Triglyceride High-Density Lipoprotein Ratios Predict Glycemia-Lowering in Response to Insulin Sensitizing Drugs in Type 2 Diabetes: A Post Hoc Analysis of the BARI 2D.
Glycemic management is central in prevention of small vessel and cardiovascular complications in type 2 diabetes. With the plethora of newer medications and recommendations for a patient centered approach, more information is necessary to match the proper drug to each patient. We showed that BARI 2D, a five-year trial designed to compare two different glycemic treatment strategies, was suitable for assessing different responses according to different phenotypic characteristics. Treatment with insulin sensitizing medications such as thiazolidinediones and metformin was more effective in improving glycemic control, particularly in the more insulin resistant patient, when compared to the insulin provision strategy using insulin and or sulfonylureas. Triglyceride and high density lipoprotein ratio (TG/HDL-cholesterol ratio) was found to be a readily available and practical biomarker that helps to identify the insulin resistant patient. These results support the concept that not all medications for glycemic control work the same in all patients. Thus, tailored therapy can be done using phenotypic characteristics rather than a "one-size-fits-all approach."


Characterization of Bartonella clarridgeiae flagellin (FlaA) and detection of antiflagellin antibodies in patients with lymphadenopathy.
Cat scratch disease (CSD) is a frequent clinical outcome of Bartonella henselae infection in humans. Recently, two case reports indicated Bartonella clarridgeiae as an additional causative agent of CSD. Both pathogens have been isolated from domestic cats, which are considered to be their natural reservoir. B. clarridgeiae and B. henselae can be distinguished phenotypically by the presence or absence of flagella, respectively. Separation of the protein content of purified flagella of B. clarridgeiae by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis indicated that the flagellar filament is mainly composed of a polypeptide with a mass of 41 kDa. N-terminal sequencing of 20 amino acids of this protein revealed a perfect match to the N-terminal sequence of flagellin (FlaA) as deduced from the sequence of the flaA gene cloned from B. clarridgeiae. The flagellin of B. clarridgeiae is closely related to flagellins of Bartonella bacilliformis and several Bartonella-related bacteria. Since flagellar proteins are often immunodominant antigens, we investigated whether antibodies specific for the FlaA protein of B. clarridgeiae are found in patients with CSD or lymphadenopathy. Immunoblotting with 724 sera of patients suffering from lymphadenopathy and 100 healthy controls indicated specific FlaA antibodies in 3.9% of the patients' sera but in none of the controls. B. clarridgeiae FlaA is thus antigenic and expressed in vivo, providing a valuable tool for serological testing. Our results further indicate that B. clarridgeiae might be a possible etiologic agent of CSD or lymphadenopathy. However, it remains to be clarified whether antibodies to the FlaA protein of B. clarridgeiae are a useful indicator of acute infection.


Differential mucin gene expression in human pancreatic and colon cancer cells.
The purpose of this study was to determine the quantity and nature of the mucins synthesized and secreted by four different pancreatic cancer cell lines. Well- to moderately-differentiated SW1990 and CAPAN-2 human pancreatic cancer cells were found to produce more high-Mr glycoprotein (HMG) than less-differentiated MIA PaCa-2 and PANC-1 cells. Most of the labelled HMG was secreted within 24 h. The results of chemical and enzymic degradation, ion-exchange chromatography and density-gradient centrifugation indicated that the HMG in SW1990 and CAPAN-2 cells has the properties expected for mucins, whereas much of the HMG in MIA PaCa-2 and PANC-1 cells may not be mucin, but proteoglycan. These results are consistent with immunoblots and Northern blots showing the presence of apomucin and apomucin mRNA in SW1990 and CAPAN-2 cells, but not in MIA PaCa-2 and PANC-1 cells. The Western blots and Northern blots also show that SW1990 and CAPAN-2 cells, like breast cancer cells, have the mammary-type apomucin and mRNA coded by the MUC1 gene, but lack the intestinal type apomucin and mRNA coded by the MUC2 gene. In contrast, the colon cancer cell lines tested in culture express apomucin and mRNA coded by MUC2 but not by MUC1.


Use of automated external defibrillators by a U.S. airline.
Passengers who have ventricular fibrillation aboard commercial aircraft rarely survive, owing to the delay in obtaining emergency care and defibrillation. In 1997, a major U.S. airline began equipping its aircraft with automated external defibrillators. Flight attendants were trained in the use of the defibrillator and applied the device when passengers had a lack of consciousness, pulse, or respiration. The automated external defibrillator was also used as a monitor for other medical emergencies, generally at the direction of a passenger who was a physician. The electrocardiogram that was obtained during each use of the device was analyzed by two arrhythmia specialists for appropriateness of use. We analyzed data on all 200 instances in which the defibrillators were used between June 1, 1997, and July 15, 1999. Automated external defibrillators were used for 200 patients (191 on the aircraft and 9 in the terminal), including 99 with documented loss of consciousness. Electrocardiographic data were available for 185 patients. The administration of shock was advised in all 14 patients who had electrocardiographically documented ventricular fibrillation, and no shock was advised in the remaining patients (sensitivity and specificity of the defibrillator in identifying ventricular fibrillation, 100 percent). The first shock successfully defibrillated the heart in 13 patients (defibrillation was withheld in 1 case at the family's request). The rate of survival to discharge from the hospital after shock with the automated external defibrillator was 40 percent. A total of 36 patients either died or were resuscitated after cardiac arrest. No complications arose from use of the automated external defibrillator as a monitor in conscious passengers. The use of the automated external defibrillator aboard commercial aircraft is effective, with an excellent rate of survival to discharge from the hospital after conversion of ventricular fibrillation. There are not likely to be complications when the device is used as a monitor in the absence of ventricular fibrillation.


Cancer stem cell vaccination confers significant antitumor immunity.
Most studies of cancer stem cells (CSC) involve the inoculation of cells from human tumors into immunosuppressed mice, preventing an assessment on the immunologic interactions and effects of CSCs. In this study, we examined the vaccination effects produced by CSC-enriched populations from histologically distinct murine tumors after their inoculation into different syngeneic immunocompetent hosts. Enriched CSCs were immunogenic and more effective as an antigen source than unselected tumor cells in inducing protective antitumor immunity. Immune sera from CSC-vaccinated hosts contained high levels of IgG which bound to CSCs, resulting in CSC lysis in the presence of complement. CTLs generated from peripheral blood mononuclear cells or splenocytes harvested from CSC-vaccinated hosts were capable of killing CSCs in vitro. Mechanistic investigations established that CSC-primed antibodies and T cells were capable of selective targeting CSCs and conferring antitumor immunity. Together, these proof-of-concept results provide a rationale for a new type of cancer immunotherapy based on the development of CSC vaccines that can specifically target CSCs.


Low-level laser therapy in different stages of rheumatoid arthritis: a histological study.
Rheumatoid arthritis (RA) is an autoimmune inflammatory disease of unknown etiology. Treatment of RA is very complex, and in the past years, some studies have investigated the use of low-level laser therapy (LLLT) in treatment of RA. However, it remains unknown if LLLT can modulate early and late stages of RA. With this perspective in mind, we evaluated histological aspects of LLLT effects in different RA progression stages in the knee. It was performed a collagen-induced RA model, and 20 male Wistar rats were divided into 4 experimental groups: a non-injured and non-treated control group, a RA non-treated group, a group treated with LLLT (780 nm, 22 mW, 0.10 W/cm(2), spot area of 0.214 cm(2), 7.7 J/cm(2), 75 s, 1.65 J per point, continuous mode) from 12th hour after collagen-induced RA, and a group treated with LLLT from 7th day after RA induction with same LLLT parameters. LLLT treatments were performed once per day. All animals were sacrificed at the 14th day from RA induction and articular tissue was collected in order to perform histological analyses related to inflammatory process. We observed that LLLT both at early and late RA progression stages significantly improved mononuclear inflammatory cells, exudate protein, medullary hemorrhage, hyperemia, necrosis, distribution of fibrocartilage, and chondroblasts and osteoblasts compared to RA group (p < 0.05). We can conclude that LLLT is able to modulate inflammatory response both in early as well as in late progression stages of RA.


Transfer from blue light or green light to white light partially reverses changes in ocular refraction and anatomy of developing guinea pigs.
Relative to the broadband white light (BL), postnatal guinea pigs develop myopia in a monochromic middle-wavelength light (ML, 530 nm) environment and develop hyperopia in a monochromic short-wavelength light (SL, 430 nm) environment. We investigated whether transfer from SL or ML to BL leads to recuperation of ocular refraction and anatomy of developing guinea pigs. Two-week-old guinea pigs were given (a) SL for 20 weeks, (b) SL recuperation (SLR, SL for 10 weeks then BL for 10 weeks), (c) ML for 20 weeks, (d) ML recuperation (MLR, ML for 10 weeks then BL for 10 weeks), or (e) BL for 20 weeks. Two weeks after transfer from ML to BL (MLR group), ocular refraction increased from 1.95 ± 0.35 D to 2.58 ± 0.24 D, and vitreous length decreased from 3.48 ± 0.06 mm to 3.41 ± 0.06 mm. Two weeks after transfer from SL to BL (SLR group), ocular refraction decreased from 5.65 ± 0.61 D to 4.33 ± 0.49 D, and vitreous length increased from 3.18 ± 0.07 mm to 3.26 ± 0.11 mm. The MLR and SLR groups had final ocular refractions that were significantly different from those of the ML and SL groups at 20 weeks (ML vs. MLR: p < 0.0001; SL vs. SLR: p < 0.0001) but were still significantly different from the BL group (BL vs. MLR: p = 0.0120; BL vs. SLR: p = 0.0010). These results suggest that recuperation was not complete after return to BL for 10 weeks.


Implementing Montessori Methods for Dementia™ in Ontario long-term care homes: Recreation staff and multidisciplinary consultants' perceptions of policy and practice issues.
Montessori-based activities use a person-centred approach to benefit persons living with dementia by increasing their participation in, and enjoyment of, daily life. This study investigated recreation staff and multidisciplinary consultants' perceptions of factors that affected implementing Montessori Methods for Dementia™ in long-term care homes in Ontario, Canada. Qualitative data were obtained during semi-structured telephone interviews with 17 participants who worked in these homes. A political economy of aging perspective guided thematic data analysis. Barriers such as insufficient funding and negative attitudes towards activities reinforced a task-oriented biomedical model of care. Various forms of support and understanding helped put Montessori Methods for Dementia™ into practice as a person-centred care program, thus reportedly improving the quality of life of residents living with dementia, staff and family members. These results demonstrate that when Montessori Methods for Dementia™ approaches are learned and understood by staff they can be used as practical interventions for long-term care residents living with dementia.


