CD4+ T cell-mediated recognition of a conserved cholesterol-dependent cytolysin epitope generates broad antibacterial immunity.

CD4+ T cell-mediated immunity against Streptococcus pneumoniae (pneumococcus) can protect against recurrent bacterial colonization and invasive pneumococcal diseases (IPDs). Although such immune responses are common, the pertinent antigens have remained elusive. We identified an immunodominant CD4+ T cell epitope derived from pneumolysin (Ply), a member of the bacterial cholesterol-dependent cytolysins (CDCs). This epitope was broadly immunogenic as a consequence of presentation by the pervasive human leukocyte antigen (HLA) allotypes DPB1∗02 and DPB1∗04 and recognition via architecturally diverse T cell receptors (TCRs). Moreover, the immunogenicity of Ply427-444 was underpinned by core residues in the conserved undecapeptide region (ECTGLAWEWWR), enabling cross-recognition of heterologous bacterial pathogens expressing CDCs. Molecular studies further showed that HLA-DP4-Ply427-441 was engaged similarly by private and public TCRs. Collectively, these findings reveal the mechanistic determinants of near-global immune focusing on a trans-phyla bacterial epitope, which could inform ancillary strategies to combat various life-threatening infectious diseases, including IPDs.

Ablation of the dystrophin Dp71f alternative C-terminal variant increases sarcoma tumour cell aggressiveness.

Alterations in Dp71 expression, the most ubiquitous dystrophin isoform, have been associated with patient survival across tumours. Intriguingly, in certain malignancies, Dp71 acts as a tumour suppressor, while manifesting oncogenic properties in others. This diversity could be explained by the expression of two Dp71 splice variants encoding proteins with distinct C-termini, each with specific properties. Expression of these variants has impeded the exploration of their unique roles. Using CRISPR/Cas9, we ablated the Dp71f variant with the alternative C-terminus in a sarcoma cell line not expressing the canonical C-terminal variant, and conducted molecular (RNAseq) and functional characterisation of the knockout cells. Dp71f ablation induced major transcriptomic alterations, particularly affecting the expression of genes involved in calcium signalling and ECM-receptor interaction pathways. The genome-scale metabolic analysis identified significant downregulation of glucose transport via membrane vesicle reaction (GLCter) and downregulated glycolysis/gluconeogenesis pathway. Functionally, these molecular changes corresponded with, increased calcium responses, cell adhesion, proliferation, survival under serum starvation and chemotherapeutic resistance. Knockout cells showed reduced GLUT1 protein expression, survival without attachment and their migration and invasion in vitro and in vivo were unaltered, despite increased matrix metalloproteinases release. Our findings emphasise the importance of alternative splicing of dystrophin transcripts and underscore the role of the Dp71f variant, which appears to govern distinct cellular processes frequently dysregulated in tumour cells. The loss of this regulatory mechanism promotes sarcoma cell survival and treatment resistance. Thus, Dp71f is a target for future investigations exploring the intricate functions of specific DMD transcripts in physiology and across malignancies.

Degradation strategies for structural characterization of insoluble synthetic polymers by mass spectrometry.

With the advent of soft ionization techniques such as electrospray (ESI) and matrix-assisted laser desorption/ionization (MALDI) to produce intact gas-phase ions from nonvolatile macromolecules, mass spectrometry has become an essential technique in the field of polymeric materials. However, (co)polymers of very high molecular weight or with reticulated architectures still escape ESI or MALDI, mainly due to solubility issues. Strategies developed to tackle such an analytical challenge all rely on sample degradation to produce low-mass species amenable to existing ionization methods. Yet, chain degradation needs to be partial and controlled to generate sufficiently large species that still contain topological or architectural information. The present article reviews the different analytical degradation strategies implemented to perform mass spectrometry of these challenging synthetic polymers, covering thermal degradation approaches in sources developed in the 2000s, off-line sample pre-treatments for controlled chemical degradation of polymeric substrates, and most recent achievements employing reactive ionization modes to perform chemolysis on-line with MS.

Prenatal treatment with preimplantation factor improves early postnatal neurogenesis and cognitive impairments in a mouse model of Down syndrome.

