The association between serum inflammatory biomarkers and incident hypertension among postmenopausal women in the Buffalo OsteoPerio Study.

Several serum inflammatory biomarkers have been associated with blood pressure and hypertension prevalence in cross-sectional studies. Few of these associations have been evaluated prospectively. We examined associations for 10 serum inflammatory biomarkers with incident hypertension among 471 postmenopausal women (mean age = 65) in the Buffalo OsteoPerio Study. Concentrations of C-reactive protein, interleukin (IL)-2, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF)-α, monocyte chemoattractant protein (MCP)-1, adiponectin, and leptin were measured using multiplexed sandwich immunoassays on fasting serum samples collected at baseline (1997-2001). Incident hypertension (195 cases) was defined as physician-diagnosed hypertension and treatment with medication identified on annual mailed health surveys during follow-up (mean 10 years). Cox regression was used to estimate hazard ratios (HR) and 95% confidence intervals (CI) between log-transformed biomarkers (per 1-SD) and hypertension. When adjusted for age, leptin was significantly associated with hypertension risk (HR = 1.55, 95% CI: 1.04, 2.29), however, the association was attenuated and not significant after adjustment for demographic and lifestyle factors, including BMI. Significant (P < 0.10) interactions were observed for smoking (never, ever) with CRP (HR: never, 1.31; ever, 0.91; P = 0.06) and MCP-1 (HR: never, 0.59; ever, 5.11; P = 0.004); for BMI (<25, ≥25) with MCP-1(HR: <25, 3.45; ≥25, 0.95; P = 0.07); for systolic BP with IL-10 (HR: <120, 0.85; 120-139, 1.11; P = 0.07); and for diastolic BP with MCP-1 (HR: <80, 1.29; 80-89, 0.84; P = 0.03) and with adiponectin (HR: <80, 0.86; 80-89, 1.50; P = 0.03). This study adds needed understanding on prospective associations between several serum inflammatory biomarkers and hypertension risk in older postmenopausal women, among whom hypertension burden is substantial.

CD133 expression in circulating hematopoietic progenitor cells.

BACKGROUND: Circulating hematopoietic progenitors (HPCs) are involved in inflammatory diseases such as atherosclerosis, the immune response to cancer, and disorders of the hematopoietic system. HPC characterization by flow cytometry typically utilizes CD34 in combination with other cell surface markers to identify cell populations that give rise to specific hematopoietic lineages. CD133, also known as prominin-1, is a cell surface protein found in HPCs that has a similar but not interchangeable expression pattern with CD34 for characterization of HPC populations. Our goal was to determine the distribution of CD133 expression within HPC sub-types amongst circulating CD34+ cells. METHODS: The quantity of different HPC populations within the CD34+ CD133+ and CD34+ CD133- fractions of venous blood obtained from healthy human subjects was measured using multicolor flow cytometry. RESULTS: The majority of circulating CD34+ cells is CD133-. CD133+ CD34+ cells express low levels of CD38, contain cell populations bearing cell surface markers of hematopoietic stem cells, multipotent progenitors, and multilymphoid progenitors, and are largely devoid of CD38 expressing lineage specified progenitors. These findings clarify the composition of circulating CD133+ CD34+ cell types in adult human subjects. © 2018 International Clinical Cytometry Society.

Association of Periodontal Disease and Edentulism With Hypertension Risk in Postmenopausal Women.

