CN106618566A - Method for positioning chronic-spinal-cord-injury scar tissue - Google Patents
Method for positioning chronic-spinal-cord-injury scar tissue Download PDFInfo
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- CN106618566A CN106618566A CN201510740062.0A CN201510740062A CN106618566A CN 106618566 A CN106618566 A CN 106618566A CN 201510740062 A CN201510740062 A CN 201510740062A CN 106618566 A CN106618566 A CN 106618566A
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- spinal cord
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Abstract
The invention discloses a method for positioning chronic-spinal-cord-injury scar tissue. The method includes the steps that 1) the scar tissue at animal spinal-cord injuries is generally positioned in cooperation with characteristics such as spinal cord structures and the hardness; 2) the head-close end and the tail-close end of the scar tissue at the animal spinal-cord injuries are positioned, and limits of the scar tissue at the animal spinal-cord injuries and normal spinal-cord tissue are determined. When the head-close end of the scar tissue is positioned, a receiving electrode is put on the head of an animal, an exciting electrode is put at the animal spinal-cord injury, and moves in the animal head direction, and the limit of the spinal-cord tissue and the scar tissue is judged through whether a somatosensory evoked potential exists or not; when the tail-close end of the scar tissue is positioned, the receiving electrode is put at animal lower limb muscles, the exciting electrode is put at the animal spinal-cord injury, and moves in the animal lower limb direction, and the limit of the spinal-cord tissue and the scar tissue is judged through whether a motor evoked potential exists or not.
Description
Technical field
The present invention relates to a kind of method of positioning chronic spinal cord lesion cicatricial tissue in biomedical sector.
Background technology
Spinal cord is the important component part of nervous centralis, and major function is to transmit the nerve information between brain and peripheral nerve,
It is also simultaneously the low-level center of many simple reflex activities.Spinal cord injury is mostly traumatic, is common in traffic accident, pounds
Wound, tumble injury, motional injury and some such as earthquake, mine disaster natural disasters.After spinal cord injury, due to damage and its
Secondary a series of pathologic-physiological reaction, including hypoxic-ischemic and organism immune response etc., cause aixs cylinder and myelin
Decompose, or even pericaryon is dead, causes gradual nerve fiber to be lost.Meanwhile, the star-like colloid of damage zone
The activation such as cell, microglia and fibroblast, propagation and extracellular matrix secretion composition, form scar group
Knit, these cicatricial tissues form barrier at damage field edge.These cicatricial tissues are damaging different times with difference
Effect.In the acute stage damaged, these cicatricial tissues can suppress the further diffusion for damaging；And in the urgency damaged
Property the phase after (several weeks are to the several months) cicatricial tissue suppress on the contrary nerve reparation, not only itself can be used as a kind of physics screen
Barrier hinders nerve fiber regeneration, and its extracellular matrix composition chondroitin sulfate proteoglycan for including
(chondroitin sulfate proteoglycan, CSPG) can suppress nerve as a kind of axon regeneration Inhibitory molecules
Regeneration.After spinal cord injury, cicatricial tissue is connected with the nerve fiber of surrounding normal, therefore how to judge scar
Tissue, is to rebuild the microenvironment for being conducive to spinal cord injury to repair to promote spinal cord injury so as to the excision cicatricial tissue of safety
The major issue that reparation faces.
The content of the invention
The technical problem to be solved is how to position the cicatricial tissue at chronic spinal cord lesion.
For solve above-mentioned technical problem, present invention firstly provides at animal spinal cord injury cicatricial tissue localization method.
