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This study provides further evidence for the impact of the aromas of plant essential oils on aspects of cognition and mood in healthy participants. One hundred and forty-four volunteers were randomly assigned to conditions of ylang-ylang aroma, peppermint aroma, or no aroma control. Cognitive performance was assessed using the Cognitive Drug Research computerized assessment battery, with mood scales completed before and after cognitive testing. The analysis of the data revealed significant differences between conditions on a number of the factors underpinning the tests that constitute the battery. Peppermint was found to enhance memory whereas ylang-ylang impaired it, and lengthened processing speed. In terms of subjective mood peppermint increased alertness and ylang-ylang decreased it, but significantly increased calmness. These results provide support for the contention that the aromas of essential oils can produce significant and idiosyncratic effects on both subjective and objective assessments of aspects of human behavior. They are discussed with reference to possible pharmacological and psychological modes of influence.

Background: Parathyroid hormone (PTH) replacement has been demonstrated to be superior to conventional treatment with calcium supplementation and vitamin D analogs for the treatment of hypoparathyroidism. In this investigation we evaluated the feasibility of using PTH microsphere encapsulation as a potential delivery system for PTH., Materials and Methods: Using the spontaneous emulsion technique, PTH microspheres were created by encapsulating PTH (1-34) in a copolymer of polyglycolic and polylactic acid (PLGA). Additional microspheres were constructed by coencapsulating calmodulin with PTH (1-34) in the PLGA microspheres. Microsphere production was confirmed using electron microscopy. PTH release was measured in vitro using an enzyme-linked immunosorbent assay. The bioactivity of PTH released from the microspheres was confirmed in vivo using a hypoparathyroid rat model by measuring serum calcium concentrations before and 3 h after subcutaneous injection of PTH microspheres., Results: PTH microsphere and PTH/calmodulin microspheres could be created using the spontaneous emulsion technique. Physiologically significant PTH release was measured in vitro for 20 days. PTH release was calcium sensitive and exhibited negative feedback. This effect was augmented by coencapsulation with calmodulin. PTH released from the microspheres caused a significant rise in serum calcium levels from an average of 6.35 (6.19-6.48 mg/dL) to 8.55 mg/dL (8.22-8.73). PTH released from the PTH/calmodulin microspheres resulted in an increase in serum calcium from a mean of 6.8 (6.7-6.9 mg/dL) to 8.1 mg/dL (7.8-8.2)., Conclusions: The PLGA microspheres can be used to provide calcium sensitive controlled release of biologically active PTH and offer a potential mean of providing biomimetic hormone replacement therapy.

The objective of the study was to test for an association between type 2 diabetes mellitus (T2DM) and body mass index (BMI) and three single nucleotide polymorphisms (SNP)s in the peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1alpha) gene. We were also interested in whether these associations differed by tertiles of diet, physical activity or presence of polymorphisms in the peroxisome proliferator-activated receptor gamma (PPAR-gamma) gene among Hispanics and Non-Hispanic Whites (NHW) from Colorado. We studied 216 Hispanic pedigrees (1850 nuclear families) and 236 NHW pedigrees (1240 families) from the San Luis Valley and Denver. We genotyped the Gly482Ser, Thr528Thr and Thr612Met polymorphisms in the PGC-1alpha gene and the Pro12Ala polymorphism of the PPAR-gamma gene. Historical physical activity (average METS/week) as well as average dietary intake over the past year was assessed by self-report. Data were analyzed using the Family Based Association Test (FBAT) as well as generalized estimating equations (GEE). We did not find any significant association between three SNPs in the PGC-1alpha gene and T2DM in Hispanics or NHW; however, using FBAT, we found the common Thr612Thr allele of the PGC-1alpha gene to be associated with T2DM among Hispanic subjects carrying the rare Pro12Ala allele of the PPAR-gamma gene (p=.003). We found similar associations when we considered a haplotype containing that allele (p=.002). However, the results of the GEE analysis did not confirm these findings: odds ratio (OR)=1.68, 95% CI (0.5, 5.2) suggesting these results may due to chance. BMI also did not show any consistent associations with the PGC-1alpha gene. In conclusion, we did not find an association between the PGC-1alpha gene and T2DM or BMI and there were no consistent interactions with diet, physical activity or the Pro12Ala polymorphism of the PPAR-gamma gene.

The order Microsporidia contains a number of ubiquitous pathogens that can infect various animals, including humans. Enterocytozoon bieneusi and Encephalitozoon intestinalis have been associated with gastrointestinal illness in humans. The effect of four disinfectants--ammonium hydroxide, hydrogen peroxide, and two commercial disinfectants containing peroxyacetic acid (Tsunami) and N-alkyl dimethyl benzyl ammonium chloride (Timsen)--on E. intestinalis spores was examined using exposure times of 1, 5, and 15 min. Spore viability was determined in vitro with RK-13 cells. Hydrogen peroxide was most efficient at inactivating microsporidial spores at all tested concentrations and treatment times, whereas ammonium hydroxide was effective only at the highest concentration at all exposure times. Tsunami (40 microg/ml) and Timsen (200 and 400 ppm) could inactivate spores when incubated for 5 and 15 min.

