CN106857274A - A kind of method for building up of dry eyes animal model - Google Patents
A kind of method for building up of dry eyes animal model Download PDFInfo
- Publication number
- CN106857274A CN106857274A CN201710094351.7A CN201710094351A CN106857274A CN 106857274 A CN106857274 A CN 106857274A CN 201710094351 A CN201710094351 A CN 201710094351A CN 106857274 A CN106857274 A CN 106857274A
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- muroid
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- support bar
- box
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- 208000003556 Dry Eye Syndromes Diseases 0.000 title claims abstract description 32
- 206010013774 Dry eye Diseases 0.000 title claims abstract description 32
- 238000010171 animal model Methods 0.000 title claims abstract description 29
- 238000000034 method Methods 0.000 title claims abstract description 27
- 208000010340 Sleep Deprivation Diseases 0.000 claims abstract description 23
- 239000007788 liquid Substances 0.000 claims abstract description 18
- 235000013305 food Nutrition 0.000 claims abstract description 8
- 230000006378 damage Effects 0.000 claims abstract description 4
- 239000003651 drinking water Substances 0.000 claims abstract description 4
- 235000020188 drinking water Nutrition 0.000 claims abstract description 4
- 235000021590 normal diet Nutrition 0.000 claims abstract description 4
- 239000000126 substance Substances 0.000 claims abstract description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 11
- 235000017166 Bambusa arundinacea Nutrition 0.000 claims description 2
- 235000017491 Bambusa tulda Nutrition 0.000 claims description 2
- 241001330002 Bambuseae Species 0.000 claims description 2
- 235000015334 Phyllostachys viridis Nutrition 0.000 claims description 2
- 239000011425 bamboo Substances 0.000 claims description 2
- 239000000463 material Substances 0.000 claims description 2
- 230000000737 periodic effect Effects 0.000 claims description 2
- 229920003023 plastic Polymers 0.000 claims description 2
- 239000004033 plastic Substances 0.000 claims description 2
- 229910001220 stainless steel Inorganic materials 0.000 claims description 2
- 239000010935 stainless steel Substances 0.000 claims description 2
- 239000008399 tap water Substances 0.000 claims description 2
- 235000020679 tap water Nutrition 0.000 claims description 2
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 2
- 239000012498 ultrapure water Substances 0.000 claims description 2
- 241001465754 Metazoa Species 0.000 description 11
- 210000002919 epithelial cell Anatomy 0.000 description 11
- 230000006698 induction Effects 0.000 description 7
- 230000028327 secretion Effects 0.000 description 7
- 210000004027 cell Anatomy 0.000 description 6
- 210000004087 cornea Anatomy 0.000 description 6
- 230000008569 process Effects 0.000 description 6
- 230000006907 apoptotic process Effects 0.000 description 4
- 230000008859 change Effects 0.000 description 4
- 238000001514 detection method Methods 0.000 description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 4
- 210000000981 epithelium Anatomy 0.000 description 4
- 206010054949 Metaplasia Diseases 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 239000007850 fluorescent dye Substances 0.000 description 3
- 210000004907 gland Anatomy 0.000 description 3
- 239000003550 marker Substances 0.000 description 3
- 230000015689 metaplastic ossification Effects 0.000 description 3
- 102100023970 Keratin, type I cytoskeletal 10 Human genes 0.000 description 2
- 101710183404 Keratin, type I cytoskeletal 10 Proteins 0.000 description 2
- 230000002159 abnormal effect Effects 0.000 description 2
- 210000004082 barrier epithelial cell Anatomy 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 230000004069 differentiation Effects 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 230000004890 epithelial barrier function Effects 0.000 description 2
- 210000002175 goblet cell Anatomy 0.000 description 2
- 238000003364 immunohistochemistry Methods 0.000 description 2
- 238000011835 investigation Methods 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 230000036385 rapid eye movement (rem) sleep Effects 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000007958 sleep Effects 0.000 description 2
- 101000972282 Homo sapiens Mucin-5AC Proteins 0.000 description 1
- 102100033420 Keratin, type I cytoskeletal 19 Human genes 0.000 description 1
- 101710183399 Keratin, type I cytoskeletal 19 Proteins 0.000 description 1
- 102000011782 Keratins Human genes 0.000 description 1
