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Background: Current percutaneous transthoracic needle biopsies (PTNB) navigation systems present challenges due to additional steps and limitations on the operating environment., Research Question: We developed a novel, registration-free navigation system for swift and precise CT-guided PTNB, eliminating the need for body surface markers and intraoperative registration. This study assesses its efficacy and safety., Methods: A prospective study was conducted on participants aged 18-80 years prepared for PTNB at two clinical centers, from December 2021 to August 2022. The primary endpoint was the success rate of biopsies within 2 needle adjustments, and the secondary endpoint was the success rate within a single adjustment. Safety endpoints were defined by adverse events occurrence., Results: The study included 98 patients (median age, 64 years, IQR 54-69 years, 71 men). The primary endpoint achieved a biopsy success rate of 98.98 %, and the secondary endpoint demonstrated 97.96 %. The overall success rate was 98.98 %, significantly exceeding the target value of 85 % (P < 0.0001). The median number of CT scans was 3, significantly fewer than predicted for the manual puncture scheme [3 (IQR 3-3) to 8 (IQR 6-8), P < 0.0001]. The average procedure duration was 18.0 min (IQR: 14.0-29.0 min). The most common adverse events were hemorrhage (14 instances) and pneumothorax (8 instances). Other adverse events included elevated blood pressure, hemoptysis, and other common events., Interpretation: Our registration-free navigation system proved to be an effective and safe system for assisting percutaneous lung biopsies in clinical practice., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Chunfu Fang, Shu-ting Shen, Yuchuan Yang, Zhe-ni Shi, Bing Niu, and Jian Liu are employees of Shanghai Simple Touch Technology Co., Ltd. All other authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: Patients with rare, pathogenic cardiomyopathy (CM) and arrhythmia variants can present with atrial fibrillation (AF). The efficacy of AF ablation in these patients is unknown., Objective: This study tested the hypotheses that: 1) patients with a pathogenic variant in any CM or arrhythmia gene have increased recurrence following AF ablation; and 2) patients with a pathogenic variant associated with a specific gene group (arrhythmogenic left ventricular CM [ALVC], arrhythmogenic right ventricular CM, dilated CM, hypertrophic CM, or a channelopathy) have increased recurrence., Methods: We performed a prospective, observational, cohort study of patients who underwent AF catheter ablation and whole exome sequencing. The primary outcome measure was ≥30 seconds of any atrial tachyarrhythmia that occurred after a 90-day blanking period., Results: Among 1,366 participants, 109 (8.0%) had a pathogenic or likely pathogenic (P/LP) variant in a CM or arrhythmia gene. In multivariable analysis, the presence of a P/LP variant in any gene was not significantly associated with recurrence (HR 1.15; 95% CI 0.84-1.60; P = 0.53). P/LP variants in the ALVC gene group, predominantly LMNA, were associated with increased recurrence (n = 10; HR 3.75; 95% CI 1.84-7.63; P < 0.001), compared with those in the arrhythmogenic right ventricular CM, dilated CM, hypertrophic CM, and channelopathy gene groups. Participants with P/LP TTN variants (n = 46) had no difference in recurrence compared with genotype-negative-controls (HR 0.93; 95% CI 0.54-1.59; P = 0.78)., Conclusions: Our results support the use of AF ablation for most patients with rare pathogenic CM or arrhythmia variants, including TTN. However, patients with ALVC variants, such as LMNA, may be at a significantly higher risk for arrhythmia recurrence., Competing Interests: Funding Support and Author Disclosures This work was supported by grants from the American Heart Association (AHA20SCG35540034 [Dr Shoemaker]) and the National Institutes of Health (R01HL155197 [Dr Shoemaker]). It was also supported by CTSA award (UL1TR000445) from the National Center for Advancing Translational Sciences. Its contents are solely the responsibility of the authors and do not necessarily represent official views of the AHA or NIH. Dr Shoemaker serves on the advisory board and has received sponsored research funds from Roche Diagnostics. Dr Kanagasundram has received speaking fees from Biosense Webster and Janssen. Dr Crossley has received consulting fees or honoraria from Bayer Healthcare, Boston Scientific, Janssen Pharmaceuticals, Medtronic, and Spectranetics. Dr Richardson has received research funding from Medtronic Inc, Abbott Inc and served as a consultant for Philips Inc and Biosense Webster. Dr Montgomery has received research funding from Medtronic Inc. Dr Ellis has received research funding from Boston Scientific, Medtronic, and Boehringer Ingelheim; and consulting fees from Medtronic, Boston Scientific, Abbott Medical, and Atricure. Dr Michaud has received consulting fees or honoraria from Boston Scientific, Medtronic, Biotronik, Abbott, and Biosense Webster. Vanderbilt receives fellowship support from Medtronic inc., Boston Scientific inc., Abbott Labs, Biotronik, and Johnson & Johnson. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: TTN encodes a sarcomeric protein called titin. Pathogenic rare variants in TTN are the most common finding in patients with atrial fibrillation (AF) and positive genetic testing., Objectives: This study sought to define the characteristics and outcomes in patients with AF and pathogenic TTN variants compared with genotype-negative patients with AF., Methods: Patients who presented initially with AF were enrolled in an AF registry. Retrospectively they underwent research sequencing for cardiomyopathy and arrhythmia genes. TTN(+) AF cases were defined as participants with pathogenic or likely pathogenic (P/LP) rare variants located in exons with high cardiac expression. They were matched 1:2 with control subjects with no P/LP variants. Phenotyping used retrospective manual chart review., Results: Among 2794 participants; 57 (2.0%) TTN(+) AF cases were identified and matched with 114 control subjects. Low QRS complex voltage was present more often in TTN(+) AF cases (18% vs 5%; P < 0.01), with no difference in PR, QRS interval, or QTc. More TTN(+) AF cases had persistent AF at enrollment (44% vs 30%; P = 0.028) and had undergone multiple cardioversions (61% vs. 37%; P < 0.01). By end of follow-up (median 8.3 years; Q1, Q3: 4.5, 13.7 years), 11% of TTN(+) AF cases developed sustained ventricular tachycardia/ventricular fibrillation, 44% left ventricular (LV) systolic dysfunction (LV ejection fraction <50%), and 47% met a combined endpoint of sustained ventricular tachycardia/ventricular fibrillation or LV systolic dysfunction., Conclusions: TTN(+) AF patients undergo more cardioversions and have more persistent forms of AF. Approximately 50% develop LV systolic dysfunction and/or malignant ventricular arrhythmias. These results highlight the need for diagnostic evaluation and management in TTN(+) patients beyond the usual care for AF., Competing Interests: Funding Support and Author Disclosures This work was supported by grants from the American Heart Association (AHA20SCG35540034 [to Dr Shoemaker] and AHA18SFRN34110369 [to Dr Roden]) and the National Institutes of Health (R01HL155197 [to Dr Shoemaker]); and was also supported by CTSA award (UL1TR000445) from the National Center for Advancing Translational Sciences. Its contents are solely the responsibility of the authors and do not necessarily represent official views of the AHA or National Institutes of Health. Dr Kanagasundram has received speaker fees from Biosense Webster and Janssen. Dr Montgomery has received research funding from Medtronic. Dr Ellis has received research funding from Boston Scientific, Medtronic, and Boehringer Ingelheim; and has received consulting fees from Medtronic, Boston Scientific, Abbott Medical, and Atricure. Dr Crossley has received consulting fees or honoraria from Bayer Healthcare, Boston Scientific, Janssen Pharmaceuticals, Medtronic, and Spectranetics. Dr Richardson has received research funding from Medtronic and Abbott; and has served as a consultant for Philips and Biosense Webster. Dr Tandri has received a research grant from Abbott. Dr Stevenson has received honoraria from Abbott, Boston Scientific, Medtronic, Johnson and Johnson, and Biotronik; and has received research funding from Adagio Medical. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Recurrence after atrial fibrillation (AF) ablation remains common. We evaluated the association between recurrence and levels of biomarkers of cardiac remodeling, and their ability to improve recurrence prediction when added to a clinical prediction model., Methods and Results: Blood samples collected before de novo catheter ablation were analyzed. Levels of bone morphogenetic protein-10, angiopoietin-2, fibroblast growth factor-23, insulin-like growth factor-binding protein-7, myosin-binding protein C3, growth differentiation factor-15, interleukin-6, N-terminal pro-brain natriuretic peptide, and high-sensitivity troponin T were measured. Recurrence was defined as ≥30 seconds of an atrial arrhythmia 3 to 12 months postablation. Multivariable logistic regression was performed using biomarker levels along with clinical covariates: APPLE score (Age >65 years, Persistent AF, imPaired eGFR [<60 ml/min/1.73m 2 ], LA diameter ≥43 mm, EF <50%; which includes age, left atrial diameter, left ventricular ejection fraction, persistent atrial fibrillation, and estimated glomerular filtration rate), preablation rhythm, sex, height, body mass index, presence of an implanted continuous monitor, year of ablation, and additional linear ablation. A total of 1873 participants were included. A multivariable logistic regression showed an association between recurrence and levels of angiopoietin-2 (odds ratio, 1.08 [95% CI, 1.02-1.15], P =0.007) and interleukin-6 (odds ratio, 1.02 [95% CI, 1.003-1.03]; P =0.02). The area under the receiver operating characteristic curve of a model that only contained clinical predictors was 0.711. The addition of any of the 9 studied biomarkers to the predictive model did not result in a statistically significant improvement in the area under the receiver operating characteristic curve., Conclusions: Higher angiopoietin-2 and interleukin-6 levels were associated with recurrence after atrial fibrillation ablation in multivariable modeling. However, the addition of biomarkers to a clinical prediction model did not significantly improve recurrence prediction.

