Twinning: Coronary Artery Disease in Monozygotic Twins.

We present the case of a patient whose monozygotic twin brother suffered a fatal myocardial infarction at the age of 40. The patient presented with similar symptoms as his brother. Given the family history, ischemic evaluation was undertaken and revealed similar coronary anatomy and severe coronary artery disease (CAD). We review the current literature regarding genetic and environmental factors regarding coronary anatomy, locations of atherosclerotic lesions, and screening in twins.

Triple-acting Lytic Enzyme Treatment of Drug-Resistant and Intracellular Staphylococcus aureus.

Multi-drug resistant bacteria are a persistent problem in modern health care, food safety and animal health. There is a need for new antimicrobials to replace over used conventional antibiotics. Here we describe engineered triple-acting staphylolytic peptidoglycan hydrolases wherein three unique antimicrobial activities from two parental proteins are combined into a single fusion protein. This effectively reduces the incidence of resistant strain development. The fusion protein reduced colonization by Staphylococcus aureus in a rat nasal colonization model, surpassing the efficacy of either parental protein. Modification of a triple-acting lytic construct with a protein transduction domain significantly enhanced both biofilm eradication and the ability to kill intracellular S. aureus as demonstrated in cultured mammary epithelial cells and in a mouse model of staphylococcal mastitis. Interestingly, the protein transduction domain was not necessary for reducing the intracellular pathogens in cultured osteoblasts or in two mouse models of osteomyelitis, highlighting the vagaries of exactly how protein transduction domains facilitate protein uptake. Bacterial cell wall degrading enzyme antimicrobials can be engineered to enhance their value as potent therapeutics.

Clinical trial participation improves outcome: a matched historical cohort study.

BACKGROUND: Well-conducted, investigator-led randomized controlled trials (RCTs) are the gold standard for evaluating the efficacy of new treatments and are a key component of evidence-based medicine. It is unclear whether participating in an RCT is beneficial to the individual before the results of RCTs are known. PURPOSE: In a matched historical cohort study, we examined whether participation in RCTs was associated with improved health outcomes. METHODS: Participants in the Action in Diabetes and Vascular Disease: Preterax and Diamicron MR Controlled Evaluation (ADVANCE), Ongoing Telmisartan Alone and in Combination with Ramipril Global Endpoint Trial (ONTARGET), or Telmisartan Randomized Assessment Study in ACE-intolerant Subjects with Cardiovascular Disease (TRANSCEND) studies and non-participant controls were selected from patients attending outpatient clinics at Middlemore Hospital between 2001 and 2003. RESULTS: A total of 251 RCT participants and 502 randomly selected patients not enrolled in a trial but who met study entry criteria were matched for age, gender, and ethnicity. There was a significant difference in all-cause mortality for trial participants versus non-participants over the study period (unadjusted relative risk reduction (RRR) = 63%; 95% confidence interval (CI) = 28%-81%) and a significant reduction in cardiovascular mortality (unadjusted RRR = 81%; 95% CI = 17%-95%) favouring RCT participants. Allowing for co-morbidity, the adjusted RRR of all-cause mortality associated with trial participation was 55% (95% CI = 10%-77%). Active treatment in an RCT was found to be less explanatory than trial participation. The adjusted RRR for cardiovascular mortality associated with active treatment in a trial was 86% (95% CI = -2% to 98%), with trial participation found to be less explanatory than active treatment. LIMITATIONS: The main limitations of this trial relate to its design as a retrospective study with a historical cohort comparison group. Limitations include lack of complete data for some patients, bias in selection of the comparison group, and the effects of confounding variables. However, the study design and analysis were planned so as to minimize these as much as possible. CONCLUSION: This study revealed significantly lower all-cause mortality among participants in industry-sponsored RCTs compared with non-participants who received routine hospital outpatient care. This effect was independent of study drug.

Cleavage specificity of Enterococcus faecalis EnpA (EF1473), a peptidoglycan endopeptidase related to the LytM/lysostaphin family of metallopeptidases.

