CN108646035A - The new opplication of insulin-like growth factor - Google Patents
The new opplication of insulin-like growth factor Download PDFInfo
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- CN108646035A CN108646035A CN201810480624.6A CN201810480624A CN108646035A CN 108646035 A CN108646035 A CN 108646035A CN 201810480624 A CN201810480624 A CN 201810480624A CN 108646035 A CN108646035 A CN 108646035A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/74—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/575—Hormones
- G01N2333/65—Insulin-like growth factors (Somatomedins), e.g. IGF-1, IGF-2
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/28—Neurological disorders
- G01N2800/2871—Cerebrovascular disorders, e.g. stroke, cerebral infarct, cerebral haemorrhage, transient ischemic event
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/50—Determining the risk of developing a disease
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/52—Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
Abstract
The present invention provides the new opplications of insulin-like growth factor, specifically provide the effect of insulin-like growth factor is as assessment test individual mescenchymal stem cell intravenous transplantation cerebral infarction, and/or implantation cell survival and/or the application for whether having functional marker.In the present invention, after cell transplantation, negative correlation is presented with cerebral infarction volume in 1 concentration of cerebral infarction group peripheral blood IGF, and positive correlation is presented with 1 concentration of infarct cortical area IGF, and positive correlation is presented with infarct cortical area IGF 1+ cell numbers.
Description
Technical field
The present invention relates to the new opplications of insulin-like growth factor, specifically exist about insulin-like growth factor
The effect of analyzing and determining mescenchymal stem cell intravenous transplantation cerebral infarction is implanted into cell survival and/or whether has work(
Application in energy.
Background technology
Mescenchymal stem cell (mesenchymal stem cells, MSC) transplanting is possible to treat a variety of diseases, such as god
Through system degenerative disease such as Parkinson, senile dementia, metabolic disease such as diabetes, auto-immune related disease such as system
Property lupus erythematosus etc..
It is required that the effect of whether MSC fed by vein survives for a variety of diseases in vivo, which generates, and MSC is immune
Source property is weaker, can be with allograft, but its existing time in people or animal body is limited, if implantation cell
All dead, curative effect will disappear quickly.Based on this, a kind of easy flexible mode is needed to judge that the MSC being implanted into animal body is
It is no still to survive and play therapeutic effect.
Two aspects are related to for the detection of MSC functions, first is whether cell survives；Second, whether cell has
Function rather than be in dormancy state.
It survives generally for MSC and still has functional detection, be that cell is marked before being implanted into cell, place
After dead animal, paraformaldehyde perfusion is fixed, and is taken brain tissue, is observed under the microscope after slice dyeing, identify the cell of implantation
Whether various cell factors are generated.This method needs to put to death animal, if necessary to put the work(to being implanted into cell in different times
It can be detected, it is necessary to which point puts to death animal in different times.In view of difference between the inter-individual difference and batch of animal, respectively
Comparison between a time point needs more experimental animal quantity that could obtain the standard deviation to tally with the actual situation.
It is by being marked to transplanted cells, using living animal that another kind detection MSC, which survives and has functional method,
The method of imaging carries out.It is to use luciferase (Luciferase) genetic marker cell or DNA, and fluorescent technique in this method
Then it is marked using fluorescent reporter group (GFP, RFP, Cyt and dyes etc.).Utilize a set of very sensitive optical detector
Device, allows the cellular activity and gene behavior that researcher can be in direct monitoring living body biological body.Pass through this system, Ke Yiguan
Survey situation of the cell in living animal body, such as the growth and transfer of tumour, the development of infectious diseases, the table of specific gene
Up to etc. biological processes.Compared to it is traditional, need to put to death experimental animal in different time points method to obtain data, it is seen that light
In-vivo imaging by being recorded in different time points to same panel, track same object observing (label cell and
Gene) movement and variation, the data of gained are more intuitive.However, this technology is to implantation cell, there are less positions
Detection sensitivity is inadequate, such as is first gathered in spleen and lungs after mescenchymal stem cell vein transplantation, can be lived by toy
Body imaging technique detects；But the quantity that the cell of vein implantation enters intracerebral by blood-brain barrier is seldom, it generally can not be by
The technology is effectively shown.Another is mainly disadvantageous in that the survival for only showing implantation cell to the technology, to being implanted into cell
The function of performance cannot be detected directly, such as whether implantation cell can not also intuitively show in various cell factors of generation etc..
