Memristive Crossbar Array-Based Probabilistic Graph Modeling.

Modern graph datasets with structural complexity and uncertainties due to incomplete information or data variability require advanced modeling techniques beyond conventional graph models. This study introduces a memristive crossbar array (CBA)-based probabilistic graph model (C-PGM) utilizing Cu0.3Te0.7/HfO2/Pt memristors, which exhibit probabilistic switching, self-rectifying, and memory characteristics. C-PGM addresses the complexities and uncertainties inherent in structural graph data across various domains, leveraging the probabilistic nature of memristors. C-PGM relies on the device-to-device variation across multiple memristive CBAs, overcoming the limitations of previous approaches that rely on sequential operations, which are slower and have a reliability concern due to repeated switching. This new approach enables the fast processing and massive implementation of probabilistic units at the expense of chip area. In this study, the hardware-based C-PGM feasibly expresses small-scale probabilistic graphs and shows minimal error in aggregate probability calculations. The probability calculation capabilities of C-PGM are applied to steady-state estimation and the PageRank algorithm, which is implemented on a simulated large-scale C-PGM. The C-PGM-based steady-state estimation and PageRank algorithm demonstrate comparable accuracy to conventional methods while significantly reducing computational costs.

Imbalance in the vWF - ADAMTS13 axis exists early in acute pancreatitis and predicts persistent organ failure and pancreatic necrosis-a prospective study.

BACKGROUND: The pathophysiology of Acute Pancreatitis (AP) may be complicated by endothelial activation. von Willebrand Factor (vWF)- ADAMTS13 axis is a marker of endothelial activation. The study aimed to investigate the axis in AP, comparing it in patients with and without persistent organ failure (OF), with and without pancreatic necrosis, and correlating it with the standard severity scores (CRP, APACHE II, BISAP, SOFA, and qSOFA) METHODS: vWF-Antigen (vWF:Ag), vWF-Collagen-Binding-Assay (vWF:CBA), and ADAMTS13 activity (ADAMTS13:act) levels were measured within 5 days of symptom onset in consecutive patients (n = 98), who were admitted with a first episode of AP (Dec 2021-May 2023). RESULTS: Of the 98 patients admitted with AP, 78(79.6 %) had no or transient OF; 20(20.4 %) had persistent OF. Age was comparable (43.73 ± 15.36 vs 38.65 ± 13.69) [mean ± SD](years), and males were predominant in both groups (70.5 % vs 80 %). Patientswith persistent OF had higher vWF:CBA(%)[323(279-486.5) vs 199.5(159.1-295.75)] and lower ADAMTS13:act(%)[35.4(23.8-56.85) vs 56.35(44.1-71.9)][median (25th - 75th percentile)](P = 0.001) than those with no or transient OF. Patients with pancreatic necrosis (n = 19) had lower ADAMTS13:act(%)[42.79 ± 18.69] than those without pancreatic necrosis (n = 18) [62.49 ± 22.64] (P < 0.01). ADAMTS13:act had a negative correlation(r = -0.2), whereas vWF:Ag and vWF:CBA had a positive correlation (r = 0.2) with the standard severity scores (P < 0.05). ADAMTS13:act could predict pancreatic necrosis [AUROC-0.737, P < 0.05] and persistent OF [AUROC-0.746, P < 0.001], while vWF:CBA could predict persistent OF [AUROC- 0.73, P < 0.001]. CONCLUSION: vWF-ADAMTS13 axis helps to predict severe disease and is associated with poor outcomes in acute pancreatitis.

The distinction of area postrema syndrome between MOGAD and NMOSD.

