SE-HOXC13-AS对胰腺癌生物学行为的影响及其临床意义

目的 探讨超级增强子(super enhancers, SE)-HOXC13-AS对胰腺癌生物学行为的影响及临床意义。方法 采用qRT-PCR法检测HOXC13-AS在胰腺癌组织和细胞中的表达,分析HOXC13-AS表达与胰腺癌临床病理特征的关系。选择siRNA转染HOXC13-AS表达量最高的PANC-1细胞,应用CCK-8法、细胞周期实验、划痕实验、Transwell实验及细胞凋亡实验检测敲低HOXC13-AS对胰腺癌细胞增殖、迁移、侵袭及凋亡的影响。应用生物数据库分析和JQ1抑制剂验证SE与HOXC13-AS之间调控关系,并通过CCK-8法、划痕实验、Transwell实验及细胞凋亡实验检测抑制SE-HOXC13-AS对胰腺癌细胞增殖、迁移、侵袭及凋亡的影响。结果 胰腺癌组织中HOXC13-AS平均表达量(4.40±6.21)显著高于癌旁组织(1.01±0.02,P<0.05),HOXC13-AS表达与胰腺癌分化程度和临床分期密切相关。与正常胰腺导管上皮细胞(0.93±0.11)相比,胰腺癌细胞株BXPC-3(2.55±0.19)、SW1990(5.49±0.92)、CF-...     

地高辛标记的大鼠p75 RNA探针的制备和应用

目的制备用地高辛标记的神经生长因子低亲和力受体(p75)的RNA探针.研究p75在海马组织中的表达.方法设计p75引物,构建p75/pGEM-T重组质粒,分别用ApaⅠ和Sac Ⅰ进行酶切得到线性化DNA片段,以Sp6和T7聚合酶转录合成酶合成地高辛标记的(dig-)正反义RNA探针.运用点膜杂交的方法检验探针的敏感度,运用该探针,通过原位杂交分析p75在海马中的表达.结果构建了p75/pGEM-T质粒,获得高效价的正、反义dig-p75 RNA探针,应用该探针发现p75 mRNA在海马中的表达.结论成功制备了地高辛标记的p75RNA探针,为进一步研究p75在海马中发育和损伤过程中的表达打下基础.     

神经营养素受体p75NTR介导的信号转导

神经营养素 (neurotrophin, NT)受体具有低亲和力的Mr为 75 000NT受体(75KDNTreceptor, p75NTR)和高亲和力的原肌球蛋白受体激酶(tropomyosinreceptorkinase, Trk)受体家族两类。二者均为跨膜蛋白受体, 参与调节以神经元为主的某些细胞的生长、分化、存活、修复以及凋亡等多种生物学效应。研究表明, 二者间的信号转导及生物学效应既相互协同又彼此拮抗, 既紧密联系又有显著区别。Trk受体一般介导“正性”信号, 如促进神经元生长、维持其存活; 而p75NTR则具有多种生物学效应, 除可促进神经元存活、生长外, 还可诱导神经元凋亡、抑…     

CXCL12／CXCR4对胰腺癌细胞生物学行为的影响

目的探讨CXCLl2／CXCR4生物轴对胰腺癌细胞增殖、侵袭等生物学行为的影响。方法体外培养胰腺癌细胞系Miapaca-2,将其分为对照组、CXCLl2组和AMD3100组。（1）采用RT—PCR检测胰腺癌细胞中CXCLl2、CXCR4、基质金属蛋白酶-2（MMP-2）、基质金属蛋白酶-9（MMP-9）和人尿激酶型纤溶酶原激活物（uPA）mRNA的表达水平;（2）采用CCK-8法检测各组细胞的增殖情况;（3）采用Transwell侵袭实验检测CXCLl2／CXCR4对胰腺癌细胞趋化活性的影响。结果胰腺癌细胞系Miapaca-2中CXCLl2mRNA未见表达,而CXCR4mRNA在胰腺癌细胞中有表达。MMP-2、MMPO和uPAmRNA在AMD3100组、对照组和CXCLl2组中的表达水平呈递增趋势,差异具有统计学意义（P〈0．05）。胰腺癌细胞的增殖和侵袭能力在CXCLl2组明显增强,而在AMD3100组得到了有效的抑制,组间差异有统计学意义（P〈0．05）。结论趋化因子CXCLl2及其受体CXCR4所构成的生物轴对胰腺癌细胞的增殖和侵袭能力发挥着重要的作用。     

