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Objectives: Given the importance of inflammation in atherosclerosis, we sought to determine if atherosclerotic plaque inflammation could be measured noninvasively in humans using positron emission tomography (PET). Background: Earlier PET studies using fluorodeoxyglucose (FDG) demonstrated increased FDG uptake in atherosclerotic plaques. Here we tested the ability of FDG-PET to measure carotid plaque inflammation in patients who subsequently underwent carotid endarterectomy (CEA). Methods: Seventeen patients with severe carotid stenoses underwent FDG-PET imaging 3 h after FDG administration (13 to 25 mCi), after which carotid plaque FDG uptake was determined as the ratio of plaque to blood activity (target to background ratio, TBR). Less than 1 month after imaging, subjects underwent CEA, after which carotid specimens were processed to identify macrophages (staining with anti-CD68 antibodies). Results: There was a significant correlation between the PET signal from the carotid plaques and the macrophage staining from the corresponding histologic sections (r = 0.70; p < 0.0001). When mean FDG uptake (mean TBR) was compared with mean inflammation (mean percentage CD68 staining) for each of the 17 patients, the correlation was even stronger (r = 0.85; p < 0.0001). Fluorodeoxyglucose uptake did not correlate with plaque area, plaque thickness, or area of smooth muscle cell staining. Conclusions: We established that FDG-PET imaging can be used to assess the severity of inflammation in carotid plaques in patients. If subsequent natural history studies link increased FDG-PET activity in carotid arteries with clinical events, this noninvasive measure could be used to identify a subset of patients with carotid atherosclerosis in need of intensified medical therapy or carotid artery intervention to prevent stroke. [ABSTRACT FROM AUTHOR]

: BackgroundHearts from non–heart-beating organ donors are not transplanted because of risk of ischemia-reperfusion injury. We tested whether pharmacologic pre-conditioning with adenosine and the Na+/H+ exchanger inhibitor, cariporide, combined with controlled reperfusion, would prevent injury in porcine hearts that had sustained 30 minutes of hypoxia/ischemia in closed-chest animals.: MethodsHearts from Yorkshire pigs (100 kg) were studied in 3 groups. Group 1 (control) hearts were surgically removed while beating. Group 2 hearts were harvested from animals made hypoxic by discontinuing mechanical ventilation for 30 minutes. Group 3 hearts were hypoxic as in Group 2, but these animals received adenosine (40 mg) and cariporide (400 mg) 10 minutes before stopping ventilation. Cardiac function in all groups was assessed ex vivo in a working heart apparatus in which pressure and flow measurements were made over 3 hours. Controlled reperfusion in Group 3 hearts used leukocyte-depleted blood perfusate containing free radical scavengers. Myocardial injury was assessed on the basis of perfusate creatine phosphokinase activity and histopathologically determined injury score.: ResultsGroups 1 and 3 hearts could be resuscitated to perform work equivalently during the entire reperfusion period and showed positive responses to increases in pre-load and norepinephrine. Group 2 hearts could not perform work. After 3 hours, Group 2 hearts showed significantly higher creatine phosphokinase and histopathologic injury scores compared to with Groups 1 and 3, which were not significantly different from each other.: ConclusionsPharmacologic pre-conditioning and controlled reperfusion effectively protect non-beating porcine hearts from injury after 30 minutes of hypoxia/ischemia in situ. [Copyright &y& Elsevier]

The COVID-19 pandemic initiated public interest in wastewater-based epidemiology (WBE). Public and private entities responded to the need to produce timely and accurate data. LuminUltra and Hach partnered to provide a rapid, field-based quantitative polymerase chain reaction (qPCR) test for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater. This study evaluates the Hach GeneCount SARS-CoV-2 Wastewater RT-qPCR Assay Kit and LuminUltra GeneCount ® Q-16 RT-PCR instrument. The Hach LuminUltra methods were compared to the Promega Wizard ® Enviro Total Nucleic Acid kit and Bio-Rad CFX Opus 96 Real-time PCR Detection System. Over a 12-week period, wastewater samples were collected weekly from seven locations in the Roanoke/Salem, VA sewersheds. Concentration and extraction of the viral RNA were followed by qPCR analysis. The target gene for detection was the nucleocapsid gene (N1) of the SARS-CoV-2 virus. Costs, ease of use, time to produce results, sample preparation, and data comparisons were considered. The comparison determined that the Hach LuminUltra method and instrument were more affordable, consumed less time, and required less technical expertise. While the new method was specific, it had low sensitivity. This evaluation suggests the Hach LuminUltra method should be reserved for limited situations requiring onsite field analysis where data accuracy is not essential., Competing Interests: The authors declare there is no conflict., (© 2024 The Authors This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (CC BY 4.0), which permits copying, adaptation and redistribution, provided the original work is properly cited (http://creativecommons.org/licenses/by/4.0/).)

