Miracle-dz/UniFAR-FTUs huggingface dataset:
A Mast Cell–Specific Receptor Is Critical for Granuloma Induced by Intrathecal Morphine Infusion

Key Points A mouse model of granuloma induced by intrathecal morphine was established. Mast cell–specific receptor MrgprB2 is essential for granuloma pathogenesis. Activation of MrgprB2 in mast cells promote granuloma-related cell recruitment. Visual Abstract Intrathecal morphine infusion is often applied to treat chronic pain related to cancer and other conditions. However, persistent pain can be caused by nerve compression because of granuloma formation. In this study, a mouse model of morphine-induced granuloma formation by intrathecal catheterization morphine infusion into the atlanto-occipital membrane of the foramen magnum was established in wild-type mice, MrgprB2 mutant (MrgprB2−/−) mice, and in mast cell–deficient W-sash c-kit mutant (KitW-sh/W-sh) mice. Heat-related pain after surgery was performed to investigate the antipain effect of morphine. H&E staining and immunofluorescence staining of the spinal cord were applied to analyze the mechanism of granuloma formation. Morphine-induced mast cell degranulation was assessed by measuring the Ca2+ influx and mediator release. Anaphylactoid reactions were measured after s.c. morphine infusion to the paws. Chemokine release by mast cells was determined by Human XL Cytokine Array. Experiments with wild-type, MrgprB2 mutant, and mast cell–deficient W-sash c-kit mutant mice demonstrated that morphine activated mast cells and inflammatory cell aggregation through MrgprB2 in intrathecal infusion sites. The chemokine production of human mast cells demonstrated that granuloma formation is correlated with chemokines release. In addition, morphine activated mouse primary mast cells and de novo chemokine synthesis via the MRGPRX2 in human LAD2 cells. We concluded that granuloma formation during intrathecal morphine infusion was associated with MrgprB2/X2. Reducing MRGPRX2 potentially blocks morphine-induced side effects, including granuloma formation.
Lipid Bilayer Thickness Measured by Quantitative DIC Reveals Phase Transitions and Effects of Substrate Hydrophilicity

Quantitative differential interference contrast microscopy is demonstrated here as a label-free method, which is able to image and measure the thickness of lipid bilayers with 0.1 nm precision. We investigate the influence of the substrate on the thickness of fluid-phase 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)-supported lipid bilayers and find a thinning of up to 10%, depending on substrate hydrophilicity, local bilayer coverage, and ionic strength of the medium. With fluorescently labeled lipid bilayers, we also observe changes in the bilayer thickness depending on the choice of fluorophore. Furthermore, liquid-ordered domains in bilayers, formed from DOPC, cholesterol, and sphingomyelin, are measured, and the corresponding thickness change between the liquid-ordered and liquid-disordered phases is accurately determined. Again, the thickness difference is found to be dependent on the presence of the fluorophore label, highlighting the need for quantitative label-free techniques.
Early Stage Glycosylation Biomarkers in Alzheimer’s Disease

Alzheimer’s disease (AD) is of great cause for concern in our ageing population, which currently lacks diagnostic tools to permit accurate and timely diagnosis for affected individuals. The development of such tools could enable therapeutic interventions earlier in the disease course and thus potentially reducing the debilitating effects of AD. Glycosylation is a common, and important, post translational modification of proteins implicated in a host of disease states resulting in a complex array of glycans being incorporated into biomolecules. Recent investigations of glycan profiles, in a wide range of conditions, has been made possible due to technological advances in the field enabling accurate glycoanalyses. Amyloid beta (Aβ) peptides, tau protein, and other important proteins involved in AD pathogenesis, have altered glycosylation profiles. Crucially, these abnormalities present early in the disease state, are present in the peripheral blood, and help to distinguish AD from other dementias. This review describes the aberrant glycome in AD, focusing on proteins implicated in development and progression, and elucidates the potential of glycome aberrations as early stage biomarkers of AD.
Glutathione Conjugation and Protein Adduction Derived from Oxidative Debromination of Benzbromarone in Mice

