Animals ; Cardiovascular Diseases ; Cardiovascular System ; Humans ; MicroRNAs

Objective To study the chemical components from the pericarps of Juglans mandshurica with anti-tumor activity.Methods It showed that pericarps of J.mandshurica had a good performance in treating cancer and low toxicity from CHCl3 and EtOAc portions by a lot of pharmacological experiments and modern clinical application in the past.Compounds were isolated by chromatography on silica gel columns and AB-8 macroporous resin.On the basis of spectral evidences including 1D-NMR and 2D-NMR,the structures of these compounds were determined.Results These compounds from CHCl3 portion were identified as 20(S)-protopanaxadiol-3-one(1),dammara-20,24-dien-3?-ol(2),galeon(3),juglanin A(4),2?,3?,23-trihydroxy olean-12-en-28-oic acid(5),2?,3?,23-trihydroxy urs-12-en-28-oic acid(6),oleanolic acid(7),ursolic acid(8).Conclusion Compound 1 is a new natural product.Compounds 2 and 5-8 are isolated firstly from the plants of Juglans L.which are due to pentacyclic triterpane.Compound 3 is isolated from the pericarps of J.mandshurica for the first time.

Objective To explore the efficiency and safety of sodium valproate intravenous therapy on refractory status epilepsy(RSE)by diazepam in Chinese adult.Methods Twenty-five patients of RSE,consecutively admitted to West China Hospital of Sichuan University from September 1999 to May 2007,received intravenous administration of sodium valproate after being refractory to intravenous diazepam and intramuscular phenobarbitone.The efficiency and side effect of sodium valproate were then evaluated.There were ten males and fifteen females with mean age of fifty-five years.The symptoms of status epilepticus(SE)included generalized tonic-clonic status epilepticus(GTCS)(14 cases),complex partial status epilepticus(CPS)(7 cases),simple partial status epilepticus(SPS)(2 cases)and myoclonic status epielepticus(MS)(2 cases).The primary diseases prior to SE were viral encephalitis(13 cases),withdrawal of antiepilepsy drug(AEDs)(5 cases),cerebral trauma and brain surgery(3 cases),cerebral infarction(2 cases),intoxication(1 case)and cerebral palsy(1 case).Among them nine patients had a history of epilepsy.Results Of all the 25 cases of RSE,twenty-two were controlled with sodium valproate within one hour,and consciousness recovered within next one hour.No significant changes were found in blood pressure,heart rate,respiratory rate and cardiac rhythm,and no encephalopathy occurred.On the other hand,the other three cases who failed to be controlled,frequency and duration of seizure were redured.Conclusions Although intravenous sodium valproate has not been approved to be the drug of first choice for SE therapy,but it is non-sedative anti-convulsion drug that can be safely used to stop different kinds of diazepam resistant SE efficiently without obvious side effect.In addition,a quick recovery of patients' consciousness may be expected within an hour afte RSE being controlled.

Objective To explore the influence of alimentary tract reconstruction on blood glucose level in gastric cancer patients complicated with type 2 diabetes mellitus. Methods Clinical data of 67 gastric cancer patients complicated with type 2 diabetes mellitus and the level of blood glucose before operation, one month, three month, six month after operation were retrospectively analyzed. BMI of 53 patients was lower than 25kg/m2,9 patients between 25～29.9 kg/m2 and 5 patients was higher than 30kg/m2 .Total gastrectomy with P-type jejunal pouch Roux-en-Y esophagojejunostomy was performed in 26 cases, and total gastrectomy with Roux-en-Y esophagojejunostomy was performed in 11 cases, distal subtotal gastrectomy with Roux-en-Y gastrojejunostomy in 30 cases. Results Operations on sixty-seven patients were all uneventfully. The mean fasting blood glucose level in the morning was 9.6±3.3 mmol/L before operation, 7.4±2.6 mmol/L one month after operation, 7.5±2.3 mmol/L three month after operation, and 7.7±2.9 mmol/L six month after operation. There were significant differences between the blood glucose level of before operation and one month after operation (P<0.05), and there were no significant differences between three month,six month and one month after operation (P>0.05). Conclusions Alimentary tract reconstruction has obviousinfluence on blood glucose level in type 2 diabetes mellitus patients. It takes about one month for reveal the effect of operation. This phenomenon is of value for clinical application. Its mechanism needs further research.

