CN110530964B - Characteristic marker of milk exosome and method for identifying milk exosome - Google Patents
Characteristic marker of milk exosome and method for identifying milk exosome Download PDFInfo
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Abstract
The invention discloses a characteristic marker of milk exosome and a method for identifying the milk exosome, wherein the marker is an N-linked sugar chain, and the molecular weight and the structural formula of the N-linked sugar chain are 1599.5656(Man)5(Gal)1(GlcNAc)3And 2192.8088(Fuc)1(Man)3(Gal)2(GlcNAc)6. The invention obtains 2 reliable characteristic N-linked sugar chain structures of the milk exosomes as the markers of the milk exosomes through screening, and the 2 characteristic N-linked sugar chains can be used for simply, conveniently and quickly identifying the exosomes, thereby having practical significance for the research of clinical targeted drug loading.
Description
Technical Field
The invention belongs to the technical field of milk exosome identification, and particularly relates to a characteristic marker of a milk exosome and a method for identifying the milk exosome.
Background
Exosomes are biological nanovesicles of about 30-150nm secreted by almost all mammalian cell types, including mast cells, dendritic cells, B lymphocytes, epithelial cells, endothelial cells, adipocytes, and neurons. Current research has established that exosomes can facilitate cellular communication by delivering functional cargo such as proteins, mrnas, and micrornas (mirnas) in many biological processes. Exosomes play an irreplaceable role not only in normal physiological processes such as immune response, cell proliferation, inflammation, lactation and neuronal function, but also in different stages of disease progression such as liver disease, neurodegenerative disease and severe tumour-related diseases. Various components of the exosome under physiological conditions and pathological conditions can be used as treatment targets, and the exosome can also be used as a drug or gene delivery carrier in diseases, and the milk exosome has the potential of becoming a perfect targeted drug carrier because of the biological source and the easily obtained advantages, and the mechanism of targeted drug loading of the exosome is probably the specific recognition of the exosome surface glycoprotein and the targeted cells, so the analysis of the N-linked sugar chain on the surface of the milk exosome is very important for researching the mechanism of the targeted drug loading of the exosome.
Although the field of the existing exosomes is rapidly developed, a plurality of obstacles still exist, such as non-standard separation method, tedious characterization method of the separated exosomes, long time consumption, no characteristic markers of the exosomes with better quality and the like, so that the milk exosomes are analyzed to find the characteristic markers, a new method is provided for rapidly and simply identifying the milk exosomes, and the method has practical significance for clinical targeted drug loading research.
Disclosure of Invention
This section is for the purpose of summarizing some aspects of embodiments of the invention and to briefly introduce some preferred embodiments. In this section, as well as in the abstract and the title of the invention of this application, simplifications or omissions may be made to avoid obscuring the purpose of the section, the abstract and the title, and such simplifications or omissions are not intended to limit the scope of the invention.
The present invention has been made in view of the above-mentioned technical drawbacks.
Therefore, as one aspect of the invention, the invention overcomes the defects in the prior art and provides a marker which is characteristic of milk exosomes.
In order to solve the technical problems, the invention provides the following technical scheme: a marker characteristic of milk exosomes, wherein: the marker is an N-linked sugar chain having a molecular weight of 1599.5656(Man)5(Gal)1(GlcNAc)3And 2192.8088(Fuc)1(Man)3(Gal)2(GlcNAc)6。
As another aspect of the invention, the invention overcomes the deficiencies of the prior art and provides a method for identifying milk exosomes.
In order to solve the technical problems, the invention provides the following technical scheme: a method for identifying milk exosomes: and carrying out mass spectrum detection on the N-linked sugar chain of the milk exosome, and verifying the characteristic N-linked sugar chain of the milk exosome as an exosome marker.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the N-linked sugar chain has a molecular weight of 1599.5656(Man)5(Gal)1(GlcNAc)3And 2192.8088(Fuc)1(Man)3(Gal)2(GlcNAc)6。
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the mass spectrometric detection of the N-linked sugar chain of the milk exosome further comprises the mass spectrometric detection of the modified N-linked sugar chain.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the modification of the N-linked sugar chain includes a pan-methylation modification of the N-linked sugar chain.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the modification of the N-linked sugar chain includes acetylhydrazine modification of a non-reducing terminal sialic acid to the N-linked sugar chain.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the modification of the N-linked sugar chain includes aniline modification of the N-linked sugar chain at the reducing end of the N-linked sugar chain.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: also comprises the extraction of milk exosomes.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the extraction of the milk exosomes comprises the step of extracting the milk exosomes by an ultracentrifugation method.