Background and objectives: Both myelin oligodendrocyte glycoprotein-IgG associated disorders (MOGAD) and neuromyelitis optica spectrum disorder (NMOSD) are demyelinating diseases of the central nervous system. They present similar clinical manifestations such as optica neuritis, myelitis and area postrema syndrome (APS). The distinctions of optica neuritis (ON) and myelitis between them have been elaborated to great length while their differences in APS remain to be elucidated. We aim to report the frequency of APS in patients with MOGAD as well as NNOSD patients, and to compare the characteristics of APS between patients with MOGAD and those with NMOSD. Methods: Seven MOG-IgG positive APS patients were retrospectively identified between 2017 and 2022. APS phenotypes have been previously described. The similarities and differences between MOGAD and NMOSD patients with APS was compared, including the frequency and duration of APS between the two diseases, and their incidences of accompanied subtentorial lesions have also been described and compared. Results: We reviewed a cohort of 218 MOG-IgG-positive patients, and 396 patients with NMOSD. 200 MOGAD patients and 332 NMOSD patients were included in this study. In the cohort, seven patients with MOG-IgG-positive antibody presented with APS were analyzed, four of whom had disease onset with APS. Of the 332 patients with NMOSD, 47 had APS attacks while 31 had APS at disease onset. In patients with MOGAD, 2 had nausea, 3 had vomiting, 5 had hiccups, and 1 patient presented with all three symptoms above. In patients with NMOSD, 70.2 % had nausea, vomiting and hiccups at the same time during APS attacks. Apart from the medulla oblongata, other subtentorial regions were also affected in 6/7 MOGAD patients while 14/47 NMOSD patients had other subtentorial regions involved. During an APS attack, the incidence of concomitant lesions in the brainstem and other regions was significantly greater in MOGAD than in the NMOSD cohort (P = 0.008*). Conclusion: APS is a rare, but not isolated clinical manifestation of MOGAD. APS happened more frequently with other supratentorial and subtentorial lesions in MOGAD. The symptoms of NVH (nausea, vomiting, hiccups) tended to happen respectively in MOGAD compared with NMOSD. The phenotype or mechanism of APS in MOGAD may differ from that in NMOSD.

The Use of Ground Coal Bottom Ash/Slag as a Cement Replacement for Sustainable Concrete Infrastructure.

Cement production requires considerable energy and natural resources, severely impacting the environment due to harmful gas emissions. Coal bottom ash (CBA) and coal boiler slag (CBS), byproducts of coal-fired powerplants having pozzolanic properties, can be mechanically ground and replace cement in concrete, which reduces waste in landfills, preserves natural resources, and reduces health hazards. This study was performed to determine the optimum cement replacement amount of ground CBA (GCBA) and ground CBS (GCBS) in concrete, which was 10% for GCBA and 5% for GCBS. GCBA-based concrete exhibited superior tensile strength, modulus of elasticity, and durability compared to the control. In the Rapid Chloride Penetration Test, 10% GCBA concrete resulted in 2026 coulombs at 56 days, compared to 3405 coulombs for the control, indicating more resistance to chloride penetration. Incorporating 2.5% nanoclay in GCBA-based concrete increased the optimum GCBA content by 5%, and the compressive strength of 15% GCBA concrete increased by 4 MPa. The mortar consisting of the finest GCBA(L1) having Blaine fineness of 3072 g/cm2 yielded the highest compressive strength (32.7 MPa). The study discovered that the compressive strength of GCBA and GCBS-based mortars increases with fineness, and meeting the recommended fineness limit in ASTM C618 enhances concrete or mortar properties.

Rheumatoid arthritis autologous synovial fluid affects the plasticity and function of peripheral and induced T regulatory cells in vitro.

The synovial fluid (SF) microenvironment in rheumatoid arthritis (RA) may alter the stability and function of Tregs. In the present study, we assessed cytokine levels and percentage of Tregs, Tregs expressing CXCR3 (Th1-like Treg), CCR6 (Th17-like Treg) in RA peripheral blood (PB) and RA-SF using fluorescence cytometry. Effect of autologous SF on plasticity and function of RA-PB Tregs (pTregs; CD4+CD25hiCD127Lo/-) and induced vimentin-pulsed Tregs (iTregsVIM) was assessed in vitro. Cytokines and percentage of Th1-like and Th17-like Tregs were higher in RA-PB than OA-PB; higher in RA-SF than osteoarthritis (OA)-SF. Compared to OA-SF exposed OA-pTregs, RA-SF exposed RA-pTregs showed higher percentage of Th1-like (11% vs 20%) and Th17-like (16% vs 36%) Tregs; higher T-bet (p = 0.0001), RORγ (p = 0.0001) and lower FOXP3 (p = 0.0001) gene expression; and diminished percentage suppression of autologous T effector cells (36% vs 74%). RA-SF exposed iTregsVIM showed increased percentage of Th1-like and Th17-like Tregs compared to iTregsVIM exposed to AB serum (8% vs 0.1%; 21% vs 0.1%). IL-2, Tocilizumab and 5-azacytidine reduced the conversion of iTregsVIM (8% vs 2.4%; 21% vs 6.9%), when used in combination. To conclude, microenvironment in the RA synovial fluid is possibly responsible for conversion of pTregs into Th-like Tregs and their functional loss. A blockade of cytokine receptors and methyl transferases could inhibit Tregs conversion, providing clinical relevance for future Tregs targeting therapies.

