CN104232581B - Purposes of the poloxamer in induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation - Google Patents
Purposes of the poloxamer in induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation Download PDFInfo
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- CN104232581B CN104232581B CN201410429405.7A CN201410429405A CN104232581B CN 104232581 B CN104232581 B CN 104232581B CN 201410429405 A CN201410429405 A CN 201410429405A CN 104232581 B CN104232581 B CN 104232581B
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Abstract
The invention discloses purposes of the poloxamer in induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation.Poloxamer is added in hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation system, can induce, promote hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation, and be proliferated fastly, erythroid differentiation is high-efficient.
Description
Technical field
The present invention relates to biomedicine field, especially hematopoietic stem/progenitor cells proliferation and erythroid progenitor cells induction differentiation neck
Domain.In particular it relates to poloxamer induction of hematopoiesis stem/progenitor cells be proliferated and/or erythroid differentiation in purposes, be used for
The culture medium of induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation divides for induction of hematopoiesis stem/progenitor cells proliferation and/or red system
The kit of change and the method for promoting hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation.
Background technique
In recent years since blood sources anxiety and blood transfusion related disease make it is intended that it is safer to find, fills
The blood sources of foot.Research shows that stem cell can be induced to differentiate into red blood cell in vitro can be used as new blood sources.In addition to
Outside mature erythrocyte, erythroid progenitor cells can also substitute clinical red blood cell transfusion, play its hematopoiesis support therapeutic effect, Er Qiehong
Be that progenitor cells can continue to be proliferated in vivo, for much need Transfusion treat patient, be expected to reduce blood transfusion number with
And the side effect that blood transfusion generates, thus, erythroid progenitor cells provides new alternative route for clinical blood supply.
However, current hematopoietic stem/progenitor cells proliferation and/or erythroid progenitor cells differentiated system, there is also proliferation and induction point
The problems such as changing low efficiency, still needs to improve.
Summary of the invention
The present invention is directed at least solve one of the technical problems existing in the prior art.For this purpose, one object of the present invention
The method for being to propose a kind of proliferation and the induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation that induce differentiation efficiency high.
It should be noted that the present invention is the following work based on inventor and completes:
Poloxamer (poloxamer) is Pluronic F68, according to polymer ethylene oxide and ring
The proportion of Ethylene Oxide, poloxamer have a series of different molecular weights and the kind containing polyoxypropylene, polyoxyethelene content.Moor Lip river
The polyoxyethylene chain of husky nurse has relative hydropathy, and polyoxypropylene chains have lipophilicity relatively, therefore are a kind of non-ionic height
Molecular surface active agent is mainly used as emulsifier and solubilizer in pharmaceutical preparation.PLURONICS F87 (poloxamer 188)
Structural formula be PEO76PPO29PEO76, with very high safety, and toxicity is low, non-stimulated anaphylaxis, no antigen,
Good biocompatibility, chemical property are stable, are not easy to cause haemolysis, and emulsifying capacity is strong.Poloxamer188 (poloxamer 407)
Structural formula be PEO100PPO65PEO100, with good surface-active and safety, activating agent middle-molecular-weihydroxyethyl is larger, hydrophilic
Property it is stronger, toxicity is minimum, it is particularly common in pharmacy as defoaming agent, solubilizer and medicine controlled releasing auxiliary material etc., be the U.S.
The drug auxiliary material that pharmacopeia newly increases.
However, the present inventor as seed cell and carries out erythroid progenitor cells using umbilical cord blood hematopoietic stem/progenitor cells
When induction differentiation research, have been surprisingly found that different cytokines combination culture medium assists with only plus compared with the culture medium of combinations of factors
PLURONICS F87 or the external evoked human umbilical cord blood mononuclear cell of poloxamer188 can obtain highly expressed erythroid progenitor cells.Moor Lip river
Husky nurse 188 and poloxamer188 can promote hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation.And the discovery, at present not yet
See relevant report.
Thus, according to an aspect of the present invention, the present invention provides poloxamers to be proliferated in induction of hematopoiesis stem/progenitor cells
And/or the purposes in erythroid differentiation.Poloxamer is added in hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation system as a result,
It can induce, promote hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation, and be proliferated fastly, erythroid differentiation is high-efficient.