Aim: In cardiomyocytes, transverse tubules (T-tubules) associate with the sarcoplasmic reticulum (SR), forming junctional membrane complexes (JMCs) where L-type calcium channels (LTCCs) are juxtaposed to Ryanodine receptors (RyR). Junctophilin-2 (JPH2) supports the assembly of JMCs by tethering T-tubules to the SR membrane. T-tubule remodelling in cardiac diseases is associated with downregulation of JPH2 expression suggesting that JPH2 plays a crucial role in T-tubule stability. Furthermore, increasing evidence indicate that JPH2 might additionally act as a modulator of calcium signalling by directly regulating RyR and LTCCs. This study aimed at determining whether JPH2 overexpression restores normal T-tubule structure and LTCC function in cultured cardiomyocytes., Methods and Results: Rat ventricular myocytes kept in culture for 4 days showed extensive T-tubule remodelling with impaired JPH2 localization and relocation of the scaffolding protein Caveolin3 (Cav3) from the T-tubules to the outer membrane. Overexpression of JPH2 restored T-tubule structure and Cav3 relocation. Depletion of membrane cholesterol by chronic treatment with methyl-β-cyclodextrin (MβCD) countered the stabilizing effect of JPH2 overexpression on T-tubules and Cav3. Super-resolution scanning patch-clamp showed that JPH2 overexpression greatly increased the number of functional LTCCs at the plasma membrane. Treatment with MβCD reduced LTCC open probability and activity. Proximity ligation assays showed that MβCD did not affect JPH2 interaction with RyR and the pore-forming LTCC subunit Cav1.2, but strongly impaired JPH2 association with Cav3 and the accessory LTCC subunit Cavβ2., Conclusions: JPH2 promotes T-tubule structural stability and recruits functional LTCCs to the membrane, most likely by directly binding to the channel. Cholesterol is involved in the binding of JPH2 to T-tubules as well as in the modulation of LTCC activity. We propose a model where cholesterol and Cav3 support the assembly of lipid rafts which provide an anchor for JPH2 to form JMCs and a platform for signalling complexes to regulate LTCC activity., (© The Author(s) 2020. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Background: Aberrant Ca 2+ handling is a prominent feature of heart failure. Elucidation of the molecular mechanisms responsible for aberrant Ca 2+ handling is essential for the development of strategies to blunt pathological changes in calcium dynamics. The peptidyl-prolyl cis - trans isomerase peptidyl-prolyl isomerase 1 (Pin1) is a critical mediator of myocardial hypertrophy development and cardiac progenitor cell cycle. However, the influence of Pin1 on calcium cycling regulation has not been explored. On the basis of these findings, the aim of this study is to define Pin1 as a novel modulator of Ca 2+ handling, with implications for improving myocardial contractility and potential for ameliorating development of heart failure., Methods and Results: Pin1 gene deletion or pharmacological inhibition delays cytosolic Ca 2+ decay in isolated cardiomyocytes. Paradoxically, reduced Pin1 activity correlates with increased sarco(endo)plasmic reticulum calcium ATPase (SERCA2a) and Na 2+ /Ca 2+ exchanger 1 protein levels. However, SERCA2a ATPase activity and calcium reuptake were reduced in sarcoplasmic reticulum membranes isolated from Pin1-deficient hearts, suggesting that Pin1 influences SERCA2a function. SERCA2a and Na 2+ /Ca 2+ exchanger 1 associated with Pin1, as revealed by proximity ligation assay in myocardial tissue sections, indicating that regulation of Ca 2+ handling within cardiomyocytes is likely influenced through Pin1 interaction with SERCA2a and Na 2+ /Ca 2+ exchanger 1 proteins., Conclusions: Pin1 serves as a modulator of SERCA2a and Na 2+ /Ca 2+ exchanger 1 Ca 2+ handling proteins, with loss of function resulting in impaired cardiomyocyte relaxation, setting the stage for subsequent investigations to assess Pin1 dysregulation and modulation in the progression of heart failure., (© 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.)

Rationale: Pathological increases in cardiac afterload result in myocyte hypertrophy with changes in myocyte electrical and mechanical phenotype. Remodeling of contractile and signaling Ca 2+ occurs in pathological hypertrophy and is central to myocyte remodeling. STIM1 (stromal interaction molecule 1) regulates Ca 2+ signaling in many cell types by sensing low endoplasmic reticular Ca 2+ levels and then coupling to plasma membrane Orai channels to induce a Ca 2+ influx pathway. Previous reports suggest that STIM1 may play a role in cardiac hypertrophy, but its role in electrical and mechanical phenotypic alterations is not well understood., Objective: To define the contributions of STIM1-mediated Ca 2+ influx on electrical and mechanical properties of normal and diseased myocytes, and to determine whether Orai channels are obligatory partners for STIM1 in these processes using a clinically relevant large animal model of hypertrophy., Methods and Results: Cardiac hypertrophy was induced by slow progressive pressure overload in adult cats. Hypertrophied myocytes had increased STIM1 expression and activity, which correlated with altered Ca 2 + -handling and action potential (AP) prolongation. Exposure of hypertrophied myocytes to the Orai channel blocker BTP2 caused a reduction of AP duration and reduced diastolic Ca 2+ spark rate. BTP2 had no effect on normal myocytes. Forced expression of STIM1 in cultured adult feline ventricular myocytes increased diastolic spark rate and prolonged AP duration. STIM1 expression produced an increase in the amount of Ca 2+ stored within the sarcoplasmic reticulum and activated Ca 2+ /calmodulin-dependent protein kinase II. STIM1 expression also increased spark rates and induced spontaneous APs. STIM1 effects were eliminated by either BTP2 or by coexpression of a dominant negative Orai construct., Conclusions: STIM1 can associate with Orai in cardiac myocytes to produce a Ca 2+ influx pathway that can prolong the AP duration and load the sarcoplasmic reticulum and likely contributes to the altered electromechanical properties of the hypertrophied heart., (© 2017 American Heart Association, Inc.)