BACKGROUND: Multiple cross-sectional epidemiologic studies have suggested an association between periodontal disease and tooth loss and hypertension, but the temporality of these associations remains unclear. The objective of our study was to evaluate the association of baseline self-reported periodontal disease and edentulism with incident hypertension. METHODS: Study participants were 36,692 postmenopausal women in the Women's Health Initiative-Observational Study who were followed annually from initial periodontal assessment (1998-2003) through 2015 (mean follow-up 8.3 years) for newly diagnosed treated hypertension. Cox proportional hazards regression with adjustment for potential confounders was used to calculate hazard ratios (HRs) and 95% confidence intervals (CIs). RESULTS: Edentulism was significantly associated with incident hypertension in crude (HR (95% CI) = 1.38 (1.28-1.49)) and adjusted (HR (95% CI) = 1.21 (1.11-1.30)) models. This association was stronger among those <60 years compared to ≥60 years (P interaction 0.04) and among those with <120 mm Hg systolic blood pressure, compared to those with ≥120 mm Hg (P interaction 0.004). No association was found between periodontal disease and hypertension. CONCLUSIONS: These findings suggest that edentulous postmenopausal women may represent a group with higher risk of developing future hypertension. As such improved dental hygiene among those at risk for tooth loss as well as preventive measures among the edentulous such as closer blood pressure monitoring, dietary modification, physical activity, and weight loss may be warranted to reduce disease burden of hypertension. Further studies are needed to clarify these results and further elucidate a potential role of periodontal conditions on hypertension risk.

Association of clinical measures of periodontal disease with blood pressure and hypertension among postmenopausal women.

BACKGROUND: Hypertension and periodontal disease are common conditions among postmenopausal women. Periodontal disease has been found associated with hypertension in previous studies, but data in postmenopausal women is limited. METHODS: We assessed the cross-sectional associations of clinically measured periodontal disease with prevalent hypertension and measured systolic blood pressure (SBP) among 1341 postmenopausal women enrolled in the Buffalo Osteoporosis and Periodontal Disease (OsteoPerio) study, an ancillary study of the Women's Health Initiative-Observational Study. RESULTS: Clinical attachment level (CAL) and number of teeth missing were positively associated with SBP among those not taking antihypertensive medication in crude and multivariable adjusted linear regression models (both P < 0.05). Alveolar crestal height (ACH) and gingival bleeding on probing were associated with higher SBP in crude but not multivariable adjusted models. Neither probing pocket depth (PPD) nor severity categories of periodontitis were associated with SBP. Number of teeth missing was significantly associated with prevalent hypertension in crude and multivariable adjusted models (OR = 1.14, per 5 teeth; P = 0.04). ACH was associated with prevalent hypertension in crude but not adjusted models. CAL, PPD, gingival bleeding, and severity of periodontitis were not significantly associated with prevalent hypertension. CONCLUSIONS: These results suggest that measures of oral health including CAL and number of teeth missing are associated with blood pressure in postmenopausal women. Prospective studies are needed to further investigate these associations and the potential underlying mechanisms for these relationships.

Myocardial and Serum Galectin-3 Expression Dynamics Marks Post-Myocardial Infarction Cardiac Remodelling.

BACKGROUND: Acute myocardial infarction (MI) causes significant changes in cardiac morphology and function. Galectin-3 is a novel and potentially therapeutically important mediator of cardiac remodelling. Myocardial and serum galectin-3 expression dynamics in response to the early cardiovascular outcomes after acute MI are not fully elucidated. METHODS: We first performed a comprehensive longitudinal microarray analyses in mice after acute MI. We then measured the serum levels of galectin-3 in a translational porcine model of coronary microembolism-induced post-ischaemic cardiac remodelling. We validated our pre-clinical studies in humans by measuring serum galectin-3 levels of 52 patients with acute ST-elevation MI (STEMI) and 11 healthy controls. We analysed galectin-3 data in relation to the development of major adverse cardiovascular outcomes (MACO). RESULTS: Of the 9,753 genes profiled at infarcted and remote myocardium at eight different time points, dynamic myocardial overexpression of galectin-3 mRNA was detected. In a pig model of diffuse myocardial damage and cardiac remodelling, galectin-3 localised to the areas of tissue damage and myocardial fibrosis, with proportionate increase of their serum galectin-3 expression levels. In humans, increased serum galectin-3 level was associated with in-hospital MACO. CONCLUSIONS: In this translational study, we demonstrated that galectin-3 is dynamically overexpressed in response to acute MI-induced cardiac remodelling. Elevated galectin-3 levels are associated with the development of in-hospital MACO.

Simultaneous measurement of human hematopoietic stem and progenitor cells in blood using multicolor flow cytometry.