In addition, the price of the reagents such as small animal living body imaging device and luciferase, substrate is more expensive.
Invention content
Whether it is an object of the present invention to provide a kind of MSC that novel detection implantation individual is internal still to survive simultaneously
And the method with function rather than in dormancy state.
The present invention is mainly by detecting insulin-like growth factor (Insulin-like Growth Factor-1, IGF-
1) mark still survived and functioned as the MSC cells of vein implantation.
IGF-1 is the single chain polypeptide containing 70 amino acid for being located at No. 12 chromogene coding, structure and pancreas
Island element is similar.Human body has multiple tissues and internal organs all to express IGF-1, and IGF-1 is mainly regulated and controled by growth hormone.Growth hormone
The transcription of IGF-1 genes can be quickly activated, while adjust the chromosome structure of IGF-1 genes, to be formed on chromosome
The action target spot of growth hormone.Other than growth hormone, estradiol, angiotensins etc. are also the activation of IGF-1 genetic transcriptions
Factor.Local organization IGF-1 is generated with autocrine or paracrine, and the IGF-1 in blood is mainly generated by liver, only few
Amount IGF-1 comes from bone and is secreted into blood, it is generally recognized that IGF-1 Biological Attribute of Industrial is played by the free IGF-1 in blood
's.
Inventor has found that the growth factor I GF-1 in peripheral blood can illustrate as indicator under study for action：
Whether there is work(to the therapeutic effect of cerebral infarction, implantation cell survival and implantation cell after the transplanting of MSC cells is
Energy.
In the present invention, the implantation cell " having function " refers to that implantation cell is not in dormancy state, but also exists
It plays a role.Effect herein is mainly 2 points：First, immunological regulation, show as inhibiting proinflammatory disease excessively high after cerebral infarction because
Son, TNF-α, IL-1 β etc.；Second is that brain tissue is promoted to generate a variety of trophism cell factors.More concern implantation of the invention herein
Cell is still promoting brain tissue to generate a variety of trophism cell factors as its functional mark of tool.
The present invention studies have shown that mescenchymal stem cell intravenous transplantation cerebral infarction, after cell transplantation, infarcted region cortex
The IGF-1 serum-concentrations raising more notable than ischemic control group of transplantation group.Peripheral blood IGF-1 concentration and cerebral infarction after the transplanting of cerebral ischemia group
Dead volume is without apparent correlation, but negative is presented with cerebral infarction volume in the cerebral infarction group peripheral blood IGF-1 concentration of cell transplantation
Guan Xing.Cerebral ischemia group transplanting after peripheral blood IGF-1 concentration with intracerebral infarct cortical area IGF-1 concentration without apparent correlation, but carefully
Positive correlation is presented with infarct cortical area IGF-1 concentration in the cerebral infarction group peripheral blood IGF-1 concentration of born of the same parents' transplanting.Cerebral ischemia group is moved
Peripheral blood IGF-1 concentration and intracerebral infarct cortical area IGF-1+ cell numbers are without apparent correlation after plant, but the hindbrain of cell transplantation
Positive correlation is presented with infarct cortical area IGF-1+ cell numbers in infarct group peripheral blood IGF-1 concentration.
