肿瘤坏死因子相关凋亡诱导配体在多种疾病中的生物学作用

肿瘤坏死因子相关凋亡诱导配体（Apo2L/TRAIL）是肿瘤坏死因子家族的一员,可以诱导表达特异性死亡受体TRAIL-R1 （DR4）或TRAIL-R2 （DR5）的细胞凋亡.TRAIL可诱导多种肿瘤细胞凋亡,对正常细胞无杀伤性,因此将其作为抗肿瘤靶向治疗候选药物之一,备受关注.近期研究发现,TRAIL既通过其受体介导细胞凋亡信号通路,又可传导非凋亡的信号通路,在自身免疫病和感染性疾病中发挥重要生物学作用.因而简单阐明TRAIL在人类多种疾病中生物学作用的研究进展,对今后TRAIL研究方向和治疗策略的选择具有指导意义.     

TRAIL对多种肿瘤细胞的杀伤作用

目的： 研究肿瘤坏死因子相关凋亡诱导配体（TRAIL）作用下多种肿瘤细胞的生长抑制效应及凋亡诱导情况。方法： 利用大肠杆菌基因工程菌表达非融合rhsTRAIL，进行目的蛋白的分离纯化，得到rhsTRAIL样品，纯度为97%。通过倒置显微镜下观察、MTT法、流式细胞仪法检测其对细胞的生长抑制和诱导凋亡情况。结果： 一定浓度的TRAIL 可有效抑制LS174-T细胞、MCF-7细胞、GLC细胞、7402细胞、Jurkut T细胞生长，其生长抑制率具剂量依赖性，且各细胞对TRAIL敏感性不同，其中Jurkat T细胞最为敏感。用2 mg/L TRAIL作用Jurkat T细胞0-72 h，6 h后细胞即发生明显凋亡，其细胞凋亡率具时间依赖性。结论： 所制备的TRAIL可抑制多种肿瘤细胞生长，并诱导Jurkat T细胞凋亡。     

杭白菊增强TRAIL基因诱导大肠癌细胞凋亡及机制的实验研究

目的：探讨杭白菊提取液对肿瘤坏死因子相关凋亡诱导配体基因（TRAIL）抑制人大肠癌细胞株DLD-1作用的影响及其可能机制。 方法： 杭白菊提取液联合重组腺病毒载体（Ad）介导的TRAIL基因作用于人大肠癌细胞株DLD-1，通过倒置显微镜、MTT比色法和流式细胞仪，研究分析其对DLD-1细胞抑制作用的效果。采用逆转录聚合酶链反应（RT-PCR）和流式细胞术检测杭白菊提取液作用前后DLD-1细胞TRAIL、TRAIL受体（TRAIL-Rs）mRNA以及细胞表面TRAIL蛋白表达的变化。 结果： Ad/hTERT-gTRAIL对DLD-1细胞的生长抑制率和凋亡率分别为31.4%和13.5%；联合杭白菊提取液后，生长抑制率和凋亡率均显著提高，达93.1%和45.4%（P<0.05）。杭白菊提取液作用后DLD-1细胞TRAIL mRNA的表达量从作用前的0.46上调至1.01, 细胞表面TRAIL蛋白表达的百分率从作用前的2.2%升高到5.0%（P<0.05）。TRAIL死亡受体（DR4、DR5）mRNA的表达量分别从0.70和0.22上调至1.10和0.83（P<0.05），而TRAIL诱骗受体（DcR1、DcR2）mRNA的表达量从0.60和1.15下调至0.19和0.78（P<0.05）。 结论： 杭白菊提取液联合重组腺病毒载体（Ad）介导的TRAIL基因(Ad/hTERT-gTRAIL)能有效诱导DLD-1细胞的凋亡。杭白菊提取液上调TRAIL死亡受体表达以及下调TRAIL诱骗受体的表达可能在增强TRAIL诱导的凋亡作用中起着重要作用。     

