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Neuroprotective drugs to protect the brain against cerebral ischemia and reperfusion (I/R) injury are urgently needed. Mammalian cell-produced recombinant human erythropoietin (rhuEPO M ) has been demonstrated to have excellent neuroprotective functions in preclinical studies, but its neuroprotective properties could not be consistently translated in clinical trials. The clinical failure of rhuEPO M was thought to be mainly due to its erythropoietic activity-associated side effects. To exploit its tissue-protective property, various EPO derivatives with tissue-protective function only have been developed. Among them, asialo-rhuEPO, lacking terminal sialic acid residues, was shown to be neuroprotective but non-erythropoietic. Asialo-rhuEPO can be prepared by enzymatic removal of sialic acid residues from rhuEPO M (asialo-rhuEPO E ) or by expressing human EPO gene in glycoengineered transgenic plants (asialo-rhuEPO P ). Both types of asialo-rhuEPO, like rhuEPO M , displayed excellent neuroprotective effects by regulating multiple cellular pathways in cerebral I/R animal models. In this review, we describe the structure and properties of EPO and asialo-rhuEPO, summarize the progress on neuroprotective studies of asialo-rhuEPO and rhuEPO M , discuss potential reasons for the clinical failure of rhuEPO M with acute ischemic stroke patients, and advocate future studies needed to develop asialo-rhuEPO as a multimodal neuroprotectant for ischemic stroke treatment.

Objectives: To determine if radial artery (RA) access compared with femoral artery (FA) access for percutaneous coronary intervention (PCI) is associated with a lower incidence of acute kidney injury (AKI)., Background: AKI results in substantial morbidity and cost following PCI. Prior studies comparing the occurrence of AKI associated with radial artery (RA) versus femoral artery (FA) access have mixed results., Methods: Using a large state-wide database, 14,077 patients (8,539 with RA and 5,538 patents with FA access) were retrospectively compared to assess the occurrence of AKI following PCI. To reduce selection bias and balance clinical data across the two groups, a novel machine learning method called a Generalized Boosted Model was conducted on the arterial access site generating a weighted propensity score for each variable. A logistic regression analysis was then performed on the occurrence of AKI following PCI using the weighted propensity scores from the Generalized Boosted Model., Results: As shown in other studies, multiple variables were associated with an increase in AKI after PCI. Only RA access (OR 0.82; 95% CI 0.74-0.91) and male gender (OR 0.80; 95% CI 0.72-0.89) were associated with a lower occurrence of AKI. Based on the calculated Mehran scores, patients were stratified into groups with an increasing risk of AKI. RA access was consistently found to have a lower risk of AKI compared with FA access across these groups of increasing risk., Conclusions: Compared with FA access, RA access is associated with an 18% lower rate of AKI following PCI. This effect was observed among different levels of risk for developing AKI. Although developed from a retrospective analysis, this study supports the use of RA access when technically possible in a diverse group of patients., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2023 Patrick S. Kietrsunthorn et al.)

Mammalian cell-produced recombinant human erythropoietin (rhuEPO M ) has been shown to be a multimodal neuroprotectant targeting an array of key pathological mechanisms in experimental stroke models. However, the rhuEPO M clinical trials were terminated due to increased risk of thrombosis, largely ascribed to its erythropoietic function. We recently took advantage of a plant-based expression system lacking sialylation capacity to produce asialo-rhuEPO P , a rhuEPO derivative without sialic acid residues. In the present study, we proved that asialo-rhuEPO P is non-erythropoietic by repeated intravenous injection (44 μg/kg bw) in mice showing no increase in hemoglobin levels and red blood cell counts, and confirmed that it is non-immunogenic by measuring humoral response after immunizing the mice. We demonstrate that it is neuroprotective in a cerebral ischemia and reperfusion (I/R) mouse model, exhibiting ~ 50% reduction in cerebral infarct volume and edema, and significant improvement in neurological deficits and histopathological outcome. Our studies further revealed that asialo-rhuEPO P , like rhuEPO M , displays pleiotropic neuroprotective effects, including restoring I/R-interrupted mitochondrial fission and fusion proteins, preventing I/R injury-induced increase in mitophagy and autophagy markers, and inhibiting apoptosis to benefit nerve cell survival. Most importantly, asialo-rhuEPO P lacking erythropoietic activity and immunogenicity holds great translational potential as a multimodal neuroprotectant for stroke treatment., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Understanding proteomic changes in the ischemic penumbra are crucial to rescue those salvageable cells and reduce the damage of an ischemic stroke. Since the penumbra region is dynamic with heterogeneous cells/tissues, tissue sampling from animal models of stroke for the molecular study is a challenge. In this study, cultured hippocampal HT22 cells under hypoxia treatment for 17.5 h with 0.69 mM low glucose (H+LG) could mimic ischemic penumbral cells since they had much higher cell viability and viable cell number compared to hypoxia without glucose (H-G) treatment. To validate established cell-based ischemic penumbral model and understand the beneficial effects of low glucose (LG), quantitative proteomics analysis was performed on H+LG, H-G, and normoxia with normal 22 mM glucose (N+G) treated cells. We identified 427 differentially abundant proteins (DAPs) between H-G and N+G and further identified 105 DAPs between H+LG and H-G. Analysis of 105 DAPs revealed that LG promotes cell survival by activating HIF1α to enhance glycolysis; preventing the dysregulations of extracellular matrix remodeling, cell cycle and division, and antioxidant and detoxification; as well as attenuating inflammatory reaction response, protein synthesis and neurotransmission activity. Our results demonstrated that this established cell-based system could mimic penumbral conditions and can be used for molecular studies., (Copyright © 2020 Li, Kittur, Hung, Li, Ge, Sane and Xie.)

