Objective:To investigate the effects of Xuebijing injection on gut microbiota and intestinal mechanical barrier in mice with lipopolysaccharide (LPS)-induced sepsis, and analyze the potential mechanism by which Xuebijing injection protects gastrointestinal tract.Methods:Twenty-four healthy and clean grade male C57BL/6N mice were divided into four groups, control group, LPS group, LPS+ 5 μl/g Xuebijing injection group (5 μl/g Xuebijing injection group), and LPS+ 10 μl/g Xuebijing injection group (10 μl/g Xuebijing injection group), with six mice in each group. A mouse model of sepsis was established by intraperitoneal injection of mice with 10 μg/g LPS. At 0 and 12 h after successful modeling, the mice were intraperitoneally injected with 5 or 10 μl/g Xuebijing injection. Blood, ileum, and colon fecal samples were collected 12 h after the second administration. ELISA was used to detect the levels of diamine oxidase (DAO), D-blood lactic acid (D-Lac), TNF-α, and IL-6. HE staining was used to observe the local ileum damage, and Chiu′s score was used to evaluate the degree of intestinal tissue damage. Immunohistochemical staining and Western blot were used to detect the expression of Claudin-1, Occludin, and zona occludins-1(ZO-1) in ileum tissues, followed by semi quantitative analysis. One-way analysis of variance was used for intergroup comparisons, and LSD or Tamhane′s T2 test was used for pairwise comparisons based on the homogeneity of variance. The diversity and species composition of mouse fecal microbiota, and the differences among groups were analyzed using 16S rRNA sequencing.Results:The levels of DAO, D-Lac, TNF-α, and IL-6 in the LPS group were higher than those in the control group (all P<0.000 1). After the intervention with Xuebijing injection, the levels of DAO, D-Lac, TNF-α, and IL-6 decreased (all P<0.05) and showed no significant differences with those in the control group (all P>0.05). Besides, 10 μl/g Xuebijing injection was more effective than 5 μl/g Xuebijing injection in reducing the concentrations (all P<0.05). Chiu′s score was higher in the LPS group than in the control group and the 10 μl/g Xuebijing injection group (both P<0.05). Western blot showed that the expression levels of Occludin, Claudin-1, and ZO-1 in the LPS group were lower than those in the control group (all P<0.01), and Xuebijing injection intervention significantly increased the expression levels of these proteins in a dose-dependent manner as compared with the LPS group (all P<0.000 1). Apart from the expression level of ZO-1, which showed no significant difference between the two Xuebijing injection groups ( P>0.05), the results of immunohistochemical staining were consistent with those of Western blot. The 16S rRNA sequencing results showed that there were differences in the Alpha and Beta diversity indices, and the composition and structure of gut microbiota among the four groups. The structure of gut microbiota in the mice treated with Xuebijing injection was similar to that in the mice of the control group and it was in a dose-dependent manner. Wilcoxon rank sum test showed that there were statistically significant differences in six gut microbiota groups at the phylum level, and 32 gut microbiota groups at the genus level among the mice of four groups (all P<0.05). Conclusions:Xuebijing injection can provide protective effects on the gastrointestinal tract by protecting the structure of gut microbiota and intestinal barrier function, and the protective effect is somewhat correlated with the drug dosage.

Objective:To construct a full-length infectious clone of enterovirus D68（EV-D68）expressing enhanced green fluorescent protein（EGFP）by reverse genetics in order to provide an efficient tool for studying the biological characteristics and screening antiviral drugs for EV-D68.Methods:Gene synthesis and overlap PCR techniques were used to construct the full-length clone plasmid pUC57-EV-D68 of EV-D68. The full-length viral sequence was then transferred into the pCAGGS plasmid to obtain the pCAGGS-EV-D68 plasmid. The EGFP gene was amplified and inserted into the pCAGGS-EV-D68 plasmid to construct the pCAGGS-EGFP-EV-D68 plasmid. Then，the two constructed plasmids were transfected into human rhabdomyosarcoma（RD）cells to rescue recombinant viruses RV-EV-D68 and RV-EGFP-EV-D68. The rescued viruses were identified using PCR，Western blot，and immunofluorescence techniques. The antiviral effect of doxycycline was evaluated using the rescued RV-EGFP-EV-D68. Statistical analysis was performed using the two independent samples t-test. Results:The recombinant virus RV-EGFP-EV-D68 capable of expressing EGFP was successfully rescued. Even after 15 serial passages，the virus retained EGFP expression with no significant difference in viral titers compared to the parental virus，indicating its stable passage in RD cells. Besides，the rescued strains exhibited similar replication characteristics to the parental virus. While at 24 and 36 h after infection，the titers of the rescued strains were significantly lower than that of the parental strain（both P<0.05）. This study demonstrated that doxycycline significantly reduced the fluorescence intensity of RV-EGFP-EV-D68-infected RD cells（ P<0.01）. Meanwhile，a negative correlation was observed between the doxycycline concentration and the fluorescence intensity，indicating that the rescued virus could be used for antiviral drug evaluation. Conclusions:This study successfully constructs an infectious clone of EV-D68 expressing EGFP. The rescued recombinant virus RV-EGFP-EV-D68 has been verified to be applicable for the evaluation of antiviral drugs.

