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OBJECTIVE@#To establish an efficient protocol for directed differentiation of human induced pluripotent stem cells (hiPSCs) into functional midbrain dopaminergic progenitor cells (DAPs) in vitro.@*METHODS@#hiPSCs were induced to differentiate into DAPs in two developmental stages. In the first stage (the first 13 days), hiPSCs were induced into intermediate cells morphologically similar to primitive neuroepithelial cells (NECs) in neural induction medium containing a combination of small molecule compounds. In the second stage, the intermediate cells were further induced in neural differentiation medium until day 28 to obtain DAPs. After CM-DiI staining, the induced DAPs were stereotactically transplanted into the right medial forebrain bundle (MFB) of rat models of Parkinson's disease (PD). Eight weeks after transplantation, the motor behaviors of PD rats was evaluated. Immunofluorescence assay of brain sections of the rats was performed at 2 weeks after transplantation to observe the survival, migration and differentiation of the transplanted cells in the host brain microenvironment.@*RESULTS@#hiPSCs passaged stably on Matrigel showed a normal diploid karyotype, expressed the pluripotency markers OCT4, SOX2, and Nanog, and were positive for alkaline phosphatase. The primitive neuroepithelial cells obtained on day 13 formed dense cell colonies in the form of neural rosettes and expressed the neuroepithelial markers (SOX2, Nestin, and PAX6, 91.3%-92.8%). The DAPs on day 28 highly expressed the specific markers (TH, FOXA2, LMX1A and NURR1, 93.3-96.7%). In rat models of PD, the hiPSCs-DAPs survived and differentiated into TH+, FOXA2+ and Tuj1+ neurons at 2 weeks after transplantation. Eight weeks after transplantation, the motor function of PD rats was significantly improved as shown by water maze test (P < 0.0001) and apomorphine-induced rotation test (P < 0.0001) compared with rats receiving vehicle injection.@*CONCLUSION@#HiPSCs can be effectively induced to differentiate into DAPs capable of differentiating into functional neurons both in vivo and in vitro. In rat models of PD, the transplanted hiPSCs-DAPs can survive for more than 8 weeks in the MFB and differentiate into multiple functional neurocytes to ameliorate neurological deficits of the rats, suggesting the potential value of hiPSCs-DAPs transplantation for treatment of neurological diseases.
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Humans , Rats , Animals , Induced Pluripotent Stem Cells , Cell Differentiation/physiology , Neurons , Parkinson Disease , Mesencephalon , Cells, CulturedABSTRACT
OBJECTIVE@#To clarify the functional effects of differential expression of ring finger and tryptophan-aspartic acid 2 (RFWD2) on dendritic development and formation of dendritic spines in cerebral cortex neurons of mice.@*METHODS@#Immunofluorescent staining was used to identify the location and global expression profile of RFWD2 in mouse brain and determine the co-localization of RFWD2 with the synaptic proteins in the cortical neurons. We also examined the effects of RFWD2 over-expression (RFWD2-Myc) and RFWD2 knockdown (RFWD2-shRNA) on dendritic development, dendritic spine formation and synaptic function in cultured cortical neurons.@*RESULTS@#RFWD2 is highly expressed in the cerebral cortex and hippocampus of mice, and its expression level was positively correlated with the development of cerebral cortex neurons and dendrites. RFWD2 expression was detected on the presynaptic membrane and postsynaptic membrane of the neurons, and its expression levels were positively correlated with the length, number of branches and complexity of the dendrites. In cultured cortical neurons, RFWD2 overexpression significantly lowered the expressions of the synaptic proteins synaptophysin (P < 0.01) and postsynapic density protein 95 (P < 0.01), while RFWD2 knockdown significantly increased their expressions (both P < 0.05). Compared with the control and RFWD2-overexpressing cells, the neurons with RFWD2 knockdown showed significantly reduced number of dendritic spines (both P < 0.05).@*CONCLUSION@#RFWD2 can regulate the expression of the synaptic proteins, the development of the dendrites, the formation of the dendritic spines and synaptic function in mouse cerebral cortex neurons through ubiquitination of Pea3 family members and c-Jun, which may serve as potential treatment targets for neurological diseases.
