WO2014202022A1 - Method for preparing rhesus macaque diabetes mellitus type 2 model - Google Patents
Method for preparing rhesus macaque diabetes mellitus type 2 model Download PDFInfo
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- WO2014202022A1 WO2014202022A1 PCT/CN2014/080412 CN2014080412W WO2014202022A1 WO 2014202022 A1 WO2014202022 A1 WO 2014202022A1 CN 2014080412 W CN2014080412 W CN 2014080412W WO 2014202022 A1 WO2014202022 A1 WO 2014202022A1
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- 241000282560 Macaca mulatta Species 0.000 title claims abstract description 37
- 238000000034 method Methods 0.000 title claims abstract description 24
- 201000000083 maturity-onset diabetes of the young type 1 Diseases 0.000 title abstract 3
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 claims abstract description 24
- 241000282693 Cercopithecidae Species 0.000 claims abstract description 20
- ZSJLQEPLLKMAKR-GKHCUFPYSA-N streptozocin Chemical compound O=NN(C)C(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O ZSJLQEPLLKMAKR-GKHCUFPYSA-N 0.000 claims abstract description 18
- 241001465754 Metazoa Species 0.000 claims abstract description 16
- ZSJLQEPLLKMAKR-UHFFFAOYSA-N Streptozotocin Natural products O=NN(C)C(=O)NC1C(O)OC(CO)C(O)C1O ZSJLQEPLLKMAKR-UHFFFAOYSA-N 0.000 claims abstract description 12
- 235000012000 cholesterol Nutrition 0.000 claims abstract description 12
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- 238000010253 intravenous injection Methods 0.000 claims abstract description 5
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- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 27
- 239000008103 glucose Substances 0.000 description 27
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- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 6
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- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- 210000000227 basophil cell of anterior lobe of hypophysis Anatomy 0.000 description 3
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- YQEZLKZALYSWHR-UHFFFAOYSA-N Ketamine Chemical compound C=1C=CC=C(Cl)C=1C1(NC)CCCCC1=O YQEZLKZALYSWHR-UHFFFAOYSA-N 0.000 description 1
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- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
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- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
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- PBGKTOXHQIOBKM-FHFVDXKLSA-N insulin (human) Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@H]1CSSC[C@H]2C(=O)N[C@H](C(=O)N[C@@H](CO)C(=O)N[C@H](C(=O)N[C@H](C(N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC=3C=CC(O)=CC=3)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC=3C=CC(O)=CC=3)C(=O)N[C@@H](CSSC[C@H](NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=3C=CC(O)=CC=3)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=3NC=NC=3)NC(=O)[C@H](CO)NC(=O)CNC1=O)C(=O)NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O)=O)CSSC[C@@H](C(N2)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](NC(=O)CN)[C@@H](C)CC)[C@@H](C)CC)[C@@H](C)O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)CC=1C=CC=CC=1)C(C)C)C1=CN=CN1 PBGKTOXHQIOBKM-FHFVDXKLSA-N 0.000 description 1
- PDJAZCSYYQODQF-UHFFFAOYSA-N iodine monofluoride Chemical compound IF PDJAZCSYYQODQF-UHFFFAOYSA-N 0.000 description 1
- 229960003299 ketamine Drugs 0.000 description 1
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- 150000002632 lipids Chemical class 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/027—New or modified breeds of vertebrates
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/02—Breeding vertebrates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
- A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2207/00—Modified animals
- A01K2207/25—Animals on a special diet
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2227/00—Animals characterised by species
- A01K2227/10—Mammal
- A01K2227/106—Primate
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2267/00—Animals characterised by purpose
- A01K2267/03—Animal model, e.g. for test or diseases
- A01K2267/035—Animal model for multifactorial diseases
- A01K2267/0362—Animal model for lipid/glucose metabolism, e.g. obesity, type-2 diabetes
Definitions
- the invention relates to a method for preparing a rhesus monkey type 2 diabetes model.
- T2DM Type 2 Diabetes Mellitus
- T2DM The pathogenesis of T2DM has many influencing factors, the process is long, and the individual responses are significantly different.
- T2DM models with insulin resistance have been successfully established through genetic screening, genetic modification and diet induction, but more and more studies have shown that small animals have huge fat and inflammatory responses. Differences, can not well simulate the pathophysiological characteristics of human metabolic diseases.
- the establishment of large animals, especially the highly anthropomorphic non-human primate T2DM model, is of great significance for the clinical transformation of T2DM research.
- the present invention provides a method for preparing a rhesus monkey type 2 diabetes model.
- the preparation method of the rhesus monkey type 2 diabetes model of the present invention comprises the following steps:
- the feed contains the following weight ratio raw materials: 78 standard monkey feeds. 15 parts of animal fats, 5 parts of sugar, 2 parts of cholesterol;
- the animal fat is lard; the sugar is sucrose.
- the standard monkey feed refers to the monkey compound feed specified in the National Standard of the Chinese Republic of China GB14924.8-2001.
- Animal fat refers to fat derived from animals, such as butter, sheep oil, and lard.
- the administration method is intravenous injection.
- the present invention establishes a feed for the Ganger type 2 diabetes model, which comprises the following raw materials by weight ratio: 78 parts of standard monkey feed, 15 parts of animal fat, 5 parts of sugar, and 2 parts of cholesterol.
- the animal fat is lard; the sugar is sucrose.
- the present invention also provides a rhesus monkey type 2 diabetes model prepared by the aforementioned method and its use in screening for a drug for treating a type 2 diabetes model.