OBJECTIVE: To analyze the demographic and clinical characteristics of inpatients with osteoporotic vertebral compression fractures (OVCF) and provide a basis for clinical prevention and treatment. METHODS: A retrospective analysis was performed on the clinical data of 744 inpatients diagnosed with OVCF between January 2017 and December 2021 who met the inclusion criteria. Among them, 146 were male and 598 were female, with age ranging from 50 to 95 years (mean, 69.37 years). The demographic characteristics (gender, age, ethnicity, occupation, regional distribution, urban-rural distribution, and seasonal incidence) and clinical features [causes of injury, history of vertebral fractures, smoking and drinking history in males, comorbidities (hypertension, diabetes, coronary atherosclerotic heart disease, cerebral infarction), body mass index (BMI), blood lipid levels, menopausal age in females, vertebral bone mineral density T-value, number of vertebral fractures, and fracture segment distribution] of OVCF patients were analyzed. Multiple linear regression was used to analyze the independent risk factors of vertebral osteoporosis. RESULTS: The demographic analysis indicated that female patients with OVCF were significantly younger than male patients ( P<0.05). Significant differences were observed in the age distribution of OVCF between males and females ( P<0.05), with the highest proportion of male patients in the 70-79 years group (37.0%) and the highest proportion of female patients in the 60-69 years group (40.0%). From 2017 to 2021, the age of onset for OVCF gradually increased, with a similar trend observed for both genders. The distribution of occupations between genders also showed significant differences ( P<0.05); with the top three occupations for males being farmers (48.6%), retirees (24.7%), and workers (13.7%), while for females, the leading occupations were farmers (51.5%), retirees (19.4%), and service workers (10.0%). Female OVCF patients had higher BMI, vertebral bone mineral density T-value, history of vertebral fractures, hypertension prevalence, and blood lipid levels compared to male patients ( P<0.05). No significant difference between the males and the females was found in ethnicity, seasonal distribution, regional distribution, urban-rural distribution, causes of injury, number of vertebral fractures, or prevalence of comorbidities (except hypertension) ( P>0.05). Among the 744 OVCF patients, a total of 1 309 vertebrae were involved, with 628 thoracic vertebrae (48.0%) and 681 lumbar vertebrae (52.0%). The most common fracture segments were L ₁ (22.5%), T ₁₂ (21.2%), followed by L ₂ (12.2%) and T ₁₁ (10.2%). No significant gender difference was observed in the distribution of fracture segments ( P>0.05). Multiple linear regression analysis indicated that older age, female, and lower BMI were independent risk factors for vertebral osteoporosis ( P<0.05). CONCLUSION The age of onset of OVCF patients is increasing year by year. The number of fractured vertebral bodies, age distribution of morbidity, occupational distribution, BMI, history of vertebral fracture, hypertension, and blood lipid levels are related to gender. The occurrence of OVCF is mainly in the thoracolumbar segment. The female, older age, and lower BMI are independent risk factors of osteoporosis.
