全反式维甲酸对神经母细胞瘤细胞系体外生长的影响

目的　体外观察全反式维甲酸 (alltransretinoicacid ,ATRA)对神经母细胞瘤细胞系 (SK N SH)细胞生长的影响。方法　SK N SH细胞持续培养在含ATRA的 96孔板中 ,采用MTT法检测ATRA的体外抗增殖效果 ,用相差倒置显微镜及透射电镜观察细胞形态变化。结果　 0 .1～ 1μmol/L浓度的ATRA对SK N SH细胞可产生明显的抗增殖作用 ;其抗增殖作用呈时间和剂量依赖关系。 1μmol/LATRA作用 12天 ,细胞生长抑制率达 77.0 3%。ATRA能诱导SK N SH细胞分化成熟 ,先是细胞出现神经树 (轴 )突样胞突 ,培养 10天后多个细胞形成类似神经节样形态结构 ,“神经节”之间形成类神经纤维 ;电镜观察细胞核浆比例下降 ,无特征性细胞凋亡形态变化。结论　ATRA通过诱导分化作用 ,而明显抑制神经母细胞瘤细胞增殖。     

节细胞神经母细胞瘤1例

患者男,59岁.因左腹、腰部胀痛,逐渐加重1个月于1998年1月8日入院.平素健康.查体:左上腹触及一6cm×5cm质硬肿块,固定、轻压痛.B超检查:左肾前上极探及一8.0cm×7.8cm,低回声不均质肿块,包膜完整、增厚.CT扫描示左侧腹膜后肿瘤,与胰、左肾关系密切.X线胸片未见心肺异常.CEA<20μg/ml.经手术证实,肿瘤位于腹膜后、胰体尾部及左肾上极之间,并侵犯左肾门及肾蒂血管,血供丰富.肿瘤为灰红色,卵圆形9cm×7cm×6cm大小,切面见大片出血、坏死,部分囊性变,伴有鱼肉样组织.肿瘤免疫组化结果:NSE( )、FN(-)、GFAP(-)、S_(100)节细胞( )、小细胞(-).病理诊断为节细胞神经母细胞瘤.     

微波辐射对人成神经母细胞瘤SH-SY5Y细胞增殖与分化的影响

目的观察微波辐射对人成神经母细胞瘤SH-SY5Y细胞增殖与分化的影响。 方法  采用10、30、50 mW/cm²辐射强度的微波辐射SH-SY5Y细胞5 min,0 mW/cm²为对照组,锥虫蓝拒染法进行细胞计数并计算增殖率;20 μmol/L全反式维甲酸黑暗下诱导分化72 h后,在光学显微镜下观察细胞分化的形态并计算分化细胞率;流式细胞仪检测细胞周期;蛋白质印迹检测细胞神经纤丝重链亚基的蛋白表达。结果微波辐射后随辐射强度增加,细胞增殖率在各时间点上均出现不同程度的梯度抑制,与对照组（0 mW/cm²）相比,10 mW/cm²辐射组在72 h时出现明显抑制;30 mW/cm²辐射组在48 h时出现明显抑制;50 mW/cm2辐射组在辐射12 h时就出现明显抑制;20 μmol/L全反式维甲酸黑暗下诱导分化72 h后,30、50 mW/cm²微波辐射组细胞分化的数量少于对照组细胞,而且细胞轴突长度短于对照组细胞;随着辐射强度的增加,G₀/G₁期的SH-SY5Y细胞比例逐渐增加,S 期细胞比例逐渐减少,G₂+M 期细胞比例变化相对较小;微波辐射后,SH-SY5Y细胞总NF-H蛋白表达无明显变化,但磷酸化的NF-H蛋白表达随辐射强度的增加逐渐降低。结论微波辐射可同时抑制SH-SY5Y细胞增殖与分化并呈现明显的剂量依赖性抑制作用;细胞周期呈G₀/G₁期阻滞;对NF-H的表达无影响,但明显抑制了NF-H的磷酸化。     

节细胞神经母细胞瘤1例

患者,男,13岁.因右侧腰痛伴低热3天,在襄樊市中医院做双肾CT示右肾上腺肿瘤于2003年6月入我院.     