Curriculum structure: principles and strategy.
This report provides general guidelines for the structure of a curriculum, followed by specific advice on the principles of learning and teaching, the process of restructuring and change leadership and management. It provides examples of several educational philosophies, including vertical and horizontal integration. It discusses the use of competence, learning outcomes, level of degree and assessment and provides a number of recommendations. It does not seek to be prescriptive of time allocation to disciplines within a curriculum. Although this report has been written primarily for those who will develop an undergraduate curriculum, the information may be sufficiently generic to apply to the recent development in graduate entry ('shortened dental' or 'accelerated') courses and to postgraduate degree planning and higher education certificate or diploma courses for other dental care professionals (auxiliaries). The report may have a European bias as progress is made to converge and enhance educational standards in 29 countries with different educational approaches - a microcosm of global collaboration.


Analysis and comparison of plasmid profiles of Borrelia burgdorferi sensu lato strains.
The relationship between plasmid profiles and genospecies of the Lyme disease borreliae was investigated by using 40 strains from diverse biological and geographical sources. The genospecies of the strains were determined by examination of rRNA gene restriction patterns with cDNA probes complementary to the 16S and 23S rRNAs of Escherichia coli. Plasmid profiles were obtained by pulsed-field gel electrophoresis. The number of plasmids per strain and the size of these plasmids ranged from 4 to 10 and from 13.3 to 57.7 kb, respectively. The strains all contained a single large plasmid of 50 to 57.7 kb, with the exception of two Borrelia garinii strains that contained two or three of the large plasmids. The large plasmids of Borrelia burgdorferi sensu stricto strains ranged in size from 51.4 to 52.7 kb and were consistently smaller than the 54.0- to 57.7-kb plasmids present in B. garinii and Borrelia afzelii. The exceptions of this observation were the two B. garinii strains with multiple large plasmids; in this case the large plasmids were 50.6 to 53 kb. Although a large degree of heterogeneity in the sizes and frequencies of occurrence of smaller plasmids was observed, there were some differences among the three genospecies. The differences in plasmids were further studied by using two BamHI DNA fragments from a 28.7-kb plasmid of B. burgdorferi sensu stricto 297 as probes. Both probes hybridized with the 27- to 29-kb plasmids of B. burgdorferi sensu stricto strains. In contrast, two patterns of hybridization were observed with B. garinii and B. afzelii. One pattern was the hybridization of both probes, with each probe hybridizing with a different plasmid. The other pattern was the hybridization of a plasmid of a strain with only one of the two probes. some strains of the three genospecies did not hybridize with either probe. Our results suggest that the plasmid profiles of B. burgdorferi sensu lato have genospecies characteristics and that the hybridization patterns of similar-sized plasmids are different for the three genospecies.


Roles of EGFR, PI3K, AKT, and mTOR in heavy metal-induced cancer.
Humans are exposed to heavy metals through a variety of occupational and non-occupational means. Growing evidence has accumulated that prolonged exposure to these heavy metals is associated with cancer occurrence at various body sites including lung, liver, bladder, colon, and skin. Much research effort has been placed on discovering the mechanisms by which heavy metals induce different kinds of cancers. Results from these mechanistic studies have varied for different metals, but increased activation of signaling pathways is often observed. This review will focus on the signaling molecules including epidermal growth factor receptor (EGFR), phosphatidyl inositol 3-kinase (PI3K), AKT, and mammalian target of rapamycin (mTOR) in carcinogenesis and cancer progression; and how these molecules are affected by the exposure to heavy metals: arsenic, chromium, nickel, and cadmium. Furthermore, drug targets for the prevention and therapy of cancers induced by heavy metals will be discussed with a focus on drugs that are currently in clinical trials for these targets.


Activation of InsP₃ receptors is sufficient for inducing graded intrinsic plasticity in rat hippocampal pyramidal neurons.
The synaptic plasticity literature has focused on establishing necessity and sufficiency as two essential and distinct features in causally relating a signaling molecule to plasticity induction, an approach that has been surprisingly lacking in the intrinsic plasticity literature. In this study, we complemented the recently established necessity of inositol trisphosphate (InsP3) receptors (InsP3R) in a form of intrinsic plasticity by asking if InsP3R activation was sufficient to induce intrinsic plasticity in hippocampal neurons. Specifically, incorporation of d-myo-InsP3 in the recording pipette reduced input resistance, maximal impedance amplitude, and temporal summation but increased resonance frequency, resonance strength, sag ratio, and impedance phase lead. Strikingly, the magnitude of plasticity in all these measurements was dependent on InsP3 concentration, emphasizing the graded dependence of such plasticity on InsP3R activation. Mechanistically, we found that this InsP3-induced plasticity depended on hyperpolarization-activated cyclic nucleotide-gated channels. Moreover, this calcium-dependent form of plasticity was critically reliant on the release of calcium through InsP3Rs, the influx of calcium through N-methyl-d-aspartate receptors and voltage-gated calcium channels, and on the protein kinase A pathway. Our results delineate a causal role for InsP3Rs in graded adaptation of neuronal response dynamics, revealing novel regulatory roles for the endoplasmic reticulum in neural coding and homeostasis.


Semi-Ownership and Sterilisation of Cats and Dogs in Thailand.
The aim of this study was to identify the prevalence of cat and dog semi-ownership in Thailand and factors that predict sterilisation. Semi-ownership was defined as interacting/caring for a companion animal that the respondent does not own, such as a stray cat or dog. A randomised telephone survey recruited 494 Thai nationals residing in Thailand. The findings revealed that 14% of respondents (n = 71) engaged in dog semi-ownership and only 17% of these dogs had been sterilised. Similarly, 11% of respondents (n = 55) engaged in cat semi-ownership and only 7% were known to be sterilised. Using Hierarchical Multiple Regression, the findings showed that 62% and 75% of the variance in intentions to sterilise semi-owned dogs and cats, respectively, was predicted by religious beliefs, and psychosocial factors such as attitudes, perceived pressure from others, and perceived behavioural control. Community awareness campaigns that approach the issue of sterilisation in a way that is consistent with cultural and religious traditions using Thai role models, such as veterinarians, may go some way in reducing stray animal population growth.


Optimized turmeric extract reduces β-Amyloid and phosphorylated Tau protein burden in Alzheimer's transgenic mice.
In a previous in vitro study, the standardized turmeric extract, HSS-888, showed strong inhibition of Aβ aggregation and secretion in vitro, indicating that HSS-888 might be therapeutically important. Therefore, in the present study, HSS-888 was evaluated in vivo using transgenic 'Alzheimer' mice (Tg2576) over-expressing Aβ protein. Following a six-month prevention period where mice received extract HSS-888 (5mg/mouse/day), tetrahydrocurcumin (THC) or a control through ingestion of customized animal feed pellets (0.1% w/w treatment), HSS-888 significantly reduced brain levels of soluble (∼40%) and insoluble (∼20%) Aβ as well as phosphorylated Tau protein (∼80%). In addition, primary cultures of microglia from these mice showed increased expression of the cytokines IL-4 and IL-2. In contrast, THC treatment only weakly reduced phosphorylated Tau protein and failed to significantly alter plaque burden and cytokine expression. The findings reveal that the optimized turmeric extract HSS-888 represents an important step in botanical based therapies for Alzheimer's disease by inhibiting or improving plaque burden, Tau phosphorylation, and microglial inflammation leading to neuronal toxicity.


The association between nurse-administered midazolam following cardiac surgery and incident delirium: an observational study.
Post-operative delirium after cardiac surgery is an adverse event that affects patients' recovery and complicates the delivery of nursing care. Numerous risk factors for delirium are uncontrollable; however, nurses' pro re nata drug administration of sedatives may be a controllable risk factor. This study examined the relationship between nurses' pro re nata administration of midazolam hydrochloride to cardiac surgery patients and the development of post-operative delirium. Observational study. Cardiac surgery intensive care and nursing units of a tertiary care center in Vancouver, Canada. 122 male and female patients requiring non-emergent surgery for coronary artery disease or valvular heart disease who did not have pre-existing cognitive impairment, severe hearing or visual impairment, substance misuse, alcohol intake exceeding 7 drinks per week, or renal impairment requiring hemodialysis. Patients were assessed for delirium, on three occasions, with the Confusion Assessment Method for the Intensive Care Unit (CAM-ICU) during the first 72 h after surgery and through reviews of physicians' notes. Risk factor and midazolam dosage data were collected from medical records. 77.9% of the patients in this sample received midazolam hydrochloride post-operatively. The prevalence of delirium ranged from 37.7% to 44.3%. Almost all of the dosages of midazolam (85-87%) were given before the first indication of delirium; that is, most of the patients had received their entire dosage before the first signs of delirium were detected. Bivariate analysis with logistic regression models revealed that for every additional milligram of midazolam administered, the patients were 7-8% more likely to develop delirium. Multivariate logistic regression models demonstrated that the magnitude of the association between midazolam dosage and delirium was not confounded by established risk factors including age and peripheral vascular disease. Nurses play an important role in the prediction, assessment and prevention of post-operative delirium. Sedatives should be administered with caution because they increase a patient's risk of developing delirium. Nurses' decisions regarding sedation administration must be informed by empirical knowledge, accurate assessment data and clear rationale with consideration of how these actions may contribute to the development of delirium.


Crystal structure of the pestivirus envelope glycoprotein E(rns) and mechanistic analysis of its ribonuclease activity.
Pestiviruses, which belong to the Flaviviridae family of RNA viruses, are important agents of veterinary diseases causing substantial economical losses in animal farming worldwide. Pestivirus particles display three envelope glycoproteins at their surface: E(rns), E1, and E2. We report here the crystal structure of the catalytic domain of E(rns), the ribonucleolytic activity of which is believed to counteract the innate immunity of the host. The structure reveals a three-dimensional fold corresponding to T2 ribonucleases from plants and fungi. Cocrystallization experiments with mono- and oligonucleotides revealed the structural basis for substrate recognition at two binding sites previously identified for T2 RNases. A detailed analysis of poly-U cleavage products using (31)P-NMR and size exclusion chromatography, together with molecular docking studies, provides a comprehensive mechanistic picture of E(rns) activity on its substrates and reveals the presence of at least one additional nucleotide binding site.


