UCMSCs-derived exosomal circHIPK3 promotes ulcer wound angiogenesis of diabetes mellitus via miR-20b-5p/Nrf2/VEGFA axis.

AIMS: Experiments confirmed that circular RNAs contributed to the pathogenesis of diabetic foot ulcers (DFUs). CircHIPK3 was upregulated in type 2 diabetes mellitus (T2DM), but its role in DFU remained unknown. Our study aimed to investigate the regulatory functions of exosomal circHIPK3 and its potential mechanisms in DFU. METHODS: Exosomal size and distribution, marker proteins, and circHIPK3 levels were evaluated by transmission electron microscope, ExoView R200, western blot, and qRT-PCR. Flow cytometry, MTT, Wound healing assays, and tube formation assays were used to assess the roles of exosomal circHIPK3 in high glucose (HG)-treated human umbilical vein endothelial cells (HUVECs). The relationships between Nrf2/VEGFA/circHIPK3 and miR-20b-5p, and between Nrf2 and VEGFA were determined by luciferase reporter assay and RNA immunoprecipitation. We used cell and mice models to investigate the mechanisms of exosomal circHIPK3 under diabetic conditions. RESULTS: CircHIPK3 was significantly upregulated in exo-circHIPK3 rather than exo-vector. Exo-circHIPK3 remarkably inhibited cell apoptosis but promoted cell proliferation, migration, and tube formation in HG-treated HUVECs. Luciferase reporter and RIP assays showed that miR-20b-5p targeted and inhibited Nrf2 and VEGFA, and circHIPK3 acted as a ceRNA of miR-20b-5p to inhibit the binding to its downstream genes Nrf2 and VEGFA. Mechanistically, circHIPK3 promoted cell proliferation, migration, and angiogenesis via downregulating miR-20b-5p to upregulate Nrf2 and VEGFA. However, the overexpressed miR-20b-5p could abolish the promoting effects of circHIPK3 overexpression on cell proliferation, migration, and tube formation under HG conditions. CONCLUSION: UCMSCs-derived exosomal circHIPK3 protected HG-treated HUVECs via miR-20b-5p/Nrf2/VEGFA axis. The exosomal circHIPK3 might be a therapeutic candidate to treat DFU.

Reference values of induced sputum cytology in healthy children in guangzhou, southern china.

OBJECTIVE: To establish normal reference values of induced sputum cytology in healthy children in southern China. METHODS: During a period from January 2010 to December 2011, a total of 580 healthy children (5-16 years of age) were approached. A total of 266 children (137 boys and 129 girls) participated in the study. Sputum induction was carried out by using 5% hypertonic saline. Cell types in the sputum were examined by using routine methods. RESULTS: Sputum induction was completed in 175 of the 266 subjects (65.79%), but 16 sputum samples were disqualified. The overall success rate was 59.77% (159/266). Macrophages and neutrophils were the predominant cell types: macrophages: median, 76.14%; interquartile range (IQR), 32.68%; and 2.5% to 97.5% percentile, 1.00% to 94.50%; neutrophils: median, 20.67%; IQR, 33.0%; and 2.5% to 97.5% percentile, 4.00% to 92.75%; eosinophils: median, 0.39%; IQR, 1.93%; and 2.5% to 97.5% percentile, 0.00% to 6.50%; and lymphocytes: median, 1.22%; IQR, 2.04%; and 2.5% to 97.5% percentile, 0.00% to 5.00%. The cell types did not differ among different age, gender, and passive smoking groups. Adverse events occurred in 4.4% (7/159) of the participants who completed the procedures but required no specific treatment to dissipate. Peak expiratory flow did not differ between those who completed the procedures compared with those who did not, suggesting that the procedure is safe and feasible in children. CONCLUSIONS: The current study represents the first attempt to develop normal reference values of induced sputum cytology in Chinese children, and could be used as a control for future studies.

[Preliminary establishment of cytological examination and the normal reference values for hypertonic saline solution-induced sputum of healthy children in Guangzhou].

