晚稻收割期与大麦播种期联因试验总结

我市在年积温5300℃等温线以北的绍虞平原稻区,三熟制连作晚稻种植以晚粳类型占优势,冬作则以大麦为主。由于晚粳稻收割适期不明确,所以收获时间较长,相应地推迟和延长了后作大麦的播种期,造成大麦产量不稳不高,从而阻碍了全年三熟产量进一步提高。为明确前作晚粳稻收割与后作大麦播种双佳适期,我们进行了晚粳稻收割期与大麦播种期联因试验。现将结果报道如下,供生产上参考。试验条件与设计试验设在本所麦、稻、稻三熟制田块。

Clinical significance of serum expression of GROβ in esophageal squamous cell carcinoma

AIM:To determine the association between serum levels of growth-related gene product β(GROβ) and clinical parameters in esophageal squamous cell carcinoma(ESCC).METHODS:Using enzyme-linked immunosorbent assay,serum GROβ levels were measured in ESCC patients(n = 72) and healthy volunteers(n = 83).The association between serum levels of GROβ and clinical parameters of ESCC was analyzed statistically.RESULTS:The serum GROβ levels were much higher in ESCC patients than in healthy controls(median:645 ng/L vs 269 ng/L,P < 0.05).Serum GROβ levels were correlated positively with tumor size,lymph node metastasis,and tumor-node-metastasis(TNM) staging,but not with gender or the histological grade of tumors in ESCC patients.The sensitivity and specificity of the assay for serum GROβ were 73.61% and 56.63%,respectively.CONCLUSION:GROβ may function as an oncogene product and contribute to tumorigenesis and metastasis of ESCC.

Cardioprotective effects of Guanxinshutong （GXST） against myocardial ischemia/ reperfusion injury in rats

Background The protective effects against reperfusion injury of cardioprotective drugs have recently been evaluated and found to be inadequate. Guanxinshutong （GXST）, a combination of the traditional herb and Mongolian medicine, is effective and safe in treating angina pectoris in clinical trials. We assess the cardioprotective effects of GXST against myocardial ischemia and reperfusion （MI/R） injury in rats and explore its possible mechanism. Methods Forty-five male Sprague Dawley rats were randomized into three groups： non-MlfR group （Sham, n = 15）, MUR group treated with vehicle （Control, n = 15） and MI/R group treated with GXST （Drug, n = 15）. MI/R was induced by ligation of the left anterior descending coronary artery （LAD） for 30 minutes, followed by 2/24 hour reperfusion in the Control and Drug groups. In the Sham group, the LAD was exposed without occlusion. GXST powder （in the Drug group） or saline （in the Control and Sham groups） were administered via direct gastric gavage from 7 day prior to surgery. Blood samples were collected from the carotid artery （10 rats each group） after 2 hours of reperfusion, to determine the levels of tumor necrosis factor-or （TNF-ct）, interleukin-1 ~ （IL-113）, interleukin-6 （IL-6） and intercellular adhesion molecule-1 （ICAM-1） using enzyme-linked immunosorbent assays. The animals were then sacrificed and the hearts were harvested for histopathology and western blot analysis. Infarct size was measured in the remaining five rats in each group after 24 hours reperfusion. Results GXST significantly decreased levels of TNF-ct, IL-1β, IL-6, ICAM-1, apoptosis index （AI） and infarct size. GXST also obviously inhibited nuclear factor kappa B （NF.r,B） activity when compared with the Control group （all P 〈 0.05）. Conclusions GXST is effective in protecting the myocardium against MI/R injury in rats. Its possible cardioprotective mechanism involves inhibition of the inflammatory response and apoptosis following MI/R injury.