Correlates of adherence to supervised exercise in patients awaiting surgical removal of malignant lung lesions: results of a pilot study.
To examine the demographic, medical, and social-cognitive correlates of adherence to a presurgical exercise training intervention in patients awaiting surgery for suspected malignant lung lesions. Pilot study, single-group, prospective design with convenience sampling. Exercise training was performed at a university research fitness center in western Canada. 19 patients awaiting surgical resection of suspected malignant lung lesions. At baseline, participants completed a questionnaire including the Theory of Planned Behavior variables of perceived behavioral control, attitude, and subjective norm, as well as medical and demographic information. Participants were asked to attend five supervised exercise sessions per week during surgical wait time (X = 8 +/- 2.4 weeks). Theory of Planned Behavior variables and exercise adherence. Adherence to the exercise intervention was 73% (range = 0%-100%). Correlates of adherence were perceived behavioral control (r = 0.63; p = 0.004) and subjective norm (r = 0.51; p = 0.014). Participants with greater than 80% adherence reported significantly higher behavioral control than participants with less than 80% adherence (X difference = 1.1; 95% confidence interval = 0.1-2.2; p = 0.035). Men had better adherence than women (X difference = 24.9%; 95% confidence interval = 0.4-49.4; p = 0.047). Perceived behavioral control and subjective norm were the strongest correlates of exercise adherence. Women could be at risk for poor exercise adherence prior to lung surgery. This information could be useful for clinicians in their attempts to improve adherence to exercise interventions in patients awaiting surgery for malignant lung lesions.


Inhibition of P. aeruginosa c-di-GMP phosphodiesterase RocR and swarming motility by a benzoisothiazolinone derivative.
Various important cellular processes in bacteria are controlled by c-di-GMP, such as motility, biofilm formation and virulence factors production. C-di-GMP is synthesized from two molecules of GTP by diguanylate cyclases (DGCs) and its actions are terminated by EAL or HD-GYP domain phosphodiesterases (PDEs), which hydrolyze c-di-GMP to linear pGpG or GMP. Thus far the majority of efforts have been dedicated to the development of inhibitors of DGCs but not PDEs. This is probably because the old view was that inhibiting any c-di-GMP PDE would lead to biofilm formation, an undesirable phenotype. Recent data however suggest that some PDEs only change the localized (not global) concentration of c-di-GMP to increase bacterial virulence and do not affect biofilm formation. A challenge therefore is to be able to develop selective PDE inhibitors that inhibit virulence-associated PDEs but not inhibit PDEs that regulate bacterial biofilm formation. Using high throughput docking experiments to screen a library of 250 000 commercially available compounds against E. coli YahA (also called PdeL), a benzoisothiazolinone derivative was found to bind to the c-di-GMP binding site of YahA with favorable energetics. Paradoxically the in silico identified inhibitor (a benzoisothiazolinone derivative) did not inhibit the hydrolysis of c-di-GMP by YahA, the model PDE that was used in the docking, but instead inhibited RocR, which is a PDE from the opportunistic pathogen P. aeruginosa (PA). RocR promotes bacterial virulence but not biofilm dispersal, making it an ideal PDE to target for anti-virulence purposes. This newly identified RocR ligand displayed some selectivity and did not inhibit other P. aeruginosa PDEs, such as DipA, PvrR and PA4108. DipA, PvrR and PA4108 are key enzymes that reduce global c-di-GMP concentration and promote biofilm dispersal; therefore the identification of an inhibitor of a PA PDE, such as RocR, that does not inhibit major PDEs that modulate global c-di-GMP is an important step towards the development of selective c-di-GMP PDEs that could have interesting biomedical applications. The identified RocR ligand could also inhibit P. aeruginosa (PAO1) swarming but not swimming or biofilm formation. Rhamnolipid production was decreased, explaining the inhibition of swarming.


Opioid pharmacology.
Mu agonists have been an important component of pain treatment for thousands of years. The usual pharmacokinetic parameters (half-life, clearance, volume of distribution) of opioids have been known for some time. However, the metabolism has, until recently, been poorly understood, and there has been recent interest in the role of metabolites in modifying the pharmacodynamic response in patients, in both analgesia and adverse effects. A number of opioids are available for clinical use, including morphine, hydromorphone, levorphanol, oxycodone, and fentanyl. Advantages and disadvantages of various opioids in the management of chronic pain are discussed. This review looks at the structure, chemistry, and metabolism of opioids in an effort to better understand the side effects, drug interactions, and the individual responses of patients receiving opioids for the treatment of intractable pain. Mu receptor agonists and agonist-antagonists have been used throughout recent medical history for the control of pain and for the treatment of opiate induced side effects and even opiate withdrawal syndromes.


Needs of people with dementia and their spousal carers: A study of those living in the community.
To identify care and support needs, as reported by people with dementia and their spousal carers living in the community in metropolitan Western Australia. An interpretive description approach was utilised. Semi-structured interviews were conducted with 10 dyads of spousal carers and people with dementia. Comparative analysis was used to develop themes regarding need, which were reviewed using two focus groups of spousal carers. Three major themes were developed through analysis of data from the interviews and were refined by the focus groups: (a) environmental enablers to support care; (b) strong caring relationships; and (c) adaptation of daily life roles. As dementia progresses, continued support to maintain participation in meaningful activity is needed. Strong caring relationships, knowledge and an understanding of dementia among health professionals, service providers and family members were key to maintaining support for the needs of people with dementia and their spousal carers.


Poly(ADP-ribose) polymerase activity contributes to peroxynitrite-induced spinal cord neuronal cell death in vitro.
Peroxynitrite, which has been implicated in secondary neuronal damage resulting from spinal cord injury, is capable of mediating several toxic interactions including inducing DNA strand breaks and activating the nuclear enzyme, poly (ADP-ribose) polymerase (PARP). In the present study we have tested the hypothesis that peroxynitrite-induced cell death in spinal cord injury is due to activation of PARP. Initially we examined whether peroxynitrite exerts toxic effects on primary cultures of spinal cord neurons and then determined whether the spinal cord neuronal cell death triggered by peroxynitrite was associated with PARP activation. Peroxynitrite dose-dependently reduced the viability of spinal cord neurons in vitro. Furthermore, peroxynitrite exposure markedly increased the number of DNA strand breaks in primary spinal cord neurons, resulting in activation of PARP. To identify whether PARP activation plays a direct role in peroxynitrite-induced neurotoxicity we assessed the effects of the PARP inhibitors, nicotinamide, 3-aminobenzamide and 5-iodo-6-amino-1,2 benzopyrone on cell viability in spinal cord neurons exposed to peroxynitrite. The presence of the PARP inhibitors in the cultures not only inhibited peroxynitrite-induced PARP activity in spinal cord neurons but also protected the cells from the deleterious actions of peroxynitrite. Therefore, our results demonstrate that peroxynitrite exerts toxic effects on spinal cord neurons in vitro at least in part through a PARP-dependent pathway.


Hemodilution tolerance in patients with coronary artery disease who are receiving chronic beta-adrenergic blocker therapy.
Hemodilution tolerance is not well defined in patients with coronary artery disease receiving beta-adrenergic blockers chronically. Ninety patients scheduled for coronary artery bypass graft (CABG) surgery were randomized to a hemodilution (n = 60) and a control group (n = 30). During midazolam-fentanyl anesthesia, hemodynamic variables, ST segment deviation, and O2 consumption were determined prior to and after 6 and 12 mL/kg isovolemic exchange of blood for 6% hydroxyethyl starch. Hemoglobin decreased from 12.6 +/- 0.2 to 9.9 +/- 0.2 g/dL (mean +/- SEM, P < 0.05). With stable filling pressures, cardiac index increased from 2.05 +/- 0.05 to 2.27 +/- 0.05 L.min-1.m-2(P < 0.05) and O2 extraction from 27.4% +/- 0.6% to 31.2% +/- 0.7% (P < 0.05), resulting in stable O2 consumption. No alterations in ST segments were observed in leads II and V5 during hemodilution. Individual increases in cardiac index and O2 extraction were not linearly related to age and left ventricular (LV) ejection fraction (P = 0.841, P = 0.799). We conclude that isovolemic hemodilution to a hemoglobin value of 9.9 +/- 0.2 g/dL is well tolerated and fully compensated in patients with coronary artery disease receiving beta-adrenergic blockers chronically. Within the investigated ranges, the compensatory mechanisms during hemodilution are largely independent of age (35-81 yr) and LV ejection fraction (26%-83%).


[Effect of prolonged post-tetanic potentiation on the activity of mediator metabolism enzymes in the hippocampus of rabbits].
The activity of acetylcholinesterase (AChE) and monoaminoxidase (MAO) in the field CA3 of rabbit's hippocampus was studied with histochemical methods during prolonged posttetanic potentiation (PTP), evoked by short (20 s) low-frequency (20/s) stimulation of n. septo-fimbrialis. Studies were conducted 20 min (group I) and 60 min (group II) after tetanization. In the group I a decrease in AChE activity was observed, especially in str. pyramidale and in the adjacent region of str. oriens. In the group II AChE activity increased, mostly in the external part of str. radiatum. MAO activity increased in both animal groups. For the group I a low (r = 0,42), but significant (P less than 0.05) correlation between PTP value and change in response to AChE was discovered. The reduction of activity 20 min after tetanization may be due to non-specific effect of tetanization as such and to a relatively short posttetanic depression. The increase in MAO activity is interpreted as an indirect confirmation of the part of the monoaminoergic systems in stabilization of plastic rearrangements elicited by tetanization.


Adaptive evolution of genomically recoded Escherichia coli.
Efforts are underway to construct several recoded genomes anticipated to exhibit multivirus resistance, enhanced nonstandard amino acid (nsAA) incorporation, and capability for synthetic biocontainment. Although our laboratory pioneered the first genomically recoded organism (Escherichia coli strain C321.∆A), its fitness is far lower than that of its nonrecoded ancestor, particularly in defined media. This fitness deficit severely limits its utility for nsAA-linked applications requiring defined media, such as live cell imaging, metabolic engineering, and industrial-scale protein production. Here, we report adaptive evolution of C321.∆A for more than 1,000 generations in independent replicate populations grown in glucose minimal media. Evolved recoded populations significantly exceeded the growth rates of both the ancestral C321.∆A and nonrecoded strains. We used next-generation sequencing to identify genes mutated in multiple independent populations, and we reconstructed individual alleles in ancestral strains via multiplex automatable genome engineering (MAGE) to quantify their effects on fitness. Several selective mutations occurred only in recoded evolved populations, some of which are associated with altering the translation apparatus in response to recoding, whereas others are not apparently associated with recoding, but instead correct for off-target mutations that occurred during initial genome engineering. This report demonstrates that laboratory evolution can be applied after engineering of recoded genomes to streamline fitness recovery compared with application of additional targeted engineering strategies that may introduce further unintended mutations. In doing so, we provide the most comprehensive insight to date into the physiology of the commonly used C321.∆A strain.