According to embodiments of the present invention, the poloxamer is PLURONICS F87 or poloxamer188.As a result, in Hematopoietic Stem
PLURONICS F87 or poloxamer188 are added in progenitor cell proliferation and/or erythroid differentiation system, can be induced, be promoted Hematopoietic Stem
Progenitor cell proliferation and/or erythroid differentiation, and be proliferated fastly, erythroid differentiation is high-efficient.
According to an embodiment of the invention, the hematopoietic stem/progenitor cells, which derive from, is selected from marrow, peripheral blood, placenta and Cord blood
At least one, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.The i.e. described hematopoietic stem/progenitor cells
Source be not particularly limited, it is thin Hematopoietic Stem ancestral can be obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation
Born of the same parents can also be directly separated from marrow, peripheral blood, placenta or Cord blood and obtain, such as directly separate Cord blood from bleeding of the umbilicus
Mononuclearcell, as hematopoietic stem/progenitor cells source.
According to another aspect of the present invention, the present invention also provides one kind for induction of hematopoiesis stem/progenitor cells proliferation and/or
The culture medium of erythroid differentiation.According to an embodiment of the invention, the culture medium includes: poloxamer.It is surprisingly found by the inventors that adopting
It is used for the culture medium culture hematopoietic stem/progenitor cells of induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation with this, can effectively lure
It leads hematopoietic stem/progenitor cells proliferation and/or is converted into erythroid progenitor cells, and proliferation is fast, erythroid differentiation is high-efficient.
According to embodiments of the present invention, the poloxamer is PLURONICS F87 or poloxamer188.Hematopoietic Stem ancestral as a result,
Cell Proliferation and/or erythroid differentiation are high-efficient.
In addition, the training according to the above embodiment of the present invention for induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation
Feeding base can also have the following additional technical features:
According to an embodiment of the invention, the concentration of the PLURONICS F87 is 0.2~2mg/mL, preferably 1mg/mL.By
This, hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation are high-efficient.
According to an embodiment of the invention, the concentration of the poloxamer188 is 0.2~2mg/mL, preferably 1mg/mL.By
This, hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation are high-efficient.
According to an embodiment of the invention, the culture medium is to be added to SCF, IGF-1, EPO, transferrins and dexamethasone
Stemspan culture medium.Be conducive to induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation as a result,.
According to an embodiment of the present invention, the concentration of the SCF is 10~300ng/mL, preferably 100ng/mL；The IGF-1
Concentration be 5~100ng/mL, preferably 40ng/mL；The concentration of the EPO is 1~20U/mL, preferably 5U/mL；It is described to turn iron egg
White concentration is 10~300 μ g/mL, preferably 100 μ g/mL；The concentration of the dexamethasone is 0.1~3 μM, preferably 1 μM.By
This, using the culture medium culture hematopoietic stem/progenitor cells, proliferation and/or erythroid differentiation are high-efficient, and effect is good.
According to embodiments of the present invention, the hematopoietic stem/progenitor cells are derived from selected from marrow, peripheral blood, placenta and Cord blood
At least one, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.The i.e. described hematopoietic stem/progenitor cells
Source is not particularly limited, and it is thin can to obtain Hematopoietic Stem ancestral by embryonic stem cell or inductive pluripotent stem cells vitro differentiation
Born of the same parents can also be directly separated from marrow, peripheral blood, placenta or Cord blood and obtain, such as directly separate Cord blood from bleeding of the umbilicus
Mononuclearcell, as hematopoietic stem/progenitor cells source.
According to another aspect of the invention, the present invention also provides one kind for induction of hematopoiesis stem/progenitor cells proliferation and/or
The kit of erythroid differentiation.According to an embodiment of the invention, containing poloxamer in the kit.Inventor sends out in surprise
It is existing, using this for induction of hematopoiesis stem/progenitor cells proliferation and/or the kit of erythroid differentiation, hematopoietic stem/progenitor cells are cultivated, it can
Effective induction of hematopoiesis stem/progenitor cells are proliferated and/or are converted into erythroid progenitor cells, and differentiation efficiency is high, and proliferation is fast.