Down syndrome (DS) is a genetic disease characterized by a supernumerary chromosome 21. Intellectual deficiency (ID) is one of the most prominent features of DS. Central nervous system defects lead to learning disabilities, motor and language delays, and memory impairments. At present, a prenatal treatment for the ID in DS is lacking. Subcutaneous administration of synthetic preimplantation factor (sPIF, a peptide with a range of biological functions) in a model of severe brain damage has shown neuroprotective and anti-inflammatory properties by directly targeting neurons and microglia. Here, we evaluated the effect of PIF administration during gestation and until weaning on Dp(16)1Yey mice (a mouse model of DS). Possible effects at the juvenile stage were assessed using behavioral tests and molecular and histological analyses of the brain. To test the influence of perinatal sPIF treatment at the adult stage, hippocampus-dependent memory was evaluated on postnatal day 90. Dp(16)1Yey pups showed significant behavioral impairment, with impaired neurogenesis, microglial cell activation and a low microglial cell count, and the deregulated expression of genes linked to neuroinflammation and cell cycle regulation. Treatment with sPIF restored early postnatal hippocampal neurogenesis, with beneficial effects on astrocytes, microglia, inflammation, and cell cycle markers. Moreover, treatment with sPIF restored the level of DYRK1A, a protein that is involved in cognitive impairments in DS. In line with the beneficial effects on neurogenesis, perinatal treatment with sPIF was associated with an improvement in working memory in adult Dp(16)1Yey mice. Perinatal treatment with sPIF might be an option for mitigating cognitive impairments in people with DS.

Human T cells recognize HLA-DP-bound peptides in two orientations.

Human leukocyte antigen (HLA) molecules present small peptide antigens to T cells, thereby allowing them to recognize pathogen-infected and cancer cells. A central dogma over the last 50+ y is that peptide binding to HLA molecules is mediated by the docking of side chains of particular amino acids in the peptide into pockets in the HLA molecules in a conserved N- to C-terminal orientation. Whether peptides can be presented in a reversed C- to N-terminal orientation remains unclear. Here, we performed large-scale identification of peptides bound to HLA-DP molecules and observed that in addition to peptide binding in an N- to C-terminal orientation, in 9 out of 14 HLA-DP allotypes, reverse motifs are found, compatible with C- to N-terminal peptide binding. Moreover, we isolated high-avidity human cytomegalovirus (CMV)-specific HLA-DP-restricted CD4+ T cells from the memory repertoire of healthy donors and demonstrate that such T cells recognized CMV-derived peptides bound to HLA-DPB1*01:01 or *05:01 in a reverse C- to N-terminal manner. Finally, we obtained a high-resolution HLA-DPB1*01:01-CMVpp65(142-158) peptide crystal structure, which is the molecular basis for C- to N-terminal peptide binding to HLA-DP. Our results point to unique features of HLA-DP molecules that substantially broaden the HLA class II bound peptide repertoire to combat pathogens and eliminate cancer cells.

Comprehensive analysis of genomic complexity in the 5' end coding region of the DMD gene in patients of exons 1-2 duplications based on long-read sequencing.

BACKGROUND: Dystrophinopathies are the most common X-linked inherited muscle diseases, and the disease-causing gene is DMD. Exonic duplications are a common type of pathogenic variants in the DMD gene, however, 5' end exonic duplications containing exon 1 are less common. When assessing the pathogenicity of exonic duplications in the DMD gene, consideration must be given to their impact on the reading frame. Traditional molecular methods, such as multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS), are commonly used in clinics. However, they cannot discriminate the precise physical locations of breakpoints and structural features of genomic rearrangement. Long-read sequencing (LRS) can effectively overcome this limitation. RESULTS: We used LRS technology to perform whole genome sequencing on three families and analyze the structural variations of the DMD gene, which involves the duplications of exon 1 and/or exon 2. Two distinct variant types encompassing exon 1 in the DMD Dp427m isoform and/or Dp427c isoform are identified, which have been infrequently reported previously. In pedigree 1, the male individuals harboring duplication variant of consecutive exons 1-2 in the DMD canonical transcript (Dp427m) and exon 1 in the Dp427c transcript are normal, indicating the variant is likely benign. In pedigree 3, the patient carries complex SVs involving exon 1 of the DMD Dp427c transcript showing an obvious phenotype. The locations of the breakpoints and the characteristics of structural variants (SVs) are identified by LRS, enabling the classification of the variants' pathogenicity. CONCLUSIONS: Our research sheds light on the complexity of DMD variants encompassing Dp427c/Dp427m promoter regions and emphasizes the importance of cautious interpretation when assessing the pathogenicity of DMD 5' end exonic duplications, particularly in carrier screening scenarios without an affected proband.