TRAIL联合紫杉醇诱导胃腺癌SGC-7901细胞凋亡作用研究

目的:观察紫杉醇联合肿瘤坏死因子相关凋亡诱导配体(TRAIL)诱导人胃腺癌SGC-7901细胞凋亡的作用及其协同作用机制。方法:将TRAIL、紫杉醇及TRAIL联合紫杉醇诱导SGC-7901细胞48小时,用流式细胞仪(FCM)检测细胞凋亡率和线粒体跨膜电位的改变;用MTT法检测SGC-7901细胞增殖反应;用免疫印迹(Westernblot)法检测TRAIL死亡受体DR4(TRAIL-R1)、DR5(TRAIL-R2)的表达变化。结果:TRAIL和/或紫杉醇对SGC-7901细胞增殖有抑制作用,两者联合用药组对细胞增殖的抑制率较单独用药明显增加(P<0.01);联合用药组细胞凋亡率较单独用药组明显增加(P<0.01);0.3μmol/L紫杉醇作用48小时后,DR4表达明显升高(P<0.05),而DR5表达没有明显改变(P>0.05)。结论:紫杉醇可协同TRAIL诱导SGC-7901细胞凋亡,DR4表达增加可能是其协同作用的机制。     

TRAIL在病毒损伤胰岛β细胞中的作用

目的：研究肿瘤坏死因子相关凋亡诱导配体（TRAIL）在病毒诱导的人胰岛β细胞损伤中的作用。方法： 用Annexin-V法检测比较柯萨奇B病毒（CVB）和风疹病毒（RV）引起的人胰岛β细胞系CM细胞凋亡，再用可溶性TRAIL受体和抗TRAIL单克隆抗体进行阻断实验。结果： 5噬斑形成单位（PFU）CVB3和CVB4作用CM细胞5 h导致30%以上CM细胞凋亡；0.01 PFU CVB3和CVB4作用CM细胞24 h导致超过80%CM细胞凋亡；10 PFU RV作用CM细胞24 h导致25%CM细胞凋亡；4种可溶性TRAIL受体以及抗TRAIL单克隆抗体对CVB所致细胞凋亡有阻断作用。结论： CM细胞对CVB更为敏感，TRAIL参与CVB诱导的CM细胞凋亡。     

hTERT启动子驱动的TRAIL诱导宫颈癌细胞的凋亡作用

目的：探讨端粒酶（hTERT）启动子驱动的肿瘤坏死因子相关凋亡诱导配体（TRAIL）对宫颈癌细胞生长抑制的作用。方法：脂质体转染将hTERT-TRAIL转染至宫颈癌细胞HeLa,RT-PCR检测TRAILmRNA的表达;通过MTT、流式细胞术检测稳定转染细胞的生长情况,电镜观察转染细胞的超微结构。结果：转染真核表达载体hTERT-TRAIL组细胞中TRAILmRNA细胞表达量高于对照组（P〈0.05）;hTERT-TRAIL组细胞对宫颈癌细胞的抑制率高于CMV-TRAIL组和对照组（P〈0.05）;hTERT-TRAIL组细胞对宫颈癌细胞的凋亡率明显高于转染CMV-TRAIL组和对照组（P〈0.01）;电镜结果显示,hTERT-TRAIL对细胞作用是以细胞凋亡为主。结论：hTERT启动子驱动的肿瘤坏死因子相关凋亡诱导配体对人宫颈癌细胞的生长有明显的抑制作用和诱导凋亡作用,为宫颈癌的研究奠定基础和临床治疗提供思路。     

