Objective To investigate the changes in eosinophil counts and the activation of microglial cells in the brain tissues of mice at different stages of Angiostrongylus cantonensis infection, and to examine the role of microglia in regulating the progression of angiostrongyliasis and unravel the possible molecular mechanisms. Methods Fifty BALB/c mice were randomly divided into the control group and the 7-d, 14-d, 21-day and 25-d infection groups, of 10 mice in each group. All mice in infection groups were infected with 30 stage III A. cantonensis larvae by gavage, and animals in the control group was given an equal amount of physiological saline. Five mice were collected from each of infection groups on days 7, 14, 21 d and 25 d post-infection, and 5 mice were collected from the control group on the day of oral gavage. The general and focal functional impairment was scored using the Clark scoring method to assess the degree of mouse neurological impairment. Five mice from each of infection groups were sacrificed on days 7, 14, 21 d and 25 d post-infection, and 5 mice from the control group were sacrificed on the day of oral gavage. Mouse brain tissues were sampled, and the pathological changes of brain tissues were dynamically observed using hematoxylin and eosin (HE) staining. Immunofluorescence staining with eosinophilic cationic protein (ECP) and ionized calcium binding adaptor molecule 1 (Iba1) was used to assess the degree of eosinophil infiltration and the counts of microglial cells in mouse brain tissues in each group, and the morphological parameters of microglial cells (skeleton analysis and fractal analysis) were quantified by using Image J software to determine the morphological changes of microglial cells. In addition, the expression of M1 microglia markers Fcγ receptor III (Fcgr3), Fcγ receptor IIb (Fcgr2b) and CD86 antigen (Cd86), M2 microglia markers Arginase 1 (Arg1), macrophage mannose receptor C-type 1 (Mrc1), chitinase-like 3 (Chil3), and phagocytosis genes myeloid cell triggering receptor expressed on myeloid cells 2 (Trem2), CD68 antigen (Cd68), and apolipoprotein E (Apoe) was quantified using real-time quantitative reverse transcription PCR (RT-qPCR) assay in the mouse cerebral cortex of mice post-infection. Results A large number of A. cantonensis larvae were seen on the mouse meninges surface post-infection, and many neuronal nuclei were crumpled and deeply stained, with a large number of bleeding points in the meninges. The median Clark scores of mouse general functional impairment were 0 (interquartile range, 0), 0 (interquartile range, 0.5), 6 (interquartile range, 1.0), 14 (interquartile range, 8.5) points and 20 (interquartile range, 9.0) points in the control group and the 7-d, 14-d, 21-d and 25-d groups, respectively (H = 22.45, P < 0.01), and the median Clark scores of mouse focal functional impairment were 0 (interquartile range, 0), 2 (interquartile range, 2.5), 7 (interquartile range, 3.0), 18 (interquartile range, 5.0) points and 25 (interquartile range, 6.5) points in the control group and the 7-d, 14-d, 21-d and 25-d groups, respectively (H = 22.72, P < 0.01). The mean scores of mice general and focal functional impairment were all higher in the infection groups than in the control group (all P values < 0.05). Immunofluorescence staining showed a significant difference in the eosinophil counts in mouse brain tissues among the five groups (F = 40.05, P < 0.000 1), and the eosinophil counts were significantly higher in mouse brain tissues in the 14-d (3.08 ± 0.78) and 21-d infection groups (5.97 ± 1.37) than in the control group (1.00 ± 0.28) (both P values < 0.05). Semi-quantitative analysis of microglia immunofluorescence showed a significant difference in the counts of microglial cells among the five groups (F = 17.66, P < 0.000 1), and higher Iba1 levels were detected in mouse brain tissues in 14-d (5.75 ± 1.28), 21-d (6.23 ± 1.89) and 25-d infection groups (3.70 ± 1.30) than in the control group (1.00 ± 0.30) (all P values < 0.05). Skeleton and fractal analyses showed that the branch length [(162.04 ± 34.10) μm vs. (395.37 ± 64.11) μm; t = 5.566, P < 0.05] and fractal dimension of microglial cells (1.30 ± 0.01 vs. 1.41 ± 0.03; t = 5.266, P < 0.05) were reduced in mouse brain tissues in the 21-d infection group relative to the control group. In addition, there were significant differences among the 5 groups in terms of M1 and M2 microglia markers Fcgr3 (F = 48.34, P < 0.05), Fcgr2b (F = 55.46, P < 0.05), Cd86 (F = 24.44, P < 0.05), Arg1 (F = 31.18, P < 0.05), Mrc1 (F = 15.42, P < 0.05) and Chil3 (F = 24.41, P < 0.05), as well as phagocytosis markers Trem2 (F = 21.19, P < 0.05), Cd68 (F = 43.95, P < 0.05) and Apoe (F = 7.12, P < 0.05) in mice brain tissues. Conclusions A. cantonensis infections may induce severe pathological injuries in mouse brain tissues that are characterized by massive eosinophil infiltration and persistent activation of microglia cells, thereby resulting in progressive deterioration of neurological functions.

