CN108042510A - A kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque and application thereof - Google Patents
A kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque and application thereof Download PDFInfo
- Publication number
- CN108042510A CN108042510A CN201810140423.1A CN201810140423A CN108042510A CN 108042510 A CN108042510 A CN 108042510A CN 201810140423 A CN201810140423 A CN 201810140423A CN 108042510 A CN108042510 A CN 108042510A
- Authority
- CN
- China
- Prior art keywords
- rapamycin
- platelet membrane
- pnp
- platelet
- coated
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/51—Nanocapsules; Nanoparticles
- A61K9/5107—Excipients; Inactive ingredients
- A61K9/5176—Compounds of unknown constitution, e.g. material from plants or animals
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
- A61K31/436—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a six-membered ring having oxygen as a ring hetero atom, e.g. rapamycin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/51—Nanocapsules; Nanoparticles
- A61K9/5107—Excipients; Inactive ingredients
- A61K9/513—Organic macromolecular compounds; Dendrimers
- A61K9/5146—Organic macromolecular compounds; Dendrimers obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyethylene glycol, polyamines, polyanhydrides
Abstract
The present invention relates to a kind of novel form of rapamycin, i.e., a kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque.The present invention also provides the preparation method and applications of the novel form.Its advantage is shown：The present invention develops a kind of coated rapamycin bionic nano particle of platelet membrane (RAP PNP) for targeting atherosclerotic plaque, rapamycin is contained using nano-carrier, nano-carrier pan coating platelet membrane, utilize the attachment proteins on platelet membrane, simulation blood platelet is to the natural ability of going back to the nest of atherosclerotic plaque, targeted delivery rapamycin is local to atherosclerotic plaque, improve drug concentration in patch, activating macrophage autophagy, inhibit plaque progression and stablize patch, the effect of improving rapamycin antiatherosclerosis, and reduce its Systemic reaction.
Description
Technical field
The present invention relates to technical field of pharmaceuticals, are a kind of platelet membranes for targeting atherosclerotic plaque specifically
Coated rapamycin bionic nano particle and its purposes in the drug for preparing treatment atherosclerosis disease.
Background technology
Atherosclerosis (AS) is a kind of chronic inflammation and metabolic disease, causes a variety of cardiovascular and cerebrovascular diseases
Pathophysiological basis and main cause have higher disability rate and the death rate.In the past few years, the treatment in relation to AS constantly obtains prominent
Broken property progress, the extensive use of antiplatelet, statins and interventional treatment (PCI) have greatly facilitated the pre- of AS patient
Afterwards.However, reverse and stablize patch be still at present most intractable basis and clinical problem, it is necessary to develop more efficiently means and
Drug therapy atherosclerosis.
Rapamycin (Rapamycin, RAP) and its derivative (everolimus, Zuo Tamosi etc.) are clinically wide at present
The immunosuppressor of general application has a variety of effects such as immunosupress, anti-inflammatory, anti-proliferate, Induces Autophagy.There is multinomial research
Authentication system application RAP and its derivative play an important role of to inhibit progression of atherosclerosis and stablize patch.Work as local concentration
When reaching certain level, rapamycin and the like can activate the autophagy process of macrophage in patch, so as to reverse plaque
And promote plaque stability (Martinet, W., De Loof, H.＆De Meyer, G.R., 2014.mTOR inhibition:a
promising strategy for stabilization of atherosclerotic
plaques.Atherosclerosis,233,601-7).In vitro study confirms the medicine needed for macrophage autophagy in activation patch
Object concentration is far longer than accessible concentration when being administered systemically.Meanwhile although systemic dosage increase also can be improved accordingly
The drug concentration of patch part, but the poisonous side effect of medicine such as rapamycin are larger, wherein caused hypercholesterolemia and high blood
Sugar can promote AS to be in progress in turn again.Clinically the local administration of rapamycin and the like generally relies on instrument such as medicine at present
Object sacculus or bracket for eluting medicament mediation.But there are certain limitations for the drug delivery of instrument mediation：(1) limitations：Only
It can guarantee that stent and the patch of the adherent part of sacculus have abundant drug, remaining coronary arteries patch, the equal nothing of whole body internal organs artery plaque
Covering.And AS is substantially general vascular conditions, artery A S treatments need to take all artery plaques into account.It is more awkward, spot
Block vulnerability has more prognostic value than hemadostewnosis degree, and intervention at present is intervened on the basis of stenotic severity, can not
Estimation endangers the Vulnerable plaque of heavier light moderate stenosis.(2) time limitation：Only can just it implement in interventional treatment,
Stent drug release is fast, lasts for hours to several weeks, and AS is substantially a kind of chronic inflammation disease, it is necessary to long-term administration.
(3) though balloon dilatation and stenting discharge the therapeutic effect with mechanical support, due to vascular dissection damage and foreign body reaction without
It doubts and increases the atheromatous plaque for starting a new round, counteract the anti-plaque efficiency of drug to a certain extent.Rapamycin application
Limitation highlighted exploitation rapamycin novel form strongly, for atherosclerotic plaque part targeted delivery rapamycin
Drug improves curative effect, and reducing toxicity has important clinical meaning.
