Direct observation of living laryngeal carcinoma cells during invasion of healthy cell formation.

After cloning of laryngeal carcinoma cells from tumor fragments obtained during surgery, the behavior of pure tumor cells was investigated under the inverted microscope. In tissue culture flasks, separately grown tumor cell layers and fibroblast layers were cocultured to visualize the invasion of malignant cells into healthy tissue. Using a time-lapse camera, the cells were recorded. The tumor cells often penetrated the healthy cell formation in a wedge-shaped cell formation. Near the tumor front, an elevated cell motility was found accompanied by continuous changes in the cell shape. Intercellular gaps were broader in the front line than in the more posterior parts of the tumor cell formation. The profound motility of the carcinoma cell enabled selected cells to leave the tumor cell formation completely and to migrate as single cells through healthy tissue. Eventually, some of the single tumor cells migrating through the fibroblast layer moved back to the tumor cell formation. Thus they left healthy appearing tissue behind which was earlier infiltrated by a malignant cell.     

白藜芦醇抑制喉癌Hep-2细胞系的生长

目的:研究白藜芦醇对喉癌细胞系Hep-2细胞生长的抑制作用。方法:在显微镜下计算细胞密度,MTT法绘制细胞生长曲线并计算细胞生长抑制率,利用软琼脂集落形成实验在倒置纤维镜下观察白藜芦醇处理Hep-2细胞前后细胞形态学变化及计算集落形成率。结果:各浓度的白藜芦醇均可使细胞生长曲线降低,细胞倍增时间延长。白藜芦醇也可以明显抑制Hep-2细胞形成集落,并发现白藜芦醇对细胞形态的影响与药物的剂量有明显的关系。结论:白藜芦醇对喉癌细胞系Hep-2细胞生长的抑制具有时间和剂量依赖性。     

Study of the mechanism of supporting cells repairing the organ of Corti in terms of cell kinetics--nuclear DNA synthesis of supporting cell of the organ of Corti in the cochlea damaged by nitromin administration

3H-thymidine autoradiography in vivo was carried out on normal mature mice, whose organs of Corti were securely damaged by Nitrogen Mustard-N-oxide (nitromin). The cell kinetics of supporting cells of the organ of Corti were examined and relation between these changes and mechanisms of supporting cells to maintain the organ of Corti was discussed in this paper. A few grains in various kinds of supporting cells in S stage, which were not found in control mice without nitromin injection, were discovered. Those were detected in Hensen's cell, Deiters' cell, Claudius' cells, and inner phalangeal cell. Mitosis was seen in one Claudius' cell in addition. Therefore some supporting cells seem to be able to synthesize DNA and proliferate in the acutely damaged organ of Corti even in mature cochlea and those in G0 stage begin to go around cell cycle as occasion demands. Although labelled supporting cells decreased over time, the portion of supporting cell is likely to change in proportion as the extension of damage in the organ of Corti. If this new dynamic change of cell kinetics in the acutely damaged organ of Corti means the reaction to repair the organ of Corti, the supporting cells seem to have a reasonable role to maintain the organ of Corti including repairment of reticular lamina.     

Small cell anaplastic carcinoma of the larynx: Review of the literature and report of a case

The case study of a patient with small cell anaplastic carcinoma (oat cell) of the larynx is detailed and the literature is reviewed. Small cell anaplastic carcinoma of the larynx has been demonstrated to be histologically identical to small cell anaplastic carcinoma of bronchogenic origin. The aggressive biologic behavior of this tumor justifies managing small cell anaplastic carcinoma as a systemic disease. Because of early widespread dissemination of tumor, surgery or radiotherapy alone or in combination have not been successful in controlling the disease. The combination of radiotherapy with chemotherapy has been shown to be the most effective approach to the treatment of small cell anaplastic carcinoma of the lung. We believe that a similar regimen should be considered the treatment of choice for small cell anaplastic carcinoma of the larynx.     