Importance: Repair of nasal septal perforations is challenging regardless of surgical technique due to their location and the health of surrounding tissue. There is currently no surgical procedure which is completely effective in the treatment of anteriorly located perforations., Objective: To report a novel method of closing anterior septal perforations using an inferiorly based mucosal rotation flap and an acellular dermal interposition graft, as well as expand upon a previous series., Design: The study includes patients who underwent surgical repair for septal perforations by the senior author between 2003 and 2015., Setting: The study took place at MetroHealth Medical Center in Cleveland, Ohio., Participants: Thirty-nine patients (15 male) with septal perforations of various size and etiology underwent endonasal repair using rotation flaps. The average age of patients was 42-years old (range 10-67years). INTERVENTION FOR CLINICAL TRIALS OR EXPOSURE FOR OBSERVATIONAL STUDIES: Five patients had perforations such that we used inferiorly based flaps, while 35 cases utilized posteriorly based flaps. Acellular dermis was used in addition to a unilateral rotation flap., Main Outcomes and Measures: The primary outcome desired was a complete closure of the septal perforation. The success, or lack thereof, was monitored after healing from surgery., Results: Thirty-seven of the forty surgical procedures demonstrated complete closure of the perforation, a 92.5% success rate. Perforations were separated based upon size. Small perforations (<1cm) had a 93.3% success rate, medium (1-2cm) 88.9%, and all seven large perforations (>2cm) were closed successfully. In addition, all five of the inferiorly based procedures resulted in complete closure of the perforation. Of the failed repairs, one required revision surgery to repair a recurring perforation, while the other two were asymptomatic following the procedure., Conclusions and Relevance: Endonasal repair using inferiorly based mucosal rotation flaps coupled with an acellular dermal interposition graft is a valid technique for the repair of septal perforations. Posterior rotation flaps are preferred due to major septal blood supply from branches of the sphenopalatine artery, but inferiorly based flaps are also viable options for repair for perforations located in the anterior septum., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Introduction: Corporal punishment (CP) is defined as the use of physical force with the intention of causing a child to experience pain but not injury for the purpose of correction or control of the child's behavior. CP has been linked to a variety of negative consequences for children, including physical abuse, eternalizing behavioral problems, and slowed cognitive development. Many American children continue to experience CP at the hands of their parents and other caregivers. The purpose of this study was to evaluate learner attitude toward CP before and after implementation of a pediatric nurse practitioner-designed educational intervention and influences upon learner attitude and beliefs about CP., Method: This study used a pre- and postsurvey design to assess learner attitude about CP before and after participation in an educational intervention. Influences upon learner attitudes and beliefs regarding CP were also described. Learners (N = 882) were health care providers., Results: Nearly all learners (n = 747; 84.7%) stated that the way their parents disciplined them influenced their attitudes toward CP. Fewer than one fifth of learners who were also parents (n = 126; 14.4%) reported that their child's health care provider had ever discussed child discipline with them. Prior to the educational intervention, more than one third of learners (n = 351; 39.88%) endorsed spanking as sometimes necessary, yet significantly fewer learners (n = 251; 28.9%; p < .001) made this statement after the educational intervention. Child discipline management was included in the health care provider education for fewer than half of learners (n = 365; 41.4%)., Discussion: The potential for experiencing CP as a child to result in negative consequences for children has been well documented, yet many American parents continue to use CP as a form of child discipline, and some pediatric health care professionals continue to endorse its use. Pediatric health care providers, including nurses and pediatric nurse practitioners, need to be educated about child discipline and CP. All pediatric health care providers need to advocate for the use of positive parenting principles and discourage the use of CP., (Copyright © 2015 National Association of Pediatric Nurse Practitioners. Published by Elsevier Inc. All rights reserved.)

