| ObjectiveHypoxia is a basic characteristic of micro - environment of solid tumors. An-giogenesis and celluar adaptation to hypoxia represent a critical step in tumori-genesis. The key factor regulating cellular O2 homeostasis is hypoxia - inducible factor -1 ( HIF - 1) , which transcriptionally regulate expression of several dozens of the target genes. HIF -1 activates transcription of the genes that are involved in crucial aspects of cancerbiology, including angiogenesis, cell survival, glucose metabolism , invasion and metastasis. Untill now vascular endothelial growth factor (VEGF) is the most prominent angiogenic growth factor, whose expression is regulated by some factors, of which hypoxia is the strongest inducer of VEGF expression. CD34 is widely used in marking vascular endothelial cell and detecting microvessel density (MVD). The specificity of CD34 antigen is higher than others. In order to evaluate the role of HIF - 1α protein in tumori-genesis of human laryngeal squamous cell carcinoma and its relationship with MVD, inmunohistochemical technique was applied to examining expression of HIF - 1α and CD34, which may provide theoretical foundation for antiangiogen-ic therapies of human laryngeal squamous cell carcinoma.Methods1. Patient selection: Tissue samples of human laryngeal squamous cell carcinoma (n =50) and normal laryngeal tissues (n = 10) were obtained from patients of the Second affiliated hospital of China Medical University between 2000 and 2003. All tumors were graded according toWorld Health Organization crite-ria( 1997 ) , male: n = 39, female: n = 11; above 60 years : n -21, below 60 years: n = 23; superglotical: n = 28, glotical: n = 20, subglotical: n = 2. For TNM stages, Tl + T2: n = 21, T3 + T4: n = 29; with lymphnode metastasis: n = 18, without lymphnode metastasis:n =32;well differentiated tumor cells; n = 23, poor differentiated tumor cells ;n =27.2. Immunohistochemistry; A total of 50 patients and 10 normal tissues formalin - fixed and paraffin embedded specimens was available for this study. Expression of HIF - la and CD34 was immuno - histochemically investigated. Paraffin sections were cut at a 4jxm thickness, after repairing antigen and blocking nonspecific reactivity, the sections were incubated with anti - HIF - 1 a poly-clonal and anti - CD34 monoclonal antibody and then with biotin - conjugated second antibody. Positive signals were detected the SABC and SP method with SABC and SP kit. Sections were also counterstained with hematoxylin.3. Result determination: The labeling indexs of HIF - 1 a were calculated microscopically field at a magnification of 400 x by countingtumor cells from 5 selected fields. Strong reactions in the nuclei were considered positive for HIF -la. Microvessels density(MVD) was determined by anti - CD34 immunostain-ing. First,select a microscopically field at a magnification of 40 x,then calculated at a magnification of 200 x by 3 selected fields.4. Statistical methods: Analysis is progressed with statistics software SPSS. Statistical significance was defined as p <0.05.Results1. The immunohistochemistry positive matter of HIF - la and CD34 assumes brown - yellow particles. In normal laryngeal tissues, there was almost no expression of HIF - la( 10% , 1/10) and a little of CD34 expression (13.5 ± 5.4). For the malignant tumors, the expression of HIF - 1 a and CD34 goes up, were (78% ,39/50) and (28.0 ± 11.3). Respectively, There were significant differences. (P<0.01)2. The positive rates of HIF - la were correlated to the depth of invasion and lymphnode metastasis. HIF - la of (T3 + T4) (89.7% ) was significantly... |