Humans ; Male ; Female ; Aged ; Middle Aged ; Retrospective Studies ; Spinal Fractures/etiology* ; Aged, 80 and over ; Osteoporotic Fractures/etiology* ; Fractures, Compression/etiology* ; Risk Factors ; Bone Density ; China/epidemiology* ; Osteoporosis/epidemiology* ; Comorbidity ; Inpatients ; Sex Factors ; Age Factors

Objective:To investigate the application of time series models in forecasting pre-hospital emergency ambulance trips in Handan City and develop the LPro ensemble model for improved prediction accuracy to support emergency resource allocation.Methods:Pre-hospital emergency data from Handan Emergency Medical Command Center (2019-2023) were retrospectively analyzed. From 324 799 original records, 289 949 valid records were included after cleaning. The training set (2019-2022: 215 918 records) included 35 527 records in 2019, 52 015 in 2020, 61 836 in 2021, and 66 540 in 2022. The validation set (2023) contained 74 031 records. ARIMA, linear trend seasonal, exponential smoothing, and Prophet models were fitted to the training set. The LPro ensemble model was constructed using MAPE-based weighting (linear trend seasonal model: 0.38, Prophet: 0.62). Performance metrics included MAPE, RMSE, MAE, and R 2. Results:Data showed annual growth (compound annual growth rate 23.27%) and seasonal patterns (October peaks, February troughs). Ambulance dispatches increased annually with monthly cyclical patterns. For 2023 validation predictions: ARIMA (MAPE 8.76%, RMSE 619, MAE 491, R 2 0.4563), linear trend seasonal (MAPE 9.83%, RMSE 671, MAE 545, R 2 0.3608), Prophet (MAPE 8.43%, RMSE 562, MAE 503, R 2 0.5513), exponential smoothing (MAPE 8.08%, RMSE 643, MAE 410, R 2 0.4124). LPro model showed superior performance (MAPE 7.05%, RMSE 491, MAE 393, R 2 0.6570), with 16.37% lower MAPE, 12.63% lower RMSE, 21.87% lower MAE, and 19.17% higher R 2 versus Prophet. Conclusion:The LPro ensemble model substantially enhances prediction accuracy and reliability, offering scientific support for emergency resource optimization and dispatch scheduling in Handan City.

Objective:To investigate the effects of Xuebijing injection on gut microbiota and intestinal mechanical barrier in mice with lipopolysaccharide (LPS)-induced sepsis, and analyze the potential mechanism by which Xuebijing injection protects gastrointestinal tract.Methods:Twenty-four healthy and clean grade male C57BL/6N mice were divided into four groups, control group, LPS group, LPS+ 5 μl/g Xuebijing injection group (5 μl/g Xuebijing injection group), and LPS+ 10 μl/g Xuebijing injection group (10 μl/g Xuebijing injection group), with six mice in each group. A mouse model of sepsis was established by intraperitoneal injection of mice with 10 μg/g LPS. At 0 and 12 h after successful modeling, the mice were intraperitoneally injected with 5 or 10 μl/g Xuebijing injection. Blood, ileum, and colon fecal samples were collected 12 h after the second administration. ELISA was used to detect the levels of diamine oxidase (DAO), D-blood lactic acid (D-Lac), TNF-α, and IL-6. HE staining was used to observe the local ileum damage, and Chiu′s score was used to evaluate the degree of intestinal tissue damage. Immunohistochemical staining and Western blot were used to detect the expression of Claudin-1, Occludin, and zona occludins-1(ZO-1) in ileum tissues, followed by semi quantitative analysis. One-way analysis of variance was used for intergroup comparisons, and LSD or Tamhane′s T2 test was used for pairwise comparisons based on the homogeneity of variance. The diversity and species composition of mouse fecal microbiota, and the differences among groups were analyzed using 16S rRNA sequencing.Results:The levels of DAO, D-Lac, TNF-α, and IL-6 in the LPS group were higher than those in the control group (all P<0.000 1). After the intervention with Xuebijing injection, the levels of DAO, D-Lac, TNF-α, and IL-6 decreased (all P<0.05) and showed no significant differences with those in the control group (all P>0.05). Besides, 10 μl/g Xuebijing injection was more effective than 5 μl/g Xuebijing injection in reducing the concentrations (all P<0.05). Chiu′s score was higher in the LPS group than in the control group and the 10 μl/g Xuebijing injection group (both P<0.05). Western blot showed that the expression levels of Occludin, Claudin-1, and ZO-1 in the LPS group were lower than those in the control group (all P<0.01), and Xuebijing injection intervention significantly increased the expression levels of these proteins in a dose-dependent manner as compared with the LPS group (all P<0.000 1). Apart from the expression level of ZO-1, which showed no significant difference between the two Xuebijing injection groups ( P>0.05), the results of immunohistochemical staining were consistent with those of Western blot. The 16S rRNA sequencing results showed that there were differences in the Alpha and Beta diversity indices, and the composition and structure of gut microbiota among the four groups. The structure of gut microbiota in the mice treated with Xuebijing injection was similar to that in the mice of the control group and it was in a dose-dependent manner. Wilcoxon rank sum test showed that there were statistically significant differences in six gut microbiota groups at the phylum level, and 32 gut microbiota groups at the genus level among the mice of four groups (all P<0.05). Conclusions:Xuebijing injection can provide protective effects on the gastrointestinal tract by protecting the structure of gut microbiota and intestinal barrier function, and the protective effect is somewhat correlated with the drug dosage.