- 108010076876 Keratins Proteins 0.000 description 1
- 102100022496 Mucin-5AC Human genes 0.000 description 1
- 108010049420 RE1-silencing transcription factor Proteins 0.000 description 1
- 208000025865 Ulcer Diseases 0.000 description 1
- 230000006472 autoimmune response Effects 0.000 description 1
- 230000006399 behavior Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 208000035475 disorder Diseases 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000004043 dyeing Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 230000008556 epithelial cell proliferation Effects 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 229940020947 fluorescein sodium Drugs 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 230000030214 innervation Effects 0.000 description 1
- 206010023332 keratitis Diseases 0.000 description 1
- 210000000110 microvilli Anatomy 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000008506 pathogenesis Effects 0.000 description 1
- 230000003950 pathogenic mechanism Effects 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 230000001991 pathophysiological effect Effects 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 231100000167 toxic agent Toxicity 0.000 description 1
- 239000003440 toxic substance Substances 0.000 description 1
- 230000009261 transgenic effect Effects 0.000 description 1
- 230000001228 trophic effect Effects 0.000 description 1
- 231100000397 ulcer Toxicity 0.000 description 1
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
- A01K1/00—Housing animals; Equipment therefor
- A01K1/02—Pigsties; Dog-kennels; Rabbit-hutches or the like
- A01K1/03—Housing for domestic or laboratory animals
- A01K1/031—Cages for laboratory animals; Cages for measuring metabolism of animals
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Zoology (AREA)
- Environmental Sciences (AREA)
- Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Clinical Laboratory Science (AREA)
- General Health & Medical Sciences (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Acyclic And Carbocyclic Compounds In Medicinal Compositions (AREA)
Abstract
The invention discloses a kind of method for building up of dry eyes animal model, its equipment therefor includes the upper lid that a box-like body and a lid are located in box-like body, box-like body is built with the liquid for not producing chemical damage to muroid in right amount, the depth of liquid is long no more than the body of muroid, more than liquid level is provided with least support bar stood for muroid；Upper lid is provided with a food rest area and and feeds pool, for muroid normal diet；Specifically include following steps：(1) muroid experimental animal is placed on support bar, covers lid, place food and drinking water, and regularly replace the liquid；(2) above-mentioned muroid experimental animal is made to stand daily in the apparatus, the time of standing is the sleep deprivation time, and rest continuously repeats the step during the muroid experimental animal is put into common rearging cage by standing after terminating, you can obtain dry eyes animal model.The present invention makes simple, and repeatability is high, model stability.
Description
Technical field
The invention belongs to medical animal modelling technique field, and in particular to a kind of method for building up of dry eyes animal model.
Background technology
Dry eyes refer to cause tear film unstable and/or eye surface different due to tear quality and quantity and dynamic (dynamical) exception
Often, and with a class disease of ophthalmic uncomfortable symptom.Dry eyes can be divided into：Evaporated strong type dry eyes, aqueous shortage type dry eyes, glued
Abnormal protein type dry eyes, tear dynamics ectype dry eyes and mixed type dry eyes.Clinical investigation shows that dry eyes are common at present
Eye surface diseases, and secondary can cause keratitis, cornea rebirth blood vessel and ulcer of the cornea etc., have a strong impact on the quality of life of people
And operating efficiency.The common pathophysiological features of dry eye morbidity have ocular epithelial barrier to destroy, tear is reduced, Conjunctival Goblet Cells
Reduction, ocular epithelial squamous epithelial metaplasia etc..Because dry eye morbidity mechanism is complicated and with diversity, the hair on dry eyes at present
A series of bases of interpretation of the cause, onset and process of an illness system, diagnosis and treatment and clinical problem still need further further investigation.