A rare case of extramedullary multiple myeloma causing cardiac tamponade secondary to a plasma cell-based pericardial effusion is described. A systematic search using PubMed (National Library of Medicine) was used to identify a further 27 cases dating back to 1970. Case characteristics, treatment strategies, and survival time following tamponade are discussed. Linear regression demonstrated a weak but statistically significant correlation between survival time following tamponade and treatment with systemic chemotherapy and steroids (β=16.8 weeks, P=.009). However, this manifestation of extramedullary multiple myeloma still conveys a dismal prognosis with a median survival following tamponade of only 6 weeks based on our review., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Recombinant human erythropoietin (rhuEPO) and asialoerythropoietin (asialo-rhuEPO) are cardioprotective. However, the protective effects of rhuEPO could not be translated into clinical practice because of its hematopoiesis-associated side effects while non-erythropoietic asialo-rhuEPO is unavailable in large quantities for clinical studies. This study was designed to investigate the cardiomyocyte protective potential of plant-produced asialo-rhuEPO (asialo-rhuEPO P ) against staurosporine (STS)-induced injury in HL-1 murine cardiomyocytes and identify cellular pathway(s) responsible for its cardioprotection., Methods: HL-1 cardiomyocytes were simultaneously treated with STS and asialo-rhuEPO P . Cellular injury, apoptosis, and cell viabilities were measured by LDH assay, Hoechst staining and trypan blue exclusion method, respectively while western blotting was used to study its effects on apoptosis and autophagy hallmarks., Results: Our results showed that 20 IU/ml asialo-rhuEPO P provided 39% protection to cardiomyocytes compared to STS-treated cells, which is 2-fold better than that of mammalian cell-produce rhuEPO (rhuEPO M ). Asialo-rhuEPO P was found to suppress activation of proapoptotic kinase Mst1 (mammalian Sterile-20-like kinase 1) and FOXO3, leading to inhibition of apoptotic pathway and restoration of autophagy as indicated by the reduction of fragmented/condensed nuclei, altered ratios of Bax/Bcl2, p-Bad/Bad, cytosol/mitochondrial cyt c and caspase-3 activation, and the restored levels of autophagy markers Beclin1, p62 and LC3B-II. Additionally, Akt was found to be activated and FOXO3 was phosphorylated on Ser253, suggesting inhibition of FOXO3 transcriptional function., Conclusions: Asialo-rhuEPO P -mediated cardioprotection occurs through activation of PI3K/Akt pathway leading to suppression of Mst1 activation and promoting cardiomyocyte survival., General Significance: Asialo-rhuEPO P could be used to modulate Mst1 activity elevated under numerous pathological states.

Pancreatic beta-cell death adversely contributes to the progression of both type I and II diabetes by undermining beta-cell mass and subsequently diminishing endogenous insulin production. Therapeutics to impede or even reverse the apoptosis and dysfunction of beta-cells are urgently needed. Asialo-rhuEPO, an enzymatically desialylated form of recombinant human erythropoietin (rhuEPO), has been shown to have cardioprotective and neuroprotective functions but with no adverse effects like that of sialylated rhuEPO. Heretofore, the anti-apoptotic effect of asialo-rhuEPO on pancreatic beta-cells has not been reported. In the current study, we investigated the cytoprotective properties of plant-produced asialo-rhuEPO (asialo-rhuEPO P ) against staurosporine-induced cell death in the pancreatic beta-cell line RIN-m5F. Our results showed that 60 IU/ml asialo-rhuEPO P provided 41% cytoprotection while 60 IU/ml rhuEPO yielded no effect. Western blotting results showed that asialo-rhuEPO P treatment inhibited both MST1 and caspase-3 activation with the retention of PDX1 and insulin levels close to untreated control cells. Our study provides the first evidence indicating that asialo-rhuEPO P -mediated protection involves the reduction of MST1 activation, which is considered a key mediator of apoptotic signaling in beta-cells. Considering the many advantages its plant-based expression, asialo-rhuEPO P could be potentially developed as a novel and inexpensive agent to treat or prevent diabetes after further performing studies in cell-based and animal models of diabetes.

Although coronary tortuosity can influence the hemodynamics of coronary arteries, the relationship between tortuosity and flow has not been thoroughly investigated partly due to the absence of a widely accepted definition of tortuosity and the lack of patient-specific studies that analyze complete coronary trees. Using a computational approach we investigated the effects of tortuosity on coronary flow parameters including pressure drop, wall shear stress, and helical flow strength as measured by helicity intensity. Our analysis considered idealized and patient-specific geometries. Overall results indicate that perfusion pressure decreases with increased tortuosity, but the patient-specific results show that more tortuous vessels have higher physiological wall shear stress values. Differences between the idealized and patient-specific results reveal that an accurate representation of coronary tortuosity must account for all relevant geometric aspects, including curvature imposed by the heart shape. The patient-specific results exhibit a strong correlation between tortuosity and helicity intensity, and the corresponding helical flow contributes directly to the observed increase in wall shear stress. Therefore, helicity intensity may prove helpful in developing a universal parameter to describe tortuosity and assess its impact on patient health. Our data suggest that increased tortuosity could have a deleterious impact via a reduction in coronary perfusion pressure, but the attendant increase in wall shear stress could afford protection against atherosclerosis.

It is estimated that the prevalence of primary adrenal insufficiency (Addison disease) is 1 in 10,000 people. There are multiple case reports and several studies that suggest a correlation between Addison disease and abnormalities of cardiac function. The pathophysiology of cardiac abnormalities in this condition is incompletely understood. This review explores what is currently known about the cardiac manifestations of Addison disease.

