Objective To observe the development of hematopoietic stem cell and the apoptosis of ICM(intermediate cell mass) in folic acid deficient zebrafish embryos and investigate the mechanism by which folic acid deficiency induces abnormal hematopoiesis.Method The folic acid deficient zebrafish model was induced by both using the dihydrofolate reductase antagonism methotrexate(MTX) and knock-down dihydrofolate reductase gene.The development of embryos was observed under microscope.The blood cells were detected by O-dianisidine staining.Whole-mount in situ hybridization and real-time PCR were performed to examine the expression of FLK-1,GATA1and GATA2.Apoptosis in intermediate cell mass(ICM) was examined by TUNEL(terminal-deoxynucleotidyl transferase mediated nick end labeling) method.Results The abnormal developments of ICMs were observed both in MTX treated embryos and DHFR knock-down embryos.O-dianisidine staining revealed that folic acid deficiency resulted in the decreasing number of blood cells.In folic acid deficient embryos,the expression of FLK-1、GATA1and GATA2 was reduced and the apoptosis in ICMs was increased.Conclusion The abnormal hematopoiesis in zebrafish induced by folic acid deficiency is related with the increasing apoptosis in ICMs and decreasing expressions of FLK-1,GATA1and GATA2.

Objective To construct a folic acid deficient model in zebrafish and to observe the axial development in folic acid deficient embryos, so as to probe the mechanism by which folic acid deficiency induces abnormal development of axis. Methods We constructed the folic acid deficient zebrafish model by both using the antagonism of dihydrofolate reductase (MTX) and knocking-down dihydrofolate reductase gene. Then we observed the axial excursion of folic acid deficient embryos at 17 hpf under microscope. We labeled and observed the positions of liver, spleen and heart by using whole-mount in situ hybridization with specific antisense RNA probes. The expressions of some genes, which are down stream factors of Nodal signal pathway and important for axial development, were detected by whole-mount in situ hybridization and Real-time PCR. Results Parts of folic acid deficient embryos had axial excursion and abnormal positions of liver, spleen and heart. The expressing intensities of ntl and gsc appeared normal in folic acid deficient embryos, but the expressing spatial patterns were abnormal, which revealed the malformation of axial mesoderm. Conclusions Folic acid deficiency induced the abnormal development of axis and the malformation of axial mesoderm. Folic acid deficiency had no obviouse effect on Nodal pathway.

Objective To study the putative mechanisms underlying fetal alcohol syndrome by comparative protein-profile analysis between normal and ethanol-treated zebrafish embryo with twodimensional electrophoresis (2-DE).Methods Zebrafish embryos were exposed in 400 mmol/L ethanol at dome stage for 3 hours,and then ethanol-induced abnormalities were observed.Proteomes of zebrafish embryos at early stages including zygote stage,dome stage,shield stage and 5-somite stage,were separated by 2-DE.The subtraction analysis method was applied to eliminate the interference from maternal derived proteins.The ethanol-treated embryos at 5-somite stage was analyzed by 2-DE,and the protein profile was compared with that generated from control embryos at the same stage.The data obtained from 2-DE analysis were verified by in-situ hybridization.Results 400 mmol/L ethanol treatment caused axial malformation (62%) and cyclopia (60%) in zebrafish embryos.The 2-DE analysis showed that the expression of Collagen2al (Col2a1) and TAR DNA binding protein (TDP) was decreased in 12 hours post fertilization (12 hpf) ethanol-treated embryos by 81% and 73%,respectively.The in-situ hybridization also demonstrated that the expression of Col2al in axial mesoderm was reduced by ethanol treatment at the same stage.But for 24 hpf ethanoltreated embryos,the expression of Col2al in axis recovered to a comparable level to that in control embryos,while the structure of neural tube was disrupted severely by ethanol exposure.Conclusions It is suggested that the expressions of Col2al and TDP were disrupted by ethanol during early stage,which might induce the zebrafish developmental abnormalities.The ethanol interference on early expression of Col2al is supposed to be one of the major reasons leading to later abnormalities of axis and neutral tube.

Objective: To investigate the effects of physical activity (PA) intensity and sedentary behavior (SB) on calcanues bone mineral density (BMD) in preschool children, so as to provide a basis for rationalizing the daily physical activity of preschool children to promote bone health. Methods: A total of 673 pre school children aged 3-6 years from nine kindergartens in Pingxiang City, Ganzhou City and Yingtan City of Jiangxi Province, were selected from September to December 2021 by using the whole stratified cluster random sampling method. The PA levels and SB were measured by using a three axis acceleration sensor, and left calcanues BMD was measured by an ultrasound bone densitometer. Multiple linear regression was used to explore the effects of changes in PA on calcanues BMD in pre school children of all ages. Results: Of the 673 preschoolers surveyed, 498 (74.0%) achieved an average of ≥60 min of moderate to vigorous physical activity (MVPA) per day, there were 265 boys (71.2%), and 233 girls ( 77.4 %). The difference between genders was not statistically significant ( χ 2=2.77, P >0.05). There was no statistically significant difference in the BMD test of the calcaneus bones of preschoolers by gender ( Z=0.42, P >0.05). The difference in BMD results of pre school children with 3, 4, 5 to 6 years was statistically significant ( H=2.65, P <0.05). Correlation analysis revealed a negative correlation between SB duration and calcaneus BMD ( r =-0.13), and a positive correlation between low intensity physical activity (LPA) duration, MVPA duration, and calcaneus BMD ( r =0.14, 0.25 ) ( P <0.05). Multivariate linear regression analysis showed that SB duration negatively correlated with calcaneus BMD, whereas LPA and MVPA duration positively correlated with calcaneus BMD ( P <0.05). Conclusions MVPA duration is positively correlated with the growth of BMD in the heel bone and negatively correlated with SB. The kindergartens can adjust their curricula according to the physical and mental developmental characteristics, gender and age differences of pre school children, increase the time of outdoor activities, and reduce the sedentary time to promote the bone health of young children.

