Umbilical cord mesenchymal stem cells (UC-MSCs) have been widely used in regenerative medicine, but there is limited research on the stability of UC-MSCs formulation during production. This study aims to assess the stability of the cell stock solution and intermediate product throughout the production process, as well as the final product following reconstitution, in order to offer guidance for the manufacturing process and serve as a reference for formulation reconstitution methods. Three batches of cell formulation were produced and stored under low temperature (2-8 ℃) and room temperature (20-26 ℃) during cell stock solution and intermediate product stages. The storage time intervals for cell stock solution were 0, 2, 4, and 6 h, while for intermediate products, the intervals were 0, 1, 2, and 3 h. The evaluation items included visual inspection, viable cell concentration, cell viability, cell surface markers, lymphocyte proliferation inhibition rate, and sterility. Additionally, dilution and culture stability studies were performed after reconstitution of the cell product. The reconstitution diluents included 0.9% sodium chloride injection, 0.9% sodium chloride injection + 1% human serum albumin, and 0.9% sodium chloride injection + 2% human serum albumin, with dilution ratios of 10-fold and 40-fold. The storage time intervals after dilution were 0, 1, 2, 3, and 4 h. The reconstitution culture media included DMEM medium, DMEM + 2% platelet lysate, 0.9% sodium chloride injection, and 0.9% sodium chloride injection + 1% human serum albumin, and the culture duration was 24 h. The evaluation items were viable cell concentration and cell viability. The results showed that the cell stock solution remained stable for up to 6 h under both low temperature (2-8 ℃) and room temperature (20-26 ℃) conditions, while the intermediate product remained stable for up to 3 h under the same conditions. After formulation reconstitution, using sodium chloride injection diluted with 1% or 2% human serum albumin maintained a viability of over 80% within 4 h. It was observed that different dilution factors had an impact on cell viability. After formulation reconstitution, cultivation in medium with 2% platelet lysate resulted in a cell viability of over 80% after 24 h. In conclusion, the stability of cell stock solution within 6 h and intermediate product within 3 h meets the requirements. The addition of 1% or 2% human serum albumin in the reconstitution diluent can better protect the post-reconstitution cell viability.

ObjectiveThis study aims to explore the potential mechanism of Danggui Niantongtang in treating knee osteoarthritis （KOA） by regulating the intestinal flora through 16S rDNA analysis. MethodThirty-six C57BL/6J mice were subjected to anterior cruciate ligament transection （ACLT） to establish a KOA model and were randomly divided into the sham surgery group， model group， low-dose Danggui Niantongtang group （0.819 g·kg^-1）， medium-dose Danggui Niantongtang group （1.638 g·kg^-1）， high-dose Danggui Niantongtang group （3.276 g·kg^-1）， and Meloxicam group （0.975 mg·kg^-1）， with 6 mice in each group. Except for the treatment groups， the sham surgery group and model group were given normal saline by gavage. After 4 weeks of continuous intervention， feces and intact knee joints of the mice were collected. Hematoxylin-eosin （HE） staining and Safranin O-Fast Green staining were performed to observe the pathological changes in knee joint tissue morphology. The 16S rDNA sequencing was used to analyze changes in the abundance and diversity of intestinal microorganisms before and after treatment， along with corresponding functional predictions. ResultHigh-dose Danggui Niantongtang and Meloxicam significantly relieved pain symptoms in KOA mice， improved the disorder of joint structure， maintained the integrity of knee articular cartilage， increased the expression of type Ⅱ collagen alpha 1 （Col2a1） in articular cartilage， and decreased the expression of matrix metalloproteinase-13 （MMP-13）. The results of 16S rDNA sequencing showed that high-dose Danggui Niantongtang could adjust the abundance and structure of intestinal microbial species. Compared with the sham surgery group， the abundance of Proteobacteria， Actinobacteria， Ruminococcus， and Bacteroides was significantly increased in the model group （P<0.05）， while in the Danggui Niantongtang group， the abundance of these four flora was significantly reduced compared with the model group. Compared with the sham surgery group， the abundance of Verrucomicrobia， Oscillospira， and Akkermansia was significantly decreased in the model group （P<0.05）， while in the Danggui Niantongtang groups， the abundance of these three flora was significantly increased compared with the model group （P<0.05）. Functional pathway prediction of differential genera revealed that species differences among groups mainly involved metabolic pathways with high abundance associated with biosynthesis and precursors， as well as energy production， including amino acid biosynthesis， nucleotide and nucleoside biosynthesis， cofactors， prosthetic groups， electron carriers， and vitamin biosynthesis. ConclusionDanggui Niantongtang can effectively protect articular cartilage and delay the progression of KOA， possibly by regulating the structure of the intestinal flora， promoting probiotics， and inhibiting the growth of harmful pathogenic bacteria.

