Production of α-amylase from gluconate and carbon dioxide by protein synthesis and secretion optimization in Cupriavidus necator H16.

Chemoautotrophic Cupriavidus necator H16 has a strong protein synthesis ability and has been used to produce intracellular protein products. However, studies optimizing its secretion system and the producing extracellular enzyme products (EEPs) are lacking. Here, we focused on investigating the feasibility of synthesizing and secreting EEPs in C. necator H16, using α-amylase as a prototype. α-Amylase expression optimization, genome modification, and secretion system engineering were performed to construct and optimize the α-amylase-producing engineering C. necator H16. Finally, the optimized engineering strain could produce α-amylase, with the α-amylase activity per unit cells reaching up to 5.54 U/OD600 using gluconate as substrate, which was 29.2-fold compared with that of initial engineering strain. Additionally, when using carbon dioxide as substrate, the α-amylase activity per unit cells of engineered strain reached 4.26 U/OD600. Overall, this study demonstrates the feasibility of developing C. necator H16 as a host for autotrophic production of α-amylase.

Technical specification for developing a clinical practice guideline for the integration of traditional Chinese medicine and Western medicine.

Developing a clinical practice guideline (CPG) for integrating traditional Chinese medicine (TCM) and Western medicine (WM) requires the accurate identification, collation, and integration of all available evidence on TCM and WM in a comprehensive, meaningful, and resource-efficient manner. This entails framing appropriate clinical questions, retrieving and synthesizing evidence from multiple resources, and providing concise and complete recommendations for specific diseases. However, some existing CPGs for integrating TCM and WM lack deep and organic integration. As the effective preparation of a CPG for integrating TCM and WM typically involves a complex set of principles, methodology, and steps, we believe that a cohesive, step-by-step guide on how to prepare a CPG for integrating TCM and WM is essential. To facilitate the design and development of a robust CPG, we present a clear and concise methodology, outlining relevant principles and procedures, supported by references for guidance. This technical specification aims to simplify the methodology for preparing a CPG for integrating TCM and WM; provide healthcare professionals and researchers with methodologically sound tools; and enhance the quality of CPGs for integrating TCM and WM. This technical specification may help elucidate this complex process, facilitate evaluation of the quality of published CPGs for integrating TCM and WM, and improve the understanding and application of recommendations for the combined and integrated use of TCM and WM in a new system.

IL-17A-neutralizing antibody ameliorates inflammation and fibrosis in rosacea by antagonizing the CXCL5/CXCR2 axis.

Rosacea is a chronic inflammatory skin disorder that can lead to fibrosis. However, the mechanisms underlying fibrosis in the later stages of rosacea have been less thoroughly investigated. Interleukin-17A (IL-17A) has been implicated in both inflammation and organ fibrosis; however, the effectiveness and mechanism of IL-17A-neutralizing antibodies in the later stages of rosacea-related fibrosis remain unclear. In this study, we induced rosacea-like lesions in mice using LL-37 and administered IL-17A-neutralizing antibodies. The results indicated that the IL-17A-neutralizing antibodies alleviated skin damage, reduced skin thickness, and decreased the secretion of inflammatory factors (TNF-α, CAMP, TLR4, P-NF-kB), angiogenesis-related factors (CD31, VEGF), and the TGF-ß1 signaling pathway, along with factors associated with epithelial-mesenchymal transition and the deposition of fibrosis-related proteins (COL1) in the rosacea-like mouse models. Furthermore, the IL-17A-neutralizing antibodies effectively diminished the expression of IL-17, IL-17R, CXCL5, and CXCR2 in the skin. Our findings demonstrate that IL-17A-neutralizing antibodies inhibit the activation of the CXCL5/CXCR2 axis in rosacea-like skin tissue, thereby ameliorating inflammation and fibrosis associated with the condition.

A novel method for quality evaluation of Radix Angelica sinensis based on molecularly imprinted electrochemical sensor.

Rapid and sensitive detection of n-butylidenephthalide (NBP) is crucial for quality control of Radix Angelica Sinensis (RAS) and its related pharmaceuticals due to their shared pharmacological effects, such as immune enhancement and anti-tumor properties. Current detection methods struggle to quantify NBP quickly and accurately. A molecularly imprinted polymer (MIP)-based electrochemical sensor has been developed, forming a film on PCN-222(Fe) via electropolymerization for the rapid and selective detection of NBP. o-Phenylenediamine (o-PD) was polymerized onto PCN-222(Fe), utilizing its high surface area and porous structure to create a high-performance MIP (MIP/PCN-222(Fe)) sensor. This sensor detects NBP binding at the molecularly imprinted sites through a redox probe, with current changes reflecting the NBP content in the sample. This sensor exhibits a strong affinity for NBP, with a linear detection range from 200 nM to 1 mM, a detection limit of 76 nM, and high specificity towards similar phthalide compounds. Experimental results show that the MIP/PCN-222(Fe) sensor can accurately quantify NBP in real samples, offering a simplified method with promising applications for RAS quality evaluation.

