Modified Ganlu Xiaodu decoction attenuates airway barrier injury in Mycoplasma pneumoniae pneumonia by inhibiting ZBP1-mediated PANoptosis.

To compare clinical and imaging features in children with macrolide-susceptible (MSMP) and macrolide-resistant Mycoplasma pneumoniae pneumonia (MRMP) in a real-world clinical setting. We retrospectively analysed paediatric M. pneumoniae (MP) cases at Hunan People's Hospital from June to December 2023, utilizing polymerase chain reaction (PCR) to categorize infections into MRMP and MSMP groups based on 23S rRNA mutations. We compared symptoms, laboratory findings, imaging results and treatment outcomes between MRMP and MSMP groups. Additionally, univariate and multivariate logistic regression analyses were performed to identify factors associated with the switch to doxycycline in MRMP patients. Of 199 children with MP, 151 were categorized as MRMP due to 23S rRNA mutations, while 48 were classified as MSMP. The MRMP group included more preschool- and school-aged children, with more severe symptoms (persistent high fever and extensive lung consolidation). MRMP patients received longer antibiotic courses, with imaging showing segmental or lobar infiltrates. Consequently, treatment was switched from azithromycin to doxycycline in 66.2% of MRMP patients. Multivariate logistic regression analysis revealed that segmental or lobar parenchymal infiltration was independently associated with an increased likelihood of receiving doxycycline (OR = 3.942, p < 0.001). These real-world data suggest that MRMP may present with more severe clinical manifestations and radiological features than MSMP among hospitalized children during an atypical post-COVID resurgence of MP. Doxycycline could be considered as the second-line therapy for MRMP patients exhibiting severe symptoms or lung consolidation when macrolides fail to achieve clinical improvement.

Purpose Macrolide-resistant Mycoplasma pneumoniae pneumonia (MPP) is characterized by prolonged fever and radiological progression despite macrolide treatment. Few studies have examined serum procalcitonin (PCT) level in children with MPP. We aimed to investigate the association of acute inflammation markers including PCT with clinical parameters in children with MPP. Methods A total of 147 children were recruited. The diagnosis of MPP relied on serial measurement of IgM antibody against mycoplasma and/or polymerase chain reaction. We evaluated the relationships between C-reactive protein (CRP), PCT, and lactate dehydrogenase (LDH) levels and white blood cell (WBC) counts, and clinical severity of the disease. We used multivariate logistic regression analysis to estimate the odds ratio for prolonged fever (>3 days after admission) and hospital stay (> 6 days), comparing quintiles 2–5 of the PCT levels with the lowest quintile. Results The serum PCT and CRP levels were higher in children with fever and hospital stay than in those with fever lasting ≤ 3 days after admission and hospital stay ≤ 6 days. CRP level was higher in segmental/lobar pneumonia than in bronchopneumonia. The LDH level and WBC counts were higher in children with fever lasting for >3 days before compared to those with fever lasting for ≤ 3 days. The highest quintile of PCT levels was associated with a significantly higher risk of prolonged fever and/or hospital stay than the lowest quintile. Conclusion Serum PCT and CRP levels on admission day were associated with persistent fever and longer hospitalization in children with MPP.

Purpose: This study was performed to investigate differences in clinical, laboratory, and radiologic features of clinical course between macrolide-resistant Mycoplasma pneumoniae (MRMP) and macrolide-sensitive Mycoplasma pneumoniae (MSMP) pneumonia in children.Methods: A total of 146 children hospitalized with Mycoplasma pneumoniae (MP) pneumonia in a tertiary hospital were enrolled in this study.Information on the characteristics was retrospectively obtained using electronic medical records.Results: The duration of fever and hospitalization was significantly longer and respiratory virus coinfection was more common in the MRMP group than in the MSMP group (47.0% vs. 24.1%,P= 0.026).No significant intergroup difference in the laboratory findings, except monocytes (%), was observed.The proportion of patients with pneumonic lesions involving more than half of the whole lung was higher (28.2% vs. 10.3%), with a poorer response to the stepwise treatment for MP pneumonia, in the MRMP group than in the MSMP pneumonia group.MRMP pneumonia in children may be associated with a higher disease burden than MSMP pneumonia.Conclusion: MRMP pneumonia in children might be associated with a more increased disease burden than MSMP pneumonia.This study suggested the need of distinguished therapeutic approaches to reduce the disease burden due to MRMP pneumonia in children.