The research of the present invention is also shown that trophic factors in the brain tissue of infarcted region, and IGF-1 is 4 days after infarct, shows within 14 days
Transplantation group raising more notable than ischemic control group, BDNF (Brain-derived neurotrophic factor, brain source nerve
Trophic factors) only there are notable raising, VEGF (Vascular endothelial growth factor, vascular endothelia on day 4
Growth factor), GDNF (Glial-derived neurotrophic factor, glial cell line-derived neurotrophic factor) transplanting before
Afterwards without significant changes.Meanwhile peripheral blood B DNF transplanting is front and back and finds no significant difference, i.e. peripheral blood B DNF can not be used as exhibition
The index whether Cerebral Infarction Treatment effect and transplanted cells are survived and functioned shown.
To, on the one hand, the present invention provides insulin-like growth factor (IGF-1) as assessment test individual mesenchyma
The effect of stem cell intravenous transplantation cerebral infarction, and/or implantation cell survival and/or whether has a functional marker
Application.
On the other hand, the present invention also provides the horizontal reagents of detection IGF-1 to prepare for assessing between test individual
The effect of mesenchymal stem cells intravenous transplantation cerebral infarction, and/or implantation cell survival and/or whether has a functional inspection
Survey the application in agent or detecting system.
Specific embodiment according to the present invention, in of the invention, the level of the IGF-1 is insulin-like growth factor
Level of peripheral blood or serum levels.
Specific embodiment according to the present invention, in of the invention, the level of the IGF-1 is insulin-like growth factor
Protein expression level.
Specific embodiment according to the present invention, in of the invention, the horizontal method for detecting IGF-1 can be in this field
Known any feasible method, for example, the methods of enzyme linked immunosorbent assay (ELISA).To detect the level of IGF-1
Reagent can be the reagent of the expression based on any feasible detection IGF-1 as known in the art.
Specific embodiment according to the present invention, in of the invention, the test individual is mammal or people.
Specific embodiment according to the present invention, in of the invention, the test individual is mescenchymal stem cell vein transplantation
Individual.
Specific embodiment according to the present invention, in of the invention, in the sample (peripheral blood or serum) from test individual
IGF-1 levels increase, and cell survival is implanted into the test individual and has function.
Specific embodiment according to the present invention, in of the invention, peripheral blood IGF-1 concentration and cerebral infarction vein transplantation MSC
Cerebral infarction volume is negatively correlated.It can be by detecting peripheral blood, you can relatively accurately assess the Infarction volume of cerebral infarction.
Specific embodiment according to the present invention, in of the invention, peripheral blood IGF-1 concentration and intracerebral cortical infarction area IGF-
1 concentration positive correlation.
Specific embodiment according to the present invention, in of the invention, peripheral blood IGF-1 concentration and intracerebral cortical infarction area generate
The MSC cell quantity positive correlations of IGF-1.It can be by detecting peripheral blood, you can relatively accurately whether assessment implantation cell survives
And generating IGF-1.
Specific embodiment according to the present invention, in of the invention, IGF-1 levels increase in the sample from test individual,
The effect of assessing test individual mescenchymal stem cell intravenous transplantation cerebral infarction is preferable.
On the other hand, the present invention also provides one kind for assessing test individual mescenchymal stem cell intravenous transplantation brain
The effect of infarct, and/or implantation cell survival and/or whether have functional detecting system (detection device), the detection
System includes：
Detection unit, the detection unit include detecting the horizontal reagent of insulin-like growth factor；
Analytic unit, the analytic unit are filled for analyzing the testing result of detection unit between assessment test individual
The effect of matter stem cell intravenous transplantation cerebral infarction, and/or implantation cell survival and/or whether have function.
Specific embodiment according to the present invention, of the invention is used to assess test individual mescenchymal stem cell vein transplantation
Whether the effect for the treatment of cerebral infarction, and/or implantation cell survival and/or have functional detecting system, are according to coming from
The height of IGF-1 levels in the sample of test individual, to assess the test individual mescenchymal stem cell intravenous transplantation cerebral infarction
Dead curative effect, and/or implantation cell survival and/or whether there is function.Wherein, IGF- in the sample from test individual
The effect of 1 horizontal raising, the test individual mescenchymal stem cell intravenous transplantation cerebral infarction, is preferable, is implanted into cell survival and tool
Functional quantity is more.
