BACKGROUND:Primary vascular endothelial cels are mostly harvested through aorta endothelial cel cultures and micro-artery endothelial cel cultures using enzyme digestion and tissue adhesion methods, and the quality and purity of harvested cels cannot meet the need for current scientific research.
OBJECTIVE:To investigate an improved extraction of primary vascular endothelial cels and the relevant identification method.
METHODS: A segment of rabbit aorta was cut to culture vascular endothelial cels using the improved extraction method in group A or using adhesion method in group B. In the group A, the vascular intima was striped out with microsurgical instruments, and digested enzymaticaly to acquire single primary cels folowed by culture in endothelial cel culture medium. In the group B, the whole vascular intima was adhered to the culture dish that was incubated in a 5%CO2, 37℃ incubator for 1 hour. Cel pelets in the two groups were culturedin vitro. Cel morphology was observed using a microscope; immunohistochemical staining was used to detect CD31, VIII factor and Vimentin protein for identification of vascular endothelial cels.
RESULTS AND CONCLUSION:The purity and number of vascular endothelial cels extracted by the improved method were higher than those by the adhesion method. Immunohistochemical findings showed positive expression of CD31 and VIII factor, but negative expression of Vimentin. These findings indicate that the improved extraction method can obtain more vascular endothelial cels with higher purity, which is of strong operability and practicality.

Objective:To evaluate the nasal safety of gastrodin nasal temperature-sensitive in situ gel through the studies on cilia toxicity in toads and mucosal irritation in rats. Methods:The toads were randomly divided into four groups, saline group, gastrodin in situ gel group, blank gel matrix group and sodium deoxycholate group, and the cilia toxicity was observed in vivo by a toad palate meth-od. The rats were randomly divided into three groups, saline group, gastrodin in situ gel group and blank gel matrix group, and the mucosal irritation was studied in rats through the observation of nasal mucosal pathological changes and behavioral indices. Results:Compared with the saline group, gastrodin in situ and blank gel matrix showed no notable effect on the cilia movement function in toads, and the effect on cilia movement of sodium deoxycholate showed statistically significant difference when compared with that of sa-line, gastrodin in situ gel and blank gel matrix (P<0. 01). During and after the treatment, no sneezing appeared in the rats. Com-pared with that in the saline group, the number of scratching nose in the gastrodin in situ gel group and blank gel matrix group in-creased (P<0. 05) without difference between the groups (P>0. 05), and after the 2-day withdrawal, that in the gastrodin in situ gel group and blank gel matrix group decreased significantly when compared with that at the last administration (P<0. 05) and showed no notable difference when compared with that in the saline group (P>0. 05). The number of inflammatory cells in the nasal mucosa in the gastrodin in situ gel group and blank gel matrix group increased complicated with congestion and cilia falling off, and after the with-drawal, the mucosal morphology in the three groups showed no significant difference. Conclusion:The local application of gastrodin in situ gel has high security, which is valuable to be studied further.

Objective To investigate the methylation status of 14-3-3σ promoter in nasopharyngeal carcinoma cell lines and the influence of de-methylation treatment on 14-3-3σ expression. Methods Methylation status of 14-3-3σ gene promoter and 14-3-3σ mRNA expression were detected by methylation specific PCR (MSP) and RT-PCR in nasopharyngeal carcinoma cell lines CNE1, CNE2,5-8F,6-10B and immortalized nonneoplastic human nasopharyngeal epithelial cell line, NP69. Four nasopharyngeal carcinoma cell lines were treated with 5-asa-2' -deoxycytidine(5-aza-2dC) in different concentration for 72 h, then 14-3-3σ promoter meth-ylation status and m RNA expression were assessed, and western-blot was performed to detect the expression of 14-3-3σ protein. Results 14-3-3σ promoter methylation was detected by MSP in all of the four nasopharyn-geal carcinoma cell lines untreated by 5-aza-2dC whereas not in the treated ones or the immortalized human na-sopharyngeal epithelial cell line, NP69. Accordingly, 14-3-3σ mRNA expression was significantly discounted in untreated nasopharyngeal carcinoma cell lines as compared with NP69. 5-aza-2dC treatment dose-depend-ently reversed 14-3-3σ promoter methylation status and consequently upregulated the expression of 14-3-3σmRNA and protein in 4 nasopharyngeal carcinoma cell lines. High-differentiated CNE1 was more sensitive to 5-aza-2dC than lowly-differentiated CNE2, 5-8F and 6-10B. Conclusion Promoter methylation directly leads to decreased 14-3-3σ gene expression in nasopharyngeal carcinoma cell lines, and 14-3-3σ promoter de-methylation perhaps indicates a new target for nasopharyngeal carcinoma treatment.