The localization method of cicatricial tissue at animal spinal cord injury provided by the present invention, including it is following 1) and 2)：
1) general location is carried out with reference to cicatricial tissue at following character pair animal spinal cord injury：At the animal spinal cord injury
Normal spinal cord be in bean curd shape, with normal spinal cord structure；Cicatricial tissue week at the animal spinal cord injury
Enclose quality fine and close partially hard, in the middle of cicatricial tissue or have slough or a cavity；
2) near head end of cicatricial tissue at the animal spinal cord injury is positioned and to described using electro physiology detector
The caudal part of cicatricial tissue is positioned at animal spinal cord injury, so that it is determined that cicatricial tissue and ridge at animal spinal cord injury
The boundary of myeloid tissue, completes the positioning of cicatricial tissue at animal spinal cord injury；
The near head end to cicatricial tissue at animal spinal cord injury carries out positioning to be included：Receiving electrode is placed in described dynamic
The head of thing, stimulating electrode is placed at the animal spinal cord injury and along described to the movement of the direction of the animal head
Stimulating electrode, judges the boundary of myeloid tissue and cicatricial tissue, if there is body-sensing by the presence or absence of SEP
Evoked ptential, at the stimulating electrode Normal spinal cord is organized as, if without SEP, the thorn
Cicatricial tissue is organized as at sharp electrode；
The caudal part to cicatricial tissue at animal spinal cord injury carries out positioning to be included：Receiving electrode is placed in described dynamic
At the lower limb muscles or anal sphincter of thing, stimulating electrode is placed at the animal spinal cord injury and along to the animal
The stimulating electrode is moved in the direction of lower limb, judges myeloid tissue with cicatricial tissue by the presence or absence of Motion Evoked Potential
Boundary, if there is Motion Evoked Potential, at the stimulating electrode Normal spinal cord is organized as, if do not transported
Dynamic Evoked ptential, at the stimulating electrode cicatricial tissue is organized as.
In said method, the electro physiology detector can be myoelectric apparatus.The myoelectric apparatus concretely Beijing Wei Dikanghong
Medical science and technology Co., Ltd product, model Keypoint 9033A07.
In said method, the animal can be mammal.The mammal can be dog or people.The dog can be ratio
Lattice dog.
To solve above-mentioned technical problem, present invention also offers the cutting method of animal spinal cord injury cicatricial tissue.
The cutting method of animal spinal cord injury cicatricial tissue provided by the present invention, including using the animal spinal cord injury
The localization method of cicatricial tissue determines the border of animal spinal cord injury cicatricial tissue, and the method excision using operation is described dynamic
Thing spinal cord injury cicatricial tissue.
In the cutting method of above-mentioned animal spinal cord injury cicatricial tissue, the animal can be mammal.The lactation is moved
Thing can be dog or people.The dog can be beasle dog.
To solve above-mentioned technical problem, present invention also offers the electro physiology detector is being prepared for positioning animal ridge
Application in the device of marrow injury region cicatricial tissue.
Above-mentioned electro physiology detector is used to position in the application at animal spinal cord injury in the device of cicatricial tissue in preparation,
Described device also includes recording the carrier of herein below：The localization method of cicatricial tissue includes right at animal spinal cord injury
The near head end of cicatricial tissue is positioned and the caudal part of cicatricial tissue at animal spinal cord injury is entered at animal spinal cord injury
Row positioning, so that it is determined that cicatricial tissue and the boundary of myeloid tissue at animal spinal cord injury, complete at animal spinal cord injury
The positioning of cicatricial tissue；
The near head end to cicatricial tissue at animal spinal cord injury carries out positioning to be included：Receiving electrode is placed in described dynamic
The head of thing, stimulating electrode is placed at the animal spinal cord injury and along described to the movement of the direction of the animal head
Stimulating electrode, judges the boundary of myeloid tissue and cicatricial tissue, if there is body-sensing by the presence or absence of SEP
Evoked ptential, at the stimulating electrode Normal spinal cord is organized as, if without SEP, the thorn
Cicatricial tissue is organized as at sharp electrode；
The caudal part to cicatricial tissue at animal spinal cord injury carries out positioning to be included：Receiving electrode is placed in described dynamic
At the lower limb muscles or anal sphincter of thing, stimulating electrode is placed at the animal spinal cord injury and along to the animal
The stimulating electrode is moved in the direction of lower limb, judges myeloid tissue with cicatricial tissue by the presence or absence of Motion Evoked Potential
Boundary, if there is Motion Evoked Potential, at the stimulating electrode Normal spinal cord is organized as, if do not transported
Dynamic Evoked ptential, at the stimulating electrode cicatricial tissue is organized as.
