荠菜属CaCA家族基因拷贝数变异与表达分化的研究

荠菜属植物是十字花科中分布较广泛的植物,其中四倍体荠菜(Capsella bursa-pastoris)是一种由祖先二倍体亲本杂交再经多倍化后形成的多倍体物种,在地域分布及多样化的逆境适应能力方面与其祖先亲本相比具有明显优势,目前四倍体荠菜对于环境适应性的研究报道还不多见.植物钙/阳离子逆转运蛋白超家族(CaCAs)是一类膜定位的离子转运蛋白,在植物抗冷、抗盐等信号通路中起到重要作用.本研究通过生物信息学及表达谱分析手段,发现四倍体荠菜CaCAs的13对亚基因组基因中有4对出现了拷贝数变异,其中3对基因在其编码的氨基酸序列和蛋白质跨膜结构方面与其直系同源基因间存在较大差异,且在冷响应表达上与拟南芥相比出现了明显偏倚.此外,与四倍体荠菜的祖先亲本Capsella rubella相比,四倍体荠菜CaCAs基因的组织表达分化较为普遍.CaCAs基因在四倍体荠菜中的拷贝数变异和表达分化现象提示了CaCAs对植物环境适应性的贡献.

帕金森病人血清分化型胚胎软骨表达基因1和肝X受体β水平的相关性研究

目的:探讨帕金森病(PD)病人血清中分化型胚胎软骨表达基因1(DEC1)和肝X受体β(LXRβ)水平及二者相关性。方法:选择PD病人50例(PD组),选择同期健康人群50名为对照(对照组)。检测2组受试者血清DEC1、LXRβ水平。结果:PD组血清DEC1水平明显高于对照组(P<0.01),LXRβ水平明显低于对照组(P<0.01)。不同性别的PD病人DEC1和LXRβ水平差异均无统计学意义(P>0.05);年龄≥60岁PD病人的LXRβ水平低于<60岁者(P<0.05);有认知功能障碍的PD病人DEC1和LXRβ水平分别明显高于和低于无认知障碍者(P<0.01);随病情分期加重,PD病人的DEC1水平逐渐升高,LXRβ水平逐渐降低(P<0.05~P<0.01)。PD病人血清DEC1水平与LXRβ水平呈负相关关系(r=-0.326,P<0.05)。结论:PD病人血清DEC1与LXRβ水平呈负相关关系,二者与PD的发病及病情进展有关。

诱导分化剂联合化疗药治疗可移植性人脑胶质瘤的实验研究

背景与目的:丙戊酸钠(valproate,VPA)不仅是经典的临床抗癫痫药物,近年来还发现其对多种肿瘤细胞都有一定的诱导分化作用,但单独应用或与化疗药联合应用对脑胶质瘤体内疗效如何尚不确定。本研究旨在探讨两者抗胶质瘤的联合效应。方法:将低分化人脑胶质瘤体外细胞系SHG-44移植于裸小鼠制作成可移植性实体瘤模型后,单用VPA以及联合应用盐酸尼莫司汀(nimustine,ACNU)进行治疗,观察移植瘤体积、病理学形态、细胞增殖周期、分化抗原表达等指标的变化。结果:VPA和ACNU联合用药组的疗效明显优于任何一种单独给药组,表现为肿瘤生长抑制,细胞异型性降低,G0/G1期细胞比例增加,胶原纤维酸性蛋白表达上调。结论: VPA联合应用化疗药能够明显增强VPA抑制人脑胶质瘤生长和诱导其向良性分化的作用。

4种作物果糖激酶基因的鉴定与比较分析

【目的】基于全基因组序列,鉴定与分析水稻、玉米、高粱和谷子4种作物的果糖激酶(fructokinase, FRK)家族基因,明确其复制起源方式、系统发生关系、序列及表达分化特征等,为深入开展FRK基因功能研究、改良作物产量和品质等提供数据。【方法】采用基因组搜索、基因组内及组间比较等多种方法,分析4种作物FRK家族成员的基因结构、保守基序、系统进化、复制扩张和基因表达分化等。【结果】鉴定到水稻13个、玉米16个、高粱14个、谷子16个FRK基因家族成员,其中水稻11个、玉米7个、高粱12个、谷子13个基因为离散复制基因,水稻、高粱和谷子各有1对ρWGD基因,玉米有8个WGD基因(1个ρWGD基因,7个mWGD基因)。该家族成员亚家族间基因结构差异大,但在亚家族内具有保守性。基因表达模式分析发现,直系同源基因间的表达模式相似,ρWGD基因对发生了明显的表达分化,而玉米物种特异的mWGD基因对的表达差异较小。【结论】探明了4种作物基因组包含的FRK家族基因,揭示了其复制起源、基因结构、序列及表达模式的演化特征。

