Objective:To establish and optimize a detection method of dibutyryl cyclic adenosine phosphate related substances.Methods:On the basis of the JP,the experimental conditions were optimized to obtain the optimal detection conditions.The column was CAPCEILPAK C18;mobile phase was pH 6.0 phosphate buffer solution-meth-anol with gradient elution;detection wavelength was 258 mm.The method validation was carried out.Results:In the system applicability solution,the separation degree of the main peak and adjacent peaks in each destruction solution was greater than 1.5;the solution was stable within 48 h.The content of each impurity showed a good line-ar relationship with the peak area in the range of 50％-150％of the limit concentration,R2＞0.999 4.In the spike experiment,the average recovery rate of N6-butyryl 3',5-cyclophosphatin sodium was 99.4％-106.6％.The average recoveries of 2'-0 butyryl 3,5-cycloadenosine monophosphate sodium were 88.8％-103.6％,and the average recoveries of cyclophosphaminate sodium were 96.9％-102.5％,which met the requirements.Three bat-ches of samples were tested,and the above impurities were detected.Conclusion:This method has high sensitivi-ty,good separation effect,accuracy and durability,and can be used for the detection of bucladesine sodium,provi-ding a basis for its quality control.

Objective To investigate whether the extracts of Patrinia heterophylla(MTH)inhibits the activation of macrophages induced by lipopolysaccharide(LPS)in vitro and the inflammatory response induced in vivo by suppressing the nuclear factor kappa B(NF-κB)signaling pathway.Methods Cellular studies:RAW264.7 cells were divided into 6 groups:control group,LPS group,LPS+Dexamethasone(DEX)group and LPS+MTH(low,medium,high dose)groups.The LPS group,LPS+DEX and LPS+MTH group were induced with a final concentration of 100 ng/ml LPS.While the LPS+DEX group was additionally treated with 5.0 ng/ml DEX,the LPS+MTH group was additionally treated with MTH(final concentrations of 0.1,0.2 and 0.4 mg/ml).The cell activity was detected using CCK-8 assay,cell invasion was detected using Transwell assay,cell apoptosis was detected using flow cytometry,and interleukin-6(IL-6),interleukin-17(IL-17)and tumor necrosis factor-α(TNF-α)were detected using real-time fluorescence quantitative PCR and enzyme-linked immunosorbent assay,respectively.The expression and secretion levels of nitric oxide(NO)in cells were detected by Griess assay,total reactive oxygen species(ROS)levels were detected by flow cytometry.The phosphorylation level of p65 and inhibitor of κB(IκB)were detected by protein immunoblotting assay.The transcriptional activity was detection by luciferase reporter gene assay.Animal studies:50 rats were randomly divided into 5 groups,10 per group,namely the control group,LPS group,LPS+DEX group,LPS+MTH low-dose group(6 g/kg)and LPS+MTH high-dose group(24 g/kg).LPS was injected into the lungs of rats,and the groups were orally administered at 36 h,24 h,12 h before modeling,and 12 h,24 h after modeling.12 h after the last administration,bronchoalveolar lavage fluid was collected,and the IL-6,IL-17,TNF-α and NF-κB signaling pathway-related indicators in the bronchoalveolar lavage fluid were measured.Results Cellular studies:Compared with the control group,the cell activity,invasion,apoptosis,IL-6,IL-17 and TNF-α mRNA and protein,NO,ROS,phosphorylation level of P536 protein S536 site and IκB protein S32 site and transcription activity of NF-κB had significantly increased in the LPS group(all P＜0.05).Compared with the LPS group,the cell activity,invasion,IL-6,IL-17,and TNF-αmRNA and protein,NO,ROS,phosphorylation level of P536 protein S536 site and IκB protein S32 site and transcription activity of NF-κB had significantly decreased in 3 LPS+MTH groups(all P＜0.05),and apoptosis was significantly increased in 3 LPS+MTH groups(all P＜0.05).All of which showed a dose-dependent trend of MTH.Animal studies:Compared with the control group,the LPS group showed a significant increase in IL-6,IL-17,TNF-α and phosphorylation levels of p65 protein at S536 site and IκB protein at S32 site(all P＜0.05).Compared with the LPS group,the 2 LPS+MTH groups showed a significant decrease in IL-6,IL-17,TNF-α and phosphorylation levels of p65 protein at S536 site and IκB protein at S32 site(all P＜0.05).These indicators showed a dose-dependent trend with MTH.Conclusion MTH can inhibit the activation of mouse macrophage RAW264.7 induced by LPS and the inflammatory response in the lungs of rats induced by LPS,which may be related to the inhibition of the NF-κB signaling pathway.