Hematopoietic stem cells are the source of all inflammatory cell types. Discovery of specific cell surface markers unique to human hematopoietic stem (HSC) and progenitor (HSPC) cell populations has facilitated studies of their development from stem cells to mature cells. The specific marker profiles of HSCs and HSPCs can be used to understand their role in human inflammatory diseases. The goal of this study is to simultaneously measure HSCs and HSPCs in normal human venous blood using multicolor flow cytometry. Our secondary aim is to determine how G-CSF mobilization alters the quantity of each HSC and HSPC population. Here we show that cells within the CD34+ fraction of human venous blood contains cells with the same cell surface markers found in human bone marrow samples. Mobilization with G-CSF significantly increases the quantity of total CD34+ cells, blood borne HSCs, multipotent progenitors, common myeloid progenitors, and megakaryocyte erythroid progenitors as a percentage of total MNCs analyzed. The increase in blood borne common lymphoid and granulocyte macrophage progenitors with G-CSF treatment did not reach significance. G-CSF treatment predominantly increased the numbers of HSCs and multipotent progenitors in the total CD34+ cell population; common myeloid progenitors and megakaryocyte erythroid progenitors were enriched relative to total MNCs analyzed, but not relative to total CD34+ cells. Our findings illustrate the utility of multicolor flow cytometry to quantify circulating HSCs and HSPCs in venous blood samples from human subjects. © 2016 International Clinical Cytometry Society.

Effects of statins on TH1 modulating cytokines in human subjects.

Background. Activation of the innate immune system by cholesterol accelerates atherosclerosis. High levels or modified forms of cholesterol stimulate release of the inflammatory cytokines IL-12 and IL-18 that synergistically stimulate T lymphocytes to produce the atherogenic cytokine interferon-γ. While activation of the innate immune system by cholesterol is well-described in animal models and human subjects with high cholesterol levels or known atherosclerotic disease, the interaction of cholesterol and lipoproteins with the innate immune system in human subjects without known atherosclerosis is less well-described. The goal of our study was to assess the TH1 modulating cytokines IL-12 p40 and IL-18, and their counter regulatory cytokines IL-18 binding protein and IL-27, to determine if their levels are linked to cholesterol levels or other factors. Methods. We performed a blinded, randomized hypothesis-generating study in human subjects without known atherosclerotic disease. We measured serum lipids, lipoprotein levels, and collected plasma samples at baseline. Subjects were randomized to two weeks of therapy with atorvastatin, pravastatin, or rosuvastatin. Lipids and cytokine levels were measured after two weeks of statin treatment. Subjects were given a four-week statin-free period. At the end of the four-week statin-free period, venous blood was sampled again to determine if serum lipids returned to within 5% of their pre-statin levels. When lipid levels returned to baseline, subjects were again treated with the next statin in the randomization scheme. IL-12, IL-18, IL-18 binding protein, and IL-27 were measured at baseline and after each statin treatment to determine effects of statin treatment on their blood levels, and identify correlations with lipids and lipoproteins. Results. Therapy with statins revealed no significant change in the levels of IL-12, IL-18, IL-18 binding protein or IL-27 levels. We found that IL-18 levels positively correlate with total cholesterol levels (r (2) = 0.15, p < 0.03), but not HDL or LDL cholesterol. In contrast, IL-12 p40 levels inversely correlated with total cholesterol (r (2) = -0.17, p < 0.008), HDL cholesterol (r (2) = -0.22, p < 0.002), and apolipoprotein A1 (r (2) = -0.21, p < 0.002). Similarly, IL-18 binding protein levels inversely correlated with apolipoprotein A1 levels (r (2) = -0.13, p < 0.02). Conclusions. Our findings suggest that total cholesterol levels positively regulate IL-18, while HDL cholesterol and apolipoprotein A1 may reduce IL-12 p40 and IL-18 binding protein levels. Additional studies in a larger patient population are needed to confirm these findings, and verify mechanistically whether HDL cholesterol can directly suppress IL-12 p40 and IL-18 binding protein levels in human subjects.