It is of the present invention for assess test individual mescenchymal stem cell intravenous transplantation cerebral infarction the effect of and/
Or be implanted into cell survival and/or whether have functional detecting system, can be virtual bench, as long as the inspection can be realized
Survey the function of unit and analytic unit.The detection unit can be include various detection reagents, kit or inspection
Survey instrument；The data analysis unit can be it is any may be implemented to carry out analyzing processing to the testing result of detection unit and
It obtains operation instrument, module or the virtual unit of starting cerebral apoplexy risk situation, such as can be in advance according to above-mentioned
It assesses the effect of IGF-1 value height is corresponded to individual mescenchymal stem cell intravenous transplantation cerebral infarction by principle, and/or implantation is thin
Born of the same parents' survival condition and/or whether with function the case where be formulated to convenient for control consult data drawing list, by the inspection of detection unit
Survey result IGF-1 values compare the effect of data drawing list can obtain individual mescenchymal stem cell intravenous transplantation cerebral infarction,
And/or it is implanted into cell survival and/or whether there is the case where function.
On the other hand, the present invention also provides a kind of treatments of the individual mescenchymal stem cell intravenous transplantation cerebral infarction of assessment
It imitates, and/or is implanted into cell survival and/or whether have functional method, this method includes：Detection individual (can come
From the sample of test individual) level of IGF-1；According to the height of individual IGF-1 levels, to assess the individual mescenchymal stem cell
The effect of intravenous transplantation cerebral infarction, and/or implantation cell survival and/or whether have function the case where.
In conclusion whether the present invention provides a kind of MSC that new detection implantation individual is internal still to survive and have
Functional rather than method in dormancy state.The method of the present invention can put repeatedly same individual in different times logical
Whether the mescenchymal stem cell for crossing acquisition venous blood detection vein transplantation plays therapeutic effect.It, can in the zooscopy experimental stage
To efficiently reduce the individual difference of animal and reduce the demand to experimental animal total amount.Transplant MSC's in Patients with Cerebral Infarction
In clinical trial, implantation cell survival can be largely indicated with the peripheries IGF-1 blood concentration and plays secretion IGF-1
Effect, and with reduce Infarction volume therapeutic effect.
Description of the drawings
Figure 1A displays are broken up to the growth conditions of the 5th generation BM-MSC.
The BM-MSC that Figure 1B-Fig. 1 G are cultivated by fluidic cell identification expresses CD90, CD73, CD44, CD105 and CD34
The analysis chart of situation.
Fig. 2 is BMSC cell transplantation group cerebral infarction volume TTC colored graphs after ischemic control group and ischemic.Wherein, picture A,
Ischemic control group.Region shown in picture B lattices is cerebral infarction core space, Striatum (no ischemic injuries) and cerebral infarction periphery
The materials position of cortical area (no ischemic injuries).
Fig. 3 A and Fig. 3 B show the result of 4d, 14d time point ELISA detection IGF-1 contents after cell transplantation.In figure, #:p
<0.05 compared with ischemia group；*:p<0.05 compared with sham-operation group.
4 days peripheral blood IGF-1 concentration is tested with cerebral infarction volume correlation after Fig. 4 A show Nonimplantation group animal cerebral infarction
As a result.
Fig. 4 B show 4 days peripheral blood IGF-1 concentration and cerebral infarction volume correlation reality after transplantation group cerebral infarction zoografting
Test result.
14 days peripheral blood IGF-1 concentration is tested with cerebral infarction volume correlation after Fig. 4 C show Nonimplantation group animal cerebral infarction
As a result.
