[Relationship between Cellular Microenvironment and Idiopathic Pulmonary Fibrosis].

Idiopathic pulmonary fibrosis(IPF)is a progressive lung disease characterized by pulmonary interstitial fibrosis and pulmonary dysfunction.Cell microenvironment is mainly composed of cell components,extracellular matrix,extracellular regulators,and liquid substances.Changes in microenvironment components are closely related to IPF.This article elaborates the roles of cell microenvironments including cytokines,mesenchymal cells,extracellular matrix,and unfolded proteins in the pathogenesis of IPF.

Whiteboard Use in Labor and Delivery: A Tool to Improve Patient Knowledge of the Name of the Delivery Provider and Satisfaction with Care.

Introduction The impact of whiteboard use in labor rooms has not previously been studied. This quality improvement study aimed to evaluate patient knowledge of their delivering physician's name and the change in patient satisfaction after the implementation of a whiteboard in labor rooms. Methods A multidisciplinary team designed a dry-erase whiteboard prompting care providers to record their names, roles and patient care information. A questionnaire was administered to patients before and after the whiteboard implementation. Patients who had a planned cesarean or vaginal birth within 1 h of admission were excluded. Categorical variables were compared using Chi square and Fisher's exact tests. A multivariable logistic regression was performed to control for confounders. Results 191 patients completed the questionnaires. Although patients were not randomized, the pre-and post-intervention groups were similar. Post-intervention, we found a significant increase in recalling the delivery resident's name [21/101 (20.8%) vs. 33/90 (36.7%), p = 0.016] and a non-significant increase in recalling the name of the attending and nurse [19/101 (18.8%) vs. 23/90 (25.6%), p = 0.296; 46/101 (45.5%) vs. 53/90 (58.9%), p = 0.082]. Post-intervention, patient satisfaction with care was significantly higher [83/101 (82.2%) vs. 83/89 (93.3%), p = 0.028]. Knowledge of the delivery resident's name was associated with higher patient satisfaction [115/137 (84%) vs. 51/53 (96%), p = 0.03] and attendance of the postpartum care visit [50.4% (69/137) vs. 64.8% (35/54), p = 0.049]. Discussion The use of a well-designed whiteboard increases laboring patients' knowledge of their delivery physician's name and may improve patient satisfaction with care on Labor and Delivery.

A Highly Conserved Salt Bridge Stabilizes the Kinked Conformation of ß2,3-Sheet Essential for Channel Function of P2X4 Receptors.

Significant progress has been made in understanding the roles of crucial residues/motifs in the channel function of P2X receptors during the pre-structure era. The recent structural determination of P2X receptors allows us to reevaluate the role of those residues/motifs. Residues Arg-309 and Asp-85 (rat P2X4 numbering) are highly conserved throughout the P2X family and were involved in loss-of-function polymorphism in human P2X receptors. Previous studies proposed that they participated in direct ATP binding. However, the crystal structure of P2X demonstrated that those two residues form an intersubunit salt bridge located far away from the ATP-binding site. Therefore, it is necessary to reevaluate the role of this salt bridge in P2X receptors. Here, we suggest the crucial role of this structural element both in protein stability and in channel gating rather than direct ATP interaction and channel assembly. Combining mutagenesis, charge swap, and disulfide cross-linking, we revealed the stringent requirement of this salt bridge in normal P2X4 channel function. This salt bridge may contribute to stabilizing the bending conformation of the ß2,3-sheet that is structurally coupled with this salt bridge and the α2-helix. Strongly kinked ß2,3 is essential for domain-domain interactions between head domain, dorsal fin domain, right flipper domain, and loop ß7,8 in P2X4 receptors. Disulfide cross-linking with directions opposing or along the bending angle of the ß2,3-sheet toward the α2-helix led to loss-of-function and gain-of-function of P2X4 receptors, respectively. Further insertion of amino acids with bulky side chains into the linker between the ß2,3-sheet or the conformational change of the α2-helix, interfering with the kinked conformation of ß2,3, led to loss-of-function of P2X4 receptors. All these findings provided new insights in understanding the contribution of the salt bridge between Asp-85 and Arg-309 and its structurally coupled ß2,3-sheet to the function of P2X receptors.

