In Drosophila, anatomically discrete dopamine neurons that innervate distinct zones of the mushroom body (MB) assign opposing valence to odors during olfactory learning. Subsets of MB neurons have temporally unique roles in memory processing, but valence-related organization has not been demonstrated. We functionally subdivided the alphabeta neurons, revealing a value-specific role for the approximately 160 alphabeta core (alphabetac) neurons. Blocking neurotransmission from alphabeta surface (alphabetas) neurons revealed a requirement during retrieval of aversive and appetitive memory, whereas blocking alphabetac only impaired appetitive memory. The alphabetac were also required to express memory in a differential aversive paradigm demonstrating a role in relative valuation and approach behavior. Strikingly, both reinforcing dopamine neurons and efferent pathways differentially innervate alphabetac and alphabetas in the MB lobes. We propose that conditioned approach requires pooling synaptic outputs from across the alphabeta ensemble but only from the alphabetas for conditioned aversion.
It is now almost forty years since the first description of learning in the fruit fly Drosophila melanogaster. Various incarnations of the classic mutagenesis approach envisaged in the early days have provided around one hundred learning defective mutant fly strains. Recent technological advances permit temporal control of neural function in the behaving fly. These approaches have radically changed experiments in the field and have provided a neural circuit perspective of memory formation, consolidation and retrieval. Combining neural perturbations with more classical mutant intervention allows investigators to interrogate the molecular and cellular processes of memory within the defined neural circuits. Here, we summarize some of the progress made in the last ten years that indicates a remarkable conservation of the neural mechanisms of memory formation between flies and mammals. We emphasize that considering an ethologically-relevant viewpoint might provide additional experimental power in studies of Drosophila memory.
Delineating breast ductal carcinoma using combined dye-enhanced wide-field polarization imaging and optical coherence tomography
Intra-operative delineation of breast cancer is a challenging problem. We used dye-enhanced wide-field polarization imaging for rapid demarcation of en face cancer margins and optical coherence tomography (OCT) for cross-sectional evaluation. Ductal carcinoma specimens were stained with methylene blue. Wide-field reflectance images were acquired at 440 and 640 nm. Wide-field fluorescence images were excited at 640 nm and registered between 660 nm and 750 nm. OCT images were acquired using a 1310 nm swept-source system. The results were validated against histopathology. Both imaging modalities provided diagnostic information on cancer margins. Combined OCT and wide-field polarization imaging shows promise for intra-operative detection of ductal breast carcinoma.
Sodium bicarbonate is central to the treatment of many poisonings. When it was placed on the FDA drug shortage list in 2012, alternative treatment strategies to specific poisonings were considered. Many hospital pharmacies, poison centers, and medical toxicologists proposed sodium acetate as an adequate alternative, despite a paucity of data to support its use in medical toxicology. The intention of this review is to educate the clinician on the use of sodium acetate and to advise them on the potential adverse events when given in excess. We conducted a literature search focused on the pharmacology of sodium acetate, its use as a buffer in pathologic acidemia and dialysis baths, and potential adverse events associated with excess sodium acetate infusion. It appears safe to replace sodium bicarbonate infusion with sodium acetate on an equimolar basis. The metabolism of acetate, however, is more complex than bicarbonate. Future prospective studies will be needed to confirm the efficacy of sodium acetate in the treatment of the poisoned patient.
The use of alternative polyadenylation sites renders integrin beta1 (Itgb1) mRNA isoforms with differential stability during mammary gland development
Integrins are heterodimeric cell-surface adhesion receptors that play a critical role in tissue development. Characterization of the full-length mRNA encoding the beta1 subunit (Itgb1) revealed an alternative functional cleavage and polyadenylation site that yields a new Itgb1 mRNA isoform 578 bp shorter than that previously reported. Using a variety of experimental and bioinformatic approaches, we found that the two Itgb1 isoforms are expressed at different levels in a variety of mouse tissues, including the mammary gland, where they are differentially regulated at successive developmental stages. The longer mRNA species is prevelant during lactation, whereas the shorter is induced after weaning. In 3D cultures, where expression of integrin beta1 protein is required for normal formation of acini, experimental blockade of the longer isoform induced enhanced expression of the shorter species which allowed normal morphological mammary differentiation. The short isoform lacks AU-rich motifs and miRNA target sequences that are potentially implicated in the regulation of mRNA stability and translation efficiency. We further determined that the AU-binding protein HuR appears to selectively stabilize the longer isoform in the mammary gland. In summary, the results of the present study identify a new regulatory instance involved in the fine-tuning of Itgb1 expression during mammary gland development and function.
