Objective : To generate heterozygous TRAF2 knockout mice, the CRISPR/Cas9 technology was successfully employed. These mice were served as a valuable model to explore the pathological mechanisms underlying inflammatory and immune disorders mediated by abnormal TNF-α-TRAF2 signaling and to develop new therapeutic targets. Methods : A vector targeting the knockout of the TRAF2 gene was constructed. Lead RNA and Cas9 Mrna were introduced into the fertilized eggs of C57BL/6JGpt mice through microinjection to mediate the TRAF2 gene mutation in mice. The mouse tail protein was extracted and the genotype of the F0 generation was determined by PCR and Western blot. TRAF2+/- mice were successfully obtained. F0 generation mice were backcrossed with C57BL/6JGpt wild-type mice to obtain stable TRAF2+/- mice for propagation and subsequent experiments. The body weight of TRAF2+/- mice was detected; Western blot was used to detect the expression of TRAF2 in the spleen, liver and kidney tissues of TRAF2+/- mice. The development of spleen, liver and kidney tissues in TRAF2+/- mice was detected by HE staining. Results : PCR identification using specific primers demonstrated that TRAF2+/- mice exhibited a target band at 679 bp. Western blot analysis results indicated that, compared with the WT group, the expression of TRAF2 in the tail protein of TRAF2+/- mice was significantly reduced(P+/- mice had a lower body weight compared to their littermate WT mice(P+/- mice was decreased(P+/- mice and WT mice. Conclusion The successful construction of TRAF2+/- mice has provided an important animal model for exploring the role of TRAF2 in developmental regulation, revealing the mechanism of inflammatory immune diseases mediated by abnormal TNF-α-TRAF2 signaling, and screening related drug targets.

Objective:This aims to investigate the diagnostic and evaluative value of MRI for lymphatic malformations in the head, neck, and facial regions of children. Methods:A retrospective analysis was conducted on the MRI imaging data of 31 cases of head, neck, and facial lymphatic malformations in children admitted to the Department of Otolaryngology, Head and Neck Surgery, Children's Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, from January 2022 to January 2024. Results:The MRI images of this group of cases primarily displayed irregular morphology（80.6%, 25/31）, thin-walled cysts（80.6%, 25/31）, and compression of surrounding tissues. The boundaries were clear（100%, 31/31）, with characteristics of invasive and drill-like growth（93.5%）. The cyst walls or internal septa exhibited high signal intensity on T1WI, low signal intensity on T2WI, and mild to moderate enhancement（100%）. The contents of the cysts showed low signal intensity on T1WI, high signal intensity on T2WI, and no enhancement（35.5%, 11/31）. Mixed signals with varying degrees of enhancement were observed in 20 cases（64.5%）. There were 29 cases of multilocular cysts（93.5%, 29/31）, and 11 cases of fluid-fluid levels（35.5%）. The MRI diagnostic accuracy for this group of cases was 100%. Conclusion:Lymphatic Malformations of head, neck and facial region in children have very characteristic features on MRI, such as typical thin wall, clear boundaries, irregular shapes, invasive growth, no enhancement, multilocular cystic masses, fluid-fluid level, etc. Furthermore, it is more appropriate for children with lymphatic malformations owing to its non-radiation and non-invasive benefits. Diagnosing lymphatic malformations in the head, neck, and facial region in children should begin with this.
Humans ; Retrospective Studies ; Lymphatic Abnormalities/diagnostic imaging* ; Magnetic Resonance Imaging ; Neck/diagnostic imaging* ; Head/diagnostic imaging* ; Face/diagnostic imaging* ; Child ; Male ; Female ; Child, Preschool ; Adolescent ; Infant

