Canagliflozin alleviates pulmonary hypertension by activating PPARγ and inhibiting its S225 phosphorylation.

Pulmonary hypertension (PH) is a progressive fatal disease with no cure. Canagliflozin (CANA), a novel medication for diabetes, has been found to have remarkable cardiovascular benefits. However, few studies have addressed the effect and pharmacological mechanism of CANA in the treatment of PH. Therefore, our study aimed to investigate the effect and pharmacological mechanism of CANA in treating PH. First, CANA suppressed increased pulmonary artery pressure, right ventricular hypertrophy, and vascular remodeling in both mouse and rat PH models. Network pharmacology, transcriptomics, and biological results suggested that CANA could ameliorate PH by suppressing excessive oxidative stress and pulmonary artery smooth muscle cell proliferation partially through the activation of PPARγ. Further studies demonstrated that CANA inhibited phosphorylation of PPARγ at Ser225 (a novel serine phosphorylation site in PPARγ), thereby promoting the nuclear translocation of PPARγ and increasing its ability to resist oxidative stress and proliferation. Taken together, our study not only highlighted the potential pharmacological effect of CANA on PH but also revealed that CANA-induced inhibition of PPARγ Ser225 phosphorylation increases its capacity to counteract oxidative stress and inhibits proliferation. These findings may stimulate further research and encourage future clinical trials exploring the therapeutic potential of CANA in PH treatment.

Asprosin contributes to vascular remodeling in hypertensive rats via superoxide signaling.

OBJECTIVE: Proliferation and migration of vascular smooth muscle cells (VSMCs) contribute to vascular remodeling. Asprosin, a newly discovered protein hormone, is involved in metabolic diseases. Little is known about the roles of asprosin in cardiovascular diseases. This study focused on the role and mechanism of asprosin on VSMC proliferation and migration, and vascular remodeling in a rat model of hypertension. METHODS AND RESULTS: VSMCs were obtained from the aortic media of 8-week-old male Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Asprosin was upregulated in the VSMCs of SHR. For in vitro studies, asprosin promoted VSMC proliferation and migration of WKY and SHR, and increased Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity, NOX1/2/4 protein expressions and superoxide production. Knockdown of asprosin inhibited the proliferation, migration, NOX activity, NOX1/2 expressions and superoxide production in the VSMCs of SHR. The roles of asprosin in promoting VSMC proliferation and migration were not affected by hydrogen peroxide scavenger, but attenuated by superoxide scavenger, selective NOX1 or NOX2 inhibitor. Toll-like receptor 4 (TLR4) was upregulated in SHR, TLR4 knockdown inhibited asprosin overexpression-induced proliferation, migration and oxidative stress in VSMCs of WKY and SHR. Asprosin was upregulated in arteries of SHR, and knockdown of asprosin in vivo not only attenuated oxidative stress and vascular remodeling in aorta and mesentery artery, but also caused a subsequent persistent antihypertensive effect in SHR. CONCLUSIONS: Asprosin promotes VSMC proliferation and migration via NOX-mediated superoxide production. Inhibition of endogenous asprosin expression attenuates VSMC proliferation and migration, and vascular remodeling of SHR.

Involvement of circRNA Regulators MBNL1 and QKI in the Progression of Esophageal Squamous Cell Carcinoma.

OBJECTIVES: To investigate the role of circRNA regulators MBNL1 and QKI in the progression of esophageal squamous cell carcinoma. BACKGROUND: MBNL1 and QKI are pivotal regulators of pre-mRNA alternative splicing, crucial for controlling circRNA production - an emerging biomarker and functional regulator of tumor progression. Despite their recognized roles, their involvement in ESCC progression remains unexplored. METHODS: The expression levels of MBNL1 and QKI were examined in 28 tissue pairs from ESCC and adjacent normal tissues using data from the GEO database. Additionally, a total of 151 ESCC tissue samples, from stage T1 to T4, consisting of 13, 43, 87, and 8 cases per stage, respectively, were utilized for immunohistochemical (IHC) analysis. RNA sequencing was utilized to examine the expression profiles of circRNAs, lncRNAs, and mRNAs across 3 normal tissues, 3 ESCC tissues, and 3 pairs of KYSE150 cells in both wildtype (WT) and those with MBNL1 or QKI knockouts. Transwell, colony formation, and subcutaneous tumorigenesis assays assessed the impact of MBNL1 or QKI knockout on ESCC cell migration, invasion, and proliferation. RESULTS: ESCC onset significantly altered MBNL1 and QKI expression levels, influencing diverse RNA species. Elevated MBNL1 or QKI expression correlated with patient age or tumor invasion depth, respectively. MBNL1 or QKI knockout markedly enhanced cancer cell migration, invasion, proliferation, and tumor growth. Moreover, the absence of either MBNL1 or QKI modulated the expression profiles of multiple circRNAs, causing extensive downstream alterations in the expression of numerous lncRNAs and mRNAs. While the functions of circRNA and lncRNA among the top 20 differentially expressed genes remain unclear, mRNAs like SLCO4C1, TMPRSS15, and MAGEB2 have reported associations with tumor progression. CONCLUSIONS: This study underscores the tumor-suppressive roles of MBNL1 and QKI in ESCC, proposing them as potential biomarkers and therapeutic targets for ESCC diagnosis and treatment.