Efficacy, tolerability, and safety of teverelix DP in patients with advanced prostate cancer: A multicenter, open-label, phase 2 trial.

BACKGROUND: Teverelix drug product (DP) is a novel injectable gonadotropin-releasing hormone antagonist. METHODS: An adaptive phase 2, open-label, multicenter trial was conducted in patients with advanced prostate cancer to evaluate the efficacy and safety of a combined subcutaneous (SC) and intramuscular (IM) loading dose regimen of teverelix DP of 120 mg SC + 120 mg IM (Group 1; N = 9) or 180 mg SC + 180 mg IM (Group 2; N = 41) administered at a single visit, followed by 6-weekly SC maintenance doses of 120 mg (Group 1) or 180 mg (Group 2), up to Day 168. The primary endpoint was the proportion of patients achieving castration levels with serum testosterone <0.5 ng/mL at Day 28 with a target castration rate of 90%. Injection sites were inspected by the investigator at every visit and reactions (ISRs) were proactively recorded. RESULTS: The target castration rate was reached in Group 2 (97.5%) but not in Group 1 (62.5%). The castration rates were not maintained to Day 42 (Group 2: 82.5%; Group 1: 50.0%). Suppression of testosterone to castrate levels occurred rapidly (median time: 2 days for both groups). Suppression of testosterone, prostate-specific antigen, follicle-stimulating hormone, and luteinizing hormone was sustained throughout the treatment period, being more prominent with the higher dose. The adverse event (AE) profile was similar between groups. The most common AEs were injection-site induration (n = 40: 80.0%), injection-site erythema (n = 35: 70.0%), and hot flush (n = 21: 42.0%). Most ISRs were Grade 1. CONCLUSION: Overall, the teverelix DP doses were generally well-tolerated but did not adequately maintain castration levels.

Pharmacogenetic determinants of tenofovir diphosphate and lamivudine triphosphate concentrations in people with HIV/HBV coinfection.

The nucleos(t)ide analogs require phosphorylation to the pharmacologically active anabolites in cells. We investigated the hypothesis that single-nucleotide polymorphisms (SNPs) in genes that encode transporters and phosphodiesterase (PDE) enzymes involved in tenofovir (TFV), disoproxil fumarate (TDF), and lamivudine (3TC) disposition will be associated with concentrations of their phosphate anabolites and virologic response. Individuals with human immunodeficiency virus (HIV) and hepatitis B virus (HBV) coinfection receiving TDF/3TC-containing antiretroviral therapy were enrolled. Steady-state TFV diphosphate (TFV-DP) and 3TC triphosphate (3TC-TP) concentrations in peripheral blood mononuclear cells (PBMCs) and dried blood spot samples were quantified. The relationship between genetic variants and TFV-DP and 3TC-TP concentrations as well as with virologic response were examined using multivariable linear regression. Of the 136 participants (median age 43 years; 63% females), 6.6% had HBV non-suppression, and 7.4% had HIV non-suppression. The multidrug resistance protein 2 (encoded by ABCC2 rs2273697) SNP was associated with 3TC-TP concentrations in PBMCs. The human organic anion transporter-1 (encoded by SLC28A2) rs11854484 SNP was associated with HIV non-suppression, and when evaluated together with SNPs with marginal associations (ABCC2 rs717620 and PDE1C rs30561), participants with two or three variants compared to those with none of these variants had an adjusted odds ratio of 48.3 (confidence interval, 4.3-547.8) for HIV non-suppression. None of the SNPs were associated with HBV non-suppression. Our study identified ABCC2 SNP to be associated with 3TC-TP concentrations in PBMCs. Also, a combination of genetic variants of drug transporters and PDE was associated with HIV non-suppression.

HLA-DP on Epithelial Cells Enables Tissue Damage by NKp44+ Natural Killer Cells in Ulcerative Colitis.

