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Objective— Transplant-associated arteriosclerosis manifests as progressive vascular neointimal expansion throughout the arterial system of allografted solid organs, and eventually compromises graft perfusion and function. Allografts placed in colony stimulating factor (CSF)-1-deficient osteopetrotic ( Csf1 op /Csf1 op ) mice develop very little neointima, a finding attributed to impaired recipient macrophage function. We examined how CSF-1 affects neointima-derived vascular smooth muscle cells, tested the significance of CSF-1 expressed in donor tissue, and evaluated the contribution of secreted versus cell surface CSF-1 isoforms in transplant-associated arteriosclerosis. Methods and Results— CSF-1 activated specific signaling pathways to promote migration, survival, and proliferation of cultured vascular smooth muscle cells. Tumor necrosis factor-α addition increased CSF-1 and CSF-1 receptor expression, and tumor necrosis factor-α-driven proliferation was blocked by anti-CSF-1 antibody. In a mouse vascular allograft model, lack of recipient or donor CSF-1 impaired neointima formation; the latter suggests local CSF-1 function within the allograft. Moreover, reconstitution of donor or recipient cell surface CSF-1, without secreted CSF-1, restored neointimal formation. Conclusion— Vascular smooth muscle cells activation, including that mediated by tumor necrosis factor-α, can be driven in an autocrine/juxtacrine manner by CSF-1. These studies provide evidence for local function of CSF-1 in neointimal expansion, and identify CSF-1 signaling in vascular smooth muscle cells, particularly cell surface CSF-1 signaling, as a target for therapeutic strategies in transplant-associated arteriosclerosis.

To determine the ability of simple hemodynamic parameters obtained at the time of cardiac catheterization to predict in-hospital mortality following ST-elevation myocardial infarction (STEMI).Hemodynamic parameters measured at the time of primary percutaneous coronary intervention (PPCI) could potentially identify high-risk patients who would benefit from aggressive hemodynamic support in the Cardiac Catheterization laboratory.This is a retrospective single-center study of 219 consecutive patients with STEMI. Left ventricular end-diastolic pressure (LVEDP), systolic blood pressure (SBP), and aortic diastolic blood pressure were obtained after successful revascularization. The prognostic ability of LVEDP, pulse pressure, and SBP/LVEDP ratio were compared to major mortality risk scores.Patients had a mean age of 60 ±14 years, were predominantly white (73%), male (64%), with anterior wall infarcts in 39%. Comorbidities included diabetes mellitus (27%), heart failure (9%), and chronic kidney disease (7%). In-hospital mortality was 9%. Patients with SBP/LVEDP ≤ 4 had increased risk of in-hospital death (32% vs. 5.3%, P 0.0001), intra-aortic balloon pump (IABP) usage (51.6% vs. 9.6%, P 0.0001) and combined endpoint of death or IABP usage (58.1% vs. 13.3%, P 0.0001) compared to patients with SBP/LVEDP 4. The area under curve (AUC) for SBP/LVEDP ratio for in-hospital mortality (0.69) was more predictive than LVEDP (0.61, P = 0.04) or pulse pressure (0.55, P = 0.02) but similar to Shock Index (ratio of heart rate to SBP) and Modified Shock Index (ratio of HR to mean arterial pressure).An SBP/LVEDP ratio ≤ 4 identified a group of STEMI patients at high risk of in-hospital death. © 2017 Wiley Periodicals, Inc.

A laser-generated-focused ultrasound (LGFU) transducer generates high-pressure (up to 20 MPa), high-frequency (>10 MHz) shock waves with a tight focal spot. In this work, we aim to demonstrate the feasibility of using LGFU transducers for sonothrombolysis in vitro. A carbon black LGFU transducer was designed, fabricated and characterized. The prototyped LGFU was applied with in-vitro thrombolysis tests involving microbubble contrast agent (MCA). A conventional piezo ultrasound transducer was used as a secondary excitation source to enhance the cavitation effect by dual-frequency excitation. The in vitro test results showed that microbubble-mediated LGFU treatment can yield the lytic rate of approximately 2 mg/min, suggesting that LGFU transducers may be useful in precision high lytic rate sonothrombolysis.

Without early revascularization, both inpatient and outpatient STEMIs have poor outcomes. Reasons for denying PCI for STEMI, however, remain uncertain. This single-center retrospective cohort study compares factors and outcomes associated with ineligibility for PCI between inpatients and outpatients following ST-elevation myocardial infarction (STEMI).A total of 1,759 STEMI patients between June 2009 and January 2015 were assessed. Individual medical records were reviewed to obtain reasons for PCI ineligibility for STEMI patients who did not receive reperfusion therapy.Compared to outpatients with STEMI (n = 1,688), inpatients (n = 71) were less likely to receive coronary angiography (60.6% vs 95.9%; P 0.001) or PCI (50.7% vs 80.9%; P 0.001), with longer ECG/door to first device activation times (97 [78, 131] vs 63 [49, 78] minutes; P 0.001). When coronary angiography was performed, however, similar rates of PCI and procedural success were seen in both groups. Principal contraindication for PCI was risk of bleeding within the inpatient population and complex coronary artery disease within the outpatient population. Total in-hospital mortality was higher in inpatient STEMIs compared to outpatients (42.2% vs 10.0%; P 0.001), but lower for patients eligible for PCI in both groups.Reasons for PCI ineligibility differ between inpatient and outpatient STEMIs. Inpatients have increased risks of bleeding, lower coronary angiography and PCI use, and higher in-hospital mortality. Especially for inpatients, specific PCI STEMI protocols that anticipate and overcome types of ineligibility and delay for cardiac catheterization may improve outcomes.