Pouteria torta: a native species of the Brazilian Cerrado as a bioindicator of glyphosate action.
In Brazil, the expansion of agricultural activity and the associated indiscriminate use of herbicides such as glyphosate is directly related to the loss of biodiversity in the Cerrado. The identification of plant species as bioindicators of herbicide action, especially species native to the area, can help in monitoring the impacts of xenobiotics in the remaining Cerrado. Thus, this study was designed to evaluate the possible use of the native Cerrado species Pouteria torta as a bioindicator of glyphosate action via changes in physiological performance. At 16 months after sowing, the effect of glyphosate was evaluated by applying the following doses: 0 (control), 25, 50, 100, 200, 400, 800, and 1200 g a.e. ha-1. In response to glyphosate, P. torta exhibited reductions in photosynthesis and chloroplastid pigment content, as well as accumulation of shikimic acid and the occurrence of chlorosis and necrosis. These changes demonstrate the high sensitivity of P. torta to glyphosate and its potential for use as a bioindicator of this herbicide.


Matrix based fertilizers reduce nitrogen and phosphorus leaching in three soils.
We compared the efficacy of matrix based fertilizers (MBFs) formulated to reduce NO3-, NH4+, and total phosphorus (TP) leaching, with Osmocoate 14-14-14, a conventional commercial slow release fertilizer (SRF) and an unamended control in three different soil textures in a greenhouse column study. The MBFs covered a range of inorganic N and P in compounds that are relatively loosely bound (MBF 1) to more moderately bound (MBF 2) and more tightly bound compounds (MBF 3) mixed with Al(SO4)3H2O and/or Fe2(SO4)3 and with high ionic exchange compounds starch, chitosan and lignin. When N and P are released, the chemicals containing these nutrients in the MBF bind N and P to a Al(SO4)3H2O and/or Fe2(SO4)3 starch-chitosan-lignin matrix. One milligram (8000 spores) of Glomus intradices was added to all formulations to enhance nutrient uptake. In all three soil textures the SRF leachate contained a higher amount of NH4+, NO3- and TP than leachate from all other fertilizers. In all three soils there were no consistent differences in the amount of NH4+, NO3- and TP in the MBF leachates compared to the control leachate. Plants growing in soils receiving SRF had greater shoot, root and total biomass than all MBFs regardless of Al(SO4)3H2O or Fe2(SO4)3 additions. Arbuscular mycorrhizal infection in plant roots did not consistently differ among plants growing in soil receiving SRF, MBFs and control treatments. Although the MBFs resulted in less plant growth in this experiment they may be applied to soils growing plants in areas that are at high risk for nutrient leaching to surface waters.


ChAdOx1 nCoV-19 (AZD1222) vaccine-induced Fc receptor binding tracks with differential susceptibility to COVID-19.
Despite the success of COVID-19 vaccines, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern have emerged that can cause breakthrough infections. Although protection against severe disease has been largely preserved, the immunological mediators of protection in humans remain undefined. We performed a substudy on the ChAdOx1 nCoV-19 (AZD1222) vaccinees enrolled in a South African clinical trial. At peak immunogenicity, before infection, no differences were observed in immunoglobulin (Ig)G1-binding antibody titers; however, the vaccine induced different Fc-receptor-binding antibodies across groups. Vaccinees who resisted COVID-19 exclusively mounted FcγR3B-binding antibodies. In contrast, enhanced IgA and IgG3, linked to enriched FcγR2B binding, was observed in individuals who experienced breakthrough. Antibodies unable to bind to FcγR3B led to immune complex clearance and resulted in inflammatory cascades. Differential antibody binding to FcγR3B was linked to Fc-glycosylation differences in SARS-CoV-2-specific antibodies. These data potentially point to specific FcγR3B-mediated antibody functional profiles as critical markers of immunity against COVID-19.


Assessing the Performance of 3 Human Immunodeficiency Virus Incidence Risk Scores in a Cohort of Black and White Men Who Have Sex With Men in the South.
Risk scores have been developed to identify men at high risk of human immunodeficiency virus (HIV) seroconversion. These scores can be used to more efficiently allocate public health prevention resources, such as pre-exposure prophylaxis. However, the published scores were developed with data sets that comprise predominantly white men who have sex with men (MSM) collected several years prior and recruited from a limited geographic area. Thus, it is unclear how well these scores perform in men of different races or ethnicities or men in different geographic regions. We assessed the predictive ability of 3 published scores to predict HIV seroconversion in a cohort of black and white MSM in Atlanta, GA. Questionnaire data from the baseline study visit were used to derive individual scores for each participant. We assessed the discriminatory ability of each risk score to predict HIV seroconversion over 2 years of follow-up. The predictive ability of each score was low among all MSM and lower among black men compared to white men. Each score had lower sensitivity to predict seroconversion among black MSM compared to white MSM and low area under the curve values for the receiver operating characteristic curve indicating poor discriminatory ability. Reliance on the currently available risk scores will result in misclassification of high proportions of MSM, especially black MSM, in terms of HIV risk, leading to missed opportunities for HIV prevention services.


Lighting as a Compensatory Strategy for Acquired Visual Deficits After Stroke: Two Case Reports.
The objective of this article was to explore the effects of enhanced lighting on the function of people with an acquired visual deficit resulting from cerebrovascular accident (CVA) in an inpatient rehabilitation facility (IRF). An ABAB design was used with two cases to assess how a short-term intervention involving lighting changes affected clients' grooming performance in an IRF. Analysis consisted of scores on the grooming section of the FIM®, times for each grooming task, and light meter readings. Both participants demonstrated improvements in function as measured by the FIM and in time to complete grooming tasks with enhanced lighting. These results suggest that individualized lighting can lead to improved grooming function in clients with CVA.


Absence of posttraumatic arthritis following intraarticular fracture in the MRL/MpJ mouse.
Posttraumatic arthritis is a frequent long-term complication of intraarticular fractures. A model of a closed intraarticular fracture in C57BL/6 mice that progresses to posttraumatic arthritis has been developed. The MRL/MpJ mouse has shown unique regenerative abilities in response to injury. The objective of this study was to determine if the MRL/MpJ mouse is protected from posttraumatic arthritis after intraarticular fractures. Intraarticular fractures were created in MRL/MpJ mice and C57BL/6 control mice (n = 16 each). Limbs were analyzed for posttraumatic arthritis 4 and 8 weeks after fracture using microfocal computed tomography bone morphology, subchondral bone thickness evaluation, and histologic evaluation of cartilage degeneration. Serum cytokines and biomarkers were measured after the mice were killed. Intraarticular fractures were successfully created in all 32 mice. In the experimental fractured limbs, C57BL/6 mice had a decrease in bone density, increased subchondral bone thickness, and increased cartilage degeneration compared with normal contralateral control limbs. In the MRL/MpJ mice, no differences in bone density, subchondral bone thickness, or histologic grading of cartilage degeneration were seen between fractured and contralateral control limbs. Cytokine analysis showed lower systemic levels of the proinflammatory cytokine interleukin-1alpha (IL-1alpha) and higher levels of the antiinflammatory cytokines IL-4 and IL-10 in the MRL/MpJ mice. This study shows that the MRL/MpJ mouse is relatively protected from posttraumatic arthritis after intraarticular fracture. Further investigation into the mechanism involved in this response will hopefully provide new insight into the pathogenesis, prevention, and treatment of posttraumatic arthritis after intraarticular fracture.


Retrospective morphometric analysis of the infraorbital foramen with cone beam computed tomography.
The aim of our study is to examine the morphometric characteristics of the infraorbital foramen (IOF) and its anatomic localization by using cone-beam computerized tomography (CBCT). In our study, the anatomic characteristics of the IOF were identified by studying retrospectively the CBCT images of one hundred twenty-five (125) patients. These characteristics were assessed statistically in terms of age, gender, and right-left side. In our study, 42% of them had IOF in circular form and 58% in oval form. Of them 66.90% of those in oval form were in oblique direction, 28.30% of them were in vertical direction, and 4.80% of them were in horizontal direction. The mean vertical diameter of the IOF was found as 3.71 ± 0.61 mm and its mean horizontal diameter was found as 3.17 ± 0.56 mm. The mean distance of IOF to midline were measured as 25.10 ± 2.17 mm, distance to upper edge of the orbita was measured as 41.91 ± 2.77 mm, distance to infraorbital edge was measured as 5.63 ± 1.77 mm, distance to the lateral nasal wall was measured as 9.32 ± 2.68 mm, distance to spina nasalis anterior was measured as 17.97 ± 3.99 mm, thickness of the soft tissue on the foramen was measured as 11.52 ± 2.40 mm, and distance between the right and the left IOF was measured as 50.20 ± 3.90 mm. Accessory foramen was found in 56.60% ratio. This study reveals that CBCT may guide for local anesthesia applications and for other invasive procedures in order to prevent the damage of the neurovascular structures during maxillofacial surgery by identifying the distances of anatomic points such as IOF having surgically importance.


Lack of association of the HMGA1 IVS5-13insC variant with type 2 diabetes in an ethnically diverse hypertensive case control cohort.
Recently, the high-mobility group A1 gene (HMGA1) variant IVS5-13insC has been associated with type 2 diabetes, but reported associations are inconsistent and data are lacking in Hispanic and African American populations. We sought to investigate the HMGA1-diabetes association and to characterize IVS5-13insC allele frequencies and linkage disequilibrium (LD) in 3,070 Caucasian, Hispanic, and African American patients from the INternational VErapamil SR-Trandolapril STudy (INVEST). INVEST was a randomized, multicenter trial comparing two antihypertensive treatment strategies in an ethnically diverse cohort of hypertensive, coronary artery disease patients. Controls, who were diabetes-free throughout the study, and type 2 diabetes cases, either prevalent or incident, were genotyped for IVS5-13insC using Taqman®, confirmed with Pyrosequencing and Sanger sequencing. For LD analysis, genotyping for eight additional HMGA1 single nucleotide polymorphisms (SNPs) was performed using the Illumina® HumanCVD BeadChip. We used logistic regression to test association of the HMGA1 IVS5-13insC and diabetes, adjusted for age, gender, body mass index, and percentage European, African, and Native American ancestry. We observed IVS5-13insC minor allele frequencies consistent with previous literature in Caucasians and African Americans (0.03 in cases and 0.04 in controls for both race/ethnic groups), and higher frequencies in Hispanics (0.07 in cases and 0.07 in controls). The IVS5-13insC was not associated with type 2 diabetes overall (odds ratio 0.98 [0.76-1.26], p=0.88) or in any race/ethnic group. Pairwise LD (r2) of IVS5-13insC and rs9394200, a SNP previously used as a tag SNP for IVS5-13insC, was low (r2=0.47 in Caucasians, r2=0.25 in Hispanics, and r2=0.06 in African Americans). Furthermore, in silico analysis suggested a lack of functional consequences for the IVS5-13insC variant. Our results suggest that IVS5-13insC is not a functional variant and not associated with type 2 diabetes in an ethnically diverse, hypertensive, coronary artery disease population. Larger, more adequately powered studies need to be performed to confirm our findings.