BACKGROUND & AIMS: Ulcerative colitis (UC) is characterized by severe inflammation and destruction of the intestinal epithelium, and is associated with specific risk single nucleotide polymorphisms in HLA class II. Given the recently discovered interactions between subsets of HLA-DP molecules and the activating natural killer (NK) cell receptor NKp44, genetic associations of UC and HLA-DP haplotypes and their functional implications were investigated. METHODS: HLA-DP haplotype and UC risk association analyses were performed (UC: n = 13,927; control: n = 26,764). Expression levels of HLA-DP on intestinal epithelial cells (IECs) in individuals with and without UC were quantified. Human intestinal 3-dimensional (3D) organoid cocultures with human NK cells were used to determine functional consequences of interactions between HLA-DP and NKp44. RESULTS: These studies identified HLA-DPA1∗01:03-DPB1∗04:01 (HLA-DP401) as a risk haplotype and HLA-DPA1∗01:03-DPB1∗03:01 (HLA-DP301) as a protective haplotype for UC in European populations. HLA-DP expression was significantly higher on IECs of individuals with UC compared with controls. IECs in human intestinal 3D organoids derived from HLA-DP401pos individuals showed significantly stronger binding of NKp44 compared with HLA-DP301pos IECs. HLA-DP401pos IECs in organoids triggered increased degranulation and tumor necrosis factor production by NKp44+ NK cells in cocultures, resulting in enhanced epithelial cell death compared with HLA-DP301pos organoids. Blocking of HLA-DP401-NKp44 interactions (anti-NKp44) abrogated NK cell activity in cocultures. CONCLUSIONS: We identified an UC risk HLA-DP haplotype that engages NKp44 and activates NKp44+ NK cells, mediating damage to intestinal epithelial cells in an HLA-DP haplotype-dependent manner. The molecular interaction between NKp44 and HLA-DP401 in UC can be targeted by therapeutic interventions to reduce NKp44+ NK cell-mediated destruction of the intestinal epithelium in UC.

Genome-wide characterization and expression profiling of E2F/DP gene family members in response to abiotic stress in tomato (Solanum lycopersicum L.).

BACKGROUND: E2F/DP (Eukaryotic 2 transcription factor/dimerization partner) family proteins play an essential function in the cell cycle development of higher organisms. E2F/DP family genes have been reported only in a few plant species. However, comprehensive genome-wide characterization analysis of the E2F/DP gene family of Solanum lycopersicum has not been reported so far. RESULTS: This study identified eight nonredundant SlE2F/DP genes that were classified into seven groups in the phylogenetic analysis. All eight genes had a single E2F-TDP domain and few genes had additional domains. Two segmental duplication gene pairs were observed within tomato, in addition to cis-regulatory elements, miRNA target sites and phosphorylation sites which play an important role in plant development and stress response in tomato. To explore the three-dimensional (3D) models and gene ontology (GO) annotations of SlE2F/DP proteins, we pointed to their putative transporter activity and their interaction with several putative ligands. The localization of SlE2F/DP-GFP fused proteins in the nucleus and endoplasmic reticulum suggested that they may act in other biological functions. Expression studies revealed the differential expression pattern of most of the SlE2F/DP genes in various organs. Moreover, the expression of E2F/DP genes against abiotic stress, particularly SlE2F/DP2 and/or SlE2F/DP7, was upregulated in response to heat, salt, cold and ABA treatment. Furthermore, the co-expression analysis of SlE2F/DP genes with multiple metabolic pathways was co-expressed with defence genes, transcription factors and so on, suggested their crucial role in various biological processes. CONCLUSIONS: Overall, our findings provide a way to understand the structure and function of SlE2F/DP genes; it might be helpful to improve fruit development and tolerance against abiotic stress through marker-assisted selection or transgenic approaches.

Coordination Structure Modulation in Group-VIB Metal Doped Ag3PO4 Augments Active Site Density for Electrocatalytic Conversion of N2 to NH3.