According to embodiments of the present invention, the poloxamer is PLURONICS F87 or poloxamer188.Hematopoietic Stem ancestral as a result,
Cell Proliferation and/or erythroid differentiation are high-efficient, and effect is good.
In addition, the examination according to the above embodiment of the present invention for induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation
Agent box can also have the following additional technical features:
According to an embodiment of the invention, the kit further includes SCF, IGF-1, EPO, transferrins and ground plug rice
Pine.
According to some embodiments of the present invention, the PLURONICS F87 kit further includes basal medium, described
Basal medium is Stemspan culture medium.
Some preferred embodiments according to the present invention, the PLURONICS F87, poloxamer188, SCF, IGF-1, EPO,
At least one of transferrins and dexamethasone are dissolved in the basal medium.Further, some tools according to the present invention
Body example, the concentration of the SCF are 10~300ng/mL, preferably 100ng/mL.Some specific examples according to the present invention, it is described
The concentration of IGF-1 is 5~100ng/mL, preferably 40ng/mL.Some specific examples according to the present invention, the concentration of the EPO are
1~20U/mL, preferably 5U/mL.Some specific examples according to the present invention, the concentration of the transferrins are 10~300 μ g/
ML, preferably 100 μ g/mL.Some specific examples according to the present invention, the concentration of the dexamethasone are 0.1~3 μM, preferably 1 μM.
Specific example according to the present invention, the concentration of the PLURONICS F87 are 0.2-2mg/mL, preferably 1mg/mL.According to the present invention
Specific example, the concentration of the poloxamer188 is 0.2~2mg/mL, preferably 1mg/mL.It is trained as a result, using the kit
Hematopoietic stem/progenitor cells are supported, proliferation and/or erythroid differentiation are high-efficient, and effect is good.
Some specific examples according to the present invention, the hematopoietic stem/progenitor cells derive from selected from marrow, peripheral blood, placenta and
At least one of Cord blood, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.
Some specific examples according to the present invention, the kit include to be described previously for induction of hematopoiesis stem/progenitor cells
The culture medium of proliferation and/or erythroid differentiation.Thereby, it is possible to effective for induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation.
In accordance with a further aspect of the present invention, the present invention also provides a kind of promotion hematopoietic stem/progenitor cells proliferation and/or red systems
The method of differentiation.According to an embodiment of the invention, this method breaks up cultivating system culture institute using the induction containing poloxamer
State hematopoietic stem/progenitor cells.It is surprisingly found by the inventors that using this method can effectively facilitate hematopoietic stem/progenitor cells proliferation and/or it is red
System's differentiation, easy to operate, at low cost, proliferation and/or induction differentiation efficiency are high, and effect is good.
According to embodiments of the present invention, the poloxamer is PLURONICS F87 or poloxamer188.Hematopoietic Stem ancestral as a result,
Cell Proliferation and/or erythroid differentiation are high-efficient, and effect is good.
According to an embodiment of the invention, containing 0.2~2mg/mL, preferably 1mg/mL pool in the induction differentiation cultivating system
Luo Shamu 188.Be conducive to the proliferation and/or erythroid differentiation of hematopoietic stem/progenitor cells as a result,.
According to a further embodiment of the invention, contain the preferred 1mg/ of 0.2~2mg/mL in the induction differentiation cultivating system
ML poloxamer188.Be conducive to the proliferation and/or erythroid differentiation of hematopoietic stem/progenitor cells as a result,.
According to an embodiment of the invention, the induction differentiation cultivating system containing poloxamer is to be described previously for
The culture medium or kit of induction of hematopoiesis stem/progenitor cells proliferation and/or erythroid differentiation.As a result, hematopoietic stem/progenitor cells proliferation and/or
Erythroid differentiation is high-efficient, effect is good.
According to an embodiment of the invention, carrying out the culture 14-21 days, preferably 14 days or 21 days.Be conducive to hematopoiesis as a result,
Stem/progenitor cells proliferation and/or erythroid differentiation.
According to some embodiments of the present invention, with 1 × 106~3 × 106/ ml, preferably 2 × 106The cell density of/ml be into
The row culture.Hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation is high-efficient, effect is good as a result,.