Cardiovascular disease has become a leading cause of death for individuals with paraplegia. This is the first clinical study in the literature to investigate the clinical outcomes and treatment of AMI patients with paraplegia. We identified AMI patients with paraplegia cohort by using principal

Pericardial constriction from tuberculous pericarditis is a dreaded but rare sequelae in developed countries. We present a case of effusive-constrictive pericarditis in a man under acute treatment for pulmonary tuberculosis. A 63 year old Korean man under acute treatment for pulmonary tuberculosis

Cardiovascular disease has become a leading cause of death for patients with paraplegia. Acute myocardial infarction in patients with paraplegia has not been described in the literature. This study investigates clinical features, management strategies, and outcomes of these patients.Acute myocardial infarction in patients with or without paraplegia was identified in the New York State Inpatient Database between 2007 and 2013. Clinical comorbidities, management strategies and their associated outcomes were compared using propensity score-matching analysis.Among 402,569 patients with acute myocardial infarction, 1400 had a concomitant diagnosis of paraplegia. Compared with those without, patients with paraplegia were younger, more likely to be black, and had a higher prevalence of hypertension, anemia, congestive heart failure, coagulopathy, and depression, but a lower prevalence of diabetes, hyperlipidemia, obesity, chronic lung disease, and renal failure. Patients with paraplegia were more likely to receive medical therapy without a diagnostic cardiac catheterization than those without (83.7% vs 64.5%, P .001). Nine percent of patients with paraplegia received revascularization, which was significantly lower than that without paraplegia. In terms of the clinical outcome, patients with paraplegia had higher in-hospital mortality than those without (22.4% vs 16.8%, P .001). Among the patients with paraplegia, the subcohort that received revascularization had lower in-hospital mortality (9.5% vs 22.0%, P .01), had shorter length of stay (13.0 vs 16.9 days, P =.08), and higher hospital charges ($130,079 vs $92,125, P .001) than those without revascularization. Furthermore, the paraplegic subcohort underwent coronary artery bypass grafting that was associated with higher in-hospital mortality (21.7% vs 1.7%, P .001), longer length of stay (24.8 vs 14.2 days, P .001), and higher hospital charges ($231,323 vs $144,449, P .01) than subcohort that received percutaneous coronary intervention.Acute myocardial infarction patients with concomitant paraplegia had distinct clinical characteristics and comorbidity profiles; were less likely to receive revascularization therapy; and had higher in-hospital mortality. Acute myocardial infarction patient with paraplegia who underwent revascularization were associated with better clinical outcomes, in particular, those who were treated with percutaneous coronary intervention had significantly lower in-hospital mortality than those treated with coronary artery bypass grafting.

OBJECTIVE— Chronic pancreatitis, characterized by pancreatic exocrine tissue destruction with initial maintenance of islets, eventually leads to insulin-dependent diabetes in most patients. Mice deficient for the transcription factors E2F1 and E2F2 suffer from a chronic pancreatitis-like syndrome and become diabetic. Surprisingly, onset of diabetes can be prevented through bone marrow transplantation. The goal of the described studies was to determine the hematopoietic cell type responsible for maintaining islets and the associated mechanism of this protection. RESEARCH DESIGN AND METHODS— Mouse models of acute and chronic pancreatitis, together with mice genetically deficient for macrophage production, were used to determine roles for macrophages in islet angiogenesis and maintenance. RESULTS— We demonstrate that macrophages are essential for preventing endocrine cell loss and diabetes. Macrophages expressing matrix metalloproteinase-9 migrate to the deteriorating pancreas. E2f1/E2f2 mutant mice transplanted with wild-type, but not macrophage-deficient colony stimulating factor 1 receptor mutant (Csf1r−/−), bone marrow exhibit increased angiogenesis and proliferation within islets, coinciding with increased islet mass. A similar macrophage dependency for islet and islet vasculature maintenance is observed during caerulein-induced pancreatitis. CONCLUSIONS— These findings demonstrate that macrophages promote islet angiogenesis and protect against islet loss during exocrine degeneration, could explain why most patients with chronic pancreatitis develop diabetes, and suggest an avenue for preventing pancreatitis-associated diabetes.

Intracerebral hemorrhage (ICH) is an infrequent but severe complication in pregnant women with hypertension. The authors describe a patient with chronic hypertension who developed superimposed preeclampsia and spontaneous ICH during the thirty‐fifth week of pregnancy. ICH was diagnosed by computed tomographic scan. She underwent successful emergent cesarean section and neurosurgical decompression of the ICH. Both intraoperative surveillance and postoperative magnetic resonance angiographic examination of the cerebral vessels failed to identify an aneurysm or arteriovenous malformation. The authors discuss the diagnosis and management in this case and review the literature regarding this challenging complication of pregnancy and preeclampsia. Controversies regarding treatment of hypertension during pregnancy are discussed in light of the impact on the management of this patient.