As a preferred embodiment of the method for identifying milk exosomes according to the present invention: the ultracentrifugation method is 135000g centrifugation for 60 min.
The invention has the beneficial effects that: the invention obtains 2 reliable characteristic N-linked sugar chain structures of the milk exosomes as the markers of the milk exosomes through screening, and the 2 characteristic N-linked sugar chains can be used for simply, conveniently and quickly identifying the exosomes, thereby having practical significance for the research of clinical targeted drug loading.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the description of the embodiments will be briefly introduced below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings based on these drawings without inventive exercise. Wherein:
FIG. 1 is a flow chart of the experiment for separating milk exosome and whey protein;
FIG. 2 is an electron microscope image of the milk exosome of the present invention;
FIG. 3 is a MS diagram of different modification methods of the N-linked sugar chain of the milk exosome of the present invention;
FIG. 4 is a MS diagram showing different modification methods of the whey protein N-linked sugar chains according to the present invention;
FIG. 5 is a MS/MS diagram of N-linked sugar chains characteristic of the milk exosomes of the present invention.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with examples are described in detail below.
In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, but the present invention may be practiced in other ways than those specifically described and will be readily apparent to those of ordinary skill in the art without departing from the spirit of the present invention, and therefore the present invention is not limited to the specific embodiments disclosed below.
Furthermore, reference herein to "one embodiment" or "an embodiment" means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one implementation of the invention. The appearances of the phrase "in one embodiment" in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments.
Extracting milk exosomes:
fresh milk (200 mL) was stored at 4 deg.C, and centrifuged at 13000g for 30min to remove fat globules and a portion of casein, and defatted milk was obtained. To the skim milk was added an equal volume of distilled water, the pH was adjusted to 4.6 with 2M HCl, and casein was precipitated. Centrifuging the turbid solution at 4 deg.C and 8000g for 40min to obtain upper layer whey. Whey was passed through a 0.22 μm filter, the filtrate was centrifuged at 135000g for 60min at 4 ℃, the supernatant was discarded, the pellet was washed 3 times with sterile PBS solution and resuspended in precooled PBS solution to obtain exosome solution.
1. Mass spectrometric analysis of milk exosomes panmethylated N-linked sugar chains:
release and modification of N-linked sugar chains:
(1) unmodified N-linked sugar chain release:
taking the milk exosome solution with the corresponding volume of 500 mu g of protein content, adding 150 mu L of 8M urea solution, and fully and uniformly mixing. 40mM ammonium bicarbonate solution was added to make the final sample volume 1 mL. Releasing the sugar chains with 0.5. mu.L of PNGase-F, and incubating at 37 ℃ for 10-12 h. The pH of the sample was adjusted to about 2 with 50% TFA, and the sample was desalted (see below for details), and lyophilized to obtain an unmodified N-linked sugar chain.
(2) N-linked sugar chain pan-methylation modification:
to the lyophilized unmodified N-linked sugar chains, 150. mu.L of DMSO was added, mixed well, transferred to a glass tube, and sonicated for 15 min. Grinding 0.5g NaOH in a dry mortar into coarse powder, quickly transferring to a glass grinder, adding 1mL DMSO, and grinding for 10-15min to obtain uniform paste. To the sample, 200. mu.L of pasty NaOH and 50. mu.L of iodomethane were added (protected from light), and the operation was repeated 3 times with shaking for 20 min. After completion of the shaking, 1mL of ultrapure water was added immediately, and the reaction was terminated by shaking for 5 min. The sample was transferred to a new glass tube, 1mL of ultrapure water was added, and the mixture was shaken for 10 min. Adding 1mL chloroform for extraction, shaking for 10min, and removing the upper aqueous phase. Adding 2mL of ultrapure water for oscillation, repeating the operation for 5 times, removing the water phase, and freeze-drying to obtain the pan-methylated modified N-linked sugar chain.
