Objective To investigate the role and potential mechanism of secreted phosphoprotein 1 (SPP1)⁺ macrophages in the formation of chronic renal allograft fibrosis. Methods The expression features of SPP1⁺ macrophages in renal allografts of chronic allograft dysfunction (CAD) patients were analyzed based on single-cell transcriptome data of renal tissues from patients with CAD. Transcription factor VIPER analysis and DoRothEA transcription factor activity analysis were performed on the single-cell transcriptome data. Renal tissue samples were collected from kidney transplant recipients, including the CAD group (n=5) and the non-renal allograft fibrosis group (CTL group, n=5). A mouse model of chronic allograft rejection was established and divided into the allogeneic kidney transplantation group (CAD group, n=3) and the syngeneic kidney transplantation group (SYN group, n=3). Hematoxylin-eosin staining was used to detect renal tissue injury in mice, and Masson staining was used to detect renal tissue fibrosis. Immunofluorescence staining was performed to detect SPP1 expression in renal tissues of transplant recipients and mouse renal allografts. Bone marrow-derived macrophages (BMDMs) were extracted from mice and subjected to hypoxia stimulation. The expression of hypoxia-inducible factor (HIF)-1α and SPP1 was detected by Western blot, and SPP1 expression was detected by flow cytometry. BMDMs were transfected with HIF-1α overexpression plasmid and HIF-1α small interfering RNA (siRNA) followed by hypoxia intervention, and the expression of HIF-1α and SPP1 was detected by Western blot. Mouse aortic endothelial cells (MAECs) were co-cultured with the supernatant of BMDMs, and the expression of endothelial-mesenchymal transition (EndMT)-related markers was detected by Western blot and immunofluorescence. Results Single-cell transcriptome analysis showed that the proportion of SPP1⁺ macrophages in renal allograft tissues was significantly higher in the CAD group than in the CTL group (P<0.05). The renal injury score and the percentage of interstitial fibrotic area in the CAD group were significantly higher than those in the SYN group (both P<0.05). Immunofluorescence staining showed that the proportion of SPP1⁺ macrophages was increased in the CAD group compared with the CTL group, and also increased in the CAD group compared with the SYN group (both P<0.05). VIPER analysis and DoRothEA transcription factor activity analysis revealed activation of the hypoxia pathway and upregulated expression of transcription factors such as HIF-1α in SPP1⁺ macrophages. SPP1 expression was elevated in BMDMs under hypoxic conditions. Knockdown of HIF-1α inhibited hypoxia-induced SPP1 protein expression, whereas overexpression of HIF-1α upregulated SPP1 protein levels. After co-culture of hypoxia-induced BMDMs with MAECs, the expression levels of EndMT-related markers were increased. Conclusions SPP1⁺ macrophages differentiated under hypoxia are significantly infiltrated in the formation of chronic renal allograft fibrosis, and may promote renal allograft fibrosis by inducing EndMT in renal vascular endothelial cells.

