PubMed Health⌕ Search

Biomedical subjects

Bruce M McManus

Publications and source records attributed to Bruce M McManus.

At least 19 recordsLinked to original sources
PubMed2007

Dysregulation of the ubiquitin-proteasome system by curcumin suppresses coxsackievirus B3 replication.

Curcumin (diferuloylmethane), a natural polyphenolic compound extracted from the spice turmeric, has been reported to have anti-inflammatory, antioxidant, and antiproliferative properties by modulating multiple cellular machineries. It inhibits several intracellular signaling pathways, including the mitogen-activated protein kinases (MAPKs), casein kinase II (CKII), and the COP9 signalosome (CSN), in various cell types. It has also been recently demonstrated that exposure to curcumin leads to the dysregulation of the ubiquitin-proteasome system (UPS). Coxsackievirus infection is associated with various diseases, including myocarditis and dilated cardiomyopathy. In searching for new antiviral agents against coxsackievirus, we found that treatment with curcumin significantly reduced viral RNA expression, protein synthesis, and virus titer and protected cells from virus-induced cytopathic effect and apoptosis. We further demonstrated that reduction of viral infection by curcumin was unlikely due to inhibition of CVB3 binding to its receptors or CVB3-induced activation of MAPKs. Moreover, gene silencing of CKII and Jab1, a component of CSN, by small interfering RNAs did not inhibit the replication of coxsackievirus, suggesting that the antiviral action of curcumin is independent of these pathways. Finally, we showed that curcumin treatment reduced both the 20S proteasome proteolytic activities and the cellular deubiquitinating activities, leading to increased accumulation of ubiquitinated proteins and decreased protein levels of free ubiquitin. We have recently demonstrated that the UPS-mediated protein degradation and/or modification plays a critical role in the regulation of coxsackievirus replication. Thus, our results suggest an important antiviral effect of curcumin wherein it potently inhibits coxsackievirus replication through dysregulation of the UPS.

Apoptosis↗
PubMed2006

Allograft vasculopathy versus atherosclerosis.

Over the last 4 decades, heart transplantation (HTx) has evolved as a mainstream therapy for heart failure. Approximately half of patients needing HTx have organ failure consequent to atherosclerosis. Despite advances in immunosuppressive drugs, long-term success of HTx is limited by the development of a particular type of coronary atherosclerosis, referred to as cardiac allograft vasculopathy (CAV). Although the exact pathogenesis of CAV remains to be established, there is strong evidence that CAV involves immunologic mechanisms operating in a milieu of nonimmunologic risk factors. The immunologic events constitute the principal initiating stimuli, resulting in endothelial injury and dysfunction, altered endothelial permeability, with consequent myointimal hyperplasia and extracellular matrix synthesis. Lipid accumulation in allograft arteries is prominent, with lipoprotein entrapment in the subendothelial tissue, through interactions with proteoglycans. The apparent endothelial "intactness" in human coronary arteries of the transplanted heart suggest that permeability and function of the endothelial barrier altered. Various insults to the vascular bed result in vascular smooth muscle cell (SMC) activation. Activated SMCs migrate from the media into the intima, proliferate, and elaborate cytokines and extracellular matrix proteins, resulting in luminal narrowing and impaired vascular function. Arteriosclerosis is a broad term that is used to encompass all diseases that lead to arterial hardening, including native atherosclerosis, postangioplasty restenosis, vein bypass graft occlusion, and CAV. These diseases exhibit many similarities; however, they are distinct from one another in numerous ways as well. The present review summarizes the current understanding of the risk factors and the pathophysiological similarities and differences between CAV and atherosclerosis.

Animals↗
PubMed2006

Novel role for integrin-linked kinase in modulation of coxsackievirus B3 replication and virus-induced cardiomyocyte injury.