Doping is considered a promising material engineering strategy in electrochemical nitrogen reduction reaction (NRR), provided the role of the active site is rightly identified. This work concerns the doping of group VIB metal in Ag3PO4 to enhance the active site density, accompanied by d-p orbital mixing at the active site/N2 interface. Doping induces compressive strain in the Ag3PO4 lattice and inherently accompanies vacancy generation, the latter is quantified with positron annihilation lifetime studies (PALS). This eventually alters the metal d-electronic states relative to Fermi level and manipulate the active sites for NRR resulting into side-on N2 adsorption at the interface. The charge density deployment reveals Mo as the most efficient dopant, attaining a minimum NRR overpotential, as confirmed by the detailed kinetic study with the rotating ring disk electrode (RRDE) technique. In fact, the Pt ring of RRDE fails to detect N2H4, which is formed as a stable intermediate on the electrode surface, as identified from in-situ attenuated total reflectance-infrared (ATR-IR) spectroscopy. This advocates the complete conversion of N2 to NH3 on Mo/Ag3PO4-10 and the so-formed oxygen vacancies formed during doping act as proton scavengers suppressing hydrogen evolution reaction resulting into a Faradaic efficiency of 54.8% for NRR.

Deletion of DP148R, DP71L, and DP96R Attenuates African Swine Fever Virus, and the Mutant Strain Confers Complete Protection against Homologous Challenges in Pigs.

The African swine fever virus (ASFV) has caused a devastating pandemic in domestic and wild swine, causing economic losses to the global swine industry. Recombinant live attenuated vaccines are an attractive option for ASFV treatment. However, safe and effective vaccines against ASFV are still scarce, and more high-quality experimental vaccine strains need to be developed. In this study, we revealed that deletion of the ASFV genes DP148R, DP71L, and DP96R from the highly virulent isolate ASFV CN/GS/2018 (ASFV-GS) substantially attenuated virulence in swine. Pigs infected with 104 50% hemadsorbing doses of the virus with these gene deletions remained healthy during the 19-day observation period. No ASFV infection was detected in contact pigs under the experimental conditions. Importantly, the inoculated pigs were protected against homologous challenges. Additionally, RNA sequence analysis showed that deletion of these viral genes induced significant upregulation of the host histone H3.1 gene (H3.1) and downregulation of the ASFV MGF110-7L gene. Knocking down the expression of H3.1 resulted in high levels of ASFV replication in primary porcine macrophages in vitro. These findings indicate that the deletion mutant virus ASFV-GS-Δ18R/NL/UK is a novel potential live attenuated vaccine candidate and one of the few experimental vaccine strains reported to induce full protection against the highly virulent ASFV-GS virus strain. IMPORTANCE Ongoing outbreaks of African swine fever (ASF) have considerably damaged the pig industry in affected countries. Thus, a safe and effective vaccine is important to control African swine fever spread. Here, an ASFV strain with three gene deletions was developed by knocking out the viral genes DP148R (MGF360-18R), NL (DP71L), and UK (DP96R). The results showed that the recombinant virus was completely attenuated in pigs and provided strong protection against parental virus challenge. Additionally, no viral genomes were detected in the sera of pigs housed with animals infected with the deletion mutant. Furthermore, transcriptome sequencing (RNA-seq) analysis revealed significant upregulation of histone H3.1 in virus-infected macrophage cultures and downregulation of the ASFV MGF110-7L gene after viral DP148R, UK, and NL deletion. Our study provides a valuable live attenuated vaccine candidate and potential gene targets for developing strategies for anti-ASFV treatment.

Social and emotional alterations in mice lacking the short dystrophin-gene product, Dp71.

BACKGROUND: The Duchenne and Becker muscular dystrophies (DMD, BMD) are neuromuscular disorders commonly associated with diverse cognitive and behavioral comorbidities. Genotype-phenotype studies suggest that severity and risk of central defects in DMD patients increase with cumulative loss of different dystrophins produced in CNS from independent promoters of the DMD gene. Mutations affecting all dystrophins are nevertheless rare and therefore the clinical evidence on the contribution of the shortest Dp71 isoform to cognitive and behavioral dysfunctions is limited. In this study, we evaluated social, emotional and locomotor functions, and fear-related learning in the Dp71-null mouse model specifically lacking this short dystrophin. RESULTS: We demonstrate the presence of abnormal social behavior and ultrasonic vocalization in Dp71-null mice, accompanied by slight changes in exploratory activity and anxiety-related behaviors, in the absence of myopathy and alterations of learning and memory of aversive cue-outcome associations. CONCLUSIONS: These results support the hypothesis that distal DMD gene mutations affecting Dp71 may contribute to the emergence of social and emotional problems that may relate to the autistic traits and executive dysfunctions reported in DMD. The present alterations in Dp71-null mice may possibly add to the subtle social behavior problems previously associated with the loss of the Dp427 dystrophin, in line with the current hypothesis that risk and severity of behavioral problems in patients increase with cumulative loss of several brain dystrophin isoforms.