Gastric cancer cells induce human CD4^+ Foxp3^+ regulatory T cells through the production of TGF-β1

AIM: To elucidate the molecular and cellular features responsible for the increase of regulatory T cells (Tregs) in gastric cancer. METHODS: The frequencies of CD4 + Foxp3 + Tregs and the level of transforming growth factor-β1 (TGF-β1) were analyzed from 56 patients with gastric cancer byflow cytometry and enzyme-linked immunosorbent assay respectively. Foxp3 gene expression was analyzed by real-time polymerase chain reaction. The gastric cancer microenvironment was modeled by establishing the coculture of gastric cancer cell line, MGC-803, with sorting CD4 + T cells. The normal gastric mucosa cell line, GES-1, was used as the control. The production of TGF-β1 was detected in supernatant of MGC and GES-1. The carboxyfluorescein diacetatesuccinimidyl ester (CFSE) dilution assay was performed to evaluate the proliferation characteristics of induced Tregs. Neutralizing anti-TGF-β1 antibody was added to the co-culture system for neutralization experiments. RESULTS: The level of serum TGF-β1 in gastric cancer patients (15.1 ± 5.5 ng/mL) was significantly higher than that of the genderand age-matched healthy controls (10.3 ± 3.4 ng/mL) (P < 0.05). Furthermore, the higher TGF-β1 level correlated with the increased population of CD4 + Foxp3 + Tregs in advanced gastric cancer (r = 0.576, P < 0.05). A significant higher frequency of CD4 + Foxp3 + Tregs was observed in PBMCs cultured with the supernatant of MGC than GES-1 (10.6% ± 0.6% vs 8.7% ± 0.7%, P < 0.05). Moreover, using the purified CD4 + CD25 T cells, we confirmed that the increased Tregs were mainly induced from the conversation of CD4 + CD25 naive T cells, and induced Tregs were functional and able to suppress the proliferation of effector T cells. Finally, we demonstrated that gastric cancer cells induced the increased CD4 + Foxp3 + Tregs via producing TGF-β1. Gastric cancer cells upregulated the production of TGF-β1 and blockade of TGF-β1 partly abrogated Tregs phenotype. CONCLUSION: Gastric cancer cell can induce Tregs development via producing TGF-β1, by which the existence of cross-talk between the tumor and immune cells might regulate anti-tumor immune responses.

Competition between TRAF2 and TRAF6 Regulates NF-κB Activation in Human B Lymphocytes

Objective To investigate the role of TNF receptor-associated factor 2 (TRAF-2) and TRAF6 in CD40-induced nuclear factor-κB (NF-κB) signaling pathway and whether CD40 signaling requires TRAF2. Methods Human B cell lines were transfected with plasmids expressing wild type TRAF2 or dominant negative TRAF2,TRAF2-shRNA,or TRAF6-shRNA. The activation of NF-κB was detected by Western blot,kinase assay,transfactor enzyme-linked immunosorbent assay (ELISA),and fluorescence resonance energy transfer (FRET). Analysis of the role of TRAF-2 and TRAF-6 in CD40-mediated NF-κB activity was examined following stimulation with recombinant CD154. Results TRAF2 induced activity of IκB-kinases (IKKα,IKKi/ε),phosphorylation of IκBα,as well as nuclear translocation and phosphorylation of p65/RelA. In contrast,TRAF6 strongly induced NF-κB activation and nuclear translocation of p65 as well as p50 and c-Rel. Engagement of CD154-induced nuclear translocation of p65 was inhibited by a TRAF6-shRNA,but conversely was enhanced by a TRAF2-shRNA. Examination of direct interactions between CD40 and TRAFs by FRET documented that both TRAF2 and TRAF6 directly interacted with CD40. However,the two TRAFs competed for CD40 binding. Conclusions These results indicate that TRAF2 can signal in human B cells,but it is not essential for CD40-mediated NF-κB activation. Moreover,TRAF2 can compete with TRAF6 for CD40 binding,and thereby limit the capacity of CD40 engagement to induce NF-κB activation.

