BACKGROUND:Bone relies on the close connection between blood vessels and bone cells to maintain its integrity.Bones are in a physiologically hypoxic environment.Therefore,the study of angiogenesis and osteogenesis in hypoxic environment is closer to the microenvironment in vivo. OBJECTIVE:To explore the influence of Wenshen Tongluo Zhitong(WSTLZT)formula on H-type bone microvascular endothelial cell/bone marrow mesenchymal stem cell co-culture system in hypoxia environment and its related mechanism. METHODS:Enzyme digestion method and flow sorting technique were used to isolate and identify H-type bone microvascular endothelial cells.Mouse bone marrow mesenchymal stem cells were isolated and obtained by bone marrow adhesion method.H-type bone microvascular endothelial cell/bone marrow mesenchymal stem cell hypoxic co-culture system was established using Transwell chamber and anoxic culture workstation.WSTLZT formula powder was used to intervene in each group at a mass concentration of 50 and 100 μg/mL.The angiogenic function of H-type bone microvascular endothelial cells in the co-culture system was evaluated by scratch migration test and tube formation test.The osteogenic differentiation ability of bone marrow mesenchymal stem cells in the co-cultured system was evaluated by alkaline phosphatase staining and alizarin red staining.The protein and mRNA expression changes of PDGF/PI3K/AKT signal axis related molecules in H-type bone microvascular endothelial cells in the co-cultured system were detected by Western Blotting and q-PCR,respectively. RESULTS AND CONCLUSION:(1)Compared with the normal oxygen group,the scratch mobility and new blood vessel length of H-type bone microvascular endothelial cells were significantly higher(P<0.05);the osteogenic differentiation capacity of bone marrow mesenchymal stem cells was higher(P<0.05);the expression of PDGF/PI3K/AKT axis-related molecular protein and mRNA increased(P<0.05)in the hypoxia group.(2)Compared with the hypoxia group,scratch mobility and new blood vessel length were significantly increased in the H-type bone microvascular endothelial cells(P<0.05);bone marrow mesenchymal stem cells had stronger osteogenic function(P<0.05);the expression of PDGF/PI3K/AKT axis-related molecular proteins and mRNA further increased(P<0.05)after treatment with different dose concentrations of WSTLZT formula.These findings conclude that H-type angiogenesis and osteogenesis under hypoxia may be related to the PDGF/PI3K/AKT signaling axis,and WSTLZT formula may promote H-type vasculo-dependent bone formation by activating the PDGF/PI3K/AKT signaling axis,thereby preventing and treating osteoporosis.

Senile osteoporosis is a growing public health challenge with significant impacts on the daily life of the elderly population as a result of its hidden nature,high prevalence,and high risk of disability.Suitable animal models that simulate senile osteoporosis are crucial for understanding its pathological mechanism and to facilitate the development of anti-osteoporosis drugs and identify new therapeutic targets.This review considers the most commonly used method for creating animal models of senile osteoporosis,analyzes their advantages and limitations,and discusses research progress in animal models in terms of evaluation indicators,to provide references for research using animal models of senile osteoporosis.

Senile osteoporosis is a growing public health challenge with significant impacts on the daily life of the elderly population as a result of its hidden nature,high prevalence,and high risk of disability.Suitable animal models that simulate senile osteoporosis are crucial for understanding its pathological mechanism and to facilitate the development of anti-osteoporosis drugs and identify new therapeutic targets.This review considers the most commonly used method for creating animal models of senile osteoporosis,analyzes their advantages and limitations,and discusses research progress in animal models in terms of evaluation indicators,to provide references for research using animal models of senile osteoporosis.

