喉咽鳞状细胞癌及其颈淋巴结微卫星位点杂合性缺失的研究

目的 :探讨喉咽鳞状细胞癌 (LPSCC)D9S171微卫星位点杂合性缺失 (LOH)的临床意义及对检测颈淋巴结微转移的应用价值。方法 :应用聚合酶链反应对 18例LPSCC及 72个颈淋巴结在 9号染色体上的D9S171微卫星位点进行扩增 ,分析D9S171微卫星位点的DNA等位基因的LOH。结果 :颈淋巴结组织D9S171位点的LOH发生率 4 0 .2 8%显著高于常规病检的阳性率 8.33% (P <0 .0 1) ;肿瘤组织中的LOH发生率与肿瘤分化程度有关 (P <0 .0 5 ) ,并且与肿瘤临床分期呈密切线性正相关性 (P <0 .0 1)。结论 :D9S171位点的LOH分析可能是检测LPSCC颈淋巴结微转移的较为敏感的手段之一 ,并可能作为判断LPSCC患者预后的指标之一。     

微切割喉鳞状细胞癌9p13-23区域微卫星杂合性缺失的研究

目的探讨喉鳞状细胞癌(简称鳞癌)在9p13-23区域微卫星(microsatellite)发生杂合性缺失(1ossofheterozygosity,LOH)的热点.方法采用显微切割法从病理切片中挑取肿瘤组织,选取位于9p13-23区域的13个高多态性微卫星引物对42例喉鳞癌组织进行聚合酶链反应和变性凝胶电泳.结果①42例喉鳞癌在9p13-23区域等位基因LOH的总发生率是97.6%(41/42).在13个微卫星引物中,LOH发生率最高者是位于9p22-23的D9S162(89.5%),其次是位于9p21的D9S171(80.0%).与p16基因紧密连锁的D9S1748的LOH发生率仅50.0%.②等位基因缺失作图分析发现42例喉鳞癌组织在9p13-23上存在2个明显的LOH较小区域,分别位于99211的D9S161～D9S171之间和9p22-23的IFNA和D9S162之间.结论喉鳞癌在9p13-23区域除抑癌基因p16以外可能还存在2个或2个以上候选抑癌基因,这些候选抑癌基因也许和p16一样与喉鳞癌的发生、发展密切相关.     

喉癌前病变及喉鳞状细胞癌微卫星杂合性缺失的变化及其意义

目的:探讨喉癌前病变及喉鳞状细胞癌病变组织上杂合性缺失(LOH)的特征及其意义。方法:选取染色体3p、9p和17p上6个多态性微卫星位点D3S1234、D9S171、D9S1748、D9S162、INFA和p53,利用聚合酶链式反应-简单序列长度多态性-银染技术,对49例喉癌前病变和喉癌组织进行LOH分析。结果:6个微卫星标记物LOH发生率分别为:单纯过度增生3.70%,轻度不典型增生10.81%,重度不典型增生26.03%,喉鳞状细胞癌38.67%。其中LOH的总检出率在不同病理组间差异有统计学意义(χ2=17.686,P<0.01),其频率随病变程度加重而明显升高。6个多态性微卫星位点中,LOH发生率最高的位点D9S171(35.00%)。结论:基因水平的改变发生在喉癌变的早期阶段,微卫星标志物可能成为喉癌前病变早期诊断的有用标志物。     

喉鳞状细胞癌中脆性组氨酸三联体基因微卫星的不稳定性和杂合性丢失

目的　探讨脆性组氨酸三联体 (fragilehistidinetriad ,FHIT)基因微卫星不稳定性(microsatelliteinstability ,MSI)和杂合性丢失 (lossofheterozygosity ,LOH)与喉鳞状细胞癌 (简称鳞癌 )发生、发展的关系。方法　采用聚合酶链式反应 简单序列长度多态性 银染技术 ,分析 4 1例喉鳞癌中FHIT基因D3S12 34和D3S130 0位点的MSI及LOH。结果　D3S12 34位点LOH发生率为 4 4 4 % (16 /36 ) ,MSI发生率为 19 4 % (7/ 36 ) ;D3S130 0位点LOH发生率为 36 4 % (12 / 33) ,MSI发生率为 2 4 2 % (8/33)。两个位点总的LOH发生率为 5 2 6 % (2 0 / 38) ,总的MSI发生率为 2 8 9% (11/ 38)。总的LOH发生率与喉鳞癌患者TNM分期、病理分级、淋巴结转移及复发有关 (P <0 0 5 ) ,总MSI发生率与喉鳞癌患者淋巴结转移有关 (P <0 0 5 )。结论　FHIT基因LOH和MSI与喉鳞癌的发生、发展有关 ,并可能为喉鳞癌的早期诊断提供新的途径和依据     