Objective To compare the clinical outcomes and retinal volume changes in patients with ischemic and non-ischemic macular edema secondary to branch retinal vein occlusion(BRVO-ME)using optical coherence tomography angiography(OCTA).Methods The clinical data of 34 ischemic BRVO-ME patients(34 eyes,ischemic group)and 21 non-ischemic BRVO-ME patients(21 eyes,non-ischemic group)were retrospectively analyzed.Patients in both groups re-ceived intravitreal injections of ranibizumab.The best corrected visual acuity(BCVA)and retinal volume of the macular ar-ea were assessed before,1 day,1 week,1 month,3 months and 6 months after the treatment.Results The BCVA(log-MAR)at 1 day after the treatment was 0.63±0.37 in the ischemic group and 0.44±0.22 in the non-ischemic group,and the difference was statistically significant(P=0.017).The retinal volumes of the outer retina,the full retina,and the Farafovea and Perifovea subdivisions of the full retina before the treatment were(6.42±1.90)mm3,(8.75±1.82)mm3,(3.20±0.87)mm3 and(5.10±0.89)mm3 in the ischemic group and(5.52±1.57)mm3,(7.83±1.56)mm3,(2.80± 0.71)mm3,and(4.66±0.77)mm3 in the non-ischemic group,respectively;1 day after treatment,they were(4.97± 1.18)mm3,(7.46±1.47)mm3,(2.62±0.60)mm3 and(4.53±0.80)mm3 in the ischemic group and(4.25±0.48)mm3,(6.58±0.56)mm3,(2.26±0.26)mm3 and(4.06±0.40)mm3 in the non-ischemic group,respectively;at 1 week after the treatment,they were(4.40±0.82)mm3,(6.90±0.85)mm3,(2.38±0.36)mm3 and(4.24±0.49)mm3 in the ischemic group and(4.04±0.35)mm3,(6.33±0.49)mm3,(2.15±0.19)mm3 and(3.95±0.35)mm3 in the non-ische-mic group,respectively,and the differences between the two groups were statistically significant(all P＜0.05).The a-mount of retinal volume change from baseline in the outer retina and the full retina was(-2.48±2.38)mm3 and(-2.54±2.38)mm3 in the ischemic group,and(-1.31±1.58)mm3 and(-1.38±1.58)mm3 in the non-ischemic group at 1 month after treatment,respectively,and the differences between the two groups were statistically significant(both P＜0.05).Conclusion Ranibizumab is effective in treating both ischemic and non-ischemic BRVO-ME.The short-term visu-al prognosis is better in the non-ischemic group than the ischemic group,and the retinal volume is higher in the ischemic group than the non-ischemic group.However,no significant difference is observed in the visual prognosis or retinal volume between the two groups after long-term treatment.

Objective:A multi-center and large sample volume study was conducted on the verification and improvement of the early established criteria for intelligent routine urinalysis validation (including the microscopic review rules and manual validation rules, referred to as intelligent criteria for short), in order to improve the clinical application of this intelligent criteria.Methods:A total of 31 456 urine specimens were collected from the inpatients and outpatients in six hospitals in China, from March to September 2019. Firstly, 3105 specimens were analyzed for preliminary verification and improvement of the intelligent criteria based on the results of the microscopic examination and manual validation. Secondly, 28 351 specimens were used to verify the clinical application of the improved intelligent criteria. All samples were manually validated as reference.Results:The approval inconsistency rate of the manual validation rules in the original intelligent criteria was 8.59% (202/2 352), and the interception inconsistency rate was 8.84% (208/2 352). The false negative rate and the microscopic review rate of the microscopic review rules were similar to the previous results. Based on an in-depth analysis of big data and the discussions by senior technicians from eight hospitals, one microscopic review rules and four manual validation rules were added, meanwhile two manual validation rule was deleted. The manual validation standards were unified. Finally, the intelligent criteria was improved. Based on the improved intelligent criteria, for microscopic review rules, the false positive rate, false negative rate (misdiagnosis rate), and microscopic review rate did not change significantly, which were 14.72% (457/3 105), 4.06% (126/3 105), and 24.73% (768/3 105), respectively. The approval inconsistency rate and the interception inconsistency rate of manual validation rules were both reduced to 0; the total manual validation rate of the intelligent criteria was 50.89% (1 580/3 105), and the auto-validation rate was 49.11% (1 525/3 105). The large sample volume verification results were consistent with the preliminary verification results of the improved intelligent criteria.Conclusion:This multi-center and large sample volume study had shown that the improved intelligent criteria had better clinical performance.