Blood platelet is the biologically active fritter kytoplasm that ripe megacaryocyte kytoplasm crack releasing gets off from marrow,
Nearest research has shown that blood platelet not only takes part in the forming process of thrombus after plaque rupture, is sent out in atherosclerosis
Important function (Wu, M.D., Atkinson, T.M.＆Lindner, J.R., 2017.Platelets are also played during exhibition
and von Willebrand factor in atherogenesis.Blood,129,1415-1419).Atherosclerosis
Local inflammation reaction causes vascular remodeling, promotes hematoblastic adherency and aggregation, and further forms blood with inflammatory cell
Platelet monocyte compound, causes the formation of vulnerable plaque, and the erosion or rupture of plaque fibrous cap cause spot is in the block to promote to coagulate
Substance exposes, and further promotes hematoblastic aggregation and forms thrombus, ultimately results in acute myocardial infarction AMI.By the work of fluorescent marker
Change blood platelet and be injected to ApoE gene knockouts (ApoE-/-) in Mice Body, it is seen that it is to going back to the nest at atherosclerotic lesion, spot
Platelet content can be as plaque progression and unstable index in block.Equally, can be sent out in the atherosclerotic plaque of people
Now due to a large amount of platelet aggregations caused by the increase of new vessels permeability, bleeding etc. inside patch.Naturally returned using blood platelet
The characteristics of nest atherosclerotic plaque, design and structure imitate hematoblastic nanoscale medicine delivery system, are coated with rapamycin drug, have
Hoping increases targeted delivery of the rapamycin to patch, improves target site drug concentration, increases curative effect, reduces side reaction.
Traditional blood platelet simulation strategy mainly utilizes the known more single small peptide for simulating platelet adhesion reaction molecule
Or ligand modified nano particle, it is difficult to really simulate blood platelet, in spot natural adhesive attraction in the block, simultaneously efficient targeting artery is athero-
Plaque (Farokhzad, O.C., 2015.Nanotechnology:Platelet mimicry.Nature,526,47-
8).Cell membrane is coated in nano grain surface by the coated bionic nano particle of cell membrane, so that nano particle is provided simultaneously with
Biomembrane and the function of synthesizing film become hot spot (Fang, R.H., Jiang, Y., Fang, the J.C.＆ of current clinical research
Zhang,L.,2017.Cell membrane-derived nanomaterials for biomedical
applications.Biomaterials,128,69-83).At present there is no contained using the coated bionic nano particle of cell membrane
The correlative study and relevant patent application of rapamycin targeted therapy atherosclerotic plaque.
According to background above, consider blood platelet to the natural targeting and targeted delivery rapamycin of patch in treatment artery
Important function in plaque disease, the present invention construct the coated rapamycin bionic nano particle (RAP- of platelet membrane
PNP), rapamycin is contained using nano-medicament carrier, platelet membrane is coated in carrier surface, by intravenous injection, realize target
It is local to delivering rapamycin to atherosclerotic plaque, intra-arterial macrophage autophagy is activated, inhibits plaque progression, increases patch
The effect of stability, raising rapamycin treatment atherosclerosis disease.
The content of the invention
First purpose of the present invention is to overcome the various deficiencies of the administration of rapamycin legacy system and coating stent of medicine
And side effect, a kind of novel form of rapamycin is provided, i.e., a kind of platelet membrane coating for targeting atherosclerotic plaque
Rapamycin bionic nano particle (RAP-PNP).
Second object of the present invention is to provide a kind of preparation method of rapamycin novel form.
Third object of the present invention is to provide a kind of application of rapamycin novel form.
To realize above-mentioned first purpose, the present invention adopts the technical scheme that：A kind of group for treating atherosclerosis
Object is closed, the composition is pan coating platelet membrane, the internal nano-carrier for containing mTOR inhibitors.
Preferably, the mTOR inhibitors are rapamycin or derivatives thereof.
The nano-carrier can be any polymer that can contain rapamycin or derivatives thereof, it is preferred that institute
The material for the nano-carrier stated is poly- acetic acid-hydroxyacetic acid.It is furthermore preferred that the nano-carrier is poly- acetic acid-hydroxyacetic acid
The nanoparticle prepared by the precipitation method.
Preferably, the grain size of the nanoparticle is 100-200nm.
Preferably, the platelet membrane is coated in nano-carrier surface after polyethyleneglycol modified.It is it is furthermore preferred that described
Polyethylene glycol be DSPE-PEG2000.
Preferably, the platelet membrane is humanized's platelet membrane, is extracted by human blood platelets.
Preferably, 1~10 × 109The nanoparticle of the platelet membrane coating 1mg of a blood platelet extraction.It is furthermore preferred that 3 × 109
The nanoparticle of the platelet membrane coating 1mg of a blood platelet extraction.
