免疫组化检测对喉癌手术切缘的安全性研究

目的：探讨从细胞的蛋白水平确定喉癌手术的安全切缘范围。方法：采用免疫组化ＬＳＡＢ或ＳＡＢＣ方法,对３６例原发性喉癌的三组标本行５种抗体检测。结果：５种抗体在喉癌组表达阳性率分别与两边缘组阳性率差异非常显著或差异显著,而近癌旁组与远癌旁组阳性率之间差异均无显著差异,５种抗体在高分化组与低分化组阳性率差异显著,ＰＣＮＡ在是否有颈淋巴结转移组的阳性率表达差异显著。     

细胞周期调控因子与喉癌癌变的相关性研究

目的 :探讨喉癌癌变过程中 Cyclin E、p2 1WAF1 /CIP1和 p5 3表达的临床病理学意义。方法 :用免疫组化检测 2 0例喉正常粘膜、40例喉不典型增生病变和 6 0例喉癌组织中 Cyclin E、p2 1WAF 1 /CIP1和 p5 3的表达。结果 :1Cyclin E阳性表达率在正常组、不典型增生组和喉癌组分别为 5 .0 % (1/ 2 0 )、2 0 .0 % (8/ 40 )、45 .0 % (2 7/ 6 0 ) ,喉癌组与正常组比较 ,其差异有极显著性意义 (P<0 .0 1) ;p2 1WAF1 /CIP1阳性表达率分别为 95 .0 % (19/ 2 0 )、75 .0 % (30 /40 )和 6 3.3% (38/ 6 0 ) ,喉癌组与正常组比较 ,其差异亦有显著性意义 (P<0 .0 5 ) ;p5 3阳性表达率分别为 0 (0 /2 0 )、30 .0 % (12 / 40 )和 6 1.7% (37/ 6 0 ) ,喉癌组与正常组比较 ,其差异有极显著性意义 (P <0 .0 1)。 2 p2 1WAF 1 /CIP1在高、中、低分化的喉癌中阳性表达率逐渐降低 ,其差异有显著性意义 (P <0 .0 5 )。 3 Cyclin E和 p5 3阳性表达显著相关 ,p2 1WAF 1 /CIP1与 p5 3阳性表达无相关性。结论 :1Cyclin E和 p5 3异常表达是喉癌发生中早期分子事件。 2p2 1WAF 1 /CIP1 表达与喉癌细胞分化程度有关。 3细胞周期因子相互调控机制在喉癌发展中起重要作用。     

声门上型喉癌手术切缘的分子学研究

目前喉癌主要是手术治疗,其根本目的在于彻底切除肿瘤并在此基础上进行喉功能重建,学者们开展各种术式治疗喉癌,但术后复发率仍有15％（Kaplan等,1983）。手术切缘是影响喉癌术后复发的主要原因,其中分布于切缘的少量癌细胞及细胞表面形态正常而内部基因已发生改变并可发展成为癌的细胞,目前常规病理检查不能完全检测到。肿瘤抑制基因p53突变是人类恶性肿瘤最常见的遗传学改变,在肿瘤中具有较高的突变率,可作为肿瘤标记物。本文应用聚合酶链反应-单链构象多态性分析（PCR—SSCP）法检测声门上型喉癌手术切除标本不同切缘部位的p53基因突变情况,     

喉癌术后局部复发与手术切缘的关系

目的 :探讨喉癌术后局部复发与手术切缘的关系。方法 :对 1991～ 1999年行手术治疗并随访满 3年以上的喉癌患者的临床资料进行回顾性分析 ,分别选取手术切缘、T分期和术后放疗等因素进行统计学分析。结果 :① 16 0例喉癌患者术后局部复发 36例 ,复发率为 2 2 .5 % ;②局部复发与肿瘤的T分期有关 ,差异有统计学意义 ;③切缘≤ 3mm组的局部复发率高于切缘 4～ 5mm组和 >5mm组 ,差异有统计学意义 ;④手术切缘≤3mm的 6 9例中 ,32例行术后放疗的局部复发率低于 37例未行放疗者 ,差异有统计学意义 ;⑤声门型喉癌 5mm的手术切缘与切缘≤ 3mm组相比 ,局部复发率较低 ,差异有统计学意义。而 5mm的手术切缘对声门上型喉癌不够安全 ,与≤ 3mm组相比 ,局部复发率相似 ,差异无统计学意义。结论 :①以 5mm作为声门型喉癌的手术切缘是相对安全的 ;②以 5mm作为声门上型喉癌的手术切缘不够安全 ,如局部条件许可 ,建议以 10mm为安全切缘 ;③对手术切缘不够者行术后放疗可降低局部复发率 ,但不主张因此而放宽对安全切缘的标准。     