- the present invention screens a method for treating a drug of a type 2 diabetes model, which comprises the following steps: a. Establishing a rhesus monkey type 2 diabetes model according to the aforementioned method;
- the modeling method of the present invention can induce hyperinsulinemia, impaired glucose tolerance and insulin resistance in rhesus monkeys, and the clinical manifestations of type 2 diabetes are simple, and the method of administration is simple and reproducible.
- Figure 1 Rhesus monkey weight abdominal circumference test
- FIG. 3 HE staining of liver tissue of rhesus monkeys, A. HE staining of liver in control group; B. HE staining of liver in experimental group (arrow indicates inflammatory cell infiltration); C. Liver HE staining in experimental group (arrow indicates hepatic cell edema) ;
- Control group monkey standard monkey feed
- Feeding method Feed 2 times a day, feeding 0.3-0.413 ⁇ 4/time*.
- Control group Monkey standard monkey feed - Commercialized monkey pellet feed (purchased from the Animal Research Institute of Sichuan Medical Science Institute, the formula is the same as the monkey compound feed specified in the National Standard of the Chinese Republic of China GB14924.8-2001).
- High fat and high cholesterol feed 2% (w/w) Edible cholesterol (Anhui Tianqi Chemical Technology Co., Ltd.), 15% (w/w) refined lard (commercially available), 5% (w/w) sucrose (commercially available), 78% (w/w) monkey standard monkey feed.
- Streptozotocin (STZ) purchased from Chengdu Yuyang High-Tech Development Co., Ltd.
- Solution A (0.1 M citric acid solution): Weigh 2.1 g of citric acid (C6H807-H20 FW: 210.14) dissolved in 100 ml of double distilled water.
- Solution B (0.1 M sodium citrate solution): 2.94 g of sodium citrate (C6H5Na307-2H20 FW: 294.10) was weighed and dissolved in 100 ml of double distilled water.
- Streptozotocin solution STZ is weighed according to animal weight ratio, and STZ is dissolved at 1% concentration with the above AB mixed working solution. The solution is required to be stored and stored at 4 ° C. The injection is completed within 30 minutes. .
- Blood glucose detector Luo Kang full vitality blood glucose detector, Luo Kang full vitality blood glucose test strips Other tests are completed in the 4th Division of the University of West China Hospital Laboratory.
- the experimental group (including STZ group and HF group) was given a rouge high cholesterol diet, and the control group was fed with regular monkey feed, fed twice a day, and the feeding amount was 0.3-0.413 ⁇ 4/time* only;
- the STZ group After giving a rouge high-cholesterol diet for 24 months, the STZ group received a small dose of STZ intravenously: 25 mg/kg each time, once a week for 2 consecutive times. The HF group and the control group were left untreated.
- IVGTT intravenous glucose tolerance test
- ketamine 50 mg/ml was intramuscularly injected at 15 mg/Kg ;
- the upper and lower limbs of the rhesus monkey are fixed on the operating table, and the saphenous vein of the lower leg of the lower leg is exposed to the saphenous vein, and the iodine fluoride is disinfected;
- Rhesus monkeys were fasted for more than 16 hours, routine anesthesia, weighing, and collecting 4 ml of lower extremity venous blood into a separate gel-promoting tube.
- the venous cannula was placed from the median vein of the elbow and the two veins were used to construct two venous channels for infusion of insulin and glucose.
- Infusion of human insulin solution (Uberin, 40 U/mL; > at a rate of 4 mU / (kg ⁇ min) within 10 min of the clamp starts to increase the blood insulin concentration rapidly, followed by 2 mU / (kg * within 11 min) Mm) Continuous infusion of the rate and simultaneous input of 20% glucose solution with an infusion pump.
- the venous blood glucose level was measured every 5 minutes, and the infusion rate of 200 mL/L glucose solution was adjusted according to the blood glucose level, so that the blood glucose level of the subject was maintained at about 4.5 mmol/L.
- Venous blood was measured every 30 min to determine serum insulin concentration.
- High-fat diet can induce glucose metabolism and lipid metabolism disorder in rhesus monkeys
- SIciamp (mg ⁇ kg - 1 ⁇ min - 1 / ⁇ / L) 0.12 ⁇ 0.05 0.07 ⁇ 0.03
- the high-fat and high-cholesterol diet caused fat ectopic viscera, and obvious lipid droplet deposition appeared in the liver and skeletal muscle of rhesus monkeys in the experimental group.
- the experimental group showed obvious diet-dependent hypercholesterolemia. Sustained hypercholesterolemia triggered compensatory changes in the corresponding serum lipid metabolism parameters, which involved accelerated cholesterol transport (lipoprotein HDL, elevated LDL). , increased conversion and excretion, increased fat breakdown and elevated triglycerides.
- the control group was normal.
- the fasting blood glucose of the experimental group increased gradually after 6 months of feeding, while the serum insulin level increased progressively, and HbAlC was significantly higher than the control group.
- HbAlC was significantly higher than the control group.
- the area under the glucose curve increased, the glucose clearance rate (Kglu) and the area under the insulin curve decreased.
- the M value and SIciamp of the clamp experiment were lower than those of the control group, and the insulin resistance index increased.
- the experimental results showed that the islet ⁇ -cell function of the rhesus monkey in the experimental group was impaired, the glucose tolerance was impaired, and the exogenous and endogenous insulin sensitivity of the peripheral tissues decreased.
- the fasting insulin level in the STZ group, the area under the blood glucose curve of the IVGTT, and the insulin resistance index (HOMA-I) were significantly higher than those in the other two groups, demonstrating hyperinsulinemia, impaired glucose tolerance, and insulin resistance.
- the modeling method of the present invention based on high-fat and high-cholesterol feed feeding, is supplemented by a small dose of STZ, which causes partial ⁇ cell cell damage, satisfies the necessary conditions for type 2 diabetes, and induces insulin resistance through lipid metabolism disorder.