Key Message: C -terminally fused Strep -tag II is removed from rhuEPO expressed in tobacco plants. The finding suggests that direct fusion of purification tags at the C -terminus of rhuEPO should be avoided. Asialo-erythropoietin (asialo-EPO), a desialylated form of EPO, is a potent tissue-protective agent. Recently, we and others have exploited a low-cost plant-based expression system to produce recombinant human asialo-EPO (asialo-rhuEPO(P)). To facilitate purification from plant extracts, Strep-tag II was engineered at the C-terminus of EPO. Although asialo-rhuEPO(P) was efficiently expressed in transgenic tobacco plants, affinity purification based on Strep -tag II did not result in the recovery of the protein. In this study, we investigated the stability of Strep-tag II tagged asialo-rhuEPO(P) expressed in tobacco plants to understand whether this fused tag is cleaved or inaccessible. Sequencing RT-PCR products confirmed that fused DNA sequences encoding Strep-tag II were properly transcribed, and three-dimensional protein structure model revealed that the tag must be fully accessible. However, Western blot analysis of leaf extracts and purified asialo-rhuEPO(P) revealed that the Strep-tag II was absent on the protein. Additionally, no peptide fragment containing Strep-tag II was identified in the LC-MS/MS analysis of purified protein further supporting that the affinity tag was absent on asialo-rhuEPO(P). However, Strep-tag II was detected on asialo-rhuEPO(P) that was retained in the endoplasmic reticulum, suggesting that the Strep-tag II is removed during protein secretion or extraction. These findings together with recent reports that C-terminally fused Strep-tag II or IgG Fc domain are also removed from EPO in tobacco plants, suggest that its C-terminus may be highly susceptible to proteolysis in tobacco plants. Therefore, direct fusion of purification tags at the C-terminus of EPO should be avoided while expressing it in tobacco plants.

Asialoerythropoietin (asialo-EPO) is a desialylated form of human glycoprotein hormone erythropoietin (EPO), which has been reported to be neuro-, cardio-, and renoprotective in animal models of organ injuries. Since the current method of production of asialo-EPO from mammalian cell-made recombinant human EPO (rhuEPO(M)) by enzymatic desialylation is not commercially viable, we and others used plant-based expression systems to produce recombinant human asialo-EPO (asialo-rhuEPO(P)). Despite achieving high expression levels in plants, its purification from plant extracts has remained a greater challenge, which has prevented studying its tissue-protective effects and translating it into clinical practice. In this study, a procedure was developed to purify asialo-rhuEPO(P) from transgenic tobacco leaf tissues in two steps: ion-exchange chromatography based on its high pI (8.75) to separate it from acidic plant proteins, and immunoaffinity chromatography to obtain pure asialo-rhuEPO(P). Using this process, up to 31% of the asialo-rhuEPO(P) could be recovered to near homogeneity from plant extracts. This work demonstrates that asialo-rhuEPO(P) expressed in tobacco plants could be purified in high yield and purity using minimal steps, which might be suitable for scale-up. Furthermore, the ion-exchange chromatography step together with the use of protein-specific antibody column could be used to purify a wide variety of basic recombinant proteins from transgenic leaf tissues., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Acute myocardial infarction and acute myeloid leukemia are rarely reported as concomitant conditions. The management of ST-elevation myocardial infarction (STEMI) in patients who have acute myeloid leukemia is challenging: the leukemia-related thrombocytopenia, platelet dysfunction, and systemic coagulopathy increase the risk of bleeding, and the administration of thrombolytic agents can be fatal. We report the case of a 76-year-old man who presented emergently with STEMI, myelodysplastic syndrome, and newly recognized acute myeloid leukemia transformation. Standard antiplatelet and anticoagulation therapy were contraindicated by the patient's thrombocytopenia and by his reported ecchymosis and gingival bleeding upon admission. He declined cardiac catheterization, was provided palliative care, and died 2 hours after hospital admission. We searched the English-language medical literature, found 8 relevant reports, and determined that the prognosis for patients with concomitant STEMI and acute myeloid leukemia is clearly worse than that for either individual condition. No guidelines exist to direct the management of STEMI and concomitant acute myeloid leukemia. In 2 reports, dual antiplatelet therapy, anticoagulation, and drug-eluting stent implantation were used without an increased risk of bleeding in the short term, even in the presence of thrombocytopenia. However, we think that a more conservative approach--balloon angioplasty with the provisional use of bare-metal stents--might be safer. Simultaneous chemotherapy for the acute myeloid leukemia is crucial. Older age seems to be a major risk factor: patients too frail for emergent treatment can die within hours or days.

Asialo-erythropoietin, a desialylated form of human erythropoietin (EPO) lacking hematopoietic activity, is receiving increased attention because of its broader protective effects in preclinical models of tissue injury. However, attempts to translate its protective effects into clinical practice is hampered by unavailability of suitable expression system and its costly and limit production from expensive mammalian cell-made EPO (rhuEPO(M)) by enzymatic desialylation. In the current study, we took advantage of a plant-based expression system lacking sialylating capacity but possessing an ability to synthesize complex N-glycans to produce cytoprotective recombinant human asialo-rhuEPO. Transgenic tobacco plants expressing asialo-rhuEPO were generated by stably co-expressing human EPO and β1,4-galactosyltransferase (GalT) genes under the control of double CaMV 35S and glyceraldehyde-3-phosphate gene (GapC) promoters, respectively. Plant-produced asialo-rhuEPO (asialo-rhuEPO(P)) was purified by immunoaffinity chromatography. Detailed N-glycan analysis using NSI-FTMS and MS/MS revealed that asialo-rhuEPO(P) bears paucimannosidic, high mannose-type and complex N-glycans. In vitro cytoprotection assays showed that the asialo-rhuEPO(P) (20 U/ml) provides 2-fold better cytoprotection (44%) to neuronal-like mouse neuroblastoma cells from staurosporine-induced cell death than rhuEPO(M) (21%). The cytoprotective effect of the asialo-rhuEPO(P) was found to be mediated by receptor-initiated phosphorylation of Janus kinase 2 (JAK2) and suppression of caspase 3 activation. Altogether, these findings demonstrate that plants are a suitable host for producing cytoprotective rhuEPO derivative. In addition, the general advantages of plant-based expression system can be exploited to address the cost and scalability issues related to its production.