Background/aims: First to determine if body mass index (BMI) or waist circumference (WC) differ by possession of the Ala allele of the peroxisome proliferator-activated receptor gamma (PPARgamma) Pro12Ala polymorphism, and second, to determine if dietary fat intake and physical activity moderate these potential relationships among Hispanic Americans from Colorado., Methods: We studied 216 Hispanic pedigrees (1,850 nuclear families) from the San Luis Valley and Denver (mean age 50 years). The families were genotyped for the Pro12Ala polymorphism of the PPARgamma gene. BMI, WC and diabetes status were measured in the clinic, and, for diabetics, self-reported BMI at diabetes diagnosis was used. Data were analyzed using the Family-Based Association Test (FBAT). We conducted overall and stratified analyses by tertiles of monounsaturated, polyunsaturated (PUFA) and saturated (SFA) fat intake, the PUFA:SFA ratio, and physical activity. We also used generalized estimating equations (GEE) to further explore the FBAT findings., Results: The average BMI and WC were approximately 29 kg/m(2) and 101 cm, respectively. The frequency of the Ala allele was 13.5%. We did not find the Ala allele to be associated with BMI or WC overall (p = 0.58 and 0.23, respectively). We did find the Ala allele to be associated with increased BMI in those with high dietary PUFA and PUFA:SFA ratio (p = 0.007 and 0.01, respectively); however, these findings were not confirmed using the GEE models., Conclusions: One previous study has reported the Ala allele to be associated with higher BMI among Hispanics; however, we did not find this association. Further, we could not conclude that dietary fat intake or physical activity altered any association between BMI or WC and the Ala allele., (Copyright 2007 S. Karger AG, Basel.)

Endothelial cells respond to vascular endothelial growth factor (VEGF) to produce new blood vessels. This process of angiogenesis makes a critical contribution during embryogenesis and also in the response to ischemia in adult tissues. We have studied the intracellular trafficking of the major VEGF receptor KDR (VEGFR2). Unlike other related growth factor receptors, we find that a significant proportion of KDR is held in an endosomal storage pool within endothelial cells. We find that KDR can be delivered to the plasma membrane from this intracellular pool and that VEGF stimulates this recycling to the cell surface. KDR recycling appears to be distinct from the previously characterized Rab4- and Rab11-dependent pathways, but, instead, KDR(+) recycling vesicles contain Src tyrosine kinase and VEGF-stimulated recycling requires Src activation. Taken together, these data show that intracellular trafficking of KDR is markedly different from other receptor tyrosine kinases and suggest that the regulation of KDR trafficking by VEGF provides a novel mechanism for controlling the sensitivity of endothelial cells to proangiogenic signals.

If the question "What is a gene?'' proves to be worth asking it must be able to elicit an answer which both recognizes and address the reasons why the concept of the gene ever seemed to be something worth getting excited about in the first place as well analyzing and evaluating the latest develops in the molecular biology of DNA. Each of the preceding papers fails to do one of these and suffers the consequences. Where Rolston responds to the apparent failure of molecular biology to make good on the desideratum of the classical gene by veering off into fanciful talk about "cybernetic genes,'' Griffiths and Stotz lose themselves in the molecular fine print and forget to ask themselves why "genes'' should be of any special interest anyway.

This project brought together a team of researchers and decision-makers to conduct policy-relevant research to support the introduction of advanced nursing practice roles in British Columbia. All team members, including decision-makers, were actively involved in the conceptualization, design, data collection, analysis and interpretation of the study. This level of engagement, coupled with ongoing knowledge translation (KT) activities, led to the implementation by stakeholders of a majority of the study's recommendations. The results have since been used to guide legislative and regulatory development and to design a nurse practitioner education program.

Statement of the Problem: Issues regarding the use of composite preheating need to be investigated so that the clinician will better understand the variables associated with this method., Purpose: To examine the multiple aspects of use of a commercial composite preheating device (Calset, AdDent Inc., Danbury, CT, USA)., Materials and Methods: Temperature values of three heating units and composite compules were obtained using a K-type thermocouple and were recorded digitally in real time. The following parameters were measured: maximum heater and composite temperature and its stability upon storage, composite temperature change when removed from the heater and injected, the effect of delivery system on ejected composite temperature, and the effect of repeated and extended preheating on composite monomer conversion (using infrared spectroscopy). Monomer conversion was measured after repeated composite cycling (from room temperature [RT] to 60 degrees C, 10x) or extended preheating (24 hours at 60 degrees C), and values were compared with composite maintained at RT (control group). Among test parameters, data (N=5 for each parameter) were analyzed using Student's t-test, analysis of variance, and the Tukey-Kramer post-hoc test where appropriate (alpha=0.05)., Results/conclusions: Two of the three tested units achieved the stated preset temperatures. Composite attained temperature values close to the heating unit. Composite temperature drop upon removal from the heater was dramatic: within 2 minutes a 50% temperature drop was noted. Heating the compule while preloaded in the syringe provided higher delivery temperatures than heating the compule separately (p < 0.00). Optimum results were achieved when preheated composite was dispensed and used as quickly as possible. Neither repeated nor extended preheating of composite significantly affected monomer conversion., Clinical Significance: Preheating composite has potential benefits, but should be used with knowledge of its limitations. Reheating of unused composite does not affect its degree of conversion, thus decreasing material waste. Heating of the composite preloaded in the delivery syringe enhances the temperature of extruded composite.

An atraumatic clavicular fracture presented after radical treatment for laryngeal carcinoma. This presented a diagnostic dilemma. The differential diagnosis included metastatic bone disease and osteomyelitis as well as post-radiotherapy complications. After investigation, the cause was thought to be a post-radiation fracture of the clavicle and to the best knowledge of the authors, this is the first ever documented in a patient who had undergone a total laryngectomy with bilateral modified radical neck dissections and post-operative radiotherapy. Cases of a fractured clavicle post-radiation have been most commonly documented in patients with breast cancer and only a few cases have been documented in patients with laryngeal cancer treated with a total laryngectomy, bilateral radical neck dissections and radiotherapy.