DRR1调节神经母细胞瘤细胞分化的作用机制

目的:探究肾细胞癌下调蛋白1（down-regulated in renal cell carcinoma 1,DRR1）在神经母细胞瘤细胞分化中的作用以及内在的分子机制。方法:在神经母细胞瘤细胞中敲减或过表达DRR1后,分别考察细胞分化状态的变化、细胞周期的变化以及利用real-time PCR的方法检测其潜在下游因子的表达变化;利用神经母细胞瘤患者数据库,分析在神经母细胞瘤患者样本中DRR1的潜在下游因子的表达与DRR1表达之间的相关性;诱导神经母细胞瘤细胞分化后,利用real-time PCR的方法检测DRR1的潜在下游因子的表达变化。结果:DRR1的表达可促进神经母细胞瘤细胞的分化;敲减DRR1促进神经母细胞瘤细胞的G1/S期的转换,过表达DRR1抑制神经母细胞瘤细胞的G1/S期的转换;DRR1的表达改变影响其潜在下游因子在神经母细胞瘤细胞中的表达;在神经母细胞瘤患者样本中DRR1的表达与其潜在下游因子的表达呈显著相关性;DRR1的潜在下游因子的表达随神经母细胞瘤细胞分化状态的变化而改变。结论:DRR1通过调控下游因子的表达调节神经母细胞瘤细胞的分化。     

神经纤毛蛋白-1与神经胶质母细胞瘤发生发展的相关性研究进展

摘 要：神经纤毛蛋白-1（neuropilin-1,NRP1）是一种广泛表达的跨膜蛋白,其在各种肿瘤组织中广泛表达,随肿瘤恶性程度的增加而上调,与肿瘤预后显著相关。胶质母细胞瘤(GBM)是脑恶性肿瘤中最常见、恶性程度最高的肿瘤类型。研究发现,NRP1对于肿瘤的调控和促进作用在神经胶质母细胞瘤（GBM）中更为显著。体内实验表明,NRP1以共受体的形式参与、调节了GBM中血管生长因子的激活,进而促进GBM的发展。全文综述了NPP1的结构与功能特点,以及NRP1参与GBM发生、发展过程的研究进展。     

TIAM1在儿童神经母细胞瘤中的研究进展

随着分子诊断技术的不断发展,精准治疗已进入恶性肿瘤的治疗范畴,靶向治疗是近年来较为热门的研究方向。神经母细胞瘤（neuroblastoma,NB）是儿童期最常见的颅外实体瘤。研究表明,很多基因参与NB的发生发展过程,T淋巴瘤侵袭转移诱导因子1（T lymphoma invasion and metastasis inducing factor 1,TIAM1）是其中之一。TIAM1主要与下游的RAC1（Ras-related C3 botu?linum toxin substrate 1）结合,作用于TrkA/TIAM1/RAC1通路,激活下游相关因子,参与调节神经元轴突的分化过程。因此,深入的研究和实验或许可以更透彻的阐明具体的机制,为NB未来的诊疗提供一个新的方向。      

原发性肾神经母细胞瘤1例

神经母细胞瘤多发生于交感神经系统，原发于肾脏非常少见。我科于近年诊断1例原发性肾神经母细胞瘤．现报道如下。     

原发颈部节细胞神经母细胞瘤1例

患儿,男,5岁,因发现左颈部肿块10 d入院.肿块无疼痛,不伴呼吸困难,无张口受限.查体:一般情况好,生命体征平稳;左颈部隆起,皮肤无红肿及破溃;可触及肿块,约7 cm×4 cm,上界至乳突尖,下界至环状软骨下缘水平,位于胸锁乳突肌深面,质硬,无压痛,表面欠光滑,边界清楚,较固定,不随吞咽运动;     

三氧化二砷对神经母细胞瘤细胞增殖的影响

目的：观察三氧化二砷体外对神经母细胞瘤细胞增殖的影响及其作用机制的探讨。方法：（１）ＭＴＴ法观察三氧化二砷对神经母细胞瘤细胞生长的影响;（２）化学染色观察实验和对照组细胞形态学改变;（３）ＰＩ染色后流式细胞仪检测实验和对照组细胞凋亡峰;（４）凝胶电泳观察凋亡细胞ＤＮＡ的“梯型”条带,结果：（１）三氧化二砷明显抑制神经母细胞瘤细胞株ＳＪ－Ｎ－ＳＨ细胞增殖,抑制作用呈剂量和作用时间依赖性,（２）通过细     

MRP1 gene expression level regulates the death and differentiation response of neuroblastoma cells.