Ezrin蛋白过表达对胰腺癌细胞系Panc-1生物学行为的影响

目的探讨Ezrin过表达对胰腺癌细胞系Panc-1生物学行为的影响。方法用过表达载体pcb6-Ezrin稳定转染Panc-1细胞,并用流式细胞仪检测细胞周期,CCK-8法检测细胞生长曲线,扫描电镜观察细胞表面形态及表面突起的变化,用未包被及包被Matrigel胶的Transwell小室分别检测细胞的运动和侵袭能力,并用免疫印迹法检测信号相关蛋白Erk1/2的变化。结果过表达Ezrin后其蛋白的表达明显升高,Panc-1细胞表面的细胞突起、微绒毛数量明显增加,运动及侵袭能力也明显增加,但对细胞的体外增殖和细胞周期无明显的影响。Ezrin的过表达能够激活Erk1/2的表达。结论 Ezrin蛋白对胰腺癌细胞的细胞突起、表面微绒毛的形成、细胞骨架及细胞的运动和侵袭发挥着重要的作用。因此,Ezrin可能在胰腺癌的进展中发挥着重要的作用,Erk1/2信号转导途径在这一过程中发挥着重要的作用。     

P162对食管癌细胞株Eca109的放射增敏作用及其对p75NTR表达的影响

 目的：研究靶向Ras-GTP酶激活蛋白SH3功能区结合蛋白(G3BP)的新药P162对人食管癌细胞株Eca109的放射增敏作用及其对p75神经营养因子受体(p75NTR)表达的影响。方法：CCK-8法检测P162对食管癌细胞株Eca109增殖抑制的影响;集落形成实验检测P162对Eca109细胞的放射增敏效应,单击多靶模型拟合细胞存活曲线并计算放射增敏比;倒置显微镜观察细胞形态学改变;流式细胞术检测p75NTR的表达。结果：P162对食管癌细胞株Eca109有增殖抑制作用,且呈时间和剂量依赖性,2.5、5.0、10 μmol/L P162对Eca109细胞的放射增敏比分别为1.54、2.35和2.33。随着照射剂量的增加,食管癌细胞中p75NTR的表达增加,经5 μmol/L P162处理的实验组中p75NTR的表达明显低于未经处理的对照组。结论：P162对Eca109细胞有放射增敏作用,并且能抑制食管癌干细胞p75NTR的表达。P162的增敏作用可能与抑制食管癌干细胞有关。     

寻常型银屑病患者皮损表皮TNF-α受体p75的表达及其意义

银屑病是一种具有遗传倾向的炎症性增殖性皮肤病 ,其发病机制尚未完全阐明 ,但其皮损表皮中有多种细胞因子包括TNF α高表达。TNF α与其细胞上的受体结合才能发挥其生物学效应 ,目前国内外尚未见有关银屑病皮损表皮p75mRNA表达与否的研究报道。为探讨TNF α受体p75在寻常型银屑病皮损表皮中的表达及其意义 ,我们采用免疫组化和RT PCR方法对进行期寻常型银屑病患者皮损表皮TNF α受体p75进行了检测。2 7例患者中男 18例 ,女 9例 ;年龄15岁～ 35岁 ;病程 1个月～ 7年。 16份正常对照皮肤取自美容整形医院。取典型皮损 2块 ,一块OCT…     