Background: Brain metastases are the most common intracranial tumour in adults, estimated to occur in up to 40% of patients with cancer. Despite being used in clinical practice for 50 years, the effectiveness of whole brain radiotherapy for the treatment of brain metastases remains uncertain., Objectives: To assess the effectiveness of whole brain radiotherapy (WBRT) on survival and quality of life. To identify whether patient performance status, number of brain metastases, extent of extracranial disease and primary site of cancer are important effect modifiers., Design: Systematic literature review., Methods: Electronic searches of four databases, augmented by hand searches of the most frequently encountered journal and assessment of the reference lists of consensus statements and all retrieved papers. Included papers underwent structured data extraction, assessment and qualitative synthesis., Results: Thirty-two primary studies were included, with a range of study designs, methodological quality, pre-treatment variables, interventions and outcome measures. From the limited evidence available, survival appeared to increase when patients were selected by performance status (survival increasing from approximately three to seven months in high performance status groups, as defined by Karnofsky performance status or Recursive Partitioning Analysis classification). The evidence suggests no survival benefit when patients with poor performance status were treated with whole brain radiotherapy. No studies undertook direct measurement of patients' quality of life. Surrogate measures of patients' quality of life, such as improvement in neurological function or improvement/maintenance of KPS > or =70, produced response rates ranging from 7 to 90%., Conclusion: The heterogeneity of study designs, quality and outcomes necessitates caution in interpreting the review findings. WBRT appears to be of benefit in higher performance status patients but not in low performance status patients. This suggests a basis for current practice, however further robust trial evidence is required.

Background: Serum antibodies against thyroglobulin (TgAbs) are common in patients with differentiated thyroid cancer (DTC) and can interfere in thyroglobulin (Tg) assays. We identified the epitopes on Tg recognized by TgAb-positive sera from patients with DTC and examined the association between epitope specificity patterns and Tg recovery., Methods: We tested 50 DTC sera for Tg epitope specificity, TgAbs, and Tg recovery. Epitope recognition was determined by use of a panel of 10 well-characterized Tg monoclonal antibodies directed against 6 Tg antigenic clusters (I-VI) in competitive reactions with test sera. Tg was measured by the Thyroglobuline IRMA (CIS bio international). Recovery of added Tg (TgREC) was determined by an in-house assay., Results: Epitope recognition was restricted to immunodominant clusters in 58% of patients, whereas the rest were either broadly heterogeneous (16%) or nonreactive (26%). Median Tg recovery did not differ between sera with restricted and unrestricted specificities (69% vs 80%; P >0.05). TgREC was inversely correlated with the total number of epitopes recognized by sera (r = -0.66; P <0.001)., Conclusions: TgAbs with both restricted and broad specificities are present in patients with DTC. TgAb interference is related to the number of epitopes recognized by sera rather than the pattern of epitope recognition.

There has been ongoing confusion about the meaning of advanced nursing practice (ANP) and the nature of ANP roles in Canada and elsewhere. A broad range of roles and titles have been adopted throughout Canada in an attempt to delineate specialized and/or advanced roles within nursing. One key objective in a recent three-phase study of ANP in British Columbia was to clarify the role and understanding of advanced nursing practice and related roles within the larger healthcare system. Our intent in this paper is to describe the understandings of ANP that emerged in Phase I of this recent study and to compare registered nurses' understandings of ANP to the characteristics and competencies identified by the Canadian Nurses Association (2002) framework. (Note: The term "nurse," as used in this paper, refers to "registered nurse.") We conclude by identifying future directions for development of advanced practice roles.

We assessed the contribution of facility-level variables to antimicrobial use in a cohort of 50 nursing homes and found that antimicrobial use was significantly correlated with the percentage of nursing home residents with feeding tubes, the number of healthcare aides, and the country of origin of the facility.

To determine factors that predispose or protect healthcare workers from severe acute respiratory syndrome (SARS), we conducted a retrospective cohort study among 43 nurses who worked in two Toronto critical care units with SARS patients. Eight of 32 nurses who entered a SARS patient's room were infected. The probability of SARS infection was 6% per shift worked. Assisting during intubation, suctioning before intubation, and manipulating the oxygen mask were high-risk activities. Consistently wearing a mask (either surgical or particulate respirator type N95) while caring for a SARS patient was protective for the nurses, and consistent use of the N95 mask was more protective than not wearing a mask. Risk was reduced by consistent use of a surgical mask, but not significantly. Risk was lower with consistent use of a N95 mask than with consistent use of a surgical mask. We conclude that activities related to intubation increase SARS risk and use of a mask (particularly a N95 mask) is protective.

As the number of older adults in acute care rises, acute care specialty nurses must gain gerontological knowledge in their nursing practice with older patients. A blend of two bodies of knowledge and competencies is required to care effectively for older adults in the hospital. To this end, a gerontological enrichment program was developed to improve care to hospitalized older patients by augmenting acute care nurses competencies with gerontological skill, knowledge, and abilities. This article will explain the implementation details, the anticipated outcomes for nurses and older acutely ill patients, the evaluation process, and the strategies that are in place to maintain the gains.

Exposure to Pb in-utero and in infancy has been associated with cognitive risk, even at low blood Pb levels of 0.48-0.96 micromol L(-1). Based on the hypothesis that wild game consumed by First Nation women of Mushkegowuk Territory contains Pb that might be transferred through blood or breast-milk to the fetus or infant, the study's objectives were to describe: (1) Pb in maternal and cord blood at birth and infant blood at 4 months, and (2) dietary influence on Pb status. Cord and maternal Pb were 0.10 +/- 0.08 and 0.11 +/- 0.06 micromol L(-1) (x +/- SD), respectively, and were significantly correlated (r = 0.77, p < 0.0001, n = 70), as was infant blood Pb (0.08 +/- 0.05 micromol L(-1)) with matched cord blood (r = 0.65, p < 0.0001, n = 30). Two cord blood samples (3%) were above 0.48 micromol L(-1). Pb in breast-milk at 0.010 +/- 0.008 micromol L(-1) (n = 25), was significantly lower than Pb in commercial formula or evaporated milk-based feedings (range 0.02-0.05 micromol L(-1), p < 0.05), and correlated with matched maternal blood Pb (r = 0.55, p < 0.005). However, in a sub-sample of infants (n = 31), blood Pb was similar for breastfed and formula-fed groups, though above the evaporated milk-fed group (p < 0.05). Maternal consumption of wild fowl, mammals and fish, estimated from the previous year, provided, respectively, 128 +/- 124, 46 +/- 68 and 8 +/- 13 MJ annum(-1). Traditional animal food intake, especially wild fowl, correlated significantly with cord blood Pb (Spearman rank correlation, p = 0.017). Although blood and milk Pb levels were largely within acceptable ranges, the presence of some elevated levels and association between blood Pb and traditional game consumption may reflect the legacy of using lead-containing ammunition.

