CN105255835B - It is a kind of to express the cell membrane particles for melting memebrane protein and its preparation and application - Google Patents
It is a kind of to express the cell membrane particles for melting memebrane protein and its preparation and application Download PDFInfo
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Abstract
The present invention relates to a kind of express to melt the cell membrane particles and its preparation and application of memebrane protein, and VSVG is expressed in cell membrane particles；Cell membrane particles wrapping biological macromolecular and organelle；Cell membrane particles do not have nucleus.Preparation method: first with expression vesicular stomatitis virus albumen plasmid pair Ad293 cell transient transfection；Trypsin digestion cell is used after transfecting 48h, then is centrifuged 3min；Gained cell suspension is placed in syringe, and syringe is installed in the syringe-driven filter for having disposed polycarbonate membrane, push needle tubing make cell suspension from filter on one side release to it is other while, obtain expression VSVG cell membrane particles.The cell membrane particles that memebrane protein is melted in expression are applied in transmitting large biological molecule and organelle.Inventive film grains are bigger, energy wrapping biological macromolecular and mitochondria, functionally transferrin matter and mitochondria.Additional chemical reagent is not added, effectively maintains VSVG activity, improves after membrane granule enters inner body and discharges mitochondria efficiency, to improve mitochondria transmission efficiency.
Description
Technical field
The present invention relates to synthetic biology technologies more particularly to one kind can merge vesicular stomatitis virus glycoprotein membrane granule
And it is prepared and the application in transmitting large biological molecule and organelle.
Background technique
The study found that certain diseases, such as phenylketonuria result in patient's appearance due to lacking relevant protease
Abnormal symptom；And other disease, such as nervus retrogression Alzheimer disease, it is commonly called as Alzheimer, with mitochondria function
There can be greatest relationship extremely.Although realizing that the purpose for the treatment of has been achieved with great progress using drug delivery system, so
And these systems can't efficiently solve existing issue.
A variety of drug delivery systems are had been set up at present, for example, nano particle can pass through packaging or conjugated
Mode realize the transmitting of therapeutic protein and polypeptide, still, complicated production process, pharmaceutical activity decline and be difficult to
The obstacle for breaking through inner body limits the extensive use of this system invariably；In addition, the transmitting of albumen can also pass through allochthon
(exosome) it is realized with microvesicle (microvesicle), to overcome the problems, such as that operating process is cumbersome；And pass through vesicular stomatitis
Viral glycoprotein (VSV-G) nanometer membrane granule and mesh is transmitted using the VSV-G viroid body that reverse transcription stromatin Gag is obtained
Albumen, overcome the obstacle of inner body and improve the efficiency of transmitting.However VSV-G nanometers of membrane granules are too small, cannot wrap up more
The size of big substance, e.g. mitochondria, mitochondria can arrive micron order.
Compared with large biological molecule transmission system, organelle transmission system progress is very slowly and limited.The study found that line
Plastochondria can spontaneously pass through cell membrane nanotubes (membrane nanotubles) and transmit between cell, and can
Restore the aerobic respiration function of mitochondrial defects type cell；Mitochondria transmitting also may be implemented in mitochondria microinjection, but tight
The operation of lattice requires and special instrument and equipment is not suitable for batch and prepares；In addition, utilizing actinomycin D (actinomycin
D cellular functional " stoning ") can be made, become the cytosome containing mitochondrial, but some cells are by drug influence, still
Retain kernel function；Studies have reported that, the mitochondria of in-vitro separation can also be effectively transmitted using cell-penetrating peptide (CPP) recently.
But transmission efficiency is relatively low.
It is time-consuming and laborious and biological since that there are transmission efficiencies is low for the delivery system that transmits large biological molecule and organelle at present
The problems such as safety, therefore, the method for establishing a kind of efficiently transmitting large biological molecule and organelle has for treating a variety of diseases
Great meaning.
Summary of the invention
The first object of the present invention is to provide a kind of cell membrane particles for melting memebrane protein of expressing, it can be achieved that large biological molecule
With effective transmitting of organelle.
