17β—雌二醇下调血管平滑肌内皮素A型受体的表达

为进一步探讨雌激素对心血管的保护作用,实验在双侧卵巢去势大鼠模型和培养的血管平滑肌细胞（VSMCs)上,观察17β－雌二醇（E2）对血管反应性及VSMCs增殖的影响,以RT－PCR和Western blot检测内皮素受体（ETAR）的表达,结果显示：去势雌性大鼠血管对内皮素（ET－1）的反应性明显增高,ETAR特异性受体阻断剂BQ123能完全阻断ET－1对VSMCs增殖的影响,E2能明显抑制ET－1对VSMCs增殖的作用,RT－PCR结果显示E2能抑制ETAR mRNA的表达,Western blot进一步证实E2能抑制ETAR蛋白表达,E2受体阻断剂Tamoxifen能部分抑制ET－1对VSMCs的增殖及ETAR的mRNA和蛋白 的表达。以上结果提示;ET－1促VSMCs增殖的效应主要是由ETAR介导的,雌激素能通过下调ETAR来抑制ET－1对VSMCs 促增殖的作用和血管对ET－1的反应,且此作用与雌激素受体有关。     

培养的血管平滑肌细胞ERα和ERβ的表达与细胞表型转变及增殖时相有关

目的探讨雌激素对血管平滑肌细胞（VSMC）增殖的双重效应机制。方法采用Westernblot、电镜形态定量及细胞计数的方法,动态检测原代培养大鼠VSMC在有或无10^-8mol/L17β-雌二醇（E2）存在下,雌激素受体（ER）α和β表达变化与细胞表型转变及增殖时相的关系。结果无E2存在时,VSMC在从收缩型向合成型转变（原代培养第0到5天）及活跃增殖（第5到12天）过程中,ERβ表达无明显变化,但ERα表达明显上升,导致ERα/ERβ比值升高。这种变化并不随VSMC表型的恢复及增殖停止而逆转。有E2存在时,ERα/ERβ比值在第5天时低于对照组,而第9天后各时点均高于对照组;这种影响与E2对不同状态VSMC的不同作用基本对应,即延长原代收缩型SMC的增殖潜伏期,但促进已发生表型转变的VSMC增殖。结论雌激素对不同表型VSMC的双重效应与表型转变前后ERα/ERβ比值变化有关。     

17β—雌二醇抑制内皮素诱导的血管平滑肌细胞增殖作用

目的和方法：利用组织块贴壁法进行大鼠VSMC培养,胰蛋白酶分散细胞法传代。实验采用第4-6代细胞。采用氚-胸腺嘧啶核苷（[^3H]-TdR)掺入和细胞计数来作为VSMC增殖的指标,以RT-PCR的方法检测ETAR的表达,观察17β-雌二醇（E2）对内皮素-I（endothelin-l,ET-1)介导的血管平滑肌细胞（VSMC）增殖反应以及对内皮素A型受体（ETAR）表达的影响。结果：ETAR特异性拮抗剂BQ123能完全阻断ET-1介导的VSMC增殖反应;E2可明显抑制ET-1促进VSMC增殖的作用,RT-PCR结果显示E2能抑制ETAR的表达,12h时抑制作用最为明显;E2受体阻断剂Tamoxifen亦能部分抑制ET-1对VSMC的增殖及ETAR的mRNA的表达。结论：ET-1促进VSMC增殖作用主要通过ETAR介导的,雌激素可通过抑制ETARmRNA表达来发挥对ET-1促进VSMC增殖的抑制作用。     

MLT抑制E2诱发的大鼠垂体PRL瘤的增生涉及雌激素受体的作用

目的：探讨褪黑素（melatonin,MLT）抑制17-β-雌二醇（17-β-estradiol,E2）诱发垂体催乳素（prolactin,PRL）瘤的增生及其机制的初始阶段,MLT对雌激素受体的作用。方法：在体实验采用每日定时给各组SD大鼠分别皮下注射不同浓度的MLT,建立MLT抑制E2诱发的垂体PRL瘤增生的动物模型。离体实验采用原代培养细胞原位杂交方法,探讨垂体PRL瘤细胞MLT受体的表达、MLT对雌激素受体（ER）表达的作用;应用电泳迁移率改变（EMSA）的方法,观察MLT对雌激素受体（ER）与雌激素反应元件（ERE）结合的效应。结果：每只大鼠每日定时皮下注射0．25或0．50mg MLT能显著抑制E2诱发的垂体PRL瘤的增生（分别P〈0．01、P〈0．05）;F4诱发的垂体PRL瘤细胞内有MLT受体MLT1a和MLT1b;给0．25mg／day／rat MLT组的大鼠垂体PRL瘤细胞内ER的表达显著减少（P〈0．01）、ER与ERE的结合量显著降低（P〈0．01）。结论：一定剂量的MLT能显著抑制E2诱发的SD大鼠垂体PRL瘤的增生,其作用机制之一可能与MLT抑制ER的表达、及其部分阻断ER与ERE的结合有关。     