Introduction: Chronic kidney disease-mineral and bone disorder (CKD-MBD) is characterized by bone abnormalities, vascular calcification, and some other complications. Although there are diagnostic criteria for CKD-MBD, in situations when conducting target feature examining are unavailable, there is a need to investigate and discover alternative biochemical criteria that are easy to obtain. Moreover, studying the correlations between the newly discovered biomarkers and the existing ones may provide insights into the underlying molecular mechanisms of CKD-MBD., Methods: We collected a cohort of 116 individuals, consisting of three subtypes of CKD-MBD: calcium abnormality, phosphorus abnormality, and PTH abnormality. To identify the best biomarker panel for discrimination, we conducted six machine learning prediction methods and employed a sequential forward feature selection approach for each subtype. Additionally, we collected a separate prospective cohort of 114 samples to validate the discriminative power of the trained prediction models., Results: Using machine learning under cross validation setting, the feature selection method selected a concise biomarker panel for each CKD-MBD subtype as well as for the general one. Using the consensus of these features, best area under ROC curve reached up to 0.95 for the training dataset and 0.74 for the perspective dataset, respectively., Discussion/conclusion: For the first time, we utilized machine learning methods to analyze biochemical criteria associated with CKD-MBD. Our aim was to identify alternative biomarkers that could serve not only as early detection indicators for CKD-MBD, but also as potential candidates for studying the underlying molecular mechanisms of the condition., (© 2024. The Author(s).)

The COVID-19 pandemic has become a global catastrophe, affecting the health and economy of the human community. It is required to mitigate the impact of pandemics by developing rapid molecular diagnostics for SARS-CoV-2 virus detection. In this context, developing a rapid point-of-care (POC) diagnostic test is a holistic approach to the prevention of COVID-19. In this context, this study aims at presenting a real-time, biosensor chip for improved molecular diagnostics including recombinant SARS-CoV-2 spike glycoprotein and SARS-CoV-2 pseudovirus detection based on one-step-one-pot hydrothermally derived CoFeBDCNH 2 -CoFe 2 O 4 MOF-nanohybrids. This study was tested on a PalmSens-EmStat Go POC device, showing a limit of detection (LOD) for recombinant SARS-CoV-2 spike glycoprotein of 6.68 fg/mL and 6.20 fg/mL in buffer and 10% serum-containing media, respectively. To validate virus detection in the POC platform, an electrochemical instrument (CHI6116E) was used to perform dose dependent studies under similar experimental conditions to the handheld device. The results obtained from these studies were comparable indicating the capability and high detection electrochemical performance of MOF nanocomposite derived from one-step-one-pot hydrothermal synthesis for SARS-CoV-2 detection for the first time. Further, the performance of the sensor was tested in the presence of Omicron BA.2 and wild-type D614G pseudoviruses., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 Elsevier B.V. All rights reserved.)