Enterococcus faecalis EnpA (EF1473) is a 1721-residue predicted protein encoded by prophage 03 that displays similarity to the staphylolytic glycyl-glycyl endopeptidases lysostaphin and LytM. We purified a catalytically active fragment of the protein, EnpA(C), comprising residues 1374-1505 and showed that the recombinant polypeptide efficiently cleaved cross-linked muropeptides generated by muramidases, but was poorly active in intact sacculi. Analysis of the products of digestion of purified dimers by mass spectrometry indicated that EnpA(C) cleaves the D-Ala-L-Ala bond formed by the D,D-transpeptidase activity of penicillin-binding proteins in the last cross-linking step of peptidoglycan synthesis. Synthetic D was identified as the minimum substrate of EnpA(C) indicating that interaction of the enzyme with the donor peptide stem of cross-linked dimers is sufficient for its activity. Peptidoglycan was purified from various bacterial species and digested with mutanolysin and EnpA(C) to assess enzyme specificity. EnpA(C) did not cleave direct cross-links, but tolerated extensive variation in cross-bridges with respect to both their length (one to five residues) and their amino acid sequence. Recognition of the donor stem of cross-linked dimers could account for the substrate specificity of EnpA(C), which is significantly broader in comparison to endopeptidases belonging to the lysostaphin family.

Epidemiological pattern of tattoo skin disease: a potential general health indicator for cetaceans.

The presence of tattoo skin disease (TSD) was examined in 1392 free-ranging and dead odontocetes comprising 17 species from the Americas, Europe, South Africa, New Zealand and Greenland. We investigated whether TSD prevalence varied with sex, age and health status. TSD was encountered in cetaceans from the Pacific and Atlantic Oceans as well as in those from the North, Mediterranean and Tasman Seas. No clear patterns related to geography and host phylogeny were detected, except that prevalence of TSD in juveniles and, in 2 species (dusky dolphin Lagenorhynchus obscurus and Burmeister's porpoise Phocoena spinipinnis), in adults was remarkably high in samples from Peru. Environmental factors and virus properties may be responsible for this finding. Sex did not significantly influence TSD prevalence except in the case of Peruvian P. spinipinnis. Generally, there was a pattern of TSD increase in juveniles compared to calves, attributed to the loss of maternal immunity. Also, in most samples, juveniles seemed to have a higher probability of suffering TSD than adults, presumably because more adults had acquired active immunity following infection. This holo-endemic pattern was inverted in poor health short-beaked common dolphins Delphinus delphis and harbour porpoises Phocoena phocoena from the British Isles, and in Chilean dolphins Cephalorhynchus eutropia from Patagonia, where adults showed a higher TSD prevalence than juveniles. Very large tattoos were seen in some adult odontocetes from the SE Pacific, NE Atlantic and Portugal's Sado Estuary, which suggest impaired immune response. The epidemiological pattern of TSD may be an indicator of cetacean population health.

LysK CHAP endopeptidase domain is required for lysis of live staphylococcal cells.

LysK is a staphylococcal bacteriophage endolysin composed of three domains: an N-terminal cysteine, histidine-dependent amidohydrolases/peptidases (CHAP) endopeptidase domain, a midprotein amidase 2 domain, and a C-terminal SH3b_5 (SH3b) cell wall-binding domain. Both catalytic domains are active on purified peptidoglycan by positive-ion electrospray ionization MS. The cut sites are identical to LytA (phi11 endolysin), with cleavage between d-alanine of the stem peptide and glycine of the cross-bridge peptide, and N-acetylmuramoyl-l-alanine amidase activity. Truncations of the LysK containing just the CHAP domain lyse Staphylococcus aureus cells in zymogram analysis, plate lysis, and turbidity reduction assays but have no detectable activity in a minimal inhibitory concentration (MIC) assay. In contrast, truncations harboring just the amidase lytic domain show faint activity in both the zymogram and turbidity reduction assays, but no detectable activity in either plate lysis or MIC assays. A fusion of the CHAP domain to the SH3b domain has near full-length LysK lytic activity, suggesting the need for a C-terminal binding domain. Both LysK and the CHAP-SH3b fusion were shown to lyse untreated S. aureus and the coagulase-negative strains. In the checkerboard assay, the CHAP-SH3b fusion achieves the same level of antimicrobial synergy with lysostaphin as the full-length LysK.

Effect of fenofibrate on amputation events in people with type 2 diabetes mellitus (FIELD study): a prespecified analysis of a randomised controlled trial.