PURPOSE: The purpose of this study was to evaluate the stability of health literacy in adults with diabetes over time. Understanding the dynamic nature of health literacy is important when tailoring health messages, especially those targeted at the management of chronic health conditions.
METHOD: This was a descriptive longitudinal study of 751 adults with diabetes randomly selected from primary care practices in the Vermont Diabetes Information System study between July 2003 and December 2007. Participants were interviewed and completed questionnaires upon entrance into the study and again 24 months later. Health literacy was measured with the Short Test for Functional Health Literacy of Adults. Participants also completed the SF-12 and the Self-Administered Comorbidity Questionnaire and self-reported their sex, income, education, marital status, race/ethnicity, health insurance, duration of diabetes, and problems with vision.
RESULTS: A significant decrease in health literacy was noted over 24 months. The largest decrease was in adults > =65 years of age and those with higher physical function at baseline. Smaller declines were noted for women and participants who were white, higher educated, poly-pharmacy users, and with fair to excellent vision.
CONCLUSIONS: Health literacy exhibits decline with increasing age among adults with diabetes. Individual variability in health literacy has implications for the best timing and approach to provide self-management education and support.
Metastatic carcinoma to the thyroid gland: a single institution 20-year experience and review of the literature
The thyroid gland is an uncommon site for metastatic disease but cases have been well-documented in the literature, particularly in autopsy series. A retrospective review of surgical pathology and autopsy pathology database for patients with metastatic carcinoma to the thyroid was performed at the University of Massachusetts Medical Center between January 1993 to January 2013. We identified a total of 10 patients with metastatic carcinoma to the thyroid; 6 were in surgical pathology specimens out of a total of 1,295 thyroid carcinoma (0.46 %) and 4 were diagnosed at autopsy out of a total of 2,117 (0.19 %) autopsy cases during this period. Cases with direct extension of the tumor into the thyroid from local primary sites such as larynx, esophagus or soft tissues of the neck were excluded. The primary tumors in these cases comprised of four lung carcinomas, three colorectal carcinomas, a renal cell carcinoma, a pleural malignant mesothelioma, and an unknown primary. Therefore, it is important to keep intrathyroidal metastases in the differential diagnosis when evaluating a thyroid nodule, particularly in patients with a previous history of malignancy. Furthermore, a literature review reveals over 1,400 cases have been previously reported, with the most common malignancies from the kidney (34 %), lung (15 %), gastrointestinal tract (14 %), and breast (14 %).
Patterns of pulmonary function tests (PFTs) and flow-volume loops among patients with clinically important tracheobronchomalacia (TBM) are not well described. Small studies suggest 4 main flow-volume loop morphologies: low maximum forced expiratory flow, biphasic expiratory curve, flow oscillations, and notching. We studied common PFT and flow-volume loop patterns among the largest prospective series of patients to date, undergoing clinical evaluation for symptomatic moderate to severe TBM. METHODS:
We conducted a retrospective analysis of prospectively collected data from patients who were referred to our Chest Disease Center from January 2002 to December 2008, with respiratory symptoms that were attributed primarily to TBM. The PFT results of 90 subjects with symptomatic moderate to severe TBM were evaluated. RESULTS:
By PFTs, 40 (44.4%) subjects had an obstructive ventilatory defect, 16 (17.8%) had a definite or highly likely restrictive ventilatory defect, 15 (16.7%) had a mixed defect, and 19 (21.1%) were within normal limits. Among 76 subjects with available flow-volume loops, the most frequent finding was low maximum forced expiratory flow, in 62 (81.6%) subjects, followed by biphasic morphology (15, 19.7%), notched expiratory loop (7, 9.2%), and expiratory oscillations (2, 2.6%). The balance of 13 subjects (17.1%) had no distinctive flow-volume loop abnormality. CONCLUSION:
PFTs and flow-volume loops are normal in a substantial number of patients with moderate to severe TBM, and should not be used to decide whether TBM is present or clinically important.