- STZ partial ⁇ cell cell damage
- the formation of a stable and standardized rhesus monkey T2DM model in a relatively short period has important practical application value.
- the modeling method of the invention forms a relatively stable and standardized rhesus monkey T2DM model in a relatively short period, and can be used for screening for type 2 diabetes drugs, which is suitable for industrial applications.
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Abstract
A method for preparing a Rhesus macaque diabetes mellitus type 2 model, comprising the following steps: (1) a Rhesus macaque is fed a high-fat and high-cholesterol feed twice per day at a feeding amount of 0.3-0.4 kg/instance/animal, where the feed comprises raw materials of the following proportions by weight: 78 parts of a standard monkey feed, 15 parts of an animal fat, 5 parts of a sugar, and 2 parts of cholesterol; and, (2) after 24 months, streptozotocin of a dosage of 25 mg/kg is administered to the Rhesus macaque once per week for 2 successive instances. By employing a scheme of long-term feeding of the high-fat and high-cholesterol feed and intravenous injection of a small dose of streptozotocin for joint modeling, a Rhesus macaque diabetes mellitus type 2 model is successfully induced.
Description
一种恒河猴 2型糖尿病模型的制备方法 技术领域 Method for preparing rhesus monkey type 2 diabetes model
本发明涉及恒河猴 2型糖尿病模型的制备方法。 The invention relates to a method for preparing a rhesus monkey type 2 diabetes model.
背景技术 Background technique
近年来, 全球糖尿病发病率增长迅速。 WHO 最新公布的数据显示, 全 球糖尿病患者人数已经超过 1.77亿, 预计到 2025年将达到 3.7 亿。 我国糖 尿病发病率也正以惊人的速度增加, 中华医学会糖尿病分会新近的流行病学 调査发现, 全国糖尿病发病率已经从 2002 年的 4%迅速提高到了现在的 11%, 此外还有 15%的人血糖异常。 我国目前糖尿病患者数量已经达到了 9100 万, 列居世界第一。 在糖尿病患者中, 90%以上为 2型糖尿病 (Type2 Diabetes Mellitus, T2DM) ,不断探索 T2MD治疗新模式及开发新型药物具有 重要的临床意义和社会价值。 In recent years, the global incidence of diabetes has grown rapidly. According to the latest data from WHO, the number of people with diabetes worldwide has exceeded 177 million, and it is expected to reach 370 million by 2025. The incidence of diabetes in China is also increasing at an alarming rate. The recent epidemiological survey of the Diabetes Branch of the Chinese Medical Association found that the national incidence of diabetes has rapidly increased from 4% in 2002 to 11% now, and 15% in addition. People have abnormal blood sugar. The number of people with diabetes in China has reached 91 million, ranking first in the world. More than 90% of patients with diabetes mellitus are type 2 Diabetes Mellitus (T2DM). It is of great clinical and social value to continuously explore new modes of T2MD treatment and develop new drugs.
T2DM 的发病过程影响因素繁多, 过程漫长, 个体反应差异明显。 在啮 齿类动物, 通过遗传筛选、 基因改造和饮食诱导已经成功建成了具有胰岛素 抵抗特征的 T2DM模型, 但越来越多的研究证明, 小动物在脂代谢、炎性反 应等方面和人体存在巨大差异, 不能很好模拟人体代谢性疾病的病理生理特 征。建立大动物,特别是高度拟人的非人灵长类 T2DM模型对 T2DM研究的 临床转化具有重要意义。 The pathogenesis of T2DM has many influencing factors, the process is long, and the individual responses are significantly different. In rodents, T2DM models with insulin resistance have been successfully established through genetic screening, genetic modification and diet induction, but more and more studies have shown that small animals have huge fat and inflammatory responses. Differences, can not well simulate the pathophysiological characteristics of human metabolic diseases. The establishment of large animals, especially the highly anthropomorphic non-human primate T2DM model, is of great significance for the clinical transformation of T2DM research.
但建立大动物 T2DM模型, 制定标准的诱导方案和评价标准具有相当 的难度。 灵长类动物中筛选自发性糖尿病动物几率很小, 数量稀少, 而转基 因技术在灵长类动物还没有成熟的技术, 而且成本高昂。 However, it is quite difficult to establish a large animal T2DM model and to develop standard induction programs and evaluation criteria. Screening for spontaneously diabetic animals in primates is rare and rare, and transgenic technology has no mature technology in primates and is costly.
发明内容 Summary of the invention
为了解决上述问题, 本发明提供了一种恒河猴 2型糖尿病模型的制备方 法。 In order to solve the above problems, the present invention provides a method for preparing a rhesus monkey type 2 diabetes model.
本发明恒河猴 2型糖尿病模型的制备方法, 包括如下歩骤: The preparation method of the rhesus monkey type 2 diabetes model of the present invention comprises the following steps:
( 1 )向恒河猴饲喂高脂高胆固醇饲料,每天饲喂 2次,饲喂量为 0.3-0.4kg/ 次 ·只,所述饲料包含如下重量配比的原料:标准猴饲料 78份、动物油脂 15 份、 糖 5份、 胆固醇 2份; (1) Feeding the high-heavy and high-cholesterol feed to the rhesus monkeys, feeding twice a day, feeding 0.3-0.4kg/time. The feed contains the following weight ratio raw materials: 78 standard monkey feeds. 15 parts of animal fats, 5 parts of sugar, 2 parts of cholesterol;
(2) 24个月后, 将剂量为 25mg/Kg的链脲菌素施用于猴, 每周施用一 次, 连续 2次。 (2) After 24 months, streptozotocin at a dose of 25 mg/kg was administered to the monkey once a week for 2 consecutive times.
