CN114748506A - Application of bone marrow mesenchymal stem cell exosome - Google Patents
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Abstract
The invention provides an application of a mesenchymal stem cell exosome in bone marrow. Liver function impairment was alleviated by injecting BMSCs-Exo into HIRI Kunming mice. Wherein miR-25-3p is remarkably reduced in HIRI liver, BMSCs and BMSCs-Exo both contain miR-25-3p, miR-25-3p expression is increased after miR-25-3p mimics (miR-25-3p agomir and miR-25-3p mimic) are respectively transferred into HIRI mice and AML12 liver cells with hypoxia/reoxygenation, the expression reduction of apoptosis and inflammation related factors is detected by qRT-PCR and Western blotting, and the liver injury is reduced by measuring the content of aspartate Aminotransferase (AST) and alanine Aminotransferase (ALT) in serum and hematoxylin-eosin (HE) staining. In addition, the expression reduction of genes and related proteins of MAPK/ERK signal channels and PI3K/AKT signal channels is detected by qRT-PCR and Western blotting. Therefore, miR-25-3p inhibits HIRI injury-induced hepatocyte apoptosis by inhibiting MAPK/ERK and PI3K/AKT signaling pathways, so that HIRI is improved, and a new idea is provided for treating HIRI.
Description
Technical Field
The invention relates to the fields of biology and medicine, in particular to application of a bone marrow mesenchymal stem cell exosome.
Background
Liver ischemia reperfusion injury (HIRI) refers to a pathological phenomenon that blood supply is restored after blocking the blood supply of liver for a period of time in the liver surgical operation process, and liver dysfunction and structural damage are not reduced but aggravated. HIRI is a common clinical problem in liver trauma treatment, liver lobe excision, liver transplantation and the like, can cause organ failure and increase the incidence rate of rejection, and has great influence on the prognosis and survival of patients. Until now, there has been a lack of clinically effective interventions. During ischemia reperfusion, cells are in an oxidative stress state, endoplasmic reticulum stress and calcium overload occur, mitochondria swelling and outer membrane fracture occur, and cytochrome C is released, so that an apoptosis signal path is promoted, and apoptosis is caused.
Mesenchymal Stem Cells (MSCs) are one of the most widely used types of stem cells in clinical practice for immunomodulation, organ reconstruction and tissue repair, and secrete exosomes (Exo) under stimulation of rest or hypoxia, stress, radiation and oxidative damage. Exosomes are considered as a new therapeutic tool for delivering functional proteins, mrnas, mirnas and lncrnas, delivering bioactive molecules to recipient cells to modulate cellular functions, with higher biosafety and more stable signaling efficiency than stem cells, and can be a powerful candidate for cell-free therapy. Research shows that the mesenchymal stem cell exosome can regulate apoptosis and reduce hepatic ischemia reperfusion injury, but the stem cell exosome contains various bioactive substances, and the research needs to be carried out deeply on the action mechanism of the stem cell exosome for reducing the hepatic ischemia reperfusion injury.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide an application of a mesenchymal stem cell exosome.
One of the purposes of the invention is realized by adopting the following technical scheme: use of a mesenchymal stem cell exosome in the preparation of a medicament for treating liver ischemia reperfusion injury, the medicament for inhibiting the MAPK/ERK signalling pathway and the PI3K/AKT signalling pathway.
In one embodiment, the bone marrow mesenchymal stem cell exosome comprises miR-25-3 p.
The invention also aims to provide application of miRNA from bone marrow mesenchymal stem cell exosome in preparation of a medicament for treating liver ischemia-reperfusion injury.
The second purpose of the invention is realized by adopting the following technical scheme: the application of miRNA derived from bone marrow mesenchymal stem cell exosome in preparing a medicament for treating liver ischemia-reperfusion injury, wherein the medicament is used for inhibiting MAPK/ERK signaling pathway and PI3K/AKT signaling pathway.
In one embodiment, the miRNA is miR-25-3 p.
The invention also aims to provide a medicament for treating the liver ischemia-reperfusion injury.
The third purpose of the invention is realized by adopting the following technical scheme: a medicament for treating ischemia reperfusion injury in the liver, the medicament comprising a component that inhibits the MAPK/ERK signaling pathway and the PI3K/AKT signaling pathway.
In one embodiment, the medicament comprises miR-25-3 p.
