CN106389394A - Construction method of cynomolgus macaque model for alcoholic liver disease - Google Patents
Construction method of cynomolgus macaque model for alcoholic liver disease Download PDFInfo
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- 238000010276 construction Methods 0.000 title claims abstract description 19
- 208000022309 Alcoholic Liver disease Diseases 0.000 title abstract description 10
- 241000282567 Macaca fascicularis Species 0.000 title abstract 4
- 238000010171 animal model Methods 0.000 claims abstract description 19
- 238000012317 liver biopsy Methods 0.000 claims abstract description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 34
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- 108010011485 Aspartame Proteins 0.000 claims description 16
- 239000000605 aspartame Substances 0.000 claims description 16
- IAOZJIPTCAWIRG-QWRGUYRKSA-N aspartame Chemical compound OC(=O)C[C@H](N)C(=O)N[C@H](C(=O)OC)CC1=CC=CC=C1 IAOZJIPTCAWIRG-QWRGUYRKSA-N 0.000 claims description 16
- 229960003438 aspartame Drugs 0.000 claims description 16
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- 235000020188 drinking water Nutrition 0.000 claims description 11
- 210000004185 liver Anatomy 0.000 claims description 11
- 208000019425 cirrhosis of liver Diseases 0.000 claims description 10
- 235000013305 food Nutrition 0.000 claims description 10
- 230000002440 hepatic effect Effects 0.000 claims description 10
- 208000010706 fatty liver disease Diseases 0.000 claims description 9
- 230000006698 induction Effects 0.000 claims description 9
- 150000002632 lipids Chemical class 0.000 claims description 9
- 239000000243 solution Substances 0.000 claims description 9
- 208000006454 hepatitis Diseases 0.000 claims description 7
- 230000001476 alcoholic effect Effects 0.000 claims description 6
- 238000002347 injection Methods 0.000 claims description 6
- 239000007924 injection Substances 0.000 claims description 6
- 208000024891 symptom Diseases 0.000 claims description 6
- 208000007082 Alcoholic Fatty Liver Diseases 0.000 claims description 5
- 206010016262 Fatty liver alcoholic Diseases 0.000 claims description 5
- 208000026594 alcoholic fatty liver disease Diseases 0.000 claims description 5
- 102000004169 proteins and genes Human genes 0.000 claims description 5
- 108090000623 proteins and genes Proteins 0.000 claims description 5
- 231100000240 steatosis hepatitis Toxicity 0.000 claims description 5
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- 206010019708 Hepatic steatosis Diseases 0.000 claims description 4
- 206010019728 Hepatitis alcoholic Diseases 0.000 claims description 4
- 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 claims description 4
- 229930006000 Sucrose Natural products 0.000 claims description 4
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- 210000000170 cell membrane Anatomy 0.000 claims description 4
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- 239000000835 fiber Substances 0.000 claims description 3
- 239000004006 olive oil Substances 0.000 claims description 3
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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 2
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- 206010061218 Inflammation Diseases 0.000 description 1
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- 208000027418 Wounds and injury Diseases 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 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
- 230000015572 biosynthetic process Effects 0.000 description 1
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- 230000035622 drinking Effects 0.000 description 1
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- 150000004665 fatty acids Chemical class 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 239000003862 glucocorticoid Substances 0.000 description 1
- 231100000234 hepatic damage Toxicity 0.000 description 1
- 231100000753 hepatic injury Toxicity 0.000 description 1
- 230000010224 hepatic metabolism Effects 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
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- 230000003834 intracellular effect Effects 0.000 description 1
- 230000003859 lipid peroxidation Effects 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 230000008818 liver damage Effects 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 230000001071 malnutrition Effects 0.000 description 1
- 235000000824 malnutrition Nutrition 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 229930189775 mogroside Natural products 0.000 description 1
- 230000017074 necrotic cell death Effects 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 208000015380 nutritional deficiency disease Diseases 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