Hyperhomocysteinemia (HHcy) impairs re-endothelialization and accelerates vascular remodeling. The role of CD34(+)/VEGF receptor (VEGFR) 2(+) progenitor cells (PCs) in vascular repair in HHcy is unknown. We studied the effect of HHcy on PCs and its role in vascular repair in severe HHcy (∼150 μM), which was induced in cystathionine-β synthase heterozygous mice fed a high-methionine diet for 8 weeks. Vascular injury was introduced by carotid air-dry endothelium denudation. CD34(+)/VEGFR2(+) cells were examined by flow cytometry. HHcy reduced bone marrow (BM) CD34(+)/VEGFR2(+) cells and suppressed replenishment of postinjury CD34(+)/VEGFR2(+) cells in peripheral blood (PB). Donor green fluorescent protein-positive PC homing to the injured vessel was reduced in HHcy after CD34(+) PCs from enhanced green fluorescent protein mice were adoptively transferred following carotid injury. CD34(+) PC transfusion partially reversed HHcy-suppressed re-endothelialization and HHcy-induced neointimal formation. Furthermore, homocysteine (Hcy) inhibited proliferation, adhesion, and migration and suppressed β1-integrin expression and activity in human CD34(+) endothelial colony-forming cells (ECFCs) isolated from PBs in a dose-dependent manner. A functional-activating β1-integrin antibody rescued Hcy-suppressed adhesion and migration in CD34(+) ECFCs. In conclusion, HHcy reduces BM CD34(+)/VEGFR2(+) generation and suppresses CD34(+)/VEGFR2(+) cell mobilization and homing to the injured vessel via β1-integrin inhibition, which partially contributes to impaired re-endothelialization and vascular remodeling., (© FASEB.)

Background: Health Information Management (HIM) and Health Informatics (HI) were very separate professions when they were first formed. However, with the increasing adoption of electronic health records, the interests of the two fields have become more aligned., Objectives: To describe the evolution of a joint master's program in health informatics(HI) and health information management (HIM)., Methods: After analyzing workforce needs, and reviewing both CAHIIM accreditation requirements and existing curricular offerings in separate programs in HIM and HI, a joint program was developed., Results: An HI master's program with a core curriculum for all students and tracks in Data Analytics, User Experience and Advanced Practice HIM was developed. A model for a comprehensive examination, based on the CAHIIM competencies, to be administered prior to and after the core curriculum was also developed., Conclusions: A core and track curriculum that incorporates HIM education as part of the Master of Science of Health Informatics provides a feasible roadmap for the future as HIM and HI become more closely aligned.

Background: To improve and learn from patient outcomes, particularly under new care models such as Accountable Care Organizations and Patient-Centered Medical Homes, requires establishing systems for follow-up and feedback., Objective: To provide post-visit feedback to physicians on patient outcomes following acute care visits., Design: A three-phase cross-sectional study [live follow-up call three weeks after acute care visits (baseline), one week post-visit live call, and one week post-visit interactive voice response system (IVRS) call] with three patient cohorts was conducted. A family medicine clinic and an HIV clinic participated in all three phases, and a cerebral palsy clinic participated in the first two phases. Patients answered questions about symptom improvement, medication problems, and interactions with the healthcare system., Patients: A total of 616 patients were included: 142 from Phase 1, 352 from Phase 2 and 122 from Phase 3., Main Measures: Primary outcomes included: problem resolution, provider satisfaction with the system, and comparison of IVRS with live calls made by research staff., Key Results: During both live follow-up phases, at least 96% of patients who were reached completed the call compared to only 48% for the IVRS phase. At baseline, 98 of 113 (88%) patients reported improvement, as well as 167 of 196 (85%) in the live one-week follow-up. In the one-week IVRS phase, 25 of 39 (64%) reported improvement. In all phases, the majority of patients in both the improved and unimproved groups had not contacted their provider or another provider. While 63% of providers stated they wanted to receive patient feedback, they varied in the extent to which they used the feedback reports., Conclusions: Many patients who do not improve as expected do not take action to further address unresolved problems. Systematic follow-up/feedback mechanisms can potentially identify and connect such patients to needed care.

Mucoceles are benign, epithelial-lined mucous cysts. Commonly mucoceles form secondary to obstruction of a sinus outflow tract or from mucosal gland entrapment from chronic infection, inflammation, iatrogenic trauma, external trauma, or neoplasm. We present a rare case of a nasal mucocele in a 37-year old male arising from a remote history of maxillofacial trauma. To our knowledge, mucoceles associated with nasal bone fractures have not been reported in the literature., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Given relatively less favorable health outcomes in rural Alabama, electronic health records (EHRs) have an even greater potential to improve quality and alleviate disparities if meaningfully used., Objectives: We examined rural-urban differences as it pertained to perceived barriers, benefits, and motivating factors of EHR implementation., Methods: We used multivariate logistic models to analyze data collected from a state-wide, self-completed survey of health information management directors in Alabama hospitals., Results: Findings from our analyses indicate that fewer rural hospitals (8%) have implemented EHRs as compared with urban hospitals (18%). Rural hospitals were 71% less likely to consider reduction in costs as a benefit of EHRs (OR = 0.29), and were 75% less likely to consider lack of structured technology as a challenge factor of EHR implementation (OR = 0.25)., Conclusion: Promotion of EHRs in rural areas is challenging but necessary. Understanding perceived barriers and motivating factors of EHR implementation among rural hospitals can inform policy decisions, especially in light of recent meaningful use initiatives.