In order to solve the above-mentioned technical problem, the present invention adopts the following technical scheme that:
A kind of to express the cell membrane particles for melting memebrane protein, the cell membrane particles express vesicular stomatitis disease on cell membrane
Malicious glycoprotein；The cell membrane particles wrapping biological macromolecular and organelle；The cell membrane particles do not have nucleus, cell
Core DNA and nucleoprotein.Vesicular stomatitis virus glycoprotein has the function of to melt film.Wrapping biological macromolecular includes mRNA,
MicroRNA, mitochondrial DNA, cytoplasm protein, the large biological molecules such as epicyte protein.
Further, the cell membrane particles package protein and mitochondria.Mitochondria can be effectively wrapped up, and is transmitted
It is functioned into target cell.
Further, the cell membrane particles size is 3 μm ~ 5 μm.In transmitting substance process, the size of nano particle
It is that comparison is suitble to transmit substance, e.g. protein and some small-molecule drugs, and particle is bigger, the efficiency of transmitting can be
Decline.But the shortcomings that nano particle size is cannot to wrap up bigger substance, e.g. mitochondria, the size of mitochondria can
To arrive micron order, in order to solve the problems, it is necessary to which the bigger particle of preparation could effectively wrap up mitochondria.This hair
Bright cell membrane particles are 3-5 μm, although decreasing in transmission efficiency, can pack more substances, protect simultaneously
Demonstrate,prove certain transmission efficiency；And wrapped up vesicular stomatitis virus albumen on membrane granule and melted memebrane protein, improve membrane granule
The efficiency for discharging mitochondria after into inner body solves package mitochondria to greatly increase the transmission efficiency of mitochondria
The problem of, realize efficiently the quickly transmitting of organelle.
The second object of the present invention is to provide a kind of preparation method expressed and melt the cell membrane particles of memebrane protein, including with
Lower step:
(1) it is transiently transfected with expression vesicular stomatitis virus albumen plasmid pair Ad293 cell；
(2) after transfecting 48h, with trypsin digestion cell, it is centrifuged the expression vesicular stomatitis virus sugar to be suspended after 3min
The cell of albumen；
(3) cell suspension obtained by step (2) is placed in syringe, and syringe is installed to the syringe needle for having disposed polycarbonate membrane
In formula filter, push needle tubing make cell suspension from filter on one side release to it is other while, acquisition expression vesicular stomatitis disease
The cell membrane particles of malicious glycoprotein.
Pancreatin is a kind of protease, by protein degradation on location, makes iuntercellular junction and cell-culture dish
Locate protein degradation, at this moment cell is due to becoming spherical shape under the tension force effect of therein cytoskeleton, so that cell be made to separate.
Further, the step (1) the following steps are included:
(11) the expression vesicular stomatitis virus albumen plasmid of 1mg is added in centrifuge tube, 50 μ l free serum cultures is added
Liquid will mix；
(12) serum-free medium of 3ml Polyjet transfection reagent and 50ml are added in another centrifuge tube, are gently shaken
It is even；Then it is added in step (12) acquired solution；Transfection cocktail is obtained, 5-10min is stored at room temperature；
(13) culture solution is sucked, and adds 1ml fresh serum free culture solution, transfection cocktail is added to Ad293 at once
Cell gently shakes up；
(14) culture solution more renewed after 12h.
Polyjet transfection reagent is a kind of DNA transfection reagent of biodegradable high molecular polymer, PolyJet examination
Agent the fixed DNA of low concentration can migrate during electrophoresis, and form transfecting complexes in 5 minutes.It can transfect
Rapid and degradable polymer, can substantially reduce cytotoxicity in this way after polymer cell endocytosis.
Further, every 1x106Ad293 cell, with the expression vesicular stomatitis virus albumen plasmid transfection of 1mg.
Further, the polycarbonate membrane pore size is 3 μm；The filter is the filtering of 25mm film changeable needle-based
Device.Being covered with diameter above polycarbonate membrane is 3 μm of circle, advantageously forms that form is complete, film is melted in expression of uniform size
The cell membrane particles of albumen.
The third object of the present invention is to provide a kind of cell membrane particles for melting memebrane protein of expressing in transmitting large biological molecule
With the application in organelle.Purpose large biological molecule and organelle are transmitted to the cell of large biological molecule and organelle defect
In.The cell membrane particles that memebrane protein is melted in expression can effectively wrap up mitochondria, and be transmitted in target cell and function.It can be with
Purpose large biological molecule and organelle are transmitted in the cell of large biological molecule and organelle defect, restore the normal function of cell
Energy.