Antimicrobial peptide protonectin disturbs the membrane integrity and induces ROS production in yeast cells.

Candidiasis is often observed in immunocompromised patients and is the 4th most common cause of bloodstream infections. However, antifungals are limited, so novel antifungal agents are urgently needed. Antimicrobial peptides (AMPs) are considered as potential alternatives of conventional antibiotics. In the present study, antimicrobial peptide protonectin was chemically synthesized and its antifungal activity and mode of action were studied. Our results showed that protonectin has potent antifungal activity and fungicidal activity against the tested fungi cells. Its action mode involved the disruption of the membrane integrity and the inducing of the production of cellular ROS. Furthermore, protonectin could inhibit the formation of biofilm and kill the adherent fungi cells. In conclusion, with the increase of fungal infection, protonectin may offer a new strategy and be considered as a potential therapeutic agent against fungal disease.

Antifungal effect and action mechanism of antimicrobial peptide polybia-CP.

The incidence of life-threatening invasive fungal infections increased significantly in recent years. However, the antifungal therapeutic options are very limited. Antimicrobial peptides are a class of potential lead chemical for the development of novel antifungal agents. Antimicrobial peptide polybia-CP was purified from the venom of the social wasp Polybia paulista. In this study, we synthesized polybia-CP and determined its antifungal effects against a series of Candidian species. Our results showed that polybia-CP has potent antifungal activity and fungicidal activity against the tested fungal cells with a proposed membrane-active action mode. In addition, polybia-CP could induce the increase of cellular reactive oxygen species production, which would attribute to its antifungal activity. In conclusion, the present study suggests that polybia-CP has potential as an antifungal agent or may offer a new strategy for antifungal therapeutic option.

Regulatory logic and pattern formation in the early sea urchin embryo.

We model the endomesoderm tissue specification process in the vegetal half of the early sea urchin embryo using Boolean models with continuous-time updating to represent the regulatory network that controls gene expression. Our models assume that the network interaction rules remain constant over time and the dynamics plays out on a predetermined program of cell divisions. An exhaustive search of two-node models, in which each node may represent a module of several genes in the real regulatory network, yields a unique network architecture that can accomplish the pattern formation task at hand--the formation of three latitudinal tissue bands from an initial state with only two distinct cell types. Analysis of an eight-gene model constructed from available experimental data reveals that it has a modular structure equivalent to the successful two-node case. Our results support the hypothesis that the gene regulatory network provides sufficient instructions for producing the correct pattern of tissue specification at this stage of development (between the fourth and tenth cleavages in the urchin embryo).

HPGD: An Intermediate Player in Microglial Polarization and Multiple Sclerosis Regulated by Nr4a1.

HPGD encodes 15-Hydroxyprostaglandin dehydrogenase catalyzing the decomposition of prostaglandin E2 and has not been reported in multiple sclerosis (MS). We previously found that Nr4a1 regulated microglia polarization and inhibited the progression of experimental autoimmune encephalomyelitis (EAE). Bioinformatics analysis suggested that HPGD might be regulated by Nr4a1. Therefore, this study aimed to explore the role of HPGD in microglia polarization and determine whether HPGD mediates the inhibition of EAE by Nr4a1. C57BL/6 mice were treated with MOG35-55 peptide to induce EAE. BV-2 cells were treated with LPS/IL-4 to induce M1/M2 polarization. We then analyzed the pathological changes of spinal cord tissue, detected the expression levels of M1/M2 genes in tissues and cells, and explored the effect of HPGD on PPARγ activation to clarify the role of HPGD in EAE. The interaction between HPGD and Nr4a1 was verified by ChIP and pull-down assay. HPGD was downregulated in the spinal cord of EAE mice and HPGD overexpression alleviated the progression of EAE. Experiments in vitro and in vivo revealed that HPGD inhibited M1 polarization, promoted M2 polarization and increased PPARγ-DNA complex level. Nr4a1 could bind to the promoter of HPGD and its overexpression increased HPGD level. HPGD overexpression (or knockdown) reversed the effect of Nr4a1 knockdown (or overexpression) on M1/2 polarization. HPGD is regulated by Nr4a1 and inhibits the progression of EAE through shifting the M1/M2 polarization and promoting the activation of PPARγ signaling pathway. This study provides potential targets and basis for the development of MS therapeutic drugs.