Clinical practice guidelines of Chinese patent medicine in China: A critical review.

OBJECTIVES: To analyze the methodology, evidence, recommendations, quality, and implementation of traditional Chinese patent medicine (CPM) guidelines. METHODS: We retrieved clinical application guidelines of CPM published from 2019 to 2022. Independent screening and data extraction were performed by two evaluators. The basic information about the guidelines, including evidence and recommendations, were extracted and statistically analyzed. Quality and implementation were evaluated using the Implementation Evaluation Tool and Appraisal of Guidelines for Research & Evaluation (AGREE) II. RESULTS: In total, 29 guidelines were analyzed, including 262 recommendations and 2308 references. All the CPM guidelines followed the principle of "evidence as a core, consensus as a supplement, and experience as a reference" and the methods provided by WHO Handbook. An average of 89 references were cited in each guideline and 8 in each recommendation. Randomized controlled trials and systematic reviews constituted 89 % and 0.9 %, respectively, of all references. Low or very low-quality evidence characterized 74.5 % and weak recommendations characterized 83.6 %. Of all recommendations, 13.7 % were based on expert consensus, and 9.5 % of strong recommendations were based on low or very low-quality evidence. The AGREE II scores for each domain were: scope and purpose (79.63 %) and editorial independence (79.27 %), followed by clarity of presentation (72.59 %), stakeholder involvement (69.99 %), rigor of development (53.97 %) and applicability (5.11 %). The implementation quality of most guidelines was either high (44.8 %) or moderate (55.2 %). CONCLUSIONS: The results for CPM guidelines were impressive in terms of methodology, quality, and implementation. However, confidence in CPM recommendations was downgraded by low quality of evidence.

Effect of macrophage-to-myofibroblast transition on silicosis.

BACKGROUND: The aim was to explore the effect of macrophage polarization and macrophage-to-myofibroblast transition (MMT) in silicosis. METHODS: Male Wistar rats were divided into a control group and a silicosis group developed using a HOPE MED 8050 dynamic automatic dusting system. Murine macrophage MH-S cells were randomly divided into a control group and an SiO2 group. The pathological changes in lung tissue were observed using hematoxylin and eosin (HE) and Van Gieson (VG) staining. The distribution and location of macrophage marker (F4/80), M1 macrophage marker (iNOS), M2 macrophage marker (CD206), and myofibroblast marker (α-smooth muscle actin [α-SMA]) were detected using immunohistochemical and immunofluorescent staining. The expression changes in iNOS, Arg, α-SMA, vimentin, and type I collagen (Col I) were measured using Western blot. RESULTS: The results of HE and VG staining showed obvious silicon nodule formation and the distribution of thick collagen fibers in the lung tissue of the silicosis group. Macrophage marker F4/80 increased gradually from 8 to 32 weeks after exposure to silica. Immunohistochemical and immunofluorescent staining results revealed that there were more iNOS-positive cells and some CD206-positive cells in the lung tissue of the silicosis group at 8 weeks. More CD206-positive cells were found in the silicon nodules of the lung tissues in the silicosis group at 32 weeks. Western blot analysis showed that the expressions of Inducible nitric oxide synthase and Arg protein in the lung tissues of the silicosis group were upregulated compared with those of the control group. The results of immunofluorescence staining showed the co-expression of F4/80, α-SMA, and Col I, and CD206 and α-SMA were co-expressed in the lung tissue of the silicosis group. The extracted rat alveolar lavage fluid revealed F4/80+α-SMA+, CD206+α-SMA+, and F4/80+α-SMA+Col I+ cells using immunofluorescence staining. Similar results were also found in MH-S cells induced by SiO2. CONCLUSIONS: The development of silicosis is accompanied by macrophage polarization and MMT.

Co-occurrence of Erdheim-Chester disease and clonally evolving acute myeloid leukemia with FLT3-ITD and PTPN11 mutations.

Erdheim-Chester disease (ECD) is a rare histiocytosis that tends to co-exist with other myeloid malignancies. Here, we use genetic and transcriptomic sequencing to delineate a case of co-occurring BRAFV600E-mutated ECD and acute myeloid leukemia (AML), followed by AML remission and relapse. The AML relapse involved the extinction of clones with KMT2A-AFDN and FLT3-ITD, and the predominance of PTPN11-mutated subclones with distinct transcriptomic features. This case report has highlighted the screening for other myeloid malignancies at the diagnosis of ECD and the clinical significance of PTPN11-mutated AML subclones that require meticulous monitoring.