The role of vascular capacitance in the coronary arteries.
When the left coronary artery was perfused with nonpulsatile pressure, the onset of diastole was accompanied by a capacitance overshoot in flow with an exponential decay back to a steady state. Time constant for that decay ranged from 55 msec when tone was present to 105 msec with maximal dilation. Since the transient resulted from a fall in tissue pressure, this represents an estimation of intramural arterial capacitance only. Transients in perfusion pressure, which would also affect epicardial arteries, yielded similar time constants. We concluded that most of the coronary capacitance resides in the small intramural vessels. Analysis of transients yielded a value for capacitance of between 0.01 and 0.05 ml/mm Hg per 100 g. We then used the data from the transients to construct coronary pressure flow curves which were free of any back flow from capacitance. When coronary tone was present, the curves indicated that flow ceased at 30 mm Hg. With maximal dilation, flow ceased at only 18 mm Hg. Long diastoles in those same hearts indicated that flow ceased at about 10 mm Hg higher pressure. Although capacitance causes critical closing pressure as determined by a long diastole to be artifactually high, critical closing pressure is still appreciable in the heart, and tone dependent. Finally, three computer models were built, one of which included only small vessel capacitances, the second, only vascular waterfalls, and the third, both of the above. Only model 3 was capable of reproducing the flow patterns which were actually seen.


Human bone marrow osteoprogenitors express estrogen receptor-alpha and bone morphogenetic proteins 2 and 4 mRNA during osteoblastic differentiation.
Understanding the mechanisms that control the proliferation and commitment of human stem cells into cells of the osteogenic lineage for the preservation of skeletal structure is of basic importance in bone physiology. This study examines some aspects of the differentiation in vitro of human bone marrow fibroblastic cells cultured in the absence (basal media) or presence of 1nM dexamethasone and 50 micrograms/ml ascorbate for 6, 10, 14, and 21 days. Northern blot analysis and in situ hybridisation with digoxygenin-labelled riboprobes for Type I collagen, osteocalcin, bone morphogenetic proteins 2 (BMP-2), and 4 (BMP-4) and the estrogen receptor alpha (ERalpha), together with immunocytochemical analysis of ERalpha expression and histochemical staining of alkaline phosphatase was performed. In basal media, alkaline phosphatase activity and collagen expressions were detected at day 6, ERalpha from day 10 and osteocalcin from day 10. In the presence of dexamethasone and ascorbate, cell proliferation and alkaline phosphatase were markedly stimulated over 10 to 14 days with a dramatic increase in the temporal expression of Type I collagen, ERalpha, and osteocalcin mRNAs in these cultures. Northern blot analysis showed cells cultured in basal media, expressed the highest levels of the mRNA for each marker protein at day 14, whereas in the presence of ascorbate and dexamethasone, the highest levels for alkaline phosphatase, ERalpha, osteocalcin, BMP-2, and BMP-4 were observed at day 21. ERalpha, BMP-2, and BMP-4 expression were found to correlate temporally with induction of the osteoblast phenotype as determined by alkaline phosphatase, collagen, and osteocalcin expression. These results give additional information on the development of the osteoblast phenotype from early fibroblastic stem cells and on the biological factors involved in this process. These studies suggest a role for estrogen and BMP-2 and -4 in the differentiation of osteoprogenitor cells.


S-nitrosoglutathione inhibits alpha1-adrenergic receptor-mediated vasoconstriction and ligand binding in pulmonary artery.
Endogenous nitric oxide donor compounds (S-nitrosothiols) contribute to low vascular tone by both cGMP-dependent and -independent pathways. We have reported that S-nitrosoglutathione (GSNO) inhibits 5-hydroxytryptamine (5-HT)-mediated pulmonary vasoconstriction via a cGMP-independent mechanism likely involving S-nitrosylation of its G protein-coupled receptor (GPCR) system. Because catecholamines, like 5-HT, constrict lung vessels via a GPCR coupled to G(q), we hypothesized that S-nitrosothiols modify the alpha1-adrenergic GPCR system to inhibit pulmonary vasoconstriction by receptor agonists, e.g., phenylephrine (PE). Rat pulmonary artery rings were pretreated for 30 min with and without an S-nitrosothiol, either GSNO or S-nitrosocysteine (CSNO), and constricted with sequential concentrations of PE (10(-8)-10(-6) M). Effective cGMP-dependence was tested in rings pretreated with soluble guanylate cyclase inhibitors {either 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or LY-83583} or G kinase inhibitor (KT-5823), and a thiol reductant [dithiothreitol (DTT)] was used to test reversibility of S-nitrosylation. Both S-nitrosothiols attenuated the PE dose response. The GSNO effect was not prevented by LY-83583, ODQ, or KT-5823, indicating cGMP independence. GSNO inhibition was reversed by DTT, consistent with S-nitrosylation or other GSNO-mediated cysteine modifications. In CSNO-treated lung protein, the alpha1-adrenergic receptor was shown to undergo S-nitrosylation in vitro using a biotin switch assay. Studies of alpha1-adrenergic receptor subtype expression and receptor density by saturation binding with 125I-HEAT showed that GSNO decreased alpha1-adrenergic receptor density but did not alter affinity for antagonist or agonist. These data demonstrate a novel cGMP-independent mechanism of reversible alpha1-adrenergic receptor inhibition by S-nitrosothiols.


The metastability of the proteome of spinal motor neurons underlies their selective vulnerability in ALS.
Amyotrophic lateral sclerosis (ALS) is a heterogeneous motor neuron disease with familial forms linked to numerous mutations in a range of genes. The resulting variant proteins, including SOD1, TDP-43, and FUS, disturb protein homeostasis in a variety of ways and lead to the formation of intracellular inclusion bodies that are characteristic of different neuropathological subtypes of the disease. These inclusions are made up of scores of proteins that do not appear at first to share obvious characteristics other than coaggregation. Recent evidence, however, suggests that these aggregating proteins can be characterized as being supersaturated in spinal motor neurons, as they exhibit cellular concentrations exceeding their solubilities. Here, we show that the average supersaturation of the entire spinal motor neuron proteome is greater than that of the ALS-resistant oculomotor neurons, suggesting that the vulnerability of spinal motor neurons is linked to the overall metastability of their proteome against aggregation. Consistently, ALS expression data suggest that affected neurons respond to pathology by transcriptional downregulation of supersaturated proteins, including specifically ion channels. These results identify a mechanism by which protein homeostasis imbalance leads to inclusion body formation in ALS, and to a disruption of other processes dependent on proteins that are supersaturated, thereby resulting in the dysfunctional excitability alterations observed in vivo.


Subdural empyema: Clinical presentations and management options for an uncommon neurosurgical emergency in a developing country.
Controversy regarding the best management strategy for subdural empyema (SDE) attests to the persisting poor outcomes for this uncommon life threatening intracranial suppurative process. Late presentation confounds the problem in developed countries. While craniotomy is commonly recommended, it is not always possible in late presentation with advanced morbidity. The aim of this study was to identify the pattern of clinical presentation and explore the outcomes following management of SDE using burr hole, aspiration, and drainage (BAAD) in resource poor settings. This is a retrospective review of prospectively collected data of 18 patients presenting with SDE over a period of 10 years from two neurosurgical centers. Data was abstracted on patients' demographic characteristics, sources of SDE, clinical presentation and site of infection, methods of diagnosis, organisms isolated, treatments received, and outcome. Collected data was entered into the Statistical Package for the Social Sciences version 17 software and subjected to descriptive analysis for all variables. Majority of the patients presented late with Glasgow Coma Scale score GCS of 9/15. Altered sensorium was noted in 14 (77.8%) of the patients, 11 (61.1%) out of the 14 patients had ≤ grade 3 of Bannister and Williams level of consciousness. The mainstay of treatment for all patients was BAAD of abscess and administration of appropriate antibiotics. Fourteen patients (77.8%) were discharged on grade A of H.W. Mauser's grading system. Three mortalities were recorded only in patients who had grade 4 Bannister and Williams grading. BAAD is the near approximated option to standard craniotomy management in a limited resource facility and it has a very good clinical outcome. However, more studies are required to draw the final conclusion.


Identification of factors associated with good response to growth hormone therapy in children with short stature: results from the ANSWER Program®.
To identify factors associated with growth in children on growth hormone (GH) therapy using data from the American Norditropin Studies: Web-enabled Research (ANSWER) Program® registry. GH-naïve children with GH deficiency, multiple pituitary hormone deficiency, idiopathic short stature, Turner syndrome, or a history of small for gestational age were eligible (N = 1,002). Using a longitudinal statistical approach, predictive factors were identified in patients with GHD for change from baseline in height standard deviation score (ΔHSDS) following 2 years of treatment. Gradual increases in ΔHSDS over time were observed for all diagnostic categories. Significant predictive factors of ΔHSDS, ranked by significance were: height velocity (HV) at 4 months > baseline age > baseline HSDS > baseline body mass index (BMI) SDS > baseline insulin-like growth factor I (IGF-I) SDS; gender was not significant. HV at 4 months and baseline BMI SDS were positively correlated, whereas baseline age, HSDS, and IGF-I SDS were negatively correlated with ΔHSDS. These results may help guide GH therapy based on pretreatment characteristics and early growth response.