CSF-1 is the primary mononuclear phagocyte and osteoclast growth factor. Autocrine regulation by CSF-1 has been reported in macrophages during inflammatory responses and in neoplastic cells. To investigate whether inflammatory disease or neoplasia was the dominant consequence of autocrine regulation by CSF-1 in CSF-1 receptor (CSF-1R)-expressing cells, we created mice that express CSF-1 under the control of the CSF-1R promoter/first intron driver [transgene TgN(Csf1r-Csf1)Ers (TgRC) mice], which have reduced thymic size, a short lifetime, and low body weight and develop osteoporosis. In 4-week-old TgRC mice, osteoclast numbers are elevated, and macrophage densities are increased in bone marrow, spleen, liver, and brain. Cultured TgRC macrophages express CSF-1 and proliferate without exogenous CSF-1 and in the presence of neutralizing antimouse CSF-1 antibody. Compared with macrophages from nontransgenic littermates, TgRC macrophages exhibit a stellate morphology, express elevated mRNAs for proinflammatory cytokines, and despite a lower, steady-state cytokine secretion, secrete elevated levels of inflammatory cytokines in response to LPS, indicating that TgRC macrophages are functionally primed through the CSF-1R. Thus, autocrine regulation of CSF-1R-expressing cells by CSF-1 leads to a severe phenotype that emphasizes the importance of the known, local production of CSF-1 in inflammatory disease.

BCL-6 is a transcription repressor frequently deregulated in non-Hodgkin’s B cell lymphomas. Its activity is also critical to germinal center development and balanced Th1/Th2 differentiation. Previous studies have suggested that NF-κB activity is suppressed in germinal center and lymphoma B cells that express high levels of BCL-6, and yet the reason for this is unknown. We report in this study that BCL-6 can bind to three sequence motifs in the 5′ regulatory region of NF-κB1 in vitro and in vivo, and repress NF-κB1 transcription both in reporter assays and in lymphoma B cell lines. BCL-6−/− mice further confirm the biological relevance of BCL-6-dependent regulation of NF-κB1 because BCL-6 inactivation caused notable increase in p105/p50 proteins in several cell types. Among these, BCL-6−/− macrophage cell lines displayed a hyperproliferation phenotype that can be reversed by NF-κB inhibitors, e.g., N-tosyl-l-phenylalanine chloromethyl ketone and SN50, a result that is consistent with increased nuclear κB-binding activity of p50 homodimer and p50/p65 heterodimer. Our results demonstrate that BCL-6 can negatively regulate NF-κB1 expression, thereby inhibiting NF-κB-mediated cellular functions.

Reperfusion times for ST-elevation myocardial infarction (STEMI) occurring in outpatients have improved significantly, but quality improvement efforts have largely ignored STEMI occurring in hospitalized patients (inpatient-onset STEMI).To define the incidence and variables associated with treatment and outcomes of patients who develop STEMI during hospitalization for conditions other than acute coronary syndromes (ACS).Retrospective observational analysis of STEMIs occurring between 2008 and 2011 as identified in the California State Inpatient Database.STEMIs were classified as inpatient onset or outpatient onset based on present-on-admission codes. Patients who had a STEMI after being hospitalized for ACS were excluded from the analysis.Regression models were used to evaluate associations among location of onset of STEMI, resource utilization, and outcomes. Adjustments were made for patient age, sex, comorbidities, and hospital characteristics. The analysis allowed for the location of inpatient STEMI to have a multiplicative rather than an additive effect for resource utilization since these measures were highly skewed.A total of 62,021 STEMIs were identified in 303 hospitals, of which 3068 (4.9%) occurred in patients hospitalized for non-ACS indications. Patients with inpatient-onset STEMI were older (mean, 71.5 [SD, 13.5] years vs 64.9 [SD, 14.1] years; P .001) and more frequently female (47.4% vs 32%; P .001) than those with outpatient-onset STEMI. Patients with inpatient-onset STEMI had higher in-hospital mortality (33.6% vs 9.2%; adjusted odds ratio (AOR), 3.05; 95% CI, 2.76-3.38; P .001), were less likely to be discharged home (33.7% vs 69.4%; AOR, 0.38; 95% CI, 0.34-0.42; P .001), and were less likely to undergo cardiac catheterization (33.8% vs 77.8%; AOR, 0.19; 95% CI, 0.16-0.21; P .001) or percutaneous coronary intervention (21.6% vs 65%; AOR, 0.23; 95% CI, 0.21-0.26; P .001). Length of stay and inpatient charges were higher for inpatient-onset STEMI (mean length of stay, 13.4 days [95% CI, 12.8-14.0 days] vs 4.7 days [95% CI, 4.6-4.8 days]; adjusted multiplicative effect, 2.51; 95% CI, 2.35-2.69; P .001; mean inpatient charges, $245,000 [95% CI, $235,300-$254,800] vs $129,000 [95% CI, $127,900-$130,100]; adjusted multiplicative effect, 2.09; 95% CI, 1.93-2.28; P .001).Patients who had a STEMI while hospitalized for a non-ACS condition, compared with those with onset of STEMI as an outpatient, were less likely to undergo invasive testing or intervention and had a higher in-hospital mortality rate.

