| Laryngeal carcinoma is the most common malignant disease that occurring in larynx and accounts for 5.7%-7.6% of all malignant tumors. A long history of tobacco use and alcohol abuse are common factors which are thought to be responsible for mutations resulting in laryngeal carcinoma. Recently, the research is focused on oncogene and tumor suppressor gene. For example, alteration of tumor suppressor gene can result in tumorigenesis. Although the level of operation,chemotherapy and radiotherapy has been improved, Probability of surviving for patients with laryngeal carcinoma is still low, especially for recurrent ones. Our study includes three parts:①Mutational analysis of the PTEN gene in laryngeal squamous cell carcinoma; ②Detection of expression of caspase-3 in laryngeal squamous cell carcinoma;③Detection of expression of PTEN, caspase-3,PCNA in recurrent laryngeal squamous cell carcinoma; 1. Mutational analysis of the PTEN gene exon5 and exon8 in laryngeal squamous cell carcinoma First we got genome from fresh human laryngeal squamous cell carcinoma(LSCC) tissue, second we designed two pairs of primer for exon5 and exon8, Then we got the cDNA of exon5 and exon8 by PCR. They are about 253bp and 233 bp.The specific cDNA was cloned into the pMD-18-T vector. Then the recombinant plasmid was digested with EcoR I, HindⅢ. The inserts with the molecular size of 310bp and 290bp were released . The recombinant plasmid was confirmed by DNA sequencing. Exon5,8 of PTEN were detected by TA cloning DNA sequencing in 7 cases. 1case of exon5, transversion GTA→GTT; 1case of exon5, transition GTA→ATA and TGT→TAT which may result in missence mutation. 1 case of exon5, transversion ATA→ATT. 1 case of exon8, transition ACA→ATA, which may result in missence mutation. 1 case of exon5, Mutation has been detected in 4 sites which has been mentioned above. 1 case of exon8, transversion ACA→ACC. Positive rate of PTEN in laryngeal carcinoma is 58.3%(35/60), Accordingly, normal laryngeal tissues and normal laryngeal tissues which were closely adjacent to carcinoma are 100%(10/10) and 95%(19/20) respectively. So we can conclude that PTEN gene may play an important role in LSCC tumorigenesis, the mutation of exon 5,8 of the PTEN gene may be related to the occurrence of LSCC, which isn't decisive.2．Expression of Caspase-3 in laryngeal carcinoma A total of 289 LSCC and corresponding 20 normal tissue,60 atypical hyperplasia of vocal fold were obtained from the department of Otolaryngology, JiLin University, which were studied on expression of Caspase-3 by immunohistochemistry method. 134 glottic type,109 supraglottic type, 27 subglottic type, 19 transglottic type; 58 T1, 61 T2, 117 T3, 53 T4 ; 203 well-differentiated cancer, 86 moderate and poor- differentiated cancer. Caspase-3 was mainly expressed in cytoplasm, seldom in nucleus. High expression was observed in normal laryngeal tissues than in LSCC. Expression of caspase-3 was relevant to clinical stage, pathological grade and lymph node metastasis. 5-year probability rate of caspase-3 positive group was higher than that of caspase-3 negative group. So we can conclude that caspase-3 may be relevant to LSCC tumorigenesis and prognosis. We also study the relation between PTEN and Caspase-3, in conclusion, expression of PTEN and Caspase-3 is down-regulated in tumorigenesis. Moreover, low expression of PTEN can reduce expression of Caspase-3 to make tumor cell apoptosis disorder, so expression of Caspase-3 is relevant to expression of PTEN. However, a further study is necessary to directly clarify regulatory roles of PTEN and expression of Caspase-3 in LSCC. 3. Detection of expression of PTEN, caspase-3,PCNA in recurrent laryngeal squamous cell carcinoma Laryngeal cancer recurrence is very common, which includes local recurrence, lymph node recurrence and distal metastasis. The first two reasons are the main reason for postoperative failure. Totally, 289 cases of LSCC from 1991 to 1998 treated in ENT department of the second clinical hosp...
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