Viral myocarditis is a major cause of sudden cardiac death in children and young adults. Among viruses, coxsackievirus B3 (CVB3) is the most common agent for myocarditis. Recently, more consideration has been given to the role of signaling pathways in pathogenesis of enteroviral myocarditis, providing new platform for identifying a new potential therapeutic target for this, so far, incurable disease. Previously, we reported on the role of the protein kinase-B/Akt in CVB3 replication and virus-induced cell injury. Here, we report on regulation of virus-induced Akt activation by the integrin-linked kinase in infected mouse cardiomyocytes and HeLa cells. This study also presents the first observation that inhibition of ILK in CVB3-infected cells significantly improves the viability of infected cells, while blocking viral replication and virus release. Complementary experiments using a constitutively active form of Akt1 revealed that the observed protective effect of ILK inhibition is dependent on the associated downregulation of virus-induced Akt activation. To our knowledge, this is the first report of such beneficial effects of ILK inhibition in a viral infection model and conveys new insights in our efforts to characterize a novel therapeutic target for treatment of enteroviral myocarditis.

Animals↗
PubMed2006

Reduced expression of vascular endothelial growth factor paralleled with the increased angiostatin expression resulting from the upregulated activities of matrix metalloproteinase-2 and -9 in human type 2 diabetic arterial vasculature.

Impaired angiogenesis could contribute to the increased incidence of coronary and peripheral artery disease in diabetic patients. Angiogenesis is initiated by vascular endothelial growth factor (VEGF), a potent angiogenic cytokine, and suppressed by angiostatin, which is generated by matrix metalloproteinase (MMP)-2 and -9 through proteolytic cleavage of plasminogen. We hypothesized that MMP-2 and -9 were upregulated in the diabetic vasculature, resulting in increased angiostatin production and reduced blood vessel formation. In diabetic internal mammary artery samples (n=32) collected from patients undergoing coronary artery bypass grafting surgery, capillary density was only 30% of that in the nondiabetic vessels (n=32), whereas VEGF expression was reduced by 48%. Diabetes upregulated the expression and the gelatinolytic activity of MMP-2 and -9. Active MMP-2 and -9 were released from diabetic arteries, but not from nondiabetic vessels, during phenylephrine-induced vasoconstriction. Diabetes enhanced transcription and protein expression of tissue inhibitor of MMP (TIMP)-1 but had an opposite effect on TIMP-2. In diabetic vessels angiostatin was increased by 62% and was positively correlated with the activities of MMP-2 and -9 (r2=0.806 and 0.742, respectively). This report indicated a strong correlation between the upregulation of MMP-2 and MMP-9 and the increased angiostatin expression in the human diabetic arterial vasculature. The enhanced angiostatin production with a reduced VEGF formation may explain the pathogenesis of impaired angiogenesis in diabetes mellitus.

Aged↗
PubMed2006

Biologically variable bypass reduces enzymuria after deep hypothermic circulatory arrest.

BACKGROUND: Renal injury is common after open-heart surgery. Cardiopulmonary bypass contributes to the problem. We compared conventional nonpulsatile perfusion (NP) to biologically variable perfusion (BVP), which uses a computer controller to restore physiological beat-to-beat variability to roller pump flow. We hypothesized BVP would decrease renal injury after deep hypothermic circulatory arrest. METHODS: Pigs were randomly assigned to either BVP (n = 9) or NP (n = 9), cooled, arrested at 18 degrees C (1 hour), reperfused, and rewarmed and maintained normothermic (3 hours). Additional pigs had NP for a similar time as above, but without circulatory arrest (n = 3), or were sham-treated without bypass (n = 3). Hemodynamics, acid-base status, temperature, and urine volumes were measured. Urinary enzyme markers of tubular injury were compared post-hoc for gamma glutamyl transpeptidase, alkaline phosphatase, and glutathione S-transferase and by urine proteomics using mass spectrometry. RESULTS: Urine output at 1 hour after arrest was 250 +/- 129 mL with BVP versus 114 +/- 66 mL with NP (p < 0.02). All three renal enzyme markers were higher with NP after arrest compared with BVP. In animals on bypass without arrest or those sham-treated, no elevations were seen in renal enzymes. Urine proteomics revealed abnormal proteins, persisting longer with NP. Biologically variable perfusion decreased cooling to 21.0 +/- 9.0 minutes versus 31.7 +/- 7.5 minutes (p < 0.002), and decreased rewarming to 22.1 +/- 3.9 minutes versus 31.2 +/- 5.1 minutes (p < 0.002). CONCLUSIONS: Biologically variable perfusion improved urine output, decreased enzymuria, and attenuated mass spectrometry urine protein signal with more rapid temperature changes. This strategy could potentially shorten bypass duration and may decrease renal tubular injury with deep hypothermic circulatory arrest.