BACKGROUND: Amputations in people with type 2 diabetes mellitus substantially impair their quality of life and impose high costs on health-care systems. Our aim was to assess the effect of fenofibrate on amputation events in a large cohort of patients with type 2 diabetes. METHODS: In the Fenofibrate Intervention and Event Lowering in Diabetes (FIELD) study, 9795 patients aged 50-75 years with type 2 diabetes were randomly assigned by computer-generated randomisation sequence to receive fenofibrate 200 mg per day (n=4895) or matching placebo (n=4900) for 5 years' duration. Information about non-traumatic amputation-a prespecified tertiary endpoint of the study-was routinely gathered. Clinicians who were masked to treatment allocation adjudicated amputations as minor or major (below or above the ankle, respectively). Amputations were also classified on the basis of whether or not large-vessel disease was present in the limb, to distinguish those related to large-artery atherosclerosis from those predominantly related to microvascular disease. Analysis was by intention to treat (ITT). The FIELD study is registered as an International Standard Randomised Controlled Trial, number ISRCTN64783481. FINDINGS: All 9795 patients were included in the ITT population. 115 patients had one or more non-traumatic lower-limb amputations due to diabetes. Previous cardiovascular disease, microvascular disease, previous non-traumatic amputation or skin ulcer, smoking, and longer duration of diabetes were more frequent in patients who had amputations during the trial than in those who had other cardiovascular events or in those who had neither event (all p<0.001 for three-way comparison). Mean lipid concentrations differed between patients who had on-study amputations and those who had other cardiovascular events or neither event, but by no more than 0.2 mmol/L. The risks of first amputation (45 vs 70 events; hazard ratio [HR] 0.64, 95% CI 0.44-0.94; p=0.02) and minor amputation events without known large-vessel disease (18 vs 34 events; 0.53, 0.30-0.94; p=0.027) were lower for patients assigned to fenofibrate than for patients assigned to placebo, with no difference between groups in risk of major amputations (24 vs 26 events; 0.93, 0.53-1.62; p=0.79). INTERPRETATION: Classic markers of macrovascular and microvascular risk were associated with lower extremity amputations in patients with type 2 diabetes. Treatment with fenofibrate was associated with a lower risk of amputations, particularly minor amputations without known large-vessel disease, probably through non-lipid mechanisms. These findings could lead to a change in standard treatment for the prevention of diabetes-related lower-limb amputations. FUNDING: Laboratoires Fournier SA (now part of Solvay Pharmaceuticals) and National Health and Medical Research Council of Australia.

Composition and proposed structure of a water-soluble glycan from the Keumsa Sangwhang Mushroom (Phellinus linteus).

Phellinus linteus is a mushroom which has been prized in Korea for its medicinal properties since ancient times. A glycan preparation from this source has shown promise in experimental anti-tumour and metastasis studies, but the glycan has not been well characterized. In the present work, the glycan fraction of a hot water extract from P. linteus (Keumsa Sangwhang Mushroom, Korea) has been isolated following ammonium sulfate precipitation, dialysis/concentration and anion exchange chromatographic steps. Analyses for monosaccharide composition showed glucose and mannose to be the major constituents. Digestion of the glycan fraction with specific glycosidases and by linkage analysis allowed us to propose a core beta(1-3) linked glucan heavily substituted via (1-6) links with beta(1-3) linked mannose chains. Significant levels of galactose and xylose, present in the glycan fraction, may be associated with this glucomannan or not. These findings are consistent with the view that a core beta(1-3)-linked glucan chain with beta(1-6) branch points is a common feature of mushroom glycans possessing anti-tumour activity.

LambdaSa1 and LambdaSa2 prophage lysins of Streptococcus agalactiae.

Putative N-acetylmuramyl-l-alanine amidase genes from LambdaSa1 and LambdaSa2 prophages of Streptococcus agalactiae were cloned and expressed in Escherichia coli. The purified enzymes lysed the cell walls of Streptococcus agalactiae, Streptococcus pneumoniae, and Staphylococcus aureus. The peptidoglycan digestion products in the cell wall lysates were not consistent with amidase activity. Instead, the structure of the muropeptide digestion fragments indicated that both the LambdaSa1 and LambdaSa2 lysins exhibited gamma-d-glutaminyl-l-lysine endopeptidase activity. The endopeptidase cleavage specificity of the lysins was confirmed using a synthetic peptide substrate corresponding to a portion of the stem peptide and cross bridge of Streptococcus agalactiae peptidoglycan. The LambdaSa2 lysin also displayed beta-d-N-acetylglucosaminidase activity.