The problem of how to reconcile the apparent "purposiveness" of the living organism with nonteleological, mechanist modes of explanation was given a certain form through most of the 20th century by a relatively decontextualized understanding of the gene as the heritable determinant of phenotypic traits. As instrumentally preformationist presuppositions about genes give way to the burgeoning elucidation of cell and molecular mechanisms of epigenesis, basic questions about the nature of complex living systems and their evolutionary origins once again come into consideration. Some suggestions are offered for a vision of the genetically recontextualized organism.

Objective: To determine modifiable risk factors for nosocomial Clostridium difficile-associated diarrhea (CDAD)., Design: Case-control study., Setting: 300-bed tertiary-care hospital., Participants: Hospital inpatients present during the 3-month study period., Methods: Case-patients identified with nosocomial CDAD over the study period were compared to two sets of control patients: inpatients matched by age, gender, and date of admission; and inpatients matched by duration of hospital stay. Variables including demographic data, comorbid illnesses, antibiotic exposure, and use of gastrointestinal medications were assessed for case- and control-patients. Conditional logistic regression was performed to identify risk factors for nosocomial CDAD., Results: 27 case-patients were identified and were compared to the two sets of controls (1:1 match for each comparison set). For the first set of controls, use of ciprofloxacin (odds ratio [OR], 5.5; 95% confidence interval [CI 95], 1.2-24.8; P=.03) was the only variable that remained significant in the multivariable model. For the second set of controls, prior exposure to cephalosporins (OR, 6.7; CI 95, 1.3-33.7; P=.02) and to ciprofloxacin (OR, 9.5; CI 95, 1.01-88.4; P=.05) were kept in the final model., Conclusions: Along with cephalosporins, prior quinolone use predisposed hospitalized patients to nosocomial CDAD. Quinolones should be used judiciously in acute-care hospitals, particularly in those where CDAD is endemic.

Glucose regulates proinsulin biosynthesis via stimulation of the translation of the preproinsulin mRNA in pancreatic beta-cells. However, the mechanism by which this occurs has remained unclear. Using recombinant adenoviruses that express the preproinsulin mRNA with defined alterations, the untranslated regions (UTRs) of the preproinsulin mRNA were examined for elements that specifically control translation of the mRNA in rat pancreatic islets. These studies revealed that the preproinsulin 5'-UTR was necessary for glucose stimulation of preproinsulin mRNA translation, whereas the 3'-UTR appeared to suppress translation. However, together the 5'- and 3'-UTRs acted cooperatively to markedly increase glucose-induced proinsulin biosynthesis. In primary hepatocytes the presence of the preproinsulin 3'-UTR led to reduced mRNA levels compared with the presence of the SV40 3'-UTR, consistent with the presence of mRNA stability determinants in the 3'-UTR that stabilize the preproinsulin mRNA in a pancreatic beta-cell-specific manner. Translation of these mRNAs in primary hepatocytes was not stimulated by glucose, indicating that regulated translation of the preproinsulin mRNA occurs in a pancreatic beta-cell-specific manner. Thus, the untranslated regions of the preproinsulin mRNA play crucial roles in regulating insulin production and therefore glucose homeostasis by regulating the translation and the stability of the preproinsulin mRNA.

Widespread application of EBPM by bedside providers is needed to demonstrate the success of pain management strategies on patient outcomes. This goal is not easy to attain and generally requires time, patience, and a multidisciplinary team approach. Implementation and evaluation of pain interventions increase awareness and knowledge of pain management strategies and can result in an overall improvement in pain management. The literature and guidelines recommend the use of specific strategies to ensure practice change. Studies suggest that a more intensive or "active" effort to alter practice is generally most successful. The pain management program should be marketed so that both the health care providers and patients are aware of the goal and resources available. It may take 3 to 5 years to infuse the change and see the improvement. Reinfusion over time also needs to be planned. Bedside practitioners need to have knowledge of the current best evidence in pain management of the critically ill patient. Barriers to implementation must be eliminated so that practitioners can conscientiously and judiciously implement strategies to relieve pain. Opinion leaders and change agents need to be available to continually champion EBPM, and prompts to ask about pain should be provided to practitioners and patients.

Purpose: To prospectively compare full-field digital mammography (FFDM) with screen-film mammography (SFM) for cancer detection in a screening population., Materials and Methods: At two institutions, 4,945 FFDM examinations were performed in women aged 40 years and older presenting for SFM. Two views of each breast were acquired with each modality. SFM and FFDM images were interpreted independently. Findings detected with either SFM or FFDM were evaluated with additional imaging and, if warranted, biopsy., Results: Patients in the study underwent 152 biopsies, which resulted in the diagnosis of 35 breast cancers. Twenty-two cancers were detected with SFM and 21 with FFDM. Four were interval cancers that became palpable within 1 year of screening and were considered false-negative findings with both modalities. The difference in cancer detection rate was not significant. FFDM had a significantly lower recall rate (11.5%; 568 of 4,945) than SFM (13.8%; 685 of 4,945) (P <.001, McNemar chi(2) model; P <.03, generalized estimating equations model). The positive biopsy rate for findings detected with FFDM (30%; 21 of 69) was higher than that for findings detected with SFM (19%; 22 of 114), but this difference was not significant., Conclusion: No difference in cancer detection rate has yet been observed between FFDM and SFM. FFDM has so far led to fewer recalls than SFM.

