疾病动物模型,淋巴瘤动物模型 z-bio 2021-03-11 387

　　(1) Animal model of chemically induced lymphoma

　　[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.

　　[Method of Modeling] Administration methods such as intraperitoneal injection, intravenous injection, oral administration or gavage, single or divided administration, can induce thymic lymphoma in multi-strain mice. At present, the most commonly used method is intraperitoneal injection: with 8ml of DMSO 92ml of 0.01mol/L PBS (pH=7.2) as the solvent, the MNU is formulated into an induction solution with a concentration of 10mg/ml, which is ready to use. SPF C57BL/6 mice of 6 to 8 weeks are used, both male and female. MNU induction solution (50mg/kg body weight) was injected intraperitoneally for a total of 2 injections (at 0 and 8 weeks respectively). After the start of the injection, continue the basic diet, and observe the general changes in the animal's weight, diet, coat color, and activity.

　　[Model features] ①The formation of mouse thymic lymphoma induced by MNU is actually a single T lymphocyte TCR gene rearrangement, which undergoes malignant transformation and monoclonal proliferation under the action of MNU, which eventually replaces normal cells in the thymus. It is a tumor derived from T cells. ②The structure of the thymus gland was destroyed under the light microscope and replaced by diffusely distributed medium-sized lymphoid tumor cells. ③It can be administered in single or divided doses such as intraperitoneal injection, intravenous injection, oral or gavage, etc., which can induce lymphoma in animals of different strains, but the incidence is different.

　　[Model evaluation and application] The method is simple and easy to implement, the relative period of tumor formation is short and the tumorigenic rate is high; chemical carcinogens often induce multi-site tumors, so they are not often used for drug screening; from the etiological point of view, it is more comparable to human tumors Approximate, so this model is often used for specific in-depth research.

　　(2) Virus-induced lymphoma animal model

　　[Modeling mechanism] Epstein-Barr virus (EBV) is a double-stranded DNA virus with a genome length of 172kb and a linear molecule in the virus particle; it has the characteristics of B lymphocytes and can pass through the EBV/c3d receptor (CD21 ) Infecting B lymphocytes, after entering B lymphocytes, their DNA will be circularized and become an additional component that can replicate outside the chromosomes of B lymphocytes, establishing recessive infections in cells, stimulating cell proliferation and transformation, and inducing diseases.

　　【Method of Modeling】

　　1. For EBV separation, select the standard cell line B95-8 that can release transforming EBV particles and transform marmoset B lymphocytes with EBV in vitro. After large-scale culture, the cells are placed for 7-10 days with starvation therapy, and the culture medium is collected. Centrifuge at 4000r/min at 4°C for 30 minutes, transfer the supernatant to another sterile centrifuge tube, centrifuge at 12 000r/min at 40C for 120-150 minutes, discard the supernatant, and add 1% fresh medium of the original culture solution. Repeatedly blow with a pipette and centrifuge at 3000r/min for 20 minutes. The supernatant was filtered with a 0.22μm disposable filter to obtain a 100-fold concentrated EBV suspension, which was aliquoted and stored at -80°C for later use.

　　2. Lymphocyte separation and inoculation of fresh blood from healthy adults, use the EBVIgA rapid detection kit to detect the VCA-IgA antibody titer. Lymphocytes (PBL) are separated with lymphocyte separation solution, and the volume of PBL is fixed to 8×10000000～10×10000000 with PRIM 1640 culture medium without calf serum. Under aseptic conditions, SCID mice were intraperitoneally injected with 1ml of PBL suspension. Three days after PBL inoculation, 0.4ml of EBV suspension was injected intraperitoneally.

　　3. In the process of model replication, the host may die due to graft-versus-host reaction, which makes the replication model lack of stability. Cyclosporin A (cyclosporin A) can be used to inhibit the above reaction. When using, dilute 1mg/ml with 0.9% NaCl injection and inject After PBL, 10 mg/(kg·d) per SCID mouse was injected intraperitoneally for 2 consecutive days, and 15 mg/(kg·d) from the third day, injected every other day, a total of 11 times.

　　【Characteristics of the model】①The formation of solid lymphocyte tumors induced by the virus, which is aggressive and lethal, belongs to non-Hodgkin's lymphoma with a high degree of malignancy. Histopathological observation showed that the tumor cells were schizophrenic. Non-cleaved cells, some are immunoblast-like morphology, and some have obvious characteristics of plasma cell differentiation. ②The application of gene probe and monoclonal antibody test results showed that the induced tumor is a human EB cell-derived tumor and contains the Epstein-Barr virus small nucleic acid molecule EBER-1, and Epstein-Barr virus particles can be observed in the nucleus of the tumor cell.

　　[Model evaluation and application] Normal cells are derived from the peripheral venous blood of healthy adults, and the specimens are easy to obtain; due to the simple structure of the virus genome, clear molecular background, easy modification and operation, short experimental period (only about 2 months), and small survival Mice have a high tumor induction rate; the induced tumors are invasive solid tumors, which are easy to observe, and their histopathological morphology and growth characteristics are in line with the biological characteristics of human malignant tumors; EBV is widespread in tumor cells, and there is a single cancer-inducing factor. EBV has a direct basis for the cause and effect of human normal cell tumors.

　　(3) Ionizing radiation-induced lymphoma animal model

　　【Modeling mechanism】Ionizing radiation can directly cause DNA damage. Promote the occurrence and development of various tumors in humans and animals. A large number of studies have shown that the thymus, as an important part of the immune system, is the "target organ" of radiation carcinogenesis. Radiation-induced thymic lymphoma in mice has also become one of the classic animal models for studying radiation carcinogenesis. no

　　Whether acute irradiation or fractional irradiation is a high-risk factor for thymic lymphoma.

　　[Modeling method] Inbred BALB/c mice, female, weighing 18-22g, using deep X-ray therapy machine for whole body irradiation, the absorbed dose rate is 0.287Gy/min, the absorbed dose is 1.75Gy, once a week, A total of four times can be molded. The model's success time is about 6 months.

　　[Model features] ①The original structure disappears and is replaced by tumor cells. The tumor cells are large, less cytoplasmic, basophilic, and have various nuclear abnormalities. There are more euchromatin in the nucleus, with obvious nucleoli, and abundant free cytoplasmic ribosomes. The endoplasmic reticulum proliferates and expands on the slippery surface, and viral particles and budding can be seen. The low degree of cell differentiation, fast growth, and vigorous proliferation are consistent with the performance of tumor cells; ②The tumor cells have infiltration and metastasis behaviors, which can be seen in the same strain of mice. Portability.

　　[Model evaluation and application] The induced tumor model is easy to operate, the target organ and the carcinogen are constant, and the induced tumor formation rate is high, which basically simulates the tumorigenesis process. However, the individual variation of the incubation period of tumors is relatively large, and it is not easy to obtain animals with a uniform disease course or tumor size for experimental treatment at the same time.