Deficiency of purinergic P2X4 receptor alleviates experimental autoimmune hepatitis in mice.

Purinergic P2X4 receptor (P2X4R) has been shown to have immunomodulatory properties in infection, inflammation, and organ damage including liver regeneration and fibrosis. However, the mechanisms and pathophysiology associated with P2X4R during acute liver injury remain unknown. We used P2X4R-/- mice to explore the role of P2X4R in three different models of acute liver injury caused by concanavalin A (ConA), carbon tetrachloride, and acetaminophen. ConA treatment results in an increased expression of P2X4R in the liver of mice, which was positively correlated with higher levels of aspartate aminotransferase and alanine aminotransferase in the serum. However, P2X4R gene ablation significantly reduced the severity of acute hepatitis in mice caused by ConA, but not by carbon tetrachloride or acetaminophen. The protective benefits against immune-mediated acute hepatitis were achieved via modulating inflammation (Interleukin (IL)-1ß, IL-6, IL-17A, interferon-γ, tumor necrosis factor-α), oxidative stress (malondialdehyde, superoxide dismutase, glutathione peroxidase, and catalase), apoptosis markers (Bax, Bcl-2, and Caspase-3), autophagy biomarkers (LC3, Beclin-1, and p62), and nucleotide oligomerization domain-likereceptorprotein 3(NLRP3) inflammasome-activated pyroptosis markers (NLRP3, Gasdermin D, Caspase-1, ASC, IL-1ß). Additionally, administration of P2X4R antagonist (5-BDBD) or agonist (cytidine 5'-triphosphate) either improved or worsened ConA-induced autoimmune hepatitis, respectively. This study is the first to reveal that the absence of the P2X4 receptor may mitigate immune-mediated liver damage, potentially by restraining inflammation, oxidation, and programmed cell death mechanisms. And highlight P2X4 receptor is essential for ConA-induced acute hepatitis.

TRPV1 analgesics disturb core body temperature via a biased allosteric mechanism involving conformations distinct from that for nociception.

Efforts on developing transient receptor potential vanilloid 1 (TRPV1) drugs for pain management have been hampered by deleterious hypo- or hyperthermia caused by TRPV1 agonists/antagonists. Here, we compared the effects of four antagonists on TRPV1 polymodal gating and core body temperature (CBT) in Trpv1+/+, Trpv1-/-, and Trpv1T634A/T634A. Neither the effect on proton gating nor drug administration route, hair coverage, CBT rhythmic fluctuations, or inflammation had any influence on the differential actions of TRPV1 drugs on CBT. We identified the S4-S5 linker region exposed to the vanilloid pocket of TRPV1 to be critical for hyperthermia associated with certain TRPV1 antagonists. PSFL2874, a TRPV1 antagonist we discovered, is effective against inflammatory pain but devoid of binding to the S4-S5 linker and inducing CBT changes. These findings implicate that biased allosteric mechanisms exist for TRPV1 coupling to nociception and CBT regulation, opening avenues for the development of non-opioid analgesics without affecting CBT.

Causal structure of oscillations in gene regulatory networks: Boolean analysis of ordinary differential equation attractors.

