Nociceptor-localized cGMP-dependent protein kinase I is a critical generator for central sensitization and neuropathic pain.

Patients with neuropathic pain often experience exaggerated pain and anxiety. Central sensitization has been linked with the maintenance of neuropathic pain and may become an autonomous pain generator. Conversely, emerging evidence accumulated that central sensitization is initiated and maintained by ongoing nociceptive primary afferent inputs. However, it remains elusive what mechanisms underlie this phenomenon and which peripheral candidate contributes to central sensitization that accounts for pain hypersensitivity and pain-related anxiety. Previous studies have implicated peripherally localized cGMP-dependent protein kinase I (PKG-I) in plasticity of nociceptors and spinal synaptic transmission as well as inflammatory hyperalgesia. However, whether peripheral PKG-I contributes to cortical plasticity and hence maintains nerve injury-induced pain hypersensitivity and anxiety is unknown. Here, we demonstrated significant upregulation of PKG-I in ipsilateral L3 dorsal root ganglia (DRG), no change in L4 DRG, and downregulation in L5 DRG upon spared nerve injury. Genetic ablation of PKG-I specifically in nociceptors or post-treatment with intervertebral foramen injection of PKG-I antagonist, KT5823, attenuated the development and maintenance of spared nerve injury-induced bilateral pain hypersensitivity and anxiety. Mechanistic analysis revealed that activation of PKG-I in nociceptors is responsible for synaptic potentiation in the anterior cingulate cortex upon peripheral neuropathy through presynaptic mechanisms involving brain-derived neurotropic factor signaling. Our results revealed that PKG-I expressed in nociceptors is a key determinant for cingulate synaptic plasticity after nerve injury, which contributes to the maintenance of pain hypersensitivity and anxiety. Thereby, this study presents a strong basis for opening up a novel therapeutic target, PKG-I, in nociceptors for treatment of comorbidity of neuropathic pain and anxiety with least side effects.

Presurgical depression and anxiety are not associated with worse epilepsy surgery outcome five years postoperatively.

PURPOSE: Anxiety and depression have been associated with poor seizure control after epilepsy surgery. This study explored the effect of presurgical anxiety or depression on two- and five-year seizure control outcomes. METHODS: Adult subjects were enrolled between 1996 and 2001 in a multicenter prospective study to evaluate outcomes of resective epilepsy surgery. A Poisson regression was used to analyze the association of depression and anxiety with surgical outcome, while adjusting for gender, age, ethnicity, number of years with seizures, and presence of mesial temporal sclerosis. RESULTS: The relative risk (RR) of presurgical depression on two-year seizure-free outcome in this cohort is 1.12 (95% confidence interval (CI), 0.84-1.49) and 1.06 (CI, 0.73-1.55) on five-year seizure free outcome. The RR of presurgical anxiety on two-year seizure outcome is 0.73 (CI, 0.50-1.07) and 0.70 (CI, 0.43-1.17) on five-year seizure outcome. When including Engel classes I and II, the RRs of presurgical depression, anxiety, or both two years after surgery were 0.96 (p=0.59), 0.73 (p<0.05), and 0.97 (p=0.70), respectively, and they were 0.97 (p=0.82), 0.84 (p=0.32), and 0.89 (p=0.15), respectively, five years after surgery. Only presurgical anxiety was associated with worse epilepsy surgery outcome two year after surgery but not at five years postsurgery. Depression was not a risk factor for poor epilepsy surgical outcome in the long term. CONCLUSION: These findings from a prospective study that utilized a standardized protocol for psychiatric and seizure outcome assessment suggest that presurgical mood disorders have no substantial impact on postsurgical seizure outcome for up to five years after surgery.

Ten-Year Prevalence and Incidence of Urinary Incontinence in Older Women: A Longitudinal Analysis of the Health and Retirement Study.