Multi-approach study on diethylhexyl phthalate and monoethylhexyl phthalate binding to lysozyme: In silico, bioactivity and surface plasmon resonance analyses.

Objective High mortality rates are linked to acute respiratory distress syndrome (ARDS), a prevalent type of respiratory failure. Amid the COVID-19 outbreak in particular, a viable defensive method is provided by traditional Chinese medicine (TCM). This study investigated whether chlorogenic acid (CGA), a primary component of honeysuckle, could protect against ARDS. Methods We employed network pharmacology to explore the honeysuckle and ARDS component-target-disease network, and enrichment function analysis to uncover the potential mechanisms of honeysuckle in treating ARDS. LPS-induced ARDS rat models (each group rats n = 6) were used for validation, and the CGA treatments group were was administered by gavage at 100 mg/kg. including flow cytometry for T cell subsets, ELISA for inflammatory factors, and neutrophil extracellular trap (NET) markers. Histological, immunofluorescence, and transmission electron microscopy analyses were conducted to evaluate CGA’s role of CGA in ARDS. mRNA sequencing and molecular docking and surface plasmon resonance (SPR) analysis were performed to determine CGA’s influence on the PI3K/AKT signalling pathway. Results We identified 144 common drug-disease targets, with honeysuckle containing 23 potentially active components. Key genes included STAT3, PIK3CA, and AKT1, which are involved in the PI3K/AKT, HIF-1, and Ras signalling pathways. Compared to the control group, in vivo studies revealed a marked diminution by CGA in cellular infiltration, oedema, and interstitial thickness observed in lungs impacted by ARDS. Furthermore, inflammatory mediators like IL-6, IL-1β, TNFα, IL-10 were lowered through CGA administration, alongside NET indicators including PAD4, citH3, and myeloperoxidase (MPO). T cell subtypes were altered during ARDS and CGA intervention. Molecular docking indicated a strong binding of CGA to PI3K and AKT1. SPR analysis further confirmed a high-affinity binding between CGA and PI3K, characterized by a low equilibrium dissociation constant (KD). Conclusion CGA alleviates ARDS by inhibiting the PI3K/AKT signaling pathway, thereby suppressing inflammation, regulating T-cell subtypes, and reducing NET formation.

70-kDa heat shock protein family is a molecular chaperone that binds to a variety of client proteins and peptides in the cytoplasm. Several studies have revealed binding motifs between 70-kDa heat shock protein family and cytoplasmic proteins by conventional techniques such as phage display library screening. However, little is known about the binding motif based on kinetic parameters determined by surface plasmon resonance analysis. We investigated the major inducible cytosolic 70-kDa heat shock protein (Hsp70)-binding motif with the human leukocyte antigen B*2702-derived peptide Bw4 (RENLRIALRY) by using a Biacore system based on surface plasmon resonance analysis. The K(D) value of Hsp70-Bw4 interaction was 1.8 x 10(-6) m. Analyses with truncated Bw4 variant peptides showed the binding motif of Hsp70 to be seven residues, LRIALRY. To further study the characteristics of this motif, 126 peptides derived from Bw4, each with single amino acid substitution, were synthesized and analyzed for Hsp70 binding affinity. Interestingly, the Hsp70 binding affinity was abrogated when the residues were substituted for by acidic (Asp and Glu) ones at any position. In contrast, if the substitute residue was aromatic (Trp, Tyr, and Phe) or an Arg residue at any position, Hsp70 binding affinity was maintained. Thus, this study presents a new binding motif between Hsp70 and peptides derived from the natural protein human leukocyte antigen B*2702 and may also elucidate some characteristics of the Hsp70 binding characteristic, enhancing our understanding of Hsp70-binding determinants that may influence diverse cellular and physiological processes.

