Miracle-dz/UniFAR-FTUs huggingface dataset:
Expression, Purification, and Biochemical Characterization of the Amino-terminal Extracellular Domain of the Human Calcium Receptor*

We purified the extracellular domain (ECD) of the human calcium receptor (hCaR) from the medium of HEK-293 cells stably transfected with a hCaR cDNA containing an isoleucine 599 nonsense mutation. A combination of lectin, anion exchange, and gel permeation chromatography yielded milligram quantities of >95% pure protein from 15 liters of starting culture medium. The purified ECD ran as an ∼78-kDa protein on SDS-polyacrylamide gel electrophoresis and was found to be a disulfide-linked dimer. Its NH2-terminal sequence, carbohydrate content, and CD spectrum were defined. Tryptic proteolysis studies showed two major sites accessible to cleavage. These studies provide new insights into the structure of the hCaR ECD. Availability of purified ECD protein should permit further structural studies to help define the mechanism of Ca2+ activation of this G protein-coupled receptor.
Proliferative response of bronchoalveolar lymphocytes to beryllium. A test for chronic beryllium disease.

STUDY OBJECTIVE to ascertain whether measuring the proliferative response of bronchoalveolar lymphocytes to beryllium salts is useful for diagnosing chronic beryllium disease. DESIGN prospective case series compared to normal volunteers and patients with sarcoidosis. SETTING university referral center. PATIENTS twenty-three consecutive beryllium workers were evaluated. Fourteen had chronic beryllium disease diagnosed on the basis of histologic evidence of a progressive pulmonary granulomatosis. Four had biopsy evidence of non-beryllium disease. Three had probable chronic beryllium disease but did not have lung biopsies. Two did not have biopsies and had basilar fibrosis on chest roentgenogram suggestive of non-beryllium lung disease. These patients were compared with 6 normal volunteers and 16 patients with sarcoidosis. MEASUREMENTS AND MAIN RESULTS bronchoalveolar lavage was done and the proliferative response of the lung cells to two beryllium salts was tested. A positive proliferative test was defined as a stimulation index of more than five on two determinations. The sensitivity of this test was 100% in the 14 patients with definite chronic beryllium disease. The specificity of the test was also 100% among the normal volunteers and the 16 patients with sarcoidosis. The test was positive in none of the four patients with biopsy evidence of non-beryllium disease, none out of two patients with lower lobe fibrosis suggestive of non-beryllium disease, and all of three patients with probable chronic beryllium lung disease. CONCLUSIONS the proliferative response of bronchoalveolar lymphocytes to beryllium appears to be a useful test for chronic beryllium disease.
Tuned Escherichia coli as a host for the expression of disulfide‐rich proteins

Disulfide‐bond formation is a major post‐translational modification and is essential for protein folding, stability, and function. This is especially true for secreted proteins, many of which possess great potential for biotechnological applications. Focusing on the use of Escherichia coli for the production of this class of proteins, we describe the mechanisms that maintain redox compartmentalization in the cell, with an emphasis on those that promote the formation and isomerization of disulfide bonds in the bacterial periplasm, while presenting parallel pathways in the eukaryotic endoplasmic reticulum. Based on these concepts, we review the use of E. coli as a cell factory for the production of heterologous disulfide‐containing proteins using either peri‐ or cytoplasmic expression and, in particular, how these compartments can be tuned to improve the yield of correctly folded recombinant proteins. Finally, we describe a few examples of the production of small disulfide‐rich proteins (protease inhibitors) to illustrate how soluble, active, and fully oxidized recombinants may be successfully obtained upon peri‐ or cytoplasmic expression in E. coli.
L‐Type Voltage‐Gated Calcium Channels Are Involved in the in Vivo and in Vitro Expression of Fear Conditioning

