CN103432572A - Rhesus monkey diabetic nephropathy model preparation method - Google Patents
Rhesus monkey diabetic nephropathy model preparation method Download PDFInfo
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Abstract
The present invention discloses a rhesus monkey diabetic nephropathy model preparation method, which comprises: (1) applying streptozocin with a dose of 80-100 mg/kg on a rhesus monkey; and (2) after a blood glucose concentration is increased to 11.1 mmol/l, applying insulin before meal, such that a rhesus monkey blood glucose concentration is 10-20 mmol/L. According to the present invention, based on the diabetes mellitus rhesus monkey, the blood glucose control method is adopted to successfully induce the rhesus monkey diabetic nephropathy model.
Description
Technical field
The present invention relates to the preparation method of Rhesus Macacus diabetic nephropathy model.
Background technology
The sickness rate of diabetes (Diabetes Mellitus, DM) is the trend of rapid growth.It is reported that now approximately there are 9,200 ten thousand DM and 1.48 hundred million DM patient in early stage in China, becomes second onset diabetes big country after the U.S..Along with the development gradually of the state of an illness increases the weight of, in these patients, approximately have the type 1 diabetes patient of 30-40% and Yue 20% type 2 diabetes mellitus patient follow serious renal function injury---diabetic nephropathy (Diabetic Nephropathy, DN).
DN is one of the most serious chronic complicating diseases of diabetics, is also the topmost cause of death of diabetics.Once significant clinical symptoms has appearred in DN patient, now selectable medicine is less, and the therapeutic effect of medicine is poor, and the state of an illness often is carries out sexual development until the renal failure in latter stage brings huge financial burden to patient family and society.Therefore furtheing investigate pathogenesis and the early stage diagnostic method of DN, explore on this basis the effective measures that delay or reverse disease process, is focus and the difficulties of current medical research.
Animal model is the basis of research mankind major disease, sets up the DN animal model highly approximate with the mankind, will obtain the effect of getting twice the result with half the effort at aspects such as research its pathogeny, early diagnosis and therapeutic interventions.Non-human primate is as the mankind's close relative, about 96% above homology aspect genetic, its morphosis, physiological function and biochemical metabolism are all closely similar with the mankind, set up a kind of non-human primate DN model genesis, early diagnosis and the new drug development etc. of studying DN are had to important scientific meaning and economic worth.
Summary of the invention
The object of the present invention is to provide the preparation method of Rhesus Macacus diabetic nephropathy model.
The preparation method of Rhesus Macacus diabetic nephropathy model of the present invention, comprise the steps:
(1) streptozotocin that is 80～100mg/kg by dosage is applied to Rhesus Macacus;
(2) after blood sugar concentration rises to 11.1mmol/l, administration of insulin before the meal, making the Rhesus Macacus blood sugar concentration is 10-20mmol/L.
The preparation method of Rhesus Macacus diabetic nephropathy model of the present invention, comprise the steps:
(1) streptozotocin that is 80～100mg/kg by dosage is applied to Rhesus Macacus;
(2) after blood sugar concentration is increased to 11.1mmol/l, the high lipid food of feeding, feed every day 2 times, scale of feeding is 0.3-0.4kg/ time, and in administration of insulin before the meal, making the Rhesus Macacus blood sugar concentration is 10-20mmol/L, the raw material that described feedstuff comprises following weight proportion: 78 parts, standard monkey feedstuff, 15 parts of animal oils, 5 parts of sugar, 2 parts, cholesterol.
Standard monkey feedstuff, refer to the monkey mixed feed that " the Chinese name standard GB/T 14924.8-2001 of republic " stipulates.
Animal oil, refer to the fat that derives from animal, as: Adeps Bovis seu Bubali, Adeps Caprae seu ovis, Adeps Sus domestica.
In step (1), described method of application is intravenous injection.
In step (2), described method of application is subcutaneous injection.
In step (2), described animal oil is Adeps Sus domestica; Described sugar is sucrose.
