Lentivirus-mediated RNA interference targeting vascular endothelial growth factor gene enhances the sensitivity of K562 cells to STI571

AIM: To investigate the effects of RNA interference (RNAi) inhibiting the expression of vascular endothelial growth factor (VEGF) gene mediated by lentiviral vector on the proliferation and apoptosis of K562 leukemic cell line. METHODS: A lentiviral vector containing short hairpin RNA (shRNA) targeting VEGF was constructed and cotransfected with the packaging plasmids mixture into 293T cells by Lipofectamine 2000. K562 cells were infected with the packaged lentivirus. The levels of VEGF mRNA and protein were detected by real-time quantitative RT- PCR, Western blotting and ELISA. Cellular proliferation was determined by trypan blue dye exclusion and MTT assay. STI571 (imatinib mesylate)-induced apoptosis was analyzed by flow cytometry. RESULTS: The lentiviral shRNA vector targeting VEGF was successfully constructed and transfected into K562 cells. The expressions of VEGF mRNA and protein in K562-shVEGF cells transfected with pRNAT-shRNA were significantly inhibited when compared with those of K562 and K562-con cells (mock transduction). The proliferation rate of K562-shVEGF cells slowed down. After STI571 treatment, the percentages of apoptotic cells in K562-shVEGF cells increased more significantly than those of K562 and K562-con cells (P<0.05). CONCLUSION: Inhibition of VEGF by lentivirus-mediated RNAi effectively inhibits proliferation and increases the sensitivity of K562 cells to STI571.     

Progress in association of RSK2 with malignancies and its inhibitors

Ribosomal S6 kinase 2 (RSK2) is a member of the p90^Rsk, which belongs to Ser/Thr kinase family. It is a downstream molecule of Ras/MAPK cascade and regulates a variety of cellular processes. RSK2 plays a key role in the cell proliferation, survival and transformation. Furthermore, the aberrant up-regulation of RSK2 was observed in different malignancies. We here briefly review the structure and function of RSK2, the relationship between RSK2 and tumor, and the existing RSK2 inhibitors.     

Influences of protein kinase C inhibitors on the expression of IL-2 and IFN-γ by human T-lymphocytes

AIM:To investigate the influences of protein kinase C(PKC) inhibitors on the expression of interleukin-2(IL-2) and interferon-γ(IFN-γ) byin vitro activated T-lymphocytes. METHODS:Double fluorescent staining together with flow cytometry was adopted to detect intracellular cytokines and to analyze the effects of H7 and gossypol on IL-2 and IFN-γ expression levels of T-lymphocytes stimulated with phorbol ester (PDB)+ionomycin(I) in the presence of monensin.RESULTS:The expression rates of IL-2 and IFN-γ of CD3⁺ T cells stimulated with PDB+I for 4 h were 16.64±2.04 and 25.81±3.53(x±s), respectively, which were significantly higher than that of control (1.06±0.22 and 3.12±0.77)(P＜0.05). Gossypol was able to inhibit the expression of IL-2 and IFN-γ significantly, with the expression rates of 2.08±0.12 and 9.01±1.90, respectively. At the presence of 50 μmol/L H7, the rates of IL-2⁺ and IFN-γ⁺ CD3⁺ T cells were 0.43±0.06 and 2.40±0.27, respectively. The effect of H7 was stronger than that of gossypol. CONCLUSION:PKC plays an important role in the expression of IL-2 and IFN-γ of CD3⁺T cells and its inhibitors H7 and gossypol exert significant inhibitory effect on the expression of these two cytokines. It is suggested that H7 and gossypol may have modulatory effect on T-cell-dependent specific immune responses by inhibiting PKC activity.     

探究城市森林公园建设对提升象山城市品位的意义

建设城市森林公园,打造环境优雅、风光奇特、空气清新、景色宜人、树木花草万紫千红的生态景观,是象山新时代发展的必然趋势。经探索研究,象山县可因地制宜打造一个集休闲、健身、生态、文化、游乐、度假于一体的城市森林公园。     

建立果品流通体系,促进果品产业化发展

山东省是水果生产大省,素有“落叶果树王国”之称。据资料统计,2000年果品产量已达966万t,仅苹果产量就达600多万t,居全国各省市苹果产量的首位。但是由于果品生产仍是以千家万户分散经营的生产方式为主,这种生产方式在信息、资金、运输、贮藏、加工等方面,很难适应不断变化的国内外大市场的需要,致使果品市场不够兴旺,销售份额不大,无果区买不到水果,产果区的水果却大量积压。随着果品产量的不断增加,产、供、销等诸多环节的矛盾与问题将会更加突出;同时,我国加入世界贸易组织,国外市场对国内市场产生了更大的冲击,这就需要加强果品流通领…     

