Acceptability of a companion patient guide to support expert consensus guidelines on surveillance mammography in older breast cancer survivors.

PURPOSE: To support shared decision-making, patient-facing resources are needed to complement recently published guidelines on approaches for surveillance mammography in breast cancer survivors aged ≥ 75 or with < 10-year life expectancy. We created a patient guide to facilitate discussions about surveillance mammography in older breast cancer survivors. METHODS: The "Are Mammograms Still Right for Me?" guide estimates future ipsilateral and contralateral breast (in-breast) cancer risks, general health, and the potential benefits/harms of mammography, with prompts for discussion. We conducted in-clinic acceptability testing of the guide by survivors and their clinicians at a National Cancer Institute-designated comprehensive cancer center, including two community practices. Patients and clinicians received the guide ahead of a clinic visit and surveyed patients (pre-/post-visit) and clinicians (post-visit). Acceptability was defined as ≥ 75% of patients and clinicians reporting that the guide (a) should be recommended to others, (b) is clear, (c) is helpful, and (d) contains a suitable amount of information. We also elicited feedback on usability and mammography intentions. RESULTS: We enrolled 45 patients and their 21 clinicians. Among those responding in post-visit surveys, 33/37 (89%) patients and 15/16 (94%) clinicians would recommend the guide to others; 33/37 (89%) patients and 15/16 (94%) clinicians felt everything/most things were clear. All other pre-specified acceptability criteria were met. Most patients reported strong intentions for mammography (100% pre-visit, 98% post-visit). CONCLUSION: Oncology clinicians and older breast cancer survivors found a guide to inform mammography decision-making acceptable and clear. A multisite clinical trial is needed to assess the guide's impact mammography utilization. TRIAL REGISTRATION: ClinicalTrials.gov-NCT03865654, posted March 7, 2019.

Comparison of Medication History Accuracy Between Nurses and Pharmacy Personnel.

PURPOSE:: To evaluate the differences in medication history errors made by pharmacy technicians, students, and pharmacists compared to nurses at a community hospital. METHODS:: One hundred medication histories completed by either pharmacy or nursing staff were repeated and evaluated for errors by a fourth-year pharmacy student. The histories were analyzed for differences in the rate of errors per medication. Errors were categorized by their clinical significance, which was determined by a panel of pharmacists, pharmacy students, and nurses. Errors were further categorized by their origin as either prescription (Rx) or over the counter (OTC). The primary outcome was the difference in the rate of clinically significant errors per medication. Secondary outcomes included the differences in the rate of clinically insignificant errors, Rx errors, and OTC errors. Differences in the types of errors for Rx and OTC medications were also analyzed. Additionally, the number of patients with no errors was compared between both groups. RESULTS:: The pharmacy group had a lower clinically significant error rate per medication (0.03 vs 0.09; relative risk [RR] = 0.66; 95% confidence interval [CI]: 0.020-0.093; P = .003). For secondary outcomes, the pharmacy group had a lower total error rate (0.21 vs 0.36, RR = 0.58; 95% CI: 0.041-0.255; P = .007), Rx error rate (0.09 vs 0.27, RR = 0.44; 95% CI: 0.071-0.292; P = .002), and OTC error rate (0.24 vs 0.46; RR = 0.52; 95% CI: 0.057-0.382; P = .009) per medication. The pharmacy group completed 20% more medication histories without Rx errors ( P = .045) and 25% more histories without OTC errors ( P = .041). CONCLUSION:: This study demonstrated that expanded use of pharmacy technicians and students improves the accuracy of medication histories in a community hospital.

Whole-genome sequences of Odocoileus hemionus deer adenovirus isolates from deer, moose and elk are highly conserved and support a new species in the genus Atadenovirus.

