胰腺癌伴神经浸润动物模型的建立

目的 建立胰腺癌伴神经浸润的动物模型.方法 32只裸鼠分为4组,每组8只.将人胰腺癌细胞SW1990、CAPAN-2、PANC1分别注射于裸鼠的坐骨神经周围,以不做任何处理的裸鼠作为对照组.观察术后裸鼠体质量变化、成瘤情况、成瘤时间、造模成功率等指标.结果 对照组裸鼠体质量增加,各癌细胞注射组裸鼠成瘤后体质量明显下降,甚至呈现恶液质,成瘤侧肢体活动受限.CAPAN-2注射组及SW1990注射组的8只裸鼠最终均成瘤,PANC1注射组只有5只成瘤.以肿瘤最长径长至约0.8 cm为时间截点,CAPAN-2注射组、PANC1注射组、SW1990注射组裸鼠的成瘤时间分别为（49.8±5.0）、（56.6±2.4）、（25.4±3.0）d.SW1990注射组成瘤时间最短,PANC1注射组成瘤时间最长,3组间成瘤时间的差异具有统计学意义（F=73.51,P＜0.01）.CAPAN-2注射组、PANC1注射组、SW1990注射组裸鼠神经浸润造模成功率分别为87.5％ （7/8）、20.0％（1/5）、50.0％ （4/8）.结论 成功建立了胰腺癌伴神经浸润动物模型,但不同的胰腺癌细胞系建立的动物模型具有不同的特点.     

与胰腺癌神经浸润相关的临床因素分析

目的 观察胰腺癌的神经浸润状况,分析与其相关的临床因素.方法 回顾性分析73例胰腺癌患者的神经浸润状况,分析神经浸润与肿瘤临床病理特征及患者生存率之间的关系.结果 73例中38例(52.1％)有神经浸润,其中6例(15.8％)为单纯胰内神经浸润,32例(84.2％)为胰内、胰外神经均浸润.神经浸润与患者性别、年龄及肿瘤病理类型、分化程度、大小、淋巴结转移均无关(P值均＞0.05),而与腹痛、脉管浸润、肿瘤组织表皮生长因子受体(EGFR)及血管内皮生长因子(VEGF)表达均显著相关(P值均＜0.01).有神经浸润患者的中位生存时间为8个月,显著短于无神经浸润患者的13个月(x2=4.69,P=0.030).结论 胰腺癌的神经浸润发生率较高,可引起明显腹痛,其与脉管浸润及肿瘤组织EGFR和VEGF表达相关,是影响胰腺癌患者术后生存率的因素之一.     

胰腺癌513例神经浸润的病理分析

目的 探讨胰腺癌神经浸润的特征及其与其他临床病理参数之间的关系.方法 光镜下观察491例胰腺导管腺癌、22例其他胰腺恶性肿瘤、41例胰腺良性病变和21例慢性胰腺炎组织中的神经浸润状况,分析其与其他病理学指标的相关性.结果 胰腺导管腺癌的神经浸润率为74%,显著高于其他类型恶性肿瘤的23%(P＜0.01).导管腺癌癌细胞通常穿越外周神经中膜到达内部的神经纤维束,有的甚至横断整根神经纤维.但神经浸润与导管腺癌的分化程度无关.52%的胰腺导管腺癌癌旁组织呈慢性炎症改变,且程度严重,远高于其他类型胰腺癌(14%)及胰腺良性病变(15%)的慢性炎症发生率(P＜0.01).胰腺导管腺癌淋巴细胞浸润神经的发生率为65%,远高于其他恶性肿瘤的36%和胰腺良性病变的22%(P＜0.01).胰腺导管腺癌的神经浸润与癌旁慢性胰腺炎症以及淋巴细胞浸润神经均相关,但与淋巴结转移无关.结论 神经浸润是胰腺导管腺癌特征性的生物学行为之一.     

Galectin-3在人胰腺癌细胞株SW1990增殖浸润中的作用

目的 研究Galectin-3（GAL-3）在人胰腺癌细胞株SWl990增殖、浸润中的作用。方法 体外培养人胰腺癌细胞株SW1990和人胰腺星状细胞（PSC），收集细胞培养上清液。ELISA、RT—PCR和Westernblot检测PSC和SW1990细胞GAL-3的表达。四唑氮蓝还原法（MTT）和流式细胞仪检测不同培养液对SW1990细胞和PSC的增殖的影响。体外侵袭试剂盒检测PSC上清液和GAL-3抗体对SW1990细胞浸润的影响。结果SW1990细胞表达GAL-3，PSC培养上清能增强其GAL-3的表达。SW1990上清液通过GAL-3促进PSC增殖，不同的培养上清液影响PSC细胞的S期比例。PSC培养上清促进SW1990细胞的增殖和浸润，加入GAL-3单抗后，这种促进作用被部分抑制，不同的培养上清液影响SW1990细胞的S期和Gz＋M期的比例。结论 GAL-3参与SW1990的增殖和浸润。     