Objective To investigate the relationship between interleukin-1β (IL-1β) and miR-185-5p in the process of joint injury in acute gouty arthritis (AGA). Methods The serum miR-185-5p levels of 89 AGA patients and 91 healthy volunteers were detected by real-time quantitative PCR. The correlation between miR-185-5p expression level and VAS score or IL-1β expression level was evaluated by Pearson correlation coefficient method. Receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value of miR-185-5p in AGA. THP-1 cells were induced by sodium urate (MSU) to construct an in vitro acute gouty inflammatory cell model. After the expression level of miR-185-5p in THP-1 cells was upregulated or downregulated by transfection of miR-185-5p mimics or inhibitors in vitro, inflammatory cytokines of THP-1 cells, such as IL-1β, IL-8 and tumor necrosis factor α (TNF-α), were detected by ELISA. The luciferase reporter gene assay was used to determine the interaction between miR-185-5p and the 3'-UTR of IL-1β. Results Compared with the healthy control group, the expression level of serum miR-185-5p in AGA patients was significantly reduced. The level of serum miR-185-5p was negatively correlated with VAS score and IL-1β expression level. The area under the curve (AUC) was 0.905, the sensitivity was 80.17% and the specificity was 83.52%. Down-regulation of miR-185-5p significantly promoted the expression of IL-1β, IL-8 and tumor necrosis factor (TNF-α), while overexpression of miR-185-5p showed the opposite results. Luciferase reporter gene assay showed that IL-1β was the target gene of miR-185-5p, and miR-185-5p negatively regulated the expression of IL-1β. Conclusion miR-185-5p alleviates the inflammatory response in AGA by inhibiting IL-1β.
Humans ; 3' Untranslated Regions ; Arthritis, Gouty/genetics* ; Interleukin-1beta/genetics* ; Interleukin-8 ; Luciferases ; MicroRNAs/genetics* ; Tumor Necrosis Factor-alpha

OBJECTIVE To study the intervention effects of Wenshen Tongluo Zhitong Recipe(WTZR)on macrophage senes-cence and senile osteoporosis.METHODS The senescent macrophage model was established using hydrogen peroxide(H2 O2)and subsequently divided into four groups:control,model,low-dose drug-treated serum,and high-dose drug-treated serum.β-galactosidase staining,Western blot and qPCR were employed to evaluate the mRNA expression of senescence markers p21 and p53.ROS staining and JC-1 staining were applied to assess mitochondrial function in macrophages.The mRNA levels of Interleukin(IL)-6,IL-10,CD206,and inducible nitric oxide synthase(iNOS)were determined by qPCR analysis.Immunofluorescence was used to evaluate argi-nase(ARG1)and iNOS protein expressions for assessing the impact of drug-containing serum on macrophage polarization.qPCR analy-sis was conducted to measure osteocalcin(OCN),collagen type Ⅰ alpha 1(Col1a1),runt-related transcription factor 2(Runx2)mRNA levels as osteoblast-related markers;ALP staining along with alizarin red staining were performed to evaluate the effect of macrophage conditioned medium treated with drug-containing serum on bone marrow mesenchymal stromal cell(BMSC)osteogenic differentiation.C57BL/6J mice were randomly allocated into four groups:control group,model group,WTZR low-dose group,and WTZR high-dose group.The senile osteoporosis(SOP)mouse model was established by D-galactose.Micro-CT scanning analyzed fe-mur microstructure while HE staining detected pathological changes in femur bone tissue samples collected from each experimental con-dition.Furthermore,Western blot was used to detect the senescence-related molecules p21 and p53 and the osteogenesis-related mark-ers OCN and Runx2,qPCR analysis measured tibial expression levels of senescence-related molecules(p21,p53)as well as macro-phage polarisation-related molecules(IL-6,iNOS,CD206,and IL-10)to assess the effect of this compound on a mouse model simula-ting SOP.RESULTS Following intervention with serum containing WTZR,there was a significant decrease in the number of senes-cent positive cells compared to the model group.Additionally,there was a notable decrease in p21 and p53 mRNA and proteins expres-sion(P＜0.05,P＜0.01,P＜0.001).Furthermore,drug-containing WTZR effectively inhibited ROS production induced by H2 O2 and mitigated mitochondrial membrane potential reduction in macrophages(P＜0.05,P＜0.001).Treatment with drug-containing WTZR re-sulted in down-regulated mRNA expression of M1-related gene iNOS(P＜0.05)while up-regulating mRNA expression of M2-related genes CD163 and CD206(P＜0.05).The drug-containing WTZR significantly reduced fluorescence intensity for iNOS(P＜0.01)while increasing ARG1(P＜0.05)fluorescence intensity.Moreover,conditioned medium from macrophages treated with drug-containing ser-um increased ALP positive cell count(P＜0.01,P＜0.001),alizarin red positive area(P＜0.05),as well as Col1a1,Runx2 and OCN mRNA expression levels(P＜0.05,P＜0.01).The Tb.N,BMD,and BV/TV were significantly higher in the WTZR group compared to the model group(P＜0.05,P＜0.01,P＜0.001);meanwhile,Tb.Sp was notably lower than that observed in the model group(P＜0.05,P＜0.01);bone trabeculae were significantly improved,increased in number and widened.Additionally,the compound could significantly inhibit the D-galactose induced up-regulation of tibial senescence-related genes and proteins p21 and p53(P＜0.05,P＜0.001,P＜0.0001),promote the expression of osteogenesis-related markers OCN and Runx2 protein(P＜0.01,P＜0.0001),promote the down-regulation of M1 related genes IL-6 and iNOS(P＜0.05),and promote the expression of M2 related genes IL-10 and CD206(P＜0.05,P＜0.01).CONCLUSION Wenshen Tongluo Zhitong Recipe may play an anti-osteoporosis effect by inhibiting macrophage senescence and promoting the osteogenic differentiation of BMSC.