微切割喉鳞状细胞癌9p13—23区域微卫星杂合性缺失的研究

目的：探讨喉鳞状细胞癌（简称鳞癌）在9p13-23区域微卫星（microsatellite)发生杂合性缺失（loss of heterozygosity,LOH)的热点。方法：采用显微切割法从病理切片中挑取肿瘤组织,选取位于9p13-23区域的13个高多态性微卫星引物对42例喉鳞癌组织进行聚合酶链反应和变性凝胶电泳。结果：（1）42例喉鳞癌在9p13-23区域等位基因LOH的总发生率是97．6％（41／42）。在13个微卫星引物中,LOH发生率最高者是位于9p22-23的D9S162（89．5％）,其次是位于9p21的D9S171（80．0％）,与p16基因紧密连锁的D9S1748的LOH发生率仅50．0％,（2）等位基因缺失作图分析发现42例喉鳞癌组织在9p13-23上存在2个明显的LOH较小区域,分别位于9p21的D9S161－D9S171之间和9p22-23的IFNA和D9S162之间。结论：喉鳞癌在9p13-23区域除抑癌基因p16以外可能还存在2个或2个以上候选抑癌基因,这些候选抑癌基因也许和p16-一样与喉鳞癌的发生,发展密切相关。     

喉癌及癌前病变等位基因不平衡性的相关研究

目的探讨喉鳞状细胞癌变过程中微卫星DNA等位基因不平衡性的特征及其意义。方法选取染色体3P、9P和17P上6个多态性微卫星位点D3S1234、D9S171、D9S1748、D9S162、INFA和D17S796,利用聚合酶链式反应一简单序列长度多态性一银染技术,对49例喉癌癌前病变和喉癌组织进行等位基因不平衡分析,统计杂合性缺失（10ssofheterozygosity,LOH）和微卫星不稳定性（microsatelliteinstability,MSI）的发生率及其与临床病理特征的相关性。结果6个微卫星标记物LOH和MSI发生率分别为：喉癌癌前病变中单纯过度增生为3．7％和14．8％,轻度不典型增生为10．8％和21．6％,重度不典型增生为26．0％和23．3％;喉鳞状细胞癌为38．7％和21．3％。其中LOH的总检出率在不同病理组间有统计学意义（X2=17．686,P=0．000）,而MSI的检出率统计学意义（X2=0．314,P〉0．05）。不同病理组间D9S171和D9S162单个位点LOH检出率有统计学意义（P=0．022,P=0．025）。在癌前病变早期MSI发生率高于LOH。结论等位基因不平衡可能参与喉癌发生发展,微卫星分析法为喉癌癌前病变的早期诊断提供新的途径。     

头颈部鳞癌患者尿酚含量测定的临床意义

目的探讨尿酚含量与头颈鳞癌临床分期、组织学分级、颈淋巴结转移之间的关系,评价它在临床检测中的意义.方法检测34例头颈鳞癌、9例良性肿瘤患者及30例正常人尿中尿酚含量,比较它在手术前后的变化,并与30例正常人的尿酚含量对照.结果正常人、良性肿瘤、鳞癌患者尿酚阳性率分别为6.67%、11.11%、52.94%,正常人明显低于鳞癌患者(P＜0.05).头颈鳞癌T1～T2期患者尿酚阳性率为77.78%,明显低于T3～T4期的45.83%(P＜0.05).结论肿瘤标志物尿酚可作为临床辅助诊断手段.     