Objective To investigate the effect of perioperative pulmonary infection in elderly patients with mild to moderate chronic obstructive pulmonary disease (COPD) undergoing general anesthesia.Methods Forty elderly patients undergoing general anesthesia and abdominal surgery, 24 males, 16 females, aged 65-81 years, ASA physical status Ⅰ-Ⅲ, BMI 19-28 kg/m2, were randomly divided into two groups (n=20 each): protective ventilation group (group PV) and conventional ventilation group (group CV).Lung protective ventilation was received in group PV: intermittent positive pressure ventilation, tidal volume 6 ml/kg (ideal body weight), positive end expiratory pressure (PEEP) 5-10 cm H2O, alveolar recruitment maneuver every 30 minutes;conventional ventilation was received in group CV: intermittent positive pressure ventilation, tidal volume 10 ml/kg (ideal body weight), without using the PEEP and alveolar recruitment maneuver.Venous blood samples for interleukin 6 (IL-6) and interleukin-8 (IL-8) were taken at five different time points: before the anesthesia induction (T1), 2 h after mechanical ventilation (T2), at the end of operation (T3), 6 h (T4) and 24 h (T5) after operation.The clinical pulmonary infection score (CPIS) was recorded at before anesthesia, days 1, 3, 5 and 7 after surgery.The incidence of postoperative pulmonary inflammation was also recorded.Results There was no statistical difference in the two groups with respect to age, body mass index, ASA physical status, intraoperative volume of infusion, estimated blood loss, urine volume, mechanical ventilation time, operation method and IL-6, IL-8 levels at T1-T5.Compared with T1, the IL-6 and IL-8 levels in two groups at T2-T5 increased significantly (P＜0.05).Compared that before anesthesia, CPIS in group CV on postoperative days 1, 3 and 5 increased significantly (P＜0.05);compared with group CV, CPIS and the incidence of postoperative pulmonary inflammation in group PV reduced significantly on postoperative days 1, 3 and 5 (P＜0.05).Conclusion Lung protective ventilation can not reduce perioperative IL-6, IL-8 levels in laparotomy elderly patients with COPD, but it can reduce the incidence of pulmonary inflammation and pulmonary infection within 5 postoperative days.

Objective To screen for new methylation association genes in HL-60 to reveal the pathogenesis of leukemia, and provide important theoretical and scientific basis for the prevention and cure of leukemia. Methods Two-dimensional fluorescence difference gel electrophoresis (F-2D-DIGE) was performed to separate the total proteins from acute myelogenous leukemia (AML) cell line HL-60 cells with or without 5-aza-2-deoxycytidine (5-aza-2-dC) treatment. Imaging software Decyder 6.5 and PDQuest were used to detect the differential expression protein spots, and matrix-assisted laser desorption/ionizaion time-of-flight mas spectrometer (MALDI-TOF MS) was adopted to identify the differential expression proteins. Results F-2D-DIGE maps of 5-aza-2-dC-untreated HL-60 and-treated HL-60 cells were established. A total of 53 differential protein spots were detected, and 35 differential proteins were successfully identified. Of the identified proteins, 32 proteins were up-regulated, and 3 proteins were down-regulated in HL-60 cells after 5-aza-2-dC treatment.Conclusion Thirty-five differential proteins may be associated with methylation in HL-60 cell line, which provides the important clues for epigenetic study of leukemia.

Animals ; Animals, Newborn ; Brain ; blood supply ; metabolism ; pathology ; Carrier Proteins ; genetics ; DNA-(Apurinic or Apyrimidinic Site) Lyase ; genetics ; Gene Expression ; Hypoxia-Ischemia, Brain ; genetics ; pathology ; In Situ Hybridization ; Protein Tyrosine Phosphatase, Non-Receptor Type 13 ; Protein Tyrosine Phosphatases ; genetics ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar

Objective To evaluate the clinical and manometric characteristics of achalasia.Meth-ods Patients diagnosed as having achalasia from July 2010 to July 2014 at our hospital were enrolled.High resolution manometry(HRM)results were analyzed.Data of Eckardt scale,MDADI and SF-36 were ana-lyzed.Results All subjects had dysphagia,35.6% (37 /104)accompanied with regurgitation,26.9%(28 /104)with heartburn and 17.3%(18 /104)with weight loss.According to HRM results and Chicago classification criteria,16.35%(17 /104)of the subjects were classified as type Ⅰ,76.92%(80 /104)as type Ⅱ and 6.73%(7 /104)as type Ⅲ.27.9%(29 /104),19.2%(20 /104)and 24.0%(25 /104)of the subjects finished Eckardt scale,MDADI and SF-36,respectively.Eckardt score was positively correlated with integrated relaxation pressure(IRP)(r =0.421,P <0.05)and MDADI physical score was negatively with IRP(r =-0.530,P <0.05).Conclusion Dysphagia often occurs as the chief complaint among acha-lasia patients.And type Ⅱ is the most common.IRP is an indicator of the severity of clinical symptoms and impairment of quality of life.

