PubMed Health⌕ Search

Biomedical subjects

B M McManus

Publications and source records attributed to B M McManus.

At least 55 records · Page 3Linked to original sources
PubMed1998

Rapid cytochrome c release, activation of caspases 3, 6, 7 and 8 followed by Bap31 cleavage in HeLa cells treated with photodynamic therapy.

Photodynamic therapy (PDT) is a clinical approach that utilizes light-activated drugs for the treatment of a variety of pathologic conditions. The initiating events of PDT-induced apoptosis are poorly defined. It has been shown for other proapoptotic stimuli that the integral endoplasmic reticulum protein Bap31 is cleaved by caspases 1 and 8, but not by caspase-3. Further, a 20 kDa Bap31 cleavage fragment is generated which can induce apoptosis. In the current report, we sought to determine whether Bap31 cleavage and generation of p20 is an early event in PDT-induced apoptosis. The mitochondrial release of cytochrome c, involvement of caspases 1, 2, 3, 4, 6, 7, 8, and 10 and the status of several known caspase substrates, including Bap31, were evaluated in PDT-treated HeLa cells. Cytochrome c appeared in the cytosol immediately following light activation of the photosensitizer benzoporphyrin derivative monoacid ring A. Activation of caspases 3, 6, 7, and 8 was evident within 1-2 h post PDT. Processing of caspases 1, 2, 4, and 10 was not observed. Cleavage of Bap31 was observed at 2-3 h post PDT. The caspase-3 inhibitor DEVD-fmk blocked caspase-8 and Bap31 cleavage suggesting that caspase-8 and Bap31 processing occur downstream of caspase-3 activation in PDT-induced apoptosis. These results demonstrate that release of mitochondrial cytochrome c into the cytoplasm is a primary event following PDT, preceding caspase activation and cleavage of Bap31. To our knowledge, this is the first example of a chemotherapeutic agent inducing caspase-8 activation and demonstrates that caspase-8 activation can occur after cytochrome c release.

Amino Acid Chloromethyl Ketones↗
PubMed1998

Overexpression of Bcl-X(L) prevents caspase-3-mediated activation of DNA fragmentation factor (DFF) produced by treatment with the photochemotherapeutic agent BPD-MA.

Photodynamic therapy (PDT) is a clinically effective cancer treatment. For human promyelocytic leukemia HL-60 cells, cleavage of pro-caspase-3 (CPP32/Yama/apopain) into its proteolytically active subunits rapidly follows the photodynamic treatment of these cells with cytotoxic levels of the photosensitizer benzoporphyrin derivative monoacid ring A and visible light. Cleavage of a recently identified cytosolic 45 kDa protein, DNA fragmentation factor (DFF), is required for endonuclease activation leading to DNA fragmentation. In the present study, DFF was rapidly processed following PDT. Overexpression of the anti-apoptotic Bcl-X(L) gene in HL-60 cells prevented PDT-induced caspase activation, DFF cleavage and DNA fragmentation. These results demonstrate for the first time an example of chemotherapeutic drug-induced activation of DFF and its regulation by Bcl-X(L).

Apoptosis↗
PubMed1998

Induction of virus-induced IDDM in virus resistant mice without lymphocyte maturation.

The role of lymphocytes in the pathogenesis of viral-induced insulin dependent diabetes mellitus (IDDM) is controversial. To better understand how a virus-induced IDDM depends on the infiltrating lymphocytes, encephalomyocarditis virus (EMCV) was inoculated intraperitoneally into three kinds of mice; virus-susceptible C57BL/6, virus-resistant 129/SV and recombination activity gene-2 (Rag2) knockout 129/SV mice. Pancreatic inflammation and beta cell necrosis were evaluated after EMCV, D variant (10(3) pfu/mouse) inoculation. On post-inoculation day 14, the lethal rates of C57BL/6, 129/SV and Rag2 knockout mice were 52, 10 and 100%, respectively. The blood glucose in Rag2KO mice on day 8 was significantly elevated as compared with 129SV mice (231 +/- 49 vs 169 +/- 32 mg/dl, P<0.05). In situ hybridization demonstrated the EMCV genome in the pancreas of Rag2 knockout and C57BL/6 mice, but not in 129/SV mice. Beta cell necrosis were more severe in Rag-2 knockout mice than in wild type 129/SV mice, but lymphocyte infiltration was less severe than C57BL/6. Pancreas in Rag2 knockout mice infected with virus were affected more severely than the virus-resistant strain of mice. Diabetogenic virus induced IDDM in virus-resistant mice without mature lymphocytes.