We present the first complete genome sequence of Odocoileus hemionus deer adenovirus 1 (OdAdV-1). This virus can cause sporadic haemorrhagic disease in cervids, although epizootics with high mortality have occurred in California. OdAdV-1 has been placed in the genus Atadenovirus, based on partial hexon, pVIII and fibre genes. Ten field isolates recovered from naturally infected mule deer (Odocoileus hemionus), white-tailed deer (Odocoileus virginiana) and moose (Alces alces) from Wyoming, black-tailed deer (Odocoileus hemionus columbianus) from California, and Rocky Mountain elk (Cervus elaphus nelsoni) from Colorado and Washington state were sequenced. The genome lengths ranged from 30 620 to 30 699 bp, contained the predicted proteins and gene organization typical of members of genus Atadenovirus, and had a high percentage of A/T nucleotides (66.7 %). Phylogenic analysis found that the closest ancestry was with ruminant atadenoviruses, while a divergence of the hexon, polymerase and penton base proteins of more than 15 % supports classification as a new species. Genetic global comparison between the 10 isolates found an overall 99 % identity, but greater divergence was found between those recovered from moose and elk as compared to deer, and a single variable region contained most of these differences. Our findings demonstrate that OdAdV-1 is highly conserved between 10 isolates recovered from multiple related cervid species, but genotypic differences, largely localized to a variable region, define two strains. We propose that the virus type name be changed to cervid adenovirus 1, with the species name Cervid atadenovirus A. Sequence data were used to develop molecular assays for improved detection and genotyping.

Characterisation of the antiproliferative constituents and activity of Ficus exasperata (Vahl) on ovarian cancer cells -a preliminary investigation.

Ovarian cancer is one of the most common gynaecological cancers today. This study therefore investigates the anticancer effects of Ficus exasperata extracts and fractions on ovarian cancer cells. The antiproliferative activity of the crude extracts (1 mg/mL) was assessed using the MTT assay on A2780 (ovarian cancer) cell line. Bio-activity guided fractionation was performed and preliminary identification was further achieved using high resolution mass spectrometry and nuclear magnetic resonance spectroscopy. All crude extracts tested exhibited antiproliferative activity except for the methanol extract which interestingly showed proliferative effects. Five fatty acids were identified from the active fractions (FB1-10 and FB1-12). FB1-12 exhibited an IC50 value of 15.20 µg/mL. The least potent fraction (FB1-4 + 5) had an IC50 value of 34.51 µg/mL. H1-HEX and H1-MET exhibited 97.2 and 97.9%, respectively, compared to control. This study therefore provides proof-of-principle that fatty acids of Ficus exasperata exhibit significant antiproliferative effects on ovarian cancer cells.

Plumage micro-organisms and preen gland size in an urbanizing context.

Urbanization of Earth's habitats has led to considerable loss of biodiversity, but the driving ecological mechanism(s) are not always clear. Vertebrates like birds typically experience urban alterations to diet, habitat availability, and levels of predation or competition, but may also be exposed to greater or more pathogenic communities of microbes. Birds have been popular subjects of urban ecological research but, to our knowledge, no study has assessed how urban conditions influence the microbial communities on bird plumage. Birds carry a large variety of microorganisms on their plumage and some of them have the capacity to degrade feather keratin and alter plumage integrity. To limit the negative effects of these feather-degrading bacteria, birds coat their feathers with preen gland secretions containing antibacterial substances. Here we examined urban-rural variation in feather microbial abundance and preen gland size in house finches (Haemorhous mexicanus). We found that, although urban and rural finches carry similar total-cultivable microbial loads on their plumage, the abundance of feather-degrading bacteria was on average three times higher on the plumage of urban birds. We also found an increase in preen gland size along the gradient of urbanization, suggesting that urban birds may coat their feathers with more preen oil to limit the growth or activity of feather-degrading microbes. Given that greater investment in preening is traded-off against other immunological defenses and that feather-degrading bacteria can alter key processes like thermoregulation, aerodynamics, and coloration, our findings highlight the importance of plumage microbes and microbial defenses on the ecology of urban birds.

Induction of Heterologous Tier 2 HIV-1-Neutralizing and Cross-Reactive V1/V2-Specific Antibodies in Rabbits by Prime-Boost Immunization.