Compared with prior art, expression of the invention melt memebrane protein cell membrane particles particle it is bigger, package and transmitting more
More substances, not only can wrapping biological macromolecular and mitochondria, and functionally can transferrin matter and mitochondria, and make
The substance of transmitting exercises normal function.Addition additional chemical reagent is not needed when preparation, effectively maintains vesicular stomatitis disease
The bioactivity of toxalbumin, the PolyJet transfection reagent that when preparation uses can fix deoxyribose core by low concentration during electrophoresis
Acid migration, and transfecting complexes are formed in 5 minutes, being formed by polymer can be after transfecting polymer cell endocytosis rapidly
And degradable polymer, cytotoxicity can be substantially reduced in this way.The cell membrane particles of memebrane protein are melted in the expression being prepared
It is uniform in size, form is complete.It is 3 μm that the present invention uses 25mm film changeable syringe-driven filter, syringe and pore size for the first time
The vesicular stomatitis virus albumen plasmid cell film membrane granule that the quick and convenient preparation diameter of polycarbonate membrane is about 3 μm, this
Kind of membrane granule can wrap up protein and mitochondria, reduce degradation of the protein mitochondria in vitro in transmittance process, although
Grain is bigger than nano particle, but can pack more substances, while guaranteeing certain transmission efficiency；And it is wrapped on membrane granule
It has wrapped up in and has melted memebrane protein vesicular stomatitis virus albumen, the efficiency that membrane granule enters release mitochondria after inner body has been improved, thus greatly
The earth improves the transmission efficiency of mitochondria, this is not only that research large biological molecule and the relevant disease of organelle defect are established
Basis, and it is significant for research cell nucleocytoplasmic relation.
Detailed description of the invention
Fig. 1 is the preparation method schematic diagram for the cell membrane particles that memebrane protein is melted in present invention expression；
Fig. 2 is the phenogram for the cell membrane particles that memebrane protein is melted in present invention expression, wherein 293/EGFP, 293/Mito, and
293/H2B respectively indicates 293 cells of expression cytoplasm green fluorescent protein, expresses 293 cells of mitochondria green fluorescent protein
With 293 cells of expression cell core H2B albumen and the fusion protein of green fluorescent protein, one row's figure of Far Left is cell split
Figure, other three rows are from left to right followed successively by the light field, green fluorescent protein and split figure of cell membrane particles；
Fig. 3 be pLP-VSVG(of the present invention express vesicular stomatitis virus albumen) and pDsRed-Expressed(express it is red
The plasmid of color fluorescin) expression obtained from cotransfection Ad293 cell melts the cell membrane particles of memebrane protein, and it is thin with Ad293 target
Fluorescence Identification figure after born of the same parents' incubation, a are the fluorescence photo of 5h after being incubated for, and b is the fluorescence photo of 12h after being incubated for；
Fig. 4 is that film is melted in expression obtained from pLP-VSVG and pDsRed-Expressed cotransfection Ad293 cell of the present invention
The Efficiency Statistics figure of the cell membrane particles of albumen；
Fig. 5 is table obtained from pLP-VSVG of the present invention and pCMV (CMC promoter)-CreER cotransfection Ad293 cell
Up to the cell membrane particles for melting memebrane protein, reporter gene with pCMV-DsRed/loxP2/DsRed(Loxp system) recipient cell incubates
Fluorescence photo after educating 48h, from left to right respectively DsRed(red fluorescent protein), EGFP(green fluorescent protein it is positive) and
Split figure；
Fig. 6 is that memebrane protein is melted in expression obtained from pLP-VSVG and pCMV-CreER cotransfection Ad293 cell of the present invention
EGFP cell and DsRed negative cells after cell membrane particles, with pCMV-DsRed/loxP2/DsRed recipient cell incubation 48h
Ration statistics qualification figure；
Fig. 7 is the green fluorescent protein that pLP-VSVG of the present invention and pMito-EGFP(expression can enter mitochondria) cotransfection