Objective:To investigate the possible mechanisms underlying the involvement of the gut microbiota metabolite trimethylamine oxide (TMAO) in the pathogenesis of chronic spontaneous urticaria (CSU) .Methods:From June 2023 to June 2024, 67 CSU patients were enrolled from the Dermatology and Cosmetic Center, the Third Affiliated Hospital of Chongqing Medical University, and 69 age-matched healthy controls were also collected at the same time. Serum TMAO levels in both groups were measured using enzyme-linked immunosorbent assay (ELISA) , and D-dimer levels were collected from the CSU patients. A degranulation model was established in rat basophilic leukemia RBL-2H3 cells using anti-DNP IgE/DNP-BSA (IgE/Ag group) ; these cells were additionally grouped to be treated with different concentrations of TMAO (IgE/Ag+10 μmol/L TMAO group, IgE/Ag + 50 μmol/L TMAO group, IgE/Ag + 100 μmol/L TMAO group) ; untreated RBL-2H3 cells served as a blank control group. To investigate the effect of the extracellular signal-regulated kinase (ERK) phosphorylation inhibitor U0126 on the action of TMAO, RBL-2H3 cells were divided into another 5 groups: blank group, IgE/Ag group, IgE/Ag + 1 μmol/L U0126 group, IgE/Ag + 100 μmol/L TMAO group, and IgE/Ag + 100 μmol/L TMAO + 1 μmol/L U0126 group. In vivo, a localized allergic reaction model was established in the ears of C57BL/6 mice using anti-DNP IgE/DNP-BSA (IgE/Ag group) , and additional groups included blank group, IgE group, IgE/Ag + solvent (DMSO) group, and IgE/Ag + 10 μg/μl TMAO group. ELISA was performed to detect levels of inflammatory mediators in cell culture supernatants and mouse serum. Toluidine blue staining was employed to observe mast cell degranulation in the cell experiment and mouse ear tissue samples, Evans blue staining to assess vascular permeability in mouse ear tissue samples, and Western blot analysis to detect the ERK phosphorylation levels. The t test was used for comparisons between two groups, and one-way analysis of variance for multiple comparisons. Results:Serum TMAO levels were significantly higher in the 67 CSU patients than in the 69 healthy controls ( t = 13.27, P < 0.001) . Among the 32 CSU patients with available data about D-dimer, serum TMAO levels were positively correlated with D-dimer levels ( r = 0.62, P < 0.001) . In RBL-2H3 cell experiments, degranulation rates were significantly higher in the IgE/Ag + 10, 50, and 100 μmol/L TMAO groups than in the IgE/Ag group; morphologically, RBL-2H3 cells treated with 10, 50, and 100 μmol/L TMAO became increasingly rounded; 50 and 100 μmol/L TMAO significantly promoted the production of β-hexosaminidase (β-Hex) , interleukin-6 (IL-6) , and tumor necrosis factor-α (TNF-α) (all P < 0.01) , and upregulated ERK phosphorylation levels ( P < 0.01) ; the levels of ERK phosphorylation, IL-6, TNF-α, and β-Hex were significantly lower in the IgE/Ag + U0126 group than in the IgE/Ag group, as well as lower in the IgE/Ag + TMAO + U0126 group than in the IgE/Ag + TMAO group (all P < 0.001) . In the mouse model of localized allergic reaction, the IgE/Ag + TMAO group showed increased vascular permeability, edema degree, and mast cell degranulation, as well as significantly elevated ERK phosphorylation levels and TNF-α expression in mouse ear tissues compared with the IgE/Ag + DMSO group (both P < 0.05) . Conclusion:Elevated serum TMAO may participate in the pathogenesis of CSU by upregulating ERK phosphorylation levels in mast cells and skin tissues, thereby promoting IgE/Ag-mediated degranulation of effector cells and production of inflammatory mediators.