CD133与喉癌肿瘤干细胞的实验研究

目的探讨CD133在人喉癌细胞系Hep-2中的表达，观察纯化的CD133阳性肿瘤细胞、CD133阴性肿瘤细胞及未分选Hep-2细胞在重症联合免疫缺陷小鼠中的成瘤性，筛选Hep-2细胞系肿瘤干细胞的表型。方法流式细胞仪检测CD133在Hep-2细胞系中的表达，免疫磁珠分选技术纯化CD133阳性肿瘤细胞，分选所得各细胞亚群细胞以及未分选细胞以一定的数量级注人重症联合免疫缺陷小鼠腹部皮下，比较其成瘤差异性。分选后的细胞进行了细胞周期的分析和HE染色观察生长形态，以排除成瘤差异由细胞周期分布不同及异源细胞引起。结果流式细胞仪示CD133在Hep-2细胞系中呈微量恒定表达，表达概率（3．15±0．83）％。免疫磁珠富集的CD133阳性肿瘤细胞并不处于细胞周期生长旺盛时相或优势分裂时相，而是与分选前周期分布相似且均匀的一类细胞；HE染色示细胞形态亦符合恶性肿瘤细胞的病理生长特性。体外成瘤试验显示16／20个CD133阳性细胞注射部位、7／20个CD133阴性细胞注射部位、10／20个未分选细胞注射部位可见肿瘤生长，统计学分析表明CD133阳性肿瘤细胞较CD133阴性细胞（χ^2=8．286，P=0．004）、未分选细胞（X2=3．956，P=0．047）在重症联合免疫缺陷小鼠体内具有很强的成瘤性。结论喉癌Hep-2细胞系中，CD133阳性癌细胞具有强的体内增殖能力，CD133为喉癌肿瘤干细胞的标志之一。     

Histologic correlation of expression of Ki-67 in squamous cell carcinoma of the glottis according to the degree of cell differentiation

IntroductionSquamous cell carcinoma is the most common neoplasm of the larynx and glottis, and its prognosis depends on the size of the lesion, level of local invasion, cervical lymphatic spread, and presence of distant metastases. Ki-67 (MKI67) is a protein present in the core, whose function is related to cell proliferation.AimTo evaluate the expression of marker Ki-67 in squamous cell carcinoma of the larynx and glottis and its correlation to pathological findings.MethodsExperimental study with immunohistochemistry analysis of Ki-67, calculating the percentage of the cell proliferation index in glottic squamous cell carcinomas.ResultsSixteen cases were analyzed, with six well-differentiated and 10 poorly/moderately differentiated tumors. There was a correlation between cell proliferation index and degree of cell differentiation, with higher proliferation in poorly/moderately differentiated tumors.ConclusionThe cell proliferation index, as measured by Ki-67, may be useful in the characterization of histological degree in glottic squamous cell tumors.     

Basaloid squamous cell carcinoma of the head and neck

PURPOSE OF REVIEW: Basaloid squamous cell carcinoma is an uncommon variant of squamous cell carcinoma and was first described as a distinct entity in 1986. Basaloid squamous cell carcinoma seems to have a poorer survival rate than classical squamous cell carcinoma. On the basis of a critical literature survey, we attempt to evaluate if basaloid squamous cell carcinoma is really more aggressive and presents a poorer outcome than squamous cell carcinoma. RECENT FINDINGS: All papers are retrospective, and most include small numbers of cases, which are further diminished when subdivided according to specific sites. Only in three studies was basaloid squamous cell carcinoma of the head and neck region compared with matched squamous cell carcinoma controls. These studies did not show a uniform tendency regarding the aggressiveness and outcome of basaloid squamous cell carcinoma. In addition, several recent papers confirmed the presumed greater aggressiveness and worse outcome, and other recent papers questioned these characteristics. SUMMARY: The presented literature survey does not permit conclusions regarding the aggressiveness and outcome of basaloid squamous cell carcinoma compared with squamous cell carcinoma. Greater numbers of basaloid squamous cell carcinoma should be studied and compared with site-matched, stage-matched, and age-matched controls of conventional squamous cell carcinoma.     