Animals↗
PubMed1998

Interaction of viral proteins with host cell death machinery.

In recent years, intense research has been directed towards understanding molecular mechanisms involved in viral pathogenesis. It is now known that many viruses manipulate host defense mechanisms to prevent apoptosis in order to maximize viral replication. Towards the end of their replication cycle, certain viruses direct the synthesis of proteins that induce apoptosis or cell lysis thereby facilitating viral release from the cell. The present review summarizes the current understanding of interactions between viral proteins and the host cell death machinery.

Animals↗
PubMed1998

Endothelium-smooth muscle interaction in cardiac transplantation.

1. The inducible isoform of nitric oxide synthase (iNOS) is expressed in human and experimental cardiac allografts and is localized to infiltrating macrophages, cardiac myocytes, endothelial cells and smooth muscle cells. A recent clinical report proposes a causal link between myocardial expression of iNOS and ventricular contractile dysfunction, a potentially graft- and life-threatening post-transplant complication. 2. Coronary blood flow is elevated in human graft recipients with biopsy proven cellular rejection, indicating that vasodilation accompanies graft rejection. In Lewis-to-F-344 coronary resistance vessels, which show intimal expression of iNOS, pressure-induced myogenic tone is significantly inhibited. Selective iNOS inhibition partially reverses the inhibition of myogenic tone, confirming that iNOS produces vasoactive nitric oxide (NO) and may mediate the rejection-induced vasodilation seen clinically. 3. Endothelial dysfunction, identified as loss of endothelium-dependent dilation, has tremendous prognostic significance in vascular diseases of multiple aetiologies. In transplantation, endothelial dysfunction predicts early cardiac allograft vasculopathy and poor clinical outcome. Lewis-to-F-344 coronary vessels develop endothelial dysfunction at 1 week post-transplantation, but this is preceded by a transient state of endothelial cell hyperfunction, with enhanced endothelial production of NO. 4. The normal interaction between endothelial and smooth muscle cells in coronary resistance vessels is critical for the regulation of coronary blood flow and the maintenance of fluid homeostasis. With allospecific expression of iNOS, the inhibition of vascular tone predicts greatly enhanced intravascular pressure in precapillary arterioles and capillaries; this would be expected to cause a net movement of fluid from the intravascular compartment into the myocardial interstitium, resulting in ventricular oedema, non-compliance and poor contractile performance.

Cell Survival↗
PubMed1998

Caspase activation and specific cleavage of substrates after coxsackievirus B3-induced cytopathic effect in HeLa cells.

Coxsackievirus B3 (CVB3), an enterovirus in the family Picornaviridae, induces cytopathic changes in cell culture systems and directly injures multiple susceptible organs and tissues in vivo, including the myocardium, early after infection. Biochemical analysis of the cell death pathway in CVB3-infected HeLa cells demonstrated that the 32-kDa proform of caspase 3 is cleaved subsequent to the degenerative morphological changes seen in infected HeLa cells. Caspase activation assays confirm that the cleaved caspase 3 is proteolytically active. The caspase 3 substrates poly(ADP-ribose) polymerase, a DNA repair enzyme, and DNA fragmentation factor, a cytoplasmic inhibitor of an endonuclease responsible for DNA fragmentation, were degraded at 9 h following infection, yielding their characteristic cleavage fragments. Inhibition of caspase activation by benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (ZVAD.fmk) did not inhibit the virus-induced cytopathic effect, while inhibition of caspase activation by ZVAD.fmk in control apoptotic cells induced by treatment with the porphyrin photosensitizer benzoporphyrin derivative monoacid ring A and visible light inhibited the apoptotic phenotype. Caspase activation and cleavage of substrates may not be responsible for the characteristic cytopathic effect produced by picornavirus infection yet may be related to late-stage alterations of cellular homeostatic processes and structural integrity.

Amino Acid Chloromethyl Ketones↗
PubMed1998

Impaired expression of atrial natriuretic peptide in diabetic rats with myocardial infarction.