Objective To investigate whether the extracts of Patrinia heterophylla(MTH)inhibits the activation of macrophages induced by lipopolysaccharide(LPS)in vitro and the inflammatory response induced in vivo by suppressing the nuclear factor kappa B(NF-κB)signaling pathway.Methods Cellular studies:RAW264.7 cells were divided into 6 groups:control group,LPS group,LPS+Dexamethasone(DEX)group and LPS+MTH(low,medium,high dose)groups.The LPS group,LPS+DEX and LPS+MTH group were induced with a final concentration of 100 ng/ml LPS.While the LPS+DEX group was additionally treated with 5.0 ng/ml DEX,the LPS+MTH group was additionally treated with MTH(final concentrations of 0.1,0.2 and 0.4 mg/ml).The cell activity was detected using CCK-8 assay,cell invasion was detected using Transwell assay,cell apoptosis was detected using flow cytometry,and interleukin-6(IL-6),interleukin-17(IL-17)and tumor necrosis factor-α(TNF-α)were detected using real-time fluorescence quantitative PCR and enzyme-linked immunosorbent assay,respectively.The expression and secretion levels of nitric oxide(NO)in cells were detected by Griess assay,total reactive oxygen species(ROS)levels were detected by flow cytometry.The phosphorylation level of p65 and inhibitor of κB(IκB)were detected by protein immunoblotting assay.The transcriptional activity was detection by luciferase reporter gene assay.Animal studies:50 rats were randomly divided into 5 groups,10 per group,namely the control group,LPS group,LPS+DEX group,LPS+MTH low-dose group(6 g/kg)and LPS+MTH high-dose group(24 g/kg).LPS was injected into the lungs of rats,and the groups were orally administered at 36 h,24 h,12 h before modeling,and 12 h,24 h after modeling.12 h after the last administration,bronchoalveolar lavage fluid was collected,and the IL-6,IL-17,TNF-α and NF-κB signaling pathway-related indicators in the bronchoalveolar lavage fluid were measured.Results Cellular studies:Compared with the control group,the cell activity,invasion,apoptosis,IL-6,IL-17 and TNF-α mRNA and protein,NO,ROS,phosphorylation level of P536 protein S536 site and IκB protein S32 site and transcription activity of NF-κB had significantly increased in the LPS group(all P＜0.05).Compared with the LPS group,the cell activity,invasion,IL-6,IL-17,and TNF-αmRNA and protein,NO,ROS,phosphorylation level of P536 protein S536 site and IκB protein S32 site and transcription activity of NF-κB had significantly decreased in 3 LPS+MTH groups(all P＜0.05),and apoptosis was significantly increased in 3 LPS+MTH groups(all P＜0.05).All of which showed a dose-dependent trend of MTH.Animal studies:Compared with the control group,the LPS group showed a significant increase in IL-6,IL-17,TNF-α and phosphorylation levels of p65 protein at S536 site and IκB protein at S32 site(all P＜0.05).Compared with the LPS group,the 2 LPS+MTH groups showed a significant decrease in IL-6,IL-17,TNF-α and phosphorylation levels of p65 protein at S536 site and IκB protein at S32 site(all P＜0.05).These indicators showed a dose-dependent trend with MTH.Conclusion MTH can inhibit the activation of mouse macrophage RAW264.7 induced by LPS and the inflammatory response in the lungs of rats induced by LPS,which may be related to the inhibition of the NF-κB signaling pathway.