Background: Experimental evidence suggests genetic variation in 4q25/PITX2 modulates pulmonary vein (PV) myocardial sleeve length. Although PV sleeves are the main target of atrial fibrillation (AF) ablation, little is known about the association between different PV sleeve characteristics with ablation outcomes., Objectives: This study sought to evaluate the association between clinical and genetic (4q25) risk factors with PV sleeve length in humans, and to evaluate the association between PV sleeve length and recurrence after AF ablation., Methods: In a prospective, observational study of patients undergoing de novo AF ablation, PV sleeve length was measured using electroanatomic voltage mapping before ablation. The sentinel 4q25 AF susceptibility single nucleotide polymorphism, rs2200733, was genotyped. The primary analysis tested the association between clinical and genetic (4q25) risk factors with PV sleeve length using a multivariable linear regression model. Covariates included age, sex, body mass index, height, and persistent AF. The association between PV sleeve length and atrial arrhythmia recurrence (>30 seconds) was tested using a multivariable Cox proportional hazards model., Results: Between 2014 and 2019, 197 participants were enrolled (median age 63 years [IQR: 55 to 70 years], 133 male [67.5%]). In multivariable modeling, men were found to have PV sleeves 2.94 mm longer than women (95% CI: 0.99-4.90 mm; P < 0.001). Sixty participants (30.5%) had one 4q25 risk allele and 6 (3.1%) had 2 alleles. There was no association between 4q25 genotype and PV sleeve length. Forty-six participants (23.4%) experienced arrhythmia recurrence within 3 to 12 months, but there was no association between recurrence and PV sleeve length., Conclusions: Common genetic variation at 4q25 was not associated with PV sleeve length and PV sleeve length was not associated with ablation outcomes. Men did have longer PV sleeves than women, but more research is needed to define the potential clinical significance of this observation., Competing Interests: Funding Support and Author Disclosures This research work was supported in part by the National Institutes of Health grant K23 HL127704 (Shoemaker). The project was also supported by CTSA award UL1TR000445 from the National Center for Advancing Translational Sciences. Its contents are solely the responsibility of the authors and do not necessarily represent official views of the National Center for Advancing Translational Sciences or the National Institutes of Health. Dr. Shoemaker serves on the advisory board; and has received sponsored research funds from Roche Diagnostics. Dr. Kanagasundram has received speakers fees from Biosense Webster and Janssen. Dr. Crossley has received consulting fees or honoraria from Bayer Healthcare, Boston Scientific, Janssen Pharmaceuticals, Medtronic, and Spectranetics. Dr. Richardson has received research funding from Medtronic Inc, Abbott Inc; and served as a consultant for Philips Inc and Biosense Webster. Dr. Ellis has received research funding from Boston Scientific, Medtronic, Boehringer Ingelheim; and consulting fees from Medtronic, Boston Scientific, Abbott Medical, and Atricure. Dr. Michaud has received consulting fees or honoraria from Boston Scientific, Medtronic, Biotronik, Abbott, and Biosense Webster. Vanderbilt receives fellowship support from Medtronic Inc, Boston Scientific Inc, Abbott Labs, Biotronik, and Johnson & Johnson. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Published by Elsevier Inc.)

Background: Experimental data suggest ryanodine receptor-mediated intracellular calcium leak is a mechanism for atrial fibrillation (AF), but evidence in humans is still needed. Propafenone is composed of two enantiomers that are equally potent sodium-channel blockers; however, (R )-propafenone is an ryanodine receptor inhibitor whereas (S )-propafenone is not. This study tested the hypothesis that ryanodine receptor inhibition with (R )-propafenone prevents induction of AF compared to (S )-propafenone or placebo in patients referred for AF ablation., Methods: Participants were randomized 4:4:1 to a one-time intravenous dose of (R )-propafenone, (S )-propafenone, or placebo. The study drug was given at the start of the procedure and an AF induction protocol using rapid atrial pacing was performed before ablation. The primary endpoint was 30 s of AF or atrial flutter., Results: A total of 193 participants were enrolled and 165 (85%) completed the study protocol (median age: 63 years, 58% male, 95% paroxysmal AF). Sustained AF and/or atrial flutter was induced in 60 participants (84.5%) receiving (R )-propafenone, 60 (80.0%) receiving (S )-propafenone group, and 12 (63.2%) receiving placebo. Atrial flutter occurred significantly more often in the (R )-propafenone (N=23, 32.4%) and (S )-propafenone (N=26, 34.7%) groups compared to placebo (N=1, 5.3%, P =0.029). There was no significant difference between (R )-propafenone and (S )-propafenone for the primary outcome of AF and/or atrial flutter induction in univariable ( P =0.522) or multivariable analysis ( P =0.199, adjusted for age and serum drug level)., Conclusions: There is no difference in AF inducibility between (R )-propafenone and (S )-propafenone at clinically relevant concentrations. These results are confounded by a high rate of inducible atrial flutter due to sodium-channel blockade., Registration: https://clinicaltrials.gov; Unique Identifier: NCT02710669.

Presentation of antigenic peptides to T-cell receptors is an essential step in the adaptive immune response. In the mouse the class Ib major histocompatibility complex molecule, H2-M3, presents bacterial- and mitochondrial-derived peptides to T-cell receptors on cytotoxic T cells. Four mitochondrial heptapeptides, differing only at residue 6, form complexes with H2-M3 which can be distinguished by T cells. No structures of relevant receptors are available. To investigate the structural basis for this distinction, crystal structures were determined and molecular dynamics simulations over one microsecond were done for each complex. In the crystal structures of the heptapeptide complexes with H2-M3, presented here, the side chains of the peptide residues at position 6 all point into the H2-M3 binding groove, and are thus inaccessible, so that the very similar structures do not suggest how recognition and initiation of responses by the T cells may occur. However, conformational differences, which could be crucial to T-cell discrimination, appear within one microsecond during molecular dynamics simulations of the four complexes. Specifically, the three C-terminal residues of peptide ligands with alanine or threonine at position 6 partially exit the binding groove; this does not occur in peptide ligands with isoleucine or valine at position 6. Structural changes associated with partial peptide exit from the binding groove, along with relevant peptide binding energetics and immunological results are discussed. Communicated by Ramaswamy H. Sarma.

Objectives: This study aimed to review the utility of quinidine in patients presenting with recurrent sustained ventricular arrhythmia (VA) and limited antiarrhythmic drug (AAD) options., Background: Therapeutic options are often limited in patients with structural heart disease and recurrent VAs. Quinidine has an established role in rare arrhythmic syndromes, but its potential use in other difficult VAs has not been assessed in the present era., Methods: We performed a retrospective analysis of 37 patients who had in-hospital quinidine initiation after multiple other therapies failed for VA suppression at our tertiary referral center. Clinical data and outcomes were obtained from the medical record., Results: Of 30 patients with in-hospital quantifiable VA episodes, quinidine reduced acute VA from a median of 3 episodes (interquartile range [IQR]: 2 to 7.5) to 0 (IQR: 0 to 0.5) during medians of 3 days before and 4 days after quinidine initiation (p < 0.001). VA events decreased from a median of 10.5 episodes per day (IQR: 5 to 15) to 0.5 episodes (IQR: 0 to 4) after quinidine initiation in the 12 patients presenting with electrical storm (p = 0.004). Among the 24 patients discharged on quinidine, 13 (54.2%) had VA recurrence during a median of 138 days. Adverse effects in 9 of the 37 patients (24.3%) led to drug discontinuation, most commonly gastrointestinal intolerance., Conclusions: In patients with recurrent VAs and structural heart disease who have limited treatment options, quinidine can be useful, particularly as a short-term therapy., Competing Interests: Funding Support and Author Disclosures The authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Copyright © 2021 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Objectives: The aims of this study were to quantify the pain and distress associated with the administration of intranasal (IN) midazolam in young children using 4 observational measures and to evaluate the degree of validity of these measures., Methods: We conducted a prospective observational pilot study. Children aged 1 to 7 years requiring IN midazolam were enrolled. Children were videotaped, and scores were assigned to baseline and administration phases using the Observational Scale of Behavioral Distress-Revised (OSBD-R), Children's Hospital of Eastern Ontario Pain Scale (CHEOPS), and the Faces-Legs-Activity-Cry-Consolability (FLACC) scale. The cry duration following administration was assessed. Interrater reliability and convergent validity were determined for all 4 measures. Internal consistency and responsivity for the OSBD-R, CHEOPS, and FLACC scales were determined., Results: We enrolled 20 children. The mean OSBD-R, CHEOPS, and FLACC scores associated with administration of IN midazolam were 27.1 (SD, 13.5), 11.5 (SD, 1.2), and 8.9 (SD, 2.7), respectively. The mean cry duration was 105.5 (SD, 68.8) seconds. The intraclass correlation coefficients for all measures ranged from 0.82 to 0.99. The Cronbach α's for the OSBD-R, CHEOPS, and FLACC were between 0.71 and 0.97. Pearson correlation coefficients for comparisons between OSBD-R, CHEOPS, and FLACC were between 0.82 and 0.96 but were between 0.32 and 0.51 for comparisons involving cry duration., Conclusions: We have identified estimates of pain and distress associated with administration of IN midazolam in young children that can be used to determine desired effect sizes for trials that study interventions to treat this pain and distress. The OSBD-R, CHEOPS, and FLACC scales are suitable choices for outcome measures., Competing Interests: Disclosure: The authors declare no conflict of interest., (Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.)