We evaluated the effects of myocardial infarction (MI) on the hemodynamics and the expression of atrial natriuretic peptide (ANP) mRNA in rats with streptozotocin-induced diabetes. Eight weeks after streptozotocin injection, the diabetic rats and age-matched nondiabetic controls underwent coronary artery ligation. One week later, the left ventricular end-diastolic pressure, systolic blood pressure, infarct size, and serum ANP levels did not differ significantly between the diabetic and nondiabetic rats. Compared with control animals without MI, the atrial ANP/beta-actin mRNA ratio in rats with MI was increased to 195% in diabetic animals and 213% in nondiabetic animals. In the left ventricle, however, the ANP/beta-actin mRNA ratio in diabetic animals with MI was increased to only 131% compared with control animals, whereas the corresponding increase in nondiabetic animals was 240% (p<0.05). Thus, the modulation of ANP mRNA expression after MI was impaired in the left ventricle, but not in the atria, of diabetic rats. A reduced myocardial expression of ANP could increase the morbidity and mortality associated with cardiovascular disorders in patients with diabetes.

Animals↗
PubMed1997

Homocyst(e)ine and coronary artery disease. Clinical evidence and genetic and metabolic background.

Many studies have demonstrated a strong association between elevated plasma total homocyst(e)ine levels and vascular diseases. Consequently, hyperhomocyst(e)inemia is now generally accepted as an independent risk factor for coronary artery disease. We critically reviewed the results of 35 human studies in which the levels of plasma total homocysteine were measured in patients with atherosclerotic diseases (n = 4338) and in controls (n = 22,593). Total homocysteine levels were consistently higher in patients than in controls. The average of this increment among 23 case-control studies was 26%. New insights into the biochemical pathways of total homocysteine metabolism, the factors that influence total homocysteine levels, genetic contributions to hyperhomocyst(e)inemia, the pathogenesis of homocyst(e)ine-induced vascular damage, and current recommendations for treatment of hyperhomocyst(e)inemia were also reviewed. Various lines of evidence now link hyperhomocyst(e)inemia with vascular diseases. Although there are no data from double-blind, placebo-controlled clinical trials of treatment for hyperhomocyst(e)inemia, the strong epidemiologic and experimental evidence argues for treatment of hyperhomocyst(e)inemia; in fact, its treatment with low doses of vitamins is thought to be safe and is inexpensive.

Clinical Medicine↗
PubMed1997

In vitro mutational and inhibitory analysis of the cis-acting translational elements within the 5' untranslated region of coxsackievirus B3: potential targets for antiviral action of antisense oligomers.

The 5' untranslated region (5'UTR) of coxsackievirus B3 (CVB3) RNA forms a highly ordered secondary structure that has been implicated in controlling initiation of viral translation by internal ribosomal entry. To test this hypothesis, synthetic bicistronic RNAs, with all or part of the 5'UTR in the intercistronic space, were translated in rabbit reticulocyte lysates. In the presence of an upstream cistron, the chloramphenicol acetyltransferase gene, designed to block ribosomal scanning, the CVB3 5'UTR was capable of directing the internal initiation of translation of the downstream reporter gene (P1), confirming the presence of an internal ribosomal entry site (IRES). This finding was further supported by the data on predicted secondary structures within the 5'UTR. Of special note, analysis of various deletion mutants demonstrated that the IRES of CVB3 is located roughly at stem-loops G, H, and I spanning nucleotides (nt) 529 and 630. The region from nt 1 to 63 (stem-loop A) also appears important, and it may be an essential binding site for translation initiation factors. Based on these findings, in vitro translation inhibition assays using RNA fragments of the 5'UTR as inhibitor were performed. Both antisense and sense RNA segments transcribed from these two cis-acting regions and the surrounding sequence of the initiation codon AUG showed strong inhibition of viral protein synthesis. Antisense molecules may inhibit translation by blocking ribosome and initiation factor binding within the 5'UTR via specific hybridization to their viral RNA target sequences, while sense sequences may function by competing with viral RNA for ribosomes and/or translation initiation factors. These cis-acting translational elements may serve as potential targets for the antiviral action of oligomers.