Subject(s)
Animals , Mice , Aspartic Acid/metabolism , Cerebral Cortex , Dendritic Spines/metabolism , Neurons/metabolism , Synapses , Tryptophan/metabolismABSTRACT
Objective To explore the clinical significance of some risk factor,such as blood calcium serum alkaline phosphatase (ALP) and hemoglobin (Hb) for predicting or diagnosing bone metastases from urinary bladder neoplasms.Methods A total of 35 patients diagnosed as urinary bladder neoplasms and bone metastasis based on the histopathologic analysis (bone metastasis group,BM),and 105 cases of age-,sex-and follow-up time matched urinary bladder neoplasms patients without bone metastasis (non bone metastasis group,NBM) were included in present study.The correlations between clinical-pathological parameters (blood calcium,ALP,Hb) and bone metastases from urinary bladder neoplasms were analyzed and the risk factors for predicting or diagnosing bone metastases were identified.Results The levels of blood calcium,ALP and Hb were significantly higher at first diagnosis and follow-up period in BM group than in NBM group (P＜0.05).Multivariate logistic regression analysis indicated that the levels of calcium,ALP and Hb at first diagnosis were the independent risk factors for predicting the later incidence of bone metastases (P＜0.05).The levels of calcium,ALP and Hb in follow-up period were the independent risk factors for diagnosing bone metastases (P＜0.05).Receiver operating characteristic (ROC) curves analysis showed that the levels of calcium,ALP,Hb may give a more accurate prediction on bone metastases (AUC:0.615,0.704,0.692;cut-off:2.55mmol/L,97.0U/L,101.5g/L,respectively) or diagnosing bone metastases (AUC:0.706,0.776,0.829;cut-off:2.46mmol/L,112.5U/L,103.5g/L,respectively).Conclusion The levels of blood calcium,ALP and Hb may be used to predict the risk of bone metastases for patients firstly diagnosed as urinary bladder neoplasms,and predict whether the patients will be complicated with bone metastases during follow-up period.
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Objectives: To investigate the status of secondary prevention, treatment and risk factors of retired elderly patients (≥80 years old) with coronary heart disease in Guangzhou. Methods: A total of 752 questionnaires were collected from the elderly patients with coronary heart disease who underwent routine cardiovascular checkup in our hospital from March to May in 2016. The questionnaire included basic situation, complications, weight, blood pressure, low density lipoprotein, glucose level and secondary prevention medications based on evidence-based medicine. Results: Antiplatelet drugs, beta blockers, statins, ACEI/ARB use rate was 55.72%, 30.98%, 25.13%, 42.69% for the secondary prevention; 21.94% patients did not take secondary prevention drugs, the combined use rate of the four drugs was 4.92%. In elderly patients with coronary heart disease, the control rate of blood pressure (<150/90 mmHg), fasting blood glucose (<7mmol/L), 2-hour postprandial blood glucose (<10 mmol/L), body mass index (<24 kg/m2) were 81.17% (470/579 cases), 77.61% (253/326 cases), 55.21% (180/326 cases) and 54.73% (266/486 cases), respectively. With LDL-C<1.8 mmol/L as the standard, percent with normal LDL-C value was 14.06% (63/448 cases). Conclusions: In this survey population, use rate of the four types of drugs for secondary prevention and the control rate of known risk factors for coronary artery disease are very low. Efforts are needed to improve rational use of the secondary prevention drugs, minimize the gaps between promote the guidelines and clinical practice, and reduce the recurrence rate of coronary heart disease in elderly patients with coronary artery disease.
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Objective To observe the effects of diacetyldianhydrogalactito (DADAG) on the proliferation and apopotosis in lung cancer cell NCI-H460.Methods MTT assay was performed to determine the half inhibitory concentration of DADAG(2.88,5.75,11.50,23.00 and 46.00 μg/mL)for the NCI-H460 cells and colony formation assay was used to detect the ability of cell proliferation; through AO/EB staining method, morphological changes of apoptotic cell under fluorescent microscope were observed;cell apoptosis was detected by flow cytometry to test the effects of DADAG on NCI-H460;RT-PCR was used to detect the effect of DADAG on Bcl-2 and Bax mRNA expres-sion levels within the cell. Results Compared with that in blank control group, cell proliferation was inhibited in the group treated with DADAG; the number of colony formation decreased and AO/EB staining results showed that cell apoptosis was characterized by typical morphological changes such as swelling, shrinking and fragmentation. Flow cytometry detection results showed that apoptosis rate was increased in the group treated with DADAG(all P<0.05).RT-PCR results indicated that the expression of pro-apoptosis gene Bax was up-regulated,while expression of anti-apoptosis gene Bcl-2 down-regulated. Conclusions DADAG exhibits the inhibitory activity in lung cancer cell NCI-H460 and can induce the apoptosis.The potential mechanism maybe associated with up-regulating the ex-pression of Bax but down-regulating the expression of Bcl-2.