Background Major advances have been made in the treatment of ST ‐elevation myocardial infarction ( STEMI ) in outpatients. In contrast, little is known about outcomes in STEMI that occur in patients hospitalized for a noncardiac condition. Methods and Results This was a retrospective, single‐center study of inpatient STEMI s from January 1, 2007, to July 31, 2011. Forty‐eight cases were confirmed to be inpatient STEMI s of a total of 139 410 adult discharges. These patients were older and more often female and had higher rates of chronic kidney disease and prior cerebrovascular events compared with 227 patients with outpatient STEMI s treated during the same period. Onset of inpatient STEMI was heralded most frequently by a change in clinical status (60%) and less commonly by patient complaints (33%) or changes on telemetry. Coronary angiography and percutaneous coronary intervention were performed in 71% and 56% of patients, respectively. The median time to obtain ECG (41 [10, 600] versus 5 [2, 10] minutes; P ECG to angiography time (91 [26, 209] versus 35 [25, 46] minutes; P ECG to first device activation ( FDA ) (129 [65, 25] versus 60 [47, 76] minutes; P STEMI . Survival to discharge was lower for inpatient STEMI (60% versus 96%; P Conclusions Patients who develop a STEMI while hospitalized for a noncardiac condition are older and more often female, have more comorbidities, have longer ECG ‐to‐ FDA times, and are less likely to survive than patients with an outpatient STEMI .

Multiple scoring systems that incorporate clinical information, vital signs and physical examination have been derived to predict mortality in patients with ST-elevation myocardial infarction (STEMI). The purpose of this study was to determine the ability of simple hemodynamic parameters obtained at

The CSF-1 receptor (CSF-1R) regulates CNS microglial development. However, the localization and developmental roles of this receptor and its ligands, IL-34 and CSF-1, in the brain are poorly understood. Here we show that compared to wild type mice, CSF-1R-deficient (Csf1r−/−) mice have smaller brains of greater mass. They further exhibit an expansion of lateral ventricle size, an atrophy of the olfactory bulb and a failure of midline crossing of callosal axons. In brain, IL-34 exhibited a broader regional expression than CSF-1, mostly without overlap. Expression of IL-34, CSF-1 and the CSF-1R were maximal during early postnatal development. However, in contrast to the expression of its ligands, CSF-1R expression was very low in adult brain. Postnatal neocortical expression showed that CSF-1 was expressed in layer VI, whereas IL-34 was expressed in the meninges and layers II–V. The broader expression of IL-34 is consistent with its previously implicated role in microglial development. The differential expression of CSF-1R ligands, with respect to CSF-1R expression, could reflect their CSF-1R-independent signaling. Csf1r−/− mice displayed increased proliferation and apoptosis of neocortical progenitors and reduced differentiation of specific excitatory neuronal subtypes. Indeed, addition of CSF-1 or IL-34 to microglia-free, CSF-1R-expressing dorsal forebrain clonal cultures, suppressed progenitor self-renewal and enhanced neuronal differentiation. Consistent with a neural developmental role for the CSF-1R, ablation of the Csf1r gene in Nestin-positive neural progenitors led to a smaller brain size, an expanded neural progenitor pool and elevated cellular apoptosis in cortical forebrain. Thus our results also indicate novel roles for the CSF-1R in the regulation of corticogenesis.

Rationale : The collateral circulation is tissue- and life-saving in obstructive arterial disease. Disappointing outcomes in clinical trials aimed at augmenting collateral growth highlight the need for greater understanding of collateral biology. Objective : The role of endothelial nitric oxide synthase (eNOS) in forming native (preexisting) collaterals and remodeling in obstructive disease are unknown or controversial issues, respectively. Methods and Results : We compared the native collateral circulation in healthy tissue and collateral remodeling after femoral artery ligation (FAL) in wild-type and eNOS-knockout (KO) mice. Perfusion after FAL fell further in adult eNOS-KOs, in association with fewer native collaterals in hindlimb (confirmed in brain). This was not attributable to impaired collateral formation in the embryo-neonate, but rather from collateral loss during growth to adulthood. Compared to wild-type, eNOS-KOs evidenced reduced collateral remodeling, angiogenesis, and flow-mediated dilation of the arterial bed supplying the collaterals, resulting in lower perfusion and greater ischemic injury at all time points over 21 days following FAL. To probe the mechanism for impaired remodeling, we performed genome-wide expression profiling of isolated, remodeling hindlimb collaterals 24 hour after FAL. Upregulation of genes encoding cytokines/chemokines, inflammatory, stress response, and cell cycle proteins was evident in wild-type mice. In contrast, expression was lower in 40 of 44 cell cycle genes in eNOS-KO mice, in association with impaired proliferation of vascular wall cells. Conclusions : Our findings suggest a novel role for eNOS in maintaining native collateral density during natural growth to adulthood and in collateral remodeling in obstructive disease, the latter through regulation of cell proliferation.