LRP16对乳腺癌MCF-7细胞增殖的影响

用Northern印迹方法检测雌二醇 (17β E2 )对LRP16mRNA表达的时间及剂量依赖性调控作用 .构建LRP16基因启动子序列调控的萤光素酶报告子 (pS0 ) ,并与雌激素受体α和 β(ERα和ERβ)表达载体共转染COS 7和MCF 7细胞后测定萤光素酶活性 .将LRP16基因的表达载体转染MCF 7细胞 ,测定过表达LRP16对细胞的生长特性的影响 .17β E2 使MCF 7细胞中LRP16mRNA表达水平增加 ,增加幅度未显示出 17β E2 培养时间和剂量的依赖性 .pS0 与ERα表达载体共转染细胞的相对萤光素酶活性较非共转染组 (对照组 )及pS0 ERβ表载体共转染组升高 5～ 10倍 .LRP16基因过表达促进MCF 7细胞的增殖 .研究表明 ,雌激素可能通过ERα上调乳腺癌MCF 7细胞LRP16基因的表达并促进细胞增殖     

过表达ER恢复雌激素对子宫内膜癌KLE细胞中Notch信号通路的调控

目的：通过观察雌激素对子宫内膜癌KLE细胞中Notch信号通路的影响,探讨过表达雌激素核受体（estrogenreceptor,ER）是否可以恢复雌激素对Notch信号通路的调控作用,继而调节细胞增殖活性。方法：MTT检测雌激素及Notch信号通路对细胞增殖活性的影响;RT．PCR及Westem．blotting检测雌激素及Notch通路抑制剂DAPT对Notch表达的影响;质粒的抽提及转染使KLE细胞中的雌激素核受体ER过表达。结果：雌激素呈剂量依赖效应促进KLE细胞的增殖活性,其中以雌激素浓度为1．0×10-9M时最明显（相对于对照组为1．25±0．026,P〈0．05）;抑制Notch信号通路的表达可以明显下调KLE细胞的增殖活性（0．76±0．02,P〈0．05）;在KLE细胞中,雌激素对Notch的表达没有明显的调控作用,但是将其雌激素核受体过表达后,雌激素可明显上调Notch的表达,并显著促进细胞的增殖活性（1．24±0．02,P〈0．05）。结论：在ER阴性的子宫内膜癌细胞中过表达ER,可以恢复雌激素对Notch信号通路的调控,从而进一步的调控细胞增殖活性。     

原癌基因FOS蛋白、雌二醇及其受体在中国林蛙不同时期精巢中的表达

用免疫细胞化学方法检测了原癌基因FOS蛋白、17β-雌二醇(E2)和雌激素受体(ER)在中国林蛙生精周期中不同时期精巢内的表达定位。结果显示:在中国林蛙生精周期的Ⅰ—Ⅴ期,E2和ER在精原细胞、精母细胞、精子细胞、精子、支持细胞和间质细胞内均有表达。在不同时期的精巢中,E2和ER在生精细胞的定位具有一致性:在Ⅱ—Ⅲ期,精子细胞的E2和ER阳性表达最强;在Ⅲ期,精子中E2和ER阳性反应强度显著高于Ⅳ—Ⅴ期(P<0.01)。在生精周期的Ⅰ—Ⅴ期,支持细胞中,E2和ER表达强度经历由强减弱再增强的变化过程。在生精周期的Ⅲ期,间质细胞中E2和ER阳性反应强度高于其他各期。除精子外的生精细胞,支持细胞和间质细胞内均有FOS阳性反应,其表达强度呈现阶段性变化。     

乳腺癌雌激素受体β不同亚型表达载体的构建及其转录活性分析

目的：构建人乳腺癌中雌激素受体B（ERβ）3种亚型ERβ1、ERβ2和ERβ5的真核表达载体,测定不同亚型ERβ的转录活性。方法：以乳腺癌细胞MCF7的cDNA为模板,PCR扩增ERβ1、ERβ2和ERβ5万基因,分别克隆到pXJ40-Mye载体,Western印迹检测克隆载体在293T细胞内的表达;将上述载体与含雌激素应答元件（ERE）的萤光素酶（Luc）报告基因载体（ERE—luc）共转293T细胞,测定各亚型ER／3的转录活性。结果：构建了Myc—ERβ1、Myc—ERβ2和Mye—ERβ5表达载体,转录活性结果显示上述表达载体均具有活性,雌激素可升高ERβ5的转录活性,不能升高Eβ2和ER5的转录活性。结论：该研究为进-步探讨不同亚型ERβ在乳腺癌中的功能奠定了基础。     