Benzbromarone (BBR), a uricosuric agent, has been known to induce hepatotoxicity, and its toxicity has a close relation to cytochrome P450–mediated metabolic activation. An oxidative debromination metabolite of BBR has been reported in microsomal incubations. The present study attempted to define the oxidative debromination pathway of BBR in vivo. One urinary mercapturic acid (M1) and one glutathione (GSH) conjugate (M2) derived from the oxidative debromination metabolite were detected in BBR-treated mice after solid phase extraction. M1 and M2 shared the same chromatographic behavior and mass spectral identities as those detected in N-acetylcysteine/GSH- and BBR-fortified microsomal incubations. The structure of M1 was characterized by chemical synthesis, along with mass spectrometry analysis. In addition, hepatic protein modification that occurs at cysteine residues (M′3) was observed in mice given BBR. The observed protein adduction reached its peak 4 hours after administration and occurred in a dose-dependent manner. A GSH conjugate derived from oxidative debromination of BBR was detected in livers of mice treated with BBR, and the formation of the GSH conjugate apparently took place earlier than the protein adduction. In summary, our in vivo work provided strong evidence for the proposed oxidative debromination pathway of BBR, which facilitates the understanding of the mechanisms of BBR-induced hepatotoxicity. SIGNIFICANCE STATEMENT This study investigated the oxidative debromination pathway of benzbromarone (BBR) in vivo. One urinary mercapturic acid (M1) and one glutathione (GSH) conjugate (M2) derived from the oxidative debromination metabolite were detected in BBR-treated mice. M1 and M2 were also observed in microsomal incubations. The structure of M1 was characterized by chemical synthesis followed by mass spectrometry analyses. More importantly, protein adduction derived from oxidative debromination of BBR (M′3) was observed in mice given BBR, and occurred in dose- and time-dependent manners. The success in detection of GSH conjugate, urinary N-acetylcysteine conjugate, and hepatic protein adduction in mice given BBR provided solid evidence for in vivo oxidative debromination of BBR. The studies allowed a better understanding of the metabolic activation of BBR.
Functional Characterization of Pharmcogenetic Variants of Human Cytochrome P450 2C9 in Korean Populations

Human cytochrome P450 2C9 is a highly polymorphic enzyme that is required for drug and xenobiotic metabolism. Here, we studied eleven P450 2C9 genetic variants—including three novel variants F69S, L310V, and Q324X—that were clinically identified in Korean patients. P450 2C9 variant enzymes were expressed in Escherichia coli and their bicistronic membrane fractions were prepared The CO-binding spectra were obtained for nine enzyme variants, indicating P450 holoenzymes, but not for the M02 (L90P) variant. The M11 (Q324X) variant could not be expressed due to an early nonsense mutation. LC-MS/MS analysis was performed to measure the catalytic activities of the P450 2C9 variants, using diclofenac as a substrate. Steady-state kinetic analysis revealed that the catalytic efficiency of all nine P450 2C9 variants was lower than that of the wild type P450 2C9 enzyme. The M05 (R150L) and M06 (P279T) variants showed high kcat values; however, their Km values were also high. As the M01 (F69S), M03 (R124Q), M04 (R125H), M08 (I359L), M09 (I359T), and M10 (A477T) variants exhibited higher Km and lower kcat values than that of the wild type enzyme, their catalytic efficiency decreased by approximately 50-fold compared to the wild type enzyme. Furthermore, the novel variant M07 (L310V) showed lower kcat and Km values than the wild type enzyme, which resulted in its decreased (80%) catalytic efficiency. The X-ray crystal structure of P450 2C9 revealed the presence of mutations in the residues surrounding the substrate-binding cavity. Functional characterization of these genetic variants can help understand the pharmacogenetic outcomes.
Efficacy of Arabica Versus Robusta Coffee in Improving Weight, Insulin Resistance, and Liver Steatosis in a Rat Model of Type-2 Diabetes

The effects of chronic coffee exposure in models of type 2 diabetes mellitus (T2D) is scarcely studied, and the efficacy of the main coffee species has never been compared. We tested the hypothesis that long-term consumption of arabica and robusta coffee may differentially delay and affect T2D development in Zucker diabetic fatty rats. Three study groups received either chow mixed with arabica or robusta instant coffee (1.8% w/w) or unsupplemented chow food for 10 weeks. Both coffee species reduced liver triglyceride content and area under the curve of fasting and postprandial insulin. At study end, plasma adiponectin, total cholesterol and high density lipoprotein levels were higher in the robust group compared with both arabica and control groups. The liver gene expression of Glucose-6-phosphatase, catalytic subunit (G6pc) and Mechanistic target of rapamycin (mTOR) in robusta and Cpt1a in both coffee groups was downregulated. In conclusion, long-term consumption of both coffee species reduced weight gain and liver steatosis and improved insulin sensitivity in a rat model of T2D. Robusta coffee was seemingly superior to arabica coffee with respect to effects on lipid profile, adiponectin level and hepatic gene expression.
Maternal Low-Fat Diet Programs the Hepatic Epigenome despite Exposure to an Obesogenic Postnatal Diet