OBJECTIVES: To measure the incidence of urinary incontinence (UI) over 10 years in older women who did not report UI at baseline in 1998, to estimate the prevalence of female UI according to severity and type, and to explore potential risk factors for development of UI. DESIGN: Secondary analysis of a prospective cohort. SETTING: Health and Retirement Study. PARTICIPANTS: Women participating in the Health and Retirement Study between 1998 and 2008 who did not have UI at baseline (1998). MEASUREMENTS: UI was defined as an answer of "yes" to the question, "During the last 12 months, have you lost any amount of urine beyond your control?" UI was characterized according to severity (according to the Sandvik Severity Index) and type (according to International Continence Society definitions) at each biennial follow-up between 1998 and 2008. RESULTS: In 1998, 5,552 women aged 51 to 74 reported no UI. The cumulative incidence of UI in older women was 37.2% (95% confidence interval (CI)=36.0-38.5%). The most common incontinence type at the first report of leakage was mixed UI (49.1%, 95% CI=46.5-51.7%), and women commonly reported their symptoms at first leakage as moderate to severe (46.4%, 95% CI=43.8-49.0%). CONCLUSION: Development of UI in older women was common and tended to result in mixed type and moderate to severe symptoms.

Final analysis of a phase II study of modified FOLFIRINOX in locally advanced and metastatic pancreatic cancer.

BACKGROUND: Modifications of FOLFIRINOX are widely used despite the absence of prospective data validating efficacy in metastatic disease (metastatic pancreatic cancer (MPC)) or locally advanced pancreatic cancer (LAPC). We conducted a multicentre phase II study of modified FOLFIRINOX in advanced pancreatic cancer to assess the impact of dose attenuation in MPC and efficacy in LAPC. METHODS: Patients with untreated MPC or LAPC received modified FOLFIRINOX (irinotecan and bolus 5-fluorouracil reduced by 25%). Adverse events (AEs) were compared with full-dose FOLFIRINOX. Response rate (RR), median progression-free survival (PFS) and median overall survival (OS) were determined. RESULTS: In total, 31 and 44 patients with LAPC and MPC were enrolled, respectively. In MPC, efficacy of modified FOLFIRINOX was comparable with FOLFIRINOX with RR 35.1%, OS 10.2 months (95% CI 7.65-14.32) and PFS 6.1 months (95% CI 5.19-8.31). In LAPC, efficacy was notable with RR 17.2%, resection rate 41.9%, PFS 17.8 months (95% CI 11.0-23.9) and OS 26.6 months (95% CI 16.7, NA). Neutropenia (P<0.0001), vomiting (P<0.001) and fatigue (P=0.01) were significantly decreased. [(18)F]-Fluorodeoxyglucose positron emission tomography imaging response did not correlate with PFS or OS. CONCLUSIONS: In this first prospective study of modified FOLFIRINOX in MPC and LAPC, we observed decreased AEs compared with historical control patients. In MPC, the efficacy appears comparable with FOLFIRINOX. In LAPC, PFS and OS were prolonged and support the continued use of FOLFIRINOX in this setting.

Viral load monitoring and antiretroviral treatment outcomes in a pediatric HIV cohort in Ghana.

BACKGROUND: HIV-infected children in sub-Saharan Africa may be at a high risk of staying on a failing first-line regimen and developing drug-resistance HIV variants due to lack of routine viral load monitoring. We investigated whether cumulative viral load, measured as viremia copy-years (VCY) could predict morbidity in a setting where viral load is not routinely monitored. METHODS: This was a single-center prospective observational longitudinal study of HIV-infected children initiating antiretroviral therapy (ART) at the Pediatric HIV/AIDS Care program at Korle-Bu Teaching Hospital in Accra, Ghana. The main outcome was morbidity measured as frequency of hospitalizations, opportunistic infections, and outpatient sick visits. The main explanatory variable was viral load measured as VCY. RESULTS: The study included 140 children who initiated ART between September 2009 and May 2013 and had at least 2 viral load measurements. There were 184 hospitalizations, with pneumonia being the most common cause (22.8 %). A total of 102 opportunistic infections was documented, with tuberculosis being the most common opportunistic infection (68 %). A total of 823 outpatient sick visits was documented, with upper respiratory infections (14.2 %) being the most common cause. Forty-four percent of our study participants had >4 log10 VCY. Children in this sub-cohort had a higher frequency of sick visits compared with those with <4 log10 VCY (p = 0.03). Only 6.5 % of children with >4 log10 VCY had been identified as treatment failure using WHO clinical and immunological treatment failure criteria. CONCLUSIONS: High level of cumulative viral load may translate to virological failure and subsequent increased all-cause morbidity. Our finding of potential utility of VCY in pediatrics warrants further investigations. VCY may be a good alternate to routine viral load measurement as its determination may be less frequent and could be personalized to save cost.

