接种根瘤菌对蚕豆／玉米间作系统产量及结瘤作用的影响

通过田间试验,研究了不同施氮水平下蚕豆接种根瘤菌GS374对蚕豆/玉米间作系统产量及蚕豆结瘤作用的影响。结果表明,不施氮处理接种根瘤菌所获得的单作或间作系统产量与不接种但施N225kghm-2的相应系统产量相当,且施N225kghm-2处理接种仍能促进蚕豆的结瘤作用。统计分析表明,与不接种根瘤菌、蚕豆单作、不施氮相比,接种、蚕豆/玉米间作、施氮均极显著地提高了蚕豆生物学产量,但只有间作能显著增加其籽粒产量;施氮显著增加玉米生物量和籽粒产量。施N225kghm-2后,蚕豆接种、间作对玉米生物量无显著影响;但不施氮时蚕豆接种显著提高了与之间作的玉米籽粒和生物学产量,增幅分别为34.3%和25.6%。接种根瘤菌显著提高了不同氮处理以籽粒产量为基础计算的土地当量比和不施氮处理以生物学产量为基础计算的土地当量比。蚕豆接种根瘤菌与不接种相比,其单株根瘤数和根瘤干重均显著增加;间作与蚕豆单作相比对根瘤数的影响较小,但显著促进了蚕豆单株根瘤干重的增加。因此,本研究认为豆科作物接种合适的根瘤菌,是进一步提高豆科/禾本科作物间作系统间作优势的又一重要途径。

大豆根瘤菌结瘤作用统计方法的建立及其对接种效果的统计分析

利用发光大豆根瘤菌ＨＮ０１ＬＣ０２接种大豆黑农３９，通过对发光根瘤在大豆根系上的数量和位置分布进行分析，对根瘤菌的结瘤作用建立了一种新型统计方法，并用这种统计方法分析了不同接种技术对豆科植物结瘤作用和植物生长的影响。

一株能在大豆上结瘤的苜蓿中华根瘤菌

苜蓿中华根瘤菌 (Sinorhizobiummeliloti)XJ96 0 77分离自新疆的苜蓿根瘤中 ,其原宿主为紫花苜蓿 (Medicagosativa)。交叉结瘤试验发现 ,它既可在苜蓿上又能在大豆上结瘤固氮。DNA (G +C)mol%分析表明 ,XJ96 0 77的DNA (G +C)mol%为 6 1 9% ,与已报道的根瘤菌属的DNA (G +C)mol%范围 (5 9%～ 6 4 % )相符。DNA同源性分析表明 ,XJ96 0 77与苜蓿中华根瘤菌USDA10 0 2 T 和 0 4 2BM的同源性分别达到 93%和 80 % ,说明XJ96 0 77归属于苜蓿中华根瘤菌。应用绿色荧光蛋白基因标记XJ96 0 77,得到重组菌株XJ96 0 77(G)。将其接种普通紫花苜蓿 ,通过激光共聚焦荧光显微镜可以检测到标记基因的表达。接种北引 1号大豆上 ,同样可以清楚地观察到标记基因在根瘤中的表达 ,从而确证了XJ96 0 77能同时在苜蓿和大豆上结瘤。通过不同品种大豆的结瘤试验 ,发现XJ96 0

结瘤因子的研究进展(英文)

结瘤因子是由根瘤菌产生的一类信号分子 ,它们在结瘤的起始阶段发挥着十分重要的作用。新近的研究结果证明结瘤因子大分子骨架上的不同侧链基团是决定细菌与宿主植物间相互识别的关键因素 ,根瘤菌细胞中一系列结瘤基因编码能够合成Lipo chito oligosaccharides(LCOs)的各种酶类 ,进而确定结瘤信号分子的特定结构。目前 ,一系列令人兴奋的实验结果表明 :LCOs不仅可促进豆科作物的生物固氮作用 ,对一些非豆科作物的细胞分裂作用等同样具有刺激作用。对根瘤菌结瘤因子的研究显然有助于进一步了解细菌与植物的相互作用机理 ,并进而为农业生产带来直接利益。本文在综述这方面的研究进展同时 ,还就瘤菌、豆科作物和结瘤信号分子之间的相互作用机理 ,以及根际促生细菌、水杨酸和结瘤信号分子之间的可能关系进行了理论分析。

