CN107753138A - A kind of construction method for the method and its animal model for assessing micro- osteogenic action - Google Patents
A kind of construction method for the method and its animal model for assessing micro- osteogenic action Download PDFInfo
- Publication number
- CN107753138A CN107753138A CN201711101302.8A CN201711101302A CN107753138A CN 107753138 A CN107753138 A CN 107753138A CN 201711101302 A CN201711101302 A CN 201711101302A CN 107753138 A CN107753138 A CN 107753138A
- Authority
- CN
- China
- Prior art keywords
- micro
- animal model
- blood
- osteogenic action
- trace element
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Abstract
The invention discloses a kind of construction method for the method and its animal model for assessing micro- osteogenic action.The construction method that the present invention assesses the animal model of micro- osteogenic action comprises the following steps：(1) solution containing preset concentration trace element ion is prepared；(2) rat calvarium is for hole；(3) blood is taken from rat tailses, and the blood of acquirement is mixed with the solution containing aimed concn trace element ion made from step (1), form blood clot；(4) blood clot is placed at skull defeci, sewing-up cut, obtains assessing the animal model of micro- osteogenic action.The present invention is used as ionophore using animal autoblood, the blood clot containing preset concentration ion can be prepared, exclude to apply influence of the bone alternate material as ionophore in existing model simultaneously, by the skeletonization effect for detecting Cranial defect, and then influence of the aimed concn trace element to osteanagenesis process is assessed, method is effective, easy.
Description
Technical field
The present invention relates to a kind of animal evaluation models, more particularly to a kind of method for assessing micro- osteogenic action and its
The construction method of animal model.
Background technology
Trace element containing numerous kinds in the bone tissue of human body, such as iron, zinc, copper, manganese, chromium, selenium, molybdenum, cobalt, fluorine.
These trace elements are much present in bone tissue in the form of an ion, and play weight to osteanagenesis, reparation, reconstruction and pathologic process
The adjustment effect wanted.On the other hand, bone alternate material, implant surface modification are carried out using trace element ion, makes bone biological
Material possesses more preferable bone guided and osteoinductive, is a kind of conventional material modification means.These yuan are usually necessarily to compare
Example is doped into bone-grafting material or planting body shows, on the premise of material basic structure characteristic is not changed, in mechanical performance, dissolving
Degree, porosity, biocompatibility etc. play important modifying function.Therefore, understand trace element ion to bone tissue again
The concentration dependant effect of raw process, for disclosing the osteogenic action effect of various concentrations ion, illustrating ion pair osteanagenesis process
Mechanism of action and explore and prepare more excellent ion modification bone alternate material, there is important science and application value.
In the influence for assessing various dose ion pair osteanagenesis process, it usually needs substitute material by some form of bone
Material carries out ion carrying, such as bone meal, ossein, artificial synthesized support.Carrying process generally includes, by ion load to these
Carrier inside, its release performance is determined, then it is placed at animal Cranial defect.By one section of healing time, material is taken out, entered
Row RNA, Protein Detection, sections observation etc., with the osteogenic action of assessment material, and then infer the effect for carrying ion.But
Because material support belongs to allosome material together in itself, the change of its surface property, porosity, biocompatibility be possible to influence into
Bone process, and material modified ion release in vivo can not accurately be measured by experiment in vitro, therefore, above method can not
The concentration dependent for studying ion pair osteanagenesis process is embodied well.
On the other hand, the influence for detecting ion pair skeletonization can also be by experiment in vitro, by the culture medium of different ions concentration
Skeletonization relevant cell is acted on, detects the change of its Bone formation-related gene, albumen, but experiment in vitro environment and the big phase of vivo environment
It is very unlike, often can not the effect of real simulation ion in vivo, reference value is relatively low.To sum up, ion pair skeletonization is detected merely
Influence there is no reliable method now.
The content of the invention
It is an object of the invention to a kind of micro- osteogenic action of assessment is provided in place of overcome the deficiencies in the prior art
Method and its animal model construction method.
To achieve the above object, the technical scheme taken of the present invention is：A kind of animal for assessing micro- osteogenic action
The construction method of model, it is comprised the steps of：
(1) solution containing preset concentration trace element ion is prepared；
(2) rat calvarium is for hole；
(3) take blood from rat tailses, and will the blood of acquirement and step (1) it is obtained containing aimed concn trace element from
The solution mixing of son, forms blood clot；
(4) blood clot is placed at skull defeci, sewing-up cut, obtains assessing the animal mould of micro- osteogenic action
Type.
As the preferred embodiment of the present invention, trace element in the step (1) is iron, zinc, copper, manganese, chromium, selenium,
Molybdenum, cobalt or fluorine etc..
As the preferred embodiment of the present invention, in the solution containing aimed concn trace element ion, solvent is
Ultra-pure water.