Influence of Sub-Daily Variation on Multi-Fractal Detrended Fluctuation Analysis of Wind Speed Time Series.

Using multi-fractal detrended fluctuation analysis (MF-DFA), the scaling features of wind speed time series (WSTS) could be explored. In this paper, we discuss the influence of sub-daily variation, which is a natural feature of wind, in MF-DFA of WSTS. First, the choice of the lower bound of the segment length, a significant parameter of MF-DFA, was studied. The results of expanding the lower bound into sub-daily scope shows that an abrupt declination and discrepancy of scaling exponents is caused by the inability to keep the whole diel process of wind in one single segment. Additionally, the specific value, which is effected by the sub-daily feature of local meteo-climatic, might be different. Second, the intra-day temporal order of wind was shuffled to determine the impact of diel variation on scaling exponents of MF-DFA. The results illustrate that disregarding diel variation leads to errors in scaling. We propose that during the MF-DFA of WSTS, the segment length should be longer than 1 day and the diel variation of wind should be maintained to avoid abnormal phenomena and discrepancy in scaling exponents.

HIV knowledge, stigma, and illness beliefs among pediatric caregivers in Ghana who have not disclosed their child's HIV status.

The majority of HIV-infected children in sub-Saharan Africa have not been informed of their HIV status. Caregivers are reluctant to disclose HIV status to their children because of concern about the child's ability to understand, parental sense of guilt, and fear of social rejection and isolation. We hypothesized that the low prevalence of pediatric HIV disclosure in Ghana is due to lack of accurate HIV information and high HIV stigma among caregivers. This is a preliminary analysis of baseline data of an HIV pediatric disclosure intervention study in Ghana ("Sankofa"). "Sankofa" - is a two-arm randomized controlled clinical trial comparing disclosure intervention plus usual care (intervention arm) vs usual care (control arm) at Korle-Bu Teaching Hospital (KBTH; control arm) and Komfo-Anokye Teaching Hospital (KATH; intervention arm). We enrolled HIV-infected children, ages 7-18 years who do not know their HIV status, and their caregivers. Baseline data of caregivers included demographic characteristics; Brief HIV Knowledge Questionnaire (HIV-KQ-18); Brief Illness Perception Questionnaire; and HIV Stigma Scale. Simple and multivariable linear regression analyses were used to assess the relationship between caregiver characteristics and HIV knowledge, stigma, and illness perception. Two hundred and ninety-eight caregivers were enrolled between January 2013 and July 2014 at the two study sites; KBTH (n = 167) and KATH (n = 131). The median age of caregivers was 41 years; 80.5% of them were female and about 60% of caregivers were HIV-positive. Seventy-eight percent of caregivers were self-employed with low household income. In both unadjusted and adjusted analyses, HIV negative status and lower level of education were associated with poor scores on HIV-KQ. HIV positive status remained significant for higher level of stigma in the adjusted analyses. None of the caregiver's characteristics predicted caregiver's illness perception. Intensification of HIV education in schools and targeted community campaigns are needed.

Sankofa pediatric HIV disclosure intervention cyber data management: building capacity in a resource-limited setting and ensuring data quality.