To realize above-mentioned second purpose, the present invention adopts the technical scheme that：A kind of targeting atherosclerotic plaque
The coated rapamycin bionic nano particle of platelet membrane preparation method, the preparation method comprises the following steps：
(1) rapamycin nanoparticle carrier is prepared；
(2) platelet membrane is prepared, and is carried out polyethyleneglycol modified；
(3) polyethyleneglycol modified platelet membrane is coated in rapamycin nanoparticle carrier surface.
To realize above-mentioned 3rd purpose, the present invention adopts the technical scheme that：A kind of targeting atherosclerotic plaque
The coated rapamycin bionic nano particle of platelet membrane prepare treatment atherosclerosis drug in application.
The technical solution adopted by the present invention：
1) synthesize rapamycin nanoparticle carrier, prepare platelet membrane, and carry out it is polyethyleneglycol modified, it is by ultrasound that blood is small
Plate film is coated in rapamycin nanoparticle carrier surface, its physicochemical property is characterized, such as grain size, current potential, form, skin covering of the surface egg
White integration, rapamycin envelop rate and drugloading rate, nano particle circulation time, drug release patterns etc..
2) using after fluorescent dye DiD marked with nanometer granule, by with collagen, fibrin and von Willebrand disease
The vitro binding assay evaluation coated bionic nano particle of platelet membrane of the factor (von Willebrand factor, vWF)
(PNP) simulate platelet adhering function ability, by with atherosclerotic plaque in mice and people's Carotid arteriosclerosis plaque
The vitro binding assay of block evaluates its affinity to atherosclerotic plaque.
3) after using fluorescent dye DiD marked with nanometer granule, the coated bionic nano particle of tail vein injection platelet membrane
(PNP), by targeting atherosclerotic plaque in the coated bionic nano granule of isolated organ imaging in evaluation platelet membrane
Targeting and specificity.
4) the coated rapamycin bionic nano particle (RAP-PNP) of platelet membrane is investigated to atherosclerotic plaque
Therapeutic effect.
5) coated rapamycin bionic nano particle (RAP-PNP) the treatment artery atherosis spot of platelet membrane is investigated
Macrophage autophagy situation further inquires into platelet membrane coating rapamycin bionic nano granule therapy Atherosclerosis in block
Change the mechanism of disease.
The invention has the advantages that：
The present invention develops a kind of coated rapamycin bionic nano of platelet membrane for targeting atherosclerotic plaque
Particle (RAP-PNP) is a kind of rapamycin (RAP) novel pharmaceutical formulation, i.e., contains rapamycin using nano-carrier, and nanometer carries
Body pan coating platelet membrane, using the attachment proteins on platelet membrane, simulation blood platelet is to the day of atherosclerotic plaque
Right ability of going back to the nest, targeted delivery rapamycin is local to atherosclerotic plaque, improves drug concentration in patch, activates macrophage
Cell autophagy the effect of inhibiting plaque progression and stablize patch, improve rapamycin antiatherosclerosis, and reduces its whole body
Property adverse reaction.
Description of the drawings
Attached drawing 1：The schematic diagram of the coated rapamycin bionic nano particle preparation (RAP-PNP) of platelet membrane.Wherein, thunder
Pa mycin is contained in nano-carrier, and after platelet membrane is polyethyleneglycol modified, nano-carrier table is coated in by electrostatic interaction
Face.
Attached drawing 2：The characterization result of the coated rapamycin nanoparticle particle of platelet membrane.Figure A is blood platelet vesica Electronic Speculum knot
Fruit, figure B are PLGA nano-carriers Electronic Speculum as a result, figure C is the coated bionic nano particle Electronic Speculum result of platelet membrane.Figure D is grain
Footpath is distributed.Figure E is Zeta potential.Figure F is shelf-stability.Figure G measures that platelet membrane is coated bionical to be received for Coomassie brilliant blue
Rice grain surface membrane protein integration, figure H are special for the coated bionic nano particle surface of western blot determination platelet membrane
Sign property memebrane protein integration.Figure I is rapamycin drugloading rate, and figure J is rapamycin envelop rate.
Attached drawing 3：Long circulating feature and the release of external rapamycin drug in the coated bionic nano granule of platelet membrane
Feature.It is PNP circulation time curves in vivo to scheme A.Figure B is rapamycin drug release patterns in vitro.
Attached drawing 4：The in-vitro simulated platelet adhering function of the coated bionic nano particle (PNP) of platelet membrane and to patch
Binding ability.It is that for PNP to the binding ability of collagen, figure B is PNP under static state to fibrin under static state to scheme A
Binding ability.It is that for PNP to the binding ability of collagen, figure D is that flow regime descends PNP to fiber egg under flow regime to scheme C
White binding ability, figure E are PNP under flow regime to the binding ability of vWF.Figure F and figure G is external PNP to rat aorta congee
The binding ability and relative value of sample patch are not coated with the common nanoparticle (NP) of platelet membrane as control.Figure H is PNP bodies
Outside with people's carotid artery atherosclerosis plaques binding ability, NP is as control.