The high lipid food of Rhesus Macacus diabetic nephropathy model of the present invention, the raw material that it comprises following weight proportion: 78 parts, standard monkey feedstuff, 15 parts of animal oils, 5 parts of sugar, 2 parts, cholesterol.
Described animal oil is Adeps Sus domestica; Described sugar is sucrose.
Animal model prepared by preceding method of the present invention purposes in the medicine of screening treatment Rhesus Macacus diabetic nephropathy.
The present invention screens the method for the medicine for the treatment of Rhesus Macacus diabetic nephropathy model, and it comprises the steps:
A, according to preceding method, set up the Rhesus Macacus diabetic nephropathy model;
B, drug candidate is applied to animal model;
C, with the medicine of the potential treatment Rhesus Macacus diabetic nephropathy of animal model evaluation.
Modeling method of the present invention can induce Rhesus Macacus the clinical symptoms of the diabetic nephropathyes such as kidney injury, Microalbuminuria to occur, and medication is simple, repeatable strong.
Obviously, according to foregoing of the present invention, according to ordinary skill knowledge and the customary means of this area, not breaking away under the above-mentioned basic fundamental thought of the present invention prerequisite, can also make modification, replacement or the change of other various ways.
The specific embodiment of form, be described in further detail foregoing of the present invention again by the following examples.But this should be interpreted as to the scope of the above-mentioned theme of the present invention only limits to following example.All technology realized based on foregoing of the present invention all belong to scope of the present invention.
The accompanying drawing explanation
Fig. 1 glycemic control situation;
Fig. 2 renal function testing result;
Fig. 3 fibrosis gene alteration situation;
Fig. 4 renal hemodynamics parameter and blood perfusion coefficient;
Blood distribution and frequency spectrum that Fig. 5 B-ultrasonic renal checks;
Fig. 6 nephridial tissue pathological change;
Fig. 7 nephridial tissue immunofluorescence.
The specific embodiment
The preparation of embodiment 1 Rhesus Macacus diabetogenous nephrosis of the present invention symptom model
1 experiment material and instrument
1.1 test sample essential information
IL-1 β antibody: abcam
TNF-a antibody: abcam
TGF-beta 1 antibodies: Bioworld
CTGF antibody: abcam
Collagen IV antibody: abcam
Fibronection antibody: Bioworld
MCP-1 antibody: ebioscience
CD68 (KP1) antibody: abcam
ENOS antibody: abcam
Total RNA from animal tissues extracts test kit: day root
IScript
tMcDNA synthetic agent box: BIO-RAD
iQ
TM 
Green?Supermix：BIO-RAD
1.2 test required key instrument, apparatus
The full vigor type of blood-sugar detecting instrument: Luo Kang blood-sugar detecting instrument, the full vigor type of Luo Kang blood sugar test paper
Fully automatic blood biochemistry analyzer (Switzerland Roche COBAS Integra400Plus, numbering MEB-02-01)
BIO-RAD CFX96 quantitative real time PCR Instrument
BIO-RAD S200PCR instrument
LEICA DM4000B fluorescence microscope
Olympus BX51 optical microscope
Puncture needle, puncture rifle: BARD
Color ultrasound; Iu22philips
Fundus fluorescein angiography: Topcon TRC.50DX(Japan opens up general Kanggong department and produces)
1.3 laboratory animal
1.3.1 experimental system
14 of Rhesus Macacus, body weight 3.5～5.5kg, intend breeding study base (production licence number: SCXK (river) 2004-013) by Chengdu, Sichuan safety animal and introduce.Enter the room and carry out tuberculin, parasite, Salmonella, the Salmonella of congratulating inspection the last week.
1.4 animal feeding management
1.4.1 quarantine/environmental adaptation
First conform and quarantine one week before the Rhesus Macacus that enters the room test, select healthy buck as animal subject, the quarantine content: the quality index whether required when ordering is consistent; Body temperature, food ration, blood biochemistry, electrolyte; The animal general state; Whether the weight of animals reaches the scope of test requirements document body weight.