Desirudin, a subcutaneously (SC) administered direct thrombin inhibitor, is indicated for prevention of venous thromboembolic events (VTEs) after total joint replacement surgery. DESIR-ABLE (multicenter trial of desirudin for the prophylaxis of thrombosis: an alternative to heparin-based anticoagulation) was a multicenter, open-label, single-arm study of hospitalized patients requiring VTE protection designed to extend the safety profile for desirudin to include a broad population of perioperative/critically ill patients. The primary end point was major bleeding. A total of 516 patients undergoing major surgery (378, 73%) or who were medically ill with prolonged immobility (138, 27%) were enrolled at 19 centers and received desirudin 15 mg Q12H. Many patients had high-risk features for bleeding and thrombosis such as thrombocytopenia (<100 x 10(9)/mL, n = 50), severe obesity (body mass index >35, n = 145), and renal impairment (creatinine clearance <60 mL/min, n = 292). There were no major bleeds and no VTE-related deaths in this study. The DESIR-ABLE demonstrated the safety of desirudin in critically ill perioperative and medical patients. Trials in specific surgical or medically ill patients are needed to confirm these findings.

Background: Doxorubicin (DOX) is associated with premature cardiovascular events including myocardial infarction. This study was performed to determine if the weekly administration of DOX influenced coronary arteriolar medial and/or adventitial wall thickening., Methods: Thirty-two male Sprague-Dawley rats aged 25.1± 2.4 weeks were randomly divided into three groups and received weekly intraperitoneal injections of normal saline (saline, n = 7), or low (1.5 mg/kg to 1.75 mg/kg, n = 14) or high (2.5 mg/kg, n = 11) doses of DOX. The animals were treated for 2-12 weeks, and euthanized at pre-specified intervals (2, 4, 7, or 10+ weeks) to obtain histopathologic assessments of coronary arteriolar lumen diameter, medial wall thickness, adventitial wall thickness, and total wall thickness (medial thickness + adventitial thickness)., Results: Lumen diameter was similar across all groups (saline: 315±34 µm, low DOX: 286±24 µm, high DOX: 242±27 µm; p = 0.22). In comparison to animals receiving weekly saline, animals receiving weekly injections of 2.5 mg/kg of DOX experienced an increase in medial (23±2 µm vs. 13±3 µm; p = 0.005), and total wall thickness (51±4 µm vs. 36±5 µm; p = 0.022), respectively. These increases, as well as adventitial thickening became more prominent after normalizing for lumen diameter (p<0.05 to p<0.001) and after adjusting for age, weight, and total cumulative DOX dose (p = 0.02 to p = 0.01). Animals receiving low dose DOX trended toward increases in adventitial and total wall thickness after normalization to lumen diameter and accounting for age, weight, and total cumulative DOX dose (p = 0.06 and 0.09, respectively)., Conclusion: In conclusion, these data demonstrate that weekly treatment of rats with higher doses of DOX increases coronary arteriolar medial, adventitial, and total wall thickness. Future studies are warranted to determine if DOX related coronary arteriolar effects are reversible or preventable, exacerbate the known cardiomyopathic effects of DOX, influence altered resting or stress-induced myocardial perfusion, or contribute to the occurrence of myocardial infarction.

A number of cutaneous disorders encountered by the dermatologist have overlapping cardiac pathology. In recent years, many genetic linkages common to pathological processes in the cutaneous and cardiovascular systems have been identified. This review will describe primary cutaneous disorders with potential cardiac manifestations, including congenital syndromes, inherited cutaneous disorders associated with later cardiovascular disease, and syndromes associated with early cardiovascular pathology. The dermatologist may be the first to diagnose cutaneous findings associated with underlying cardiovascular disease; therefore, it is of prime importance for the dermatologist to be aware of these associations and to direct the appropriate workup., (Copyright © 2012 American Academy of Dermatology, Inc. Published by Mosby, Inc. All rights reserved.)

Recombinant hirudins (desirudin, lepirudin) are direct thrombin inhibitors administered as anticoagulants for heparin-induced thrombocytopenia (HIT) and venous thromboembolism (VTE) prophylaxis. Although these small polypeptides are widely used, concern exists over reports of antigenicity. In the largest study of r-hirudin immunogenicity to-date, we evaluated the prevalence, quantity and specificity of IgG immune responses to desirudin (15 mg SC q12h for as long as clinically required) in 245 surgical and medically-ill subjects enrolled in DESIRABLE, a multicenter, open-label, clinical trial of hospitalized patients requiring VTE prophylaxis. Sera obtained before and 30 days after desirudin administration were analyzed for IgG anti-desirudin by immunoenzymetric assay using immobilized desirudin to bind desirudin-reactive antibody and peroxidase conjugated monoclonal-anti-human IgG Fc to detect bound IgG antibody. Of 245 study subjects, 19 (7.7%) were antibody "responders" (>2-fold increase in IgG antibody levels with >50% inhibition by desirudin 30 days post-treatment). There were no differences between responders and non-responders in incidence of clinical outcomes or bleeding-related adverse events. Forty-six patients had detectable desirudin-reactive IgG antibody prior to treatment, with no significant increase in antibody levels after exposure and no increase in clinical events. The origin of pre-existing hirudin-reactive IgG antibody requires further investigation involving suspected anti-thrombin-thrombin interactions. These results indicate a low potential for immunogenicity, with <8% of patients developing IgG antibodies after desirudin administration for VTE prophylaxis. In contrast to reports on lepirudin, production of anti-hirudin antibodies to desirudin has no apparent effect on clinical events., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Unlabelled: Erythropoietin (EPO) is a glycoprotein hormone that displays both hematopoietic and tissue-protective functions by binding to two distinct receptors. Recombinant human EPO (rhuEPO) is widely used for the treatment of anemia, but its use for tissue protection is limited because of potentially harmful increases in red blood cell mass when higher doses of rhuEPO are used. Recent studies have shown that asialoerythropoietin (asialo-rhuEPO), a desialylated form of rhuEPO, lacks hematopoietic activity, but retains cytoprotective activity. Currently, a small amount of asialo-rhuEPO is produced by enzymatic desialylation of rhuEPO. The prohibitive cost of rhuEPO, however, is a major limitation of this method. Plants have the ability to synthesize complex N-glycans, but lack enzymatic activities to add sialic acid and β1,4-galactose to N-glycan chains. Plants could be genetically engineered to produce asialo-rhuEPO by introducing human β1,4-galactosyltransferase. The penultimate β1,4-linked galactose residues are important for in vivo biological activity. In this proof of concept study, we show that tobacco plants co-expressing human β1,4-galactosyltransferase and EPO genes accumulated asialo-rhuEPO. Purified asialo-rhuEPO binds to an Erythrina cristagalli lectin column, indicating that its N-glycan chains bear terminal β1,4-galactose residues and that the co-expressed GalT is functionally active. Asialo-rhuEPO interacted with the EPO receptor (EPOR) with similar affinity as rhuEPO, implying that it was properly folded. The strategy described here provides a straightforward way to produce asialo-rhuEPO for research and therapeutic purposes., Key Message: N-glycosylation pathway in tobacco plants could be genetically engineered to produce a tissue-protective cytokine, asialoerythropoietin (a desialylated form of human hormone erythropoietin).

