p53、p21、p27、p16在口腔鳞癌中的表达及与预后的关系

目的:研究细胞周期相关蛋白p53、p21、p27、p16在口腔鳞癌组织中的表达及其与预后的相关性,寻找预后判断的有效生物标志物。方法:用免疫组织化学的方法检测随访资料完整的110例口腔鳞癌患者术后石蜡切片中p53、p21、p27、p16的表达,通过目标蛋白阳性染色细胞比例和阳性细胞中蛋白的染色强度判定各蛋白的表达水平,应用SAS 9.0软件中的Kaplan-Meier分析p53、p21、p27、p16蛋白表达水平与口腔鳞癌预后的相关性,筛选和确定与口腔鳞癌预后相关的细胞周期蛋白。结果:110例口腔鳞癌标本蛋白检测和统计分析结果表明,p53、p27、p16蛋白的表达水平与口腔鳞癌术后生存时间并无显著的相关性;p21表达与口腔鳞癌的术后生存率呈显著的相关性(P<0.05)。结论:口腔鳞癌患者癌组织标本的p21表达水平与预后呈正相关,p21有望成为口腔鳞癌预后判断的候选生物标志物。     

大鼠三叉神经脊束尾侧亚核内Cdk5/p39的表达与活性

目的:探讨周期依赖性蛋白激酶5(Cdk5)2种激活剂p35和p39在出生后各发育阶段大鼠三叉神经脊束尾侧亚核内的表达变化.方法:采用Western印迹分析、免疫沉降和Cdk5活性分析,检测出生后各发育阶段大鼠三叉神经脊束尾侧亚核内Cdk5的活性变化和Cdk5、p39的表达水平.采用SPSS11.0软件包中的两两比较和扩展t检验进行统计学分析.结果:在新生大鼠三叉神经脊束尾侧亚核组织中,p39表达较低;出生后2～3周时,p39的表达最高;到成体时,其表达降低到与新生大鼠三叉神经脊束尾侧亚核组织中的相同水平.在出生后各发育阶段,大鼠三叉神经脊束尾侧亚核内Cdk5的表达一致,而Cdk5的活性逐渐减弱,新生大鼠三叉神经脊束尾侧亚核内的Cdk5活性(115.5Kepm)比成体中的Cdk5活性(19.0 Kcpm)约高6倍,各组之间均有显著差异(P<0.01).结论:在出生后各发育阶段,大鼠三叉神经脊束尾侧亚核内p39的表达与p35表达结果在时间上有差异,Cdk5/p39和Cdk5/p35在出生后各发育阶段大鼠三叉神经脊束尾侧亚核内可能扮演不同的角色.     

口腔扁平苔藓组织中p73和p14^ARF基因表达与癌变的关系

目的探讨p73、p14^ARF在口腔扁平苔藓组织中的表达与癌变的关系。方法采用免疫组化方法,分别检测口腔扁平苔藓、口腔鳞状细胞癌和正常口腔粘膜各30例组织中p73、p14^ARF的表达情况;采用美国Media Cybernetics公司的Image Plus5.0专业图像分析软件进行IOD值（累计光密度值）的测量。结果p73表达的IOD值在正常口腔粘膜、口腔扁平苔藓及口腔鳞状细胞癌中不断增高,而p14ARF表达的IOD值在正常口腔粘膜、口腔扁平苔藓及口腔鳞状细胞癌中却逐渐下降。两组间有统计学意义（P〈0.05）。p73、p14^ARF在口腔扁平苔藓、正常口腔粘膜、口腔鳞状细胞癌组织中的表达呈负性相关性（P〈0.05）。结论p73、p14^ARF这两个基因在OLP的癌变过程中具有一定的作用。     

成釉细胞瘤中细胞周期素D1及其抑制因子和hTERT的表达

目的研究端粒酶逆转录酶（ hTERT）、细胞周期素D（1 cyclin D1）、周期素依赖激酶抑制因子p16INK4、p21WAF1 mRNA和p27KIP1蛋白在人成釉细胞瘤（ ABs）中的表达。方法用原位杂交和免疫组化SP法分别检测ABs中hTERT、cyclin D1、p16INK4、p21WAF1 mRNA和p27KIP1蛋白的表达。结果hTERT mRNA在ABs中有较强表达,51例为阳性表达,cyclin D1 mRNA在ABs中23例为阳性表达,而p16INK4、p21WAF1 mRNA和p27KIP1蛋白在ABs中分别有17、12、9例为阳性表达。伴随ABs的复发和恶变,hTERT、cyclin D1 mRNA阳性率逐渐上升,p16INK4、p21WAF1 mRNA和p27KIP1蛋白阳性表达丢失。经Kendall相关性分析得出,hTERT mRNA与p16INK4、p21WAF1 mRNA、p27KIP1蛋白之间在ABs中rk分别为- 0.587、- 0.652、- 0.783,均有统计学意义（ P<0.001）。结论hTERT在ABs中的活性可能与p16INK4、p21WAF1 mRNA、p27KIP1蛋白丢失等多种因子共同调控有关。     