Modulation of glycosaminoglycan levels in tree shrew sclera during lens-induced myopia development and recovery.

PURPOSE: In juvenile tree shrews, positioning a negative-power lens in front of an eye produces a hyperopic shift in refractive state and causes a compensatory increase in axial length over several days so that the eye is myopic when the lens is removed. During negative lens compensation, the scleral extracellular matrix is remodeled. A biomechanical property of the sclera, creep rate, increases; during recovery from induced myopia, the creep rate decreases below normal levels. Changes in glycosaminoglycan (GAG) levels, including those of hyaluronan, may participate in these changes in creep rate and, in turn, participate in controlling the axial length and refractive state. This study investigated the unsulfated and sulfated GAG composition of the sclera during compensation for a -5 diopter (D) lens and during recovery. METHODS: Capillary electrophoresis was used to assess the relative levels (ng/mg dry scleral weight) of unsulfated GAGs (hyaluronan [HA] and chondroitin [C0S]), sulfated GAGs (chondroitin-4-sulfate [C4S], chondroitin-6-sulfate [C6S], and dermatan sulfate [DS]) in the sclera of groups of tree shrews (n = 5 per group) that wore a monocular -5 D lens for 1, 2, 4, or 11 days or had 11 days of -5 D lens wear followed by 1, 2, or 4 days of recovery from lens wear. The fellow eye served as an untreated control. Groups of normal and plano lens-treated animals provided age-matched values. RESULTS: Expressed as a fraction of dry weight, levels of HA were lower after 1, 4, and 11 days of -5 D lens wear. Levels of C0S, C6S, and C4S were significantly lower after 4 and 11 days of lens wear. After 1 and 2 days of recovery, GAG levels in the treated eyes were not significantly different from those in control eyes. After 4 recovery days, HA levels were lower, but the levels of all other GAGs were not different in the recovering and control eyes. Some binocular changes also occurred. CONCLUSIONS: The rapid differential decrease in HA levels during negative lens compensation and the absence of any difference after just 1 day of recovery suggest that HA levels may play a previously unrecognized early role in regulating the biomechanical property (creep rate) of the sclera. The reduced levels of the other GAGs, which occur when creep rate is at its peak elevation, and their rapid return to normal after 1 day of recovery suggest that they may also participate in regulating this biomechanical property of the sclera.

Endopeptidase and glycosidase activities of the bacteriophage B30 lysin.

Synthetic peptides corresponding to portions of group B streptococcal peptidoglycan were used to show that the endopeptidase activity of bacteriophage B30 lysin cleaves between D-Ala in the stem peptide and L-Ala in the cross bridge and that the minimal peptide sequence cleaved is DL-gamma-Glu-Lys-D-Ala-Ala-Ala. The only glycosidase activity present is that of N-acetyl-beta-D-muramidase.

Psychedelic sacraments.

The use of psychedelic substances within a context that emphasizes religious experiences and aims to provide spiritual insights is not a new phenomenon. However, the proscription of these substances in most modern societies leads to such use now typically occurring in an underground and idiosyncratic manner that often leaves individuals on their own with regard to the interpretation and integration of their experiences and insights. In contrast, the numerous examples in the ethnographic and the historical literature indicate that many cultures independently developed similar frameworks for using these substances for both individually and socially beneficial purposes and arrived at similar conclusions as to which of the substances available to them were the most appropriate for these purposes. This article focuses on a special type of socially sanctioned framework called a "sacrament" and contrasts this with other, more idiosyncratic forms of psychedelic use. It discusses how this framework helps to structure and channel the experiences induced by these substances, thereby increasing the likelihood of individually constructive and socially integrative experiences.

Demonstration of a hyperglycemia-driven pathogenic abnormality of copper homeostasis in diabetes and its reversibility by selective chelation: quantitative comparisons between the biology of copper and eight other nutritionally essential elements in normal and diabetic individuals.