Objective To explore the effects of murinecytomegalovirus(MCMV)infected mouse embryonic fibroblasts(MEF)on the proliferation and activation of regulatory T cell in vitro. Methods A co-culture system of T cell and MCMV infected MEF(T-MEF MCMV)was established.The viral load of supernatant was determined by plaque assay.The proliferation of T cells was observed with cell counting.The proportion of CD4+ CD25+ cells was measured by flow cytometry.The levels of Foxp3 protein were measured by Western blot.Reverse transcription polymerase chain reaction(RT-PCR)assay was utilized tO determine whether MCMV infection of MEF influenced the level of transforming growth factor(TGF)-β mRNA.The level of TGF-β protein in supernatant was measured by double-antibody sandwich enzyme linked immunosorbent assay(ELISA).The difference between T-MEF MCMV group and control group was assessed by one-way ANOVA.Results When T cells were co-cultured with MCMV infected MEF for 1 day and 3 days,the viral load in supernatant decreased.But when co-culture lasted for 6 days,the antiviral effect obviously diminished,as the viral load[(5.58±0.67)×105 PFU/mL]of the experimental group showed no statistic difference with MEF MCMV control group[(6.05±0.34)×105PFU/mL].When co-cultured with MCMV infected MEF for 3 days,T cell increased from pre-culture level of[(2.02±0.05)×106/mL]to(2.25± 0.13)×106/mL(P＜0.05).But when co-culture lasted for 6 days,the number of T eelI returned to (2.08±0.14)×106/mL,which had no statistic difference with that of co-culture for 3 days system. Both the expressions of Foxp3 protein and the proportion of CD4+ CD25+ Foxp3+ Treg cells in T-MEF MCMV group were up-regulated as the infection time extended,which were two times and three times of the control group level,respectively.The mRNA level(A value)of TGF-β in MCMV infected MEF increased from baseline of 1.09±0.13 to 3.15±0.54 on day 3 after infection.The expression of TGF-β in supernatant 3 days after infection was(3.85±0.32) μg/L,which was significantly higher than that before infection[(1.74±0.14)μg/L,P＜0.05].Conclusions Activated T cells have antiviral effect.However,the function of T cells is rapidly inhibited after activation,which may be due to the expression of Foxp3 mRNA induced by MCMV infected MEF and increased CD4+ CD25+ Treg proportion of the co-cultured T cells.TGF-β level is significantly increased after CMV infection,which may be an important mechanism of Treg proliferation.MCMV may manipulate Treg to evade specific immune elimination and,as a result,to cause CMV replication.

6 cases(2 males to 4 females) who have received mandibula reconstruction with free vascularized fibular flaps in our hospital were studied retrospectively.The average length of the fibula grafts was 8.0 cm(range from 7.0 to 14 cm). 3 cases received primary insertion of osteointegrated dental implants into the free fibula flap. The mean follow-up of six cases of patients was 3 months to 1 year. The success rate was 100%. All patients spoke clearly. 4 patients could eat normally, the other two could eat soft food. 5/6 of patients with the recovery of facial appearance was satisfied. All patients were able to walk normally. No ankle unstability was reported. Vascularized free fibula flap is confirmed to be one of the optimal methods for mandible reconstruction by our study.

AIM: To explore the effect of bone marrow-derived mesenchymal stem cells (BMSCs) combined with vitamin E on the inflammatory reaction in acute kidney injury ( AKI) rats.METHODS:Gentamicin was used to in-duce AKI and the rats were treated with BMSCs combined with vitamin E.After treatment, the rat plasma and kidney tis-sues were collected, and the expression of inflammatory factors at mRNA and protein levels was detected by real-time quan-titative PCR and ELISA.RESULTS:After the treatment with BMSCs combined with vitamin E, the inflammatory proteins were down-regulated in the plasma and the renal tissues.Compared with single treatment group, the decreases in the in-flammatory proteins were more obvious in combined treatment group.CONCLUSION: The method of BMSCs combined with vitamin E takes the anti-inflammatory effect on AKI, indicating a new and potential mode in clinical application for AKI therapy.

AIM: To investigate whether efficient production of neuron-like cells from embryonic stem cells(ESCs) can be indued by astrocyte-conditioned medium(ACM) in vitro.METHODS: Based on the 4-/4+ protocol established by Bain,two groups were studied: ATRA group,ATRA with ACM group.ESCs were induced into neuron-like cells by means of three-step differentiation in vitro.The totipotency of ESCs was identified by the observation of cells' morphology and the formations of teratoma in immunocomprised mice.The cell differentiation was evaluated continuously by detection of the cellular specific markers of neural stem cell, neurons and astrocytes such as nestin,NF-200,NSE and GFAP using immuno-histochemistry assay.RESULTS:(1) The ESC-D3 cells kept the ability of differentiation into cellular derivations of all three primary germ layers after continuous passage culture.(2) The ratio of NF-200 and NSE positive cells in the cells induced by ATRA with ACM was higher than that in the cells induced by ATRA only.(3) Finally,the positive rate of the neuron-like cells was up to 73.5% in the group induced by ATRA with ACM.CONCLUSION: The ESCs are induced into neuron-like cells with high purity and efficiency by ATRA with ACM.