Background: Niacin is suboptimally used in patients because it causes flushing and erythema. These side effects have been attributed to release of the vasodilating prostaglandin D2, generated in a reaction catalyzed by cyclooxygenase-1. Aspirin reduces but does not completely eliminate these side effects. Because some patients are resistant to its antiplatelet effects, we hypothesized that patients with persistent niacin-induced erythema might be aspirin resistant., Methods: Platelet function studies (via the use of a whole-blood platelet function aggregometer [VerifyNow; Accumetrics, San Diego, CA] with end points of aspirin reaction unit [ARUs] and P2Y12 reaction units [PRUs]) were performed on 32 healthy, drug-free subjects before and after 324 mg of aspirin. A niacin skin test with the use of topical methylnicotinate was also performed before and after the administration of aspirin. Responses to methyl nicotinate were assessed by a reaction score and by counting the time to first visible redness (TTR)., Results: All subjects had an expected decrease in arachidonic acid induced platelet response (ARU 642.8 ± 47.20 before to 431.5 ± 41.1 after aspirin, P < .0001) without a significant change in the PRU. The reaction score and TTR were prolonged by aspirin at methylnicotinate concentrations ≥ 0.001 M. Although no subject had aspirin resistance (defined as ARU > 550), there was considerable variability in skin responses with erythema elicited in all subjects at the greatest concentrations. There was no difference in the ARU for subjects with TTR values above and below the mean, indicating that aspirin resistance does not explain the variation in skin responses to a topical niacin derivative., Conclusion: Aspirin resistance is unlikely to be a significant contributor to the persistent erythema and flushing in niacin-treated patients., (Copyright © 2011 National Lipid Association. Published by Elsevier Inc. All rights reserved.)

Ischemic heart disease remains one of the most common causes of mortality in developed countries. Recently, stem cell therapy is being considered for treating ischemic heart diseases. On the other hand, there has been evidence of chondro-osteogenic mass formation after stem cell injection in the heart. In a recent publication, Chiavegato et al. (J Mol Cell Cardiol. 42 (2007) 746-759) has suggested that amniotic fluid-derived stem (AFS) cells cause chondro-osteogenic masses in the infarcted heart. The goal of the current study was to further examine the formation of such masses, specifically, the role of AFS cells in this process. Our results confirm the presence of similar bone-like masses in the left ventricular wall of infarcted rats; however, this phenomenon occurred independent of AFS cell injection into the myocardium. Moreover, AFS cell injection did not increase the presence of chondro-osteogenic masses. Echocardiographic analysis of large infarctions in rats frequently revealed the presence of echogenic structures in the left ventricular wall. We further demonstrated a significant relationship between the infarction size and chondro-osteogenic formation and subsequent decrease in cardiac function. Collectively, our study indicates that chondro-osteogenic differentiation can take place in infarcted rat heart independent of cell injection. These results have significant implications for future design and testing of stem cell therapy for treatment of cardiac muscle diseases., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Cardiac rupture after a myocardial infarction is an uncommon event with devastating consequences. Although the clinical features of rupture have been described, the genetic and molecular influences on this outcome in patients are less certain. In mice, at least 17 genetic models have been developed that enhance or suppress the likelihood of rupture postmyocardial infarction. The purpose of this review is to describe these recent advances, recognizing that nearly all of the information has been obtained from mouse models of free wall rupture. Although it is probable that the same genetic determinants apply to septal and papillary muscle rupture, the possibility remains that there are unique modulators of risk for rupture at differing anatomic sites within the heart. It is likely that the candidate genes also influence rupture in humans, although this conclusion must be confirmed. The mouse models will be helpful to direct future proteomic and genomic studies in patients and may already suggest certain fundamental pathways. For example, the essential role of collagen production and stabilization postmyocardial infarction may direct therapies to enhance collagen cross-linking and limit its degradation as a strategy to reduce rates of rupture and enhance myocardial healing.

Background: Kidney failure (stage 5 chronic kidney disease [CKD]) is an independent risk factor for stent thrombosis (ST). Moderate (stage 3-4) CKD and proteinuria are both associated with adverse cardiovascular events, including worse outcomes after myocardial infarction (MI). Whether moderate CKD and proteinuria increase the risk of ST after MI is not known. This study evaluated the risk of ST associated with moderate CKD and dipstick proteinuria., Methods: We retrospectively analyzed clinical and laboratory data from 956 non-stage 5 CKD patients who were admitted with MI and received intracoronary stenting. Clinical follow-up was collected at 1 year for definite or probable ST, as well as for all-cause mortality, nonfatal MI or death, and target vessel revascularization or coronary artery bypass graft surgery., Results: After adjustment for multiple clinical and biochemical covariates, patients with both estimated glomerular filtration rate (GFR) of 15 to 59 mL min(-1) 1.73 m(-2) and > or =30 mg/dL dipstick proteinuria had increased cumulative incidence of ST (hazard rate [HR] 3.69, 95% CI 1.54-8.89), all-cause mortality (HR 2.68, 95% CI 1.34-5.37), and nonfatal MI or death (HR 3.20, 95% CI 1.77-5.81) at 1 year. In addition, estimated GFR of 15 to 59 mL min(-1) 1.73 m(-2) was a significant independent predictor of ST (HR 2.61, 95% CI 1.33-5.10). Dipstick proteinuria > or =30 mg/dL was associated with a trend toward increased risk for all outcomes., Conclusions: In an acute MI population, moderate CKD was identified as a novel prognostic marker for ST. In addition, patients with both decreased GFR and proteinuria had higher incidences of all-cause mortality and nonfatal MI or death than patients with either condition alone.