Objective:To investigate long-term auditory changes and characteristics of Alport syndrome（AS） patients with different degrees of renal injury. Methods:Retrospectively analyzing clinical data of patients diagnosed AS from January 2007 to September 2022, including renal pathology, genetic detection and hearing examination. A long-term follow-up focusing on hearing and renal function was conducted. Results:This study included 70 AS patients, of which 33（25 males, 8 females, aged 3.4-27.8 years） were followed up, resulting in a loss rate of 52.9%.The follow-up period ranged from 1.1to 15.8 years, with 16 patients followed-up for over 10 years. During the follow-up, 10 patients presenting with hearing abnormalities at the time of diagnosis of AS had progressive hearing loss, and 3 patients with new hearing abnormalities were followed up, which appeared at 5-6 years of disease course. All of which were sensorineural deafness. While only 3 patients with hearing abnormalities among 13 patients received hearing aid intervention. Of these patients,7 developed end-stage renal disease（ESRD）, predominantly males （6/7）. The rate of long-term hearing loss was significantly different between ESRD group and non-ESRD group（P=0.013）. There was no correlation between the progression of renal disease and long-term hearing level（P>0.05）. kidney biopsies from 28 patients revealed varying degrees of podocyte lesion and uneven thickness of basement membrane. The severity of podocyte lesion was correlated with the rate of long-term hearing loss（P=0.048）, and there was no correlation with the severity of hearing loss（P>0.05）. Among 11 cases, theCOL4A5mutationwas most common （8 out of 11）, but there was no significant correlation between the mutation type and hearing phenotype（P>0.05）. Conclusion:AS patients exhibit progressive hearing loss with significant heterogeneity over the long-term.. THearing loss is more likely to occur 5-6 years into the disease course. Hearing abnormalities are closely related to renal disease status, kidney tissue pathology, and gene mutations, emphasizing the need for vigilant long-term hearing follow-up and early intervention.
Male ; Child ; Female ; Humans ; Nephritis, Hereditary/pathology* ; Retrospective Studies ; Kidney ; Deafness ; Hearing Loss/genetics* ; Kidney Failure, Chronic/pathology* ; Mutation