A common approach to the modeling of gene regulatory networks is to represent activating or repressing interactions using ordinary differential equations for target gene concentrations that include Hill function dependences on regulator gene concentrations. An alternative formulation represents the same interactions using Boolean logic with time delays associated with each network link. We consider the attractors that emerge from the two types of models in the case of a simple but nontrivial network: a figure-8 network with one positive and one negative feedback loop. We show that the different modeling approaches give rise to the same qualitative set of attractors with the exception of a possible fixed point in the ordinary differential equation model in which concentrations sit at intermediate values. The properties of the attractors are most easily understood from the Boolean perspective, suggesting that time-delay Boolean modeling is a useful tool for understanding the logic of regulatory networks.

Clinical, imaging features and outcomes of patients with anti-GFAP antibodies: a retrospective study.

Objective: To evaluate and compare the clinical features, imaging, overlapping antibodies, and prognosis of pediatric and adult patients with anti-GFAP antibodies. Methods: This study included 59 patients with anti-GFAP antibodies (28 females and 31 males) who were admitted between December 2019 and September 2022. Results: Out of 59 patients, 18 were children (under 18 years old), and 31 were adults. The overall cohort's median age at onset was 32 years old, 7 for children, and 42 for adults. There were 23 (41.1%) patients with prodromic infection, 1 (1.7%) patient with a tumor, 29 (53.7%) patients with other non-neurological autoimmune diseases, and 17 (22.8%) patients with hyponatremia. Fourteen (23.7%) patients had multiple neural autoantibodies, with the AQP4 antibody being the most common. Encephalitis (30.5%) was the most common phenotypic syndrome. Common clinical symptoms included fever (59.3%), headache (47.5%), nausea and vomiting (35.6%), limb weakness (35.6%), and disturbance of consciousness (33.9%). Brain MRI lesions were primarily located in the cortex/subcortex (37.3%), brainstem (27.1%), thalamus (23.7%), and basal ganglia (22.0%). Spinal cord MRI lesions often involved the cervical and thoracic spinal cord. There was no statistically significant difference in the MRI lesion site between children and adults. Out of 58 patients, 47 (81.0%) had a monophasic course, and 4 died. The last follow-up showed that 41/58 (80.7%) patients had an improved functional outcome (mRS <3), and children were more likely than adults to have no residual disability symptoms (p = 0.001). Conclusion: There was no statistically significant difference in clinical symptoms and imaging findings between children and adult patients with anti-GFAP antibodies; Patients with anti-GFAP antibodies may present with normal MRI findings or delayed MRI abnormalities, and patients with overlapping antibodies were common. Most patients had monophasic courses, and those with overlapping antibodies were more likely to relapse. Children were more likely than adults to have no disability. Finally, we hypothesize that the presence of anti-GFAP antibodies is a non-specific witness of inflammation.

Comparison of glial fibrillary acidic protein-immunoglobulin G-associated myelitis with myelin oligodendrocyte glycoprotein-immunoglobulin G-associated myelitis.

Objective: Glial fibrillary acidic protein-immunoglobulin G (GFAP-IgG)-associated myelitis and myelin oligodendrocyte glycoprotein-IgG (MOG-IgG)-associated myelitis have rarely been compared. Therefore, this study aimed to explore the clinical, laboratory, and imaging features of them to identify the differences. Methods: Overall, 14 and 24 patients with GFAP-IgG-and MOG-IgG-associated myelitis, respectively, were retrospectively screened and included in the study. Results: Among the 14 patients with GFAP-IgG-associated myelitis, the condition was more common in males (71.4%), with a median age of onset of 36.5 years, and more common in adults than in children (35.7%). In contrast, among the 24 patients with MOG-IgG-associated myelitis, the condition was equally divided between males and females, with a median age of onset of 9.5 years and more in children (66.7%) than in adults. The median age of onset of GFAP-IgG-associated myelitis was later than that of the MOG-IgG group. Isolated myelitis was rare in both groups. Elevated cerebrospinal fluid (CSF) protein levels were more prevalent in patients with GFAP-IgG-associated myelitis (64.3%) than in those with MOG-IgG-associated myelitis (16.7%) (p < 0.05), whereas patchy gadolinium enhancement of the cerebral lesion site was less common in patients with GFAP-IgG-associated myelitis than in those with MOG-IgG associated myelitis (p < 0.05). Six patients had a combination of other neurological autoantibodies, the specific mechanism of the overlapping antibodies remains unclear. Conclusion: Cerebrospinal fluid analysis and gadolinium enhanced MRI examination may help to distinguish the two kinds of myelitis.