Implementation of post-discharge malaria chemoprevention (PDMC) in Benin, Kenya, Malawi, and Uganda: stakeholder engagement meeting report.

A Stakeholder engagement meeting on the implementation of post-discharge malaria chemoprevention (PDMC) in Benin, Kenya, Malawi, and Uganda was held in Nairobi, Kenya, on 27 September 2023. Representatives from the respective National Malaria Control Programmes, the World Health Organization (WHO) Geneva, Africa Regional and Kenya offices, research partners, non-governmental organizations, and the Medicines for Malaria Venture participated. PDMC was recommended by the WHO in June 2022 and involves provision of a full anti-malarial treatment course at regular intervals during the post-discharge period in children hospitalized with severe anaemia in areas of moderate-to-high malaria transmission. The WHO recommendation followed evidence from a meta-analysis of three clinical trials and from acceptability, delivery, cost-effectiveness, and modelling studies. The trials were conducted in The Gambia using monthly sulfadoxine-pyrimethamine during the transmission season, in Malawi using monthly artemether-lumefantrine, and in Kenya and Uganda using monthly dihydroartemisinin-piperaquine, showing a significant reduction in all-cause mortality by 77% (95% CI 30-98) and a 55% (95% CI 44-64) reduction in all-cause hospital readmissions 6 months post-discharge. The recommendation has not yet been implemented in sub-Saharan Africa. There is no established platform for PDMC delivery. The objectives of the meeting were for the participating countries to share country contexts, plans and experiences regarding the adoption and implementation of PDMC and to explore potential delivery platforms in each setting. The meeting served as the beginning of stakeholder engagement within the PDMC Saves Lives project and will be followed by formative and implementation research to evaluate alternative delivery strategies in selected countries. Meeting highlights included country consensus on use of dihydroartemisinin-piperaquine for PDMC and expansion of the target group to "severe anaemia or severe malaria", in addition to identifying country-specific options for PDMC delivery for evaluation in implementation research. Further exploration is needed on whether the age group should be extended to school-age children.

Decolorization of RhB on three-dimensional porous CeO2/LaFeO3/SrTiO3 catalyst via photo-fenton-like catalysis.

The catalysts with three-dimensional porous (3DP) CeO2, LaFeO3 and SrTiO3 are synthesized by sol-gel method and chemical precipitation method. The resulting multi-component 3DP CeO2/LaFeO3/SrTiO3 composite material featured a high specific surface area (26.08 m2/g), which can provide more surface active sites to improve adsorption capacity and catalytic performance. The photocatalytic, Fenton-like, photo-Fenton-like performance of the catalyst are studied on decolorization of RhB under UV irradiation, respectively. 3DP CeO2/LaFeO3/SrTiO3 exhibits high catalytic performance. Compared with photocatalytic or Fenton-like performance, 3DP CeO2/LaFeO3/SrTiO3 catalyst exhibits higher photo-Fenton-like performance, facilitating efficient decolorization of the rhodamine B. Moreover, the initial reaction rate on decolorization of RhB with 3DP CeO2/LaFeO3/SrTiO3 is 10.55, 5.52, 3.67 and 1.51 times higher than that with SrTiO3, LaFeO3, 3DP CeO2 and 3DP CeO2/LaFeO3, respectively. Meanwhile, 3DP LaFeO3/CeO2/SrTiO3 has a wider pH usage range in the synergistic reaction. Finally, a catalytic mechanism for the decolorization of rhodamine B is proposed. The continuous cycling of Fe3+/Fe2+ and Ce4+/Ce3+ and the production of active substances are achieved under the photo-Fenton-like effect of the catalyst.

Matrix gla protein, a potential marker of tissue remodelling and physiological ageing of the gut in crohn's disease.