Objective: To investigate the milk drinking behavior and bone mineral density level of pupils in Hainan Province, and to explore the correlation between bone mineral density and milk drinking behavior, in order to provide scientific basis for promoting the healthy development of bones in children and adolescents. Methods: In November 2021, a cross sectional survey including demographic characteristics, milk intake, unhealthy eating behavior, physical activity and sleep was conducted among 696 students from grades 3 to 5 in Sanya and Baisha, Hainan by stratified cluster random sampling, and bone mineral density at the distal 1/3 of the right forearm was measured by dual energy X-ray absorptiometry. t-test was used to compare the differences in bone mineral density among different milk drinking behaviors of pupils, and multiple linear regression was used to analyze the correlation between milk consumption and bone mineral density. Results: About 25.3% students consumed milk daily and 13.9% consumed ≥ 300 g of milk daily. The mean bone mineral density at the distal 1/3 of the right forearm was (0.237±0.041)g/cm 2. The bone mineral density was greater in the group with daily milk intake than in the group without daily milk intake [(0.250± 0.037 )(0.204±0.034) g/cm 2 , t=15.00, P <0.01], and the bone mineral density was greater in the group with daily average milk intake ≥300 g than in the group with daily average milk intake <300 g [(0.284±0.036)(0.229±0.037)g/cm 2, t=13.48, P < 0.01 ]. Multiple linear regression analysis showed that daily average milk intake was positively correlated with bone mineral density, with a correlation coefficient ( β=0.020, t=21.46, P <0.01). Conclusion Milk consumption among pupils is inadequate, and milk drinking behavior has a positive impact on bone mineral density, so effective milk drinking intervention should be carried out to promote children s bone development.

Objective To compare the difference in dose of general anesthesia between pastoral patients and urban pa-tients.Methods The patients who performed abdominal surgery under general anesthesia in Peoples hospital of Bortala Mongo-lian Autonomous Prefecture from January 1st to April 30th 2022 were included and divided into pastoral patient group and ur-ban patient group according to the residential place of patients.Anesthesia and surgery were carried out according to hospital regulations.The general characteristics of all patients were recorded and the dosages of general anesthesia drugs were compared between the two groups.Results There was no significant difference between the two groups in general and intraoperative con-ditions.Patients'performances were stable,and no severe adveres reaction was observed.The dosages of Propofol,Remifen and Rocuronium in pastoral patients were higher than that in urban patients(all P＜0.05).Conclusion The dosage of general anes-thesia for patients in pastoral areas is higher,which ensures the stability of anesthesia depth and vital signs of patients,creates good conditions for surgery and promotes rapid recovery of patients.

Objective:To observe the recovery of exercise tolerance among stroke survivors using the cardiopulmonary exercise test (CPET) and analyze the factors influencing it.Methods:A total of 81 stroke survivors hospitalized in the Department of Rehabilitation Medicine of our hospital in year 2018 and 2019 were selected. The general clinical data of all patients were collected retrospectively and combined with the data from CPET to establish a data set. Version 25.0 of the SPSS software was used for multiple linear regressions analysis, with the peak oxygen uptake as the dependent variable, and the age, Brunnstrom stage, peak power, peak heart rate, peak respiratory exchange ratio, peak breathing reserve, peak ventilation per minute, the slope of the VE-VCO 2 curve (ΔVE/ΔVCO 2) and peak end tidal partial pressure of carbon dioxide as independent variables. Results:Stroke survivors at Brunnstrom stage III Ⅲ, Ⅳ and Ⅴ or higher decreased sequentially with their age, but their peak oxygen uptake increased gradually. The multiple linear regression model constructed by " stepwise method" showed that the fitted multiple linear regression equation was statistically significant ( F=100.228, P<0.001). Moreover, the average peak power, peak heart rate, peak ventilation per minute and the slope of the VE-VCO 2 curve were all found to be significant independent predictors of peak oxygen uptake in these stroke survivors. Conclusions:Skeletal muscle power, cardiac function, pulmonary ventilation and ventilation efficiency are useful independent predictors of the exercise tolerance of stroke survivors.