胰腺癌淋巴管生成及其与生物学行为关系

目的研究胰腺癌淋巴管生成的机制、分布特征及其与胰腺癌临床病理学的关系。方法应用免疫组化双标记方法检测42例胰腺癌和12例癌旁胰腺组织中血管内皮生长因子-C（VEGF—C）及肾小球足突细胞黏蛋白（Podoplanin）的表达。结果42例胰腺癌组织VEGF—C阳性率为61．9％,12例癌旁胰腺组织中VEGF—C阳性表达率为58．3％,两者差异无显著性（x^2=0．050,P〉0．05）。VEGF-C阳性率与胰腺癌的淋巴结转移有密切关系（x^2=4．822,P〈0．05）,而与胰腺癌大小、组织学分级、神经浸润及临床病理分期无关（P〉0．05）。胰腺癌组织中可见明确的Podoplanin阳性淋巴管,其数目和分布具有明确的异质性。肿瘤边缘组织中淋巴管数最多,其次为肿瘤表浅部,而肿瘤中心区最少;在形态上,肿瘤边缘可以见到较多管腔扩张的淋巴管,而肿瘤中心及表浅淋巴管多为闭锁的条索状或点状。癌旁胰腺中Podoplanin阳性淋巴管分布在呈慢性炎症的胰腺组织周围,多数呈扩张状态。42例胰腺癌组织中LVD为7．67±1．25,12例癌旁胰腺组织中LVD为7．85±0．93,二者差别无显著性（t=0．639,P〉0．05）。胰腺癌组织中LVD与淋巴结转移及临床病理分期有关（t=7．076,6．803,P〈0．01）,而与胰腺癌大小、组织学分级和神经浸润无关（P〉0．05）。胰腺癌组织中VEGF-C的表达与LVD间呈正相关性（r=0．509,P〈0．05）。结论胰腺癌及癌旁胰腺组织中VEGF—C高表达,促进淋巴管增生,可能是临床上胰腺癌早期发生淋巴结转移的重要机制之一。     

p75神经营养素受体介导的信号传递

p75神经营养素受体(p75~(NTR))除可增强神经营养素与Trks受体作用外,还可与神经营养素结合启动另外的信号传递通路。首先,活化的p75~(NTR)可激活胞内的转录因子NF-κB;其次,p75~(NTR)与神经营养素结合,可促进神经细胞内鞘磷脂的水解、神经酰胺的释放,导致细胞程序性死亡。此外,p75~(NTR)还表现出对配体的选择性,而且它与Trks之间存在着相互作用的关系。     

神经元的发育过程中p75NTR和sortilinr的表达研究

目的 神经元发育过程中需要p75NTR和sortilin的参与.方法 体外培养新生SD大鼠海马神经元,通过细胞免疫组织化学方法观察这一过程中p75NTR、sortilin的表达情况.结果 用出生0～1h新生SD大鼠培养海马神经元,培养3d的神经元经neun免疫组化染色鉴定,其阳性率可达到90％以上.观察发现体外培养的海马神经元生长有明显的阶段性.经细胞免疫组织化学染色发现,p75NTR、sortilin在神经元发育过程中均有表达.结论 体外培养的海马神经元生长有明显的阶段性,p75NTR、sortilin在神经元发育过程中均有表达.     

MiRNA-29-2-5p对人胰腺癌细胞SW1990迁移能力的影响

目的研究miR-29b-2-5p在胰腺癌细胞系SW1990中迁移的作用。方法荧光定量PCR方法检测miR-29b-2-5p在胰腺癌细胞系SW1990中的过表达情况,Transwell小室迁移实验和细胞划痕实验观察miR-29b-2-5p对胰腺癌细胞系SW1990的迁移能力的影响。结果在胰腺癌细胞系SW1990中过表达miR-29b-2-5p后,miR-29b-2-5p表达显著上调,胰腺癌细胞系的迁移能力明显下降。通过Targetscan软件预测及免疫蛋白印迹法检测发现miR-29b-2-5p的靶蛋白AKT表达水平明显降低。结论 miR-29b-2-5p显著抑制胰腺癌细胞的迁移能力可能与抑制AKT表达相关,为解明胰腺癌迁移机制及针对胰腺癌迁移的新靶点开发提供依据。     

Bone mesenchymal stem cells for gene transfer of NGF to the adult rat brain: rescue the NGFR p75 positive neurons from fimbria-fornix lesion-induced degeneration