It is demonstrated experimentally that the cicatricial tissue using the localization method of cicatricial tissue at the animal spinal cord injury of the present invention to animal
After being positioned and being carried out the excision of cicatricial tissue, it is not changed in the behaviouristics of the front animal of cicatricial tissue excision, feels
Do not change with plane of movement, the resection operation for showing cicatricial tissue does not cut off normal myeloid tissue, not to normal yet
Myeloid tissue cause damage, beasle dog is not had undesirable effect, illustrate the excision hand for being successfully made cicatricial tissue
Art.And the mark CSPG of cicatricial tissue is positive in the cicatricial tissue for cutting off, show that the cicatricial tissue for cutting off is scar
Tissue；And neurofilament NF coloration results find not finding that neurofilament NF is dyeed in the cicatricial tissue for cutting off, show what is cut off
Neurofilament is not contained in cicatricial tissue.It is demonstrated experimentally that using the positioning of cicatricial tissue at the animal spinal cord injury of the present invention
Method can successfully be positioned to the cicatricial tissue of animal, and can carry out cutting for cicatricial tissue according to the positioning result
Remove.
Description of the drawings
The CSPG testing results of cicatricial tissues of the Fig. 1 to cut off in the resection operation of beasle dog cicatricial tissue.
The NF coloration results of cicatricial tissues of the Fig. 2 to cut off in the resection operation of beasle dog cicatricial tissue.
The CSPG testing results of the cicatricial tissue cut off in the resection operation of Fig. 3 behaviour cicatricial tissues.
The NF coloration results of the cicatricial tissue cut off in the resection operation of Fig. 4 behaviour cicatricial tissues.
Specific embodiment
The present invention is further described in detail with reference to specific embodiment, the embodiment for being given only for
The present invention is illustrated, rather than in order to limit the scope of the present invention.
Experimental technique in following embodiments, if no special instructions, is conventional method.
Material used, reagent etc. in following embodiments, if no special instructions, commercially obtain.
Beasle dog in following embodiments is Nanjing An Limo Science and Technology Ltd.s Products.
The positioning of cicatricial tissue and excision at embodiment 1, beasle dog spinal cord injury
From 13 1-1.5 one full year of life female beagle dogs, body weight 7-9kg.
First, the excision of cicatricial tissue
After beasle dog is anaesthetized, T7-T9 vertebral plates exposing spinal cord tissue is opened, the spinal cord of T8 section beasle dogs is cut off into 2-3mm
(due to spinal cord retraction, the length between two broken ends of fractured bone is about 5mm), obtain the full transection lesion model of dog acute spinal.Will
The same day for carrying out Transected Spinal Cord trauma surgery is designated as operation 0 day.It is postoperative to carry out anti-inflammation routine care, Clean room
Interior raising.
After operation 2 months, the positioning and resection operation of cicatricial tissue are carried out, concrete grammar is as follows：Beasle dog is anaesthetized,
T7-T9 vertebral plates exposing spinal cords tissue is opened, the positioning for animal spinal cord injury cicatricial tissue being carried out substantially using naked eyes,
Myeloid tissue is in bean curd shape, and color whitens, with normal spinal cord structure；Quality is fine and close partially hard around cicatricial tissue,
There are slough or cavity in cicatricial tissue centre, and color is partially red.Then using myoelectric apparatus, (Beijing Wei Dikanghong is medical
Science and Technology Ltd.'s product, model Keypoint 9033A07) cicatricial tissue is further positioned：Contrast
The near head end of cicatricial tissue carries out positioning at lattice dog spinal cord injury includes：Receiving electrode is placed in into the head of beasle dog, will
Stimulating electrode is placed at beasle dog spinal cord injury and moves stimulating electrode along to the direction of beasle dog head, is lured by body-sensing
The presence or absence of generating position judges the boundary of myeloid tissue and cicatricial tissue, if there is SEP, at stimulating electrode
Be organized as myeloid tissue, if without SEP, at stimulating electrode cicatricial tissue is organized as；Contrast lattice
The caudal part of cicatricial tissue carries out positioning at dog spinal cord injury includes：Receiving electrode is placed at the lower limb muscles of beasle dog,
Stimulating electrode is placed at beasle dog spinal cord injury and stimulating electrode is moved along to the direction of beasle dog lower limb, by motion
The presence or absence of Evoked ptential judges the boundary of myeloid tissue and cicatricial tissue, if there is Motion Evoked Potential, stimulating electrode
Place is organized as myeloid tissue, if without Motion Evoked Potential, at stimulating electrode cicatricial tissue is organized as.