Effect of Helicobacter pylori cdrA on interleukin-8 secretions and nuclear factor kappa B activation

AIM： To investigate genetic diversity of Helicobacter pylori （H. pylorl） cell division-related gene A （cdrA） and its effect on the host response.METHODS： Inactivation of H. py/ori cdrA, which is involved in ceil division and morphological elonga- tion, has a role in chronic persistent infections. Ge- netic property of H. pylori cdrA was evaluated using polymerase chain reaction and sequencing in 128 （77 American and 51 Japanese） clinical isolates obtained from 48 and 51 patients, respectively. Enzyme-linked immunosorbent assay was performed to measure in- terleukin-8 （IL-8） secretion with gastric biopsy speci- mens obtained from American patients colonized with cdrA-positive or -negative strains and AGS cells co- cultured with wild-type HPK5 （cdrA-positive） or its de- rivative HPKT510 （cdrA-disruptant）. Furthermore, the cytotoxin-associated gene A （cagA） status （transloca- tion and phosphorylation） and kinetics of transcription factors [nuclear factor-kappa B （NF-~：B） and inhibition kappa B] were investigated in AGS cells co-cultured with HPK5, HPKT510 and its derivative HPKSCA （cagA- disruptant） by western blotting analysis with immuno- precipitation. RESULTS： Genetic diversity of the H. pylori cdrA gene demonstrated that the cdrA status segregated into two categories including four allele types, cdrA-positive （al- lele types, I and 11 ） and cdrA-negative （allele types; 111 and IV） categories, respectively. Almost all Japanese isolates were cdrA-positive （ 1 ： 7.8% and 11 ： 90.2%）, whereas 16.9% of American isolates were cdrA-positive （11） and 83.1% were cdrA-negative （nl： 37.7% and IV： 45.5%）, indicating extended diversity of cdrA in individual American isolates. Comparison of each isolate from different regions （antrum and corpus） in the stomach of 29 Americans revealed that cdrA status was identical in both isolates from different regions in 17 cases. However, 12 cases had a different cdrA al- lele and 6 of them exhibited a different cdrA category between two regions in the stomach. Furthermore, in 5 of the 6 cases possessing a different cdrA category, cdrA-negative isolate existed in the corpus, suggesting that cdrA-negative strain is more adaptable to coloni- zation in the corpus. IL-8 secretions from AGS revealed that IL-8 levels induced by a cdrA-disrupted HPKT510 was significantly lower （P 〈 0.01） compared to wild- type HPK5： corresponding to 50%-60% of those of wild-type HPK5. These data coincided with in vivo data that an average value of IL-8 in biopsy specimens from cdrA-positive and cdrA-negative groups was 215.6 and 135.9 pg/mL, respectively. Western blotting analysis documented that HPKT510 had no effect on CagA translocation and phosphorylation, however, nuclear accumulation of NF-κB was lower by HPKT510 com- pared to HPK5. CONCLUSION： Colonization by a cdrA-negative or cdrA-dysfunctional strain resulted in decreased IL-8 production and repression of NF-κB, and hence, atten- uate the host immunity leading to persistent infection.

Changes of Nerve Growth Factor in Amniotic Fluid and Correlation with Ventriculomegaly

Objective To detect the change of nerve growth factor (NGF) level in human amniotic fluid during gestation, and to explore the relationship between this change and fetal ventriculo-megaly (VM). Methods The studied subjects (collected from 2004 to 2007) were divided into four groups, including the second-trimester pregnancy group (n=113), third-trimester pregnancy group (n=110), fetal cerebral VM group (n=12), and healthy control group (n=12) which matched with the VM group in gestational weeks. The amniotic fluid specimens were obtained during amniocentesis or cesarean section. The NGF levels in amniotic fluid were detected with en-zyme-linked immunosorbent assay. Results A significantly negative correlation was found between gestational age and the NGF level in amniotic fluid (r= 0.6149, P<0.0001). The NGF level in patients with fetal VM was significantly lower than that in healthy controls (33.95±29.24 pg/mL vs. 64.73±16.21 pg/mL, P=0.024). Conclusion NGF levels in amniotic fluid may be a sensitive marker for fetal VM.

