| Purpose: To study the association of P16, VEGF and MVD with the biological behavior of human breast cancers. 87cases of breast cancers were examined in terms of the original tissues of VEGF, P16 protein, and MVD by adopting the method of immunohistochemistry and pattern quantitative analysis in order to make clear the relationship between them and the clinical pathology parameters, and explore the function they play in breast cancer occurrence, development and tumor vascular forming. Subjects and instruments: This was conducted in the General Surgery Department No 1, No2 and No3 Hospital of Jilin University. 87 removed samples of breast cancers from the year 2000 to 2003 were used. The patients were not treated in any way before the surgery and were diagnosed by several surgeons as flung cancers. According to the WHO, the tissues are classified into soked tube cancer (54cases), invasive lobular carcinoma (33cases), the latter one including marrowing carcinoma, pituita adenocarcinoma, glandulose ductal carcinoma and secrete cancer. The levels of the tissues are determined regarding the standard of Bloomand Richardson (1). All the patients are females, from 32 to 68, the average age is 49.8. TNM stages (the standard used in 1988): stage I + stage II, 56 cases; stage III + stage IV, 31 cases. The cross of the tumor: 1-8 cm, among whom 46 cases with metastasis in the lymph node, and 41 without. After all the samples are elected and standardized, they are ready for the normal HE coloring and immunohistochemistry coloring. Immunohistochemistry coloring agents: Ultrasensirive S-P kit , single clone antibiotic CD34, VEGF and P16, all are instantly applied and all are purchased from Maixin Biological Technology Development Corporation, Fuzhou; Ag；liguid. S-P method is used in the process of immunohistochemistry coloring, and PBS is used instead of sample one to do the negative comparison, while the segment tissue of the breast cancer CD34 positive, VEGF positive, and P16 positive are used to do the positive comparison. Results: Diagnose interior, exterior and nuclear of the tumor cells in the vascular endothelial cells with particles of brown and their colors are deeper than the background as positive ones. Method of quantity-fixing in terms of the shapes: 1. Method of capillary quantity-fixing: Referring to the method reported by Weidner, choose the areas with the most microvasculars with the help of 100 times microscope, count the number of the microvasculars colored into brown by CD34 in 5 different areas with 200 times perspective, taking the average number as the MVD. The unit is /200 x, the result is expressed by (±s). The formation of microvascular is not based on whether there are vasculars or not, or if there are red cells in the vasculars. The isolated positive single endothelial cells or cell clusters are considered as one vascular, and the blood vessels with more than 8 red cells and with thicker muscles are not counted. 2. Method of VEGF-counting: Evaluated by the amount of density of the color and the percentage of the positive cells. (a) Density of the color: the negative one is 0, the weak positive is 1, the positive is 2 and the strong positive is 3. (b) Percentage of the positive cells: the non-positive cells are 0, the one < 25% is 1, those between 26-50% are 2, the ones >50% are 3. The biggest number of a+b is 6, 0-2 are considered as negative and 3-6 positive. 3. Method of P16-counting: Choose 5 high-perspectives under the microscope for each section of the organic tissue, and take the average value after counting the number of the positive cells. Those more than 10% of the positive cells within each perspective are positive, and negative if they are less than 10%. Method of statistics: Adopting the method of t-testing, x2 testing and analysis of variance respectively according to the types of the dada. Results: 1. There are 55.17% (48-87) whose P16 is positive among the 87 breast cancer cases, the expression of P16 protein is main...
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