Targeting of KOR by famotidine promotes OPC maturation differentiation and CNS remyelination via STAT3 signaling pathway.

This study aims to identify FDA-approved drugs that can target the kappa-opioid receptor (KOR) for the treatment of demyelinating diseases. Demyelinating diseases are characterized by myelin sheath destruction or formation that results in severe neurological dysfunction. Remission of this disease is largely dependent on the differentiation of oligodendrocyte precursor cells (OPCs) into mature oligodendrocytes (OLGs) in demyelinating lesions. KOR is an important regulatory protein and drug target for the treatment of demyelinating diseases. However, no drug targeting KOR has been developed due to the long clinical trials for drug discovery. Here, a structure-based virtual screening was applied to identify drugs targeting KOR among 1843 drugs of FDA-approved drug libraries, and famotidine was screen out by its high affinity cooperation with KOR as well as the clinical safety. We discovered that famotidine directly promoted OPC maturation and remyelination using the complementary in vitro and in vivo models. Administration of famotidine was not only effectively enhanced CNS myelinogenesis, but also promoted remyelination. Mechanically speaking, famotidine promoted myelinogenesis or remyelination through KOR/STAT3 signaling pathway. In general, our study provided evidence of new clinical applicability of famotidine for the treatment of demyelinating diseases for which there is currently no effective therapy.

The gut microbiota contributes to methamphetamine-induced reproductive toxicity in male mice.

Methamphetamine (METH) is a psychostimulant drug belonging to the amphetamine-type stimulant class, known to exert male reproductive toxicity. Recent studies suggest that METH can disrupt the gut microbiota. Furthermore, the gut-testis axis concept has gained attention due to the potential link between gut microbiome dysfunction and reproductive health. Nonetheless, the role of the gut microbiota in mediating the impact of METH on male reproductive toxicity remains unclear. In this study, we employed a mouse model exposed to escalating doses of METH to assess sperm quality, testicular pathology, and reproductive hormone levels. The fecal microbiota transplantation method was employed to investigate the effect of gut microbiota on male reproductive toxicity. Transcriptomic, metabolomic, and microbiological analyses were conducted to explore the damage mechanism to the male reproductive system caused by METH. We found that METH exposure led to hormonal disorders, decreased sperm quality, and changes in the gut microbiota and testicular metabolome in mice. Testicular RNA sequencing revealed enrichment of several Gene Ontology terms associated with reproductive processes, as well as PI3K-Akt signaling pathways. FMT conveyed similar reproductive damage from METH-treated mice to healthy recipient mice. The aforementioned findings suggest that the gut microbiota plays a substantial role in facilitating the reproductive toxicity caused by METH, thereby highlighting a prospective avenue for therapeutic intervention in the context of METH-induced infertility.

Knowledge and attitude of spouses of puerperas towards breastfeeding.