Obesity and metabolic disease present a danger to long-term health outcomes. It has been hypothesized that epigenetic marks established during early life might program individuals and have either beneficial or harmful consequences later in life. In the present study, we examined whether maternal diet alters DNA methylation and whether such modifications persist after an obesogenic postnatal dietary challenge. During gestation and lactation, male Sprague-Dawley rats were exposed to either a high-fat diet (HF; n = 10) or low-fat diet (LF; n = 10). After weaning, all animals were fed a HF diet for an additional nine weeks. There were no differences observed in food intake or body weight between groups. Hepatic DNA methylation was quantified using both methylated DNA immunoprecipitation sequencing (MeDIP-seq) and methylation-sensitive restriction enzyme sequencing (MRE-seq). Overall, 1419 differentially methylated regions (DMRs) were identified. DMRs tended to be located in CpG shores and were enriched for genes involved in metabolism and cancer. Gene expression was measured for 31 genes in these pathways. Map3k5 and Igf1r were confirmed to be differentially expressed. Finally, we attempted to quantify the functional relevance of intergenic DMRs. Using chromatin contact data, we saw that conserved DMRs were topologically associated with metabolism genes, which were associated with differential expression of Adh5, Enox1, and Pik3c3. We show that although maternal dietary fat is unable to reverse offspring weight gain in response to a postnatal obesogenic diet, early life diet does program the hepatic methylome. Epigenetic alterations occur primarily in metabolic and cancer pathways and are associated with altered gene expression, but it is unclear whether they bear consequence later in life.
Utility of B-Type Natriuretic Peptide for Detecting Cardiac Involvement in Immunoglobulin Amyloidosis.

A useful biomarker for detecting cardiac amyloidosis (CA) has not been fully established. We aimed to investigate the utility of several biomarkers to detect CA in patients with amyloid light-chain (AL) amyloidosis.We examined the plasma levels of B-type natriuretic peptide (BNP), N-terminal fragment of the pro-brain natriuretic peptide (NT-proBNP), high-sensitivity cardiac troponin T (hs-cTnT), serum amyloid A, and the difference between kappa and lambda free light chain (dFLC) between CA patients (n = 30, 47.6%) and non-CA patients (n = 33, 52.4%). Levels of BNP were significantly higher in the CA group compared to the non-CA group (1200.0 versus 224.0 pg/mL, P = 0.001). From the ROC analysis, the sensitivity and specificity of BNP for detecting CA (with a cut-off value of 412 pg/mL) were 83% and 70%, respectively, and the area under the receiver operating curve was 0.75 (95% CI 0.61-0.90, P < 0.001) in all AL amyloidosis patients (n = 63). In contrast, other markers such as NT-proBNP, hs-cTnT, serum amyloid A, and dFLC were not useful for detecting CA in AL amyloidosis patients. Additionally, in the Cox proportional hazard analysis, BNP was a predictor of all-cause mortality (hazard ratio 3.266, 95% confidence interval 1.498-7.119, P = 0.003). BNP is a useful biomarker for detecting cardiac involvement and predicting prognosis in AL amyloidosis patients.
MiR-203 regulates JAK-STAT pathway in affecting pancreatic cancer cells proliferation and apoptosis by targeting SOCS3.

