Patient experience in systemic lupus erythematosus: development of novel patient-reported symptom and patient-reported impact measures.

BACKGROUND: Comprehensive assessment of systemic lupus erythematosus (SLE) and its treatment requires patient-reported outcome (PRO) measures to capture impacts and fluctuating symptoms. The objective of this study was to develop PROs, in accordance with the Food and Drug Administration (FDA) PRO Guidance, to assess fluctuations in SLE symptoms and its impact. METHODS: Following independent review board approval, six US rheumatology practices recruited patients with SLE to participate in concept elicitation (CE) interviews, in order to identify important SLE symptoms and their impacts. The SLE Symptom Severity Diary (SSD) and SLE Impact Questionnaire (SIQ) were drafted based on CE interview results and clinician input. The PROs were revised based on patient feedback from cognitive debriefing (CD) interviews, clinician feedback, and a translatability assessment. RESULTS: Forty-one patients completed CE interviews. Commonly-reported symptoms included fatigue (98%), joint pain (93%), and rash (88%). The most frequently reported impact was difficulty with chores/housework (61%). Eighteen patients completed CD interviews. The PROs were considered comprehensive, clear, and relevant.The SSD contains 17 items assessing energy/vitality, joint and muscle pain/stiffness/swelling, flu-like symptoms, cognition, numbness/tingling, skin symptoms and hair loss using an 11-point numeric response scale and a 24-h recall period (with the exception of hair loss). It also evaluates steroid status and dose. The SIQ contains 50 items, uses a 5-point Likert scale and a 7-day recall period, to assess disease impacts including patients' ability to make plans, work, and physical/social/emotional functioning. CONCLUSION: The SSD and SIQ are comprehensive SLE-specific PROs developed in accordance with the FDA PRO Guidance. Following assessment of their measurement properties, they may be useful in clinical studies and clinical practice to measure fluctuations in, and the impact of, symptoms in patients with SLE.

Response to belimumab among patients with systemic lupus erythematosus in clinical practice settings: 24-month results from the OBSErve study in the USA.

OBJECTIVE: To examine disease activity and clinical outcomes, and describe overall patterns of systemic lupus erythematosus (SLE) care in patients who received belimumab in a real-world clinical setting. METHODS: This observational cohort study was conducted in US clinical practices. Rheumatologists (n=92) identified adults with SLE who had received ≥8 infusions of belimumab plus standard of care (SoC). Physicians assessed disease outcomes at 6-month intervals using patient medical charts, for up to 24 months. The primary outcome was physician-assessed change in SLE disease. Other outcomes included change in steroid use, laboratory tests and healthcare resource utilisation (HCRU). RESULTS: Of 501 patients (intent-to-treat population (ITT)), 446 were female, mean age was 43.3 years and 98% had moderate/severe disease activity at baseline (first dose of belimumab). Data for 277 patients who completed 24 months of belimumab treatment were available. Among the ITT, a ≥50% improvement in overall clinical response between baseline and month 6 was reported for 48.7% of patients; continued improvement was seen at all subsequent 6-month intervals relative to the previous timepoint. The percentage of patients with moderate/severe disease also decreased at each timepoint. At baseline, 77.0% of patients received steroids at a mean (SD) prednisone equivalent dose of 19.9 (14.39) mg/day, which decreased to 8.4 (7.35) mg/day at month 6 and 6.1 (9.31) mg/day at month 24. Abnormal laboratory values typically associated with SLE also demonstrated improvements at month 6, which continued through 24 months. HCRU decreased over the duration of the study. CONCLUSIONS: Patients with SLE who received belimumab plus SoC for up to 24 months demonstrated improvements in disease severity and laboratory values and a reduction in steroid use and HCRU as early as month 6. Improvements continued through 24 months, providing evidence of reduced disease activity among patients taking belimumab in real-world clinical practice.