Abstract: Fear conditioning, a behavioral model of fear learning and cue‐related anxiety, causes enhanced neuronal transmission in the thalamic to lateral amygdala pathway. 1,2 In the expression phase of learned fear, this increased transmission recorded in vitro is revealed in increased amplitudes of excitatory postsynaptic currents (EPSCs) and occlusion of paired‐pulse facilitation (PPF) implicating a presynaptic increase in transmitter release. Here we examined the contribution of L‐type calcium channels in fear conditioning. We measured the effect of nimodipine (Nim, 1.5–20 mg/kg), an L‐type calcium channel antagonist, on fear‐potentiated startle in which startle was assessed in animals receiving paired or unpaired tone and foot shock. Nim administered intraperitoneally blocked fear‐potentiated startle but not baseline startle in a dose‐dependent manner. We also analyzed the effect of Nim (10 μM) in vitro on synaptic facilitation of EPSCs and PPF in slices from naïve control, unpaired control, and fear‐conditioned animals. In neurons from naïve control animals, Nim had no effect on EPSC amplitude or PPF, but in slices from fear‐conditioned rats, Nim reduced EPSC amplitude, suggesting the recruitment of L‐type calcium channels within the fear‐conditioning pathway. Nim increased PPF in slices from fear‐conditioned animals, suggesting that L‐type calcium channels may contribute to increased probability of release in fear conditioning. In slices from unpaired animals, Nim decreased synaptic transmission but had little effect on PPF, suggesting that stress or contextual fear learning may induce L‐type channel activity in fear‐conditioned and unpaired control animal groups. We also analyzed protein expression of the α1C and α1D L‐type calcium channel subunits isolated from the amygdala and found that α1C protein was significantly increased in fear‐conditioned animals. These findings suggest that L‐type calcium channels play a role in the amygdala in cued fear conditioning and have important implications in the treatment of anxiety and in emotional learning and plasticity.
High levels of plasma IL‐10 and expression of IL‐10 by keratinocytes during visceral leishmaniasis predict subsequent development of post‐kala‐azar dermal leishmaniasis

Some patients develop post‐kala‐azar dermal leishmaniasis (PKDL) after they have been treated for the systemic infection kala‐azar (visceral leishmaniasis). It has been an enigma why the parasites cause skin symptoms after the patients have been successfully treated for the systemic disease. We report here that PKDL development can be predicted before treatment of visceral leishmaniasis, and that IL‐10 is involved in the pathogenesis. Before treatment of visceral leishmaniasis, Leishmania parasites were present in skin which appeared normal on all patients. However, IL‐10 was detected in the keratinocytes and/or sweat glands of all patients who later developed PKDL (group 1) and not in any of the patients who did not develop PKDL (group 2). Furthermore, the levels of IL‐10 in plasma as well as in peripheral blood mononuclear cell culture supernatants were higher in group 1 than in group 2.
A large multicenter study of methicillin-susceptible and methicillin-resistant Staphylococcus aureus prosthetic joint infections managed with implant retention.

BACKGROUND Several series predicting the prognosis of staphylococcal prosthetic joint infection (PJI) managed with debridement, antibiotics, and implant retention (DAIR) have been published, but some of their conclusions are controversial. At present, little is known regarding the efficacy of the different antibiotics that are used or their ability to eliminate methicillin-resistant S. aureus (MRSA) infection. METHODS This was a retrospective, multicenter, observational study of cases of PJI by S. aureus that were managed with DAIR (2003-2010). Cases were classified as failures when infection persistence/relapse, death, need for salvage therapy, or prosthesis removal occurred. The parameters that predicted failure were analyzed with logistic and Cox regression. RESULTS Out of 345 episodes (41% men, 73 years), 81 episodes were caused by MRSA. Fifty-two were hematogenous, with poorer prognoses, and 88% were caused by methicillin-susceptible S. aureus (MSSA). Antibiotics were used for a median of 93 days, with similar use of rifampin-based combinations in MSSA- and MRSA-PJI. Failure occurred in 45% of episodes, often early after debridement. The median survival time was 1257 days. There were no overall prognostic differences between MSSA- and MRSA-PJI, but there was a higher incidence of MRSA-PJI treatment failure during the period of treatment (HR 2.34), while there was a higher incidence of MSSA-PJI treatment failure after therapy. Rifampin-based combinations exhibited an independent protective effect. Other independent predictors of outcome were polymicrobial, inflammatory, and bacteremic infections requiring more than 1 debridement, immunosuppressive therapy, and the exchange of removable components of the prosthesis. CONCLUSIONS This is the largest series of PJI by S. aureus managed with DAIR reported to date. The success rate was 55%. The use of rifampin may have contributed to homogenizing MSSA and MRSA prognoses, although the specific rifampin combinations may have had different efficacies.
Hypoglycemia: A Potent Stimulus to Secretion of Growth Hormone

