Aim To identify the underlying target of bullatine A(BA)against rheumatoid arthritis(RA)u-sing limited proteolysis-small molecule mapping(LiP-SMap)drug target proteomics and to provide a scientif-ic basis for clinical application of Aconiti brachypodi Radix in the treatment of RA.Methods LiP-SMap drug target proteomics was employed to perform bioin-formatics analysis for comparing and validating the dif-ferential protein expression after BA intervention.A collagen-induced arthritis(CIA)model was estab-lished in DBA/1 mice using bovine type Ⅱ collagen.The mice were then divided into the CIA model group,methotrexate-positive control group(MTX group),and BA groups(10 mg·kg-1 and 20 mg·kg-1)based on their clinical scores.After drug intervention,the thera-peutic efficacy against RA was assessed by joint index scores and foot thickness measurements.Histopatholog-ical changes in the arthritic joints of CIA mice were e-valuated using hematoxylin and eosin(HE)staining.Enzyme-linked immunosorbent assay(ELISA)was employed to detect inflammatory cytokines interleukin-17(IL-17)and total IgG and IgG3 anti-collagen-spe-cific antibodies levels from the serum of CIA mice.Flow cytometry was used to detect the expression levels of intracellular Th17 cells(IL-17+CD4+T cells)and Th1 cells(IFN-γ+CD4+T cells).Fluorescent quanti-tative PCR was performed to detect the expression of genes related to differential proteins.Results The proteomic analysis identified Serpinb1a as a protein with strong binding affinity to BA,and KEGG enrich-ment analysis indicated IL-17 signaling pathway was a crucial pathway of BA in against RA.BA treatment significantly reduced clinical scores and foot thickness,improved local arthritis symptoms in CIA mice,and al-leviated inflammatory cell infiltration into arthritic joints(P＜0.05).Differential protein validation re-sults showed that BA had strong affinity with Serpinb1a(-5.92 kJ·mol-1)and downregulated the expres-sion of Serpinb1a mRNA.Furthermore,the administra-tion of BA markedly reduced serum IL-17 A levels from CIA mice,inhibited the expression of intracellular IL-17 A and IFN-γ cytokines in splenic CD4+T cells(P＜0.05),and significantly downregulated the transcrip-tional expression of IL-17F(P＜0.05).Conclusion BA exhibits therapeutic effects on collagen-induced arthritis,and its mechanism of action may involve the regulation of Serpinb1a and the IL-17 signaling path-way.

AIM:To investigate the effects of macrophage galactose-type lectin 1(Mgl1)gene deletion on he-matopoietic stem/progenitor cells(HSPCs)under steady-state conditions and inflammation.METHODS:Mice were di-vided into a control group(wild-type)and an experimental group(Mgl1 gene-deleted).Flow cytometry was used to ana-lyze the proportions of various hematopoietic cell lineages in the peripheral blood and bone marrow of both groups,assess-ing the impact of Mgl1 gene deletion on steady-state hematopoiesis(n=3～4).Transplantation and colony-forming assays were utilized to evaluate the effects of Mgl1 gene deletion onthe repopulation capacity and colony-forming ability of HSPCs(n=5).The LPS-induced inflammation model was employed to examine the effects of Mgl1 gene deletion on the inflamma-tory response of HSPCs both in vitro and in vivo(n=5～8).Western blot and RT-qPCR were conducted to analyze the alter-ations in signaling pathways regulated by Mgl1 in the inflammatory response of HSPCs(n=3).RESULTS:(1)Mgl1 gene deletion had no significant effecton steady-state hematopoiesis(P>0.05).(2)Mgl1 gene deletion promoted inflam-mation-induced cell differentiation of HSPCs(P<0.01).(3)Mgl1 gene deletion accelerated the exhaustion of HSPCs un-der prolonged inflammatory conditions(P<0.01).(4)Mgl1 was found to regulate the inflammatory response of HSPCs via the NF-κB signaling pathway.CONCLUSION:Mgl1 gene deletion enhances the inflammatory response of HSPCs via the NF-κB signaling pathway.