Randomised clinical trial: vercirnon, an oral CCR9 antagonist, vs. placebo as induction therapy in active Crohn's disease.

BACKGROUND: Many patients with active Crohn's disease do not adequately respond to therapies, highlighting the need for new treatments. AIMS: To conduct a randomised, double-blind, placebo-controlled phase 3 study to assess the efficacy and safety of vercirnon, an oral inhibitor of CC chemokine receptor-9, for the treatment of patients with moderately-to-severely active Crohn's disease. METHODS: Patients with a Crohn's Disease Activity Index (CDAI) of 220-450, plus evidence of active disease (endoscopically confirmed or elevation of both C-reactive protein and faecal calprotectin), who had failed corticosteroid or immunosuppressant therapy were enrolled. Patients were equally randomised to receive placebo, vercirnon 500 mg once daily or vercirnon 500 mg twice daily. The primary endpoint was clinical response, defined as a 100-point decrease in CDAI from baseline to week 12. RESULTS: Six hundred and eight patients were randomised. Patient characteristics and baseline demographics were similar among the groups. The proportions of patients achieving a clinical response were 25.1%, 27.6% and 27.2% for placebo, once daily and twice daily respectively; treatment differences were not significant (2.5%; 95% confidence interval, CI -6.1% to 11.0%, P = 0.546 for once daily vs. placebo, and 2.1%; 95% CI -6.5% to 10.7%, P = 0.648 for twice daily vs. placebo). Adverse events were reported in 69.8%, 73.3% and 78.1% with serious adverse events in 8.9%, 5.9%, and 6.0% of patients in the placebo, once-daily and twice-daily groups, respectively. CONCLUSIONS: We did not demonstrate efficacy of vercirnon as an induction therapy in patients with moderately-to-severely active Crohn's disease; its effect in maintenance therapy was not addressed.

Isoflurane-induced post-conditioning in senescent hearts is attenuated by failure to activate reperfusion injury salvage kinase pathway.

BACKGROUND: We investigated the cardioprotective effects of isoflurane administered at the onset of reperfusion in senescent rat in vivo, and the activation of the reperfusion injury salvage kinase (RISK) pathway to address a possible mechanism underlying age-related differences. METHODS: Male Wistar rats were assigned to age groups (young, 3-5 months; old, 20-24 months), and randomly selected to receive isoflurane (1 minimum alveolar concentration) or not for 3 min before and 2 min after reperfusion (ISO postC). Rats were subjected to coronary occlusion for 30 min followed by 2 h of reperfusion. Western blot analysis was used to assess the phosphorylation of extracellular signal-regulated kinase (ERK1/2), Akt, and GSK3ß 15 min after reperfusion. RESULTS: Brief administration of isoflurane 3 min before and 2 min after the initiation of early reperfusion reduced infarct size (56 ± 8% of left ventricular area at risk, mean ± standard deviation) compared with controls (68 ± 4%) in young rats, but had no effect in old rats (56 ± 8% in ISO postC and 56 ± 10% in control, respectively). Phosphorylation of ERK1/2, Akt, and GSK3ß were increased in the young ISO postC group but not in the old ISO postC group compared with control groups of the respective ages. CONCLUSIONS: We demonstrated that isoflurane post-conditions the heart in young but not in senescent rats. Failure to activate RISK pathway may contribute to attenuation of isoflurane-induced post-conditioning effect in senescent rats.

Titrated propofol induction vs. continuous infusion in children undergoing magnetic resonance imaging.