Objective: To analyze the clinical , immunological and genetic features of a child with BCL11B mutation induced neurodevelopmental disorder. Methods: The clinical data and genetic test of a child with BCL11B mutation hospitalized in the Department of Rheumatology and Immunology in Children′s Hospital of Chongqing Medical University in December 2018 were extracted and analyzed. The literature was searched with "BCL11B mutation" and "immunodeficiency 49" as key words in Chinese databases and Pubmed until January 2019 was reviewed. Results: A male patient aged 3 years and 11 months with facial dysmorphisms and delayed language and motor development was admitted due to neurodevelopmental retardation over two years. Laboratory tests showed normal human immunoglobulin (IgG 12.90 g/L, IgA 1.02 g/L, IgM 1.15 g/L, IgE 532 000 U/L), Trec (228) and proliferation of T and B cells. The lymphocyte subsets revealeda reduced percentage of B cells (0.108) but normal absolute numbers (0.574×10^-3/L), and an increased percentage (0.828) as well as absolute numbers (4.415×10^-3/L) of T cells. A heterozygous BCL11B mutation was detected by sanger sequencing, showing a de novo frameshift mutation c.1887_c.1893delCGGCGGG in exon 4. Two papers were found which were all in English, with total of 14 patients(13 patients with complete information). Thirteen mutations were reposed, including 7 frameshift, 2 nonsense, 2 missense, and 2 chromosomal rearrangements; Thirteen patients had heterozygous mutations. All patients had delayed language and motor development and facial dysplasia which were mainly hypertelorism, thin eyebrows and small palpebral fissures. Some patients had dental anomalies, ametropia and allergy, and a few were combined with immune impairment, but without overt signs of immunodeficiency. Only one patient had multisystem anomalies and profound immune deficiency. Conclusions BCL11B is essential for development of the nervous and the immune system. In this study, the de novo mutation of BCL11B gene resulted in neurodevelopmental and immunological disorders.

Objective:To analyze the clinical and immunological characteristics of a patient with activated phosphoinositide 3-kinase δ syndrome 2 (APDS2).Methods:A retrospective analysis of clinical data, immune-related gene sequencing, imaging and laboratory findings of a patient with APDS2 admitted to Children′s Hospital of Chongqing Medical University was performed. The absolute and relative numbers of peripheral lymphocyte subsets, immune cell subsets and phenotypes were detected by flow cytometry with the age matched healthy child or the patient's father as a control.Results:A female patient aged 6 years and 4 months old was firstly admitted due to paleness over one month and cough for 7 days in June 2017. The IgA (<0.067 g/L) decreased while the IgM (2.55 g/L) increased. The abdominal ultrasound found hepatomegaly (subcostal 1.7 cm) and splenomegaly (subcostal 3.6 cm), and gene sequencing revealed a heterozygous mutation in the PIK3R1 gene c.1425+1G>A. After the treatment with prednisone which was initiated with a dose of 10 mg/times, 3 times/d and continued and tapered over 7 months, the IgM decreased to normal (1.72 g/L), and the hepatomegaly (subcostal 0 cm) and splenomegaly (subcostal 0.5 cm) were improved. The patient was readmitted due to pale and sallow complexion for half a month in July 2019. The percentage of naive CD4 +T (0.386) and naive CD8 +T cells (0.271) were decreased while the percentage of terminally differentiated effector memory CD8 +T cells (0.377) and transitional B cells (0.223) were increased. The mean fluorescence intensity (MFI) of phosphorylated protein kinase B (AKT) in CD3 +T, CD4 +T and CD8 +T cells were higher in the patient (4 125, 5 213, 3 497) than those in her father (3 434, 3 312, 3 058). The percentage of follicular helper T cell (Tfh) (0.299), Th1 (0.491) and Th1-like cells (0.438) in the patient were higher than those in the healthy control (0.156,0.313,0.303), while the percentage of Th17 (0.126) and Th17-like cells (0.188) were lower than those in the healthy control (0.198, 0.315). And the percentage of CD57 in the patient (0.306) was also higher than that in the healthy control (0.246). Conclusions:The humoral immunity and cellular immunity of APDS2 patient are impaired to varying degrees. The steroid can improve the lymphoproliferation and autoimmune hemolytic anemia in this case.