Prevalence of pediatric HIV disclosure is low in resource-limited settings. Innovative, culturally sensitive, and patient-centered disclosure approaches are needed. Conducting such studies in resource-limited settings is not trivial considering the challenges of capturing, cleaning, and storing clinical research data. To overcome some of these challenges, the Sankofa pediatric disclosure intervention adopted an interactive cyber infrastructure for data capture and analysis. The Sankofa Project database system is built on the HUBzero cyber infrastructure ( https://hubzero.org ), an open source software platform. The hub database components support: (1) data management - the "databases" component creates, configures, and manages database access, backup, repositories, applications, and access control; (2) data collection - the "forms" component is used to build customized web case report forms that incorporate common data elements and include tailored form submit processing to handle error checking, data validation, and data linkage as the data are stored to the database; and (3) data exploration - the "dataviewer" component provides powerful methods for users to view, search, sort, navigate, explore, map, graph, visualize, aggregate, drill-down, compute, and export data from the database. The Sankofa cyber data management tool supports a user-friendly, secure, and systematic collection of all data. We have screened more than 400 child-caregiver dyads and enrolled nearly 300 dyads, with tens of thousands of data elements. The dataviews have successfully supported all data exploration and analysis needs of the Sankofa Project. Moreover, the ability of the sites to query and view data summaries has proven to be an incentive for collecting complete and accurate data. The data system has all the desirable attributes of an electronic data capture tool. It also provides an added advantage of building data management capacity in resource-limited settings due to its innovative data query and summary views and availability of real-time support by the data management team.

Multiple types of motives don't multiply the motivation of West Point cadets.

Although people often assume that multiple motives for doing something will be more powerful and effective than a single motive, research suggests that different types of motives for the same action sometimes compete. More specifically, research suggests that instrumental motives, which are extrinsic to the activities at hand, can weaken internal motives, which are intrinsic to the activities at hand. We tested whether holding both instrumental and internal motives yields negative outcomes in a field context in which various motives occur naturally and long-term educational and career outcomes are at stake. We assessed the impact of the motives of over 10,000 West Point cadets over the period of a decade on whether they would become commissioned officers, extend their officer service beyond the minimum required period, and be selected for early career promotions. For each outcome, motivation internal to military service itself predicted positive outcomes; a relationship that was negatively affected when instrumental motives were also in evidence. These results suggest that holding multiple motives damages persistence and performance in educational and occupational contexts over long periods of time.

Mood, anxiety, and incomplete seizure control affect quality of life after epilepsy surgery.

OBJECTIVE: We examined the complex relationship between depression, anxiety, and seizure control and quality of life (QOL) outcomes after epilepsy surgery. METHODS: Seven epilepsy centers enrolled 373 patients and completed a comprehensive diagnostic workup and psychiatric and follow-up QOL evaluation. Subjects were evaluated before surgery and then at 3, 6, 12, 24, 48, and 60 months after surgery. Standardized assessments included the Quality of Life in Epilepsy Inventory-89, Beck Depression Inventory (BDI), and Beck Anxiety Inventory (BAI). A mixed-model repeated-measures analysis was used to analyze associations of depression, anxiety, seizure outcome, and seizure history with overall QOL score and QOL subscores (cognitive distress, physical health, mental health, epilepsy-targeted) prospectively. RESULTS: The groups with excellent and good seizure control showed a significant positive effect on the overall QOL compared to the groups with fair and poor seizure control. The BDI and BAI scores were both highly and negatively associated with overall QOL; increases in BDI and BAI scores were associated with decreased overall QOL score. CONCLUSIONS: Depression and anxiety are strongly and independently associated with worse QOL after epilepsy surgery. Interestingly, even partial seizure control, controlling for depression and anxiety levels, improved QOL. Management of mood and anxiety is a critical component to postsurgical care.

Parotid gland lymphoma: prognostic analysis of 2140 patients.