In normal subjects, hypoglycemia produces an abrupt and sustained rise in levels of human growth hormone in plasma. This effect is independent of insulin, glucagon, or epinephrine. Prolonged fasting is accompanied by a rise in the hormone level in plasma. Measurement of this hormone after induced hypoglycemia is a specific test for pituitary somatotropic function.
Extended enoxaparin monotherapy for acute symptomatic pulmonary embolism

We investigated the efficacy and safety of extended enoxaparin monotherapy in symptomatic patients with acute pulmonary embolism (PE). We randomized 40 patients in a 1:1 allocation to enoxaparin monotherapy (1 mg/kg twice daily for 10-18 days, and then 1.5 mg/kg once daily until day 90) (n = 20) or to enoxaparin 1.0 mg/kg twice daily as a bridge to warfarin with a target international normalized ratio of 2.0-3.0 for 90 days (at least 10 doses of enoxaparin overlapping with warfarin for at least 4 days) (n = 20). All patients underwent echocardiography, cardiac troponin I (TnI), and brain natriuretic peptide testing to identify patients with an increased likelihood of adverse clinical outcomes. The end-points were newly diagnosed deep venous thrombosis (DVT) or PE and bleeding events through day 90. In 15 patients on extended enoxaparin therapy, we used repeated measure analysis of variance (ANOVA) to investigate differences in anti-Xa levels obtained at 2, 4, 8 and 12 weeks. The patients’ mean age was 52 ± 17 years; the most common comorbidities were obesity (58%), hypertension (30%), concomitant DVT (30%) and cancer (15%). Twelve (30%) patients had elevated cardiac TnI >0.1 mg/l and 11 (28%) had moderate or severe right ventricular dysfunction on echocardiography. Ten (25%) patients received thrombolysis with a continuous infusion of 100 mg alteplase prior to randomization. During a 90-day follow-up, one patient from the enoxaparin monotherapy group suffered symptomatic distal DVT; one from the warfarin group had recurrent symptomatic PE (p= 1.0). None of the study patients had major hemorrhage; two warfarin group patients had minor bleeding compared with none in the enoxaparin monotherapy group (p= 0.49). Repeated measure ANOVA did not reveal significant differences in anti-Xa levels over time (p= 0.217). In patients with acute symptomatic PE, extended enoxaparin monotherapy is feasible and warrants further investigation in a large clinical trial.
Teaching Effectiveness in Fluid Discipline: The Case of Cost Management Education in India

The paper has been organised initially to briefly examine the fluid characteristic of cost management discipline through its historical developments. It then proceeds to trace the existing status of cost management education, encompassing substantive issues on text contents, sequencing and teaching methods in Indian business schools. The Paper finally seeks to achieve its objective by recognising and integrating purpose-based application, manufacturing orientation and descriptive theory development.
Selection and Tuning of a Fast and Simple Phase-Contrast Microscopy Image Segmentation Algorithm for Measuring Myoblast Growth Kinetics in an Automated Manner