Dissolved organic matter composition and fluorescence characteristics of the river affected by coal mine drainage.

Coal mine drainage (CMD) discharged into surface waters results in serious environmental pollution risk to rivers, lakes, and reservoirs. Coal mine drainage generally contains a variety of organic matter and heavy metals due to coal mining activities. Dissolved organic matter (DOM) plays an important role in the physicochemical and biological processes of many aquatic ecosystems. In this study, the investigations were carried out in the dry and wet seasons in 2021 to assess the characteristics of DOM compounds in coal mine drainage and the CMD-affected river. The results indicated that the pH of CMD-affected river pressed close to coal mine drainage. Besides, coal mine drainage lowered DO by 36% and increased total dissolved solids by 19% in the CMD-affected river. Coal mine drainage decreased absorption coefficient a(350) and absorption spectral slope S275-295 of DOM in the CMD-affected river; hence, DOM molecular size increased with decreasing S275-295. Three-dimensional fluorescence excitation-emission matrix spectroscopy and parallel factor analysis identified humic-like C1, tryptophan-like C2, and tyrosine-like C3 in the CMD-affected river and coal mine drainage. DOM in the CMD-affected river mainly originated from microbial and terrestrial sources, with strong endogenous characteristics. The ultra-high-resolution Fourier transform ion cyclotron resonance mass spectrometry analysis revealed that coal mine drainage had a higher relative abundance of CHO (44.79%), with a higher unsaturation degree of DOM. Coal mine drainage decreased the AImod,wa, DBEwa (double bond equivalents), Owa, Nwa, and Swa values and increased the relative abundance of the O3S1 species with DBE of 3 and carbons number range of 15-17 at the CMD inlet to the river channel. Moreover, coal mine drainage with the higher protein content increased the protein content of water at the CMD inlet to the river channel and the downstream river. DOM compositions and proprieties in coal mine drainage were investigated to further understand the influence of organic matter on heavy metals in future study.

Naringenin confers protection against experimental autoimmune encephalomyelitis through modulating the gut-brain axis: A multiomics analysis.

Multiple sclerosis (MS) is a disease of the central nervous system that involves the immune system attacking the protective covering of nerve fibers. This disease can be influenced by both environmental and genetic factors. Evidence has highlighted the critical role of the intestinal microbiota in MS and its animal model, experimental autoimmune encephalomyelitis (EAE). The composition of gut microflora is mainly determined by dietary components, which, in turn, modulate host homeostasis. A diet rich in naringenin at 0.5% can effectively mitigate the severity of EAE in mice. However, there is little direct data on the impact of naringenin at optimal doses on EAE development, as well as its intestinal microbiota and metabolites. Our study revealed that 2.0% naringenin resulted in the lowest clinical score and pathological changes in EAE mice, and altered the gene expression profiles associated with inflammation and immunity in spinal cord tissue. We then used untargeted metabolomics and 16S rRNA gene sequences to identify metabolites and intestinal microbiota, respectively. Naringenin supplementation enriched gut microbiota in EAE mice, including increasing the abundance of Paraprevotellaceae and Comamonadaceae, while decreasing the abundance of Deltaproteobacteria, RF39, and Desulfovibrionaceae. Furthermore, the changes in gut microbiota affected the production of metabolites in the feces and brain, suggesting a role in regulating the gut-brain axis. Finally, we conducted a fecal transplantation experiment to validate that gut microbiota partly mediates the effect of naringenin on EAE alleviation. In conclusion, naringenin has potential immunomodulatory effects that are influenced to some extent by the gut microbiome.