Keratinocyte growth factor gene therapy ameliorates ulcerative colitis in rats

AIM:To investigate the effect of keratinocyte growth factor(KGF) gene therapy in acetic acid-induced ulcerative colitis in rat model.METHODS:The colitis of Sprague-Dawley rats was induced by intrarectal infusion of 1 mL 5%(v/v) acetic acid.Twenty-four hours after exposed to acetic acid,rats were divided into three experimental groups:control group,attenuated Salmonella typhimurium Ty21a strain(SP) group and SP strain carrying human KGF gene(SPK) group,and they were separately administered orally with 10% NaHCO3,SP or SPK.Animals were sacrificed and colonic tissues were harvested respectively on day 3,5,7 and 10 after administration.Weights of rats,colonic weight/length ratio and stool score were evaluated.Histological changes of colonic tissues were examined by hematoxylin and eosin(HE) staining method.The expression of KGF,KGF receptor(KGFR) and TNF-α were measured either by enzyme-linked immunosorbent assay or Western blotting.Immunohistochemistry was used to detect the cellular localization of KGFR and Ki67.In addition,superoxide dismutase(SOD) activity and malondialdehyde(MDA) contents in the homogenate were measured.RESULTS:Body weight and colonic weight/length ratio were declined in SPK group compared with SP and control groups(body weight:272.78 ± 17.92 g vs 243.72 ± 14.02 g and 240.68 ± 12.63 g,P < 0.01;colonic weight/length ratio:115.76 ± 7.47 vs 150.32 ± 5.99 and 153.67 ± 5.50 mg/cm,P < 0.01).Moreover,pathological changes of damaged colon were improved in SPK group as well.After administration of SPK strain,KGF expression increased markedly from the 3rd d,and remained at a high level till the 10th d.Furthermore,KGFR expression and Ki67 expression elevated,whereas TNF-α expression was inhibited in SPK group.In the group administered with SPK,SOD activity increased significantly(d 5:26.18 ± 5.84 vs 18.12 ± 3.30 and 18.79 ± 4.74 U/mg,P < 0.01;d 7:35.48 ± 3.35 vs 22.57 ± 3.44 and 21.69 ± 3.94 U/mg,P < 0.01;d 10:46.10 ± 6.23 vs 25.35 ± 4.76 and 27.82 ± 6.42 U/mg,P < 0.01) and MDA contents decreased accordingly(d 7:7.40 ± 0.88 vs 9.81 ± 1.21 and 10.45 ± 1.40 nmol/mg,P < 0.01;d 10:4.36 ± 0.62 vs 8.41 ± 0.92 and 8.71 ± 1.27 nmol/mg,P < 0.01),compared with SP and control groups.CONCLUSION:KGF gene therapy mediated by attenuated Salmonella ameliorates ulcerative colitis induced by acetic acids,and it may be a safe and effective treatment for ulcerative colitis.

Emodin promoted pancreatic claudin-5 and occludin expression in experimental acute pancreatitis rats

AIM:To investigate the effect of emodin on pancreatic claudin-5 and occludin expression,and pancreatic paracellular permeability in acute pancreatitis(AP).METHODS:Experimental pancreatitis was induced by retrograde injection of 5% sodium taurocholate into the biliopancreatic duct.Emodin was injected via the external jugular vein 0 or 6 h after induction of AP.Rats from sham operation and AP groups were injected with normal saline at the same time.Samples of pancreas were obtained 6 or 12 h after drug administration.Pancreatic morphology was examined with hematoxylin and eosin staining.Pancreatic edema was estimated by measuring tissue water content.Tumor necrosis factor(TNF)-α and interleukin(IL)-6 level were measured by enzyme-linked immunosorbent assay.Pancreatic paracellular permeability was assessed by tissue dye extravasation.Expression of pancreatic claudin-5 and occludin was examined by immunohistology,quantitative real-time reverse transcriptase polymerase chain reaction and western blotting.RESULTS:Pancreatic TNF-α and IL-6 levels,wet/dry ratio,dye extravasation,and histological score were significantly elevated at 3,6 and 12 h following sodium taurocholate infusion;treatment with emodin prevented these changes at all time points.Immunostaining of claudin-5 and occludin was detected in rat pancreas,which was distributed in pancreatic acinar cells,ductal cells and vascular endothelial cells,respectively.Sodium taurocholate infusion significantly decreased pancreatic claudin-5 and occludin mRNA and protein levels at 3,6 and 12 h,and that could be promoted by intravenous administration of emodin at all time points.CONCLUSION:These results demonstrate that emodin could promote pancreatic claudin-5 and occludin expression,and reduce pancreatic paracellular permeability.