We have previously reported a strong correlation between poor prognosis in childhood neuroblastoma (NB) patients and high-level expression of the transmembrane efflux pump, Multidrug Resistance-associated Protein (MRP1), in NB tumour tissue. In this study, we inhibited the endogenous expression of MRP1 in 2 different NB tumour cell lines by stably transfecting an MRP1 antisense expression vector (MRP-AS). Compared with control cells, MRP-AS transfectant cells demonstrated a higher proportion of dead and morphologically apoptotic cells, spontaneous neuritogenesis, and, increased synaptophysin and neurofilament expression. Bcl-2 protein expression was markedly reduced in MRP-AS cells compared to controls. Conversely, we found that the same NB tumour cell line overexpressing the full-length MRP1 cDNA in sense orientation (MRP-S) demonstrated resistance to the neuritogenic effect of the differentiating agent, all-trans-retinoic acid. Taken together, the results suggest that the level of MRP1 expression in NB tumour cells may influence the capacity of NB cells for spontaneous regression in vivo through cell differentiation and death.     

Targeting Notch pathway induces growth inhibition and differentiation of neuroblastoma cells

High-risk neuroblastoma is a severe pediatric tumor characterized by poor prognosis. Understanding the molecular mechanisms involved in tumor development and progression is strategic for the improvement of pharmacological therapies. Notch was recently proposed as a pharmacological target for the therapy of several cancers and is emerging as a new neuroblastoma-related molecular pathway. However, the precise role played by Notch in this cancer remains to be studied extensively. Here, we show that Notch activation by the Jagged1 ligand enhances the proliferation of neuroblastoma cells, and we propose the possible use of Notch-blocking γ-secretase inhibitors (GSIs) in neuroblastoma therapy. Two different GSIs, Compound E and DAPT, were tested alone or in combination with 13-cis retinoic acid (RA) on neuroblastoma cell lines. SH-SY5Y and IMR-32 cells were chosen as paradigms of lower and higher malignancy, respectively. Used alone, GSIs induced complete cell growth arrest, promoted neuronal differentiation, and significantly reduced cell motility. The combination of GSIs and 13-cis RA resulted in the enhanced growth inhibition, differentiation, and migration of neuroblastoma cells. In summary, our data suggest that a combination of GSIs with 13-cis RA offers a therapeutic advantage over a single agent, indicating a potential novel therapy for neuroblastoma.     

Interleukin 13 participates in terminal differentiation of esophageal squamous cell carcinoma cells

BackgroundIn China, esophageal squamous cell carcinoma (ESCC) accounts for more than 90% of all esophageal cancer cases. Interleukin 13 (IL-13) was widely reported to play a key role in tumor progression. Our previous study reported that IL-13 was a favorable predictive marker for the overall survival of esophageal squamous cell carcinoma (ESCC) patients, but how IL-13 contributes to ESCC progression remains unknown. This study aims to explore the role of IL-13 and its underlying downstream molecular mechanisms in ESCC progression.MethodsTissue microarrays including 262 primary ESCC tumor tissues were collected and analyzed. The expression of IL-13 in ESCC tumor tissue was detected with immunohistochemistry staining (IHC). Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to qualify the expressions of KRT13, KRT4 and 15-lipoxygenase-1 (15-LOX-1) in cultured ESCC cell lines with recombinant IL-13 treatment.ResultsIL-13 was expressed in the esophageal epithelium cells and ESCC tumor cells. High IL-13 expression in ESCC tumor cells predicted a good prognosis for patients. Recombinant human IL-13 raised KRT13 and 15-LOX-1 mRNA levels, but lowered KRT4 mRNA level 15-LOX-1 in ESCC cells in vitro.ConclusionsIn summary, our study suggests that IL-13 might improve the prognosis of ESCC by promoting the terminal differentiation of ESCC cells. This may offer potential new therapeutic target for early treatment of ESCC.     

Inhibition of Akt sensitises neuroblastoma cells to gold(III) porphyrin 1a, a novel antitumour drug induced apoptosis and growth inhibition

Background:

Gold(III) porphyrin 1a is a new class of anticancer drug, which inhibits cell proliferation of wide range of human cancer cell lines and induces apoptosis in human nasopharyngeal carcinoma cells. However, the underlying signalling mechanism by which gold(III) porphyrin 1a modifies the intracellular apoptosis pathways in tumour cells has not been explained in detail in neuroblastoma cells.