Background: Cellular treatments for repairing diseased tissues represent a promising clinical strategy. Umbilical cord tissue-derived cells (UTC) are a unique source of cells with a low immunogenic profile and potential for tissue repair. By using UTC from miniature swine, we previously demonstrated that despite their low immunogenic phenotype, UTC could induce an immune response under certain inflammatory conditions and after multiple subcutaneous (SC) injections. Given that repeat dosing of cells may be necessary to achieve a lasting therapeutic benefit, in this study, we examined approaches to avoid an immune response to multiple SC injections of UTC., Methods: By using in vitro and in vivo measures of sensitization to SC cellular injections, we assessed the effects of varying the location of administration site, prolongation of timing between injections, and use of immunosuppressive treatments on repeated cellular injections in Massachusetts General Hospital major histocompatibility complex-defined miniature swine., Results: Although under normal conditions, a single SC injection of major histocompatibility complex-mismatched UTC did not induce a detectable immune response, multiple SC injections of UTC demonstrated rapid humoral and cell-mediated immune responses. Avoidance of an immune response to repeat SC injection was achieved by concurrent immunosuppression with each dose of UTC., Conclusions: UTC and other similar cell types believed to be nonimmunogenic have the potential to induce immune responses under certain conditions. These studies provide important considerations and guidelines for preclinical studies investigating allogeneic cellular therapies.

This manuscript reports on five cases of spontaneous myelogenous leukemia, similar to human disease, occurring within highly inbred, histocompatible sublines of Massachusetts General Hospital (MGH) MHC-defined miniature swine. In cases where a neoplasm was suspected based on clinical observations, samples were obtained for complete blood count, peripheral blood smear, and flow cytometric analysis. Animals confirmed to have neoplasms were euthanized and underwent necropsy. Histological samples were obtained from abnormal tissues and suspect lesions. The phenotype of the malignancies was assessed by flow cytometric analysis of processed peripheral blood mononuclear cells and affected tissues. Five cases of spontaneous myeloid leukemia were identified in adult animals older than 30 months of age. All animals presented with symptoms of weight loss, lethargy, and marked leukocytosis. At autopsy, all animals had systemic disease involvement and presented with severe hepatosplenomegaly. Three of the five myelogenous leukemias have successfully been expanded in vitro. The clustered incidence of disease in this closed herd suggests that genetic factors may be contributing to disease development. Myelogenous leukemia cell lines established from inbred sublines of MGH MHC-defined miniature swine have the potential to be utilized as a model to evaluate therapies of human leukemia., (Copyright 2009 Elsevier B.V. All rights reserved.)

MHC class II (MHCII) genes have been implicated in the regulation of T lymphocyte responses. However, the mechanism of MHCII-driven regulation remains unknown. Matching for MHCII between donors and recipients of allografts favors regulatory T cell tolerance to transplants and provides a unique opportunity to study this regulation. In this study, we investigated MHCII regulation using transfer of donor MHCII genes in recipients of cardiac allografts. Transfer of MHCII IA(b) genes in the bone marrow of CBA mice (H-2(k)) prior to the grafting of IA(b+) fully allogeneic C57BL/6 (B6, H-2(b)) heart transplants resulted in donor-specific tolerance associated with long-term survival of B6, but not third-party, allografts without sustained immunosuppression. Strikingly, the majority of accepted heart transplants (>170 d) were devoid of allograft vasculopathy. Further studies indicated that intracellular IA(b) initiated the tolerogenic process, which was mediated by regulatory T cells (Tregs) that polarized antigraft responses to Th2 cytokine producers. This mechanism seems to be unique to MHCII genes, because previous MHC class I gene-based therapies failed to produce Tregs. These results demonstrate the key role of MHCII in the induction of Tregs. They also underscore a potential mechanism of specific inactivation of T cells in this model; when activated by IA(b+) grafts, IA(b)-specific Tregs repress the entire alloresponse to C57BL/6 transplants (including MHC I and minor Ags), thus mediating T cell tolerance.

In a rat model of left ventricular pressure overload hypertrophy with biventricular failure, we studied the effects of intracoronary delivery of mesenchymal stem cells (MCS) upon right ventricular hemodynamic performance, profiles of local inflammation and apoptosis, and determinants of extracellular matrix remodeling. Sprague-Dawley rats underwent aortic banding and were followed by echocardiography. After a decrease in left ventricular fractional shortening of 25% from the baseline (relative 50% reduction), animals were randomized to an intracoronary injection of MSC (n=28) or PBS (n=20). Right ventricular hemodynamic assessment and measurement of local inflammatory markers, proapoptotic factors, and determinants of extracellular matrix remodeling were performed on post-transplantation day 7, 14, 21 or 28. MSC injection improved right ventricular systolic function in the MSC group compared to the control group (mean+/-SD, max dP/dt 772+/-272 mm Hg/s vs. 392+/-132 at 28 days, P<0.01). Diastolic function was similarly improved (mean+/-SD, max -dP/dt -558+/-171 mm Hg/s vs. -327+/-131 at 28 days, P<0.05). Right ventricular levels of IL-1, IL-6, TNF-alpha, bax, bak and p38 were significantly decreased in the MSC treated animals. Expression of MMP-3, MMP-6, MMP-9, TIMP-1 and TIMP-3 declined in the MSC group compared with controls after 28 days. In this model of left ventricular pressure overload hypertrophy and biventricular failure, intracoronary delivery of MSC was associated with an improvement in the right ventricular hemodynamic performance, profiles of local inflammation and apoptosis, and determinants of extracellular matrix remodeling.