Lactoferrin (LF) is an iron-binding secretory protein, which is distributed in the secondary granules of polynuclear lymphocytes as well as in the milk produced by female mammals. Although it has multiple functions, for example antimicrobial, immunomodulatory, antiviral, and anti-tumor metastasis activities, the receptors responsible for these activities are not fully understood. In this study, the binding epitopes for human LF were first isolated from a hexameric random peptide library displayed on T7 phage. Interestingly, two of the four isolated peptides had a representative cell adhesion motif, Arg-Gly-Asp (RGD), implying that human LF interacts with proteins with the RGD motif. We found that human LF bound to the RGD-containing human extracellular matrix proteins, fibronectin and vitronectin. Furthermore, human LF inhibited cell adhesion to these matrix proteins in a concentration-dependent manner but not to the RGD-independent cell adhesion molecule like laminin or collagen. These results indicate that a function of human LF is to block the various interactions between the cell surface and adhesion molecules. This may explain the multifunctionality of LF.

Motherwort synergized with oxytocin for postpartum hemorrhage prevention: Integrated clinical efficacy and mechanism exploration.

Gut barrier injury in sepsis is a major contributor to distant organ dysfunction and bad clinical outcomes. Enteric glia-derived S-nitrosoglutathione (GSNO) has been recognized as a novel modulator of gut barrier integrity. In this study, we tested the potential therapeutic effect and mechanism of exogenous GSNO on endotoxin-induced inflammatory response and intestinal barrier injury in a rat model of endotoxemia. Male Sprague-Dawley rats were randomly assigned to four groups as follows: control (saline only), GSNO, lipopolysaccharide (LPS), and LPS + GSNO. Femoral venous injection of LPS (10 mg/kg) or saline was followed by GSNO (1 mg/kg) or saline injection 15 minutes later. Distal ileum tissues and blood were harvested after 3 hours of LPS/saline injection. The intestinal barrier function was measured histologically and by intestinal permeability to fluorescein isothiocyanate dextran. The ultrastructural change of the epithelial tight junction was observed using transmission electron microscope, and the expression level of tight junction protein ZO-1 was analyzed using immunofluorescence and Western blot. Systemic and intestinal inflammation was measured by analyzing the tumor necrosis factor and interleukin 1β levels in plasma and distal ileum tissue, respectively. The levels of nuclear factor κB (NF-κB) and myosin light-chain kinase in the distal ileum were measured by Western blot. Compared with the endotoxemic rats, the addition of GSNO reduced the intestinal injury observed in histologic sections, decreased permeability to fluorescein isothiocyanate dextran, attenuated damage of the junction between epithelia, and protected against the LPS-induced expression decrease of ZO-1. Furthermore, addition of GSNO reduced plasma and intestinal tumor necrosis factor and interleukin 1β levels as well as inhibited the LPS-induced up-regulation of myosin light-chain kinase expression and NF-κB p65 level in the intestine. The data indicate that GSNO protects against the LPS-induced systemic inflammatory response and attenuated intestinal inflammation and epithelial barrier injury in rats, possibly through the inhibition of the NF-κB pathway.

Membrane-protein interaction plays key roles in a wide variety of biological processes. Although various methods have been employed to measure membrane binding of soluble proteins, a robust high-throughput assay that is universally applicable to all proteins is lacking at present. Here we report a new fluorescence quenching assay utilizing enhanced green fluorescence protein (EGFP)-fusion proteins and a lipid containing a dark quencher, N-dimethylaminoazobenzenesulfonyl-phosphatidylethanolamine (dabsyl-PE). The EGFP fluorescence emission intensity showed a large decrease (i.e., >50%) when EGFP-fusion proteins bound the vesicles containing 5 mol% dabsyl-PE. This simple assay, which can be performed using either a cuvette-based spectrofluorometer or a fluorescence plate reader, allowed rapid, sensitive, and accurate determination of lipid specificity and affinity for various lipid binding domains, including two pleckstrin homology domains, an epsin N-terminal homology domain, and a phox homology domain. The assay can also be applied to high-throughput screening of small molecules that modulate membrane binding of proteins.