Fig. 4 D show 14 days peripheral blood IGF-1 concentration and cerebral infarction volume correlation reality after transplantation group cerebral infarction zoografting
Test result.
4 days peripheral blood IGF-1 concentration and intracerebral infarct cortical area IGF-1 are dense after Fig. 5 A display Nonimplantation group animal cerebral ischemias
Spend correlation experimental result.
Fig. 5 B show 4 days peripheral blood IGF-1 concentration and infarct cortical area IGF-1 concentration after transplantation group cerebral infarction zoografting
Correlation experimental result.
Fig. 5 C show 14 days peripheral blood IGF-1 concentration and cerebral infarction dead zone concentration dependence reality after Nonimplantation group animal cerebral infarction
Test result.
Fig. 5 D show 14 days peripheral blood IGF-1 concentration and cerebral infarction dead zone concentration dependence after transplantation group cerebral infarction zoografting
Experimental result.
Fig. 6 shows BM-MSC the and IGF-1 double labelled staining results of transplantation group rat layer infarcted region CM-Dil labels.Its
In, picture A：The anti-IGF-1 antibody mediated immunities dyeing in rats with cerebral ischemia infarcted region；Picture B：BM- is marked in the same visual field CM-DiI
MSC.Picture C：The merging image of IGF-1, CM-Dil and DAPI, picture D：Picture C institutes collar region enlarged drawing, engineer's scale=50
μm。
Fig. 7 A show 4 days peripheral blood IGF-1 concentration and intracerebral infarct cortical area IGF-1+ after Nonimplantation group animal cerebral ischemia
Cell data/coherency experimental result.
4 days peripheral blood IGF-1 concentration and infarct cortical area IGF-1+ are thin after Fig. 7 B display transplantation group cerebral infarction zoograftings
Born of the same parents' data/coherency experimental result.
Fig. 7 C show 14 days peripheral blood IGF-1 concentration and cerebral infarction dead zone IGF-1+ cell numbers after Nonimplantation group animal cerebral ischemia
Correlation experimental result.
Fig. 7 D show 14 days peripheral blood IGF-1 concentration and cerebral infarction dead zone IGF-1+ cells after transplantation group cerebral infarction zoografting
Data/coherency experimental result.
Fig. 8 A, Fig. 8 B show transplantation group 4 days, 14 days peripheral blood BDNF contents transplant front and back result of variations.
Fig. 9 A to Fig. 9 D respectively illustrate after transplanting four kinds of trophic factors BDNF, GDNF, IGF- in 4 days infarcted region brain tissues
1, the changes of contents of VEGF.
Figure 10 A to Figure 10 D respectively illustrate after transplanting four kinds of trophic factors BDNF, GDNF in 14 days infarcted region brain tissues,
The changes of contents of IGF-1, VEGF.
Specific embodiment
In order to which technical characteristic, purpose and the advantageous effect to the present invention are more clearly understood, in conjunction with specific implementation
Example and technical scheme of the present invention is carried out described further below, it should be understood that these examples be merely to illustrate the present invention rather than
It limits the scope of the invention.In embodiment, each Starting reagents material is commercially available, test method without specific conditions
For conventional method known to fields and normal condition, or according to the condition proposed by apparatus manufacturer.
Embodiment 1
1. cell collection and culture identification
Pass on 3-5 for when BM-MSC growth conditions it is good, can be evenly distributed in culture dish bottom (Figure 1A).Pass through
Flow cytometry can detect that cultivated BM-MSC can high expression CD90, CD73, CD44, CD105 (Figure 1B-figure
1F), the positive rate for detecting cell is all higher than 90%；But CD34 (Fig. 1 G) is not expressed.
Specific method please refers to document：Zhao Chunsong etc., " human umbilical cord mesenchymal stem cells drench peripheral blood of patients with Parkinson disease
The inhibition of bar hyperplasia ",《Capital University of Medical Sciences's journal》, 2014,35 (3), 344-352.