Because cocaine- and amphetamine-regulated transcript (CART) coexists with alpha-melanocyte stimulating hormone (alpha-MSH) in the arcuate nucleus neurons and we have recently demonstrated that alpha-MSH innervates TRH-synthesizing neurons in the hypothalamic paraventricular nucleus (PVN), we raised the possibility that CART may also be contained in fibers that innervate hypophysiotropic thyrotropin-releasing hormone (TRH) neurons and modulate TRH gene expression. Triple-labeling fluorescent in situ hybridization and immunofluorescence were performed to reveal the morphological relationships between pro-TRH mRNA-containing neurons and CART- and alpha-MSH-immunoreactive (IR) axons. CART-IR axons densely innervated the majority of pro-TRH mRNA-containing neurons in all parvocellular subdivisions of the PVN and established asymmetric synaptic specializations with pro-TRH neurons. However, whereas all alpha-MSH-IR axons in the PVN contained CART-IR, only a portion of CART-IR axons in contact with pro-TRH neurons were immunoreactive for alpha-MSH. In the medial and periventricular parvocellular subdivisions of the PVN, CART was co-contained in approximately 80% of pro-TRH neuronal perikarya, whereas colocalization with pro-TRH was found in <10% of the anterior parvocellular subdivision neurons. In addition, >80% of TRH/CART neurons in the periventricular and medial parvocellular subdivisions accumulated Fluoro-Gold after systemic administration, suggesting that CART may serve as a marker for hypophysiotropic TRH neurons. CART prevented fasting-induced suppression of pro-TRH in the PVN when administered intracerebroventricularly and increased the content of TRH in hypothalamic cell cultures. These studies establish an anatomical association between CART and pro-TRH-producing neurons in the PVN and demonstrate that CART has a stimulatory effect on hypophysiotropic TRH neurons by increasing pro-TRH gene expression and the biosynthesis of TRH.

The crystal structure of recombinant wild-type green fluorescent protein (GFP) has been solved to a resolution of 1.9 A by multiwavelength anomalous dispersion phasing methods. The protein is in the shape of a cylinder, comprising 11 strands of beta-sheet with an alpha-helix inside and short helical segments on the ends of the cylinder. This motif, with beta-structure on the outside and alpha-helix on the inside, represents a new protein fold, which we have named the beta-can. Two protomers pack closely together to form a dimer in the crystal. The fluorophores are protected inside the cylinders, and their structures are consistent with the formation of aromatic systems made up of Tyr66 with reduction of its C alpha-C beta bond coupled with cyclization of the neighboring glycine and serine residues. The environment inside the cylinder explains the effects of many existing mutants of GFP and suggests specific side chains that could be modified to change the spectral properties of GFP. Furthermore, the identification of the dimer contacts may allow mutagenic control of the state of assembly of the protein.

We have generated transgenic (Tg) lines of zebrafish in which the green fluorescent protein (GFP)-encoding gfp cDNA is driven by the Xenopus laevis ef1 alpha enhancer/promoter; Tg embryos from most of these lines show detectable fluorescence throughout their body. We have investigated the copy number of the Tg genes in fluorescent and non-fluorescent lines, in order to determine how this affects the production of detectable levels of GFP in the zebrafish embryo. Additionally, we have injected purified recombinant GFP into embryos to determine the intracellular GFP concentration required for detection, both when all of the cells in the embryo contain GFP and when only a few do.

The diagnostic accuracy of transtelephonic pacemaker monitoring (TTM) has been quantified in a retrospective study involving 369 patients in three U.S. cardiac centers. Using existing medical records, TTM findings in a total of 413 reports were judged for equivalence to the findings of subsequent physical examinations in pacemaker clinics. This study found TTM follow-up testing to have a sensitivity of 94.6%, specificity of 98.5%, positive predictive value of 93.3%, and negative predictive value of 98.8%. The study also documents the clinical utility of TTM in identifying various modes of pacemaker malfunctions and instances of significant arrhythmia.

Type 1 diabetes mellitus is caused by severe insulin deficiency secondary to the autoimmune destruction of pancreatic beta cells. Patients need to be controlled by periodic insulin injections to prevent the development of ketoacidosis, which can be fatal. Sustained, low-level expression of the rat insulin 1 gene from the liver of severely diabetic rats was achieved by in vivo administration of a recombinant retroviral vector. Ketoacidosis was prevented and the treated animals exhibited normoglycemia during a 24-hr fast, with no evidence of hypoglycemia. Histopathological examination of the liver in the treated animals showed no apparent abnormalities. Thus, the liver is an excellent target organ for ectopic expression of the insulin gene as a potential treatment modality for type 1 diabetes mellitus by gene therapy.

Mutations in the RIPE3a element have shown it to be crucial for efficient tissue-specific expression of the insulin gene. In order to isolate factors binding to this element, we used a labeled RIPE3 probe to screen an expression library derived from a hamster insulinoma cell line. We isolated a clone encoding beta-cell E-box transcriptional activator1 (BETA 1). This clone is a member of the class A subfamily of the helix-loop-helix superfamily of transcriptional activators, as determined both by sequence analysis and by functional association with a class B member (myogenin). This clone is related to, but distinct from, other clones isolated from the same library which are also capable of binding RIPE3a. Analysis showed these additional clones to be the hamster homologs of E12 and E47 (German, M. S., Blaner, M. A., Nelson, C., Moss, L. G., and Rutter, W. J. (1991) Mol. Endocrinol. 5, 292-299). Antibodies were raised against BETA 1 and against a common epitope of E12 and E47 to determine which proteins were contained in the native RIPE3a binding complex. Using these antibodies, we were able to separate the complex into major and minor fractions which contained either E12/47 or BETA 1, respectively. Thus, these two gene products are found in separate fractions of the tissue-specific binding activity and are therefore both likely to be important in insulin gene regulation.