Production of lymphokine-activated lymphocytes. Lysis of human head and neck squamous cell carcinoma cell lines

Lymphokine-activated killer cells are thought to be important mediators of host tumor defense. In the present study, the cytotoxic potential of lymphokine-activated lymphocytes against different head and neck squamous cell carcinoma cell lines was investigated. Lymphokine-activated killer cells were derived from peripheral blood lymphocytes. Effector peripheral blood lymphocyte cell suspensions were incubated in the presence or absence of recombinant interleukin-2. Cytotoxicity of incubated cells or fresh peripheral blood lymphocytes was determined in a 3-hour chromium 51 release assay. Target cell lines included K562 (a natural killer-sensitive target) and the following head and neck squamous cell carcinoma cell lines: Cal 27, UMSCC-1, UMSCC-8, UMSCC-16, UMSCC-19, and UMSCC-22a. Fresh peripheral blood lymphocytes and peripheral blood lymphocytes cultured in the absence of added interleukin-2 demonstrated minimal cytotoxic effects against the squamous cell carcinoma targets. In contrast, these fresh and incubated lymphocytes showed significant cytotoxic effects against K562. Cells preincubated in the presence of interleukin-2 demonstrated a statistically significant increase in cytotoxic effects against K562 and all squamous cell carcinoma targets. These investigations support the possible role of lymphokine-activated killer cells in host defense against squamous cell carcinoma. In vitro natural killer cell activity against head and neck squamous cell carcinoma cell lines is low; however, significant lymphokine-activated killer cell cytotoxicity is present.     

中药土贝母对Hep-2细胞株诱导分化的研究

目的：为了观察土贝母对Ｈｅｐ－２细胞株诱导调亡的机制。方法：应用ＭＴＴ比色法和流式细 胞仪测定细胞周期的变化，并应用电子显微镜技术观察诱导分化后亚细胞结构。结果：土贝母制剂对 Ｈｅｐ－２细胞株有明显的抑制作用，且呈剂量依赖性，流式细胞仪分析，不同的药物浓度和不同的作用时 间凋亡的发生差异均有显著意义，Ｇ１期细胞堆积，Ｓ期细胞减少，Ｇ２／Ｍ期细胞相对增多，亚细胞结构， 细胞线粒体肿胀变性，核固缩。结论：土贝母制剂能阻上Ｇ０／Ｇ１期细胞向Ｓ期转化进程，抑制Ｈｅｐ－２增 殖，促进Ｈｅｐ－２细胞凋亡。     

Qualitative and quantitative observations of spiral ganglion development in the rat

The postnatal development of the spiral ganglion cells in the rat was studied from birth until the adult stage. At birth, a single population of ganglion cells is present. Some of them are surrounded by one or two layers of satellite cell processes. With maturation, the satellite cell processes increase in number around the cell body and its processes. At the end of the first postnatal week, two important events occur. The first is the appearance of myelin lamellae between the 4th and the 6th postnatal day in both ganglion cell processes, and between the 6th and the 8th day in the cell body. The second event is the appearance of a new type of cell (the Type II spiral ganglion cell) on the 6th to the 8th day postpartum. At this stage, the Type II cell is mainly characterized by densely packed neurofilamentous structures in the cytoplasm. Comparison between the myelination of the cell body and its processes reveals three main differences! There is a time lag of approximately 2 days between the onset of myelination in the cell body and in its processes. The kinetics of myelination are different in the cell processes and in the cell body. The myelination of the cell body starts slowly, whereas it is very fast in the processes. Later, the kinetics of myelination decrease in the processes, and increase in the cell body. At all stages including the adult, the fibers have a myelin sheath composed of more lamellae than the cell body. These observations are discussed with respect to development in other species.     

Maspin蛋白在鼻咽癌放射敏感CNE2细胞株中的表达分析

目的探讨放射敏感鼻咽癌CNE2细胞株在放射条件下丝氨酸蛋白酶抑制剂Maspin蛋白的反应性变化,分析鼻咽癌放射作用的分子机制。方法采用MTT比色法检测放射线对鼻咽癌CNE2细胞株生长的影响,胎盼蓝染色法检测CNE2细胞增殖,流式细胞术检测CNE2细胞放射后的凋亡及周期改变,Western blotting分析Maspin蛋白的表达。结果鼻咽癌CNE2细胞株放射后,细胞周期重新分布,出现较多的G2/M期细胞以及较少的G0/G1期细胞;增殖受到抑制、细胞凋亡明显、Maspin蛋白表达上调,且其上调与细胞凋亡率增高呈现一致性改变,也与细胞周期分布改变相关。结论鼻咽癌CNE2细胞对放射线敏感、容易被诱发凋亡;细胞周期参与了放射诱导的CNE2放射反应调节;放射提高了CNE2细胞的Maspin蛋白表达,提示该蛋白参与了CNE2细胞的放射反应;Maspin蛋白表达提高涉及CNE2细胞周期和凋亡改变,是一个值得深入研究的鼻咽癌放射敏感相关分子。     