Myocardial necrosis is now recognized as a common feature of thrombotic thrombocytopenic purpura (TTP), usually due to platelet plugging in the microvasculature. Despite reports that TTP patients with myocardial damage have higher morbidity and mortality, there are no established guidelines for managing these high-risk patients. The universal occurrence of thrombocytopenia and variable findings including renal dysfunction present unique challenges in this setting. Established therapies including plasma exchange and immunosuppression are the mainstay of therapy for all TTP patients. For the subset of patients with myocardial damage, therapy with more potent antiplatelet drugs, agents that enhance NO availability and alleviate vasospasm and drugs that suppress von Willebrand factor levels may have additional benefit. However, clinical trials are needed to optimize therapy for this subset of TTP patients.

Background: Several case reports and series have described myocardial infarctions (MIs) in patients hospitalized for thrombotic thrombocytopenic purpura (TTP). The exact magnitude and outcome of this complication are unknown., Methods: Electronic medical records for patients admitted to Wake Forest University Baptist Medical Center were examined from 1996 to 2005. Those patients having a diagnosis of TTP during the hospitalization period were included in the analysis. Only patients' initial episodes of TTP were analyzed. Baseline cardiac and TTP risk factors were documented. Outcomes analyzed included MIs, arrhythmias, development of congestive heart failure and death., Results: Eighty-five patients diagnosed with TTP were identified with 13 (15.3%) having MIs, as defined by an elevation of cardiac enzymes. Median troponin I value was 5.9 ng/mL (range 3.7-8.8 ng/mL). Twelve patients had non-ST segment elevation MIs and one had ST segment elevation. Two of 13 patients who had echocardiographic analysis had documented wall motion abnormalities. There was no difference between non-MI and MI patients in cardiac risk factors, prior cardiac events, history of thromboembolic disease or heart failure. There was no in-hospital mortality difference., Conclusion: MI is an important complication of TTP, identified in 15.3% of patients in our study. Routine cardiovascular evaluation with cardiac enzymes, electrocardiography, and telemetry is warranted in acute TTP patients. Appropriate intervention is yet to be determined.

Human tissue transglutaminase (TGM2) is a calcium-dependent crosslinking enzyme involved in the posttranslational modification of intra- and extracellular proteins and implicated in several neurodegenerative diseases. To find specific inhibitors to TGM2, two structurally diverse chemical libraries (LOPAC and Prestwick) were screened. We found that ZM39923, a Janus kinase inhibitor, and its metabolite ZM449829 were the most potent inhibitors with IC(50) of 10 and 5 nM, respectively. In addition, two other inhibitors, including tyrphostin 47 and vitamin K(3), were found to have an IC(50) in the micromolar range. These agents used in part a thiol-dependent mechanism to inhibit TGM2, consistent with the activation of TGM2 by reduction of an intramolecular disulfide bond. These inhibitors were tested in a polyglutamine-expressing Drosophila model of neurodegeneration and found to improve survival. The TGM2 inhibitors we discovered may serve as valuable lead compounds for the development of orally active TGM2 inhibitors to treat human diseases.

Background: After myocardial infarction (MI), biomarkers can be helpful to identify patients who might benefit from more intensive therapies. The prothrombin time-derived fibrinogen (PTDF) assay is widely available and relatively inexpensive. We determined whether PTDF predicts events in patients with MI and compared this assay with brain natriuretic peptide (BNP) and C-reactive protein (CRP)., Methods: We retrospectively analyzed data from 915 patients admitted with MI. Follow-up was collected at 1 year for major adverse cardiac events (MACE) defined as death from any cause, nonfatal MI or death, target vessel revascularization, or coronary artery bypass grafting., Results: Patients in the fourth quartile of PTDF were older and had more risk factors but fewer ST-elevation MI and lower peak troponin values. The fourth quartiles of PTDF, CRP, and BNP were associated with increased MACE compared with the first quartiles with hazard ratios of 2.08 (1.30-3.33), 1.94 (1.22-3.07), and 2.56 (1.57-4.18), respectively, findings that remained significant after adjustment. When outcomes by strata of PTDF were examined, CRP failed to add additional prognostic value. Higher BNP levels predicted MACE in the upper but not lower stratum of PTDF., Conclusion: In patients with MI, PTDF is a predictor of MACE at 1 year, with equivalent value compared to BNP and CRP. With low PTDF levels, neither BNP nor CRP adds prognostic value. At elevated PTDF values, higher BNP, but not CRP, identifies a higher-risk population. Therefore, PTDF can be substituted for CRP, with BNP being useful in the presence of elevated PTDF.

Prognostic markers are needed to identify patients with acute coronary syndrome (ACS) who are at high risk for adverse events. Although the search for new biomarkers is quite active, prognostic information is available from routine hematologic tests, such as the complete blood count. For example, elevated white blood cell counts during ACS are associated with increased risk of mortality, heart failure, shock, and left ventricular dysfunction. Anemia is associated with increased risk of mortality, whereas elevated platelet counts predict poorer clinical and angiographic outcomes. In this review, we summarize the literature regarding the use of clinical hematology tests including white blood cell count, hemoglobin and hematocrit values, and platelet count as prognostic markers in patients with ACS, and we describe potential mechanisms to explain these associations.