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- 229940013618 stevioside Drugs 0.000 description 1
- OHHNJQXIOPOJSC-UHFFFAOYSA-N stevioside Natural products CC1(CCCC2(C)C3(C)CCC4(CC3(CCC12C)CC4=C)OC5OC(CO)C(O)C(O)C5OC6OC(CO)C(O)C(O)C6O)C(=O)OC7OC(CO)C(O)C(O)C7O OHHNJQXIOPOJSC-UHFFFAOYSA-N 0.000 description 1
- 235000019202 steviosides Nutrition 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 238000002560 therapeutic procedure Methods 0.000 description 1
- 230000009385 viral infection Effects 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/02—Halogenated hydrocarbons
-
- 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
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/0004—Screening or testing of compounds for diagnosis of disorders, assessment of conditions, e.g. renal clearance, gastric emptying, testing for diabetes, allergy, rheuma, pancreas functions
- A61K49/0008—Screening agents using (non-human) animal models or transgenic animal models or chimeric hosts, e.g. Alzheimer disease animal model, transgenic model for heart failure
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Epidemiology (AREA)
- Zoology (AREA)
- Environmental Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Endocrinology (AREA)
- Medicinal Chemistry (AREA)
- Urology & Nephrology (AREA)
- Rheumatology (AREA)
- Pathology (AREA)
- Gastroenterology & Hepatology (AREA)
- Diabetes (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Toxicology (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Engineering & Computer Science (AREA)
- Acyclic And Carbocyclic Compounds In Medicinal Compositions (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Fodder In General (AREA)
Abstract
The invention belongs to a construction method of an animal model, and in particular relates to a construction method of a cynomolgus macaque model for alcoholic liver disease. The construction method of the cynomolgus macaque model for alcoholic liver disease comprises the following steps: screening experimental animals, feeding the screened experimental animals with high-glucose and high-fat feed, and conducting liver biopsy and acquisition of the determined model. The construction method of the cynomolgus macaque model for alcoholic liver disease provided by the invention is short in duration and high in success rate; the construction method is suitable for researching the occurrence mechanism of the alcoholic liver disease and is convenient for screening drugs for treating the alcoholic liver disease and for evaluating the efficacy of the drugs on the alcoholic liver disease; and the construction method has a good application prospect.
Description
Technical field
The invention belongs to the construction method of animal model is and in particular to arrive a kind of structure of machin Alcoholic Liver Disease Model
Method.
Background technology
With China's rapid development of economy, the consumption of resident's alcohol is also continuously increased, and related problem is also got over to drinking
Come more prominent.Normal person at 24 hours in vivo can metabolism 120g ethanol, but for a long time, excessively drink and can exceed the metabolizable energy of body
Power, may result in AML.The toxic action of the generation of this disease mainly ethanol and its metabolin-acetaldehyde causes, especially
It is that acetaldehyde is the most obvious to hepatocellular toxic action, the two directly or indirectly leads to the wide array of pathologies of liver：Degeneration of liver cells (wine
Essence fatty liver), downright bad (hepatitis), fibrillatable (liver fibrosis), cirrhosis, liver cancer can be developed into when serious.
Clinically, the treatment of AML lacks targetedly target treatment method, clinically can only be passive
(prevent and treat malnutrition with nutritional support and controlled inflammation with glucocorticoid) based on anti symptom treatment.
Therefore, set up a kind of animal model closer to mankind's AML, especially build and there is alcoholic liver
The animal model of pathology process, using its mechanism of this Research of Animal Model for Study, and then finds new therapy target and medicine, is
There is very important basic medical theory and clinical value.
The information being disclosed in this background section is merely intended to increase the understanding of the general background to the present invention, and should not
Recognize when being considered or imply in any form that this information structure has been the prior art well known to persons skilled in the art.
Content of the invention
It is an object of the invention to provide a kind of construction method of AML animal model, the feature of the method be
Build animal model process, gradually realize alcoholic liver pathology process (include alcoholic fatty liver, alcoholic hepatitis,
Alcoholic fibrillatable etc.).