Objective To investigate the reliability and validity of the assessment tools of Brief ICF Core Sets for Arthroplasty of Knee Osteoarthritis in Perioperative Period(ICSAKOPP). Methods From May,2022 to April,2023,320 patients undergoing knee arthroplasty were selected in Peking University Third Hospital,China-Japan Friendship Hospital,Peking University First Hospital and Chinese PLA General Hospital.Trained assessors used Brief ICSAKOPP to evaluate all enrolled patients before arthroplasty,three days(±one day)after arthroplasty,three weeks(±one week)after arthroplasty,and three months(±one month)after ar-throplasty.Western Ontario and McMaster Universities Osteoarthritis Index(WOMAC)scores were recorded at the same time.Five professionals were asked to score all the items of Brief ICSAKOPP,and the content validity index(CVI)was caculated. Results A total of 64 cases were dropped down.CVI of all the items of the Brief ICSAKOPP were above 0.8,with a av-erage CVI of the scale of 0.938.The Cronbach's α coefficient of the Brief ICSAKOPP was 0.813.There was a moderate correlation(r=0.681,P<0.001)between the overall Brief ICSAKOPP and WOMAC scores,as well as body functional dimension score(r=0.668,P<0.001)and activities and participation dimension score(r=0.657,P<0.001). Conclusion Brief ICSAKOPP is good in content validity,internal consistency reliability and criterion validity.

Objective To compare the differences in the levels of lactate dehydrogenase(LDH),creatine kinase(CK),and neuron-specific enolase(NSE)in the cerebrospinal fluid(CSF)between the patients with purulent meningitis(PM)and those with tuberculous meningitis(TBM)and evaluate the utility of the three biomarkers in the differentiation of PM and TBM.Methods Thirty-five PM patients and 45 TBM patients who attended the Department of Neurology from December 2019 to December 2021 were enrolled in this study.The CSF samples were collected from the patients before treatment.The levels of LDH,CK,NSE,and glucose,protein,and chloride in CSF were measured and compared between PM and TBM patients.The correlation between the biomarkers in CSF was analyzed by Pearson correlation coefficient.Linear regression analysis was performed to analyze the risk factors for the occurrence of TBM.Receiver operating characteristic(ROC)curves were plotted to evaluate the discriminative value of CSF LDH,CK,NSE alone or in combination in differential diagnosis of PM and TBM.Results Patient age,gender,clinical features such as fever,pathological signs,vomiting,limb twitching,and CSF levels of glucose and protein did not show significant difference between TBM patients and PM patients(P＞0.05).TBM was associated with significantly higher LDH and NSE levels in CSF and significantly lower CK level and chloride in CSF compared to PM(P＜0.05).Pearson correlation analysis showed that LDH level in CSF was negatively correlated with CK and chloride(r＜0,P＜0.05),and positively correlated with NSE(r＞0,P＜0.05).CK was negatively correlated with NSE(r＜0,P＜0.05),and positively correlated with chloride(r＞0,P＜0.05).NSE was negatively correlated with chloride(r＜0,P＜0.05).Linear regression analysis showed that LDH,CK,NSE,and chloride levels in CSF were all associated with the occurrence of TBM(P＜0.05).ROC curves demonstrated that the AUCs of CSF levels of LDH,CK,NSE alone and in combination for differentiation between PM and TBM were 0.849(95％CI:0.768-0.930),0.858(95％CI:0.779-0.937),0.851(95％CI:0.771-0.931),and 0.954(95％CI:0.911-0.996),respectively.Conclusions The clinical features are similar for PM and TBM patients.However,the levels of LDH,CK,and NSE in CSF are different between PM and TBM.LDH,CK,NSE in combination may improve the differential diagnosis between PM and TBM.