Objective: Pain and distress associated with intranasal midazolam administration can be decreased by administering lidocaine before intranasal midazolam (preadministered lidocaine) or combining lidocaine with midazolam in a single solution (coadministered lidocaine). We hypothesized coadministered lidocaine is non-inferior to preadministered lidocaine for decreasing pain and distress associated with intranasal midazolam administration., Methods: Randomized, outcome assessor-blinded, noninferiority trial. Children aged 6 months to 7 years undergoing laceration repair received intranasal midazolam with preadministered or coadministered lidocaine. Pain and distress were evaluated with the Observational Scale of Behavioral Distress-Revised (OSBD-R) (primary outcome; non-inferiority margin 1.8 units) and the Children's Hospital of Eastern Ontario Pain Scale (CHEOPS) and Faces, Legs, Activity, Cry, Consolability (FLACC) scales and cry duration (secondary outcomes). Secondary outcomes also included adverse events, clinician and caregiver satisfaction, and pain and distress associated with intranasal lidocaine administration., Results: Fifty-one patients were analyzed. Mean OSBD-R scores associated with intranasal midazolam administration were 6.4 (95% confidence interval [CI] 5, 7.8) and 7 (95% CI 5.2, 8.9) units for preadministered and coadministered lidocaine, respectively. The difference of 0.6 (95% CI -1.7, 2.8) units represented an inconclusive non-inferiority determination. CHEOPS and FLACC scores and cry duration were similar between groups. OSBD-R, CHEOPS, and FLACC scores and cry duration associated with intranasal lidocaine administration were 3.8, 9.9, and 6 units, and 56 seconds, respectively. Clinicians considered coadministered lidocaine easier to administer., Conclusion: Pain and distress associated with intranasal midazolam administration were similar when using coadministered or preadministered lidocaine, but our non-inferiority determination was inconclusive. Administration of intranasal lidocaine by itself was associated with a measurable degree of pain and distress.Keywords: intranasal, midazolam, anxiolysis, sedation, emergency department, emergency medicine, pain, distress, pediatric, lidocaine, laceration., Competing Interests: None of the authors have any conflicts of interest to disclose., (© 2020 The Authors. JACEP Open published by Wiley Periodicals LLC on behalf of the American College of Emergency Physicians.)

The prognostic significance of the preoperative electrocardiogram (ECG), particularly intraventricular conduction delays (IVCDs), on postoperative outcomes among patients undergoing noncardiac surgery is uncertain. In a retrospective cohort, we evaluated the risk associated with preoperative IVCDs on in-hospital death and postoperative myocardial infarction (POMI). The 152,479 patients who underwent noncardiac surgery were categorized by preoperative electrocardiographic findings: normal (36.1%), left bundle branch block (LBBB, 1.2%), right bundle branch block (2.9%), nonspecific IVCD (3.3%), and any other ECG abnormality (56.5%). The primary and secondary outcomes were postoperative in-hospital mortality and POMI, respectively. In multivariable-adjusted models, compared with normal ECGs, each electrocardiographic abnormality category was associated with increased risk of postoperative death: LBBB odds ratio (OR) 1.89 (95% confidence interval [CI] 1.35 to 2.65), right bundle branch block OR 1.73 (95% CI 1.33 to 2.24), nonspecific IVCD OR 1.95 (95% CI 1.53 to 2.48), and other abnormal ECG OR 1.94 (95% CI 1.68 to 2.25). ECGs with conduction delays did not confer increased risk of postoperative death compared with other ECG abnormalities. Moreover, receiver operating characteristic analysis of models incorporating demographic and co-morbidity data demonstrated marginal additive benefit of any electrocardiographic data. Risk of POMI was not significantly increased among ECGs with conduction delays compared with both normal and other abnormal ECGs. In conclusion, patients with intraventricular conduction disease, including LBBB, on preoperative ECG are not at greater risk of postoperative in-hospital death or POMI compared with patients with other ECG abnormalities. Furthermore, any preoperative electrocardiographic abnormalities, including intraventricular delays, provide marginal clinical utility beyond demographic and clinical history for predicting postoperative in-hospital death or POMI., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: There are two major classes of cardiac tissue models: the ionic model and the FitzHugh-Nagumo model. During computer simulation, each model entails solving a system of complex ordinary differential equations and a partial differential equation with non-flux boundary conditions. The reproducing kernel method possesses significant applications in solving partial differential equations. The derivative of the reproducing kernel function is a wavelet function, which has local properties and sensitivities to singularity. Therefore, study on the application of reproducing kernel would be advantageous., Objective: Applying new mathematical theory to the numerical solution of the ventricular muscle model so as to improve its precision in comparison with other methods at present., Methods: A two-dimensional reproducing kernel function inspace is constructed and applied in computing the solution of two-dimensional cardiac tissue model by means of the difference method through time and the reproducing kernel method through space., Results: Compared with other methods, this method holds several advantages such as high accuracy in computing solutions, insensitivity to different time steps and a slow propagation speed of error. It is suitable for disorderly scattered node systems without meshing, and can arbitrarily change the location and density of the solution on different time layers., Conclusions: The reproducing kernel method has higher solution accuracy and stability in the solutions of the two-dimensional cardiac tissue model.