2. Mass spectrometry of the N-linked sugar chains modified at both ends of the milk exosomes:
(1) n-linked sugar chain non-reducing end sialic acid acethydrazide modification and sugar chain release
Assembling a 10kD ultrafiltration membrane into a collecting pipe for collecting waste liquid in the ultrafiltration process, adding a milk exosome solution with the protein content of 500 mug in a corresponding volume, centrifuging for 15min at 14000g, concentrating the solution to the bottom of an ultrafiltration membrane tube, and discarding an effluent liquid. Adding 150 μ L of 8M urea, mixing, centrifuging at 14000g for 15min, and discarding the effluent; adding 300 μ L of 8mol/L urea, centrifuging at 14000g for 15min, and discarding the effluent; adding 100mM DTT solution to make the final concentration of the solution in the ultrafiltration membrane be 10mM, mixing well, incubating at 56 deg.C for 45min in a dry thermostat, centrifuging at 14000g for 15min in a desktop centrifuge after reaction, and discarding the effluent; adding 100mM IAM solution, fully sucking and blowing the solution in the ultrafiltration membrane to make the final concentration of the solution be 20mM, uniformly mixing, paying attention to the light-proof operation of IAM, uniformly mixing, placing the ultrafiltration tube in a dark environment, standing for reaction for 45min, centrifuging 14000g for 15min after the reaction is finished, and discarding the effluent liquid; adding 150 mu L of ultrapure water, fully mixing, centrifuging for 15min at 14000g, repeating the step for three times to clean IAM in the solution to avoid the influence on the subsequent reaction; after the washing is finished, 100 mu L of 1mol/L acethydrazide, 20 mu L of 1mol/L hydrochloric acid and 20 mu L of 2mmol/L EDC are added, the mixture is fully blown, sucked and uniformly mixed, an ultrafiltration tube is placed in a 120-turn table to ensure the suspension reaction of the protein, and the reaction is carried out for 4 hours at room temperature; centrifuging at 14000g for 15min after the reaction is finished, discarding the effluent, adding 150 μ L of 40mmol/L NH4HCO3Thoroughly blowing, sucking, mixing, centrifuging at 14000g for 15min, and adding NH4HCO3Repeatedly cleaning the solution for 3 times; taking out the ultrafiltration tube, transferring the ultrafiltration tube into a clean collecting tube, releasing the sugar chain by using 0.5 mu L of PNGase F, and incubating for 10-12 h at 37 ℃; centrifuging at 14000g for 15min, reserving the effluent, adding 150 mu L of ultrapure water into an ultrafiltration membrane to suck the heavy-suspended and precipitated protein, centrifuging at 15000g for 15min, repeating for 2 times, fully collecting the N-linked sugar chain, reserving the effluent in the collection tube, and freeze-drying to obtain the N-linked sugar chain with sialic acid modified by acethydrazide.
(2) Aniline modification of N-linked sugar chain reducing end
To the acethydrazide-modified N-linked sugar chain was added 10. mu.L of an aniline solution, 25. mu.L of 1mol/L NaCNBH3And fully blowing and sucking. Reacting in a shaking table at 75 ℃ for 30min, and freeze-drying to obtain the N-linked sugar chain modified by the reducing end of aniline.
The method of desalting (Clean up) treatment of the primary N-linked sugar chain sample was:
(1) salt removal by pan-methylation
Dissolve the sample with 500 μ L of 0.1% trifluoroacetic acid (V/V); taking Waters
Vac,1cc tc 18Cartridges, adding 1mL of pure acetonitrile, washing the column repeatedly 3 times, 1mL of 80% acetonitrile/0.1% trifluoroacetic acid (V/V), washing the column repeatedly 3 times, 1mL of 0.1% trifluoroacetic acid (V/V), washing the column repeatedly 3 times; adding the sample, collecting effluent, and repeating the sample loading for 1 time; 1mL of 0.1% trifluoroacetic acid (V/V) was added and the column was washed repeatedly 3 times; 1mL of 1% formic acid (V/V) was added and the column was washed 1 time; after completion of the washing, 400. mu.L of 50% acetonitrile/0.1% formic acid (V/V) was added to elute the N-linked sugar chain, and the effluent was collected and freeze-dried.