苜蓿中华根瘤菌042BMnoeA基因的克隆、敲除与功能的初步分析

通过PCR扩增获得了 0 4 2BM的noeA基因。该基因与苜蓿中华根瘤菌 (Sinorhizobiummeliloti) 10 2 1noeA的同源性为 99% ,而其NoeA与 10 2 1NoeA的相似性为 97%。还发现其NoeA与中慢生根瘤菌 (Mesorhizobiumsp .)BNC1可能的SAM_依赖性的甲基转移酶相似性为 32 % ,而其 30 3～ 36 2氨基酸区域与大肠杆菌 (Escherichiacoli)的核糖体 5 0S亚基的L11蛋白甲基转移酶 (PrmA)的 16 0～ 2 2 0氨基酸区域的相似性达到 4 1%。通过插入卡那盒 ,敲除noeA ,获得突变株 0 4 2BMA_Km。与苜蓿中华根瘤菌 0 4 2BM相比 ,敲除noeA的突变株在普通紫花、保定、宁夏、百发和傲汉苜蓿品种上的结瘤数、根瘤鲜重和植株地上部分的干重都有不同程度的增加 ,而在秘鲁苜蓿品种上的结瘤数和植株地上部分的干重明显下降 ,在皇后和美国杂花苜蓿品种上则没有明显的变化。

Combined effects of Cantide and chemotherapeutic drugs on inhibition of tumor cells' growth in vitro and in vivo

AIM: To investigate the combination effect of hTERT antisense oligonucleotide 'Cantide' and three chemotherapeutic drugs (cisplatin, 5-fluorouracil (5-FU) and adriamycin (ADM)) on inhibiting the proliferation of HepG2, BGC and A549 cell lines in vitro, and to investigate the efficacy of Cantide used in combination with cisplatin (DDP) in vivo. METHODS: Cantide was transfected into these tumor cells by Lipofectin, and cell growth activity was calculated by microcytotoxicity assay. In vivo study, cells of HepG2 were implanted in Balb/c nude mice for 4 d. Then Cantide, DDP and Cantide+DDP were given intra peritonea Ily for 24 d respectively. The body weights of the tumor-bearing animals and their tumor mass were measured later to assess the effect of combination therapy in the nude mice. To evaluate the interaction of Cantide and these chemotherapeutic drugs, SAS software and Jin Zhengjun method were used. RESULTS: Combination treatments with 0.1μmol/L Cantide reduced the IC50 of DDP, 5-FU and ADM from 1.07, 4.15 and 0.29 μg/mL to 0.25,1.52 and 0.12 μg/mL respectively. The inhibition ability of DDP, 5-FU and ADM respectively in combination with Cantide in these tumor cells was higher than that of these drugs alone (P<0.0001). And synergism (Q≥1.15)was observed at the lower concentration of DDP (≤1μg/mL) and ADM (≤0.1 μg/mL) with combination of Cantide. In vivo, combination treatment with Cantide and DDP produced the greater growth inhibition of human liver carcinoma cells HepG2 in nude mice (0.65±0.19 g tumor) compared with that when only one of these drugs was used (Cantide group: 1.05±0.16 g tumor, P= 0.0009<0.001; DDP group: 1.13±0.09 g tumor, P= 0.0001<0.001). CONCLUSION: These findings indicate that Cantide may enhance therapeutic effectiveness of chemotherapeutic drugs over a wide range of tumor cells in vitro, and the combination use of Cantide and DDP can produce much higher inhibition rates, as compared with when either of these drugs was used only in vivo.