Global intracoronary infusion of allogeneic cardiosphere-derived cells improves ventricular function and stimulates endogenous myocyte regeneration throughout the heart in swine with hibernating myocardium.

BACKGROUND: Cardiosphere-derived cells (CDCs) improve ventricular function and reduce fibrotic volume when administered via an infarct-related artery using the "stop-flow" technique. Unfortunately, myocyte loss and dysfunction occur globally in many patients with ischemic and non-ischemic cardiomyopathy, necessitating an approach to distribute CDCs throughout the entire heart. We therefore determined whether global intracoronary infusion of CDCs under continuous flow improves contractile function and stimulates new myocyte formation. METHODS AND RESULTS: Swine with hibernating myocardium from a chronic LAD occlusion were studied 3-months after instrumentation (n = 25). CDCs isolated from myocardial biopsies were infused into each major coronary artery (∼ 33 × 10(6) icCDCs). Global icCDC infusion was safe and while ∼ 3% of injected CDCs were retained, they did not affect ventricular function or myocyte proliferation in normal animals. In contrast, four-weeks after icCDCs were administered to animals with hibernating myocardium, %LADWT increased from 23 ± 6 to 51 ± 5% (p<0.01). In diseased hearts, myocyte proliferation (phospho-histone-H3) increased in hibernating and remote regions with a concomitant increase in myocyte nuclear density. These effects were accompanied by reductions in myocyte diameter consistent with new myocyte formation. Only rare myocytes arose from sex-mismatched donor CDCs. CONCLUSIONS: Global icCDC infusion under continuous flow is feasible and improves contractile function, regresses myocyte cellular hypertrophy and increases myocyte proliferation in diseased but not normal hearts. New myocytes arising via differentiation of injected cells are rare, implicating stimulation of endogenous myocyte regeneration as the primary mechanism of repair.

Fractalkine (CX3CL1), GM-CSF and VEGF-a levels are reduced by statins in adult patients.

BACKGROUND: Fractalkine (CX3CL1) promotes migration and adhesion of lymphocytes and monocytes to inflamed tissues. Prior studies show a role for CX3CL1 in atherosclerosis. The relationship between inflammatory cytokines, cholesterol, and CX3CL1 levels in human subjects without known coronary artery disease is not well characterized. The goal of our study was to assess baseline CX3CL1 levels, and after modulation of cholesterol levels by statins to determine if CX3CL1 is linked to cholesterol levels or inflammatory stimuli. METHODS: We performed a blinded, randomized hypothesis generating study in human subjects without known coronary artery disease treated sequentially with three statins of differing potencies. Fractalkine (CX3CL1), GM-CSF, VEGF-A, other chemokines, and lipid levels were measured. Mechanistic studies of CX3CL1 induction by LDL cholesterol and TNFα in cultured human endothelial cells were performed using real-time PCR. RESULTS: Therapy with statins reduced total and LDL cholesterol levels as expected. CX3CL1 levels were significantly reduced from no statin control levels (89.9 ± 18.5 pg/mL) after treatment with atorvastatin (60.0 ± 7.8 pg/mL), pravastatin (54.2 ± 7.0 pg/mL) and rosuvastatin (65.6 ± 7.3 pg/mL) (χ (2)(2) = 17.4, p ≤ 0.001). Cholesterol is not a known regulator of CX3CL1. We found GM-CSF (r(2) = 0.524; p < 0.005) and VEGF-A (r(2) = 0.4; p < 0.005) levels were highly and positively correlated with CX3CL1. Total (r(2) = 0.086) and LDL cholesterol (r(2) = 0.059) levels weakly correlated with CX3CL1 levels. Finally, we tested whether LDL cholesterol could induce CX3CL1, GM-CSF, and VEGF-A in human endothelial cells, versus TNFα. LDL cholesterol alone resulted in small, non-significant increases in CX3CL1 and GM-CSF, while TNFα resulted in > 10-fold induction. CONCLUSIONS: Our findings suggest that while statins suppress CX3CL1 levels, inflammatory cytokines may be the major regulator of CX3CL1 levels rather than cholesterol itself. Additional studies in a larger patient population are needed to confirm these findings, determine if CX3CL1 levels reflect inflammation levels, and potentially add to standard risk factors in prediction of atherosclerotic disease events.