UNLABELLED: Poxvirus prime-protein boost used in the RV144 trial remains the only immunization strategy shown to elicit a modest level of protection against HIV-1 acquisition in humans. Although neutralizing antibodies (NAb) were generated, they were against sensitive viruses, not the more resistant "tier 2" isolates that dominate circulating strains. Instead, risk reduction correlated with antibodies recognizing epitopes in the V1/V2 region of HIV-1 envelope glycoprotein (Env). Here, we examined whether tier 2 virus NAb and V1/V2-specific non-NAb could be elicited by a poxvirus prime-gp120 boost strategy in a rabbit model. We studied two clade B Envs that differ in multiple parameters, including tissue origin, neutralization sensitivity, and presence of the N197 (N7) glycan that was previously shown to modulate the exposure of conserved epitopes on Env. We demonstrate that immunized rabbits generated cross-reactive neutralizing activities against >50% of the tier 2 global HIV-1 isolates tested. Some of these activities were directed against the CD4 binding site (CD4bs). These rabbits also generated antibodies that recognized protein scaffolds bearing V1/V2 sequences from diverse HIV-1 isolates and mediated antibody-dependent cellular cytotoxicity. However, there are subtle differences in the specificities and the response rates of V1/V2-specific antibodies between animals immunized with different Envs, with or without the N7 glycan. These findings demonstrate that antibody responses that have been correlated with protection against HIV-1 acquisition in humans can be elicited in a preclinical model by a poxvirus prime-gp120 boost strategy and that improvements may be achievable by optimizing the nature of the priming and boosting immunogens. IMPORTANCE: The only vaccine approach shown to elicit any protective efficacy against HIV-1 acquisition is based on a poxvirus prime-protein boost regimen (RV144 Thai trial). Reduction of risk was associated with nonneutralizing antibodies targeting the V1/V2 loops of the envelope protein gp120. However, the modest efficacy (31.2%) achieved in this trial highlights the need to examine approaches and factors that may improve vaccine-induced responses, including cross-reactive neutralizing activities. We show here that rabbits immunized with a novel recombinant vaccinia virus prime-gp120 protein boost regimen generated antibodies that recognize protein scaffolds bearing V1/V2 sequences from diverse HIV-1 isolates and mediated antibody-dependent cellular cytotoxicity. Importantly, immunized rabbits also showed neutralizing activities against heterologous tier 2 HIV-1 isolates. These findings may inform the design of prime-boost immunization approaches and help improve the protective efficacy of candidate HIV-1 vaccines.

Immunogenicity of reduced dose priming schedules of serogroup C meningococcal conjugate vaccine followed by booster at 12 months in infants: open label randomised controlled trial.