2. cerebral infarction volume compares
Experiment packet：
(1) sham-operation group, animal expose arteria carotis communis by opening cranium exposure right side arteria cerebri media, cervical incision, but not
To blood vessel carry out folder close or coagulation operation.
(2) ischemia group (being equal to cerebral infarction group), animal right side arteria cerebri media are blocked after coagulation, homonymy arteria carotis communis
Folder closes 30 minutes, that is, completes cerebral ischemic model and make.Folder gives 1ml physiological saline in 30 minutes after the completion of closing through tail vein.
(3) transplantation group, cerebral ischemic model making is identical as ischemia group, after the completion of folder closes arteria carotis communis 30 minutes, interval 30
Minute is implanted into MSC cells through vein.MSC cells are placed in physiological saline, total amount 1ml.
The time point of 2d dyes the infarct face of BM-MSC transplanting after detection ischemic control group and ischemic by TTC after the transfer
Product comparison.It understands that the damage of dMCAO rat cerebral infarctions is predominantly located at cortex, slightly involves caudate nucleus and hippocampus (Fig. 2).After ischemic
BM-MSC transplantation groups cerebral infarction volume is substantially reduced compared with ischemic control group, and the cerebral infarction average external volume of ischemic control group is 29.46
± 5.76%, and the cerebral infarction average external volume of BM-MSC transplantation groups is 22.68 ± 5.51% (Fig. 2) after ischemic.
3. the concentration mensuration of peripheral blood IGF-1
The time point of 4d, 14d after cell transplantation, every group 10 only at mould and successful implantation rat with 10% chloraldurate
Brain is taken immediately after deep anesthesia, and infarct core space brain tissue and infarcted region periphery striatum is taken to be used as detection sample respectively,
The PBS being pre-chilled with 4 DEG C washes off bloodstain, and PBS is sopped up with filter paper, and addition is fully homogenized with brain tissue homogenate's liquid under the conditions of 0-4 DEG C,
Then 30min is centrifuged under the conditions of 4 DEG C with the rotating speed of 15000r/min, takes supernatant.With the total egg of BCA protein quantification kit measurements
Then Bai Liang is added dilution according to the amount of 40 μ g brain total proteins of every hole and is made into 100 μ l samples, it is detected with ELISA kit
The content of middle trophic factors IGF-1.
Testing result is shown referring to Fig. 3 A and Fig. 3 B, it is seen that two time points, the IGF-1 blood of infarcted region cortex transplantation group
Clear concentration ratio ischemic control group significantly increases.
4. transplantation group peripheral blood IGF-1 concentration is negatively correlated with cerebral infarction volume
By to the peripheral blood IGF-1 concentration of each animal and the analysis of cerebral infarction volume, present invention discover that cerebral ischemia
Group 4 days peripheral blood IGF-1 concentration of infarct with cerebral infarction volume without apparent correlation (Fig. 4 A), but after cell transplantation, transplantation group
Negative correlation (Fig. 4 B) is presented with cerebral infarction volume in peripheral blood IGF-1 concentration.
Also have within 14 days after transplanting a similar situation, i.e., Nonimplantation group peripheral blood IGF-1 concentration with cerebral infarction volume without apparent phase
Closing property (Fig. 4 C), and transplantation group has correlation (Fig. 4 D).
5. transplantation group peripheral blood IGF-1 concentration is related to intracerebral concentration
The present invention compared the peripheral blood IGF-1 concentration of each animal and the concentration of intracerebral infarct cortical area IGF-1, hair
4 days peripheral blood IGF-1 concentration (is schemed with intracerebral infarct cortical area IGF-1 concentration without apparent correlation after existing Nonimplantation group cerebral ischemia
5A), but after cell transplantation, positive correlation (figure is presented with infarct cortical area IGF-1 concentration in transplantation group peripheral blood IGF-1 concentration
5B)。
Also have within 14 days after transplanting a similar situation, i.e., Nonimplantation group peripheral blood IGF-1 concentration and cerebral infarction dead zone IGF-1 concentration without
Apparent correlation (Fig. 5 C), and transplantation group has apparent positive correlation (Fig. 5 D).