IGF-I CA repeat polymorphisms, especially the allele containing CA19 repeats, have been reported to be associated with the risk for various types of cancers. However, the results still remain controversial and ambiguous. This meta-analysis was performed to evaluate the association between IGF-I CA19 repeat polymorphisms and the risk of cancer. Total 18 studies with IGF-I CA19 repeat genotyping on 9,873 patients and 15,607 controls were analyzed. We used random-effects model with a pooled OR of 0.69 (95% CI = 0.60-0.79) for the recessive genetic model, 0.97 (95% CI = 0.86-1.10) for the dominant genetic model, 0.99 (95% CI = 0.86-1.14) for the homozygote comparison and 1.06 (95% CI = 0.91-1.23) for the heterozygote comparison. In the subgroup analysis of recessive model, OR (95% CI) was 0.65 (0.52-0.80) in breast cancer, 0.68 (0.53-0.86) in prostate cancer, and 0.71 (0.52-0.96) in Caucasian. In conclusion, IGF-1 CA19 repeat polymorphisms are unlikely to be a major determinant of susceptibility to cancer. However, the subgroup analysis of recessive model indicates that IGF-I CA19 repeat polymorphisms may reduce the risk of certain types of cancer or in a specific population.

Introduction: Cardiac implantable electronic device (CIED) infections are potentially preventable complications associated with high morbidity, mortality, and cost. A recently developed bio-absorbable antibacterial envelope (TYRX™-A) might prevent CIED infections in high-risk subjects. However, data regarding safety and efficacy have not been published., Methods and Results: In a single-center retrospective cohort study, we compared the prevalence of CIED infections among subjects with ≥2 risk factors treated with the TYRX™-A envelope (N = 135), the nonabsorbable TYRX™ envelope (N = 353), and controls who did not receive an envelope (N = 636). Infection was ascertained by individual chart review. The mean (95% confidence interval) number of risk factors was 3.08 (2.84-3.32) for TYRX™-A, 3.20 (3.07-3.34) for TYRX™, and 3.09 (2.99-3.20) for controls, P = 0.3. After a minimum 300 days follow-up, the prevalence of CIED infection was 0 (0%) for TYRX™-A, 1 (0.3%) for TYRX™, and 20 (3.1%) for controls (P = 1 for TYRX™-A vs. TYRX™, P = 0.03 for TYRX™-A vs. controls, and P = 0.002 for TYRX™ vs. controls). In a propensity score-matched cohort of 316 recipients of either envelope and 316 controls, the prevalence of infection was 0 (0%) and 9 (2.8%), respectively, P = 0.004. When limited to 122 TYRX™-A recipients and 122 propensity-matched controls, the prevalence of CIED infections was 0 (0%) and 5 (4.1%), respectively, P = 0.024., Conclusions: Among high-risk subjects, the TYRX™-A bio-absorbable envelope was associated with a very low prevalence of CIED related infections that was comparable to that seen with the nonabsorbable envelope., (© 2015 Wiley Periodicals, Inc.)

β-Glycosidase from Thermococcus kodakarensis KOD1 is a hyperthermophilic enzyme with β-glucosidase, β-mannosidase, β-fucosidase and β-galactosidase activities. Sequence alignment with other β-glycosidases from hyperthermophilic archaea showed two unique active site residues, Gln77 and Asp206. These residues were represented by Arg and Asp in all other hyperthermophilic β-glycosidases. The two active site residues were mutated to Q77R, D206N and D206Q, to study the role of these unique active site residues in catalytic activity and to alter the substrate specificity to enhance its β-glucosidase activity. The secondary structure analysis of all the mutants showed no change in their structure and exhibited in similar conformation like wild-type as they all existed in dimer form in an SDS-PAGE under non-reducing conditions. Q77R and D206Q affected the catalytic activity of the enzyme whereas the D206N altered the catalytic turn-over rate for glucosidase and mannosidase activities with fucosidase activity remain unchanged. Gln77 is reported to interact with catalytic nucleophile and Asp206 with axial C2-hydroxyl group of substrates. Q77R might have made some changes in three dimensional structure due to its electrostatic effect and lost its catalytic activity. The extended side chains of D206Q is predicted to affect the substrate binding during catalysis. The high-catalytic turn-over rate by D206N for β-glucosidase activity makes it a useful enzyme in cellulose degradation at high temperatures., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Humoral immunity against diverse pathogens is rapidly elicited from natural antibody repertoires of limited complexity. But the organizing principles underlying the antibody repertoires that facilitate this immunity are not well-understood. We used HER2 as a model immunogen and reverse-engineered murine antibody response through constructing an artificial antibody library encoded with rudimentary sequence and structural characteristics learned from high throughput sequencing of antibody variable domains. Antibodies selected in vitro from the phage-displayed synthetic antibody library bound to the model immunogen with high affinity and specificities, which reproduced the specificities of natural antibody responses. We conclude that natural antibody structural repertoires are shaped to allow functional antibodies to be encoded efficiently, within the complexity limit of an individual antibody repertoire, to bind to diverse protein antigens with high specificity and affinity. Phage-displayed synthetic antibody libraries, in conjunction with high-throughput sequencing, can thus be designed to replicate natural antibody responses and to generate novel antibodies against diverse antigens.

Scientists are interested in understanding the molecular origin of protein thermostability and thermoactivity for possible biotechnological applications. The enzymes from extremophilic organisms have been of particular interest in the last two decades. β-glycosidase, Tkβgly is a hyperthermophilic enzyme from Thermococcus kodakarensis KOD1. Tkβgly contains two conserved cysteine residues, C88 and C376. The protein tertiary structure obtained through homology modeling suggests that the C88 residue is located on the surface whereas C376 is inside the protein. To study the role of these cysteine residues, we substituted C88 and C376 with serine residues through site-directed mutagenesis. The wild-type and C376S protein existed in dimeric form and C88S in monomeric form, in an SDS-PAGE gel under non-reducing conditions. Optimal temperature experiments revealed that the wild-type was active at 100 °C whereas the C88S mutant exhibited optimal activity at 70 °C. The half-life of the enzyme at 70 °C was drastically reduced from 266 h to less than 1 h. Although C88 was not present in the active site region, the kcat/Km of C88S was reduced by 2-fold. Based on the structural model and biochemical properties, we propose that C88 is crucial in maintaining the thermostability and thermoactivity of the Tkβgly enzyme.

Vascular endothelial growth factor (VEGF) polymorphisms, specifically +405G/C (rs2010963), reportedly influence the risk for various digestive cancers. However, the consequences of these polymorphisms remain controversial and ambiguous. Therefore, we performed a meta-analysis of 11 studies with VEGF +405G/C genotyping on 2,862 patients and 3,028 controls using the random effects model. We obtained a pooled odds ratio (OR) of 1.04 (95% confidence interval (CI) = 0.86-1.26) for the recessive genetic model, 1.07 (95% CI = 0.81-1.42) for the dominant genetic model, 1.09 (95% CI = 0.81-1.47) for the homozygote comparison, and 1.03 (95% CI = 0.83-1.27) for the heterozygote comparison. In the subgroup analysis of the recessive model, the OR was 1.20 (95% CI = 1.02-1.40) in colorectal cancer. These results show that VEGF +405G/C polymorphisms are unlikely to be a major determinant of susceptibility to digestive cancer. Furthermore, the subgroup analysis of recessive model indicates that VEGF +405G/C polymorphisms increase the risk for colorectal cancer.