2nd, the detection of cicatricial tissue excision
1st, beasle dog neurological deficit score
To further determine that whether the excision of cicatricial tissue causes wound to beasle dog, in beasle dog cicatricial tissue hand is cut off
Beasle dog neurological deficit score is carried out respectively within 7 days within preoperative 1 day with Post operation.In evaluation process, beasle dog is opened at one
Move freely in the environment put, scored by two observers.Observer is familiar with score by rules but does not know the hand of each beasle dog
Art situation.It is divided into 0-14 levels, 14 points is that function is normal, and 0 point completely loses for function.Olby standards of grading
Such as table 1, appraisal result such as table 2.
Table 1, Olby standards of grading
| Fraction | Behavior |
| 0 | Without any hind limb motor or deep pain |
| 2 | Without any hind limb motor, but afterbody autogenic movement |
| 3 | Hind leg is unable to load-bearing, only one joint (hip joint) motion |
| 4 | Hind leg is unable to load-bearing, and more than one joint motions, run duration is less than 50% |
| 5 | Hind leg is unable to load-bearing, and more than one joint motions, run duration is more than 50% |
| 6 | Hind leg energy load-bearing, but the load-bearing time is less than 10% |
| 7 | Hind leg energy load-bearing, but the load-bearing time is between 10-50% |
| 8 | Hind leg energy load-bearing, the load-bearing time is more than 50% |
| 9 | The load-bearing of hind leg 100%, but muscular strength do not recover completely, and gait wrong time is more than 90% |
| 10 | The load-bearing of hind leg 100%, but muscular strength do not recover completely, gait stagger the time between 50-90% |
| 11 | The load-bearing of hind leg 100%, but muscular strength do not recover completely, and gait wrong time is less than 50% |
| 12 | Muscular strength is normal, but hind leg gait is chaotic, and wrong time is more than 50% |
| 13 | Muscular strength is normal, but hind leg gait is chaotic, and wrong time is less than 50% |
| 14 | Normal gait |
Table 2, Olby appraisal results
| Beasle dog is numbered | Preoperative scoring | Postoperative scoring |
| 1 | 3 | 3 |
| 2 | 3 | 3 |
| 3 | 3 | 3 |
| 4 | 4 | 4 |
| 5 | 3 | 3 |
| 6 | 3 | 3 |
| 7 | 3 | 3 |
| 8 | 3 | 3 |
| 9 | 3 | 3 |
| 10 | 3 | 3 |
| 11 | 4 | 4 |
| 12 | 3 | 3 |
| 13 | 3 | 3 |
Neurological deficit score result shows that the behaviouristics of dog is not changed in, and shows the excision of cicatricial tissue before and after excision scar
Operation do not cut off normal myeloid tissue, also not to normal myeloid tissue cause damage, bad is not caused to beasle dog
Affect, illustrate the resection operation for being successfully made cicatricial tissue.
2nd, the histological stain of cicatricial tissue
48 hours are fixed in the paraformaldehyde of cicatricial tissue 4% cut off during step one is performed the operation.FFPE, section,
5 μm of thickness, the slide bonding die processed using Poly-D-lysine, drying obtains cicatricial tissue section, so
Carry out the detection and neurofilament NF dyeing of the mark CSPG of cicatricial tissue respectively afterwards.
The detection method of the mark CSPG of cicatricial tissue is as follows：(1) after cicatricial tissue section being washed into one time with PBS, use
Lowlenthal serum (Beijing Bioisystech Co., Ltd of Zhong Shan Golden Bridge product, article No. is ZLI-9021) confining liquid is closed
20-40min；(2) lowlenthal serum is sucked, PBS is washed twice, the water around section is cleaned with blotting paper；(3) add
200 μ L are mixed with the anti-CSPG antibody (1 from mouse:200) (Abcam, ab11570) on biopsy tissues,
Make liquid that myeloid tissue is completely covered；Overnight incubation at (4) 4 DEG C；(5) PBS is cleaned two or three time；(6) according to two anti-explanations
Anti- (the Alexa of invitrogen companies of fluorescence two of book dilution488Donkey Anti-Mouse IgG
(H+L), article No. is CA21202s), it is incubated at room temperature 1 hour, notice that lucifuge is operated；(7) PBS is washed twice, and lucifuge is certainly
So dry；(8) it is added dropwise after the mountant containing DAPI, seals up cover glass, basis of microscopic observation, photograph.