Background: We studied a novel animal model of pressure overload hypertrophy in transition to heart failure following ascending aortic constriction. We sought to assess chronologic changes in hemodynamic parameters, echocardiographic signs of left ventricular (LV) remodeling, exercise tolerance, and profiles of systemic and local inflammation., Materials and Methods: A cohort of Sprague Dawley rats underwent aortic constriction proximal to the innominate artery and were followed by echocardiography. A group of animals were euthanized 20 wk after aortic constriction, before any detectable decline in fractional shortening (normal fractional shortening (FS) or control group; n = 6). When additional animals reached an absolute 25% decline in fractional shortening, they were randomized to be euthanized on d 0 (25% downward arrow FS group; n = 5), or d 21 (>25% downward arrow FS group; n = 6). Hemodynamic and echocardiographic assessment, swim testing to exhaustion, and measurement of systemic and local inflammatory markers was performed at each time interval., Results: An absolute decline of 25% in FS after aortic constriction was observed between 24 and 28 wk for most animals. The transition from compensated to decompensated hypertrophy was associated with markedly decreased dP/dt(max) and dP/dt(min), increased LV end-systolic diameter and LV end-diastolic diameter, stabilization of LV free wall diameter, decreased exercise performance and up-regulation in expression of interleukin-1, interleukin-6, tumor necrosis factor-alpha, and atrial natriuretic peptide. All animals developed heart failure., Conclusions: This study demonstrates that proximal aortic constriction in young rats represents an excellent experimental model of pressure overload hypertrophy that may be useful for testing the efficacy of novel therapies for the treatment of heart failure.

Changes in ventricular extracellular matrix (ECM) composition of pressure overload hypertrophy determine clinical outcomes. The effects of mesenchymal stem cell (MSC) transplantation upon determinants of ECM composition in pressure overload hypertrophy have not been studied. Sprague-Dawley rats underwent aortic banding and were followed by echocardiography. After an absolute decrease in fractional shortening of 25% from baseline, 1 x 10(6) MSC (n = 28) or PBS (n = 20) was randomly injected intracoronarily. LV protein analysis, including matrix metalloproteinases (MMP-2, MMP-3, MMP-6, MMP-9) and tissue inhibitors of metalloproteinases (TIMP-1, TIMP-2, TIMP-3), was performed after sacrifice on postoperative day 7, 14, 21 or 28. Left ventricular levels of MMP-3, MMP-6, MMP-9, TIMP-1 and TIMP-3 were demonstrated to be decreased in the MSC group compared with controls after 28 days. Expression of MMP-2 and TIMP-2 remained relatively stable in both groups. Successful MSCs delivery was confirmed by histological analysis and visualization of labelled MSCs. In this model of pressure overload hypertrophy, intracoronary delivery of MSCs during heart failure was associated with specific changes in determinants of ECM composition. LV reverse remodeling was associated with decreased ventricular levels of MMP-3, MMP-6, MMP-9, TIMP-1 and TIMP-3, which were upregulated in the control group as heart failure progressed. These effects were most significant at 28 days following injection., ((c) 2008 John Wiley & Sons, Ltd.)

Phlegmasia cerulea dolens is a devastating complication of massive deep venous thrombosis, which is clinically characterized by massive lower extremity tissue edema and subsequent arterial insufficiency. These experiments evaluated the local tissue effects of acute global venous obstruction combined with partial arterial ischemia. Experiments were performed to assess the effects of heparin on the cytokine response to simultaneous venous and partial arterial obstruction. Murine hind limbs were subjected to conditions of unilateral venous occlusion and partial tourniquet limb ischemia, which was confirmed by laser Doppler imaging (LDI). Mice underwent either hind limb venous obstruction with intravenous unfractionated heparin (200IU/kg) or intravenous saline 5min before venous occlusion. Sham-treated mice were subjected to anesthesia alone without venous occlusion. After 3hr, the mice were killed and tissue was harvested for measurement of edema (wet to dry weight ratio, W/D), muscle viability, indices of local thrombosis (thrombin-antithrombin complex [TAT]), and cytokine analysis for growth-related oncogene-1 (GRO-1) and interleukin-6 (IL-6, protein via enzyme-linked immunoassay and mRNA via reverse transcriptase polymerase chain reaction). Bleeding time and volume were documented in saline- and heparin-treated mice to confirm systemic anticoagulation. Administration of intravenous heparin resulted in a marked increase in bleeding time and volume. LDI confirmed venous obstruction and ongoing arterial inflow. Venous obstruction resulted in severe visible edema that correlated with a significantly higher W/D ratio but was not associated with a significant decrease in muscle viability. GRO-1 and IL-6 protein and mRNA levels were significantly elevated in the venous occlusion group compared to sham. Heparin therapy significantly decreased TAT3 levels but did not alter the profile of GRO-1 or IL-6 protein levels seen with venous occlusion. Venous occlusion with partial ischemia induces a unique and potent local cytokine expression. Heparin therapy did not ameliorate the cytokine response. These data indicate that heparin therapy does not modulate the cytokine response to venous obstruction.