BACKGROUND: The inactive dephosphorylated and uncarboxylated form of the matrix Gla protein (dp-ucMGP) has been shown to be increased in plasma of inflammatory bowel disease (IBD) patients. Our aim was to assess if the plasmatic level of dp-ucMGP could reflect disease endoscopic activity, presence of strictures and cumulative structural bowel damage in Crohn's disease (CD) patients. METHODS: The plasmatic level of dp-ucMGP was measured in a monocentric cohort of prospectively recruited patients. The analysis was done by chemiluminescent immunoassay on blood samples collected the day of a planned ileocolonoscopy. In addition to classical clinical data (gender, age, body mass index (BMI), disease duration, current treatment), endoscopic data (disease location, Crohn's Disease Endoscopic Index of Severity (CDEIS), mucosal healing (MH), presence of 9 CD lesion types) and biological markers (faecal calprotectin and C-reactive protein (CRP)) were collected. The association between dp-ucMGP level and Lémann index was also investigated. Univariate linear regression was used to investigate the relationship between dp-ucMGP level and different parameters collected. RESULTS: A total of 82 ileocolonoscopies and dp-ucMGP assays were performed in 75 CD patients (45 females; 37 ileocolonic, 19 ileal and 19 colonic diseases) between October 2012 and November 2019. A total of 24 patients (29.3%) showed MH. The dp-ucMGP levels were not associated with MH, CDEIS, faecal calprotectin or CRP levels. Plasmatic dp-ucMGP levels increased significantly with age (p = 0.0032), disease duration (p = 0.0033), corticosteroids use (p = 0.019) and tended to increase in patients with intestinal strictures (p = 0.086) but not with the Lémann index. CONCLUSION: The significant increase of plasmatic dp-ucMGP levels with age, disease duration and the trend observed in patients with non-ulcerated strictures may suggest that this extracellular matrix protein could be a marker of tissue remodelling and physiological ageing of the gut.

Penicimides A and B, two novel diels-alder [4 + 2] cycloaddition ergosteroids from Penicillium herquei.

Two novel naturally occurring [4 + 2] Diels-Alder cycloaddition ergosteroids (1 and 2), three undescribed oxidized ergosteroids (3-5), and eleven known analogs (6-16) were isolated from Penicillium herquei. Compounds 1 and 2 represent the first reported cycloadducts of a steroid with 1,4,6-trimethyl-1,6-dihydropyridine-2,5-dione or 4,6-dimethyl-1,6-dihydropyridine-2,5-dione to date. Compound 3 is the C-15 epimer of (22E,24R)-9α,11ß-dihydroxyergosta-4,6,8(14),22-tetraen-3-one (14). The chemical structures of these compounds were elucidated through widespread spectroscopic analyses, mainly including HRESIMS and 1D and 2D NMR data, calculated 13C NMR-DP4+ analysis, and electronic circular dichroism (ECD) data analyses. Biological evaluations of Compounds 1-16 revealed that 3, 9-11, and 15 inhibited the production of NO in LPS-induced RAW264.7 cells with an IC50 value from 7.37 ± 0.69 to 38.9 ± 2.25 µM (the positive control dexamethasone IC50: 9.54 ± 0.71 µM). In addition, Compound 3 exhibited a potent inhibitory effect on the secretion of the proinflammatory cytokines TNF-α and IL-6, the transcription level of the proinflammatory macrophage markers TNF-α, and the expression of the iNOS protein.

Preparation and epitope analysis of monoclonal antibodies against African swine fever virus DP96R protein.

BACKGROUND: Many proteins of African swine fever virus (ASFV, such as p72, p54, p30, CD2v, K205R) have been successfully expressed and characterized. However, there are few reports on the DP96R protein of ASFV, which is the virulence protein of ASFV and plays an important role in the process of host infection and invasion of ASFV. RESULTS: Firstly, the prokaryotic expression vector of DP96R gene was constructed, the prokaryotic system was used to induce the expression of DP96R protein, and monoclonal antibody was prepared by immunizing mice. Four monoclonal cells of DP96R protein were obtained by three ELISA screening and two sub-cloning; the titer of ascites antibody was up to 1:500,000, and the monoclonal antibody could specifically recognize DP96R protein. Finally, the subtypes of the four strains of monoclonal antibodies were identified and the minimum epitopes recognized by them were determined. CONCLUSION: Monoclonal antibody against ASFV DP96R protein was successfully prepared and identified, which lays a foundation for further exploration of the structure and function of DP96R protein and ASFV diagnostic technology.