Paeoniflorin mitigates MMP-12 inflammation in silicosis via Yang-Yin-Qing-Fei Decoction in murine models.

BACKGROUND: Silicosis presents a significant clinical challenges and economic burdens, with Traditional Chinese Medicine (TCM) emerging as a potential therapeutic avenue. However, the precise effects and mechanisms of TCM in treating silicosis remain uncertain and subject to debate. OBJECTIVE: The study aims to elucidate the therapeutic role and mechanisms of the Yang-Yin-Qing-Fei Decoction (YYQFD) and its key component, paeoniflorin, in silicosis using a murine model. METHODS: Silicotic mice were treated with YYQFD, pirfenidone (PFD), or paeoniflorin. RAW264.7 cells and mouse lung fibroblasts (MLF) were stimulated with silica, matrix metalloproteinase-12 (MMP-12), or TGF-ß1, followed by treatment with paeoniflorin, PFD, or relevant inhibitors. YYQFD constituents were characterized using High-Performance Liquid Chromatography (HPLC). Lung fibrosis severity was assessed via histopathological examination, micro-CT imaging, lung functions, and Western blot analysis. Transcriptome sequencing and bioinformatics analysis were employed to delineate the gene expression profile and target genes modulated by YYQFD in silicosis. RESULTS: Treatment with YYQFD ameliorated silica-induced lung fibrosis. Transcriptome sequencing identified MMP-12 as a potential common target of YYQFD and PFD. Additionally, a potential pro-inflammatory role of MMP-12, regulated by silica-induced TLR4 signaling pathways, was revealed. Paeoniflorin, one of the most distinctive compounds in YYQFD, attenuated silica-induced MMP-12 increase and its derived inflammatory factors in macrophages through a direct binding effect. Notably, paeoniflorin treatment exerted anti-fibrotic effects by inhibiting MMP-12-derived inflammatory factors and TGF-ß1-induced myofibroblast differentiation in silica-exposed mice. CONCLUSIONS: This study underscores paeoniflorin as one of the most principal bioactive compounds in YYQFD, highlighting its capacity to attenuate lung inflammation driven by macrophage-derived MMP-12 and reduce lung fibrosis both in vivo and in vitro.

Mitochondria-Targeted Prodrug Nanoassemblies for Efficient Ferroptosis-Based Therapy via Devastating Ferroptosis Defense Systems.

Ferroptosis is a form of regulated cell death accompanied by lipid reactive oxygen species (ROS) accumulation in an iron-dependent manner. However, the efficiency of tumorous ferroptosis was seriously restricted by intracellular ferroptosis defense systems, the glutathione peroxidase 4 (GPX4) system, and the ubiquinol (CoQH2) system. Inspired by the crucial role of mitochondria in the ferroptosis process, we reported a prodrug nanoassembly capable of unleashing potent mitochondrial lipid peroxidation and ferroptotic cell death. Dihydroorotate dehydrogenase (DHODH) inhibitor (QA) was combined with triphenylphosphonium moiety through a disulfide-containing linker to engineer well-defined nanoassemblies (QSSP) within a single-molecular framework. After being trapped in cancer cells, the acidic condition provoked the structural disassembly of QSSP to liberate free prodrug molecules. The mitochondrial membrane-potential-driven accumulation of the lipophilic cation prodrug was delivered explicitly into the mitochondria. Afterward, the thiol-disulfide exchange would occur accompanied by downregulation of reduced glutathione levels, thus resulting in mitochondria-localized GPX4 inactivation for ferroptosis. Simultaneously, the released QA from the hydrolysis reaction of the adjacent ester bond could further devastate mitochondrial defense and evoke robust ferroptosis via the DHODH-CoQH2 system. This subcellular targeted nanoassembly provides a reference for designing ferroptosis-based strategy for efficient cancer therapy through interfering antiferroptosis systems.

Long-Term Outcomes of Autologous Stem Cell Transplantation in Patients with Newly Diagnosed POEMS Syndrome.