Background: We have previously reported the successful induction of renal allograft tolerance in non-human primates using a nonmyeloablative conditioning regimen to produce a mixed-chimeric state in the recipient. In the present study, we applied this same technique to lung allotransplantation in cynomolgus monkeys., Methods: Nine pairs of fully major histocompatibility complex (MHC)-mismatched cynomolgus monkeys were used. The conditioning regimen consisted of total body irradiation, thymic irradiation, and antithymocyte globulin. The recipients underwent lung and bone marrow transplantation, followed by anti-CD154 monoclonal antibody (mAb), and a 1-month course of cyclosporine. The regimen included anti-CD8 mAb in the last 5 recipients and alpha 1-antitripsin in the last 3 recipients. The results were compared with 8 recipients that received kidney allografts using the same regimen., Results: Transient chimerism developed in all lung recipients, as was previously seen in the kidney recipients. Nonetheless, the lung recipients rejected their allografts significant earlier than the kidney recipients (P < .01)., Conclusions: Despite the successful induction of mixed chimerism in recipients of fully MHC-mismatched lung allografts, we have not observed long-term graft survival, as has been seen in an analogous kidney model. Strategies to overcome this problem include organ-specific modifications of the transplant regimen.

We studied the effects of indirect allorecognition on the induction and maintenance phases of tolerance in miniature swine cotransplanted with heart and kidney allografts. MHC class I-mismatched heart and kidney grafts were cotransplanted in recipients receiving CyA for 12 days. Recipients were unimmunized or immunized with a set of donor-derived or control third-party MHC class I peptides either 21 days prior to transplantation or over 100 days after transplantation. T-cell proliferation, delayed type hypersensitivity reaction (DTH) and antibody production were assessed. All animals injected with donor MHC class I peptides developed potent indirect alloresponses specific to the immunizing peptides. While untreated recipients developed stable tolerance, all animals preimmunized with donor allopeptides rejected kidney-heart transplants acutely. In contrast, when peptide immunization was delayed until over 100 days after kidney-heart transplantation, no effects were observed on graft function or in vitro measures of alloimmunity. Donor peptide immunization prevented tolerance when administered to recipients pre transplantation but did not abrogate tolerance when administered to long-term survivors post transplantation. This suggests that the presence of T cells activated via indirect allorecognition represent a barrier to the induction but not the maintenance of tolerance.

The use of alpha1,3-galactosyltransferase gene-knockout (GalT-KO) swine donors in discordant xenotransplantation has extended the survival of cardiac xenografts in baboons following transplantation. Eight baboons received heterotopic cardiac xenografts from GalT-KO swine and were treated with a chronic immunosuppressive regimen. The pathologic features of acute humoral xenograft rejection (AHXR), acute cellular xenograft rejection (ACXR) and chronic rejection were assessed in the grafts. No hyperacute rejection developed and one graft survived up to 6 months after transplantation. However, all GalT-KO heart grafts underwent graft failure with AHXR, ACXR and/or chronic rejection. AHXR was characterized by interstitial hemorrhage and multiple thrombi in vessels of various sizes. ACXR was characterized by TUNEL(+) graft cell injury with the infiltration of T cells (including CD3 and TIA-1(+) cytotoxic T cells), CD4(+) cells, CD8(+) cells, macrophages and a small number of B and NK cells. Chronic xenograft vasculopathy, a manifestation of chronic rejection, was characterized by arterial intimal thickening with TUNEL(+) dead cells, antibody and complement deposition, and/or cytotoxic T-cell infiltration. In conclusion, despite the absence of the Gal epitope, acute and chronic antibody and cell-mediated rejection developed in grafts, maintained by chronic immunosupression, presumably due to de novo responses to non-Gal antigens.

Background: We hypothesized that sarcoplasmic reticulum Ca2+ ATPase pump (SERCA-2a) gene delivery would have beneficial effects upon exercise capacity and markers of inflammation in the setting of heart failure., Materials and Methods: A pressure-overload model of experimental heart failure was used in rats. Following a decrease in fractional shortening of >or=25%, animals underwent intracoronary adenoviral-mediated gene transfection using SERCA-2a. Heart failure animals were randomized to receive the SERCA-2a gene, the beta galactosidase (control) gene, or followed without any further intervention. Exercise and hemodynamic testing were performed, and myocardial and systemic markers of inflammation were assayed after 7 and 21 d., Results: Animals receiving Ad.SERCA-2a showed an increase in exercise tolerance (499.0 +/- 14.9 versus 312.8 +/- 10.5 s, P < 0.0001) relative to Ad.Gal group. Groups treated with Ad.SERCA-2a had significantly decreased serum levels of the inflammatory markers interleukin-1, interleukin-6, and tumor necrosis factor-alpha compared with Ad.Gal-treated animals. Serum levels of atrial natriuretic peptide were decreased in animals receiving Ad.SERCA-2a compared with animals receiving Ad.Gal at day 7 (0.35 +/- 0.03 versus 0.52 +/- 0.11 pg/mL, P = 0.001). Myocardial levels of the proapoptotic protein bax were reduced in Ad.SERCA-2a -treated animals compared with those receiving Ad.Gal at day 7 (protein level/actin: 0.24 +/- 0.05 versus 0.33 +/- 0.04, P = 0.04) and day 21 (protein level/actin: 0.61 +/- 0.04 versus 0.69 +/- 0.01, P = 0.001)., Conclusions: Genetic modulation of heart failure using the SERCA-2a gene was associated with improvement in cardiac function and exercise capacity as well as improvements in heart-failure associated inflammatory markers.