大鼠肝癌发生过程中肿瘤坏死因子相关的凋亡诱导配体及核转录因子-κB的表达

目的:探讨肿瘤坏死因子相关的凋亡诱导配体(TRAIL)及核转录因子κB(NF-κB)在大鼠肝癌发生过程巾的表达及意义.方法:应用免疫组织化学SP法,对二乙基亚硝胺(DEN)诱发的大鼠肝癌发生过程中TRAIL及NF-κB的动态表达进行检测.结果:二乙基亚硝胺诱发的肝癌为肝细胞癌,大鼠肝癌癌变过程大致经过肝细胞损伤期、肝细胞增生-硬化期和肝细胞癌变期3个阶段.在正常大鼠肝、损伤期及增生硬化期偶见少量肝细胞呈TRAIL阳性表达,进入癌变期,TRAIL阳性细胞数量增多,主要位于癌结节内.在正常大鼠肝,偶见少量肝细胞呈NF-κB阳性表达,随着肝癌发生发展,阳性表达细胞逐渐增多,至诱癌晚期,可见大量NF-κB阳性表达细胞,均比正常肝组织表达高.结论:TRAIl及NF-κB可能与肝癌的发生发展密切相关.     

TRAIL及其受体DR5与动脉粥样硬化关系的初步研究

目的探讨肿瘤坏死因子(INF)相关的凋亡诱导配体(TRAIL)及其受体DR5与动脉粥样硬化(AS)之间的关系。方法冠心病(CAD)组61例,正常对照组22例。应用酶联免疫吸附法测定血浆可溶性TRAIL(sTRAIL)和可溶性DR5(sDR5)水平。免疫组织化学测定冠状动脉TRAIL和DR5蛋白表达情况。结果CAD组血浆sTRAIL和sDR5水平均显著性升高(P<0.001,P<0.05)。3支血管病变组和双支血管病变组血浆sTRAIL和sDR5水平均分别显著高于单支血管病变组和正常对照组(P<0.01,P<0.01,P<0.01,P<0.05)。TRAIL和DR5主要表达于平滑肌细胞的胞质,TRAIL还可表达于AS中的巨噬细胞。AS组冠状动脉的TRAIL和DR5表达明显高于非AS组(P<0.01,P<0.05)。结论TRAIL及其受体DR5可能参与了AS的进展,其浓度越高,冠状动脉病变越重。     

乙型肝炎病毒X基因促TRAIL诱导Huh7细胞凋亡的机制

目的探讨乙型肝炎病毒(hepatitis B virus,HBV)X基因(HBX)促肿瘤坏死因子相关凋亡诱导配体(TNF relatedapoptosis inducing ligand,TRAIL)蛋白诱导人肝癌细胞Huh7凋亡的机制。方法应用脂质体转染法将pcDNA3.1-HBX、pcDNA3.1分别转染Huh7细胞,建立稳定表达HBX的细胞模型Huh7-HBX和空载体对照细胞模型Huh7-3.1;运用TRAIL蛋白分别作用Huh7-HBX细胞、Huh7-3.1及Huh7细胞后,采用CCK8、流式细胞术检测细胞增殖和凋亡情况;Western blot检测死亡受体4(deathreceptor 4,DR4)和死亡受体5(death receptor 5,DR5)的表达情况;分光光度法检测细胞caspase8和caspase3活性。结果 RT-PCR及Western blot证实成功构建细胞株Huh7-HBX;CCK8和流式细胞术检测结果均显示Huh7-HBX细胞的凋亡率明显高于Huh7-3.1及Huh7(P<0.05);Western blot显示Huh7-HBX细胞的DR4、DR5的表达量明显高于Huh7-3.1及Huh7细胞(P<0.05);caspase活性检测结果显示,Huh7-HBX细胞caspase8及caspase3的活性明显高于Huh7-3.1及Huh7(P<0.05)。结论 HBX能够通过上调Huh-7细胞表面死亡受体DR4、DR5的表达,进而促进TRAIL蛋白诱导的细胞凋亡,为进一步研究HBX的促凋亡机制提供实验依据。     

