口腔鳞癌及癌前病变组织中p27、p53蛋白的表达

目的　探讨 p2 7、p5 3蛋白表达在口腔鳞癌发生发展中的意义。 方法　应用免疫组化S P法分别检测 9例口腔正常黏膜 ,11例单纯性增生、2 6例癌前病变及 5 4例鳞癌组织中p2 7、p5 3蛋白的表达。 结果　p2 7蛋白在口腔正常黏膜和单纯性增生组织中呈高表达 ,在癌前病变和鳞癌组织中高 (低 )表达率分别为 6 1 5 % (38 5 % )、2 5 9% (6 1 1% ) ,在鳞癌中阴性表达率为 13% ;p2 7蛋白的表达与鳞癌的组织分化程度、临床分期相关 (P <0 0 5 )。p5 3蛋白在正常黏膜、单纯性增生及轻、中度不典型增生中未见表达 ,在重度不典型增生和鳞癌中可见 2 8 6 %和 4 8 1%的阳性表达 ,二者差异有高度显著性 (P <0 0 1) ;在鳞癌中 p5 3蛋白表达与组织分化程度相关 (P <0 0 5 ) ;p2 7和p5 3表达在鳞癌中呈负相关 (P <0 0 1)。 结论　p2 7蛋白表达的减少在口腔鳞癌的发生发展中起着重要作用 ,并与其预后因素密切相关。p5 3蛋白的表达在癌前病变向鳞癌转变过程中起重要作用。综合分析 p2 7、p5 3表达有助于口腔鳞癌的早期诊断和患者预后的估计。     

细胞周期素依赖性激酶2及p27^kip1与血管瘤

目的：进一步探讨细胞周期素依赖性激酶2（CDK2）及p27^kip1在血管瘤发生、发展及退化过程中的作用机制。方法：采用免疫组织化学SP法检测49例皮肤毛细血管瘤增生期、退化期及正常皮肤组织中CDK2和p27^kip1的表达水平；采用HPIAS-1000高清晰度彩色病理图文报告管理系统，对CDK2和p27^kip1表达的平均光密度和阳性面积率进行图像分析。结果：增生期组CDK2的表达明显高于退化期组和正常皮肤组织组；增生期血管瘤内皮细胞p27^kip1的表达显著低于退化期血管瘤内皮细胞。结论：p27^kip1能抑制血管瘤内皮细胞的增殖，在血管瘤的退化过程中起了重要作用；而CDK2能促进血管瘤内皮细胞增殖，在血管瘤的增生过程中起了重要作用。     

细胞周期素依赖性激酶2及p27Kipl与血管瘤

目的:进一步探讨细胞周期素依赖性激酶2(CDK2)及p27Kipl.在血管瘤发生、发展及退化过程中的作用机制.方法:采用免疫组织化学SP法检测49例皮肤毛细血管瘤增生期、退化期及正常皮肤组织中CDK2和p27Kipl的表达水平;采用HPIAS-1000高清晰度彩色病理图文报告管理系统,对CDK2和p27Kipl表达的平均光密度和阳性面积率进行图像分析.结果:增生期组CDK2的表达明显高于退化期组和正常皮肤组织组;增生期血管瘤内皮细胞p27Kipl的表达显著低于退化期血管瘤内皮细胞.结论:p27Kipl能抑制血管瘤内皮细胞的增殖,在血管瘤的退化过程中起了重要作用;而CDK2能促进血管瘤内皮细胞增殖,存血管瘤的增生过程巾起了重要作用.     

肺鳞癌及癌前病变PCNA、p16、p21、p53表达及细胞凋亡观察

目的 :观察肺癌癌变过程中细胞增殖与凋亡活性的改变 ,分析它们在肺癌发生发展过程中的意义并探讨其调控基因的作用。方法 :收集 86例支气管黏膜活检标本 ,采用免疫酶标法对各组病例进行PCNA、p5 3、p2 1、p16蛋白表达检测 ,用 3′ OH末端DNA原位标记技术进行凋亡细胞检测。结果 :在正常与鳞化组 ,PCNA只在极个别细胞中表达 ,p2 1、p5 3未见表达 ,p16阳性表达率分别为 10 0 %、93 3% ,凋亡细胞阳性率为 (2 9± 5 6 ) %、(2 4± 4 7) %。轻、中度非典型增生组 ,PCNA也只在少数细胞中表达 ,p2 1、p5 3没有表达或极少数细胞表达 ,p16表达率分别为 91 7%、83 3% ,凋亡细胞阳性率分别为 (2 2 0± 4 3) %和 (18 5± 3 7) %。在重度非典型增生与高分化鳞癌组织中PCNA阳性率显著增加 ,而凋亡细胞阳性率却明显减少 ,p2 1、p5 3表达增加 ,p16显著下降。结论 :支气管黏膜癌变是一个由量变到质变的过程 ,而中到重度非典型增生是这一过程中的关键性阶段 ,不仅增殖活性增加 ,而且凋亡活性下降 ,它们的调控基因p16、p2 1、p5 3也发生了显著的变化 ,可能起着重要的调节作用。     