OBJECTIVE: To investigate the extent of knowledge about breastfeeding and attitudes towards infant feeding among spouses of puerperas at the time of discharge from hospital, and explore the factors influencing spousal attitudes toward breastfeeding. METHODS: We conducted a questionnaire survey among 204 spouses of puerperas who were admitted in the maternity wards at a tertiary hospital in Shaanxi Province between October 2021 and December 2021. Respondents who fulfilled the inclusion criteria were identified using convenient sampling. RESULTS: (1) The score of breastfeeding knowledge among spouses prior to discharge from the hospital was (10.56 ± 3.78), with an accuracy rate of 59.6%, and the lowest accuracy rate was for Item 1 "Newborns should be fed on time, not on demand" (42.6%) and Item 5 "Breastfeeding can prevent infant rickets" (49.5%). (2) The average score of spouses' infant feeding attitudes was (58.15 ± 5.55), and the lowest scoring was for Item 17 "Daily urine volume of infants is a reliable indicator to judge whether they get enough breast milk" (1.99 ± 1.14). (3) Generalized linear model analysis showed a more positive attitude (higher score) among spousal attitudes towards infant feeding in those who had received breastfeeding education [OR = 4.588, 95% CI (0.160 ∼ 3.598)] and those with a master's degree or above [OR = 18.278, 95% CI (3.471 ∼ 9.346)]. CONCLUSION: (1) Spouses that received breastfeeding education and those that had a Masters Degree and above had more positive attitude towards infant feeding. (2) Medical staff should focus on puerperas'spouses with degrees below master's level who had not received breastfeeding education. We recommend using a variety of education methods to enable them to acquire more knowledge on breastfeeding and develop a more positive attitude towards breastfeeding, which will further enhance spousal support for breastfeeding, thus positivizing postpartum co-parenting attitudes and improving the rate of exclusive breastfeeding.

Azaphilone pigments from the marine-derived Penicillium sclerotium UJNMF 0503 and their neuroprotective potential against H2O2-induced cell apoptosis through modulating PI3K/Akt pathway.

Azaphilones represent a particular group of fascinating pigments from fungal source, with easier industrialization and lower cost than the traditional plant-derived pigments, and they also display a wide range of pharmacological activities. Herein, 28 azaphilone analogs, including 12 new ones, were obtained from the fermentation culture of a marine fungus Penicillium sclerotium UJNMF 0503. Their structures were elucidated by MS, NMR and ECD analyses, together with NMR and ECD calculations and biogenetic considerations. Among them, compounds 1 and 2 feature an unusual natural benzo[d][1,3]dioxepine ring embedded with an orthoformate unit, while 3 and 4 represent the first azaphilone examples incorporating a novel rearranged 5/6 bicyclic core and a tetrahydropyran ring on the side chain, respectively. Our bioassays revealed that half of the isolates exhibited neuroprotective potential against H2O2-induced injury on RSC96 cells, while compound 13 displayed the best rescuing capacity toward the cell viability by blocking cellular apoptosis, which was likely achieved by upregulating the PI3K/Akt signaling pathway.

Co-culture of RhoA-overexpressed Microtia Chondrocytes and Adipose-Derived Stem Cells in the Construction of Tissue-engineered Ear-shaped Cartilage.

Microtia is a congenital auricle dysplasia with a high incidence and tissue engineering technology provides a promising strategy to reconstruct auricles. We previously described that the engineered cartilage constructed from microtia chondrocytes exhibited inferior levels of biochemical and biomechanical properties, which was proposed to be resulted from the decreased migration ability of microtia chondrocytes. In the current study, we found that Rho GTPase members were deficient in microtia chondrocytes. By overexpressing RhoA, Rac1 and CDC42, respectively, we further demonstrated that RhoA took great responsibility for the decreased migration ability of microtia chondrocytes. Moreover, we constructed PGA/PLA scaffold-based cartilages to verify the chondrogenic ability of RhoA overexpressed microtia chondrocytes, and the results showed that overexpressing RhoA was of limited help to improve the quality of microtia chondrocyte engineered cartilage. However, co-culture of ADSCs significantly improved the biochemical and biomechanical property of engineered cartilage. Especially, co-culture of RhoA overexpressed microtia chondrocytes and ADSCs produced an excellent effect on the wet weight, cartilage-specific extracellular matrix and biomechanical property of engineered cartilage. Furthermore, we presented that co-culture of RhoA overexpressed microtia chondrocytes and ADSCs combined with human ear-shaped PGA/PLA scaffold and titanium alloy stent fabricated by CAD/CAM and 3D printing technology effectively constructed and maintained auricle structure in vivo. Collectively, our results provide evidence for the essential role of RhoA in microtia chondrocytes and a developed strategy for the construction of patient-specific tissue-engineered auricular cartilage.

Applicability of the generalized wind profile model over mountainous forests.