Cytotrophoblasts are the specialized epithelial cells of the placenta. During the first trimester, a subpopulation of chorionic villus cytotrophoblasts differentiates along an invasive pathway and penetrates the maternal endometrium, decidua and spiral arterioles. Cytotrophoblast invasiveness declines rapidly during the second half of gestation. Isolated cytotrophoblasts of different gestational ages retain this differential invasiveness in culture. To determine whether the properties of integrin receptors for extracellular matrix molecules differ between invasive and non-invasive cytotrophoblasts, detergent extracts of isolated cytotrophoblasts of different gestational ages, and of first-trimester villous fibroblasts, were immunoprecipitated with subunit-specific anti-beta 1 integrin antibodies. Striking alterations in electrophoretic mobility were observed in beta 1 integrins from first-trimester cytotrophoblasts, as compared with those from term cytotrophoblasts or first-trimester villous fibroblasts, suggesting a cell-type-specific, temporally regulated alteration in glycosylation. Treatment of total first-trimester cytotrophoblast beta 1 integrins or the isolated alpha 5/beta 1 fibronectin receptor with endo-beta-galactosidase restored electrophoretic mobility to control levels, suggesting the presence of polylactosamine-bearing oligosaccharides. Further analysis by enzyme digestion and lectin-affinity chromatography suggested that they consisted of at least three antennae and short, sialylated lactosamine units. These oligosaccharides did not affect the affinity of the first-trimester cytotrophoblast fibronectin receptor for fibronectin. However, this receptor bound more strongly to wheat germ agglutinin than control fibronectin receptor and resisted elution by high concentrations of sugar hapten, requiring ionic detergent for removal. These results suggested that the altered glycosylation affected the conformation of the fibronectin receptor.

Several studies indicate that mammographic evaluation of the breast is inhibited by both submammary and submuscular implants, regardless of implant composition. Two hundred radiographs were evaluated following subpectoral placement of 25 combination gel/saline and 25 saline-filled tissue expanders. Within 3 months after augmentation, mammograms were performed using the standard and displacement techniques; patients with inflated implants were followed by a repeat displacement study (with the expander deflated). After completion of the mammogram, expanders were reinflated. Our experience demonstrates that better visualization of breast tissue may be accomplished by implant-deflated views in patients with small amounts of breast tissue, tight skin envelopes, or capsular contraction. In each of these circumstances, the displaced view failed to adequately demonstrate glandular tissue. The presence of the inflated implant impairs mammographic visualization in a number of ways, including implant opacity, impaired automatic exposure control, interference with breast compressibility, and implant displacement. These effects are consistently present in small breasts (< an estimated 300 g), in breasts with grade III or IV capsular contracture, and in breasts with minimal skin laxity (i.e., nonptotic breasts). Breast compression was compromised to a greater degree in the presence of grade III and IV capsular contracture; however, deflation of an adjustable implant optimized radiographic interpretation to a high degree. The deflatable saline implant yields a less obscured evaluation of the augmented breast. We suggest that an extended-use deflatable device (i.e., tissue expander) be considered as an option in our search for a safe and reliable implant for augmentation mammoplasty, particularly in patients with small breasts, tight skin envelopes, or a propensity for capsular contracture.

The authors describe the process and outcome of implementing a research-based pain management protocol in four adult critical care units at a large, Midwestern tertiary care center. The project was initiated and directed by members of the divisional research committee. Strategies used to change practice included determining if pain management was a problem via quality assessment monitors, surveying nurses regarding their knowledge and attitude toward pain management, educating staff members about the research base for the practice change, using change champions in each unit, and developing a core group of nurses in each unit to facilitate the change. Outcomes of this research utilization project include a 41% decline in the number of patients in pain, a 44% decline in pain intensity, and improvement in nurses knowledge about pain.

Body composition analysis is an important component of nutritional assessment in cystic fibrosis (CF). No gold standard of measurement exists, and techniques applicable to healthy populations may be unsuitable for CF patients. We assessed lean body mass (LBM) in 12 children with CF by skinfold (SK) measurements, bioelectrical impedance analysis (BIA), and dual-photon absorptiometry (DPA) and repeated these measures in 10 subjects 6 mo later. SK and DPA measures in eight older CF patients and eight healthy controls were compared to evaluate any effect of disease on estimates of LBM by use of DPA. Good agreement between the measures was seen at baseline and 6 mo by use of concordance plots. However, the limits of agreement between measures ranged up to 19% of SK-derived LBM measures (baseline: SK and DPA, 2.63 to -3.93 kg; SK and BIA, 2.36 to -1.24 kg; BIA and DPA, 1.88 to -4.28 kg; 6 mo: SK and DPA, 2.10 to -3.58 kg; SK and BIA, 6.28 to -5.49 kg; BIA and DPA, 5.53 to -7.79 kg). The change in LBM over 6 mo did not correlate among the three measures. Only BIA change in LBM correlated with weight change (r = 0.716, P < 0.02), probably due to the inclusion of weight in the regression equations for determining LBM from impedance. The relationship between SK and DPA measures did not differ between the CF and control groups, suggesting that there was no effect of disease on the DPA measure. The results suggest that none of these methods is precise enough to follow short-term changes in the nutritional status of CF patients longitudinally.