Objective To investigate the current situation and influencing factors of latent tuberculosis infection(LTBI)among close contacts of positive etiology pulmonary tuberculosis(PTB)patients,provide basis for formula-ting intervention measures for LTBI.Methods A multi-stage stratified cluster random sampling method was used to select close contacts of positive etiology PTB patients from 39 districts and counties in Chongqing City as the study objects.Demographic information was collected by questionnaire survey and the infection of Mycobacterium tuberculosis was detected by interferon gamma release assay(IGRA).The influencing factors of LTBI were analyzed by x2 test and binary logistic regression model.Results A total of 2 591 close contacts were included,the male to female ratio was 0.69∶1,with the mean age of(35.72±16.64)years.1 058 cases of LTBI were detected,Myco-bacterium tuberculosis latent infection rate was 40.83％.Univariate analysis showed that the infection rate was dif-ferent among peoples of different age,body mass index(BMI),occupation,education level,marital status,wheth-er they had chronic disease or major surgery history,whether they lived together with the indicator case,and whether the cumulative contact time with the indicator case ≥250 hours,difference were all statistically significant(all P＜0.05);infection rate presented increased trend with the increase of age and BMI(both P＜0.001),and decreased trend with the increase of education(P＜0.05).Logistic regression analysis showed that age 45-54 years old(OR=1.951,95％CI:1.031-3.693),age 55-64 years old(OR=2.473,95％CI:1.279-4.781),other occupations(OR=0.530,95％CI:0.292-0.964),teachers(OR=0.439,95％CI:0.242-0.794),students(OR=0.445,95％CI:0.233-0.851),junior high school education or below(OR=1.412,95％CI:1.025-1.944),BMI＜18.5 kg/m2(OR=0.762,95％CI:0.586-0.991),co-living with indicator cases(OR=1.621,95％CI1.316-1.997)and cumu-lative contact time with indicator cases ≥250 hours(OR=1.292,95％CI:1.083-1.540)were the influential fac-tors for LTBI(all P＜0.05).Conclusion The close contacts with positive etiology PTB have a high latent infection rate of Mycobacterium tuberculosis,and it is necessary to pay attention to close contacts of high age,farmers,and frequent contact with patients,and take timely targeted interventions to reduce the risk of occurrence of disease.