Neurofilament NF colouring methods are as follows：(1) after cicatricial tissue section being washed into one time with PBS, with lowlenthal serum (Beijing
Bioisystech Co., Ltd of Zhong Shan Golden Bridge product, article No. is ZLI-9021) confining liquid closing 20-40min；(2) suck
Lowlenthal serum, PBS is washed twice, and the water around section is cleaned with blotting paper；(3) 200 μ L are added from mouse
anti-NF(1:200) (Beijing Bioisystech Co., Ltd of Zhong Shan Golden Bridge product, article No. is ZM-0198) is in section
Tissue, makes liquid that myeloid tissue is completely covered；Overnight incubation at (4) 4 DEG C；(5) PBS is cleaned two or three time；(6) according to
Anti- (the Alexa of invitrogen companies of fluorescence two of two anti-specification dilutions594Donkey Anti-Rabbit
IgG (H+L), article No. is A21207), it is incubated at room temperature 1 hour, notice that lucifuge is operated；(7) PBS is washed twice, lucifuge
Naturally dry；(8) it is added dropwise after the mountant containing DAPI, seals up cover glass, basis of microscopic observation, photograph.
As a result show that the mark CSPG positives (Fig. 1) of cicatricial tissue in the cicatricial tissue of excision shows the scar for cutting off
Trace is organized as cicatricial tissue；And neurofilament NF coloration results find not finding that neurofilament NF is dyeed in the cicatricial tissue for cutting off
(Fig. 2), show not containing neurofilament in the cicatricial tissue for cutting off.
The chronic spinal cord lesion scar excision of embodiment 2, people
According to《Medical and health organization carries out clinical research projects management measures》Carry out clinical research etc. policy.Choose
5 complete cross-section Patients of Spinal, 5 complete cross-section Patients of Spinal damage sections scope neck 6 to chest 12, damage
Wound is -32 months 2 months apart from 0.5cm-4.5cm, trauma time, does not there is the complication that other are serious.This 5
Spinal cord injury is carried out after the complete cross-section Patients of Spinal signature Informed Consent Form of name according to the method for the step one of embodiment 1
The resection operation of place's cicatricial tissue.
The ASIA methods of marking that spinal cord injury scar excision Post operation is adopted international standards for 7 days is scored, and finds 5
Do not change with plane of movement with postoperative sensation before name operation in patients, illustrate resection organization for cicatricial tissue, spinal cord injury
Scar excision operation in do not cut off normal myeloid tissue, also not to normal myeloid tissue cause damage.
48 hours will be fixed in the paraformaldehyde of cicatricial tissue 4% of spinal cord injury scar excision surgery excision.FFPE,
Section, 5 μm of thickness, the slide bonding die processed using Poly-D-lysine, drying obtains cicatricial tissue section,
Then according to the method for the step 2 of embodiment 1 carries out respectively the detection of the mark CSPG of cicatricial tissue and neurofilament NF dyes
Color.As a result show, as a result show, the mark CSPG positives (Fig. 3) of cicatricial tissue, table in the cicatricial tissue of excision
The cicatricial tissue of bright excision is cicatricial tissue；And neurofilament NF coloration results find not finding god in the cicatricial tissue for cutting off
Organizine NF dyes (Fig. 4), shows not containing neurofilament in the cicatricial tissue for cutting off.