Animals↗
PubMed1997

Human leukocyte antigen class II associations in patients with idiopathic dilated cardiomyopathy. Myocarditis Treatment Trial Investigators.

BACKGROUND: Idiopathic dilated cardiomyopathy (IDC) is a disease of unknown etiology for which immune abnormalities, possibly related to viral infections, are suspected but unproven. Previous serologic studies have reported associations between human leukocyte antigen DR4 and IDC. A molecular study of human leukocyte antigen associations was undertaken in patients with IDC to further explore the possibility of susceptibility markers of genetically determined disease. METHODS AND RESULTS: In this study, 36 patients from the Myocarditis Treatment Trial (32 IDC and 4 myocarditis patients) were examined using restriction fragment length polymorphism analysis and polymerase chain reaction amplification with sequence-specific primers to perform class II typing. All 4 myocarditis patients were DQ5 positive and 3 possessed the allele DQB1*0501. In the IDC group, the frequency of human leukocyte antigen DR4 was similar to that reported in the normal population. In addition, there was no excess prevalence of any molecularly defined DR4 alleles (0401-0419). There was an increase in the frequency of DR12 in IDC patients. The frequencies of the alleles DQB1 *0503 and DQB1*0301 and/or *0304 were also increased in IDC patients versus the normal population. CONCLUSION: The molecular studies point to a relationship between the DQ locus and IDC.

Adult↗
PubMed1997

Myocarditis as systemic disease: new perspectives on pathogenesis.

1. Myocarditis may be an early indicator of or may subsequently lead to dilated cardiomyopathy in humans. This hypothesis has evolved from research on viruses that induce myocarditis, wherein the coxsackie B group viruses (CVB) in the family Picornaviridae are the most common known viral infectants of heart muscle. 2. Many competing hypotheses exist as to the pathogenesis of CVB3-induced myocarditis, including direct virus-induced myocyte damage and immunopathological disease with autoimmune sequelae. Evidence to support the direct-damage and viral RNA-persistence hypothesis is derived from in situ hybridization and gene amplification studies. 3. Recent use of terminal deoxynucleotidyl transferase-mediated nick-end labelling indicates that this injury in target organs is largely non-apoptotic in nature. Most apoptotic bodies in cardiac tissue are derived from immune cells. 4. Beyond infection of heart muscle, CVB3 can also associate with, infect and persist in cells of immune origin. The CVB3 localizes to follicles in spleens and lymph nodes of the murine host and this particular localization may continue in mice susceptible to more aggressive myocarditis. Whether virus-immune cell association in these compartments is advantageous (or essential) to the host in the evolution of anti-viral immune responses or whether it is more advantageous to the virus in immunosuppression of the host is not known. 5. We suggest that CVB3 can directly perturb or alter the immune response, thereby delaying viral clearance from vulnerable systemic organs. Both host and viral genetic factors can influence susceptibility, persistence and disease progression. 6. Picornaviruses use a unique method for the initiation of translation, involving the internal binding of the ribosome on a sequence element of the 5' untranslated region, termed an internal ribosome entry site (IRES). 7. The IRES of CVB3 is located at approximately stem loops G, H and I, spanning nucloetides 530 and 630. Arrest of host translation is also a feature of picornavirus infection. Such regulation of host cell translation machinery no doubt fosters viral replication at the expense of the host cell. 8. Differences between cell types in the mechanisms, along with those at other key steps in the viral life cycle and in signalling via kinase pathways, may determine viral tropism and cellular destruction and the physiological outcome of neighbouring cells.

Animals↗
PubMed1997

Complement component 3 interactions with coxsackievirus B3 capsid proteins: innate immunity and the rapid formation of splenic antiviral germinal centers.

Innate immunity is central to the clearance of pathogens from hosts as well as to the definition of acquired immune responses (D. T. Fearon, and R. M. Locksley, Science 272:50-53, 1996). Coxsackievirus B3 (CVB3), a human cardiopathic virus, was evaluated for the ability to activate the alternative and classical pathway of complement. CVB3 proteins interact with complement component 3 (C3, a soluble protein effector of innate immunity) after either in vitro exposure to mouse serum or in vivo murine infection and activate the alternative pathway of complement. In addition, we demonstrate that viral antigen retention and localization in germinal centers is dependent on C3, while virus antigen retention in extrafollicular regions in the spleen is not. In vivo depletion of native C3 abolished the rapid formation of virus-specific germinal centers (by day 3 post-CVB3 infection) in the absence of serum anti-CVB3 antibodies. These studies demonstrate that innate immune mechanisms, such as C3 interaction with CVB3, are essential for splenic antiviral germinal center formation in naive (antigen nonsensitized) mice resistant (C57BL/6J strain) and susceptible (A/J strain) to CVB3-induced myocarditis.