Objective:To explore the influence of initial high-risk cytogenetic abnormalities on the outcomes of children with acute myeloid leukemia (AML) after post-transplant Cyclophosphamide (PTCy)-based allogeneic hematopoietic stem cell transplantation (allo-HSCT).Methods:A retrospective cohort study.AML children who underwent PTCy-based allo-HSCT after the first complete remission at Wuhan Children′s Hospital, Tongji Medical College, Huazhong University of Science and Technology between April 2017 and April 2024 were enrolled.Patients were divided into intermediate-risk and high-risk groups based on their initial cytogenetic features.These patients were further divided into complex karyotype, 11q23 rearrangement, and other karyotype groups.Clinical characteristics and survival outcomes were compared among these groups.Measurement and count data were analyzed using Wilcoxon rank-sum/Kruskal-Wallis and χ2 tests, respectively.Survival and risk factor analyses were performed using Kaplan-Meier and Cox proportional hazards methods, respectively. Results:A total of 51 AML children who underwent allo-HSCT were included in this study.The median age at transplantation was 3.2 years and the median follow-up time was 4.6 years.There were 26 cases in the intermediate-risk group and 25 cases in the high-risk group; 8 cases in the complex karyotype group, 14 cases in the 11q23 rearrangement group, and 29 cases in the other karyotype groups.By the end of the follow-up on November 30, 2024, 11 patients relapsed, 8 patients died, and 13 patients developed grades Ⅱ-Ⅳ acute graft-versus-host disease (GVHD).The 3-year overall survival (OS), relapse-free survival (RFS), and grades Ⅱ-Ⅳ acute GVHD-free and relapse-free survival (GRFS) were 84.0% (95% CI: 74.4%-94.8%), 74.5% (95% CI: 63.4%-87.5%), and 58.8% (95% CI: 46.7%-74.0%), respectively.The 3-year OS of the high-risk group was significantly lower than that of the intermediate-risk group (71.8% vs.96.2%, P=0.022), while differences in 3-year RFS and GRFS between the 2 groups were not statistically significant (68.0% vs.80.8%, P=0.400; 52.0% vs.65.4%, P=0.420).The 3-year OS, RFS and GRFS of the complex karyotype group were significantly lower than those of 11q23 rearrangement and other karyotype groups (50.0% vs.85.7%, 93.1%, P=0.009; 37.5% vs.85.7%, 79.3%, P=0.022; 25.0% vs.64.3%, 65.5%, P=0.049).Multivariate analysis showed that a complex karyotype was an independent prognostic factor affecting 3-year OS and GRFS [OS: HR=6.79 (95% CI: 1.13-43.80), P=0.044; GRFS: HR=3.72(95% CI: 1.13-12.20), P=0.030]. Conclusions:High-risk cytogenetic features are significant predictors of survival outcomes in pediatric AML patients undergoing PTCy-based allo-HSCT.

Aim To investigate whether protein inhibitor of activated STAT3(PIAS3)deficiency exacerbates the occurrence and development of atherosclerosis(As)in female ApoE knockout mice.Methods PIAS3 gene knockout mice with ApoE-/-background(PIAS3-/-/ApoE-/-)and their littermate PIAS3+/+/ApoE-/-mice were bred and fed with a high-fat/high-cholesterol diet for 12 weeks to induce As.Body weight(every week)and plasma lipid levels including to-tal cholesterol,triglyceride,high density lipoprotein cholesterol and non-high density lipoprotein cholesterol(every 4 weeks)of the mice were measured.Oil red O staining,HE staining,immunohistochemistry staining and immunofluores-cence staining were performed on mouse aortic tree and frozen sections of aortic root to evaluate the area,cellular composi-tion and stability of As plaques.Moreover,the expression of estrogen receptor α(ERα)and its co-localization with vas-cular smooth muscle cells(VSMC)in plaques were determined by immunofluorescence staining.Results Compared with PIAS3+/+/ApoE-/-mice,PIAS3-/-/ApoE-/-mice showed no significant differences in body weight,major organ weight(heart,liver,spleen,kidney and epididymal fat)and plasma lipid levels;however,PIAS3 deficiency promoted the forma-tion of As in female PIAS3-/-/ApoE-/-mice.Compared with PIAS3+/+/ApoE-/-mice,PIAS3-/-/ApoE-/-mice showed an increased lipid accumulation and a decreased VSMC content in As plaques(P＜0.05),leading to a decrease in plaque stability.In addition,the expression of ERα in the As plaques of PIAS3-/-/ApoE-/-mice was significantly downregulated(P＜0.05),and there was a obvious co-localization between ERα and VSMC.The reduction of VSMC content in PIAS3-/-/ApoE-/-mouse plaques might lead to a decrease of ERα expression,thereby weakening the anti-As effect of es-trogen.Conclusion PIAS3 deficiency exacerbates the formation of As plaques in female PIAS3-/-/ApoE-/-mice,which might be due to the regulatory effect of PIAS3 on ERα expression in plaques.