A cell proteomic approach for the detection of secretable biomarkers of invasiveness in oral squamous cell carcinoma

OBJECTIVE: To identify potential biomarkers of invasiveness in oral squamous cell carcinoma. DESIGN: A pilot proteomic study for the identification of secreted and cleaved proteins that can serve as potential biomarkers for head and neck carcinoma invasiveness. SUBJECTS: Two primary cell lines and their variants were established from 2 oral squamous cell carcinoma human tissue samples with distinct invasive phenotypes. The cell lines were confirmed to maintain the invasive capacity of the original cancer when implanted into the tongues of immunocompromised RAG-2/gamma(c) mice. INTERVENTIONS: Invasiveness was assessed by the capacity of cells to invade through a matrigel matrix using the Boyden chamber assay and correlated with the invasiveness seen clinically and histologically in patients. In parallel, cell lines were grown in serum-free conditioned medium, which was then used to identify secreted and/or cleaved proteins that emanate from cancer cells, using 2-dimensional gel electrophoresis and matrix-assisted laser desorption-ionization combined with tandem mass spectrometry. RESULTS: The invasion assays revealed a correlation between cell migration capacity through matrigel matrix and the aggressive phenotype seen in the clinical and histopathological assessments. More than 50 proteins were identified as being differentially secreted in media between the least and the more aggressive cell lines (P < .05). These include proteins that regulate cell metabolism, cell structure, cell adhesion, and cell motility, as well as proteins with undefined function. CONCLUSIONS: We report a sensitive and clinically relevant approach to screen for secreted biomarkers of oral squamous cell carcinoma invasiveness using proteomic technology. Both high- and low-abundant secreted proteins were identified and can represent potential biomarkers for oral cancer.     

Primary description of a new entity, renal cell-like carcinoma of the nasal cavity: van Meegeren in the house of Vermeer

BACKGROUND: Few sinonasal malignancies can manifest, histologically, as clear cell neoplasia. The most likely such tumor to be encountered is metastatic renal cell carcinoma. Primary sinonasal tumors that can appear as clear cell malignancies include squamous cell carcinoma and mucoepidermoid carcinoma. Primary salivary clear cell carcinoma occurs almost exclusively in the oral cavity and has not been described in the nasal cavity. OBJECTIVE: To report a unique sinonasal clear cell malignancy that mimicked metastatic renal carcinoma. STUDY DESIGN: Case report. OUTCOME MEASUREMENTS: Radiography, histology, histochemistry, immunohistochemistry, and electron microscopy. RESULTS: Histologically, the tumor was identical to renal cell carcinoma. No evidence of renal malignancy was found by abdominal computed tomographic scan or gadolinium-enhanced magnetic resonance imaging. Histochemistry confirmed the presence of tumor glycogen but no mucin. Immunohistochemistry confirmed strong expression of low- and high-molecular-weight keratin and S100, and no vimentin expression. Electron microscopy showed tumor myofibroblastic differentiation and cytoplasmic glycogen, neutral lipid vacuoles, and cholesterol. CONCLUSIONS: There was no clinical evidence of renal cell carcinoma. The immunohistochemical and ultrastructural findings were inconsistent with the diagnosis of renal cell carcinoma and showed features also inconsistent with the diagnosis of primary salivary clear cell carcinoma. We therefore conclude that this tumor represents a new and distinct entity, notable in its presentation as a "counterfeit renal cell carcinoma."     