OBJECTIVES/HYPOTHESIS: Assess the demographic, clinical, and pathologic features of patients with parotid gland lymphoma and their prognostic importance using US population-based data. STUDY DESIGN: Retrospective cohort study. METHODS: Patients were selected from the Surveillance, Epidemiology, and End Results program database between the years of 1973 and 2008, and individual characteristics were compared using univariate and multivariate Cox proportional hazards models. Kaplan-Meier survival curves were constructed and log-rank tests were performed. RESULTS: We identified 2,140 patients with primary parotid gland lymphoma. Hodgkin lymphoma was found in 3.5% of patients. More common were non-Hodgkin lymphoma subtypes: marginal zone B-cell lymphoma, follicular lymphoma, and diffuse large B cell lymphoma accounted for 27.9%, 25.8%, and 23.7% of cases, respectively. Survival was decreased with patient age over 50 years, increasing stage, male gender, non-Hodgkin histology, and status other than married. Of the patients, 72.0% received some form of surgery, and 136 patients had facial nerve sacrifice during parotidectomy. CONCLUSIONS: Non-Hodgkin lymphoma is the predominant type of lymphoma seen in the parotid gland. Patient and histologic features determine survival, and surgery is often performed. Facial nerve sacrifice, which is contraindicated given the systemic nature of lymphoma and the role of chemotherapy and radiation in its treatment, is reported in 6.4% of patients with parotid gland lymphoma. LEVEL OF EVIDENCE: 2b.

Polycystic ovary syndrome in adolescence: impaired glucose tolerance occurs across the spectrum of BMI.

CONTEXT: Polycystic ovary syndrome (PCOS) is a chronic condition with metabolic manifestations spanning the reproductive years. OBJECTIVE: We sought to examine glucose metabolism, irrespective of the presence of obesity in a cohort of adolescent girls with PCOS. DESIGN: One hundred adolescents were assessed for PCOS in a multi-specialty adolescent PCOS program. PCOS was diagnosed by Androgen Excess Society criteria. Oral glucose tolerance testing (OGTT), homeostatic model assessment of insulin resistance, and androgen and lipid profiles were performed for those meeting criteria. RESULTS: Sixty-six adolescents (mean age 15.8 ± 0.2 yrs, range 13.0-18.6) had confirmed PCOS, and were eligible for inclusion in our analysis. Abnormal glucose metabolism was present in 12 of 66 (18.2%) subjects: 2 (3.0%) impaired fasting glucose, 10 (15.2%) impaired glucose tolerance (IGT), and 1 (1.5%) type 2 diabetes. IGT was the most common abnormality, occurring with equal frequency in obese (OB, mean body mass index (BMI) 36.9 ± 0.8 kg/m(2) ) and non-obese (NOB, mean BMI 24.5 ± 0.6 kg/m(2) ) adolescents (p = 0.3). In a subgroup analysis, NOB adolescents with IGT (NOB-IGT) had similar mean 2-h insulin, high density lipoprotein, C-reactive protein, and testosterone levels to the OB cohort despite marked differences in BMI (p < 0.001) and % body fat (p = 0.002). However, the NOB-IGT group had a lower mean fasting insulin level than the OB cohort (p = 0.04). CONCLUSION: Abnormal glucose metabolism is highly prevalent in adolescents with PCOS. In particular, IGT occurs across the spectrum of BMI. A screening OGTT should be considered for adolescents diagnosed with PCOS, independently of their BMI.

Surgical residency and attrition: defining the individual and programmatic factors predictive of trainee losses.

