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1.  Comparison of Markers and Functional Attributes of Human Adipose-Derived Stem Cells and Dedifferentiated Adipocyte Cells from Subcutaneous Fat of an Obese Diabetic Donor 
Advances in Wound Care  2014;3(3):219-228.
Objective: Adipose tissue is a robust source of adipose-derived stem cells (ADSCs) that may be able to provide secreted factors that promote the ability of wounded tissue to heal. However, adipocytes also have the potential to dedifferentiate in culture to cells with stem cell-like properties that may improve their behavior and functionality for certain applications.
Approach: ADSCs are adult mesenchymal stem cells that are cultured from the stromal vascular fraction of adipose tissue. However, adipocytes are capable of dedifferentiating into cells with stem cell properties. In this case study, we compare ADSC and dedifferentiated fat (DFAT) cells from the same patient and fat depot for mesenchymal cell markers, embryonic stem cell markers, ability to differentiate to adipocytes and osteoblasts, senescence and telomerase levels, and ability of conditioned media (CM) to stimulate migration of human dermal fibroblasts (HDFs).
Innovation and Conclusions: ADSCs and DFAT cells displayed identical levels of CD90, CD44, CD105, and were CD34- and CD45-negative. They also expressed similar levels of Oct4, BMI1, KLF4, and SALL4. DFAT cells, however, showed higher efficiency in adipogenic and osteogenic capacity. Telomerase levels of DFAT cells were double those of ADSCs, and senescence declined in DFAT cells. CM from both cell types altered the migration of fibroblasts. Despite reports of ADSCs from a number of human depots, there have been no comparisons of the ability of dedifferentiated DFAT cells from the same donor and depot to differentiate or modulate migration of HDFs. Since ADSCs were from an obese diabetic donor, reprogramming of DFAT cells may help improve a patient's cells for regenerative medicine applications.
doi:10.1089/wound.2013.0452
PMCID: PMC3955970  PMID: 24669358
2.  Phase II Trial of Neoadjuvant/adjuvant Imatinib Mesylate for Advanced Primary and Metastatic/recurrent Operable Gastrointestinal Stromal Tumors: Long-term Follow-up Results of Radiation Therapy Oncology Group 0132 
Annals of surgical oncology  2011;19(4):1074-1080.
Background
Imatinib inhibits the KIT and PDGFR tyrosine kinases, resulting in its notable antitumor activity in gastrointestinal stromal tumor (GIST). We previously reported the early results of a multi-institutional prospective trial (RTOG 0132) using neoadjuvant/adjuvant imatinib either in primary resectable GIST or as a planned preoperative cytoreduction agent for metastatic/recurrent GIST.
Methods
Patients with primary GIST (≥5 cm, group A) or resectable metastatic/recurrent GIST (≥2 cm, group B) received neoadjuvant imatinib (600 mg/day) for approximately 2 months and maintenance postoperative imatinib for 2 years. We have now updated the clinical outcomes including progression-free survival, disease-specific survival, and overall survival at a median follow-up of 5.1 years, and we correlate these end points with duration of imatinib therapy.
Results
Sixty-three patients were originally entered (53 analyzable: 31 in group A and 22 in group B). Estimated 5-year progression-free survival and overall survival were 57% in group A, 30% in group B; and 77% in group A, 68% in group B, respectively. Median time to progression has not been reached for group A and was 4.4 years for group B. In group A, in 7 of 11 patients, disease progressed >2 years from registration; 6 of 7 patients with progression had stopped imatinib before progression. In group B, disease progressed in 10 of 13 patients >2 years from registration; 6 of 10 patients with progressing disease had stopped imatinib before progression. There was no significant increase in toxicity compared with our previous shortterm analysis.
Conclusions
This long-term analysis suggests a high percentage of patients experienced disease progression after discontinuation of 2-year maintenance imatinib therapy after surgery. Consideration should be given to studying longer treatment durations in intermediate- to high-risk GIST patients.
doi:10.1245/s10434-011-2190-5
PMCID: PMC3800166  PMID: 22203182
3.  Identification of the Plasmodium berghei resistance locus 9 linked to survival on chromosome 9 
Malaria Journal  2013;12:316.
Background
One of the main causes of mortality from severe malaria in Plasmodium falciparum infections is cerebral malaria (CM). An important host genetic component determines the susceptibility of an individual to develop CM or to clear the infection and become semi-immune. As such, the identification of genetic loci associated with susceptibility or resistance may serve to modulate disease severity.