^125Ⅰ粒子短时低剂量率照射对胰腺癌Capan-2细胞神经浸润的影响

目的 观察^125I粒子短时低剂量率照射对胰腺癌Capan-2细胞神经浸润的影响,并探讨其分子机制.方法 建立胰腺癌Capan-2细胞和大鼠背根神经节（DRG）共培养及Capan-2或DRG单培养模型.通过125I粒子低剂量率照射平板对3种模型进行照射,以相应未照射模型作为对照.倒置显微镜下观察癌细胞、DRG的生长,图像分析软件计算神经突和癌细胞集落占据的表面积,ELISA法检测细胞培养上清液和基质胶溶解液中神经生长因子（NGF）及转化生长因子α（TGF-α）浓度,RT-PCR法检测胰腺癌Capan-2细胞神经营养因子-3（NT-3）mRNA表达.结果 共培养模型中DRG发出的神经突向癌细胞定向、集中生长,而癌细胞沿神经突的方向生长.经125I粒子照射后这种定向、集中和互逆的生长受到一定程度的抑制.共培养组第5天所增加的神经突表面积为290.15±12.08,较DRG单培养组的124.83±6.96显著增加（P＜0.01）,经照射后的神经突表面积减少到201.53±12.20（P ＜0.01）;所增加的Capan-2细胞表面积为300.47±12.99,较Capan-2细胞单培养组的199.30±8.60显著增加（P＜0.01）,经照射后的Capan-2细胞表面积减少到202.35±7.97 （P ＜0.01）.共培养组不表达NT-3mRNA,经照射后NT-3mRNA表达量为0.68±0.04（P ＜0.05）.共培养组培养上清液中NGF及TGF-α浓度分别为（27.56 ±13.73）、（40.86±20.73） ng/ml,经照射后分别升高到（94.98±33.80）、（157.54±83.76） ng/ml,差异有统计学意义（P＜0.05或＜0.01）.共培养组基质胶溶解液中NGF及TGF-α浓度分别为（60.42 ±33.03）、（64.39±21.52） ng/ml,经照射后分别升高到（132.52±53.01）、（138.38±83.58） ng/ml,其中NGF的差异有统计学意义（P＜0.05）.结论 125I粒子短时低剂量率照射可以抑制胰腺癌和神经的交互作用,其机制可能与癌细胞促神经浸润介质NGF、TGF-α和NT-3等表达上调有关.     

胰腺癌神经浸润的分子机制

胰腺癌是一类高度恶性的肿瘤,目前只有约7％的胰腺癌能被早期诊断,由于难以早期诊断及频繁发生的局部或者远处转移,只有15％～20％的患者最终可以接受根治术。尽管近年来对胰腺癌的研究在基础和临床方面均取得进展,但是胰腺癌的预后仍然严峻,总体5年生存率不到5％。     

神经生长因子及其受体与胰腺癌神经浸润转移的关系

胰腺癌恶性程度高,预后差,5年生存率很低,究其原因主要是由于它特殊的生物学行为,即高浸润性和高转移率,其中以神经浸润发生率最高。文献报道,胰腺癌神经浸润率达64%～100%[1]。因此,深入探讨胰腺癌浸润转移的分子生物学机制,对预防胰腺癌的浸润转移、提高患者预后具有重要的意     

Galectin-3在人胰腺癌细胞株SW1990增殖浸润中的作用

目的 研究Galectin-3(GAL-3)在人胰腺癌细胞株SW1990增殖、浸润中的作用.方法 体外培养人胰腺癌细胞株SW1990和人胰腺星状细胞(PSC),收集细胞培养上清液.ELISA、RT-PCR和Western blot检测PSC和SW1990细胞GAL-3的表达.四唑氮蓝还原法(MTT)和流式细胞仪检测不同培养液对SW1990细胞和PSC的增殖的影响.体外侵袭试剂盒检测PSC上清液和GAL-3抗体对SW1990细胞浸润的影响.结果 SW1990细胞表达GAL-3,PSC培养上清能增强其GAL-3的表达.SW1990上清液通过GAL-3促进PSC增殖,不同的培养上清液影响PSC细胞的S期比例.PSC培养上清促进SW1990细胞的增殖和浸润,加入GAL-3单抗后,这种促进作用被部分抑制,不同的培养上清液影响SW1990细胞的S期和G2+M期的比例.结论 GAL-3参与SW1990的增殖和浸润.     