BACKGROUND: Propofol is the popular intravenous (i.v.) anaesthetic for paediatric sedation because of its rapid onset and recovery. We compared the efficacy and safety of a single dose and conventional infusion of propofol for sedation in children who underwent magnetic resonance imaging (MRI). METHODS: This was a double-blind, randomized-controlled study. One hundred and sixty children were assigned to group I (single dose) or II (infusion). Sedation was induced with i.v. propofol 2 mg/kg, and supplemental doses of propofol 0.5 mg/kg were administered until adequate sedation was achieved. After the induction of sedation, we treated patients with a continuous infusion of normal saline at a rate of 0.3 ml/kg/h in group I and the same volume of propofol in group II. In case of inadequate sedation, additional propofol 0.5 mg/kg was administered and the infusion rate was increased by 0.05 ml/kg/h. Induction time, sedation time, recovery time, additional sedation and adverse events were recorded. RESULTS: Recovery time was significantly shorter in group I compared with group II [0 (0-3) vs. 1 (0-3), respectively, P<0.001]. Group I (single dose) had significantly more patients with recovery time 0 compared with group II (infusion) (65/80 vs. 36/80, respectively, P<0.001). Induction and sedation times were not significantly different between groups. There was no significant difference in the frequency of additional sedation and adverse events between groups. CONCLUSION: A single dose of propofol without a continuous infusion can provide appropriate sedation in children undergoing MRI for <30 min.

Effects of fentanyl pretreatment on the QTc interval during propofol induction.

Prolongation of the corrected QT (QTc) interval is associated with various anaesthetic drugs. The QTc prolongation may become more exacerbated during laryngoscopy and intubation, which is possibly caused by sympathetic stimulation. The aim of this study was to investigate the effects of fentanyl on the QTc interval during propofol induction in healthy patients. The patients were randomly allocated to receive either fentanyl (n = 25) or saline (n = 25) before induction. The QTc interval was significantly prolonged immediately after intubation in control group compared to preceding values, but it did not change in the fentanyl group. The number of patients with the prolonged QTc interval exceeding 20 ms immediately after intubation compared to the baseline values was 14 in the control group and seven in the fentanyl group. In conclusion, pretreatment with fentanyl 2 microg x kg(-1) significantly attenuated QTc prolongation associated with laryngoscopy and tracheal intubation during propofol induction.

The safety and efficacy of adding etanercept to methotrexate or methotrexate to etanercept in moderately active rheumatoid arthritis patients previously treated with monotherapy.

OBJECTIVE: To determine if adding etanercept (ETN) to methotrexate (MTX) or MTX to ETN for 52 weeks in rheumatoid arthritis (RA) patients with moderate disease activity provides higher efficacy. METHODS: All patients (n = 227) received open-label ETN 25 mg subcutaneously twice-weekly and MTX orally up to 20 mg weekly for 52 weeks and had completed a 3-year study in which patients received MTX, ETN or combination therapy. Endpoints were based on Disease Activity Score (DAS) and European League Against Rheumatism (EULAR) responses. RESULTS: Patients previously receiving combination therapy (Combination group; n = 96) had a lower disease activity at baseline. The mean DAS for those previously receiving MTX (ETN-added group; n = 55) and previously receiving ETN (MTX-added group; n = 76) were in the moderate disease activity range at baseline; Combination patients had a low disease activity. The greatest increase in DAS remission rates from baseline to week 52 was in the ETN-added group (23.6% to 41.8%, p<0.01), although Combination (37.6% to 50.0%, p<0.01) and MTX-added (26.7% to 36.8%, p = NS) also demonstrated improvements. DAS low disease activity and EULAR responses showed similar results. No new safety issues were identified. CONCLUSION: RA patients who were partial responders to long-term MTX or etanercept monotherapy obtained a higher efficacy with combination therapy. Responses achieved by patients with combination therapy after 3 years in the previous study were sustained or improved during the fourth year of treatment. This trial supports the higher therapeutic effect of combination treatment with etanercept and MTX in RA patients with moderate disease activity despite monotherapy with one of the two agents.

Efficacy and safety of rofecoxib 12.5 mg and celecoxib 200 mg in two similarly designed osteoarthritis studies.

