AIM:To construct and breed infertility model mice with ATP/GTP binding carboxypeptidase 1(Agtpbp1)gene knockout homozygote(Agtpbp1-/-)by CRISPR/Cas9 technology,so as to provide an animal model for the subsequent exploration of the pathogenesis of Agtpbp1 gene in male sterility.METHODS:The CRISPR/Cas9 technology was used to obtain Agtpbp1 gene knockout heterozygote(Agtpbp1+/-)mice according to the data of the main protein functional region of Agtpbp1 gene and combined with Cas9 nuclease.The obtained Agtpbp1+/-mice were mated,and their offspring were genotyped by PCR and agarose gel electrophoresis.The expression of Agtpbp1 at different levels was detected by RT-PCR,Western blot and immunohistochemical staining to support the identification results.The HE staining was used to observe the mouse cerebellum and eyeball structure to analyze the effect of Agtpbp1 gene knockout on Purkinje cells and photoreceptor cells.The symptoms of ataxia in mice were observed in combination with behavioral tests.The growth of mice was observed,and the changes of testicular tissue volume and weight of male mice were analyzed.The HE staining was used to observe the changes of testicular structure,and PAS staining was used to observe the changes of testicular germ cell cycle.Finally,sperm analyzer was used to analyze the sperm motility,so as to analyze the growth and develop-ment of the mice.RESULTS:The male and female Agtpbp1+/-mice could continue to mate,and three genotypes,Agtpbp1 wild-type(Agtpbp1+/+),Agtpbp1+/-and Agtpbp1-/-,were obtained.The genotypes of the offspring mice were successfully identified by PCR.The results of RT-PCR,Western blot and immunohistochemical staining verified the successful con-struction of Agtpbp1-/-mouse model at different levels(P<0.05).The results of HE staining showed that Purkinje cells were lost in the cerebellum of Agtpbp1-/-mice and the number of photoreceptor cells in the eyeball was reduced.Behavioral tests confirmed that Agtpbp1-/-mice had ataxia symptoms such as motor dysfunction and uncoordinated movements.Com-pared with control group,the testicular volume and weight of Agtpbp1-/-mice were significantly reduced.The results of HE staining showed a very small amount of sperm in the testis of Agtpbp1-/-mice.Combined with the sperm analyzer,it was ob-served that the sperm motility,vitality and movement rate of Agtpbp1-/-mice were significantly lower than those of the con-trol mice.Testicular sections with PAS staining showed cell cycle arrest of the sperm from Agtpbp1-/-mice.CONCLU-SION:In this study,Agtpbp1 knockout mice were successfully bred.The deletion of Agtpbp1 caused the arrest of sper-matogenic cell differentiation and the decrease in sperm motility in adult male mice,resulting in infertility.At the same time,it provides a new experimental tool for further exploring the molecular mechanism of Agtpbp1-induced male sterility.

Objective:To explore the effect and mechanism of Andrias davidianus skin mucopolysaccharides (ASMP) on full-thickness skin defect wound healing in diabetic mice. Methods:This study was an experimental study. The ASMP with polysaccharide content of (70.0±0.3)% was prepared; the proliferation activity of human umbilical vein endothelial cells (HUVECs) was detected by cell counting kit-8, showing that the optimal concentration of ASMP was 0.05 mg/mL. The HUVECs were taken and divided into blank control group, vascular endothelial growth factor (VEGF) group, and ASMP group according to the random number table method (the same grouping method below), which were cultured with conventional medium and the media containing 50 ng/mL VEGF and 0.05 mg/mL ASMP, respectively, and then cultured under hypoxic (with volume fraction of oxygen being 5%) and normal-oxygen conditions for 12 hours, and the length of tube formation was observed. Human monocytic leukemia cells were induced with phorbol ester to differentiate into M0 macrophages. These cells were then divided into blank control group, lipopolysaccharide (LPS) group, and ASMP group, which were cultured respectively using conventional medium, LPS-containing medium followed by conventional medium, and LPS-containing medium followed by 0.05 mg/mL ASMP-containing medium. After 48 hours of culture, the expressions of CD86 and CD206 proteins (expressed as relative fluorescence intensity, the same