紫杉醇对人喉鳞状细胞癌Hep-2放射增敏作用的实验研究

目的 在体外用细胞生物学实验技术了解紫杉醇联合直线加速器X射线照射对Hep-2细胞株的作用，以及紫杉醇对Hep-2细胞株细胞周期的影响，并探讨其作用机制。方法 ①用MTT法计算Hep-2细胞经直线加速器照射组；先紫杉醇作用12、18、24、30h后再直线加速器照射组；先直线加速器照射再联合紫杉醇作用12、18、24、30h组各组细胞生长抑制率。②应用流式细胞仪检测Hep-2细胞经紫杉醇作用12、18、24、30h后各组细胞周期分布。结果 ①先紫杉醇作用再直线加速器照射组细胞生长抑制率较高（各组均值达到87．51％～89．64％），与先照射再联合紫杉醇组细胞生长抑制率（各组均值为57．84％～64．34％）及单纯照射组细胞生长抑制率（均值为41．90％）相比有统计学意义（P〈0．01）。②用紫杉醇后Hep-2细胞株细胞周期停滞于G2／M期，G2／M期停滞于24-30h达到峰值（24．65％～25．15％），并在用药后30h观察到凋亡峰。结论 紫杉醇有放射增敏作用，其可能机制是使细胞生长停滞于射线敏感期G2／M期，从而加强射线对肿瘤细胞的杀伤效应。     

大鼠椭圆囊感觉上皮细胞长期培养体系的建立及毛细胞标记物表达的意义

目的：建立椭圆囊感觉上皮细胞（USEC）长期培养体系，为内耳毛细胞再生研究提供稳定的活性细胞。方法：取1d龄wistar大鼠，在显微镜下取出椭圆囊上皮，嗜热菌蛋白酶处理，获得具活性的纯椭圆囊感觉上皮，胰蛋白酶＋胶原酶消化，培养传代，传25代。取第25代USEC在倒置显微镜、透射电镜下对USEC的生长特征、形态结构进行观察；免疫细胞化学检测细胞角蛋白18、波形蛋白、Brn3．a及Calretinin的表达。RT—PCR检测毛细胞的特征性标记物AchRa9、MyosinVIIamRNA的表达。结果：USEC细胞培养达6个月传25代，第25代USEC均呈扁平、多角形、核大而圆的上皮细胞形态，细胞之间连接紧密，形成单层时均呈“铺路石样”外观，可见dome结构。该细胞表达角蛋白18和不表达波形蛋白，微绒毛丰富，细胞间连接紧密，提示其上皮起源。第25代USEC表达毛细胞的特征性标志物Brn3．a、Calretinin及AchRa9、Myosin Ⅶa mRNA，表明长期培养USEC仍具有毛细胞的前体细胞的特性。结论：本实验成功建立了USEC长期培养体系。长期培养的USEC性状未发生明显改变，表达上皮细胞及毛细胞的特征性标志物，具有毛细胞前体细胞的特征。这为毛细胞再生的机制及内耳分子、基因研究提供了稳定的细胞来源。     

EGCG对鼻咽癌细胞株CNE-2及相关基因表达的影响

目的:研究EGCG对鼻咽癌细胞系CNE-2的增殖与凋亡作用,探讨其对细胞增殖与凋亡相关基因Bcl-2、Bax、Caspase-3表达的影响。方法:应用MTT实验、流式细胞仪检测细胞的增殖与周期;Hoechst33258荧光染色法检测细胞凋亡;RT-PCR检测Bcl-2、Bax、Caspase-3表达。结果:EGCG明显抑制CNE-2细胞的增殖,诱导其凋亡,该作用具有剂量-效应关系。EGCG抑制CNE-2细胞Bcl-2表达,诱导CNE-2细胞Bax、Caspase-3表达。结论:EGCG在体外具有抗鼻咽癌细胞的作用,此作用可能是通过调节细胞增殖与凋亡基因的表达实现的。     