The foundation of animal model is the pathogenesis for studying dry eyes, explores effective and practical means for the treatment of method.Mesh
It is preceding it has been reported that the method for building up of dry eyes animal model include：Made by trophic factor, environmental factor, toxicant, medicine
With, change animality hormone secretion level, the innervation of removal lachrymal gland or ocular, induction lachrymal gland produce autoimmune response,
Surgical removal lachrymal gland and transgenic animals etc..The foundation of these dry eyes animal models generally require technical conditions higher or compared with
Time long, and variability is larger.The use of different animals model is respectively provided with certain limitation, has with clinical practice situation
Certain gap.
The content of the invention
It is an object of the invention to overcome prior art defect, there is provided a kind of method for building up of dry eyes animal model.
Technical scheme is as follows：
A kind of method for building up of dry eyes animal model, its equipment therefor includes that a box-like body and a lid are located at box-like body
On upper lid, built with the liquid for not producing chemical damage to muroid in right amount, the depth of liquid is no more than muroid for box-like body
Body is long, more than liquid level is provided with least support bar stood for muroid；Upper lid is provided with a food rest area and and feeds water
Area, for muroid normal diet；Specifically include following steps：
(1) muroid experimental animal is placed on support bar, covers lid, place food and drinking water, and periodically more
Change the liquid；
(2) above-mentioned muroid experimental animal is made to stand daily in the apparatus 1~24h, the time of standing is sleep deprivation
Be put into the muroid experimental animal in common rearging cage after terminating rest by time, standing, is kept quite during rest, it is to avoid bother
Muroid experimental animal rests, and continuously repeats the step 1~100d, you can obtain dry eyes animal model.
In a preferred embodiment of the invention, the cross section of the support bar for circle, a diameter of 0.01~
10cm。
In a preferred embodiment of the invention, the material of the support bar be wooden, bamboo, plastics, irony or
Stainless steel.
In a preferred embodiment of the invention, the relatively described box-like body of the support bar is fixed.
In a preferred embodiment of the invention, the liquid is water.
It is further preferred that the liquid is ordinary tap water, pure water or ultra-pure water.
In a preferred embodiment of the invention, the liquid level in support bar distance be 1~5cm.
In a preferred embodiment of the invention, the time of the periodic replacement of the step (1) is 1~2d.
Beneficial effects of the present invention：
1st, the present invention allows muroid experimental animal to stand on the support bar close to the water surface, and animal is needed with certain pin grip
Balance could be kept on support bar, when animal enters rapid-eye-movement sleep, holotonia declines, immediately from support bar
Upper landing so that face be stained with or the water surface that is dipped into below support bar and wake up with a start suddenly, this behavior can repeated, and selectivity is broken
The rapid-eye-movement sleep of animal is broken, sleep deprivation is caused.
2nd, the present invention makes simple, and repeatability is high, model stability.
3rd, the dry eyes animal model that the present invention sets up can strictly control the time that sleep lacks, beneficial to grinding for the dry eyes course of disease
Study carefully, and inducement is single controllable, and other influences factor can be avoided to produce influence to experimental result.
4th, in model Induction Process, animal pattern engenders that the characteristic of ocular epithelial barrier destruction changes to the present invention
Become：Reduced (proved by methods such as HE dyeing, fluorescein sodium colorings) including corneal epithelial cell, cell arrangement is disorderly, cell
Connection reduces (methods such as the immunohistochemistry of mark are connected by cell to be proved), the reduction of epithelial cell proliferation ability and (leads to
Crossing the methods such as the immunohistochemistry of ability of cell proliferation mark proves).
5th, in model Induction Process, animal pattern conjunctival epithelial cell, corneal epithelial cell engender squama to the present invention
The pathological characteristicses of columnar epithelium metaplasia change：Including cell normal differentiation mark such as conjunctival epithelium Specific marker keratin
K19 expression is reduced, and the positive expression of abnormal differentiation mark, such as skin epithelial cell Specific marker keratin occurs
The expression of K10.
6th, in model Induction Process, the goblet cell in animal pattern conjunctival epithelial cell is gradually decreased the present invention, can
Confirmed with being dyeed with Specific marker MUC5AC immuning tissues.