Satellite telemetry reveals higher fishing mortality rates than previously estimated, suggesting overfishing of an apex marine predator.
Overfishing is a primary cause of population declines for many shark species of conservation concern. However, means of obtaining information on fishery interactions and mortality, necessary for the development of successful conservation strategies, are often fisheries-dependent and of questionable quality for many species of commercially exploited pelagic sharks. We used satellite telemetry as a fisheries-independent tool to document fisheries interactions, and quantify fishing mortality of the highly migratory shortfin mako shark (Isurus oxyrinchus) in the western North Atlantic Ocean. Forty satellite-tagged shortfin mako sharks tracked over 3 years entered the Exclusive Economic Zones of 19 countries and were harvested in fisheries of five countries, with 30% of tagged sharks harvested. Our tagging-derived estimates of instantaneous fishing mortality rates (F = 0.19-0.56) were 10-fold higher than previous estimates from fisheries-dependent data (approx. 0.015-0.024), suggesting data used in stock assessments may considerably underestimate fishing mortality. Additionally, our estimates of F were greater than those associated with maximum sustainable yield, suggesting a state of overfishing. This information has direct application to evaluations of stock status and for effective management of populations, and thus satellite tagging studies have potential to provide more accurate estimates of fishing mortality and survival than traditional fisheries-dependent methodology.


(15)N NMR Spectroscopy, X-ray and Neutron Diffraction, Quantum-Chemical Calculations, and UV/vis-Spectrophotometric Titrations as Complementary Techniques for the Analysis of Pyridine-Supported Bicyclic Guanidine Superbases.
Pyridine substituted with one and two bicyclic guanidine groups has been studied as a potential source of superbases. 2-{hpp}C5H4N (I) and 2,6-{hpp}2C5H3N (II) (hppH = 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine) were protonated using [HNEt3][BPh4] to afford [I-H][BPh4] (1a), [II-H][BPh4] (2), and [II-H2][BPh4]2 (3). Solution-state (1)H and (15)N NMR spectroscopy shows a symmetrical cation in 2, indicating a facile proton-exchange process in solution. Solid-state (15)N NMR data differentiates between the two groups, indicating a mixed guanidine/guanidinium. X-ray diffraction data are consistent with protonation at the imine nitrogen, confirmed for 1a by single-crystal neutron diffraction. The crystal structure of 1a shows association of two [I-H](+) cations within a cage of [BPh4](-) anions. Computational analysis performed in the gas phase and in MeCN solution shows that the free energy barrier to transfer a proton between imino centers in [II-H](+) is 1 order of magnitude lower in MeCN than in the gas phase. The results provide evidence that linking hpp groups with the pyridyl group stabilizes the protonation center, thereby increasing the intrinsic basicity in the gas phase, while the bulk prevents efficient cation solvation, resulting in diminished pKa(MeCN) values. Spectrophotometrically measured pKa values are in excellent agreement with calculated values and confirm that I and II are superbases in solution.


Genetic control of lifespan: studies from animal models.
The premise that there are genes that wield a strong influence on longevity has, until recently, not been a popular one and there has been no concerted effort to find such genes. However, the finding that single genes can have large effects on the lifespans of yeast, worms and flies raises the possibility that individual genes in mammals may similarly have relatively large effects on longevity. Recent advances in mammalian genetics, many associated with the large-scale efforts to sequence the human and mouse genomes, have accelerated the search for longevity genes, principally in mice. Here, we review results using animal models that have recently shed light on genes regulating longevity and ageing patterns. A large number of genetically defined strains of mice are available and this, together with their established history of use in genetic research and their relatively short lifespans, has made murine models particularly useful. We also review our own work in which genes regulating mouse lifespan and those regulating cell cycling of haematopoietic progenitor cells have been mapped to the same locations in the genome. These results suggest that some of the same genes affect both traits, and further suggest a cause-and-effect relationship between cumulative cell-cycle activity and longevity of an organism.


Prevention of perinatal group B streptococcal disease. Revised guidelines from CDC.
Group B streptococcus (GBS) remains a leading cause of serious neonatal infection despite great progress in perinatal GBS disease prevention in the 1990s. In 1996, CDC, in collaboration with other agencies, published guidelines for the prevention of perinatal group B streptococcal disease (CDC. Prevention of perinatal group B streptococcal disease: a public health perspective. MMWR 1996;45[RR-7]:1-24). Data collected after the issuance of the 1996 guidelines prompted reevaluation of prevention strategies at a meeting of clinical and public health representatives in November 2001. This report replaces CDC's 1996 guidelines. The recommendations are based on available evidence and expert opinion where sufficient evidence was lacking. Although many of the recommendations in the 2002 guidelines are the same as those in 1996, they include some key changes: * Recommendation of universal prenatal screening for vaginal and rectal GBS colonization of all pregnant women at 35-37 weeks' gestation, based on recent documentation in a large retrospective cohort study of a strong protective effect of this culture-based screening strategy relative to the risk-based strategy * Updated prophylaxis regimens for women with penicillin allergy * Detailed instruction on prenatal specimen collection and expanded methods of GBS culture processing, including instructions on antimicrobial susceptibility testing * Recommendation against routine intrapartum antibiotic prophylaxis for GBS-colonized women undergoing planned cesarean deliveries who have not begun labor or had rupture of membranes * A suggested algorithm for management of patients with threatened preterm delivery * An updated algorithm for management of newborns exposed to intrapartum antibiotic prophylaxis Although universal screening for GBS colonization is anticipated to result in further reductions in the burden of GBS disease, the need to monitor for potential adverse consequences of intrapartum antibiotic use, such as emergence of bacterial antimicrobial resistance or increased incidence or severity of non-GBS neonatal pathogens, continues, and intrapartum antibiotics are still viewed as an interim strategy until GBS vaccines achieve licensure.


Gram-scale production of recombinant microbial enzymes in shake flasks.
Heterologous production of recombinant proteins is a cornerstone of microbiological and biochemical research as well as various biotechnological processes. Yields and quality of produced proteins have a tremendous impact on structural and enzymology studies, development of new biopharmaceuticals and establishing new biocatalytic processes. Majority of current protocols for recombinant protein expression in Escherichia coli exploit batch cultures with complex media, often providing low yields of the target protein due to oxygen transfer limitation, rapid depletion of carbon sources and pH changes during the cultivation. Recently introduced EnBase technology enables fed-batch-like cultivations in shake flasks with continuous glucose release from a soluble starch. In this study, we critically compare the yields of fourteen model enzymes in E. coli cultured in a novel semi-defined medium and in a complex medium. Significant improvements of the volumetric yields 2-31 times were observed for all tested enzymes expressed in enzymatic fed-batch-like cultures with no adverse impact on enzyme structure, stability or activity. Exceptional yields, higher than 1 g of protein per liter of culture, were obtained with six enzymes. We conclude that the novel semi-defined medium tested in this study provides a robust improvement of protein yields in shake flasks without investment into costly bioreactors.


Links between bereavement due to sudden death and academic functioning: Results from a nationally representative sample of adolescents.
Bereavement due to sudden loss may disrupt positive adjustment among youth, yet few studies have examined the age at which youth are most likely to first encounter sudden loss, the co-occurrence of sudden loss with other traumatic events, and the independent effects of sudden loss on academic functioning. Data were analyzed from the National Comorbidity Survey-Adolescent Supplement (N = 10,148, Mage = 15.18, 51.1% female). Youth reported on whether they had experienced sudden loss (along with 17 other traumatic events), the age at which they had first experienced sudden loss, and multiple indicators of academic functioning. Sudden loss was the most frequently occurring traumatic event among youth; approximately 30% of adolescents reported at least one sudden loss in their lifetime. Youth were most likely to have first experienced sudden loss during middle adolescence (15 to 16 years of age). Although sudden loss co-occurred with several other traumas, about 10% of youth reported experiencing only sudden loss. After accounting for demographic characteristics and other traumatic events, experiencing sudden loss was associated with lower academic achievement, lower ability to concentrate and learn, less enjoyment of school, lower school belongingness, and lower beliefs that teachers treat youth fairly. Sudden loss is common among adolescents and has important implications for school functioning. Schools may improve academic functioning by adopting routine screening for sudden loss and assessing potential need for bereavement-informed mental health services. (PsycINFO Database Record


Treat to target in psoriatic arthritis-evidence, target, research agenda.
This article summarises the evidence on treating to target (T2T) in psoriatic arthritis (PsA). The evidence from the recent European League Against Rheumatism (EULAR) literature search in SpA is reviewed. This article found no studies in SpA in 2011 which compared a T2T approach against standard care in a randomised controlled trial (RCT). One of the issues raised in PsA is what target to use. The minimal disease activity (MDA) criteria were developed specifically for PsA and have been validated in observational and interventional cohorts. These have now been utilised in the Tight Control of PsA (TICOPA) study comparing T2T to standard care in early PsA providing the first evidence that T2T in PsA can improve outcomes. To translate this into clinical practice in PsA, patient education and feasibility are key. Future research must address the optimal therapies to be used within a T2T framework which will need to take subtype of PsA into account.


Time estimation during sleep relates to the amount of slow wave sleep in humans.
Humans have the ability to estimate the amount of time that has elapsed during sleep (time estimation ability; TEA) that enables a subset of individuals to wake up at a predetermined time without referring to a watch or alarm clock. Although previous studies have indicated sleep structure as a key factor that might influence TEA during sleep, which sleep parameters could affect the TEA has not been clarified. We carried out an experimental study in which 20 healthy volunteers participated in six time estimation trials during the 9-h nighttime sleep (NS) experiment or daytime sleep (DS) experiment. The time estimation ratio (TER, ratio of the subjective estimated time interval to actual time interval) decreased significantly from the first to the sixth trial in both the NS and DS experiments. TER correlated positively with slow wave sleep (SWS) in both experiments, suggesting that SWS was a determining factor in accurate time estimation, irrespective of circadian phase they slept. No other sleep parameters showed steady influence on TEA. The present findings demonstrate that longer period of SWS is associated with the longer sleep time they subjectively experienced during sleep.


Neuroprotective effect of a CNTF-expressing lentiviral vector in the quinolinic acid rat model of Huntington's disease.
Neurodegenerative diseases represent promising targets for gene therapy approaches provided effective transfer vectors. In the present study, we evaluated the effectiveness of LacZ-expressing lentiviral vectors with two different internal promoters, the mouse phosphoglycerate kinase 1 (PGK) and cytomegalovirus (CMV), to infect striatal cells. The intrastriatal injection of lenti-beta-Gal vectors lead to 207, 400 +/- 11,500 and 303,100 +/- 4,300 infected cells in adult rats, respectively. Importantly, the beta-galactosidase activity was higher in striatal extracts from PGK-LacZ-injected animals as compared to CMV-LacZ animals. The efficacy of the system was further examined with a potential therapeutic gene for the treatment of Huntington's disease, the human ciliary neurotrophic factor (CNTF). PGK-LacZ- or PGK-CNTF-expressing viruses were stereotaxically injected into the striatum of rats, 3 weeks later the animals were unilaterally lesioned with 180 nmol of quinolinic acid (QA). Control animals displayed 148 +/- 43 apomorphine-induced rotations ipsilateral to the lesion 5 days postlesion as compared to 26 +/- 22 turns/45 min in the CNTF-treated group. The extent of the striatal damage was significantly diminished in the CNTF-treated rats as indicated by the 52 +/- 9.7% decrease of the lesion volume and the sparing of DARPP-32, ChAT and NADPH-d neuronal populations. These results further establish that lentiviruses may represent an efficient gene delivery system for the screening of therapeutic molecules in Huntington's disease.