Blinds and severe visual impairments can utilize tactile sticks to assist their walking. However, they cannot fully understand the dangling objects in front of their walking routes. This research proposed a mobile real-time dangling objects sensing (RDOS) prototype, which is located on the cap to sense any front barrier. This device utilized cheap ultrasonic sensor to act as another complement eye for blinds to understand the front dangling objects. Meanwhile, the RDOS device can dynamically adjust the sensor's front angle that is depended on the user's body height and promote the sensing accuracy. Meanwhile, two major required algorithms, height-angle measurement and ultrasonic sensor alignment, are proposed with this prototype. The research team also integrated the RDOS device prototype with mobile Android devices by communicating with Bluetooth to record the walking route.

Interleukin-12 (IL-12) is a pleiotropic cytokine which bridges innate and adaptive immunity in defense against pathogens. IL-12 proved to be an effective and successful adjuvant to enhance both the innate and adaptive immune responses and could be applicable for a rationale vaccine formulation in fish against pathogen infection. We have cloned the p35 and p40 cDNAs of IL-12 from orange-spotted grouper (Epinephelus coioides). Grouper IL-12 most resembles with sea bass orthologues; moderate to low identity with other teleost and mammalian counterparts. The structural model of grouper IL-12 heterodimer revealed NC(141)F three amino acid patch of grouper p35, which is present in teleost p35 but absent in mammalian and avian p35, and is spatially nearby the conserved cysteine residue located at A-helix of p35 to form a disulfide bond when the 14aa peptide located at loop 1 of grouper p35 was aligned with human corresponding exon 4, instead of exon 5. The results indicated that the loss of this 3aa patch during evolution was compensated by the duplication of exon 4 in mammalian p35 to gain another cysteine residue to form a disulfide bond, evidenced by chicken p35 which does not contain NCF corresponding 3-aa patch nor exon 4 duplication. Accordingly, the inter-chain disulfide bond of IL-12 heterodimer is conserved from teleost to mammalian IL-12. A single chain grouper IL-12 (scgIL-12) construct linked by (G4S)3 was successfully expressed in baculovirus-insect cell system; its identity has been confirmed by LC/MS/MS. In addition, the biological activity of recombinant scgIL-12 (rscgIL-12) are demonstrated for its stimulation of PBL proliferation, chemotactic migration, induction of TNF-α gene expression and a plausible adjuvant effect of prolonged protection against parasite infection in fish. We illustrated the first time in lower vertebrate that grouper IL-12 possesses both cytokine and chemokine activities., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Three new transition metal complexes [Mn2(DCA)2(bipy)2]·5H2O (1), [M2(DCA)2(bipy)2(H2O)]·10H2O(M=Ni(II)(2);Zn(II)(3)), (DCA=demethylcantharate, 7-oxabicyclo[2,2,1]heptane-2,3-dicarboxylate, C8H8O5) were synthesized and characterized by elemental analysis, molar conductance, infrared spectra and X-ray diffraction techniques. Each metal ion was six-coordinated in complexes. Complex 1 has a Mn2O2 center. Complexes 2 and 3 have asymmetric binuclear structure. Great amount of intermolecular hydrogen-bonding and π-π(*) stacking interactions were formed in these complex structures. The DNA-binding properties of complexes were investigated by electronic absorption spectra and viscosity measurements. The DNA binding constants Kb/(Lmol(-1)) were 1.71×10(4) (1), 2.62×10(4) (2) and 1.59×10(4) (3) at 298 K. The complexes could quench the intrinsic fluorescence of bovine serum albumin (BSA) strongly through static quenching. The protein binding constants Ka/(L mol(-1)) were 7.27×10(4) (1), 4.55×10(4) (2) and 7.87×10(4) L mol(-1) (3) and binding site was one. The complexes bind more tightly with DNA and BSA than with ligands. Complexes 1 and 3 had stronger inhibition ratios than Na2(DCA) against human hepatoma cells (SMMC-7721) lines and human gastric cancer cells (MGC80-3) lines in vitro. Complex 3 showed the strongest antiproliferative activity against SMMC-7721 (IC50=29.46±2.12 μmol L(-1)) and MGC80-3 (IC50=27.02±2.38 μmol L(-1)), which shows potential in anti-cancer drug development., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: Diabetic retinopathy is one of the most common microvascular complications of diabetes and one of the major causes of adult visual impairment in national surveys in Taiwan. This study aimed to identify the healthcare costs of Taiwan's National Health Insurance program on behalf of diabetic patients with stable or progressive retinopathy., Methods: A retrospective cohort study was conducted with 4,988 medication-using diabetic retinopathy subjects >or= 40 years of age under National Health Insurance Program coverage between 2000 and 2004. Study cohort subjects were recorded as having diabetic retinopathy according to ICD-9-CM codes. States of diabetic retinopathy were strategically divided into stable and progressive categories according to subjects' conditions at follow-up in 2004. Expenditures were calculated and compared for the years 2000 and 2004., Results: During the 4-year follow-up (2000 through 2004), 4,116 subjects (82.5%) of 4,988 diabetic subjects were in the stable category, and 872 (17.5%) were in the progressive category. Average costs of those in the normal category increased by US $48 from US $1921 in 2000 to US $1969 in 2004 (p = 0.594), whereas costs for those progressing from normal to non-proliferative diabetic retinopathy (NPDR) or proliferative diabetic retinopathy (PDR) increased by US $1760, from US $1566 in 2000 to US $3326 in 2004 (p < 0.001). The PDR category had the highest average costs at US $3632 in 2000. The NPDR-to-PDR category experienced the greatest increase in costs at US $3482, from US $2723 in 2000 to US $6204 in 2004 (p = 0.042), and the greatest percentage of increase at 2.3% (2.2% when adjusted by comparing to normal category)., Conclusions: This large-scale longitudinal study provides evidence that increased healthcare costs are associated with progressive diabetic retinopathy among diabetic NHI enrollees in Taiwan.

The grouper iridovirus (GIV) belongs to the family Iridoviridae, whose genome contains an antiapoptotic B-cell lymphoma (Bcl)-2-like gene. This study was carried-out to understand whether GIV blocks apoptosis in its host. UV-irradiated grouper kidney (GK) cells underwent apoptosis. However, a DNA fragmentation assay of UV-exposed GK cells after GIV infection revealed an inhibition of apoptosis. The UV- or heat-inactivated GIV failed to inhibit apoptosis, implying that a gene or protein of the viral particle might contribute to an apoptosis inhibitory function. The DNA ladder assay for GIV-infected GK cells after UV irradiation confirmed that apoptosis inhibition was an early process which occurred as early as 5 min post-infection. A GIV-Bcl sequence comparison showed distant sequence similarities to that of human and four viruses; however, all possessed the putative Bcl-2 homology (BH) domains of BH1, BH2, BH3, and BH4, as well as a transmembrane domain. Northern blot hybridization showed that GIV-Bcl transcription began at 2 h post-infection, and the mRNA level significantly increased in the presence of cycloheximide or aphidicolin, indicating that this GIV-Bcl is an immediate-early gene. This was consistent with the Western blot results, which also revealed that the virion carries the Bcl protein. We observed the localization of GIV-Bcl on the mitochondrial membrane and other defined intracellular areas. By immunostaining, it was proven that GIV-Bcl-expressing cells effectively inhibited apoptosis. Taken together, these results demonstrate that GIV inhibits the promotion of apoptosis by GK cells, which is mediated by the immediate early expressed viral Bcl gene.