Methods:

Cell proliferation and apoptosis were determined by measuring 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Annexin V binding, respectively. Western blot assay was used to detect proteins involved in apoptotic and Akt pathways. In vivo tumour growth was assessed by inoculating tumour cells to nude mice subcutaneously, and gold(III) porphyrin 1a was administrated intravenously.

Results:

This study assessed the antitumour effect and mechanism of gold(III) porphyrin 1a on neuroblastoma in vitro and in vivo. Gold(III) porphyrin 1a displayed a growth inhibition and induction of apoptosis in neuroblastoma cells effectively in vitro, which was accompanied with release of cytochrome c and Smac/DIABLO and caspases activation. Further studies indicated that gold(III) porphyrin 1a inhibited X-linked inhibitor of apoptosis (XIAP). However, we found that gold(III) porphyrin 1a can induce a survival signal, Akt activation within minutes and could last for at least 24 h. To further confirm association between activation of Akt and the effectiveness of gold(III) porphyrin 1a, neuroblastoma cells were treated with API-2, an Akt-specific inhibitor. API-2 sensitised cells to gold(III) porphyrin 1a-induced apoptosis and growth inhibition.

Conclusion:

These results suggested that Akt may be considered as a molecular ‘brake'' that neuroblastoma cells rely on to slow down gold(III) porphyrin 1a-induced apoptosis and antiproliferation. Gold(III) porphyrin 1a is a mitochondrial apoptotic stimulus but also activates Akt, suggesting an involvement of Akt in mediating the effectiveness to growth inhibition and apoptosis by gold(III) porphyrin 1a and that inhibition of Akt can enhance the anticancer activity of gold(III) porphyrin 1a in neuroblastoma.     

激活NGF/trkA信号传导通路引起NB细胞良性分化的研究

目的 :观察神经生长因子 (nervegrowthfactor ,NGF)与其高亲和性受体 (trkA)信号传导通路的激活 ,对神经母细胞瘤 (neu roblastoma ,NB)良性分化的影响。方法 :利用脂质体转染法将trkAcDNA重组质粒转染入人神经母细胞瘤SH SY5Y细胞系中 ,应用RT PCR技术鉴定转染后基因的表达 ,比较转染前后SH SY5Y细胞的分化程度及表达trkA的SH SY5Y细胞用NGF处理前后的分化程度。结果 :成功转染trkA基因并在SH SY5Y细胞中稳定表达 ,对照组及空载体组trkAPCR率分别为 0 43± 0 0 73 8和 0 44± 0 0 9,实验组为 2 69± 0 2 2 ,实验组与前两组比较差异有统计学意义 ,P <0 0 1;转染后SH SY5Y细胞的分化程度明显增加 ,用NGF处理后 ,其分化程度进一步增加。分化细胞的百分率实验组 1为 3 7 69± 1 75 ,实验组 2为 2 6 2 8± 1 3 4,对照组 18 43 5± 1 2 5 ,空载体组 17 61± 1 69。实验组 1与对照组及空载体组相比差异有统计学意义 ,P <0 0 1;实验组 2与对照组及空载体组相比差异有统计意义 ,P <0 0 5 ;实验组 2和实验组 1相比差异有统计学意义 ,P <0 0 1。结论 :NGF/trkA信号传导通路的激活能够引起NB细胞的良性分化 ,这一传导通路的激活可能是NB细胞良性逆转的重要因素。     

Nucleolar protein PES1 is a marker of neuroblastoma outcome and is associated with neuroblastoma differentiation

Neuroblastoma (NB) is a childhood malignant tumor that arises from precursor cells of the sympathetic nervous system. Spontaneous regression is a phenomenon unique to NBs and is caused by differentiation of tumor cells. PES1 is a multifunctional protein with roles in both neural development and ribosome biogenesis. Various kinds of models have revealed the significance of PES1 in neurodevelopment. However, the roles of PES1 in NB tumorigenesis and differentiation have remained unknown. Here we show that NB cases with MYCN amplification and clinically unfavorable stage (INSS stage 4) express higher levels of PES1. High PES1 expression was associated with worse overall and relapse‐free survival. In NB cell lines, PES1 knockdown suppressed tumor cell growth and induced apoptosis. This growth inhibition was associated with the expression of NB differentiation markers. However, when the differentiation of NB cell lines was induced by the use of all‐trans retinoic acid, there was a corresponding decrease in PES1 expression. Pes1 expression of tumorspheres originated from MYCN transgenic mice also diminished after the induction of differentiation with growth factors. We also reanalyzed the distribution of PES1 in the nucleolus. PES1 was localized in the dense fibrillar component, but not in the granular component of nucleoli. After treatment with the DNA‐damaging agent camptothecin, this distribution was dramatically changed to diffuse nucleoplasmic. These data suggest that PES1 is a marker of NB outcome, that it regulates NB cell proliferation, and is associated with NB differentiation.     

