BACKGROUND: Group pre-test has confirmed that amnion endothelial cell conditioned medium can induce human umbilical cord blood mesenchymal stem cells into dopaminergic neuron-like cells. In this process, neurotrophic factors and their receptors may play an important role. OBJECTIVE: To study the function of neurotrophic factors secreted by amniotic epithelial cells in the differentiation of human umbilical cord blood mesenchymal stem cells into neurons.METHODS: P1 human umbilical cord blood mesenchymal stem cells at 2×10~8 /L were incubated and assigned to 3 group. Control group was added with HG-DMEM medium. Induction group received human amniotic epithelial cell medium. Blocking agent group underwent blocking agent K252a fluid, and the incubated was conducted at 36 ℃ for 40 minutes, and then amniotic epithelial cell medium was added. Immunofluorescence chemistry was used to determine neuron specific enolase and dopamine transporter expression in human umbilical cord blood mesenchymal stem cells. Real-time quantitative PCR was employed to detect neuron specific enolase, dopamine transporter and tyrosine hydroxylase expression in human umbilical cord blood mesenchymal stem cells. RESULTS AND CONCLUSION: Nerve growth factor and brain-derived neurotrophic factor were observed in human amniotic supernatant. P1 human umbilical cord blood mesenchymal stem cells expressed Trka and Trkb. Forty-eight hours following induction, compared with the control group, positive expression of neuron specific enolase and dopamine transporter was significantly increased in the induction and blocking agent groups (P < 0.05), especially in the induction group (P < 0.05). Neuron specific enolase, dopamine transporter and tyrosine hydroxylase mRNA levels were significantly greater in the induction and blocking agent groups compared with the control group (P < 0.01), and each gene mRNA levels were significantly greater in the induction group than in the blocking agent group (P < 0.01). Results verified that neurotrophic factor in the human amniotic epithelial cells plays important effects on differentiation of human umbilical cord blood mesenchymal stem cells into neurons. The promotion effects are mediated by activating Trk receptor.

Objective To explore the impacts of catechol-O-Methyltransferase (COMT) gene polymorphisms on the outcomes of methylphenidate treatment for attention deficit hyperactivity disorder (ADHD) in children. Methods One hundred seventy-seven ADHD children of Chinese Han descent received open-labelled dose titration with methylpheni?date to achieve optimal response in 2~4 weeks. The behavior changes were evaluated by using ADHD diagnostic scale (parent version) before and after treatment. COMT gene rs4680 (Val158Met) and rs165599 were genotyped using fluores?cent real-time PCR. The genotype distribution and treatment outcomes including remission, response and non-response were analyzed. Results The treatment response differed significantly among patients with different genotypes of rs4680 (P<0.01). Thirty-eight (40.9%) patients with G/G genotype had remission and 19 (20.4%) had good response;Thirty-five (48.6%) patients with G/A genotype had remission and 16 (22.2%) had good response. In contrast, only one patients with A/A genotype had remission and none of them had good response. The changes of the ADHD diagnostic scale total scores, inattention scores and hyperactivity impulsive scores were statistically different among different genotypes of rs4680 (P<0.05). The hyperactivity impulsive scores were significantly higher in the G allele carrier than A allele carrier (P=0.04). The treatment response was not significantly different among different rs165599 genotypes (P>0.05). Conclu?sion COMT gene rs4680 (Val158Met) polymorphism is associated with methylphenidate response in a Han Chinese popu?lation. Patients with G allele is more likely to benefit from methylphenidate treatment in comparison with A/A genotype.

Objective To study the associations between neo-adjuvant chemotherapy (NACT)sensitivity and expression of relative proliferation and apoptosis genes in cervical cancer tissues. Furthermore,the potential roles of relative genes expression as monitor in the chemotherapy against cervical cancer tissues were studied. Methods Fifteen pathologically proved Ⅱ A stage cervical cancer patients with HPV 16 positive were divided into effective group and non-effective group according to the clinical response to NACT, the changes of HPV16-E6, p53 genes and proteins expressions were measured by real-time quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blot. At the same time, MTT assay was used to detected proliferative activity to paclitaxel liposome and carboplatin (paraplatin) in HeLa cells. Results In effective group, the overall response rate was 73.3 % (11/15) and the HPV16-E6 mRNA and protein level after NACT was significantly decreased than that before NACT (t=3.359, t=3.614, P＜0.05), while p53mRNA and protein level increased significantly (t =5.852, t=2.838, P ＜0.05). However, there have no significant different between pre-NACT and Post-NACT of the two genes in the non-effective group. After using chemotherapy HeLa cells growth decreased. Conclusion HPV16-E6 and p53 expreesion level are useful paraneters in evaluating NACT response in cervical cancer tissues. The detection of the two genes expression is a method for predicting efficacy in NACT.