OBJECTIVE Janus kinase (JAK)- signal transducer and transcriptional activator (STAT) pathway overactivation is closely related to tumorigenesis. Cytokine signal transduction inhibitor 3 (SOCS3) is a negative regulator of JAK-STAT. It is shown that miR-203 is significantly elevated in the pancreatic cancer tissues. The bioinformatics analysis revealed a targeted binding site between miR-203 and the 3'-UTR of SOCS3 mRNA. This study investigated the role of miR-203 in regulating SOCS3 expression and the proliferation and apoptosis of the pancreatic cancer cells. PATIENTS AND METHODS Quantitative Real Time-PCR (qRT-PCR) was used to detect the expressions of miR-203 and SCOS3 mRNA in tumor tissues and paracancerous tissues. The Dual-Luciferase reporter gene assay was adopted to validate the target interaction between miR-203 and SOCS3. The PANC-1 cells were cultured in vitro and divided into miR-NC group and miR-203 inhibitor group followed by an analysis of the expressions of SOCS3, p-JAK2, and p-STAT3, cell apoptosis by flow cytometry, and cell proliferation by EdU staining. RESULTS Compared with the adjacent tissues, miR-203 expression was significantly increased, while SOCS3 mRNA level was significantly declined in the tumor tissues of pancreatic cancer patients. There was a targeted regulatory relationship between miR-203 and SOCS3 mRNA. Compared with those in HPDE6-C7 cells, miR-203 level was upregulated, whereas SOCS3 mRNA and the protein expressions were reduced in pancreatic cancer PANC-1 and BXPC3 cells. The transfection of miR-203 inhibitor significantly increased SOCS3 mRNA and the protein levels, decreased p-JAK2 and p-STAT3 protein expressions, enhanced cell apoptosis, and inhibited cell proliferation in the pancreatic cancer PANC-1 cells. CONCLUSIONS Increased miR-203 expression and reduced SOCS3 level are associated with the pathogenesis of pancreatic cancer. MiR-203 can regulate the proliferation and apoptosis of the pancreatic cancer cells by targeting the inhibited SOCS3 expression and regulating the JAK-STAT pathway activity.
Home initiation of chronic non-invasive ventilation in COPD patients with chronic hypercapnic respiratory failure: a randomised controlled trial

Introduction Chronic non-invasive ventilation (NIV) has become evidence-based care for stable hypercapnic COPD patients. While the number of patients increases, home initiation of NIV would greatly alleviate the healthcare burden. We hypothesise that home initiation of NIV with the use of telemedicine in stable hypercapnic COPD is non-inferior to in-hospital NIV initiation. Methods Sixty-seven stable hypercapnic COPD patients were randomised to initiation of NIV in the hospital or at home using telemedicine. Primary outcome was daytime arterial carbon dioxide pressure (PaCO2) reduction after 6 months NIV, with a non-inferiority margin of 0.4 kPa. Secondary outcomes were health-related quality of life (HRQoL) and costs. Results Home NIV initiation was non-inferior to in-hospital initiation (adjusted mean difference in PaCO2 change home vs in-hospital: 0.04 kPa (95% CI −0.31 to 0.38 kPa), with both groups showing a PaCO2 reduction at 6 months compared with baseline (home: from 7.3±0.9 to 6.4±0.8 kPa (p<0.001) and in-hospital: from 7.4±1.0 to 6.4±0.6 kPa (p<0.001)). In both groups, HRQoL improved without a difference in change between groups (Clinical COPD Questionnaire total score-adjusted mean difference 0.0 (95% CI −0.4 to 0.5)). Furthermore, home NIV initiation was significantly cheaper (home: median €3768 (IQR €3546–€4163) vs in-hospital: median €8537 (IQR €7540–€9175); p<0.001). Discussion This is the first study showing that home initiation of chronic NIV in stable hypercapnic COPD patients, with the use of telemedicine, is non-inferior to in-hospital initiation, safe and reduces costs by over 50%. Trial registration number NCT02652559.
Many human RNA viruses show extraordinarily stringent selective constraints on protein evolution