Science Letters:IGFBP7 plays a potential tumor suppressor role against colorectal carcinogenesis with its expression associated with DNA hypomethylation of exon 1

Insulin-like growth factor binding-protein-7 （IGFBP7） was obtained from our previous colonic adenocarcinoma （CRC） and normal mucosa suppression subtraction hybridization （SSH） cDNA libraries. By RT-PCR and immunohistochemistry, we found that IGFBP7 was overexpressed in CRC tissue compared to normal tissue. However, our in vitro experiments performed in 10 CRC cell lines showed that IGFBP7 expressed only in SW480 and Caco2 cell lines, which implied an underlying reversible regulatory mechanism. Using methylation-specific PCR （MSP） and bisulfite sodium PCR （BSP）, we found that its expression was associated with DNA hypomethylation of exonl. This was further supported by the in vitro study which showed restored IGFBP7 expression after demethylation agent 5-aza-2＇-deoxycytidine treatment. Correlation analysis between IGFBP7 expression and prognosis indicated that overexpression of IGFBP7 in CRC tissue correlated with favourable survival. Investigation of the functional role of IGFBP7 through transfection studies showed that IGFBP7 protein could inhibit growth rate, decrease colony formation activity, and induce apoptosis in RKO and SW620 cells, suggesting it a potential tumor suppressor protein in colorectal carcinogenesis. In conclusion, our study clearly demonstrated that IGFBP7 plays a potential tumor suppressor role against colorectal carcinogenesis and its expression is associated with DNA hypomethylation of exon 1.

ANTITUMOR EFFECTS OF MONOCLONAL ANTIBODY FAB’FRAGMENT—CONTAINING IMMUNOCONJUGATES

Objective.Using monoclonal antibody (mAb) Fab′ fragment to develop mAb immunoconjugates for cancer. Methods.Fab′ fragment of mAb 3A5 was prepared by digestion of the antibody with pepsin and then reduced by dithiothreitol (DTT),while Fab′ fragment of mAb 3D6 was obtained by digestion of the antibody with ficin and subsequently reduced by β mercaptoethanol.The conjugation between Fab′ fragment and pingyangmycin (PYM),an antitumor antibiotic,was mediated by dextran T 40.Immunoreactivity of Fab′ PYM conjugates with cancer cells was determined by ELISA,and the cytotoxicity of those conjugates to cancer cells was determined by clonogenic assay.Antitumor effects of the Fab′ PYM conjugates were evaluated by subcutaneously transplanted tumors in mice. Results.The molecular weight of Fab′ fragment was approximately 53 kD,while the average molecular weight of Fab′ PYM conjugate was 170 kD.The Fab′ PYM conjugates showed immunoreactivity with antigen relevant cancer cells and selective cytotoxicity against target cells.Administered intravenously,Fab′ PYM conjugates were more effective against the growth of tumors in mice than free PYM and PYM conjugated with intact mAb. Conclusion.Fab′ PYM conjugate may be capable of targeting cancer cells and effectively inhibiting tumor growth,suggesting its therapeutic potential in cancer treatment.