Methodology
The Plasmodium berghei mouse model for experimental cerebral malaria (ECM) reproduces several disease symptoms seen in human CM, and two different phenotypes, a susceptible (FVB/NJ) and a resistant mouse strain (DBA/2J), were examined.
Results
FVB/NJ mice died from infection within ten days, whereas DBA/2J mice showed a gender bias: males survived on average nineteen days and females either died early with signs of ECM or survived for up to three weeks. A comparison of brain pathology between FVB/NJ and DBA/2J showed no major differences with regard to brain haemorrhages or the number of parasites and CD3+ cells in the microvasculature. However, significant differences were found in the peripheral blood of infected mice: For example resistant DBA/2J mice had significantly higher numbers of circulating basophils than did FVB/NJ mice on day seven. Analysis of the F2 offspring from a cross of DBA/2J and FVB/NJ mice mapped the genetic locus of the underlying survival trait to chromosome 9 with a Lod score of 4.9. This locus overlaps with two previously identified resistance loci (char1 and pymr) from a blood stage malaria model.
Conclusions
Survival best distinguishes malaria infections between FVB/NJ and DBA/2J mice. The importance of char1 and pymr on chromosome 9 in malaria resistance to P. berghei was confirmed. In addition there was an association of basophil numbers with survival.
doi:10.1186/1475-2875-12-316
PMCID: PMC3848760  PMID: 24025732
Plasmodium berghei; Experimental cerebral malaria; Quantitative trait locus; Malaria; Basophil; Chromosome mapping; Mouse
4.  The Nucleus Accumbens as a Potential Target for Central Poststroke Pain 
Mayo Clinic Proceedings  2012;87(10):1025-1031.
Although deep brain stimulation (DBS) has been found to be efficacious for some chronic pain syndromes, its usefulness in patients with central poststroke pain (CPSP) has been disappointing. The most common DBS targets for pain are the periventricular gray region (PVG) and the ventralis caudalis of the thalamus. Despite the limited success of DBS for CPSP, few alternative targets have been explored. The nucleus accumbens (NAC), a limbic structure within the ventral striatum that is involved in reward and pain processing, has emerged as an effective target for psychiatric disease. There is also evidence that it may be an effective target for pain. We describe a 72-year-old woman with a large right hemisphere infarct who subsequently experienced refractory left hemibody pain. She underwent placement of 3 electrodes in the right PVG, ventralis caudalis of the thalamus, and NAC. Individual stimulation of the NAC and PVG provided substantial improvement in pain rating. The patient underwent implantation of permanent electrodes in both targets, and combined stimulation has provided sustained pain relief at nearly 1 year after the procedure. These results suggest that the NAC may be an effective DBS target for CPSP.
doi:10.1016/j.mayocp.2012.02.029
PMCID: PMC3498057  PMID: 22980165
CPSP, central poststroke pain; DBS, deep brain stimulation; ECT, electroconvulive therapy; ICL, intercommissural line; MCS, motor cortex stimulation; NAC, nucleus accumbens; PFC, prefrontal cortex; PVG, periventricular gray region; VC, ventralis caudalis of the thalamus
5.  RTOG 0247: A Randomized Phase II Study of Neoadjuvant Capecitabine and Irinotecan or Capecitabine and Oxaliplatin with Concurrent Radiation Therapy for Patients with Locally Advanced Rectal Cancer 
Purpose
To evaluate rate of pathologic complete response (pCR) and toxicity of two neoadjuvant chemoradiation (chemoRT) regimens for T3/T4 rectal cancer in a randomized phase II study.
Methods and Materials
Patients with T3 or T4 rectal cancer < 12 cm from the anal verge were randomized to preoperative RT (50.4 Gy in 1.8 Gy fractions) with (1) concurrent capecitabine (1200 mg/m2/d M-F) and irinotecan (50 mg/m2 weekly × 4 doses) (arm 1), or (2) concurrent capecitabine (1650 mg/m2/d M-F) and oxaliplatin (50 mg/m2 weekly × 5 doses) (arm 2). Surgery was performed 4–8 weeks after chemoRT, and adjuvant chemotherapy 4–6 weeks after surgery. The primary endpoint was pCR rate, requiring 48 evaluable patients per arm.
Results
146 patients were enrolled. Protocol chemotherapy was modified due to excessive GI toxicity after treatment of 35 patients; 96 were assessed for the primary endpoint—final regimen described above. Patient characteristics were similar for both arms. Following chemoRT, tumor downstaging was 52% and 60%, and nodal downstaging (excluding N0 patients) was 46% and 40%, for arms 1 and 2, respectively. The pCR rate for arm 1 was 10% and for arm 2 was 21%. For arms 1 and 2, respectively, preop chemoRT grade 3/4 hematologic toxicity was 9% and 4%, and grade 3/4 non-hematologic toxicity was 26% and 27%.