1.4.2 animal feeding
The SYXK(river) 2003-030), rustless steel mouse cage sub-cage rearing for every of Rhesus Macacus Rhesus Macacus is raised in Chinese native medicine security assessment centers conventional animal room, national Chengdu (experimental animal room occupancy permit number:, the raising condition meets GB GB14925-2001,21 ± 5 ℃ of room temperatures (temperature difference per day≤4 ° C), relative humidity 55 ± 15%, within 12 hours, light and shade replaces, rate of ventilation 8～10 times/hour.
1.4.3 animal identification method
Rhesus Macacus is by the common labelling of label on chest card and cage
1.5 experiment grouping
Grouping comprises normal group (3), after the modeling of experimental group A(STZ diabetes, by subcutaneous injection insulin 15 minutes before the meal every day, control blood glucose<10mmol/l, 3), after the modeling of experimental group B(STZ diabetes, by subcutaneous injection insulin 15 minutes before the meal every day, control blood glucose in 10-20mmol/l, 5), after the modeling of experimental group C(STZ diabetes, by subcutaneous injection insulin 15 minutes before the meal every day, control blood glucose in 10-20mmol/l, and the high lipid food of feeding, 3).
Feed formula:
Experimental group A, B: monkey standard monkey feedstuff---business-like monkey pellet (purchased from Sichuan Academy of Medical Sciences's Institute of Botany, formula is identical with the monkey mixed feed of " the Chinese name standard GB/T 14924.8-2001 of republic " regulation).
Experimental group C: high lipid food: 15% refined lard (commercially available), 5% sucrose (commercially available), 2% cholesterol (commercially available), 78% monkey standard monkey feedstuff.
The mode of feeding: feed every day 2 times, scale of feeding be 0.3-0.4kg/ time only.
2 experimental techniques
2.1 the animal general status is observed
Observe the situations such as animal appearance sign (hair, eye and the mucosa that comprise animal), behavioral activity, mental status, glandular secretion, breathing, diet, Excreta, injection site, detect the weight of animals, blood pressure and food ration.
2.2 modeling method
Experimental group A, B:
More than animal different fasting 12h, after venoclysis normal saline 100ml dilatation, then inject the STZ (80mg/kg) prepared before administration;
Give conventional monkey forage feed, feed every day 2 times, scale of feeding is 0.3-0.4kg/ time;
Detect animal fasting glucose (FBG), FBG by subcutaneous injection insulin 15 minutes before the meal every day, controls blood sugar concentration higher than after 11.1mmol/l in continuous 2 days.
Experimental group C:
More than animal different fasting 12h, after venoclysis normal saline 100ml dilatation, then inject the STZ (80mg/kg) prepared before administration;
Give conventional monkey forage feed, feed every day 2 times, scale of feeding is 0.3-0.4kg/ time;
Detect animal fasting glucose (FBG), FBG by subcutaneous injection insulin 15 minutes before the meal every day, controls blood sugar concentration higher than after 11.1mmol/l in continuous 2 days, gives with high lipid food simultaneously and feeds, and feeds every day 2 times, and scale of feeding is 0.3-0.4kg/ time.