Loss of cilia suppresses cyst growth in genetic models of autosomal dominant polycystic kidney disease
Kidney cysts occur following inactivation of polycystins in otherwise intact cilia or following complete removal of cilia by inactivation of intraflagellar transport-related proteins. We investigated the mechanisms of cyst formation in these two distinct processes by combining conditional inactivation of polycystins with concomitant ablation of cilia in developing and adult kidney and liver. We found that loss of intact cilia suppressed cyst growth following inactivation of polycystins and that the severity of cystic disease was directly related to the length of time between the initial loss of the polycystin proteins and the subsequent involution of cilia. This cilia-dependent cyst growth was not explained by activation of the MAPK/ERK, mTOR or cAMP pathways and is likely to be distinct from the mechanism of cyst growth following complete loss of cilia. These data establish the existence of a new pathway defined by polycystin-dependent inhibition and cilia-dependent activation that promotes rapid cyst growth.
Extent and severity of coronary artery disease by coronary CT angiography is associated with elevated left ventricular diastolic pressures and worsening diastolic function
BACKGROUND: Patients with flow-limiting coronary stenoses exhibit elevated left ventricular end-diastolic pressure (LVEDP) and abnormal left ventricular (LV) relaxation.
OBJECTIVE: We investigated the relationship of extent and severity of coronary artery disease (CAD) by coronary CT angiography (CTA) to LVEDP and measures of LV diastolic dysfunction.
METHODS: We identified consecutive patients undergoing coronary CTA and transthoracic echocardiography who were assessed for diastolic function. CAD was evaluated on a per-patient, per-vessel, and per-segment basis for intraluminal diameter stenosis by using an 18-segment model (0 = none, 1 = 1%-49%, 2 = 50%-69%, and 3 = 70%-100%) and summed over segments to obtain overall coronary plaque burden (segment stenosis score [SSS]; maximum = 54). Transthoracic echocardiography evaluated mitral inflow E wave-to-A wave ratio, tissue Doppler early mitral annual tissue velocity axial excursion, stage of diastolic dysfunction, and LV dimensions and estimated LVEDP from the ratio of mitral inflow velocity to early mitral annular (medial) tissue velocity.
RESULTS: Four hundred seventy-eight patients (57% women; mean age, 57.9 +/- 14.6 years; 24.9% prior CAD) comprised the study population. Increasing per-patient maximal coronary stenosis, number of vessels with obstructive stenosis, and SSS were associated with increased LVEDP. The prevalence of advanced diastolic dysfunction increased with greater number of obstructive vessels. In multivariable analyses, SSS was associated with increased LVEDP (0.8 mm Hg per tertile increase in SSS, 0.5-1.1; P < .001); reduced E' axial excursion (-0.3; 95% confidence interval [CI], -0.5 to -0.1; P = .001), increased LV mass index (1.6 g/m(2) per tertile increase in SSS; P = .04), and increased relative wall thickness (0.005; 95% CI, 0.004-0.009; P = .03), with consistent relationships persisting even among persons with per-patient maximal stenosis < 50% and LV ejection fraction >>= 55%.
CONCLUSIONS: Extent and severity of obstructive as well as nonobstructive CAD by coronary CTA are associated with increased LVEDP and measures of diastolic dysfunction. Elsevier Inc. All rights reserved.