Objective:To construct a full-length infectious clone of enterovirus D68（EV-D68）expressing enhanced green fluorescent protein（EGFP）by reverse genetics in order to provide an efficient tool for studying the biological characteristics and screening antiviral drugs for EV-D68.Methods:Gene synthesis and overlap PCR techniques were used to construct the full-length clone plasmid pUC57-EV-D68 of EV-D68. The full-length viral sequence was then transferred into the pCAGGS plasmid to obtain the pCAGGS-EV-D68 plasmid. The EGFP gene was amplified and inserted into the pCAGGS-EV-D68 plasmid to construct the pCAGGS-EGFP-EV-D68 plasmid. Then，the two constructed plasmids were transfected into human rhabdomyosarcoma（RD）cells to rescue recombinant viruses RV-EV-D68 and RV-EGFP-EV-D68. The rescued viruses were identified using PCR，Western blot，and immunofluorescence techniques. The antiviral effect of doxycycline was evaluated using the rescued RV-EGFP-EV-D68. Statistical analysis was performed using the two independent samples t-test. Results:The recombinant virus RV-EGFP-EV-D68 capable of expressing EGFP was successfully rescued. Even after 15 serial passages，the virus retained EGFP expression with no significant difference in viral titers compared to the parental virus，indicating its stable passage in RD cells. Besides，the rescued strains exhibited similar replication characteristics to the parental virus. While at 24 and 36 h after infection，the titers of the rescued strains were significantly lower than that of the parental strain（both P<0.05）. This study demonstrated that doxycycline significantly reduced the fluorescence intensity of RV-EGFP-EV-D68-infected RD cells（ P<0.01）. Meanwhile，a negative correlation was observed between the doxycycline concentration and the fluorescence intensity，indicating that the rescued virus could be used for antiviral drug evaluation. Conclusions:This study successfully constructs an infectious clone of EV-D68 expressing EGFP. The rescued recombinant virus RV-EGFP-EV-D68 has been verified to be applicable for the evaluation of antiviral drugs.

Objective:To explore the effect of Baduanjin on gait parameters and serum nerve growth factor in Parkinson disease (PD) patients with freezing of gait(FOG).Methods:From December 2021 to December 2022, thirty-eight PD patients with FOG who met the inclusion and exclusion criteria were randomly divided into observation group ( n=18) and control group ( n=20) by random number table.The patients in both two groups received 4 weeks of drug therapy combined with basic rehabilitation treatment respectively, and the patients in observation group received additional Baduanjin training.Efficacy was evaluated 1 day before intervention and after 4 weeks of intervention through unified Parkinson's disease rating scale-Ⅱ(UPDRS-Ⅱ) item 14, freezing of gait questionnaire (FOGQ), gait starting time, gait cycle, stride length, dynamic plantar peak pressure and average pressure, while the levels of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor(GDNF) in peripheral blood of patients were tested.SPSS 23.0 software was used to conduct Chi-square test, paired t-test, independent sample t-test and Mann-Whitney U test. Results:Before treatment, there were no significant differences in score of UPDRS-Ⅱ item 14, FOGQ score, gait starting time, gait cycle, stride length, dynamic planar peak pressure, average pressure, peripheral blood BDNF level and GDNF level between the two groups ( t=-0.542, 0.562, 0.490, 0.674, 0.440, 0.606, -0.835, -0.873, -0.250, all