Loss of protein homeostasis is a hallmark of cellular senescence, and ribosome pausing plays a crucial role in the collapse of proteostasis. However, our understanding of ribosome pausing in senescent cells remains limited. In this study, we utilized ribosome profiling and G-quadruplex RNA immunoprecipitation sequencing techniques to explore the impact of RNA G-quadruplex (rG4) on the translation efficiency in senescent cells. Our results revealed a reduction in the translation efficiency of rG4-rich genes in senescent cells and demonstrated that rG4 structures within coding sequence can impede translation both in vivo and in vitro. Moreover, we observed a significant increase in the abundance of rG4 structures in senescent cells, and the stabilization of the rG4 structures further exacerbated cellular senescence. Mechanistically, the RNA helicase DHX9 functions as a key regulator of rG4 abundance, and its reduced expression in senescent cells contributing to increased ribosome pausing. Additionally, we also observed an increased abundance of rG4, an imbalance in protein homeostasis, and reduced DHX9 expression in aged mice. In summary, our findings reveal a novel biological role for rG4 and DHX9 in the regulation of translation and proteostasis, which may have implications for delaying cellular senescence and the aging process.
G-Quadruplexes ; Cellular Senescence ; Ribosomes/genetics* ; Humans ; Animals ; Mice ; DEAD-box RNA Helicases/genetics* ; Protein Biosynthesis ; RNA/chemistry* ; Neoplasm Proteins

BACKGROUND: Hepatic inflammatory cell accumulation and the subsequent systematic inflammation drive acute-on-chronic liver failure (ACLF) development. Previous studies showed that the vagus nerve exerts anti-inflammatory activity in many inflammatory diseases. Here, we aimed to identify the key molecule mediating the inflammatory process in ACLF and reveal the neuroimmune communication arising from the vagus nerve and immunological disorders of ACLF. METHODS: Proteomic analysis was performed and validated in ACLF model mice or patients, and intervention animal experiments were conducted using neutralizing antibodies. PNU-282987 (acetylcholine receptor agonist) and vagotomy were applied for perturbing vagus nerve activity. Single-cell RNA sequencing (scRNA-seq), flow cytometry, immunohistochemical and immunofluorescence staining, and clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) technology were used for in vivo or in vitro mechanistic studies. RESULTS: The unbiased proteomics identified C-X-C motif chemokine ligand 9 (CXCL9) as the greatest differential protein in the livers of mice with ACLF and its relation to the systematic inflammation and mortality were confirmed in patients with ACLF. Interventions on CXCL9 and its receptor C-X-C chemokine receptor 3 (CXCR3) improved liver injury and decreased mortality of ACLF mice, which were related to the suppressing of hepatic immune cells' accumulation and activation. Vagus nerve stimulation attenuated while vagotomy aggravated the expression of CXCL9 and the severity of ACLF. Blocking CXCL9 and CXCR3 ameliorated liver inflammation and increased ACLF-associated mortality in ACLF mice with vagotomy. scRNA-seq revealed that hepatic macrophages served as the major source of CXCL9 in ACLF and were validated by immunofluorescence staining and flow cytometry analysis. Notably, the expression of CXCL9 in macrophages was modulated by vagus nerve-mediated cholinergic signaling. CONCLUSIONS Our novel findings highlighted that the neuroimmune communication of the vagus nerve-macrophage-CXCL9 axis contributed to ACLF development. These results provided evidence for neuromodulation as a promising approach for preventing and treating ACLF.
Animals ; Mice ; Chemokine CXCL9/metabolism* ; Vagus Nerve/physiology* ; Acute-On-Chronic Liver Failure/metabolism* ; Humans ; Male ; Mice, Inbred C57BL ; Proteomics ; Flow Cytometry ; Receptors, CXCR3/metabolism*

Bone grafting is a primary method for treating bone defects. Among various graft materials, xenogeneic bone substitutes are widely used in clinical practice due to their abundant sources, convenient processing and storage, and avoidance of secondary surgeries. With the advancement of domestic production and the limitations of imported products, an increasing number of bone filling or grafting substitute materials isentering clinical trials. Relevant experts have drafted this consensus to enhance the management of medical device clinical trials, protect the rights of participants, and ensure the scientific and effective execution of trials. It summarizes clinical experience in aspects, such as design principles, participant inclusion/exclusion criteria, observation periods, efficacy evaluation metrics, safety assessment indicators, and quality control, to provide guidance for professionals in the field.
Humans ; Bone Substitutes/therapeutic use* ; Randomized Controlled Trials as Topic/methods* ; Consensus ; Bone Transplantation ; Research Design