below) were measured by immunofluorescence, and the mRNA expression levels of arginase-1 (Arg1) and CD206 were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction. Eighteen male C57 mice aged 8-10 weeks were used, and diabetic model was successfully established using streptozotocin combined with a high-fat and high-sugar diet. Full-thickness skin defect wounds were created on the backs of the mice, and the mice were divided into blank control group, alginate dressing group, and ASMP group (with 6 mice in each group), which were treated with physiological saline, alginate dressing, and ASMP, respectively. Wound healing was observed on post injury day (PID) 3, 7, 10, and 14, and the wound healing rates of mice were calculated. On PID 7, the expressions of CD31 and CD206 proteins in the wound tissue of mice were observed by immunofluorescence. On PID 14, the thickness of granulation tissue in wounds of mice was observed by hematoxylin-eosin staining. The sample size for all experiments was 3.Results:After 12 hours of culture in normal-oxygen condition, compared with that in blank control group, the tube formation length of HUVECs in VEGF and ASMP groups was significantly increased (with q values of 10.08 and 16.91, respectively, P<0.05). After 12 hours of culture in hypoxic condition, compared with that in blank control group, the tube formation length of HUVECs in VEGF and ASMP groups was significantly increased (with q values of 11.61 and 16.91, respectively, P<0.05); compared with that in VEGF group, the tube formation length of HUVECs in ASMP group was significantly increased ( q=5.30, P<0.05). After 48 hours of culture, the relative fluorescence intensity of CD206 protein in M0 macrophages in ASMP group was 31.90±1.76, significantly higher than 1.00±0.25 in blank control group and 2.21±0.42 in LPS group (with q values of 50.75 and 48.75, respectively, both P values <0.05); the relative fluorescence intensity of CD86 protein was 5.82±0.63, significantly lower than 53.73±4.61 in LPS group ( q=30.90, P<0.05). After 48 hours of culture, the mRNA expressions of Arg1 and CD206 in M0 macrophages in ASMP group were significantly higher than those in blank control group (with q values of 35.02 and 13.09, respectively, P<0.05) and LPS group (with q values of 32.24 and 11.24, respectively, P<0.05). On PID 3, there was no statistically significant difference in intercomparison in the wound healing rate of mice among the blank control, alginate dressing, and ASMP groups ( P>0.05). Compared with those in blank control group, the wound healing rates of mice in alginate dressing group on PID 10 and 14 were significantly increased (with q values of 11.76 and 12.50, respectively, P<0.05), and the wound healing rates of mice in ASMP group on PID 7, 10, and 14 were significantly increased (with q values of 5.84, 15.90, and 14.96, respectively, P<0.05); compared with those in alginate dressing group, the wound healing rates of mice in ASMP group on PID 7 and 10 were significantly increased (with q values of 4.77 and 4.14, respectively, P<0.05). On PID 7, the relative fluorescence intensity of CD31 protein in wound tissue of mice in alginate dressing and ASMP groups was significantly stronger than that in blank control group (with q values of 7.63 and 16.85, respectively, P<0.05); the relative fluorescence intensity of CD31 protein in wound tissue of mice in ASMP group was significantly stronger than that in alginate dressing group ( q=9.22, P<0.05). On PID 7, the relative fluorescence intensity of CD206 protein in wound tissue of mice in alginate dressing and ASMP groups was significantly stronger than that in blank control group (with q values of 8.76 and 29.36, respectively, P<0.05), and the relative fluorescence intensity of CD206 protein in wound tissue of mice in ASMP group was significantly stronger than that in alginate dressing group ( q=20.61, P<0.05). On PID 14, the wound granulation tissue of mice in ASMP group was thicker compared with that in blank control group and alginate dressing group. Conclusions:ASMP can significantly enhance the ability of new blood vessel formation and optimize the immune microenvironment by promoting HUVEC tube formation as well as inducing macrophages to polarize toward the M2 type, thereby accelerating full-thickness skin defect wound healing in diabetic mice.