Ku蛋白在皮肤血管瘤组织中表达及其临床意义

目的:探讨Ku蛋白在皮肤血管瘤组织中的表达及其临床意义。方法:收集深圳市龙华医院病理科2009~2013年皮肤血管瘤存档蜡块20例,其中男性15例,女性5例。采用免疫组织化学S-P法检测20例皮肤血管瘤及周围正常组织5例中Ku蛋白表达水平,采用HPIAS-1000图文报告管理系统对Ku蛋白的表达进行图像分析,并用SPSS 13.0软件对各组免疫组织化学反应阳性颗粒的平均光密度、阳性面积率等做单因素方差分析和SNK(q)检验。结果:Ku蛋白在皮肤血管瘤增生期组织中呈高表达,而在退化期和正常皮肤组织中呈低表达。皮肤血管瘤增生期组织中Ku蛋白的表达明显高于退化期和正常皮肤组织。结论:Ku蛋白在皮肤血管瘤的发生和发展中起重要作用。     

胃癌及其癌前病变组织中p53、p73蛋白的表达

胃癌的发生、发展涉及多种原癌基因的激活和抑癌基因的失活。我们应用免疫组化SP法检测正常胃黏膜、各级胃黏膜不典型增生、早期及进展期胃癌组织中p53、p73蛋白的表达,以探讨胃癌发生、发展的规律,寻找胃癌早期诊断和判断预后的标志物,为胃癌的早期诊治及预后评估提供可靠依据     

非小细胞肺癌p63基因的表达及其与3号染色体位点变化的关系

目的　探讨非小细胞肺癌 p6 3基因的蛋白表达水平及其与定位在 3号染色体 2 7～ 2 9区域改变的关系。 方法　应用比较基因组杂交 (CGH)技术对 70例原发性肺鳞状细胞癌和肺腺癌标本进行染色体不平衡性分析。采用组织芯片技术构建12 2例原发性非小细胞肺癌石蜡包埋标本组织芯片 ,并采用免疫组织化学方法检测p6 3蛋白表达情况。比较 p6 3蛋白表达及其与 3号染色体末端改变的关系。结果　CGH分析结果发现 ,30例鳞状细胞癌 2 4例出现 3q2 7～ 2 9区域DNA拷贝数目的增加 ,4 0例腺癌仅 8例发现 3q2 7～ 2 9区域DNA获得。p6 3免疫组化染色结果显示 :5 0例 (84 75 % )鳞状细胞癌免疫组化染色为阳性 ;3例大细胞肺癌中 2例 (6 6 6 6 % )为阳性反应 ;腺癌中仅有 1例 (1 6 7% )为阳性。p6 3蛋白的阳性表达率与患者的年龄、性别、肿瘤的分级、肿瘤的转移以及生存率无关 (P >0 0 5 )。p6 3免疫组化阳性率与 3q2 7～ 2 9区域的改变比较结果显示 :p6 3免疫组化阳性反应与 3号染色体长臂 2 7～ 2 9区域的DNA扩增呈正相关 (P <0 0 1)。结论　p6 3基因的扩增与肺鳞状细胞癌发生和发展有密切的关系     

乳腺癌p53和p27蛋白的表达及意义

目的　探讨p5 3蛋白和p2 7蛋白在乳腺癌中的表达及意义。方法　应用免疫组化S -P法检测 6 7例乳腺癌中p5 3蛋白和p2 7蛋白的表达情况。结果　 6 7例乳腺癌中p5 3蛋白和p2 7蛋白的阳性表达率分别为 5 8 3%和 5 9 7% ,p5 3蛋白的表达与乳腺腋窝淋巴结转移有相关性 (P <0 0 1) ,p2 7蛋白的表达也与乳腺腋窝淋巴结转移有关 (P <0 0 5 ) ,乳腺癌p2 7和p5 3蛋白表达之间无明显相关。结论　p5 3蛋白的过表达和p2 7蛋白的失表达均与乳腺癌发生发展相关 ,二者可作为判断乳腺癌预后的有用指标     