Effects of the peritoneal dialysate biocompatibility on the 2', 5'-oligoadenylate synthetase activity

AIM: To examine the 2', 5'-oligoadenylate synthetase activity and the peritoneal pathological changes when using commercially dialysates in order to reveal effects of the peritoneal dialysate biocompatibility on the immune function and morphological changes. METHODS: (1) CAPD animal models were made using dialysates. (2) Dynamic measurements of 2', 5'-oligoadenylate synthetase activity were made by paper chromatography. (3)The pathological changes of the peritoneal mesothelial cells were examined by light microscope. RESULTS: 2', 5'-oligoadenylate synthetase activities in both experimental (dialysis solution groups) and control group were increased and the mesothelial cell began to exfoliate and become thicker with the infiltration of inflammatory cells at different degrees in the experimental groups, while in the Baxter group, no active inflammation was observed. When the peritoneal dialysis stopped, activity of 2', 5'-oligoadenylate synthetase began to be normal in control group and continued to decline in experimental groups. CONCLUSION: Prolonged immune suppression existed with long-term peritoneal dialysis, causing pathological changes of the peritoneum at certain degrees. Different dialysis solutions resulted in different degrees of immuno-supression and peritoneal damages, which was relevant to the biological compatibility of the peritoneal dialysate.     

Nitrogenous compounds enhance the growth of petunia and reprogram biochemical changes against the adverse effect of salinity

This study aimed to determine ways to improve the growth and salt tolerance of petunia. Effects of polyamines (PAs; spermidine [Spd], spermine [Spm], and putrescine [Put]) and a nitric oxide (NO) donor (sodium nitroprusside [SNP]) were investigated. Initially, we screened petunia cultivars against sodium chloride (0–125 mM). The petunia cultivar Hurrah Red was identified as salt-sensitive cultivar in the basis of salt-effect on seed germination, fresh weight, and root length of seedlings. Treatment of Hurrah Red shoots with nitrogenous compounds improved the number, length, and fresh weight of roots, as well as the length and fresh weight of shoots over those of the control. Furthermore, plantlets rooted in an optimal concentration of Spd (34.5 µM), Spm (24.8 µM), Put (62.1 µM), and SNP (3.9 µM) were treated with 200 mM NaCl for 3 days to assess their tolerance level. Salt-affected plantlets showed higher level of lipid peroxidation, reduced catalase (CAT), peroxidase (POD), polyphenol oxidase (PPO) activities and decreased contents of photosynthetic pigments and polyphenol than those of the control. PAs and SNP treatments significantly elevated photosynthetic pigments, enhanced antioxidant enzymes, and decreased lipid peroxidation in salt-stressed plantlets. Moreover, the growth and salt-tolerance response of petunia was highest when plantlets were exposed to SNP, followed by levels on treatment with Put, Spm, and Spd. Thus, the findings of this study suggested that treatment with exogenous SNP, Put, Spm, and Spd could protect petunia plants against soil salinity and improve their commercial production.     

Three kinds of antisense oligodeoxynucleotide enhance the sensitivity of leukemia cell K562 to cisplatin

AIM: To investigate whether antisense oligodeoxynucleotides of hTERT、bcl-2 and c-myc could enhance the sensitivity of leukemia cell K562 to cisplatin. METHODS: The inhibiting effects of cisplatin and cisplatin plus antisense oligodeoxynucleotide on K562 cells were determined by MTT. RESULTS: The inhibiting rate of 20 μmol/L cisplatin to K562 cell is 17.17%±1.36% and it becomes 25.41%±1.77%, 26.18%±1.43% and 28.29%±1.05%, respectively, as combinated with antisense oligodeoxynucleotide of hTERT, bcl-2 or c-myc. CONCLUSION: These results indicated that antisense oligodeoxynucleotides of hTERT, bcl-2 and c-myc enhanced efficacy of cisplatin in K562 leukemic cells.     