As the preferred embodiment of the present invention, take bone drill standby to rat progress calvarium using 5mm in the step (2)
Hole.
As the preferred embodiment of the present invention, the forming method of blood clot is in the step (3)：Target rich will be contained
After the solution of degree trace element ion is well mixed with blood, it is stored at room temperature, forms blood clot.
As the preferred embodiment of the present invention, in the step (3), the solution containing aimed concn trace element ion
Volume ratio with blood is 1:9；Such as：The wherein volume of trace element ion solution is 20 μ l, the rat tailses blood of collection
Volume be 180 μ l.
The present invention also provides a kind of method for assessing micro- osteogenic action, and methods described is：Using in claim
State the animal model that method structure assesses micro- osteogenic action, according to setting time point by the blood clot in animal model or
Skull defeci position is taken out, and carries out skeletonization, osteoclastic correlated performance characterizes.
As the preferred embodiment of the method for the present invention for assessing micro- osteogenic action, the correlated performance table
Levy in the amount formed for the expression of detection skeletonization and osteoclastic related gene, the new bone of observation or osteoid and tissue morphology at least
It is a kind of.
Compared with prior art, beneficial effects of the present invention are：The present invention is by by aimed concn trace element ion
Solution is mixed with blood, after standing forms blood clot, is placed at Cranial defect, and blood clot or skull group are taken out in particular point in time
Knit, by carrying out efficiency evaluation to the osteogenic action of animal model the methods of qRT-PCR and Micro-CT, can not only so prepare
Blood clot containing certain concentration ion, by detecting the skeletonization effect of Cranial defect, and then it is right to assess aimed concn trace element
The influence of osteanagenesis process, method are effective, easy.Meanwhile method of the invention can also exclude to substitute using bone in existing model
Influence of the material as ionophore, detect concentration dependent of the simple trace element to osteanagenesis process.
Brief description of the drawings
Fig. 1 is surgical procedure of blood clot of the present invention implantation containing trace element ion solution at Rat calvarial defect
Figure；
Fig. 2 is that the Micro-CT at the postoperative 4 weeks Rat calvarial defects of the present invention rebuilds figure；
Fig. 3 is the postoperative 2 days rat blood clots BMP2 of present invention gene expression dose figure；
Fig. 4 is the postoperative 2 days rat blood clots BMP6 of present invention gene expression dose figure；
Fig. 5 is the postoperative 2 days rat blood clots VEGF of present invention gene expression dose figure；
Fig. 6 is the postoperative 2 days rat blood clots RANKL of present invention gene expression dose figure；
Fig. 7 is the postoperative 2 days rat blood clots MCSF of present invention gene expression dose figure；
Fig. 8 is the postoperative 2 days rat blood clots OPG of present invention gene expression dose figure.
Embodiment
For the object, technical solutions and advantages of the present invention are better described, below in conjunction with the drawings and specific embodiments pair
The present invention further illustrates.
Embodiment 1
The method that the present embodiment assesses micro- osteogenic action comprises the following steps：
(1) the micro- solion containing aimed concn is prepared
Trace element is fluorine, and concentration of sodium fluoride is respectively concentration A in the Fluorinse of preparation:1mg/L, concentration B：
10mg/L, control group：0mg/L (i.e. control group is ultra-pure water).
(2) rat calvarium is for hole；
The healthy experimental rat of 6-8 weeks is taken, 1% amobarbital 0.4-0.5ml/100g anesthetized rats are injected intraperitoneally；Shave
Head hair；Row stringer otch after iodophor disinfection, soft tissue is separated to expose calvarium；Take bone drill in calvarium for hole with 5mm.
(3) rat tailses take blood, and the blood of acquirement is mixed with the solion containing aimed concn trace element, formed
Blood clot；
2ml front end of the syringe needle is cut off with scissors；Iodophor disinfection rat tailses, tail point 1-2mm is cut off, is received with front end of the syringe needle
Collect the μ l of rat tailses blood 180；20 μ l fluoride solutions are added, and are well mixed, are stored at room temperature about 10min, form blood clot.
(4) blood clot is placed at skull defeci, sewing-up cut.
Gauze removes the serum on blood clot, by the small heart tamponade of blood clot at skull defeci, sewing-up cut.
Surgical procedure in wherein step 1-4 is as shown in figure 1, A-H is expressed as：A. experimental rat；B. art area exposes；
C-D. skull defeci hole shape is prepared；E. afterbody takes blood；F. it is mixed into solion to be measured；G. blood clot is placed；H. wound sutures.