人胰腺癌鸡胚模型的建立及其肿瘤生物学观察

目的建立人胰腺癌鸡胚移植模型,为临床胰腺癌研究提供一种简便实用的工具.方法将人胰腺癌细胞系SW1990细胞接种到鸡胚绒毛尿囊膜(CAM)上,动态观察移植瘤的形态及肿瘤生物学特性,分析影响移植瘤成活的因素,并对瘤组织行组织学检查.结果成功建立了人胰腺癌鸡胚移植模型,移植瘤组织学结构与人胰腺癌相似;接种癌细胞数量影响接种成瘤率,接种癌细胞数量越大,成瘤率越高;移植瘤可诱发血管生成并向肿瘤呈放射状集中.结论将SW1990细胞接种到鸡胚上建立鸡胚移植胰腺癌是可行的,本模型可动态观察胰腺癌生长,模拟其在体内生长情况,为胰腺癌研究提供一种简便实用的工具.     

神经母细胞瘤体外细胞系的构建方法

目的 对神经母细胞瘤(NB)进行体外培养，探讨神经母细胞瘤(NB)体外建系的方法。方法 NB患儿新鲜手术标本，采用组织块培养法、酶消化培养法建立原代细胞系；用选择性酶消化法和机械刮除法进行细胞纯化；从细胞形态学、神经特异性烯醇化酶染色、软琼脂克隆形成实验三方面对细胞进行初步鉴定，证实所培养细胞是否为NB细胞。结果 两种细胞培养法均可培养出NB细胞，纯化后可得形态均一的细胞系。结论 可采用适当的细胞培养方法建立神经母细胞瘤的体外细胞系。     

Pleiotrophin promotes perineural invasion in pancreatic cancer

Perineural invasion(PNI)in pancreatic cancer is an important cause of local recurrence,but little is known about its mechanism.Pleiotrophin(PTN)is an important neurotrophic factor.It is of interest that our recent experimental data showed its involvement in PNI of pancreatic cancer.PTN strongly presents in the cytoplasm of pancreatic cancer cells,and high expression of PTN and its receptor may contribute to the high PNI of pancreatic cancer.Correspondingly,PNI is prone to happen in PTN-positive tumors.We thus hypothesize that,as a neurite growth-promoting factor,PTN may promote PNI in pancreatic cancer.PTN is released at the time of tumor cell necrosis,and binds with its highaffinity receptor,N-syndecan on pancreatic nerves,to promote neural growth in pancreatic cancer.Furthermore,neural destruction leads to a distorted neural homeostasis.Neurons and Schwann cells produce more N-syndecan in an effort to repair the pancreatic nerves.However,the abundance of N-syndecan attracts further PTN-positive cancer cells to the site of injury,creating a vicious cycle.Ultimately,increased PTN and N-syndecan levels,due to the continuous nerve injury,may promote cancer invasion and propagation along the neural structures.Therefore,it is meaningful to discuss the relationship between PTN/N-syndecan signaling and PNI in pancreatic cancer,which may lead to a better understanding of the mechanism of PNI in pancreatic cancer.     

Midkine promotes perineural invasion in human pancreatic cancer

AIM:To investigate midkine(MK)and syndecan-3protein expression in pancreatic cancer by immunohistochemistry,and to analyze their correlation with clinicopathological features,perineural invasion,and prognosis.METHODS:Pancreatic cancer tissues(including adequately sized tumor tissue samples and tissue samples taken from areas less than 2.0 cm around the tumor)were taken from 42 patients who were undergoing a partial duodenopancreatectomy.MK and syndecan-3proteins were detected by immunohistochemistry using a standardized streptavidin-peroxidase method,and analyzed for their correlation with clinicopathological features,perineural invasion,and prognosis.Associations of neural invasion with aggressive characteristics of pancreatic cancer and the presence of perineural invasion were assessed by two independent observers blinded to the patient status.RESULTS:MK and syndecan-3 were found in 26(61.9%)and 24(57.1%)specimens,respectively.MK and syndecan-3 expression was associated with perineural invasion(P=0.018 and 0.031,respectively).High MK expression was closely associated with advanced tumor,node and metastasis stage(P=0.008),lymph node metastasis(P=0.042),and decreased postoperative survival at 3years(51.0%vs 21.8%,P=0.001).Syndecan-3 levels were correlated with tumor size(P=0.028).Patients who were syndecan-3 negative had a higher cumulative survival rate than those who were positive,but the difference was not significant(44.0%vs 23.0%,P>0.05).CONCLUSION:MK and syndecan-3 are frequently expressed in pancreatic cancer and associated with perineural invasion.High expression of MK and syndecan-3may contribute to the highly perineural invasion and poor prognosis of human pancreatic cancer.     