2.3 hematological examination
Detect index: red blood cell count(RBC) (RBC, capacitance method), hemoglobin (HGB, the HiCN method), red cell volume (HCT, the Power Capacity point-score), mean corpuscular volume (MCV, calculate), mean corpuscular hemoglobin (MCH, calculate), mean corpuscular hemoglobin concentration (MCHC) (MCHC, calculate), reticulocyte count (RET, fluorescent staining method), numeration of leukocyte (WBC, laser method) and the classification (neutrophilic granulocyte NEU, lymphocyte LYM, mononuclear cell MONO, eosinophilic granulocyte EOS, basophilic granulocyte BASO, laser method), platelet count (PLT, capacitance method), reticulocyte (RET, Liu Shi+fluorescent staining method), prothrombin time (PT, the chromophoric substrate method), activated partial thrombin time (APTT, the chromophoric substrate method).Aspartate amino transferase (AST, IFCC P-5 '-P method), alanine aminotransferase (ALT, IFCC P-5 '-P method), gamma glutamyltransferase (GGT, L-gamma-glutamyl-3-carboxyl-paranitroanilinum method), creatine phosphokinase (CK, the HK-G6PD method), alkali phosphatase (ALP, the 4-NPP method), lactic acid dehydrogenase (LDH, the IFCC method), blood urea nitrogen (Urea, the Urease-GLDH-Kinetic method), total protein (TP, the Biuret method), albumin (ALB, the BCG method), blood glucose (GLU, the GLK-G6PDH method), total bilirubin (TBIL, the Diazo method), creatinine (Crea, the JAFF method), T-CHOL (CHOL, the CHOD-P method), triglyceride (TG, the GPO-PAP method), Na ion concentration (Na+, ion selective electrode method), potassium concentration (K+, ion selective electrode method), chlorine ion concentration (Cl-, ion selective electrode method) etc.
Detection time: give the STZ Isosorbide-5-Nitrae, each survey of week once, survey once by later every 1～February.
Method: detect from the blood sampling of Rhesus Macacus back leg great saphenous vein.Hematological examination is with blood with the EDTA anticoagulant, and clotting time uses blood with the sodium citrate anticoagulant.
2.4 fasting glucose (Fasting blood glucose, FBG) and post-prandial glycemia (Postprandial blood glucose, PBG) detect
Detection time: detect weekly FBG(points in the mornings 9 for 2 times, on an empty stomach after 12h) 2h and after the PBG(feed).
Detection method: get finger or toe peripheral blood, detect by the Roche blood glucose meter.
2.5 intravenous glucose tolerance experiment (IVGTT)
Vein gives 50% glucose injection (0.5g/kg body weight), within 0,1,3,5,10,30,60 and 120 minutes after injecting, measures blood sugar level, checks after glucose injection 0,1,3,5 simultaneously, the insulin level (RIA or ELISA) in 30 minutes blood samples
2.6 empty stomach glycolated hemoglobin, serum insulin and C peptide (C-peptide, C-P) horizontal detection
Detection time: got serum every 4 weeks and detect.
Detection method: radioimmunology (RIA) test kit, enzyme linked immunosorbent detection method (ELISA)
2.7 insulinize
Empty stomach and post-prandial glycemia monitoring, according to kind, dosage form and the consumption of blood sugar level adjustment diabetes monkey subcutaneous injection insulin.Therapeutic effect is observed in 24 hours blood glucose monitoring, determines optimal case.Regular promoting the circulation of blood routine and biochemical analysis.
2.8 examination of ocular fundus
Detection time: when before administration and after administration, the observation period finishes.
Assay method: Rhesus Macacus is through ketalar anesthesia (intramuscular injection, 15mg/kg), U.S. Dolly mydriasis (eye drip, 1 droplet /), with binocular indirect ophthalmoscope, checked, all imaging evaluations all carry out (the Animal Anesthesia method is with reference to Rhesus Macacus surgery anesthesia SOP) under the general anesthesia state.Observe shape, size, the color and luster of papilla of optic nerve with ophthalmofundoscope, whether edge is clear; Macula area have or not edema, hemorrhage, ooze out and the pigment disorder; Retina has or not edema, oozes out, hemorrhage, peel off and new vessels etc.
2.9 kidney B ultrasonic and renal angiography
Color ultrasound observes the kidney form and blood perfusion is analyzed hemodynamic index and blood perfusion coefficient.Contrast agent SonoVue (Bracco SpA, Milan, Italy) injects 1ml(5mg/ml by great saphenous vein), then with the 2ml normal saline flushing.Do afterwards left kidney, midfeather is not less than 10 minutes again, midway flash explosion in addition.