阿司匹林增强TRAIL诱导的HepG-2细胞凋亡

目的:研究阿司匹林能否增强肿瘤坏死因子相关凋亡诱导配体(TRAIL)诱导的HepG-2细胞凋亡,并初步探讨其作用机制。方法:HepG-2细胞放入含15%胎牛血清的DMEM培养液中培养,改良苯酚-品红染色观察细胞形态变化,MTT法检测细胞增殖程度,TUNEL法检测细胞凋亡指数,RT-PCR检测细胞中survivinmRNA的表达水平,FCM检测细胞凋亡率和细胞周期。结果:不同浓度的阿司匹林联合TRAIL实验组细胞增殖抑制率明显高于空白对照组及TRAIL和阿司匹林单独用药组,且细胞凋亡指数也明显提高,凋亡率与阿司匹林的浓度呈现剂量依赖关系,联合用药组凋亡相关基因survivin的表达与空白对照组和TRAIL和阿司匹林单用组相比明显下调。结论:阿司匹林能够增强TRAIL诱导的HepG-2细胞凋亡,其作用机制可能与survivin基因的表达下调有关。     

T cells and eosinophils in bronchial smooth muscle cell death in asthma

Background Bronchial smooth muscle cells (SMC) proliferate, express adhesion molecules, secrete cytokines and thus efficiently contribute to the pathogenesis of asthma.
Objective The aim of the study was to investigate whether, and by which mechanism, T cells and eosinophils can cause death of airway SMC.
Methods The T cell- and eosinophil-induced cell death was analysed in primary human bronchial SMC cultures as well as in bronchial biopsy specimens from non-asthmatic and asthmatic individuals.
Results Bronchial SMC death showed characteristic morphological features of apoptosis in 3–6 days cultures with inflammatory cytokines (IFN-γ, TNF-α), soluble death ligands [sFasL, TNF-related apoptosis-inducing ligand (TRAIL)] and activated T-helper type 1 (Th1) and Th2 cell supernatants. The recombinant eosinophil cationic protein induced SMC necrosis within 1 h. Resting SMC expressed the death receptors TNFR1, TNFR2, Fas, TRAILR1, TRAILR2 and membrane FasL as a death-inducing ligand. IFN-γ and TNF-α up-regulated TNFR1, TNFR2, Fas and membrane FasL on SMC. TNF-α up-regulated TRAILR1 and TRAILR2; sFasL up-regulated TNFR2. The intracellular caspase-3 activation in SMC was significantly increased by IFN-γ, sFasL, TRAIL, Th1 and Th2 cell supernatants. Increased expression of TRAIL in asthmatics, but not in non-asthmatic individuals was demonstrated in situ . The apoptosis receptors TRAILR1 and TRAILR2 were expressed in SMC and epithelial cells both in healthy and asthmatic biopsies. Prominent apoptosis of SMC was observed in fatal asthma, but not intermittent asthma biopses.
Conclusion The demonstration of bronchial SMC death both by apoptosis and necrosis indicates the essential role of T cells and eosinophils in the bronchial tissue injury particularly in the severe asthma.     

Galectin-3 does not reliably distinguish benign from malignant thyroid neoplasms

AIMS: To determine whether galectin-3 is a sensitive indicator of thyroid malignancy. It has been suggested as a potential marker for differentiating thyroid carcinoma from benign or non-neoplastic lesions in preoperative fine-needle aspirates (FNAs). METHODS: Galectin-3 protein expression was assessed by immunohistochemistry in formalin-fixed thyroid tissues from 124 patients with histological diagnoses of papillary carcinoma (n = 38), follicular carcinoma (n = 19), follicular adenoma (n = 32) and dominant nodules of multinodular goitre (n = 35). Expression of galectin-3 was also assessed by Western blotting in 24 fresh thyroid tissues. RESULTS: Galectin-3 expression was observed in the majority of carcinomas (papillary 92%; follicular 74%). However, a large proportion of follicular adenomas (72%) and multinodular goitres (57%) also expressed galectin-3. In addition, galectin-3 expression was observed in epithelial cells of normal thyroid tissue and Hashimoto's thyroiditis. Galectin-3 immunopositivity was significantly greater in papillary carcinomas than in dominant nodules or follicular adenomas (P < 0.0001, P = 0.0005, respectively). However, galectin-3 expression was no greater in follicular carcinomas than in follicular adenomas (P = 0.8735). Western blotting analysis confirmed both the specificity of the antiserum and expression of galectin-3 in multinodular goitres, follicular adenomas/carcinomas and papillary carcinomas. CONCLUSION: The data demonstrate that galectin-3 is not a reliable immunohistochemical marker to distinguish benign from malignant thyroid follicular lesions.     