BACKGROUND: Voluntary resident attrition remains problematic despite recent changes in postgraduate general surgery training, including reduction of work hours. STUDY DESIGN: We conducted a prospective study of all postgraduate year (PGY)-1 and -2 trainees on the 2008 American Board of Surgery resident roster (ABS-RR) who completed the National Study of Expectations and Attitudes of Residents in Surgery (NEARS) survey after the American Board of Surgery In-Training Examination (ABSITE) in 2008 or 2009. RESULTS: Among 2,222 PGY-1 and -2 residents on the 2008 ABS-RR, 2,033 completed the NEARS survey in 2008 or 2009 (91.5%). The only demographic or programmatic variables associated with voluntary attrition on univariate analysis were PGY-1 status (9.4% risk vs 4.5% risk for PGY-2, p < 0.001) and program location (p = 0.03). Response differences (p < 0.01) were noted in 23 survey items. In multivariate modeling, PGY-2 status was protective against voluntary attrition (p < 0.001, hazard ratio [HR] 0.41), while programs located outside of the South (Northeast: p = 0.006, HR 2.39; Midwest: p = 0.01, HR 2.37; West: p = 0.10, HR 1.76) were associated with higher attrition. The attrition group more frequently reported that they had considered leaving training (p < 0.001, HR 2.59), that the personal cost of training was too great (p < 0.001, HR 2.89), that they were dissatisfied with their operative experience (p = 0.002, HR 1.89), and that they were not committed to completing their training (p < 0.001, HR 3.96). Using the estimated regression coefficient for each variable in the multivariate models, we calculated a risk score for individual residents; these scores were used to construct covariate-adjusted survivorship functions. CONCLUSIONS: Resident attitudes, PGY-1 status, and program location are most frequently associated with voluntary attrition. Our risk score calculation represents a novel potential tool for programs to quantify deficiencies in the training experience of residents, and develop targeted strategies to limit disaffection and improve resident retention.

FOLFIRINOX for locally advanced and metastatic pancreatic cancer: single institution retrospective review of efficacy and toxicity.

Although FOLFIRINOX significantly increases survival in metastatic pancreatic cancer (MPC) compared to gemcitabine (Conroy et al. N Engl J Med 364:1817-1825, 2011), toxicities have tempered enthusiasm for its use in full doses. To assess the impact of dose attenuations on toxicity and efficacy, we reviewed our institution's experience with FOLFIRINOX in locally advanced pancreatic cancer (LAPC) and MPC. We performed a retrospective review of dose, toxicity, and efficacy of FOLFIRINOX in all patients with LAPC and MPC treated between June 2010 and July 2011 at Yale. Toxicities in all patients and response rate (RR) and survival in previously untreated MPC were compared to data reported by Conroy. Overall survival (OS) and progression-free survival were estimated by Kaplan-Meier method. Thirty-five patients were treated (16 LAPC; 19 MPC). Twenty-nine patients received dose attenuations with the first cycle. Median relative doses of irinotecan and bolus fluorouracil were less than those reported by Conroy (64 vs. 81 % and 66 vs. 82 %, respectively). RR was 50 % in LAPC and 47 % in MPC, and the latter did not differ significantly from the RR reported by Conroy (p = 0.19). OS at 6 and 12 months in MPC was comparable to OS reported by Conroy. Grade 3/4 toxicities were less than reported by Conroy, including fatigue (p = 0.009) and neutropenia (p < 0.0001). Nine patients experienced transient dysarthria during irinotecan administration. Our findings validate the efficacy and tolerability of FOLFIRINOX in LAPC and MPC and suggest that dose attenuations of irinotecan and bolus fluorouracil improve tolerability without compromising efficacy.

Human POLB gene is mutated in high percentage of colorectal tumors.

Previous small scale sequencing studies have indicated that DNA polymerase ß (pol ß) variants are present on average in 30% of human tumors of varying tissue origin. Many of these variants have been shown to have aberrant enzyme function in vitro and to induce cellular transformation and/or genomic instability in vivo, suggesting that their presence is associated with tumorigenesis or its progression. In this study, the human POLB gene was sequenced in a collection of 134 human colorectal tumors and was found to contain coding region mutations in 40% of the samples. The variants map to many different sites of the pol ß protein and are not clustered. Many variants are nonsynonymous amino acid substitutions predicted to affect enzyme function. A subset of these variants was found to have reduced enzyme activity in vitro and failed to fully rescue pol ß-deficient cells from methylmethane sulfonate-induced cytotoxicity. Tumors harboring variants with reduced enzyme activity may have compromised base excision repair function, as evidenced by our methylmethane sulfonate sensitivity studies. Such compromised base excision repair may drive tumorigenesis by leading to an increase in mutagenesis or genomic instability.