Objective To conduct a reliability and validity analysis of the quantitative diagnostic criteria for hysterosalpingography(HSG)in clinical trials.Methods Questionnaire survey method was used for this study.A test questionnaire scale was constructed based on the imaging data of 10 patients who had received HSG.Fifteen medical workers engaged in HSG-related clinical and scientific research work were invited to score the test questionnaire according to quantitative diagnostic criteria.Internal consistency testing,correlation analysis,and exploratory factor analysis were used to evaluate the reliability and validity of the quantified diagnostic criteria.Results The overall Cronbach's α coefficient for this quantitative diagnostic criteria was 0.834.The Cronbach's α coefficients for the three evaluation functions,including tubal patency,tubal adhesion and pelvic adhesion,were 0.722,0.627 and 0.724 respectively.The Spearman-Brown split-half reliability coefficient was 0.859,and the Guttman split-half coefficient was 0.830.The inter-rater reliability measured by the Kendall W coefficient was 0.806(P＜0.05).The analysis indicated that the evaluation function for pelvic adhesion was suitable for factor analysis(KMO value=0.573,Bartlett Spherical test P=0.003).Two common factors were extracted(characteristic root being set at 1),and the cumulative variance interpretation rate was up to 74.74％.These factors were named as"items with low ambiguity"and"items with potential ambiguity"respectively.Conclusion The overall reliability and validity of this quantitative diagnostic criteria are satisfactory,it can meet the requirements of HSG-related clinical trials and has the potential for clinical promotion.

Objective: To explore the longitudinal associations between dietary macronutrients and lung function in schoolaged children, so as to provide the nutritional research evidence for promoting children s lung health. Methods: In November 2021, two primary schools located in Chengdu, Sichuan Province were selected from the Southwest China Childhood Nutrition and Growth (SCCNG) cohort by a stratified cluster random sampling method, enrolling a total of 1 112 school aged children aged 8 to 13 years. At baseline, the dietary and sociodemographic characteristics of the children were assessed. One year later, the forced vital capacity (FVC) of the children was measured and converted into Z scores (FVC- Z ), while the vital capacity index (VCI) was also calculated. Generalized linear regression analysis was employed to examine the associations between dietary macronutrients and lung function, considering interactions with gender and age, followed by stratified analysis. Results: After adjusting for confounding factors, the analysis results of the generalized linear regression model showed that the carbohydrate energy ratio was negatively correlated with FVC- Z ( β =-0.02) and VCI ( β =-0.16), while the fat energy ratio showed a positive correlation with FVC- Z ( β =0.03) and VCI ( β =0.23) ( P <0.05). The protein energy ratio was positively correlated with FVC- Z ( β =0.09) and VCI ( β =0.60) specifically in girls ( P <0.05). Additionally, there was an interaction effect of age on the associations between macronutrients and lung function ( P <0.01); in children aged 8-9 and 10-11, the carbohydrate energy supply ratio was negatively correlated with FVC- Z ( β =-0.04, -0.03) and VCI ( β =-0.29, -0.21), and fat energy supply ratio was positively correlated with FVC- Z ( β =0.07, 0.05) and VCI ( β =0.46, 0.32) ( P <0.05). Conclusions There are age and sex differences in the association of dietary macronutrients with lung function, with a low carbohydrate, high fat diet promoting lung function in children. Additionally, protein intake appears to have a positive influence on the lung function of girls. The early school age period may represent a critical window for dietary interventions aimed at promoting lung health.