Conclusions
Preoperative chemoRT with capecitabine plus oxaliplatin for distal rectal cancer has significant clinical activity (10/48 pCRs) and acceptable toxicity. This regimen is currently being evaluated in a phase III randomized trial (NSABP R04).
doi:10.1016/j.ijrobp.2011.05.027
PMCID: PMC3208721  PMID: 21775070
Neoadjuvant; chemotherapy; radiation; rectal; cancer
6.  Clk/STY (cdc2-Like Kinase 1) and Akt Regulate Alternative Splicing and Adipogenesis in 3T3-L1 Pre-Adipocytes 
PLoS ONE  2013;8(1):e53268.
The development of adipocytes from their progenitor cells requires the action of growth factors signaling to transcription factors to induce the expression of adipogenic proteins leading to the accumulation of lipid droplets, induction of glucose transport, and secretion of adipokines signaling metabolic events throughout the body. Murine 3T3-L1 pre-adipocytes sequentially express all the proteins necessary to become mature adipocytes throughout an 8–10 day process initiated by a cocktail of hormones. We examined the role of Clk/STY or Clk1, a cdc2-like kinase, in adipogenesis since it is known to be regulated by Akt, a pivotal kinase in development. Inhibition of Clk1 by a specific inhibitor, TG003, blocked alternative splicing of PKCβII and expression of PPARγ1 and PPARγ2. SiRNA depletion of Clk1 resulted in early expression of PKCβII and sustained PKCβI expression. Since Clk1 is a preferred Akt substrate, required for phosphorylation of splicing factors, mutation of Clk1 Akt phosphorylation sites was undertaken. Akt sites on Clk1 are in the serine/arginine-rich domain and not the kinase domain. Mutation of single and multiple sites resulted in dysregulation of PKCβII, PKCβI, and PPARγ1&2 expression. Additionally, adipogenesis was blocked as assessed by Oil Red O staining, adiponectin, and Glut1 and 4 expression. Immunofluorescence microscopy revealed that Clk1 triple mutant cDNA, transfected into pre-adipocytes, resulted in excluding SRp40 (SFSR6) from co-localizing to the nucleus with PFS, a perispeckle specific protein. This study demonstrates the role of Akt and Clk1 kinases in the early differentiation of 3T3-L1 cells to adipocytes.
doi:10.1371/journal.pone.0053268
PMCID: PMC3537621  PMID: 23308182
7.  Housing Characteristics and their Influence on Health-Related Quality of Life in Persons Living with HIV in Ontario, Canada: Results from the Positive Spaces, Healthy Places Study 
AIDS and Behavior  2012;16(8):2361-2373.
Although lack of housing is linked with adverse health outcomes, little is known about the impacts of the qualitative aspects of housing on health. This study examined the association between structural elements of housing, housing affordability, housing satisfaction and health-related quality of life over a 1-year period. Participants were 509 individuals living with HIV in Ontario, Canada. Regression analyses were conducted to examine relationships between housing variables and physical and mental health-related quality of life. We found significant cross-sectional associations between housing and neighborhood variables—including place of residence, housing affordability, housing stability, and satisfaction with material, meaningful and spatial dimensions of housing—and both physical and mental health-related quality of life. Our analyses also revealed longitudinal associations between housing and neighborhood variables and health-related quality of life. Interventions that enhance housing affordability and housing satisfaction may help improve health-related quality of life of people living with HIV.
doi:10.1007/s10461-012-0284-0
PMCID: PMC3481053  PMID: 22903401
Housing; Housing affordability; Housing satisfaction; Health-related quality of life; HIV
8.  Target Selection and Determination of Function in Structural Genomics 
Iubmb Life  2003;55(4-5):249-255.
Summary
The first crucial step in any structural genomics project is the selection and prioritization of target proteins for structure determination. There may be a number of selection criteria to be satisfied, including that the proteins have novel folds, that they be representatives of large families for which no structure is known, and so on. The better the selection at this stage, the greater is the value of the structures obtained at the end of the experimental process. This value can be further enhanced once the protein structures have been solved if the functions of the given proteins can also be determined. Here we describe the methods used at either end of the experimental process: firstly, sensitive sequence comparison techniques for selecting a high-quality list of target proteins, and secondly the various computational methods that can be applied to the eventual 3D structures to determine the most likely biochemical function of the proteins in question.
doi:10.1080/1521654031000123385
PMCID: PMC3366504  PMID: 12880206
Structural genomics; target selection; function from structure; functional annotation
9.  CXCR4 Expression in Prostate Cancer Progenitor Cells 
PLoS ONE  2012;7(2):e31226.