Abstract Acquiring and processing phase-contrast microscopy images in wide-field long-term live-cell imaging and high-throughput screening applications is still a challenge as the methodology and algorithms used must be fast, simple to use and tune, and as minimally intrusive as possible. In this paper, we developed a simple and fast algorithm to compute the cell-covered surface (degree of confluence) in phase-contrast microscopy images. This segmentation algorithm is based on a range filter of a specified size, a minimum range threshold, and a minimum object size threshold. These parameters were adjusted in order to maximize the F-measure function on a calibration set of 200 hand-segmented images, and its performance was compared with other algorithms proposed in the literature. A set of one million images from 37 myoblast cell cultures under different conditions were processed to obtain their cell-covered surface against time. The data were used to fit exponential and logistic models, and the analysis showed a linear relationship between the kinetic parameters and passage number and highlighted the effect of culture medium quality on cell growth kinetics. This algorithm could be used for real-time monitoring of cell cultures and for high-throughput screening experiments upon adequate tuning.
Differential Requirements for OX40 Signals on Generation of Effector and Central Memory CD4+ T Cells1

Memory T cells can be divided into effector memory (TEM) and central memory (TCM) subsets based on their effector function and homing characteristics. Although previous studies have demonstrated that TCR and cytokine signals mediate the generation of the two memory subsets of CD8+ T cells, the mechanisms for generation of the CD4+ TEM and TCM cell subsets are unknown. We found that OX40-deficient mice showed a marked reduction in the number of CD4+ TEM cells, whereas the number of CD4+ TCM cells was normal. Adoptive transfer experiments using Ag-specific CD4+ T cells revealed that OX40 signals during the priming phase were indispensable for the optimal generation of the CD4+ TEM, but not the CD4+ TCM population. In a different transfer experiment with in vitro established CD4+CD44highCD62Llow (TEM precursor) and CD4+CD44highCD62Lhigh (TCM precursor) subpopulations, OX40-KO TEM precursor cells could not survive in the recipient mice, whereas wild-type TEM precursor cells differentiated into both TEM and TCM cells. In contrast, TCM precursor cells mainly produced TCM cells regardless of OX40 signals, implying the dispensability of OX40 for generation of TCM cells. Nevertheless, survival of OX40-KO TEM cells was partially rescued in lymphopenic mice. During in vitro recall responses, the OX40-KO TEM cells that were generated in lymphopenic recipient mice showed impaired cytokine production, suggesting an essential role for OX40 not only on generation but also on effector function of CD4+ TEM cells. Collectively, the present results indicate differential requirements for OX40 signals on generation of CD4+ TEM and TCM cells.
Age Pensioner Profiles: A Longitudinal Study of Income, Assets and Decumulation

Using eight years of data drawn from the records of Australia’s Centrelink agency, we describe the income, asset and decumulation patterns of over 10,000 age pensioners. Analysis of this longitudinal data set shows that age pensioners, on average, preserve both financial and residential wealth, consuming conservatively and, ultimately, passing on substantial bequests. While younger households do run down financial wealth early in retirement, older households generally maintain their assessable asset balances, and some even manage to save. The largest falls in assets are linked to changes in household structure due to death or the breakdown of a relationship. So, as in many other developed countries, age pensioners in Australia appear to ‘under-consume’, holding on to assets, and even building a buffer, well into their later years.
The Joint Mortality of Couples in Continuous Time

This paper introduces a probabilistic framework for the joint survivorship of couples in the context of dynamic stochastic mortality models. The death of one member of a couple can have either deterministic or stochastic effects on the other; our new framework gives an intuitive and flexible pairwise cohort-based probabilistic mechanism that can account for both. It is sufficiently flexible to allow modelling of effects that are short-term (called the broken-heart effect) and/or long-term (named life circumstances bereavement). In addition, it can account for the state of health of both the surviving and the dying spouse and can allow for dynamic and asymmetric reactions of varying complexity. Finally, it can accommodate the pairwise dependence of mortality intensities before the first death. Analytical expressions for bivariate survivorship in representative models are given, and their sensitivity analysis is performed for benchmark cases of old and young couples. Simulation and estimation procedures are provided that are straightforward to implement and lead to consistent parameter estimation on synthetic dataset of 10000 pairs of death times for couples.
Global Variance Term Premia and Intermediary Risk Appetite