优选地, 所述动物油脂是猪油; 所述糖是蔗糖。 Preferably, the animal fat is lard; the sugar is sucrose.
标准猴饲料, 是指《中华人名共和国国家标准 GB14924.8-2001》规定的 猴配合饲料。
动物油脂, 是指来源于动物的脂肪, 如, 牛油、 羊油、 猪油。 The standard monkey feed refers to the monkey compound feed specified in the National Standard of the Chinese Republic of China GB14924.8-2001. Animal fat refers to fat derived from animals, such as butter, sheep oil, and lard.
歩骤 (2 ) 中, 所述的施用方式是静脉注射。 In the step (2), the administration method is intravenous injection.
本发明建立恒河 2型糖尿病模型的饲料, 它包含如下重量配比的原料: 标准猴饲料 78份、 动物油脂 15份、 糖 5份、 胆固醇 2份。 The present invention establishes a feed for the Ganger type 2 diabetes model, which comprises the following raw materials by weight ratio: 78 parts of standard monkey feed, 15 parts of animal fat, 5 parts of sugar, and 2 parts of cholesterol.
优选地, 所述动物油脂是猪油; 所述糖是蔗糖。 Preferably, the animal fat is lard; the sugar is sucrose.
本发明还提供了前述方法制备得到的恒河猴 2型糖尿病模型及其在筛选 治疗 2型糖尿病模型的药物中的用途。 The present invention also provides a rhesus monkey type 2 diabetes model prepared by the aforementioned method and its use in screening for a drug for treating a type 2 diabetes model.
本发明筛选治疗 2型糖尿病模型的药物的方法, 它包括如下歩骤: a、 按照前述方法, 建立恒河猴 2型糖尿病模型; The present invention screens a method for treating a drug of a type 2 diabetes model, which comprises the following steps: a. Establishing a rhesus monkey type 2 diabetes model according to the aforementioned method;
b、 将候选药物施用于动物模型; b. administering the candidate drug to the animal model;
c、 用动物模型评价潜在的治疗 2型糖尿病的药物。 c. Use animal models to evaluate potential drugs for the treatment of type 2 diabetes.
本发明造模方法可以诱导恒河猴出现高胰岛素血症、 糖耐量受损和胰岛 素抵抗, 出现 2型糖尿病的临床表现, 给药方法简单, 可重复性强。 The modeling method of the present invention can induce hyperinsulinemia, impaired glucose tolerance and insulin resistance in rhesus monkeys, and the clinical manifestations of type 2 diabetes are simple, and the method of administration is simple and reproducible.
显然,根据本发明的上述内容,按照本领域的普通技术知识和惯用手段, 在不脱离本发明上述基本技术思想前提下,还可以做出其它多种形式的修改、 替换或变更。 It is apparent that various other modifications, substitutions and changes can be made in the form of the above-described embodiments of the present invention without departing from the spirit and scope of the invention.
以下通过实施例形式的具体实施方式, 对本发明的上述内容再作进一歩 的详细说明。 但不应将此理解为本发明上述主题的范围仅限于以下的实例。 凡基于本发明上述内容所实现的技术均属于本发明的范围。 The above description of the present invention will be further described in detail by way of specific embodiments of the embodiments. However, the scope of the above-mentioned subject matter of the present invention should not be construed as being limited to the following examples. Any technique implemented based on the above description of the present invention is within the scope of the present invention.
附图说明 DRAWINGS
图 1 恒河猴体重腹围检测; Figure 1 Rhesus monkey weight abdominal circumference test;
图 2 恒河猴肝脏、 肌肉组织油红 0染色; Figure 2 Rhesus liver and muscle tissue oil red 0 staining;
图 3 恒河猴肝脏组织 HE染色, A. 对照组肝脏 HE染色; B. 实验组 肝脏 HE染色 (箭头示炎性细胞浸润); C. 实验组肝脏 HE染色 (箭头示肝 细胞轻度水肿); Figure 3 HE staining of liver tissue of rhesus monkeys, A. HE staining of liver in control group; B. HE staining of liver in experimental group (arrow indicates inflammatory cell infiltration); C. Liver HE staining in experimental group (arrow indicates hepatic cell edema) ;
图 4 恒河猴脂代谢指标检测; Figure 4 Detection of lipid metabolism indicators in rhesus monkeys;
图 5 恒河猴糖代谢相关指标; Figure 5 Rhesus monkey sugar metabolism related indicators;
图 6 高脂饲养 18m后 IVGTT和高胰岛素血 -正常血糖钳夹实验结果; 图 7 胰岛功能损伤相关指标。 Figure 6 IVGTT and high insulin blood-normal blood glucose clamp test results after 18m of high-fat feeding; Figure 7 Related indicators of islet function damage.
具体实施方式 detailed description
实施例 1 本发明恒河猴 2型糖尿病征模型的制备 Example 1 Preparation of Rhesus Monkey Type 2 Diabetes Model of the Invention
1实验材料和仪器 1 experimental materials and instruments
1.1实验材料 1.1 Experimental materials
6月龄恒河猴 8只(雌雄各半), 无 B疱疹病毒、 猴逆转录病毒、 猴白血 病病毒和免疫缺陷病毒感染, 购自成都平安动物繁育基地。
实验组: STZ组 (HF+STZ组, 4只) 给予髙脂高胆固醇饮食 24月后通 过静脉注射小剂量链脲菌素诱导 2型糖尿病, 高脂组(HF组, 4只)仅给予 高胆固醇饮食喂养。 Eight 6-month-old rhesus monkeys (male and female), no B virus, monkey retrovirus, monkey leukemia virus and immunodeficiency virus infection, purchased from Chengdu Pingan Animal Breeding Base. Experimental group: STZ group (HF+STZ group, 4 rats) After giving a rouge high cholesterol diet 24 months after intravenous injection of small doses of streptozotocin induced type 2 diabetes, high fat group (HF group, 4) only given high Cholesterol diet feeding.