Objective To investigate the effects and regulatory mechanisms of carboxyamidotriazole orotate(CTO)on the metabolism and inflammatory mediators of glioma-associated microglia(GAM).Methods Tumor volume was regularly monitored by in vivo imaging,and histological examination was performed to detect the extent of tumor in-filtration;non-targeted metabolomics analysis was used to detect the level of tricarboxylic acid cycle metabolites in cells;seahorse cell energy measurement method was used to detect the oxygen consumption rate(OCR)and extra-cellular acidification rate(ECAR)of cells;immunofluorescence was used to detect the degree of hypoxia in cells;quantitative PCR was used to detect the mRNA level of pro-cancer mediators M1/M2 in cells;Western blot was used to detect the protein level of hypoxia-inducible factor-1α(HIF-1α)and programmed death receptor-ligand 1(PD-L1).Results CTO inhibited the tumor progression in mice,and down-regulated the oxidative phosphorylation level and improved cell hypoxia in vitro(P＜0.01).It also downregulated the expression of pro-oncogenic mediators iNos,Arg-1,Il-10,and Irf4 in GAM(P＜0.01).When combined with lactate dehydrogenase inhibitor stiripentol(STP),CTO-induced enhancement of glycolysis and upregulation of PD-L1 expression in GAM was attenuated(P＜0.01),and the expressions of Arg-1 and Il-10 was further downregulated(P＜0.000 1).Conclusions CTO down-regulates the expression of several oncogenic genes in GAM and inhibits tumor progression in mice.Combined use of lactate dehydrogenase inhibitors can weaken the adverse effect of CTO and reduce the transcriptional level of GAM oncogenic mediators.

Objective To explore the impact of electrostimulation biofeedback therapy combined with pelvic floor muscle training on postpartum pelvic floor function in first vaginal birth women.Methods A retrospective analysis was conducted on the clinical data of in first vaginal birth women.The control group(n=49)received pelvic floor muscle training,while the observation group(n=49)received electrostimulation biofeedback therapy in addition to pelvic floor muscle training.Com-parisons were made between the two groups before treatment and 10 weeks after treatment in terms of uterine artery blood flow indicators[peak systolic velocity(PSV),resistance index(RI),and pulsa-tility index(PI)of uterine artery],serological indicators[C-reactive protein(CRP),interleukin-6(IL-6)],pelvic floor ultrasound indicators(bladder neck mobility,anteroposterior diameter of levator hi-atal area,levator hiatal volume),pelvic floor function[Pelvic Floor Distress Inventory-20(PFDI-20)],pelvic floor muscle strength,and quality of life[Short Form-36 Health Survey(SF-36)].The costs of the two treatments were compared at the end of treatment.Results Ten weeks after treatment,PSV,muscle potential values of type Ⅰ and type Ⅱ muscles,and SF-36 scores were significantly increased in both groups compared with those before treatment,with the observation group showing higher values than the control group(P＜0.05).RI,PI,CRP,IL-6,bladder neck mobility,anteroposterior di-ameter of levator hiatal area,levator hiatal volume,and PFDI-20 scores were decreased in both groups compared with those before treatment,with the observation group showing lower values than the control group(P＜0.05).After treatment,the treatment cost in the observation group was significantly higher than that in the control group(P＜0.05).Conclusion Electrostimulation biofeedback ther-apy combined with pelvic floor muscle training can effectively improve postpartum uterine artery blood flow indicators,serological indicators,pelvic floor ultrasound indicators,pelvic floor muscle strength,and promote pelvic floor function recovery in first vaginal delivery women.