OBJECTIVE: To compare the lower osteoarthritis (OA) dose of rofecoxib to the recommended dose of celecoxib in two identically designed studies. METHODS: Patients with knee OA were randomized (2:2:1 ratio: rofecoxib 12.5 mg once daily (qd), celecoxib 200 mg qd, or placebo, respectively). The primary endpoint was patient global assessment of response to therapy (PGART) averaged over 6 weeks on a five-point scale. Rofecoxib would be declared at least as effective as celecoxib if the lower bound of the 95% confidence interval (95% CI) for difference in means was no lower than -0.5. Additional endpoints included Pain and Physical Function subscales of the Western Ontario and McMaster (WOMAC) OA Index. Adverse experiences (AEs) were recorded and combined from the two studies for analysis. RESULTS: Study 1 enrolled 395 patients (rofecoxib, n = 160; celecoxib, n = 157; placebo, n = 78). Study 2 enrolled 413 patients (rofecoxib, n = 159; celecoxib, n = 169; placebo, n = 85). Rofecoxib 12.5 mg was at least as effective as celecoxib 200 mg by PGART (Study 1 difference -0.09 [95% CI: -0.32, 0.14] and Study 2 difference 0.02 [95% CI: -0.20, 0.24]), and both were significantly (p < 0.001) more effective than placebo. Comparable efficacy was also seen for WOMAC Pain and Physical Function subscales with the active treatments. There was a significantly higher (p < 0.05) incidence of serious AEs with celecoxib than rofecoxib or placebo, none of which was drug-related. There were no significant differences in the pre-specified measurements of safety including drug-related AEs or discontinuations due to AEs, and the medications demonstrated similar safety as assessed by spontaneous reporting. CONCLUSIONS: Rofecoxib 12.5 mg and celecoxib 200 mg provided comparable efficacy over 6 weeks, and both were significantly more efficacious than placebo. The medications demonstrated similar safety compared to one another and placebo. The primary limitations of these studies were that they were only 6 weeks long and were powered for efficacy. Therefore, conclusions about long-term safety cannot be inferred.

Combined Fenton-MF process increases acrylonitrile removal.

The Fenton oxidation process is possessed of the advantages of both oxidation and coagulation processes. In addition to these functions, Fenton's reagent is also a typical initiator of polymerization. The application of the Fenton-microfiltration process for removal of acrylonitrile (AN), which is the major raw material for manufacturing ABS reins, was investigated. As for Fenton oxidation, in the range of pH 2 to pH 4, AN removal efficiency increased as the pH increased. In experiment of the same initial molar ratio of [FeSO4]0/[H2O2]0, the higher dosage can obtain the higher removal efficiency. At pH 4, the AN removal increased as the [H2O2]0 increased for each [FeSO4]0. Acrylic acid and acrylamide were detected in the solution after Fenton oxidation. On the other hand, acrylamide, polyacrylamide, and polyacrylic acid exist in the precipitate after the Fenton oxidation of AN solution. Moreover, it was also found that the operational mode is an important factor of the combined Fenton-MF process.

Rofecoxib versus codeine/acetaminophen in postoperative dental pain: a double-blind, randomized, placebo- and active comparator-controlled clinical trial.