Technical scheme provided by the present invention is：
A kind of construction method of AML animal model, comprises the following steps：
(1) machin choosing male 8-10kg is as research object, in addition to normal nursing basal feed, suitable at the 1st week
It should be allowed in the stage voluntarily to drink the drinking water containing Aspartame, starting from the 2nd week to press drinking water increases weekly 10%
Alcohol is freely drunk by it, filters out to the 4th weekend and drinks the machin that 30% alcohol is more than 50mL daily as experimental animal；
(2) started to replace the Aspartame alcoholic solution with 10% sucrose from the 5th week, except 30% alcohol is fed in continuation
Outward, additionally add 20% leaf fat in basal feed；
(3) foundation of alcoholic fatty liver model：Fed using high sugar, high lipid food, bar freely drunk by 30% alcohol
Under part, raised for the 12nd weekend 1., carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin performance significantly fat
Liver symptom；
(4) foundation of alcoholic hepatitis, Liver Fibrosis Model：On the basis of 1., it is continuing with high sugar, high lipid food is fed
Support, 30% alcohol freely drinks condition, and lumbar injection carbon tetrachloride solution, 2 times a week, per injection amount is 0.25mL/
kg；At 16th weekend, carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin shows obvious liver plasma membrane and breaks
The hepatitis symptom split, infiltrating；At 24th weekend, carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin performance is bright
Aobvious liver fibrosis symptom.
Preferably, the mass percent of Aspartame is in the drinking water containing Aspartame described in step (1)
0.1-1%.
Preferably, the mass percent of Aspartame is 0.5% in the described drinking water containing Aspartame.
Preferably, the basal feed described in step (1) is protein 15-20%, carbohydrate 50-60%, fat
Fat 5%, crude fibre 4-6%, other 9-26%.
Preferably, the high sugar described in step (4), high lipid food are on the basis of basal feed, sugar is risen to by 3%
8-10%, fat rises to 20-25% by 5%.
Preferably, the carbon tetrachloride solution described in step (4) is that carbon tetrachloride is dispersed in olive oil, its content
For 10%v/v.
Compared with prior art, the present invention has the advantages that：
(1) the construction method time of the machin Alcoholic Liver Disease Model of the present invention is short, and success rate is high, can be used for studying
The genesis mechanism of AML, is easy to screen the medicine for the treatment of AML and evaluates medicine to this AML
Drug effect, has a good application prospect.
(2) animal has the nature detesting alcohol, therefore passes through luring of sweetener at the 1st week, promotes machin can actively take the photograph
Food ethanol.In order to avoid part sweetener, liver is damaged protective effect (as sucrose, glucose etc. are provided that hepatic mitochondria energy,
Mogroside, stevioside etc. are avoided that hepatocellular damage), modeling initial stage preferred Aspartame is sweetener, so can
Lure machin actively to ingest alcohol, can avoid to hepatocellular protective effect again.
(3) caused by the generation of AML is mainly ethanol and its toxic action of metabolin-acetaldehyde, especially acetaldehyde
It is many on hepatocellular impact：Impact mitochondria to the generation of atriphos (ATP), the biosynthesis of protein and
Excretion be obstructed, micro-duct injury, make albumen, fat excretion obstacle and in liver cell accumulate.And affecting hepatocellular initial stage, sugarcane
Sugar, glucose, fat, protein etc. can resist hepatocellular loss.Alcoholic fatty liver model is to be drawn by alcohol in early stage
On the premise of playing slight liver damage, increasing sugared part, the ratio of fat in feed, thus increasing the burden of liver, promoting fat in liver
Intracellular rapid accumulation, forms fatty liver.
(4) simple sugared, the higher fatty acid nursing of alcohol+height in most cases can only induce steatosis, wants to reach tighter
The hepatic injury (as hepatitis, liver fibrosis etc.) of weight, generally requires the second ripple and hits, i.e. secondary damage.The mode that second ripple hits
Have a lot, including drug factors, hormone, liver microsomalinducer, genetic manipulation and virus infection etc..The preferred carbon tetrachloride of the present invention
The means hit for the second ripple, because carbon tetrachloride quickly can strengthen lipid peroxidation after liver metabolism, cause in liver cell
The denaturation of matter net, mitochondria, golgiosome or even cell membrane and necrosis, form hepatitis, and the later stage is easier to cause liver fibrosis.
Brief description
Accompanying drawing 1 shows obvious fatty liver symptom for machin；
Accompanying drawing 2 shows obvious hepatitis symptom for machin；
Accompanying drawing 3 shows obvious liver fibrosis symptom for machin.