Objective To explore the efficacy and safety of dapagliflozin in patients with paroxysmal atrial fibrillation (PAF) combined with heart failure with preserved ejection fraction (HFpEF). Methods A total of 120 patients with PAF combined with HFpEF treated at Jinshan Hospital of Fudan University from July 2022 to July 2023 were selected and randomly divided into the dapagliflozin group (n＝60, standard treatment combined with dapagliflozin) and the control group (n＝60, standard treatment combined with placebo). After 12 months of follow-up, the Kansas City Cardiomyopathy Questionnaire-Total Symptom Score (KCCQ-TSS), PAF duration, recurrence rate and frequency of PAF, left atrial diameter, left ventricular end-systolic diameter, left ventricular end-diastolic diameter, left ventricular ejection fraction, P-wave dispersion, blood pressure, plasma N-terminal pro-brain natriuretic peptide (NT-proBNP), estimated glomerular filtration rate (eGFR), and glycated hemoglobin A_1C (HbA_1C) were compared between the two groups. Cardiovascular outcomes and adverse events were observed. Results A total of 10 patients lost to follow-up, and 110 patients were included in the analysis (55 in each group). After 12 months of treatment, the KCCQ-TSS in the dapagliflozin group was significantly higher than that in the control group ([61.68±2.65] points vs [44.98±4.76] points, P＜0.001). The PAF duration in the dapagliflozin group was significantly shorter than that in the control group ([144±18] min vs [270±24] min, P＝0.045). After treatment, frequency of PAF, NT-proBNP levels, left ventricular end-systolic diameter, left ventricular end-diastolic diameter, left atrial diameter, P-wave dispersion, and HbA_1C levels showed statistical differences between the two groups (P＜0.05). The heart failure readmission rate and PAF recurrence rate in the dapagliflozin group were significantly lower than those in the control group (P＜0.05). There was no significant difference in the incidence of adverse events between the two groups during treatment. Conclusions Dapagliflozin improves patients’ quality of life, reduces PAF duration and recurrence rate, decreases heart failure readmission rate, lowers NT-proBNP levels, reverses cardiac remodeling, and demonstrates favorable safety in patients with PAF combined with HFpEF.