2.10B the lower renal biopsy with gun-biopsy of super guiding
Go to ventricumbent position after Rhesus Macacus intramuscular injection ketamine and safe and sound anesthesia, drape after the preserved skin sterilization, point of puncture is selected left upper pole of kidney or the lower utmost point, avoids blood vessel and renal pelvis.BARD kidney lancet puncture aspiration, put into respectively the EP pipe of 10% paraformaldehyde and RNA latter and preserve.All fixing organizations carry out HE, Masson, PAS normal dyeing, and immunofluorescence dyeing (CTGF, eNOS, IL-1 β, IV Collagen Type VI, fibronectin FN, TGF-β, MCP-1, TNF-α, CD68) changes in micro-Microscopic observation histopathology.RNA latter uses fluorescence quantitative PCR detection smad2, smad3, smad4, smad7, actin, NF-κ b gene expression after preserving tissue extraction RNA reverse transcription.
2.11 liquid-phase chip detects
Get the blood plasma of the different courses of disease, detect the factor (IL-1, IL-6, MCP-1, TNF-α, IL-17, IL-18)
2.12 urine detection
Rhesus Macacus is put into metabolic cage, collects urina sanguinis and twenty-four-hour urine, detects routine urinalysis, urinaryalbumin and creatinine.
3 experimental results
3.1 glycemic control situation
As shown in Figure 1, the blood glucose of each experimental group all remains in the scope of predesignating.
3.2 renal function detects
As shown in Figure 2, uroscopy shows that experimental group B and C started to have occurred Microalbuminuria (MAU) at 36th month, and, along with course of disease passing has increase trend.Tri-groups of serum creatinines of blood 42 months ABC of biochemical demonstration have been compared notable difference with blood urea nitrogen with normal group.
3.3 blood distribution and frequency spectrum that B-ultrasonic renal checks
As shown in Figure 5, early stage kidney vascular bed resistance increment, thus blood flow in kidney is changed, along with aggravation, arteriosclerotic vessel wall elasticity lowers, luminal stenosis, and peripheral vascular resistance increases, renal blood perfusion reduces, make the kidney blood flow be low stream, high-impedance state, wherein, B group and C group pathological change very obvious, C organizes the most obvious.
3.4 renal hemodynamics parameter and blood perfusion coefficient
As shown in Figure 4, with matched group, compare, tri-groups of hemodynamic parameter drag indexs of ABC (RI) increase, lowest speed diastasis (EDV) reduces, and area under the blood perfusion coefficient curve (AUC) reduces, the difference maximum of C group and matched group, secondly, A organizes the difference minimum to the B group.
3.5 nephridial tissue pathological change
As shown in Figure 6, compare the A group with normal group without obviously changing, BC group inflammatory cell is invaded the deposition of profit, proliferation of mesangial cells, basement membrane thickened, collagen and glycogen, and the C group is more remarkable.
3.6 nephridial tissue immunofluorescence
As shown in Figure 7, the nephridial tissue immunofluorescence shows experimental group BC and normal group and tests that the A group is compared CTGF, eNOS, IL-1, IV Collagen Type VI, fibronectin (FN), TGF-β expresses increase, and C is more remarkable at experimental group, MCP1 and TNF-alpha expression also appear in experimental group C simultaneously.
3.7 fibrosis gene alteration
As shown in Figure 3, Real-time PCR shows that experimental group B and C started to occur that Smad2 and Smad3mRNA express at 24th month, and experimental group C is more remarkable, and occurs that at 42nd month Smad7 and NF-κ b mrna expression increase.
3.8 ophthalmologic examination
Except one (05539) has the eyes cataract fails to check, all the other two external eyes are without extremely, and conjunctiva is without congestion and edema, corneal transparency, and the anterior chamber is without extremely, pupil moderate loose large (all dripped and used mydriatic).Crystal is transparent, and vitreous body is without muddiness.Optical fundus: eyes leopard stricture of vagina shape optical fundus, the papilla of optic nerve color is light red, C/D=0.3-0.7, A:V=1:2, visible sieve aperture and blood vessel fluctuation.Have no hemorrhage, ooze out, microangioma and edema, central fovea of macula is reflective clear.