SPP1、DEC1、C1QTNF6蛋白与口腔鳞状细胞癌患者临床病理指标及预后的关系

目的:探讨口腔鳞状细胞癌患者血清重组人分泌型磷蛋白1(SPP1)、软骨分化的表达基因1(DEC1)和补体C1q/肿瘤坏死因子相关蛋白6(C1QTNF6)的表达水平与其临床病理指标及预后的关系。方法:免疫组织化学染色法和电化学发光免疫分析法检测88例口腔鳞状细胞癌患者癌组织和血清中SPP1、DEC1和C1QTNF6蛋白的表达水平;Pearson相关性分析和Kaplan-Meier生存分析法分析患者血清SPP1、DEC1和C1QTNF6蛋白表达水平与其临床病理指标的相关性和对患者预后的影响。多因素Cox回归法分析影响口腔鳞状细胞癌患者预后的危险因素。结果:免疫组织化学染色结果显示口腔鳞状细胞癌患者癌组织中SPP1、DEC1和C1QTNF6蛋白的阳性表达率较癌旁组织分别增加了1.94、2.98和2.35倍(P<0.05或P<0.01);电化学发光免疫分析法结果显示口腔鳞状细胞癌患者血清SPP1、DEC1和C1QTNF6蛋白表达水平较正常人分别上调了8.61、6.20和4.03倍(P<0.05或P<0.01);Pearson相关性分析结果显示患者血清SPP1、DEC1和C1QTNF6蛋白表达水平与患者发生多灶嗜神经侵犯、肿瘤浸润程度、淋巴结转移、较高的TNM分期呈正相关(P<0.05);Kaplan-Meier生存分析结果显示,血清SPP1、DEC1和C1QTNF6蛋白高表达水平的口腔鳞状细胞癌患者总生存率低;多因素Cox回归分析结果表明多灶嗜神经侵犯、肿瘤高浸润度、淋巴结转移和高的TNM分期是影响预后的危险因素(P<0.05)。结论:口腔鳞状细胞癌患者癌组织和血清中SPP1、DEC1和C1QTNF6蛋白表达水平升高,且与患者发生多灶嗜神经侵犯、肿瘤浸润和淋巴结转移、较高的TNM分期呈正相关,而与患者预后呈负相关。

Pilot Study of Molecular Mechanism on Vasculogenic Mimicry in Bi-directional Differentiated Malignant Tumors

Objective: To investigate the role of collagen IV and PAS positive substancesecreted by tumor cells in vasculogenic mimicry (VM) and the effects of VM on tumor cells expressingVEGF. Methods: 158 cases of bi-direction differential malignant tumor specimens withparaffin-embedded were enrolled into our study and made tissue microarray which were dual-stainedwith CD31-PAS and stained with collagen IV. The difference of the areas and distribution withpattern surrounded by between CD31 and PAS positive respectively were identified via grid-counting,as well as the difference of VEGF expression with VE absent and present. Results: The basementmembrane of VM was both PAS and collagen IV positive. VEGF expression in the bi-directiondifferential malignant tumor was higher VM-absent than VM-present and the difference wasstatistically significance in malignant melanoma and alveolar rhabdomyosarcoma (P 【 0.05).Conclusion: PAS positive substance and collagen IV compose the wall of VE and VE could provide theoxygen and nutrition for tumor growth and progression.