Sellers of variance swaps earn time-varying risk premia for their exposure to realized variance, the level of variance swap rates, and the slope of the variance swap curve. To measure risk premia, we estimate a dynamic term structure model that decomposes variance swap rates into expected variances and term premia. Empirically, we document a strong global factor structure in variance term premia across the U.S., U.K., Europe, and Japan. We further show that variance term premia are negatively correlated with the risk appetite of hedge funds, broker-dealers, and mutual funds. Our results support the hypothesis that financial intermediaries are marginal investors in the variance swap market.
Three-Dimensional Dosimetry for Radiation Safety Estimates from Intrathecal Administration

Intrathecal administration is of growing interest for drug delivery, and its utility is being increasingly investigated through imaging. In this work, the 3-dimensional Voxel-Based Internal Dosimetry Application (VIDA) and 4D Extended Cardiac Torso Phantom (XCAT) were extended to provide radiation safety estimates specific to intrathecal administration. Methods: The 3-dimensional VIDA dosimetry application Monte Carlo simulation was run using a modified XCAT phantom with additional and edited cerebrospinal fluid (CSF) regions to produce voxel-level absorbed dose per unit cumulated activity maps for 9 selected source regions. Simulation validation was performed to compare absorbed dose estimates for common organs in a preexisting dosimetry tool (OLINDA/EXM). Dynamic planar imaging data were acquired in 6 healthy subjects using administered volumes of 5 or 15 mL (n = 3 each) of 185 MBq of 99mTc-diethylenetriaminepentaacetic acid. Absorbed dose was estimated for each subject using the intrathecal-specific dosimetry application. Results: Simulation results were within 6% of OLINDA estimates for common organs. Absorbed dose estimates were highest (0.3–0.8 mGy/MBq) in the lumbar CSF space. A whole-body effective dose estimate of 0.003 mSv/MBq was observed. An administered volume dependency was observed with a 15-mL volume, resulting in lower absorbed dose estimates for several intrathecal and nonintrathecal regions. Conclusion: The intrathecal-specific VIDA implementation enables tailored dosimetry estimation for regions most relevant in intrathecal administration. Absorbed doses are highly localized to CSF and spinal regions and should be taken into consideration when designing intrathecal imaging studies. A potentially interesting relationship was observed between absorbed dose and administered volume, which merits further investigation.
Association of the −2510A/G chemokine (C–C motif) ligand 2 polymorphism with knee osteoarthritis in a Korean population

Objective: To investigate the possible association between polymorphisms [the −2510A/G promoter polymorphism (rs1024611) and the Cys35Cys coding polymorphism (rs4586) in exon 2] of the chemokine (C–C motif) ligand 2 (CCL2) gene and knee osteoarthritis (OA) in a Korean population. Methods: DNA was obtained from 153 Korean primary knee OA patients and 270 healthy controls. CCL2 genomic variants (−2510A/G and Cys35Cys polymorphisms) were detected by polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP). In additional, the effect of −2510A/G on CCL2 transcription was examined, using a luciferase reporter gene construct transfected into HMC‐1 cells. Results: The −2510A/G promoter polymorphism was associated with OA [genotype frequency, p = 0.041; allele frequency, p = 0.017, odds ratio (OR) = 1.45, 95% confidence interval (CI) = 1.07–1.96]. Significant association was observed between the G carrier of the −2510A/G promoter polymorphism and primary knee OA patients (p = 0.021, OR = 2.25, 95% CI = 1.12–4.52). The G carrier of the −2510A/G promoter polymorphism was also associated with both clinically subtyped OA patients (OA patients with functionally poor index and radiographically severe OA patients). However, no significant difference was found in the Cys35Cys polymorphism. Haplotype frequency analysis revealed a significant difference (χ2 = 8.98, p = 0.030). The CCL2 serum level of subjects with the G carrier (290.0±87.5 pg/mL) of the −2510A/G promoter polymorphism was statistically higher than that of subjects with the non‐G carrier (161.5±48.3 pg/mL). The luciferase activity was significantly greater from interleukin (IL)‐1β‐induced cells transfected with constructs containing G at position −2510. Conclusions: The G carrier of the −2510A/G promoter polymorphism was found to be associated with primary knee OA, and could be a susceptibility factor in the development of primary knee OA in the Korean population.