8th, it is of the invention in model Induction Process, animal pattern corneal epithelial cell top layer apoptosis cells, and its number
Amount gradually increases.
9th, in model Induction Process, the aqueous lacrimal secretion of animal pattern is gradually decreased the present invention with time lengthening.
10th, the purposes of the model that the present invention sets up is to lack induction dry eye model by sleeping, and is conducive to dry eye morbidity
The research of mechanism；For being screened to Dry eye treatment related drugs and scheme, being identified；Lack phase for sleep deprivation or sleep
Research in terms of the pathomechanism of related disorders and treatment.
Brief description of the drawings
Fig. 1 is the structural representation of equipment therefor of the present invention.
Fig. 2 is the fluorescent staining figure of mouse ocular after sleep deprivation different time in the embodiment of the present invention 1.
Fig. 3 is the aqueous lacrimal secretion situation map of mouse after sleep deprivation different time in the embodiment of the present invention 1.
Fig. 4 is the corneal epithelial surfaces stereoscan photograph of mouse after sleep deprivation different time in the embodiment of the present invention 1.
Fig. 5 is the corneal epithelial cell apoptosis detection figure of mouse after sleep deprivation different time in the embodiment of the present invention 1.
Fig. 6 is the corneal epithelial cell Phenotypic examination figure of mouse after sleep deprivation different time in the embodiment of the present invention 1.
Specific embodiment
Technical scheme is further detailed and described below by way of specific embodiment combination accompanying drawing.
Embodiment 1：
A kind of method for building up of dry eyes animal model, its equipment therefor is (as shown in Figure 1) to include a box-like body and a lid
The upper lid in box-like body is located at, built with suitable quantity of water, the depth of water is long no more than the body of muroid, more than the water surface 1 for box-like body
~5cm is provided with two support bars stood for mouse (relatively described box-like body is fixed)；Upper lid is provided with food placement
Pool is fed in area and, for mouse normal diet；Specifically include following steps：
(1) mouse is placed on support bar, covers lid, place food and drinking water, and change every 1~2d
Water, to keep cleaning；
(2) mouse is made to stand daily in the apparatus, the time of standing is sleep deprivation time, every afternoon 13:00-
17:00 stands terminate after the muroid experimental animal is put into rest 4h in common rearging cage, kept quite during rest, it is to avoid beat
Mouse rest is disturbed, remaining time carries out sleep deprivation, after sleep deprivation different time, such as 1 day, 3 days, 5 days etc., entered to mouse
The fluorescent staining of row ocular, lacrimal secretion detection, breakup time of tear film detection, judge mouse dry eyes situation, you can obtain dry eyes animal
Model.
As shown in Figures 2 to 6, Fig. 2 is the fluorescent staining of mouse ocular after sleep deprivation different time to concrete outcome, is pointed out
With the extension of sleep deprivation time, the fluorescent color of mouse cornea epithelium is more and more.Fig. 3 is for after sleep deprivation different time
The aqueous lacrimal secretion situation of mouse, pointed out sleep deprivation after 2 days, and the aqueous lacrimal secretion of mouse is to decline nearly 50%, and is tieed up
Hold in low secretion level.Fig. 4 be sleep deprivation different time after mouse corneal epithelial surfaces scanning electron microscopic observation, point out with
The extension of sleep deprivation time, the microvillus of mouse cornea epithelial surface is shorter and shorter and more and more rare.Fig. 5 is sleep deprivation
The corneal epithelial cell apoptosis detection of mouse, points out with the extension of sleep deprivation time, mouse cornea epithelium after different time
The apoptosis of cell is in increase trend.Fig. 6 be sleep deprivation different time after mouse corneal epithelial cell Phenotypic examination, point out with
The extension of sleep deprivation time, the expression of mouse cornea epithelial cell K10 gradually strengthens, and shows as squamous metaplasia.
The above, only presently preferred embodiments of the present invention, therefore can not according to this limit the scope of present invention implementation, i.e.,
The equivalence changes made according to the scope of the claims of the present invention and description and modification, all should still belong in the range of the present invention covers.