Attached drawing 5：To the targeting and specificity of atherosclerotic plaque in mice in PNP bodies.Figure A is tail vein injection
PNP, cycle 2 it is small when after, isolated organ image checking PNP is to the targeting and specificity of mouse patch.Figure B is relative fluorescence number
Value.It is combination situations of the micro- sem observation PNP of frozen section row to patch to scheme C.Control group is atherosclerosis mouse+PBS
Group, atherosclerosis mouse+NP groups and normal mouse+PNP groups.
Attached drawing 6：Coated rapamycin bionic nano particle (RAP-PNP) the treatment rat aorta atherosis of platelet membrane
The therapeutic effect of patch.A is schemed for treatment artery atherosclerotic plaque wholistic therapy effect, and figure B is patch entire area.
Attached drawing 7：The main pathology of characteristic aortic root patch and immunofluorescence dyeing, wherein oil red observation patch face
Necrosis area in patch is observed in product, HE dyeing, and collagen content in patch, CD68 dyeing observation patches are observed in Masson dyeing
Fibroblast content in patch is observed in interior macrophage content, α-SMA dyeing.
Attached drawing 8：Coated rapamycin bionic nano particle (RAP-PNP) the treatment rat aorta atherosis of platelet membrane
The therapeutic effect of patch.Figure D, E, F, G, H and I are respectively aortic root atherosclerotic plaque area, opposite tube chamber face
Product, necrosis area, collagen content, the relative value of macrophage content and fibroblast content.Control group is PBS
Group, free rapamycin drug group, common rapamycin nanoparticle particle (RAP-NP) treatment group and the coated thunder pa of platelet membrane
Mycin bionic nano particle (RAP-PNP) treatment group.
Attached drawing 9：Influences of the RAP-PNP to macrophage autophagy in atherosclerotic plaque in mice.Figure A is CD68 (marks
Remember macrophage) and autophagy marker protein LC3-II immunofluorescence figure.Figure B detects for Western blot (Western Blot)
The expression of autophagy labelled protein p62 and LC3-II in patch is schemed the relative expression that C and figure D is respectively LC3-II and p62 and is contained
Amount.Scheme the transmission electron microscope picture that E is autophagic vacuole in RAP-PNP group macrophages.Control group is PBS groups, free rapamycin drug
Group, common rapamycin nanoparticle particle (RAP-NP) treatment group and the coated rapamycin bionic nano particle of platelet membrane
(RAP-PNP) treatment group.
Specific embodiment
It elaborates with reference to embodiment to specific embodiment provided by the invention.
Following embodiment prepares a kind of coated rapamycin bionic nano particle of platelet membrane, and it is athero- to artery to inquire into it
The targeting ability of plaque and the therapeutic effect of targeted therapy patch.
The experimental data statistical method that following embodiment uses：Multiple-group analysis uses ANOVA methods.
Embodiment 1：Synthesize the coated rapamycin bionic nano particle of platelet membrane
Synthesis schematic diagram is shown in Fig. 1.After 800 μ g rapamycins, 20mg PLGA are dissolved in 1ml acetone, it is injected into 2ml water,
The PLGA nano particles of rapamycin are contained using the preparation of the nanoparticle precipitation method under vacuo.It is used rich in human blood platelets blood plasma
After EDTA anti-freezings, the remaining haemocyte of 100g centrifugation 20min removals.Supernatant addition EDTA and prostacyclin is taken to inhibit blood platelet and swash
Living, 800g centrifugation 20min, supernatant discarding, blood platelet is used to be resuspended containing the PBS of EDTA and protease inhibitors.3×109It is a
Blood platelet is used to be coated with 1mg PLGA nano particles.Blood platelet prepares platelet membrane using multigelation method.Utilize liquid nitrogen flash freezer
It after blood platelet, dissolves at room temperature, after being repeated 3 times, 4000g centrifugations are anti-using the PBS solution containing protease inhibitors
After after backwashing is washed, platelet membrane is resuspended with water.Platelet membrane is incubated 30min at 37 DEG C with DSPE-PEG2000 and is repaiied for polyethylene glycol
Decorations, rear to add in the PLGA nano particles for containing rapamycin, ultrasonic 2min is coated with for platelet membrane, and ultrasound condition is：
52KHz, 100W.To study the external and internal behavior tracer of the coated bionic nano particle (PNP) of platelet membrane, fluorescence is used
Dyestuff DiD marks PNP.The PNP preparation methods of fluorescent marker are same as above, it is only necessary to substitute rapamycin with third through a certain amount of DiD
After ketone dissolving nano particle is prepared with method.