The study was to evaluate the therapeutic benefit of transplanted bone mesenchymal stem cells (BMSCs) transfected never growth factor (NGF) gene and GFP gene (as a reporter gene), in treating the rat with fimbria-fornix lesion. After transduction of NGF gene via recombinant retroviral vectors into the rat BMSCs, BMSCs were therefore transformed into the GFP-NGF positive BMSCs, nearly 100% of BMSCs expressed NGF, and then transplanted into basal forebrain of rat with fimbria-fornix lesion. After 2 weeks post-transplantation, the GFP-NGF positive BMSCs survive and fuse in vivo with astroglia or NGFR p75 positive neurons in the basal forebrain, no evidence of transdifferentiation was observed in this study. The number of NGFR p75 positive neurons in basal forebrain of NGF group was significantly higher than those of the void plasmid group (p < 0.05) or the PBS group (p < 0.01). These results indicate that the GFP-NGF positive BMSCs provide, by way of paracrine, NGF that effectively perform the functions of neuroprotection, which cell fusion may be also contribute to.     

Nerve growth factor receptor (p75 NTR) and pattern of invasion predict poor prognosis in oral squamous cell carcinoma

Aims:  To evaluate the expression of p75 neurotrophin receptor (p75^NTR) in oral squamous cell carcinoma (OSCC). The results were related to tumour node metastasis (TNM) stage, World Health Organization (WHO) grade, invasive front grading (IFG) and prognosis.
Methods and results:  Immunohistochemically, the expression of p75^NTR was assessed in 53 T1-T2 OSCCs. Clinical data were recorded prospectively. The end-point was disease-free survival. All tumours expressed p75^NTR, and this expression, both in central/superficial tumour areas and at the invasive front, was associated with poor prognosis ( P  = 0.03 and P  = 0.02) (log rank test). Tumours with marked cellular dissociation (IFG parameter) had more recurrences than tumours with collective tumour cell invasion ( P  = 0.03). In tumours showing both p75^NTR at the invasive front and marked tumour cell dissociation, the average risk of recurrence was increased about 17 times (Cox regression analysis) compared with tumours with low p75^NTR expression and collective invasion. Traditional prognostic systems were of no prognostic significance.
Conclusion:  p75^NTR was expressed in all OSCCs. p75^NTR expression and the pattern of invasion were significantly associated with a poor prognosis in OSCCs, and both were better prognostic factors than traditional prognostic parameters. The combination of p75^NTR expression and the pattern of invasion strongly increased precision in the identification of tumours with poor disease-free survival.     

Expression of p75NGFR,a Proliferative and Basal Cell Marker,in the Buccal Mucosa Epithelium during Re-epithelialization

We investigated the expression of p75^NGFR, a proliferative and basal cell marker, in the mouse buccal mucosa epithelium during wound healing in order to elucidate the role of epithelial stem cells. Epithelial defects were generated in the epithelium of the buccal mucosa of 6-week-old mice using CO₂ laser irradiation. BrdU was immediately administered to mice following laser irradiation. They were then sacrificed after 1, 3, 7, and 14 days. Paraffin sections were prepared and the irradiated areas were analyzed using immunohistochemistry with anti-p75^NGFR, BrdU, PCNA, and CK14 antibodies. During re-epithelialization, PCNA (–)/p75^NGFR (+) cells extended to the wound, which then closed, whereas PCNA (+)/p75^NGFR (+) cells were not observed at the edge of the wound. In addition, p75^NGFR (–)/CK14 (+), which reflected the presence of post-mitotic differentiating cells, was observed in the supra-basal layers of the extended epithelium. BrdU (+)/p75^NGFR (+), which reflected the presence of epithelial stem cells, was detected sparsely in buccal basal epithelial cells after healing, and disappeared after 7 days. These results suggest that p75^NGFR (+) keratinocytes are localized in the basal layer, which contains oral epithelial stem cells, and retain the ability to proliferate in order to regenerate the buccal mucosal epithelium.     