ObjectiveTo explore the evolution of medication patterns and syndrome-herb associations of traditional Chinese medicine （TCM） in treating inflammatory bowel disease （IBD）， providing a theoretical foundation for precise syndrome differentiation and treatment in clinical practice. MethodsMedical case literature on TCM treatment of IBD from 1960 to 2024 was retrieved to establish a database. Frequency statistics， cluster analysis， change point detection， and association rule mining were employed to comprehensively analyze the syndrome distribution， therapeutic methods， medication patterns， and their temporal variations. ResultsA total of 685 medical cases were included. Common syndromes were dampness-heat （66.42%） and spleen deficiency （56.20%）. Primary therapeutic methods included heat clearing （63.65%）， spleen invigorating （47.45%）， and dampness draining （36.79%）. High-frequency herbs included Coptidis Rhizoma （354）， Paeoniae Radix Alba （303）， Aucklandiae Radix （292）， Codonopsis Radix （253）， and Glycyrrhizae Radix et Rhizoma （244）. Initial prescription clustering revealed three core therapeutic method combinations： heat clearing and detoxifying （represented by Baitouweng Tang）， spleen invigorating and Qi reinforcing （represented by Shenling Baizhusan）， and cold-heat regulation （represented by Wumeiwan combined with Shaoyao tang）. Temporal analysis identified 2008 as a key transition point in TCM treatment of IBD， with significantly increased usage frequency of heat-clearing and dampness-drying herbs such as Fraxini Cortex， Phellodendri Chinensis Cortex， Sophorae Flavescentis Radix， and Scutellariae Radix as well as hemostatic herbs such as carbonized Sanguisorbae Radix， Bletillae Rhizoma， Agrimoniae Herba， and Notoginseng Radix et Rhizoma. Follow-up efficacy analysis showed median improvement rates of 64.0% at the first follow-up， 76.0% at the second follow-up， and 78.7% at the third follow-up. Syndrome-drug association analysis revealed specific herb pairs with significant therapeutic advantages， including Notoginseng Radix et Rhizoma + Coicis Semen， Sanguisorbae Radix + Coptidis Rhizoma， and Codonopsis Radix + Aconii Lateralis Radix Praeparaia. ConclusionTCM medication patterns for treating IBD demonstrate distinct temporal evolution characteristics， with significantly increased usage frequency of herbs such as Fraxini Cortex， Notoginseng Radix et Rhizoma， and Agrimoniae Herba. Significant therapeutic method-herb associations and syndrome-herb association patterns exist， with the formation of specific herb pairs， providing evidence-based support for precise syndrome differentiation and treatment of IBD.

BACKGROUND: Macrophage polarization anomalies and dysfunction play a crucial role in the pathogenesis of immune thrombocytopenia (ITP). Itaconate is a Krebs cycle-derived immunometabolite synthesized by myeloid cells to modulate cellular metabolism and inflammatory responses. This study aimed to evaluate the immunoregulatory effects of an itaconate derivative on macrophages in patients with ITP. METHODS: Peripheral blood-derived macrophages from patients with ITP and healthy controls were treated with 4-octyl itaconate (4-OI), a derivative of itaconate that can penetrate the cell membrane. Macrophage polarization, antigen-presenting functions, and phagocytic capability were measured via flow cytometry and enzyme-linked immunosorbent assay (ELISA). Macrophage glycolysis in patients with ITP and the metabolic regulatory effect of 4-OI were detected using a Seahorse XFe96 Analyzer. An active murine model of ITP was used to evaluate the therapeutic effects of 4-OI in vivo . RESULTS: 4-OI reduced the levels of CD80 and CD86 in M1 macrophages and suppressed the release of tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 pro-inflammatory cytokines, suggesting that 4-OI could hinder the polarization of macrophages toward an M1 phenotype. We found that 4-OI pretreated M1 macrophages reduced the proliferation of CD4 + T cells and promoted the differentiation of regulatory T cells. In addition, after 4-OI treatment, the phagocytic capacity of M1 macrophages toward antibody-coated platelets decreased significantly in patients with ITP. In addition, the glycolytic function of M1 macrophages was elevated in individuals with ITP compared to those in healthy controls. 4-OI treatment downregulated glycolysis in M1 macrophages. The glycolysis inhibitor 2-deoxy-d-glucose (2-DG) also inhibited the polarization of M1 macrophages and restored their functions. In vivo , 4-OI treatment significantly increased platelet counts in the active ITP murine model. CONCLUSIONS Itaconate derivative 4-OI inhibited M1 macrophage polarization and restored impaired functions through metabolic reprogramming. This study provides a novel therapeutic option for ITP.
Macrophages/metabolism* ; Humans ; Animals ; Succinates/pharmacology* ; Mice ; Male ; Female ; Adult ; Middle Aged ; Flow Cytometry ; Tumor Necrosis Factor-alpha/metabolism* ; Enzyme-Linked Immunosorbent Assay ; Purpura, Thrombocytopenic, Idiopathic/metabolism* ; Glycolysis/drug effects* ; Metabolic Reprogramming