Cell-Based Assay to Detect the Autoantibody Serostatus in Patients with Neuromyelitis Optica Spectrum Disorder (NMOSD).

Cell-based assay (CBA) is an immunofluorescence assay that is extensively used for the confirmatory diagnosis of inflammatory demyelinating diseases of the central nervous system, like neuromyelitis optica spectrum disorder (NMOSD). Detecting the type of autoantibody present in the sera of the patients is the primary goal. CBA is the most sensitive and recommended detection method among all similar tools. Briefly, serum autoantibody is screened by transfecting specific cells seeded on cover glasses with full-length specific antigen fused with green fluorescent protein (GFP), followed by treating them with the patient serum used here as the source of primary antibody. The autoantibody-treated cells are further labeled with a rhodamine-conjugated secondary antibody. The co-localization of GFP and rhodamine is visualized by confocal microscopy, and the intensity of fluorescence is evaluated to determine the presence of autoantibody. A detailed protocol to screen antibodies against AQP4 and MOG in human sera using this method is described.

Cell-based vs enzyme-linked immunosorbent assay for detection of anti-Tribbles homolog 2 autoantibodies in Chinese patients with narcolepsy.

STUDY OBJECTIVES: Narcolepsy type 1 is attributed to a deficiency in cerebrospinal fluid orexin and is considered linked to autoimmunity. The levels of anti-Tribbles homolog 2 (TRIB2) autoantibodies are elevated in the sera of some patients with narcolepsy with cataplexy. Additionally, injecting mice with serum immunoglobulin from patients with narcolepsy with positive anti-TRIB2 antibodies can induce hypothalamic neuron loss and alterations in sleep patterns. Consequently, we hypothesized the existence of a potential association between anti-TRIB2 antibodies and narcolepsy. To test this possibility, we used cell-based assays (CBAs) and enzyme-linked immunosorbent assays (ELISAs) to detect the presence of anti-TRIB2 antibodies in Chinese patients with narcolepsy. METHODS: We included 68 patients with narcolepsy type 1, 39 patients with other central disorders of hypersomnolence, and 43 healthy controls. A CBA and a conventional ELISA were used to detect anti-TRIB2 antibody levels in patients' sera. RESULTS: CBA was used to detect serum anti-TRIB2 antibodies in Chinese patients with narcolepsy, and the results were negative. However, when the ELISA was used, only 2 patients with narcolepsy type 1 had TRIB2 antibody titers higher than the mean titer plus 2 standard deviations of the healthy controls. CONCLUSIONS: In our study, ELISA identified TRIB2 autoantibodies in sera of patients with narcolepsy where CBA failed to demonstrate them. Contrary to our hypothesis, this intriguing finding deserves further research to elucidate the potential association between TRIB2 and narcolepsy type 1. Exploring the implications of TRIB2 autoantibodies in narcolepsy and disparate outcomes between ELISA and CBA could provide crucial insights. CITATION: Zhong X, Yuan Y, Zhan Q, et al. Cell-based vs enzyme-linked immunosorbent assay for detection of anti-Tribbles homolog 2 autoantibodies in Chinese patients with narcolepsy. J Clin Sleep Med. 2024;20(6):941-946.

Sulforaphane effectively inhibits HBV by altering Treg/Th17 immune balance and the MIF-macrophages polarizing axis in vitro and in vivo.