G protein-coupled receptor kinase 2 (GRK2) participates in the phosphorylation and desensitization of G protein-coupled receptor (GPCR), impacting various biological processes such as inflammation and cell proliferation. Dysregulated expression and activity of GRK2 have been reported in multiple cells in rheumatoid arthritis (RA). However, whether and how GRK2 regulates synovial hyperplasia and fibroblast-like synoviocytes (FLSs) proliferation is poorly understood. In this study, we investigated the regulation of GRK2 and its biological function in RA. We found that GRK2 transmembrane activity was increased in FLSs of RA patients and collagen-induced arthritis (CIA) rats. Additionally, we noted a positive correlation between high GRK2 expression on the cell membrane and serological markers associated with RA and CIA. Immunoprecipitation-mass spectrometry and pull-down analyses revealed tumor necrosis factor receptor-associated factor 2 (TRAF2) as a novel substrate of GRK2. Furthermore, surface plasmon resonance (SPR) and molecular docking assays determined that the C-terminus of GRK2 binds to the C-terminus of TRAF2 at the Gln340 residue. GRK2 knockdown and the GRK2 inhibitor CP-25 attenuated synovial hyperplasia and FLS proliferation in CIA both in vitro and in vivo by decreasing GRK2 membrane expression and activity. Mechanistically, increased GRK2 transmembrane activity contributed to the recruitment of TRAF2 on the cell membrane, promoting GRK2-TRAF2 interactions that facilitate the recruitment of the E3 ubiquitin ligase TRIM47 to TRAF2. This enhanced TRAF2 Lys63 polyubiquitylation and induced nuclear factor (NF)-κB activation, leading to synovial hyperplasia and abnormal proliferation of FLSs. Our study provides a mechanistic and preclinical rationale for further evaluation of GRK2 as a therapeutic target for RA.

Mitochondrial DNA (mtDNA) acts as a damage-associated molecular pattern to activate the stimulator of interferon genes (STING) signaling in macrophages, promoting tissue inflammation. However, its role in acute myocardial infarction (AMI) remains unclear. Macrophage-specific Sting1 knockout mice were used to validate STING's pathological role in AMI. Cardiac and liver mtDNA were used to activate macrophages in co-culture systems with cardiomyocytes to assess fibrosis and hypertrophy. Panaxatriol saponin (PTS) was tested for its ability to block mtDNA-driven macrophage activation and subsequent cardiomyocyte damage. STING-PTS binding ability was analyzed. AMI rats received PTS to evaluate its effects on myocardial inflammation and ventricular remodeling. In vivo, macrophage-specific Sting1 knockout reduced myocardial inflammation and injury after AMI. In vitro, mtDNA-activated macrophages induced cardiomyocyte fibrosis and hypertrophy through STING signaling. PTS suppressed mtDNA-driven macrophage activation by directly binding STING, thereby blocking inflammatory cascades. In AMI rats, PTS treatment attenuated acute inflammation and reversed ventricular remodeling. These findings establish the mtDNA-STING axis in macrophages as a critical driver of post-AMI inflammation and identify pharmacological STING inhibition with PTS as a promising therapeutic strategy. The study bridges genetic validation with translational applications, highlighting macrophage STING as a novel target for ischemic heart disease management.

Objective:To explore the association between greenspace morphology and the risk of severe fever with thrombocytopenia syndrome (SFTS) in Shandong Province.Methods:The SFTS cases data and land use data in Shandong province from 2018 to 2022 were collected, 6 landscape indices were calculated through Fragstats 4.2 software, and a negative binomial regression model was used to quantitatively evaluate the effect. Meteorological and socioeconomic factors during the same period were controlled as covariates. Subgroup analysis were conducted by age, gender and occupation. The percentage of relative risk[(RR-1)×100%] with 95% confidence interval (95% CI) was used as effect estimates. Results:There was a significant association between greenspace morphology and the risk of SFTS. Every 1% increase in the percentage, the mean area, fragmentation, aggregation, connectedness, and complexity of the shape of greenspace will lead to an increase in the risk of SFTS by 4.28% (95% CI: 2.88%-5.68%), 3.56% (95% CI: 1.06%-6.12%), 2.43% (95% CI: 1.38%-3.61%), 2.22% (95% CI: 1.39%-3.11%), 0.41% (95% CI: 0.26%-0.56%), 0.26% (95% CI: 0.10%-0.41%), respectively. There were statistically significant differences in the association between greenspace morphology and the risk of SFTS among different age and occupational subgroups. The elderly and farmers were the more susceptible to the changes in the percentage of greenspace and the complexity of the shape of greenspace. Conclusions:Greenspace morphology is related to the risk of SFTS. The elderly and farmers are vulnerable populations.