Tumor progenitor cells represent a population of drug-resistant cells that can survive conventional chemotherapy and lead to tumor relapse. However, little is known of the role of tumor progenitors in prostate cancer metastasis. The studies reported herein show that the CXCR4/CXCL12 axis, a key regulator of tumor dissemination, plays a role in the maintenance of prostate cancer stem-like cells. The CXCL4/CXCR12 pathway is activated in the CD44+/CD133+ prostate progenitor population and affects differentiation potential, cell adhesion, clonal growth and tumorigenicity. Furthermore, prostate tumor xenograft studies in mice showed that a combination of the CXCR4 receptor antagonist AMD3100, which targets prostate cancer stem-like cells, and the conventional chemotherapeutic drug Taxotere, which targets the bulk tumor, is significantly more effective in eradicating tumors as compared to monotherapy.
doi:10.1371/journal.pone.0031226
PMCID: PMC3281066  PMID: 22359577
10.  Phase II and Coagulation Cascade Biomarker Study of Bevacizumab with or without Docetaxel in Patients with Previously Treated Metastatic Pancreatic Adenocarcinoma 
Purpose
Treatment options are limited for advanced pancreatic cancer progressive after gemcitabine therapy. The vascular endothelial growth factor (VEGF) pathway is biologically important in pancreatic cancer, and docetaxel has modest anti-tumor activity. We evaluated the role of the anti-VEGF antibody bevacizumab as second-line treatment for patients with metastatic pancreatic cancer.
Design
Patients with metastatic adenocarcinoma of the pancreas who had progressive disease on a gemcitabine-containing regimen were randomized to receive bevacizumab alone or bevacizumab in combination with docetaxel.
Results
Thirty-two patients were enrolled; 16 to bevacizumab alone (Arm A) and 16 to bevacizumab plus docetaxel (Arm B). Toxicities were greater in Arm B with the most common grade 3/4 nonhematologic toxicities including fatigue, diarrhea, dehydration and anorexia. No confirmed objective responses were observed. At 4 months, 2/16 patients in Arm A and 3/16 in Arm B were free from progression. The study was stopped according to the early stopping rule for futility. Median PFS and OS were 43 days and 165 days in Arm A and 48 days and 125 days in Arm B. Elevated D-dimer levels and thrombin-antithrombin complexes were associated with decreased survival and increased toxicity.
Conclusion
Bevacizumab with or without docetaxel does not have antitumor activity in gemcitabine-refractory metastatic pancreatic cancer. Baseline and on-treatment D-dimer and thrombin-antithrombin complex levels are associated with increased toxicity and decreased survival.
doi:10.1097/COC.0b013e3181d2734a
PMCID: PMC3030655  PMID: 20458210
11.  Developing an algorithm for pulse oximetry derived respiratory rate (RRoxi): a healthy volunteer study 
Objective The presence of respiratory information within the pulse oximeter signal (PPG) is a well-documented phenomenon. However, extracting this information for the purpose of continuously monitoring respiratory rate requires: (1) the recognition of the multi-faceted manifestations of respiratory modulation components within the PPG and the complex interactions among them; (2) the implementation of appropriate advanced signal processing techniques to take full advantage of this information; and (3) the post-processing infrastructure to deliver a clinically useful reported respiratory rate to the end user. A holistic algorithmic approach to the problem is therefore required. We have developed the RROXI algorithm based on this principle and its performance on healthy subject trial data is described herein.
Methods Finger PPGs were collected from a cohort of 139 healthy adult volunteers monitored during free breathing over an 8-min period. These were subsequently processed using a novel in-house algorithm based on continuous wavelet transform technology within an infrastructure incorporating weighted averaging and logical decision making processes. The computed oximeter respiratory rates (RRoxi) were then compared to an end-tidal CO2 reference rate (\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ {\text{RR}}_{{{\text{ETCO}}_{ 2} }} $$\end{document}).
Results\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ {\text{RR}}_{{{\text{ETCO}}_{ 2} }} $$\end{document} ranged from a lowest recorded value of 2.97 breaths per min (br/min) to a highest value of 28.02 br/min. The mean rate was 14.49 br/min with standard deviation of 4.36 br/min. Excellent agreement was found between RRoxi and \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$ {\text{RR}}_{{{\text{ETCO}}_{ 2} }} $$\end{document}, with a mean difference of −0.23 br/min and standard deviation of 1.14 br/min. The two measures are tightly spread around the line of agreement with a strong correlation observable between them (R2 = 0.93).