Age-associated accumulation of CMV-specific CD8+ T cells expressing the inhibitory killer cell lectin-like receptor G1 (KLRG1).
Large clonal expansions of peripheral CD8+ T cells carrying receptors for single epitopes of CMV and EBV are common in the elderly and may be associated with an immune risk phenotype predicting mortality. Here we show that the frequency of CD8+ T cells expressing the inhibitory killer cell lectin-like receptor G1 (KLRG1), a marker of cells unable to undergo further clonal expansion, was markedly elevated in CD8+ T cells from old donors. Moreover, tetramer staining revealed that the elevated frequency of CMV-specific CD8+ T cells in the elderly was due to an accumulation of cells bearing this dominant negative receptor. The fraction of CMV-specific T cells able to secrete interferon-gamma after specific antigenic stimulation was significantly lower in the elderly than in the young, although the total number of functional cells was comparable. Therefore, the majority of the clonally expanded virus-specific CD8+ cells in the elderly was dysfunctional. Thus, T cell responses are altered in the aged by an accumulation of replicatively senescent dysfunctional T cells carrying receptors for persistent herpes viruses. The presence of clonal expansions of such virus-specific cells may shrink the available repertoire for other antigens and contribute to the increased incidence of infectious disease in the elderly.


Spontaneous bacterial cell lysis and biofilm formation in the colon of the Cape Dune mole-rat and the laboratory rabbit.
A wide range of techniques, including high-throughput DNA sequencing methods, have been applied to the evaluation of the normal intestinal flora. However, the inability to grow many of those species in culture imposes substantial constraints on the techniques used to evaluate this important community. The presence of biofilms in the normal gut adds further complexity to the issue. In this study, a flow cytometric analysis was used to separate intact bacterial cells, cell debris, and other particulate matter based on bacteria-specific staining and particle size. In addition, an analysis of biofilm formation using fluorescent light microscopy was conducted. Using these approaches, the ratio of bacterial cell debris to intact bacterial cells as a measure of spontaneous lysis of bacterial cells in the gut of the Cape dune mole-rat (Bathyergus suillus) and the laboratory rabbit (Oryctolagus cuniculus) was examined, and the degree of biofilm formation was semi-quantitatively assessed. The results suggest that the degree of spontaneous cell lysis was greater in the appendix than in the cecum in both the mole-rat and the rabbit. Further, the results point toward extensive epithelial-associated biofilm formation in the proximal mole-rat and rabbit large bowel, although the biofilms may be less structured than those found in laboratory rodents and in humans.


Are we ready for a new epidemic of under recognized liver disease in South Asia especially in Pakistan? Non alcoholic fatty liver disease.
Nonalcoholic fatty liver disease (NAFLD) is increasingly recognized as an important public health problem nowadays. NAFLD encompass a variety of liver pathologies including simple steatosis, NASH, fibrosis, cirrhosis and finally cancer. It is associated with obesity, metabolic syndrome, dyslipidaemia, Insulin resistance (IR) and type 2 diabetes. It is the most common chronic liver disease in USA and considered to be increasing in Asia Pacific region including South Asia however there is no community based study from Pakistan. Customarily NAFLD had been regarded as a benign disease; however clinical as well as epidemiological studies had contradicted this belief because approximately 20% of the patients with NAFLD had NASH which has propensity to develop cirrhosis and ultimately to HCC. The diagnosis of NAFLD is made most of the times incidentally on abdominal imaging which is done for other purposes. Despite its prevalence, treatment options are very limited. However modification of risk factors such as dyslipidemia, diabetes control and weight reduction does help in NAFLD. Fatty liver results due to lack of physical activity; hence foremost step to manage such patients would be to develop the healthy life style. We need population based studies in our country so that we can protect our population from a new epidemic.


Diagnosis and Management of Atopic Dermatitis for Primary Care Providers.
To provide primary care providers an up-to-date approach to the diagnosis and management of atopic dermatitis (AD). PubMed, Cochrane, and MEDLINE databases were searched for relevant meta-analyses, randomized controlled trials, systematic reviews, and observational studies about the diagnosis and management of AD. AD is a common chronic skin disease characterized by immune dysregulation and skin barrier dysfunction. It has a substantial impact on quality of life across all ages. Despite this, AD is often still undertreated due to inaccurate diagnoses, even by many experienced dermatologists. Many primary care providers may not be aware of its commonly associated medical and psychiatric comorbidities or be familiar with new treatment options. In this clinical review, we describe the pathophysiology, epidemiology, diagnosis, and treatment of AD, specifically highlighting commonly used therapies and novel medications.


Online schools and children with special health and educational needs: comparison with performance in traditional schools.
In the United States, primary and secondary online schools are institutions that deliver online curricula for children enrolled in kindergarten through 12th grade (K-12). These institutions commonly provide opportunities for online instruction in conjunction with local schools for students who may need remediation, have advanced needs, encounter unqualified local instructors, or experience scheduling conflicts. Internet-based online schooling may potentially help children from populations known to have educational and health disadvantages, such as those from certain racial or ethnic backgrounds, those of low socioeconomic status, and children with special health care needs (CSHCN). To describe the basic and applied demographics of US online-school users and to compare student achievement in traditional versus online schooling environments. We performed a brief parental survey in three states examining basic demographics and educational history of the child and parents, the child's health status as measured by the CSHCN Screener, and their experiences and educational achievement with online schools and class(es). Results were compared with state public-school demographics and statistical analyses controlled for state-specific independence. We analyzed responses from 1971 parents with a response rate of 14.7% (1971/13,384). Parents of online-school participants were more likely to report having a bachelor's degree or higher than were parents of students statewide in traditional schools, and more of their children were white and female. Most notably, the prevalence of CSHCN was high (476/1971, 24.6%) in online schooling. Children who were male, black, or had special health care needs reported significantly lower grades in both traditional and online schools. However, when we controlled for age, gender, race, and parental education, parents of CSHCN or black children reported significantly lower grades in online than in traditional schooling (adjusted odds ratio [aOR] 1.45, 95% confidence interval [CI] 1.29-1.62 for CSHCN, P < .001; aOR 2.73, 95% CI 2.11-3.53 for black children, P < .001.) In contrast, parents with a bachelor's degree or higher reported significantly higher online-school grades than traditional-school grades for their children (aOR 1.45, 95% CI 1.15-1.82, P < .001). The demographics of children attending online schools do not mirror those of the state-specific school populations. CSHCN seem to opt into online schools at a higher rate. While parents report equivalent educational achievement in online and traditional classrooms, controlling for known achievement risks suggests that CSHCN and black children have lower performance in online than in traditional schools. Given the millions of students now in online schools, future studies must test whether direct assistance in online schools, such as taking individualized education plans into consideration, will narrow known disparities in educational success. Only then can online schools emerge as a true educational alternative for at-risk populations.


Identification and physiological activity of survival factor released from cardiomyocytes during ischaemia and reperfusion.
We carried out a screening of survival factors released from cells exposed to simulated ischaemia and reperfusion (sI/R) using the embryonic rat heart-derived cell line, H9c2 cells, and examined the physiological role of the identified factor. The culture medium supernatant of H9c2 cells exposed to sI/R was separated by column chromatography and the fractions examined for survival activity. The protein with survival activity was identified by mass spectrometry, and its physiological role was examined in the models of ischaemia. Cell survival activity was detected in at least three fractions of the cell supernatant collected during sI/R and subjected to a series of column chromatographic steps. Among the proteins measured by mass spectrometry and western blotting, a p36 protein identified as a glycolytic enzyme, lactate dehydrogenase muscle subunit (M-LDH), showed strong survival activity. H(2)O(2)-induced intracellular calcium overload in H9c2 cells and irregular Ca(2+) transients in adult rat cardiomyocytes were both found to be inhibited by pretreatment with M-LDH. M-LDH also lowered the frequency and amplitude of early afterdepolarizations induced by H(2)O(2) in adult rat cardiomyocytes and suppressed the ischaemia-reperfusion-induced reduction of cardiac output from mouse working heart preparations. M-LDH was found to increase the phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2), which plays a role in H9c2 cell survival. M-LDH released from cardiomyocytes after hypoxia and reoxygenation has a role in protecting the heart from oxidative stress-induced injury through an intracellular signal transduction pathway involving ERK1/2.


In situ resistance, not immigration, supports invertebrate community resilience to drought intensification in a Neotropical ecosystem.
While future climate scenarios predict declines in precipitations in many regions of the world, little is known of the mechanisms underlying community resilience to prolonged dry seasons, especially in 'naïve' Neotropical rainforests. Predictions of community resilience to intensifying drought are complicated by the fact that the underlying mechanisms are mediated by species' tolerance and resistance traits, as well as rescue through dispersal from source patches. We examined the contribution of in situ tolerance-resistance and immigration to community resilience, following drought events that ranged from the ambient norm to IPCC scenarios and extreme events. We used rainshelters above rainwater-filled bromeliads of French Guiana to emulate a gradient of drought intensity (from 1 to 3.6 times the current number of consecutive days without rainfall), and we analysed the post-drought dynamics of the taxonomic and functional community structure of aquatic invertebrates to these treatments when immigration is excluded (by netting bromeliads) or permitted (no nets). Drought intensity negatively affected invertebrate community resistance, but had a positive influence on community recovery during the post-drought phase. After droughts of 1 to 1.4 times the current intensities, the overall invertebrate abundance recovered within invertebrate life cycle durations (up to 2 months). Shifts in taxonomic composition were more important after longer droughts, but overall, community composition showed recovery towards baseline states. The non-random patterns of changes in functional community structure indicated that deterministic processes like environmental filtering of traits drive community re-assembly patterns after a drought event. Community resilience mostly relied on in situ tolerance-resistance traits. A rescue effect of immigration after a drought event was weak and mostly apparent under extreme droughts. Under climate change scenarios of drought intensification in Neotropical regions, community and ecosystem resilience could primarily depend on the persistence of suitable habitats and on the resistance traits of species, while metacommunity dynamics could make a minor contribution to ecosystem recovery. Climate change adaptation should thus aim at identifying and preserving local conditions that foster in situ resistance and the buffering effects of habitat features.