Rationale: Collaterals are arteriole-to-arteriole anastomoses that connect adjacent arterial trees. They lessen ischemic tissue injury by serving as endogenous bypass vessels when the trunk of 1 tree becomes narrowed by vascular disease. The number and diameter (“extent”) of native (preexisting) collaterals, plus their amount of lumen enlargement (growth/remodeling) in occlusive disease, show remarkably wide variation among inbred mouse strains (eg, C57BL/6 and BALB/c), resulting in large differences in tissue injury in models of occlusive disease. Evidence suggests similar large differences exist among healthy humans. Objective: To identify candidate loci responsible for genetic-dependent collateral variation. Methods and Results: Cerebral collateral number and diameter were determined in 221 C57BL/6×BALB/c F2 progeny, followed by linkage analysis to identify quantitative trait loci (QTL) for collateral number and diameter. Four QTL were obtained for collateral number, including epistasis between 2 loci. A QTL that was identical to the strongest QTL for collateral number on chromosome 7 (logarithm of the odds [LOD]=29, effect size=37%) was also mapped for collateral diameter (LOD=17, effect size=30%). Chromosome substitution strain analysis confirmed this locus. We also obtained a unique QTL on chromosome 11 for collateral remodeling after middle cerebral artery occlusion. Association mapping within the chromosome 7 QTL interval using collateral traits measured for 15 inbred strains delineated 172-kbp ( P =0.00002) and 290-kbp ( P =0.0004) regions on chromosome 7 containing 2 and 7 candidate genes, respectively. Conclusions: We conclude that collateral extent and remodeling are unique, highly heritable complex traits, with 1 QTL predominantly affecting native collateral number and diameter.

The evolution of drug-eluting stents (DES), effective periprocedural antithrombotic therapy, and advanced interventional techniques have fueled the surge of percutaneous coronary interventions. Stent thrombosis remains a serious complication of coronary artery stent implantation. Long-term antiplatelet therapy is required to prevent stent thrombosis, especially following DES implantation. Discontinuation of antiplatelet therapy (particularly clopidogrel) is the strongest independent risk factor for the development of stent thrombosis. Bleeding complications, most of which arise from the upper gastrointestinal (GI) tract, are the major limiting factors for antiplatelet therapy. The association of aspirin with the increased risk of upper GI bleeding has been well established. Peptic ulcer bleeding and Helicobacter pylori infection are the 2 most important risk factors for aspirin-associated GI bleeding complications. Endoscopy (for both surveillance and potential intervention), performed either emergently or semi-electively, is the primary tool for definitive management of GI bleeding. Considering the increase in GI bleeding risk seen with prolonged antiplatelet therapy, adjunctive proton pump inhibitor therapy and/or eradication of H. pylori infection might be beneficial for DES patients on long-term antiplatelet therapy.

The evolution of drug-eluting stents (DES), effective periprocedural antithrombotic therapy, and advanced interventional techniques have fueled the surge of percutaneous coronary interventions. Stent thrombosis remains a serious complication of coronary artery stent implantation. Long-term antiplatelet therapy is required to prevent stent thrombosis, especially following DES implantation. Discontinuation of antiplatelet therapy (particularly clopidogrel) is the strongest independent risk factor for the development of stent thrombosis. Bleeding complications, most of which arise from the upper gastrointestinal (GI) tract, are the major limiting factors for antiplatelet therapy. The association of aspirin with the increased risk of upper GI bleeding has been well established. Peptic ulcer bleeding and Helicobacter pylori infection are the 2 most important risk factors for aspirin-associated GI bleeding complications. Endoscopy (for both surveillance and potential intervention), performed either emergently or semi-electively, is the primary tool for definitive management of GI bleeding. Considering the increase in GI bleeding risk seen with prolonged antiplatelet therapy, adjunctive proton pump inhibitor therapy and/or eradication of H. pylori infection might be beneficial for DES patients on long-term antiplatelet therapy.

CSF-1 is the major regulator of tissue macrophage development and function. A GM-CSF-dependent, CSF-1 receptor (CSF-1R)-deficient F4/80hiMac-1+Gr1–CD11c+ bone marrow macrophage (BMM) line (MacCsf1r−/−) was developed to study the roles of the eight intracellular CSF-1R tyrosines phosphorylated upon receptor activation. Retroviral expression of the wild-type CSF-1R rescued the CSF-1-induced survival, proliferation, differentiation, and morphological characteristics of primary BMM. Mutation of all eight tyrosines failed to rescue, whereas the individual Y → F mutants (544, 559, 697, 706, 721, 807, 921, 974) rescued these CSF-1-inducible phenotypes to varying degrees. The juxtamembrane domain Y559F and activation loop Y807F mutations severely compromised proliferation and differentiation, whereas Y706, Y721F, and Y974F mutations altered morphological responses, and Y706F increased differentiation. Despite their retention of significant in vitro tyrosine kinase activity, Y559F and Y807F mutants exhibited severely impaired in vivo receptor tyrosine phosphorylation, consistent with the existence of cellular mechanisms inhibiting CSF-1R tyrosine phosphorylation that are relieved by phosphorylation of these two sites. The MacCsf1r−/− macrophage line will facilitate genetic and proteomic approaches to CSF-1R structure/function studies in the major disease-related CSF-1R-expressing cell type.