The rapid emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants that evade immunity elicited by vaccination has posed a global challenge to the control of the coronavirus disease 2019 (COVID-19) pandemic. Therefore, developing countermeasures that broadly protect against SARS-CoV-2 and related sarbecoviruses is essential. Herein, we have developed a lipid nanoparticle (LNP)-encapsulated mRNA (mRNA-LNP) encoding the full-length Spike (S) glycoprotein of SARS-CoV-2 (termed RG001), which confers complete protection in a non-human primate model. Intramuscular immunization of two doses of RG001 in Rhesus monkey elicited robust neutralizing antibodies and cellular response against SARS-CoV-2 variants, resulting in significantly protected SARS-CoV-2-infected animals from acute lung lesions and complete inhibition of viral replication in all animals immunized with low or high doses of RG001. More importantly, the third dose of RG001 vaccination elicited effective neutralizing antibodies against current epidemic XBB and JN.1 strains and similar cellular response against SARS-CoV-2 Omicron variants (BA.1, XBB.1.16, and JN.1) were observed in immunized mice. All these results together strongly support the great potential of RG001 in preventing the infection of SARS-CoV-2 variants of concern (VOCs).

Periodontal disease is a risk factor for many systemic diseases such as Alzheimer's disease and adverse pregnancy outcomes. Cleft palate (CP), the most common congenital craniofacial defect, has a multifaceted etiology influenced by complex genetic and environmental risk factors such as maternal bacterial or virus infection. A prior case-control study revealed a surprisingly strong association between maternal periodontal disease and CP in offspring. However, the precise relationship remains unclear. In this study, the relationship between maternal oral pathogen and CP in offspring was studied by sonicated P. gingivalis injected intravenously and orally into pregnant mice. We investigated an obvious increasing CP (12.5%) in sonicated P. gingivalis group which had inhibited osteogenesis in mesenchyme and blocked efferocytosis in epithelium. Then glycolysis and H4K12 lactylation (H4K12la) were detected to elevate in both mouse embryonic palatal mesenchyme (MEPM) cells and macrophages under P. gingivalis exposure which further promoted the transcription of metallopeptidase domain17 (ADAM17), subsequently mediated the shedding of transforming growth factor-beta receptor 1 (TGFBR1) in MEPM cells and mer tyrosine kinase (MerTK) in macrophages and resulted in the suppression of efferocytosis and osteogenesis in palate, eventually caused abnormalities in palate fusion and ossification. The abnormal efferocytosis also led to a predominance of M1 macrophages, which indirectly inhibited palatal osteogenesis via extracellular vesicles. Furthermore, pharmacological ADAM17 inhibition could ameliorate the abnormality of P. gingivalis-induced abnormal palate development. Therefore, our study extends the knowledge of how maternal oral pathogen affects fetal palate development and provides a novel perspective to understand the pathogenesis of CP.
Animals ; Female ; Porphyromonas gingivalis ; Pregnancy ; Mice ; Cleft Palate/etiology* ; Glycolysis

Laccases (LACs), belonging to the multicopper oxidase family, are closely associated with various biological functions including lignin synthesis and responses to biotic and abiotic stresses in plants. However, few studies have reported the laccase genes in China rose (Rosa chinensis). Prickles cause difficulties to the management and harvest of R. chinensis and have become a trait concerned in the breeding. To investigate the expression patterns of laccase genes in roses, we cloned a laccase gene from an ancient variety R. chinensis 'Old Blush' and named it RcLAC15. The expression level of RcLAC15 in prickles was significantly higher than those in roots, stems, and leaves. Fifty-eight laccase genes were identified in the genome of R. chinensis, and bioinformatics analysis revealed that RcLAC15 was a homolog of AtLAC15, predicting that RcLAC15 was a stable hydrophilic protein without transmembrane structures. The recombinant expression vector pBI121-proRcLAC15:: GUS was introduced into Arabidopsis, and GUS staining results showed that the RcLAC15 promoter specifically drove GUS gene expression at the edges of Arabidopsis leaves. In summary, RcLAC15 is a gene specifically expressed in the prickles of R. chinensis. This discovery provides a reference for exploring the biological functions of laccase genes in the prickles of R. chinensis.
Laccase/metabolism* ; Rosa/enzymology* ; Cloning, Molecular ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism* ; Arabidopsis/metabolism* ; Plants, Genetically Modified/metabolism*