Cell turnover in neuromasts of zebrafish larvae

The numbers and positions of cells undergoing cell death and proliferation in the neuromasts of 10 day old zebrafish larvae were assessed to investigate the ability of supporting cells to differentiate into hair cells. Evaluations of cell death and proliferation showed that a subpopulation of cells located in the centre of the neuromast undergo cell death, and a different subpopulation located at the periphery proliferate. This suggests that cell death of hair cells and proliferation of mantle supporting cells occurs as part of normal development, creating constant turnover of hair cells. We show that the caspase inhibitor zVADfmk reduces cell death while the aminoglycoside neomycin specifically induces an increased amount of cell death in the central population of cells. Both of these treatments affect the rate of proliferation of the peripheral subpopulation of cells in the neuromast suggesting that a feedback mechanism occurs regulating cell death and proliferation. We propose that the dying population of cells are hair cells and the proliferating cells are 'mantle' supporting cells, which is in agreement with previous observations suggesting that supporting cells can give rise to hair cells following hair cell death.     

Determination of hair cell degeneration and hair cell death in neomycin treated cultures of the neonatal rat cochlea

The spatial-temporal course of hair cell degeneration and hair cell death was examined in the mammalian cochlea following aminoglycoside treatment. Organotypic cultures were established from postnatal rats (P3) and treated with 1 mM neomycin sulfate for 12-48 h and analyzed using a live/dead assay under epifluorescence microscopy. Live hair cells were labeled with calcein, a probe whose fluorescence and cellular retention depends upon intracellular esterase activity and cell-membrane integrity, respectively. Hair cell death was determined by ethidium homodimer-1, a probe that can enter cells with compromised cell membranes only. Inside the cell it binds to DNA. Hair cell morphology was also examined using phalloidin labeling, scanning electron microscopy and semi-thin section analysis. Results showed that hair cell degeneration and hair cell death occurred in a time dependent gradient from base to apex. After 48 h of neomycin treatment, most apical hair cells survived while most basal hair cells died. Calcein labeling provides a sensitive functional assay for measuring hair cell survival.     

Functions of the mastoid cell system: auto-regulation of temperature and gas pressure

This article presents a new approach to understanding the physiological functions of the mastoid cell system. It is suggested that the cell system, in combination with the continuous blood flow through the adjacent large vessels, makes up a compound functional unit that serves to protect the sensitive vestibular part of the inner ear from inadequate stimulation by external temperature changes. By virtue of the large surface area of the cell system mucosa with respect to the enclosed gas volume, the mastoid cell system may also work as a pressure regulator. Variations of the bi-directional exchange of fluid over the capillary network in the mucosa will change the size of the lumen that is available for the gas in the cell system. Volumes of gas and fluid can thus be exchanged to keep the intratympanic pressure within physiological limits. The process is most effective in a cell system with a high area-to-volume ratio.     

mAb-EGFR功能化修饰的金纳米棒靶向光热作用对人咽鳞状细胞癌FaDu细胞株和293T细胞株的细胞毒性分析

目的：探讨mAb-EGFR功能化修饰的金纳米棒靶向光热作用对人咽鳞状细胞癌FaDu细胞株和对非肿瘤细胞的细胞毒性,初步明确特定吸光度的金纳米棒在体外实验中所需剂量。方法：在CTAB体系下合成实验所需金纳米棒并且完成EGFR单克隆抗体功能化修饰,用紫外分光光度计及电镜表征,以体外培养的人咽鳞状细胞癌FaDu细胞和非肿瘤细胞293T细胞系为实验材料,用Western blot免疫印迹法定性比较两株细胞EGFR的表达,及明确细胞内吞金纳米棒。设定不同剂量金纳米棒浓度（15%、25%、35%）,乳酸脱氢酶法比较金纳米棒对FaDu细胞株和293T细胞株毒性差异及初步明确实验剂量。结果：金纳米棒对于FaDu细胞株的细胞毒性高于293T细胞株（P〈0.01）,在293T细胞株实验组中金纳米棒剂量浓度15%、25%两组间无明显差异,而这两组与35%剂量比较均差异有统计学意义（P〈0.01）。结论：mAb-EGFR功能化修饰的金纳米棒在近红外激光照射时对人咽鳞状细胞癌细胞作用表现出比非肿瘤细胞更高的细胞毒性,对于特定吸光度功能化修饰的金纳米棒光热作用杀伤人咽鳞状细胞癌FaDu细胞株的安全剂量浓度为15%~25%。