Cigarette smoke extract induces COX-2 expression via a PKCalpha/c-Src/EGFR, PDGFR/PI3K/Akt/NF-kappaB pathway and p300 in tracheal smooth muscle cells.
Exposure to cigarette smoke extract (CSE) leads to airway or lung inflammation, which may be mediated through cyclooxygenase-2 (COX-2) expression and its product prostaglandin E2 (PGE2) synthesis. The aim of this study was to investigate the molecular mechanisms underlying CSE-induced COX-2 expression in human tracheal smooth muscle cells (HTSMCs). Here, we describe that COX-2 induction is dependent on PKCalpha/c-Src/EGFR, PDGFR/PI3K/Akt/NF-kappaB signaling in HTSMCs. CSE stimulated the phosphorylation of c-Src, EGFR, PDGFR, and Akt, which were inhibited by pretreatment with the inhibitor of PKCalpha (Gö6976 or Gö6983), c-Src (PP1), EGFR (AG1478), PDGFR (AG1296), or PI3K (LY294002). Moreover, CSE induced a significant increase in COX-2 expression, which was reduced by pretreatment with these inhibitors or transfection with siRNA of PKCalpha, Src, or Akt. Furthermore, CSE-stimulated NF-kappaB p65 phosphorylation and translocation were also attenuated by pretreatment with Gö6976, PP1, AG1478, AG1296, or LY294002. CSE-induced COX-2 expression was also mediated through the recruitment of p300 associated with NF-kappaB in HTSMCs, revealed by coimmunoprecipitation and Western blot analysis. In addition, pretreatment with the inhibitors of NF-kappaB (helenalin) and p300 (garcinol) or transfection with p65 siRNA and p300 siRNA markedly inhibited CSE-regulated COX-2 expression. However, CSE-induced PGE2 generation was reduced by pretreatment with the inhibitor of COX-2 (NS-398). These results demonstrated that in HTSMCs, CSE-induced COX-2-dependent PGE2 generation was mediated through PKCalpha/c-Src/EGFR, PDGFR/PI3K/Akt leading to the recruitment of p300 with NF-kappaB complex.


From Flexner to competencies: reflections on a decade and the journey ahead.
This article is a sequel to one published in 2002 only a few years after the initiation of the shift to competency-based medical education (CBME). The authors reflect on the major forces that have influenced the movement and tipped the balance toward widespread adoption of CBME in the United States, primarily in graduate medical education. These forces include regulatory bodies, international counterparts, and the general public. The authors highlight the most important lessons learned over the decade. These include (1) the need for standardization of language to develop a shared vision of the path ahead, (2) the power of direct observation in assessment, (3) the challenge of developing meaningful measures of performance, (4) desired outcomes as the starting point for curriculum development, (5) dependence on reflection in the development of expertise, (6) the need for exploiting the role of learners in their learning, and (7) competent clinical systems as the required learning environment for producing competent physicians.The authors speculate on why this most recent attempt to shift to CBME differs from previous aborted attempts. They conclude by explaining how the recent lessons learned inform the vision of what successful implementation of CBME would look like, and discussing the importance of milestones, entrustable professional activities, and an integrated, rather than a reductionist, approach to assessment of competence. The fundamental question at each step along the way in implementing CBME should be "How do we improve medical education to provide better care for patients?"


TnSeq of Mycobacterium tuberculosis clinical isolates reveals strain-specific antibiotic liabilities.
Once considered a phenotypically monomorphic bacterium, there is a growing body of work demonstrating heterogeneity among Mycobacterium tuberculosis (Mtb) strains in clinically relevant characteristics, including virulence and response to antibiotics. However, the genetic and molecular basis for most phenotypic differences among Mtb strains remains unknown. To investigate the basis of strain variation in Mtb, we performed genome-wide transposon mutagenesis coupled with next-generation sequencing (TnSeq) for a panel of Mtb clinical isolates and the reference strain H37Rv to compare genetic requirements for in vitro growth across these strains. We developed an analytic approach to identify quantitative differences in genetic requirements between these genetically diverse strains, which vary in genomic structure and gene content. Using this methodology, we found differences between strains in their requirements for genes involved in fundamental cellular processes, including redox homeostasis and central carbon metabolism. Among the genes with differential requirements were katG, which encodes the activator of the first-line antitubercular agent isoniazid, and glcB, which encodes malate synthase, the target of a novel small-molecule inhibitor. Differences among strains in their requirement for katG and glcB predicted differences in their response to these antimicrobial agents. Importantly, these strain-specific differences in antibiotic response could not be predicted by genetic variants identified through whole genome sequencing or by gene expression analysis. Our results provide novel insight into the basis of variation among Mtb strains and demonstrate that TnSeq is a scalable method to predict clinically important phenotypic differences among Mtb strains.


Biochemical properties and base excision repair complex formation of apurinic/apyrimidinic endonuclease from Pyrococcus furiosus.
Apurinic/apyrimidinic (AP) sites are the most frequently found mutagenic lesions in DNA, and they arise mainly from spontaneous base loss or modified base removal by damage-specific DNA glycosylases. AP sites are cleaved by AP endonucleases, and the resultant gaps in the DNA are repaired by DNA polymerase/DNA ligase reactions. We identified the gene product that is responsible for the AP endonuclease activity in the hyperthermophilic euryarchaeon, Pyrococcus furiosus. Furthermore, we detected the physical interaction between P. furiosus AP endonuclease (PfuAPE) and proliferating cell nuclear antigen (PCNA; PfuPCNA) by a pull-down assay and a surface plasmon resonance analysis. Interestingly, the associated 3'-5' exonuclease activity, but not the AP endonuclease activity, of PfuAPE was stimulated by PfuPCNA. Immunoprecipitation experiments using the P. furiosus cell extracts supported the interaction between PfuAPE and PfuPCNA in the cells. This is the first report describing the physical and functional interactions between an archaeal AP endonuclease and PCNA. We also detected the ternary complex of PfuPCNA, PfuAPE and Pfu uracil-DNA glycosylase. This complex probably functions to enhance the repair of uracil-containing DNA in P. furiosus cells.


Reduced resuscitation fluid volume for second-degree burns with delayed initiation of ascorbic acid therapy.
Oxygen radicals may play an important role in injury due to thermal burns. High-dose antioxidant ascorbic acid (vitamin C, Cenolete, Abbott Laboratory, Abbott Park, Ill) therapy reduces edema of burned and unburned tissue, lipid peroxidation, and subsequent resuscitation fluid volume requirement in experimental burn models. To determine the hemodynamic effects of delayed initiation (2 hours after injury) of antioxidant therapy in patients with second-degree burns. Experimental study. MATERIALS AND INTERVENTION: Burns over 70% of body surface area were produced by subxiphoid immersion of 12 guinea pigs in 100 degrees C water for 3 seconds. The animals were resuscitated with Ringer's lactated solution according to the Parkland formula (4 mL/kg for 1% of burned body surface area during the first 24 hours) from 0.5 to 2 hours following injury, after which the resuscitation fluid volume was reduced to 25% of the Parkland formula. Animals received Ringer's lactated solution to which ascorbic acid (340 mg/kg during the first 24 hours) was added (vitamin C group [n = 6]) or Ringer's lactated solution only (control group [n = 6]). Heart rates, mean arterial blood pressure, cardiac output, hematocrit level, and water content in burned and unburned tissue were measured before injury and at intervals thereafter. There were no significant differences in heart rates (P = .29) and blood pressures (P = .53) between the 2 groups throughout the 24-hour study period. No animal died. The vitamin C group showed significantly lower hematocrit levels (P < .05) and significantly higher cardiac output values (P < .05) at 7 hours following burn injury and at intervals thereafter (P < .001). With delayed initiation of high-dose ascorbic acid therapy, the 24-hour fluid resuscitation volume was reduced to 32.5% of the Parkland formula, while maintaining adequate cardiac output values.


Both the establishment and maintenance of neuronal polarity require the activity of protein kinase D in the Golgi apparatus.
Neuronal polarization requires coordinated regulation of membrane trafficking and cytoskeletal dynamics. Several signaling proteins are involved in neuronal polarization via modulation of cytoskeletal dynamics in neurites. However, very little is known about signaling proteins in the neuronal soma, which regulate polarized membrane trafficking and neuronal polarization. Protein kinase D (PKD) constitutes a family of serine/threonine-specific protein kinases and is important in regulating Golgi dynamics and membrane trafficking. Here, we show that two members of the PKD family, PKD1 and PKD2, are essential for the establishment and maintenance of neuronal polarity. Loss of function of PKD with inhibitor, dominant negative, and short interfering RNA disrupts polarized membrane trafficking and induces multiple axon formation. Gain of function of PKD can rescue the disruption of polarized membrane trafficking and neuronal polarity caused by cytochalasin D, which results in F-actin depolymerization. PKD1 and PKD2 are also found to be involved in the maintenance of neuronal polarity, as evidenced by the conversion of preexisting dendrites into axons on PKD inhibition. Unlike other polarity proteins, PKD does not interact with the cytoskeleton in neurites. Instead, PKD regulates neuronal polarity through its activity in the Golgi apparatus. These data reveal a novel mechanism regulating neuronal polarity in the Golgi apparatus.


Differential effectiveness of microbially induced resistance against herbivorous insects in Arabidopsis.
Rhizobacteria-induced systemic resistance (ISR) and pathogen-induced systemic acquired resistance (SAR) have a broad, yet partly distinct, range of effectiveness against pathogenic microorganisms. Here, we investigated the effectiveness of ISR and SAR in Arabidopsis against the tissue-chewing insects Pieris rapae and Spodoptera exigua. Resistance against insects consists of direct defense, such as the production of toxins and feeding deterrents and indirect defense such as the production of plant volatiles that attract carnivorous enemies of the herbivores. Wind-tunnel experiments revealed that ISR and SAR did not affect herbivore-induced attraction of the parasitic wasp Cotesia rubecula (indirect defense). By contrast, ISR and SAR significantly reduced growth and development of the generalist herbivore S. exigua, although not that of the specialist P. rapae. This enhanced direct defense against S. exigua was associated with potentiated expression of the defense-related genes PDF1.2 and HEL. Expression profiling using a dedicated cDNA microarray revealed four additional, differentially primed genes in microbially induced S. exigua-challenged plants, three of which encode a lipid-transfer protein. Together, these results indicate that microbially induced plants are differentially primed for enhanced insect-responsive gene expression that is associated with increased direct defense against the generalist S. exigua but not against the specialist P. rapae.