Chronic cough relief by allosteric modulation of P2X3 without taste disturbance.

P2X receptors are cation channels that sense extracellular ATP. Many therapeutic candidates targeting P2X receptors have begun clinical trials or acquired approval for the treatment of refractory chronic cough (RCC) and other disorders. However, the present negative allosteric modulation of P2X receptors is primarily limited to the central pocket or the site below the left flipper domain. Here, we uncover a mechanism of allosteric regulation of P2X3 in the inner pocket of the head domain (IP-HD), and show that the antitussive effects of quercetin and PSFL2915 (our nM-affinity P2X3 inhibitor optimized based on quercetin) on male mice and guinea pigs were achieved by preventing allosteric changes of IP-HD in P2X3. While being therapeutically comparable to the newly licensed P2X3 RCC drug gefapixant, quercetin and PSFL2915 do not have an adverse effect on taste as gefapixant does. Thus, allosteric modulation of P2X3 via IP-HD may be a druggable strategy to alleviate RCC.

Rethinking and Improving Feature Pyramids for One-stage Referring Expression Comprehension.

Referring Expression Comprehension (REC) is an important task in the vision-and-language community, since it is an essential step for many cross-modal tasks such as VQA, image retrieval and image caption. To obtain a better trade-off between speed and accuracy, existing researches usually follow a one-stage paradigm, where this task can be considered as a language-conditioned object detection task. Meanwhile, previous one-stage REC frameworks provide many different research perspectives, such as the strategies of fusion, the stage of fusion and the design of detection head. Surprisingly, these works mostly ignore the value of integrating multi-level features and even only apply single-scale features to locate the target. In this paper, we focus on rethinking and improving feature pyramids for one-stage REC. By experimental validations, we first prove that although multi-scale fusion is an effective approach for improving performance, the mature neck structures from object detection (e.g., FPN, BFN and HRFPN) have a limited impact on this task. Further, we visualize the outputs of FPN and find the underlying reason is that these coarse-grained FPN fusion strategies suffer from semantic ambiguity problem. Based on the above insights, we propose a new Language-Guided FPN (LG-FPN) method, which can dynamically allocate and select the fine-grained information by stacking language-gate and union-gate. A large number of contrastive and ablative experiments show that our LG-FPN is an effective and reliable module that can adapt to different visual backbones, fusion strategies and detection heads. Finally, our method achieves state-of-the-art performance on four referring expression datasets.

Pyranone Derivatives With Antitumor Activities, From the Endophytic Fungus Phoma sp. YN02-P-3.

Two new pyranone derivatives phomapyrone A (2) and phomapyrone B (3), one new coumarin 11S, 13R-(+)-phomacumarin A (1), three known pyranones (4-6), together with three known amide alkaloids fuscoatramides A-C (7-9), as well as 9S, 11R-(+)-ascosalitoxin (10) were isolated from the endophytic fungus Phoma sp. YN02-P-3, which was isolated from the healthy leaf tissue of a Paulownia tree in Yunnan Province, China. Their structures were elucidated using extensive NMR spectroscopic and HRESIMS data and by comparing the information with literature data. In addition, all compounds were tested for their cytotoxicity activity against human tumor cell lines, and the results showed that new compounds 1-3 showed moderate inhibitory activity against the HL-60 cell line with IC50 values of 31.02, 34.62, and 27.90 µM, respectively.

Effect of biochar derived from biogas residue on methane production during dry anaerobic fermentation of kitchen waste.