头颈鳞癌的颈淋巴结转移与nm23—H1基因的相关性研究

目的：研究转移抑制基因nm23-H1的表达产物NDPK（二磷酸核苷激酶）在人头颈鳞癌原发灶中的表达及其意义。方法：应用S－P染色方法检测106例头颈鳞癌原发灶组织中nm23-H1/NDPK的表达,结果:nm23-H1/NDPK表达在未发生颈淋巴结转移的头颈鳞癌原发灶中的表达阳性率为75．41％（46／61）,在已发生颈淋巴结转移的头颈鳞癌原发灶中的表达阳性率为35．56％（16／45）,两者差异有显著性（P＜0．05）,结论：nm23-H1/NDPK表达与头颈鳞癌的颈淋巴结转移与否呈显著负相关,可能在头颈鳞癌的颈淋巴结转移过程中起负性调控作用,可用于预估颈淋巴结转移出现的风险。     

头颈鳞癌血管生成与颈淋巴结转移相关性研究

目的 :探讨头颈鳞癌血管生成与其颈淋巴结转移的关系以及血管内皮生长因子(VEGF)在头颈鳞癌血管生成中的作用。方法 :应用免疫组化SABC法检测 5 8例头颈鳞癌组织中微血管密度 (IMVD)及VEGF的表达。结果 :5 8例头颈鳞癌组织中IMVD为 2 3.93± 8.77,肿瘤分化程度 ,高与中、高与低间 ,IMVD差异有显著性意义 (均P <0 .0 5 ) ;中与低间 ,差异无显著性意义 (P >0 .0 5 )。颈淋巴结转移组IMVD(2 7.92± 9.11)明显高于非转移组 (2 0 .6 9± 7.0 8) ,其差异有显著性意义 (P <0 .0 1)。癌组织中VEGF表达与瘤内IMVD呈正相关 (rs=0 .4 87,P <0 .0 1)。结论 :瘤内IMVD可作为预测头颈鳞癌颈淋巴结转移的一个重要指标 ;VEGF可促进头颈鳞癌血管生成。     

喉癌、喉咽癌脆性组氨酸三联体基因杂合性缺失和微卫星不稳定

目的 探讨脆性组氨酸三联体(fragile histidine triad,FHIT)基因杂合性缺失(loss of heterozygosity,LOH)和微卫星不稳定(microsatellite instability,MSI)与喉癌、喉咽癌发生发展的关系.方法 选取FHIT基因内的三个微卫星位点D3S1234、D3S4103、D3S1300进行PCR扩增,应用单链长度多态性分析-银染技术进行LOH和MSI分析.结果 ①D3S4103、D3S1234、D3S1 300的杂合率分别为76.32%、71.05%和78.95%;②喉癌、喉咽癌总LOH发生率分别是:D3S4103(48.28%)、D3S1234(37.04%)、D3S1300(33.33%);③喉癌、喉咽癌D3S1234、D3S1300、D3S4103的总LOH发生率与患者年龄、性别、吸烟、肿瘤分化程度、肿瘤部位、T分期和肿瘤复发与否的关系均无统计学意义(P＞0.05);④喉癌、喉咽癌总MSI发生率分别是:D3S1234(18.42%)、D3S1300(28.95%)、D3S4103(21.05%);⑤喉癌、喉咽癌FHIT基因D3S1234、D3S1 300、D3S41 03的总MSI发生率与患者年龄、性别、吸烟史、肿瘤部位、肿瘤分化程度和T分期的关系无统计学意义(P＞0.05);⑥复发喉癌、喉咽癌病例FHIT基因总MSI发生率为83.33%,原发病例总MSI发生率为30.77%,复发病例MSI阳性率显著高于原发病例,差异具有统计学意义(P=0.004).结论 ①FHIT基因在喉癌、喉咽癌中既存在杂合性丢失,又存在微卫星不稳定,但以前者为主;②FHIT基因参与了喉癌、喉咽癌的发生,可能是喉癌、喉咽癌候选抑癌基因之一;③FHIT基因微卫星不稳定与喉癌、喉咽癌复发可能相关.     