The analytical equation based on Monin-Obukhov (M-O) similarity theory (i.e., wind profile equation) has been adopted since 1970s for using in the prediction of wind vertical profile over flat terrains, which is mature and accurate. However, its applicability over complex terrains remains unknown. This applicability signifies the accuracy of the estimations of aerodynamic parameters for the boundary layer of non-flat terrain, such as zero-displacement height (d) and aerodynamic roughness length (z0), which will determine the accuracy of frequency correction and source area analysis in calculating carbon, water, and trace gas fluxes based on vorticity covariance method. Therefore, the validation of wind profile model in non-flat terrain is the first step to test whether the flux model needs improvement. We measured three-dimensional wind speed data by using the Ker Towers (three towers in a watershed) at Qingyuan Forest CERN in the Mountainous Region of east Liaoning Province, and compared them with data from Panjin Agricultural Station in the Liaohe Plain, to evaluate the applicability of a generalized wind profile model based on the Monin-Obukhov similarity theory on non-flat terrain. The results showed that the generalized wind profile model could not predict wind speeds accurately of three flux towers separately located in different sites, indicating that wind profile model was not suitable for predicting wind speeds in complex terrains. In the leaf-off and leaf-on periods, the coefficient of determination (R2) between observed and predicted wind speeds ranged from 0.12 to 0.30. Compared to measured values, the standard error of the predicted wind speeds was high up to 2 m·s-1. The predicted wind speeds were high as twice as field-measured wind speed, indicating substantial overestimation. Nevertheless, this model correctly predicted wind speeds in flat agricultural landscape in Panjin Agricultural Station. The R2 between observed wind speeds and predicted wind speed ranged from 0.90 to 0.93. The standard error between observed and predicted values was only 0.5 m·s-1. Results of the F-test showed that the root-mean-square error of the observed and predicted wind speeds in each secondary forest complex terrain was much greater than that in flat agricultural landscape. Terrain was the primary factor affecting the applicability of wind profile model, followed by seasonality (leaf or leafless canopy). The wind profile model was not applicable to the boundary-layer flows over forest canopies in complex terrains, because the d was underestimated or both the d and z0 were underestimated, resulting in inaccurate estimation of aerodynamic height.

Investigation of Metabolic and Inflammatory Disorder in the Aging FGF21 Knockout Mouse.

Aging is a physiological condition accomplished with persistent low-grade inflammation and metabolic disorders. FGF21 has been reported to act as a potent longevity determinant, involving inflammatory response and energy metabolism. In this study, we engineered aging FGF21 knockout mice of 36-40 weeks and observed that FGF21 deficiency manifests a spontaneous inflammatory response of lung and abnormal accumulation of lipids in liver. On one hand, inflamed state in lungs and increased circulating inflammatory cytokines were found in FGF21 knockout mice of 36-40 weeks. To evaluate the ability of FGF21 to suppress inflammation, a subsequent study found that FGF21 knockout aggravated LPS-induced pulmonary exudation and inflammatory infiltration in mice, while exogenous administration of FGF21 reversed these malignant phenotypes by enhancing microvascular endothelial junction. On the other hand, FGF21 knockout induces fatty liver in aging mice, characterized by excessive accumulation of triglycerides within hepatocytes. Further quantitative metabolomics and lipidomics analysis revealed perturbed metabolic profile in liver lacking FGF21, including disrupted glucose and lipids metabolism, glycerophospholipid metabolism, and amino acid metabolism. Taken together, this investigation reveals the protective role of FGF21 during aging by weakening the inflammatory response and balancing energy metabolism.

Potassium affects the association between dietary intake of vitamin C and NAFLD among adults in the United States.

INTRODUCTION: Although the association between nonalcoholic fatty liver disease (NAFLD) and vitamin C has been well studied, the effects of dietary potassium intake on this relationship are still unclear. Thus, this study aimed to determine the effects of dietary potassium intake on the association between vitamin C and NAFLD. METHODS: We performed a cross-sectional learn about with 9443 contributors the usage of 2007-2018 NHANES data. Multiple logistic regression evaluation has been utilized to check out the affiliation of dietary vitamin C intake with NAFLD and advanced hepatic fibrosis (AHF). Subsequently, we plotted a smoothed match curve to visualize the association. Especially, the analysis of AHF was conducted among the NAFLD population. In addition, stratified evaluation used to be developed primarily based on demographic variables to verify the steadiness of the results. Effect amendment by way of dietary potassium intake used to be assessed via interplay checks between vitamin C and NAFLD in the multivariable linear regression. RESULTS: In this cross-sectional study, we found that vitamin C was negatively related to NAFLD and AHF. The relationship between vitamin C and NAFLD was different in the low, middle and high potassium intake groups. Furthermore, potassium intake significantly modified the negative relationship between vitamin C and NAFLD in most of the models. CONCLUSION: Our research showed that potassium and vitamin C have an interactive effect in reducing NAFLD, which may have great importance for clinical medication.