Acute Kidney Injury↗
PubMed2006

Versican: signaling to transcriptional control pathways.

Versican, a chondroitin sulfate proteoglycan, is one of the main components of the extracellular matrix, which provides a loose and hydrated matrix during key events in development and disease. Versican participates in cell adhesion, proliferation, migration, and angiogenesis, and hence plays a central role in tissue morphogenesis and maintenance. In addition, versican contributes to the development of a number of pathologic processes including atherosclerotic vascular diseases, cancer, tendon remodeling, hair follicle cycling, central nervous system injury, and neurite outgrowth. Versican is a complex molecule consisting of modular core protein domains and glycosaminoglycan side chains, and there are various steps of synthesis and processes regulating them. Also, there is differential temporal and spatial expression of versican by multiple cell types and in different developmental and pathological time frames. To fully appreciate the functional roles of versican as it relates to changing patterns of expression in development and disease, an in depth knowledge of versican's biosynthetic processing is necessary. The goal of this review is to evaluate the current status of our knowledge regarding the transcriptional control of versican gene regulation. We will be focusing on the signal transduction pathways, promoter regions, cis-acting elements, and trans-factors that have been characterized.

Animals↗
PubMed2005

Mathematical modelling to centre low tidal volumes following acute lung injury: a study with biologically variable ventilation.

BACKGROUND: With biologically variable ventilation [BVV--using a computer-controller to add breath-to-breath variability to respiratory frequency (f) and tidal volume (VT)] gas exchange and respiratory mechanics were compared using the ARDSNet low VT algorithm (Control) versus an approach using mathematical modelling to individually optimise VT at the point of maximal compliance change on the convex portion of the inspiratory pressure-volume (P-V) curve (Experimental). METHODS: Pigs (n = 22) received pentothal/midazolam anaesthesia, oleic acid lung injury, then inspiratory P-V curve fitting to the four-parameter logistic Venegas equation F(P) = a + b[1 + e-(P-c)/d]-1 where: a = volume at lower asymptote, b = the vital capacity or the total change in volume between the lower and upper asymptotes, c = pressure at the inflection point and d = index related to linear compliance. Both groups received BVV with gas exchange and respiratory mechanics measured hourly for 5 hrs. Postmortem bronchoalveolar fluid was analysed for interleukin-8 (IL-8). RESULTS: All P-V curves fit the Venegas equation (R2 > 0.995). Control VT averaged 7.4 +/- 0.4 mL/kg as compared to Experimental 9.5 +/- 1.6 mL/kg (range 6.6 - 10.8 mL/kg; p < 0.05). Variable VTs were within the convex portion of the P-V curve. In such circumstances, Jensen's inequality states "if F(P) is a convex function defined on an interval (r, s), and if P is a random variable taking values in (r, s), then the average or expected value (E) of F(P); E(F(P)) > F(E(P))." In both groups the inequality applied, since F(P) defines volume in the Venegas equation and (P) pressure and the range of VTs varied within the convex interval for individual P-V curves. Over 5 hrs, there were no significant differences between groups in minute ventilation, airway pressure, blood gases, haemodynamics, respiratory compliance or IL-8 concentrations. CONCLUSION: No difference between groups is a consequence of BVV occurring on the convex interval for individualised Venegas P-V curves in all experiments irrespective of group. Jensen's inequality provides theoretical proof of why a variable ventilatory approach is advantageous under these circumstances. When using BVV, with VT centred by Venegas P-V curve analysis at the point of maximal compliance change, some leeway in low VT settings beyond ARDSNet protocols may be possible in acute lung injury. This study also shows that in this model, the standard ARDSNet algorithm assures ventilation occurs on the convex portion of the P-V curve.

Algorithms↗
PubMed2005

Regulation of the versican promoter by the beta-catenin-T-cell factor complex in vascular smooth muscle cells.