Background and Aims:Carotid plaque instability is a critical pathological basis for ischemic stroke.Identifying key molecular markers to evaluate plaque stability has important clinical implications.Recent studies have emphasized the regulatory roles and predictive value of long non-coding RNAs(lncRNAs)in plaque stability.In our previous transcriptome sequencing analysis of human stable and unstable carotid plaques,we identified lncRNA C-C motif chemokine ligand 3 antisense RNA 1(CCL3-AS1)as significantly upregulated in unstable plaques,suggesting a potential association with plaque instability.Therefore,this study aimed to validate CCL3-AS1 expression in an expanded plaque sample cohort and to explore its role and underlying molecular mechanism in carotid plaque destabilization.Methods:Carotid plaque specimens were obtained from patients undergoing carotid endarterectomy and classified into stable and unstable groups(n=15 per group)based on HE and Sirius red staining.qRT-PCR was used to validate the expression of candidate lncRNA CCL3-AS1.The localization and co-expression of CCL3-AS1 with macrophages in plaques were determined by RNA fluorescence in situ hybridization(FISH)combined with immunofluorescence staining.In vitro,THP1-derived macrophages were transduced with lentivirus or treated with antisense oligonucleotides(ASO)to overexpress or knock down CCL3-AS1,respectively,and the expression levels of inflammatory cytokines and matrix metalloproteinases(MMPs)were assessed.In vivo,an unstable carotid plaque model was established by tandem ligation of the right carotid artery in apolipoprotein E-deficient(ApoE-/-)mice,followed by local overexpression of CCL3-AS1.The effects on plaque morphology,macrophage infiltration,and MMP-9 expression were evaluated.Additionally,bioinformatic prediction using the catRAPID v2.1 omics platform was performed to identify potential RNA-binding proteins interacting with CCL3-AS1.RNA stability assays and RNA-binding protein immunoprecipitation(RIP)were conducted to verify the regulatory mechanism of MMP-9 expression.Results:CCL3-AS1 was significantly upregulated in unstable carotid plaques and was predominantly localized to the cytoplasm of plaque-infiltrating macrophages.In vitro,overexpression of CCL3-AS1 markedly increased the expression of MCP-1,TNF-α,IL-1β,iNOS,and MMP-9 in macrophages,whereas knockdown had the opposite effect.In the ApoE-/-mouse model of unstable carotid plaques,CCL3-AS1 overexpression led to fibrous cap rupture,increased infiltration of pro-inflammatory macrophages,enhanced MMP-9 secretion,and promoted plaque instability.Co-expression analysis revealed a strong correlation between CCL3-AS1 and MMP-9 expression(r=0.89,P=0.001).RNA stability assays demonstrated that CCL3-AS1 delayed the degradation of MMP-9 mRNA.Bioinformatic prediction identified heterogeneous nuclear ribonucleoprotein K(hnRNP-K)as a potential binding partner of CCL3-AS1.RIP and FISH co-localization confirmed the interaction,suggesting that CCL3-AS1 enhances MMP-9 mRNA stability through binding to hnRNP-K,thereby promoting its expression.Conclusion:As a macrophage-enriched inflammatory lncRNA,CCL3-AS1 may promote carotid plaque instability by enhancing MMP-9 expression via hnRNP-K-mediated mRNA stabilization.This lncRNA represents a potential molecular target for early intervention and stratification of ischemic stroke.