对照组: 给予猴标准猴饲料 Control group: monkey standard monkey feed
饲喂方式: 每天饲喂 2次, 饲喂量为 0.3-0.41¾/次*只。 Feeding method: Feed 2 times a day, feeding 0.3-0.413⁄4/time*.
饲料配方: Feed formula:
对照组: 猴标准猴饲料一一商业化的猴颗粒饲料 (购自四川省医学科学 院动物研究所, 配方与《中华人名共和国国家标准 GB14924.8-2001》规定的 猴配合饲料相同)。 Control group: Monkey standard monkey feed - Commercialized monkey pellet feed (purchased from the Animal Research Institute of Sichuan Medical Science Institute, the formula is the same as the monkey compound feed specified in the National Standard of the Chinese Republic of China GB14924.8-2001).
实验组: 高脂高胆固醇饲料: 2% (w/w) 食用胆固醇 (安徽天启化工 科技有限公司), 15% (w/w) 精炼猪油 (市售), 5% (w/w) 蔗糖 (市售), 78% (w/w) 猴标准猴饲料。 Experimental group: High fat and high cholesterol feed: 2% (w/w) Edible cholesterol (Anhui Tianqi Chemical Technology Co., Ltd.), 15% (w/w) refined lard (commercially available), 5% (w/w) sucrose (commercially available), 78% (w/w) monkey standard monkey feed.
1.2 主要试剂 1.2 main reagents
链脲菌素 (STZ): 购自成都宇洋高科技发展有限公司 Streptozotocin (STZ): purchased from Chengdu Yuyang High-Tech Development Co., Ltd.
1.3 主要溶液的配制 1.3 Preparation of the main solution
柠檬酸钠缓冲液配制: Sodium citrate buffer preparation:
A液 (0.1M枸橼酸溶液): 称取 2.1g枸橼酸 (C6H807-H20 FW: 210.14 ) 溶于 100ml双蒸水中。 B液 (0.1M枸橼酸钠溶液): 称取 2.94g枸橼酸钠 (C6H5Na307-2H20 FW: 294.10 ) 溶于 100ml双蒸水中。 Solution A (0.1 M citric acid solution): Weigh 2.1 g of citric acid (C6H807-H20 FW: 210.14) dissolved in 100 ml of double distilled water. Solution B (0.1 M sodium citrate solution): 2.94 g of sodium citrate (C6H5Na307-2H20 FW: 294.10) was weighed and dissolved in 100 ml of double distilled water.
工作液: 用时取 28 mlA液加 22 mlB液 (按照 1.32: 1的比例) ,双蒸水 稀释到 100ml,调 PH=4.5。 Working solution: Take 28 ml of A solution and 22 ml of B solution (according to the ratio of 1.32:1), dilute to 100 ml with double distilled water, adjust pH=4.5.
链脲菌素溶液: 按动物重量比称取 STZ, 用上述 AB混合工作液以 1% 的浓度溶解 STZ, 随用随配, 该液要求 4°C配置、 保存, 尽量在 30分钟内注 射完毕。 Streptozotocin solution: STZ is weighed according to animal weight ratio, and STZ is dissolved at 1% concentration with the above AB mixed working solution. The solution is required to be stored and stored at 4 ° C. The injection is completed within 30 minutes. .
1.4 主要仪器 1.4 Main instruments
血糖检测仪: 罗康全活力型血糖检测仪, 罗康全活力型血糖试纸 其他检测均在四 ) 11大学华西医院检验科完成。 Blood glucose detector: Luo Kang full vitality blood glucose detector, Luo Kang full vitality blood glucose test strips Other tests are completed in the 4th Division of the University of West China Hospital Laboratory.
2 实验方法 2 Experimental methods
2. 1诱导造糖尿病模型 2. 1 induced diabetes model
实验组 (包括 STZ组和 HF组) 给予髙脂高胆固醇饮食喂养, 对照组给 予常规猴饲料喂养, 每天饲喂 2次, 饲喂量为 0.3-0.41¾/次*只; The experimental group (including STZ group and HF group) was given a rouge high cholesterol diet, and the control group was fed with regular monkey feed, fed twice a day, and the feeding amount was 0.3-0.413⁄4/time* only;
给予髙脂高胆固醇饮食 24月后, STZ组静脉给予小剂量 STZ: 每次 25 mg/Kg, 每周一次, 连续 2次, HF组和对照组不作处理。 After giving a rouge high-cholesterol diet for 24 months, the STZ group received a small dose of STZ intravenously: 25 mg/kg each time, once a week for 2 consecutive times. The HF group and the control group were left untreated.
