动物模型,闭塞性脉管炎 z-bo 2020-08-06 474

　　Thromboangiitis obliterans is a peripheral vascular disease dominated by autoimmunity, smoking, cold and other external factors and sex hormones are the main cause of the disease. The following describes the method of using the above-mentioned pathogenic factors to replicate the animal model of thromboangiitis obliterans.

　　(1) Copy method

　　1) A model of thromboangitis obliterans in rats by tobacoo and immune extract

　　Wistar rats weighing 200-250g, 6-8 months old. Before modeling, the rats adapt to the environment for 1 week. The hind limbs of the rats were depilated, and the immune extracts from vasculitis (TAO) patients were subcutaneously injected around the hind limb arteries of the rats, each time 0.1ml, after subcutaneous injection, skin hills were formed, once every 4 days. Place the rats injected with the TAO immune extract for the first time in a drying container, and use a plastic tube to introduce the smoke formed by the slow combustion of tobacco in the combustion can into the drying container, 15 minutes each time, 1-2 times a day. Then, 25% of the tobacco extract was injected subcutaneously around the hind limb arteries of rats, once daily. The modeling cycle is 2 months in total. After modeling, blood was collected to detect changes in whole blood and plasma viscosity and erythrocyte sedimentation rate; the middle, small arteriovenous and surrounding muscle tissues of the distal end of the affected limb of the rat were taken, fixed with 10% formaldehyde, and embedded in paraffin. Make conventional tissue sections, stain with HE, and observe under light microscope. The arteries below the knee joints of the affected limbs of the rats were sharply cut about 5 mm, and they were cut longitudinally and flattened to make a size of 5 mm2. Wash the blood in the lumen carefully with physiological saline and place it in a 2.5% glutaraldehyde fixative solution pre-cooled at 4°C. After fixation for 12 hours, wash it with 0.1mol/L phosphate buffer (pH 7.2) for 2 hours (change in between) 2~3 times new solution), then fix with 1% osmium acid for a second time, wash with double distilled water for 2h (change 2~3 times in the middle of the new solution), dehydrate with rising gradient ethanol, replace with isoamyl acetate, low vacuum critical The spots are dried, platinum-plated on the IB-5 ion sputtering apparatus, and observed and photographed with a scanning electron microscope.

　　2) A model of thromboangitis obliterans in rats by cold exposure and immune extract

　　Wistar rats weighing 200-250g, 6-8 months old. Before modeling, the rats adapt to the environment for 1 week. The hind limbs of the rats were depilated, and the immune extracts from vasculitis (TAO) patients were subcutaneously injected around the hind limb arteries of the rats, each time 0.1ml, after subcutaneous injection, skin hills were formed, once every 4 days. The hind limbs of the rats injected with the TAO immune extract for the first time were placed in a mixture of ice and water at 0°C, and the freezing time was increased from 15, 20, and 30 minutes. After each rat adapted to this temperature, the experiment was 30 minutes/d for a total of 2 months. Pathological specimen preparation and detection indicators are the same as the rat model of vasculitis obliterans with immune extract and tobacco.

　　3) A model of thromboangitis obliterans in rats by immune extract and sexial hormone (A model of thromboangitis obliterans in rats by immune extract and sexial hormone)

　　Wistar rats weighing 200-250g, 6-8 months old. Before modeling, the rats adapt to the environment for 1 week. The hind limbs of the rats were depilated, and the immune extracts from vasculitis (TAO) patients were subcutaneously injected around the hind limb arteries of the rats, each time 0.1ml, after subcutaneous injection, skin hills were formed, once every 4 days. The rats that received the first injection of TAO immune extract were injected intramuscularly with testosterone propionate 1 mg/kg, once a day, to promote their mating with sterilized female rats. The modeling process was 2 months in total. In the first week of modeling, there were no significant changes in the model rats. In the second week, the skin color of the extremities of the model rats was flushed, and there were no late symptoms such as toe ulceration and gangrene during the entire modeling process. Pathological specimen preparation and detection indicators are the same as the rat model of vasculitis obliterans with immune extract and tobacco.

　　4) A immune model of thromboangitis obliterans (A immune model of thromboangitis obliterans)

　　Weighing 200-250g, 6-8 months old male Wistar rat. The hind limbs of the rats were depilated, and the immune extracts from vasculitis (TAO) patients were subcutaneously injected around the hind limb arteries of the rats, each time 0.1ml, after subcutaneous injection, skin hills were formed, once every 4 days. The immune extract was mixed with incomplete Freund's adjuvant (1:1) 0.1ml, and injected subcutaneously into the inner side of the rat limb. The test cycle, pathological specimen preparation and detection indicators are the same as the rat model of vasculitis obliterans caused by immune extract and tobacco.

　　5) A model of thromboangitis obliterans in rats by tobacoo, cold exposure, sexy hormone and immunity

　　Weighing 200-250g, 6-8 months old male Wistar rat. The hind limbs of the rats were depilated, and the immune extracts from vasculitis (TAO) patients were subcutaneously injected around the hind limb arteries of the rats, each time 0.1ml, after subcutaneous injection, skin hills were formed, once every 4 days. Then give (1), (2), (3), (4) all the operations of the various factors in the modeling method, and the modeling process totals 2 months. Pathological specimen preparation and detection indicators are the same as the rat model of vasculitis obliterans with immune extract and tobacco.

　　(2) Model characteristics Among the above five modeling methods, the tobacco plus winter plus sex hormone plus immune comprehensive model has the highest success rate, the most obvious staging of the disease, and the shortest cycle; the immune extract plus sex hormone has the lowest success rate and the least disease , The stage of disease progression is not obvious in the short term. The immune extract plus tobacco rat model of vasculitis obliterans has a higher success rate for early TAO inflammation symptoms, but a lower success rate for late symptoms. The immune extract plus chilled rat vasculitis obliterans model is simpler and shorter than the model in which the necrotic tissue plasma of TAO patients is injected as the antigen to immunize the animal; but it is not as good as the immune extract plus tobacco model rat model Time and quantification are accurate.

　　(3) Comparative medicine Thromboangiitis obliterans is an autoimmune peripheral vascular disease. Smoking, cold and other external factors and differences in sex hormones are the main causes of this disease. The above-mentioned model animals created for its etiology all showed the corresponding signs of TAO, with naked eyes flushing, swelling, and late toe gangrene and toe ulceration. Pathological examination and scanning of the middle and small arteries and veins of the hind limbs of the model rats The pathological specimens taken by electron microscopy were basically the same as those taken from amputations of confirmed TAO patients. There are different degrees of intimal hyperplasia, fresh or muscularized thrombus, intact and thickened inner elastic membrane, and different degrees of vascular inflammation; hemorheological examination also showed that the various indicators of the model rats were significantly increased, which is inconsistent with the clinical The symptoms, signs, and pathological changes of TAO patients are consistent with the TAO syndrome differentiation type that has been clinically summarized for many years. In the vasculitis obliterans model caused by the above five methods, the success rate of male mice is significantly higher than that of female mice, which is the same as the clinical incidence.