OBJECTIVE: To determine whether the immunogenicity of a single dose infant priming schedule of serogroup C meningococcal (MenC) conjugate vaccine is non-inferior to a two dose priming schedule when followed by a booster dose at age 12 months. DESIGN: Phase IV open label randomised controlled trial carried out from July 2010 until August 2013 SETTING: Four centres in the United Kingdom and one centre in Malta. PARTICIPANTS: Healthy infants aged 6-12 weeks followed up until age 24 months. INTERVENTIONS: In the priming phase of the trial 509 infants were randomised in a 10:10:7:4 ratio into four groups to receive either a single MenC-cross reacting material 197 (CRM) dose at 3 months; two doses of MenC-CRM at 3 and 4 months; a single MenC-polysaccharide-tetanus toxoid (TT) dose at 3 months; or no MenC doses, respectively. Haemophilus influenzae type b (Hib)-MenC-TT vaccine was administered to all infants at 12 months of age. All infants also received the nationally routinely recommended vaccines. Blood samples were taken at age 5, 12, 13, and 24 months. MAIN OUTCOME MEASURE: MenC serum bactericidal antibody assay with rabbit complement (rSBA) one month after the Hib-MenC-TT vaccine. Non-inferiority was met if the lower 95% confidence limit of the difference in the mean log10 MenC rSBA between the single dose MenC-CRM and the two dose MenC-CRM groups was >-0.35. RESULTS: The primary objective was met: after a Hib-MenC-TT booster dose at 12 months of age the MenC rSBA geometric mean titres induced in infants primed with a single MenC-CRM dose were not inferior to those induced in participants primed with two MenC-CRM doses in infancy (660 (95% confidence interval 498 to 876) v 295 (220 to 398)) with a corresponding difference in the mean log10 MenC rSBA of 0.35 (0.17 to 0.53) that showed superiority of the single over the two dose schedule). Exploration of differences between the priming schedules showed that one month after Hib-MenC-TT vaccination, MenC rSBA ≥ 1:8 was observed in >96% of participants previously primed with any of the MenC vaccine schedules in infancy and in 83% of those who were not vaccinated against MenC in infancy. The MenC rSBA geometric mean titres induced by the Hib-MenC-TT boost were significantly higher in children who were primed with one rather than two MenC-CRM doses in infancy. Only priming with MenC-TT, however, induced robust MenC bactericidal antibody after the Hib-MenC-TT booster that persisted until 24 months of age. CONCLUSIONS: MenC vaccination programmes with two MenC infant priming doses could be reduced to a single priming dose without reducing post-boost antibody titres. When followed by a Hib-MenC-TT booster dose, infant priming with a single MenC-TT vaccine dose induces a more robust antibody response than one or two infant doses of MenC-CRM. Bactericidal antibody induced by a single Hib-MenC-TT conjugate vaccine dose at 12 months of age (that is, a toddler only schedule), without infant priming, is not well sustained at 24 months. Because of rapid waning of MenC antibody, programmes using toddler only schedules will still need to rely on herd protection to protect infants and young children.Trial registration Eudract No: 2009-016579-31; NCT01129518; study ID: 2008_06 (http://clinicaltrials.gov).

Immunological effect of administration of sequential doses of Haemophilus influenzae type b and pneumococcal conjugate vaccines in the same versus alternating limbs in the routine infant immunisation schedule: an open-label randomised controlled trial.

BACKGROUND: The use of different limbs for the administration of sequential doses of an intradermal rabies vaccine was shown to result in reduced vaccine immunogenicity. We aimed to assess whether this phenomenon also occurs with routine infant vaccines. METHODS: In this open-label, randomised, controlled study, eligible healthy infants 6-12 weeks of age recruited through five clinical trials units (four in the UK and one in Malta) were randomly assigned in a 1:1 ratio to two vaccination groups: consistent limb or alternating limb. Infants in the consistent limb group received the diphtheria-tetanus-acellular pertussis-inactived polio-Haemophilus influenzae type b combined vaccine (DTaP-IPV-Hib) at 2, 3, and 4 months of age, and the pneumococcal conjugate vaccine (PCV13) at 2, 4, and 12 months, all administered to the right leg. Infants in the alternating limb group received DTaP-IPV-Hib in the left leg at 2 months and in the right leg at 3 and 4 months; and PCV13 in the left leg at 2 months, in the right leg at 4 months, and in the left arm at 12 months. All infants in both groups received the combined H influenzae type b and capsular group C Neisseria meningitidis tetanus toxoid conjugate vaccine (Hib-MenC-TT), administered in the left leg at 12 months. Randomisation was achieved by randomly generated codes, with permuted block size of 30, and was stratified by study site. Group allocation was not masked from study staff and parents of participants after enrolment, but group allocation was masked from laboratory staff assessing blood samples. The current study was a prespecified secondary objective of a parent phase 4 trial that assessed the induction of immunity following varying schedules of vaccination with glyco-conjugate capsular group C Neisseria meningitidis (Men C) vaccines in infancy. The objective of the current study was to compare the immunogenicity and reactogenicity of vaccines delivered in either consistent or alternating limbs. Immunogenicity was assessed by comparing serum IgG geometric mean concentrations at 5, 12, 13, and 24 months, analysed per protocol. This study is registered with ClinicalTrials.gov, number NCT01129518. FINDINGS: Between July 5, 2010, and Aug 1, 2013, we enrolled 509 infants and randomly allocated them to the consistent limb group (n=254) or the alternating limb group (n=255). Anti-H influenzae type b anti-polyribosylribitol phosphate IgG geometric mean concentrations were lower in the consistent limb group than in the alternating limb group at 5 months (consistent limb 0·41 µg/mL [95% CI 0·31-0·54] vs alternating limb 0·61 µg/mL [0·45-0·82]; p=0·0268) and at 12 months (0·35 µg/mL [0·28-0·43] vs 0·50 µg/mL [0·40-0·62]; p=0·0136). Anti-tetanus toxoid antibody IgG geometric mean concentrations were lower in the consistent limb group (1·63 IU/mL [95% CI 1·40-1·90]) than in the alternating limb group (2·30 IU/mL [1·97-2·68]) at 13 months (p=0·0008) and at 24 months (0·44 IU/mL [0·37-0·52] vs 0·61 IU/mL [0·51-0·73]; p=0·0074). Anti-pneumococcal IgG geometric mean concentrations were similar between both groups at all timepoints. The proportions of participants who had adverse events did not differ between the two groups. INTERPRETATION: Use of different (alternating) limbs for sequential doses of routine infant vaccines does not reduce, and might enhance, immunogenicity. The underlying mechanism for this finding warrants further research. FUNDING: NIHR Oxford Biomedical Research Centre and GlaxoSmithKline Biologicals.