(5) efficiency evaluation of the animal model of micro- skeletonization effect is assessed
A, put to death rat within postoperative 4 weeks, animal skull tissue is taken out, fixed using 4% paraformaldehyde, row micro-CT is swept
Retouch and three-dimensional reconstruction, observe skeletonization situation at skull defeci.As a result as shown in Fig. 2 the different skeletonization situations organized at skull defeci
Notable ion concentration dependence is presented, A, B composition bone situation are significantly better than control group and (refer to the solion concentration in blood clot
Be zero, i.e. ultra-pure water), illustrate that this In vivo model can preferably embody ion skeletonization situation.
B, after being implanted into 48 hours, animal is put to death, coating is cut, experimental group and control group part lack of skull blood clot is taken
Go out, and extract RNA therein, carry out qRT-PCR, detection regulation and control skeletonization (BMP-2, BMP-6, VEGF), it is osteoclastic (RANKL,
MCSF, OPG) related gene expression.As a result as shown in figures 3-8, * represents P in figure<0.05, * * represents P<0.01, * * *
Represent P<0.001.From Fig. 3-8, A composition bone related genes BMP2, BMP6, VEGF and osteoclastic related gene RANKL, MCSF
Significantly rise, B composition bone related genes BMP2, BMP6 and suppresses osteoclastic gene OPG and significantly raises, prompt the fluorine of A group concentration from
The Osteoblastic activity that son promotes is driven by BMP2, BMP6, VEGF, while osteoclastic reconstruction activity enlivens, and the fluorine ion of B group concentration promotes
Osteoblastic activity driven by BMP2, BMP6, while osteoclastic activity is suppressed significantly, i.e., based on simple Osteoblastic activity, this is explains
Physiological mechanism under bright fluorine ion skeletonization concentration dependent provides foundation.
Finally, it should be noted that the above embodiments are merely illustrative of the technical solutions of the present invention rather than the present invention is protected
The limitation of scope is protected, although being explained in detail with reference to preferred embodiment to the present invention, one of ordinary skill in the art should
Understand, technical scheme can be modified or equivalent substitution, without departing from the essence of technical solution of the present invention
And scope.
Claims (8)
1. a kind of construction method for the animal model for assessing micro- osteogenic action, it is characterised in that comprise the steps of：
(1) solution containing preset concentration trace element ion is prepared；
(2) rat calvarium is for hole；
(3) blood is taken from rat tailses, and aimed concn trace element ion will be contained made from the blood of acquirement and step (1)
Solution mixes, and forms blood clot；
(4) blood clot is placed at skull defeci, sewing-up cut, obtains assessing the animal model of micro- osteogenic action.
2. the construction method of the animal model of micro- osteogenic action is assessed according to claim 1, it is characterised in that institute
It is iron, zinc, copper, manganese, chromium, selenium, molybdenum, cobalt or fluorine to state the trace element in step (1).
3. the construction method of the animal model of micro- osteogenic action is assessed according to claim 1, it is characterised in that institute
State in the solution containing aimed concn trace element ion, solvent is ultra-pure water.
4. the construction method of the animal model of micro- osteogenic action is assessed according to claim 1, it is characterised in that institute
State in step (2) takes bone drill to carry out calvarium to rat for hole using 5mm.
5. the construction method of the animal model of micro- osteogenic action is assessed according to claim 1, it is characterised in that institute
The forming method for stating blood clot in step (3) is：Solution containing aimed concn trace element ion is well mixed with blood
Afterwards, it is stored at room temperature, forms blood clot.
6. the construction method of the animal model of micro- osteogenic action is assessed according to claim 1, it is characterised in that institute
State in step (3), the volume ratio of solution and blood containing aimed concn trace element ion is 1:9.
A kind of 7. method for assessing micro- osteogenic action, it is characterised in that methods described is：Using claim 1~6 times
One methods described structure assesses the animal model of micro- osteogenic action, according to setting time point by the blood in animal model
Grumeleuse or skull defeci position are taken out, and carry out skeletonization, osteoclastic correlated performance characterizes.