Compared with the prior art, the invention has the beneficial effects that: the invention discovers that miRNAmiR-25-3p from the exosome of the human mesenchymal stem cell inhibits the hepatocyte apoptosis induced by HIRI injury through inhibiting MAPK/ERK and PI3K/AKT signal channels, thereby being capable of relieving the hepatic ischemia reperfusion injury. Therefore, the present invention provides a new concept for the treatment of HIRI.
Drawings
FIG. 1A is a diameter size result graph of a ZetaView assay exosome provided by an embodiment of the present invention;
FIG. 1B is a diagram illustrating morphological features of exosomes displayed by transmission electron microscopy according to an embodiment of the present invention;
FIG. 1C is a diagram showing the result of Western blot detection of an exosome marker protein provided in the present invention;
FIG. 2 is a graph showing the results of the ALT and AST contents in the serum after injecting BMSCs exosomes into HIRI mice and perfusing the BMSCs exosomes for 3h, 6h and 9h respectively;
FIG. 3 is a graph showing the results of the content of miR-25-3p in HIRI mouse liver and its control CK, BMSCs-Exo treated HIRI mouse group and its control PBS treated HIRI mouse group, detected by qRT-PCR according to the embodiment of the present invention;
FIGS. 4A and 4B are graphs showing the results of Tunel detecting apoptosis of liver tissue according to the embodiment of the present invention; wherein, white cells in fig. 4A represent apoptotic cells, and gray cells represent normal cells;
FIG. 4C is a graph showing the results of qRT-PCR assay of the content of GSDMD and NLRP3mRNA in liver tissue according to the embodiment of the present invention;
FIGS. 4D and 4E are graphs showing the result of Western blot detection of Caspase1 content in liver tissue according to the embodiment of the present invention; wherein lanes 1 and 2 of FIG. 4D are normal saline NS, lanes 3 and 4 are miR-25-3p agomir;
FIG. 4F is a graph showing the results of qRT-PCR assay of AML12 cell inflammation-associated gene content according to the present invention;
FIG. 5A is a graph showing the results of the content of ALT and AST in serum after treating mice with ischemia-reperfusion injury with miR-25-3p mimic provided by the embodiment of the invention;
FIG. 5B is a graph of the results of HE staining of liver tissues after treatment of liver ischemia-reperfusion mice with the miR-25-3p mimic provided in the embodiments of the present invention;
FIGS. 6A and 6B are graphs showing the results of Western blot detection of the expression of p-AKT and p-ERK in liver tissues according to the embodiment of the present invention; wherein lanes 1 and 2 in fig. 6A represent NS, and lanes 3 and 4 represent agomir;
FIG. 6C is a graph showing the results of qRT-PCR assay for the levels of mRNA for PI3K, Raf, Ras and MEK genes in liver tissue according to an embodiment of the present invention;
FIG. 6D is a graph showing the results of qRT-PCR assay of PI3K and MEK gene mRNA content in AML12 hypoxic reoxygenation hepatocytes, provided by an embodiment of the present invention.
Detailed Description
The present invention is further described with reference to the accompanying drawings and the detailed description, and it should be noted that, in the case of no conflict, any combination between the embodiments or technical features described below may form a new embodiment. In the following examples, reagents used were all analytical grade and were commercially available unless otherwise indicated. Experimental procedures not specifically identified herein are generally carried out under conventional conditions such as those described in the molecular cloning guidelines, published by scientific Press 2002, edited by J. SammBruk et al, or under conditions recommended by the manufacturer. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. Moreover, any methods and materials similar or equivalent to those described herein can be used in the practice of the present invention.
First, experiment method
1. Collection, purification and identification of BMSCs exosomes
Human mesenchymal stem cells (BMSCs) are purchased from Wuhan Pronoch Life technologies, Inc. (CP-H166), cultured in DMEM medium containing 10% fetal calf serum, washed with PBS when the cells reach 70% -80% fusion, inoculated into serum-free medium at a ratio of 1:3 for culturing for 48H, collected at 4 ℃, centrifuged at 300g for 10min, and kept; centrifuging again for 10min at 2000g, and keeping the supernatant; centrifuging at 10000g for 30min, and keeping supernatant; after filtration using a 0.22 μm filter, the precipitate was retained by centrifugation at 100000g for 2 h. Adding PBS, mixing, centrifuging for 2h under 100000g, and keeping precipitate, wherein the precipitate is relatively pure exosome. Add 100. mu.L PBS to exosomes and freeze-store in-80 ℃ refrigerator. The particle analyzer detects exosome diameters. Exosome morphology was observed by transmission electron microscopy. And identifying the exosome surface marker by using Western blot.