Objective To compare the sensitivities of the stimulation effect of human cord blood mesenchymal stem cells(CB-MSCs) and CB-MSCs of neuronal differentiation to lymphocytes(LCs) detected with carboxyfluorescein didcetate(CFSE) and MTT. Methods To prepare LCs from SD rat and divided into four group stimulating cells: 1. CB-MSCs;2. Dif-CB-HSCs;3. SH-SY5Y(positive control);4. Auto-LC(negative control).Stimulating cells were respectively Co-cultured with LCs. The proliferation of LCs was detected with MTT and CFSE ( n =3). Results CB-MSCs and Dif-CB-HSCs stimulated LCs to proliferate more weakly than positive control detected with MTT and CFSE. The quantity of proliferation of lymphocytes Co-cultured with CB-MSCs and Dif-CB-HSCs were higher than that of Auto-LC detected with CFSE. But MTT OD value of CB-MSCs and Dif-CB-HSCs was a little lower than that of Auto-LC. Statistical analysis results showed no significant difference.Conclusion CFSE can reflect proliferation status of lymphocytes better than MTT. CFSE shows more advantages in practical use.

Objective To study the effects of amniotic epithelial cells conditioned medium on the differentiation of bone marrow stromal cells into neural cells. Methods Bone marrow stromal cells and amniotic epithelial cells were isolated and cultured in vitro,then the cell surface antigen was detected by flow cytometry and the expressions of nestin and ki67 were detected by immunofluorescence staining method.When the cells were co-cultured with amniotic epithelial cells conditioned medium,the morphological character of cells was observed by inverse phase-contrast microscope,and the expressions of NSE(neurone specific enolase),TH(tyrosine hydroxylase) and DAT(dopamine transporter) were detected by immunofluorescence staining method. Results Amniotic epithelial cells conditioned medium had obvious inductive effect on bone marrow stromal cell's neural differentiation.Conclusion The amniotic epithelial cells conditioned medium may have inductive effect neuron-like cell's differentiation and dopaminergic neuron-like cell's differentiation of bone marrow stromal cells in vitro.

Objective To investigate the function of amnion endothelial cell in the differentiation of human umbilical cord blood stem cells into dopaminergic neurons.Methods Primary human amnion endothelial cells were separated and cultured in vitro;the conditioned medium(CM) was prepared through high speed centrifugation.The cord blood mesenchymal stem cells of P_1 passage were induced by the conditioned medium,and the mophology of cells was observed under the inverted phase contrast microscope.The expression of tyrosine hydroxylase(TH)and dopamine transportor(DAT) of the induced cord blood mesenchymal stem cells were detected by immunocytochemistry staining method and immunoblotting(Western blotting).Results The masculine rate of TH and DAT of the cord blood mesenchymal stem cells of P_1 passage which were induced by amnion endothelial cells conditioned medium was higher than that of the control group,with a significant difference(P

Objective:To study the association between set shifting in ADHD and NRXN1 gene.Methods:According to the diagnostic standard of the Diagnostic and Statistical Manual of Mental Disorders,Fourth Edition (DSM-Ⅳ).Totally 756 Han Chinese ADHD children and 133 Han Chinese unaffected children were involved in the analysis.Set shifting,including number connection time (NOTIM),number connection error times (NOERR),number and letter alternant connection time (LETIM),number and letter altemant error times (LEERR),and shifting time(each value was the difference between LETIM and NOTIM),was recorded by trail making test (TMT).Two SNPs (Single Nucleotide Polymorphisms) of NRXN1 gene,rs1592728 and rs4971652,were chose to detect genotype using Sequenom Mass ARRAY system by real time quantitative polymerase chain reaction.Linear regression analysis was applied to explore the influence of set shifting,then,stratified analysis was used to study the association between set shifting and rs1592728 as well as rs4971652 in ADHD cases and controls separately.Results:Linear regression analysis showed that there was a negative correlation between set shifting and month (β =0.42,P ＜0.001),IQ (β =0.34,P ＜ 0.001),group (β =0.08,P =0.004),GG genotype of rs4971652 (β =0.06,P =0.039).Among ADHD children,there was a negative relationship between set shifting and month (β =0.46,P ＜0.001),IQ (β =0.32,P ＜ 0.001),GG genotype of rs4971652 (β =0.07,P =0.018),a positive association was found between set shifting and ADHDSUB (β =0.06,P =0.033),set shifting damaged higher with ADHD-Ⅰ children than ADHD-C children.While,in controls,set shifting was in inverse relation with month (β =0.25,P =0.002) and IQ (β =0.40,P ＜ 0.001).Conclusion:It suggests that the association between shift in ADHD children and polymorphism of NRXN1 gene is existed,set shifting deficit less seriously in GG genotype.