6. infarct cortex is implanted into the double labelling of the red fluorescence and IGF-1 of cell BM-MSC
The red fluorescence of implantation cell BM-MSC is dispersed in distribution in cerebral infarction dead zone cortex.IGF-1 immunohistochemical staining results
It was found that the green fluorescence (the picture A in Fig. 6) of IGF-1 immunohistochemical stainings and the CM-DiI red being implanted into entrained by cell are glimmering
Light is in infarcted region as it can be seen that having 30-40% to carry the CM-Dil red fluorescences (picture in Fig. 6 in the cell of expression IGF-1
B).It finds that the immunofluorescence dyeing of IGF-1 is primarily present in infarct periphery cortical area by simple stain photo, also has in infarcted region
There is (the picture A in Fig. 6) in staining positive cells.The cell of these double labellings can preferably be overlapped with DAPI nuclear targetings
(picture C, picture D in Fig. 6).
7. transplantation group peripheral blood IGF-1 concentration is related to infarcted region cortex IGF-1+ cell quantities
The present invention compared the peripheral blood IGF-1 concentration and intracerebral infarct cortical area IGF-1+ cell numbers of each animal,
It was found that after cerebral ischemia group animal infarct 4 days peripheral blood IGF-1 concentration with intracerebral infarct cortical area IGF-1+ cell numbers without apparent phase
Closing property (Fig. 7 A), but after cell transplantation, transplantation group animal peripheral blood IGF-1 concentration is in infarct cortical area IGF-1+ cell numbers
Existing positive correlation (Fig. 7 B).
Also there are within 14 days after transplanting similar situation, i.e. Nonimplantation group peripheral blood IGF-1 concentration and cerebral infarction dead zone IGF-1+ cells
Number is without apparent correlation (Fig. 7 C), and transplantation group has apparent positive correlation (Fig. 7 D).
8. trophic factors changes of contents in transplantation group peripheral blood and infarcted region brain tissue
The present invention also has detected peripheral blood B DNF (Brain-derived neurotrophic factor, brain source nerve
Trophic factors) the front and back variation of transplanting, (Fig. 8 A), 14 days (Fig. 8 B) all do not find that transplantation group and Nonimplantation group have 4 after transplanting
Significant difference, i.e. peripheral blood B DNF can not be used as whether displaying survives and play to Cerebral Infarction Treatment effect and transplanted cells
The index of function.
The present invention also has detected the content of four kinds of trophic factors in the brain tissue of infarcted region, including BDNF, IGF-1, VEGF
(Vascular endothelial growth factor, vascular epithelial growth factor), GDNF (Glial-derived
Neurotrophic factor, glial cell line-derived neurotrophic factor).Wherein IGF-1 is 4 days after infarct, shows to transplant within 14 days
Group raising more notable than ischemic control group, BDNF only have notable raising, VEGF, GDNF transplanting front and back without significant changes (figure on day 4
9A to Fig. 9 D, Figure 10 A to Figure 10 D).
Claims (10)
1. the effect of insulin-like growth factor is as assessment test individual mescenchymal stem cell intravenous transplantation cerebral infarction,
And/or it is implanted into cell survival and/or whether has the application of functional marker.
2. the horizontal reagent for detecting insulin-like growth factor is being prepared for assessing the starting cerebral apoplexy illness wind of test individual
The detection agent of danger or the application in detecting system.
3. application according to claim 2, wherein the level of the insulin-like growth factor is insulin-like growth factor
The level of peripheral blood or serum levels of son.