We recently showed that treatment with the Cu(II)-selective chelator, trientine, alleviates heart failure in diabetic rats, improves left ventricular hypertrophy in humans with type 2 diabetes, and increases urinary Cu excretion in both diabetic rats and humans compared with nondiabetic control subjects. In this study, we characterized the homeostasis of Cu and eight other nutritionally essential elements in diabetes under fully residential conditions in male subjects with type 2 diabetes and age-matched control subjects. We then probed elemental balance with oral trientine in a parallel-group, placebo-controlled study in these subjects. Before treatment, there were no detectable between-group differences in the balance of any element, although urinary output of several elements was greater in diabetic subjects. Mean extracellular superoxide dismutase (EC-SOD) activity was elevated in diabetic subjects, and its activity correlated strongly with the interaction between [Cu]serum and HbA1c. Trientine caused the Cu balance to become negative in diabetic subjects through elevated urinary Cu losses and suppressed elevated EC-SOD. Basal urinary Cu predicted urinary Cu losses during treatment, which caused extraction of systemic Cu(II). We suggest that cardiovascular complications in diabetes might be better controlled by therapeutic strategies that focus on lowering plasma glucose and loosely bound systemic Cu(II).

Relationships between polychlorinated biphenyls and health status in harbor porpoises (Phocoena phocoena) stranded in the United Kingdom.

To investigate possible relationships between polychlorinated biphenyl (PCB) exposure and infectious disease mortality in harbor porpoises (Phocoena phocoena) in United Kingdom waters, summed blubber concentrations of 25 chlorobiphenyl congeners (sigma25CB) in healthy harbor porpoises that died of acute physical trauma (mainly by-catch; n = 175) were compared with sigma25CB values in animals that died of infectious disease (n = 82). The infectious disease group had significantly greater sigma25CB values (mean, 27.6 mg/kg lipid) than the physical trauma group (mean, 13.6 mg/kg lipid; p < 0.001). This association occurred independently of other potentially confounding variables, including age, sex, two indices of nutritional status, season, region, and year found. Total blubber PCB levels (as Aroclor 1254) were also calculated, enabling direct comparison with a proposed threshold for adverse health effects (including immunosuppression) in marine mammals of 17 mg/kg lipid. In porpoises with total PCB levels greater than 17 mg/kg lipid (n = 154), total PCB levels were significantly higher in the infectious disease group compared to the physical trauma group (p < 0.001). This association was no longer significant in porpoises with total PCB levels of less than 17 mg/kg lipid (n = 103; p > 0.55). These findings are consistent with a causal (immunotoxic) relationship between PCB exposure and infectious disease mortality, and they provide a framework for future quantitative risk-assessment analyses of porpoise populations of known size and PCB exposure.

Regeneration of the heart in diabetes by selective copper chelation.

Heart disease is the major cause of death in diabetes, a disorder characterized by chronic hyperglycemia and cardiovascular complications. Although altered systemic regulation of transition metals in diabetes has been the subject of previous investigation, it is not known whether changed transition metal metabolism results in heart disease in common forms of diabetes and whether metal chelation can reverse the condition. We found that administration of the Cu-selective transition metal chelator trientine to rats with streptozotocin-induced diabetes caused increased urinary Cu excretion compared with matched controls. A Cu(II)-trientine complex was demonstrated in the urine of treated rats. In diabetic animals with established heart failure, we show here for the first time that 7 weeks of oral trientine therapy significantly alleviated heart failure without lowering blood glucose, substantially improved cardiomyocyte structure, and reversed elevations in left ventricular collagen and beta(1) integrin. Oral trientine treatment also caused elevated Cu excretion in humans with type 2 diabetes, in whom 6 months of treatment caused elevated left ventricular mass to decline significantly toward normal. These data implicate accumulation of elevated loosely bound Cu in the mechanism of cardiac damage in diabetes and support the use of selective Cu chelation in the treatment of this condition.

The bifunctional peptidoglycan lysin of Streptococcus agalactiae bacteriophage B30.