Cholesterol and hematopoietic stem cells: inflammatory mediators of atherosclerosis.

Atherosclerosis causing heart attack and stroke is the leading cause of death in the modern world. Therapy for end-stage atherosclerotic disease using CD34(+) hematopoietic cells has shown promise in human clinical trials, and the in vivo function of hematopoietic and progenitor cells in atherogenesis is becoming apparent. Inflammation plays a central role in the pathogenesis of atherosclerosis. Cholesterol is a modifiable risk factor in atherosclerosis, but in many patients cholesterol levels are only mildly elevated. Those with high cholesterol levels often have elevated circulating monocyte and neutrophil counts. How cholesterol affects inflammatory cell levels was not well understood. Recent findings have provided new insight into the interaction among hematopoietic stem cells, cholesterol, and atherosclerosis. In mice, high cholesterol levels or inactivation of cholesterol efflux transporters have multiple effects on hematopoietic stem cells (HSPCs), including promoting their mobilization into the bloodstream, increasing proliferation, and differentiating HSPCs to the inflammatory monocytes and neutrophils that participate in atherosclerosis. Increased levels of interleukin-23 (IL-23) stimulate IL-17 production, resulting in granulocyte colony-stimulating factor (G-CSF) secretion, which subsequently leads to HSPC release into the bloodstream. Collectively, these findings clearly link elevated cholesterol levels to increased circulating HSPC levels and differentiation to inflammatory cells that participate in atherosclerosis. Seminal questions remain to be answered to understand how cholesterol affects HSPC-mobilizing cytokines and the role they play in atherosclerosis. Translation of findings in animal models to human subjects may include HSPCs as new targets for therapy to prevent or regress atherosclerosis in patients.

Regional myocardial sympathetic denervation predicts the risk of sudden cardiac arrest in ischemic cardiomyopathy.

OBJECTIVES: The PAREPET (Prediction of ARrhythmic Events with Positron Emission Tomography) study sought to test the hypothesis that quantifying inhomogeneity in myocardial sympathetic innervation could identify patients at highest risk for sudden cardiac arrest (SCA). BACKGROUND: Left ventricular ejection fraction (LVEF) is the only parameter identifying patients at risk of SCA who benefit from an implantable cardiac defibrillator (ICD). METHODS: We prospectively enrolled 204 subjects with ischemic cardiomyopathy (LVEF ≤35%) eligible for primary prevention ICDs. Positron emission tomography (PET) was used to quantify myocardial sympathetic denervation ((11)C-meta-hydroxyephedrine [(11)C-HED]), perfusion ((13)N-ammonia) and viability (insulin-stimulated (18)F-2-deoxyglucose). The primary endpoint was SCA defined as arrhythmic death or ICD discharge for ventricular fibrillation or ventricular tachycardia >240 beats/min. RESULTS: After 4.1 years follow-up, cause-specific SCA was 16.2%. Infarct volume (22 ± 7% vs. 19 ± 9% of left ventricle [LV]) and LVEF (24 ± 8% vs. 28 ± 9%) were not predictors of SCA. In contrast, patients developing SCA had greater amounts of sympathetic denervation (33 ± 10% vs. 26 ± 11% of LV; p = 0.001) reflecting viable, denervated myocardium. The lower tertiles of sympathetic denervation had SCA rates of 1.2%/year and 2.2%/year, whereas the highest tertile had a rate of 6.7%/year. Multivariate predictors of SCA were PET sympathetic denervation, left ventricular end-diastolic volume index, creatinine, and no angiotensin inhibition. With optimized cut-points, the absence of all 4 risk factors identified low risk (44% of cohort; SCA <1%/year); whereas ≥2 factors identified high risk (20% of cohort; SCA ∼12%/year). CONCLUSIONS: In ischemic cardiomyopathy, sympathetic denervation assessed using (11)C-HED PET predicts cause-specific mortality from SCA independently of LVEF and infarct volume. This may provide an improved approach for the identification of patients most likely to benefit from an ICD. (Prediction of ARrhythmic Events With Positron Emission Tomography [PAREPET]; NCT01400334).