Background and Aims:Carotid plaque instability is a critical pathological basis for ischemic stroke.Identifying key molecular markers to evaluate plaque stability has important clinical implications.Recent studies have emphasized the regulatory roles and predictive value of long non-coding RNAs(lncRNAs)in plaque stability.In our previous transcriptome sequencing analysis of human stable and unstable carotid plaques,we identified lncRNA C-C motif chemokine ligand 3 antisense RNA 1(CCL3-AS1)as significantly upregulated in unstable plaques,suggesting a potential association with plaque instability.Therefore,this study aimed to validate CCL3-AS1 expression in an expanded plaque sample cohort and to explore its role and underlying molecular mechanism in carotid plaque destabilization.Methods:Carotid plaque specimens were obtained from patients undergoing carotid endarterectomy and classified into stable and unstable groups(n=15 per group)based on HE and Sirius red staining.qRT-PCR was used to validate the expression of candidate lncRNA CCL3-AS1.The localization and co-expression of CCL3-AS1 with macrophages in plaques were determined by RNA fluorescence in situ hybridization(FISH)combined with immunofluorescence staining.In vitro,THP1-derived macrophages were transduced with lentivirus or treated with antisense oligonucleotides(ASO)to overexpress or knock down CCL3-AS1,respectively,and the expression levels of inflammatory cytokines and matrix metalloproteinases(MMPs)were assessed.In vivo,an unstable carotid plaque model was established by tandem ligation of the right carotid artery in apolipoprotein E-deficient(ApoE-/-)mice,followed by local overexpression of CCL3-AS1.The effects on plaque morphology,macrophage infiltration,and MMP-9 expression were evaluated.Additionally,bioinformatic prediction using the catRAPID v2.1 omics platform was performed to identify potential RNA-binding proteins interacting with CCL3-AS1.RNA stability assays and RNA-binding protein immunoprecipitation(RIP)were conducted to verify the regulatory mechanism of MMP-9 expression.Results:CCL3-AS1 was significantly upregulated in unstable carotid plaques and was predominantly localized to the cytoplasm of plaque-infiltrating macrophages.In vitro,overexpression of CCL3-AS1 markedly increased the expression of MCP-1,TNF-α,IL-1β,iNOS,and MMP-9 in macrophages,whereas knockdown had the opposite effect.In the ApoE-/-mouse model of unstable carotid plaques,CCL3-AS1 overexpression led to fibrous cap rupture,increased infiltration of pro-inflammatory macrophages,enhanced MMP-9 secretion,and promoted plaque instability.Co-expression analysis revealed a strong correlation between CCL3-AS1 and MMP-9 expression(r=0.89,P=0.001).RNA stability assays demonstrated that CCL3-AS1 delayed the degradation of MMP-9 mRNA.Bioinformatic prediction identified heterogeneous nuclear ribonucleoprotein K(hnRNP-K)as a potential binding partner of CCL3-AS1.RIP and FISH co-localization confirmed the interaction,suggesting that CCL3-AS1 enhances MMP-9 mRNA stability through binding to hnRNP-K,thereby promoting its expression.Conclusion:As a macrophage-enriched inflammatory lncRNA,CCL3-AS1 may promote carotid plaque instability by enhancing MMP-9 expression via hnRNP-K-mediated mRNA stabilization.This lncRNA represents a potential molecular target for early intervention and stratification of ischemic stroke.

Objective:To explore the association between greenspace morphology and the risk of severe fever with thrombocytopenia syndrome (SFTS) in Shandong Province.Methods:The SFTS cases data and land use data in Shandong province from 2018 to 2022 were collected, 6 landscape indices were calculated through Fragstats 4.2 software, and a negative binomial regression model was used to quantitatively evaluate the effect. Meteorological and socioeconomic factors during the same period were controlled as covariates. Subgroup analysis were conducted by age, gender and occupation. The percentage of relative risk[(RR-1)×100%] with 95% confidence interval (95% CI) was used as effect estimates. Results:There was a significant association between greenspace morphology and the risk of SFTS. Every 1% increase in the percentage, the mean area, fragmentation, aggregation, connectedness, and complexity of the shape of greenspace will lead to an increase in the risk of SFTS by 4.28% (95% CI: 2.88%-5.68%), 3.56% (95% CI: 1.06%-6.12%), 2.43% (95% CI: 1.38%-3.61%), 2.22% (95% CI: 1.39%-3.11%), 0.41% (95% CI: 0.26%-0.56%), 0.26% (95% CI: 0.10%-0.41%), respectively. There were statistically significant differences in the association between greenspace morphology and the risk of SFTS among different age and occupational subgroups. The elderly and farmers were the more susceptible to the changes in the percentage of greenspace and the complexity of the shape of greenspace. Conclusions:Greenspace morphology is related to the risk of SFTS. The elderly and farmers are vulnerable populations.