To sum up, the related symptoms of diabetic nephropathy does not appear in experimental group A, and the typical clinical symptom of diabetic nephropathy has appearred in experimental group B and C:
1, experimental group B and C started to have occurred Microalbuminuria (MAU) and along with the course of disease is passed increase trend is arranged at 36th month;
2, nephridial tissue HE, Masson, the PAS demonstration of dyeing: experimental group A is without pathological changes, experimental group B and C kidney occur that since 24th month inflammatory cell invades the deposition of profit, proliferation of mesangial cells, basement membrane thickened, collagen and glycogen, and find that local glomerular sclerosis appearred in experimental group C at 42nd month.Same time nephridial tissue immunofluorescence shows experimental group BC in normal group and tests that the A group is compared CTGF, eNOS, IL-1, IV Collagen Type VI, fibronectin (FN), TGF-β expresses increase, and C is more remarkable at experimental group, MCP1 and TNF-alpha expression also appear in experimental group C simultaneously;
3, Real-time PCR shows: experimental group B and C started to occur that Smad2 and Smad3mRNA express at 24th month, experimental group C is more remarkable, and occurred that at 42nd month Smad7 and NF-κ b mrna expression increase;
4, renal angiography shows, the blood perfusion coefficient AUC of experimental group B and C and hydromechanics parameter drag index RI significantly increase, diastasis, lowest speed EDV reduced (P<0.05), arteriosclerotic vessel wall elasticity lowers, luminal stenosis, peripheral vascular resistance increases, and renal blood perfusion reduces, and makes the kidney blood flow be low stream, high-impedance state;
The present invention is 10-20mmol/L by the blood sugar concentration scope of long-term control diabetes Rhesus Macacus, make the diabetes Rhesus Macacus occur that the nephridial tissue inflammatory cell invades the typical clinical symptom of the diabetic nephropathyes such as the deposition of profit, proliferation of mesangial cells, basement membrane thickened, collagen and glycogen, kidney injury, Microalbuminuria, Rhesus Macacus diabetic nephropathy modeling success of the present invention is described.
The present invention is 10-20mmol/L by the blood sugar concentration scope of long-term control diabetes Rhesus Macacus, the mode of taking in conjunction with the Long-term High-fat fat, make the various diabetic nephropathy classical symptoms of diabetes Rhesus Macacus more more remarkable than the mode of controlling separately blood glucose, illustrate that the mode of the two Combined Treatment is more excellent.
The medicine of embodiment 2 use model discrimination treatment of the present invention diabetic nephropathy
A, the Rhesus Macacus diabetic nephropathy model of setting up according to embodiment 1 method;
B, drug candidate is applied to animal model;
C, observe the affect situation of drug candidate on the various indexs of metabolism syndrome, estimate the medicine of potential treatment diabetic nephropathy disease.
To sum up, modeling method of the present invention is by the mode of long-term control blood glucose, induce Rhesus Macacus the clinical symptoms of diabetic nephropathy to occur, in addition, long-term glycemic control is taken in the mode of combining with high fat and also can be induced Rhesus Macacus the clinical symptoms of diabetic nephropathy to occur, and symptom is more remarkable, illustrate that two kinds of modes of the present invention all can effectively set up the Rhesus Macacus diabetic nephropathy model.
Claims (9)
1. the preparation method of a Rhesus Macacus diabetic nephropathy model, is characterized in that: comprise the steps:
(1) streptozotocin that is 80～100mg/kg by dosage is applied to Rhesus Macacus;
(2) after blood sugar concentration rises to 11.1mmol/l, administration of insulin before the meal, making the Rhesus Macacus blood sugar concentration is 10-20mmol/L.