Specific embodiment
With reference to concrete example, the present invention is further detailed explanation.
A kind of construction method of AML animal model, comprises the following steps：
(1) machin choosing male 8-10kg is as research object, in addition to normal nursing basal feed, suitable at the 1st week
It should be allowed in the stage voluntarily to drink the drinking water containing Aspartame, starting from the 2nd week to press drinking water increases weekly 10%
Edible ethanol is allowed to freely drink, and filters out to the 4th weekend and drinks the machin that 30% alcohol is more than 50ml daily as test
Animal；In the described drinking water containing Aspartame, the mass percent of Aspartame is 0.1-1%；Described basal feed
For protein 15-20%, carbohydrate 50-60%, fat 5%, crude fibre 4-6%, other 9-26%；
(2) started to replace the Aspartame alcoholic solution with 10% sucrose from the 5th week, except 30% alcohol is fed in continuation
Outward, additionally add 20% leaf fat in basal feed；
(3) foundation of alcoholic fatty liver model：Fed using high sugar, high lipid food, bar freely drunk by 30% alcohol
Under part, raised for the 12nd weekend 1., carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin performance significantly fat
Liver symptom, referring to Fig. 1；Described high sugar, high lipid food are on the basis of basal feed, and sugar rises to 8-10% by 3%, fat
Rise to 20-25% by 5%；
(4) foundation of alcoholic hepatitis, Liver Fibrosis Model：On the basis of 1., it is continuing with high sugar, high lipid food is fed
Support, 30% alcohol freely drinks condition, and lumbar injection carbon tetrachloride solution, 2 times a week, per injection amount is 0.25mL/
kg；At 16th weekend, carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin shows obvious liver plasma membrane and breaks
The hepatitis symptom split, infiltrating, referring to Fig. 2；At 24th weekend, carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, eat crab
Monkey shows obvious liver fibrosis symptom, referring to Fig. 3；Described carbon tetrachloride solution is that carbon tetrachloride is dispersed in olive oil
In, its content is 10%v/v.
The description of the aforementioned specific illustrative embodiment to the present invention illustrate that and illustration purpose.These descriptions
It is not wishing to limit the invention to disclosed precise forms, and it will be apparent that according to above-mentioned teaching, can much be changed
And change.The purpose of selecting and describing the exemplary embodiment is that explaining that the certain principles of the present invention and its reality should
With so that those skilled in the art be capable of and utilize the present invention various different exemplary and
Various different selections and change.The scope of the present invention is intended to be limited by claims and its equivalents.
Claims (6)
1. a kind of construction method of AML animal model is it is characterised in that comprise the following steps：
(1) machin choosing male 8-10kg, as research object, in addition to normal nursing basal feed, adapted to rank at the 1st week
Duan Rangqi voluntarily drinks the drinking water containing Aspartame, starts from the 2nd week to press the alcohol increasing weekly 10% drinking water
Allow it freely to drink, filter out to the 4th weekend and drink the machin that 30% alcohol is more than 50mL daily as experimental animal；
(2) started to replace the Aspartame alcoholic solution with 10% sucrose from the 5th week, in addition to continuing to feed 30% alcohol,
The leaf fat of extra interpolation 20% in basal feed；
(3) foundation of alcoholic fatty liver model：Fed using high sugar, high lipid food, under the conditions of 30% alcohol is freely drunk,
Raised for the 12nd weekend 1., carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin shows obvious fatty liver condition
Shape；
(4) foundation of alcoholic hepatitis, Liver Fibrosis Model：On the basis of 1., it is continuing with high sugar, high lipid food is fed,
30% alcohol freely drinks condition, and lumbar injection carbon tetrachloride solution, and 2 times a week, per injection amount is 0.25mL/kg；
At 16th weekend, carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin performance obvious liver plasma membrane rupture, leaching
The hepatitis symptom of profit；At 24th weekend, carry out liver biopsy through B ultrasonic induction, hepatic tissue is dyeed with HE, machin shows obvious liver
Fibrotic symptoms.