BACKGROUND: Hepatitis B virus (HBV) infection is a major public health problem. After HBV infection, viral antigens shift the immune balance in favor of viral escape. Sulforaphane (SFN) is a traditional Chinese medicine.It regulates multi-biological activities, including anti-inflammation, anticancer, and antiviral. However, few studies reported that SFN can inhibit HBV infection before. METHODS: An immunocompetent HBV CBA/CaJ mouse model and a co-culture model were used to explore the effect of SFN on HBV and whether SFN altered the immune balance after HBV infection. RESULTS: We found that SFN was able to reduce HBV DNA, cccDNA, HBsAg, HBeAg, and HBcAg levels in serum and liver tissues of HBV-infected mice. In vitro and in vivo experiments showed that SFN could significantly increase the expression of Cd86 and iNOS and inhibit the expression of Arg1 on macrophages after HBV infection. After SFN administration, Th17 markers in liver tissue and serum were significantly increased. There was no significant changes in the proportion of Treg cells in peripheral blood, but a significant increase in the proportion of Th17 cells and decrease of the Treg/Th17 ratio. Using a network pharmacology approach, we predicted macrophage migration inhibitory factor (MIF) as a potential target of SFN and further validated that MIF expression was significantly increased after HBV infection and SFN significantly inhibited MIF expression both in vitro and in vivo. There was an upward trend in HBV markers (p>0.05) after MIF overexpression. Overexpression of MIF combined with the use of SFN resulted in a significant reversion in the expression of HBV markers and polarization of macrophages towards the M1 phenotype. CONCLUSION: Our results indicated that immunocompetent HBV CBA/CaJ mouse model is a good model to evaluate HBV infection. SFN could inhibit the expression of HBV markers, promote polarization of macrophages towards the M1 phenotype after HBV infection, change the proportion of Treg and Th17 cells. Our findings demonstrate that SFN inhibit HBV infection by inhibiting the expression of MIF and promoting the polarization of macrophages towards the M1 phenotype, which illustrates a promising therapeutic approach in HBV infection.

Establishment and visual analysis of CBA/J-Pde6bY347Y/Y347X and C3H/HeJ-Pde6bY347Y/Y347X mice.

In CBA/J and C3H/HeJ mice, retinitis pigmentosa is inherited as an autosomal-recessive trait due to a mutation in Pde6b, which encodes cGMP phosphodiesterase subunit b. In these strains, the Y347X mutation in Pde6b leads to the upregulation of cGMP levels, increased Ca2+ influx induces rod death, and the outer segment and rod cells entirely disappeared by 35 days after birth. In the present study, we utilized the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) 9-mediated gene editing to repair the Y347X mutation in CBA/J and C3H/HeJ mice. Evaluation of the established CBA/J-Pde6bY347Y/Y347X and C3H/HeJ-Pde6bY347Y/Y347X mice, which were confirmed to have normal retinal layers by live fundoscopic imaging and histopathological analysis, revealed improved visual acuity based on the visual cliff and light/dark latency tests. Furthermore, our analyses revealed that the visible platform test was a more effective tool for testing visual behavior in these mice. The results suggest that the established strains can serve as control groups for CBA/J and C3H/HeJ in ophthalmology studies involving retinitis pigmentosa.

Age-related changes of auditory sensitivity across the life span of CBA/CaJ mice.

The inbred mouse strain CBA/CaJ is a frequently used animal model of age-related hearing loss in humans. These mice display significant hearing loss at a relatively advanced age, similar to most humans, with progressive loss of hearing as the mouse continues to age. While important descriptions of hearing loss in this mouse strain at multiple ages have previously been published, shortcomings persist in the data for hearing over the lifespan of the mouse. Therefore, we analyzed auditory brainstem response threshold data from records maintained by our research group to yield an extensive database of thresholds over nearly the entire life span of the CBA/CaJ mouse (from 79 to 1085 days). Data was collected from in-house bred mice of CBA/CaJ stock, initially from The Jackson Laboratory. Data was collected using BiosigRZ software and TDT System III hardware. Thresholds were routinely measured in conjunction with behavioral and electrophysiological experiments; only responses from baseline or experimentally naïve animals were analyzed. The resulting data set comprised 376 female mice and 441 males. At the lowest and highest frequencies (8 & 32 kHz), initial thresholds were just under 30 dB SPL and increased slowly until they were significantly different at 16-18 months compared to 1-3 months age, with the difference increasing over subsequent ages. At the middle frequencies (12 & 16 kHz), initial thresholds were just under 20 dB SPL and increased until they became different from initial at 16-18 months. At 24 kHz, initial thresholds were just above 20 dB and became different from initial at 13-16 months of age. The rate of change of thresholds with age were similar for all frequencies until about 30 months of age, when 32 kHz threshold changes lagged behind other frequencies. Generally, CBA/CaJ mice in our colony display relatively low thresholds until approximately 16 months of age, depending on frequency. After 16-18 months, thresholds become significantly worse. After approximately 20-22 months thresholds increase linearly with age.