BACKGROUND: In recent studies of acute pain and primary dysmenorrhea, rofecoxib, a nonsteroidal anti-inflammatory drug that selectively targets the cyclooxygenase-2 enzyme, was found to be similar in efficacy to ibuprofen and naproxen sodium. OBJECTIVE: The purpose of this study was to determine the analgesic efficacy of a single oral dose of rofecoxib 50 mg compared with the combination of codeine 60 mg/acetaminophen 600 mg in a model of postsurgical dental pain. METHODS: In this double-blind, placebo- and active comparator-controlled, parallel-group study, patients experiencing moderate or severe pain after the surgical extraction of > or = 2 third molars, at least 1 of which was a mandibular impaction, were randomized to receive placebo, rofecoxib 50 mg, or codeine 60 mg/acetaminophen 600 mg. Patient evaluations of pain intensity, pain relief, and global assessments were recorded throughout the 24-hour period after dosing. The 2-stopwatch method was used to determine time to confirmed perceptible pain relief. The primary end point assessing overall analgesic effect was total pain relief over 6 hours (TOPAR6). Secondary end points were patient global assessment of response to therapy (PGART) at 6 hours, onset of analgesia, peak analgesic effect, and duration of analgesia. RESULTS: A total of 393 patients were enrolled; 182 received rofecoxib, 180 received codeine/acetaminophen, and 31 received placebo. The overall analgesic effect of rofecoxib 50 mg was greater than that of codeine 60 mg/acetaminophen 600 mg for TOPAR6 (12.4 vs 7.0; P < 0.001) and PGART at 6 hours (P < 0.001). The onset of analgesic effect was similar for rofecoxib and codeine/acetaminophen. Peak analgesic effect as measured by peak pain relief scores during the first 6 hours was significantly greater in the rofecoxib group compared with the codeine/acetaminophen group (P < 0.001), as was the duration of analgesic effect measured by the time to rescue analgesia (9.6 hours vs 2.3 hours, P < 0.001). Adverse events were reported in 33.0%, 46.1%, and 32.3% of patients treated with rofecoxib, codeine/acetaminophen, and placebo, respectively. The most common adverse events were nausea (6.0%, 25.0%, and 9.7%, respectively) and vomiting (3.8%, 18.3%, and 6.5%, respectively). Significantly more patients in the codeine/acetaminophen group than in the rofecoxib group experienced adverse events overall (P < 0.050) and nausea in particular (P < 0.001). CONCLUSION: In this study of moderate to severe postoperative dental pain, the analgesic efficacy of rofecoxib 50 mg was greater than that of codeine/acetaminophen, with a lower incidence of adverse events and nausea.

Overexpression of and RNA interference with the CCAAT enhancer-binding protein on long-term facilitation of Aplysia sensory to motor synapses.

In the marine mollusk Aplysia, the CCAAT/enhancer-binding protein, ApC/EBP, serves as an immediate early gene in the consolidation of long-term facilitation in the synaptic connection between the sensory and motor neurons of the gill-withdrawal reflex. To further examine the role of ApC/EBP as a molecular switch of a stable form of long-term memory, we cloned the full-length coding regions of two alternatively spliced forms, the short and long form of ApC/EBP. Overexpression of each isoform by DNA microinjection resulted in a l6-fold increase in the expression of the coinjected luciferase reporter gene driven by an ERE promoter. In addition, when we overexpressed ApC/EBP in Aplysia sensory neurons, we found that the application of a single pulse of 5-HT that normally induced only short-term facilitation now induced long-term facilitation. Conversely, when we attempted to block the synthesis of native ApC/EBP by microinjecting double-strand RNA or antisense RNA, we blocked long-term facilitation in a sequence-specific manner. These data support the idea that ApC/EBP is both necessary and sufficient to consolidate short-term memory into long-term memory. Furthermore, our results suggest that this double-strand RNA interference provides a powerful tool in the study of the genes functioning in learning and memory in Aplysia by specifically inhibiting both the constitutive and induced expression of the genes.

Early passive mobilization after digital nerve repair and grafting in a fresh cadaver.