We have cloned FSH-beta cDNA from duck pituitary gland by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end (RACE) methods. The cloned duck FSH-beta cDNA contains 1909-bp nucleotides including 396-bp of open-reading frame and 1491-bp of 3'-untranslational region. The open-reading frame encodes a 131-amino acid protein with a putative 20-amino acid signal peptide and a putative 111-amino acid mature protein. The deduced amino acid sequence shows a remarkable similarity (94-98%) to those of other avian FSH-beta subunits; while it exhibits lower similarities with those of turtles (82-84%), mammals (63-71%), and amphibians (53-57%). The structural model analysis of duck FSH suggests that the cysteine-knot and beta-strands for maintaining the specific structural frame, and the "seat-belt" loop for specific binding to FSH receptor have been conserved in tetrapodian FSH-betas.

A cDNA encoding thyroid stimulating hormone beta-subunit (TSHbeta) was cloned from pituitary of the Chinese soft-shell turtle, Pelodiscus sinensis, and its regulation of mRNA expression was investigated for the first time in reptile. The Chinese soft-shell turtle TSHbeta cDNA was cloned from pituitary RNA by reverse transcription and polymerase chain reaction (RT-PCR), and rapid amplification cDNA end (RACE) methods. The Chinese soft-shell turtle TSHbeta cDNA consists of 580-bp nucleotides, including 67-bp nucleotides of 5'-untranslated region (UTR), 402-bp of the open reading frame, and 97-bp of 3'-UTR followed by a 14 poly (A) trait. It encodes a precursor protein molecule of 133 amino acids with a putative signal peptide of 19 amino acids and a putative mature protein of 114 amino acids. The number and position of 12 cysteine residues, presumably forming six disulfide bonds, one putative asparagine-linked glycosylation site, and six proline residues that are found at positions for changing the backbone direction of the protein have been conserved in the turtle as in other vertebrate groups. The deduced amino acid sequence of the Chinese soft-shell turtle TSHbeta mature protein shares identities of 82-83% with birds, 71-72% with mammals, 49-57% with amphibians, and 44-61% with fish. The Chinese soft-shell turtle pituitaries were incubated in vitro with synthetic TRH (TSH-releasing hormone), thyroxine and triiodothyronine at doses of 10(-10) and 10(-8)M. TRH stimulated, while thyroid hormones suppressed, TSHbeta mRNA levels in dose-related manner. The sequences of cDNA and its deduced peptide of TSHbeta as well as the regulation of its mRNA level were reported for the first time in reptile.

The cDNAs encoding beta-subunits of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) have been cloned from the pituitary of snakehead fish, Channa maculata, and the three-dimensional structural models of the encoded FSH and LH were investigated. The cloned cDNAs, including 5'-untranslated region (UTR), open-reading frame, and 3'-UTR followed by a poly(A) tail, were obtained by reverse transcription-polymerase chain reaction and rapid amplification of cDNA end methods. The open-reading frames of FSH-beta cDNA encodes a 120-amino acid protein with a signal peptide of 18 amino acids and a mature protein of 102 amino acids; while LH-beta cDNA encodes a 140-amino acid protein with a signal peptide of 33 amino acids and a mature protein of 115 amino acids. The amino acid sequence identities of snakehead fish FSH-beta and LH-beta in comparison with other fish are 27.8-81.9% and 45.2-88.8%, respectively; while in comparison with tetrapods are 26.2-28.9% and 37.5-51.2%, respectively. Both FSH-beta and LH-beta of snakehead fish resemble most to those of Perciformes, implying their closer phylogenetic relationship. All 12 cysteine residues are conserved in snakehead fish LH-beta; while 11 cysteine residues are conserved in its FSH-beta. The third cysteine is absent in snakehead fish FSH-beta; instead, a positionally shifted cysteine residue is present at the N-terminus, as found in some phylogenetic related fish. The structure models of snakehead fish FSH and LH, constructed by using the crystal structures of human FSH and human chorionic gonadotropin as respective template, showed that the positionally shifted N-terminal cysteine residue of snakehead fish FSH-beta likely can substitute the third cysteine to form a disulfide bond with the 12th cysteine.

Follicle-stimulating hormone (FSH) is a member of the pituitary glycoprotein hormone family. These hormones are composed of two dissimilar subunits, alpha and beta. Very little information is available regarding the nucleotide and amino acid sequence of FSHbeta in reptilian species. For better understanding of the phylogenetic diversity and evolution of FSH molecule, we have isolated and sequenced the complementary DNA (cDNA) encoding the Chinese soft-shell turtle (Pelodiscus sinensis, Family of Trionychidae) FSHbeta precursor molecule by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end (RACE) methods. The cloned Chinese soft-shell turtle FSHbeta cDNA consists of 602-bp nucleotides, including 34-bp nucleotides of the 5'-untranslated region (UTR), 396-bp of the open reading frame, and 3'-UTR of 206-bp nucleotides. It encodes a 131-amino acid precursor molecule of FSHbeta subunit with a signal peptide of 20 amino acids followed by a mature protein of 111 amino acids. Twelve cysteine residues, forming six disulfide bonds within beta-subunit and two putative asparagine-linked glycosylation sites, are also conserved in the Chinese soft-shell turtle FSHbeta subunit. The deduced amino acid sequence of the Chinese soft-shell turtle FSHbeta shares identities of 97% with Reeves's turtle (Family of Bataguridae), 83-89% with birds, 61-70% with mammals, 63-66% with amphibians and 40-58% with fish. By contrast, when comparing the FSHbeta with the beta-subunits of the Chinese soft-shell turtle luteinizing hormone and thyroid stimulating hormone, the homologies are as low as 38 and 39%, respectively. A phylogenetic tree including reptilian species of FSHbeta subunits, is presented for the first time. Out of various tissues examined, FSHbeta mRNA was only expressed in the pituitary gland and can be up-regulated by gonadotropin-releasing hormone in pituitary tissue culture as estimated by fluorescence real-time PCR analysis.

Natural disasters present significant potential for injuries and death. Unlike the experience of Hurricane Andrew that destroyed a vast surface area in the rural countryside, Typhoon Nari in Taipei proved that significant damages from natural disasters also can happen to modern health care systems in urban areas. To ameliorate such damages, specific structural, nonstructural, and administrative issues must be taken into account. Such issues include the location of the health facility, design of the infrastructure, storage of equipment and machines, maintenance, medical, and nonmedical operations. Specific considerations, such as early evacuation and securing the safety of the patients before a disaster, should be emphasized. Recovery plans that determine how soon medical service can be restored to the community should also be established. Emphasis on emergency-response preparedness, mitigation procedures, and recovery efforts should all be included in a comprehensive emergency plan against the destruction caused by natural hazards.