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OBJECTIVE@#To evaluate soluble interleukin-2 receptor alpha chain (sIL-2Rα, sCD25) in serum for the determination of rheumatoid arthritis (RA) activity.@*METHODS@#Peripheral blood was collected from 108 patients with RA, 39 patients with osteoarthritis (OA) and 50 healthy control subjects, and synovial fluids were from 40 patients with RA. The sera from the patients with RA, the disease control group (osteoarthritis), the healthy control group, and the synovial fluids of the RA patients were detected by enzyme-linked immunosorbent assay (ELISA).The clinical manifestations and laboratory parameters of the patients with RA were recorded and the correlation with the serum sCD25 level was analyzed.@*RESULTS@#The serum sCD25 concentration in RA group was (2 886±1 333) ng/L, the serum sCD25 concentration in OA group was (2 090±718) ng/L, and the serum sCD25 concentration in healthy group was (1 768±753) ng/L. The serum sCD25 level in the patients with RA was significantly higher than that in the disease controls and healthy controls (P<0.001). Sensitivity of serum sCD25 in the diagnosis of RA was 66.1% and specificity was 83.0%;serum sCD25 levels and erythrocyte sedimentation rate (r=0.321, P=0.001), C-reactive protein (r=0.446, P<0.001), DAS28 score (r=0.324, P<0.001), joint tenderness count (r=0.203, P=0.024), D-dimer levels (r=0.383, P<0.001), age (r=0.24, P=0.007), IgG (r=0.207, P=0.028), HRF-IgG (r=0.345, P=0.034) showed a significant positive correlation, and disease duration (r=-0.206, P=0.021) showed a negative correlation with sCD25;In patients with rheumatoid arthritis, the positive rates of serum ESR, CRP, and sCD25 were 14.3% (2 cases), 14.3% (2 cases), and 71.4% (10 cases) in the low disease activity group. The positive rates of serum ESR, CRP and sCD25 in the moderate disease activity group were 94.2% (49 cases), 82.7% (43 cases), and 86.5% (45 cases). The positive rates of serum ESR, CRP, and sCD25 in the high disease activity group were 100% (42 cases), 95.2% (40 cases), and 90.5% (38 cases);36 cases of ESR and/or CRP were negative (about 33.3%) in 108 patients, serum sCD5 levels of 17 cases in these 36 cases (about 47.2%)increased, of which 14 cases (about 82.4%) had a DAS28 score higher than 3.2.@*CONCLUSION@#The serum sCD25 has a high specificity for diagnosis of RA and a poor sensitivity. The serum level is closely related to the activity of RA, indicating that sCD25 may be involved in the inflammatory process of RA and may become a new inflammatory marker of RA. It is more meaningful for detection of serum sCD25 when RA is active, but ESR and/or CRP is negative.
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Humans , Arthritis, Rheumatoid/pathology , Biomarkers/analysis , Blood Sedimentation , C-Reactive Protein , Case-Control Studies , Interleukin-2 Receptor alpha Subunit/analysis , Osteoarthritis , Synovial Fluid/chemistryABSTRACT
Objective: To explore the prevalence and risk factors of orthostatic hypotension (OH) in elderly hypertension patients. Methods: A total of 532 retired hypertension patients elder than 65 years in Guangzhou military region were enrolled. The patients were divided into 2 groups: Hypertension group, n=414 and Hypertension combining OH (H+OH) group, n=118. The patient's age (65-79、≥ 80), hypertension grade (Grade 1-3) and complication status were studied. The risk factors for H+OH prevalence were analyzed by multivariate Logistic regression analysis. Results: The incidence rate of H+OH was 22.2％ (118/532). In H+OH group, the ratios of elderly and very elderly patients were 6.7％ and 23.1％, P<0.05 and the ratios of OH occurrence for hypertension grade 1, 2 and 3 were 12.6％, 23.3％ and 25.2％ respectively, P<0.05. Multivariate Logistic regression analysis presented that systolic blood pressure (BP) in supine position, BP at immediate standing, heart rate in supine position, heart rate after 2 minutes standing and chronic cardiac insufficiency were the impact factors for H+OH occurrence, P<0.05. Conclusion: In elderly hypertension patients, incidence of OH was increasing with age elevating; H+OH has been related to age, severity of hypertension and chronic cardiac insufficiency.
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Objective: To explore the prevalence and risk factors of orthostatic hypotension (OH) in elderly hypertension patients. Methods: A total of 532 retired hypertension patients elder than 65 years in Guangzhou military region were enrolled. The patients were divided into 2 groups: Hypertension group, n=414 and Hypertension combining OH (H+OH) group, n=118. The patient's age (65-79、≥ 80), hypertension grade (Grade 1-3) and complication status were studied. The risk factors for H+OH prevalence were analyzed by multivariate Logistic regression analysis. Results: The incidence rate of H+OH was 22.2％ (118/532). In H+OH group, the ratios of elderly and very elderly patients were 6.7％ and 23.1％, P<0.05 and the ratios of OH occurrence for hypertension grade 1, 2 and 3 were 12.6％, 23.3％ and 25.2％ respectively, P<0.05. Multivariate Logistic regression analysis presented that systolic blood pressure (BP) in supine position, BP at immediate standing, heart rate in supine position, heart rate after 2 minutes standing and chronic cardiac insufficiency were the impact factors for H+OH occurrence, P<0.05. Conclusion: In elderly hypertension patients, incidence of OH was increasing with age elevating; H+OH has been related to age, severity of hypertension and chronic cardiac insufficiency.