In the clinical management of combined tendon and nerve injuries, there are competing treatment strategies. Isolated tendon injuries should be rapidly mobilized after repair to prevent adhesion formation, whereas isolated nerve repairs are usually immobilized to prevent disruption and to allow axon regrowth. Recommendations in the published literature for the management of combined tendon and nerve injuries are vague and advise up to 3 weeks of immobilization. The goals of this study were to determine which length of nerve gap resulted in rupture of a repair following postoperative mobilization with the modified Duran protocol and with unrestricted motion and to determine whether nerve grafts are at risk of rupture after mobilization. A total of 100 digital nerves from 10 cadaver hands were tested with the modified Duran and the unsplinted protocols. Each digital nerve on each hand was sequentially resected and repaired at five progressively larger gap lengths after testing with both protocols. The mean nerve gaps at which disruption occurred were significantly different between the splinted (9.7 +/- 0.8 mm, n = 100) and unsplinted (7.3 +/- 1.9 mm, n = 100) protocols (t test, p < 0.001). One hundred percent of repairs remained intact, with up to 5 mm of resection with the modified Duran protocol (n = 100) and with up to 2.5 mm of resection with the unsplinted protocol (n = 100). All nerve grafts remained intact after mobilization within a dorsal-blocking splint (n = 100). Considering mechanical integrity of the nerve repair only, these data suggest that early mobilization with tendon protocols may be considered after a nerve injury to avoid the detrimental tendon sequelae that result from immobilization. The adequacy of functional recovery of mobilized nerves is yet to be determined.

Multivariate correlations of geochemical binding phases of heavy metals in contaminated river sediment.

Distributions of geochemical binding phases of seven heavy metals (Cr, Cu, Co, Zn, Ni, Pb, and Cd) in sediment cores taken from six heavily polluted sites of the Ell-Ren River in Southern Taiwan were studied. Sequential extraction procedures (SEP) were used to determine the variations of heavy metal binding phases (exchangeable, bound to carbonates, bound to manganese-oxides, bound to iron-oxides, and bound to organic matter) in different sediment depths. Multivariate analyses were used to explore the correlations among these geochemical binding phases of heavy metals. Results showed that the total amounts of various binding phases of heavy metals significantly varied with sediment depth, but their binding behaviors in various phases did not significantly change with depth. The organic matter content in the sediments increased with increasing Fe-oxide content. In addition, the binding affinities of carbonates with Zn, Pb, and Ni were higher than the affinities of carbonates with the other heavy metals. The binding affinity of Fe-oxides with Cr was higher than the affinities of Fe-oxides with the other heavy metals. Both correlation matrixes and principal component analyses demonstrated that distributions of Cu, Zn, Ni, and Cd had significant correlations with each other in both different depth horizons and various geochemical binding phases. The results indicate that these heavy metals might be discharged from the same pollution sources in the past, and also showed stable geochemical binding behaviors with the high silt sediment. However, Co had a poor correlation with the other six heavy metals in various binding phases, except with organic matter. Binding behaviors of Pb in the phases of bound to carbonates and exchangeable were different from the other six heavy metals. Cu was inversly correlated with the other six heavy metals in its binding behavior with reducible phases (Fe-/Mn-oxides).

Wet air oxidation of a direct dye solution catalyzed by CoAlPO4 -5. Performance assessment and kinetic study.

Wet air oxidation (WAO) of a prepared direct dye solution was tested by using the CoAIPO4 -5 catalyst. Addition of CoAIPO4 -5 could effectively improve rate of color removal and the activation energy of color removal could decrease from about 110 kJ/mole to about 75 kJ/mole as the catalyst loading was increased from 0.0 g/L to 3.0 g/L. Performance of WAO on color removal would somewhat increase with catalyst loading at 145 degrees C whereas the effect of catalyst loading was not significant at 135 degrees C. With no addition of CoAlPO4 -5, the chemical oxygen demand (COD) value was low. This was due to difficulty of exactly measuring the true COD value of dye solution if the dye was not degraded. Via CoAIPO4 -5, COD of dye solution could be effectively decreased. The rate of COD removal would increase with catalyst loading, oxygen pressure and reaction temperature. Furthermore, a maximum COD value observed, which was due to catalyzed degradation of dye molecule via CoAlPO4 -5, could be characterized by a consecutive reaction scheme. Kinetic study of color removal is expressed as follows: rate = k x [dye](0.8) x W(0.5) x P(n) (145 degrees C) or rate = k x [dye](0.8) x W(0) x P(n) (135 degrees C); where k means rate constant, [] means concentration, W means catalyst loading, P means oxygen pressure and n means uncertain number.