Atherosclerotic coronary artery disease (CAD) comprises a broad spectrum of clinical entities that include asymptomatic subclinical atherosclerosis and its clinical complications, such as angina pectoris, myocardial infarction (MI) and sudden cardiac death. CAD continues to be the leading cause of death in industrialized society. The long-recognized familial clustering of CAD suggests that genetics plays a central role in its development, with the heritability of CAD and MI estimated at approximately 50% to 60%. Understanding the genetic architecture of CAD and MI has proven to be difficult and costly due to the heterogeneity of clinical CAD and the underlying multi-decade complex pathophysiological processes that involve both genetic and environmental interactions. This review describes the clinical heterogeneity of CAD and MI to clarify the disease spectrum in genetic studies, provides a brief overview of the historical understanding and estimation of the heritability of CAD and MI, recounts major gene discoveries of potential causal mutations in familial CAD and MI, summarizes CAD and MI-associated genetic variants identified using candidate gene approaches and genome-wide association studies (GWAS), and summarizes the current status of the construction and validations of genetic risk scores for lifetime risk prediction and guidance for preventive strategies. Potential protective genetic factors against the development of CAD and MI are also discussed. Finally, GWAS have identified multiple genetic factors associated with an increased risk of in-stent restenosis following stent placement for obstructive CAD. This review will also address genetic factors associated with in-stent restenosis, which may ultimately guide clinical decision-making regarding revascularization strategies for patients with CAD and MI.

CSF-1, the major regulator of macrophage (Mφ) development, has three biologically active isoforms: a membrane-spanning, cell surface glycoprotein, a secreted glycoprotein, and a secreted proteoglycan. We hypothesized that there are shared and unique roles of individual CSF-1 isoforms during renal inflammation. To test this, we evaluated transgenic mice only expressing the cell surface or precursors of the secreted CSF-1 isoforms for Mφ accumulation, activation, and Mφ-mediated tubular epithelial cell (TEC) apoptosis during unilateral ureteral obstruction. The only difference between secreted proteoglycan and secreted glycoprotein CSF-1 isoforms is the presence (proteoglycan) or absence (glycoprotein) of an 18-kDa chondroitin sulfate glycosaminoglycan. We report that 1) cell surface CSF-1 isoform is sufficient to restore Mφ accumulation, activation, and TEC apoptosis to wild-type levels and is substantially more effective than the secreted CSF-1 isoforms; 2) the chondroitin sulfate glycosaminoglycan facilitates Mφ accumulation, activation, and TEC apoptosis; 3) increasing the level of secreted proteoglycan CSF-1 in serum amplifies renal inflammation; and 4) cell-cell contact is required for Mφ to up-regulate CSF-1-dependent expression of IFN-γ. Taken together, we have identified central roles for the cell surface CSF-1 and the chondroitin sulfate chain on secreted proteoglycan CSF-1 during renal inflammation.

Langerhans cells (LCs) are the only dendritic cells of the epidermis and constitute the first immunological barrier against pathogens and environmental insults. The factors regulating LC homeostasis remain elusive and the direct circulating LC precursor has not yet been identified in vivo. Here we report an absence of LCs in mice deficient in the receptor for colony-stimulating factor 1 (CSF-1) in steady-state conditions. Using bone marrow chimeric mice, we have established that CSF-1 receptor-deficient hematopoietic precursors failed to reconstitute the LC pool in inflamed skin. Furthermore, monocytes with high expression of the monocyte marker Gr-1 (also called Ly-6c/G) were specifically recruited to the inflamed skin, proliferated locally and differentiated into LCs. These results identify Gr-1(hi) monocytes as the direct precursors for LCs in vivo and establish the importance of the CSF-1 receptor in this process.

The primary macrophage growth factor, colony-stimulating factor-1 (CSF-1), is homodimeric and exists in 3 biologically active isoforms: a membrane-spanning, cell-surface glycoprotein (csCSF-1) and secreted glycoprotein (sgCSF-1) and proteoglycan (spCSF-1) isoforms. To investigate the in vivo role of the chondroitin sulfate glycosaminoglycan (GAG) chain of spCSF-1, we created mice that exclusively express, in a normal tissue-specific and developmental manner, either the secreted precursor of spCSF-1 or the corresponding precursor in which the GAG addition site was mutated. The reproductive, hematopoietic tooth eruption and tissue macrophage defects of CSF-1-deficient, osteopetrotic Csf1op/Csf1op mice were corrected by transgenic expression of the precursors of either sgCSF-1 or spCSF-1. Furthermore, in contrast to the transgene encoding csCSF-1, both failed to completely correct growth retardation, suggesting a role for csCSF-1 in the regulation of body weight. However, spCSF-1, in contrast to sgCSF-1, completely resolved the osteopetrotic phenotype. Furthermore, in transgenic lines expressing different concentrations of sgCSF-1 or spCSF-1, spCSF-1 more efficiently corrected Csf1op/Csf1op defects of tooth eruption, eyelid opening, macrophage morphology, and B-cell deficiency than sgCSF-1. These results indicate an important role of the CSF-1 chondroitin sulfate proteoglycan in in vivo signaling by secreted CSF-1. (Blood. 2006;107:786-795)

In mammals, the fetal liver is the first site of definitive erythropoiesis-the generation of mature, enucleated red cells. The functional unit for definitive erythropoiesis is the erythroblastic island, a multicellular structure composed of a central macrophage surrounded by erythroblasts at various stages of differentiation. Targeted disruption of the retinoblastoma (Rb) tumour suppressor gene in the mouse leads to embryonic death caused by failure of erythroblasts to enucleate. The erythroid defect has been attributed to loss of Rb in cells that support erythropoiesis, but the identity of these cells is unknown. Here we show that Rb-deficient embryos carry profound abnormalities of fetal liver macrophages that prevent physical interactions with erythroblasts. In contrast, wild-type macrophages bind Rb-deficient erythroblasts and lead them to terminal differentiation and enucleation. Loss of Id2, a helix-loop-helix protein that mediates the lethality of Rb-deficient embryos, rescues the defects of Rb-deficient fetal liver macrophages. Rb promotes differentiation of macrophages by opposing the inhibitory functions of Id2 on the transcription factor PU.1, a master regulator of macrophage differentiation. Thus, Rb has a cell autonomous function in fetal liver macrophages, and restrains Id2 in these cells in order to implement definitive erythropoiesis.

