Objective To explore the method of primary isolation, cultivation and identification of rat brain microvessel pericytes. Methods The brain tissue of 10 3 week-old Wistar rats was separated sterilely. The brain microvessel fragments were separated using two-step enzyme digestion and one-step gradient centrifugation and were seeded in 35-mm dishes for primary culture. The cell morphology was observed by phase contrast microscopy; the immunofluorescence assay was used to identify the associated antigns, such as the α-smooth muscle actin (α-SMA), neuron-glial antigen 2 (NG2), von Willebrand factor (vWF), and glial fibrillary acidic protein (GFAP). Methyl thiazolyl tetrazolium was used to determine the cell growth curve. Results Pericytes climbed out from the adherent brain microvascular fragments around,showing polygonal, and the cell fusion was 95％ after 12-14 days. Immunofluorescence staining revealed that the molecular markers of the pericytes α-SMA and NG2 related antigens showed double positive, while the vWF and GFAP related antigens showed double negative and the cultured cells were confirmed as brain microvascular pericytes. The growth rate of primary cells was slower. The passage cells entered into logarithmic growth phase after 36 to 60 hours and entered into plateau phase after 72 to 108 hours. Conclusions This method may successfully isolate rat brain microvascular pericytes with higher purity.

Objective To establish a stable in vitro model of blood-brain barrier (BBB) simulating in vivo state using the primary-cultured rat brain microvascular endothelial cells (BMVECs) and pericytes.Methods The primary rat BMVECs and pericytes were isolated,purified and cultured.The isolated cells were identified by immunocytochemical staining method.An in vitro model of BBB was constructed using Transwell inserts (pore size 0.4 μm) coculture.Its barrier function was evaluated by the 4-hour leakage test,tight junction protein identification,transendothelial resistance detection,and permeability test.The difference between the cocultured model and simple BMVEC model across the membrane resistance values,and the permeability difference of the small molecule sodium fluorescein (Na-F) were compared.Results Confluent BMVEC monolayers demonstrated a typical cobblestone appearance and the pericytes displayed irregular shape and overlapping growth.Immunodouble labeling technique identification showed that the pericytes positively expressed α-srmooth muscle actin (α-SMA) and neuron-glial antigen 2 (NG2); after the fusion of cocultured model endothelial cells,the surface leakage test became positive; immnocytochemical staining shows that a continuous and dense tight junction formed between the endothelial cells; compared to the BMVEC model,the transendothelial electrical resistance of the cocultured model increased significantly (190.762 ± 10.326 Ω/cm2 vs.96.503 ± 8.012 Ω/cm2; t=- 24.489,P ＜0.01),and the permeability decreased significantly (56.149％ ± 3.572％ of the single endothelial model; t =19.330,P ＜ 0.01 ).Conclusions The primary isolated rat BMVECs and pericytes cocultured the morphology,structure and barrier function of in vitro model have the basic characteristics of BBB,and they have provided a useful tool for the research of BBB.

Objective To explore the protective effects of quercetin on damage induced by oxidative stress and to clari?fy its molecular mechanism. Methods Chang liver cell cultures were randomly divided into control groups, H2O2 group and 3 doses of quercetin groups. Cell survival rate was detected with MTT. Cell apoptotic rate was measured by FACS(Fluores?cence-activated cell sorting). Intracellular reactive oxygen species (ROS) level in Chang liver cells were tested by flow cy?tometer. The DCF fluorescence intensity of DCFH-DA-stained intracellular ROS was observed by fluorescence microscope. The levels of malondialdehyde (MDA), superoxide dismutase(SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) were determined in liver cells using commercial available kits. The expression of Nrf2 were detected by Western blot. Re?sults Compared with control, cell survival rate and levels of SOD, CAT and GSH-Px decreased significantly in H2O2 group (P < 0.05 ),while cell appotosis rate, content of MDA and mean fluorescence intensity(MFI) increased in H2O2 group (P <0.05). In comparison with H2O2, expression of Nrf2 protein was higher in all three quercetin treatment groups (P<0.05). Con?clusion Quercetin protected Chang liver cells from H2O2-induced oxidative stress, which may be caused by the increased ex?pressions of down stream antioxidant genes via activating the Nrf2-ARE signaling pathway.

