embryonic vascularization. Apelin receptor (Aplnr) and its endogenous ligand apelin play an important role in cardiovascular development. However, the role of apelin signaling in stem cell recruitment after ischemia is unknown. OBJECTIVE: To investigate the role of apelin signaling in recruitment after ischemia. METHODS AND RESULTS: Aplnr was specifically expressed in circulating cKit+/Flk1+ cells but not in circulating Sca1+/Flk1+ and Lin+ cells. cKit+/Flk1+/Aplnr+ cells increased significantly early after myocardial ischemia but not after hind limb ischemia, indicative of an important role for apelin/Aplnr in cell recruitment during the nascent biological repair response after myocardial damage. In line with this finding, apelin expression was upregulated in the infarcted myocardium. Injection of apelin into the ischemic myocardium resulted in accelerated and increased recruitment of cKit+/Flk1+/Aplnr+ cells to the heart. Recruited Aplnr+/cKit+/Flk1+ cells promoted neovascularization in the peri-infarct area by paracrine activity rather than active transdifferentiation, resulting into cardioprotection as indicated by diminished scar formation and improved residual cardiac function. Aplnr knockdown in the bone marrow resulted in aggravation of myocardial ischemia-associated damage, which could not be rescued by apelin. CONCLUSIONS: We conclude that apelin functions as a new and potent chemoattractant for circulating cKit+/Flk1+/Aplnr+ cells during early myocardial repair, providing myocardial protection against ischemic damage by improving neovascularization via paracine action.

Mast cells induce vascular smooth muscle cell apoptosis via a toll-like receptor 4 activation pathway.

OBJECTIVE: Activated mast cells (MCs) release chymase, which can induce vascular smooth muscle cell (VSMC) apoptosis leading to plaque destabilization. Because the mechanism through which MCs release chymase in atherosclerosis is unknown, we studied whether MC-associated VSMC apoptosis is regulated by toll-like receptor 4 (TLR4) signaling. METHODS AND RESULTS: Local recruitment and activation of MCs reduced VSMC content specifically in the cap region of vulnerable plaques in apolipoprotein E knockout mice. Cotreatment with the TLR4 antagonist Bartonella quintana lipopolysaccharide prevented this VSMC loss, suggesting an important role for TLR4 signaling in MC-induced VSMC apoptosis. Coculture of VSMCs with MCs activated by the TLR4 agonist Escherichia coli lipopolysaccharide increased VSMC apoptosis. Apoptosis was inhibited by TLR4 and chymase blockers, indicating that TLR4 signaling is involved in chymase release in MCs. This pathway was mediated via interleukin-6 because interleukin-6 promoted MC-associated VSMC apoptosis, which was inhibited by blocking chymase release. In addition, TLR4 activation in MCs induced interleukin-6 production, which was reduced by preincubation with either B. quintana lipopolysaccharide or an anti-TLR4 antibody. CONCLUSIONS: We show that MCs promote VSMC apoptosis in vivo. In addition, TLR4 signaling is important in chymase release in MCs and, therefore, in plaque destabilization by regulating VSMC apoptosis.

PDGF-induced migration of vascular smooth muscle cells is inhibited by heme oxygenase-1 via VEGFR2 upregulation and subsequent assembly of inactive VEGFR2/PDGFRß heterodimers.

OBJECTIVE: In cardiovascular regulation, heme oxygenase-1 (HO-1) activity has been shown to inhibit vascular smooth muscle cell (VSMC) proliferation by promoting cell cycle arrest at the G1/S phase. However, the effect of HO-1 on VSMC migration remains unclear. We aim to elucidate the mechanism by which HO-1 regulates PDGFBB-induced VSMC migration. METHODS AND RESULTS: Transduction of HO-1 cDNA adenoviral vector severely impeded human VSMC migration in a scratch, transmembrane, and directional migration assay in response to PDGFBB stimulation. Similarly, HO-1 overexpression in the remodeling process during murine retinal vasculature development attenuated VSMC coverage over the major arterial branches as compared with sham vector-transduced eyes. HO-1 expression in VSMCs significantly upregulated VEGFA and VEGFR2 expression, which subsequently promoted the formation of inactive PDGFRß/VEGFR2 complexes. This compromised PDGFRß phosphorylation and impeded the downstream cascade of FAK-p38 signaling. siRNA-mediated silencing of VEGFA or VEGFR2 could reverse the inhibitory effect of HO-1 on VSMC migration. CONCLUSIONS: These findings identify a potent antimigratory function of HO-1 in VSMCs, a mechanism that involves VEGFA and VEGFR2 upregulation, followed by assembly of inactive VEGFR2/PDGFRß complexes that attenuates effective PDGFRß signaling.