Associations between trans fatty acids and systemic immune-inflammation index: a cross-sectional study.

BACKGROUND: Previous studies have demonstrated that trans fatty acids (TFAs) intake was linked to an increased risk of chronic diseases. As a novel systemic inflammatory biomarker, the clinical value and efficacy of the systemic immune-inflammation index (SII) have been widely explored. However, the association between TFAs and SII is still unclear. Therefore, the study aims to investigate the connection between TFAs and SII in US adults. METHODS: The study retrieved data from the National Health and Nutrition Examination Survey (NHANES) for the years 1999-2000 and 2009-2010. Following the exclusion of ineligible participants, the study encompassed a total of 3047 individuals. The research employed a multivariate linear regression model to investigate the connection between circulating TFAs and SII. Furthermore, the restricted cubic spline (RCS) model was utilized to evaluate the potential nonlinear association. Subgroup analysis was also conducted to investigate the latent interactive factors. RESULTS: In this investigation, participants exhibited a mean age of 47.40 years, with 53.91% of them being female. Utilizing a multivariate linear regression model, the independent positive associations between the log2-transformed palmitelaidic acid, the log2 transformed-vaccenic acid, the log2-transformed elaidic acid, the log2-transformed linolelaidic acid, and the log2-transformed-total sum of TFAs with the SII (all P < 0.05) were noted. In the RCS analysis, no nonlinear relationship was observed between the log2-transformed palmitelaidic acid, the log2 transformed-vaccenic acid, the log2-transformed elaidic acid, the log2-transformed linolelaidic acid, the log2-transformed-total sum of TFAs and the SII (all P for nonlinear > 0.05). For the stratified analysis, the relationship between the circulating TFAs and the SII differed by the obesity status and the smoking status. CONCLUSIONS: A positive association was investigated between three types of TFA, the sum of TFAs, and the SII in the US population. Additional rigorously designed studies are needed to verify the results and explore the potential mechanism.

Impaired meningeal lymphatic drainage in Listeria monocytogenes infection.

Previous studies have demonstrated an association between lymphatic vessels and diseases caused by bacterial infections. Listeria monocytogenes (LM) bacterial infection can affect multiple organs, including the intestine, brain, liver and spleen, which can be fatal. However, the impacts of LM infection on morphological and functional changes of lymphatic vessels remain unexplored. In this study, we found that LM infection not only induces meningeal and mesenteric lymphangiogenesis in mice, but also impairs meningeal lymphatic vessels (MLVs)-mediated macromolecules drainage. Interestingly, we found that the genes associated with lymphatic vessel development and function, such as Gata2 and Foxc2, were downregulated, suggesting that LM infection may affect cellular polarization and valve development. On the other hand, photodynamic ablation of MLVs exacerbated inflammation and bacterial load in the brain of mice with LM infection. Overall, our findings indicate that LM infection induces lymphangiogenesis and may affect cell polarization, cavity formation, and valve development during lymphangiogenesis, ultimately impairing MLVs drainage.

The Efficacy & Molecular Mechanisms of a Terpenoid Compound Ganoderic Acid C1 on Corticosteroid-Resistant Neutrophilic Airway Inflammation: In vivo and in vitro Validation.