The cytoplasmic amino terminus of HCN1, the primary full-length HCN isoform expressed in trout saccular hair cells, was found by yeast two-hybrid protocols to bind the cytoplasmic carboxyl-terminal domain of a protocadherin 15a-like protein. HCN1 was immunolocalized to discrete sites on saccular hair cell stereocilia, consistent with gradated distribution expected for tip link sites of protocadherin 15a. HCN1 message was also detected in cDNA libraries of rat cochlear inner and outer hair cells, and HCN1 protein was immunolocalized to cochlear hair cell stereocilia. As predicted by the trout hair cell model, the amino terminus of rat organ of Corti HCN1 was found by yeast two-hybrid analysis to bind the carboxyl terminus of protocadherin 15 CD3, a tip link protein implicated in mechanosensory transduction. Specific binding between HCN1 and protocadherin 15 CD3 was confirmed with pull-down assays and surface plasmon resonance analysis, both predicting dependence on Ca(2+). In the presence of calcium chelators, binding between HCN1 and protocadherin 15 CD3 was characterized by a K(D) = 2.39 x 10(-7) m. Ca(2+) at 26.5-68.0 microm promoted binding, with K(D) = 5.26 x 10(-8) m (at 61 microm Ca(2+)). Binding by deletion mutants of protocadherin 15 CD3 pointed to amino acids 158-179 (GenBank accession number XP_238200), with homology to the comparable region in trout hair cell protocadherin 15a-like protein, as necessary for binding to HCN1. Amino terminus binding of HCN1 to HCN1, hypothesized to underlie HCN1 channel formation, was also found to be Ca(2+)-dependent, although the binding was skewed toward a lower effective maximum [Ca(2+)] than for the HCN1 interaction with protocadherin 15 CD3. Competition may therefore exist in vivo between the two binding sites for HCN1, with binding of HCN1 to protocadherin 15 CD3 favored between 26.5 and 68 microm Ca(2+). Taken together, the evidence supports a role for HCN1 in mechanosensory transduction of inner ear hair cells.

IntroductionGu-Ben-Ke-Chuan (GBKC) decoction, a well-known prescription composed of seven herbs, has been widely used for treating chronic bronchitis (CB). However, the pharmacological constituents of GBKC and the underlying mechanisms by which these components act on CB remain unclear.MethodsUltra-high-pressure liquid chromatography coupled with linear ion trap–Orbitrap tandem mass spectrometry (UHPLC-LTQ-Orbitrap) was first employed to rapidly identify compounds from GBKC. Thereafter, network pharmacology and molecular docking analyses were performed to identify the potential active constituents, candidate targets, and major pathways. Finally, the affinities between the key compounds and targets were verified via surface plasmon resonance (SPR) analysis. In addition, the anti-inflammatory effect of GBKC was verified using an LPS-induced inflammatory cell model based on the predicted results.ResultsA total of 53 major compounds were identified in the GBKC decoction. After network pharmacology-based virtual screening, 141 major targets and 39 main compounds were identified to be effective in the treatment of CB. The major targets were highly enriched in the tumor necrosis factor (TNF) signaling pathway, suggesting that GBKC could attenuate the inflammatory response in patients with CB. Furthermore, molecular docking results indicated that 20 pairs of components and target proteins relevant to the TNF pathway exhibited notable interactions. Among them, eight compound-target pairs exhibited good affinity as per SPR analysis. In addition, the production of interleukin 6 and TNF-α in LPS-induced MH-S cells was suppressed after GBKC treatment.ConclusionThis study successfully clarified the mechanism of action of GBKC against CB, which demonstrated that the integrated strategy described above is reliable for identifying the active compounds and mechanisms responsible for the pharmacological activities of GBKC decoction.

Sweroside, the effective component of jianwei shoutai pills, alleviates recurrent pregnancy loss by activating cAMP signaling pathway in maternal-fetal interface.

Poster Board III-457 Surface Plasmon Resonance (SPR) Analysis Detects Clinically Significant Low Avidity HPA-1a Antibodies Not Identified by Conventional Serology.

A numerical analysis of the fiber-optic surface plasmon resonance (SPR) sensor with crescent shape of metal coating is presented. The crescent shape of metal coating usually occurs during one side metal deposition on the cylindrical fiber. Here, for analysis of the performance of fiber-optic SPR sensors with such asymmetric metal coating, the method of three-dimensional (3D) ray-tracing and theoretical calculation of electromagnetic reflection and transmission at the metal film are applied simultaneously, which result in 3D analysis with reduced time consumption and data loads. We investigate the characteristic of fiber-optic SPR sensor with asymmetric metal layer, comparing with those of the symmetric metal coated fiber, and discuss the asymmetry effect of the sensors for practical sensing and provide the information for improving sensing capability.