Claims (8)
1. a kind of method for building up of dry eyes animal model, it is characterised in that：Its equipment therefor includes that a box-like body and a lid set
Upper lid in box-like body, built with the liquid for not producing chemical damage to muroid in right amount, the depth of liquid is or not box-like body
Body more than muroid is long, more than liquid level is provided with least support bar stood for muroid；Upper lid is provided with food placement
Pool is fed in area and, for muroid normal diet；Specifically include following steps：
(1) muroid experimental animal is placed on support bar, covers lid, place food and drinking water, and regularly replace institute
State liquid；
(2) above-mentioned muroid experimental animal is made to stand daily in the apparatus 1~24h, the time of standing is the sleep deprivation time,
Be put into the muroid experimental animal in common rearging cage after terminating rest by standing, is kept quite during rest, it is to avoid bother muroid
Experimental animal rests, and continuously repeats the step 1~100d, you can obtain dry eyes animal model.
2. method for building up as claimed in claim 1, it is characterised in that：The cross section of the support bar is circle, a diameter of
0.01~10cm.
3. method for building up as claimed in claim 1, it is characterised in that：The material of the support bar be wooden, bamboo, plastics,
Irony or stainless steel.
4. method for building up as claimed in claim 1, it is characterised in that：The relatively described box-like body of the support bar is fixed.
5. method for building up as claimed in claim 1, it is characterised in that：The liquid is water.
6. method for building up as claimed in claim 5, it is characterised in that：The liquid is ordinary tap water, pure water or ultra-pure water.
7. method for building up as claimed in claim 1, it is characterised in that：The liquid level is 1~5cm in the distance of support bar.
8. method for building up as claimed in claim 1, it is characterised in that：The time of the periodic replacement of the step (1) be 1~
2d。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710094351.7A CN106857274A (en) | 2017-02-21 | 2017-02-21 | A kind of method for building up of dry eyes animal model |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710094351.7A CN106857274A (en) | 2017-02-21 | 2017-02-21 | A kind of method for building up of dry eyes animal model |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106857274A true CN106857274A (en) | 2017-06-20 |
Family
ID=59167261
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710094351.7A Pending CN106857274A (en) | 2017-02-21 | 2017-02-21 | A kind of method for building up of dry eyes animal model |
Country Status (1)
| Country | Link |
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| CN (1) | CN106857274A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110558240A (en) * | 2019-10-11 | 2019-12-13 | 浙江中医药大学 | Sleep deprivation instrument and establishment method applied to internal heat animal model |
| CN110618276A (en) * | 2019-06-21 | 2019-12-27 | 厦门大学 | Molecular marker set for detecting sleep deprivation-induced dry eye and application thereof |
| CN113424772A (en) * | 2021-06-18 | 2021-09-24 | 李鹏军 | Animal sleep deprivation device |
-
2017
- 2017-02-21 CN CN201710094351.7A patent/CN106857274A/en active Pending
Non-Patent Citations (4)
| Title |
|---|
| S.HAKKI ONEN,ETC.: "Effects of rapid eye movement (REM) sleep deprivation on pain sensitivity in the rat", 《BRAIN RESEARCH》 * |
| YOUNG BOK LEE, ETC.: "Sleep Deprivation Reduces Tear Secretion and Impairs the Tear Film", 《IOVS》 * |
| 张维胜等: "水池平台技术在睡眠研究中的应用", 《临床精神医学杂志》 * |
| 李德明: "睡眠剥夺的心理生理影响", 《心理学动态》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110618276A (en) * | 2019-06-21 | 2019-12-27 | 厦门大学 | Molecular marker set for detecting sleep deprivation-induced dry eye and application thereof |
| CN110558240A (en) * | 2019-10-11 | 2019-12-13 | 浙江中医药大学 | Sleep deprivation instrument and establishment method applied to internal heat animal model |
| CN110558240B (en) * | 2019-10-11 | 2022-02-11 | 浙江中医药大学 | Sleep deprivation instrument and establishment method applied to internal heat animal model |
| CN113424772A (en) * | 2021-06-18 | 2021-09-24 | 李鹏军 | Animal sleep deprivation device |
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