CCBE1 is essential for mammalian lymphatic vascular development and enhances the lymphangiogenic effect of vascular endothelial growth factor-C in vivo.

RATIONALE: Collagen- and calcium-binding EGF domains 1 (CCBE1) has been associated with Hennekam syndrome, in which patients have lymphedema, lymphangiectasias, and other cardiovascular anomalies. Insight into the molecular role of CCBE1 is completely lacking, and mouse models for the disease do not exist. OBJECTIVE: CCBE1 deficient mice were generated to understand the function of CCBE1 in cardiovascular development, and CCBE1 recombinant protein was used in both in vivo and in vitro settings to gain insight into the molecular function of CCBE1. METHODS AND RESULTS: Phenotypic analysis of murine Ccbe1 mutant embryos showed a complete lack of definitive lymphatic structures, even though Prox1(+) lymphatic endothelial cells get specified within the cardinal vein. Mutant mice die prenatally. Proximity ligation assays indicate that vascular endothelial growth factor receptor 3 activation appears unaltered in mutants. Human CCBE1 protein binds to components of the extracellular matrix in vitro, and CCBE1 protein strongly enhances vascular endothelial growth factor-C-mediated lymphangiogenesis in a corneal micropocket assay. CONCLUSIONS: Our data identify CCBE1 as a factor critically required for budding and migration of Prox-1(+) lymphatic endothelial cells from the cardinal vein. CCBE1 probably exerts these effects through binding to components of the extracellular matrix. CCBE1 has little lymphangiogenic effect on its own but dramatically enhances the lymphangiogenic effect of vascular endothelial growth factor-C in vivo. Thus, our data suggest CCBE1 to be essential but not sufficient for lymphangiogenesis.

Ets2 determines the inflammatory state of endothelial cells in advanced atherosclerotic lesions.

RATIONALE: Neovascularization is required for embryonic development and plays a central role in diseases in adults. In atherosclerosis, the role of neovascularization remains to be elucidated. In a genome-wide microarray-screen of Flk1+ angioblasts during murine embryogenesis, the v-ets erythroblastosis virus E26 oncogene homolog 2 (Ets2) transcription factor was identified as a potential angiogenic factor. OBJECTIVES: We assessed the role of Ets2 in endothelial cells during atherosclerotic lesion progression toward plaque instability. METHODS AND RESULTS: In 91 patients treated for carotid artery disease, Ets2 levels showed modest correlations with capillary growth, thrombogenicity, and rising levels of tumor necrosis factor-α (TNFα), monocyte chemoattractant protein 1, and interleukin-6 in the atherosclerotic lesions. Experiments in ApoE(-/-) mice, using a vulnerable plaque model, showed that Ets2 expression was increased under atherogenic conditions and was augmented specifically in the vulnerable versus stable lesions. In endothelial cell cultures, Ets2 expression and activation was responsive to the atherogenic cytokine TNFα. In the murine vulnerable plaque model, overexpression of Ets2 promoted lesion growth with neovessel formation, hemorrhaging, and plaque destabilization. In contrast, Ets2 silencing, using a lentiviral shRNA construct, promoted lesion stabilization. In vitro studies showed that Ets2 was crucial for TNFα-induced expression of monocyte chemoattractant protein 1, interleukin-6, and vascular cell adhesion molecule 1 in endothelial cells. In addition, Ets2 promoted tube formation and amplified TNFα-induced loss of vascular endothelial integrity. Evaluation in a murine retina model further validated the role of Ets2 in regulating vessel inflammation and endothelial leakage. CONCLUSIONS: We provide the first evidence for the plaque-destabilizing role of Ets2 in atherosclerosis development by induction of an intraplaque proinflammatory phenotype in endothelial cells.

NIRS and IVUS for characterization of atherosclerosis in patients undergoing coronary angiography.