A comprehensive approach to developing a multi-epitope vaccine against Mycobacterium tuberculosis: from in silico design to in vitro immunization evaluation.

Introduction: The Bacillus Calmette-Guérin (BCG) vaccine, currently used against tuberculosis (TB), exhibits inconsistent efficacy, highlighting the need for more potent TB vaccines. Materials and methods: In this study, we employed reverse vaccinology techniques to develop a promising multi-epitope vaccine (MEV) candidate, called PP13138R, for TB prevention. PP13138R comprises 34 epitopes, including B-cell, cytotoxic T lymphocyte, and helper T lymphocyte epitopes. Using bioinformatics and immunoinformatics tools, we assessed the physicochemical properties, structural features, and immunological characteristics of PP13138R. Results: The vaccine candidate demonstrated excellent antigenicity, immunogenicity, and solubility without any signs of toxicity or sensitization. In silico analyses revealed that PP13138R interacts strongly with Toll-like receptor 2 and 4, stimulating innate and adaptive immune cells to produce abundant antigen-specific antibodies and cytokines. In vitro experiments further supported the efficacy of PP13138R by significantly increasing the population of IFN-γ+ T lymphocytes and the production of IFN-γ, TNF-α, IL-6, and IL-10 cytokines in active tuberculosis patients, latent tuberculosis infection individuals, and healthy controls, revealing the immunological characteristics and compare the immune responses elicited by the PP13138R vaccine across different stages of Mycobacterium tuberculosis infection. Conclusion: These findings highlight the potential of PP13138R as a promising MEV candidate, characterized by favorable antigenicity, immunogenicity, and solubility, without any toxicity or sensitization.

Twenty-four-month interim results from a prospective, single-arm clinical trial evaluating the performance and safety of cellular bone allograft in patients undergoing lumbar spinal fusion.

BACKGROUND: Autologous bone grafts are the gold standard for spinal fusion; however, harvesting autologous bone can result in donor site infection, hematomas, increased operative time, and prolonged pain. Cellular bone allografts (CBAs) are a viable alternative that avoids the need for bone harvesting and may increase fusion success alone or when used as an adjunct material. The present study examined the efficacy and safety of CBA when used as an adjunct graft material to lumbar arthrodesis. METHODS: A prospective, single-arm, multicenter clinical trial (NCT02969616) was conducted in adult subjects (> 18 years of age) undergoing lumbar spinal fusion with CBA graft (CBA used as primary (≥ 50% by volume), with augmentation up to 50%). Radiographic fusion status was assessed by an independent review of dynamic radiographs and CT scans. Clinical outcomes were assessed with the Oswestry Disability Index (ODI), and Visual Analog Scales (VAS) score for back and leg pain. Adverse events were assessed through the 24-month follow-up period. The presented data represents an analysis of available subjects (n = 86) who completed 24 months of postoperative follow-up at the time the data was locked for analysis. RESULTS: Postoperative 24-month fusion success was achieved in 95.3% of subjects (n = 82/86) undergoing lumbar spinal surgery. Clinical outcomes showed statistically significant improvements in ODI (46.3% improvement), VAS-Back pain (75.5% improvement), and VAS-Leg pain (85.5% improvement) (p < 0.01) scores at Month 24. No subject characteristics or surgical factors were associated with pseudoarthrosis. A favorable safety profile with a limited number of adverse events was observed. CONCLUSIONS: The use of CBA as an adjunct graft material showed high rates of successful lumbar arthrodesis and significant improvements in pain and disability scores. CBA provides an alternative to autograft with comparable fusion success rates and clinical benefits. TRIAL REGISTRATION: NCT02969616.

Evaluation of Toxicity Equivalency Factors of Tetrodotoxin Analogues with a Neuro-2a Cell-Based Assay and Application to Puffer Fish from Greece.