Activation of a heterologously expressed octopamine receptor coupled only to adenylyl cyclase produces all the features of presynaptic facilitation in aplysia sensory neurons.

Short-term behavioral sensitization of the gill-withdrawal reflex after tail stimuli in Aplysia leads to an enhancement of the connections between sensory and motor neurons of this reflex. Both behavioral sensitization and enhancement of the connection between sensory and motor neurons are importantly mediated by serotonin. Serotonin activates two types of receptors in the sensory neurons, one of which is coupled to the cAMP/protein kinase A (PKA) pathway and the other to the inositol triphosphate/protein kinase C (PKC) pathway. Here we describe a genetic approach to assessing the isolated contribution of the PKA pathway to short-term facilitation. We have cloned from Aplysia an octopamine receptor gene, Ap oa(1), that couples selectively to the cAMP/PKA pathway. We have ectopically expressed this receptor in Aplysia sensory neurons of the pleural ganglia, where it is not normally expressed. Activation of this receptor by octopamine stimulates all four presynaptic events involved in short-term synaptic facilitation that are normally produced by serotonin: (i) membrane depolarization; (ii) increased membrane excitability; (iii) increased spike duration; and (iv) presynaptic facilitation. These results indicate that the cAMP/PKA pathway alone is sufficient to produce all the features of presynaptic facilitation.

The cytoplasmic domain of human FcgammaRIa alters the functional properties of the FcgammaRI.gamma-chain receptor complex.

The gamma/zeta-chain family of proteins mediate cell activation for multiple immunoglobulin receptors. However, the recognition that these receptors may have distinct biologic functions suggests that additional signaling elements may contribute to functional diversity. We hypothesized that the cytoplasmic domain (CY) of the ligand binding alpha-chain alters the biological properties of the receptor complex. Using macrophage FcgammaRIa as a model system, we created stable transfectants expressing a full-length or a CY deletion mutant of human FcgammaRIa. Both receptors functionally associate with the endogenous murine gamma-chain. However, we have established that the CY of FcgammaRIa directly contributes to the functional properties of the receptor complex. Deletion of the FcgammaRIa CY leads to slower kinetics of receptor-specific phagocytosis and endocytosis as well as lower total phagocytosis despite identical levels of receptor expression. Deletion of the CY also converts the phenotype of calcium independent FcgammaRIa-specific phagocytosis to a calcium-dependent phenotype. Finally, deletion of the CY abrogates FcgammaRIa-specific secretion of interleukin-6 but does not affect production of interleukin-1beta. These results demonstrate a functional role for the CY of FcgammaRIa and provide a general model for understanding how multiple receptors that utilize the gamma-chain can generate diversity in function.

Human cloned alpha1A-adrenoceptor isoforms display alpha1L-adrenoceptor pharmacology in functional studies.