2. 2 恒河猴静脉葡萄糖耐受实验 2. 2 Rhesus monkey intravenous glucose tolerance test
通过静脉葡萄糖耐受实验(The intravenous glucose tolerance test, IVGTT )
测试动物胰腺功能, 了解血糖波动范围,排除动物患有自发性糖尿病的可能。 动物禁食 10-12h, 不禁水; Through the intravenous glucose tolerance test (IVGTT) Test animal pancreatic function, understand the range of blood glucose fluctuations, and rule out the possibility of animals having spontaneous diabetes. Animals fasted for 10-12h, can not help but water;
根据动物购买时体重, 按 15mg/Kg肌肉注射氯氨酮 (50 mg/ml); According to the body weight at the time of purchase, ketamine (50 mg/ml) was intramuscularly injected at 15 mg/Kg ;
待动物麻醉后, 准确称量体重, 并记录; After the animals are anesthetized, the weight is accurately weighed and recorded;
将恒河猴上下肢固定于手术台上, 下肢小腿后侧备皮暴露大隐静脉, 碘 氟消毒; The upper and lower limbs of the rhesus monkey are fixed on the operating table, and the saphenous vein of the lower leg of the lower leg is exposed to the saphenous vein, and the iodine fluoride is disinfected;
20G留滞针穿剌大隐静脉, 5ml注射器抽血 6ml (分别做生化、 血常规、 血糖和空腹胰岛素, 并标记为 0分钟数值); 20G residual needle through the great saphenous vein, 5ml syringe blood 6ml (respectively biochemical, blood routine, blood glucose and fasting insulin, and marked as 0 minutes);
立即用 5ml注射器静脉推注 50%葡萄糖高渗溶液 0.5g/Kg (在 30sec内推 完),并开始记时,再注射生理盐水 5ml, 向留滞针内推肝素(250 U/ml) lml; 分别于记时后的第 1, 3, 5, 10, 30min时采血 1.5ml, 放入红头采血管, 测定胰岛素水平; 同时分别于第 0, 1, 3, 5, 10, 30, 60, 120 min时测定 血糖; Immediately use a 5ml syringe to inject a 50% glucose hypertonic solution 0.5g / Kg (within 30 seconds), and start recording, then inject 5ml of normal saline, push the heparin (250 U / ml) into the stagnation needle lml 1.5 ml of blood was collected at the 1st, 3rd, 5th, 10th, and 30th minutes after the time of recording, and placed in the red blood collection tube to measure the insulin level; and at 0, 1, 3, 5, 10, 30, 60, respectively. Blood glucose was measured at 120 min;
检测结束, 将动物放回饲养笼中, 观察动物状况至苏醒。 At the end of the test, the animals were placed back in the cage and the animals were observed to awaken.
2.3 高胰岛素 -正常血糖钳夹试验 2.3 high insulin - normal blood glucose clamp test
恒河猴空腹 16h以上, 常规麻醉, 称重、 采集下肢静脉血 4ml注入分离 胶促凝管。 Rhesus monkeys were fasted for more than 16 hours, routine anesthesia, weighing, and collecting 4 ml of lower extremity venous blood into a separate gel-promoting tube.
取平卧位, 逆心方向行一侧手臂静脉穿剌, 置入留置导管, 以生理盐水 维持通道,以备取血测定血糖。 不采血时缓慢静滴生理盐水, 采血前临时关闭 输液器。 通过电热垫加热以维持手的温度在 50〜60°C。 Take the supine position, put the arm vein in the opposite direction of the heart, put in the indwelling catheter, and maintain the channel with normal saline to prepare blood for blood sugar measurement. Slowly instill saline without blood collection, and temporarily close the infusion set before blood collection. Heat the pad by heating to maintain the temperature of the hand at 50 to 60 °C.
从另一侧肘正中静脉置入静脉插管利用三通管组成 2条静脉通道以备 输注胰岛素和葡萄糖。 钳夹开始 10 min内以 4 mU / ( kg · min)速率输注人 胰岛素溶液(优泌林, 40U/mL;>, 使血液胰岛素浓度迅速升高, 随后 llOmin 内以 2 mU/ ( kg * mm)速率持续输注, 并同时用输液泵输入 20% 葡萄糖液。 The venous cannula was placed from the median vein of the elbow and the two veins were used to construct two venous channels for infusion of insulin and glucose. Infusion of human insulin solution (Uberin, 40 U/mL; > at a rate of 4 mU / (kg · min) within 10 min of the clamp starts to increase the blood insulin concentration rapidly, followed by 2 mU / (kg * within 11 min) Mm) Continuous infusion of the rate and simultaneous input of 20% glucose solution with an infusion pump.
在此期间每 5 min测一次静脉血糖值, 根据血糖值调整 200 mL/L萄糖 液输注率, 使受试者血糖值维持在 4.5mmol/L左右。每 30 min采静脉血测定 血清胰岛素浓度。 During this period, the venous blood glucose level was measured every 5 minutes, and the infusion rate of 200 mL/L glucose solution was adjusted according to the blood glucose level, so that the blood glucose level of the subject was maintained at about 4.5 mmol/L. Venous blood was measured every 30 min to determine serum insulin concentration.
当血糖水平变化小于 10% 时, 视为达到稳态, 稳态持续约 60 min后结 束试验。 检测胰岛素、 C肽的血标本离心分离血清后置 - 20°C冰箱保存, 统 根据 Steel's 公式计算钳夹试验过程中外周组织葡萄糖代谢率 (glucose disposal rate, M), 计算公式: M=钳夹试验最后 40 min每分钟每公斤体重的 葡萄糖输注量 +(G80— G120) X 0.0625, 其中 G80和 G120分别为钳夹试验 80min和 120 min时的血糖浓度 (mg I dl)。 根据 M值计算胰岛素敏感指数 SIClamp = M/ Δ I / fat-free mass (Kg), M是稳态时的葡萄糖输注率, Δ Ι 是稳
态时胰岛素水平与实验前的基础胰岛素水平的差值 0iIU/ml)。 When the blood glucose level changes by less than 10%, it is considered to reach a steady state, and the steady state lasts for about 60 minutes and the test is terminated. The blood samples of insulin and C-peptide were centrifuged and stored in a refrigerator at 20 ° C. The glucose disposal rate (M) of the peripheral tissues during the clamp test was calculated according to Steel's formula. The formula is: M = clamp The glucose infusion per kilogram of body weight per minute for the last 40 min of the test + (G80 - G120) X 0.0625, where G80 and G120 were the blood glucose concentrations (mg I dl) at 80 min and 120 min, respectively, in the clamp test. Calculate the insulin sensitivity index SIClamp = M / Δ I / fat-free mass (Kg) according to the M value, M is the glucose infusion rate at steady state, Δ Ι is stable The difference between the insulin level at the time and the basal insulin level before the experiment was 0 iIU/ml).