喉癌手术切缘的临床判别

目的探讨喉癌临床手术的安全切缘.方法分析1996年6月至2001年6月山东大学齐鲁医院及烟台毓璜顶医院耳鼻咽喉科手术治疗167例喉癌患者的临床资料,术前全面评估肿瘤,依据肿瘤特点分别采用甲状软骨中份旁侧入路,环甲膜入路,气管入路,咽侧入路,会厌谷入路,会厌根入路,根据肿瘤的原发部位、分期与特点切除范围相应不同;记录肿瘤扩展范围、复发、喉外侵犯、淋巴结转移的分布,统计复发率及生存率.结果 3、5年失访分别为3例和4例,按复发和死亡计.3、5年复发率为15/122(12.3%)、23/94(24.5%),生存率为110/122(90.2%)、73/94(77.7%);19例复发中单纯局部复发11例,单纯颈部复发5例,颈部和局部复发3例,颈部复发8例中,Ⅱ、Ⅲ、Ⅳ区、气管旁、锁骨上分别为3、 2、 1、 1、 1例.淋巴结转移为231/1332,声门区13/198,声门上区110/554、跨声门型103/532,声门下区5/48;其中Ⅱ、Ⅲ、Ⅳ、V区分别为121、78、13、9枚,喉前淋巴结6枚,气管旁淋巴结4枚.喉外侵犯甲状软骨板19例,甲状腺11例,气管6例,接力肌4例,胸骨舌骨肌2例,舌根9例.前连合、浸润性生长的深面切缘、会厌前间隙和舌根、披裂周围、软骨切缘以及Ⅱ区、喉前、气管旁淋巴结、甲状腺容易出现肿瘤残留和复发.结论 全面评估肿瘤、操作技术熟练,依据原发部位、分期、范围与生长特点采用适合的入路,充分暴露肿瘤,避免在肿瘤中操作,可有效地减少阳性切缘的发生.     

喉鳞状细胞癌与原癌基因eIF4E的相关性研究

目的研究原癌基因eIF4E(真核细胞翻译起始因子4E)在喉鳞状细胞癌与喉良性病变中的表达,探讨eIF4E与喉鳞状细胞癌的相关性。方法取喉鳞状细胞癌组织标本蜡块40例,取声带息肉组织标本蜡块20例作为对照。采用免疫组化SP法,用eIF4E多克隆抗体检测eIF4E在喉癌组织和声带息肉组织中的表达,并进行统计学分析。结果喉鳞状细胞癌eIF4E的表达高于声带息肉组织中的表达(P<0.01)。eIF4E在喉鳞状细胞癌中的表达与喉癌患者的临床分期、病理分级及患者年龄无明显相关性(P>0.05)。结论原癌基因eIF4E在喉鳞状细胞癌中的表达高于声带息肉,而与喉癌组织的病理分级和临床分期及患者的年龄无相关性。     

喉鳞状细胞癌手术切缘分子水平亚健康与预后关系研究

目的：从分子水平判定喉鳞状细胞癌（LSCC）切除范围的彻底性及与预后的关系。方法：用免疫组织化学方法检测23例LSCC患者手术切缘组织中癌基因EGFR和抑癌基因p53、p27以及与凋亡有关的Bcl-2蛋白的表达情况，并通过随访观察其与肿瘤复发的关系。结果：23例LSCC患者中20例手术切缘组织细胞病理检测为阴性，其中p53阳性3例（15％），p27阳性11例（55％），Bcl-2阳性7例（35％），EGFR阳性3例（15％）。有4例肿瘤局部复发，其p27均为阴性；2例Bcl-2阳性；p53和EGFR均为阴性。结论：用免疫组织化学方法检测LSCC手术切缘EGFR、p53、p27、Bcl-2水平来判断手术范围是可行的，其中p27蛋白表达较为重要，但应同时检测多种基因。     

Jab1和p27蛋白在喉鳞状细胞癌中的表达及二者关系

细胞周期的失调是肿瘤发生的重要原因。p27为CIP／KIP家族的主要成员，具有阻滞细胞通过G1期或S期的关卡作用，其蛋白表达水平及活性改变与肿瘤形成有关。目前研究发现，Jab1能特异性地与p27发生相互作用，使p27由胞核穿梭移动到胞质，通过泛素-蛋白酶体系统加速p27的降解而降低细胞内p27的含量。     

p53蛋白在喉鳞癌及癌旁组织中表达的研究

应用抗ｐ５３单克隆抗休琢ＬＳＡＢ免疫组织化学染色法检测喉鳞癌，喉癌旁组织及声带息肉ｐ５３蛋白的表达。结果显示：２５例喉鳞癌呈阳性反应，４例距肿瘤边缘〈０．５ｃｍ的喉癌旁组织呈弱阳以应，２０例距肿瘤边缘〉２ｃｍ的喉癌旁组织及１６例声带息肉均为阴性。     