OBJECTIVE: To establish the method of cytological examination and the normal reference values for hypertonic saline solution-induced sputum of healthy children (age range from 5 to 15 years) with physical examination in Guangzhou. METHOD: A total of 352 children, 5 to 15 years old, were enrolled from primary school and middle school in Guangzhou from January to December, 2010. All subjects completed a standardized questionnaire on the presence of respiratory, allergic symptoms and family history, the medical history and the physical examination was performed by doctors, lung function (forced expiratory volume at 1 s in predicted normal, FEV(1)%) was determined. There were 266 healthy children (137 males, 129 females) who were selected and undergone hypertonic saline solution induction of sputum, and cytological examination was performed. Hypertonic saline (5%) was nebulized and inhaled for 15 - 30 min. No expectoration within 30 min was defined as failure, and the procedure was terminated. The part of opaque and higher density sputum samples was detected by cytology. The proportion of neutrophils, lymphocytes, eosinophils, macrophages and monocytes was calculated. This study was approved by the institutional Ethics Review Committee of First Affiliated Hospital of Guangzhou Medical College. Informed consent was obtained from the legal guardians of all participants following a detailed description of the purpose and potential benefits of the study. RESULT: There were 175 subjects' induced sputum specimens (175/266, 65.8%), non-qualified sputum samples were obtained from 16 of the subjects. The proportions of median (IQR) of lymphocytes were 0.012 (0.020), 95%CI were ranged from 0.015 to 0.022; neutrophils 0.207 (0.330), 95%CI 0.266 - 0.356 macrophages 0.761 (0.327), 95%CI 0.607 - 0.699; eosinophils 0.004 (0.019), 95%CI 0.013 - 0.022. There were no significant differences in proportions of cytological findings of female or male, different age groups and second-hand smoking or not (all P > 0.05). The incidence of adverse event was 4.40% (7/159). CONCLUSION: The method and the preliminary data may be used for research, diagnosis and treatment of patients with chronic cough and airway inflammation.

[Effects of acute lymphoblastic leukemia children bone marrow mesenchymal stem cells on drug resistance of K562/A02 cell line].

The aim of study was to investigate the effect of acute lymphoblastic leukemia (ALL) children bone marrow mesenchymal stem cells (MSC) on resistance of K562/A02 cells and its mechanism. MSC obtained from bone marrow of AL children were cultured and identified. The co-culture of MSC and K562/A02 and the culture of K562/A02 cell suspension alone was performed, of which 2 kinds of cells were treated with same concentration of adriamycin (ADM), and the rate of apoptosis was detected by flow cytometry, bcl-2 and bax of K562/A02 were detected by RT-PCR, while mdr1 gene level was detected by FQ-PCR. The results indicated that the MSC separation and proliferation were viable and steady. The apoptosis rate of the K562/A02 cells co-cultured with MSC was 1.97 ± 0.11%, while apoptosis rate of the K562/A02 cells cultured alone was 8.38 ± 0.29%, there was significant difference (p < 0.05). As compared with the K562/A02 cells cultured alone, the bcl-2 gene expression in K562/A02 cells co-cultured with MSC obviously increased; ratio of bcl-2/bax was obviously enhanced. The mdr1 gene level in K562/A02 co-cultured with MSC was no statistical different from K562/A02 cultured alone (p > 0.05), which suggested that adhesion co-cultured with MSC did not induce mdr1 expression higher than the culture of suspension. It is concluded that the MSC of ALL children can escape the leukemia cells from proapoptotic effect of drugs, the resistance of K562/A02 to ADM may be involved in enhancement of bcl-2 gene expression of K562/A02 cells co-cultured with MSC, but not in relation to mdr1 gene in K562/A02 cells themselves.

[Preliminary study on the safety and pharmacodynamic action of low dose L-asparaginase].

OBJECTIVE: To investigate the safety and therapeutic effect of low dose (1000 U/m(2)) L-asparaginase (L-Asp) in the treatment of children with acute lymphoblastic leukemia (ALL). METHODS: Six patients were treated with low dose L-Asp after previously suffered severe side effects from standard dose L-Asp (5000 - 10,000 U/m(2)). Twenty-eight blood samples were obtained randomly from 5 of them. Plasma asparagine concentration was detected by reverse phase-high performance liquid chromatography (RP-HPLC). RESULTS: All the patients treated with low dose L-Asp showed no any toxic symptoms. The plasma asparagine levels in the patients were all above 5 micromol/L except case 4 (4.91 micromol/L) before receiving L-Asp, and were all decreased below 0.5 micromol/L five days after receiving low dose L-Asp, except case 3 (3.70 micromol/L), the results being like that of receiving standard dose L-Asp. CONCLUSION: Low dose L-Asp has definite efficacy for childhood ALL, while avoids serious side effects from standard dose L-Asp.