DP7-C/mir-26a system promotes bone regeneration by remodeling the osteogenic immune microenvironment.

OBJECTIVE: This study investigates the DP7-C/miR-26a complex as a stable entity resulting from the combination of miR-26a with the immunomodulatory peptide DP7-C. Our focus is on utilizing DP7-C loaded with miR-26a to modulate the immune microenvironment in bone and facilitate osteogenesis. METHODS: The DP7-C/miR-26a complex was characterized through transmission electron microscopy, agarose electrophoresis, and nanoparticle size potentiometer analysis. Transfection efficiency and cytotoxicity of DP7-C were assessed using flow cytometry and the CCK-8 assay. We validated the effects of DP7-C/miR-26a on bone marrow mesenchymal stem cells (BMSCs) and macrophages RAW 264.7 through gene expression and protein synthesis assays. A comprehensive evaluation of appositional bone formation involved micro-CT imaging, histologic analysis, and immunohistochemical staining. RESULTS: DP7-C/miR-26a, a nanoscale, and low-toxic cationic complex, demonstrated the ability to enter BMSCs and RAW 264.7 via distinct pathways. The treatment with DP7-C/miR-26a significantly increased the synthesis of multiple osteogenesis-related factors in BMSCs, facilitating calcium nodule formation in vitro. Furthermore, DP7-C/miR-26a promoted M1 macrophage polarization toward M2 while suppressing the release of inflammatory factors. Coculture studies corroborated these findings, indicating significant repair of rat skull defects following treatment with DP7-C/miR-26a. CONCLUSION: The DP7-C/miR-26a system offers a safer, more efficient, and feasible technical means for treating bone defects.

A retrospective study of the effects of a vasopressor bolus on systolic slope (dP/dt) and dynamic arterial elastance (Eadyn).

BACKGROUND: To enhance the utility of functional hemodynamic monitoring, the variables systolic slope (dP/dt) and dynamic arterial elastance (Eadyn) are calculated by the Hypotension Prediction Index (HPI) Acumen® Software. This study was designed to characterize the effects of phenylephrine and ephedrine on dP/dt and Eadyn. METHODS: This was a retrospective, non-randomized analysis of data collected during two clinical studies. All patients required intra-operative controlled mechanical ventilation and had an indwelling radial artery catheter connected to an Acumen IQ sensor. Raw arterial pressure waveform data was downloaded from the patient monitor and all hemodynamic measurements were calculated off-line. The anesthetic record was reviewed for bolus administrations of either phenylephrine or ephedrine. Cardiovascular variables prior to drug administration were compared to those following vasopressor administrations. The primary outcome was the difference for dP/dt and Eadyn at baseline compared with the average after the bolus administration. All data sets demonstrated non-normal distributions so statistical analysis of paired and unpaired data followed the Wilcoxon matched pairs signed-rank test or Mann-Whitney U test, respectively. RESULTS: 201 doses of phenylephrine and 100 doses of ephedrine were analyzed. All data sets are reported as median [95% CI]. Mean arterial pressure (MAP) increased from 62 [54,68] to 78 [76,80] mmHg following phenylephrine and from 59 [55,62] to 80 [77,83] mmHg following ephedrine. Stroke volume and cardiac output both increased. Stroke volume variation and pulse pressure variation decreased. Both drugs produced significant increases in dP/dt, from 571 [531, 645] to 767 [733, 811] mmHg/sec for phenylephrine and from 537 [509, 596] to 848 [779, 930] mmHg/sec for ephedrine. No significant changes in Eadyn were observed. CONCLUSION: Bolus administration of phenylephrine or ephedrine increases dP/dt but does not change Eadyn. dP/dt demonstrates potential for predicting the inotropic response to phenylephrine or ephedrine, providing guidance for the most efficacious vasopressor when treating hypotension. TRIAL REGISTRATION: Data was collected from two protocols. The first was deemed to not require written, informed consent by the Institutional Review Board (IRB). The second was IRB-approved (Effect of Diastolic Dysfunction on Dynamic Cardiac Monitors) and registered on ClinicalTrials.gov (NCT04177225).