Objective: To investigate the prevalence of postmenopausal osteoporosis (PMOP) and analyze its influencing factors among women at ages of 50 to 59 years in Dali Bai Autonomous Prefecture, Yunnan Province, so as to provide insights into the prevention of PMOP among menopausal women. Methods: Bai Ethnic menopausal women at ages of 50 to 59 years who received healthy examination at the Center of Healthy Examination, Dali Prefecture People's Hospital from June 2017 to May 2021 were selected as the study subjects, and subjects' demographic characteristics, living habits, history of diseases, family history of osteoporosis and history of parturition were collected using self-designed questionnaires. The height, body weight and bone density were measured, and fasting blood glucose, vitamin D3, blood lipids and liver functions were detected. The factors affecting the development of PMOP were identified using a multivariable logistic regression model. Results: Totally 2 000 questionnaires were allocated, and 1 584 valid questionnaires were recovered, with an effective recovery rate of 79.20%. The respondents had a mean age of ( 56.22±2.61 ) years, and mean body mass index ( BMI ) of ( 24.62±2.35 ) kg/m2. There were 497 respondents ( 31.38% ) with a family history of osteoporosis, and the prevalence of PMOP was 20.64%. Multivariable logistic regression analysis identified age ( OR=1.135, 95%CI: 1.074-1.196 ), age of menarche ( OR=1.138, 95%CI: 1.059-1.217 ), duration of menopause (OR=1.425, 95%CI: 1.228-1.622), number of parturition ( >2, OR=5.036, 95%CI: 2.972-7.101 ), smoking ( OR=2.594, 95%CI: 1.767- 3.421 ), alcohol consumption ( OR=2.051, 95%CI: 1.503-2.598 ), family history of osteoporosis ( OR=2.540, 95%CI: 1.769-3.311 ), hypertension ( OR=1.492, 95%CI: 1.406-1.578 ), diabetes ( OR=1.774, 95%CI: 1.581-1.967 ), total cholesterol ( OR=1.483, 95%CI: 1.251-1.716 ), triacylglycerol ( OR=1.801, 95%CI: 1.576-2.026 ), low-density lipoprotein cholesterol ( OR=1.614, 95%CI: 1.498-1.731 ), fasting blood glucose ( OR=1.192, 95%CI: 1.077-1.307 ), BMI ( OR=0.934, 95%CI: 0.862-0.993 ), outdoor activity ( ≥1 time/week, OR: 0.413-0.549, 95%CI: 0.329-0.637 ), age of menopause ( OR=0.909, 95%CI: 0.841-0.977 ), daily intake of calcium ( ≥600 mg, OR: 0.493-0.644, 95%CI: 0.389-0.786 ), vitamin D3 level ( ≥20 ng/mL, OR: 0.604-0.719, 95%CI: 0.523-0.853 ) and high-density lipoprotein cholesterol ( OR=0.658, 95%CI: 0.550-0.767 ) as factors affecting the development of PMOP. Conclusions The prevalence of PMOP in Dali Bai Autonomous Prefecture is similar to the nationwide level in China, and old age, smoking, alcohol consumption, a family history of osteoporosis and high blood lipid levels may increase the risk of PMOP.