The inhibitory effect of selective cyclooxygenase-2 inhibitor NS-398 on the cholangiocarcinoma line (QBC939)

Objective： We studied the inhibitory effect of human cholangiocarcinoma line （QBC939） treated by selective cyclooxygenase-2 （COX-2） inhibitor NS-398 and provided the theoretical foundation for the clinical practice. Methods： Selec- tive COX-2 inhibitor NS-398 on the growth suppression was evaluated by MTT method. The apoptotic rate was quantified and cell cycle by flow cytometry （FCM）. Invasive ability was detected by Transwell. The expression of vascular endothelial growth factor （VEGF） and COX-2 in cholangiocarcinoma line was determined by enzyme-linked immunosorbent assay （ELISA）. Results： NS-398 could be time-dose dependently inhibited the growth, induced apoptosis, invasived ability, S phase was inhibited, down-regulate the expression of VEGF and COX-2 in cholangiocarcinoma line （QBC939）. Conclusion： NS398 can inhibit proliferation of cholangiocarcinoma cell line QBC939 and inhibit the invasive ability and induce its apoptosis in vitro, which may contributed to the COX-2 dependent pathway to reduce the release of VEGF.

Relationship between serum IL-18 and VEGF levels in patients with prostate cancer

Objective： The aim of our study was to analyze intedeukin-18 （IL-18） and vascular endothelial growth factor （VEGF） serum levels in patients with prostate cancer before and after operation and the possible correlation between IL-18 and VEGF serum levels. Methods： Serum IL-18 and VEGF levels were measured by enzyme-linked immunosorbent assay （ELISA） in 36 patients with prostate cancer before and after radical prostatectomy and in 25 healthy controls. Results： Serum IL-18 and VEGF levels were significantly higher in patients with prostate cancer before operation with respect to healthy controls （P 〈 0.05）, with a highly significant correlation between IL-18 and VEGF （R = 0.800, P = 0.017）. It was significantly reduced in IL-18 and VEGF after operation. IL-18 and VEGF serum concentrations were correlated with the clinicalopathologi- cal status of patients with prostate cancer. Conclusion： It is correlative with serum IL-18 and VEGF. Serum IL-18 and VEGF levels may be useful prognostic marker in patients with prostate cancer.

Clinical study on IL-8,TNF-α,T-cell subgroup in serum of patients with rectal cancer

Objective:The aim of our study was to investigate the immune status of patients with rectal cancer and its relationship with clinicopathological features.Methods:The serum levels of interleukin-8(IL-8),tumor necrosis factor(TNF-α) and T-cell subgroup contents were measured using a double-antibody sandwich assay of ELISA in 43 patients with rectal cancer,and compared with the normal health adults.Results:In patients with rectal cancer,the serum levels of CD4,CD4/CD8 of T-cell subgroup in peripheral blood were significantly lower than the control group(P < 0.01),which gradually decreased with increase of Dukes stage;but the levels of CD8,IL-8 and TNF-α were higher than the control group,which gradually increased with increase of Dukes stage.Conclusion:The immunocompromice exists in patients with rectal cancer,there is a correlation between the contents of T-cell subgroup,IL-8 and TNF-α in serum and the Dukes stage of rectal cancer.Therefore immunotherapy can be used in patients with rectal cancer.