(2) Desalting process for sugar chain modification by other methods
Dissolve the sample with 500 μ L of 0.1% trifluoroacetic acid (V/V); supelclean from SUPELCOTM EMI-CarbTMSPE Tube, adding 1mL pure acetonitrile, repeatedly washing the column for 3 times, 1mL 80% acetonitrile/0.1% trifluoroacetic acid (V/V), repeatedly washing the column for 3 times, 1mL 0.1% trifluoroacetic acid (V/V), repeatedly washing the column for 3 times; adding the sample, collecting effluent, and repeating the sample loading for 1 time; 1mL of 0.1% trifluoroacetic acid (V/V) was added and the column was washed repeatedly 3 times; after completion of the washing, 400. mu.L of 80% acetonitrile/0.1% trifluoroacetic acid (V/V) was added to elute the N-linked sugar chain, and the effluent was collected and freeze-dried.
MALDI-TOF-Mass Spectroscopy of N-linked sugar chain samples:
the N-linked sugar chains of the milk exosomes were resolved by Ultraflextreme MALDI-TOF/TOF tandem mass spectrometry of Bruker Daltonics. And (3) acquiring the mass spectrum data of the sugar chain in FlexAnalysis software, taking mass spectrum peaks which have signal-to-noise ratio of more than 6 and are identified by at least three times of experiments, and carrying out subsequent analysis. Sugar chain structures were simultaneously analyzed manually in conjunction with glycothornbech software.
In order to obtain more information on sialylated sugar chains, the present invention separates glycoproteins using an ultrafiltration membrane and performs acetohydrazide modification on sialylated sugar chains of proteins, and the labeled sugar chains are then released on the ultrafiltration membrane and detected by mass spectrometry. Mass spectral data for N-linked sugar chains with signal to noise ratio >6 were annotated with GlycoWorkbench software.
The mass spectrometric data analysis of the N-linked sugar chain distribution of the exosomes obtained in the present invention are as follows:
(1) the discovery of 4 methods identified a total of 90N-linked sugar chains;
(2) of all 90N-linked sugar chains, 7 sugar chain structures among them were identified by four methods, 2 of which appear only in milk exosomes, and these 2 sugar chain structures were able to serve as N-linked sugar chains characteristic of exosomes; there are 19 sugar chain structures identified by three methods, 14 of which are present only in exosomes; the structure was identified by two methods in 25 sugar chain species, and 39 sugar chain structures were identified by only one method;
(3) the acetylhydrazine group modifies the terminal sialic acid of sugar chains so that the number of N-linked sugar chains identified is at most 51, followed by the pan-methylation modification method, and 49 are identified.
The characteristic N-linked sugar chain of 2 exosomes, the structural formula is as follows:
sugar chain structure 1 (G1): 1599.5656(Man)5(Gal)1(GlcNAc)3
Sugar chain structure 2 (G2): 2192.8088(Fuc)1(Man)3(Gal)2(GlcNAc)6
GlcNAc represents N-acetylglucosamine, Man represents mannose, Gal represents galactose, Fuc represents fucose, (Neu5Ac represents N-acetylneuraminic acid, Neu5Gc represents N-glycolylneuraminic acid).
In the structural formula, GlcNAc represents N-acetylglucosamine, Man represents mannose, Gal represents galactose, GalNAc represents N-acetylgalactosamine, and Fuc represents fucose.
Specifically, the structure of each sugar chain is as follows:
wherein the content of the first and second substances,
the expression Man is used for representing the Man,
represents a compound represented by GlcNAc or a compound represented by GlcNAc,
represents Fuc, and O represents Gal
The 2 structures of G1 and G2 are exosomes identified collectively using the above four methods, which are different from the N-linked sugar chains of whey protein (the N-linked sugar chains contained only in exosomes and not in whey protein in milk), and this 2-structure has reliability as a characteristic sugar chain of exosomes.
The invention obtains 2 reliable N-linked carbohydrate chain structures which are characteristic of milk exosomes through screening; by utilizing the characteristic N-linked sugar chain, the exosome can be identified simply and rapidly, and the method has practical significance for the clinical targeted drug-loading research.
It should be noted that the above-mentioned embodiments are only for illustrating the technical solutions of the present invention and not for limiting, and although the present invention has been described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, which should be covered by the claims of the present invention.