Evaluation of the induction of immune memory following infant immunisation with serogroup C Neisseria meningitidis conjugate vaccines--exploratory analyses within a randomised controlled trial.

AIM: We measured meningococcal serogroup C (MenC)-specific memory B-cell responses in infants by Enzyme-Linked Immunospot (ELISpot) following different MenC conjugate vaccine schedules to investigate the impact of priming on immune memory. METHODS: Infants aged 2 months were randomised to receive 1 or 2 doses of MenC-CRM197 at 3 or 3 and 4 months, 1 dose of MenC-TT at 3 months, or no primary MenC doses. All children received a Haemophilus influenzae type b (Hib)-MenC booster at 12 months. Blood was drawn at 5, 12, 12 months +6 days and 13 months of age. RESULTS: Results were available for 110, 103, 76 and 44 children from each group respectively. Following primary immunisations, and prior to the 12-month booster, there were no significant differences between 1- or 2-dose primed children in the number of MenC memory B-cells detected. One month following the booster, children primed with 1 dose MenC-TT had more memory B-cells than children primed with either 1-dose (p = 0.001) or 2-dose (p<0.0001) MenC-CRM197. There were no differences in MenC memory B-cells detected in children who received 1 or 2 doses of MenC-CRM197 in infancy and un-primed children. CONCLUSIONS: MenC-specific memory B-cell production may be more dependent on the type of primary vaccine used than the number of doses administered. Although the mechanistic differences between MenC-CRM197 and MenC-TT priming are unclear, it is possible that structural differences, including the carrier proteins, may underlie differential interactions with B- and T-cell populations, and thus different effects on various memory B-cell subsets. A MenC-TT/Hib-MenC-TT combination for priming/boosting may offer an advantage in inducing more persistent antibody. TRIAL REGISTRATION: EU Clinical Trials Register 2009-016579-31 ClinicalTrials.gov NCT01129518.

Water VBAC: exploring a new frontier for women's autonomy.