Introduction: Patients with non-O blood groups have higher plasma von Willebrand factor (vWF) levels than those with type O. vWF mediates platelet adhesion, aggregation and thrombosis. These considerations likely explain the prior observations that non-O patients have higher rates of arterial and venous thromboembolic events. However, the effect of blood group status on size of MI, procedural findings and outcomes after PCI for MI have not been reported., Methods: We analyzed 1198 patients who underwent percutaneous coronary intervention for acute myocardial infarction between 10/03 and 8/06, and who had ABO blood group status and clinical follow-up., Results and Conclusions: Patients with O blood type were slightly older (62 +/- 13 vs. 60 +/- 13 years; p = 0.017) had a higher prevalence of hypercholesterolemia (67% vs. 58%; p = 0.002), and had a higher burden of atherosclerosis with more vascular disease (17% vs. 13%; p = 0.017) and higher prevalence of previous PCI (22% vs. 17%; p = 0.025). Non-O blood group patients had larger infarcts as measured by median peak troponin (33 vs. 24; p = 0.037), total CK (721 vs. 532; p = 0.012) and CK-MB (101 vs. 68; p = 0.010). At PCI, non-O patients had increased visible thrombus and reduced TIMI flow pre-procedure. However, there were no differences in procedural success, in-hospital blood transfusion or occurrence of MACE at 1 year follow-up. Our data demonstrate that non-O compared to O blood groups patients have higher thrombus burden despite less extensive atherosclerosis. Nevertheless, outcomes at 1 year were similar.

Nephrogenic systemic fibrosis (NSF), also known as nephrogenic fibrosing dermopathy (NFD), occurs in renal failure patients after gadolinium contrast exposure. The fibrosis of the dermis and subcutaneous septae accompanies fibrosis of other organs, including the heart, liver, lungs, and muscle. The fibrotic skin demonstrates increased dermal collagen, fibroblasts, and mucin. The mechanism by which gadolinium is associated with fibrosis is not known. We tested the hypothesis that upregulation of transglutaminases contributes to the fibrosis seen in the organs, including skin, of renal failure patients exposed to gadolinium contrast. We performed immunohistochemical studies using antibodies to transglutaminase-2, factor XIIIa, transglutaminase isopeptide, and the histiocyte marker CD68 on five archived skin biopsies of NSF. The results indicate that the dermal fibroblasts and histiocytes of NSF express transglutaminase-2, CD68, factor XIIIa, and transglutaminase isopeptide, indicating increased expression and/or activation of transglutaminases in NSF. We recommend further research into the use of transglutaminase inhibitors in the treatment and prevention of NSF.

Myocarditis is a serious and life-threatening illness, which can pose a significant diagnostic dilemma at presentation. We report 2 patients with clinical myocarditis who had distinctive findings at coronary computed tomography. Both patients demonstrated delayed myocardial enhancement with iodinated contrast. The morphologic features of the enhancement were similar to the myocardial enhancement with gadolinium contrast on magnetic resonance imaging recently described in patients with myocarditis, and different from the enhancement patterns seen in patients with myocardial infarction. These findings are not previously reported in the literature. As computed tomography is more widely available for use in the acute setting than magnetic resonance imaging, these findings may have significant clinical utility and warrant further investigation.

FKBP-12 mediates the immunosuppressive actions of FK506 and rapamycin, and modulates the activities of the ryanodine, IP3 and type 1 TGF-ss receptors. Additionally, FKBP-12 possesses cis-trans peptidylprolyl isomerase (rotamase) activity. We have discovered that recombinant FKBP-12 readily forms a dimer and a small amount of trimer under nonreducing conditions. A mutant with substitution at the sole cysteine residue of FKBP-12 (C23S) did not form dimers or trimers. Using mutants with 5% or less rotamase activity, the formation of dimers was independent of enzymatic activity. The formation of trimers was abrogated by a F36Y substitution, even though dimer formation was preserved. Dimers were also observed with native FKBP-12 that was detached from rabbit skeletal muscle ryanodine receptors using FK590. The multimers of FKBP-12 could interact with molecular targets distinctly from the FKBP-12 monomer, for example, by facilitating the assembly of multimeric receptors or coordinating the activity of receptor subunits.

Background: The treatment of cocaine-related acute coronary syndromes presents unique challenges. Although percutaneous coronary intervention in cocaine abusers appears to be safe in the short term, longer-term outcomes have not been reported. We postulated that cocaine use would be associated with increased risk for stent thrombosis., Methods: We report 30-day and 9-month clinical outcomes including stent thrombosis, myocardial infarction, repeat revascularization, and death in 71 cocaine abusers who underwent percutaneous coronary intervention at our institution (66 of whom received a stent) compared with 3216 control patients. Propensity score-matched analysis was performed to control for statistical bias present in nonrandomized study populations., Results: Stent thrombosis occurred in 5 (7.6%) of the 66 stented cocaine abusers during the 9-month follow-up period compared to a 0.6% rate of stent thrombosis in the control database, a highly statistically significant difference (P < .001). In the propensity analysis, stent thrombosis occurred in 4 stented cocaine abusers and 0 of 70 matched controls (6.2% vs 0%; P = .04) throughout the 9-month follow-up period. There was no significant difference in overall rates of myocardial infarction, death, or repeat revascularization at 9 months., Conclusions: Because of the increased risk of stent thrombosis, consideration should be given to a more conservative approach in cocaine abusers who present with acute coronary syndromes.