Objective : To analyze the initial displacement of the upper central incisor and stress distribution of periodontal ligament under different torque values of upper incisors under the action of a four-curved auxiliary arch to provide a reliable basis for the safety of clinical application of four-curved auxiliary arches. Methods : A three-dimensional finite element model for torque control of upper anterior teeth with a homemade quadrilateral auxiliary arch was established. Four different states were analyzed: molar ligation without extraction space (group A), microimplant ligation without extraction space (group B), molar recovery with extraction space closure (group C) (the adductive traction force was set at 115 g) and microimplant recovery with extraction space closure (group D) (the adductive traction force was set at 115 g). When four types of torque (0.5 N, 1.0 N, 1.5 N, and 2.0 N) were applied. The initial displacement of upper central incisors and the stress distribution of periodontal ligament in 16 groups (A1-A4, B1-B4, C1-C4, D1-D4) were observed. Results : Under different conditions, as the strength of the four-curve auxiliary arch increases, the maxillary anterior teeth has crown labial inclination and a root lingual inclination. The displacement of the incisor tip increases with the increase in the loading force of the torque auxiliary arch, and the displacement of the incisor root apex increases as the force increases. The difference in incisor-apex displacement distance in A1-A4, B1-B4, C1-C4, D2 and D4 groups increased as the torque force increases, while the difference between the D3 group and D1 and D2 groups decreased slightly. The stress of the cervical periodontal ligament of the upper central incisor did not exceed the stress of the periodontal ligament in the following groups: A1, A2, B1, B2, B3, C1, C2, D1, and D2. The stress of the lip side of the upper central incisor did exceed the stress of the periodontal ligament in the following groups: A3, A4, B4, C3, C4, D3, and D4. In other words, when using the four-curved auxiliary arch as an implant anchorage, the force applied in the absence of extraction space should not exceed 1.5 N, and the force applied in the adduction of extraction space should not exceed 1.0 N. When using the nonimplant anchorage, the force applied in the absence of extraction space and the adduction of extraction space should not exceed 1.0 N. In addition, the range of force should not exceed the maximum stress of the periodontal ligament in the cervical region such that the effective and safe torque movement can be achieved. Under other stress conditions, the stress of the labial and cervical periodontal ligament of the upper central incisor exceeded the stress value (2.6 × 10-2MPa). The stress value of periodontal ligament was 2.6 × 10-2MPa in all groups. Conclusion A four-curved auxiliary arch has a significant effect on the upper anterior teeth, and the use of microimplants can better control root movement such that the crown of upper central incisors cannot be excessively lip inclined.

Objective:To optimize the preparation technology of rutin nanoemulsion. Methods:The pseudoternary phase diagram of rutin nanoemulsion under the condition of different Kmwas drawn. With the drug loading and the particle size as the independent var-iables,and the percent of oil and the weight ratio of surfactant to cosurfactant as the dependent variables, the blank formula of rutin nanoemulsion was optimized by central composite design-response surface method. Results:The optimal formula was as follows:the ra-tio of ethyl oleate, cremopher RH40,1,2-propanediol and water was 7: 13: 5: 25. The average particle size of the prepared rutin nanoemulsion was 26.51 nm,and the drug loading was 8.97 mg·ml-1. Conclusion:The central composite design-response surface method can obtain a good formula of rutin nanoemulison,and the model has a good predictive function.