BACKGROUND: although Vaginal Birth After Caesarean section (VBAC) has been promoted successfully as one means of reducing the caesarean section rate, the practice of VBAC using water immersion (Water VBAC) is restricted. Very little valid, reliable research evidence is available on this birth method, although initial small-scale audits indicate that Water VBAC has no adverse effect on maternal and neonatal outcomes. METHOD: in-depth semi-structured interviews were carried out with a purposive sample of eight women who had undergone Water VBAC in one midwife-led unit. The interviews aimed to explore their reasons for requesting this birthing method, and their experience of the process. An interpretative phenomenological analytical approach was adopted. FINDINGS: the women pursued Water VBAC for two main reasons: in order to prevent a repeat of the obstetric events that previously led to a caesarean section, and to counteract their previous negative birth experiences. The women reported improved physical and psychological outcomes from their Water VBAC experience when compared with their previous experience of caesarean section. Three main themes emerged: 'minimising', 'maximising' and 'managing'. Water VBAC entailed an attempt to minimise the medicalisation of the women's childbirth experience. This was achieved by limiting medical staff input in favour of midwife-led care, which was believed to minimise negative physical and psychological experiences. Correspondingly, Water VBAC was perceived as maximising physical and psychological benefits, and as a means of allowing women to obtain choice and assert control over their labour and birth. The women planning a Water VBAC believed they had to manage the potential risks associated with Water VBAC, as well as manage the expectations and behaviour of friends, family and the health care professionals involved in their care. CONCLUSIONS: for the women participating in this research, actively pursuing Water VBAC constituted a means of asserting their autonomy over the childbirth process. The value accorded to being able to exercise choice and control over their childbearing experience was high. These women's accounts indicated that information-giving and shared decision-making require improvement, and that inconsistencies in the attitudes of health care professionals need to be addressed.

Calcium sensing receptor mutations implicated in pancreatitis and idiopathic epilepsy syndrome disrupt an arginine-rich retention motif.

Calcium sensing receptor (CaSR) mutations implicated in familial hypocalciuric hypercalcemia, pancreatitis and idiopathic epilepsy syndrome map to an extended arginine-rich region in the proximal carboxyl terminus. Arginine-rich motifs mediate endoplasmic reticulum retention and/or retrieval of multisubunit proteins so we asked whether these mutations, R886P, R896H or R898Q, altered CaSR targeting to the plasma membrane. Targeting was enhanced by all three mutations, and Ca(2+)-stimulated ERK1/2 phosphorylation was increased for R896H and R898Q. To define the role of the extended arginine-rich region in CaSR trafficking, we independently determined the contributions of R890/R891 and/or R896/K897/R898 motifs by mutation to alanine. Disruption of the motif(s) significantly increased surface expression and function relative to wt CaSR. The arginine-rich region is flanked by phosphorylation sites at S892 (protein kinase C) and S899 (protein kinase A). The phosphorylation state of S899 regulated recognition of the arginine-rich region; S899D showed increased surface localization. CaSR assembles in the endoplasmic reticulum as a covalent disulfide-linked dimer and we determined whether retention requires the presence of arginine-rich regions in both subunits. A single arginine-rich region within the dimer was sufficient to confer intracellular retention comparable to wt CaSR. We have identified an extended arginine-rich region in the proximal carboxyl terminus of CaSR (residues R890 - R898) which fosters intracellular retention of CaSR and is regulated by phosphorylation. Mutation(s) identified in chronic pancreatitis and idiopathic epilepsy syndrome therefore increase plasma membrane targeting of CaSR, likely contributing to the altered Ca(2+) signaling characteristic of these diseases.

Calcium-sensing receptor biosynthesis includes a cotranslational conformational checkpoint and endoplasmic reticulum retention.

Metabolic labeling with [(35)S]cysteine was used to characterize early events in CaSR biosynthesis. [(35)S]CaSR is relatively stable (half-life approximately 8 h), but maturation to the final glycosylated form is slow and incomplete. Incorporation of [(35)S]cysteine is linear over 60 min, and the rate of [(35)S]CaSR biosynthesis is significantly increased by the membrane-permeant allosteric agonist NPS R-568, which acts as a cotranslational pharmacochaperone. The [(35)S]CaSR biosynthetic rate also varies as a function of conformational bias induced by loss- or gain-of-function mutations. In contrast, [(35)S]CaSR maturation to the plasma membrane was not significantly altered by exposure to the pharmacochaperone NPS R-568, the allosteric agonist neomycin, or the orthosteric agonist Ca(2+) (0.5 or 5 mm), suggesting that CaSR does not control its own release from the endoplasmic reticulum. A CaSR chimera containing the mGluR1alpha carboxyl terminus matures completely (half-time of approximately 8 h) and without a lag period, as does the truncation mutant CaSRDelta868 (half-time of approximately 16 h). CaSRDelta898 exhibits maturation comparable with full-length CaSR, suggesting that the CaSR carboxyl terminus between residues Thr(868) and Arg(898) limits maturation. Overall, these results suggest that CaSR is subject to cotranslational quality control, which includes a pharmacochaperone-sensitive conformational checkpoint. The CaSR carboxyl terminus is the chief determinant of intracellular retention of a significant fraction of total CaSR. Intracellular CaSR may reflect a rapidly mobilizable "storage form" of CaSR and/or may subserve distinct intracellular signaling roles that are sensitive to signaling-dependent changes in endoplasmic reticulum Ca(2+) and/or glutathione.