野生型p53对肺癌细胞生长的抑制作用

目的　探讨外源性野生型p5 3对肺癌细胞生长的抑制作用 .方法　通过向肺癌细胞中分别转染携带有荧光蛋白基因的野生型P5 3基因和突变型P5 3基因 ,而后用荧光显微镜观察荧光蛋白表达 ;中性红和MTT染料摄入法测定细胞生长活性 .结果　p5 3蛋白与绿色荧光蛋白之融合蛋白表达定位在胞浆和胞膜 ;经中性红和MTT两种方法测定 ,外源性野生型p5 3对肺癌细胞生长具有抑制作用 ;OD值分别为 0 .71± 0 .0 81(OD540 )和 0 .6 5± 0 .0 5 9(OD490 ) ,均低于对照组 (p <0 .0 5 ) .结论　p5 3外显子 5～ 8可在肺癌细胞中获得表达并对细胞生长活性有抑制作用 .     

新疆维吾尔族与汉族结直肠癌组织P53表达的差异

目的　研究新疆维吾尔族 (维族 )和汉族、浙江汉族结直肠癌组织中 p5 3、Ki 6 7、COX 2蛋白的表达 ,探讨其表达上的差异。方法　应用免疫组化S P法对 14 4例结直肠癌组织 (新疆维族 38例和汉族 5 2例、浙江汉族 5 4例 )中p5 3、Ki 6 7、COX 2蛋白表达进行检测。结果　 3组结直肠癌患者性别构成差异无统计学意义。维族结直肠癌发病年龄早于两组汉族 (浙江汉族、新疆汉族和维族病人的年龄分布分别为 5 7 7± 12 6、5 7 0± 11 5、5 1 4± 11 2岁 ,经方差分析 ,F =3 5 2 0 ,P <0 0 5 )。组织学类型 :浙江汉族病人管状腺癌占 82 7% ,新疆汉族和维族病人乳头状腺癌所占的比例高于浙江汉族 ,3组间差异有显著性(P <0 0 5 )。结直肠癌间分化程度 :新疆汉族高于其他两组 (P <0 0 1)。调整年龄、性别因素后经多因素Logistic回归模型分析 ,民族与结直肠癌p5 3蛋白的阳性表达有关。维族 p5 3阳性率高于汉族 ,比值比为 2 4 92 (95 %CI 1 0 4 6～ 5 939)。新疆汉族和维族结直肠癌与Ki 6 7和COX 2蛋白表达无相关性。结论　维族结直肠癌 p5 3蛋白表达高于汉族病人 ,其意义值得进一步探讨。     

Patterns of p53 and Ki-67 protein expression in epithelial dysplasia from the floor of the mouth

Oral squamous cell carcinoma develops through a series of precancerous stages manifested at the microscopic level as epithelial dysplasia. Mutation of the p53 tumour suppressor gene is thought to be an important component of oral carcinogenesis. p53 regulates cell proliferation and DNA repair by inhibiting the cell cycle at G1/S; loss of p53 function may therefore lead to aberrant cell kinetics. To date, no studies have examined the relationship between p53 protein and alterations in cell kinetics in oral epithelial dysplasia from a single anatomical site. Serial sections were studied from 40 routinely processed biopsy specimens of epithelial dysplasia from the floor of the mouth. The expression of p53 protein was determined by immunohistochemistry and cell proliferation was studied by immunostaining for the cell cycle-dependent protein Ki-67. The number of positive cells per millimetre of basement membrane was determined using computer image analysis and compared with site-matched normal controls. The mean p53 labelling index (LI) in normal mucosa was low, 3·48±0·92 [mean±95 per cent confidence interval (CI)], and increased sharply in the transition from mild (42·49±21·71) to moderate (104·86±51·39) epithelial dysplasia. The mean p53 LI for severe dysplasia was 119·09±56·50. Differences were also observed in the distribution of p53-positive cells between grades of dysplasia, with the development of compact p53-positive foci in severe dysplasia. Mean proliferative indices, as determined by Ki-67 expression, were significantly associated with grade of epithelial dysplasia. Furthermore, there was a significant correlation between p53 LI and Ki-67 score (r²=0·37, P=0·01). It is concluded that altered p53 protein expression is probably an early event in oral carcinogenesis in the floor of the mouth and is associated with dysregulation of cell proliferation at this site. © 1997 John Wiley & Sons, Ltd.     