The bacteriophage VP3 is used in a phage-biotyping scheme as one of the typing phages of Vibrio cholerae O1 biotype El Tor strains. Here, we have sequenced and analyzed its genome. The genome consists of 39,481 bp with an overall G + C content of 42.6 %. Fifty-two open reading frames (ORFs) were predicted. Within the genome, 17 highly conserved phage promoters and 6 rho-independent terminators were predicted. When assessed with Rluc as a reporter gene, 12 of 16 cloned VP3 promoters showed activity in the host strain V. cholerae biotype El Tor. Based on the temporal expression pattern detected using reverse transcription PCR (RT-PCR), VP3 ORFs can be classed into four groups, arranged according to their order in the VP3 genome. Terminators T1 and T6 are presumed to work efficiently. Sequencing of the typing phage VP3 of V. cholerae reveals its evolutionary subdivisions from the members of T7-like phages of Escherichia coli. Knowledge of VP3 expands the known host range of T7-like phages and will promote understanding the different infection mechanisms used by members of this genus.
Activation of the Nlrp3 inflammasome in infiltrating macrophages by endocannabinoids mediates beta cell loss in type 2 diabetes
Type 2 diabetes mellitus (T2DM) progresses from compensated insulin resistance to beta cell failure resulting in uncompensated hyperglycemia, a process replicated in the Zucker diabetic fatty (ZDF) rat. The Nlrp3 inflammasome has been implicated in obesity-induced insulin resistance and beta cell failure. Endocannabinoids contribute to insulin resistance through activation of peripheral CB1 receptors (CB(1)Rs) and also promote beta cell failure. Here we show that beta cell failure in adult ZDF rats is not associated with CB(1)R signaling in beta cells, but rather in M1 macrophages infiltrating into pancreatic islets, and that this leads to activation of the Nlrp3-ASC inflammasome in the macrophages. These effects are replicated in vitro by incubating wild-type human or rodent macrophages, but not macrophages from CB(1)R-deficient (Cnr1(-/-)) or Nlrp3(-/-) mice, with the endocannabinoid anandamide. Peripheral CB(1)R blockade, in vivo depletion of macrophages or macrophage-specific knockdown of CB(1)R reverses or prevents these changes and restores normoglycemia and glucose-induced insulin secretion. These findings implicate endocannabinoids and inflammasome activation in beta cell failure and identify macrophage-expressed CB(1)R as a therapeutic target in T2DM.
Pathogenic Escherichia coli are genetically diverse and encompass a broad variety of pathotypes, such as enteroaggregative E. coli (EAEC) or enterohemorrhagic E. coli (EHEC), which cause distinct clinical syndromes. The historically large 2011 German outbreak of hemolytic uremic syndrome (HUS), caused by a Shiga-toxin producing E. coli (STEC) of the serotype O104:H4, illustrated the emerging importance of non-O157 STEC. STEC O104:H4, with features characteristic of both enteroaggregative E. coli and enterohemorrhagic E. coli, represents a unique and highly virulent pathotype. The German outbreak both allowed for the evaluation of several potential therapeutic approaches to STEC-induced HUS and emphasizes the importance of early and specific detection of both O157 and non-O157 STEC.
Identification of new drug targets is vital for the advancement of drug discovery against Mycobacterium tuberculosis, especially given the increase of resistance worldwide to first- and second-line drugs. Because traditional target-based screening has largely proven unsuccessful for antibiotic discovery, we have developed a scalable platform for target identification in M. tuberculosis that is based on whole-cell screening, coupled with whole-genome sequencing of resistant mutants and recombineering to confirm. The method yields targets paired with whole-cell active compounds, which can serve as novel scaffolds for drug development, molecular tools for validation, and/or as ligands for co-crystallization. It may also reveal other information about mechanisms of action, such as activation or efflux. Using this method, we identified resistance-linked genes for eight compounds with anti-tubercular activity. Four of the genes have previously been shown to be essential: AspS, aspartyl-tRNA synthetase, Pks13, a polyketide synthase involved in mycolic acid biosynthesis, MmpL3, a membrane transporter, and EccB3, a component of the ESX-3 type VII secretion system. AspS and Pks13 represent novel targets in protein translation and cell-wall biosynthesis. Both MmpL3 and EccB3 are involved in membrane transport. Pks13, AspS, and EccB3 represent novel candidates not targeted by existing TB drugs, and the availability of whole-cell active inhibitors greatly increases their potential for drug discovery.