急性ST段抬高型心肌梗死PCI术后血小板CD40L及血小板指标临床相关性分析

目的 探讨急性ST段抬高型心肌梗死（STEMI）患者经皮冠状动脉介入治疗（PCI）术后血小板CID40L及血小板指标变化的临床意义.方法 选取河北大学附属医院2013年4月至2014年4月接诊的急性ST段抬高型心肌梗死行PCI术患者63例为研究组;另选同期行健康体检者50名为对照组.两组皆采取流式细胞技术进行血小板CD40L检测,并测定血小板计数（PLT）、血小板体积分布宽度（PDW）、血小板平均体积（MPV）、血小板压积（PCT）、肌钙蛋白Ⅰ（Tn-Ⅰ）等,应用Person相关分析法分析血小板CD40L及血小板指标临床相关性,其中研究组检测时间包括术前与术后,而对照组健康者则仅为入组时.结果 ①研究组术前与对照组入组时比较,CD40L及Tn-Ⅰ明显升高（P＜0.05）.②研究组患者术后与对照组入组时比较,CD40L明显升高[（8.97±0.73）MFI比（0.80±0.27）MFI],差异有统计学意义（P.＜0.05）.研究组患者术后Tn-Ⅰ明显高于对照组入组时[（18.74±2.65） μg/L比（0.010±0.008）μg/L,P＜0.05];MPV、PDW数值高于对照组入组时（P＜0.05）;而两组患者PLT、PCT未见统计学差异（P＞0.05）.③研究组患者术后CD40L、Tn-Ⅰ、MPV、PDW较术前明显升高.④PCI术后患者血小板CD40L水平与MPV及PDW呈正相关（P＜0.05）.结论 急性ST段抬高型心肌梗死行PCI患者的血小板形态改变属于血小板活化的主要特点,而血小板指标可能对血小板CD40L的表达产生影响,从而引发凝血及炎症反应,应加强重视.

Expression and Phylogenetic Analysis of Pea Actin Isoforms

Pea ( Pisum sativum Linn.) actin gene family contains at least three isoforms (PEAcⅠ, PEAcⅡand PEAcⅢ), and the DNA sequence of these isoforms show high similarity in the coding regions and significant divergence in the untranslated regions. RT_PCR and Southern blotting using 3′_untranslated region (3′_UTR) as specific probe revealed that pea isoactin genes were expressed in roots, stems, leaves, tendrils, pollen and juvenile siliques, but displayed different patterns of transcript accumulation. Two_fold serial dilution electrophoresis showed PEAcⅠ mRNA preferentially accumulated in rapidly developing tissues: it peaked in seven days' stem; remained at a rather high level in leaves within a month but decreased significantly later; varied a little in tendrils and reached a median and a very low level respectively in juvenile siliques and in pollen. PEAcⅡ displayed somewhat similar expression pattern to PEAcⅠ. The observed differences in sequences and transcript accumulation patterns suggest that the individual pea actin genes may differ in their transcriptional regulation and cellular function. Phylogenetic tree of actins showed that pea actin isoforms are as diverged from each other as they are from other plant actins, and pea actins might have originated from a common ancestor before the divergence of the dicot and monocot plants.

Cloning and Differential Gene Expression of Two Catalases in Suaeda salsa in Response to Salt Stress

Two different cDNA clones (Sscat1 and Sscat2) encoding catalase, the primary important H2O2-scavenging enzyme, were isolated from a AZap-cDNA library constructed from a 400 mmol/L NaCl-treated library of Suaeda salsa ( L.) Pall aerial tissue. Sscat1 (1.7 kb) contains a full open reading frame of 492 amino acids and Sscat2 (1.1 kb) is a partial clone. BLAST analysis indicates that the two clones share 71.9% identity in nucleotide sequence and 75% identity in deduced amino acid sequence within the last 287 amino acid residues of Sscat1. Southern blotting analysis showed that Sscat1 is multicopy in S. salsa genome, while Sscat2 is a single copy gene. Northern blotting analysis showed a rapid increase in the steady-level of both genes in roots after 48 It salt treatment, but only Sscat1 was induced in salinity treated leaves. Time-course analysis carried out in leaves confirmed that Sscat1 was induced by salt stress, in contrast to Sscat2. These implied that the expression of Sscat1 and Sscat2 genes are differentially regulated in S. salsa. The activity of total catalase is dramatically increased in response to salt stress.

