诱发性淋巴瘤,动物造模 z-bo 2022-07-01 467

　　(1) Chemically induced lymphoma animal model [modeling mechanism] The most commonly used inducer is N-methylnitrosourea (MNU). Its carcinogenic mechanism is related to the formation of O6-methylguanine, which causes GyA point mutations during DNA replication. [Modeling method] Single strain or intraperitoneal injection, intravenous injection, oral or gavage divided doses may induce multi-strain mouse thymic lymphoma. Currently, the most commonly used method is intraperitoneal injection: use 8 ml 0.01 mol/LPBS (pH = 7.2) DMSO 92 ml as a solvent, and mix MNU with an induction solution at a concentration of 10 mg/ml. And can be used immediately. Use 6-8 weeks old SPFC57BL/6 mice, male and female. MNU induction solution (50 mg/kg body weight) was injected intraperitoneally for a total of 2 injections (0 and 8 weeks respectively). After the injection, continue the basic diet and observe the general changes in animal body weight, diet, coat color and activity.

　　[Characteristics of the model] ①The formation of mouse thymic lymphoma induced by MNU is actually the rearrangement of the TCR gene of a single T lymphocyte. The effect of MNU leads to malignant transformation and monoclonal proliferation, and the cells that replace the normal thymus are derived from T cells. Tumor. (2) The structure of the thymus gland was destroyed with an optical microscope, and the lymphoid tumor cells of medium size were spread. (3) Single or divided doses such as intraperitoneal injection, intravenous injection, oral and tube feeding can be carried out. Animals of different strains may induce lymphoma, but the incidence is different.

　　[Evaluation and Application of Model] This method is simple and easy to implement, the relative period of tumor formation is short, and the tumor formation rate is high. Chemical carcinogens are often used for drug screening because they often induce multiple tumors. From the etiological point of view, tumors related to the human body are relatively similar, so this model is usually used for some detailed studies. (2) Virus-induced lymphoma animal model [Modeling mechanism] Epstein-Barr virus (EBV) is a double-stranded DNA virus with a genome length of 172 kb, which contains linear molecules in viral particles and B lymph nodes. The characteristic of spherical cells is that they infect B lymphocytes through the EBV/c3d receptor (CD21). Once in the B lymphocyte, its DNA is circularized and becomes another component of the B lymphocyte, which can replicate outside the chromosomes, thereby establishing a recessive infection in the cell. It stimulates cell growth and transformation, and induces diseases.

　　[Modeling method]

　　"1. For EBV isolation, select B95-8 cells, a standard cell line that will release transformed EBV particles and can be used to transform mar monkey B lymphocytes with EBV in vitro. After large-scale cultivation, the cells are placed in starvation therapy for 7-10 days, and the medium is collected. Centrifuge at 4000/min for 30 minutes at 4°C, transfer the supernatant to another sterile centrifuge tube, centrifuge at 12000/min for 120-150 minutes at 40°C, discard the supernatant and discard Original culture. Reuse by adding 1% fresh liquid medium. Centrifuge with a pipette and centrifuge at 3000r/min for 20 minutes. The supernatant was filtered through a 0.22 μm disposable filter to obtain a 100x concentrated EBV suspension. Distribute and store at -80°C for future use.

　　2. Isolate lymphocytes and inoculate with fresh blood from healthy adults, and use EBVIgA rapid detection kit to detect VCA-IgA antibody titer. Lymphocyte separation solution (PBL) is used to separate lymphocytes (PBL). In PRIM1640 medium without calf serum, the volume of PBL is fixed at 8×10000000 to 10×10000000. Under aseptic conditions, 1 ml of PBL suspension was injected intraperitoneally into SCID mice. Three days after PBL inoculation, 0.4 ml of EBV suspension was injected intraperitoneally.

　　3. During model replication, the host may die due to grafting and host reaction, which will make the replication model unstable. Cyclosporin A (Cyclosporin A) can be used to block the above reaction. If used, inject 0.9% NaCl to dilute to 1 mg/ml, and then inject PBL. Then, each SCID mouse was injected intraperitoneally with 10 mg/(kg·d) for 2 consecutive days, and 15 mg/(kg·d) was injected intraperitoneally from the third day, for a total of 11 injections every other day.

　　[Characteristics of the model] ① An aggressive and lethal virus that belongs to high-malignant non-Hodgkin's lymphoma forms solid lymphocyte tumors. Histopathological observation showed that the tumor cells were large cracks. Some are uncut, some have an immunogenic morphology, and some have characteristics of plasma cell differentiation. (2) The result of detection with gene probes and monoclonal antibodies is that the induced tumor is a tumor derived from human EB cells, which contains the small Epstein-Barr virus nucleic acid molecule EBER-1, and love is observed in the tumor cell nucleus. Perstein-Barr virus particles. .. [Model Evaluation and Application] Normal cells are derived from peripheral venous blood of healthy adults, and samples are easy to obtain. Due to the simple structure of the virus genome, clear molecular background, easy modification and operation, and short experiment period (about 2 months). Surviving mice have a high tumor induction rate. The induced tumors are aggressive solid tumors, which are easy to observe, and their histopathological morphology and growth characteristics are the biological characteristics of human malignant tumors. Consistent with... EBV is widely distributed in tumor cells and has a single cancer-inducing factor. The direct basis for the causal relationship between human normal cell tumors induced by EBV can be obtained.

　　(III) Animal model of lymphoma induced by ionizing radiation

　　[Modeling mechanism] Ionizing radiation can directly cause DNA damage. Promote the development of various tumors in humans and animals. Many studies have shown that the thymus is an important part of the immune system and the "target organ" of radiation carcinogenesis. Radiation-induced thymic lymphoma in mice has also become one of the classic animal models for studying the carcinogenic effects of radiation. Acute or fractional irradiation is a high risk factor for thymic lymphoma.

　　[Modeling method] Male and female, inbred BALB/c mice weighing 18-22g, using a whole body X-ray therapy device, female, absorbed dose rate is 0.287Gy/min, absorbed dose is 1.75Gy, 4 times a week One-time molding. The success period of this model is approximately 6 months.

　　[Characteristics of the model] ① The tumor cells that have lost their original structure and replaced tumor cells are large, with a small number of cells, basophils, and diverse nuclei, with a lot of euchromatin and clear nucleoli. In addition, the abundant growth of the endoplasmic reticulum and the absence of cytoplasmic ribosomes on the smooth surface, visible virus particles and budding phenomena, low cell differentiation, fast growth, and vigorous growth are related to tumor cells. 2) Tumor cells are transplantable and have invasion and metastasis behavior in mice of the same strain.

　　[Model Evaluation and Application] The induced tumor model is easy to operate, has constant target organs and carcinogens, has a high induced tumor formation rate, and basically simulates the development process of tumors. However, the incubation period of tumors varies greatly among individuals, and it is not easy to simultaneously obtain animals with the same disease course or tumor size for experimental treatment.