Conclusions These data indicate that RRoxi represents a viable technology for the measurement of respiratory rate of healthy individuals.
doi:10.1007/s10877-011-9332-y
PMCID: PMC3268017  PMID: 22231359
Respiratory rate; Pulse oximeter; Continuous monitoring
12.  Bioinformatics Training Network (BTN): a community resource for bioinformatics trainers 
Briefings in Bioinformatics  2011;13(3):383-389.
Funding bodies are increasingly recognizing the need to provide graduates and researchers with access to short intensive courses in a variety of disciplines, in order both to improve the general skills base and to provide solid foundations on which researchers may build their careers. In response to the development of ‘high-throughput biology’, the need for training in the field of bioinformatics, in particular, is seeing a resurgence: it has been defined as a key priority by many Institutions and research programmes and is now an important component of many grant proposals. Nevertheless, when it comes to planning and preparing to meet such training needs, tension arises between the reward structures that predominate in the scientific community which compel individuals to publish or perish, and the time that must be devoted to the design, delivery and maintenance of high-quality training materials. Conversely, there is much relevant teaching material and training expertise available worldwide that, were it properly organized, could be exploited by anyone who needs to provide training or needs to set up a new course. To do this, however, the materials would have to be centralized in a database and clearly tagged in relation to target audiences, learning objectives, etc. Ideally, they would also be peer reviewed, and easily and efficiently accessible for downloading. Here, we present the Bioinformatics Training Network (BTN), a new enterprise that has been initiated to address these needs and review it, respectively, to similar initiatives and collections.
doi:10.1093/bib/bbr064
PMCID: PMC3357490  PMID: 22110242
Bioinformatics; training; end users; bioinformatics courses; learning bioinformatics
13.  Efficient Use and Recycling of the Micronutrient Iodide in Mammals 
Biochimie  2010;92(9):1227-1235.
Daily ingestion of iodide alone is not adequate to sustain production of the thyroid hormones, tri- and tetraiodothyronine. Proper maintenance of iodide in vivo also requires its active transport into the thyroid and its salvage from mono- and diiodotyrosine that are formed in excess during hormone biosynthesis. The enzyme iodotyrosine deiodinase responsible for this salvage is unusual in its ability to catalyze a reductive dehalogenation reaction dependent on a flavin cofactor, FMN. Initial characterization of this enzyme was limited by its membrane association, difficult purification and poor stability. The deiodinase became amenable to detailed analysis only after identification and heterologous expression of its gene. Site-directed mutagenesis recently demonstrated that cysteine residues are not necessary for enzymatic activity in contrast to precedence set by other reductive dehalogenases. Truncation of the N-terminal membrane anchor of the deiodinase has provided a soluble and stable source of enzyme sufficient for crystallographic studies. The structure of an enzyme•substrate co-crystal has become invaluable for understanding the origins of substrate selectivity and the mutations causing thyroid disease in humans.
doi:10.1016/j.biochi.2010.02.013
PMCID: PMC2888766  PMID: 20167242
Deiodinase; Flavoprotein; Reductive Dehalogenation; Iodide Metabolism; Thyroid
14.  Investing in Threatened Species Conservation: Does Corruption Outweigh Purchasing Power? 
PLoS ONE  2011;6(7):e22749.
In many sectors, freedom in capital flow has allowed optimization of investment returns through choosing sites that provide the best value for money. These returns, however, can be compromised in countries where corruption is prevalent. We assessed where the best value for money might be obtained for investment in threatened species that occur at a single site, when taking into account corruption. We found that the influence of corruption on potential investment decisions was outweighed by the likely value for money in terms of pricing parity. Nevertheless global conservation is likely to get best returns in terms of threatened species security by investing in “honest” countries than in corrupt ones, particularly those with a high cost of living.
doi:10.1371/journal.pone.0022749
PMCID: PMC3144939  PMID: 21818383
15.  Social determinants of health associated with hepatitis C co-infection among people living with HIV: results from the Positive Spaces, Healthy Places study 
Open Medicine  2011;5(3):e132-133.
Background
Social determinants of health (SDOH) may influence the probability of people living with HIV also being infected with hepatitis C virus (HCV). We compared the SDOH of adults co-infected with HCV/HIV with that of HIV mono-infected adults to identify factors independently associated with HCV infection.