Increased p53 immunoreactivity in proliferative inflammatory atrophy of prostate is related to focal acute inflammation

Proliferative inflammatory atrophy (PIA) of prostate has been proposed as a precursor lesion of prostate cancer. The aim of the current study was to evaluate the expression of p53 protein in PIA lesions and to investigate the relationship between p53 staining and Ki‐67, glutathione S‐transferase‐π (GSTP1) and cyclooxygenase‐2 (COX‐2) immunohistochemical expression. The results revealed that p53 nuclear immunostaining appeared in PIA lesions in 2.1±3.4% (mean±SD) of the basal and 0.9±2.3% of the luminal epithelial cells. Both these values were significantly higher than those in normal‐appearing acini (p<0.0001). Increased p53 expression in luminal cells was related to focal infiltration of polymorphonuclear leucocytes. A positive correlation between p53 expression and Ki‐67 was found in COX‐2‐positive PIA lesions (r=0.610, p<0.0001). Half of the p53‐positive epithelial cells expressed diffuse GSTP1 immunostaining in the same lesions. The present study demonstrates an increased p53 expression in PIA lesions, and inflammation, especially acute inflammation, may play a role in the induction of p53 over‐expression, particularly as cells in PIA lesions are known to have a reduced defence against DNA damage.     

Increased proliferative activity and p53 expression in normal glandular breast tissue after radiation therapy

Radiation used in breast-conserving therapy (BCT) for early breast cancer, to eradicate residual malignant cells after tumour resection, induces DNA damage and cell death. Little is known about the effect of the commonly used doses of radiation therapy on normal breast tissue. Under physiological conditions, p53 plays a role in maintaining genomic stability and regulating progression through the cell cycle. In normal glandular breast tissue, p53 expression is very low, as is proliferative activity. The purpose of this study was to investigate the levels of p53 expression and proliferative activity in non-malignant glandular epithelium of the breast after BCT. The immunohistochemical expression of p53 and Ki-67 was semiquantitatively correlated in non-malignant glandular epithelium in biopsies before and after BCT in 24 women with breast cancer. In 18 cases, a recurrence was diagnosed and in the remaining cases, the clinical suspicion was not histologically confirmed. In addition, in six cases with contralateral breast cancer, the same immunohistochemical evaluation was performed in tissue from both breasts. The mean interval between the two surgical interventions was 50 months. The percentage of p53 immunoreactive epithelial cells in normal breast tissue was significantly (P<0·01) higher after radiation therapy than before in the ipsilateral side (0·2 per cent±0·3 and 4·6 per cent ±4·5, respectively). Ki-67 immunoreactivity was also significantly increased (P<0·001) after radiation therapy, from 0·6 per cent to an average of 4·8 per cent in epithelial cells. In contrast, in the patients with contralateral breast cancer, the levels of p53 and Ki-67 immunoreactivity in the normal glandular breast tissue were comparable to the ipsilateral side (P=0·7 and P=0·1, respectively). In conclusion, increased expression of p53 and Ki-67 is present in normal glandular breast tissue, even 2–5 years after radiation therapy. © 1998 John Wiley & Sons, Ltd.     

Characterization of specific p63 and p63-N-terminal isoform antibodies and their application for immunohistochemistry

The TP63 gene gives rise to protein isoforms with different properties and functions due to the presence (TAp63) or absence (ΔNp63) of an N-terminal p53-like transactivation domain. Immunohistochemistry for p63 has clinical value for certain tumour types, but investigations have been hampered by a lack of well characterized antibodies and the inability to discriminate between these N-terminal isoforms with opposite functional properties. We have extensively characterized a series of monoclonal antibodies to recombinant human TAp63 and two commercial p63 monoclonals by Western blot, immunostaining and phage display epitope mapping. Twenty-eight of 29 (96.6 %) novel monoclonals that recognized all p63 isoforms showed substantial cross-reactivity with p73, as did the commercial antibody, 4A4. One novel clone, PANp63-6.1, showed slight cross-reaction with p73 by Western blotting but not immunohistochemistry and the SFI-6 monoclonal did not cross-react with p73 or p53. Phage display revealed that the PANp63-6.1 epitope has one amino acid difference between p63 and p73, the 4A4 epitope is identical in both, whereas the SFI-6 epitope is unique to p63, accounting for these findings. We also produced and characterized a TAp63-specific clone that does not recognize p53 or p73, and we prepared polyclonal sera specific for ΔNp63 isoforms. Immunohistochemistry demonstrated that TAp63 is expressed in a variety of epithelial and other cell types during development, often in a converse pattern to ΔNp63, but has a very limited expression in normal adult tissues and is independent of ΔNp63. TAp63 was expressed in 17.6 % of squamous cancers of cervix that expressed p63, unlike normal cervix where TAp63 was not expressed. TAp63 did not associate with proliferative index, but cervical carcinomas with TAp63 expression showed improved survival. These data highlight the need for rigorous antibody characterization and indicate that p63-isoform identification may improve the clinical value of p63 expression analyses.     