Objective:To explore the application value of laparoscopic contrast enhanced ultrasound in laparoscopic surgery for patients with hepatocellular carcinoma combined with cirrhosis.Methods:The clinical data of 71 patients with hepatocellular carcinoma combined cirrhosis from February 2018 to February 2020 in Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology were retrospectively analyzed. The patients underwent preoperative enhanced CT and multi-parameter MRI examination, followed by laparoscopic partial hepatectomy, and intraoperative laparoscopic contrast enhanced ultrasound examination. Based on histological examination and follow-up results, the diagnostic efficacy of preoperative imaging and preoperative imaging combined with intraoperative laparoscopic contrast enhanced ultrasound in patients with hepatocellular carcinoma combined with cirrhosis was compared.Results:Among the 71 patients, 69 completed laparoscopic surgery and 2 converted to open surgery. One hundred and ten HCC lesions were diagnosed by preoperative imaging examination, 105 lesions were detected by intraoperative ultrasound among them, of which 98 lesions were diagnosed as HCC by intraoperative laparoscopic contrast enhanced ultrasound. There were no statistically significant difference in sensitivity, specificity, accuracy, positive predictive value and negative predictive value between preoperative imaging and preoperative imaging combined with intraoperative laparoscopic contrast enhanced ultrasound in the diagnosis of malignant liver lesions: 94.4% (102/108) vs. 99.1% (107/108), 81.0% (34/42) vs. 66.7% (28/42), 90.6% (136/150) vs. 90.0% (135/150), 92.7% (102/110) vs. 88.4% (107/121) and 85.0% (34/40) vs. 96.6% (28/29), P>0.05. Laparoscopic contrast enhanced ultrasound revealed an additional 11 suspected malignant lesions, of which 5 lesions were histologically confirmed as HCC. Seven patients underwent surgical strategy changes. Conclusions:Laparoscopic contrast enhanced ultrasound in patients with HCC combined with cirrhosis during laparoscopic surgery can be used to detect, identify, accurately locate of the lesions and modify the surgical plan.

Objective To explore the public attitude towards kidney xenotransplantation in China by constructing and validating the prediction model based on xenotransplantation questionnaire. Methods A convenient sampling survey was conducted among the public in China with the platform of Wenjuanxing to analyze public acceptance of kidney xenotransplantation and influencing factors. Using random distribution method, all included questionnaires (n=2 280) were divided into the training and validation sets according to a ratio of 7:3. A prediction model was constructed and validated. Results A total of 2 280 questionnaires were included. The public acceptance rate of xenotransplantation was 71.3%. Multivariate analysis showed that gender, marital status, resident area, medical insurance coverage, religious belief, vegetarianism, awareness of kidney xenotransplantation and whether on the waiting list for kidney transplantation were the independent influencing factors for public acceptance of kidney xenotransplantation (all P<0.05). The area under the curve (AUC) of receiver operating characteristic (ROC) of the prediction model in the training set was 0.773, and 0.785 in the validation set. The calibration curves in the training and validation sets indicated that the prediction models yielded good prediction value. Decision curve analysis (DCA) suggested that the prediction efficiency of the model was high. Conclusions In China, public acceptance of kidney xenotransplantation is relatively high, whereas it remains to be significantly enhanced. The prediction model based on questionnaire survey has favorable prediction efficiency, which provides reference for subsequent research.