Methods
In this cross-sectional study, face-to-face interviews were conducted with 509 HIV-infected adults affiliated with or receiving services from community-based AIDS service organizations (CBAOs). The primary outcome measure was self-reported HCV infection status. Chi-square, Student’s t tests, and Wilcoxon rank-sum tests were performed to compare SDOH of HCV/HIV co-infected participants with that of HIV mono-infected participants. Multivariable hierarchical logistic regression was used to identify factors independently associated with HCV co-infection.
Results
Data on 482 (95 HCV/HIV co-infected and 387 HIV mono-infected) adults were analyzed. Compared with participants infected with HIV only, those who were co-infected with HIV and HCV were more likely to be heterosexual, Aboriginal, less educated and unemployed. They were more likely to have a low income, to not be receiving antiretroviral treatment, to live outside the Greater Toronto Area (GTA), to use/abuse substances, experience significant depression, and utilize addiction counselling and needle-exchange services. They also were more likely to report a history of homelessness and perceived housing-related discrimination and to have moved twice or more in the previous 12 months. Factors independently associated with HCV/HIV co-infection were history of incarceration (odds ratio [OR] 8.81, 95% CI 4.43–17.54), history of homelessness (OR 3.15, 95% CI 1.59–6.26), living outside of the GTA (OR 3.13, 95% CI 1.59–6.15), and using/abusing substances in the past 12 months (OR 2.05, 95% CI 1.07–3.91).
Conclusion
Differences in SDOH exist between HIV/HCV co-infected and HIV mono-infected adults. History of incarceration, history of homelessness, substance use, and living outside the GTA were independently associated with HCV/HIV co-infection. Interventions that reduce homelessness and incarceration may help prevent HCV infection in people living with HIV.
PMCID: PMC3205830  PMID: 22046224
17.  Comparison of two modern vaccines and previous influenza infection against challenge with an equine influenza virus from the Australian 2007 outbreak 
Veterinary Research  2009;41(2):19.
During 2007, large outbreaks of equine influenza (EI) caused by Florida sublineage Clade 1 viruses affected horse populations in Japan and Australia. The likely protection that would be provided by two modern vaccines commercially available in the European Union (an ISCOM-based and a canarypox-based vaccine) at the time of the outbreaks was determined. Vaccinated ponies were challenged with a representative outbreak isolate (A/eq/Sydney/2888-8/07) and levels of protection were compared. A group of ponies infected 18 months previously with a phylogenetically-related isolate from 2003 (A/eq/South Africa/4/03) was also challenged with the 2007 outbreak virus. After experimental infection with A/eq/Sydney/2888-8/07, unvaccinated control ponies all showed clinical signs of infection together with virus shedding. Protection achieved by both vaccination or long-term immunity induced by previous exposure to equine influenza virus (EIV) was characterised by minor signs of disease and reduced virus shedding when compared with unvaccinated control ponies. The three different methods of virus titration in embryonated hens’ eggs, EIV NP-ELISA and quantitative RT-PCR were used to monitor EIV shedding and results were compared. Though the majority of previously infected ponies had low antibody levels at the time of challenge, they demonstrated good clinical protection and limited virus shedding. In summary, we demonstrate that vaccination with current EIV vaccines would partially protect against infection with A/eq/Sydney/2888-8/07-like strains and would help to limit the spread of disease in our vaccinated horse population.
doi:10.1051/vetres/2009067
PMCID: PMC2790087  PMID: 19863903
equine influenza; vaccine; protection; outbreak; antibody
18.  The prevalence of neuropathic pain: Clinical evaluation compared with screening tools in a community population 
Pain medicine (Malden, Mass.)  2009;10(3):586-593.
Background
Neuropathic pain is reported to be common based on studies from specialty centers and survey studies. However, few prevalence estimates have been completed in a community population using clinical evaluation.
Objective
To develop an estimate of the prevalence of neuropathic pain in community dwelling adults.
Methods
Data from a mailed survey (n=3575 community respondents), telephone interview (n=905), and a clinical examination (n=205) were linked to estimate the population prevalence of neuropathic pain. Using the clinical examination as the “gold” standard, estimates from several screening tools were developed and adjusted to the Olmsted County, Minnesota adult population.
Results
The estimated community prevalence of neuropathic pain from the clinical examination (gold standard) was 9.8%. Most other estimates were lower, including a 3.0% population prevalence using the Berger criteria and 8.8% using the S-LANSS. Only the prevalence rate based on self-report of nerve pain was higher (12.4%). Overlap among the groups each tool identified as having “neuropathic predominant pain” was only modest and the groups had significantly different rates of depressive symptoms, anxiety, limited functional ability and use of complementary and alternative medicine (CAM).