Determination of lymph node micrometastases in patients with supraglottic carcinoma

CONCLUSIONS: The frequency of loss of heterozygosity (LOH) at D9S 171 microsatellite locus on 9p21 may serve as an available method to evaluate occult micrometastases in laryngeal squamous cell carcinoma. High frequency of LOH was associated with a decreased probability of survival time. OBJECTIVE: To explore an available and sensitive method to detect cervical lymph node micrometastases in patients with laryngeal squamous cell carcinoma, the frequency of LOH at D9S171 microsatellite locus on 9p21 was studied. PATIENTS AND METHODS: Twenty samples from supraglottic cancer and 182 lymph nodes from neck dissections were examined by LOH comparing immunohistochemical (IHC) staining using cytokeratin 19 (CK19), and hematoxylin and eosin (H&E) staining. The frequency of lymph node metastasis and the clinical relevance were analysed. RESULTS: The frequency of LOH was 37.4% of lymph nodes and all of the primary tumors. Occult micrometastases were present in 9 of 20 cases; 23.6% of lymph nodes were positive for CK19 by IHC; 16.5% of lymph nodes were positive by H&E. There was a highly significant difference among the three methods. The highest rate of positive lymph nodes was at level II of the neck. There was a highly significant difference between overall survival time and lymph node metastasis with LOH and CK19 analysis.     

The role of genetic susceptibility in head and neck squamous cell carcinoma

Our research is an additional genetic study to uncover the molecular mechanisms involved in head and neck squamous cell carcinoma (HNSCC) pathogenesis by studying loss of heterozygosity (LOH) and microsatellite instability (MSI) in both premalignant and malignant patients and to highlight the genotype of HNSCC in Upper Egypt. Patients with HNSCC from various parts of the world may have unique genotypes and this is the first genetic study of HNSCC in Sohag 500 KM to the south of Cairo. We performed a prospective study of 41 patients with precancerous and 79 patients with cancerous laryngeal, esophageal, nasopharyngeal, nasal and oral lesions, and 50 controls (The control patients were cases admitted for ear surgery or simple nasal surgery, from whom we took biopsy from mucosal lining of nasopharynx). The present study included 170 individuals who were admitted to the Ear, Nose and Throat department, Sohag University Hospital, Sohag, in Egypt in the period between April 2001 and March 2003. Samples which were taken by punch biopsy were frozen and stored at −80°C and were subjected to histopathological examination. We investigated LOH and MSI by using six microsatellite markers located at chromosomes 3, 5, 9, and 17. The markers used were D3S1286, D9S171, D9S753, D17S654, D17S695, and CFS1-R. LOH was in all premalignant and malignant lesions at 5q33.3-q34 and 13% of Controls. LOH at 17p21 was absent in all premalignant lesions and was found in 53% of malignant lesions and 12.4% of Controls. In premalignant lesions, LOH was at 3pter-3p24.2 (73% of cases), at 9p21 (46%), at 9q21.1-22.3 (37%), and at 17p13 (37%). These percents increased in malignant lesions to 87, 80, 67, and 63%, respectively. They were 14, 19.4, 17, and 19% in controls. Examination of LOH could improve diagnosis, adds additional confidence, in HNSCC by DNA extraction from suspicious lesions in high-risk groups (smokers and alcoholics) and LOH at 3p/9p seems to be of particular value for early detection and definition of progression risk. If there are high percent of LOH at these chromosomes, active intervention should be done (chemoprevention and regular follow up head and neck examination for very early detection and management).     

Loss of heterozygosity of 3p21 and 9p21 in head and neck squamous cell carcinomas and its prognostic implication]

To examine whether genetic factors influence the prognosis of cancer patients, several microsatellite markers were used to determine the allelic loss of certain areas of the genome. Three microsatellite markers, D3S1067, IFNA and D9S171 were used to study the loss of heterozygosity (LOH) of 3p21 and 9p21 in 93 head and neck squamous cell carcinomas. Of 57 informative cases, LOH was detected in 27 of 57 (47%) DNA samples obtained from cancer specimens when at least one marker was used. The frequency of LOH was not correlated with the clinical factors. However, the frequency of LOH was significantly higher in the recurrent cases than in the non-recurrent cases, and patients with 3p21 and/or 9p21 LOH tended to survive for a shorter period of time. These results suggested that the allelic loss at 3p21 and/or 9p21 could be correlated with the prognosis of the patients, and that it was a novel prognostic factor independent of other clinical factors concerning head and neck cancers. LOH at 3p21 and/or 9p21 may help to identify head and neck cancer patients with a poor prognosis, who need an intensive postoperative follow-up protocol, or who are suitable for novel investigational therapeutic approaches.     