喉癌手术安全切缘的免疫组化研究

目的：从基因蛋白水平角度研究喉癌手术安全切缘。方法：应用 Ｌ Ｓ Ａ Ｂ免疫组化法,检测了ｒａｓ、ｃｍ ｙｃ、ｐ５３ 癌基因在３０ 例喉癌,边缘区,癌旁０．５、１．０、１．５、２．０ ｃｍ 粘膜处和 ４ 例正常喉粘膜中的表达情况。结果：三种癌基因产物的单独及联合表达阳性率依正常粘膜→癌旁２．０ ｃｍ →１．５ ｃｍ →１．０ ｃｍ →０．５ ｃｍ →边缘区→癌顺序递增,且癌基因蛋白的表达在距肿瘤０．５ ｃｍ 以内的癌旁组织与癌旁组织１．０ ｃｍ 处有显著性差异（ Ｐ ＜ ０．０５）。结论：距肿瘤０．５ ｃｍ 内的癌旁组织应视为癌前期病变,以切缘距肿瘤０．５ ｃｍ 作为喉癌手术安全切缘的标准较为合适     

p27蛋白和细胞周期素D1在喉鳞状细胞癌中的表达及意义

目的 :探讨喉癌及癌旁组织中抑癌基因p2 7及癌基因细胞周期素D1的表达状况 ,以及p2 7与细胞周期素D1的表达与喉癌临床特征之间的关系。方法 :用免疫组织化学S P法对 38例喉癌组织 ,癌旁组织进行p2 7及细胞周期素D1检测。结果 :p2 7及细胞周期素D1表达在喉癌和癌旁组织之间各有统计学差异。p2 7及细胞周期素D1表达与颈淋巴结转移和患者预后有相关性 (P <0 .0 5 ) ;且大多数受检喉癌组织中 ,p2 7与细胞周期素D1呈反方向表达。结论 :检测p2 7与细胞周期素D1有助于判断喉癌预后。     

Expression of p53, p21 CIP1/WAF1 and eIF4E in the adjacent tissues of oral squamous cell carcinoma: establishing the molecular boundary and a cancer progression model

The present study evaluated the expression of key molecules and the status of DNA in both oral squamous cell carcinoma (OSCC) and adjacent tissues to establish a molecular surgical boundary and provide a cancer progression model. Biopsy samples from 50 OSCC patients were divided into T (cancer), P1 (0–0.5 cm), P2 (0.5–1 cm), P3 (1–1.5 cm) and P4 (1.5–2 cm) groups based on the distances from the visible boundary of the primary focus. Twenty samples of normal mucosa were used as controls. We used immunohistochemical staining and flow cytometry to evaluate p53, p21 ^CIP1/WAF1 , eIF4E and Ki-67 expression and to determine DNA status, respectively. Sub-mucosal invasion was present in the P1 and P2 groups as determined by haematoxylin and eosin staining. Mutant p53 expression decreased gradually from cancerous to normal mucosae, whereas p21 ^CIP1/WAF1 expression displayed an opposite trend. eIF4E expression decreased from cancerous to normal mucosae. Ki-67 expression, the heteroploidy ratio, S-phase fraction and proliferative index decreased gradually with the distance from the tumour centre. Based on these results, we suggest that the resection boundary in OSCC surgery should be beyond 2 cm from the tumour. Additionally, the adjacent tissues of the primary focus could be used as a model for assessing cancer progression.     

喉鳞状细胞癌组织中骨桥蛋白和p53的表达及其临床意义

目的 研究人喉鳞状细胞癌（LSCC）及其癌旁相对正常组织中骨桥蛋白 (OPN)和p53表达水平,并探讨两者的相互关系及其临床病理学意义。方法 采用免疫组织化学SP法,对44例LSCC及癌旁组织手术切除标本常规制作石蜡包埋切片,检测OPN和p53的表达,结合喉癌患者的临床病理学特征进行分析。结果 ①44例喉癌组织中OPN和p53表达阳性率分别为75.0%（33例）和65.9%（29例）,癌旁相对正常组织中分别为13.6%（6例）和2.3%（1例）;②OPN的表达与颈部淋巴结转移、临床分期、病理组织学分级有关,与性别、年龄、原发部位无相关性;③OPN和p53的表达呈显著正相关（r＝0. 360,P＝0.016）。结论 OPN在喉癌组织中的表达高于癌旁相对正常组织,可能参与了喉癌的发生、发展和转移,有可能成为判断预后的重要生物学标记物。     