Conclusions
The estimated rates and personal characteristics of community residents with “neuropathic pain” varies widely depending on the tools used to identify neuropathic pain. None of the screening tools compared well to clinical evaluation. The differences in the groups identified by alternative screening methods become of major importance when reporting neuropathic pain epidemiology, studying therapies for neuropathic pain or attempting to translate neuropathic pain research into clinical practice.
doi:10.1111/j.1526-4637.2009.00588.x
PMCID: PMC2964880  PMID: 20849570
neuropathic pain; pain; prevalence; population based; screening tools; study populations; clinical practice
20.  Ultra low-dose naloxone and tramadol/acetaminophen in elderly patients undergoing joint replacement surgery: A pilot study 
OBJECTIVE:
A pilot study was conducted to assess whether both the rationale and feasibility exist for future randomized clinical trials to evaluate the combined use of naloxone infusion and tramadol/acetaminophen as opioid-sparing drugs in elderly patients undergoing lower extremity joint replacement surgery.
DESIGN:
Ten patients 70 years of age or older undergoing either total knee (n=7) or total hip (n=3) arthroplasty were treated prospectively. Each patient received two tablets of tramadol/acetaminophen (Tramacet; Janssen-Ortho Inc, Canada) preoperatively and every 6 h postoperatively, as well as a naloxone infusion started preoperatively at 0.25 μg/kg/h and continued up to 48 h postoperatively. In addition, standard intraoperative care was provided with 0.2 mg of intrathecal morphine, 1.4 mL of 0.75% bupivacaine, and an intra-articular infiltration of 100 mL of 0.3% ropivacaine and 30 mg of ketorolac, as well as standard postoperative morphine via patient-controlled analgesia orders and celecoxib 200 mg twice daily for five days.
OUTCOME MEASURES:
Compared with seven historical controls, also 70 years of age or older, who had undergone either a total knee (n=4) or total hip (n=3) arthroplasty, postoperative opioid use was reduced by 80%. Except for transient nausea and vomiting in 40% and 20% of patients, respectively, the 10 patients on tramadol/acetaminophen and naloxone tolerated the new regimen without difficulty.
CONCLUSION:
Consequently, a randomized, double-blinded clinical trial comparing standard therapy versus standard therapy plus these two drugs seems warranted. In such a trial, it would require approximately 20 subjects per treatment arm to detect a 80% decrease in morphine use.
PMCID: PMC2706645  PMID: 19532850
Analgesia; Elderly; Naloxone; Opioid-sparing; Total hip arthroplasty; Total knee arthroplasty; Tramacet; Tramadol
21.  Proteomic Analyses of Pancreatic Cyst Fluids 
Pancreas  2009;38(2):e33-e42.
Objectives
There are currently no diagnostic indicators that are consistently reliable, obtainable, and conclusive for diagnosing and risk-stratifying pancreatic cysts. Proteomic analyses were performed to explore pancreatic cyst fluids to yield effective diagnostic biomarkers.
Methods
We have prospectively recruited 20 research participants and prepared their pancreatic cyst fluids specifically for proteomic analyses. Proteomic approaches applied were: 1) MALDI-TOF (matrix-assisted laser-desorption-ionization time-of-flight) mass spectrometry peptidomics with LC/MS/MS (HPLC-tandem mass spectrometry) protein identification. 2) 2D gel electrophoresis. 3) GeLC/MS/MS (tryptic digestion of proteins fractionated by SDS-PAGE and identified by LC/MS/MS).
Results
Sequencing of over 350 free peptides showed that exopeptidase activities rendered peptidomics of cyst fluids unreliable; Protein nicking by proteases in the cyst fluids produced hundreds of protein spots from the major proteins, making 2D gel proteomics unmanageable; GeLC/MS/MS revealed a panel of potential biomarker proteins that correlated with CEA (carcinoembryonic antigen).
Conclusions
Two homologs of amylase, solubilized molecules of four mucins, four solubilized CEACAMs (CEA-related cell adhesion molecules), and four S100 homologs, may be candidate biomarkers to facilitate future pancreatic cyst diagnosis and risk-stratification. This approach required less than 40 microliters of cyst fluid per sample, offering the possibility to analyze cysts smaller than 1 cm diameter.