Introduction: Neutrophil predominant airway inflammation is associated with severe and steroid-resistant asthma clusters. Previously, we reported efficacy of ASHMI, a three-herb TCM asthma formula in a steroid-resistant neutrophil-dominant murine asthma model and further identified Ganoderic Acid C1 (GAC1) as a key ASHMI active compound in vitro. The objective of this study is to investigate GAC1 effect on neutrophil-dominant, steroid-resistant asthma in a murine model. Methods: In this study, Balb/c mice were systematically sensitized with ragweed (RW) and alum and intranasally challenged with ragweed. Unsensitized/PBS challenged mice served as normal controls. Post sensitization, mice were given 4 weeks of oral treatment with GAC1 or acute dexamethasone (Dex) treatment at 48 hours prior to challenge. Pulmonary cytokines were measured by ELISA, and lung sections were processed for histology by H&E staining. Furthermore, GAC1 effect on MUC5AC expression and on reactive oxygen species (ROS) production in human lung epithelial cell line (NCI-H292) was determined by qRT-PCR and ROS assay kit, respectively. Computational analysis was applied to select potential targets of GAC1 in steroid-resistant neutrophil-dominant asthma. Molecular docking was performed to predict binding modes between GAC1 and Dex with TNF-α. Results: The result of the study showed that chronic GAC1 treatment, significantly reduced pulmonary inflammation (P < 0.01-0.001 vs Sham) and airway neutrophilia (P < 0.01 vs Sham), inhibited TNF-α, IL-4 and IL-5 levels (P < 0.05-0.001 vs Sham). Acute Dex treatment reduced eosinophilic inflammation and IL-4, IL-5 levels, but had no effect on neutrophilia and TNF-α production. GAC1 treated H292 cells showed decreased MUC5AC gene expression and production of ROS (P < 0.001 vs stimulated/untreated cells). Molecular docking results showed binding energy of complex GAC1-TNF was -10.8 kcal/mol. Discussion: GAC1 may be a promising anti-asthma botanical drug for treatment of steroid-resistant asthma.

Pseudorabies virus manipulates mitochondrial tryptophanyl-tRNA synthetase 2 for viral replication.

The pseudorabies virus (PRV) is identified as a double-helical DNA virus responsible for causing Aujeszky's disease, which results in considerable economic impacts globally. The enzyme tryptophanyl-tRNA synthetase 2 (WARS2), a mitochondrial protein involved in protein synthesis, is recognized for its broad expression and vital role in the translation process. The findings of our study showed an increase in both mRNA and protein levels of WARS2 following PRV infection in both cell cultures and animal models. Suppressing WARS2 expression via RNA interference in PK-15 â€‹cells led to a reduction in PRV infection rates, whereas enhancing WARS2 expression resulted in increased infection rates. Furthermore, the activation of WARS2 in response to PRV was found to be reliant on the cGAS/STING/TBK1/IRF3 signaling pathway and the interferon-alpha receptor-1, highlighting its regulation via the type I interferon signaling pathway. Further analysis revealed that reducing WARS2 levels hindered PRV's ability to promote protein and lipid synthesis. Our research provides novel evidence that WARS2 facilitates PRV infection through its management of protein and lipid levels, presenting new avenues for developing preventative and therapeutic measures against PRV infections.

Biological characteristics of Cordyceps militaris single mating-type strains.

Cordyceps militaris has been extensively cultivated as a model cordyceps species for commercial purposes. Nevertheless, the problems related to strain degeneration and breeding technologies remain unresolved. This study assessed the physiology and fertility traits of six C. militaris strains with distinct origins and characteristics, focusing on single mating-type strains. The results demonstrated that the three identified strains (CMDB01, CMSY01, and CMJB02) were single mating-type possessing only one mating-type gene (MAT1-1). In contrast, the other three strains (CMXF07, CMXF09, and CMMS05) were the dual mating type. The MAT1-1 strains sourced from CMDB01, CMSY01, and CMJB02 consistently produced sporocarps but failed to generate ascospores. However, when paired with MAT1-2 strains, the MAT1-1 strains with slender fruiting bodies and normal morphology were fertile. The hyphal growth rate of single mating-type strains (CMDB01, CMSY01, and CMJB02) typically surpassed that of dual mating-type strains (CMXF07, CMXF09, and CMMS05). The growth rates of MAT1-2 and MAT1-1 strains were proportional to their ratios, such that a single mating-type strain with a higher ratio exhibited an increased growth rate. As C. militaris matured, the adenosine content decreased. In summary, the C. militaris strains that consistently produce sporocarps and have a single mating type are highly promising for production and breeding.

Trx4, a novel thioredoxin protein, is important for Toxoplasma gondii fitness.