2. the preparation method of a Rhesus Macacus diabetic nephropathy model, is characterized in that: comprise the steps:
(1) streptozotocin that is 80～100mg/kg by dosage is applied to Rhesus Macacus;
(2) after blood sugar concentration is increased to 11.1mmol/l, the high lipid food of feeding, feed every day 2 times, scale of feeding is 0.3-0.4kg/ time, and in administration of insulin before the meal, making the Rhesus Macacus blood sugar concentration is 10-20mmol/L, the raw material that described feedstuff comprises following weight proportion: 78 parts, standard monkey feedstuff, 15 parts of animal oils, 5 parts of sugar, 2 parts, cholesterol.
3. preparation method according to claim 1 and 2, it is characterized in that: in step (1), described method of application is intravenous injection.
4. preparation method according to claim 1 and 2, it is characterized in that: in step (2), described method of application is subcutaneous injection.
5. preparation method according to claim 2, it is characterized in that: in step (2), described animal oil is Adeps Sus domestica; Described sugar is sucrose.
6. a feedstuff of setting up the Rhesus Macacus diabetic nephropathy model, is characterized in that: the raw material that it comprises following weight proportion: 78 parts, standard monkey feedstuff, 15 parts of animal oils, 5 parts of sugar, 2 parts, cholesterol.
7. feedstuff according to claim 6, it is characterized in that: described animal oil is Adeps Sus domestica; Described sugar is sucrose.
8. the purposes of the animal model that prepared by the described method of claim 1～4 any one in the medicine of screening treatment Rhesus Macacus diabetic nephropathy.
9. a method of screening the medicine for the treatment of Rhesus Macacus diabetic nephropathy model, it is characterized in that: it comprises the steps:
A, according to the described method of claim 1～4 any one, set up the Rhesus Macacus diabetic nephropathy model;
B, drug candidate is applied to animal model;
C, with the medicine of the potential treatment Rhesus Macacus diabetic nephropathy of animal model evaluation.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014202024A1 (en) * | 2013-06-21 | 2014-12-24 | 四川大学华西医院 | Method for preparation of rhesus monkey diabetic nephropathy model |
| CN105726568A (en) * | 2016-04-11 | 2016-07-06 | 陕西科技大学 | Method for constructing diabetic nephropathy animal model |
| CN107469069A (en) * | 2017-08-18 | 2017-12-15 | 四川普莱美生物科技集团有限公司 | Modeling method, preparation, the preparation method of preparation, rhesus macaque pulmonary fibrosis model and its application of rhesus macaque pulmonary fibrosis model |
| CN115176758A (en) * | 2022-07-18 | 2022-10-14 | 四川省医学科学院.四川省人民医院实验动物研究所 | Method for establishing non-human primate type 1 diabetes model |
Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US9897565B1 (en) | 2012-09-11 | 2018-02-20 | Aseko, Inc. | System and method for optimizing insulin dosages for diabetic subjects |
| US9171343B1 (en) | 2012-09-11 | 2015-10-27 | Aseko, Inc. | Means and method for improved glycemic control for diabetic patients |
| US9233204B2 (en) | 2014-01-31 | 2016-01-12 | Aseko, Inc. | Insulin management |
| US9486580B2 (en) | 2014-01-31 | 2016-11-08 | Aseko, Inc. | Insulin management |
| US11081226B2 (en) | 2014-10-27 | 2021-08-03 | Aseko, Inc. | Method and controller for administering recommended insulin dosages to a patient |