4. application according to claim 2 or 3, wherein the level of the insulin-like growth factor is Insulin-Like life
The protein expression level of the long factor.
5. application according to claim 1 or 2, wherein the test individual is mammal or people.
6. application according to claim 1 or 2, wherein insulin-like growth factor water in the sample from test individual
It is flat to increase, it is implanted into cell survival in the test individual and there is function.
7. application according to claim 1 or 2, wherein：
Peripheral blood insulin-like growth factor concentration and cerebral infarction vein transplantation MSC cerebral infarctions volume are negatively correlated；
Peripheral blood insulin-like growth factor concentration and the area IGF-1 concentration positive correlations of intracerebral cortical infarction；Or
Peripheral blood insulin-like growth factor concentration and intracerebral cortical infarction area generate the MSC cell quantity positive correlations of IGF-1.
8. application according to claim 1 or 2, wherein：Insulin-like growth factor water in sample from test individual
The effect of flat raising, assessment test individual mescenchymal stem cell intravenous transplantation cerebral infarction, is preferable.
9. one kind is for the effect of assessing test individual mescenchymal stem cell intravenous transplantation cerebral infarction, and/or implantation cell
Survival condition and/or whether has functional detecting system, which includes：
Detection unit, the detection unit include detecting the horizontal reagent of insulin-like growth factor；
Analytic unit, for the analytic unit for analyzing the testing result of detection unit, assessment test individual mesenchyma is dry
The effect of cells i transplantation treatment cerebral infarction, and/or implantation cell survival and/or whether have function.
10. detecting system according to claim 9, wherein according to the height of IGF-1 levels in the sample from test individual
It is low, the effect of to assess the test individual mescenchymal stem cell intravenous transplantation cerebral infarction, and/or implantation cell survival
And/or whether there is function, wherein：
Peripheral blood insulin-like growth factor concentration and cerebral infarction vein transplantation MSC cerebral infarctions volume are negatively correlated；
Peripheral blood insulin-like growth factor concentration and the area IGF-1 concentration positive correlations of intracerebral cortical infarction；Or
Peripheral blood insulin-like growth factor concentration and intracerebral cortical infarction area generate the MSC cell quantity positive correlations of IGF-1.
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Citations (3)
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| WO2004090112A2 (en) * | 2003-04-01 | 2004-10-21 | United States Of America Department Of Veteran's Affairs | Stem-cell, precursor cell, or target cell-based treatment of multi-organ failure and renal dysfunction |
| US20080254019A1 (en) * | 2004-03-31 | 2008-10-16 | Yukio Kato | Therapeutic Agents and Therapeutic Methods For Treating Injured Tissue |
| CN101384702A (en) * | 2004-03-22 | 2009-03-11 | 奥西里斯治疗公司 | Mesenchymal stem cells and uses therefor |
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| WO2004090112A2 (en) * | 2003-04-01 | 2004-10-21 | United States Of America Department Of Veteran's Affairs | Stem-cell, precursor cell, or target cell-based treatment of multi-organ failure and renal dysfunction |
| CN101384702A (en) * | 2004-03-22 | 2009-03-11 | 奥西里斯治疗公司 | Mesenchymal stem cells and uses therefor |
| US20080254019A1 (en) * | 2004-03-31 | 2008-10-16 | Yukio Kato | Therapeutic Agents and Therapeutic Methods For Treating Injured Tissue |
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| XIAOBO LI，ET AL: "Intravenously Delivered Allogeneic Mesenchymal Stem Cells Bidirectionally Regulate Inflammation and Induce Neurotrophic Effects in Distal Middle Cerebral Artery Occlusion Rats Within the First 7 Days After Stroke", 《CELL PHYSIOL BIOCHEM》 * |
| 赵鸿: "加味丹参饮含药血清体外诱导的BMSCs移植对心肌IRI模型大鼠血管新生影响的研究", 《中国博士学位论文全文数据库 医药卫生科技辑》 * |
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