BACKGROUND: To successfully replicate within the host cell, Toxoplasma gondii employs several mechanisms to overcome the host cell defenses and mitigate the harmful effects of the free radicals resulting from its own metabolic processes using effectors such as thioredoxin proteins. In this study, we characterize the location and functions of a newly identified thioredoxin in T. gondii, which was named Trx4. METHODS: We characterized the functional role of Trx4 in T. gondii Type I RH and Type II Pru strains by gene knockout and studied its subcellular localization by endogenous protein HA tagging using CRISPR-Cas9 gene editing. The enzyme-catalyzed proximity labeling technique, the TurboID system, was employed to identify the proteins in proximity to Trx4. RESULTS: Trx4 was identified as a dense granule protein of T. gondii predominantly expressed in the parasitophorous vacuole (PV) and was partially co-localized with GRA1 and GRA5. Functional analysis showed that deletion of trx4 markedly influenced the parasite lytic cycle, resulting in impaired host cell invasion capacity in both RH and Pru strains. Mutation of Trx domains in Trx4 in RH strain revealed that two Trx domains were important for the parasite invasion. By utilizing the TurboID system to biotinylate proteins in proximity to Trx4, we identified a substantial number of proteins, some of which are novel, and others are previously characterized, predominantly distributed in the dense granules. In addition, we uncovered three novel proteins co-localized with Trx4. Intriguingly, deletion of trx4 did not affect the localization of these three proteins. Finally, a virulence assay demonstrated that knockout of trx4 resulted in a significant attenuation of virulence and a significant reduction in brain cyst loads in mice. CONCLUSIONS: Trx4 plays an important role in T. gondii invasion and virulence in Type I RH strain and Type II Pru strain. Combining the TurboID system with CRISPR-Cas9 technique revealed many PV-localized proximity proteins associated with Trx4. These findings suggest a versatile role of Trx4 in mediating the processes that occur in this distinctive intracellular membrane-bound vacuolar compartment.

Generic and accurate prediction of retention times in liquid chromatography by post-projection calibration.

Retention time predictions from molecule structures in liquid chromatography (LC) are increasingly used in MS-based targeted and untargeted analyses, providing supplementary evidence for molecule annotation and reducing experimental measurements. Nevertheless, different LC setups (e.g., differences in gradient, column, and/or mobile phase) give rise to many prediction models that can only accurately predict retention times for a specific chromatographic method (CM). Here, a generic and accurate method is present to predict retention times across different CMs, by introducing the concept of post-projection calibration. This concept builds on the direct projections of retention times between different CMs and uses 35 external calibrants to eliminate the impact of LC setups on projection accuracy. Results showed that post-projection calibration consistently achieved a median projection error below 3.2% of the elution time. The ranking results of putative candidates reached similar levels among different CMs. This work opens up broad possibilities for coordinating retention times between different laboratories and developing extensive retention databases.

Less common phenotypes of myelin oligodendrocyte glycoprotein antibody-related diseases in children deserve more attention.

BACKGROUND: To facilitate the identification of less common clinical phenotypes of myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) in children. METHODS: We retrospectively reviewed medical records of 236 patients with MOGAD. The following phenotypes were considered to be typical for MOGAD: ADEM, ON, TM, and NMOSD. Less common onset clinical phenotypes were screened out; their clinical and magnetic resonance imaging (MRI), diagnosis, treatment, and prognosis were summarized and analyzed. RESULTS: 16 cases (6.8%) presented as cortical encephalitis, with convulsions, headache, and fever as the main symptoms. 15 cases were misdiagnosed in the early period. 13 cases (5.5%) showed the overlapping syndrome of MOGAD and anti-N-methyl-D aspartate receptor encephalitis (MNOS), with seizures (92.3%) being the most common clinical symptom. 11 cases (84.6%) showed relapses. The cerebral leukodystrophy-like phenotype was present in seven cases (3.0%), with a recurrence rate of 50%. Isolated seizures without any findings on MRI phenotype was present in three cases (1.3%), with the only clinical symptom being seizures of focal origin. Three cases (1.3%) of aseptic meningitis phenotype presented with prolonged fever. CONCLUSION: 40/236 (16.9%) of children with MOGAD had less common phenotypes. Less common clinical phenotypes of pediatric MOGAD are susceptible to misdiagnosis and deserve more attention. IMPACT: This is the first comprehensive analysis and summary of all less commonl clinical phenotypes of MOGAD in children, while previous studies have only focused on a specific phenotype or case reports. We analyzed the characteristics of MOGAD in children and further revealed the reasons why these less common clinical phenotypes are prone to misdiagnosis and deserve more attention. Our research on treatment has shown that early detection of MOG antibodies and early treatment are of great significance for improving the prognosis of these patients.