Objective To explore the predictive value of inflammatory markers for stroke-associated pneumonia(SAP)in patients with acute ischemic stroke(AIS)based on the nomogram model.Methods According to whether pneumonia occurred,259 AIS patients were divided into SAP group(81 cases)and non-SAP group(178 cases).The clinical data of the two groups were compared.The systemic inflammatory response index(SIRI),systemic immunoinflammatory index(SII)and neutrophil to lymphocyte ratio(NLR)were calculated according to the formula.The variables with statistically significant differences were included in the multivariate binary Logistic regression model to screen out the independent risk factors for SAP in AIS patients.The independent risk factors were used to construct a predictive model,and the predictive ability of the two models,which only included traditional factors and included inflammatory indicators at the same time,was further compared from the aspects of discrimination,calibration,clinical practicability and so on.Reclassification analysis was used to evaluate the extent to which the nomogram model improved the predictive value of SAP risk in AIS patients.Results Compared with those in the non-SAP group,the rates of smoking,diabetes,dysphagia,leukocytes,neutrophils,lymphocytes,triglyceride level,NIHSS score on admission,SIRI,SII and NLR were significantly increased in the SAP group,and the rate of hypertension was decreased(all P＜0.05).Diabetes mellitus(OR =2.505,95%CI:1.070-5.850,P =0.034),dysphagia(OR =3.492,95%CI:1.501-8.119,P =0.004),NIHSS score on admission(OR = 1.310,95%CI:1.188-1.446,P＜0.001),SIRI(OR =2.417,95%CI:1.327-4.401,P =0.008),NLR(OR =1.434,95%CI:1.101-1.860,P =0.007)were independent risk factors for SAP in AIS patients.The area under the curve was 0.788(95%CI:0.725-0.852,P＜0.001)for the prediction model without inflammatory factors and 0.884(95%CI:0.838-0.930,P＜0.001)for the prediction model with independent risk factors.The calibration curve showed a good consistency between the predicted risk and the observed results.The decision curve showed that the model had a significant net benefit for predicting SAP.In addition,by calculating the net reclassification index(NRI)and the comprehensive discriminant improvement index(IDI),it was found that the nomogram model had a significant improvement in predicting the risk of SAP in AIS patients.Internal verification also proves the reliability of the nomogram model.Conclusions SIRI and NLR are independent predictors of SAP in AIS patients on admission.Adding SIRI and NLR to the traditional model can significantly improve the ability to identify the risk of SAP occurrence in AIS patients.

BACKGROUND:The use of mesenchymal stem cels in the field of tissue engineering for osteoarticular injury repair is a very promising tool since these cels are readily expandable and able to differentiate into chondrocytes. Abundant evidence suggests that microRNAs play critical roles in chondrogenic differentiation of mesenchymal stem cels.
OBJECTIVE:To observe the chondrogenic effect of human bone marrow mesenchymal stem cels transfected with lentiviral vectors bearing miR-221-3p/222-3p inhibition, thereby provding new strategies for cartilage injury.
METHODS: miRNA microarray technology was applied to detect microRNAs expression profiles at three different stages of chondrogenic differentiation induction after transforming growth factor-β3 treatment and verified by real-time fluorescence quantitative PCR (RT-qPCR). Human bone marrow mesenchymal stem cels were infected with lentivirus bearing miR-221-3p/222-3p inhibition. After co-suppressing the expression of miR-221/222-3p, cel counting kit-8 was used to determine the cel proliferation, the differentiation of bone marrow mesenchymal stem cels towards chondrocytes was verified by type II colagen protein expression through immunohistochemistry and glycosaminoglycan accumulation was also elevated by sarranine O staining. RT-PCR was used to detect type II colagen and aggrecan mRNA expression at 21 days of chondrogenic induction.
RESULTS AND CONCLUSION: The expression of miR-221-3p/222-3p was inhibited after Lv-miR221-3p/222-3p inhibition co-transfected into bone marrow mesenchymal stem cels. microRNA microarray and RT-qPCR results showed that the expression of miR-221-3p/222-3p was declined significantly at the anaphase of chondrogenic differentiation. The expression levels of chondrogenic markers, Aggrecan and type II colagen were significantly increased in the miR-221-3p/222-3p inhibition group and cel proliferation was also inhibited significantly compared with non-transduced cels or transduced with the empty lentiviral vector group. miR-221-3p/222-3p knockdown in bone marrow mesenchymal stem cels could inhibit proliferation but promote chondrogenic differentiation of bone marrow mesenchymal stem cels.