POEMS (polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin changes) syndrome is a rare form of plasma cell dyscrasia often treated with high-dose chemotherapy and autologous stem cell transplantation (ASCT). ASCT has resulted in satisfactory and sustained therapeutic outcomes. However, a substantial number of patients eventually experience disease progression, requiring second-line treatment. Therefore, it would be of further benefit to identify patients who will acquire the best long-term survival after ASCT. The aim of this study was to fully reveal the outcomes of patients undergoing ASCT in a large series with long-term follow-up. Long-term outcomes of 239 patients with newly diagnosed POEMS syndrome undergoing ASCT at a single center were evaluated retrospectively. Rates of hematologic complete response (CRH) and vascular endothelial growth factor (VEGF) complete response (CRV) were 57.3% and 68.6%, respectively, with 90.5% of patients achieving an overall clinical response. At a median follow-up of 94 months, the 5-year overall survival (OS) rate was 92.8%, and the 5-year time to next-line treatment (TTNT) rate was 72.2% (median TTNT, 96 months). Patients achieving CRH (5-year TTNT rate, 82.5% versus 60.7%; P < .0001) or CRV (5-year TTNT rate 83.7% versus 54.2%; P < .0001) had better survival outcomes compared to non-CR group patients. Dual hematologic and VEGF complete responses carry further benefit for survival (median TTNT, 129 months versus 68 months; P < .0001). Seven cases of second primary malignancy were recorded, all of which were solid tumors. Front-line ASCT resulted in excellent long-term survival in patients with POEMS syndrome, with the best survival observed in those achieving dual hematologic and VEGF CRs.

Exercise training inhibits macrophage-derived IL-17A-CXCL5-CXCR2 inflammatory axis to attenuate pulmonary fibrosis in mice exposed to silica.

Exposure to crystalline silica leads to health effects beyond occupational silicosis. Exercise training's potential benefits on pulmonary diseases yield inconsistent outcomes. In this study, we utilized experimental silicotic mice subjected to exercise training and pharmacological interventions, including interleukin-17A (IL-17A) neutralizing antibody or clodronate liposome for macrophage depletion. Findings reveal exercise training's ability to mitigate silicosis progression in mice by suppressing scavenger receptor B (SRB)/NOD-like receptor thermal protein domain associated protein 3 (NLRP3) and Toll-like receptor 4 (TLR4) pathways. Macrophage-derived IL-17A emerges as primary source and trigger for silica-induced pulmonary inflammation and fibrosis. Exercise training effectively inhibits IL-17A-CXC motif chemokine ligand 5 (CXCL5)-Chemokine (C-X-C motif) Receptor 2 (CXCR2) axis in silicotic mice. Our study evidences exercise training's potential to reduce collagen deposition, preserve elastic fibers, slow pulmonary fibrosis advancement, and enhance pulmonary function post silica exposure by impeding macrophage-derived IL-17A-CXCL5-CXCR2 axis.

ISRIB inhibits the senescence of type II pulmonary epithelial cells to alleviate pulmonary fibrosis induced by silica in mice.

The role and mechanisms of integrated stress response inhibitor (ISRIB) on silicosis are still not well defined. In the present study, the effects of ISRIB on cellular senescence and pulmonary fibrosis in silicosis were evaluated by RNA sequencing, micro-computed tomography, pulmonary function assessment, histological examination, and Western blot analysis. The results showed that ISRIB significantly reduced the degree of pulmonary fibrosis in mice with silicosis and reduced the expression of type I collagen, fibronectin, α-smooth muscle actin, and transforming growth factor-ß1. Both in vivo and in vitro results showed that ISRIB reversed the expression of senescence-related factors ß-galactosidase, phosphor-ataxia telangiectasia mutated, phosphor-ataxia telangiectasia and Rad3-related protein, p-p53, p21, p16, and plasminogen activator inhibitor type 1. The aforementioned results were consistent with the sequencing results. These findings implied that ISRIB might reduce the degree of pulmonary fibrosis in mice with silicosis by inhibiting the cellular senescence of alveolar epithelial cell type II.

A Label-Free and Antibody-Free Molecularly Imprinted Polymer-Based Impedimetric Sensor for NSCLC-Cells-Derived Exosomes Detection.

In this study, a label-free and antibody-free impedimetric biosensor based on molecularly imprinting technology for exosomes derived from non-small-cell lung cancer (NSCLC) cells was established. Involved preparation parameters were systematically investigated. In this design, with template exosomes anchored on a glassy carbon electrode (GCE) by decorated cholesterol molecules, the subsequent electro-polymerization of APBA and elution procedure afforded a selective adsorption membrane for template A549 exosomes. The adsorption of exosomes caused a rise in the impedance of the sensor, so the concentration of template exosomes can be quantified by monitoring the impedance of GCEs. Each procedure in the establishment of the sensor was monitored with a corresponding method. Methodological verification showed great sensitivity and selectivity of this method with an LOD = 2.03 × 103 and an LOQ = 4.10 × 104 particles/mL. By introducing normal cells and other cancer cells derived exosomes as interference, high selectivity was proved. Accuracy and precision were measured, with an obtained average recovery ratio of 100.76% and a resulting RSD of 1.86%. Additionally, the sensors' performance was retained at 4 °C for a week or after undergoing elution and re-adsorption cycles seven times. In summary, the sensor is competitive for clinical translational application and improving the prognosis and survival for NSCLC patients.