According to an embodiment of the invention, the hematopoietic stem/progenitor cells derive from marrow, peripheral blood, placenta and Cord blood extremely
Few one kind, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.The i.e. described hematopoietic stem/progenitor cells come
Source is not particularly limited, and can obtain hematopoietic stem/progenitor cells by embryonic stem cell or inductive pluripotent stem cells vitro differentiation,
It can also be directly separated and obtain from marrow, peripheral blood, placenta or Cord blood, such as separation Cord blood is single directly from bleeding of the umbilicus
Nucleus, as hematopoietic stem/progenitor cells source.Hematopoietic stem/progenitor cells proliferation and/or erythroid differentiation is high-efficient, effect is good as a result,
The erythroid progenitor cells of a large amount of high quality can effectively be obtained.
Wherein it should be noted that expression way " induction of hematopoiesis stem/progenitor cells proliferation " used by herein had both included
Hematopoietic stem/progenitor cells proliferation also includes proliferation of the hematopoietic stem/progenitor cells to the intermediate cell during erythroid differentiation, such as red system
The proliferation of progenitor cells, i.e. poloxamer can also induce the proliferation of erythroid progenitor cells.Similarly, poloxamer can " induction be made
Blood stem/progenitor cells are to erythroid differentiation ", i.e., can either induction of hematopoiesis stem/progenitor cells break up to erythroid progenitor cells, moreover it is possible to further induction
Erythroid progenitor cells is divided into erythroid cells.
Additional aspect and advantage of the invention will be set forth in part in the description, and will partially become from the following description
Obviously, or practice through the invention is recognized.
Detailed description of the invention
Above-mentioned and/or additional aspect of the invention and advantage will become from the description of the embodiment in conjunction with the following figures
Obviously and it is readily appreciated that, in which:
Fig. 1 show according to an embodiment of the present invention, through it is external evoked culture 21 days after cell form,
Wherein,
Figure 1A shows the form of control group cell after external evoked 21 days,
Figure 1B shows the form that experimental group cell after external evoked 21 days of PLURONICS F87 is added in culture medium；
Fig. 2 shows that according to an embodiment of the present invention flow cytomery is red through external evoked 21 days cells
It is progenitor cell surface mark expression；
Fig. 3 shows according to an embodiment of the present invention, the form of cell after external evoked 14 days,
Wherein,
Fig. 3 A shows the form of control group cell after external evoked 14 days,
Fig. 3 B, which is shown, adds the experimental group of poloxamer188 through the form of external evoked 14 days cells in culture medium；
Fig. 4 shows according to an embodiment of the present invention, the red system of the external evoked 14 days cells of flow cytomery
Progenitor cell surface mark expression.
Specific embodiment
The solution of the present invention is explained below in conjunction with embodiment.It will be understood to those of skill in the art that following
Embodiment is merely to illustrate the present invention, and should not be taken as limiting the scope of the invention.Particular technique or item are not specified in embodiment
Part, it described technology or conditions or is carried out according to the literature in the art according to product description.Agents useful for same or instrument
Production firm person is not specified in device, and being can be with conventional products that are commercially available.
Embodiment 1
1, material:
Cord blood (please provide Cord Blood-Derived, source Beijing Cord Blood Bank, acquisition units Tongzhou healthcare hospital for women & children)；
Tissue Culture Flask (U.S., Corning)；
CO2Cell incubator (U.S., 3111 type of Thermo Scientific, Model)；
Erythroid progenitor cells induced medium: it is added to the Stemspan of SCF, IGF-1, EPO, transferrins and dexamethasone
(wherein the concentration of SCF is 100ng/mL, and the concentration of IGF-1 is 40ng/mL, and the concentration of EPO is 5U/mL, transferrins for culture
Concentration is 100 μ g/mL, and the concentration of dexamethasone is 1 μM).