A randomized controlled trial of mindfulness-based stress reduction for women with early-stage breast cancer receiving radiotherapy
PURPOSE: To testthe relative effectiveness of a mindfulness-based stress reduction program (MBSR) compared with a nutrition education intervention (NEP) and usual care (UC) in women with newly diagnosed early-stage breast cancer (BrCA)undergoing radiotherapy.
METHODS: Datawere available from a randomized controlled trialof 172 women, 20 to 65 years old, with stage I or II BrCA. Data from women completing the 8-week MBSR program plus 3 additional sessions focuses on special needs associated with BrCA were compared to women receiving attention control NEP and UC. Follow-up was performed at 3 post-intervention points: 4 months, and 1 and 2 years. Standardized, validated self-administered questionnaires were used to assess psychosocial variables. Descriptive analyses compared women by randomization assignment. Regression analyses, incorporating both intention-to-treat and post hoc multivariable approaches, were used to control for potential confounding variables.
RESULTS: A subset of 120 women underwent radiotherapy; 77 completed treatment prior to the study, and 40 had radiotherapy during the MBSR intervention. Women who actively received radiotherapy (art) while participating in the MBSR intervention (MBSR-art) experienced a significant (P < .05) improvement in 16 psychosocial variables compared with the NEP-art, UC-art, or both at 4 months. These included health-related, BrCA-specific quality of life and psychosocial coping, which were the primary outcomes, and secondary measures, including meaningfulness, helplessness, cognitive avoidance, depression, paranoid ideation, hostility, anxiety, global severity, anxious preoccupation, and emotional control.
CONCLUSIONS: MBSR appears to facilitate psychosocial adjustment in BrCA patients receiving radiotherapy, suggesting applicability for MBSR as adjunctive therapy in oncological practice.
Sonic hedgehog mediates the proliferation and recruitment of transformed mesenchymal stem cells to the stomach
Studies using Helicobacter-infected mice show that bone marrow-derived mesenchymal stem cells (MSCs) can repopulate the gastric epithelium and promote gastric cancer progression. Within the tumor microenvironment of the stomach, pro-inflammatory cytokine interferon-gamma (IFNgamma) and Sonic hedgehog (Shh) are elevated. IFNgamma is implicated in tumor proliferation via activation of the Shh signaling pathway in various tissues but whether a similar mechanism exists in the stomach is unknown. We tested the hypothesis that IFNgamma drives MSC proliferation and recruitment, a response mediated by Shh signaling. The current study uses transplantation of an in vitro transformed mesenchymal stem cell line (stMSC(vect)), that over-expresses hedgehog signaling, in comparison to non-transformed wild-type MSCs (wtMSCs), wtMSCs transfected to over-express Shh (wtMSC(Shh)), and stMSCs transduced with lentiviral constructs containing shRNA targeting the Shh gene (stMSC(ShhKO)). The effect of IFNgamma on MSC proliferation was assessed by cell cycle analysis in vitro using cells treated with recombinant IFNgamma (rmIFNgamma) alone, or in combination with anti-Shh 5E1 antibody, and in vivo using mice transplanted with MSCs treated with PBS or rmIFNgamma. In vitro, IFNgamma significantly increased MSC proliferation, a response mediated by Shh that was blocked by 5E1 antibody. The MSC population collected from bone marrow of PBS- or IFNgamma-treated mice showed that IFNgamma significantly increased the percentage of all MSC cell lines in S phase, with the exception of the stMSCs(ShhKO) cells. While the MSC cell lines with intact Shh expression were recruited to the gastric mucosa in response to IFNgamma, stMSCs(ShhKO) were not. Hedgehog signaling is required for MSC proliferation and recruitment to the stomach in response to IFNgamma.
BACKGROUND: Cells secrete different types of membrane vesicles (MVs), which may act as important entities in normal human physiology and in various pathological processes. The established methods for quantification of MVs require purification or preanalytical handling of samples with labeling moieties. AIM: The authors' aim was to develop a method for high-throughput, labeling-free quantification of nonpurified MVs.
MATERIALS and METHODS: Scanning ion occlusion sensing technology, which relies on the detection of particles upon their movement through a nanopore, was investigated for the ability to quantify nanosized MVs ( < 400 nm) in bodily fluids and cell culture supernatants.