Molecular analysis of surgical margins in head and neck squamous cell carcinoma patients

OBJECTIVES/HYPOTHESIS: Molecular analysis of surgical margins is playing an increasingly important role in establishing surgical margins. Most markers lack the sensitivity and ease of applicability for effective clinical use. To date, the proto-oncogene eIF4E (4E) is elevated in 100% of head and neck squamous cell carcinoma tumors and is of prognostic value in predicting recurrence. In a retrospective study, 4E overexpression in the margins appeared to be a more sensitive predictor of recurrence when compared with p53. The goal was to confirm this finding in a prospective study and also to compare the expression of matrix metalloproteinase-9 (MMP-9) to 4E expression in tumors and margins. Other objectives were to determine which of these markers have prognostic significance in predicting recurrence and elucidate whether there is any additional benefit to analysis of surgical margins with a combination of the three molecular markers. STUDY DESIGN: A prospective study was performed on all patients who consecutively underwent primary surgical resection between 1998 and 1999 for head and neck squamous cell carcinoma. Patient and tumor characteristics were reviewed, and time to recurrence was noted. METHODS: Paraffin-embedded sections of tumors and all histologically tumor-free margins were analyzed for the presence or absence of 4E, p53, and MMP-9 with immunohistochemical analysis. Patients were followed according to the institution's head and neck cancer protocol, and time to recurrence was noted. RESULTS: Ninety-eight percent of tumors overexpressed 4E, 65% overexpressed p53, and 92% overexpressed MMP-9. Of the 52 patients with tumor-free margins, 52%, 46%, and 54% had positive margins for 4E, p53, and MMP-9, respectively. Although no significant correlation between 4E and p53 expression was seen in the margins (P =.16), a significant correlation between 4E and MMP-9 expression was noted (P =.0002). However, when expression of 4E and p53 in the margins of only the patients who overexpressed p53 in the tumors was compared (n = 34), there was a significant correlation (P =.04). There was also a significant difference in the disease-free interval between patients with 4E-positive and 4E-negative margins (P =.003). This difference in time to recurrence was not significant for the p53-positive versus the p53-negative group (P =.18) but approached significance when MMP-9 was used as a marker (P =.07). Although the univariate analysis showed that stage, nodal disease, grade, and 4E expression in the margins were significantly associated with disease-free interval, in the Cox multiple regression analysis, only 4E expression in the margin was significantly associated with disease-free interval (P =.01). CONCLUSIONS: The era for molecular analysis of surgical margins is here. Although a significant correlation was seen between 4E and MMP-9, overexpression of 4E appears to be a significant predictor of recurrence when compared with the well-studied tumor suppressor gene p53 and a relatively novel marker, MMP-9.     

Ki-67 (MIB1), p53, and Lewis-X (LeuM1) as prognostic factors of recurrence in T1 and T2 laryngeal carcinoma

OBJECTIVES: Recently published data suggest a prognostic value of immunohistochemical proliferation markers for limited laryngeal carcinoma. Previous studies have reported contrasting findings on this issue. In this context, different treatment modalities may be responsible for contradictory findings. To study the relationship between proliferative activity--expressed by the immunohistochemical labeling index of proliferation-associated markers Ki-67 (MIB1), Lewis-X (LeuM1), and proliferating cell nuclear antigen (PCNA) and by p53 status--and treatment failure in a matched-pair study on recurrent and nonrecurrent T1 and T2 glottic carcinoma having received primary transoral laser surgery. METHODS: Twenty-one patients with tumor recurrence were randomly selected and matched with 26 patients with nonrecurrent disease regarding histopathological grading and age. MIB1 staining was used to determine the Ki-67 labeling index, and LeuM1 staining for detecting the Lewis-X antigen; immunohistochemistry determined the p53 status and PCNA labeling index. RESULTS: The Ki-67 labeling index was significantly (P = .001) higher in tumors from patients who had treatment failure (mean = 20.02%) than in patients who did not fail treatment ("nonfailures") (mean = 9.95%). Carcinoma with a Ki-67 (MIB1) labeling index above the median (15%) of the general study population showed a mean time to relapse of 23 months (n = 21), compared with 50 months for cases (n = 26) below the median (P = .016). PCNA labeling index correlated less impressively with tumor recurrence (mean = 28.59% for treatment failures, mean = 21.75% for nonfailures, P = .022). Positive detection of the Lewis-X antigen was significantly associated with recurrence (P = .015) and time to relapse (P = .006). Status of p53 was not a significant prognostic factor. CONCLUSION: The Ki-67 (MIB1) labeling index may be associated with early relapse of limited laryngeal carcinoma treated with transoral laser surgery. Since the prognostic relevance of Ki-67 seems to be different for radiological and surgical concepts of treatment, Ki-67 might become useful as criterion of therapy selection. The Lewis-X antigen, for the first time used on laryngeal carcinoma, seems to be a strong prognostic marker deserving further investigations.     