The cell membrane particles of memebrane protein are melted in expression obtained from Ad293 cell, the fluorescence photo of 5h and 12h after being incubated for target cell
Fig. 8 is that memebrane protein is melted in expression obtained from pLP-VSVG and pMito-EGFP cotransfection Ad293 cell of the present invention
The Efficiency Statistics figure of cell membrane particles transmitting；
Fig. 9 is mitochondria after present invention removal ethidium bromide (EB), pyruvic acid (pyruvate), uridine (uridine)
Deficient cell HeLa Rho0(mitochondrial defects type) it is incubated for VSV-G cell membrane particles, and in the 5th day progress mitochondria shape
State fluorescent staining qualification figure is from left to right followed successively by light field, mitochondria staining reagent and split figure；
Figure 10 is mitochondria after present invention removal ethidium bromide (EB), pyruvic acid (pyruvate), uridine (uridine)
Deficient cell HeLa Rho0 and VSV-G cell membrane particles were incubated for, and in the 5th day progress mitochondrial membrane potential fluorescent staining mirror
Fixed figure, is from left to right followed successively by light field, green, red and split figure；
Figure 11 is mitochondria after the present invention removes ethidium bromide (EB), pyruvic acid (pyruvate), uridine (uridine)
Deficient cell HeLa Rho0 and VSV-G cell membrane particles are incubated for, and the quantity quantitative analysis figure of cell was carried out at the 5th day；
Figure 12 is mitochondria after the present invention removes ethidium bromide (EB), pyruvic acid (pyruvate), uridine (uridine)
Deficient cell HeLa Rho0 and VSV-G cell membrane particles are incubated for, and the quantitative analysis of mitochondrial membrane potential was carried out at the 5th day
Figure；
Wherein R represents red in picture, and G represents green, and B represents blue.
Specific embodiment
To make the object, technical solutions and advantages of the present invention clearer, the present invention is made into one below in conjunction with attached drawing
Step ground detailed description.
Embodiment 1
It is a kind of to express the preparation for melting the cell membrane particles (PMVs) of memebrane protein.Every 1x106Ad293 cell, with the table of 1mg
Up to vesicular stomatitis virus albumen (pLP-VSVG) plasmid transfection.
Preparation method is added the pLP-VSVG plasmid of 1 μ g in a 1.5ml centrifuge tube as shown in Figure 1:, be added 50 μ l without
Serum free culture system liquid mixes；3 μ l Polyjet transfection reagents and the serum-free training of 50ml are added inside new 1.5ml centrifuge tube
Nutrient solution, and it is added in plasmid solution, it mixes well, obtains transfection cocktail, be stored at room temperature 5-10min, suck culture
Culture solution in plate cell, and 1ml fresh medium is added, transfection cocktail is added to cell at once, is gently shaken up.12h
The culture solution more renewed afterwards, digestion obtains cell suspension 800g and is centrifuged 3min after 48h.Under normal circumstances, more in order to obtain
Cell can at most transfect 6x10 simultaneously6Cell, with 5ml culture solution be resuspended.
Pore size it is that 3 μm of polycarbonate membrane is placed in 25mm film changeable syringe-driven filter, and by filter
It screws, draws cell suspension in syringe with the syringe of 10ml, cell is firmly ,ed use on one side shifting while other onto from filter
The centrifuge tube of 15ml is collected.
The fresh cultured for adding 5ml again repeats the above steps.
Obtained VSV-G membrane granule 1000g centrifugation 10min is collected, supernatant is lightly drawn, leaves the training of general 100ml
Nutrient solution, and gently shake centrifuge tube, obtains that size is relatively uniform, about 3 μm of VSV-G cell membrane particles.
The PMVs size being prepared is 3 μm ~ 5 μm, and vesicular stomatitis virus glycoprotein, package life are expressed on cell membrane
Object macromolecular and organelle endocytic receptor cell are merged by the film under low ph value, and the substance discharged in cytoplasma membrane particle can
For transmitting in large biological molecule and organelle.Purpose large biological molecule and organelle are transmitted to large biological molecule and cell
In the cell of device defect.
Embodiment 2
The characterization of memebrane protein cell membrane particles is melted in expression.