舌鳞状细胞癌中p73、p63和p53基因的表达

目的：研究p73、p63和p53基因蛋白在舌鳞状细胞癌中的表达及其意义。方法：用免疫组织化学S－P法检测64例舌鳞状细胞癌组织标本p73、p63和p53基因蛋白表达。结果：舌鳞癌中p73、p63和p53基因蛋白阳性率分别为61％、92％和50％。p73蛋白表达与T分类(r=0.344,P=0.057)、区域淋巴结转移（r=0.334,p=0.007)、肿瘤侵袭方式（r=0.340,P=0.060)和不良预后（r=-0.473,P=0.000)相关。p63与所有临床病理特征和预后无关。p53与区域淋巴结转移（r=0.263,P=0.036)和不良预后（r=-0.342,P=0.013)相关。结论：p73和p53表达有助于判断舌鳞癌的生物学行为和预后。p63可能是鳞状上皮源性肿瘤的标记物。     

ToPoⅡα在口腔鳞癌PTP方案化疗中的指导意义

目的:探讨口腔鳞癌中ToPoⅡα的表达水平与临床PTP方案(顺铂+替尼泊甙+平阳霉素/博莱霉素)化疗效果之间的相关性,初步评价其表达水平是否可以指导临床PTP的应用。方法:采用回顾性研究的方法检测了46例原发口腔鳞癌标本化疗前ToPoⅡα与p53表达情况,评价其与临床化疗效果的相关性;对比分析其中18例标本化疗前后ToPoⅡα的表达变化。同时,采用Western印迹法分析8种口腔鳞癌细胞系内ToPoⅡα表达水平与化疗药物替尼泊甙的半数抑制浓度之间的关系。结果:46例口腔鳞癌患者中ToPoⅡα均有不同程度的表达,其表达水平与化疗疗效呈显著正相关;体外实验也证实8种细胞系中ToPoⅡα的表达水平与口腔鳞癌细胞系对替尼泊甙的敏感性显著相关。与化疗前相比,化疗后口腔鳞癌标本中ToPoⅡα的表达明显下降,提示有继发性耐药产生的可能性。p53的表达水平与化疗疗效无显著相关性,也与ToPoⅡα的表达水平没有相关性。结论:实验结果表明,ToPoⅡα的表达水平能够指导临床应用PTP化疗方案治疗口腔鳞癌,而p53的表达水平对临床化疗效果提示作用不明显。     

抑癌基因p53基因克隆和表达

目的：本研究对抑癌基因ｐ５３进行基因克隆和表达以获得ｐ５３基因产物。方法：采用基因克隆和重组技术将Ｐ５３ｃ ＤＮＡ编码区重组到融合蛋白表达载体 ｐＧＥＸ－２Ｔ中,用免疫斑点杂交技术检测 ｐ５３融合蛋白在大肠杆菌中的表达。结果：我们获得了ｐ３基因融合蛋白表达质粒ｐＧＥＸ－ｐ５３并在大肠杆菌中获得了表达。结论：ｐＧＥＸ－２Ｔ是一个有效的融合基因表达载体,适合于ｐ５３基因的表达。用ｐＧＥＸ－ｐ５３可以很方便地获得ｐ５３蛋白,对研究Ｐ５３基因在口腔癌发生中的作用具有应用价值。     

放射线对口腔鳞状细胞癌细胞中核因子-κB p65表达的影响

目的:探讨不同剂量放射线对口腔鳞状细胞癌(OSCC)细胞中核因子-κB(nuclear factor-κB,NF-κB)p65信号转导蛋白表达的影响.方法:采用人舌鳞状细胞癌Tca8113细胞,分为6个剂量组,分别为0y、2、4、6、8、10 Gy,在照射后的不同时间点(1,3,6,10,24,48 h)应用免疫细胞化学检测NF-κB p65的表达情况,应用Leica Qwin Plus图像分析系统对图像进行分析,测其灰度值,并应用SPSS 10.0软件对数据进行X2检验.结果:免疫细胞化学方法显示不同剂量组的平均灰度值与0 Gy组相比均为P ＜0.05,差异均具有统计学意义,而不同时间点组的平均灰度值相互比较,3 h组的平均灰度值与其他时间点组相比均为P ＜0.05,差异有显著性.结论:放射线可以激活NF-κB p65信号转导通路,此信号转导通路在肿瘤的放射治疗中可能具有重要意义.     