The coated bionic nano granular size of platelet membrane is homogeneous under Electronic Speculum, good dispersion, and pan coating platelet membrane is complete
Whole, in regular spherical (as shown in Figure 2 C), grain size/potential measurement instrument the results show RAP-PNP grain sizes are 145.7 ± 6.69nm
(as shown in Figure 2 D), current potential is -25.3 ± 2.12mv.PNP surface membrane proteins content and significant Membrane protein's amount and blood platelet
Vesica is identical (as shown in Fig. 2 G and H).By centrifuging the bag for measuring nanoprecipitation drug content and determining RAP-PNP nano particles
Envelope rate and drugloading rate, drugloading rate are 3.6% (shown in Fig. 2 I), and envelop rate is up to 88.9% (shown in Fig. 2 J).The internal blood medicine of PNP is dense
Degree is shown in Fig. 3 A, and drug release in vitro curve is shown in Fig. 3 B, can extend nano particle circulation time after platelet membrane coating and slow down medicine
Object rate of release.
Embodiment 2：PNP simulates platelet adhering function in vitro
By external PNP static with simulating blood with combinations such as collagen, fibrin, vWF respectively under dynamic condition
The adhesion function of platelet.
Under PNP static states with collagen and fibrinous combination：Collagen or fiber egg are added in 96 orifice plates
Bai Hou adds in the PNP of DiD marks, 37 DEG C of incubation 5min, and after PBS cleaning, PNP that DMSO dissolvings combine, it is glimmering that microplate reader measures its
Luminous intensity.As shown in 4A and B, PNP has stronger combination power to collagen and fibrin, is not to be coated with blood platelet respectively
8.3 times and 9.6 times of film nano particle (NP), there are significant differences between the two.
Under PNP dynamical states and the combination of collagen, fibrin and vWF：Using glass slide prepare collagen and
Fibrin is positioned in parallel-plate flow cavity, and the PNP of DiD marks flows through parallel-plate flow cavity with 500S-1, after PBS cleaning,
Confocal laser scanning microscope.As shown in Fig. 4 C and D, current intelligence, NP is to collagen or fibrin without apparent knot
It closes, and PNP and collagen and fibrin have preferable combination, the red fluorescence and the collagen egg under light field for representing PNP
There are good common location relations between bletilla fibrin.After making HUVECs creep plates, inflammatory factor stimulating endothelial cell makes
Cell membrane surface expression vWF, endothelial cell creep plate is positioned in parallel-plate flow cavity, PNP flows through parallel-plate with 500S-1
Flow cavity, row vWF immunofluorescence dyeings after PBS cleaning are observed under laser confocal microscope.As shown in Figure 4 E, NP and vWF without
It is apparent to combine, and PNP has good common location relation with the vWF (green) for being expressed in cell membrane surface.
The platelet membrane for showing PNP pan coatings based on the above results is that nanoparticle possesses the knot similar to blood platelet
The ability of collagen, fibrin and vWF are closed, can preferably simulate hematoblastic adhesion function in vitro.
Embodiment 3：PNP is combined in vitro with atherosclerotic plaque
The external binding ability of PNP and atherosclerotic plaque in mice：ApoE-/-After mouse High-fat diet 8 weeks,
Remove the PNP that sustainer is marked with DiD to be incubated altogether, the imaging of row isolated organ, as Fig. 4 F and G, PNP group fluorescence intensity apparently higher than
Control group NP, and be distributed mainly on patch and enrich region (truncus brachiocephalicus and subclavian artery separate part), illustrate that PNP is moved with mouse
Pulse atherosclerosis patch has stronger combination in vitro.
PNP and the external binding ability of people's carotid artery atherosclerosis plaques：What people's carotid plaques marked after taking out with DiD
PNP is incubated altogether, row frozen section after OCT embeddings, confocal laser scanning microscope, it is seen that PNP groups have more red fluorescence
(PNP) adhere to, and NP groups have no apparent red fluorescence (as shown at figure 4h).
In summary the experiment results show that PNP has atherosclerotic plaque stronger combination in vitro, to spot
Block possesses good targeting ability.
Embodiment 4：PNP is to the internal targeting ability of atherosclerotic plaque in mice
For observation PNP bodies in atherosclerotic plaque in mice combination situation, ApoE-/-Mouse High-fat diet 8 weeks
Afterwards, mouse is put to death after the PNP (0.5mg/ is only) through tail vein injection DiD marks, 2h, sustainer row isolated organ is taken to be imaged, it is main
Artery root row OCT is embedded, and makes distribution situations of the frozen section observation PNP in atherosclerotic plaque.Such as Fig. 5 A and B institutes
Show, PNP groups fluorescence intensity is distributed mainly on patch and enriches region apparently higher than control group NP groups and PNP+ normal mouse groups
(truncus brachiocephalicus and subclavian artery separate part), PNP groups fluorescence intensity are 46.67 ± 4.60*106, NP groups are 9.37 ± 1.16*
106, PNP+ normal mouses group is 6.56 ± 0.75*106.PNP groups fluorescence intensity is 4.98 times of NP groups.Aortic root is embedded
Row frozen section dyeing observation afterwards, it is seen that PNP groups have more red fluorescence (representing nano particle) to assemble at patch, and NP
Group and PNP+ normal mouse groups have no apparent red fluorescence (as shown in Figure 5 C).