A group B streptococcal (GBS) bacteriophage lysin gene was cloned and expressed in Escherichia coli. The purified recombinant enzyme, calculated to have a molecular mass of 49 677 Da, lysed GBS cells. The susceptibility of GBS cells to lysis by the enzyme depended upon the growth stage at which they were harvested, with early exponential phase cells most sensitive. Calcium ions enhanced the activity of the enzyme. The enzyme also lysed other beta-haemolytic streptococci, including groups A, C, E and G streptococci, but not common oral streptococci, including Streptococcus mutans. The generation of both reducing activity and N-terminal alanine residues during lysis indicated that the lysin is a bifunctional enzyme, possessing both glycosidase and endopeptidase activities. This is consistent with the presence of two conserved sequence domains, an Acm (acetylmuramidase) domain associated with lysozyme activity, and a CHAP (cysteine, histidine-dependent amidohydrolases/peptidases) domain associated with endopeptidase activity. Site-directed mutagenesis of conserved cysteine and histidine residues in the CHAP domain and conserved aspartate and glutamate residues in the Acm domain confirmed their importance for lysozyme and endopeptidase activity respectively.

Achieving normothermia in patients with febrile subarachnoid hemorrhage: feasibility and safety of a novel intravascular cooling catheter.

INTRODUCTION: Fever is common and difficult to control in patients with subarachnoid hemorrhage (SAH). We have previously shown an inverse relationship between fever and outcome in patients with SAH. MATERIALS/METHODS: This was a prospective, single-arm, feasibility trial in which nine patients with SAH underwent temperature management using an intravascular cooling catheter (ICC) to restore and maintain 24 hours of normothermia (36.5 degrees+/-0.2 degrees C). Enrollment occurred after development of a fever of at least 38.3 degrees C within 7 days of SAH that was refractory to acetaminophen treatment. The ICC was placed at the bedside through an introducer sheath via the femoral vein into the inferior vena cava (IVC). Portable X-ray confirmed placement. RESULTS: Normothermia was achieved in seven of the nine patients treated (78%); it was achieved in 100% of the patients with a 14F catheter (n=4) and in 60% of the patients with a 9F catheter (n=5). The two patients not reaching normothermia were not adequately treated for shivering. All other patients reached normothermia irrespective of intubation status. Overall, normothermia was well tolerated and not discontinued because of discomfort or adverse events. Two incidences of deep vein thrombosis were diagnosis by ultrasound that were not associated with clinical sequelae, and IVC filters were placed. No unanticipated adverse events occurred. DISCUSSION: We have demonstrated that fever can be safely and effectively controlled in patients with SAH for at least 24 hours using an ICC. Future studies are needed to assess the effect of such sustained therapy on outcome in patients with SAH.

Hallucinogens and redemption.

This article examines drug substitution with regard to hallucinogens (ayahuasca, ibogaine, peyote and LSD) set within the concept of redemption. The model examines both religious and secular approaches to the contemporary use of hallucinogens in drug substitution, both by scientists and in religious settings worldwide. The redemptive model posits that the proper use of one psychoactive substance within a spiritual or clinical context helps to free an individual from the adverse effects of their addiction to another substance and thus restores them as functioning members of their community or group. Data is drawn from the U.S., Brazil, Peru, and West Africa. Two principle mechanisms for this are proposed: the psychological mechanism of suggestibility is examined in terms of the individual reaching abstinence goals from addictive substances such as alcohol and opiates. Neurophysiological and neurochemical mechanisms to understand the efficacy of such substitution are highlighted from ongoing research on hallucinogens. Research by two of the authors with the Uñaio do Vegetal (UDV) Church in Brazil is examined in terms of the model.

The hyaluronan lyase of Streptococcus pyogenes bacteriophage H4489A.

Many pathogenic streptococci produce extracellular hyaluronan lyases which are thought to aid the spread of the organism in host tissues. In addition, several phages of group A streptococci are known to synthesize a bound form of hyaluronidase. It has been suggested that the function of this hyaluronidase is to facilitate penetration of the hyaluronan capsule by phage and thus to gain access for the phage to the cell surface of the host streptococcus [Hynes, Hancock and Ferretti (1995) Infect. Immun. 63, 3015-3020]. In the present work, the hyaluronidase of Streptococcus pyogenes bacteriophage H4489A, expressed in E. coli, has been purified and characterized. The enzyme was shown to be a lyase with a distributive action pathway. Unlike most bacterial hyaluronidases that have been characterized, the phage enzyme was found to specifically cleave hyaluronan, which adds credence to the view that its function is to digest the hyaluronan capsule of the host organism. This bacteriophage lyase may provide a practical alternative to the lyase from Streptomyces hyalurolyticus as a reagent for the specific cleavage of hyaluronan.