RESULTS: Scanning ion occlusion sensing allowed for rapid and easy measurement of the concentration of MVs in all biological fluids tested.
CONCLUSION: Scanning ion occlusion sensing technology enables the quantification of MVs in biological samples without the requirement of MV isolation and/or labeling. This offers a highly valuable addition to the currently used repertoire of MV quantification methods.
Optimal selection of patients for elective abdominal aortic aneurysm repair based on life expectancy
OBJECTIVE: Elective abdominal aortic aneurysm (AAA) repair is beneficial when rupture is likely during a patient's expected lifetime. The purpose of this study was to identify predictors of long-term mortality after elective AAA repair for moderately sized AAAs ( < 6.5-cm diameter) to identify patients unlikely to benefit from surgery.
METHODS: We analyzed 2367 elective infrarenal AAA ( < 6.5 cm) repairs across 21 centers in New England from 2003 to 2011. Our main outcome measure was 5-year life-table survival. Cox proportional hazards analysis was used to describe associations between patient characteristics and 5-year survival.
RESULTS: During the study period, 1653 endovascular AAA repairs and 714 open AAA repairs were performed. Overall, 5-year survival rates were similar by procedure type (75% endovascular repair, 80% open repair; P = .14). Advanced age greater than or equal to 75 years (hazard ratio [HR], 2.0; P < .01) and age > 80 years (HR, 2.6; P < .01), coronary artery disease (HR, 1.4; P < .04), unstable angina or recent myocardial infarction (HR, 4.6; P < .01), oxygen-dependent chronic obstructive pulmonary disease (HR, 2.7; P < .01), and estimated glomerular filtration rate < 30 mL/min/1.73 m(2) (HR, 2.8; P < .01) were associated with poor survival. Aspirin (HR, 0.8; P < .03) and statin (HR, 0.7; P < .01) use were associated with improved survival. We used these risk factors to develop risk strata for low-risk, medium-risk, and high-risk groups with survival, respectively, of 85%, 69%, and 43% at 5 years (P < .001).
CONCLUSIONS: More than 75% of patients with moderately sized AAAs who underwent elective repair in our region survived 5 years, but 4% were at high risk for 5-year mortality. Patients with multiple risk factors, especially age > 80 years, unstable angina, oxygen-dependent chronic obstructive pulmonary disease, and estimated glomerular filtration rate < 30 mL/min/1.73 m(2), are unlikely to achieve sufficient long-term survival to benefit from surgery, unless their AAA rupture risk is very high.
Expectant management of cystotomy at the time of midurethral sling placement: a retrospective case series
INTRODUCTION AND HYPOTHESIS: Cystotomy is one of the most common complications of retropubic midurethral sling placement. Some centers manage cystotomy with prolonged catheter drainage, and there are few published studies evaluating this practice. The purpose of this study is to review postoperative outcomes of patients who experienced cystotomy at the time of sling placement and did not undergo prolonged catheter drainage.
METHODS: This is a retrospective review of all patients undergoing midurethral sling placement complicated by a cystotomy at the University of Rochester between 2004 and 2009. Outpatient and inpatient records were reviewed and data collected include demographics, intraoperative details, voiding trial results, postoperative complications, and voiding function. Descriptive statistics were performed.
RESULTS: Between 2004 and 2009, 30 subjects experienced a cystotomy of the 374 subjects that had a midurethral sling placed, all by a suprapubic approach. There were 25 patients who underwent a voiding trial on the day of surgery and 20 (80 %) were discharged home without prolonged drainage. Five subjects (20 %) had urinary retention and were discharged with an indwelling catheter. All five successfully voided within 4 days of discharge. No subject required subsequent catheterization for any reason and at the 6-week postoperative evaluation all subjects denied voiding dysfunction or irritative bladder symptoms. No subject required additional intervention and postoperative complications were rare.
CONCLUSIONS: In this study, the majority of subjects experiencing a cystotomy during midurethral sling placement were successfully discharged home the day of surgery without catheter drainage. The results suggest that prolonged catheter drainage after a cystotomy during midurethral sling placement may be unnecessary.