Sun-exposure- and aging-dependent p53 protein accumulation results in growth advantage for tumour cells in carcinogenesis of nonmelanocytic skin cancer

 Three hundred and sixteen patients with nonmelanocytic skin cancer, including 46 cases of Bowen’s disease (BOD), 134 cases of squamous cell carcinoma (SCC), and 136 cases of basal cell carcinoma (BCC), were examined immunohistochemically using monoclonal antibody DO-7 to assess p53 protein accumulation related to sun exposure and ageing, and growth and differentiation of skin cancer and its precursors. The rates of p53 immunostaining of BOD, SCC and BCC were 80.4%, 76.1% and 70.6%, respectively. p53-positive cells were present not only in cancer nests, but also in dysplastic and even morphologically normal epidermis adjoining cancers. Sun exposure was statistically correlated with the p53 immunostaining scores in morphologically normal epidermis of the three skin cancers and in cancer nests of SCC and BCC. The positivity and score of p53 protein often differed significantly among the three types of cancer, especially in regions of dysplasia. Interestingly, differentiation of SCC was correlated with individual p53 scores for dysplasia and cancer nests, especially for dysplasia. BOD, as the precursor of SCC, demonstrated the strongest p53 expression. Furthermore, 12.3% cases with p53 negative cancer nests showed p53-positive reaction in dysplasia and in morphologically normal epidermis. It seems that the accumulation of p53 protein plays a part in precancerous lesions and in the genesis of more highly differentiated types of skin cancer and affects mainly the growth of tumour cells rather than their differentiation. For BCC, however, age was significantly related to p53 expression. Our findings suggest that overexpression of p53 in normal skin and cancer nests of SCC and BCC is significantly related to sun exposure, that the expression of p53 in BCC is an age-dependent process, and that the early accumulation of p53 protein may be a useful predictor for the detection of nonmelanocytic skin cancer. Received: 7 July 1998 / Accepted: 5 November 1998     

Detection of loss of heterozygosity of p53 gene in paraffin-embedded breast cancers by non-isotopic PCR–SSCP

A rapid non-isotopic PCR-SSCP (polymerase chain reaction-single-stranded conformation polymorphism) method was developed in this study to detect polymorphism and loss of heterozygosity (LOH) of p53 in formalin-fixed and paraffin-embedded samples of normal breast tissue and of breast cancer. p53 expression was also examined by immunohistochemistry. In 35 paired samples, heterozygosity in exon 4 of p53 was detected in 17 cases (49 per cent) and LOH of the p53 gene in breast cancer tissues was observed in 7 out of 15 informative cases (47 per cent). The correlation of LOH of p53 with positive p53 immunostaining did not reach statistical significance, but all immunostaining-positive tumours among informative cases had LOH of p53. The results support the hypothesis that in most cases the allelic deletion of p53 may uncover the ‘recessive mutation’ in the remaining allele. However, LOH of p53 was more frequent than positive immunostaining and was significantly associated with poor differentiation of breast cancer (P<0·05). The results suggest that the allelic deletion of p53 may also contribute to the development and progression of breast cancer by reducing the amount of normal p53 protein. These results show that non-isotopic PCR-SSCP is a simple, fast, and effective method for detecting polymorphism and LOH of the p53 gene, which is especially useful for retrospective studies.     

Expression of p73 in normal skin and proliferative skin lesions

The p73 gene is a member of the p53 gene family and the structure and functions of p73 protein are similar to those of p53. However, these two proteins have different roles. In the present study, p73 protein was found immunohistochemically to be distributed in the basal cells of the epidermis, columnar basal cells in the hair follicle and peripheral cells without lipid droplets in the sebaceous and meibomian glands; it was expressed strongly in tumor cells in basal cell carcinomas and in the basal cell-like cells in seborrheic keratosis, and weakly or negatively in the squamous cell-like cells in seborrheic keratosis and in the tumor cells in squamous cell carcinomas. No relationship was detected between p73 and p53 protein distribution and between p73 protein expression and the proliferative potential, as shown by the Ki-67 immunopositive cell ratio. The present study shows that p73 protein is likely to play important roles in skin differentiation rather than proliferation or carcinogenesis of the skin.