Pharmacochaperone-mediated rescue of calcium-sensing receptor loss-of-function mutants.

The calcium sensing receptor (CaSR) is a Family C/3 G protein-coupled receptor that translates changes in extracellular Ca(2+) into diverse intracellular signals. Loss-of-function mutations in human CaSR cause familial hypocalciuric hypercalcemia and neonatal severe hyperparathyroidism. CaSR must navigate a number of endoplasmic reticulum quality control checkpoints during biosynthesis, including a conformational/functional checkpoint. Here we examine the biosynthesis of 25 CaSR mutations causing familial hypocalciuric hypercalcemia /neonatal severe hyperparathyroidism using immunoprecipitation, biotinylation, and functional assays. We define classes of CaSR mutants based on their biosynthetic profile. Class I CaSR mutants are not rescued to the plasma membrane. To dissect the organellar compartments that class I mutants can access, we engineered a cleavage site for the proprotein convertase furin into the extracellular domain of wild-type CaSR and class I mutants. Based on absence or presence of cleavage fragments, we find most mutants are degraded from the endoplasmic reticulum (no furin-mediated cleavage), whereas others access the Golgi (furin-mediated cleavage) before degradation. Class II CaSR mutants show increased expression and/or enhanced plasma membrane localization upon treatment with MG132 or the pharmacochaperone NPS R-568, permitting assay of functional activity. Of the 10 CaSR mutants that exhibit plasma membrane localization, only two did not show enhanced functional activity after rescue with NPS R-568. The established approaches can be used with current and newly identified CaSR mutations to identify the location of biosynthetic block and to determine the likelihood of rescue by allosteric agonists.

Effect of effort pain after upper abdominal surgery on two independent measures of respiratory function.

STUDY OBJECTIVE: To determine how effort pain interacts with changing pulmonary function after upper abdominal incisions. DESIGN: Prospective, case-controlled study. SETTING: Academic teaching hospital. PATIENTS: 34 ASA physical status I, II, and III patients recovering from elective, major incisional, upper abdominal surgery. MEASUREMENTS: Manometry (maximal inspiratory and expiratory pressure) and spirometry (forced vital capacity, forced expiratory volume during the first second, peak expiratory flow) for three postoperative days. Pain scores (Visual Analog Pain Scale; VAS) at rest and after the manometric or spirometric efforts. MAIN RESULTS: Effort pain during either manometry or spirometry was greater than pain at rest on the first postoperative day. Maximal respiratory pressure concomitantly recovered with pain during daily efforts (slopes: -0.429 and -0.278% max/mm VAS; P < 0.05). Spirometric measurements showed minimal improvement. CONCLUSION: The direct relationship between resolution of pain with effort and direct measures of respiratory muscle effort using manometry, but not those obtained less directly by spirometry, suggests that assessing interactions between pain and effort requires a direct, quantifiable measure of effort.

DNA prime/protein boost immunization against HIV clade C: safety and immunogenicity in mice.