Objective:To explore the effect and mechanism of Andrias davidianus skin mucopolysaccharides (ASMP) on full-thickness skin defect wound healing in diabetic mice. Methods:This study was an experimental study. The ASMP with polysaccharide content of (70.0±0.3)% was prepared; the proliferation activity of human umbilical vein endothelial cells (HUVECs) was detected by cell counting kit-8, showing that the optimal concentration of ASMP was 0.05 mg/mL. The HUVECs were taken and divided into blank control group, vascular endothelial growth factor (VEGF) group, and ASMP group according to the random number table method (the same grouping method below), which were cultured with conventional medium and the media containing 50 ng/mL VEGF and 0.05 mg/mL ASMP, respectively, and then cultured under hypoxic (with volume fraction of oxygen being 5%) and normal-oxygen conditions for 12 hours, and the length of tube formation was observed. Human monocytic leukemia cells were induced with phorbol ester to differentiate into M0 macrophages. These cells were then divided into blank control group, lipopolysaccharide (LPS) group, and ASMP group, which were cultured respectively using conventional medium, LPS-containing medium followed by conventional medium, and LPS-containing medium followed by 0.05 mg/mL ASMP-containing medium. After 48 hours of culture, the expressions of CD86 and CD206 proteins (expressed as relative fluorescence intensity, the same below) were measured by immunofluorescence, and the mRNA expression levels of arginase-1 (Arg1) and CD206 were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction. Eighteen male C57 mice aged 8-10 weeks were used, and diabetic model was successfully established using streptozotocin combined with a high-fat and high-sugar diet. Full-thickness skin defect wounds were created on the backs of the mice, and the mice were divided into blank control group, alginate dressing group, and ASMP group (with 6 mice in each group), which were treated with physiological saline, alginate dressing, and ASMP, respectively. Wound healing was observed on post injury day (PID) 3, 7, 10, and 14, and the wound healing rates of mice were calculated. On PID 7, the expressions of CD31 and CD206 proteins in the wound tissue of mice were observed by immunofluorescence. On PID 14, the thickness of granulation tissue in wounds of mice was observed by hematoxylin-eosin staining. The sample size for all experiments was 3.Results:After 12 hours of culture in normal-oxygen condition, compared with that in blank control group, the tube formation length of HUVECs in VEGF and ASMP groups was significantly increased (with q values of 10.08 and 16.91, respectively, P<0.05). After 12 hours of culture in hypoxic condition, compared with that in blank control group, the tube formation length of HUVECs in VEGF and ASMP groups was significantly increased (with q values of 11.61 and 16.91, respectively, P<0.05); compared with that in VEGF group, the tube formation length of HUVECs in ASMP group was significantly increased ( q=5.30, P<0.05). After 48 hours of culture, the relative fluorescence intensity of CD206 protein in M0 macrophages in ASMP group was 31.90±1.76, significantly higher than 1.00±0.25 in blank control group and 2.21±0.42 in LPS group (with q values of 50.75 and 48.75, respectively, both P values <0.05); the relative fluorescence intensity of CD86 protein was 5.82±0.63, significantly lower than 53.73±4.61 in LPS group ( q=30.90, P<0.05). After 48 hours of culture, the mRNA expressions of Arg1 and CD206 in M0 macrophages in ASMP group were significantly higher than those in blank control group (with q values of 35.02 and 13.09, respectively, P<0.05) and LPS group (with q values of 32.24 and 11.24, respectively, P<0.05). On PID 3, there was no statistically significant difference in intercomparison in the wound healing rate of mice among the blank control, alginate dressing, and ASMP groups ( P>0.05). Compared with those in blank control group, the wound healing rates of mice in alginate dressing group on PID 10 and 14 were significantly increased (with q values of 11.76 and 12.50, respectively, P<0.05), and the wound healing rates of mice in ASMP group on PID 7, 10, and 14 were significantly increased (with q values of 5.84, 15.90, and 14.96, respectively, P<0.05); compared with those in alginate dressing group, the wound healing rates of mice in ASMP group on PID 7 and 10 were significantly increased (with q values of 4.77 and 4.14, respectively, P<0.05). On PID 7, the relative fluorescence intensity of CD31 protein in wound tissue of mice in alginate dressing and ASMP groups was significantly stronger than that in blank control group (with q values of 7.63 and 16.85, respectively, P<0.05); the relative fluorescence intensity of CD31 protein in wound tissue of mice in ASMP group was significantly stronger than that in alginate dressing group ( q=9.22, P<0.05). On PID 7, the relative fluorescence intensity of CD206 protein in wound tissue of mice in alginate dressing and ASMP groups was significantly stronger than that in blank control group (with q values of 8.76 and 29.36, respectively, P<0.05), and the relative fluorescence intensity of CD206 protein in wound tissue of mice in ASMP group was significantly stronger than that in alginate dressing group ( q=20.61, P<0.05). On PID 14, the wound granulation tissue of mice in ASMP group was thicker compared with that in blank control group and alginate dressing group. Conclusions:ASMP can significantly enhance the ability of new blood vessel formation and optimize the immune microenvironment by promoting HUVEC tube formation as well as inducing macrophages to polarize toward the M2 type, thereby accelerating full-thickness skin defect wound healing in diabetic mice.