Model for end-stage liver disease score predicts adverse events related to ventricular assist device therapy.

BACKGROUND: The Model for End-stage Liver Disease (MELD) score is a marker of multisystem organ dysfunction. It has been used to predict outcomes in patients undergoing hepatic and nonhepatic interventions. End-stage heart disease exhibits a varying degree of multiorgan dysfunction, which impacts the adverse events related to ventricular assist device (VAD) therapy. Our aim for the present study was to investigate the value of MELD score in predicting adverse events related with VAD therapy. METHODS: Data were collected on demographics, clinical characteristics, MELD score; Interagency Registry for Mechanically Assisted Circulatory Support-defined VAD adverse events within the first 6 months, and survival from VAD recipients (n=286; from 1996 to 2009). Univariable, multivariable, and Cox regression analyses were performed using SAS software (SAS Institute, Cary, NC). RESULTS: The mean MELD score was 14.4±5.9. Actuarial incidence of infections, bleeding events, and cardiovascular dysfunction at 6 months was 65.4%, 52.1%, and 45.6%, respectively. Multivariable Cox proportional hazards model (controlling for gender, type of device, diagnosis, intention to treat, urgency, and inotropic use) confirmed that MELD score predicted mortality, respiratory, and renal dysfunction at 6 months (p<0.01). CONCLUSIONS: Preoperative MELD score is predictive of mortality, respiratory, and renal dysfunction at 6 months after controlling for gender, type of device, diagnosis, intention to treat, urgency, and inotropic use. The MELD score may be used as a quantitative tool to assess the adverse events associated with VAD therapy.

Comprehensive analysis of variables affecting delayed gastric emptying following pancreaticoduodenectomy.

INTRODUCTION: Delayed gastric emptying (DGE) occurs commonly following pancreaticoduodenectomy (PD), and criteria for its clinical diagnosis have recently been standardized by an international consensus group. We evaluated 24 operative and peri-operative variables to assess which were independent risk factors for DGE development. Secondarily, we also examined DGE incidence over time and evaluated the consensus guidelines' ability to diagnose DGE in patients with complicated post-operative courses. METHODS: A prospective, single-surgeon database of 235 patients undergoing PD at an academic tertiary center was retrospectively reviewed and DGE was assessed per published guidelines. RESULTS: DGE occurred in 42 patients overall (17.9 %), with incidence falling from 30.0 to 9.1% during the study period. Post-operative abscess, pancreatic fistula formation, pulmonary comorbidity, and increased intraoperative blood loss were found to be independent risk factors (p<0.05) for DGE on multivariate analysis. Changes in operative technique, such as pylorus preservation, did not associate with DGE. In a separate analysis, when patients with confounding post-operative events such as re-intubation or re-laparotomy were excluded, DGE incidence was 11.9 %. CONCLUSIONS: Perturbation of the operative bed by a secondary complication seems to be the dominant risk for DGE development. The consensus guidelines for DGE diagnosis, while indispensable, may overestimate DGE incidence.

Differential hypoxic regulation of hypoxia-inducible factors 1alpha and 2alpha.

The hypoxia-inducible transcription factors (HIF) 1α and HIF-2α play a critical role in cellular response to hypoxia. Elevated HIF-α expression correlates with poor patient survival in a large number of cancers. Recent evidence suggests that HIF-2α appears to be preferentially expressed in neuronal tumor cells that exhibit cancer stem cell characteristics. These observations suggest that expression of HIF-1α and HIF-2α is differentially regulated in the hypoxic tumor microenvironment. However, the underlying mechanisms remain to be fully investigated. In this study, we investigated the transcriptional regulation of HIF-1α and HIF-2α under different physiologically relevant hypoxic conditions. We found that transcription of HIF-2α was consistently increased by hypoxia, whereas transcription of HIF-1α showed variable levels of repression. Mechanistically, differential regulation of HIF-α transcription involved hypoxia-induced changes in acetylation of core histones H3 and H4 associated with the proximal promoters of the HIF-1α or HIF-2α gene. We also found that, although highly stable under acute hypoxia, HIF-1α and HIF-2α proteins become destabilized under chronic hypoxia. Our results have thus provided new mechanistic insights into the differential regulation of HIF-1α and HIF-2α by the hypoxic tumor microenvironment. These findings also suggest an important role of HIF-2α in the regulation of tumor progression under chronic hypoxia.