The proteoglycan versican is pro-atherogenic and central to vascular injury and repair events. We identified the signaling pathways and promoter elements involved in regulation of versican expression in vascular smooth muscle cells. Phosphatidylinositol 3-kinase inhibitor, LY294002, significantly decreased versican-luciferase (Luc) promoter activity and endogenous mRNA levels. We further examined the roles of protein kinase B and glycogen synthase kinase (GSK)-3beta, downstream effectors of phosphatidylinositol 3-kinase, in the regulation of versican transcription. Co-transfection of dominant negative and constitutively active protein kinase B constructs with a versican-Luc construct decreased and increased promoter activity, respectively. Inhibition of GSK-3beta activity by LiCl augmented accumulation of beta-catenin and caused induction of versican-Luc activity as well as versican mRNA levels. Beta-catenin has no DNA binding domain, therefore it cannot directly induce transcription of the versican promoter. Software analysis of the versican promoter revealed two potential binding sites for T-cell factors (TCFs), proteins that confer transcriptional activation of beta-catenin. Electrophoretic mobility shift and supershift assays revealed specific binding of human TCF-4 and beta-catenin to oligonucleotides corresponding to a potential TCF binding site in the versican promoter. In addition to binding assays, we directly assessed the dependence of versican promoter activity on TCF binding sites. Site-directed mutagenesis of the TCF site located -492 bp relative to the transcription start site markedly diminished versican-Luc activity. Co-transfection of TCF-4 with versican-Luc did not increase promoter activity, but addition of beta-catenin and TCF-4 significantly stimulated basal versican promoter activity. Our findings suggest that versican transcription is predominantly mediated by the GSK-3beta pathway via the beta-catenin-TCF transcription factor complex in smooth muscle cells, wherein such regulation contributes to the normal or aberrant formation of provisional matrix in vascular injury and repair events.

Animals↗
PubMed2005

Strategies of conditional gene expression in myocardium: an overview.

The use of specialized reporter genes to monitor real-time, tissue-specific transgene expression in animal models offers an opportunity to circumvent current limitations associated with the establishment of transgenic mouse models. The Cre-loxP and the tetracycline (Tet)-inducible systems are useful methods of conditional gene expression that allow spatial (cell-type-specific) and temporal (inducer-dependent) control. Most often, the alpha-myosin heavy chain (alpha-MHC) promoter is used in these inducible systems to restrict expression of reporter genes and transgenes to the myocardium. An overview of each inducible system is described, along with suggested reporter genes for real-time, noninvasive imaging in the myocardium. Effective gene delivery of the inducible gene expression system is carried out by lentiviral vectors, which offer high transduction efficiency, long-term transgene expression, and low immunogenicity. This chapter outlines the packaging of myocardium-specific inducible expression systems into lentiviral vectors, in which a transgene and a reporter gene are transduced into cardiomyocytes. In doing so, transgene and reporter expression can be monitored/tracked with bioluminescence imaging (BLI) and positron emission tomography (PET).

Animals↗
PubMed2005

Bone marrow-derived recipient cells in murine transplanted hearts: potential roles and the effect of immunosuppression.

Currently, there is intense debate regarding the origin of reparative cells in injured hearts and vasculature. To determine the contribution of recipient bone marrow (BM)-derived cells to the regeneration of cells in the vasculature of transplanted hearts and to examine the effect of immunosuppression on this phenomenon, we evaluated the fate of green fluorescent protein (GFP)-positive recipient BM cells in non-GFP-expressing cardiac allografts. C57BL/6 BM-GFP chimeric recipients underwent cardiac transplantation. Allografts were immunosuppressed with tacrolimus for 14 or 30 days post-transplantation or were saline treated. Hearts were excised and stained with markers for endothelial cells (EC) or smooth muscle cells (SMC). Colocalization with BM-derived recipient cells was evaluated using confocal microscopy with three-dimensional image analysis. Immunosuppression with tacrolimus did not affect the frequency of recipient BM-derived cell chimerism as EC or SMC phenotypes. A higher frequency of EC chimerism was found at 14 days as compared to 30 days post-transplantation in allograft hearts. BM-derived recipient cells are recruited to areas of donor vascular injury with intercalation of recipient EC and SMC in the setting of ongoing alloimmune recognition of the allograft. Our findings confirm that immunosuppression with tacrolimus does not affect the frequency of recipient BM-derived cell repopulation at an early time point 14 days post-transplantation. EC repopulation by BM-derived recipient cells was found to be an early event in transplanted allograft hearts, which decreased in frequency over time.

Animals↗
PubMed2005

Granzyme B induces endothelial cell apoptosis and contributes to the development of transplant vascular disease.