高血压大鼠心脏和血管MAPK磷酸酶-1表达的改变及对平滑肌细胞增殖的影响

目的和方法：比较自发性高血压大鼠（SHR）和对照（WKY）大鼠心脏和主动脉丝裂素活化蛋白激酶磷酸酶－1（MKP－1）及细胞外信号调节激酶（ERK－1）的表达,并观察用磷酸钙共沉淀方法转染MKP－1基因对血管紧张素Ⅱ（Ang Ⅱ）刺激平滑肌细胞（VSMC）^3H－胸腺叫啶（^3H-TdR)掺入的影响,以探讨MKP－1在细胞增殖中的调节作用。结果：①与WKY大鼠相比,SHR心脏和主动脉MKP－1呈低表达,分别降低53％和45％（P均＜0．01）;而SHR心脏和主动脉ERK－1呈明显高表达（P均＜0．01）,SHR心脏和主动脉ERK－1与MKP－1蛋白比值明显高于WKY。②AngⅡ 10^-7mol/L刺激VSMC增殖较对照组增加257％（P＜0．01）,转染野生型MKP－1基因细胞可使AngⅡ刺激的^3H-TdR掺入较未转染的细胞降低63％（P＜0．05）,转染突变型MKP－1基因和转染空载体的VSMC对AngⅡ的刺激与单纯AngⅡ组相比无明显抑制作用（P＞0．05）。结论：SHR心血管组织中促增殖肥大的ERK－1表达较其失活的MKP－1占优势,并且MKP－1可显著抑制AngⅡ的VSMC增殖。     

17β-雌二醇和跑台运动对去卵巢大鼠骨组织及子宫ERα蛋白表达影响的对比研究

目的：雌激素受体α（ERα）作为雌激素调节骨效应的主要受体在雌激素对骨量和骨代谢的调节中发挥重要的作用,为此本研究比较了17β-雌二醇（E2）和跑台运动对去卵巢大鼠骨组织和子宫ERα蛋白表达影响的异同。方法：将40只健康3月龄雌性SD大鼠,按体重分层后随机分为假手术、去卵巢、雌激素和运动四个组。手术1周后,雌激素组大鼠每周按体重颈部皮下注射三次17β-雌二醇,每次25μg／kg体重。运动组每周进行4次45min、速度18m／min、坡度5°的跑台训练。连续给药或运动处理14周后。采用放射免疫法和免疫组织化学法分别检测血清E2水平以及胫骨和子宫ERα蛋白表达的变化。结果：大鼠去卵巢后,子宫重量、子宫重量指数和血清E2水平显著下降,补充外源性17β-雌二醇后,三项指标均显著增加;但运动处理只能增加血清E2水平,对子宫重量和子宫重量指数均无显著影响。子宫EIh蛋白免疫组化结果显示：ERα在去卵巢组子宫内膜腔上皮、腺上皮及基质中的表达显著低于假手术组,而在雌激素组中的表达高于去卵巢组,运动组各部位ERα表达虽有所增加,但是增加的程度远低于雌激素组。胫骨近端ERα蛋白免疫组化结果显示：去卵巢后,胫骨近端骨骺端软骨细胞的细胞核内ERα表达减少,运动和雌激素干预后,胫骨近端ERα表达增加。结论：运动和雌激素处理均能刺激去卵巢大鼠子宫和骨组织ERα蛋白的表达,但运动处理无雌激素处理增加子宫重量的副作用。可见在绝经后骨质疏松的预防中,运动处理可能要优于雌激素处理。     

Expression of cancer stem cell marker during 4-nitroquinoline 1-oxide-induced rat tongue carcinogenesis