Background Interleukin-6 (IL-6) is a pleiotropic cytokine involving in inflammation and wound healing.Previous report found that IL-6 increases phosphorylated STAT3 (p-STAT3) level and promotes corneal epithelial wound healing by stimulating migration.However,the essential role of IL-6 in corneal epithelial wound healing and the expression changes in diabetic mellitus remains unknown.Objective This study was to explore the roles of IL-6 in corneal epithelial proliferation and wound healing in both normal and diabetic mice.Methods Fifty-two normal C57BL/6 mice were randomized into normal control group (32 mice) and diabetic group (20 mice).Type 1 diabetic mellitus was induced by intraperitoneal injections of 50 mg/kg streptozotocin once per day for consecutive 5 days in the mice of the diabetic group.Whole corneal epithelium was scraped in all mice,and the corneal epithelial defect area was examined by fluorescein staining in 24,48 and 72 hours after corneal epithelium removal.Recombinant mouse IL-6 or anti-IL-6 blocking antibody of 5 μl were subconjunctivally injected according to the grouping and contrasted with PBS injection group or isotype control antibody group,respectively.TKE2 cells,a mouse corneal epithelial stem/progenitor cell line,were trypsinized and incubated in the KSFM with different concentrations of IL-6 or without IL-6,and colony formation efficency (CFE) was examined by crystal violet staining.The expressions of △NP63 and Ki67,specific makers of stem cells,were detected by immunofluorescine technology.The expressions of △NP63,Ki67 and p-STAT3 proteins were assayed in the cells by Western blot,respectively.The expression of IL-6 mRNA and protein in the regenerated corneal epithelium was detected by real time quantitative PCR and ELISA.The use and care of the mice complied with the Statement of Association for Research in Vision and Ophthalmology.Results The percentage of residual corneal epithelium defect area with initial detect area was gradually shrinked over time after PBS and IL-6 injection in both normal control mice and diabetic mice,and the percentage of residual corneal epithelium defect area was significantly reduced in the IL-6 injected group compared with the PBS injected group (normal control group:Fgroup =19.982,P ＜ 0.01;Ftime =589.350,P ＜ 0.01;Diabetic group:Fgroup =25.411,P＜0.01;Ftime =334.807,P＜0.01).The CFE was (13.23± 1.12)％,(15.87± 1.30)％,(21.69±1.62)％,(25.33±1.28)％ and (18.67±1.54)％ in the blank control group and 10,20,50,100 ng/ml IL-6-treated groups,respectively,showing a gradual increase of CFE dependent upon IL-6 concetrations (F =35.547,P＜0.01).The expressions of △NP63,Ki67,p-STAT3 proteins in the cells were gradually increased over time after 50 ng/ml IL-6 treated for 5,10,15,30 and 60 minutes,and the relative expression level of the cytokines was significnatly higher in the IL-6 cultured groups than that without IL-6 culture group (all at P＜0.05).The relative expression of IL-6 mRNA in the regenerated corneal epithelilum was 0.45±0.21 and 1.00±0.16 in the diabetic group and normal control group,respectively,and compared with the normal control group,the expression of IL-6 mRNA reduced by 56％ (t=3.42,P=0.03).The content of IL-6 protein in regenerated corneal epithelium of the diabetic group was (257±12) ng/μl,which was significantly lower than (323 ± 17) ng/μl of the normal control group (t =5.60,P＜0.01).Conclusions IL-6 promotes the proliferation and regeneration of corneal limbal stem cells to repair defected corneal epithelium by activating STAT3 signaling pathway in both normal and diabetic mice,while the blocking of endogenous IL-6 impairs the corneal epithelial cell activation and wound healing.

Objective To detect the mutations of Von Hippel-Lindau (VHL) gene via analyzing the prevalence of family members of VHL syndrome,clinical diagnosis and treatment,and gene analysis of patients with hemangioblastoma.methods All members of the VHL syndrome family members improved all relevant tests and plotted the family map.5 ml peripheral blood was extracted for gene sequencing,and the sequencing Result s were compared with the reported mutations of VHL gene in NCBI database.Result s(1)Analysis of clinical data of four members of the family:Ⅰ-2,Ⅱ-1,Ⅱ-5 suffering from central nervous system hemangioblastoma, Ⅱ-3 with pancreatic,retinopathy and pheochromocytoma,and Ⅱ-5 also combined with kidney,pancreatic lesions.The second generation of patients in the family have been treated surgically.(2)Gene sequencing Result s showed that all subjects in the test had the same mutation:exon2 109 sequence ATATCACACTGCCA was deleted and termination codon UGA appeared in exon 502.Conclusion Through the mutations of the VHL syndrome family,it is found that the family mutation type is a new mutation.For patients with central nervous system hemangioblastoma-based should be suspected of the disease and improve the family history survey.Once the diagnosis of familial VHL syndrome patients are confirmed,it is necessary to inform the other members of the family for clinical screening,and carry out genetic testing to reduce the harm of the disease to the greatest extent.