EGF-receptors in human normal and pathological thyroid tissue

Expression of the epidermal growth factor receptor (EGFR) was studied in cryosections from human thyroid tissues. Normal tissue (4 cases), nodular goitre (12), toxic goitre (9), adenoma (9), follicular carcinoma (1), papillary carcinoma (7) and poorly differentiated carcinoma (1) were used for immunohistochemistry. Northern blot analysis was performed in two nodular goitres, three adenomas, two papillary carcinomas, one follicular carcinoma and the adjacent normal tissue in five cases as well as in two cell lines from anaplastic carcinomas. Epidermal growth factor receptor immunoreactivity was detected in all tissues examined. The amount of EGFR mRNA did not differ between normal and abnormal tissues. However, the EGFR staining was weaker in normal thyroid tissue compared to the adjacent neoplastic areas suggesting an upregulation at the posttranslational level in the latter. A strong staining was also seen in hyperfunctioning thyroid glands. The EGFR location was mainly basal or basolateral in all thyroid tissues with normal histology and in toxic diffuse goitre. Pericellular and sometimes cytoplasmatic staining was seen in neoplastic tissues. In nodular goitre the staining was both basal, lateral and apical and varied in intensity. Our data suggest that a non-polarized location of EGFR probably indicates a loss of the normal epithelial cell polarity and could be interpreted as an early sign of dedifferentiation. Furthermore, a role for the EGFR is proposed, not only in the development of thyroid neoplasias but also in goitre formation.     

Functional expression of tumor necrosis factor-related apoptosis-inducing ligand in human colonic adenocarcinoma cells

TNF-related apoptosis-inducing ligand (TRAIL) can induce apoptosis in various transformed cell lines. Therefore, we investigated TRAIL sensitivity, TRAIL-induced nuclear factor-kappaB (NF-kappaB) activation, and expression of TRAIL in human colonic adenocarcinoma cell lines (HT-29, LS180, SK-CO-1). All four TRAIL receptors (TRAIL-R1 through TRAIL-R4) are expressed in these cell lines. TRAIL sensitivity was assessed by assay of cell viability. Cancer cell viabilities were 83 +/- 3.1% (HT-29), 90 +/- 4.3% (LS180), and 88 +/- 6.3% (SK-CO-1) at 24 hours after the addition of 100 ng/ml TRAIL, indicating that these cell lines were relatively resistant to TRAIL. Activation of NF-kappaB was variably influenced by TRAIL administration, with no consistent tendency among the cell lines, indicating that TRAIL-induced NF-kappaB activation might be cell-type dependent. In contrast, TRAIL was expressed in the human colonic adenocarcinoma cell lines by Western blotting and RT-PCR. Increased expression of TRAIL on tumor cells was observed by flow cytometry after cytokine stimulation (IFN-gamma, TNF-alpha) or the addition of chemotherapeutic agents (camptothecin, doxolubicin hydrochloride). TRAIL on HT-29 cells was functional and able to induce apoptosis in Jurkat cells. Jurkat cell viability was increased by the addition of TRAILR1-R4-Fc. In the presence of various cytokines or chemotherapeutic agents, functional TRAIL is expressed on the surface of tumor cells, and this expressed TRAIL might contribute to tumor immune privilege by inducing apoptosis of activated human lymphocytes.     