Embodiment 5：Coated rapamycin bionic nano particle (RAP-PNP) the treatment rat aorta of platelet membrane is athero- hard
The effect of changing patch
RAP-PNP administering modes and grouping：ApoE-/-After mouse High-fat diet 8 weeks, 1 targeted therapy is randomly divided into
Group and 3 control groups：(a) targeted therapy RAP-PNP groups；(b) RAP-NP control groups；(c) dissociate RAP control groups；(d) PBS pairs
According to group.RAP dosage is 0.5mg/kg, and every two days tail vein injections relevant nanometer drug is treated 4 weeks altogether.Free RAP drugs use
Absolute ethyl alcohol, castor oil and water (mass ratio 1：1：5) dissolve, for tail vein injection.
After tail vein injection drug therapy 4 weeks, mouse is put to death, Saline perfusion divides from aortic root to common iliac artery
Sustainer is taken out at fork, row substantially the Sudan's red colouring assesses Aortic Plaque area distributions.Aortic root prepares frost and cuts
Necrosis area in patch is observed in piece, respectively row oil red dyeing observation plaque area, HE dyeing, and Masson dyes glue in observation patch
Former protein content, CD68 immunofluorescence dyeings observation macrophage content, α-SM A immunofluorescence dyeings observation patch are interior into fibre
Tie up cell content.
As shown in Fig. 6 A and B, more patch is formed in PBS control group sustainer, dissociate RAP and non-targeted treatment group RAP-
NP can reduce the formation of atherosclerotic plaque, and the patch gross area drops to 34.8 ± 2.64% respectively from 50.1 ± 6.34%
And 40.0 ± 5.89%.The coated rapamycin bionic nano particle of platelet membrane can further increase the treatment effect of rapamycin
Fruit, the RAP-PNP group patch gross areas drop to 14.6 ± 2.91%.Fig. 7-Fig. 8 I are the pathology of aortic root patch, are immunized
Fluorescent staining and corresponding numerical value.Oil red staining reaction plaque area.It is similar to substantially plaque area result, dissociate RAP and
RAP-NP can reduce aortic root plaque area, and rapamycin treatment atherosclerotic plaque has been further amplified in RAP-PNP
The effect of block.RAP group aortic roots plaque area reduces 18.8%, RAP-NP groups and reduces 14.4%, and RAP-PNP groups are reduced
34.4%, RAP group aortic root Lumen Area increase to 45.2 ± 3.23%, RAP-NP from 36.3 ± 3.73% (PBS groups)
Group is that 42.3 ± 6.18%, RAP-PNP groups are 61.8 ± 5.70%.Significant difference is respectively provided between RAP-PNP groups and each group (such as
Fig. 7-Fig. 8 E).Necrosis area in HE dyeing observation patches, RAP-PNP groups substantially reduce bad in patch compared with other each groups
Unleavened dough is accumulated (as shown in Fig. 7 and Fig. 8 F).Meanwhile RAP-PNP dramatically increases collagen content in patch (such as Fig. 7 and Fig. 8 G institutes
Show), reduce macrophage content (as shown in Fig. 7 and Fig. 8 H), and improve fibroblast content (as shown in Fig. 7 and Fig. 8 I),
So as to increase the stability of patch.
The above results show that RAP-PNP can improve rapamycin treatment atherosclerotic plaque curative effect, inhibit patch into
It opens up and stablizes patch.
Embodiment 6：Macrophage autophagy in RAP-PNP activation patches
After treating 4 weeks, above-mentioned aortic root pathological section, row CD68 and autophagy marker protein LC3-II immunofluorescences are taken
Macrophage autophagy situation in patch is observed in dyeing altogether；Aortic tissue row Western Blot are taken, detect autophagy marker protein
P62 and LC3-II expressions detect patch inner cell autophagy situation；Truncus brachiocephalicus artery row transmissioning electric mirror checking is taken, passes through sight
Autophagic vacuole (double membrane structure) assessment macrophage autophagy situation is examined, inquires into the mechanism that RAP-PNP improves curative effect.Such as Fig. 9 A
It is shown, in RAP-PNP groups, autophagy marker protein LC3-II expression rise in macrophage, and free RAP groups and RAP-NP groups
Have no significant change.Compared to other groups, p62 expression declines Western Blot the results show RAP-PNP groups, and LC3-I to
LC3-II transformation rises, prompt RAP-PNP that can activate autophagy (Fig. 9 B-D).Transmission electron microscope is by observing autophagic vacuole (duplicature knot
Structure) the activation increase of RAP-PNP group macrophages autophagy is further demonstrated (shown in Fig. 9 E), prompt RAP-PNP targeted delivery thunders
Pa mycin is local to atherosclerotic plaque, improves local drug concentration, and activating macrophage autophagy process is mould so as to improve thunder pa
Plain medication effect inhibits plaque progression and stablizes patch.
The above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
Member, on the premise of the method for the present invention is not departed from, can also make several improvement and supplement, these are improved and supplement also should be regarded as
Protection scope of the present invention.