Long-Term Administration of LL-37 Can Induce Irreversible Rosacea-like Lesion.

Rosacea is a chronic inflammatory skin disease whose late manifestations have not yet been clearly reported in animal models. The objective of this study is to describe the skin lesions and major histopathological changes in a rosacea-like phenotype in mice induced by prolonged LL-37 administration and furthermore, to assess the potential of long-term LL-37 administration in inducing irreversible rosacea-like skin lesion models. Balb/c mice were continuously injected intradermally with LL-37 every 12 h to induce a rosacea-like phenotype. After LL-37 injections were administered for 20 consecutive days, the area of rosacea-like lesions gradually expanded in the first 13 days, then entered a stable phase. Haematoxylin and eosin (H&E) and Van Gieson's staining showed a high degree of inflammatory cell aggregation, thickening of the epidermis and dermis, and collagen deposition in large quantities. The results of immunofluorescence staining and Western blotting showed that the expression of α-SMA, TNF-α, vimentin, and COL1 in the skin of mice was significantly upregulated. Short-term LL-37 administration induced rosacea-like lesions that only featured the aggregation of inflammatory factors and thickening of the epidermis, whereas no collagen hyperplasia was observed, and a full recovery was noticed. However, rosacea-like skin lesions induced by long-term LL-37 administration did not completely recover. Our study compares rosacea-like lesions induced by short-term versus long-term LL-37 administration, and the results suggest that irreversible rosacea-like lesions can be induced by long-term LL-37 administration.

Quercetin Alleviates Pulmonary Fibrosis in Silicotic Mice by Inhibiting Macrophage Transition and TGF-ß-Smad2/3 Pathway.

Silicosis is a pulmonary disease caused by the inhalation of silica. There is a lack of early and effective prevention, diagnosis, and treatment methods, and addressing silicotic fibrosis is crucial. Quercetin, a flavonoid with anti-carcinogenic, anti-inflammatory, and antiviral properties, is known to have a suppressive effect on fibrosis. The present study aimed to determine the therapeutic effect of quercetin on silicotic mice and macrophage polarity. We found that quercetin suppressed silicosis in mice. It was observed that SiO2 activated macrophage polarity and the macrophage-to-myofibroblast transition (MMT) by transforming the growth factor-ß (TGF-ß)-Smad2/3 signaling pathway in silicotic mice and MH-S cells. Quercetin also attenuated the MMT and the TGF-ß-Smad2/3 signaling pathway in vivo and in vitro. The present study demonstrated that quercetin is a potential therapeutic agent for silicosis, which acts by regulating macrophage polarity and the MMT through the TGF-ß-Smad2/3 signaling pathway.

Kinesin family member 3A induces related diseases via wingless-related integration site/ß-catenin signaling pathway.

Kinesin family member 3A is an important motor protein that participates in various physiological and pathological processes, including normal tissue development, homeostasis maintenance, tumor infiltration, and migration. The wingless-related integration site/ß-catenin signaling pathway is essential for critical molecular mechanisms such as embryonic development, organogenesis, tissue regeneration, and carcinogenesis. Recent studies have examined the molecular mechanisms of kinesin family member 3A, among which the wingless-related integration site/ß-catenin signaling pathway has gained attention. The interaction between kinesin family member 3A and the wingless-related integration site/ß-catenin signaling pathway is compact and complex but is fascinating and worthy of further study. The upregulation and downregulation of kinesin family member 3A influence many diseases and patient survival through the wingless-related integration site/ß-catenin signaling pathway. Therefore, this review mainly focuses on describing the kinesin family member 3A and wingless-related integration site/ß-catenin signaling pathways and their associated diseases.

Signaling pathways, microenvironment, and targeted treatments in Langerhans cell histiocytosis.

Langerhans cell histiocytosis (LCH) is an inflammatory myeloid malignancy in the "L-group" histiocytosis. Mitogen-activated protein kinase (MAPK) pathway activating mutations are detectable in nearly all LCH lesions. However, the pathogenic roles of MAPK pathway activation in the development of histiocytosis are still elusive. This review will summarize research concerning the landscape and pathogenic roles of MAPK pathway mutations and related treatment opportunities in Langerhans cell histiocytosis. Video abstract.