Kitchen wastes (KW) dramatically increasing with population and economy enhancing, and dry anaerobic fermentation was used to treat it. However, the large amount of biogas residue severely restricted the application of dry anaerobic fermentation, because the high total solid might lead to the system failure. Therefore, it is urgent to find appropriate way to improve the efficiency of dry anaerobic fermentation and reduce the great amount of biogas residue. In this study, a tentative experiment was conducted to investigate the effect of biochar prepared from biogas residue on the performance of dry anaerobic fermentation system. The results showed that almost half of the biogas residue was reduced and converted into biochar. At the presence of biochar, methane yield was 308.6 mL/gVS, which was 10.5% higher than that of control. Compared to the system without biochar, the highest volatile fatty acid (VFA) concentration was 19.3% higher and the percentage of acetate and valerate was 25.3％ and 12.8％, while it was 16.3% and 22.0% in the control, suggesting that biochar accelerated the degradation of VFA. Bacteria community diversity increased, Fastidiosipila and Proteiniphilum enriched at the presence of biochar, which might accelerate the hydrolysis and acidification of KW. Hydrogenotrophic methanogens was dominated and syntrophic acetate oxidation was the primary pathway to produce methane. This study developed a new recycle route for improving the efficiency of dry anaerobic fermentation while reducing the large amount of biogas residue generated from dry anaerobic fermentation.

Predictors of Influenza Vaccination among Chinese Middle School Students Based on the Health Belief Model: A Mixed-Methods Study.

Influenza vaccination rates among Chinese middle school students are low. This study aims to explore the influencing factors of vaccination among middle school students and promote vaccination. We conducted a mixed-methods study, integrating a questionnaire survey among 9145 middle school students in four cities in China and semi-structured interviews with 35 middle school students to understand their attitudes and perceptions toward vaccination based on the Health Belief Model. We found the overall vaccination rate was 38.2% (3493/9145), with students in Beijing, boarding at school, or senior high school showing higher values than their counterparts (p < 0.05). Multiple logistic regression results showed that non-boarding (OR = 0.46, 95%CI: 0.42−0.51) and perceived barriers (OR = 0.97, 95%CI: 0.96−0.98) were unfavorable factors for influenza vaccination, whereas perceived susceptibility (OR = 1.07, 95%CI: 1.05−1.08), perceived benefits (OR = 1.02, 95%CI: 1.01−1.04), cues to action (OR = 1.08, 95%CI: 1.05−1.11), and self-efficacy (OR = 1.04, 95%CI: 1.02−1.07) were facilitators. Qualitative results indicated that positive health beliefs, school, and the home environment contribute to vaccination. In conclusion, the influenza vaccination rate among middle school students remains low. The concerns about the safety and potential side effects of vaccines are the main barriers to vaccination, underscoring the need for strengthening communication, education, and information among students and their teachers/parents.

Dynamic recognition of naloxone, morphine and endomorphin1 in the same pocket of µ-opioid receptors.

Morphine, the most widely used analgesic, relieves severe pain by activating the µ-opioid receptor (MOR), whereas naloxone, with only slight structural changes compared to morphine, exhibits inhibitory effect, and is used to treat opioid abuse. The mechanism by which the MOR distinguishes between the two is unclear. Molecular dynamics (MD) simulations on a 1-µs time scale and metadynamics-enhanced conformational sampling are used here to determine the different interactions of these two ligands with MOR: morphine adjusted its pose by continuously flipping deeper into the pocket, whereas naloxone failed to penetrate deeper because its allyl group conflicts with several residues of MOR. The endogenous peptide ligand endomorphin-1 (EM-1) underwent almost no significant conformational changes during the MD simulations. To validate these processes, we employed GIRK4S143T, a MOR-activated Gßγ-protein effector, in combination with mutagenesis and electrophysiological recordings. We verified the role of some key residues in the dynamic recognition of naloxone and morphine and identified the key residue I322, which leads to differential recognition of morphine and naloxone while assisting EM-1 in activating MOR. Reducing the side chain size of I322 (MORI322A) transformed naloxone from an inhibitor directly into an agonist of MOR, and I322A also significantly attenuated the potency of MOR on EM-1, confirming that binding deep in the pocket is critical for the agonistic effect of MOR. This finding reveals a dynamic mechanism for the response of MOR to different ligands and provides a basis for the discovery of new ligands for MOR at the atomic level.