Child ; Humans ; Mpox (monkeypox)/prevention & control* ; Disease Outbreaks

Objective To explore the value of serum microRNAs-145(miR-145)and P53 antibody detection in the evaluation of therapeutic effect and prognosis in the treatment of breast cancer with Long Bei Xiaoyao Powder.Methods Ninety cases of breast cancer patients admitted to Beijing Aerospace General Hospital from January 2020 to January 2021 were selected as the study subjects.The patients were randomly divided into an observation group and a control group by random number table method,with 45 cases in each group.All of the patients in the two groups received individualized comprehensive treatment,and additionally the observation group was treated with Long Bei Xiaoyao Powder.The course of treatment covered three months.Real-time fluorescence quantitative polymerase chain reaction(PCR)method was used to detect serum miR-145 expression level,and enzyme-linked immunosorbent assay(ELISA)was used to detect serum P53 antibody expression level.The differences in the pre-and post-treatment expression levels of serum miR-145 and P53 antibodies in the two groups were compared,the relationship between the serum miR-145 and P53 antibody expression level and the poor prognosis of the patients in the observation group was explored by logistic regression analysis,and receiver operating characteristic curve(ROC)was drawn to analyze the value of the serum miR-145 and P53 antibody detection in the prediction of poor prognosis in the observation group.Results(1)After treatment,the detection value of serum miR-145 in the two groups of patients showed an increasing trend(P＜0.05),while that of the P53 antibody showed a decreasing trend(P＜0.05).The intergroup comparison showed that the detection value of serum miR-145 in the observation group was significantly higher than that of the control group,while the detection value of P53 antibody was significantly lower than that of the control group,the differences being all statistically significant(P＜0.05).(2)In the observation group,the detection value of serum miR-145 in the patients with good prognosis was significantly higher than that in the patients with poor prognosis,and the detection value of serum P53 antibody in the patients with good prognosis was significantly lower than that in the patients with poor prognosis,the differences being all statistically significant(P＜0.05).(3)Logistic regression analysis showed that high expression of miR-145 was a protective factor for the patients with poor prognosis in the observation group,and high expression of P53 antibody was a risk factor for the patients with poor prognosis in the observation group(P＜0.05).(4)The area under the ROC curve(AUC)for the prediction of prognosis of the patients in the observation group by the combination of serum miR-145 and P53 antibody was the largest(0.877,95%CI:0.769-0.985),and its sensitivity and specificity were 93.75%and 93.10%,respectively,with statistically significant differences(P＜0.05).Conclusion Low expression of serum miR-145 and high expression of serum P53 antibody are presented in the breast cancer patients with poor prognosis,and Long Bei Xiaoyao Powder exerts satisfactory therapeutic effect in treating breast cancer,which is conducive to enhancing the expression level of serum miR-145 and reducing the expression level of serum P53 antibody.

Aim To screen out the KCNQ channel o-peners and evaluate the antiepileptic activity.Methods The high throughput screening(HTS)method of Rb+efflux assay was used to identify the active com-pound of KCNQ opener;the preferred compound QO-7 2 was selected to test the pharmacological action in multiple animal models;through the analysis of behav-ioral and EEG,combined with the observation of gener-al pharmacological experiments,the efficacy and safety of the drug were preliminarily evaluated,and the mech-anism was explored.Results By HTS we identified three series compounds with high activity,a total of 51 compounds.In the results,the QO-72 ig or ip in differ-ent doses showed significant anticonvulsant activity in the MES and PTZ induced acute epilepsy models,the anticonvulsant protection rate significantly increased(P＜0.05,0.01)and the seizure threshold was signif-icantly extended(P＜0.01).In chronic epilepsy model,the seizure ranks and duration significantly de-creased in the QO-72 treatment groups(P＜0.01)and the antiepileptic protection rates significantly increased in the higher dose(P＜0.01).Compared with PTZ group,the amplitude,seizure wave duration and power density of EEG were reduced significantly in QO-72 treatment groups(P＜0.05,0.01).Besides,rotarod,spontaneous activity and cooperative sleep tests of mice by ig at 16 times,ip at 8 times of therapeutic dose had confirmed that the QO-72 had no central side effect.Further mechanism studies were performed on the QO-72 treated animals,the outcomes revealed that there was a significant elevation in GABA(P＜0.01)in hippocampus,but there was no significant change in Glu(P＞0.05).Conclusions The compound QO-72 shows significant antiepileptic activity in the MES and PTZ models;the mechanism is not only related to o-pening KCNQ channels,but also to increasing the con-tent of inhibitory neurotransmitter GABA in brain.