2. the construction method of AML animal model according to claim 1 is it is characterised in that institute in step (1)
In the drinking water containing Aspartame stated, the mass percent of Aspartame is 0.1-1%.
3. AML animal model according to claim 2 construction method it is characterised in that described contain Ah
In the drinking water of this Ba Tian, the mass percent of Aspartame is 0.5%.
4. the construction method of AML animal model according to claim 1 is it is characterised in that institute in step (1)
The basal feed stated is protein 15-20%, carbohydrate 50-60%, fat 5%, crude fibre 4-6%, other 9-26%.
5. the construction method of AML animal model according to claim 1 is it is characterised in that institute in step (4)
The high sugar stated, high lipid food are on the basis of basal feed, and sugar rises to 8-10% by 3%, and fat rises to 20-25% by 5%.
6. the construction method of AML animal model according to claim 1 is it is characterised in that institute in step (4)
The carbon tetrachloride solution stated is that carbon tetrachloride is dispersed in olive oil, and its content is 10%v/v.
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107519153A (en) * | 2017-09-29 | 2017-12-29 | 四川省人民医院 | A kind of method that administration by gavage establishes rhesus macaque Liver Fibrosis Model |
| CN107693507A (en) * | 2017-09-29 | 2018-02-16 | 四川省人民医院 | A kind of method that intraperitoneal injection establishes rhesus macaque Liver Fibrosis Model |
| CN107801691A (en) * | 2017-12-06 | 2018-03-16 | 凯斯艾生物科技（苏州）有限公司 | A kind of chronic fat hepatitis rat model of non-alcoholic and its construction method and purposes |
| CN107912366A (en) * | 2017-12-06 | 2018-04-17 | 江苏珂玛麒生物科技有限公司 | A kind of chronic fat hepatitis non-human primate model of non-alcoholic and its construction method and purposes |
| CN111134240A (en) * | 2020-01-18 | 2020-05-12 | 湖北天勤生物科技有限公司 | Liquid feed and method for constructing alcoholic liver disease model of cynomolgus monkey |
| CN111631188A (en) * | 2019-03-01 | 2020-09-08 | 广西中医药大学 | Method for rapidly inducing alcoholic fatty liver model by single factor |
| CN114668077A (en) * | 2020-12-24 | 2022-06-28 | 常州鼠一鼠二生物科技有限公司 | Non-alcoholic fatty liver model feed for cynomolgus monkeys as well as preparation method and application of non-alcoholic fatty liver model feed |
| CN114731985A (en) * | 2022-03-29 | 2022-07-12 | 华南理工大学 | Construction method of metabolism-related fatty liver disease non-human primate model |
| CN116019054A (en) * | 2022-12-30 | 2023-04-28 | 江苏珂玛麒生物科技有限公司 | Construction method of cynomolgus monkey NASH model |
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| CN107912366B (en) * | 2017-12-06 | 2020-12-11 | 江苏珂玛麒生物科技有限公司 | Non-alcoholic chronic steatohepatitis non-human primate model and construction method and application thereof |
| CN111631188B (en) * | 2019-03-01 | 2022-04-22 | 广西中医药大学 | Method for rapidly inducing alcoholic fatty liver model by single factor |
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| CN111134240A (en) * | 2020-01-18 | 2020-05-12 | 湖北天勤生物科技有限公司 | Liquid feed and method for constructing alcoholic liver disease model of cynomolgus monkey |
| CN114668077A (en) * | 2020-12-24 | 2022-06-28 | 常州鼠一鼠二生物科技有限公司 | Non-alcoholic fatty liver model feed for cynomolgus monkeys as well as preparation method and application of non-alcoholic fatty liver model feed |
| CN114731985A (en) * | 2022-03-29 | 2022-07-12 | 华南理工大学 | Construction method of metabolism-related fatty liver disease non-human primate model |
| CN114731985B (en) * | 2022-03-29 | 2023-09-26 | 华南理工大学 | Construction method of non-human primate model of metabolic-related fatty liver disease |
| CN116019054A (en) * | 2022-12-30 | 2023-04-28 | 江苏珂玛麒生物科技有限公司 | Construction method of cynomolgus monkey NASH model |
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