新辅助化疗前后PCNA和p53在口腔鳞癌中的表达及其对化疗敏感性的预测研究

目的:探讨新辅助化疗前后增殖细胞核抗原(PCNA)和抑癌基因p53在口腔鳞癌组织中的表达水平及其与化疗疗效的关系.方法:采用SP免疫组化法,对106例口腔鳞癌患者新辅助化疗前后的PCNA和p53表达水平进行检测,分析其表达水平与化疗疗效的关系.结果:①化疗前口腔鳞癌组织中PCNA高表达率为50.94%,显著高于化疗后23.58%(P<0.05);化疗前PCNA高表达患者化疗有效率为62.96%,显著高于PCNA低表达患者的有效率30.77%(P<0.05);②化疗前p53在口腔鳞癌中的阳性表达率为44.34%,显著高于化疗后21.60%(P<0.05);化疗前口腔鳞癌p53表达阴性患者的有效率为55.93%,显著高于p53表达阳性患者的有效率36.17%(P<0.05).结论:新辅助化疗可明显降低PCNA和p53在口腔鳞癌组织中的表达水平,PCNA和p53的表达水平与新辅助化疗疗效具有显著的相关性,认为PCNA和p53的表达水平有可能成为口腔鳞癌化疗疗效的预测指标.     

p63在口腔扁平苔藓中的表达

目的探讨p53家族新成员p63在口腔扁平苔藓（OLP）中的表达，其与OLP发生、发展的关系以及是否可作为一种标志物预测OLP的转归。方法采用免疫组织化学方法检测30例012中p63蛋白的表达，同时采用RT—PCR方法分析p63编码的两大类转录产物TAp63和ANp63的mRNA水平，并且与口腔鳞状细胞癌（OSCC）及正常口腔黏膜（NOM）进行比较。结果p63蛋白在NOM、OLP和OSCC组中均有表达，与NOM比较，其累积光密度（IOD）值在OLP中下调，在OSCC中明显上调。在糜烂萎缩型OLP中，p63蛋白表达高于非糜烂型（P〈0．001）。RT-PCR检测结果显示，TAp63的mRNA水平在OLP中表现为中等。略低于NOM．明显高于OSCC；ANp63的mRNA水平在NOM和OLP中极低．其转录扩增带大多数检测不到．但在OSCC组都能检测到．而且呈高水平表达。TAp63的mRNA水平在糜烂萎缩型OLP中低于非糜烂型（P=0．018），ANp63则高于非糜烂型（P=0．049）。结论p63与OLP的发病机制有关，其中TAp63和ANp63分别起不同的作用。p63表达的高低可能可以作为预测OLP是否有高风险癌变的检测指标。     

Relationship of p53 overexpression to other cell cycle regulatory proteins in oral squamous cell carcinoma

Aberrations of the p53 gene and the overexpression of its protein are described in a variety of neoplasms, including oral and other head and neck cancers. Here we report the association of p53 (over)expression with a downstream cell cycle inhibitor p21/waf 1 in oral squamous cell carcinoma (SCC). The loss of expression of p16 and p27, two other cyclin-dependent kinase (cdk) inhibitors, was also examined. In this panel of tumours, 10/24 carcinomas were p53-immunopositive. Heterogeneous expression of p21 and p27 was seen in 10/24 SCC and 9/16 SCC, respectively, and this was not correlated to p53 status. The expression of p21 and p27 in these SCCs suggests the existence of mechanisms by which some growing tumour cells may tolerate these cell cycle inhibitors; eight SCCs lacked expression of both inhibitors but only two of these cancers overexpressed p53, suggesting that accumulation of p21/p27 can be independent of the functional status of the p53 gene. Data do not support a clear example of a phenotype that shows an overexpression of p53 with downregulation of p21 or p27 leading to cell cycle alterations. Furthermore, only three SCCs were p16-negative and p53-positive. This suggests that these two tumour suppressors may act in separate pathways.     