| WO2016069475A1 (en) | 2014-10-27 | 2016-05-06 | Aseko, Inc. | Subcutaneous outpatient management |
| US9886556B2 (en) | 2015-08-20 | 2018-02-06 | Aseko, Inc. | Diabetes management therapy advisor |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2005009119A1 (en) * | 2003-07-23 | 2005-02-03 | Mcw Research Foundation, Inc. | Rat model of diabetic nephropathy |
| CN1785433A (en) * | 2004-12-09 | 2006-06-14 | 上海凯曼生物科技有限公司 | Preparation method of Type II diabetes animal model |
| CN101095958A (en) * | 2007-07-04 | 2008-01-02 | 山西医科大学第一医院 | Method for preparing 2 type diabetes rat model |
| CN101637405A (en) * | 2008-07-31 | 2010-02-03 | 四川大学华西医院 | Method for preparing diabetic animal model |
| CN101804211A (en) * | 2009-02-13 | 2010-08-18 | 四川大学华西医院 | Establishment of rhesus autoimmune 1 type diabetes model |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101766149A (en) * | 2010-01-29 | 2010-07-07 | 东南大学 | Preparation method of 2-type diabetic nephropathy model |
| CN103432572B (en) * | 2013-06-21 | 2018-09-18 | 四川大学华西医院 | A kind of preparation method of rhesus monkey diabetic nephropathy model |
-
2013
- 2013-06-21 CN CN201310250003.6A patent/CN103432572B/en active Active
-
2014
- 2014-06-20 WO PCT/CN2014/080414 patent/WO2014202024A1/en active Application Filing
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2005009119A1 (en) * | 2003-07-23 | 2005-02-03 | Mcw Research Foundation, Inc. | Rat model of diabetic nephropathy |
| CN1785433A (en) * | 2004-12-09 | 2006-06-14 | 上海凯曼生物科技有限公司 | Preparation method of Type II diabetes animal model |
| CN101095958A (en) * | 2007-07-04 | 2008-01-02 | 山西医科大学第一医院 | Method for preparing 2 type diabetes rat model |
| CN101637405A (en) * | 2008-07-31 | 2010-02-03 | 四川大学华西医院 | Method for preparing diabetic animal model |
| CN101804211A (en) * | 2009-02-13 | 2010-08-18 | 四川大学华西医院 | Establishment of rhesus autoimmune 1 type diabetes model |
Non-Patent Citations (7)
| Title |
|---|
| 匡德宣: "不同剂量链脲菌素对恒河猴某些常规生理指标的影响", 《中国实验动物学报》 * |
| 张艳春等: "高能量膳食诱导食蟹猴2型糖尿病模型的研究", 《动物医学进展》 * |
| 徐传磊等: "建立食蟹猴糖尿病模型的研究", 《广州中医药大学学报》 * |
| 朱华 等: "STZ诱导恒河猴糖尿病动物模型的建立", 《中国比较医学杂志》 * |
| 李璇 等: "链佐星-弗氏完全佐剂制作大鼠糖尿病肾病模型", 《南京中医药大学学报》 * |
| 王保伟 等: "高脂饲料喂养时间及链脲佐菌素剂量对实验型2型糖尿病大鼠造模的影响", 《卫生研究》 * |
| 赵艳霞: "糖尿病肾病合并模性肾病模型构建及其机制研究", 《中国优秀硕士学位论文全文数据库》 * |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014202024A1 (en) * | 2013-06-21 | 2014-12-24 | 四川大学华西医院 | Method for preparation of rhesus monkey diabetic nephropathy model |
| CN105726568A (en) * | 2016-04-11 | 2016-07-06 | 陕西科技大学 | Method for constructing diabetic nephropathy animal model |
| CN105726568B (en) * | 2016-04-11 | 2019-01-29 | 陕西科技大学 | A kind of construction method of diabetic nephropathy animal model |
| CN107469069A (en) * | 2017-08-18 | 2017-12-15 | 四川普莱美生物科技集团有限公司 | Modeling method, preparation, the preparation method of preparation, rhesus macaque pulmonary fibrosis model and its application of rhesus macaque pulmonary fibrosis model |
| CN115176758A (en) * | 2022-07-18 | 2022-10-14 | 四川省医学科学院.四川省人民医院实验动物研究所 | Method for establishing non-human primate type 1 diabetes model |
| CN115176758B (en) * | 2022-07-18 | 2024-03-26 | 四川省医学科学院.四川省人民医院实验动物研究所 | Method for establishing non-human primate model of type 1 diabetes |
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