2. Laboratory animal
SPF grade male Kunming mice, weighing 20-25g, were purchased from Schlekshirta laboratory animals Ltd of Hunan (production permit: SCXK (Hunan) 2021-. The mice were fed with normal mouse feed and filtered tap water was freely added. The room temperature is 22 +/-1 ℃, the humidity is 65% +/-5%, and the illumination/darkness is alternated for 12 h. Animal protocols were approved and approved by the animal ethics committee of the Guilin medical college. The experimental process is carried out strictly according to the national standard of the people's republic of China (GB/T35892-.
3. Sources of miR-25-3p mimetics
A mimic of miR-25-3p mimic for cell and its control miR-25-3p mimic NC, and a mimic of miR-25-3p for animal, miR-25-3p agomir, were purchased from Sharp Biotech, Guangzhou. Physiological saline (NS) was used as a control for miR-25-3p agomir. .
4. Preparation of mouse liver ischemia reperfusion injury model
The mice were divided into experimental groups (agomir) and control groups (NS), 3 mice per group. 24h before surgery each mouse was injected by tail vein with 13nmol agomir diluted with 100 μ l of saline or 100 μ l of saline. Before operation, the patient is fasted for 12 hours, and water is freely drunk. 4% chloral hydrate was anesthetized by intraperitoneal injection at 0.1ml/10g, and the limbs were fixed with an adhesive tape. Sterilizing with 70% ethanol, and cutting skin layer and muscle layer from the middle of abdomen to the xiphoid process. The left and middle lobes of the liver were exposed and the hepatic artery and portal vein of the middle and left lobes were clamped closed with a non-invasive vascular clamp. During the clipping process, the incision was covered with wet gauze, and the mouse was placed on a constant temperature heating pad at 37 ℃ for heat preservation. And (3) after the continuous ischemia is finished for 60min, quickly taking out the vascular clamp, recovering the blood flow of the ischemic liver, suturing the abdominal muscle and the skin layer by layer, and disinfecting the incision. Mice were harvested at 6h reperfusion.
5. Cell hypoxia reoxygenation model
Mouse hepatocytes (AML12) were purchased from the China academy of sciences type culture Collection Committee cell Bank (SCSP-550). The cells were cultured in DMEM medium containing 10% fetal bovine serum. The cells are spread in a 24-well plate 24h in advance according to the density of 1.5 multiplied by 105 to 2.5 multiplied by 105 per well. When the cell coverage rate reaches about 60-70%, miR-25-3p mimic and a control miR-25-3p mimic NC 100nM are transfected. After 24h the cells were incubated with PBS under hypoxic conditions (5% CO2, 1% O2 and 94% N2, 37 ℃) for 6h, followed by 12h under normal conditions.
6. Serological index detection
Blood is collected from the eyeballs of the mice after 6h of reperfusion, the mice are stood for 30min at room temperature, then the centrifugation is carried out at 1800g for 15min and 4 ℃, the upper serum is taken, the kit of Nanjing institute of bioengineering is utilized to measure the levels of aspartate Aminotransferase (AST) and alanine Aminotransferase (ALT) in the serum, and an enzyme-labeling instrument with the wavelength of 510mm is utilized to measure the OD value of each hole.
7. Tunel experiment
Liver tissue was fixed with 4% paraformaldehyde for 48h, gradient alcohol dehydrated, paraffin embedded, and sectioned with paraffin microtome to a thickness of 4 μm. Baking in an oven at 60 deg.C for 4.5h, dewaxing, and gradient alcohol hydration. Treating the tissues with Proteinase K at 37 ℃ for 20min, adding 50 μ l of TUNEL reaction mixture to each tissue, and reacting for 1h in a wet box at 37 ℃ in the dark; dripping 100ul of DAPI staining solution, and incubating the incubation box at constant temperature in dark place for 20min at room temperature; and (5) sealing by using a water-soluble sealing agent. The TUNEL positive rate was calculated by observing and photographing with a fluorescence microscope at 488nm for green and 596nm for red.