Claims (9)
1. A marker characteristic of milk exosomes, characterized by: the marker is an N-linked sugar chain having a molecular weight of 1599.5656(Man)5(Gal)1(GlcNAc)3Or 2192.8088(Fuc)1(Man)3(Gal)2(GlcNAc)6。
2. A method of identifying milk exosomes, characterized by: performing mass spectrum detection on the N-linked sugar chain of the milk exosome, and verifying the characteristic N-linked sugar chain of the milk exosome as an exosome marker, wherein the molecular weight and the structural formula of the characteristic N-linked sugar chain are 1599.5656(Man)5(Gal)1 (GlcNAc)3Or 2192.8088(Fuc)1 (Man)3(Gal)2(GlcNAc)6。
3. A method of identifying milk exosomes according to claim 2, characterized in that: the mass spectrometric detection of the N-linked sugar chain of the milk exosome further comprises the mass spectrometric detection of the modified N-linked sugar chain.
4. A method of identifying milk exosomes according to claim 3, characterized in that: the modification of the N-linked sugar chain includes a pan-methylation modification of the N-linked sugar chain.
5. A method of identifying milk exosomes according to claim 3, characterized in that: the modification of the N-linked sugar chain includes acetylhydrazine modification of a non-reducing terminal sialic acid to the N-linked sugar chain.
6. A method of identifying milk exosomes according to claim 3, characterized in that: the modification of the N-linked sugar chain includes aniline modification of the N-linked sugar chain at the reducing end of the N-linked sugar chain.
7. A method of identifying milk exosomes according to claim 2 or 3, characterized in that: also comprises the extraction of milk exosomes.
8. The method of identifying milk exosomes according to claim 7, characterized in that: the extraction of the milk exosomes comprises the step of extracting the milk exosomes by an ultracentrifugation method.
9. The method of identifying milk exosomes according to claim 8, characterized in that: the ultracentrifugation method is 135000g centrifugation for 60 min.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106814187A (en) * | 2017-01-16 | 2017-06-09 | 杭州联川基因诊断技术有限公司 | Application of the free excretion body in periphery in liquid biopsy tumour diagnostic reagent is prepared |
| CN107796939A (en) * | 2016-08-30 | 2018-03-13 | 希森美康株式会社 | Sample analysis box and its manufacture method, with and application thereof |
| CN108168977A (en) * | 2018-01-08 | 2018-06-15 | 南通大学附属医院 | Extraction, detection method and the kit of excretion body biomarker MMP-13 |
| CN108315440A (en) * | 2018-03-29 | 2018-07-24 | 华南农业大学 | A kind of miRNA markers and its application for distinguishing buffalo and milk milk |
| CN109837306A (en) * | 2019-03-20 | 2019-06-04 | 江南大学附属医院（无锡市第四人民医院） | Contain the excretion body and its preparation method and application of miRNA-204-5p |
-
2019
- 2019-09-05 CN CN201910835803.1A patent/CN110530964B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107796939A (en) * | 2016-08-30 | 2018-03-13 | 希森美康株式会社 | Sample analysis box and its manufacture method, with and application thereof |
| CN106814187A (en) * | 2017-01-16 | 2017-06-09 | 杭州联川基因诊断技术有限公司 | Application of the free excretion body in periphery in liquid biopsy tumour diagnostic reagent is prepared |
| CN108168977A (en) * | 2018-01-08 | 2018-06-15 | 南通大学附属医院 | Extraction, detection method and the kit of excretion body biomarker MMP-13 |
| CN108315440A (en) * | 2018-03-29 | 2018-07-24 | 华南农业大学 | A kind of miRNA markers and its application for distinguishing buffalo and milk milk |
| CN109837306A (en) * | 2019-03-20 | 2019-06-04 | 江南大学附属医院（无锡市第四人民医院） | Contain the excretion body and its preparation method and application of miRNA-204-5p |
Non-Patent Citations (4)
| Title |
|---|
| C Escrevente.Protein glycosylation of exosomes from ovarian carcinoma cells: structures and biological roles.《Structures and biological roles》.2011, * |
| JF Rakus et al..New technologies for glycomic analysis: toward a systematic understanding of the glycome.《Annual review of analytical chemistry》.2011, * |
| Sophia G. Antimisiaris et al..Exosomes and Exosome-Inspired Vesicles for Targeted Drug Delivery.《pharmaceutics》.2018, * |
| 牛奶消化前后外泌体的变化及其对细胞增殖的影响;徐小艳 等;《同济大学学报（医学版）》;20181217;第39-43页 * |
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