Tetrodotoxin (TTX) is a potent marine neurotoxin involved in poisoning cases, especially through the consumption of puffer fish. Knowledge of the toxicity equivalency factors (TEFs) of TTX analogues is crucial in monitoring programs to estimate the toxicity of samples analyzed with instrumental analysis methods. In this work, TTX analogues were isolated from the liver of a Lagocephalus sceleratus individual caught on South Crete coasts. A cell-based assay (CBA) for TTXs was optimized and applied to the establishment of the TEFs of 5,11-dideoxyTTX, 11-norTTX-6(S)-ol, 11-deoxyTTX and 5,6,11-trideoxyTTX. Results showed that all TTX analogues were less toxic than the parent TTX, their TEFs being in the range of 0.75-0.011. Then, different tissues of three Lagocephalus sceleratus individuals were analyzed with CBA and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The obtained TEFs were applied to the TTX analogues' concentrations obtained by LC-MS/MS analysis, providing an indication of the overall toxicity of the sample. Information about the TEFs of TTX analogues is valuable for food safety control, allowing the estimation of the risk of fish products to consumers.

Effect of growth temperature on self-rectifying BaTiO3/ZnO heterojunction for high-density crossbar arrays and neuromorphic computing.

In the quest for high-density integration and massive scalability, ferroelectric-based devices provide an achievable approach for nonvolatile crossbar array (CBA) architecture and neuromorphic computing. In this report, ferroelectric-semiconductor (Pt/BaTiO3/ZnO/Au) heterojunction-based devices are demonstrated to exhibit nonvolatile and synaptic characteristics. In this study, the ferroelectric (BaTiO3) layer was modulated at various growth temperatures of 350 °C, 450 °C, 550 °C and 650 °C. Growing temperature in the ferroelectric layer has a significant impact on resistive switching. The ferroelectricity of the BaTiO3 thin film enhanced by increasing temperature causes a substantial shift in the interface state density at heterojunction interface, which is crucial for self-rectification. Furthermore, this self-rectifying property advances to reduce the crosstalk problem without any selector device. Enhanced resistive switching and neuromorphic applications have been demonstrated using BaTiO3 heterostructure devices at 550 °C. The dynamic ferroelectric polarization switching in this heterojunction demonstrated linear conductance change in artificial synapses with 91 % recognition accuracy. Ferroelectric polarization reversal with a depletion region at the heterojunction interface is the responsible mechanism for the switching in these devices. Thus, these findings pave the way for designing low power high-density crossbar arrays and neuromorphic application based on ferroelectric-semiconductor heterostructures.

High level of agreement in a fixed vs. live cell-based assay for antibodies to myelin oligodendrocyte glycoprotein in a real-world clinical laboratory setting.

Introduction: As recognition of myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease becomes more widespread, the importance of appropriately ordering and interpreting diagnostic testing for this antibody increases. Several assays are commercially available for MOG testing, and based on a few small studies with very few discrepant results, some have suggested that live cell-based assays (CBA) are superior to fixed CBA for clinical MOG antibody testing. We aimed to determine the real-world agreement between a fixed and live CBA for MOG using two of the most commonly available commercial testing platforms. Methods: We compared paired clinical samples tested at two national clinical reference laboratories and determined the real-world agreement between the fixed CBA and live CBA. Results: Of 322 paired samples tested on both platforms, 53 were positive and 246 were negative by both methodologies (agreement 92.9%, Cohen's kappa 0.78, [0.69-0.86]). Spearman correlation coefficient was 0.80 (p < 0.0001). Of the discrepant results, only 1 of 14 results positive by the live CBA had a titer greater than 1:100, and only 1 of 9 results positive by the fixed CBA had a titer of greater than 1:80. Lower titers on the fixed CBA correlate to higher titers on the live CBA. Conclusion: Overall, there is excellent agreement between fixed and live CBA for MOG antibody testing in a real-world clinical laboratory setting. Clinicians should be aware of which method they use to assess any given patient, as titers are comparable, but not identical between the assays.

Detection of Antibodies against the Acetylcholine Receptor in Patients with Myasthenia Gravis: A Comparison of Two Enzyme Immunoassays and a Fixed Cell-Based Assay.