One of the theories regarding oral carcinogenesis is that the tumor growth is initiated from cancer stem cells (CSCs) that self-renew and give rise to differentiated tumor cells, like stem cells do in normal tissues. The most common methods of CSC identification are based on CSC marker expression in carcinogenesis. This study examined the expression of CD133 and CD44, the most commonly used CSC biomarkers in oral squamous cell sarcoma (SCC), with the goal of identifying molecular biomarkers whose expression is associated with the multistep oral carcinogenesis. The expression of CD133, CD44, proliferating cell nuclear antigen (PCNA), and Cytokeratin (CK) was examined by Western blot analysis and confirmed by immunohistochemistry in a 4-nitroquinoline 1-oxide-induced rat tongue carcinogenesis model. Also, the expression of aldehyde dehydrogenase 1 (ALDH1), OCT-4 and Nanog were investigated for alteration of cancer cell stemness by Western blot. Along with the progress of multistep carcinogenesis, there were slight increases of CD133 and CD44 expression in the dysplasia group compared with normal rats. However, CD133 protein level was significantly overexpressed in SCC. The expression of PCNA and CK were low in normal group, but sequentially increased in SCC. ALDH1, Nanog and OCT-4 expression were significantly increased according to SCC grade during carcinogenesis. The findings indicate that CD133 is useful in identifying oral CSCs, which suggests that CD133 may serve as a predictor to identify CSCs with a high risk of oral cancer development.     

Molecular signaling in cancer stem cells of tongue squamous cell carcinoma: Therapeutic implications and challenges

Head and neck squamous cell carcinoma is the seventh most common cancer worldwide with high mortality rates. Amongst oral cavity cancers, tongue carcinoma is a very common and aggressive oral cavity carcinoma. Despite the implementation of a multimodality treatment regime including surgical intervention, chemo-radiation as well as targeted therapy, tongue carcinoma shows a poor overall 5-year survival pattern, which is attributed to therapy resistance and recurrence of the disease. The presence of a rare population, i.e., cancer stem cells (CSCs) within the tumor, are involved in therapy resistance, recurrence, and distant metastasis that results in poor survival patterns. Therapeutic agents targeting CSCs have been in clinical trials, although they are unable to reach into therapy stage which is due to their failure in trials. A more detailed understanding of the CSCs is essential for identifying efficient targets. Molecular signaling pathways, which are differentially regulated in the CSCs, are one of the promising targets to manipulate the CSCs that would provide an improved outcome. In this review, we summarize the current understanding of molecular signaling associated with the maintenance and regulation of CSCs in tongue squamous cell carcinoma in order to emphasize the need of the hour to get a deeper understanding to unravel novel targets.     

Effects of flavonoids on tongue squamous cell carcinoma

Squamous cell carcinoma (SCC) of the tongue is associated with tobacco use, alcohol abuse, and human papillomavirus (HPV) infections. While clinical outcomes have recently improved for HPV‐positive patients in general, 50% of patients suffering from tongue cancer die within 5 years of being diagnosed. Flavonoids are secondary plant metabolites with a wide range of biological activities including antioxidant, anti‐inflammatory, and anticancer activities. Flavonoids have generated high interest as therapeutic agents owing to their low toxicity and their effects on a large variety of cancer cell types. In this literature review, we evaluate the actions of flavonoids on SCC of the tongue demonstrated in both in vivo and in vitro models.     

Beta-keratin expression in avian tongue cell aggregates.

Rotation-mediated aggregation was used to test the capacity of early embryonic tongue cells from different regions of the tongue to reorganize histotypically and terminally differentiate in vitro. Cells dissociated from the anterior ventral (AV) or posterior ventral (PV) surface of the 12 day embryonic tongue were cultured 6 days, fixed, and embedded in paraffin. Indirect immunofluorescence microscopy was used to demonstrate the occurrence and distribution of beta-keratins, the presence of which represented regional and differentiation specific markers of late tongue development. Aggregates of AV cells were organized into a beta-keratin-producing stratified epithelium. Similar PV aggregates formed only an alpha-keratin-producing stratified epithelium. The epithelial cells of these tongue tissues constructs expressed alpha- and beta-keratins in a manner consistent with the temporal and histotypical expression of keratins observed in vivo.     

Results of irradiation in cancer of the lip,tongue and ear

CANCER OF THE LIP: The primary lesion can be controlled by irradiation in approximately 80 per cent of cases. For lesions with metastases there is only about a 25 per cent chance of five-year arrest (irradiation of the primary lesion followed by excision of involved nodes). CANCER OF THE TONGUE: Lesions in the anterior two-thirds are controllable by irradiation in about 50 per cent of cases if the nodes are not involved; the salvage is only about 15 per cent if the nodes are involved (nodes treated surgically). Lesions in the posterior third of the tongue are seldom controlled in the author's experience. CANCER OF THE EAR (AURICLE): Five-year arrest of basal-cell lesions should be attained by irradiation in about 80 per cent of cases; of squamous-cell lesions in about 60 per cent. If the lesion is extensive, radiation does not offer a superior cosmetic result to operation and entails danger of late chondronecrosis. Therefore extensive lesions are probably best treated surgically. In either event, it appears probable that results of irradiation can be improved by the use of more adequate fields and greater fractionation.     