A major potential side effect of heparin is immunogenicity, eliciting antibody development to a protein complex comprised of platelet factor 4 and heparin. Nevertheless, the clinical implications of heparin antibody positive patients remain broad, ranging from no apparent clinical consequences to life-threatening arterial and venous thromboemboli. The "Iceberg Model" has been proposed to depict this spectrum, with a relatively large population of antibody-positive patients forming the base of the iceberg, a smaller population of thrombocytopenic patients in the middle and a limited number of patients with thrombocytopenia and thrombosis comprising the apex. An underlying assumption of this model is that thrombosis occurs only in settings of relative or absolute thrombocytopenia. However, several recent studies suggest that antibody formation to platelet factor 4/heparin complexes, even in the absence of thrombocytopenia, may be associated with thrombotic events. In this review, we summarize these data, consider potential mechanisms for thrombosis, and suggest recommendations for testing and management of antibody-positive patients.

The ability of clopidogrel to inhibit platelet function in patients with congestive heart failure (CHF) was proved by the PLUTO-CHF trial. We retrospectively analyzed platelet characteristics with respect to CHF etiology, class, and ejection fraction in patients enrolled in the PLUTO-CHF study. Twenty-five patients were divided by CHF etiology, severity, and ejection fraction. All patients received aspirin 325 mg for at least 1 month prior to screening. Platelet function studies were performed at baseline and after 30 days of therapy. There were no differences in platelet parameters dependent on clinical characteristics of CHF, except for a significant (P = 0.023) decrease in platelet/endothelial cell adhesion molecule 1 (PECAM-1) expression in the New York Heart Association class III-IV due to the higher baseline values. Therapy with clopidogrel resulted in a significant inhibition of platelet activity assessed by ADP-induced and epinephrine-induced aggregation, closure time, expression of PECAM-1, glycoprotein Ib, glycoprotein IIb/IIIa antigen, glycoprotein IIb/IIIa activity with PAC-1, CD151, and reduced formation of platelet-leukocyte conjugates when compared with baseline. Clopidogrel provides antiplatelet protection in the broad spectrum of patients with CHF independently of its etiology, severity, or myocardial contractility. This uniform platelet inhibition with clopidogrel may be an important consideration in designing future large-scale clinical trials.

Millions of women still use postmenopausal hormone therapy (HT). We genotyped 2090 women in Heart and Estrogen/progestin Replacement Study for functional polymorphisms in GP1BA and GP6 and assessed the coronary heart disease (CHD) event rate over 5.8 years of follow-up. In patients receiving placebo, there was an increased CHD death/myocardial infarction (MI)/unstable angina (UA) event rate in carriers of the GP1BA -5C allele (adjusted [adj] P = .006). HT increased the hazard ratio (HR) of CHD events in patients with the GP1BA -5TT genotype by 16% and reduced the HR in patients with the TC+CC genotypes by 46% (adj interaction P < .001). HT reduced the HR in patients with the GP6 13254TT genotype by 17% but increased the HR in patients with the TC+CC genotypes by 35% (adj interaction P < .001). Furthermore, HT increased the HR of CHD events in patients with the GP1BA -5TT plus GP6 13254TC+CC genotypes by 57% and reduced the HR in patients with the GP1BA -5TC+CC plus GP6 13254TT genotypes by 55% (adj interaction P < .001). In postmenopausal women with established CHD, these polymorphisms of platelet genes were predictors of CHD events and significantly modified the effects of HT on CHD risk. It will be important to replicate these findings in other studies.

Background/aims: The transglutaminases (TG2 and factor XIIIa) may contribute to the stability of arteries by cross-linking a variety of substrates, including extracellular matrix proteins and protease inhibitors. The preferred substrates have never been determined, however., Methods: We used an amine donor, 5-biotinamidopentylamine, that is covalently linked to acceptor glutamine residues, to determine transglutaminase substrates in carotid endarterectomy tissue., Results: The incorporation of 5-biotinamidopentylamine was calcium dependent, resulting in the labeling of several proteins that were detected by streptavidin-peroxidase and purified over a monomeric avidin affinity column. A major band of 42 kDa that was eluted from the column was sequenced along with 2 additional bands (80 and 65 kDa), revealing an internal fragment of vimentin, transferrin and albumin, respectively. Vimentin dimers were detected in 5 out of 5 carotid plaque homogenates., Conclusions: Vimentin is a major arterial substrate for transglutaminases. It has previously been shown that TG2 activity and vimentin contribute to vasomotor activity of arteries. Furthermore, transglutaminases (both TG2 and factor XIIIa), as well as vimentin, regulate structural alterations (inward remodeling) that occur in response to low flow states. Transglutaminase-mediated vimentin dimerization produces a novel unifying pathway by which vasodilatory and remodeling responses may be regulated., (Copyright 2007 S. Karger AG, Basel.)

Objective: We tested associations of the growth factors VEGF, FGF-2, HGF and PDGF-BB with coronary artery calcium scores and cardiovascular events (CVD) in type 2 diabetes mellitus (T2DM)., Methods: A cross-sectional study selected 40 frequency matched (by age, gender and race) subjects with T2DM from the first (0-111) and the third (> 1400) coronary artery calcium (CAC) score tertiles in the Diabetes Heart Study (DHS), in which 36 were with and 41 were without history of CVD events. Plasma levels of VEGF, FGF-2, HGF and PDGF-BB were measured in all subjects., Results: None of the growth factors was significantly different between the first and third CAC score tertiles. Mean plasma FGF-2 and PDGF-BB levels were significantly higher in the group without prior CVD events compared with the group with prior CVD events [mean(95%CI): 219.20 (194.42-247.15) vs. 152.93 (135.64-172.43) pg/ml, p=0.03] and [mean(95%CI): 106.70 (89.12-127.74) vs. 61.56 (50.91-74.44) pg/ml, p=0.03], respectively. Subgroup analysis in the first CAC tertile showed a significantly higher PDGF-BB levels in those without compared with those with CVD events [mean (95%CI): 208.36 (190.57-228.15) vs. 102.93 (80.64-125.21) pg/ml, p=0.004]., Conclusion: Plasma growth factor levels were not significantly different between the extremes of CAC scores in T2DM. However, low plasma levels of PDGF-BB and FGF-2 are associated with prior cardiovascular events in T2DM. Studies are needed to confirm our results and also to establish temporality of this association.