The complete complementary DNA (cDNA) of proopiomelanocortin (POMC), a common precursor of opioid hormone beta-endorphin, melanotropin (MSH), and corticortropin (ACTH), was cloned and sequenced from pituitary and hypothalamus of mud turtle (Pelodiscus sinensis) by RT-PCR and rapid amplification of cDNA end (RACE) methods. Two transcripts of POMC mRNAs with different polyadenylation sites were observed. Both transcripts had an open reading frame encoding a 261-amino acid peptide containing nine dibasic amino acids (pair of Arg and Lys), putative proteolytic cleavage sites for processing to functional peptides. All the functional peptide fragments of mud turtle POMC, gamma-MSH, alpha-MSH, ACTH, beta-MSH, and beta-endorphin, are flanked by dibasic residues as found in other tetrapods, implying that it could be processed to give rise to all members of POMC-derived peptides. The deduced amino acid sequences of mud turtle POMC displays 63-67% identity with amphibian, 59% with chicken, 48-53% with mammals, and 37-59% identity with fish. However, functional peptide fragments are much more conserved than overall sequence and intervening fragments. In addition to pituitary and brain, mud turtle POMC mRNAs are also expressed in many peripheral tissues, such as skin, thyroid, and testis. This is the first report on the complete sequence of cDNA nucleotides and deduced amino acids of POMC in reptile.

Follicle-stimulating hormone (FSH) is a member of pituitary glycoprotein hormones that are composed of two dissimilar subunits, alpha and beta. Very little information is available regarding the nucleotide and amino acid sequence of FSH-beta in avian species. For better understanding of the phylogenic diversity and evolution of FSH molecule, we have isolated and sequenced the complete complementary DNA (cDNA) encoding chicken FSH-beta precursor molecule by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end (RACE) methods. The cloned chicken FSH-beta cDNA consists of 2457-bp nucleotides, including 44-bp nucleotides of the 5'-untranslated region (UTR), 396 bp of the open reading frame, and an extraordinarily long 3'-UTR of 2001-bp nucleotides followed by a poly(A)((16)) tail. It encodes a 131-amino-acid precursor molecule of FSH-beta-subunit with a signal peptide of 20 amino acids followed by a mature protein of 111 amino acids. Twelve cysteine residues, forming six disulfide bonds within beta-subunit and two putative asparagine-linked glycosylation sites, are also conserved in the chicken FSH-beta-subunit. Four proline residues, presumably responsible for changing the backbone direction of protein structure, are conserved in chicken FSH-beta-subunit as well. The nucleotide sequence of chicken FSH-beta cDNA shows high homology with quail FSH-beta cDNA, 97% homology in the open reading frame, and 85% homology in the 3'-UTR. The deduced amino acid sequence of chicken FSH-beta-subunit shows a remarkable similarity to other avian FSH-beta-subunits, 98% homology with quail, and 93% homology with ostrich, whereas a lower similarity (66 to 70%) is noted when compared with mammalian FSH-beta-subunits. By contrast, when comparing with the beta-subunits of chicken luteinizing hormone and thyroid-stimulating hormone, the homologies are as low as 37 and 40%, respectively. FSH-beta mRNA was only expressed in pituitary gland out of various tissues examined and can be up-regulated by gonadotropin-releasing hormone in pituitary tissue culture as estimated by real-time quantitative PCR., ((C)2002 Elsevier Science (USA).)

The biopotencies of pituitary gonadotropins (GTHs) from various vertebrate classes were examined in an in vitro rat Sertoli cell bioassay which was previously established for mammalian follicle-stimulating hormones (FSHs). Potencies of the gonadotropins were determined by incubation of Sertoli cells obtained from 10-day-old rats, with increasing doses of GTHs, which resulted in dose-related and parallel estradiol-17 beta formation converted from added 19-hydroxy-androstenedione. In general, mammalian (human and ovine) FSHs were most potent, avian (chicken, turkey, and ostrich) FSHs were intermediate, and reptilian (snapping turtle) and amphibian (bullfrog) FSHs were the least potent. By contrast, mammalian and bullfrog luteinizing hormones (LHs) were inactive or negligibly active in this assay; avian LHs possessed one-fifth to one-half of the potency of the FSH preparations of the same species. The data suggest that the assay is specific for FSHs from mammalian and amphibian species and is relatively specific for FSHs from avian species. Both snapping turtle FSH and LH exhibited low and similar potencies in this bioassay. Black silver carp GTH was also active in this assay, although its potency was much lower. The present study has demonstrated that the immature rat Sertoli cell aromatase assay in vitro is useful for measurement of FSH contents in mammalian species and of FSH activity in diverse nonmammalian species. It also provides an approach for the investigation of structure-function relations of gonadotropin in diverse vertebrate species in relation to phylogenic patterns and specificity of hormone-receptor interaction. The findings from the present study imply that the binding sites of vertebrate FSHs share a certain degree of homology and that the binding sites of FSH receptors on Sertoli cells from immature rat have a relatively low degree of animal class specificity.

In the present studies, an inverse relationship was observed between LTB4 and LXA4 formation when the concentration of LTA4 was increased from 50 microM to 250 microM. LTB4 was the major product at low concentration of LTA4 where as LXA4 was the major product at high concentration. This suggests that LTA4 hydrolase activity is present in MC and activated by low concentrations of LTA4. At high concentrations of LTA4, 15- or 12-lipoxygenase (LO) in MC may participat in LXs formation. Radioimmunoassays on HETEs MC activated with A23187 (3uM) were in the rank order of 15-HETE greater than 12-HETE greater than 5-HETE. Incubations of MC with 15-HPETE failed to produce any LX-like material. These results suggest that 1) 5-LO activity is not expressed in MC under these conditions, but LTA4 hydrolase activity is present in MC, 2) there are major activities of 12-LO and 15-LO in MC that are responsible for the generation of LXA4, LXB4 and HETES, 3) the production of LX and 12- or 15-LO products of AA may counterregulate the production of LTB4 and may modulate the response of the MC to inflammatory stimuli.

Gonadotropins (GTH) were isolated from pituitary glands of grass carp and their properties investigated. Acetone-dried pituitaries were extracted with ammonium acetate-ethanol and were then passed through cation exchanger, anion exchanger, and gel filtration column. Two glycoprotein gonadotropins (DE I and DE II) were isolated of MW 56,000 and 53,000 daltons, respectively, as determined by gel filtration technique. Isoelectric points of the two GTHs also differed from each other. Sialic acid contents were 3.64% and 1.47% for DE I and DE II, respectively. Both GTH's stimulated, in a similar manner, testicular androgen and ovarian estradiol synthesisin vitro.