2, erythroid progenitor cells induces
According to the method for the present invention, erythroid progenitor cells induction is carried out to human umbilical cord blood mononuclear cell, the specific steps are as follows:
Separation obtain human umbilical cord blood mononuclear cell after, be respectively adopted be added to 0.2mg/mL, 1mg/mL, 1.5mg/mL,
The erythroid progenitor cells induced medium (i.e. experiment be divided into four groups) of the PLURONICS F87 of 2mg/mL, using 24 porocyte culture plates,
37 degrees Celsius, 5%CO2The incubator culture human umbilical cord blood mononuclear cell, every hole volume of culture is 1ml, and cell density is
2x106/ml.Wherein, it tests not add the erythroid progenitor cells induced medium of PLURONICS F87 as compareing.
In vitro in Induction Process, the form of each phase cell is observed, and detects cell survival rate, cell number and cell table
Face mark.Specifically:
(1) inverted microscope observes cell
It being operated using inverted microscope (Olympus 7S100 type), inventor observes, at cell culture the 4th day, control
Group cell is in being dispersed in circle, differing in size, and cell number is more, as incubation time extends homogeneity cytosis；It is husky that pool Lip river is added
The cell aggregation of the experimental group of nurse 188 is agglomerating, is in tufted, not of uniform size.Wherein, to add the reality of 1mg/mL PLURONICS F87
It tests for group, the form of cell is as shown in Figure 1 after external evoked culture 21 days.
(2) cell biology detects
For adding the experimental group of 1mg/mL PLURONICS F87, cell survival rate, cell number detection, Yi Jili are carried out
With the expression of erythroid progenitor cells surface marker of the flow cytometer measurement through external evoked cell.Wherein, external evoked
14 days cell survival rates and cell number testing results see the table below 1.The detection of expression of cell surface marker is tested: taking induction the 21st
It cell is collected by centrifugation, and two pipes are divided into after brine, and it is anti-to be separately added into anti-human CD71-PE, CD235a-FITC
Body and Isotype control mouse IgG1, k-PE、IgG2b, k- FITC antibody, 4 DEG C, 30min, flow cytomery CD71, CD235a's
Expression, concrete outcome are shown in Table 2 and Fig. 2.
The external evoked cell survival rate of table 1 and cell number detection
The detection of the external evoked cell surface marker of table 2
Note:*The significant difference compared with the culture medium that PLURONICS F87 is not added
By table 1, table 2 and Fig. 2 it is found that PLURONICS F87 is added in culture medium, mononuclearcell can be promoted to red system ancestral
Cell induction differentiation and amplification.
Embodiment 2
1, material:
Cord blood (source Beijing Cord Blood Bank, acquisition units Tongzhou healthcare hospital for women & children)；
Tissue Culture Flask (U.S., Corning)；
CO2Cell incubator (U.S., 3111 type of Thermo Scientific, Model)；
Erythroid progenitor cells induced medium: it is added to the Stemspan of SCF, IGF-1, EPO, transferrins and dexamethasone
(wherein the concentration of SCF is 100ng/mL, and the concentration of IGF-1 is 40ng/mL, and the concentration of EPO is 5U/mL, transferrins for culture
Concentration is 100 μ g/mL, and the concentration of dexamethasone is 1 μM).
2, erythroid progenitor cells induces
According to the method for the present invention, erythroid progenitor cells induction is carried out to human umbilical cord blood mononuclear cell, the specific steps are as follows:
Separation obtain human umbilical cord blood mononuclear cell after, be respectively adopted be added to 0.2mg/mL, 1mg/mL, 1.5mg/mL,
The erythroid progenitor cells induced medium (i.e. experiment is divided into four groups) of the poloxamer188 of 2mg/mL, it is Celsius using 24 orifice plates, 37
Degree, 5%CO2The incubator culture human umbilical cord blood mononuclear cell, the volume of culture in every hole are 1ml, cell density 2x106/
ml.Wherein, it tests not add the erythroid progenitor cells induced medium of poloxamer188 as compareing.
In vitro in Induction Process, the form of each phase cell is observed, and detects cell survival rate, cell number and cell table
Face mark.Specifically:
(1) inverted microscope observes cell
It being operated using inverted microscope (Olympus 7S100 type), inventor observes, at cell culture the 4th day, control
For group cell in being dispersed in circle, differing in size, cell number is more, and as incubation time extends homogeneity cytosis, it is husky that pool Lip river is added
The cell aggregation of the experimental group of nurse 407 is agglomerating, is in tufted, not of uniform size.Wherein, to add the reality of 1mg/mL poloxamer188
It tests for group, the form of cell is as shown in Figure 3 after external evoked culture 14 days.