8. Hematoxylin-eosin (HE) staining experiment
Liver tissue was fixed with 4% paraformaldehyde for 48h, gradient alcohol dehydrated, paraffin embedded, sectioned with paraffin microtome, thickness 4 μm. Baking in an oven at 60 deg.C for 4.5 hr, dewaxing, and gradient alcohol hydration. Staining with hematoxylin for 5min, differentiating with hydrochloric acid ethanol for 30sec, and staining with eosin for 5 min. Gradient alcohol dehydration and neutral gum sealing. And (4) observing by using an optical microscope.
9. Real-time quantitative PCR (qRT-PCR) experiments
mRNA and miRNA were extracted from AML12 cells or liver tissues using UNIQ-10 column Trizol Total RNA extraction kit from Biotechnology engineering (Shanghai) Ltd. And measuring an OD260 value and an OD260/280 ratio by using a spectrophotometer, calculating the RNA concentration according to the OD260, and carrying out 1% agarose gel electrophoresis to identify the integrity of the total RNA. The mirnas and mrnas were Reverse transcribed into cdnas using Maxima Reverse Transcriptase (Thermo Scientific). qRT-PCR was performed on mRNA using 2 XSG Fast qPCR Master Mix (High Rox, B639273, BBI, ABI) and RiboBio miRNA detection was performed on miRNA according to the instructions for use. The relative expression level of the gene was determined by the 2-DDCT method. Endogenous controls for mRNA and miRNA expression were GAPDH and U6, respectively. The sequences of the primers are as follows:
10. western blot experiment
Adding protease inhibitor and RIPA lysate into liver tissue or cell to extract total protein. Protein concentration was determined using BCA protein quantification kit (Solarbio). Selecting separation gels with different concentrations (10% -15%) according to the molecular weight of the protein to carry out electrophoretic separation, activating a PVDF membrane by using methanol, installing a sandwich structure, transferring the protein onto the PVDF membrane through electrophoresis, and sealing 5% skim milk for 1h at room temperature. PBST membrane washing three times, primary antibody 4 degrees C were incubated overnight, secondary antibody 37 degrees C were incubated for 1 h. The membrane was washed three times with PBST and the protein bands were detected with ECL kit (Millipore). The gel imaging system scans and quantitatively analyzes the optical density values of the bands.
11. Transmission electron microscope
2% paraformaldehyde-2.5% glutaraldehyde pre-fixed hepatocytes, rinsing with cold buffer solution for 15min × 3 times, fixing 1% osmic acid at normal temperature for 2h, gradient ethanol dehydration, epoxypropane replacement twice, soaking epoxypropane and Epon 812 resin mixture overnight, embedding Epon 812 resin, polymerizing at 35 ℃ for 24h, 12h at 45 ℃, 12h at 55 ℃ and 24h at 60 ℃, positioning semi-thin sections, performing glass-knife 50nm ultrathin section, performing double electronic staining on uranyl acetate and lead citrate, and observing by a JEM-1230 type transmission electron microscope. After cell digestion, the cells are prepared into suspension for embedding and slicing, exosome is dripped on a carrier copper net with the aperture of 2nm by an exosome electron microscope, and the carrier copper net is kept stand for 2min, 3 percent phosphotungstic acid is negatively dyed for 5min, and observed by a JEM-1230 transmission electron microscope.
12. Statistical analysis
The analysis is carried out by SPSS 25.0 statistical software, and the difference is shown as P <0.05, which has statistical significance.
Second, experimental results
1. Isolation and characterization of BMSCs exosomes
Collecting the supernatant of the BMSCs cell culture of P3 generation, carrying out differential centrifugation, and detecting the particle size of the obtained exosome by a particle analyzer. FIG. 1A is a diameter size result graph of exosomes detected by ZetaView; FIG. 1B is a diagram showing morphological features of exosomes using transmission electron microscopy; FIG. 1C is a graph showing the results of detection of marker proteins by Western blotting. The specific results are as follows: ZetaView detected particle diameters in the purified exosomes to be about 60-200 nm. The diameter of the exosome is about 60-200nm as shown by a transmission electron microscope, the low electron density component can be seen in the cell, the exosome is circular or elliptical, and the cell membrane structure is complete. Western blot showed CD9 and CD63 positive and Calnexin negative in exosomes.