Claims (10)
1. a kind of composition for treating atherosclerosis, which is characterized in that the composition for pan coating platelet membrane,
Inside contains the nano-carrier of rapamycin or derivatives thereof.
2. composition according to claim 1, which is characterized in that the material of the nano-carrier is poly- acetic acid-hydroxyl
Acetic acid.
3. composition according to claim 2, which is characterized in that the nano-carrier leads to for poly- acetic acid-hydroxyacetic acid
Cross the nanoparticle of precipitation method preparation.
4. composition according to claim 3, which is characterized in that the grain size of the nanoparticle is 100-200nm.
5. composition according to claim 1, which is characterized in that the platelet membrane is coated with after polyethyleneglycol modified
In nano-carrier surface.
6. composition according to claim 5, which is characterized in that the polyethylene glycol is DSPE-PEG2000.
7. composition according to claim 1, which is characterized in that the platelet membrane is humanized's platelet membrane, by
Human blood platelets extracts.
8. composition according to claim 7, which is characterized in that the nano-carrier leads to for poly- acetic acid-hydroxyacetic acid
Cross the precipitation method preparation nanoparticle, 1~10 × 109The nanoparticle of the platelet membrane coating 1mg of a blood platelet extraction.
9. according to the preparation method of any compositions of claim 1-8, which is characterized in that the preparation method includes
Following steps：
(1) rapamycin nanoparticle carrier is prepared；
(2) platelet membrane is prepared, and is carried out polyethyleneglycol modified；
(3) polyethyleneglycol modified platelet membrane is coated in rapamycin nanoparticle carrier surface.
10. according to application of any compositions of claim 1-8 in the drug for preparing treatment atherosclerosis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810140423.1A CN108042510A (en) | 2018-02-11 | 2018-02-11 | A kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810140423.1A CN108042510A (en) | 2018-02-11 | 2018-02-11 | A kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN108042510A true CN108042510A (en) | 2018-05-18 |
Family
ID=62125959
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810140423.1A Pending CN108042510A (en) | 2018-02-11 | 2018-02-11 | A kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108042510A (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111110874A (en) * | 2019-11-14 | 2020-05-08 | 中国药科大学 | Radionuclide-labeled platelet membrane vesicle and preparation method and application thereof |
| CN111184700A (en) * | 2019-04-10 | 2020-05-22 | 重庆大学 | Erythrocyte membrane encapsulated rapamycin nanoparticle, preparation method and application thereof |
| CN114796488A (en) * | 2022-03-30 | 2022-07-29 | 华中农业大学 | Bionic nano-particles with antithrombotic function and preparation method and application thereof |
| CN115337468A (en) * | 2022-08-23 | 2022-11-15 | 苏州中天医疗器械科技有限公司 | Cell-targeting rapamycin drug balloon and preparation method and application thereof |
| CN117229423A (en) * | 2023-11-10 | 2023-12-15 | 北京科技大学 | Polypeptide nano material for binding collagen and preparation method and application thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6352833B1 (en) * | 1999-04-28 | 2002-03-05 | Michael E. Mendelsohn | Methods for discovery of vasoactive compounds for the nitric oxide-cyclic GMP signal pathway |
| CN101224187A (en) * | 2008-01-31 | 2008-07-23 | 中国医学科学院生物医学工程研究所 | Dual functional polymer nanometer glue bunch and preparing method and application in preparing medicine for treating second narrow blood vessel |
| CN105203752A (en) * | 2015-11-05 | 2015-12-30 | 上海中山医疗科技发展公司 | Magnetic platelet adsorption particle and preparation method thereof |
| US20170136219A1 (en) * | 2015-11-18 | 2017-05-18 | Angioscore, Inc. | Balloon angioplasty catheter coating to encourage vessel repair and further reduce restenosis |
| CN107115314A (en) * | 2011-06-02 | 2017-09-01 | 加利福尼亚大学董事会 | The nano particle and application method of film encapsulating |
-
2018
- 2018-02-11 CN CN201810140423.1A patent/CN108042510A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6352833B1 (en) * | 1999-04-28 | 2002-03-05 | Michael E. Mendelsohn | Methods for discovery of vasoactive compounds for the nitric oxide-cyclic GMP signal pathway |
| CN101224187A (en) * | 2008-01-31 | 2008-07-23 | 中国医学科学院生物医学工程研究所 | Dual functional polymer nanometer glue bunch and preparing method and application in preparing medicine for treating second narrow blood vessel |
| CN107115314A (en) * | 2011-06-02 | 2017-09-01 | 加利福尼亚大学董事会 | The nano particle and application method of film encapsulating |
| CN105203752A (en) * | 2015-11-05 | 2015-12-30 | 上海中山医疗科技发展公司 | Magnetic platelet adsorption particle and preparation method thereof |