The detection of serum anti-acetylcholine receptor (AChR) antibodies is currently an important tool for diagnosing myasthenia gravis (MG) since they are present in about 85% of MG patients. Many serological tests are now available. Nevertheless, results from these tests can be different in some patients. The aim of this study is to compare the sensitivity of a commercially available fixed cell-based assay (F-CBA) to that of enzyme-linked immunosorbent assay (ELISA) kits for anti-AChR detection in patients with a diagnosis of MG. Overall, 143 patients with a confirmed MG diagnosis were included in the study. The detection and measurement of serum anti-AChR antibodies were performed by three analytical methods, namely, a competitive ELISA (cELISA), an indirect ELISA (iELISA), and an F-CBA, according to the manufacturers' instructions. Anti-AChR antibody titers were positive in 94/143 (66%) using the cELISA, in 75/143 (52%) using the iELISA and in 61/143 (43%) using the F-CBA (adult and/or fetal). Method agreement, evaluated by concordant pairs and Cohen's kappa, was as follows: cELISA-iELISA: 110/143 (77%), k = 0.53 (95%CI 0.40-0.66); cELISA-F-CBA: 108/143 (76%), k = 0.53 (95%CI 0.41-0.66); iELISA-F-CBA: 121/143 (85%), k = 0.70 (95%CI 0.57-0.80). Our findings show that the cELISA has better analytical performance than the iELISA and F-CBA. However, the iELISA and F-CBA show the highest concordance.

Photon- and Singlet-Oxygen-Induced Cis-Trans Isomerization of the Water-Soluble Carotenoid Crocin.

Studying the cis-trans isomerization process in crocin (CR), one of the few water-soluble carotenoids extracted from saffron, is important to better understand the physiological role of cis-carotenoids in vivo and their potential as antioxidants in therapeutic applications. For that, cis-trans isomerization of both methanol- and water-dissolved CR was induced by light or thermally generated singlet oxygen (1O2). The kinetics of molecular concentrations were monitored by both high-performance liquid chromatography (HPLC) and non-destructive spectrophotometric methods. These last made it possible to simultaneously follow the cis-trans isomerization, the possible bleaching of compounds and the amount of thermally generated 1O2. Our results were in accordance with a comprehensive model where the cis-trans isomerization occurs as relaxation from the triplet state of all-trans- or 13-cis-CR, whatever is the way to populate the CR triplet state, either by photon or 1O2 energy transfer. The process is much more (1.9 to 10-fold) efficient from cis to trans than vice versa. In H2O, a 1O2-induced bleaching effect on the starting CR was not negligible. However, the CR "flip-flop" isomerization reaction could still occur, suggesting that this process can represent an efficient mechanism for quenching of reactive oxygen species (ROS) in vivo, with a limited need of carotenoid regeneration.

Biopharmaceutics Risk Assessment-Connecting Critical Bioavailability Attributes with In Vitro, In Vivo Properties and Physiologically Based Biopharmaceutics Modeling to Enable Generic Regulatory Submissions.

Quality risk assessment following ICH Q9 principles is an important activity to ensure optimal clinical efficacy and safety of a drug product. Typically, risk assessment is focused on product performance wherein critical material attributes, formulation variables, and process parameters are evaluated from a manufacturing perspective. Extending ICH Q9 principles to biopharmaceutics risk assessment to identify factors that can impact in vivo performance is an upcoming area. This is evident by recent regulatory trends wherein a new term critical bioavailability attributes (CBA) has been coined to identify such factors. Although significant work has been performed for biopharmaceutics risk assessment for new molecules, there is a need for harmonized biopharmaceutics risk assessment workflow for generic submissions. In this manuscript, we attempted to provide a framework for performing biopharmaceutics risk assessment for generic regulatory submissions. A detailed workflow for performing biopharmaceutics risk assessment includes identification of initial CBA (iCBA), their confirmatory evaluation followed by definition of the control strategy. Tools for biopharmaceutics risk assessment, i.e., bio-discriminatory dissolution method and physiologically based biopharmaceutics modeling (PBBM) were discussed from a practical perspective. Furthermore, a case study for CBA evaluation using PBBM modeling for an extended-release product for regulatory submission has been described using the proposed workflow. Finally, future directions of integrating CBA evaluation, biopharmaceutics risk assessment to the FDA Knowledge Aided Structured Assessment (KASA) initiative, the necessity of risk assessment templates, and knowledge sharing between industry and academia are discussed. Overall, the work described in this manuscript can facilitate and provide guidance for biopharmaceutics risk assessment for generic submissions.

