| Objective: Adenoid cystic carcinoma (ACC) is also named cylindroma. It is one of the most common malignant tumors of the head and neck involving salivary glands. It is generally infiltrating growth with remarkable recurrence and metastasis. This is accounted for histology by its infiltrating capacity and distinct propensity for invasions along nerves and vessels. It is difficult to be entirely resected surgically. Moreover, it is characterized by a high incidence of local recurrence after surgery. So the long-term prognosis of patients is dismal. At present, the therapy for ACC is still to enucleate through an incision directly over the mass. There has not been a potentially effective therapy for recurrent ACC. It grows predominantly with fibroblasts as it relapses. Therefore, it is much more difficult to establish a model of recurrent ACC in nude mice. Up to now, the successful model of recurrent ACC in nude mice has not been reported in the documents. In this study, we successfully established a model of recurrent ACC by using subcutaneous transplantation of intact tumor tissue in nude mice. It could provide an ideal model for clinic therapy and basic research of recurrent ACC. Materials and Methods: 1 Materials: (1) Sample collection: The sample was obtained from a female patient, 45 yeas old, with palatal bump. Clinical diagnosis was recurrent ACC. Pathological diagnosis is adenoid cystic carcinoma relapsed in palate. Under aseptic condition, the tumor tissue was obtained from surgery sample. (2) Animal: 5 SPF grade, 4~6 week-old female Balb/c nude mice were used in this study. 2 Methods: (1) Sample disposal: Under aseptic condition, pieces of tumor tissue obtained from surgery were placed in RPMI-1640 supplemented with streptomycin (500μg/ml) and penicillin (1000U/ml) for 30 minutes, normal tissue and necrotic tissue were removed. The tissues were minced into fine pieces again with fine ophthalmologic scissors and reserved in RPMI-1640. (2) Anesthesia: Every mouse was injected 0.2~0.3ml pentobarbitone solution into abdominal cavity with the concentration of 2.8mg/ml. (3) Transplanting procedure: Under anesthesia condition, the transplanting operation was performed. Small 1-mm3 pieces of recurrent ACC tissue were transplanted subcutaneously in the left anterior back and the right posterior back through a skin incision. At last, incisions were sutured with5/0 absorbable suture. (4) Passaged: When the transplanted tumors were measured approximately 5mm, the mice were killed by cervical dislocation. Under aseptic condition, tumor was stripped and transferred to the next generation nude mouse. (5) Histological observation: The samples of recurrent ACC obtained from the patient and thetransplanted tumor in nude mice were fixed in 10% neutral buffered formalin, embedded in paraffin and cut in 5μm sections which were stained with hematoxylin-eosin. Sections were observed under a multiple functional microscope AX-80. (6) Immunohistochemistry: The sections were de-paraffinized and put into the water. Then they were incubated with the following reagents: Goat serum , primary antibody (CK8 and Actin , 1:50), secondary antibody (1:100), the dilution of ABC(1:100). Subsequently, the sections were stained with DAB and also observed under multiple functional microscope AX-80. (7) Cell culture: The fresh sample of the 3rd transplanted tumor was minced into fine pieces with scissors and put into culture flask. After the tumor cells were free from the clusters, they were harvested and evaluated by Chromosome analysis. (8) Chromosome analysis: The transplanted tumor cells were terminated in the middle period of cell division by colchicine and karyotype was observed. Results: 1. Tumor Growth: Transplantation of human recurrent ACC to nude mouse was successfully established. During 2 years period, the 4th generation of tumor was recorded. A total of 5 Balb/c nude mice were transplanted with recurrent ACC. The average latent period, tumor doubling-time and passage interval was 44.33 days, 11.33 days, and 149.1 days respectively. The...
|
PDF Full Text Request