Photooxidation in Leaves of Facultative CAM Plant Sedum spectabile at C_3 and CAM Mode

The switch from C-3 to CAM pathway was induced by water stress in a C-3/CAM intermediate plant Sedum spectabile Boreau. Typical CAM criteria were observed upon 15 d of withholding water. Leaf delta(13)C value (-%) and water content showed a linear correlation fashion. Chlorophyll fluorescence parameters and antioxidative capacity were altered by water stress. Phi(PSII) and q(P) were reduced by 50% and 34% of the control, respectively, while NPQ rose ca. 180%. SOD activity and ability to scavenge DPPH. free radical went down but membrane permeability changed slightly. However, when an additional photooxidation by MV with high PPFD was carried out with leaf discs from watered (C-3 mode) and drought plants (CAM mode), q(P) and Phi(PSII) in leaves at induced CAM mode stage continuously decreased to a very low level. High 1 - q(P) value (0.86) and 1 - q(P)/NPQ ratio (>1) indicated the presence of high reduction state and unbalance of light energy budget. Together with the marked loss of membrane integral, it was evidenced that photooxidative damage was more serious in the induced CAM mode than in the C-3 mode. No advantage of photooxidation tolerance was found at the induced CAM expression stage of the facultative CAM plant, as compared with its C-3 mode stage, and also with the constitutive CAM plants reported previously. The differences in photooxidation sensitivity between the inducible CAM expressing plant and the constitutive CAM plant were discussed.

玉柏石松中甾体化合物对体外培养成骨细胞活性的影响

为研究玉柏石松中甾体化合物豆甾烷-3-酮-21-羧酸(SA)对体外培养小鼠成骨细胞系MC3T3-E1活性的影响,用Alamar Blue法检测了成骨细胞增殖率,碱性磷酸酶试剂盒检测了细胞中碱性磷酸酶活性,茜素红染色检测了成骨细胞矿化水平,荧光定量PCR检测了成骨细胞骨分化相关基因的表达.结果显示:8μmol/L和16μmol/L的SA处理细胞8 d能抑制成骨细胞碱性磷酸活性;处理细胞16 d能提高骨细胞矿化水平.SA抑制成骨早期分化相关基因(Runx-2和Osterix)的表达,促进骨基质蛋白OPN和骨重建相关转录因子(Jun-D,Fra-1和Fra-2)的表达.故SA具有促进骨折愈合的成骨活性,可能通过促进相关转录因子表达,骨折断面旧骨的吸收和骨基质钙化等方式完成.

大规模原位杂交筛选爪蟾发育调控基因

应用半自动化的系统表达筛选 ,研究了 2 4 0 0个爪蟾EST克隆在胚胎发育中的表达模式 ,从筛选出的 62 4个分化表达克隆中 。

Magnetic Resonance Imaging and Biological Markers in Pituitary Adenomas with Invasion of the Cavernous Sinus Space

Objective: To investigate the predictability of MRI and the possiblebiological markers of cavernous sinus invasion of pituitary adenomas associated with fourphenomenas: angiogenesis, cell proliferation, apoptosis and matrix metalloproteinase. Methods: Weevaluated 45 patients with pituitary adenoma according to the MRI, surgical findings and theimmunohistochemistry staining of tumor tissues. Results: The results have shown that the sensitivityof MRI for predicting cavernous sinus invasion in this prospective study was 60%, its specificity85%, its positive predictive value 83.33%, negative predictive value 62.96%. 45 specimens ofpituitary adenomas were analyzed for expression of F8, VEGF, Ki-67, c-myc, Bcl-2, nm23 and MMP-9immunoreactivity using immunoperoxidase staining. MVD was assessed using F8-related antigen. Theresults have shown that MVD of invasive pituitary adenomas was significantly higher than that ofnoninvasive (P 【 0.001). There was an association between the invasion of pituitary adenomas andKi-67 LI (P = 0.039) or the expression of VEGF (P 【 0.001) and MMP-9 (P 【 0.001). But c-myc LI andBcl-2 expression have no association with invasiveness of pituitary adenomas (P = 0.061 versus P =0.201). On the other hand, there is an inverse relationship between nm23 expression and tumorinvasion (P 【 0.001). Conclusion: Parasellar extension of pituitary adenomas through the medial wallof the cavernous sinus is diagnosed at surgery, and with sensitive gadolinium-enhanced MRI, itsextent can be partly determined by radiology. Although our study has shown that MVD and theexpression of VEGF, Ki-67, nm23 and MMP-9 have associations with invasiveness of pituitary adenomas,they are lack of specificity. These markers can only provide some useful information.