OBJECTIVE: To investigate the mechanism of circular RNA RAD18 (CircRAD18 ) in regulating daunorubicin (DNR) resistance in acute myeloid leukemia (AML) cells through the miR-185-5p/hepatoma-derived growth factor ( HDGF) axis. METHODS: Real-time fluorescence quantitative PCR and immunoblotting were applied to detect the expression of CircRAD18 , miR-185-5p, and HDGF in human AML cell lines HL-60, U937, and human AML drug-resistant cell line KG1a. KG1a cells were cultured in vitro and randomly divided into control group, DNR group, DNR+negative control group, DNR+CircRAD18 knockdown group, and DNR+CircRAD18 knockdown+miR-185-5p inhibitor group. After transfection, real-time fluorescence quantitative PCR and immunoblotting were applied to detect the expression of CircRAD18 , miR-185-5p, and HDGF of cells, CCK-8 method and Ki-67 immunofluorescence staining were applied to detect cell proliferation, flow cytometry was applied to detect cell apoptosis, and immunoblotting was applied to detect the expression of cell proliferation, apoptosis and drug resistance related proteins in each group. The double luciferase reporter gene experiment was applied to detect the targeting regulation of CircRAD18 on miR-185-5p, and miR-185-5p on HDGF in KG1a cells. RESULTS: Compared with HL-60 and U937 cells, the expression of CircRAD18 , and HDGF mRNA and protein in KG1a cells increased (all P <0.05), while miR-185-5p decreased ( P <0.05). Compared with the control group, the CircRAD18 expression, HDGF mRNA and protein expression, cell viability, proliferation rate, and PCNA, Bcl-2, BCRP, and P-gp protein expression in the DNR+CircRAD18 knockdown group decreased (all P <0.05), while miR-185-5p expression, apoptosis rate, and Bax protein expression increased (all P <0.05). There were no obvious changes in all indicators of cells in the DNR group compared with control group ( P >0.05). Compared with the DNR group, the CircRAD18 expression, HDGF mRNA and protein expression, cell viability, proliferation rate, PCNA, Bcl-2, BCRP, and P-gp protein expression in the DNR+CircRAD18 knockdown group decreased (all P < 0.05), while miR-185-5p expression, apoptosis rate, and Bax protein expression increased (all P < 0.05). There were no obvious changes in all indicators of cells in the DNR+negative control group compared with DNR group (P >0.05). Compared with the DNR+CircRAD18 knockdown group, the HDGF mRNA and protein expression, cell viability, proliferation rate, PCNA, Bcl-2, BCRP, and P-gp protein expression in the DNR+CircRAD18 knockdown+miR-185-5p inhibitor group increased (all P < 0.05), while miR-185-5p expression, apoptosis rate, and Bax protein expression decreased (all P < 0.05). CircRAD18 was able to target and down-regulate the expression of miR-185-5p in KG1a cells, and miR-185-5p was able to target and down-regulate the HDGF expression. CONCLUSION Knocking down CircRAD18 can reduce HDGF expression by up-regulating miR-185-5p, thereby weakening DNR resistance in AML cells, inhibiting KG1a cell proliferation under DNR treatment, and promoting apoptosis.
Humans ; MicroRNAs/metabolism* ; Leukemia, Myeloid, Acute ; Daunorubicin/pharmacology* ; Drug Resistance, Neoplasm ; Apoptosis ; RNA, Circular ; Intercellular Signaling Peptides and Proteins/metabolism* ; Cell Proliferation ; HL-60 Cells ; Cell Line, Tumor

OBJECTIVE: To investigate the effects of Bushen Huoxue Granule on the ubiquitin-proteasome system (UPS) in an in vitro model of Parkinson's disease. METHODS: After treated with 1-methyl-4-phenylpyridinium (MPP⁺, 1 mmol/L) for 24 h, the cells were incubated with drug-free serum, Madopar-containing serum or Bushen Huoxue Granule-containing serum (BCS, 5%, 10%, and 20%) for another 24 h. The levels of α-synuclein (α-syn), tyrosine hydroxylase (TH) and UPS-related proteins were detected by Western blot. The expression levels of α-syn in PC12 cells were also analyzed by Western blot after treated with proteasome inhibitor MG132 and WT-α-syn plasmid transfection, respectively, as well as the alterations induced by subsequent BCS intervention. Immunocytochemistry was performed to determine the changes in α-syn phosphorylation at serine 129 (pSer129-α-syn) expression. The 20S proteasome levels were measured by enzyme-linked immunosorbnent assay. RESULTS: BCS (volume fraction ⩽20%) intervention could alleviate the MMP⁺-induced cell viability decrease (P<0.05). In the MPP⁺ treated cells, α-syn was up-regulated, while TH and proteins of UPS such as ubiquitin (Ub), Ub binding with Ub-activating enzyme (UBE1), Parkin and Ub C-terminal hydrolase-1 (UCHL-1) were down-regulated (P<0.05). BCS intervention could attenuate the above changes (P<0.05). The activity of BCS on blocking α-syn accumulation was weakened by MG132 (P<0.05). While α-syn level was significantly increased in cells transfected with plasmid, and reduced by BCS intervention (P<0.05). pSer129-α-syn was increased in MPP⁺-induced PC12 cells, whereas decreased by later BCS intervention (P<0.05). The 20S proteasome activity of MPP⁺-induced PC12 cells was decreased, but increased after BCS intervention (P<0.05). CONCLUSION BCS intervention protected UPS function, increased 20S proteasome activity, promoted pathological α-syn clearance, restored cell viability, and reversed the damage caused by MPP⁺ in the in vitro model of Parkinson's disease.