Eight patients with mild to severe tardive dyskinesia (TD) were treated for 12 weeks with buspirone in dosages of up to 180 mg/day in an open-label study. Changes in TD severity were assessed by the Abnormal Involuntary Movement Scale. The rater was blind to buspirone dosage. Buspirone was well tolerated by most subjects. A within-subjects comparison of pretreatment and post-treatment Abnormal Involuntary Movement Scale scores revealed a mean improvement of 4.4 (p < 0.01). Improvement was also observed in neuroleptic-induced extrapyramidal side effects such as parkinsonism and akathisia. Scores on the Hamilton Rating Scale for Anxiety and the Brief Psychiatric Rating Scale did not change during the 12-week study. The results of this open-label pilot study suggest that relatively high doses of buspirone may be efficacious in the treatment of TD.

At the authors' institution, needle localization of breast lesions with a braided hook wire involves the wire being cut 1-2 cm from the point of entry before dissection, to avoid contamination of the sterile field with the nonsterile portion of wire. During dissection, the wire is brought through the skin into the area of dissection. In one patient, fragments of wire filaments were left within the breast. Braided hook wires must be cut cleanly, the cut surface should be wiped before dissection, and the surgical area should be cleansed before closure.

Relationships among nutritional status and skeletal and respiratory muscle function were examined in 16 children with cystic fibrosis (CF) and mild lung disease (FEV1 95 +/- 16% predicted). Subjects were randomly assigned to receive (or not) noninvasive nutritional supplementation at 25% of normal energy recommendations for 6 mo. Skeletal muscle strength and power were similar to those of healthy children as were respiratory muscle strength and endurance. Stepwise-regression analysis indicated that changes in skeletal muscle strength and energy intake correlated significantly with growth [weight (kg) = 1.90 - 0.60 (Tanner Stage) + 0.49 (maximum voluntary strength (Nm) + 0.03 (energy intake, % RNI), r = 0.76, P < 0.05], though body composition, protein biochemistry, muscle power, respiratory muscle strength, and use of dietary supplements did not. Thus, changes in skeletal muscle strength may be a functional index of changes in nutritional status in CF. Dietary supplementation per se was not associated with functional improvement.

To study the molecular regulation of voltage-dependent Ca2+ channels (VDCCs) in the beta-cell, we have cloned a cDNA for the alpha 1-subunit from a hamster insulin-secreting cell line (HIT-T15). The cDNA (HCa3a) encodes a 1610-amino acid protein with four repeating membrane domains and an overall structure characteristic of other alpha 1-subunits. Although the cDNA shows a high degree of sequence homology (97%) with a rat brain alpha 1-subunit (RB alpha 1), the C-terminal 15 amino acids of HCa3a share no similarity with any cloned alpha 1 protein. High stringency Northern blot analysis revealed a single transcript of approximately 8.6 kilobases in HIT cells and hamster pancreas. A similarly sized species was detected in hamster brain, heart, and skeletal muscle. Using polymerase chain reaction and a primer set unique to HCa3a, this alpha 1 isoform was found to be expressed in islet cell lines derived from rat, mouse, and hamster. The HIT cell alpha 1-subunit is also expressed in discrete regions of the rat central nervous system, including the cortex, cerebellum, hypothalamus, and brain stem. The expression of two alpha 1 isoforms (HCa3a and cardiac) in the HIT cell underscores the possible complexity of VDCCs in the regulation of beta-cell signal transduction. With its widespread tissue distribution, HCa3a does not conform to the current classification system used for L-type VDCCs; this suggests that an alternative system of classification is required.

Systematic mutational analysis previously identified two primary regulatory elements within a minienhancer (-247 to -198) of the rat insulin I promoter that are critical for transcriptional activity. The Far box (-241 to -232) and the FLAT element (-222 to -208) synergistically upregulate transcription and, together, are sufficient to confer tissue-specific and glucose-responsive transcriptional activity on a heterologous promoter. Detailed analysis of the FLAT element further revealed that, in addition to the positive regulatory activity it mediates in tandem with the Far box, it is a site for negative regulatory control. A portion of the FLAT element bears considerable sequence similarity to the consensus binding site for hepatocyte nuclear factor 1 alpha (HNF1 alpha; LF-B1) a liver-enriched homeodomain-containing transcription factor. Here we show that the HNF1-like site within the FLAT element exhibited positive transcriptional activity in both HepG2 and HIT cells and bound similar, but distinguishable, nuclear protein complexes in the respective nuclear extracts. Screening of a hamster insulinoma cDNA library with a PCR-derived probe encompassing the DNA-binding domain of rat HNF1 alpha resulted in isolation of a hamster HNF1 alpha (hHNF1 alpha) cDNA homolog. Specific antiserum identified the HNF1 alpha protein as one component of a specific FLAT-binding complex in HIT nuclear extracts. Expression of the hHNF1 alpha cDNA in COS cells resulted in transactivation of reporter constructs containing multimerized segments of the rat insulin I minienhancer. Thus, HNF1 alpha, one component of a DNA-binding complex involved in transcriptional regulation of the rat insulin I gene, may play a significant role in nonhepatic as well as hepatic gene transcription.

The pancreatic beta cell makes several unique gene products, including insulin, islet amyloid polypeptide (IAPP), and beta-cell-specific glucokinase (beta GK). The functions of isolated portions of the insulin, IAPP, and beta GK promoters were studied by using transient expression and DNA binding assays. A short portion (-247 to -197 bp) of the rat insulin I gene, the FF minienhancer, contains three interacting transcriptional regulatory elements. The FF minienhancer binds at least two nuclear complexes with limited tissue distribution. Sequences similar to that of the FF minienhancer are present in the 5' flanking DNA of the human IAPP and rat beta GK genes and also the rat insulin II and mouse insulin I and II genes. Similar minienhancer constructs from the insulin and IAPP genes function as cell-specific transcriptional regulatory elements and compete for binding of the same nuclear factors, while the beta GK construct competes for protein binding but functions poorly as a minienhancer. These observations suggest that the patterns of expression of the beta-cell-specific genes result in part from sharing the same transcriptional regulators.