Objectives:This study aimed to analyze the clinical and molecular characteristics of carbapenem-resistant Enterobacteriaceae (CRE) bloodstream infection (BSI) in patients with hematological diseases and to explore prognostic risk factors.Methods:This retrospective study included patients with hematologic diseases with CRE BSI at the Institute of Hematology and Blood Diseases Hospital from January 2015 to December 2022. The clinical features, carbapenemase test results, antimicrobial treatments, and outcomes were analyzed.Results:A total of 120 patients developed CRE BSI. Escherichia coli (58/120, 48.3%) was the most prevalent Enterobacteriaceae, followed by Klebsiella pneumoniae (52/120, 43.3%). A total of 93 CRE strains were tested for carbapenemase, of which 75 strains produced carbapenemase (metalloenzyme: 51 strains; serine enzyme: 24 strains). The 30-day mortality rate after BSI was 24.2% (29/120). Univariate analysis revealed significantly lower mortality in patients treated with the ceftazidime-avibactam-containing regimen than in those treated with other antibiotics (7.8% vs 36.2%, P<0.001). Moreover, initiating active therapy within 24 h of BSI onset significantly reduced mortality (15.0% vs 33.3%, P=0.019). The proportion of patients with CRE colonization receiving active therapy within 12 and 24 h was significantly higher compared with patients without colonization (12 h: 14.5% vs 34.1%, P=0.012; 24 h: 40.8% vs 65.9%, P=0.008). Multivariate analysis revealed that septic shock ( HR=24.436, 95% CI 4.148 - 143.966, P<0.001) and pulmonary infection ( HR=9.346, 95% CI 2.718-32.140, P<0.001) were independent risk factors for death within 30 days. Appropriate therapy was initiated within 24 h ( HR=0.225, 95% CI 0.059 - 0.851, P=0.028), and treatment with the ceftazidime-avibactam-containing regimen ( HR=0.082, 95% CI 0.018-0.362, P=0.001) significantly reduced mortality. Conclusion:The prognosis of CRE BSI in patients with hematological diseases is poor. Timely, appropriate therapy and receipt of a ceftazidime-avibactam-containing regimen can improve survival and prognosis.

Objectives:This study aimed to analyze the clinical and molecular characteristics of carbapenem-resistant Enterobacteriaceae (CRE) bloodstream infection (BSI) in patients with hematological diseases and to explore prognostic risk factors.Methods:This retrospective study included patients with hematologic diseases with CRE BSI at the Institute of Hematology and Blood Diseases Hospital from January 2015 to December 2022. The clinical features, carbapenemase test results, antimicrobial treatments, and outcomes were analyzed.Results:A total of 120 patients developed CRE BSI. Escherichia coli (58/120, 48.3%) was the most prevalent Enterobacteriaceae, followed by Klebsiella pneumoniae (52/120, 43.3%). A total of 93 CRE strains were tested for carbapenemase, of which 75 strains produced carbapenemase (metalloenzyme: 51 strains; serine enzyme: 24 strains). The 30-day mortality rate after BSI was 24.2% (29/120). Univariate analysis revealed significantly lower mortality in patients treated with the ceftazidime-avibactam-containing regimen than in those treated with other antibiotics (7.8% vs 36.2%, P<0.001). Moreover, initiating active therapy within 24 h of BSI onset significantly reduced mortality (15.0% vs 33.3%, P=0.019). The proportion of patients with CRE colonization receiving active therapy within 12 and 24 h was significantly higher compared with patients without colonization (12 h: 14.5% vs 34.1%, P=0.012; 24 h: 40.8% vs 65.9%, P=0.008). Multivariate analysis revealed that septic shock ( HR=24.436, 95% CI 4.148 - 143.966, P<0.001) and pulmonary infection ( HR=9.346, 95% CI 2.718-32.140, P<0.001) were independent risk factors for death within 30 days. Appropriate therapy was initiated within 24 h ( HR=0.225, 95% CI 0.059 - 0.851, P=0.028), and treatment with the ceftazidime-avibactam-containing regimen ( HR=0.082, 95% CI 0.018-0.362, P=0.001) significantly reduced mortality. Conclusion:The prognosis of CRE BSI in patients with hematological diseases is poor. Timely, appropriate therapy and receipt of a ceftazidime-avibactam-containing regimen can improve survival and prognosis.