2. BMSCs exosomes can improve liver damage caused by HIRI
In order to detect the influence of BMSCs exosomes on HIRI mice, 100 mu g of BMSCs exosomes are dissolved in 100 mu L of PBS, the dissolved BMSCs exosomes are injected into the HIRI mice through tail veins, and the dissolved BMSCs exosomes are respectively perfused for 3h, 6h and 9h to detect the ALT and AST contents in serum through eyeball blood sampling. Referring to fig. 2, the results show that at three time points of 3h, 6h, and 9h reperfusion, both ALT and AST were lower in BMSCs exosome injected group than in PBS injected group.
3. MiR-25-3p participating in BMSCs exosome to alleviate HIRI injury
In order to determine the correlation between miR-25-3p and BMSCs exosome to reduce HIRI injury, the content of miR-25-3p in the HIRI mouse liver and the HIRI mouse group treated by the control CK, BMSCs-Exo and the HIRI mouse group treated by the control PBS is detected by qRT-PCR. Referring to FIG. 3, it can be seen that miR-25-3p is expressed in HIRI liver less than CK group; BMSCs and BMSCs-Exo both contain miR-25-3 p; after BMSCs-Exo is transferred into HIRI mice through tail veins, the content of miR-25-3p in liver tissues is increased. Therefore, miR-25-3p is present in BMSCs and exosomes thereof, and is involved in lightening HIRI injury by BMSCs exosomes.
4. MiR-25-3p for inhibiting apoptosis
The miR-25-3p mimic agomir can enable miR-25-3p in an animal to be over-expressed, the miR-25-3p mimic agomir is injected into a HIRI mouse body through a tail vein, a liver is taken for 6h after reperfusion to prepare a paraffin section, a Tunel experiment is used for detecting the apoptosis condition, and DAPI is used for marking cell nucleus. Referring to fig. 4A and 4B, the results show that the experimental group (the agomir group) mice have a lower Tunel-positive cell rate than the saline-injected group (the NS group). Referring to fig. 4C, qRT-PCR assay showed that the amount of GSDMD and NLRP3 mRNA in the liver tissue was also lower in the experimental group than in the saline group. Referring to fig. 4D and 4E, the amount of Caspase1 protein in the liver tissue was lower in the experimental group than in the NS group. The miR-25-3p simulant micic can enable miR-25-3p in cells to be over-expressed, the miR-25-3p micic and a control NC are transferred into AML12 hypoxia reoxygenation hepatocytes, and the content of inflammation-related genes is detected by qRT-PCR (quantitative reverse transcription-polymerase chain reaction), please refer to FIG. 4F, and the results show that the contents of TNF-alpha, IL-6, IL-1b and IL-18 are lower than those of a control group, and the contents of TNF-alpha and IL-1b are remarkably different.
5. MiR-25-3p for alleviating liver injury
Serum ALT and AST levels generally reflect the degree of hepatocyte damage. After miR-25-3p agomir is transferred into a HIRI mouse, blood is collected by eyeballs, and the ALT and AST content in serum is detected. Referring to 5A, the results show that the ALT and AST content of the miR-25-3p group is lower than that of the control NS group, and the ALT content is remarkably reduced. Please refer to fig. 5B, the liver tissue structure was observed to find that the lobular structure of the experimental group was complete, the hepatic cell cords were arranged relatively neatly, and the cytoplasm was uniform, while the lobular structure of the control NS group was unclear, the hepatic cell arrangement was disordered and increased in different degrees, and vacuolar degeneration and inflammatory infiltration occurred.
6. miR-25-3p regulates MAPK/ERK and PI3K/AKT signal pathways
In order to clarify the signal path of miR-25-3p for regulating apoptosis, the liver tissues of mice in an experimental group and a control group are taken for detection. Referring to FIG. 6A and FIG. 6B, Western blot results show that the overexpression of miR-25-3p obviously inhibits the expression of p-AKT and p-ERK. Referring to FIG. 6C, the qRT-PCR results show that PI3K, Raf, Ras and MEK levels were also lower than the control. After transfer of AML12 cells into miR-25-3p imic, please refer to FIG. 6D, qRT-PCR also showed a decrease in PI3K and MEK mRNA levels. Wherein Raf, Ras, MEK and ERK are related factors of MAPK/ERK signaling pathway, and PI3K and AKT are related factors of PI3K/AKT signaling pathway. Thus, miR-25-3p can inhibit MAPK/ERK signaling pathway and PI3K/AKT signaling pathway.