Claims (10)
1. at animal spinal cord injury cicatricial tissue localization method, including it is following 1) and 2)：
1) general location is carried out with reference to cicatricial tissue at following character pair animal spinal cord injury：At the animal spinal cord injury
Normal spinal cord be in bean curd shape, with normal spinal cord structure；Cicatricial tissue week at the animal spinal cord injury
Enclose quality fine and close partially hard, in the middle of cicatricial tissue or have slough or a cavity；
2) near head end of cicatricial tissue at the animal spinal cord injury is positioned and to described using electro physiology detector
The caudal part of cicatricial tissue is positioned at animal spinal cord injury, so that it is determined that cicatricial tissue and ridge at animal spinal cord injury
The boundary of myeloid tissue, completes the positioning of cicatricial tissue at animal spinal cord injury；
The near head end to cicatricial tissue at animal spinal cord injury carries out positioning to be included：Receiving electrode is placed in described dynamic
The head of thing, stimulating electrode is placed at the animal spinal cord injury and along described to the movement of the direction of the animal head
Stimulating electrode, judges the boundary of myeloid tissue and cicatricial tissue, if there is body-sensing by the presence or absence of SEP
Evoked ptential, at the stimulating electrode Normal spinal cord is organized as, if without SEP, the thorn
Cicatricial tissue is organized as at sharp electrode；
The caudal part to cicatricial tissue at animal spinal cord injury carries out positioning to be included：Receiving electrode is placed in described dynamic
At the lower limb muscles or anal sphincter of thing, stimulating electrode is placed at the animal spinal cord injury and along to the animal
The stimulating electrode is moved in the direction of lower limb, judges myeloid tissue with cicatricial tissue by the presence or absence of Motion Evoked Potential
Boundary, if there is Motion Evoked Potential, at the stimulating electrode Normal spinal cord is organized as, if do not transported
Dynamic Evoked ptential, at the stimulating electrode cicatricial tissue is organized as.
2. method according to claim 1, it is characterised in that：The electro physiology detector is myoelectric apparatus.
3. method according to claim 1 and 2, it is characterised in that：The animal is mammal.
4. method according to claim 3, it is characterised in that：The mammal is dog or people.
5. method according to claim 4, it is characterised in that：The dog is beasle dog.
6. the cutting method of animal spinal cord injury cicatricial tissue, including using arbitrary described method in claim 1-5
Determine the border of animal spinal cord injury cicatricial tissue, using the method for operation the animal spinal cord injury cicatricial tissue is cut off.
7. method according to claim 6, it is characterised in that：The animal is mammal.
8. method according to claim 7, it is characterised in that：The mammal is dog or people.
9. method according to claim 8, it is characterised in that：The dog is beasle dog.
10. electro physiology detector described in claim 1 is preparing the dress for positioning cicatricial tissue at animal spinal cord injury
Application in putting.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101856221A (en) * | 2010-05-12 | 2010-10-13 | 中国科学院深圳先进技术研究院 | Detection system for determining damage degree and damage positions of spinal cord |
| KR20110094889A (en) * | 2010-02-18 | 2011-08-24 | 서승우 | Measuring method for damage of spinal cord in accordance with spine distraction |
| US8137285B1 (en) * | 2008-08-26 | 2012-03-20 | Rhythmlink International, Llc | Monopolar stimulation probe system |
| JP5278941B2 (en) * | 2007-11-13 | 2013-09-04 | 国立大学法人広島大学 | Intraoperative evoked potential diagnosis support system |
| CN103948370A (en) * | 2014-05-15 | 2014-07-30 | 中国医学科学院生物医学工程研究所 | Spinal cord injury condition monitoring method based on somatosensory evoked potential signal |
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2015
- 2015-11-03 CN CN201510740062.0A patent/CN106618566A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP5278941B2 (en) * | 2007-11-13 | 2013-09-04 | 国立大学法人広島大学 | Intraoperative evoked potential diagnosis support system |
| US8137285B1 (en) * | 2008-08-26 | 2012-03-20 | Rhythmlink International, Llc | Monopolar stimulation probe system |
| KR20110094889A (en) * | 2010-02-18 | 2011-08-24 | 서승우 | Measuring method for damage of spinal cord in accordance with spine distraction |
| CN101856221A (en) * | 2010-05-12 | 2010-10-13 | 中国科学院深圳先进技术研究院 | Detection system for determining damage degree and damage positions of spinal cord |
| CN103948370A (en) * | 2014-05-15 | 2014-07-30 | 中国医学科学院生物医学工程研究所 | Spinal cord injury condition monitoring method based on somatosensory evoked potential signal |
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