Animals↗
PubMed1997

"Tall oil"-derived phytosterols reduce atherosclerosis in ApoE-deficient mice.

We investigated the effects of a "tall oil"-derived phytosterol mixture (TODPM) on the formation of atherosclerotic lesions in apoE-deficient mice. TODPM was added at 2% (wt/wt) to the chow of nine mice; the control group had six animals. The diet of all animals contained 9% (wt/wt) fat and 0.15% (wt/wt) cholesterol. After 4 weeks, plasma total cholesterol levels were significantly reduced in the TODPM-treated mice (26.6 versus 42.0 mmol/L, P < .0001). The mean body weight of the TODPM-supplemented group was significantly higher at week 5 and throughout the study (29.4 versus 27.7 g, P < .05). The experiment was terminated at 18 weeks. Histological examination showed mature atherosclerotic lesions composed of foam cells underlying the endothelium, a mosaic of extracellular glycosaminoglycans, numerous apparently proliferative smooth muscle cells, and foci of cholesterol clefts in the control animals. By contrast, the TODPM-treated mice showed only early lesions containing mainly superficial foam cells. As assessed by morphometry, the lesion area in the aortic sinuses of TODPM-treated animals was less than half that of control animals (P < .0001). This reduced lesion area was accompanied by a substantial reduction in all lesional components, reflecting a delay in progression of atheromatous changes. A strong positive correlation (r = .69, P < .01) was found between plasma total cholesterol levels and lesion area in the aortic sinuses. TODPM also prevented the occurrence of xanthomatosis. We conclude that supplementation of a cholesterol-enriched diet with TODPM significantly lowers plasma cholesterol and retards development of atherosclerosis in apoE-deficient mice, suggesting a therapeutic potential for the mixture of phytosterols studied.

Animals↗
PubMed1997

Correction of hypertriglyceridemia and impaired fat tolerance in lipoprotein lipase-deficient mice by adenovirus-mediated expression of human lipoprotein lipase.

Humans homozygous or heterozygous for mutations in the lipoprotein lipase (LPL) gene demonstrate significant disturbances in plasma lipoproteins, including raised triglyceride (TG) and reduced HDL cholesterol levels. In this study we explored the feasibility of adenovirus-mediated gene replacement therapy for LPL deficiency. A total of 5 x 10(9) plaque-forming units (pfu) of an E1/E3-deleted adenovirus expressing either human LPL (Ad-LPL) or the bacterial beta-galactosidase gene (Ad-LacZ) as a control were administered to mice heterozygous for targeted disruption in the LPL gene (n = 57). Peak expression of total postheparin plasma LPL activity was observed at day 7 in Ad-LPL mice versus Ad-LacZ controls (834 +/- 133 vs 313 +/- 89 mU/mL, P < .01), and correlated with human-specific LPL activity (522 +/- 219 mU/mL) and mass (9214 +/- 782 ng/mL), a change that was significant to 14 and 42 days, respectively. At day 7, plasma TGs were significantly reduced relative to Ad-LacZ mice (0.17 +/- 0.07 vs 1.90 +/- 0.89 mmol/L, P < .01) but returned to endogenous levels by day 42. Ectopic liver expression of human LPL was confirmed by in situ hybridization analysis and from raised LPL activity and mass in liver homogenates. Analysis of plasma lipoprotein composition revealed a marked decrease in VLDL-derived TGs. Severely impaired oral and intravenous fat-load tolerance in LPL-deficient mice was subsequently corrected after Ad-LPL administration and closely paralleled that observed in wild-type mice. These findings suggest that liver-targeted adenovirus-mediated LPL gene transfer offers an effective means for transient correction of altered lipoprotein metabolism and impaired fat tolerance due to LPL deficiency.

Adenoviridae↗