Studies of the influence of the osteoclast on bone development, in particular on mineralization and the formation of the highly organized lamellar architecture of cortical bone by osteoblasts, have not been reported. We therefore examined the micro- and ultrastructure of the developing bones of osteoclast-deficient CSF-1R-nullizygous mice (Csf1r−/− mice). Introduction: Colony-stimulating factor-1 receptor (CSF-1R)-mediated signaling is critical for osteoclastogenesis. Consequently, the primary defect in osteopetrotic Csf1r−/− mice is severe osteoclast deficiency. Csf1r−/− mice therefore represent an ideal model system in which to investigate regulation by the osteoclast of osteoblast-mediated bone formation during development. Materials and Methods: Bones of developing Csf1r−/− mice and their littermate controls were subjected to X-ray analysis, histological examination by light microscopy and transmission electron microscopy, and a three-point bending assay to test their biomechanical strength. Bone mineralization in embryonic and postnatal bones was visualized by double staining with alcian blue and alizarin red. Bone formation by osteoblasts in these mice was also examined by double-calcein labeling and in femoral anlagen transplantation experiments. Results and Conclusions: Frequent spontaneous fractures and decreased strength parameters (ultimate load, yield load, and stiffness) in a three-point bending assay showed the biomechanical weakness of long bones in Csf1r−/− mice. Histologically, these bones have an expanded epiphyseal chondrocyte region, a poorly formed cortex with disorganized collagen fibrils, and a severely disturbed matrix structure. The mineralization of their bone matrix at secondary sites of ossification is significantly reduced. While individual osteoblasts in Csf1r−/− mice have preserved their typical ultrastructure and matrix depositing activity, the layered organization of osteoblasts on the bone-forming surface and the direction of their matrix deposition toward the bone surface have been lost, resulting in their abnormal entrapment by matrix. Moreover, we also found that (1) osteoblasts do not express CSF-1R, (2) the bone defects in Csf1r−/− embryos develop later than the development of osteoclasts in normal embryos, and (3) the transplanted Csf1r−/− femoral anlagen develop normally in the presence of wildtype osteoclasts. These results suggest that the dramatic bone defects in Csf1r−/− mice are caused by a deficiency of the osteoclast-mediated regulation of osteoblasts and that the osteoclast plays an important role in regulating osteoblastic bone formation during development, in particular, in the formation of lamellar bone.

The primary macrophage growth factor, colony-stimulating factor 1 (CSF-1), is expressed as a secreted glycoprotein or proteoglycan found in the circulation or as a biologically active cell surface glycoprotein (csCSF-1). To investigate the in vivo roles of csCSF-1, we created mice that exclusively express csCSF-1, in a normal tissue-specific and developmental manner, by transgenic expression of csCSF-1 in the CSF-1-deficient osteopetrotic (Csf1op/Csf1op) background. The gross defects of Csf1op/Csf1op mice, including growth retardation, failure of tooth eruption, and abnormal male and female reproductive functions were corrected. Macrophage densities in perinatal liver, bladder, sublinguinal salivary gland, kidney cortex, dermis, and synovial membrane were completely restored, whereas only partial or no restoration was achieved in adult liver, adrenal gland, kidney medulla, spleen, peritoneal cavity, and intestine. Residual osteopetrosis, significantly delayed trabecular bone resorption in the subepiphyseal region of the long bone, and incomplete correction of the hematologic abnormalities in the peripheral blood, bone marrow, and spleens of CSF-1-deficient mice were also found in mice exclusively expressing csCSF-1. These data suggest that although csCSF-1 alone is able to normalize several aspects of development in Csf1op/Csf1op mice, it cannot fully restore in vivo CSF-1 function, which requires the presence of the secreted glycoprotein and/or proteoglycan forms. (Blood. 2004;103:1114-1123)

Colony-stimulating factor 1 (CSF-1) regulates the survival, proliferation, and differentiation of mononuclear phagocytes. It is expressed as a secreted glycoprotein or proteoglycan found in the circulation or as a biologically active cell-surface glycoprotein. To investigate tissue CSF-1 regulation, CSF-1–nullCsf1op/Csf1opmice expressing transgenes encoding the full-length membrane-spanning CSF-1 precursor driven by 3.13 kilobases of the mouse CSF-1 promoter and first intron were characterized. Transgene expression corrected the gross osteopetrotic, neurologic, weight, tooth, and reproductive defects ofCsf1op/Csf1opmice. Detailed analysis of one transgenic line revealed that circulating CSF-1, tissue macrophage numbers, hematopoietic tissue cellularity, and hematopoietic parameters were normalized. Tissue CSF-1 levels were normal except for elevations in 4 secretory tissues. Skin fibroblasts from the transgenic mice secreted normal amounts of CSF-1 but also expressed some cell-surface CSF-1. Also, lacZ driven by the same promoter/first intron revealed β-galactosidase expression in hematopoietic, reproductive, and other tissue locations proximal to CSF-1 cellular targets, consistent with local regulation by CSF-1 at these sites. These studies indicate that the 3.13-kilobase promoter/first intron confers essentially normal CSF-1 expression. They also pinpoint new cellular sites of CSF-1 expression, including ovarian granulosa cells, mammary ductal epithelium, testicular Leydig cells, serous acinar cells of salivary gland, Paneth cells of the small intestine, as well as local sites in several other tissues.