PC12 Cells ; alpha-Synuclein/metabolism* ; Rats ; Animals ; 1-Methyl-4-phenylpyridinium/toxicity* ; Proteasome Endopeptidase Complex/metabolism* ; Drugs, Chinese Herbal/pharmacology* ; Ubiquitin/metabolism* ; Cell Survival/drug effects* ; Phosphorylation/drug effects* ; Tyrosine 3-Monooxygenase/metabolism*

OBJECTIVE: To investigate the relationship between physical activity and genetic risk and their combined effects on the risk of developing chronic obstructive pulmonary disease. METHODS: This prospective cohort study included 318,085 biobank participants from the UK. Physical activity was assessed using the short form of the International Physical Activity Questionnaire. The participants were stratified into low-, intermediate-, and high-genetic-risk groups based on their polygenic risk scores. Multivariate Cox regression models and multiplicative interaction analyses were used. RESULTS: During a median follow-up period of 13 years, 9,209 participants were diagnosed with chronic obstructive pulmonary disease. For low genetic risk, compared to low physical activity, the hazard ratios ( HRs) for moderate and high physical activity were 0.853 (95% confidence interval [ CI]: 0.748-0.972) and 0.831 (95% CI: 0.727-0.950), respectively. For intermediate genetic risk, the HRs were 0.829 (95% CI: 0.758-0.905) and 0.835 (95% CI: 0.764-0.914), respectively. For participants with high genetic risk, the HRs were 0.809 (95% CI: 0.746-0.877) and 0.818 (95% CI: 0.754-0.888), respectively. A significant interaction was observed between genetic risk and physical activity. CONCLUSION Moderate or high levels of physical activity were associated with a lower risk of developing chronic obstructive pulmonary disease across all genetic risk groups, highlighting the need to tailor activity interventions for genetically susceptible individuals.
Humans ; Pulmonary Disease, Chronic Obstructive/epidemiology* ; Exercise ; Male ; Female ; Middle Aged ; Prospective Studies ; Aged ; Genetic Predisposition to Disease ; Risk Factors ; United Kingdom/epidemiology* ; Incidence ; Adult

Objective To explore the relationship between serum Niemann-pick type C1 like protein1(NPC1L1)and propro-tein convertase subtilisin/kexin type 9(PCSK9)levels and the risk of type 2 diabetes mellitus(T2DM)in Mongolian residents.Methods A total of 72 Mongolian patients with T2DM treated in Peking University Cancer Hospital,Inner Mongolia Hospital and the Affiliated Hospital of Inner Mongolia Medical University from June 2022 to June 2024 were selected as the T2DM group,and 81 healthy people in the same period were selected as the control group.LASSO model and multivariate Logistic regression model were used to screen the risk factors of disease onset.Multiple linear regression was used to analyze the correlation between NPC1L1 and PCSK9 levels and insulin function.Restricted cubic spline(RCS)model was used to analyze the dose-response relationship between NPC1L1 and PCSK9 levels and the incidence of T2DM,and explored the interaction between NPC1L1 and PCSK9 levels on the incidence of T2DM.Results NPC1L1(3.11±0.80 ng/L)and PCSK9(10.63±0.79 ng/L)in T2DM group were significantly higher than those in the control group(0.52±0.22 ng/L,3.21±0.17 ng/L),and the differences were statisti-cally significant(t=27.982,82.443,all P<0.05).NPC1L1(OR=2.458,95%CI=2.364～2.594,P<0.05)and PCSK9(OR=2.905,95%CI=2.541～3.528)were risk factors for T2DM(all P<0.001).The results of multiple linear regression analysis showed that as NPC1L1 and PCSK9 levels increased,FINS,HbA1c,C-P and OGTT levels also increased accordingly.With the increase of NPC1L1 and PCSK9 levels,insulin function also decreased(all P<0.05).The results of RCS model showed that with the increase of NPC1L1 and PCSK9 levels,the probability of T2DM incidence also increased(χ2=22.334,25.537,all P<0.001).No significant interaction was found between NPC1L1,PCSK9 levels and islet function indexes(P>0.05).Conclusion The levels of NPC1L1 and PCSK9 are closely related to the risk of T2DM in Mongolian residents.With the increase of NPC1L1 and PCSK9 levels,the incidence probability of T2DM increases.