Interferon-gamma cooperates with retinoic acid and phorbol ester to induce differentiation and growth inhibition of human neuroblastoma cells.

The prognosis of patients with advanced stages of neuroblastoma with N-myc amplification remains poor despite escalated therapy, a situation that has called for alternative therapeutic approaches. Neuroblastoma cells, which represent immature peripheral neuronal cells, treated with certain physiologic and nonphysiologic agents such as retinoic acid (RA), phorbol esters and interferons (IFN) in vitro undergo cellular differentiation and stop to divide, a process that mimics normal neuronal development. Such "differentiation therapy" using RA after autologous bone marrow transplantation has recently given encouraging results in neuroblastoma patients with advanced disease. Considering approaches for improved differentiation therapy, we investigated possible synergistic effects of combining agents known to influence neuroblastoma growth and differentiation in vitro. Our results show that combined treatment with IFN-gamma and RA or the phorbol ester 12-O-tetradecanoyl-phorbol acetate (TPA) had synergistic or enhancing effects on morphologic differentiation and neurite outgrowth in 5 of 5 neuroblastoma cell lines, 3 of which expressed very high levels of N-myc mRNA due to N-myc amplification. The combinations RA+IFN-gamma or TPA+IFN-gamma also enhanced induced growth inhibition in all 5 cell lines, in several cases resulting in complete growth arrest under conditions where cells stimulated with either agent alone continued to grow. The phenotypic effects of the combined RA+IFN-gamma or TPA+IFN-gamma treatments were in most, but not all, investigated cases accompanied by moderate reductions in N-myc expression, suggesting that the cooperative signals may counteract N-Myc activity at several levels. The cooperativity between IFN-gamma and other differentiation signals may be relevant for approaches to improve the therapy for high-risk neuroblastoma with N-myc-amplification.     

Vascular endothelial growth factor and programmed death-1 pathway inhibitors in renal cell carcinoma

Advanced renal cell carcinoma has historically carried a poor prognosis with very limited treatment options. However, in recent years, the treatment landscape has changed drastically, with many new therapeutic options and improved survival for patients. Novel treatments consist of molecularly targeted agents against the vascular endothelial growth factor (VEGF) pathway as well as the immune checkpoint inhibitors, which stimulate an antitumor immune response. Recent strategy has focused on the development of combination therapy with the use of VEGF inhibitors and immune checkpoint inhibitors in the first-line setting. As more treatments are approved and the options for therapy expand further, there is a growing need for predictive biomarkers to personalize treatment choices for individual patients. Prospective clinical trials comparing the sequencing of treatments are needed to help determine the best therapeutic approach.     

NeuroD1 promotes neuroblastoma cell growth by inducing the expression of ALK

Neuroblastoma is derived from the sympathetic neuronal lineage of neural crest cells, and is the most frequently observed of the extracranial pediatric solid tumors. The neuronal differentiation factor, NeuroD1, has previously been shown to promote cell motility in neuroblastoma by suppressing the expression of Slit2. Here we report that NeuroD1 is also involved in the proliferation of neuroblastoma cells, including human cell lines and primary tumorspheres cultured from the tumor tissues of model mice. Interestingly, the growth inhibition of neuroblastoma cells induced by knockdown of NeuroD1 was accompanied by a reduction of ALK expression. ALK is known to be one of the important predisposition genes for neuroblastoma. The phenotype resulting from knockdown of NeuroD1 was suppressed by forced expression of ALK and, therefore, NeuroD1 appears to act mainly through ALK to promote the proliferation of neuroblastoma cells. Furthermore, we showed that NeuroD1 directly bound to the promoter region of ALK gene. In addition, the particular E‐box in the promoter was responsible for NeuroD1‐mediated ALK expression. These results indicate that ALK should be a direct target gene of NeuroD1. Finally, the expressions of NeuroD1 and ALK in the early tumor lesions of neuroblastoma model mice coincided in vivo. We conclude that the novel mechanism would regulate the expression of ALK in neuroblastoma and that NeuroD1 should be significantly involved in neuroblastoma tumorigenesis.