Lipid peroxidation and antioxidant status in colorectal cancer

AIM: Reactive oxygen species (ROS) can induce carcinogenesis via DNA injury. Both enzymatic and non-enzymatic parameters participate in cell protection against harmful influence of oxidative stress. The aim of the present study was to assess the levels of final lipid peroxidation products like malondialdehyde (MDA) and 4-hydroxy-2-nonenal (4-HNE) in primary colorectal cancer. Moreover, we analysed the activity of main antioxidative enzymes, superoxide dismutase (Cu, Zn-SOD), catalase (CAT), glutathione peroxidase (GSH-Px) and glutathione reductase (GSSRG-R) and the level of non-enzymatic antioxidants (glutathione, vitamins C and E). METHODS: Investigations were conducted in 81 primary colorectal cancers. As a control, the same amount of sample was collected from macroscopically unchanged colon regions of the most distant location to the cancer. Homogenisation of specimens provided 10% homogenates for our evaluations. Activity of antioxidant enzymes and level of glutathione were determined by spectrophotometry. HPLC revealed levels of vitamins C and E and served as a method to detect terminal products of lipid peroxidation in colorectal cancer. RESULTS: Our studies demonstrated a statistically significant increase in the level of lipid peroxidation products (MDA-Adc. muc.-2.65±0.48 nmol/g, Adc.G3-2.15±0.44 nmol/g, clinical IV stage 4.04±0.47 nmol/g, P<0.001 and 4-HNE-Adc.muc. -0.44±0.07 nmol/g, Adc.G3-0.44±0.10 nmol/g, clinical IV stage 0.52±0.11 nmol/g, P<0.001) as well as increase of Cu,Zn-SOD (Adc.muc.-363±72 U/g, Adc.G3-318?8 U/g, clinical IV stage 421±58 U/g, P<0.001), GSH-Px (Adc.muc. -2143±623 U/g, Adc.G3-2005±591 U/g, clinical IV stage 2467±368 U/g, P<0.001) and GSSG-R (Adc.muc.-880±194 U/g, Adc.G3-795±228 U/g, dinical IV stage 951±243 U/g, P<0.001) in primary tumour comparison with normal colon (MDA-1.39±0.15 nmol/g, HNE-0.29±0.03 nmol/g, Cu, Zn-SOD-117±25 U/g, GSH-Px-1723±189 U/g, GSSG-R-625±112 U/g) especially in mucinous and G3-grade adenocarcinomas as well as clinical IV stage of colorectal cancer. We also observed a decrease of CAT activity (Adc.muc. -40±14 U/g, clinical IV stage 33±18 U/g vs 84±17 U/g, P<0.001) as well as a decreased level of reduced glutathione (clinical IV stage 150±48 nmol/g vs 167±15 nmol/g, P<0.05) and vitamins C and E (vit. C-clinical IV stage 325±92 nmol/g vs 513?4 nmol/g, P<0.001; vit. E-clinical IV stage 13.3±10.3 nmol/g vs 37.5±5.2 nmol/g). CONCLUSION: Colorectal carcinogenesis is associated with serious oxidative stress and confirms that gradual advancement of oxidative-antioxidative disorders is followed by progression of colorectal cancer.

Roles of CREB-binding protein (CBP)/p300 in respiratory epithelium tumorigenesis

CREB-binding protein （CBP） and its homologue p300 are transcriptional co-activators of various sequence-specific transcription factors that are involved in a wide array of cellular activities, such as DNA repair, cell growth, differentia- tion and apoptosis. Several studies have suggested that CBP and p300 might be considered as tumour suppressors, with their prominent role being the cross-coupling of distinct gene expression patterns in response to various stimuli. They exert their actions mainly via acetylation of histones and other regulatory proteins （e.g. p53）. A major paradox in CBP/ p300 function is that they seem capable of contributing to various opposed cellular processes. Respiratory epithelium tumorigenesis represents a complex process of multi-step accumulations of a gamut of genetic and epigenetic aberrations. Transcription modulation through the alternate formation of activating and repressive complexes is the ultimate converging point of these derangements, and CBP/p300 represents key participants in this interplay. Thus, illumination of their molecular actions and interactions could reveal new potential targets for pharmacological interventions in respiratory epithelium carcinogenesis.

Indomethacin suppresses growth of colon cancer via inhibition of angiogenesis in vivo

AIM: It has been reported that regular consumption of nonsteroidal anti-inflammatory drugs like indomethacin decreases the incidence and mortality rate of a number of gastrointestinal cancers. We aimed to explore the efficacy and possible mechanisms of indomethacin on tumor growth and tumor angiogenesis of human colon cancer xenografts in nude mice. METHODS: MTT (thiazolyl blue) assay was used to assess the effect of indomethacin on cultured human colorectal cancer cell line HCT116. HCT116 cells were inoculated subcutaneously into BALB/c-nu/nu mice. After oral administration of indomethacin, 3 mg/kg·d for 4 wk, animals were sacrificed by cervical dislocation. Immunohistochemical staining was employed to determine the microvessel density (MVD) and vascular endothelial growth factor (VEGF) expression in tumor tissues. RESULTS: Indomethacin, a non-selective COX inhibitor, significantly decreased the viability of HCT116 cells in a dose-dependent manner (P<0.05) with 50% inhibition at approximately 318.2±12.7 μmol/L Growth of HCT116 cell tumor was significantly suppressed by indomethacin. The tumor volume was significantly decreased in the treated group (458.89±32.07 mm3) compared to the control group (828.21±31.59 mm3) (P<0.05). The MVD of the treated group (19.50±5.32) was markedly decreased compared to the control group (37.40±4.93) (P<0.001). The VEGF expression of the treated group (1.19±0.17) was obviously reduced as compared to the control group (1.90±0.48) (P<0.01). The decrease in MVD was positively correlated with the decrease of VEGF expression (rs = 0.714, P<0.05). We did not see gastrointestinal complications in the treated group and no differences were noted in the body weight of the mice between the two groups throughout the study CONCLUSION: Indomethacin can significantly decrease the viability of cultured HCT116 cells and retard human colorectal HCT116 cell tumor growth via inhibiting tumor angiogenesis, which might be through reduction of VEGF expression.