Objectives: In a rat model of pressure overload hypertrophy, we studied the effects of intracoronary delivery of mesenchymal stem cells on hemodynamic performance, exercise capacity, systemic inflammation, and left ventricular reverse remodeling., Methods: Sprague-Dawley rats underwent aortic banding and were followed up by echocardiographic scanning. After a decrease in fractional shortening of 25% from baseline, animals were randomized to intracoronary injection of mesenchymal stem cells (MSC group; n = 28) or phosphate-buffered saline solution (control group; n = 20). Hemodynamic and echocardiographic assessment, swim testing to exhaustion, and measurement of inflammatory markers were performed before the rats were humanely killed on postoperative day 7, 14, 21, or 28., Results: Injection of mesenchymal stem cells improved systolic function in the MSC group compared with the control group (mean +/- standard deviation: maximum dP/dt 3048 +/- 230 mm Hg/s vs 2169 +/- 97 mm Hg/s at 21 days and 3573 +/- 741 mm Hg/s vs 1363 +/- 322 mm Hg/s at 28 days: P < .001). Time to exhaustion was similarly increased in the MSC group compared with controls (487 +/- 35 seconds vs 306 +/- 27 seconds at 28 days; P < .01). Serum levels of interleukins 1 and 6, tumor necrosis factor-alpha, and brain natriuretic peptide-32 were significantly decreased in animals treated with mesenchymal stem cells. Stem cell transplantation improved left ventricular fractional shortening at 21 and 28 days. Left ventricular end-systolic and end-diastolic diameters were also improved at 28 days., Conclusions: In this model of pressure overload hypertrophy, intracoronary delivery of mesenchymal stem cells during heart failure was associated with an improvement in hemodynamic performance, maximal exercise tolerance, systemic inflammation, and left ventricular reverse remodeling. This study suggests a potential role of this treatment strategy for the management of hypertrophic heart failure resulting from pressure overload.

Background: Impaired contractile reserve in chronic MR results from load-independent, myocyte contractile abnormalities., Aims: Investigate the mechanisms of contractile dysfunction in chronic mitral valve regurgitation (MR)., Methods: Mild MR was produced in eight dogs followed by pacing induced left ventricular (LV) dilatation over eight months. In-vivo LV dP/dt was measured at several pacing rates. Contractile function was measured in isolated LV trabeculae and myocytes at several stimulation rates and during changes in extracellular [Ca2+]. Identical studies were performed with six control dogs., Results: Chronic MR resulted in a preserved ejection fraction with decreased dP/dt (p<0.01). LV trabeculae demonstrated significantly lower developed force and a negative force-frequency relation with chronic MR (p<0.05). Myocytes exhibited a negative shortening-frequency relationship in both groups with a greater decline with chronic MR (p<0.001) paralleled by decreases in peak [Ca2+](i) transients. Increases in extracellular [Ca2+] abrogated the defects in force generation in trabeculae from animals with chronic MR., Conclusion: Even with a preserved EF, chronic severe MR results in a significant reduction in intrinsic contractile function and reserve. Functional impairment was load-independent reflecting a predominant defect in calcium cycling rather than impaired peak force generating capacity due to myofibrillar attenuation.

Objectives: African Americans (AA) suffer excess hypertension-related health outcomes and their blood pressures (BPs) are widely reported to be less controlled than European Americans (EA). Intensity of hypertension treatment may play a role. We examined whether AA with treated hypertension received less-intense medication regimens than EA, as reflected in the number of antihypertensive medication classes., Design: Cross-sectional observation of baseline information from the REasons for Geographic And Racial Differences in Stroke cohort. Participants were recruited by telephone in 2003-2005, completed a telephone survey, and had BP measured and medications recorded during an in-home visit. The study's outcome was the number of classes of antihypertensive medications., Setting: U.S. national cohort study with oversampling from high stroke mortality regions. Participants were self-identified AA or EA, > or =45 years old, living in the community, and balanced on AA race and sex by design., Participants: 8960 individuals with treated hypertension., Results: Mean age was 68.0 +/- 8.6 years. AA were poorer and less educated than EA, and had worse BP control (63.5% BP < 140/ 90 mm Hg for AA, 74.0% for EA, P < .01), yet they were on more classes of BP medication (24.1% on > or =3 classes, vs. 16.9%, P < .01). AA were taking an average of 0.138 more antihypertensive medication classes than otherwise similar EA (P < .01). More intense treatment persisted across all age, sex, education, income and BP groups., Conclusions: AA were more intensely treated for hypertension than EA. Further study to identify action strategies to eliminate racial differences in hypertension outcomes is warranted.