The colony stimulating factor-1 (CSF-1) receptor (CSF-1R) directly regulates the development of Paneth cells (PC) and influences proliferation and cell fate in the small intestine (SI). In the present study, we have examined the role of CSF-1 and the CSF-1R in the large intestine, which lacks PC, in the steady state and in response to acute inflammation induced by dextran sulfate sodium (DSS). As previously shown in mouse, immunohistochemical (IHC) analysis of CSF-1R expression showed that the receptor is baso-laterally expressed on epithelial cells of human colonic crypts, indicating that this expression pattern is shared between species. Colons from Csf1r null and Csf1(op/op) mice were isolated and sectioned for IHC identification of enterocytes, enteroendocrine cells, goblet cells and proliferating cells. Both Csf1r(-/-) and Csf1(op/op) mice were found to have colon defects in enterocytes and enteroendocrine cell fate, with excessive goblet cell staining and reduced cell proliferation. In addition, the gene expression profiles of the cell cycle genes, cyclinD1, c-myc, c-fos, and c-myb were suppressed in Csf1r(-/-) colonic crypt, compared with those of WT mice and the expression of the stem cell marker gene Lgr5 was markedly reduced. However, analysis of the proliferative responses of immortalized mouse colon epithelial cells (lines; Immorto-5 and YAMC) indicated that CSF-1R is not a major regulator of colonocyte proliferation and that its effects on proliferation are indirect. In an examination of the acute inflammatory response, Csf1r(+/-) male mice were protected from the adverse affects of DSS-induced colitis compared with WT mice, while Csf1r(+/-) female mice were significantly less protected. These data indicate that CSF-1R signaling plays an important role in colon homeostasis and stem cell gene expression but that the receptor exacerbates the response to inflammatory challenge in male mice.

The CSF-1R is selectively expressed on mononuclear phagocytes, osteoclasts and cells of the female reproductive tract. CSF-1 is normally expressed in a wide variety of cell types, excluding those expressing the CSF-1 receptor (CSF-1R). Autocrine/paracrine regulation by CSF-1 plays an important role in the development of disease in mouse models of acute myeloid leukemia and mammary tumor progression to metastasis. To determine the consequences of autocrine regulation by CSF-1 in the absence of other interventions, we generated transgenic mice that expressed CSF-1 in CSF-1R bearing cells. In contrast to mice that expressed the CSF-1R in CSF-1-expressing cells, which had no gross phenotype at 1 year of age, mice expressing CSF-1 under the control of mouse CSF-1R promoter and second intron (TgRC mice), had a short lifetime, starting to lose weight and mobility after 3 weeks of age. CSF-1 mRNA was expressed in peritoneal macrophages (PM) and sequence analysis of the RT-PCR products verified the expression of a chimeric mRNA of the CSF-1R exon 2, 3 and CSF-1 mRNAs under control of the CSF-1R promoter. Quantitative real-time RT-PCR analysis revealed that CSF-1 mRNA expression was increased by 19-fold in PM and 980-fold in bone marrow derived macrophages (BMM) from a TgRC founder mouse, compared to these populations from non-transgenic mice. Consistent with autocrine regulation by CSF-1, in contrast to BMM from non-transgenic mice, TgRC BMM proliferated in the absence of CSF-1. However, there was no elevation of the circulating CSF-1 in TgRC mice, indicating that transgene-regulated expression of CSF-1 only affected local CSF-1 concentrations. X-ray and histological analysis demonstrated that TgRC mice developed osteoporosis with increased numbers of femoral TRAP-positive osteoclasts. Macrophage densities were significantly increased in bone marrow, liver, spleen and brain. In several tissues, there were local regions of increased macrophage density, consistent with effects of locally produced CSF-1 on macrophage survival and/or proliferation. Also, macrophages in several tissues had a more dendritic appearance than in non-transgenic mice, suggesting that they may be activated. TgRC PM, cultured in the presence of CSF-1 exhibited a highly dendritic morphology with multiple protruding pseudopods also suggesting an activated state. Studies of the macrophage cytokine expression profile by the real-time quantitative RT-PCR revealed elevated levels of mRNA for the proinflammatory cytokines, IL-1β (33-fold) (in BMM) and IL-10 (10-fold) and IL-6 (2.6-fold) (in PM), consistent with the priming effect of CSF-1 on proinflammatory cytokine release by macrophages. These studies indicate that inappropriate local expression of CSF-1 by or near CSF-1R-expressing cells can result in activation of macrophages and osteoclasts and severe disease. Supported by NIH grant CA32551 (ERS) and an ASH Fellow Scholar Award and Leukemia and Lymphoma Society Special Fellow Award (X-MD).