8. the method according to claim 7 for assessing micro- osteogenic action, it is characterised in that the correlated performance table
Levy in the amount formed for the expression of detection skeletonization and osteoclastic related gene, the new bone of observation or osteoid and tissue morphology at least
It is a kind of.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711101302.8A CN107753138A (en) | 2017-11-09 | 2017-11-09 | A kind of construction method for the method and its animal model for assessing micro- osteogenic action |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711101302.8A CN107753138A (en) | 2017-11-09 | 2017-11-09 | A kind of construction method for the method and its animal model for assessing micro- osteogenic action |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107753138A true CN107753138A (en) | 2018-03-06 |
Family
ID=61273346
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201711101302.8A Pending CN107753138A (en) | 2017-11-09 | 2017-11-09 | A kind of construction method for the method and its animal model for assessing micro- osteogenic action |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107753138A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113711993A (en) * | 2021-09-01 | 2021-11-30 | 杭州越波生物科技有限公司 | Method for quantitatively evaluating bone regeneration repair capacity of rat skull after bone defect by utilizing Micro-CT |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5171579A (en) * | 1991-10-11 | 1992-12-15 | Genetics Institute, Inc. | Formulations of blood clot-polymer matrix for delivery of osteogenic proteins |
| CN1471412A (en) * | 2000-06-29 | 2004-01-28 | ����ϳɼ������ô�˾ | Composition and method for the repair and regeneration of cartilage and other tissues |
| US20110293581A1 (en) * | 2009-01-16 | 2011-12-01 | Jun Lee | Bone-regenerating composition containing angiogenin |
-
2017
- 2017-11-09 CN CN201711101302.8A patent/CN107753138A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5171579A (en) * | 1991-10-11 | 1992-12-15 | Genetics Institute, Inc. | Formulations of blood clot-polymer matrix for delivery of osteogenic proteins |
| CN1471412A (en) * | 2000-06-29 | 2004-01-28 | ����ϳɼ������ô�˾ | Composition and method for the repair and regeneration of cartilage and other tissues |
| US20110293581A1 (en) * | 2009-01-16 | 2011-12-01 | Jun Lee | Bone-regenerating composition containing angiogenin |
Non-Patent Citations (5)
| Title |
|---|
| 《刘庆思名老中医传承精粹》编委会编: "《刘庆思名老中医传承精粹》", 31 August 2013, 广东人民出版社 * |
| 封伟等: "微量元素在种植体骨结合中应用研究进展", 《中国实用口腔科杂志》 * |
| 谭祖键等: "复方骨基质胶修复骨缺损实验研究", 《中华创伤杂志》 * |
| 郑铁生等: "《临床生物化学检验》", 31 August 2015, 北京：中国医药科技出版社 * |
| 郭现辉等: "自体静脉血凝块和异体骨修复兔桡骨缺损的比较", 《承德医学院学报》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113711993A (en) * | 2021-09-01 | 2021-11-30 | 杭州越波生物科技有限公司 | Method for quantitatively evaluating bone regeneration repair capacity of rat skull after bone defect by utilizing Micro-CT |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107921062A (en) | Tissue repair and renovation process | |
| CN106421880B (en) | A kind of Sepia polysaccharide/chitosan styptic sponge and its preparation method and application | |
| CN107224617B (en) | Hydrogel taking spleen extracellular matrix as raw material and preparation method thereof | |
| Wang et al. | Effects of local infiltration of insulin around titanium implants in diabetic rats | |
| CN102266585B (en) | Biological composite patch for female pelvic floor and manufacturing method thereof | |
| CN106075582A (en) | A kind of engineering blood vessel support and construction method thereof | |
| AU2011380375A1 (en) | Method for preparing edge-rigidized artificial biological valve | |
| CN106729984A (en) | A kind of Isin glue collagen repairs sponge and preparation method thereof | |
| CN104911427A (en) | Mg-Ca-Sr-Zn magnesium alloy as well as preparation method and application thereof | |
| CN106823008A (en) | Bone renovating material and preparation method thereof | |
| CN104225667A (en) | Angiogenesis-facilitating temperature-sensitive hydrogel powder and temperature-sensitive hydrogel prepared from same | |
| Szarek et al. | Lizard tail spinal cord: a new experimental model of spinal cord injury without limb paralysis | |
| CN107753138A (en) | A kind of construction method for the method and its animal model for assessing micro- osteogenic action | |
| CN104971386B (en) | Silk-fibroin timbering material and preparation method thereof | |
| CN110354307A (en) | Based on the albumen sericin gel and its preparation method and application for turning vectors containing human platelet-derived growth gene silk | |
| Hayakawa et al. | Facial nerve regeneration with bioabsorbable collagen conduits filled with collagen filaments: An experimental study | |
| CN110038162A (en) | It is a kind of to have the function of modulating vascular cell growth effect silk fibroin material and preparation method thereof | |
| CN103721294A (en) | Quick construction preparation method of human epidermal tissues | |
| CN110124104A (en) | A kind of composition being used to prepare myocardium sticking patch and its application | |
| CN109867802A (en) | A kind of preparation method of thermal response Extracellular Matrix for Replacement of Bladder bioactivity hydrogel | |
| JP4526152B2 (en) | Prothrombin time measuring reagent | |
| CN106317446A (en) | Preparation method of silk fiber fibroin hemostasis material | |
| Beck-Broichsitter et al. | Endocultivation: continuous application of rhBMP-2 via mini-osmotic pumps to induce bone formation at extraskeletal sites | |
| CN110478089A (en) | A kind of vascularization neuralization osteogenic activity bracket suitable for large segmental bone defect reparation | |
| Cornel et al. | Homeostasis of blood parameters and inflammatory markers analysis during bone defect healing after scaffolds implantation in mice calvaria defects |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20180306 |
|
| RJ01 | Rejection of invention patent application after publication |