Nuclear expression of S100A4 calcium-binding protein increases cholangiocarcinoma invasiveness and metastasization.

UNLABELLED: Cholangiocarcinoma (CCA) carries a severe prognosis because of its strong invasiveness and early metastasization. In several patients, otherwise eligible for surgical resection, micrometastasis are already present at the time of surgery. The mechanisms responsible for CCA invasiveness are unclear. S100A4, a member of the S100 family of small Ca(2+)-binding proteins, is expressed in mesenchymal cells, regulates cell motility in several cell types, and is expressed in some epithelial cancers. Thus, we aimed to study the role of S100A4 in CCA invasiveness and metastasization. The expression of S100A4 was studied by immunohistochemistry in 93 human liver samples of CCA patients undergoing surgical resection and correlated with metastases development (67 cases) and patient survival following surgery using log rank tests and multivariate analysis. S100A4 expression was studied in EGI-1 and TFK-1, human CCA cell lines with and without nuclear S100A4 expression, respectively. Metastatic properties of CCA cells were assessed by xenotransplantation in severe combined immunodeficiency (SCID) mice after transduction with lentiviral vectors encoding firefly luciferase gene. Proliferation, motility (wound healing), invasiveness (Boyden chamber), and metalloproteinases (MMPs) secretion were studied in CCA cells, with or without lentiviral silencing of S100A4. Nuclear expression of S100A4 by neoplastic ducts was a strong predictor of metastasization and reduced survival after resection (P < 0.01). EGI-1 CCA cells showed stronger metastatic properties than TFK-1 when xenotransplanted in SCID mice. S100A4-silenced EGI-1 cells showed significantly reduced motility, invasiveness, and MMP-9 secretion in vitro, without changes in cell proliferation. CONCLUSION: Nuclear S100A4 identifies a subset of CCA patients with a poor prognosis after surgical resection. Nuclear expression of S100A4 increases CCA cells invasiveness and metastasization, indicating that S100A4 may also represent a potential therapeutic target.

Allosteric aptamers: targeted reversibly attenuated probes.

Aptamers are unique nucleic acids with regulatory potentials that differ markedly from those of proteins. A significant feature of aptamers not possessed by proteins is their ability to participate in at least two different types of three-dimensional structure: a single-stranded folded structure that makes multiple contacts with the aptamer target and a double-helical structure with a complementary nucleic acid sequence. We have made use of this structural flexibility to develop an aptamer-based biosensor (a targeted reversibly attenuated probe, TRAP) in which hybridization of a cis-complementary regulatory nucleic acid (attenuator) controls the ability of the aptamer to bind to its target molecule. The central portion of the TRAP, between the aptamer and the attenuator, is complementary to a target nucleic acid, such as an mRNA, which is referred to as a regulatory nucleic acid (regNA) because it regulates the activity of the aptamer in the TRAP by hybridization with the central (intervening) sequence. The studies reported here of the ATP-DNA TRAP suggest that, as well as inhibiting the aptamer, the attenuator also acts as a structural guide, much like a chaperone, to promote proper folding of the TRAP such that it can be fully activated by the regDNA. We also show that activation of the aptamer in the TRAP by the complementary nucleic acid at physiological temperatures is sensitive to single-base mismatches. Aptamers that can be regulated by a specific nucleic sequence such as in an mRNA have potential for many in vivo applications including regulating a particular enzyme or signal transduction pathway or imaging gene expression in vivo.