3实验结果 3 experimental results
3.1高脂饮食可以诱导恒河猴糖代谢、 脂代谢紊乱 3.1 High-fat diet can induce glucose metabolism and lipid metabolism disorder in rhesus monkeys
实验结果如图 1~6和表 1所示: The experimental results are shown in Figures 1 to 6 and Table 1:
表 1 IVGTT和钳夹实验中 (18m;>实验组和对照组葡萄糖代谢变量的均值 Table 1 IVGTT and clamp experiment (18m;> mean value of glucose metabolism variables in experimental group and control group
Variables Control (n =8) HFHCD (n =8) Variables Control (n = 8) HFHCD (n = 8)
KGlu(%) 1.73±0.37 0.70±0.33*** K Glu (%) 1.73±0.37 0.70±0.33***
AUC-insulin((JU/ L* min _1) 387.24±122.39 105.50±66.04*** AUC-insulin((JU/ L* min _1 ) 387.24±122.39 105.50±66.04***
M value(mg« kg— 1 · min _1) 16.35±2.06 13.78±1.21 M value(mg« kg— 1 · min _1 ) 16.35±2.06 13.78±1.21
SIciamp (mg · kg - 1 · min - 1 /μΐυ/ L) 0.12±0.05 0.07±0.03 SIciamp (mg · kg - 1 · min - 1 /μΐυ / L) 0.12±0.05 0.07±0.03
HOMA-IR 0.85±0.55 1.55±1.12 HOMA-IR 0.85±0.55 1.55±1.12
Note: HFHCD vs. Control: *** P <0.001.HFD: 高脂高胆固醇词养组, control: 对照组 Note: HFHCD vs. Control: *** P <0.001.HFD: high fat and high cholesterol word group, control: control group
根据图 1~6和表 1可以看出, 对照组的糖代谢和脂代谢均正常, 实验组 的糖代谢和脂代谢则出现异常: According to Figures 1 to 6 and Table 1, it can be seen that the glucose metabolism and lipid metabolism of the control group were normal, and the glucose metabolism and lipid metabolism of the experimental group were abnormal:
1、高脂高胆固醇饮食使内脏发生脂肪异位,在实验组恒河猴的肝脏和骨 骼肌中出现明显的脂滴沉积。 1. The high-fat and high-cholesterol diet caused fat ectopic viscera, and obvious lipid droplet deposition appeared in the liver and skeletal muscle of rhesus monkeys in the experimental group.
2、实验组出现明显的饮食依赖的高胆固醇血症,持续的高胆固醇血症引 发了相应血清脂代谢参数的代偿性改变, 涉及到胆固醇的运输加速 (脂蛋白 HDL, LDL的升高)、 转化排泄增强、 脂肪的分解增加和甘油三酯的升高。 2. The experimental group showed obvious diet-dependent hypercholesterolemia. Sustained hypercholesterolemia triggered compensatory changes in the corresponding serum lipid metabolism parameters, which involved accelerated cholesterol transport (lipoprotein HDL, elevated LDL). , increased conversion and excretion, increased fat breakdown and elevated triglycerides.
3、对照组正常, 实验组空腹血糖自喂养 6个月后呈逐渐升高趋势, 而血 清胰岛素水平进行性增加, HbAlC明显高于对照组。高糖剌激下葡萄糖曲线 下面积升高, 葡萄糖清除率 (Kglu) 和胰岛素曲线下面积下降, 钳夹实验的 M值和 SIciamp低于对照组, 胰岛素抵抗指数增加。 3. The control group was normal. The fasting blood glucose of the experimental group increased gradually after 6 months of feeding, while the serum insulin level increased progressively, and HbAlC was significantly higher than the control group. Under the high glucose stress, the area under the glucose curve increased, the glucose clearance rate (Kglu) and the area under the insulin curve decreased. The M value and SIciamp of the clamp experiment were lower than those of the control group, and the insulin resistance index increased.
实验结果说明, 实验组恒河猴的胰岛 β细胞功能受损, 糖耐量受损, 外 周组织对外源性和内源性的胰岛素敏感性下降。 The experimental results showed that the islet β-cell function of the rhesus monkey in the experimental group was impaired, the glucose tolerance was impaired, and the exogenous and endogenous insulin sensitivity of the peripheral tissues decreased.
3.2小剂量 STZ可以诱导胰岛功能损伤, 加速恒河猴 2型糖尿病发生 静脉注射 STZ 1个月后, 恒河猴胰岛功能的各项指标如图 7所示: 与对 照组相比, STZ组恒河猴出现明显的血糖升高, 达到 2型糖尿病诊断标准; 3.2 Low-dose STZ can induce islet function damage, accelerate the rhesus monkey type 2 diabetes after intravenous injection of STZ 1 month, the rhesus monkey islet function indicators as shown in Figure 7: compared with the control group, STZ group constant Rhesus monkeys showed significant elevation of blood glucose and reached the diagnostic criteria for type 2 diabetes;
STZ组的空腹胰岛素水平、 IVGTT的血糖曲线下面积和胰岛素抵抗指数 (HOMA-I )都明显高于其他两组, 证明有高胰岛素血症、糖耐量受损和胰 岛素抵抗的发生。 The fasting insulin level in the STZ group, the area under the blood glucose curve of the IVGTT, and the insulin resistance index (HOMA-I) were significantly higher than those in the other two groups, demonstrating hyperinsulinemia, impaired glucose tolerance, and insulin resistance.