Electronic "expression" of the inward rectifier in cardiocytes derived from human-induced pluripotent stem cells.

BACKGROUND: Human-induced pluripotent stem cell (h-iPSC)-derived cardiac myocytes are a unique model in which human myocyte function and dysfunction are studied, especially those from patients with genetic disorders. They are also considered a major advance for drug safety testing. However, these cells have considerable unexplored potential limitations when applied to quantitative action potential (AP) analysis. One major factor is spontaneous activity and resulting variability and potentially anomalous behavior of AP parameters. OBJECTIVE: To demonstrate the effect of using an in silico interface on electronically expressed I(K1), a major component lacking in h-iPSC-derived cardiac myocytes. METHODS: An in silico interface was developed to express synthetic I(K1) in cells under whole-cell voltage clamp. RESULTS: Electronic I(K1) expression established a physiological resting potential, eliminated spontaneous activity, reduced spontaneous early and delayed afterdepolarizations, and decreased AP variability. The initiated APs had the classic rapid upstroke and spike and dome morphology consistent with data obtained with freshly isolated human myocytes as well as the readily recognizable repolarization attributes of ventricular and atrial cells. The application of 1 µM of BayK-8644 resulted in anomalous AP shortening in h-iPSC-derived cardiac myocytes. When I(K1) was electronically expressed, BayK-8644 prolonged the AP, which is consistent with the existing results on native cardiac myocytes. CONCLUSIONS: The electronic expression of I(K1) is a simple and robust method to significantly improve the physiological behavior of the AP and electrical profile of h-iPSC-derived cardiac myocytes. Increased stability enables the use of this preparation for a controlled quantitative analysis of AP parameters, for example, drug responsiveness, genetic disorders, and dynamic behavior restitution profiles.

LDL cholesterol modulates human CD34+ HSPCs through effects on proliferation and the IL-17 G-CSF axis.

BACKGROUND: Hypercholesterolemia plays a critical role in atherosclerosis. CD34+ CD45dim Lineage- hematopoietic stem/progenitor cells (HSPCs) give rise to the inflammatory cells linked to atherosclerosis. In mice, high cholesterol levels mobilize HSPCs into the bloodstream, and promote their differentiation to granulocytes and monocytes. The objective of our study was to determine how cholesterol levels affect HSPC quantity in humans. METHODS: We performed a blinded, randomized hypothesis generating study in human subjects (n=12) treated sequentially with statins of differing potencies to vary lipid levels. CD34+ HSPC levels in blood were measured by flow cytometry. Hematopoietic colony forming assays confirmed the CD34+ population studied as HSPCs with multlineage differentiation potential. Mobilizing cytokine levels were measured by ELISA. RESULTS: The quantity of HSPCs was 0.15 ± 0.1% of buffy coat leukocytes. We found a weak, positive correlation between CD34+ HSPCs and both total and LDL cholesterol levels (r(2)=0.096, p < 0.025). Additionally, we tested whether cholesterol modulates CD34+ HSPCs through direct effects or on the levels of mobilizing cytokines. LDL cholesterol increased cell surface expression of CXCR4, G-CSFR affecting HSPC migration, and CD47 mediating protection from phagocytosis by immune cells. LDL cholesterol also increased proliferation of CD34+ HSPCs (28 ± 5.7%, n=6, p < 0.03). Finally, the HSPC mobilizing cytokine G-CSF (r(2)=0.0683, p < 0.05), and its upstream regulator IL-17 (r(2)=0.0891, p < 0.05) both correlated positively with LDL cholesterol, while SDF-1 levels were not significantly affected. CONCLUSIONS: Our findings support a model where LDL cholesterol levels positively correlate with CD34+ HSPC levels in humans through effects on the levels of G-CSF via IL-17 promoting mobilization of HSPCs, and by direct effects of LDL cholesterol on HSPC proliferation. The findings are provocative of further study to determine if HSPCs, like cholesterol levels, are linked to CVD events.