NITRIC OXIDE AND VEGF EXPRESSION IN ISCHEMIC-HYPOXIC RETINOPATHY

Objective To evaluate nitric oxide(NO)and vascular endothelial growth factor(VEGF)in vitreous humor and blood samples in patients with proliferative diabetic retinopathy(PDR)and in patients with branch retinal vein occlusion(BRVO).Methods NO concentrations were determined by using the Greiss reaction in plasma and vitreous humor samples.VEGF levels were assayed by ELISA.The patients in the studies were divided into four groups:16 patients with PDR,5 patients with BRVO,11 patients with rhegmatogenous retinal detachment(RRD),and 10 patients with idiopathic macular hole(IMH).Results The vitreous fluid levels of NO were significantly higher in patients with PDR(15.2μmol/L,4.6-50.9μmol/L)than those in the other three groups(F=5.13,P=0.005).The concentrations of VEGF were significantly higher in patients with PDR and BRVO(1507.2 μg/mL,50.71-3722.0μg/ml;838.8μg/mL,159.6-3328.0μg/mL)than those in the other two groups(F=6.84,P=0.0008),but highest in PDR(T=3.92,P=0.001).There was no significant difference between NO and VEGF in serum in four groups.There was no correlation between concentrations of NO and VEGF in four groups whatever in vitreous or plasma(all P>0.05).Conclusion The results suggest that higher levels of NO and VEGF may be related to the angiogenesis in DR.

Enhanced plasma factor Ⅷ activity in mice via cysteine mutation using dual vectors

Hemophilia A is caused by a genetic mutation in coagulation factor VIII （FVIII） gene and gene therapy is considered to be a promising strategy for its treatment. We recently demonstrated that co-delivery of two vectors expressing M662C mutated heavy and D1828C mutated light chain genes of B-domain-deleted coagulation factor VIII （BDD-FVIII） leads to inter-chain disulfide cross-linking and improved heavy chain secretion in vitro. In this study, co-injection of both M662C and D1828C mutated BDD-FVIII gene expression vectors into mice resulted in increased heavy chain secretion and coagulation activity in plasma in vivo. Approximately （239＋_56） ng mL-1 above endogenous levels of transgenic FVIII heavy chain was found in mouse plasma using a chain-specific ELISA. For FVIII coagulation activity, approximately （1.09＋_0.25） IU mL-1 above en- dogenous levels were detected in co-injected transgenic mouse plasma using a chromogenic assay. These data demonstrate that inter-chain disulfide bonds likely increase heavy chain secretion and coagulation activity in the plasma of transgenic mice with an improved efficacy of a dual-vector delivery of BDD-FVIII gene. These findings support our ongoing efforts to develop a gene therapy for hemophilia A treatment using dual-AAV vectors.

Cytocompatibility of regenerated silk fibroin film:a medical biomaterial applicable to wound healing

Objective： To explore the feasibility of using regenerated silk fibroin membrane to construct artificial skin substitutes for wound healing, it is necessary to evaluate its cytocompatibility. Methods： The effects of regenerated silk fibroin film on cytotoxicity, adhesion, cell cycle, and apoptosis of L929 cells, growth and vascular endothelial growth factor （VEGF） expression of ECV304 cells, and VEGF, angiopoietin-1 （Ang-1）, platelet-derived growth factor （PDGF） and fibroblast growth factor 2 （FGF2） expression of WI-38 cells were assessed by 3-（4,5）-dimethylthiahiazo （-z-yl）-3,5-di-phenytetrazoliumromide （MTT） assay, viable cell counting, flow cytometry （FCM）, and enzyme-linked immunosorbant assay （ELISA）. Results： We showed that the regenerated silk fibroin film was not cytotoxic to L929 cells and had no adverse influence on their adhesion, cell cycle or apoptosis; it had no adverse influence on the growth and VEGF secretion of ECV304 cells and no effect on the secretion of VEGF, Ang-1, PDGF and FGF2 by WI-38 cells. Conclusion： The regenerated silk fibroin film should be an excellent biomaterial with good cytocompatibility, providing a framework for reparation after trauma in clinical applications.