| US20170136219A1 (en) * | 2015-11-18 | 2017-05-18 | Angioscore, Inc. | Balloon angioplasty catheter coating to encourage vessel repair and further reduce restenosis |
Non-Patent Citations (4)
| Title |
|---|
| XIAOLI WEI ETAL: "Nanoparticle Functionalization with Platelet Membrane Enables Multi-Factored Biological Targeting and Detection of Atherosclerosis", 《ACS NANO》 * |
| 吴镭、平其能主编: "《药剂学发展与展望》", 31 August 2002 * |
| 窦寅: "新型雷帕霉素微粒释药系统的构建及其抗动脉粥样硬化作用研究", 《中国博士论文全文数据库医药卫生科技辑》 * |
| 郭慧玲主编: "《药剂学》", 28 February 2014, 广州中山大学出版社 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111184700A (en) * | 2019-04-10 | 2020-05-22 | 重庆大学 | Erythrocyte membrane encapsulated rapamycin nanoparticle, preparation method and application thereof |
| CN111110874A (en) * | 2019-11-14 | 2020-05-08 | 中国药科大学 | Radionuclide-labeled platelet membrane vesicle and preparation method and application thereof |
| CN114796488A (en) * | 2022-03-30 | 2022-07-29 | 华中农业大学 | Bionic nano-particles with antithrombotic function and preparation method and application thereof |
| CN115337468A (en) * | 2022-08-23 | 2022-11-15 | 苏州中天医疗器械科技有限公司 | Cell-targeting rapamycin drug balloon and preparation method and application thereof |
| CN115337468B (en) * | 2022-08-23 | 2024-01-30 | 苏州中天医疗器械科技有限公司 | Cell targeting rapamycin medicine balloon, and preparation method and application thereof |
| CN117229423A (en) * | 2023-11-10 | 2023-12-15 | 北京科技大学 | Polypeptide nano material for binding collagen and preparation method and application thereof |
| CN117229423B (en) * | 2023-11-10 | 2024-02-06 | 北京科技大学 | Polypeptide nano material for binding collagen and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108042510A (en) | A kind of coated rapamycin bionic nano particle of platelet membrane for targeting atherosclerotic plaque and application thereof | |
| Song et al. | Platelet membrane-coated nanoparticle-mediated targeting delivery of Rapamycin blocks atherosclerotic plaque development and stabilizes plaque in apolipoprotein E-deficient (ApoE−/−) mice | |
| WO2019141267A1 (en) | Recombinant bionic nanocarrier delivery system for targeting active cd44 molecule, preparation method therefor, and uses thereof | |
| US20180243482A1 (en) | Anti-thrombogenic grafts | |
| CN107669632B (en) | Drug carrier, micelle, drug preparation, preparation method and application thereof | |
| Singh et al. | Nanobiomaterials for vascular biology and wound management: A review | |
| JP2006523507A (en) | Methods and compositions for treating myocardial disease symptoms | |
| Yao et al. | Application of a mechanically responsive, inflammatory macrophage-targeted dual-sensitive hydrogel drug carrier for atherosclerosis | |
| Cheng et al. | Influence of DL-β-hydroxybutyric acid on cell proliferation and calcium influx | |
| He et al. | Bifunctional Cx43 mimic peptide grafted hyaluronic acid hydrogels inhibited tumor recurrence and stimulated wound healing for postsurgical tumor treatment | |
| Huang et al. | Exploring naturally occurring ivy nanoparticles as an alternative biomaterial | |
| CN107802646A (en) | A kind of anti-tumor medicine | |
| Chen et al. | Fabrication of Tβ4-Exosome-releasing artificial stem cells for myocardial infarction therapy by improving coronary collateralization | |
| CN106714820A (en) | Selectin and icam/vcam peptide ligand conjugates | |
| CN112704686B (en) | Preparation method of calcium phosphate-rapamycin composite medicine, preparation method of medicine coating balloon and medicine coating balloon | |
| JP2002512640A (en) | Local delivery of sustained therapeutics | |
| Tao et al. | Nitric oxide-driven nanomotors with bowl-shaped mesoporous silica for targeted thrombolysis | |
| CN106699896A (en) | Tumor killing polypeptide capable of being self-assembled into hydrogel and application thereof | |
| CN111346070A (en) | Macrophage vesicle-loaded nano-drug preparation and application thereof in pharmacy | |
| CN108815134B (en) | Preparation and application of biological camouflage targeted nano drug delivery system for treating ischemic stroke | |
| Li et al. | Drug-Delivery Nanoplatform with Synergistic Regulation of Angiogenesis–Osteogenesis Coupling for Promoting Vascularized Bone Regeneration | |
| Wang et al. | Bacteria-inspired transformable nanoparticle targets and covers residual tumor against bladder cancer recurrence | |
| Wang et al. | Injectable decellularized extracellular matrix hydrogel loaded with exosomes encapsulating curcumin for prevention of cardiac fibrosis after myocardial infarction | |
| JP6873261B2 (en) | Drugs for targeting the heart, including tannic acid | |
| Hao et al. | Biomimetic and responsive nanoparticles loading JQ1 for dual-targeting treatment of vascular restenosis via multiple actions |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20180518 |