(2) cell biology detects
For adding the experimental group of 1mg/mL poloxamer188, cell survival rate, cell number detection, Yi Jili are carried out
With the expression of erythroid progenitor cells surface marker of the flow cytometer measurement through external evoked cell.Wherein, external evoked
14 days cell survival rates and cell number testing results see the table below 3.The detection of expression of cell surface marker is tested: taking induction the 14th
It cell is collected by centrifugation, and two pipes are divided into after brine, and it is anti-to be separately added into anti-human CD71-PE, CD235a-FITC
Body and Isotype control mouse IgG1,k-PE、IgG2b,k- FITC antibody, 4 DEG C, 30min, flow cytomery CD71, CD235a's
Expression, concrete outcome are shown in Table 4 and Fig. 4.
The external evoked cell survival rate of table 3 and cell number detection
The detection of the external evoked cell surface marker of table 4
Note:*The significant difference compared with the culture medium that poloxamer188 is not added
By table 3, table 4 and Fig. 4 it is found that poloxamer188 is added in culture medium, mononuclearcell can be remarkably promoted to red
It is progenitor cells induction differentiation and amplification.
In the description of this specification, reference term " one embodiment ", " some embodiments ", " example ", " specifically show
The description of example " or " some examples " etc. means specific features, structure, material or spy described in conjunction with this embodiment or example
Point is included at least one embodiment or example of the invention.In the present specification, schematic expression of the above terms are not
Centainly refer to identical embodiment or example.Moreover, particular features, structures, materials, or characteristics described can be any
One or more embodiment or examples in can be combined in any suitable manner.
Although an embodiment of the present invention has been shown and described, it will be understood by those skilled in the art that: not
A variety of change, modification, replacement and modification can be carried out to these embodiments in the case where being detached from the principle of the present invention and objective, this
The range of invention is defined by the claims and their equivalents.
Claims (41)
1. poloxamer is poloxamer improving the purposes in hematopoietic stem/progenitor erythroid differentiation efficiency, the poloxamer
188 or poloxamer188.
2. purposes according to claim 1, which is characterized in that the hematopoietic stem/progenitor, which derives from, is selected from marrow, periphery
At least one of blood, placenta and Cord blood, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.
3. poloxamer is preparing the purposes in culture medium, the culture medium is for improving hematopoietic stem/progenitor erythroid differentiation
Efficiency, the culture medium are the Stemspan culture medium for being added to SCF, IGF-1, EPO, transferrins and dexamethasone,
The poloxamer be PLURONICS F87 or poloxamer188,
The concentration of the PLURONICS F87 is 0.2~2mg/mL,
The concentration of the poloxamer188 is 0.2~2mg/mL.
4. purposes according to claim 3, which is characterized in that the concentration of the PLURONICS F87 is 1mg/mL.
5. purposes according to claim 3, which is characterized in that the concentration of the poloxamer188 is 1mg/mL.
6. purposes according to claim 3, which is characterized in that the concentration of the SCF is 10~300ng/mL.
7. purposes according to claim 6, which is characterized in that the concentration of the SCF is 100ng/mL.
8. purposes according to claim 3, which is characterized in that the concentration of the IGF-1 is 5~100ng/mL.
9. purposes according to claim 8, which is characterized in that the concentration of the IGF-1 is 40ng/mL.
10. purposes according to claim 3, which is characterized in that the concentration of the EPO is 1~20U/mL.
11. purposes according to claim 10, which is characterized in that the concentration of the EPO is 5U/mL.
12. purposes according to claim 3, which is characterized in that the concentration of the transferrins is 10~300 μ g/mL.
13. purposes according to claim 12, which is characterized in that the concentration of the transferrins is 100 μ g/mL.
14. purposes according to claim 3, which is characterized in that the concentration of the dexamethasone is 0.1~3 μM.
15. purposes according to claim 14, which is characterized in that the concentration of the dexamethasone is 1 μM.
16. purposes according to claim 3, which is characterized in that the hematopoietic stem/progenitor is derived from selected from marrow, outside
At least one of all blood, placenta and Cord blood, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.