Retracted: CRNDE promotes cell tongue squamous cell carcinoma cell growth and invasion through suppressing miR-384

Tongue squamous cell carcinoma (TSCC) is the most common type of oral cancer and is an aggressive head and neck malignancy. Increasing studies have demonstrated that long noncoding RNAs (lncRNAs) play important roles in diverse biological cell processes, such as cell development, fate decisions, cell differentiation, cell migration, and invasion. In our study, we showed that long noncoding RNA colorectal neoplasia differentially expressed (CRNDE) expression was upregulated in TSCC cell lines and tissues. Overexpression of CRNDE increased the TSCC cell proliferation, cell cycle, and cell invasion. Moreover, ectopic expression of CRNDE inhibited the miR-384 expression in the SCC1 cell and increased the Kirsten Ras (KRAS), cell division cycle 42, and insulin receptor substrate 1 expression, which were the direct target genes of miR-384. We demonstrated that the miR-384 expression was downregulated in the TSCC samples compared with the paired adjacent nontumor samples. The expression of CRNDE was negatively correlated with the expression of miR-384 in the TSCC samples. Overexpression of miR-384 suppressed TSCC cell proliferation, cell cycle, and invasion. Furthermore, we demonstrated that CRNDE promoted TSCC cell proliferation and invasion through inhibiting miR-384 expression. These results suggested that CRNDE acts as an oncogene in the development of TSCC, which partially occurs through inhibiting miR-384 expression.     

Notch1 regulates tongue cancer cells proliferation,apoptosis and invasion

Objectives: Notch1 regulates tumor biology in a complex, context-dependent manner. The roles of Notch1 in tongue cancer are still controversial. The aim of this study is to investigate the roles of Notch1 in tongue cancer.

Materials and Methods: The expression of Notch1 was tested between tongue cancer and normal samples by using immunohistochemistry. Tongue cancer cells were transfected with siRNA or plasmid, respectively. Cell proliferation, apoptosis, migration and invasion ability were tested in appropriate ways. The subcutaneous tumor model was established to observe the tumor growth.

Results: Notch1 was upregulated in tongue carcinoma tissues and the expression of Notch1 was related with tumor stage and differentiation. Overexpression of Notch1 could increase tongue cancer cells proliferation, invasion and migration. But inhibited the expression of Notch1 could decrease cells proliferation, invasion and migration and promote cell apoptosis in vitro and in vivo.

Conclusion: Our results prove that the oncogenic role of Notch1 in tongue cancer and provide the direction of targeted therapy of tongue cancer.     

SOD2 is a C-myc target gene that promotes the migration and invasion of tongue squamous cell carcinoma involving cancer stem-like cells

Our previous studies revealed that manganese superoxide dismutase (SOD2) contributes to the migration and invasion of tongue squamous cell carcinoma (TSCC). The purpose of the current study was to further clarify the mechanisms of SOD2 in the migration and invasion of TSCC. Side population (SP) cells were used as cancer stem-like cells and further assessed by sphere and colony formation assays, and the expression of stem cell markers (Bmi1, Nanog and ABCG2). We found that UM1 cells (TSCC cells with increased SOD2 expression, migration and invasion abilities) possessed a higher proportion of SP cells, sphere and colony formation, and expressed a higher level of stem cell markers compared to UM2 cells (reduced SOD2 expression, migration and invasion abilities). SOD2 expression as well as migration and invasion abilities were enhanced in SP cells compared to non-SP cells. Knockdown of SOD2 in UM1 cells or SP cells inhibited the migration and invasion abilities, reduced sphere and colony formation, and the expression of stem cell markers. Direct binding of the C-myc protein to the SOD2 promoter was demonstrated by chromatin immunoprecipitation and luciferase assays. Knockdown of C-myc in UM1 cells inhibited SOD2 expression as well as migration and invasion abilities. Our results indicate that cancer stem-like cells play an important role in the migration and invasion of TSCC. SOD2 is a direct target gene of C-myc and C-myc-SOD2-mediated migration and invasion of TSCC involve cancer stem-like cells.     

Plasma cell granuloma of the tongue. Report of a case

A case of plasma cell granuloma of the tongue in an otherwise symptomless 48-year-old caucasian female is reported. The polyclonal nature of the plasmocytes was revealed by immunostaining of kappa and lambda light chains. Electron microscopic observations showed typical mature plasmocytes. A parasitic etiology of this type of lesion is suggested.