Nerve growth factor regulates CD133 function to promote tumor cell migration and invasion via activating ERK1/2 signaling in pancreatic cancer

BackgroundPerineural invasion (PNI) is extremely high frequency among the various metastatic routes in pancreatic cancer. Nerve growth factor, secreted by astroglial cells, exerts effects on tumor invasion in some cancer cells, but its function on migration and invasion in pancreatic cancer is still unclear. In the present study, we determined the effects of NGF on modulating tumor cell metastatic potential and invasion activity and explored its mechanisms in pancreatic cancer.MethodsNGF and CD133 expression were detected in tumor tissues using immunohistochemical analysis and Western blotting analysis. The effects of NGF on the regulation of CD133 expression and the promotion of cancer migration and invasion were investigated using wound healing and matrigel transwell assay. A related mechanism that NGF regulates CD133's function via activating ERK1/2 signaling also was observed.ResultsNGF/CD133 is overexpressed in human pancreatic cancer and promotes the migration and invasion of human pancreatic cancer cells through the activation of the ERK/CD133 signaling cascade. NGF/ERK signaling modulates the cancer cell EMT process, migration and invasion through the regulation of CD133 expression and its subcellular localization.ConclusionsNGF/CD133 signaling initiated the migration and invasion of pancreatic cancer cells. NGF/CD133 might be an effective and potent therapeutic target for pancreatic cancer metastasis, particularly in PNI.     

胰腺星状细胞对胰腺癌AsPC-1细胞侵袭转移的影响

目的 探讨胰腺星状细胞(PSCs)对胰腺癌细胞侵袭转移的影响及基质细胞因子-I(SDF-1)在此过程中的作用.方法 常规分离、培养PSCs,收集及浓缩PSCs条件培养液(PSC-CM),应用不同浓度的PSC-CM、抗SDF-1抗体(anti-SDF-1)及两者联合应用处理AsPC-1细胞,采用MTT法检测AsPC-1细胞的增殖,Transwell小室检测细胞迁移,体外侵袭实验观察细胞侵袭能力.结果 对照组及0.25、0.5、1 μg/μl PSC-CM组AsPC-1细胞增殖的吸光度值(A490)分别为0.437 ±0.041、0.472±0.048、0.553±0.057、0.690±0.051,PSC-CM呈剂量依赖性促进细胞增殖,其中0.5、1μg/μl PSC-CM组与对照组间以及0.5 μg/μl与1μg/μl PSC-CM组间的差异具有统计学意义(P值均＜0.05).对照组、anti-SDF-1组、PSC-CM组及PSC-CM± anti-SDF-1组细胞增殖的A490值分别为0.407±0.028、0.416±0.030、0.629±0.048、0.481±0.049;穿膜细胞数分别为(35.3±7.1)、(34.8±5.6)、(140.9±12.7)、(56.5±5.9)个;侵袭细胞数分别为(27.1±2.9)、(29.1±4.2)、(81.5±8.2)、(46.4±4.4)个.anti-SDF-1组与对照组间的差异无统计学意义,PSC-CM组细胞的增殖、迁移、侵袭能力较对照组显著增强(P＜0.05或P＜0.01),PSC-CM± anti-SDF-1组细胞的增殖、迁移、侵袭能力较PSC-CM组显著降低,但仍显著高于对照组(P＜0.05或P＜0.01).结论 PSC-CM可促进AsPC-1的增殖、迁移及侵袭,其机制为部分通过SDF-1/CXCR4受体配体系统.     

The complementary role of lymphovascular invasion and perineural invasion in the TNM staging process of rectal cancer

The aim of this study is to clarify the association between lymphovascular invasion (LVI) and/or perineural invasion (PNI) and the clinical characteristics and prognostic importance of rectal cancer, to provide a basis for early adjuvant treatment of rectal cancer. We retrospectively analyzed patients diagnosed with rectal cancer. This study involved rectal cancer tissue samples were obtained by surgical methods. Data on histological form, tumor classification, tumor size, gross growth pattern, blood and lymphatic vessel invasion, and PNI of the slice by HE staining were obtained from pathological examination. Immunohistochemical analysis of tissue samples was performed to determine p53 and EGFR expressions. There were 330 rectal cancer patients included in the study. LVI and/or PNI can be used as a high-risk factor for the prognosis of rectal cancer, predict prognostic survival, and guide adjuvant therapy. The detection rates of LVI and PNI were 32.1% and 16.1%. Differentiation grade, Union for International Cancer Control staging, tumor-lymph node-metastasis staging are significantly related to LVI or PNI. Multivariate logistic regression analysis shows that poor differentiation and N ≥ 1 can be used as independent risk factors and predictive factors for LVI. At the same time, poor differentiation and T > 3 is an independent risk factor for PNI. Only poor differentiation is the risk factor for poor prognosis in Cox risk regression analysis. In addition, the simultaneous occurrence of LVI and PNI is an independent prognostic factor.