Objective To study the distribution of surfactant protein A2 (SP-A2) haplotype and its association with preterm respiratory distress syndrome (RDS) susceptibility in a local Chinese cohort.Methods Using population base and case-control study cohorts,genotyping for four single nucleotide polymorphism (SNP) was performed,rs1059046,rs17886395,rs1965707,rs1965708,in 80 term infants,50 preterm infants with RDS (RDS group) and 50 preterm infants without RDS(control group) by using TaqMan (R) real-time polymerase chain reaction.Infants in RDS group and control group were matched according to gender and gestational age.Frequency of each haplotype was compared between preterm infants with RDS and without RDS,term infants and preterm infants without RDS.Results Most common haplotypes in term infants were 1A0,1A5,1A1.In each preterm infants groups with RDS and without RDS,1A0,1 A5,1 A1 were also the most common haplotypes.Among these three common haplotypes,frequency of 1A0 was lower in preterm infants,including RDS group and control group,than that in term infants.No significant difference was found between preterm groups with RDS and without RDS (P ＞ 0.05),neither in preterm infants with gestational age ≥32 or ＜ 32 weeks.Haplotype 1A0 frequency(0.542) in term infants was significantly higher than that in preterm infants ＜ 32 weeks without RDS (0.329) (x2 =6.06,P =0.01).Conclusions SP-A2 haplotype distribution in a local Chinese population shows ethnic characteristics to some extent.Only SP-A2 does not contribute to the susceptibility for preterm RDS.

OBJECTIVETo determine the protective effect of pomegranate polyphenols on cardiac function in rats with myocardial ischemia/reperfusion (I/R) injury and explore the possible mechanism.

METHODSFifty SD rats were randomized into 5 equal groups, including a sham-operated group, an I/R model group, and 3 pomegranate polyphenol dose groups (150, 300, and 600 mg/kg). The rats were subject to a 45-min left main coronary artery occlusion followed by a 180-min reperfusion to induce myocardial I/R injury except for the rats those in the sham-operated group. The cardiac functions were monitored continuously during the experiment. At the end of the reperfusion, arterial blood samples were obtained to measure plasma contents of lactate dehydrogenase (LDH), creatine kinase (CK), malondialdehyde (MDA) and the activities of superoxide dismutase (SOD). Triphenyltetrazolium chloride (TTC) staining was used to evaluate the infarct size.

RESULTSCompared with the sham-operated group, all the rats with I/R showed significantly decreased left ventricular systolic pressure (LVSP), maximal rates of increase/decrease of the left ventricular pressure (±dp/dtmax) (P<0.01) and significantly increased left ventricular end-diastolic pressure (LVEDP) (P<0.01). Compared with the I/R model group, all the 3 pomegranate polyphenol groups had significantly improved cardiac function (P<0.05), decreased plasma contents of CK, LDH and MDA (P<0.01), increased SOD activities (P<0.01), and obviously reduced infarct size (P<0.01).

CONCLUSIONPomegranate polyphenols can protect the cardiac function of rats with I/R injury probably in association with their actions in enhancing oxygen free radical scavenging activity and decreasing lipid peroxidative damage of the myocardial tissues.

Animals ; Creatine Kinase ; blood ; Heart ; drug effects ; L-Lactate Dehydrogenase ; blood ; Lipid Peroxidation ; Male ; Malondialdehyde ; blood ; Myocardial Reperfusion Injury ; metabolism ; physiopathology ; Myocardium ; metabolism ; Oxidative Stress ; drug effects ; Polyphenols ; pharmacology ; Punicaceae ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species ; metabolism ; Superoxide Dismutase ; metabolism

50% of that before treatment; (3)Normal control group( n =10). The urinary retinol binding protein(RBP) and N acetal D amino glucosidase(NAG)were measured by ELISA. The urinary osmosis, 24 h urinary protein excretion and serum creatinine (Cr) were measured. Results: (1) The urinary RBP[(0.54?0.19) mg/L], NAG[(112.84?42.82) U/L] and osmosis [(553.62?248.91) mmol/L] in PNS patients were significantly higher than normal control group ( P

OBJECTIVEWe studied the licorice provenance rule under the manual simulative water stress conditions, discussed geographical variation pattern and the ecology mechanism and laid the theoretical basis for the licorice seed regionalization and the seed allocation.

METHODOn the spot we investigated and collected seed materials. Setting up water potential gradient by PEG, we observed the seed germination characteristic through the experiment of indoor germination.

RESULTThe sensitivity of various provenance seed germinative energy show quite remarkable difference for water potential reducing. Along with geographic longitude increasing, the seed germinates variation sensitivity increasing gradually to the water stress. The results of climatic factors' correlation analysis indicated that provenance seed germinate energy to the water stress is sensitive, in which the annual average temperature is low and annual precipitation is abundant. But it is insensitive in which the annual precipitation is little, the annual average temperature, average temperature in July and the annual average ground temperature is high.

CONCLUSIONUnder the water stress condition the licorice provenance seed germination characteristic exists remarkable geographical variation. Geographical variation is the result of natural selection.