17. a kind of for improving the kit of hematopoietic stem/progenitor erythroid differentiation efficiency, which is characterized in that in the kit
Containing poloxamer, SCF, IGF-1, EPO, transferrins and dexamethasone,
The poloxamer be PLURONICS F87 or poloxamer188,
The concentration of the PLURONICS F87 is 0.2~2mg/mL,
The concentration of the poloxamer188 is 0.2~2mg/mL.
18. kit according to claim 17, which is characterized in that further include basal medium, the basis training
Supporting base is Stemspan culture medium.
19. kit according to claim 18, which is characterized in that the PLURONICS F87, poloxamer188 are extremely
One of few and SCF, IGF-1, EPO, transferrins and dexamethasone are dissolved in the basal medium.
20. kit according to claim 19, which is characterized in that the concentration of the SCF is 10~300ng/mL.
21. kit according to claim 20, which is characterized in that the concentration of the SCF is 100ng/mL.
22. kit according to claim 19, which is characterized in that the concentration of the IGF-1 is 5~100ng/mL.
23. kit according to claim 22, which is characterized in that the concentration of the IGF-1 is 40ng/mL.
24. kit according to claim 19, which is characterized in that the concentration of the EPO is 1~20U/mL.
25. kit according to claim 24, which is characterized in that the concentration of the EPO is 5U/mL.
26. kit according to claim 19, which is characterized in that the concentration of the transferrins is 10~300 μ g/
mL。
27. kit according to claim 26, which is characterized in that the concentration of the transferrins is 100 μ g/mL.
28. kit according to claim 19, which is characterized in that the concentration of the dexamethasone is 0.1~3 μM.
29. kit according to claim 28, which is characterized in that the concentration of the dexamethasone is 1 μM.
30. kit according to claim 17, which is characterized in that the concentration of the PLURONICS F87 is 1mg/mL.
31. kit according to claim 17, which is characterized in that the concentration of the poloxamer188 is 1mg/mL.
32. kit according to claim 17, which is characterized in that the hematopoietic stem/progenitor derive from selected from marrow,
At least one of peripheral blood, placenta and Cord blood, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.
33. a kind of method for improving hematopoietic stem/progenitor erythroid differentiation efficiency, which is characterized in that utilize and contain poloxamer
Hematopoietic stem/progenitor described in induction differentiation cultivating system culture, contain in the induction differentiation cultivating system SCF, IGF-1,
EPO, transferrins and dexamethasone,
The poloxamer be PLURONICS F87 or poloxamer188,
Contain 0.2~2mg/mL PLURONICS F87 in the induction differentiation cultivating system,
Contain 0.2~2mg/mL poloxamer188 in the induction differentiation cultivating system.
34. according to the method for claim 33, which is characterized in that moored in the induction differentiation cultivating system containing 1mg/mL
Luo Shamu 188.
35. according to the method for claim 33, which is characterized in that moored in the induction differentiation cultivating system containing 1mg/mL
Luo Shamu 407.
36. according to the method for claim 33, which is characterized in that cultivating system is broken up in the induction containing poloxamer
For the described in any item culture mediums of claim 3-16 or the described in any item kits of claim 17-32.
37. according to the method for claim 33, which is characterized in that carry out the culture 7-21 days.
38. according to the method for claim 37, which is characterized in that carry out the culture 14 days or 21 days.
39. according to the method for claim 33, which is characterized in that with 1 × 106~3 × 106The cell density of/ml carries out institute
State culture.
40. according to the method for claim 39, which is characterized in that with 2 × 106The cell density of/ml carries out the culture.
41. according to the method for claim 33, which is characterized in that the hematopoietic stem/progenitor is derived from selected from marrow, outside
At least one of all blood, placenta and Cord blood, or obtained by embryonic stem cell or inductive pluripotent stem cells vitro differentiation.
Priority Applications (1)
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| poloxamers:a pharmaceutical excipients with therapeutic behaviors;hitesh r. patel1 et al.;《international journal of pharmtech research》;20091231;第11卷(第2期);299-303 * |
| 泊洛沙姆407的研究进展;高丽新等;《亚太传统医药》;20120131;第8卷(第1期);177-178 * |
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