Exposing new taxonomic variation with inflammation - a murine model-specific genome database for gut microbiome researchers.

BACKGROUND: The murine CBA/J mouse model widely supports immunology and enteric pathogen research. This model has illuminated Salmonella interactions with the gut microbiome since pathogen proliferation does not require disruptive pretreatment of the native microbiota, nor does it become systemic, thereby representing an analog to gastroenteritis disease progression in humans. Despite the value to broad research communities, microbiota in CBA/J mice are not represented in current murine microbiome genome catalogs. RESULTS: Here we present the first microbial and viral genomic catalog of the CBA/J murine gut microbiome. Using fecal microbial communities from untreated and Salmonella-infected, highly inflamed mice, we performed genomic reconstruction to determine the impacts on gut microbiome membership and functional potential. From high depth whole community sequencing (~ 42.4 Gbps/sample), we reconstructed 2281 bacterial and 4516 viral draft genomes. Salmonella challenge significantly altered gut membership in CBA/J mice, revealing 30 genera and 98 species that were conditionally rare and unsampled in non-inflamed mice. Additionally, inflamed communities were depleted in microbial genes that modulate host anti-inflammatory pathways and enriched in genes for respiratory energy generation. Our findings suggest decreases in butyrate concentrations during Salmonella infection corresponded to reductions in the relative abundance in members of the Alistipes. Strain-level comparison of CBA/J microbial genomes to prominent murine gut microbiome databases identified newly sampled lineages in this resource, while comparisons to human gut microbiomes extended the host relevance of dominant CBA/J inflammation-resistant strains. CONCLUSIONS: This CBA/J microbiome database provides the first genomic sampling of relevant, uncultivated microorganisms within the gut from this widely used laboratory model. Using this resource, we curated a functional, strain-resolved view on how Salmonella remodels intact murine gut communities, advancing pathobiome understanding beyond inferences from prior amplicon-based approaches. Salmonella-induced inflammation suppressed Alistipes and other dominant members, while rarer commensals like Lactobacillus and Enterococcus endure. The rare and novel species sampled across this inflammation gradient advance the utility of this microbiome resource to benefit the broad research needs of the CBA/J scientific community, and those using murine models for understanding the impact of inflammation on the gut microbiome more generally. Video Abstract.

Algorithm to improve the diagnosis of paraneoplastic neurological syndromes associated with SOX1 antibodies.

SOX1 antibodies (SOX1-abs) are associated with paraneoplastic neurological syndromes (PNS) and small cell lung cancer (SCLC). In many clinical laboratories SOX1-abs are determined by commercial line blots without confirmation by cell-based assay (CBA) with HEK293 cells expressing SOX1. However, the diagnostic yield of commercial line blots is low and the accessibility to the CBA, that is not commercially available, limited. Here, we evaluated if the addition of the band intensity data of the line blot and the immunoreactivity in a tissue-based assay (TBA) improve the diagnostic performance of the line blot. We examined serum of 34 consecutive patients with adequate clinical information that tested positive for SOX1-abs in a commercial line blot. Samples were also assessed by TBA and CBA. SOX1-abs were confirmed by CBA in 17 (50%) patients, all (100%) had lung cancer (SCLC in 16) and 15/17 (88%) had a PNS. In the remaining 17 patients the CBA was negative and none had PNS associated with lung cancer. TBA was assessable in 30/34 patients and SOX1-abs reactivity was detected in 15/17 (88%) with positive and in 0/13 (0%) with negative CBA. Only 2 (13%) of the 15 TBA-negative patients were CBA-positive. The frequency of TBA-negative but CBA-positive increased from 10% (1/10) when the band intensity of the line blot was weak to 20% (1/5) in patients with a moderate or strong intensity band. Confirmation by CBA should be mandatory for samples (56% in this series) not assessable (4/34; 12%) or negative in the TBA (15/34; 44%).