China ; Dehydration ; Ecosystem ; Geography ; Germination ; Glycyrrhiza uralensis ; physiology ; Seeds ; physiology

In this article, human umbilical vein endothelial cells(HUVECs)were induced by lipopolysaccharides(LPS)to establish an in vitro inflammation model to further verify the anti-inflammation effects of benserazide hydrochloride and to explore the molecular mechanisms involving in the anti-inflammation and anti-atherosclerosis of benserazide hydrochloride. The experiments were divided into blank groups(PBS+0. 5% FBS DMEM medium), model group ［LPS(500 μg/mL)+ 0. 5% FBS DMEM medium］ and drug group ［LPS(500 μg/mL)+benserazide hydrochloride+0. 5% FBS DMEM medium］. Western blot, ELISA and qPCR were used to detect the protein and mRNA expression levels of inflammatory cytokines SAP, TNF-α and MCP-1 in HUVECs cells. The expression levels of p65/p-p65, p38/p-p38, IκBα/p-IκBα, AKT/p-AKT and the nuclear translocations of p65, p38 and IκBα were detected by Western blot. The results showed that benserazide hydrochloride(1×10-9, 1×10-10, 1×10-11 mol/L)could significantly inhibit the protein and mRNA expression of pro-inflammatory cytokines SAP, TNF-α and MCP-1. Besides, it could down-regulate the protein expression of p65/p-p65, p38/p-p38, IκBα/p-IκBα and AKT/p-AKT in the signal pathway while inhibiting the nuclear translocation of p65, p38 and IκBα, thereby inhibiting the transcriptional activity of the related genes.

Objective To study the influencing factors of clinical outcome in elderly acute ischemic stroke (AIS) patients after intravenous thrombolysis.Methods One hundred and fifty-one AIS patients admitted to our hospital for intravenous thrombolysis were divided into good outcome group (n=77) and poor outcome group (n=74) according to their modified Rankin scale score 3 months after the onset of AIS.The baseline data,thrombolysis time window,NIHSS score and ischemic stroke typing before thrombolysis,early symptom improvement,cerebral hemorrhage after thrombolysis were compared between the two groups.Results The serum levels of blood glucose and CRP,NIHSS score≥9 before thrombolysis,incidence of AF and cerebral hemorrhage were significantly higher in poor outcome group than in good outcome group (P＜ 0.05,P＜0.01).Multivariate logistic regression analysis showed that NIHSS score,OCSP typing,blood glucose before thrombolysis,24 h symptom improvement were the independent influencing factors of clinical outcome in elderly AIS patients (OR =1.262,95 ％ CI:1.075-1.482,P =0.005;OR =0.203,95％CI:0.066-0.628,P=0.006;OR=1.264,95％CI:1.042-1.532,P=0.017;OR=25.764,95％CI:5.131-129.361,P=0.000).Conclusion NIHSS score,OCSP typing,blood glucose before thrombolysis and 24 h symptom improvement are the independent influencing factors of clinical outcome in elderly AIS patients after intravenous thrombolysis.

Objective To investigate the dynamic changes of symptom clusters among nasopharyngeal carcinoma patients receiving radiotherapy and provide evidence for intervention clinically.Methods The data were collected at three time points that were before radiotherapy,during the radiotherapy and after radiotherapy, by surveying 273 first visit of nasopharyngeal carcinoma patients from May 201 4 to January 201 5 with a self-designed general information questionnaire and the Chinese version of the M.D.Anderson symptom inventory and observing the changes of symptom clusters in our hospital.One-way analysis of variance and the exploratory factor analysis were applied to observe the incidence of symptoms and the changes of symptom clusters during radiotherapy.Results The statistics revealed that the incidence and intensity of all the symptoms at 3 time points were significantly increased(P <0.05),except the symptom of amnesia.There were 3 symptom clusters among 3 time points,named as gastrointestinal symptom cluster,sickness symptom cluster and somatic symptom cluster,but the composition and the variance were different at different time point.Gastrointestinal symptom cluster was made of nausea and vomiting,adding poor appetite only before radiotherapy.The gastrointestinal symptom cluster was most prominent before radiotherapy.Sickness symptom cluster consisted of drowsiness, numbness symptom before radiotherapy, while it consisted of shortness of breath, numbness, difficulty remembering symptoms in other time.Somatic symptom cluster mainly consisted of pain and fatigue,dry mouth symptom joining it during and after radiotherapy.Conclusions With the treatment of radiotherapy,the incidence and intensity of symptoms are significantly enhanced,and the constitution of symptom clusters is different at different stage.Therefore,it is important for clinical nurses to strengthen dynamic assessment of all symptoms,monitor the changes and composition of symptom clusters,thus establish corresponding management measures to improve patients′quality of life.