Objective:To investigate two cases of rare pathogenic genes,initiation codon mutations in HBA2 gene,combined with Southeast Asian deletion and their family members to understand the relationship of HBA2:c.2T＞C and HBA2:c.2delT mutations with clinical phenotype.Methods:The peripheral blood of family members was obtained for blood cell analysis and capillary electrophoresis hemoglobin analysis.Gap-PCR and reverse dot blotting(RDB)were used to detect common types of mutations in α-thalassaemia gene.Sanger sequencing was used to analyze HBA1 and HBA2 gene sequence.Results:Two proband genotypes were identified as--SEA/αα with HBA2:c.2T＞C and--SEA/αα with HBA2:c.2delT.HBA2:c.2T＞C/WT and HBA2:c.2delT/WT was detected in family members.They all presented with microcytic hypochromic anemia.Conclusion:When HBA2:c.2T＞C and HBA2:c.2delT are heterozygous that can lead to static α-thalassemia phenotype,and when combined with mild α-thalassemia,they can lead to the clinical manifestations of hemoglobin H disease.This study provides a basis for genetic counseling.

Objective:To investigate the characteristics of thalassemia gene types in children in Liuzhou,Guangxi.Methods:A total of 822 children suspected thalassemia aged from 1 day to 14 years who were admitted to our hospital from January 2019 to April 2022 were collected.Gap-PCR and PCR combined with reverse dot blot hybridization were used to detect α-and β-thalassemia genes.Results:Among 822 children,561 thalassemia carriers were detected,with a detection rate of 68.25％.Among them,303 cases were detected with α-thalassemia,and the most common genotype was--SEA/αα(163 cases),followed by-α3.7/αα(37 cases)and αcsα/αα(26 cases),44 cases with HbH disease.240 cases were detected with β-thalassemia,with a detection rate of 29.20％,and the most common genotype was βCD41-42/β N(112 cases),followed by βCD17/βN(75 cases)and βIVS-Ⅱ-654/β N(11 cases),11 cases with moderate to severe β-thalassemia.18 cases were detected with α β-thalassemia,with a detection rate of 2.19％,and--SEA/αα complex βCD41-42/βN was the most common genotype(4 cases).In Zhuang and Han populations,the detection ratio of-α3.7α/αα in α-thalassemia was the same(both 12.50％).While,the other main types such as--SEA/αα,αCSα/αα and-α4.2α/αα had certain differences.In β-thalassemia,CD41-42 and CD17 were the main genotypes detected in Han and Zhuang.Conclusion:In Liuzhou of Guangxi autonomous region,α-thalassemia is the main type in children,with a detection rate of 68.25％,and--SEA/αα is the most common genotype in mild thalassemia,followed by βCD41-42/βN.The detection rate of moderate to severe α-and β-thalassemia is relatively high.There are certain differences in the distribution of thalassemia among different ethnic groups.

Objective To determine the effects of treadmill training on the structure of hippocampal myelin and cognitive function in rats exposed to acute plateau hypoxia.Methods With 30 SPF-grade female SD rats (aged 6-8 weeks,weighing 200-220 g),6 of them were used for observation of myelin structure after injury,and the remaining 24 rats were randomly divided into control group,hypobaric hypoxia group and treadmill training group (n=8).The rats in above experimental groups were placed in a low-pressure oxygen chamber at an altitude of 6000 m for 7 consecutive days,and the rats of the control group were placed in the confined chamber for the same period without hypoxia.Then,the rats of the treadmill training group received a 4-week treadmill training scheme since the day after hypoxia.Finally,all the rats were tested for cognitive function with open field test (OFT)and Morris water maze (MWM).Transmission electron microscopy (TEM) was used to observe the changes of demyelination in the hippocampus. The expression of oligodendrocyte transcription factor 2 (Olig2)and myelin basic protein (MBP )in the hippocampal CA1 and CA3 regions was measured by immunofluorescence staining and Western blotting.Results Behavioral tests showed that the number into the central area,total distance,distance ratio in OFT and the number of platform crossings and distance to the target area in MWM were reduced in the hypobaric hypoxia group than the control group (P<0.05 ),while these indexes were increased in the treadmill training group than in the hypobaric hypoxia group (P<0.05).Immunofluorescence staining indicated that the number of Olig2 positive cells per unit area and the mean fluorescence intensity of MBP in the CA1 and CA3 regions were significantly lessen in the hypobaric hypoxia group than the control group (P<0.05 ),while these indicators were higher in the treadmill training group than the hypobaric hypoxia group (P<0.05 ).Western blotting displayed that the expression levels of Olig2 and MBP in the hippocampus were obviously lower in the hypobaric hypoxia group than the control group (P<0.01 ),while the levels were increased in the treadmill training group than the hypobaric hypoxia group (P<0.01 ).Conclusion Treadmill training promotes the number of the oligodendrocyte spectrum cells in CA1 and CA3 regions,enhances the expression of myelin-related proteins and improves myelin repair in hippocampus of hypobaric hypoxia rats,and thereby ameliorates hypoxia-induced anxiety-like behaviors and memory dysfunction.