doi:10.1097/MPA.0b013e318193a08f
PMCID: PMC2681236  PMID: 19136908
Pancreatic cyst fluid; biomarkers; proteomic; mucin; S100; CEACAM
22.  Ocean currents help explain population genetic structure 
Management and conservation can be greatly informed by considering explicitly how environmental factors influence population genetic structure. Using simulated larval dispersal estimates based on ocean current observations, we demonstrate how explicit consideration of frequency of exchange of larvae among sites via ocean advection can fundamentally change the interpretation of empirical population genetic structuring as compared with conventional spatial genetic analyses. Both frequency of larval exchange and empirical genetic difference were uncorrelated with Euclidean distance between sites. When transformed into relative oceanographic distances and integrated into a genetic isolation-by-distance framework, however, the frequency of larval exchange explained nearly 50 per cent of the variance in empirical genetic differences among sites over scales of tens of kilometres. Explanatory power was strongest when we considered effects of multiple generations of larval dispersal via intermediary locations on the long-term probability of exchange between sites. Our results uncover meaningful spatial patterning to population genetic structuring that corresponds with ocean circulation. This study advances our ability to interpret population structure from complex genetic data characteristic of high gene flow species, validates recent advances in oceanographic approaches for assessing larval dispersal and represents a novel approach to characterize population connectivity at small spatial scales germane to conservation and fisheries management.
doi:10.1098/rspb.2009.2214
PMCID: PMC2871860  PMID: 20133354
seascape genetics; dispersal; pelagic larvae; isolation by distance; derived oceanographic distance
23.  Protein function annotation by homology-based inference 
Genome Biology  2009;10(2):207.
Where information on homologous proteins is available, progress is being made in automated prediction of protein function from sequence and structure.
With many genomes now sequenced, computational annotation methods to characterize genes and proteins from their sequence are increasingly important. The BioSapiens Network has developed tools to address all stages of this process, and here we review progress in the automated prediction of protein function based on protein sequence and structure.
doi:10.1186/gb-2009-10-2-207
PMCID: PMC2688287  PMID: 19226439
24.  A Conserved Salt Bridge in the G-Loop of Multiple Protein Kinases is Important for Catalysis and for in Vivo Lyn Function 
Molecular cell  2009;33(1):43-52.
The glycine-rich G-loop controls ATP binding and phosphate transfer in protein kinases. Here we show that the functions of Src family and Abl protein tyrosine kinases require an electrostatic interaction between oppositely charged amino acids within their G loops that is conserved in multiple other phylogenetically distinct protein kinases from plants to humans. By limiting G-loop flexibility, it controls ATP binding, catalysis and inhibition by ATP-competitive compounds such as Imatinib. In WeeB mice, mutational disruption of the interaction results in expression of a Lyn protein with reduced catalytic activity, and in perturbed B cell receptor signaling. Like Lyn-/- mice, WeeB mice show profound defects in B cell development and function and succumb to autoimmune glomerulonephritis. This demonstrates the physiological importance of the conserved G-loop salt bridge and at the same time distinguishes the in vivo requirement for the Lyn kinase activity from other potential functions of the protein.
doi:10.1016/j.molcel.2008.12.024
PMCID: PMC2683036  PMID: 19150426
25.  Phase II Trial of Neoadjuvant/Adjuvant Imatinib Mesylate (IM) for Advanced Primary and Metastatic/Recurrent Operable Gastrointestinal Stromal Tumor (GIST) – early results of RTOG 0132 
Journal of surgical oncology  2009;99(1):42-47.
Background:
Therapy for gastrointestinal stromal tumors (GIST) has changed significantly with the use of Imatinib Mesylate (IM). Despite the success of this drug in metastatic GIST, disease progression remains a perplexing clinical issue suggesting the need for multimodality management. There have been no prospective studies either evaluating the neoadjuvant use of IM in primary GIST or as a preoperative cytoreduction agent for metastatic GIST.
Methods:
RTOG 0132 was a prospective phase II study evaluating safety and efficacy of neoadjuvant IM (600 mg/day) for patients with primary GIST or the preop use of IM in patients with operable metastatic GIST. The trial continued postop IM for 2 years.
Results:
63 patients were entered (52 analyzable), 30 patients with primary GIST (Group A) and 22 with metastatic GIST (Group B). Response (RECIST) in Group A was (7% partial, 83% stable, 10% unknown), in Group B (4.5% partial, 91% stable, 4.5% progression). Two year progression free survival (Group A 83%, Group B 77%). Estimated overall survival (Group A 93%, Group B 91%). Complications of surgery and IM toxicity were minimal.
Conclusion:
This trial represents the first prospective report of preop IM in GIST. This approach is feasible, requires multidisciplinary consultations, is not associated with notable postop complications.
doi:10.1002/jso.21160
PMCID: PMC2606912  PMID: 18942073
GIST; neoadjuvant imatinib; locally advanced GIST; metastatic GIST

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