Pathologic hair-pulling (trichotillomania) has been described in the dermatologic and psychiatric literature for the last century, but has become the focus of increased attention in the last few years. Once thought to be either a wholly benign condition or a symptom of obsessive-compulsive disorder, more recently hair-pulling has been noted in the context of numerous types of psychopathology, and has been reported to respond to several different types of intervention, both psychological and somatic. Estimates of its prevalence have varied widely. Because of the disparate conclusions of the literature on this condition, a more careful assessment of diagnostic validity is recommended.

Myelin turnover has been studied in the 25-day-old HPH-5 mouse, a phenylalanine hydroxylase-deficient mouse mutant. The half-life of the fast component of myelin decreased from 15 days in control mice to 4.5 days at blood phenylalanine levels of 2.5 mmol/L. The slow component of myelin seems also to be affected by the high phenylalanine level. These observations confirm similar observations obtained with chemically induced models of hyperphenylalaninaemia and are therefore due to the hyperphenylalaninaemia per se, independently of the inhibitors of phenylalanine hydroxylase. An intermediate blood level of phenylalanine (0.7 mmol/L) likewise seems to interfere with myelin metabolism, although to a lesser degree.

Cell-specific expression of the insulin gene is dependent on a conserved 8-basepair sequence, GCCATCTG, present in two copies in the 5' flanking DNA of the rat insulin 1 gene (Nir and Far elements). A protein factor with well characterized binding affinities binds to this sequence and is unique to the nuclei of insulin-producing cells. Using the Nir element as a probe to screen a hamster insulinoma cDNA expression library, we cloned two cDNA inserts that encode two related helix-loop-helix DNA-binding proteins: Syrian hamster Pan-1 (shPan-1) and Syrian hamster Pan-2 (shPan-2). These clones have minimal differences from the previously reported human E47/E12 and rat PAN (rPan) DNA-binding proteins. In vitro translated protein products of both clones bound the insulin gene promoter Nir and far elements as well as the E2 elements of the mu heavy chain and kappa light chain immunoglobulin genes. Treating insulinoma cell nuclear extract with antiserum selectively directed to each of the two shPan proteins demonstrated the presence of each form of shPan in separate DNA-binding complexes, which together form the previously described, cell-specific, Nir element-binding complex. We conclude that shPan-1 and shPan-2 are the hamster homologs of the ubiquitous E47/E12 and rPan proteins, but form parts of distinct DNA-binding complexes apparently found only in the nuclei of insulin-producing cells.

In the pancreatic beta cells the proximal step in sulfonylurea signal transduction is the binding of these clinically important drugs to high-affinity receptors in the beta cell membrane. Using HIT cells as a model system, we have established an extremely close correlation between the affinity of binding of glyburide and its analog, iodoglyburide, and the activation of various steps in stimulus-secretion coupling--inhibition of 86Rb+ efflux, increase in [Ca2+]i resulting from gating of voltage-gated calcium channels by cell depolarization, and the exocytosis of insulin. Two different L-type channel cDNAs have been identified in an HIT cell library, one neuroendocrine in type and one more cardiac-like. A HIT cell membrane protein of Mr 140,000, which we believe to be the high-affinity sulfonylurea receptor, can be covalently linked to 5(125)-iodo-2-hydroxyglyburide by ultraviolet irradiation. The receptor has been solubilized and retains binding activity and the same rank order of displacement of the 5(125)-iodo-2-hydroxyglyburide as observed with the native receptor. The Mr 140,000 protein has been partially purified and the amino acid sequences of three proteolytic fragments have been used to design oligonucleotides to screen HIT cell cDNA libraries. Since the binding constant of glyburide or iodoglyburide is closely correlated with the ability of these compounds to inhibit the ATP-sensitive K+ channel, increase [Ca2+]i, and elicit insulin secretion, we have identified the Mr 140,000 protein as the sulfonylurea receptor. Expression of the cloned cDNA should allow us to test this hypothesis directly.

To study the regulation of insulin gene expression by physiological regulators, primary cultures of rat islet cells were transfected with portions of the rat insulin I gene 5'-flanking sequence linked to the reporter gene chloramphenicol acetyltransferase (CAT). Incubation of the cells in increasing glucose concentrations led to a parallel increase in both CAT activity and CAT mRNA levels. Pretreatment of the cells with the beta-cell-specific toxin streptozotocin reduced CAT activity 97%. Beta-Cell-specific expression of CAT was also demonstrated by co-staining the transfected cells with antisera to both CAT and insulin. Experiments showing a reduction in the response to glucose in the presence of the calcium channel blocker verapamil suggest that calcium plays a role in the glucose response, possibly via regulation of factors interacting with this limited portion of the insulin gene.

We screened a HIT (hamster insulin-secreting tumor) cell cDNA library constructed in lambda gt11 with a Go-specific oligonucleotide probe and isolated six recombinant phages. The inserts of these phages encoded two forms of alpha o, called here alpha o1 and alpha o2. The deduced amino acid sequence of alpha o1 is identical in all of its 354 amino acids to that reported previously for rat and bovine alpha o; that of alpha o2, also of 354 amino acids, is identical to alpha o1 up to and including amino acid 248 and differs thereafter in 26 amino acids. At the nucleotide level, alpha o1 and alpha o2 are identical up to and including the second base of the codon that specifies amino acid 243 and differs thereafter in 88 nucleotides of the remaining open reading frame and has no similarity to alpha o1 in its 3'-untranslated region. We propose that alpha o1 and alpha o2 result as a consequence of alternative splicing of a single alpha o transcript. Northern analysis with specifically designed oligonucleotides indicates that both forms of alpha o are expressed in normal tissues, e.g. brain. After in vitro transcription and translation, the peptides encoded in the alpha o1 and alpha o2 cDNAs could be ADP-ribosylated by pertussis toxin in the presence of added beta gamma dimers. The count of distinct G proteins keeps increasing.