Anticancer effects of oligomeric proanthocyanidins on human colorectal cancer cell line, SNU-C4

AIM: Oligomeric proanthocyanidins (OPC), natural polyphenolic compounds found in plants, are known to have antioxidant and anti-cancer effects. We investigated whether the anti-cancer effects of the OPC are induced by apoptosis on human colorectal cancer cell line, SNU-C4.METHODS: Colorectal cancer cell line, SNU-C4 was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum. The cytotoxic effect of OPC was assessed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenylt-etrazolium bromide (MTT) assay. To find out the apoptotic cell death, 4, 6-diamidino-2-phenylindole (DAPI) staining,terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, reverse transcriptionpolymerase chain reaction (RT-PCR), and caspase-3 enzyme assay were performed.RESULTS: In this study, cytotoxic effect of OPC on SNUC4 cells appeared in a dose-dependent manner. OPC treatment (100 μg/mL) revealed typical morphological apoptotic features. Additionally OPC treatment (100 μg/mL)increased level of BAX and CASPASE-3, and decreased level of BCL-2 mRNA expression. Caspase-3 enzyme activity was also significantly increased by treatment of OPC (100 μg/mL) compared with control.CONCLUSION: These data indicate that OPC caused cell death by apoptosis through caspase pathways on human colorectal cancer cell line, SNU-C4.

Laparoscopic versus open right hemicolectomy with curative intent for colon carcinoma

AIM: Laparoscopic surgery, especially laparoscopic rectal surgery, for colorectal cancer has been developed considerably. However, due to relatively complicated anatomy and high requirements for surgery techniques, laparoscopic right colectomy develops relatively slowly. This study was designed to compare the outcomes of laparoscopic right hemicolectomy (LRH) with open right hemicolectomy (ORH) in the treatment of colon carcinoma. METHODS: Between September 2000 and February 2003, 30 patients with colon cancer who underwent LRH were compared with 34 controls treated by ORH in the same period. All patients were evaluated with respect to surgery related complications, postoperative recovery, recurrence and metastasis rate, cost-effectiveness and survival. RESULTS: Among 30 LRH, 2 (6.7%) were converted to open procedure. No significant differences were observed in terms of mean operation time, blood loss, post-operative complications, and hospital cost between LRH and ORH groups. Mean time for bowel movement, hospital stay, and time to resum?early activity in the LRH group were significantly shorter than those in the ORH group (2.24±0.56 vs 3.25±1.29 d, 13.94?.5 vs 18.25±5.96 d, 3.94±1.64 vs 5.45±1.82 d respectively, P<0.05). As to the lymph node yield, the specimen length and total cost for operation and drugs, there was no significant difference between the two groups. Local recurrence rate and metachronous metastasis rate had no marked difference between the two groups. Cumulative survival probability at 40 mo in LRH group (76.50%) was not obviously different compared to the ORH group (74.04%). CONCLUSION: LRH in patients with colon cancer has statistically and clinically significant advantages over ORH. Thus, LRH can be regarded as a safe and effective procedure.

Formation of Nodular Structures and Nitrogen Fixation by Rhizobia on Oilseed Rape Roots Following Treatment with Pectinolytic Bacteria

Nodular structures were formed by rhizobia on oilseed rape roots following treatment with pectinolytic bacteria. Nodules developed within 50 days.Photomicrograph of nodule cells showed that the capsulated bacteria were intrarellular.Rhizobia reisolated from the root nodules retained not only the ability of nodulation but also the characteristic of resistance to 100 μg neomycin mL-1.A low nitrogenaSe activity of the nodules was determined by the method of acetylene reduction.