喉鳞状细胞癌中BAG-1和 p53基因的表达及其临床意义

目的：研究BAG-1和p53基因在喉鳞状细胞癌组织中的表达,探讨BAG-1在喉鳞状细胞癌发生、发展中的作用及与p53的关系。方法：运用免疫组织化学SP技术,检测40例喉鳞状细胞癌组织,27例喉癌旁正常组织及20例声带息肉组织中BAG-1与p53的表达。结果：BAG-1在喉癌中表达阳性率为72．5％,较喉癌旁组织（18．5％）、声带息肉组织（20．0％）高（P〈0．05）,其表达与预后呈负相关（P〈0．05）;p53在喉癌中表达阳性率为65．0％,而在喉癌旁组织和声带息肉组织中均无表达,与在喉癌组织中的表达相比差异有统计学意义（P〈0．05）;其表达与病理分化程度、颈淋巴结转移及预后呈负相关（P〈0．05）;在喉癌组织中BAG-1与p53表达之间未见显著性相关（P〉0．05）。结论：BAG-1和p53基因的过度表达与喉鳞状细胞癌的发生、发展存在一定的关系。BAG-1可作为早期诊断喉癌的一种生物标记,尤其是细胞核BAG-1定位可作为一种不良预后的预测因子;p53表达与喉癌颈淋巴结转移有关,可单独作为喉癌的预后指标。     

喉鳞状细胞癌中细胞周期蛋白E和p27异常表达的临床意义

目的：探讨喉鳞状细胞癌（LSCC）中细胞周期蛋白E（CyclinE）和p27蛋白的表达情况及其临床意义,旨在寻找影响LSCC患者预后的新指标。方法：采用免疫组织化学ElivisionTM法检测160例LSCC和20例正常喉黏膜组织（NLT）中CyclinE和p27的表达情况;统计分析CyclinE和p27蛋白之间及其二者与LSCC患者临床病理因素和总体生存期之间的关系。结果：LSCC组织中CyclinE和p27阳性率分别为62．50％（100／160）和41．25％（66／160）,NLT中CyclinE和p27阳性率分别为35％（7／20）和70％（14／20）（P〈0．05）;CyclinE和p27与LSCC患者淋巴结转移和PTNM分期有关（P〈0．05）,与年龄、性别及肿瘤直径、部位和分化程度无关（P〉0．05）;CyclinE和p27呈显著负相关,r=-0．767（P％0．05）;Cox多因素分析显示淋巴结转移、PTNM分期、CyclinE和p27是影响LSCC患者预后的独立危险因素。Kaplan-Meier生存分析显示LSCC患者的总体生存率为36．9％（P〈0．05）;CyclinE阳性组患者5年生存率为8％,阴性组为80％（P〈O．05）;而p27蛋白阳性组患者5年生存率为77．27％,阴性组为5．32％（P〈O．05）。结论：CyclinE基因的激活同时伴有p27基因的失活可能是LSCC发生发展和侵袭转移的分子基础,联合检测CyclinE和p27对判断LSCC患者预后有重要意义。     

喉癌手术切缘中Survivin和Caspase-3及p53的表达意义及相关性研究

目的:研究Survivin、Caspase-3及p53在喉鳞状细胞癌手术切缘中的表达意义及相关性.方法:116例患者术后按肿瘤复发情况分成4组,分别为:复发≤1年组,＞1～3年组,＞3～＜5年组,≥5年组(包括5年后无复发),应用免疫组织化学方法检测手术切缘Survivin、Caspase-3及p53的表达情况.结果:Survivin在4组中的表达分别为92.9%、85.0%、50.0%及21.1%(P＜0.01)Caspase-3在4组中的表达分别为50.0%、50.0%、66.7%及89.5%(P＜0.01);p53在4组中的表达分别为57.1%、37.5%、45.8%及18.4%(P＜0.05);Survivin、Caspase-3及p53在4组的表达均有统计学意义.结论:喉癌手术切缘中Survivin、Caspase-3及p53的表达在喉鳞状细胞癌的发生、发展中可能起重要作用,并对喉鳞状细胞癌的早期诊断和预后判断有意义.