Objective:To evaluate the efficacy and safety of low-dose post-transplant cyclophosphamide (PTCY) combined with anti-thymocyte globulin (ATG) in preventing graft-versus-host disease (GVHD) in children with β-thalassemia after hematopoietic stem-cell transplantation (HSCT).Method:A retrospective analysis was conducted on 42 children with transfusion-dependent β-thalassemia who underwent HSCT at Zhongshan Hospital, Xiamen University between March 2019 and June 2023. Based on donor source, recipients were grouped into the haploidentical donor group (Haplo-RD, 10 cases) and the unrelated donor group (UD, 32 cases). The UD group was further subdivided into HLA 8/10 matched (2 cases), HLA 9/10 matched (15 cases), and fully HLA-matched (15 cases). The conditioning regimen included fludarabine, busulfan, cyclophosphamide, and thiotepa. GVHD prophylaxis consisted of ATG (4.5 mg/kg), cyclophosphamide (25 mg/kg for 2 days), cyclosporine A (CsA), and mycophenolate mofetil (MMF). Engraftment, GVHD incidence, survival, mortality, and virus reactivation rates were evaluated.Result:The median age was 6 years (range, 2~12). All patients achieved hematopoietic reconstitution. The median times to neutrophil and platelet engraftment were 11 days (range, 10~15) days and 12 days (range, 6~31), respectively. All recipients had >95% peripheral blood STR chimerism by day 30. Grade Ⅲ~Ⅳ acute GVHD occurred in 3 recipients (7.14%), and chronic GVHD occurred in 5 recipients (11.90%) -1 case extensive, 4 cases limited. Both overall survival (OS) and disease-free survival (DFS) were 92.86%. All children in the Haplo-RD group achieved DFS. Eight patients developed cytomegalovirus (CMV) viremia (no CMV disease), with a reactivation rate of 19.05%, and 9 recipients had BK virus-related urinary tract infections (6 cases in the UD group and 3 cases in the Haplo-RD group), for a total incidence of 21.43%.Conclusion:The combination of low-dose PTCY and ATG is a safe and effective strategy to prevent GVHD following haploidentical or unrelated donor HSCT in pediatric β-thalassemia. It is associated with reduced infection and viral reactivation post-transplant and contributes to high survival rates.

AIM:To construct and breed infertility model mice with ATP/GTP binding carboxypeptidase 1(Agtpbp1)gene knockout homozygote(Agtpbp1-/-)by CRISPR/Cas9 technology,so as to provide an animal model for the subsequent exploration of the pathogenesis of Agtpbp1 gene in male sterility.METHODS:The CRISPR/Cas9 technology was used to obtain Agtpbp1 gene knockout heterozygote(Agtpbp1+/-)mice according to the data of the main protein functional region of Agtpbp1 gene and combined with Cas9 nuclease.The obtained Agtpbp1+/-mice were mated,and their offspring were genotyped by PCR and agarose gel electrophoresis.The expression of Agtpbp1 at different levels was detected by RT-PCR,Western blot and immunohistochemical staining to support the identification results.The HE staining was used to observe the mouse cerebellum and eyeball structure to analyze the effect of Agtpbp1 gene knockout on Purkinje cells and photoreceptor cells.The symptoms of ataxia in mice were observed in combination with behavioral tests.The growth of mice was observed,and the changes of testicular tissue volume and weight of male mice were analyzed.The HE staining was used to observe the changes of testicular structure,and PAS staining was used to observe the changes of testicular germ cell cycle.Finally,sperm analyzer was used to analyze the sperm motility,so as to analyze the growth and develop-ment of the mice.RESULTS:The male and female Agtpbp1+/-mice could continue to mate,and three genotypes,Agtpbp1 wild-type(Agtpbp1+/+),Agtpbp1+/-and Agtpbp1-/-,were obtained.The genotypes of the offspring mice were successfully identified by PCR.The results of RT-PCR,Western blot and immunohistochemical staining verified the successful con-struction of Agtpbp1-/-mouse model at different levels(P<0.05).The results of HE staining showed that Purkinje cells were lost in the cerebellum of Agtpbp1-/-mice and the number of photoreceptor cells in the eyeball was reduced.Behavioral tests confirmed that Agtpbp1-/-mice had ataxia symptoms such as motor dysfunction and uncoordinated movements.Com-pared with control group,the testicular volume and weight of Agtpbp1-/-mice were significantly reduced.The results of HE staining showed a very small amount of sperm in the testis of Agtpbp1-/-mice.Combined with the sperm analyzer,it was ob-served that the sperm motility,vitality and movement rate of Agtpbp1-/-mice were significantly lower than those of the con-trol mice.Testicular sections with PAS staining showed cell cycle arrest of the sperm from Agtpbp1-/-mice.CONCLU-SION:In this study,Agtpbp1 knockout mice were successfully bred.The deletion of Agtpbp1 caused the arrest of sper-matogenic cell differentiation and the decrease in sperm motility in adult male mice,resulting in infertility.At the same time,it provides a new experimental tool for further exploring the molecular mechanism of Agtpbp1-induced male sterility.