P>0.05). After treatment, compared with the control group, dynamic plantar peak pressure (control group (14.26±3.23) N/cm 2, observation group (11.40±4.13) N/cm 2, t=-2.389, P=0.022) and plantar average pressure (control group (3.34±0.72) N/cm 2, observation group (2.79±0.81) N/cm 2, t=-2.209, P=0.034) of the observation group were significantly decreased (both P<0.05). There were no significant differences in UPDRS-Ⅱ item 14, FOGQ score, gait starting time, gait cycle, stride length, BDNF and GDNF concentrations in peripheral blood between the two groups after treatment (all P>0.05). The difference between pre-treatment and post-treatment of FOGQ score (control group 1.00 (0.00, 1.00) , observation group 2.00 (0.75, 3.00), Z=-2.547, P=0.011), gait starting time (control group -1.04 (-1.86, -0.47)s, observation group -2.34 (-3.41, -1.03) s, Z=-2.280, P=0.023), gait cycle (control group 0.29 (0.08, 0.58)s, observation group 0.35 (0.16, 1.00) s, Z=-2.748, P=0.006), stride length(control group 0.19 (0.14, 0.24) m, observation group 0.26 (0.23, 0.38)m, Z=-1.360, P=0.005), the dynamic plantar peak pressure (control group -4.11 (-5.87, -2.57) N/cm 2, observation group -8.44 (-10.12, -4.81) N/cm 2, Z=-3.333, P=0.001) and average pressure (control group -0.55 (-1.00, -0.03) N/cm 2, observation group -1.11 (-1.51, -0.66) N/cm 2, Z=-2.062, P=0.009) in the observation group were better than those in the control group.After treatment, the BDNF level in peripheral blood in observation group was higher than before treatment( t=-2.315, P=0.033). Conclusion:Baduanjin can improve frozen gait score and gait parameters in PD patients with FOG, which may be related to the increase of peripheral blood BDNF.

Objective To investigate the role of RNA-binding motif protein X-linked(RBMX)in regulating the proliferation,migration,invasion and glycolysis in human bladder cancer cells.Methods A lentivirus vectors system and RNA interference technique were used to construct bladder cancer 1376 and UC-3 cell models with RBMX overexpression and knockdown,respectively,and successful cell modeling was verified using RT-qPCR and Western blotting.Proliferation and colony forming ability of the cells were evaluated using EdU assay and colony-forming assay,and cell migration and invasion abilities were determined using Transwell experiment.The expressions of glycolysis-related proteins M1 pyruvate kinase(PKM1)and M2 pyruvate kinase(PKM2)were detected using Western blotting.The effects of RBMX overexpression and knockdown on glycolysis in the bladder cancer cells were assessed using glucose and lactic acid detection kits.Results RT-qPCR and Western blotting confirmed successful construction of 1376 and UC-3 cell models with RBMX overexpression and knockdown.RBMX overexpression significantly inhibited the proliferation,clone formation,migration and invasion of bladder cancer cells,while RBMX knockdown produced the opposite effects.Western blotting results showed that RBMX overexpression increased the expression of PKM1 and decreased the expression of PKM2,while RBMX knockdown produced the opposite effects.Glucose consumption and lactate production levels were significantly lowered in the cells with RBMX overexpression(P<0.05)but increased significantly following RBMX knockdown(P<0.05).Conclusion RBMX overexpression inhibits bladder cancer progression and lowers glycolysis level in bladder cancer cells by downregulating PKM2 expression,suggesting the potential of RBMX as a molecular target for diagnosis and treatment of bladder cancer.