The recombinant alpha1A-adrenoceptor displays a distinct pharmacological profile ('classical alpha1A-adrenoceptor') in homogenate binding assays, but displays the properties of the so-called alpha1L-adrenoceptor in functional studies in whole cells at 37 degrees C. As three splice variants of the human alpha1A-adrenoceptor have been described previously (alpha1A-1, alpha1A-2 and alpha1A-3), we have compared their functional pharmacological profiles, when expressed stably in Chinese hamster ovary (CHO-K1) cells (antagonist inhibition of noradrenaline-stimulated [3H]inositol phosphates accumulation). A fourth, novel isoform (alpha1A-4) has also been studied: alpha1A-4 mRNA predominates in several human tissues including prostate, liver, heart and bladder. In homogenate binding studies, all four isoforms displayed essentially identical affinity profiles, with prazosin (1-(4-amino-6,7-dimethoxy-2-quinazolinyl)-4-(2-furoyl)piperazine), tamsulosin (5-[2-[[2-(2-ethoxyphenoxy)ethyl]-amino]propyl]-2-methoxybenzen esulfonamide), RS-17053 (N-[2-(2-cyclopropylmethoxyphenoxy)ethyl]-5-chloro-alpha,alphad imethyl-1H-indole-3-ethanamine hydrochloride), WB 4101 ((2,6-dimethoxyphenoxyethyl)aminomethyl-1,4-benzodioxane hydrochloride) and 5-Me-urapidil (5-methyl-6[[3-[4-(2-methoxyphenyl)-1-piperazinyl]propyl]amino]-1,3-d imethyuracil) all displaying subnanomolar affinities. In functional studies, noradrenaline accelerated [3H]inositol phosphates production with potencies (p[A]50) of between 5.8 and 6.6. The affinities of prazosin, RS-17053, WB 4101 and 5-Me-urapidil, at antagonizing responses to noradrenaline, were reduced by approximately 10-fold (cf. binding data), while those for tamsulosin and indoramin (N-[1-[2-(1H-indol-3-yl)ethyl]-4-piperidinyl]benzamide) remained constant or increased, consistent with the previously described alpha1L-adrenoceptor. Thus, all four human recombinant alpha1A-adrenoceptor isoforms display the pharmacology of the alpha1L-adrenoceptor when studied in functional assays, consistent with the hypothesis that the putative alpha1L-adrenoceptor represents a functional phenotype of the alpha1A-adrenoceptor.

Cloning and expression of a G protein-linked acetylcholine receptor from Caenorhabditis elegans.

We have isolated a cDNA clone from the nematode Caenorhabditis elegans that encodes a protein of greatest sequence similarity to muscarinic acetylcholine receptors. This gene codes for a polypeptide of 682 amino acids containing seven putative transmembrane domains. The amino acid identities, excluding a highly variable middle portion of the third intracellular loop, to the human m1-m5 receptors are 28-34%. When this cloned receptor was coexpressed with a G protein-gated inwardly rectifying K+ channel (GIRK1) in Xenopus oocyte, acetylcholine was able to elicit the GIRK current. This acetylcholine-induced current was substantially inhibited by the muscarinic antagonist atropine in a reversible manner. However, another muscarinic agonist oxotremorine and antagonists scopolamine and pirenzepine had little or negligible effects on this receptor. Taken together, these results suggest that the cloned gene encodes a G protein-linked acetylcholine receptor that is most similar to but pharmacologically distinct from muscarinic acetylcholine receptors.

Cloning and functional characterization of a Caenorhabditis elegans muscarinic acetylcholine receptor.

A cDNA clone encoding a muscarinic acetylcholine receptor (mAChR) has been isolated from the nematode Caenorhabditis elegans. The nematode mAChR, consisted of 585 amino acids, displays a high degree of amino acid sequence homology to other invertebrate and vertebrate mAChRs. Excluding a highly variable middle portion of the third intracellular loop, the C. elegans mAChR shares about 51% amino acid sequence identity with a Drosophila mAChR and 42-44% identity with human m1-m5 mAChR subtypes. Comparison of the cDNA sequence with the corresponding genomic sequence reveals that the C. elegans mAChR gene contains ten introns, eight of them in the coding region. Pharmacological profiles of the C. elegans mAChR expressed in Chinese hamster ovary (CHO) cells were shown to be similar to those of mammalian counterparts, indicating that ligand binding domains of the receptor have been conserved during evolution. When this cloned receptor was expressed in Xenopus oocytes, acetylcholine evoked a transient Cl- current. Furthermore, activation of the receptor with oxotremorine, acetylcholine or carbachol resulted in the stimulation of phosphatidylinositol metabolism in CHO cells, suggesting that the receptor is coupled to phospholipase C activation.