The induction of both cellular and humoral immunity is an important goal for vaccine development against HIV. As a step towards the development of an efficacious vaccine against HIV clade C, the world's most prevalent strain, a combination DNA prime/protein boost immunization strategy was tested. A DNA expression vector was prepared encoding a codon-optimized env gene derived from a pediatric HIV clade C isolate, 1084i. Mice were immunized with HIV1084i env-encoding DNA, then boosted with homologous 1084i gp160. HIV1084i Env-specific T-cell responses were induced with DNA vaccination alone, but the strongest cellular immune responses were seen after boosting with gp160. Immunization with gp160 alone induced high-titer antibodies but required two inoculations. In contrast, high-titer antibodies were seen after a single 1084i gp160 boost in DNA-primed animals. All animals given gp160 inoculations, whether DNA primed or not, developed neutralizing antibodies reactive with HIV1084i and a macaque-passaged simian/human immunodeficiency construct, SHIV-1084ip. The results demonstrate the utility of this DNA prime/protein boost approach to generate cellular immunity, as well as neutralizing antibodies, against HIV clade C env antigens.

C-F Bond activation at Ni(0) and simple reactions of square planar Ni(II) fluoride complexes.

The reaction of Ni(COD)(2)(COD = 1,5-cyclooctadiene) with triethylphosphine and pentafluoropyridine in hexane has been shown previously to yield trans-[NiF(2-C(5)NF(4))(PEt(3))(2)](1a) with a preference for reaction at the 2-position of the heteroaromatic. The corresponding reaction with 2,3,5,6-tetrafluoropyridine was shown to yield trans-[NiF(2-C(5)NF(3)H)(PEt(3))(2)](1b). In this paper, we show that reaction of Ni(COD)(2) with triethylphosphine and pentafluoropyridine in THF yields a mixture of 1a and 1b. Competition reactions of Ni(COD)(2) with triethylphosphine in the presence of mixtures of heteroaromatics in hexane reveal a kinetic preference of k(pentafluoropyridine):k(2,3,5,6-tetrafluoropyridine)= 5.4:1. Treatment of 1a and 1b with Me(3)SiN(3) affords trans-[Ni(N(3))(2-C(5)NF(4))(PEt(3))(2)](2a) and trans-[Ni(N(3))(2-C(5)NHF(3))(PEt(3))(2)](2b), respectively. The complex trans-[Ni(NCO)(2-C(5)NHF(3))(PEt(3))(2)](3b) is obtained on reaction of with Me(3)SiNCO and by photolysis of under CO, while trans-[Ni(eta(1)-C [triple bond CPh)(2-C(5)NF(4))(PEt(3))(2)](4a) is obtained by reaction of phenylacetylene with 1a. Addition of KCN, KI and NaOAc to complex 1a affords trans-[Ni(X)(2-C(5)NF(4))(PEt(3))(2)](5a X = CN, 6a X = I, 7a X = OAc), respectively. The PEt(3) groups of complex are readily replaced by addition of 1,2-bis(dicyclohexylphosphino)ethane (dcpe) to produce [NiF(2-C(5)F(4)N)(dcpe)](8a). Addition of dcpe to trans-[Ni(OTf)(2-C(5)F(4)N)(PEt(3))(2)](10a), however, yields the salt [Ni(2-C(5)F(4)N)(dcpe)(PEt(3))](OTf)(9a) by substitution of only one PEt(3) and displacement of the triflate ligand. The structures of 2b, 4a, 7a and 8a were determined by X-ray crystallography. The influence of different ancillary ligands on the bond lengths and angles of square-planar nickel structures with polyfluoropyridyl ligands is analysed.

Waldenström's macroglobulinemia.

Waldenström's macroglobulinemia (WM) is a rare monoclonal gammopathy. Its clinical signs and symptoms include fatigue, weakness, hepatomegaly, splenomegaly, lymphadenopathy, and neuropathies. Patients with WM have a circulating tumor marker, the monoclonal immunoglobulin M protein. High levels of this protein can produce hyperviscosity syndrome, which often is characterized by bleeding from the mucous membranes of the nose and mouth. Asymptomatic patients with WM usually are not treated. Treatment of symptomatic patients and patients with relapsed WM may include alkylating agents, particularly chlorambucil; purine nucleoside analogs, such as fludarabine and cladribine; and, most recently, the use of rituximab. With knowledge about this unusual disease, oncology nurses can provide better care and education for patients with WM.