Rheumatoid arthritis(RA)is a chronic autoimmune disease of unknown etiology that can cause joint destruction and deformity.As a small molecule cytokine,the chemokine C-X-C motif chemokine ligand 12(CXCL12)regulates the pathogenesis of rheumatoid arthritis by binding to the specific receptor CXC chemokine receptor 4(CXCR4).Therefore,based on the bio-logical characteristics of CXCL12 and CXCR4,this paper intro-duces the pathogenesis of CXCL12/CXCR4 in RA and summari-zes the progress in RA-related research,with the aim of providing clinical value for understanding the pathogenesis of RA and de-veloping novel therapeutic targets.

Objective:To estimate the radiation dose to caregivers and the public from 177Lu- peptide receptor radionuclide therapy (PRRT) for pediatric neuroblastoma patients and determine the duration of contact restrictions, in order to provide a reference for relevant radiation protection measures. Methods:A retrospective study was conducted by collecting data from 18 pediatric neuroblastoma patients, aged between 3 and 13 (6.72±2.72), who received 177Lu-1, 4, 7, 10-tetraazacyclododecane-1, 4, 7, 10-tetraacetic acid-D-Phe1-Tyr3-Thr8-octreotide (DOTATATE) treatment at the Nuclear Medicine Department of Shandong Cancer Hospital from June 2023 to July 2023. Absorbed dose rate in air at 0, 0.1, 0.5, 1 and 2 m from the patients was measured using a radiation-survey at 1, 4, 24, 48 and 96 h after administration. The whole-body region of interest was delined using HERMES software. Subsequently, curve regression fitting was performed using a biexponential function model. By incorporating hypothesized social contact durations, the effective doses received by family members and the public in contact with patients were estimated. Additionally, MIM software was used to outline the whole-body VOI to obtain the total volume of lesions, and the Pearson or Spearman correlation coefficient was employed to analyze the relationship between the absorbed dose rate in air and clinical indicators as well as the total volume of lesions. Results:The 177Lu-DOTATATE administration dose was (4 353.42±1 451.51) MBq. All patients were discharged from hospital 24 h after 177Lu-DOTATATE administration. At the time of discharge, patients had excreted (76.70±3.99)% of the administered activity, and the absorbed dose rate in air at 0.1, 1 and 2 m from the patients were (32.74±6.98), 3.68(3.01, 4.70) and (1.22±0.51) μSv/h, respectively. After being discharged, the radiation doses to caregivers from children aged 2-5 years and 5-13 years were (2.47±1.80) mSv and (0.88±0.47) mSv, respectively. The contact restriction duration was 2 d for nighttime sleeping with family members and 1 d for contact with other children. On the day of discharge, patients should limit their time on public transportation within 4 h and do not need to restrict private transportation. Conclusions:To ensure the effective dose kept within the safety limits stipulated by current regulations, it is necessary to implement contact restrictions for patients’ family members and the public. After implementing preventive measures, 177Lu-DOTATATE treatment is a safe radionuclide therapeutic option.