Role of microsatellites instability in carcinogenesis of postcricoid carcinoma on top of plummer-vinson syndrome

To develop a molecular pattern that might help in understanding carcinogenesis of postcricoid carcinoma (PCC) on top of Plummer-Vinson syndrome (PVS) in a prospective controlled study. Twenty-four patients with PVS were diagnosed and followed up over a 4?year period, during which eight of them showed malignant change to PCC. Twenty volunteers free of neoplastic diseases were included as a control group. In the two groups, DNA extraction from mononuclear peripheral blood cells, and analysis of loss of heterozygosity (LOH) and microsatellite instability (MSI) using six paired simple tandem repeats (STRs) primers were done. The molecular weight of each STRs locus was scored and statistical correlations were performed. LOH occurred in 55.6 and 72.9% of PVS and PCC cases compared to 25% of control group. At loci D17S695, D9S753 and D9S171, LOH occurred in 54.2, 66.7, and 70.8% of PVS cases; and in 62.5% of PCC cases for each locus compared to 15, 25 and 45% of control cases. D3S1286 and CFS1-R displayed the highest frequency of LOH in PCC (100% for each) while recorded in 58.3 and 33.3% in PVS compared to 30 and 0% in control cases. Certain genetic events tend to occur as early and late events in malignant change of PVS to PCC. Detection of these events may help in understanding carcinogenesis and in early detection of malignancy. CFS1-R is the most informative marker of tumor progression.     

Elective treatment of the neck for second primary tumors of the head and neck

The aim of this study was to define the role of elective neck dissection in patients with a second N0 head and neck squamous cell carcinoma (HNSCC). We carried out a retrospective study in 74 patients with a second N0 HNSCC treated with an elective neck dissection. Thirteen patients (17.6 %) had occult neck node metastases. The risk of occult neck nodes was low for patients with a second glottic tumor (0 %), and for patients with non-glottic T1–T2 tumors who had received previous radiotherapy in the neck (5.3 %). Patients with non-glottic locally advanced tumors (T3–T4) and non-glottic T1–T2 tumors who had not received previous radiotherapy in the neck had a risk of occult neck nodes of 28.1 and 33.3 %, respectively. Elective neck dissection could be omitted in patients with glottic tumors and in patients with an early tumor (T1–T2) who had received previous radiotherapy in the neck.     

头颈部鳞状细胞癌远处转移的相关因素分析

目的探讨头颈肿瘤远处转移的相关影响因素。方法对532例头颈部原发鳞状细胞癌患者的临床病理资料进行回顾性分析。选择性别、年龄、临床分期、T分级、N分级、原发癌部位、原发癌浸润深度、原发癌病理分级、有无颈淋巴结转移、颈阳性淋巴结数目、颈淋巴结转移累及区域、颈阳性淋巴结破膜情况等临床病理因素，用)(2检验和Logistic回归进行单因素和多因素分析，并用．Kaplan-Meier法对发生远隔部位转移患者进行生存分析。结果在532例头颈部原发鳞状细胞癌患者中，60例(11．3％)发生远处转移。单因素分析显示，临床分期(P=0．0126)、T分级(P=0．0082)、原发癌部位(P=0．0011)、原发癌浸润深度(P=0，0005)、有无颈淋巴结转移(P=0．0057)、颈阳性淋巴结数目(P=0．0149)、颈淋巴结转移累及区域(P=0．0034)、颈阳性淋巴结破膜情况(P=0．0118)与发生远处转移有关。多因素分析结果表明，仅原发癌部位、原发癌浸润深度与发生远处转移明显相关。用Kaplan-Meier法进行生存分析，结果显示60例发生远隔部位转移患者的1年生存率、3年生存率、5年生存率分别为51．7％、13．3％、6．5％。结论原发肿瘤部位和浸润深度是发生远处转移的共同决定性因素。而原发癌临床分期、T分级和有无颈淋巴结转移是头颈鳞癌远处转移的影响因素，但不是导致远处转移的初始和根本因素。喉癌、下咽癌以及原发癌侵犯肌肉、骨或软骨患者易发生远处转移。