mRNA expression and DNA methylation in three key genes involved in caste differentiation in female honeybees(Apis mellifera)

In honeybee （Apis mellifera） colonies, queens and workers are altemative forms of the adult female honeybee that develop from genetically identical zygotes but that depend on differential nourishment. Queens and workers display distinct morphologies, anatomies and behavior, better known as caste differentiation. Despite some basic insights, the exact mechanism responsible for this phenomenon, especially at the molecular level, remains unclear although some progress has been achieved. In this study, we examined mRNA levels of the TOR （target of rapamycin） and Dnmt3 （DNA methyltransferase 3） genes, closely related to caste differentiation in honeybees. We also investigated mRNA expression of the S6K （similar to RPS6-p70-protein kinase） gene linked closely to organismal growth and development in queen and worker larvae （1-day and 3-day old）. Last, we investigated the methylation status of these three genes in corresponding castes. We found no difference in mRNA expression for the three genes between 1st instar queen and worker larvae; however, 3rd instar queen larvae had a higher level of TOR mRNA than worker larvae. Methylation levels of all three genes were lower in queen larvae than worker larvae but the differences were not statistically significant. These findings provide basic data for broadening our understanding of caste differentiation in female honeybees.

Correlation between expression and differentiation of endocan in colorectal cancer

AIM: To investigate the expression frequency of endocan in colorectal cancer and analyze the relationship between endocan expression and clinical parameters and to study the role of endocan in colorectal carcinogenesis. METHODS: Expression of endocan in 72 tumor tissue samples of colorectal cancer as well as in 27 normal mucous membrane tissue samples was analyzed using in situ hybridization, immunohistochemistry on tissue microarray, Western blot and reverse-transcript polymerase chain reaction (RT-PCR). RESULTS: The expression of endocan was higher in normal colon and rectum tissue samples than in cancerous tissue samples (mRNA = 92.6%, protein = 36%), and was lower in colorectal cancer tissue samples (mRNA = 70.4%, protein = 36.1%). No correlation was found between staining intensity and clinical parameters such as sex, age, tumor size andTNM stage. However, the expression of endocan was positively correlated with the tissue differentiation in colorectal cancer. CONCLUSION: The expression of endocan is down- regulated in colorectal cancer and is positively correlated with the tissue differentiation in colorectal cancer, suggesting that the expression of endocan is associated with development and differentiation of colorectal cancer.

Molecular Characterization of Four ADF Genes Differentially Expressed in Cotton

Actin depolymerizing factor （ADF）, highly conserved in all eukaryotic cells, is a low molecular mass of actin-binding protein, which plays a key role in modulating the polymerizing and depolymerizing of the actin filaments. Four cDNAs （designated GhADF2, GhADF3, GhADF4, and GhADF5, respectively） encoding ADF proteins were isolated from cotton （Gossypium hirsutum） fiber cDNA library. GhADF2 cDNA is 705 bp in length and deduces a protein with 139 amino acids. GhADF3 cDNA is 819 bp in length and encodes a protein of 139 amino acids. GhADF4 cDNA is 804 bp in length and deduces a protein with 143 amino acids. GhADF5 cDNA is 644 bp in length and encodes a protein of 141 amino acids. The molecular evolutionary relationship of these genes was analyzed by means of bioinformatics. GhADF2 is closely related to GhADF3 （99% identity） and PetADF2 （89% identity）. GhADF4 is closely related to AtADF6 （78% identity）, and GhADF5 is closely related to AtADF5 （83% identity）. These results demonstrated that the plant ADF genes are highly conserved in structure. RT-PCR analysis showed that GhADF2 is predominantly expressed in fiber, whereas, GhADF5 is mainly expressed in cotyledons. On the other hand, it seems that GhADF3 and GhADF4 have no tissue specificity. Expression levels of different ADF genes may vary considerably in the same cell type, suggesting that they might be involved in regulating tissue development of cotton and the each ADF isoform may diverge to form the functional difference from the other ADFs during evolution.