Endothelial cell death induced by cytotoxic T cells is a key initiating event in the development of transplant vascular disease (TVD), the leading cause of late solid organ transplant failure. We studied the role of the granzyme B (GrB) pathwaye, which is one of the main mechanisms by which T cells induce apoptosis of allogeneic targets, in the pathogenesis of TVD. Granzyme B, in combination with perforin (pfn), induced apoptosis of cultured endothelial cells. In hearts transplanted into GrB knockout (GrB-KO) mice, there was a similar level of vasculitis as compared to WT mice, indicating that GrB does not affect immune infiltration into allograft arteries. However, there was a significant reduction in luminal narrowing of allograft arteries from GrB-KO mice as compared to WT recipients. These results indicate that GrB plays a role in endothelial cell death in allograft arteries and in the resultant development of TVD.

Animals↗
PubMed2005

Pyrrolidine dithiocarbamate reduces coxsackievirus B3 replication through inhibition of the ubiquitin-proteasome pathway.

Coxsackievirus B3 (CVB3) is one of the most common pathogens for viral myocarditis. The lack of effective therapeutics for CVB3-caused viral diseases underscores the importance of searching for antiviral compounds. Pyrrolidine dithiocarbamate (PDTC) is an antioxidant and is recently reported to inhibit ubiquitin-proteasome-mediated proteolysis. Previous studies have shown that PDTC inhibits replication of rhinovirus, influenza virus, and poliovirus. In the present study, we report that PDTC is a potent inhibitor of CVB3. Coxsackievirus-infected HeLa cells treated with PDTC showed a significant reduction of CVB3 viral RNA synthesis, viral protein VP1 expression, and viral progeny release. Similar to previous observation that divalent ions mediate the function of PDTC, we further report that serum-containing copper and zinc are required for its antiviral activity. CVB3 infection resulted in massive generation of reactive oxygen species (ROS). Although PDTC alleviated ROS generation, the antiviral activity was unlikely dependent on its antioxidant effect because the potent antioxidant, N-acetyl-L-cysteine, failed to inhibit CVB3 replication. Consistent with previous reports that PDTC inhibits ubiquitin-proteasome-mediated protein degradation, we found that PDTC treatment led to the accumulation of several short-lived proteins in infected cells. We further provide evidence that the inhibitory effect of PDTC on protein degradation was not due to inhibition of proteasome activity but likely modulation of ubiquitination. Together with our previous findings that proteasome inhibition reduces CVB3 replication (H. Luo, J. Zhang, C. Cheung, A. Suarez, B. M. McManus, and D. Yang, Am. J. Pathol. 163:381-385, 2003), results in this study suggest a strong antiviral effect of PDTC on coxsackievirus, likely through inhibition of the ubiquitin-proteasome pathway.

Antioxidants↗
PubMed2005

Stress-activated protein kinases are involved in coxsackievirus B3 viral progeny release.

Stress-activated protein kinases (SAPKs), consisting of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38 MAPK), are activated upon various environmental stimuli, including viral infections. Cellular survival and death signaling events following coxsackievirus B3 (CVB3) infection have been studied in relationship to viral replication, but the role of SAPKs has not been scrutinized. In this study, we found that the phosphorylation of JNK1/2 and p38 MAPK was increased during active replication of CVB3 and that their phosphorylation was independent of CVB3-induced caspase activation or production of reactive oxygen species. The roles of these kinases in CVB3 infection were further evaluated using specific inhibitors: SP600125 for JNK1/2 and SB203580 for p38 MAPK. JNK1/2 inhibitors reduced CVB3-induced phosphorylation of activating transcription factor 2, and the p38 MAPK inhibitor reduced CVB3-induced phosphorylation of heat shock protein 27. Although inhibition of these kinases by specific inhibitors did not affect CVB3 viral protein synthesis, inhibition of p38 MAPK but not of JNK1/2 resulted in significant reduction of viral progeny release, suppression of CVB3-induced cell death, and blockage of CVB3-induced caspase-3 activation in infected cells. We conclude that SAPK pathways play critical roles in the life cycle of CVB3, particularly in viral progeny release.

Cell Survival↗
PubMed2005

Gamma interferon-inducible protein 10 induces HeLa cell apoptosis through a p53-dependent pathway initiated by suppression of human papillomavirus type 18 E6 and E7 expression.