Combined cytoplasmic and membranous EGFR and p53 overexpression is a poor prognostic marker in early stage oral squamous cell carcinoma

J Oral Pathol Med (2012) 41 : 559–567 Objective: Our aim was to evaluate the expression of several molecules that regulate growth, the cell cycle and signalling pathways including EGFR, p53, p16 and p27 in oral squamous cell carcinomas (OSCC). We examined their utility as prognostic markers by relating to clinicopathological characteristics and the clinical outcome. Patients and methods: Using tissue microarray technology, we analysed 67 primary OSCC and examined immunohistochemical expression of EGFR, p53, p16 and p27. Multivariate analysis was conducted to examine their role in survival. Results: Many of the markers were highly expressed in these cancers. Membranous EGFR expression in 95.2%, both membrane and cytoplasm expression in 35%, p53 expression in 61.6%, p27 expression in 89.5% and p16 expression in 27.9% of cases. In the multivariate analysis, independent prognostic influence of a lower overall survival was determined only for advanced tumour stage (P < 0.001), p53 overexpression (P = 0.004), EGFR cytoplasm and membrane co‐expression location (P = 0.002) and p16 reduced expression (P = 0.002). When considering a subgroup of early stage tumours, p53 overexpression (P = 0.028) and combined membranous and cytoplasm EGFR co‐expression (P = 0.039) were indicators of a lower overall survival. For disease‐free survival, in addition to these three factors, the histological grade (P = 0.011) showed independent prognostic values. Conclusion: The independent value of EGFR subcellular location (cytoplasm and membrane) and p53 overexpression in overall survival even in early stages of OSCC suggests that these markers may serve as reliable biological markers to identify high‐risk subgroups and to guide therapy.     

Detection of antibodies against p63 and p73 isoforms in sera from patients diagnosed with oral lichen planus

BACKGROUND: Oral lichen planus (OLP) is a chronic inflammatory disease of oral mucosa. Despite numerous publications and intense research, the etiology of OLP is still unknown, however, autoimmunity as a possible causative factor has been discussed. METHODS: In the present study sera from 20 patients clinically and histologically diagnosed with OLP were analyzed for antibodies directed toward p53, p63, and p73 using Western blot. RESULTS: Sera from two patients reacted with all six p63 isoforms, and one also with p73. The strongest reaction was noted against the TAp63beta protein, which is the most potent transactivator of all p63 proteins and is implicated in the differentiation of stratified epithelia. CONCLUSIONS: This is the first demonstration of antibodies directed against all p63 and some p73 isoforms in sera from patients diagnosed with OLP.     

p16和p53蛋白在口腔白斑和鳞状细胞癌中表达的比较研究

目的比较p16和p53在口腔白斑和口腔鳞状细胞癌中异常表达的情况。方法对17例健康者的口腔粘膜、60例白斑患者和40例口腔鳞状细胞癌患者的病损组织进行了p16和p53蛋白的免疫组化染色。结果正常口腔粘膜、白斑单纯增生、白斑异常增生和口腔鳞状细胞癌的p16阳性率分别为100％、90％、60％和35％，p53蛋白的阳性率分别为12％、27％、50％和73％，p16阳性率和细胞染色强度指数（stainning intensity index，SII）分别与口腔粘膜组织恶性度的增高显著负相关；p53阳性率和S11分别与口腔粘膜组织恶性度的增高显著正相关。p16和p53在不同组织中的阳性率显著负相关；p16和p53在白斑异常增生的协同失活率达23％，在口腔鳞状细胞癌中达到42．5％。p16阳性率、p16SII、p53SII及两种基因均异常的比率在口腔鳞状细胞癌无淋巴结转移和口腔鳞状细胞癌有淋巴结转移患者之间均有显著差异。结论p16可作为早期监视口腔白斑恶变、监测口腔鳞状细胞癌发展进程和判断口腔鳞状细胞癌预后的分子生物学指标。p16和p53失活在白斑癌变和口腔鳞状细胞癌的发展过程中可能有叠加效应。     

Prognostic value of cyclin D1, p27, and p63 in oral leukoplakia

BACKGROUND: Studies on the expression of genes regulating cell proliferation and apoptosis are essential to help better understand the severity and possible malignant transformation of oral leukoplakia. METHODS: The characteristics of cyclin D1, p27, and p63 were investigated in this microscopic study, complementing our previous results with Ki67, p53, and the apoptosis index. Clinical and histologic as well as immunohistochemical studies were carried out on oral leukoplakia of 18 patients. Homogenous, or non-homogenous (nodular or speckled) and erythroleukoplakia were determined clinically. Pathologic classification was performed according to the degree of dysplasia. Immunoperoxidase reaction for cyclin D1, p27, and p63 was carried out on the biopsy specimens and the positivity of the reactions was calculated for 1000 epithelial cells. RESULTS: The expression of cyclin D1 increased in parallel with the severity of leukoplakia. The p27 index was 14-16% in homogenous and nodular leukoplakias but it was substantially lower to 1-2% in erythroleukoplakia. The p63 index was 10% in homogenous, 5% in nodular or speckled, but nearly 20% in erythroleukoplakia, on the average. CONCLUSION: These results suggest that the characteristic expression of cyclin D1, p27, and p63 in various forms of leukoplakia may be of prognostic value.     