Structure of the functional interleukin-2 receptor: Evidence for the association of human p55 and murine p75 molecules in a mouse T cell line

The structural basis of the high affinity interleukin-2 receptorwhich was previously reconstituted in a cultured murine T cellline, EL4 by expressing either wild-type Tac antigen complementaryDNA (cDNA) or a chimeric cDNA was characterized. The chimericcDNA encodes a membrane portion whose extracellular portionconsists of that of Tac antigen whereas transmembrane and cytoplasmicportions consists of those the human insulin beta chain. TheTac antigen/anti-Tac antibody complex was treated by chemicalcrosslinking reagents, purified by goat anti-mouse immunoglobulin(lg), and was analysed by SDS–PAGE. We here demonstrated the presence in mouse EL4 transfectantsof a novel membrane protein which is closely associated withthe products of transfected cDNAs in the absence of interleukln-2.The protein is 75 kDa in size and is detected in cells whichexpress high affinity interieukln-2 receptor but not in cellswhich only express low affinity interleukin-2 receptor. Thetransmembrane region and the cytoplasmic region of Tac antigenis not necessary for the formation of the complex consistingof Tac antigen and 75 kDa molecule, indicating that a murine75 kDa molecule associates with Tac antigen extra-cellularly.     

SFRP5基因沉默促进人类胰腺癌细胞系PANC-1增殖与迁移

目的探讨慢病毒介导短发夹RNA(shRNA)沉默SFRP5基因对人类胰腺癌细胞系PANC-1细胞增殖、侵袭和迁移能力的影响。方法构建靶向SFRP5基因特异性shRNA慢病毒载体并转染人胰腺癌PANC-1细胞系,以空白质粒转染阴性对照组,未处理细胞做为空白对照组。用real-time PCR及Western blot检测转染前后SFRP5 RNA以及蛋白的表达;CCK-8实验检测细胞体外增殖能力;使用Transwell小室实验分析细胞侵袭能力;细胞划痕实验分析细胞迁移能力。结果成功建立稳定转染shRNA-SFRP5胰腺癌PANC-1细胞株。SFRP5病毒转染组与阴性对照组及空白对照组相比细胞的增殖能力明显增加(P0.01);SFRP5病毒转染组的细胞侵袭、迁移能力明显高于阴性对照组及空白对照组(P0.01)。结论 SFRP5慢病毒干扰载体能有效抑制SFRP5基因在人胰腺癌PANC-1细胞中的表达,进而促进细胞的增殖、侵袭和迁移。     

Manumycin通过诱导细胞凋亡抑制人胰腺导管癌细胞Panc-1活性

目的：观察manumycin对人胰腺导管癌细胞Panc-1的抑制效应，并探讨其诱导细胞凋亡是否经p38MAPK介导。 方法： 用MTT法检测manumycin对Panc-1细胞的抑癌作用。用caspase-3活性检测试剂盒定量检测manumycin诱导细胞凋亡的水平及评估特异性的p38MAPK抑制剂SB203580对它的影响。 结果： 经manumycin(6 μmol/L、18 μmol/L、54 μmol/L)处理Panc-1细胞24 h,对Panc-1细胞生长具有明显的抑制作用，其抑制率分别为8.9%、21.9%和67.0%，其中后二者的细胞活性与对照组相比有显著差异(P<0.01)，呈量效关系。用药24 h的IC50为34.7 μmol/L。同时，此药物可明显增加caspase-3的活性，且这一效应可部分地被p38抑制剂SB203580阻断。 结论： Manumycin可通过诱导Panc-1细胞凋亡而产生抑癌作用，p38MAPK是manumycin诱导细胞凋亡的通路之一。     

Diverse functions of the p75 neurotrophin receptor

The pan-neurotrophin receptor p75NTR belongs to a large family of receptors, which includes tumor necrosis factor receptors, Fas and approximately 25 other members. The p75NTR is the first receptor to be cloned molecularly. Recent years have seen the emergence of a consensus regarding the signaling pathways activated by p75NTR and its potential biological function, although receptor characterization had not been targeted for some years. We now know that p75NTR has surprisingly diverse effects, ranging from cell death to regulation of axon elongation. This diversity can be explained by the complex formation of p75NTR with other receptors and multiple signaling molecules that interact with the intracellular domain of p75NTR.