Gamma interferon-inducible protein 10 (IP10) is a member of the CXC family of chemokines. By differential mRNA display, we have demonstrated the upregulation of IP10 in coxsackievirus B3 (CVB3)-infected mouse hearts. Functional characterization of the IP10 gene in IP10-transfected Tet-On HeLa cells has found that IP10 induced cell apoptosis and inhibited viral replication. In the characterization of the IP10-induced apoptotic pathway, we found that overexpression of IP10 upregulated p53 and resulted in altered expression of p53-responsive genes such as the p21Cip1, p27kip1, NF-kappaB, Bax, and PUMA genes and the mitochondrial translocation of Bax. However, transduction of the IP10 cells with adenovirus expressing dominant negative p53 not only ablated p53-triggered gene expression but also abolished IP10-induced apoptosis and restored CVB3 replication to the control levels. These data suggest a novel mechanism by which IP10 inhibits viral replication through the induction of host cell death via a p53-mediated apoptotic pathway. We also found that constantly high-level expression of p53 in these tumor cells is attributed to the IP10-induced suppression of human papillomavirus E6 and E7 oncogene expression. Taken together, these data reveal not only a previously unrecognized link between chemokine IP10 and p53 in antiviral defense but also a mechanism by which IP10 inhibits tumor cell growth.

Animals↗
PubMed2005

Methods for examining stem cells in post-ischemic and transplanted hearts.

Currently, the tenet that heart muscle cells are terminally differentiated and incapable of self-repair is being challenged. Recent experimental observations suggest that both endogenous and exogenous stem cell populations have the potential to regenerate damaged areas within the heart. These findings hold promise for new therapeutic strategies to treat cardiovascular diseases, including common conditions like myocardial infarction and transplant vascular disease (TVD). In this chapter, we focus on the study of endogenous stem cells in the context of their role in modulation of cardiovascular diseases, including ischemic heart disease and TVD. Specific experimental models and methods used to study the phenomena of endogenous bone marrow-derived stem cell migration and potential differentiation are also described.

Animals↗
PubMed2005

Affymetrix oligonucleotide analysis of gene expression in the injured heart.

Microarrays have helped researchers gain much insight into gene expression profiles in the context of many diseases including those in the injured heart. Our genomic investigations have been focused on elucidation of host gene responses to enterovirus infection. We have gained valuable technical expertise in using Affymetrix oligonucleotide arrays, also known as GeneChips, and cDNA spotted arrays to probe differential gene expression in both cultured cells and in heart tissue. Here, we provide a technique-focused supplement to the Affymetrix GeneChip Expression Analysis Manual for sample preparation, processing, and array hybridization. We provide expanded explanations to highlight important points within the existing protocol and offer variations to standard procedures when appropriate. For investigators using myocardial tissues for microarray experiments, we further address the necessity of and methods for in situ flushing of the vasculature, tissue homogenization, and considerations for limits of expression detection in rare cells. It is our intention to provide useful technical information, based on our experience, to assist those researchers using Affymetrix GeneChips in their own genomic research.

Animals↗
PubMed2005

Coxsackievirus-induced myocarditis: new trends in treatment.

Myocarditis is a common inflammatory heart disease in children and young adults that may result in chronically dilated cardiomyopathy. Coxsackievirus B3 is the major etiologic agent of this disease. Current treatments for patients with viral myocarditis are almost entirely supportive. In recent years, some promising therapeutic candidates have emerged, including novel treatments and improvements of existing drugs. Among these are molecules that specially target virus entry, such as pleconaril, WIN 54954 and CAR-Fc; nucleic acid-based antiviral agents that inhibit viral translation and/or transcription, such as antisense oligodeoxynucleotide and short interfering RNA; and immunomodulatory agents that augment the host-protective immune responses to effectively clear viruses from target tissues, including interferons and immunoglobulins. In addition, certain new antiviral strategies, still in the early stages, include modulation of signal transduction pathways responsible for viral replication using enzyme inhibitors, which have revealed potential therapeutic targets for viral myocarditis. Finally, the progress in cellular cardiomyoplasty for end-stage therapy, in particular the preliminary clinical trials, is also discussed with respect to its potential future application.

Antiviral Agents↗