OBJECTIVES: The aim of this study was to compare the findings of near-infrared spectroscopy (NIRS), intravascular ultrasound (IVUS) virtual histology (VH), and grayscale IVUS obtained in matched coronary vessel segments of patients undergoing coronary angiography. BACKGROUND: Intravascular ultrasound VH has been developed to add tissue characterization to the grayscale IVUS assessment of coronary plaques. Near-infrared spectroscopy is a new imaging technique able to identify lipid core-containing coronary plaques (LCP). METHODS: We performed NIRS and IVUS-VH pullbacks in a consecutive series of 31 patients with a common region of interest (ROI) between 2 side branches. For each ROI, we analyzed the chemogram blocks by NIRS, plaque area and plaque burden by grayscale IVUS, and tissue types by IVUS-VH. The chemogram block is a summary metric of a 2-mm vertical slice of the chemogram. The value ranges from 0 to 1 according to the presence of lipids and represents the probability of LCP with a color scale from red (low probability) through orange and tan to yellow (high probability). RESULTS: Plaque area (mm(2)) increases as percentage VH derived-necrotic core (NC) content (4.6 ± 2.7 vs. 7.4 ± 3.5 vs. 8.6 ± 3.4 vs. 7.9 ± 3.3, grouped in percentage NC quartiles, p<0.001) and chemogram block probability color bin thresholds increase (4.9 ± 3.8 red, 7.3 ± 3.6 orange, 8.1 ± 3.4 tan, and 8.7 ± 3.4 yellow, p<0.001). The correlation between the block chemogram detection of lipid core and percentage NC content by VH was weak (r=0.149). Correction for the presence of calcium does not improve this correlation. CONCLUSIONS: Larger plaque area by grayscale IVUS was more often associated with either elevated percentage VH-NC or LCP by NIRS; however, the correlation between the detection of LCP by NIRS and necrotic core by VH is weak.

Rheb is essential for murine development.

Ras homolog enriched in brain (Rheb) couples growth factor signaling to activation of the target of rapamycin complex 1 (TORC1). To study its role in mammals, we generated a Rheb knockout mouse. In contrast to mTOR or regulatory-associated protein of mTOR (Raptor) mutants, the inner cell mass of Rheb(-/-) embryos differentiated normally. Nevertheless, Rheb(-/-) embryos died around midgestation, most likely due to impaired development of the cardiovascular system. Rheb(-/-) embryonic fibroblasts showed decreased TORC1 activity, were smaller, and showed impaired proliferation. Rheb heterozygosity extended the life span of tuberous sclerosis complex 1-deficient (Tsc1(-/-)) embryos, indicating that there is a genetic interaction between the Tsc1 and Rheb genes in mouse.

Dendritic cell function in transplantation arteriosclerosis is regulated by heme oxygenase 1.

RATIONALE: Heme oxygenase (HO)1 is an important modulator of physiological function with cytoprotective properties. Although HO1 has previously been associated with an improved survival of the vascular allograft in rat models in response to pharmaceutical induction of HO1 the exact mechanism by which HO1 exerts it protective function remains to be elucidated. OBJECTIVE: We sought to define the role of HO1 in dendritic cells (DCs) function that governs the alloimmune response underlying the development of transplantation associated vasculopathy. METHODS AND RESULTS: Loss of HO1 in DCs or by small interfering RNA silencing resulted in major histocompatibility complex class II (MHCII) upregulation by CIITA- driven transcriptional regulation and by STAT1 (signal transducers and activators of transcription 1) phosphorylation. As a result, increased MHCII alloantigen presentation by HO1(-/-) DCs directed the primary T-cell response preferentially toward a CD4(+) T-cell, rather than a CD8(+) T-cell reaction. In a murine model for transplantation arteriosclerosis, adoptive transfer of HO1(-/-) DCs before allograft transplantation was indeed associated with pronounced intragraft CD4(+) T-cell infiltration and increased IgG deposition, suggestive of an accelerated development of vasculopathy toward the chronic phase. The role of HO1 in DC-mediated T cell activation was further validated by inhibition of endogenous HO1 in allograft recipients. Inhibition of HO1 in DCs aggravated transplant arteriosclerosis development, by increasing intima hyperplasia, and by activation of a CD4(+) T cells allograft response, mediated by MHCII upregulation. CONCLUSIONS: These findings demonstrate that HO1 plays an important role in the genetic regulation of the vascular alloimmune response elicited by DCs.