Objective:To explore the potential of integrating virtual surgery with three-dimensional (3D) printed guides in the surgical management of mandibular benign tumors and subsequent reconstruction of bone defects.Methods:A retrospective analysis was conducted on the clinical data of patients who underwent computer-assisted resection and vascularized fibular flap reconstruction for benign mandibular tumors at the Department of Oral and Maxillofacial Surgery, First Affiliated Hospital of Zhengzhou University, from June 2013 to December 2020. According to the utilization of guide plates for mandibular and fibular osteotomy during surgical procedures or not, the patients were categorized into two cohorts: a guide plate cohort and a non-guide plate cohort. In the guide plate group, custom-designed gudie plates based on virtual surgical plans were fabricated using 3D printing technology and employed intraoperatively; In the non-guide plate group, surgery was exclusively performed based on virtual surgical plan and prebent titanium plate without any supplementary plating. The measured outcomes included fibular flap osteotomy, operation duration, and clinical flap survival. Computed tomography images obtained one week post-surgery were utilized to assess the intersegmental commissure degree between fibular segments as well as between fibular segments and mandible, commissure degree between fibular segments and prebent titanium plate, and condyle position. The satisfaction of patients with their facial appearance was evaluated 6 months after the surgery using a visual analogue scale. Statistical analysis was conducted using SPSS 21.0 software. Independent sample t-tests was utilized to compare the duration of operation and and postoperative evaluation of facial appearance, the Chi-square tests was utilized for condyle position, commissure degrees among interactions involving fibular segments, prebent titanium plates, bone segments( P<0.05 denoted statistical significance). Results:A total of 30 patients were enrolled, comprising 17 males and 13 females, with a median age of 24 years (16-64 years). The preparation process of fibular flaps proceeded smoothly. The required length of fibula was measured as (14.1 ± 1.9) cm (5.7-18.1 cm), while the number of fibular segments ranged from 2 to 4, averaging at approximately 2.9 ± 0.6. The mandibular defects were repaired using a single-layer fibula in 12 cases, a vascularized folded fibula in 7 cases and a combination of vascularized and non-vascularized fibula in 11 cases. The operation time for the guide plate group was recorded as ( 335.9 ± 64.0) min (240-433 min), while it was observed to be (470.7 ± 140.5 ) min (280-680 min) for the non-guide plate group.The postoperative follow-up duration ranged from 9 to 23 months, with an average period of 11 months. All fibular flaps demonstrated clinical survival. The number of patients with good commissure degree between fibular and mandibular segments, between prebent titanium plate and fibular and mandibular segments and the position of condyle were 15, 15 and 13 cases in guide plate group, 10, 13 and 11 cases in non-guide plate group respectively. The statistical analysis revealed a significant difference ( P<0.05) in the degree of commissure between the fibular and the mandibular segments (15/15 vs. 10/15) in the two groups. Both groups exhibited high levels of satisfaction regarding their postoperative facial appearance at the 6 months follow-up, observed to be 9.6±0.5 and 9.3±0.5 respectively, and the statisticla analysis revealed non-significant difference ( P>0.05). Conclusion:The integration of virtual surgery with 3D printed guide plates can effectively reduce operative time and improve precision in the repair and reconstruction of free-fibular flaps following resection of benign tumors of the mandible.

Objective To investigate the role of RNA-binding motif protein X-linked(RBMX)in regulating the proliferation,migration,invasion and glycolysis in human bladder cancer cells.Methods A lentivirus vectors system and RNA interference technique were used to construct bladder cancer 1376 and UC-3 cell models with RBMX overexpression and knockdown,respectively,and successful cell modeling was verified using RT-qPCR and Western blotting.Proliferation and colony forming ability of the cells were evaluated using EdU assay and colony-forming assay,and cell migration and invasion abilities were determined using Transwell experiment.The expressions of glycolysis-related proteins M1 pyruvate kinase(PKM1)and M2 pyruvate kinase(PKM2)were detected using Western blotting.The effects of RBMX overexpression and knockdown on glycolysis in the bladder cancer cells were assessed using glucose and lactic acid detection kits.Results RT-qPCR and Western blotting confirmed successful construction of 1376 and UC-3 cell models with RBMX overexpression and knockdown.RBMX overexpression significantly inhibited the proliferation,clone formation,migration and invasion of bladder cancer cells,while RBMX knockdown produced the opposite effects.Western blotting results showed that RBMX overexpression increased the expression of PKM1 and decreased the expression of PKM2,while RBMX knockdown produced the opposite effects.Glucose consumption and lactate production levels were significantly lowered in the cells with RBMX overexpression(P<0.05)but increased significantly following RBMX knockdown(P<0.05).Conclusion RBMX overexpression inhibits bladder cancer progression and lowers glycolysis level in bladder cancer cells by downregulating PKM2 expression,suggesting the potential of RBMX as a molecular target for diagnosis and treatment of bladder cancer.