Regulation of Apo2L/tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in thyroid carcinoma cells

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL)/Apo2 ligand selectively kills neoplastic cells, including thyroid carcinoma cells (Mitsiades et al: Thyroid carcinoma cells are resistant to FAS-mediated apoptosis but sensitive to tumor necrosis factor-related apoptosis-inducing ligand. Cancer Res 2000, 60:4122-41299). We investigated the mechanisms regulating Apo2L/TRAIL-induced apoptosis in thyroid carcinoma cells, as well as the impact of insulin-like growth factor (IGF)-1, interferon-gamma, and TNF-alpha. We found that the emergence of resistance to Apo2L/TRAIL, after prolonged incubation with this cytokine, was associated with increased levels of FLICE inhibitory protein (FLIP), and was overcome by cycloheximide and bisindolylmaleimide, that specifically down-regulated FLIP expression, as well as by transfection of a FLIP anti-sense oligonucleotide. IGF-1 activated Akt; up-regulated the caspase inhibitors FLIP, cIAP-2, XIAP, and survivin; and attenuated Apo2L/TRAIL-induced apoptosis. This effect was inhibited by the IGF-1 receptor neutralizing antibody aIR3, the PI-3K inhibitor wortmannin, and the heat shock protein-90 chaperone inhibitor geldanamycin. Transfection of constitutively active Akt protected from TRAIL. Conversely, interferon-gamma and TNF-alpha had a sensitizing effect. We conclude that FLIP may negatively regulate Apo2L/TRAIL-induced apoptosis in thyroid carcinomas. Microenvironmental paracrine survival factors, such as IGF-1, up-regulate caspase inhibitors, including FLIP, and protect from Apo2L/TRAIL in a PI-3K/Akt-dependent manner. T helper-1 cytokines and compounds that selectively abrogate the IGF-1 signaling pathway may be helpful adjunct agents in Apo2L/TRAIL-based anti-cancer therapeutic regimens.     

Adenovirus-mediated TRAIL expression and downregulation of Bcl-2 expression suppresses non-small cell lung cancer growth in vitro and in vivo

Primary or acquired resistance to current treatment methods remains a major factor in clinical oncology and may be caused by failures in apoptosis programs. TNF-related apoptosis-inducing ligand (TRAIL) can induce apoptosis in several human cancer cell types. However, not all cancer cells are susceptible to TRAIL and mechanisms of resistance and new strategies to enhance sensitivity are an area of intense investigation. Therefore, we investigated whether TRAIL resistance is due to Bcl-2 levels. In this study, we generated an adenoviral vector, Ad5.TRAIL/siBcl2, that permitted co-expression of shRNA against Bcl-2 and the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) therapeutic gene from a cytomegalovirus promoter. Infection with Ad5.TRAIL/siBcl2 resulted in significant cytotoxicity in non-small cell lung cancer (NSCLC) cells in vitro. In contrast, it had no effect on a normal lung cell line, WI-38. Impressively, treatment of the established NSCLC tumor model with Ad5.TRAIL/siBcl2 resulted in significant tumor regression, compared with other adenoviruses. This potent antitumor activity induced by Ad5.TRAIL/siBcl2 was due to strong inhibition of Bcl-2 and high expression of TRAIL. Thus, this study may provide a framework for future clinical applications of Ad5.TRAIL/siBcl2 in lung tumor gene therapy.     

Galectin Labeling of Cells from Paraffinized Tissues May Serve as a Diagnostic Tool in the Detection and Classification of Thyroid Carcinomas

This study seeks to determine whether the relative levels of attachment to galectins 1 and 3 of cells from thyroid tissues embedded in paraffin blocks can differentiate thyroid tumors from normal tissues. A total of 48 thyroid paraffin sample blocks from 4 groups of patients were analyzed: 12 samples served as controls, 12 samples were from patients with thyroid adenoma, 12 samples were from patients with thyroid follicular carcinoma, and 12 samples were from patients with thyroid papillary carcinoma. The relative attachment of cells to galectins 1 and 3 antigens was determined using the InnoCyte? ECM Cell Adhesion kit at different cell sample concentrations. All of the samples from thyroid tissue preparations showed attachment to galectins 1 and 3. The samples from tissues with a diagnosis of adenoma, follicular and papillary carcinoma showed an increased adherence to galectins 1 and 3 relative to the controls. Significant differences were found between the means of the adherent cells from the adenomas compared with the follicular and papillary carcinoma samples. When the outcomes from the galectins 1 and 3 cell surface binding were compared, no statistical differences were found. The cells from adenoma and carcinoma samples show more adhesion to galectins 1 and 3 than cells from the control samples. The samples prepared from follicular and papillary carcinomas show more cells adherent to galectins 1 and 3 than those from the adenomas.     