Significance The nonsynonymous substitutions (dN)-to-synonymous substitutions (dS) ratio in protein-coding genes is commonly used to study the mechanisms of gene evolution. To understand why RNA viruses show large variations in dN/dS, we studied the dN/dS ratios in 21 human RNA viruses, 8 human DNA viruses, and 17 mammals. Eighteen RNA viruses, but only 2 DNA viruses and no mammals, showed a genome-average dN/dS < 0.10. Thus, many human RNA viruses exhibited extraordinarily stringent selective constraints on protein evolution. Our among-gene and among-species comparisons revealed that both positive selection and population size play significant roles in the dN/dS variation among genes and species. This study clarified several controversial issues and increased our understanding of the mechanisms of RNA virus evolution. How negative selection, positive selection, and population size contribute to the large variation in nucleotide substitution rates among RNA viruses remains unclear. Here, we studied the ratios of nonsynonymous-to-synonymous substitution rates (dN/dS) in protein-coding genes of human RNA and DNA viruses and mammals. Among the 21 RNA viruses studied, 18 showed a genome-average dN/dS from 0.01 to 0.10, indicating that over 90% of nonsynonymous mutations are eliminated by negative selection. Only HIV-1 showed a dN/dS (0.31) higher than that (0.22) in mammalian genes. By comparing the dN/dS values among genes in the same genome and among species or strains, we found that both positive selection and population size play significant roles in the dN/dS variation among genes and species. Indeed, even in flaviviruses and picornaviruses, which showed the lowest ratios among the 21 species studied, positive selection appears to have contributed significantly to dN/dS. We found the view that positive selection occurs much more frequently in influenza A subtype H3N2 than subtype H1N1 holds only for the hemagglutinin and neuraminidase genes, but not for other genes. Moreover, we found no support for the view that vector-borne RNA viruses have lower dN/dS ratios than non–vector-borne viruses. In addition, we found a correlation between dN and dS, implying a correlation between dN and the mutation rate. Interestingly, only 2 of the 8 DNA viruses studied showed a dN/dS < 0.10, while 4 showed a dN/dS > 0.22. These observations increase our understanding of the mechanisms of RNA virus evolution.
Uncovering Structural and Molecular Dynamics of ESAT-6:β2M Interaction: Asp53 of Human β2-Microglobulin Is Critical for the ESAT-6:β2M Complexation

Key Points Asp53 residue of β2M is important in forming a complex with Met93 of ESAT-6. SM09 and SM15 rescued ESAT-6–mediated downregulation of MHC class I Ag presentation. SM09 and SM15 inhibited survival of M. tuberculosis inside the macrophage. ESAT-6 is a small secreted protein of Mycobacterium tuberculosis involved in the ESAT-6 secretion system (ESX-1)–mediated virulence and pathogenesis. The protein interacts with β2M, causing downregulation of MHC class I Ag presentation, which could be one of the mechanisms by which it favors increased survival of the bacilli inside the host. In an earlier study, we have shown that the C-terminal region of ESAT-6 is crucial for its interaction with β2M. However, the interface of β2M involved in interaction with ESAT-6 and detailed physicochemical changes associated with ESAT-6:β2M complexation are not fully defined. In this study, using computational and site-directed mutagenesis studies, we demonstrate the presence of strong noncovalent hydrophobic interactions between ESAT-6 and β2M in addition to the vital hydrogen bonding between the aspartate residue (Asp53) of β2M and methionine (Met93) of ESAT-6. Docking-based high-throughput virtual screening followed by 16-point screening on microscale thermophoresis resulted in the identification of two potent inhibitors (SM09 and SM15) that mask the critical Met93 residue of ESAT-6 that is required for ESAT-6:β2M interaction and could rescue cell surface expression of β2M and HLA in human macrophages as well as MHC class I Ag presentation suppressed by ESAT-6 in peritoneal macrophages isolated from C57BL/6 mice. Both SM09 and SM15 significantly inhibited intracellular survival of M. tuberculosis in human macrophages. Further, we characterized the physicochemical properties involved in the ESAT-6:β2M complexation, which may help in understanding host–pathogen interactions.
Understanding the molecular mechanisms of bisphenol A action in spermatozoa

Bisphenol A (BPA) is an endocrine-disrupting chemical that is capable of interfering with the normal function of the endocrine system in the body. Exposure to this chemical from BPA-containing materials and the environment is associated with deleterious health effects, including male reproductive abnormalities. A search of the literature demonstrated that BPA, as a toxicant, directly affects the cellular oxidative stress response machinery. Because of its hormone-like properties, it can also bind with specific receptors in target cells. Therefore, the tissue-specific effects of BPA mostly depend on its endocrine-disrupting capabilities and the expression of those particular receptors in target cells. Although studies have shown the possible mechanisms of BPA action in various cell types, a clear consensus has yet to be established. In this review, we summarize the mechanisms of BPA action in spermatozoa by compiling existing information in the literature.