p53 gene status and expression of p53, MDM2, and p14ARF proteins in ameloblastomas

BACKGROUND: To clarify the roles of the p53-MDM2-p14(ARF) cell cycle regulation system in oncogenesis and cytodifferentiation of odontogenic tumors, p53 gene status and expression of p53, MDM2, and p14(ARF) proteins was analyzed in ameloblastomas as well as tooth germs. METHODS: Paraffin sections of 16 tooth germs and 46 benign and 5 malignant ameloblastomas were examined immunohistochemically for the expression of p53, MDM2, and p14(ARF) proteins. Frozen tissue samples of 10 benign ameloblastomas and 1 malignant (metastasizing) ameloblastoma were analyzed by direct DNA sequencing to detect p53 gene alteration. RESULTS: Immunohistochemical reactivity for p53 was detected in 2 of 13 tooth germs, 13 of 29 ameloblastomas, and 5 of 5 malignant ameloblastomas, and the expression ratio of p53 in tooth germs was significantly lower than those in benign and malignant ameloblastomas. Direct DNA sequencing showed no alteration of p53 gene exons 5-8 in any sample of 10 benign ameloblastomas and 1 metastasizing ameloblastoma. Expression of MDM2 and p14(ARF) was detected in all samples of normal and neoplastic odontogenic epithelium, and the expression ratios in tooth germs tended to be lower than those in benign and malignant ameloblastomas. In ameloblastomas, expression of p53, MDM2, and p14(ARF) was significantly higher in plexiform cases than in follicular cases. Markedly decreased reactivity for p53, MDM2, and p14(ARF) was detected in keratinizing and granular cells in ameloblastoma subtypes. Basal cell ameloblastoma showed slightly higher reactivity for p53, MDM2, and p14(ARF) as compared with other subtypes. CONCLUSION: Elevated expression of p53, MDM2, and p14(ARF) in benign and malignant ameloblastomas suggests that alteration of the p53-MDM2-p14(ARF) cascade is involved in oncogenesis and/of malignant transformation of odontogenic epithelium. p53 gene status implied that p53 mutation might play a minor role in neoplastic changes of odontogenic epithelium. Immunoreactivity for p53, MDM2, and p14(ARF) in ameloblastoma variants suggests that these factors might be associated with tissue structuring and cytodifferentiation of ameloblastomas.     

Vigabatrin-induced modification of Ki-67 expression in gingival epithelium: immunohistochemical study of a short series

OBJECTIVE: To study the expression and role in vigabatrin (VGB)-induced gingival enlargement of Ki-67 antigen and p27KIP1, p21WAF1, and p53, proteins that activate or inhibit cell-cycle progression. MATERIALS AND METHODS: Six patients treated with VGB for partial epileptic seizures refractory to classic anticonvulsant treatment were studied. Gingival biopsies were taken from four of these patients for immunohistochemical studies; 10 control biopsies from individuals with healthy gingiva and 10 from patients with periodontal disease were also evaluated. RESULTS: Four of the six patients presented some degree of gingival enlargement (mild or moderate). Nuclear expression of Ki-67 was elevated (mean of 894 positive cells/mm2 in VGB-induced gingival enlargement vs. 391 cells/mm2 in controls with healthy gingiva and 425 cells/mm2 in controls with periodontal disease) (p < 0.01, analysis of variance: anova), and nuclear expression of cyclin-dependent kinase (cdk) inhibitors p27KIP1 and p21WAF1 was reduced. The patients with gingival enlargement presented inflammatory infiltrate in lamina propria, mainly composed of T lymphocytes (CD3+) and plasma cells (CD38+), which was even more intense than in the biopsies of patients with periodontal disease. CONCLUSION: The overexpression of antigen Ki-67 and slight underexpression of cdk-inhibitors p27KIP1 and p21WAF1 suggest that VGB induced an increase in cell proliferation and contributed, together with concomitant periodontal disease, to the gingival enlargement.