Objective To investigate the correlation between lipoprotein-associated phospholipase A2(Lp-PLA2),hemoglobin glycotion index(HGI)and the severity of type 2 diabetic retinopathy.Methods A total of 120 patients with type 2 diabetes mellitus who were hospitalized in the General Hospital of Hunan Medical University from January 2021 to June 2022 were selected and divided into type 2 diabetes without retinopathy group(NDR group,n=45),non-proliferative diabetic retinopathy group(NPDR group,n=45)and prolifera-tive diabetic retinopathy group(PDR group,n=30).In the same period,50 healthy subjects were randomly selected as control group.The clinical data,Lp-PLA2 and HGI levels of each group were collected and com-pared,and the correlation between different HGI and Lp-PLA2 levels and the severity of the disease was ana-lyzed.Results The levels of Lp-PLA2 and HGI in NDR,NPDR and PDR groups were higher than those in control group(P＜0.05).The proportion of patients with NPDR and PDR in high-Lp-PLA group(H-Lp-PLA2 group)and high-HGI group(H-HGI group)were higher than those in low-HGI group(L-HGI group)and low-Lp-PLA2 group(L-Lp-PLA2 group),and the differences were statistically significant(P＜0.05).Multiple Logistic regression analysis showed that Lp-PLA2 and HGI were risk factors for DR(P＜0.05),and were positively correlated with the severity of type 2 diabetic retinopathy(P＜0.05).Conclusion HGI and Lp-PLA2 are closely related to the severity of retinopathy in type 2 diabetes mellitus,and may be used as new indicators to predict the severity of the disease.

Objective To investigate the influencing factors for early hematoma enlargement(HE)in patients with spontaneous intracerebral hemorrhage(SICH).Methods A total of 311 pa-tients with SICH were selected as research subjects.Based on the occurrence of HE,they were divided into enlargement group and non-enlargement group.Clinical assessment indicators of patients with SICH were collected,and general information between the two groups was compared.The influencing factors for the occurrence of HE in patients with SICH were analyzed.Results Based on the presence of HE at a second CT examination within 6 to 12 hours,311 patients with SICH were divided into en-largement group(n=97)and non-enlargement group(n=214).Univariate analysis revealed signifi-cant differences between the two groups in terms of initial hematoma volume,regularity of hematoma margin,Glasgow Coma Scale(GCS)score,coagulation function,as well as the presence of swirl sign,lobulation sign,black hole sign,blend sign,lymphocyte count and neutrophil count(P＜0.05).Multivariate analysis indicated that GCS score,initial hematoma volume,history of antithrom-botic drug use,abnormal coagulation function and intracranial CT signs were independent influencing factors for HE(P＜0.05).Conclusion The occurrence of HE in patients with SICH is closely re-lated to GCS score,initial hematoma volume,history of antithrombotic drug use,abnormal coagulation function and intracranial CT signs.

Objective To quantitatively evaluate structural changes in the TMD disc using magnetic resonance T2 mapping technology at open and closed mouth positions,and assess its significance.Methods 127 newly diagnosed patients with TMD and 51 controls in the Department of Oral and Maxillofacial Surgery of the Affiliated Hospital of Yunnan University from June 2023 to January 2024 were prospectively collected.All subjects underwent routine TMJ sequence and T2 mapping sequence scans.After scanning,the pseudo-color images were processed,and T2 values of the joint disc(anterior and posterior bands)were measured at open and closed mouth positions to analyze their correlation with TMD disc displacement.Results The T2 values of the posterior zone of the joint disc in both open and closed positions were higher than those in the anterior zone of the joint disc in the case group(P<0.05);for different anterior displacement degrees of the joint disc,T2 values showed a progressive increase from the normal position group to the mild and moderate anterior displacement groups,while the severely displaced group demonstrated a decreasing trend(P<0.01);joint clicking and occlusal abnormalities were independent risk factors for non-reducible anterior joint disc displacement in TMD patients.Conclusion Scans during both mouth-opening and closing positions can be used to assess joint disc injury in TMD;T2 mapping technology can quantitatively and sensitively reflect changes in the biochemical composition of joint disc microstructure.