Ca~(2+)-ATPase抑制剂对采后草莓呼吸速率和花青苷含量的影响

用Ca2+-ATPase抑制剂钒酸钠(SO)和曙红(EB)浸泡处理转红期草莓果实(cv.Darselect)后表明,2者能提高果实呼吸速率,迅速促进外层果肉的花青苷生成,增加其可溶性糖含量,髓部果肉的花青苷生成和可溶性糖的变化较慢,而对果实的失重率影响不大。因此,Ca2+-ATPase抑制剂具有促进果实成熟衰老的作用。     

Primary determination for activity and expression of Stx2a'-LHRH chimeric toxin

AIM:To express chimeric toxin Stx2a'-LHRH and to investigate the cytotoxic activity of recombinant toxin Stx2a'-LHRH to human carcinoma cells.METHODS:Stx2a'-LHRH sequences that added the restriction endonucleases NcoⅠ and EcoRⅠ at the 5' and 3 ends were amplified by PCR and digested with appropriate restriction enzymes. The digested fragment was subcloned into the vector obtatined by digestion of plasmid pET-28a(+) with NcoⅠ and EcoRⅠ. E. coli BL21 (DE3) cells were transformed with plasmids of interst and cultured in LB medium containing ampicillin. Expression of the recombinant protein was induced by the addition of isopropylthio-β-D-galactoside (IPTG). The cytotoxity of Stx2a'-LHRH to Hep-2 cells was observed under the microscopy. RESULTS:Recombitant plasmid pET-SL was constructed successfully and the clones expressing pET-SL stablely were obtained. A special electrophoretic band in SDS-PAGE (a glycoprotein of 28kD) was noted. Stx2a'-LHRH killed Help-2 cells clearly. CONCLUSION:In this study, construction of chimeric toxin Stx2a'-LHRH and its expression were described. Moreover, it has obvious cytotoxity to Hep-2 cell. These finding could open up new vistas in the study of targeted durgs.     

The relationship between the activity of phospholipase A2 and chronic hypoxic pulmonary hypertension

AIM:To study the relationship between the activity of phospholipase A₂ (PLA₂) and chronic hypoxic pulmonary hypertension. METHODS:29 healthy SD rats were randomly divided into normal control group, chronic hypoxic group and hypoxia plus Polidatin (PD) group. The model of rat chronic hypoxic pulmonary hypertension was made by method of intermittent isobaric hypoxia for 21 days. The mean pulmonary arterial pressure (mPAP) was measured by inserting a microcatheter into the pulmonary artery. RESULTS:After exposing hypoxia for 21 days, the mPAP, R/L+S, the PLA₂ activity, TXB₂, MDA in plasma and lung homogenate increased significantly, while 6-k-PGF_1α, SOD decreased significantly. Pretreatment with PD could relieve the changes mentioned above.CONCLUSION:PLA₂ plays an important inducing role through its metabolic products and the interactional radicals in the formation of chronic hypoxic pulmonary hypertension.     

Effects of cyclooxygenase-2 inhibitors on gastric epithelial cell proliferating and gastric healing following hydrochloric acid-induced injury in rats

AIM: To clarify the effects of specific and non-specific cyclooxygenase-2 (COX-2) inhibitors on gastric epithelial cell proliferating and gastric healing following acid-induced damage. METHODS: Male Sprague-Dawley rats were given 1 mL of 0.6 mol/L hydrochloric acid (HCl) into the stomach. Ten minutes after the administration of the acid, the animals were given NS-398 (COX-2 inhibitor) or indomethacin. Levels of COX-1 and COX-2 in the gastric mucosa before and after HCl-administration were analyzed using western blotting and immunohistochemical staining. Proliferating cell nuclear antigen (PCNA) was detected using immunohistochemistry for epithelial cell proliferation. Gastric lesion index (LI) was assessed using planimetry. RESULTS: Expression of COX-2 was enhanced mainly in surface epithelial cells and neck cells following HCl-administration. At 24 h following acid administration, PCNA labeling index (PCNA-LI) was (22.72±4.33) % and (21.98±5.18) % in the groups treated with 40 mg/kg of NS-398 and indomethacin respectively, which was significantly lower than that in the control group [ (34.46±3.61) %, P< 0.05 ]; LI was (1.28±0.58) % and (1.16±0.56) % in the groups treated with 4 mg/kg and 40 mg/kg of NS-398, respectively, which was significantly higher than that in the control group [ (0.58±0.24) %, P< 0.05 ]. CONCLUSIONS: The present study demonstrated that cyclooxygenase-2 inhibitors delayed gastric mucosal healing by suppressing expansion of the mucosal proliferative zone. These results provide evidence that cyclooxygenase-2 plays an important role in gastric mucosal regeneration.     