Sox10-MCS5 enhancer dynamically tracks human oligodendrocyte progenitor fate.

In this study, we sought to establish a novel method to prospectively and dynamically identify live human oligodendrocyte precursor cells (OPCs) and oligodendrocyte lineage cells from brain dissociates and pluripotent stem cell culture. We selected a highly conserved enhancer element of the Sox10 gene, known as MCS5, which directs reporter expression to oligodendrocyte lineage cells in mouse and zebrafish. We demonstrate that lentiviral Sox10-MCS5 induced expression of GFP at high levels in a subpopulation of human CD140a/PDGFαR-sorted OPCs as well as their immature oligodendrocyte progeny. Furthermore, we show that almost all Sox10-MCS5:GFP(high) cells expressed OPC antigen CD140a and human OPCs expressing SOX10, OLIG2, and PDGFRA mRNAs could be prospectively identified using GFP based fluorescence activated cells sorting alone. Additionally, we established a human induced pluripotent cell (iPSC) line transduced with the Sox10-MCS5:GFP reporter using a Rex-Neo cassette. Similar to human primary cells, GFP expression was restricted to embryoid bodies containing both oligodendrocyte progenitor and oligodendrocyte cells and co-localized with NG2 and O4-positive cells respectively. As such, we have developed a novel reporter system that can track oligodendrocyte commitment in human cells, establishing a valuable tool to improve our understanding and efficiency of human oligodendrocyte derivation.

Recipient selection in cardiac transplantation: contraindications and risk factors for mortality.

Currently the only acknowledged, definitive treatment for refractory heart failure is heart transplantation (HTx). During the past 10 years, selection criteria for heart transplant recipients have been developed that use an analysis of risk factors associated with mortality, which were identified by consensus opinion and by single-center and multi-center database review. A number of other studies also have been designed to evaluate specific risk factors for transplant such as advanced age, diabetes, and sex. This review identifies variables that continue to provoke controversy during the candidate selection process or variables that have changed from absolute to relative contraindications for HTx. Clinicians may use the data summarized in this review as a guide to making decisions about patient candidacy for HTx. One could conclude from this analysis that a more formalized and objective scale to select patients and to assess risk of death after HTx is necessary. Moreover, as alternative therapies to HTx become reality, a better instrument for triaging patients to one form of therapy or another may be necessary.

Nerve growth factor-mediated increases in protein methylation occur predominantly at type I arginine methylation sites and involve protein arginine methyltransferase 1.

Nerve growth factor (NGF)-specific signal transduction leads to changes in protein methylation during neuronal differentiation of PC12 cells (Cimato et al. [1997] J. Cell Biol. 138:1089-1103). In the present work, we demonstrate that, among NGF-regulated proteins, arginine methylation is more prevalent than carboxylmethylation. Type I protein arginine methyltransferase (PRMT) activity produces asymmetric dimethylation of the terminal guanidinonitrogen of arginines in substrate proteins, particularly glycine and arginine-rich (GAR) segments of proteins. Several GAR peptides were used to assay for methyltransferase activity and to compete with endogenous cellular proteins for the PRMT activity in PC12 cell extracts. Peptides derived from fibrillarin and nucleolin, as well as a synthetic GAR peptide containing a repetitive GRG motif, are each extremely effective at blocking in vitro methylation of the NGF-regulated PC12 cell methylated proteins. Myelin basic protein, a substrate for type II PRMT, selectively inhibits a 45 kDa protein but is a much less effective inhibitor of total methylation at an equimolar concentration. In addition, the fibrillarin- and nucleolin-derived peptides were used to detect elevated PRMT activity in homogenates of NGF-treated PC12 cells. Finally, immunoprecipitation of PRMT1 from PC12 cells provides the first demonstration of an NGF-activated methyltransferase and implicates PRMT1 in NGF signal transduction.