Transfect the plasmid of 1 ug pEGFP-N1(expressing green fluorescent protein respectively to Ad293 cell transfecting), pMito-
EGFP, pH2B-EGFP(express nucleoprotein H2B and green fluorescent protein fusion protein plasmid), 48 hours after transfection, and by upper
Cell membrane particles obtained from the extruded film mode of the use of embodiment 1 are stated, are observed under Laser Scanning Confocal Microscope.In addition also pass through
The technologies such as DNA isolation technics, real-time round pcr and western blotting are characterized.From having transfected pEGFP-
Cell membrane particles obtained in the cell of N1, pMito-EGFP, pH2B-EGFP, these three plasmids mark cell cytoplasm respectively, carefully
Born of the same parents' mitochondria and nucleus.Membrane granule includes mitochondrial DNA, cell RNA and miRNA, cytoplasmic protein as can be seen from Figure 2
And memebrane protein, but do not include nucleus DNA, Nuclear extract matter.Therefore membrane granule wraps up various large biological molecules and cell
Device, but do not have nucleus.
Embodiment 3
The monitoring and functional verification of memebrane protein cell membrane particles transferrin matter are melted in expression.
With the plasmid of 1mg pLP-VSVG and 1 ug pDsRed-Expressed(expression red fluorescent protein) cotransfection
Ad293 cell, 48h after transfection obtain VSV-G cell membrane particles using 1 extruded film of above-described embodiment, and are added to 1x104
In the middle of Ad293, vitellophag after 5h is put into 35mm and is copolymerized on burnt culture dish, point is contaminating core in 2h and 12h DAPI, and
It is observed under Laser Scanning Confocal Microscope.With 1mg pLP-VSVG and 1mg pCMV (CMC promoter)-CreER (Cre recombinase and female
Hormone receptor fusion protein) cotransfection Ad293 cell, 48h after transfection, using above-mentioned extruded film acquisition VSV-G cell membrane particles, and
And it is added to 1x104In pCMV-DsRed/loxP2/DsRed (reporter gene of Cre-Loxp system) recipient cell, training
The 4- hydroxyl tamoxifen (4-HT) of nutrient solution addition 10mM.48h carries out fluorescence microscope.It can be seen that and incubate from Fig. 3 and Fig. 4
It after educating 5h, is observed under laser confocal microscope, red fluorescence is presented in about 40% cell, and general 12h, membrane granule are released
It puts on protein to target cell matter.It can be seen that PMVs can be delivered successfully CreER fusion protein, CreER from Fig. 5 and Fig. 6
Fusion protein can enter nucleus from cytoplasm, and DNA recombination is carried out between the site loxP, turn recipient cell from red
For green.In fluorescence microscopy microscopic observation after 48h, the cell of discovery about 18% is under the action of tamosifen (4-HT) from red
Discoloration is green.The visible membrane granule of the above results can be successfully entered target cell, and after entrance cell, membrane granule can successfully discharge
Red protein matter, it was demonstrated that VSV-G cell membrane particles effectively transferrin matter and can play biological function.
Embodiment 4
The monitoring and functional verification of VSV-G membrane granule transmitting mitochondria.
Can enter the green fluorescent protein of mitochondria with 1mg pLP-VSVG and 1mg pMito-EGFP(expression) cotransfection
Ad293 cell, 48h after transfection obtain VSV-G cell membrane particles using 1 extruded film of above-described embodiment, and are added to 1x104
In the middle of Ad293, vitellophag after 5h is put into 35mm and is copolymerized on burnt culture dish, divides in 2h and 12h DAPI(4', 6- bis-
Amidino groups -2-phenylindone) dye core, and observed under Laser Scanning Confocal Microscope.It can be seen that from Fig. 7 to Fig. 8 after being incubated for 5h, in laser
It is observed under Laser Scanning Confocal Microscope, green fluorescence is presented in about 20% cell, and after general 12 hours, the line grain of green occurs
Body structure.
Ad293 cell is transfected with 1mg pLP-VSVG, it is thin to obtain VSV-G using 1 extruded film of above-described embodiment by 48h after transfection
After birth particle, and it is added to 1x104HeLa Rho0(mitochondrial defects type) in the middle of cell, in addition, removing in cell
Ethidium bromide, pyruvic acid, uridine replace fresh cultured after being incubated for 12h.1mM MitoTracker(line grain is used after 5 days respectively
Body staining reagent) and DAPI dyed, and observed under Laser Scanning Confocal Microscope；In addition, in order to detect cell membrane potential,
It is dyed with 5mg/ml JC-1 line (mitochondrial membrane potential staining reagent), film potential is expressed as red shading value and green luminosity
Ratio between value, and it is for statistical analysis with ImageJ software.Finally, carrying out the statistics of cell quantity with blood counting chamber.