Objective To explore the relationship between tumor necrosis factor (TNF) gene polymorphisms in donors and recipients and the incidence and severity of acute graft-versus-host diseases (aGVHD) after unrelated allogeneic hematopoietic stem cell transplantation (alIo-HSCT). Methods Single nucleotide polymorphisms (SNPs) of TNFα-238 (G/A), TNFα-857 (C/T), TNFα-863 (C/A), TNFα-1031 (T/C), TNFβ + 252 (A/G) were analyzed by Multiplex SNaPshot analysis in 76 pairs of donors and recipients. Results Transplantation involving donors with TNFα-857 CC genotype resulted in a higher incidence of grade Ⅱ-Ⅳ aGVHD than donors with CT genotype (91.3% vs 8. 7% , P =0. 039). In the 23 patients with grade Ⅱ-Ⅳ aGVHD, no patients had TNFβ +252 AA genotype, 19 (82.6%) had GA genotype and 4 (17.4%) had GG genotype. There was a significant difference in the distribution pattern of the TNFβ +252 (AA, GA and GG) genotypes in these patients (P =0.03). There was no significant association of TNFα-238 (G/A), TNFα-863 (C/A) and TNFα-1031 (T/C) polymorphisms with the risk of aGVHD. Conclusion These results suggest donor TNFα-857 CC genotype is related to a higher incidence of grade Ⅱ -Ⅳ aGVHD, and patients with TNFβ +252 AA genotype have protection against the risk of grade Ⅱ -Ⅳ aGVHD.

OBJECTIVE: To investigate the significance and relationship between the expression of FOXC1 and clinicopathological features, and to explore its correlation with E-cadherin. METHOD: Immunohistochemical SP method was used to detected the expression of FOXC1 in nasopharyngeal carcinoma tissues and nasopharyngitis tissues. RESULT: (1) Immunoreaction to FOXC1 was mainly located in nucleus of nasopharyngeal carcinoma cells. The positive expression rate of FOXC1 in nasopharyngeal carcinoma tissues was 85.3% (81/95), which was significantly higher than that in nasopharyngitis tissues (59.4%) (P < 0.05). (2) The expression of FOXC1 was not related to patients' age and gender, clinical stage of cancer and lymph node metastasis (P > 0.05). (3) There was a correlation between the expression of FOXC1 and down-regulated expression of E-cadherin in nasopharyngeal carcinoma tissues (P < 0.05). CONCLUSION FOXC1 may play an important role in generation and progression of nasopharyngeal carcinoma, there may be a correlation between the expression of FOXC1 and down-regulated expression of E-cadherin, also FOXC1 may play an important role in the process of EMT in nasopharyngeal carcinoma by regulating E-cadherin.
Adolescent ; Adult ; Aged ; Antigens, CD ; Cadherins ; metabolism ; Carcinoma ; Female ; Forkhead Transcription Factors ; metabolism ; Humans ; Male ; Middle Aged ; Nasopharyngeal Carcinoma ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Nasopharyngitis ; metabolism ; Young Adult

AIMTo study the protective role of endogenous carbon monoxide to lung and kidney tissues during septic shock and its mechanism.

METHODSA rat model of CLP was built by using the method of CLP. The malondialdehyde (MDA) content and the activity of superoxide dematase (SOD) in blood, lung and kidney were detected by immunohistochemical technique and light microscope.

RESULTSPathological changes of lung and kidney in CLP + Hemin group were lighter than CLP group, inflammatory reaction and lipid peroxidation were also lighter.

CONCLUSIONEndogenous CO can protect lung and kidney from the oxidative injury. It can suppress in flammation and the oxidative injury caused by activated inflammatory cells, it is probably an important mechanism of its protective effects.

Animals ; Carbon Monoxide ; physiology ; Hemin ; pharmacology ; Kidney ; metabolism ; pathology ; Lipid Peroxidation ; Lung ; metabolism ; pathology ; Male ; Malondialdehyde ; analysis ; Rats ; Rats, Sprague-Dawley ; Shock, Septic ; metabolism ; pathology ; Superoxide Dismutase ; metabolism