实验结果说明, 本发明方法造成部分 β 细胞细胞损伤, 诱导恒河猴高
胰岛素血症、 糖耐量受损和胰岛素抵抗, 出现 2型糖尿病的临床表现, 成功 建立了恒河猴 2型糖尿病模型。 实施例 2 用本发明模型筛选治疗 2型糖尿病的药物 The experimental results show that the method of the present invention causes partial β cell cell damage and induces rhesus monkeys to be high. Insulinemia, impaired glucose tolerance and insulin resistance, the clinical manifestations of type 2 diabetes, successfully established a rhesus monkey type 2 diabetes model. Example 2 Screening for the treatment of type 2 diabetes using the model of the present invention
a、 按照实施例 1方法建立的恒河猴 2型糖尿病模型; a. Rhesus type 2 diabetes model established according to the method of Example 1;
b、 将候选药物施用于动物模型; b. administering the candidate drug to the animal model;
c、观察候选药物对 2型糖尿病的各种指标的影响情况,评价潜在的治疗 2型糖尿病的药物。 综上, 本发明造模方法, 高脂高胆固醇饲料喂养的基础上辅以小剂量 STZ, 既造成部分 β 细胞细胞损伤, 满足 2型糖尿病的必要条件, 又能通 过脂代谢紊乱诱导胰岛素抵抗, 在相对短的周期内形成较稳定和规范的恒河 猴 T2DM模型, 具有重要的实际应用价值。 工业应用性 c. Observe the effects of drug candidates on various indicators of type 2 diabetes and evaluate potential drugs for the treatment of type 2 diabetes. In summary, the modeling method of the present invention, based on high-fat and high-cholesterol feed feeding, is supplemented by a small dose of STZ, which causes partial β cell cell damage, satisfies the necessary conditions for type 2 diabetes, and induces insulin resistance through lipid metabolism disorder. The formation of a stable and standardized rhesus monkey T2DM model in a relatively short period has important practical application value. Industrial applicability
本发明造模方法,在相对短的周期内形成较稳定和规范的恒河猴 T2DM 模型, 可用于 2型糖尿病药物的筛选, 适于工业应用。
The modeling method of the invention forms a relatively stable and standardized rhesus monkey T2DM model in a relatively short period, and can be used for screening for type 2 diabetes drugs, which is suitable for industrial applications.
Claims
1、一种恒河猴 2型糖尿病模型的制备方法,其特征在于:包括如下歩骤:1. A method for preparing a rhesus monkey type 2 diabetes model, which is characterized by: including the following steps:
( 1 )向恒河猴饲喂高脂高胆固醇饲料,每天饲喂 2次,饲喂量为 0.3-0.4kg/ 次 ·只,所述饲料包含如下重量配比的原料:标准猴饲料 78份、动物油脂 15 份、 糖 5份、 胆固醇 2份; (1) Feed high-fat and high-cholesterol feed to rhesus monkeys twice a day, and the feeding amount is 0.3-0.4kg/time·animal. The feed contains raw materials with the following weight ratio: 78 parts of standard monkey feed , 15 parts of animal fat, 5 parts of sugar, 2 parts of cholesterol;
(2 ) 24个月后, 将剂量为 25mg/kg的链脲菌素施用于猴, 每周施用一 次, 连续 2次。 (2) After 24 months, streptozotocin at a dose of 25 mg/kg was administered to the monkeys once a week for 2 consecutive times.
2、 根据权利要求 1所述的制备方法, 其特征在于: 歩骤 (1 ) 中, 所述 动物油脂是猪油; 所述糖是蔗糖。 2. The preparation method according to claim 1, characterized in that: in step (1), the animal fat is lard; and the sugar is sucrose.
3、 根据权利要求 1所述的制备方法, 其特征在于: 歩骤 (2 ) 中, 所述 的施用方式是静脉注射。 3. The preparation method according to claim 1, characterized in that: in step (2), the administration method is intravenous injection.
4、一种建立恒河猴 2型糖尿病模型的饲料, 其特征在于: 它包含如下重 量配比的原料: 标准猴饲料 78份、 动物油脂 15份、糖 5份、 胆固醇 2份。 4. A feed for establishing a rhesus monkey type 2 diabetes model, characterized in that: it contains raw materials in the following weight ratio: 78 parts of standard monkey feed, 15 parts of animal fat, 5 parts of sugar, and 2 parts of cholesterol.
5、 根据权利要求 4所述的饲料, 其特征在于: 所述动物油脂是猪油; 所 述糖是蔗糖。 5. The feed according to claim 4, characterized in that: the animal fat is lard; and the sugar is sucrose.
6、 权利要求 1~3任意一项所述的方法制备得到的恒河猴 2型糖尿病模 型。 6. The rhesus monkey type 2 diabetes model prepared by the method of any one of claims 1 to 3.
7、 权利要求 6所述动物模型在筛选治疗 2型糖尿病的药物中的用途。 7. Use of the animal model of claim 6 in screening drugs for treating type 2 diabetes.
8、一种筛选治疗 2型糖尿病模型药物的方法, 其特征在于: 它包括如下 歩骤: 8. A method for screening drugs for treating type 2 diabetes model, characterized by: It includes the following steps:
a、 按照权利要求 1~3任意一项所述方法, 建立恒河猴 2型糖尿病模型; b、 将候选药物施用于动物模型; a. Establish a rhesus monkey type 2 diabetes model according to the method described in any one of claims 1 to 3; b. Apply the candidate drug to the animal model;
c、 用动物模型评价潜在的治疗 2型糖尿病的药物。
c. Use animal models to evaluate potential drugs for the treatment of type 2 diabetes.
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