Objective To investigate the clinical characteristics and prognosis of pneumococcal meningitis(PM),and drug sensitivity of Streptococcus pneumoniae(SP)isolates in Chinese children.Methods A retrospective analysis was conducted on clinical information,laboratory data,and microbiological data of 160 hospitalized children under 15 years old with PM from January 2019 to December 2020 in 33 tertiary hospitals across the country.Results Among the 160 children with PM,there were 103 males and 57 females.The age ranged from 15 days to 15 years,with 109 cases(68.1% )aged 3 months to under 3 years.SP strains were isolated from 95 cases(59.4% )in cerebrospinal fluid cultures and from 57 cases(35.6% )in blood cultures.The positive rates of SP detection by cerebrospinal fluid metagenomic next-generation sequencing and cerebrospinal fluid SP antigen testing were 40% (35/87)and 27% (21/78),respectively.Fifty-five cases(34.4% )had one or more risk factors for purulent meningitis,113 cases(70.6% )had one or more extra-cranial infectious foci,and 18 cases(11.3% )had underlying diseases.The most common clinical symptoms were fever(147 cases,91.9% ),followed by lethargy(98 cases,61.3% )and vomiting(61 cases,38.1% ).Sixty-nine cases(43.1% )experienced intracranial complications during hospitalization,with subdural effusion and/or empyema being the most common complication[43 cases(26.9% )],followed by hydrocephalus in 24 cases(15.0% ),brain abscess in 23 cases(14.4% ),and cerebral hemorrhage in 8 cases(5.0% ).Subdural effusion and/or empyema and hydrocephalus mainly occurred in children under 1 year old,with rates of 91% (39/43)and 83% (20/24),respectively.SP strains exhibited complete sensitivity to vancomycin(100% ,75/75),linezolid(100% ,56/56),and meropenem(100% ,6/6).High sensitivity rates were also observed for levofloxacin(81% ,22/27),moxifloxacin(82% ,14/17),rifampicin(96% ,25/26),and chloramphenicol(91% ,21/23).However,low sensitivity rates were found for penicillin(16% ,11/68)and clindamycin(6% ,1/17),and SP strains were completely resistant to erythromycin(100% ,31/31).The rates of discharge with cure and improvement were 22.5% (36/160)and 66.2% (106/160),respectively,while 18 cases(11.3% )had adverse outcomes.Conclusions Pediatric PM is more common in children aged 3 months to under 3 years.Intracranial complications are more frequently observed in children under 1 year old.Fever is the most common clinical manifestation of PM,and subdural effusion/emphysema and hydrocephalus are the most frequent complications.Non-culture detection methods for cerebrospinal fluid can improve pathogen detection rates.Adverse outcomes can be noted in more than 10% of PM cases.SP strains are high sensitivity to vancomycin,linezolid,meropenem,levofloxacin,moxifloxacin,rifampicin,and chloramphenicol.[Chinese Journal of Contemporary Pediatrics,2024,26(2):131-138]