TNF相关的凋亡诱导配体与肿瘤

TNF相关的凋亡诱导配体（TNF-related apoptosis-inducing ligand, TRAIL），属于TNF 超家族成员，又称为Apo-2L，TRAIL能诱导多种肿瘤细胞的凋亡，而正常的细胞却对其不敏感，TRAIL主要通过与其受体结合激活caspase-8，启动非线粒体和线粒体依赖途径导致细胞凋亡。部分肿瘤细胞对TRAIL的敏感性较差，其原因与TRAIL的受体、信号转导途径激酶以及相关蛋白存在密切联系。     

Adenoviral-mediated transfer of p53 gene enhances TRAIL-induced apoptosis in human hepatocellular carcinoma cells

p53 is a tumor suppressor protein with numerous biological functions including transformation, regulation of cell growth, differentiation and apoptosis. The TNF-related apoptosis-inducing ligand (TRAIL) can induce apoptosis in various transformed cell lines. We investigated the effects of combining wild-type p53 gene transduction by adenoviral infection (Ad-p53) with addition of TRAIL on cell death, expression levels of TRAIL receptors (TRAIL-R1, TRAIL-R2), FLICE inhibitory protein (FLIP) and X-linked inhibitor of apoptosis protein (XIAP) on human hepatocellular carcinoma (HCC) cell lines. HCC cell death was increased by combination of Ad-p53 infection and addition of TRAIL compared to either alone. Western blotting demonstrated decreased TRAIL-R1 and TRAIL-R2 levels after infection with Ad-p53. FLIP levels decreased in Huh7 cells and Hep3B cells, and XIAP levels decreased in all three HCC cell lines after infection with Ad-p53. Thus, death of HCC cells due to combined p53 gene transduction and exogenous TRAIL may be due to down regulation of FLIP or XIAP.     

Frequency and Distribution of DNA fragmentation in Hashimoto’s thyroiditis and development of papillary thyroid carcinoma

Downregulation of apoptosis and high expression of bcl-2 play an important role in the development of follicular lymphoma. However, little is known about apoptosis in thyroid disease, particularly with respect to the development of papillary carcinoma from Hashimoto’s thyroiditis. To study the early stages of cell death in various types of thyroid disease, surgical specimens from 31 patients including Hashimoto’s thyroiditis (HT,n=7), papillary carcinoma (PC,n=12), Hashimoto’s thyroiditis with papillary carcinoma (HTPC,n=5), and Graves’ disease (GD,n=7) were examined by anin situ nucleotidyl transferase assay (ISNTA), which detects DNA fragmentation. Control normal thyroid tissue (NT,n=7) was obtained from surgically resected papillary thyroid carcinomas sampled away from the primary tumor. An immunohistochemical (IHC) method was used to detect bcl-2 expression. Positive ISNTA nuclei in thyroid follicular cells or tumor cells per section were counted in all parenchymal areas, excluding areas of lymphocyte aggregates. The intensity of bcl-2 staining was graded on a scale of 1+ to 3+. The number of ISNTA-positive thyroid follicular cells was a significantly higher in HT compared to GD. In addition, there was significantly lower number of ISNTA positive non-neoplastic thyroid follicular cells in HTPC compared to HT alone. Strong expression of bcl-2 was found in all cases of GD and NT, but much less bcl-2 staining was seen in HT. There was moderate expression of bcl-2 in HTPC and PC. These findings suggest that (1) DNA fragmentation of the thyroid follicular cells plays an important role in the thyroid injury in HT but not in GD, (2) expression of bcl-2 may overcome the apoptosis in GD but not in HT, and (3) downregulation of DNA fragmentation of the follicular cells in Hashimoto’s thyroiditis associated with papillary carcinoma may suggest an important mechanism for tumor pathogenesis.