Objective:To evaluate the efficacy and safety of low-dose post-transplant cyclophosphamide (PTCY) combined with anti-thymocyte globulin (ATG) in preventing graft-versus-host disease (GVHD) in children with β-thalassemia after hematopoietic stem-cell transplantation (HSCT).Method:A retrospective analysis was conducted on 42 children with transfusion-dependent β-thalassemia who underwent HSCT at Zhongshan Hospital, Xiamen University between March 2019 and June 2023. Based on donor source, recipients were grouped into the haploidentical donor group (Haplo-RD, 10 cases) and the unrelated donor group (UD, 32 cases). The UD group was further subdivided into HLA 8/10 matched (2 cases), HLA 9/10 matched (15 cases), and fully HLA-matched (15 cases). The conditioning regimen included fludarabine, busulfan, cyclophosphamide, and thiotepa. GVHD prophylaxis consisted of ATG (4.5 mg/kg), cyclophosphamide (25 mg/kg for 2 days), cyclosporine A (CsA), and mycophenolate mofetil (MMF). Engraftment, GVHD incidence, survival, mortality, and virus reactivation rates were evaluated.Result:The median age was 6 years (range, 2~12). All patients achieved hematopoietic reconstitution. The median times to neutrophil and platelet engraftment were 11 days (range, 10~15) days and 12 days (range, 6~31), respectively. All recipients had >95% peripheral blood STR chimerism by day 30. Grade Ⅲ~Ⅳ acute GVHD occurred in 3 recipients (7.14%), and chronic GVHD occurred in 5 recipients (11.90%) -1 case extensive, 4 cases limited. Both overall survival (OS) and disease-free survival (DFS) were 92.86%. All children in the Haplo-RD group achieved DFS. Eight patients developed cytomegalovirus (CMV) viremia (no CMV disease), with a reactivation rate of 19.05%, and 9 recipients had BK virus-related urinary tract infections (6 cases in the UD group and 3 cases in the Haplo-RD group), for a total incidence of 21.43%.Conclusion:The combination of low-dose PTCY and ATG is a safe and effective strategy to prevent GVHD following haploidentical or unrelated donor HSCT in pediatric β-thalassemia. It is associated with reduced infection and viral reactivation post-transplant and contributes to high survival rates.

We report a case of a male patient who developed persistent fever and central nervous system symptoms after eating raw snails for 10 days. The patient was diagnosed with Angiostrongyliasis depended on the clinical presentation, epidemiological history, and etiological results. The patient recovered after receiving albendazole anthelmintic and dexamethasone anti-inflammatory therapy. This article incorporates literature review to sort out the diagnosis and treatment of this patient, in order to provide feasible reference for clinicians.

We report a case of a male patient who developed persistent fever and central nervous system symptoms after eating raw snails for 10 days. The patient was diagnosed with Angiostrongyliasis depended on the clinical presentation, epidemiological history, and etiological results. The patient recovered after receiving albendazole anthelmintic and dexamethasone anti-inflammatory therapy. This article incorporates literature review to sort out the diagnosis and treatment of this patient, in order to provide feasible reference for clinicians.