An Experimental Study of Radiation Therapy Combined with Hyperihermia on Angiogenesis of Hepatocellular Carcinoma in Mice
|School||Taishan Medical College|
|Keywords||Hepatic carcinoma Radiotherapy Hyperthermia Angiogenesis Vascular endothelial growth factor Microvessel density|
Purpose entities tumor in its development process occurs in a large number of tumor angiogenesis, blood vessel formation is the basis of tumor cell growth, invasion and metastasis. The primary liver angiogenesis rich one of the characteristics of the tumor. The high incidence of primary liver cancer, but the treatment less effective. In this study, using radiotherapy combined with hyperthermia experimental study of liver cancer in mice with H22 tumor angiogenesis using immunohistochemical methods to detect groups of tumor-bearing mice tumor vascular endothelial growth factor (VEGF) and microvessel density (MVD) change hematoxylin - eosin staining (HE) staining the tumor pathological changes; electron microscopy cell ultrastructure changes of radiation therapy combined with hyperthermia in nude mice tumor angiogenesis theory for the clinical treatment of liver cancer basis and new therapies. Method under aseptic conditions decimation H22 tumor mice ascites, 1500r/min conditions centrifuged 20min, the supernatant was discarded, adjusted sterile PBS for 2 × 107/ml. Take 0.2ml cell suspension was injected into Kunming mice right hind limb subcutaneous. Two weeks after planting, the subcutaneous group therapy spherical tumor began to grow about 1 cm in diameter classes. No significant difference in tumor size before grouping. Take 40 tumor-bearing mice, hyperthermia radiotherapy (joint group A, group), the radiotherapy group (B), the hyperthermia group (C) and the control group (D group), each with a random number table is divided into 10. Radiation therapy combined with hyperthermia group -60 long-range radiation therapy machine with Xinhua Medical Instrument Factory production drilling radiotherapy. Treatment anesthetized tumor-bearing mice at the same time fixed in plexiglass plate. The tumor-bearing legs fixed traction within the radiation field, radiation field edge beyond the tumor margin of at least 5 mm the the tumor surface coverage thickness of 5mm compensation film to increase the surface dose radiation field 15cm × 15 cm, source skin distance 60cm, Dose rate 81.05cGy of / min, single radiation dose 3Gy, once daily treatment for 10 consecutive days, the total radiation dose was 30Gy. Within half an hour after the end of radiotherapy given hyperthermia. With a constant temperature water bath ANTITUMOR Bureau give 41.5 ℃ ± 0.2 ℃ hyperthermia, each 50 minutes; weekly treatment, a total of 4 times. Radiotherapy group and hyperthermia group was given a single treatment. The control group was the control. During treatment every other day with a vernier caliper measurement of tumor volume, measured by balance weight. The first 5 days after the end of treatment, the mice were sacrificed by cervical method. The complete anatomical mouse tumor tissue, electron microscopy specimens drawn; measured using an analytical balance neoplasia heavy: immunohistochemical staining tumor MVD value of VEGF positive expression rate; HE staining of tumor tissue pathological changes observed with an optical microscope; tumor cells using electron microscopy ultrastructure. Variance analysis with SAS8.1 statistical package, whether the statistically significant difference between the comparison groups corresponding data. Results of radiation therapy combined with hyperthermia group (combination group, A group), the radiotherapy group (B), hyperthermia group (C) and the control group (D) tumor growth rate, respectively (9.39 mm3 / d 12.02mm3 / d , 15.91mm3 / d, vs.21.18mm3 / d, F = 3.23, P = 0.037); inhibition rates were (74.8%, 58% and 41.8%, respectively), the inhibitory effect of the combination group. No significant difference in body weight of mice (F = 0.57, P = 0.639). The mean MVD value of the combined group, radiotherapy group, hyperthermia group and control group were 35.2 ± 5.1,48.5 ± 4.6,58.1 ± 7.4 and 69.7 ± 9.6 (F = 44.24, P = 0.001), the difference was statistically significant. The joint group, radiotherapy group, hyperthermia group and control group, VEGF-positive rate was 11.4% 4-3.8%, 16.9% ± 3.9%, 20.8% ± 3.1%, 25.5% ± 6.4%, the difference between statistical significance (F = 17.42 P lt; 0.05), statistically significant. HE pathological evaluation: A group of a large number of tumor cell nuclei concentrated, tumor cells arranged in loose, separated from the neighboring cells, obviously less eosinophilic cytoplasm, sinusoids significantly reduced; radiotherapy group visible swelling of the tumor cells, nuclear pyknosis small part of the tumor cell nuclei , the tumor sinusoids less; hyperthermia group visible the tumor center point sheet ischemic necrosis, some tumor cell nuclei pyknotic cells arranged loosely rich blood flow of the tumor periphery; D group tumor tumor cells thrive, trabecular like closely spaced nuclear See mitotic. The tumor edge blood Doufeng Fu; oncology centers exuberant tumor growth ischemic necrosis. Projection electron microscopy: A group shows visible number of apoptotic and necrotic cells, nuclear condensation, nuclear fragmentation. Group B tumor cell atypia smaller cell morphology tend to be normal. C were observed in a small number of apoptotic (nuclear fragmentation cells), tumor cell nuclei and nucleoli, heterochromatin more organelles in the cytoplasm more obvious atypia. Group D atypia of tumor cells, the proliferation of the more common active tumor cells, tumor cell disorder, large nuclei, irregular shape, large and obvious nucleoli, nuclear showed the profiled organelles nuclear chromatin The mitotic visible. Conclusion radiotherapy combined with hyperthermia generated tumor vascular tumor-bearing mice, the experimental study, found that radiotherapy combined with hyperthermia can significantly inhibit tumor MVD value and VEGF positive rate of immunohistochemical staining; groups of tumor-bearing mice tumor volume body weight in the treatment of different time measurement found that radiotherapy combined with hyperthermia tumor inhibition rate; no significant difference in body weight, no significant increase in toxicity; pathological combination of tumor tissue HE staining and electron microscopic observation, the combination group significantly inhibited tumor angiogenesis to kill tumor cells also increase or promote tumor cell apoptosis. Confirmed that radiotherapy combined with hyperthermia synergistic anti-hepatoma growth effect, part of the experiment the theoretical basis for the choice of liver cancer treatment strategies.