Aim To identify the underlying target of bullatine A(BA)against rheumatoid arthritis(RA)u-sing limited proteolysis-small molecule mapping(LiP-SMap)drug target proteomics and to provide a scientif-ic basis for clinical application of Aconiti brachypodi Radix in the treatment of RA.Methods LiP-SMap drug target proteomics was employed to perform bioin-formatics analysis for comparing and validating the dif-ferential protein expression after BA intervention.A collagen-induced arthritis(CIA)model was estab-lished in DBA/1 mice using bovine type Ⅱ collagen.The mice were then divided into the CIA model group,methotrexate-positive control group(MTX group),and BA groups(10 mg·kg-1 and 20 mg·kg-1)based on their clinical scores.After drug intervention,the thera-peutic efficacy against RA was assessed by joint index scores and foot thickness measurements.Histopatholog-ical changes in the arthritic joints of CIA mice were e-valuated using hematoxylin and eosin(HE)staining.Enzyme-linked immunosorbent assay(ELISA)was employed to detect inflammatory cytokines interleukin-17(IL-17)and total IgG and IgG3 anti-collagen-spe-cific antibodies levels from the serum of CIA mice.Flow cytometry was used to detect the expression levels of intracellular Th17 cells(IL-17+CD4+T cells)and Th1 cells(IFN-γ+CD4+T cells).Fluorescent quanti-tative PCR was performed to detect the expression of genes related to differential proteins.Results The proteomic analysis identified Serpinb1a as a protein with strong binding affinity to BA,and KEGG enrich-ment analysis indicated IL-17 signaling pathway was a crucial pathway of BA in against RA.BA treatment significantly reduced clinical scores and foot thickness,improved local arthritis symptoms in CIA mice,and al-leviated inflammatory cell infiltration into arthritic joints(P＜0.05).Differential protein validation re-sults showed that BA had strong affinity with Serpinb1a(-5.92 kJ·mol-1)and downregulated the expres-sion of Serpinb1a mRNA.Furthermore,the administra-tion of BA markedly reduced serum IL-17 A levels from CIA mice,inhibited the expression of intracellular IL-17 A and IFN-γ cytokines in splenic CD4+T cells(P＜0.05),and significantly downregulated the transcrip-tional expression of IL-17F(P＜0.05).Conclusion BA exhibits therapeutic effects on collagen-induced arthritis,and its mechanism of action may involve the regulation of Serpinb1a and the IL-17 signaling path-way.

AIM:To investigate the effects of macrophage galactose-type lectin 1(Mgl1)gene deletion on he-matopoietic stem/progenitor cells(HSPCs)under steady-state conditions and inflammation.METHODS:Mice were di-vided into a control group(wild-type)and an experimental group(Mgl1 gene-deleted).Flow cytometry was used to ana-lyze the proportions of various hematopoietic cell lineages in the peripheral blood and bone marrow of both groups,assess-ing the impact of Mgl1 gene deletion on steady-state hematopoiesis(n=3～4).Transplantation and colony-forming assays were utilized to evaluate the effects of Mgl1 gene deletion onthe repopulation capacity and colony-forming ability of HSPCs(n=5).The LPS-induced inflammation model was employed to examine the effects of Mgl1 gene deletion on the inflamma-tory response of HSPCs both in vitro and in vivo(n=5～8).Western blot and RT-qPCR were conducted to analyze the alter-ations in signaling pathways regulated by Mgl1 in the inflammatory response of HSPCs(n=3).RESULTS:(1)Mgl1 gene deletion had no significant effecton steady-state hematopoiesis(P>0.05).(2)Mgl1 gene deletion promoted inflam-mation-induced cell differentiation of HSPCs(P<0.01).(3)Mgl1 gene deletion accelerated the exhaustion of HSPCs un-der prolonged inflammatory conditions(P<0.01).(4)Mgl1 was found to regulate the inflammatory response of HSPCs via the NF-κB signaling pathway.CONCLUSION:Mgl1 gene deletion enhances the inflammatory response of HSPCs via the NF-κB signaling pathway.

Objective:To investigate the prognosis and influencing factors of endoscopic surgery for early glottic carcinoma.Methods:In this retrospective study, we applied the Cox proportional hazards regression model and the random survival forest model to analyze the clinical characteristics of 385 patients [362 males, 23 females, age ranging from 33 to 91 years (62.0±9.6)] who visited the Sixth Medical Center of the General Hospital of the People's Liberation Army from January 2009, to December 2022 and diagnosed with early glottic carcinoma, encompassing variables such as age, gender, T stage, surgical approach, pathological typing, etc. The primary evaluation indicators were overall survival(OS) and disease-free survival rates (DFS). The follow-up duration ranged from 30 to 5,557 days (with a median follow-up time of 1,596 days).Results:After a three-year follow-up, the OS rate for the 385 patients was 95.83%, while the DSF rate was 82.98%. The Cox proportional hazards regression analysis revealed age ( HR=2.35, 95% CI: 1.75 to 3.15, P<0.001) and T staging ( HR=1.59, 95% CI: 1.13 to 2.23, P=0.019) as predominant factors affecting the OS and DFS. The random survival forest model identified poor tumor differentiation, and high expression of P53 and Ki-67 as predictors of inferior prognosis. Conclusion:Endoscopic surgery for early glottic carcinoma yields favorable short-term OS and reduces short-term recurrence rates, with T-stage emerging as a pivotal factor influencing recurrence. Tumors with poor differentiation and elevated expression of P53 may be indicative of an increased risk of recurrence.