Higher CO_2-insufflation pressure inhibits the expression of adhesion molecules and the invasion potential of colon cancer cells

AIM： To investigate the influence of CO2-insufflation pressure on adhesion, invasion and metastatic potential of colon cancer cells based on adhesion molecules expression. METHODS： With an/n vitro artificial pneumoperitoneum model, SW1116 human colon carcinoma cells were exposed to CO2-insufflation in 5 different pressure groups： 6 mmHg, 9 mmHg, 12 mmHg, 15 mmHg and control group, respectively for 1 h. Expression of E-cadherin, ICAM-I, CD44 and E-selectin was meas- ured at 0, 12, 24, 48 and 72 h after CO2-insufflation using flow cytometry. The adhesion and invasion capacity of SW1116 cells before and after exposure to CO2-insufflation was detected by cell adhesion/invasion assay in vitro. Each group of cells was injected intraperitoneally into 16 BALB/C mice. The number of visible abdominal cavity tumor nodules, visceral metas-tases and survival of the mice were recorded in each group. RESULTS： The expression of E-cadherin, ICAM-1, CD44 and E-selectin in SWl116 cells were changed significantly following exposure to CO2 insufflation at different pressures （P 〈 0.05）. The expression of E-cadherin, CD44 and ICAM-1 decreased with increasing CO2-insufflation pressure. The adhesive/ invasive cells also decreased gradually with increasing pressure as determined by the adhesion/invasion assay. In animal experiments, the number of abdominal cavity tumor nodules in the 15 mmHg group was also significantly lower than that in the 6 mmHg group （29.7± 9.91 vs 41.7±14.90, P = 0.046）. However, the survival in each group was not statistically different. CONCLUSION： CO2-insufflation induced a temporary change in the adhesion and invasion capacity of cancer cells in vitro. Higher CO2-insufflation pressure inhibited adhesion, invasion and metastatic potential in vitro and in vivo, which was associated with reduced expression of adhesion molecules.

Efficient generation of hepatocyte-like cells from human induced pluripotent stem cells

Human induced pluripotent stem （iPS） cells are similar to embryonic stem （ES） cells, and can proliferate intensively and differentiate into a variety of cell types. However, the hepatic differentiation of human iPS cells has not yet been reported. In this report, human iPS cells were induced to differentiate into hepatic cells by a stepwise protocol. The expression of liver cell markers and liver-related functions of the human iPS cell-derived cells were monitored and compared with that of differentiated human ES cells and primary human hepatocytes. Approximately 60% of the differentiated human iPS cells at day 7 expressed hepatic markers alpha fetoprotein and Alb. The differentiated cells at day 21 exhibited liver cell functions including albumin Asecretion, glycogen synthesis, urea production and inducible cytochrome P450 activity. The expression of hepatic markers and fiver-related functions of the iPS cellderived hepatic ceils were comparable to that of the human ES cell-derived hepatic cells. These results show that human iPS cells, which are similar to human ES cells, can be efficiently induced to differentiate into hepatocyte-like cells.

Reference Gene Selection for Normalization of PCR Analysis in Chicken Embryo Fibroblast Infected with H5N1 AIV

Chicken embryo fibroblasts (CEFs) are among the most commonly used cells for the study of interactions between chicken hosts and H5N1 avian influenza virus (AIV).In this study,the expression of eleven housekeeping genes typically used for the normalization of quantitative real-time PCR (QPCR) analysis in mammals were compared in CEFs infected with H5N1 AIV to determine the most reliable reference genes in this system.CEFs cultured from 10-day-old SPF chicken embryos were infected with 100 TCID50 of H5N1 AIV and harvested at 3,12,24 and 30 hours post-infection.The expression levels of the eleven reference genes in infected and uninfected CEFs were determined by real-time PCR.Based on expression stability and expression levels,our data suggest that the ribosomal protein L4 (RPL4) and tyrosine 3-monooxygenase tryptophan 5-monooxygenase activation protein zeta polypeptide (YWHAZ) are the best reference genes to use in the study of host cell response to H5N1 AIV infection.However,for the study of replication levels of H5N1 AIV in CEFs,the β-actin gene (ACTB) and the ribosomal protein L4 (RPL4) gene are the best references.