Discovery cell quantity be can be seen that from Fig. 9 to Figure 12 one times more than control group, Mitochondrial Shape is closer to than control group
Normal cell, and mitochondrial membrane potential in anoxic has more three times than control group.The visible membrane granule of experimental result can be successfully entered target
Cell, and after entrance cell, membrane granule can successfully discharge mitochondria, and mitochondria can replicate in target cell.It is above-mentioned
As a result it VSV-G cell membrane particles are demonstrated can effectively transmit mitochondria and concurrently wave biological function.
Above disclosed is only presently preferred embodiments of the present invention, cannot limit the right of the present invention with this certainly
Range, therefore equivalent changes made in accordance with the claims of the present invention, are still within the scope of the present invention.
Claims (5)
1. a kind of express the cell membrane particles for melting memebrane protein, which is characterized in that the cell membrane particles express water on cell membrane
Blister stomatitis viral glycoprotein；The cell membrane particles wrapping biological macromolecular and organelle；The cell membrane particles do not have
Nucleus；The organelle includes mitochondria；The cell membrane particles size is 3 μm~5 μm.
2. the preparation method that the cell membrane particles of memebrane protein are melted in a kind of expression as described in claim 1, which is characterized in that including
Following steps:
(1) it is transiently transfected with expression vesicular stomatitis virus albumen plasmid pair Ad293 cell；
(2) after transfecting 48h, with trypsin digestion cell, it is centrifuged the expression vesicular stomatitis virus glycoprotein to be suspended after 3min
Cell；
(3) cell suspension obtained by step (2) is placed in syringe, and syringe is installed to the needle-based mistake for having disposed polycarbonate membrane
In filter, push needle tubing make cell suspension from filter on one side release to it is other while, acquisition expression vesicular stomatitis virus sugar
The cell membrane particles of albumen；
The polycarbonate membrane pore size is 3 μm；The filter is 25mm film changeable syringe-driven filter.
3. according to claim 2 express the preparation method for melting the cell membrane particles of memebrane protein, which is characterized in that the step
Suddenly (1) the following steps are included:
(11) the expression vesicular stomatitis virus albumen plasmid of 1 μ g is added in centrifuge tube, 50 μ l serum-free mediums are added will
It mixes；
(12) serum-free medium of 3 μ l Polyjet transfection reagents and 50 μ l are added in another centrifuge tube, are gently shaken up；So
It is added in step (12) acquired solution afterwards；Transfection cocktail is obtained, 5-10min is stored at room temperature；
(13) culture solution is sucked, and adds 1ml fresh serum free culture solution, transfection cocktail is added to Ad293 cell at once,
Gently shake up；
(14) culture solution more renewed after 12h.
4. according to claim 3 express the preparation method for melting the cell membrane particles of memebrane protein, which is characterized in that every 1x106
Ad293 cell, with the expression vesicular stomatitis virus albumen plasmid transfection of 1 μ g.
5. a kind of expression as described in claim 1 melts the cell membrane particles of memebrane protein in transmitting large biological molecule and organelle
Using the diagnosing and treating using non-disease.
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Non-Patent Citations (4)
| Title |
|---|
| Preparation of virosomes coated with the vesicular stomatitis virus glycoprotein as efficient gene transfer vehicles for animal cells;Shoji J等;《Microbiol Immunol.》;20041231;第48卷(第3期);摘要 * |
| Preparation, characterization, and surface immobilization of native vesicles obtained by mechanical extrusion of mammalian cells;Huawen Wu等;《Integr. Biol.》;20120412;第686页右栏第4段至第687页左栏第2段、第688页右栏第1段、第691段左栏第4段 * |
| Protein Transfer Into Human Cells by VSV-G-induced Nanovesicles;Philippe-Emmanuel Mangeot等;《The American Society of Gene & Cell Therapy》;20110930;第19卷(第9期);摘要和第1658页右栏倒数第9-10行、第1664页左栏第5段、第1658页右栏倒数第14行 * |
| Treatment of human cells derived from MERRF syndrome by peptide-mediated mitochondrial delivery;JUI-CHIH CHANG等;《Cytotherapy》;20131231;1580-1596 * |
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