Relationship between activity of matrix metalloproteinases-2 and invasion,metastasis of breast cancer

AIM: To investigate relationship between activity of matrix metalloproteinases-2 (MMP-2, 72 kD) and invasion, metastasis of breast cancer. METHODS: Useing zymography and computer software assisted analysis, the activitive levels of MMP-2 (72 kD) in tissues from breast cancer were measeured. RESULTS: Mean activitive levels of MMP-2 72 kD (13.93±3.60) in breast cancer were lower than those in benign disease (21.43±8.31), P<0.05. There was no difference (P>0.05) in MMP-2 62 kD+72 kD of benign and malignant disease, but MMP-2 62 kD (13.83±4.53) and MMP-2 62 kD/62 kD+72 kD(0.48) respectively were significantly higher in malignant disease (P<0.01). It was also found that MMP-2 62 kD/62 kD+72 kD were apparently higher in invasive carcinomas (0.48) and lymph node metastases (0.61), P<0.01, respectively. CONCLUSION: These results demonstrated that a clear relationship between MMP-2 activity and the invasion and metastasis of breast carcinoma.     

Managing faba bean residues to enhance the fruit quality of the melon (Cucumis melo L.) crop

Mulching and/or topsoil incorporation of plant residues from green manure legumes can increase cropping system sustainability, and can supply and retain nutrients. Two field experiments were conducted in Italy over the period of 2006–2007, to investigate the effect of faba bean grown in a temperate environment to fruit morphology, quality and chemical composition of the subsequent melon (Cucumis melo L.) crop. Flowering faba bean plants were managed as mulch on the melon plant rows and cut and incorporated into the soil, in comparison with cultivated soil (control) and black plastic mulch.     

重视“五度一防”提升果品产业整体效益

分析着色度、糖度、硬度及环境温湿度和人工机械伤害等对果品质量的影响及常见原因,提出提高果品着色度、糖度、硬度和改善环境温湿度状况及防止人工机械伤害,提高果品产业整体效益的措施。     

Design and immune activity detection of long peptide vaccines targeting lung cancer antigen COX-2

AIM:To design long peptides based on cytotoxic T-lymphocytes (CTL) epitope prediction for lung cancer antigen cyclo-oxygenase-2 (COX-2) and to study the immune activity of the long peptides. METHODS:HLA-A2 epitopes from COX-2 protein were predicted by NetCTL 1.2, SYFPEITHI and IEDB. The CTL epitope-concentrated area was analyzed, and the appropriate length of long peptides were designed. In vitro activity experiments were used to verify the immune activity of the long peptides. ELISPOT assay and intracellular cytokine staining assay were used to investigate the ability of the peptide to induce specific restricted CTLs and release of interferon-γ (IFN-γ). The ability of the peptides to induce T-cell response was investigated by lactate dehydrogenase (LDH) and CFSE cytotoxicity assay in vitro. RESULTS:ELISPOT and intracellular cytokine staining assay showed P315-338 and P375-401 were able to induce specific CTLs and higher levels of IFN-γ release. The results of LDH and CFSE cytotoxicity assays showed the CTLs induced by P315-338 and P375-401 lysed the target cells. CONCLUSION:Two long peptides pointing to lung cancer antigen COX-2 are successfully identified, which could be used as immunotherapy vaccine in future.     

Effects of halothane and sevoflurane on the function,metabolism and Ca2＋-ATPase activity of ischemic myocardium

AIM: To study the effects of 1. 5 MAC halothane and sevoflurane on ischemic myocardium. METHODS: The isolated rat heart were perfused with halothane and sevoflurane and HR, LVEDP, LVDP, +dp/dt, -dp/dt, coronary flow (CF), the myocardial ATP content and Ca²⁺-ATPase activity were determined before and 10 min and 25 min after ischemia. In the meantime, LVP was recorded during 25 min ischemia. RESULTS: 1. 5MAC sevoflurane significantly increased CF in normal isolated rat hearts. Both halothane and sevoflurane depressed myocardial contractile function, increased normal myocardial energy storage. After 10 min ischemia, the decrease of myocardial ATP content were slowed down by halothane and sevoflurane, especially halothane. During 25 min of ischemia, the onset time of contracture was significantly delayed, and the contracture intensity was alleviated by halothane, but not sevoflurane. CONCLUSION: Halothane has better protective effect on ischemic myocardium than sevoflurane through preventing the decrease of myocardial ATP content and Ca²⁺-ATPase activity during ischemia.