Expression pattern of epithelial cell adhesion molecule on normal and malignant colon tissues

AEM: To investigate the expression pattern of epithelial cell adhesion molecule (Ep-CAM) on normal and malignant colon tissues to evaluate its diagnostic and therapeutic significance. METHODS: cDNA encoding Ep-CAM extracellular domain was doned by reverse transcription-polymerase chain reaction (RT-PCR) from excised malignant colon tissues and inserted into a glutathione S-transferase (GST)-tagged vector. Ep-CAM-GST fusion protein was induced by isopropyl-p-D-thiogalactopyranoside (IPTG) and purified with glutathione-sepharose. The Ep-CAM-GST fusion protein was mixed with Freund's adjuvant and Balb/c mice were immunized with it. Sp2/0 myeloma cells were fused with the spleen cells of the immunized mice. After having selected by indirect ELISA, the anti-Ep-CAM monoclonal antibodies (MAbs) were generaled and the corresponding ascites were obtained. Finally, the human colon carcinoma tissue array prepared from seventy individual patients was stained with the anti-Ep-CAM MAbs. RESULTS: The isdated Ep-CAM cDNA sequence was identical to the data in GenBank. The expressed fusion protein was almost soluble and had a molecular weight (MW) of 53 ku. Four MAbs against Ep-CAM were obtained and designated as FMU-Epl, FMU-Ep2, FMU-Ep3 and FMU-Ep4 respectively. Among them, FMU-Ep4 could recognize the natural Ep-CAM on Colo205 and SW480 cells, and all of them could be used for immunohistochemical staining of tissue sections. It was fbund that Ep-CAM was distributed differently in normal and various malignant colon tissues, induding squamous cell carcinoma, signet-ring cell carcinoma and adenocarcinoma. In normal colon gland epithelia, Ep-CAM antigen was mainly distributed on the basolateral membrane and in the region between the basolateral membrane and the cytoplastic part near the nuclei, whereas the expression pattern of colon malignancies was mainly on the whole surface of epithelia and the expression was much higher than the normal colon tissues. The staining pattern of tissue array showed in adenocarcinoma and papillary adenocarcinoma, and the expression of Ep-CAM was increased from grade I to grade Ⅲ. CONCLUSION: MAbs against Ep-CAM might be useful for research on the structure and function of Ep-CAM and may have diagnostic and therapeutic value to various colon carcinomas.

不要轻信微生物肥料的夸大宣传——忠告广大农民朋友

随着我国化肥工业的发展,近几年微生物肥料、微量元素肥料、植物生长调节剂等一批新型肥料随之兴起。特别是微生物肥料在一些乡镇企业发展很快,推广应用面积不断扩大,并被炒得很热。

Effect of bone marrow-derived monocytes transfected with RNA of mouse colon carcinoma on specific antitumor immunity

AIM: To investigate the effect of bone marrow-derived monocytes transfected with RNA of CT-26 (a cell line of mouse colon carcinoma) on antitumor immunity. METHODS: Mouse bone marrow-derived monocytes were incubated with mouse granulocyte macrophage colony stimulating factor (mGM-CSF) in vitro, and the purity of monocytes was detected by flow cytometry. Total RNA of CT-26 was obtained by TRIzol's process, and monocytes were transfected by TransMessenger in vitro. The activity of cytotoxic T lymphocytes (CTL) in vivo was estimated by the modified lactate dehydrogenase (LDH) release assay. Changes of tumor size in mice and animal's survival time were observed in different groups. RESULTS: Monocytes from mouse bone marrow were successfully incubated, and the positive rate of CDllb was over 95%. Vaccination of the monocytes transfected with total RNA induced a high level of specific CTL activity in vivo, and made mice resistant to the subsequent challenge of parental tumor cells. In vivo effects induced by monocytes transfected with total RNA were stronger than those induced by monocytes pulsed with tumor cell lysates. CONCLUSION: Antigen presenting cells transfected with total RNA of CT-26 can present endogenous? tumor antigens, activate CTL, and effectively induce specific antitumor immunity.