7. Small knot
In the embodiment of the invention, the experiment and analysis show that miR-25-3p expresses in the liver of a HIRI mouse and can reduce HIRI after a BMSCs exosome is injected into the HIRI mouse, and the experiment verifies that miR-25-3p exists in the BMSCs and exosome and participates in the BMSCs exosome to reduce HIRI injury.
After miR-25-3p is transfected into a mouse hepatocyte AML12 hypoxia reoxygenation model, qRT-PCR shows that the expression levels of inflammatory factors TNF-alpha, IL-6, IL-1 beta and IL-18 are reduced. miR-25-3p can also cause the expression quantity of apoptosis-related factors Caspase-1, NLRP3 and GSDMD in liver tissues of HIRI mice to be reduced. Therefore, miR-25-3p plays a role in inhibiting inflammation and apoptosis in HIRI, and miR-25-3p can reduce liver injury by detecting ALT and AST contents in serum and observing liver tissue structure.
Some studies have found that down-regulation of cellular communication network factor 1 (CCN 1) reduces ALT, AST levels, MPO activity, IL-6 and TNF-alpha levels in C57BL/6HIRI mice, inhibits MEK/ERK signaling pathways, and reduces inflammatory responses and liver injury. And researches also find that the expression of the BDNF is improved by inhibiting the expression of MEK-ERK-CREB after cerebral ischemia/reperfusion injury by the brain protease I (CH-I), thereby playing a role in neuroprotection and improving the nerve behavior function of a rat with cerebral middle artery embolism. Thus, inhibition of the MEK/ERK signaling pathway is shown to reduce the inflammatory response and to protect ischemic/reperfused organs. In addition, research finds that Octreotide (OCT) can inhibit p62 expression by inhibiting PI3K/AKT/mTOR/S757-ULK1 signal channel, improve the expression of Beclin-1, ATG7 and LC3, cause liver autophagy, maintain a series of antioxidant and anti-inflammatory cascade reactions, and play a role in protecting liver damaged by ischemia-reperfusion. Meanwhile, the PI3K/AKT pathway is involved in the regulation of apoptosis. And researches show that TLR4 is activated after spinal cord injury, and the expression of lncRNA-F630028O10Rik is promoted. The lncRNA is taken as a ceRNA of miR-1231-5p/Col1a1 axis, and the apoptosis of microglia after spinal cord injury is enhanced by activating a PI3K/AKT pathway. Thus, inhibition of the PI3K/AKT signaling pathway is shown to reduce the inflammatory response and protect ischemic/reperfused organs. Thus, inhibition of the MEK/ERK signaling pathway and the PI3K/AKT signaling pathway may reduce the inflammatory response and protect ischemic/reperfused organs.
Experiments show that miR-25-3p reduces the content of Raf, Ras, MEK and ERK of MAPK/ERK signal pathways and also reduces the content of PI3K/AKT signal pathways, namely PI3K and AKT. Therefore, miR-25-3p inhibits the MAPK/ERK signaling pathway and the PI3K/AKT signaling pathway.
In conclusion, miR-25-3p inhibits HIRI injury-induced hepatocyte apoptosis by inhibiting MAPK/ERK and PI3K/AKT signal pathways, so that HIRI is improved, and a new idea is provided for treating HIRI.
The above embodiments are only preferred embodiments of the present invention, and the scope of the present invention should not be limited thereby, and any insubstantial changes and substitutions made by those skilled in the art based on the present invention are intended to be covered by the claims.
Claims (6)
1. Use of a mesenchymal stem cell exosome for the preparation of a medicament for treating hepatic ischemia reperfusion injury, the medicament for inhibiting the MAPK/ERK signalling pathway and the PI3K/AKT signalling pathway.
2. The use of claim 1, wherein said bone marrow mesenchymal stem cell exosomes comprise miR-25-3 p.
3. The application of miRNA derived from mesenchymal stem cell exosomes in preparation of a medicine for treating liver ischemia-reperfusion injury is characterized in that the medicine is used for inhibiting a MAPK/ERK signaling pathway and a PI3K/AKT signaling pathway.
4. The use of claim 3, wherein the miRNA is miR-25-3 p.
5. A medicament for the treatment of hepatic ischemia reperfusion injury comprising a component which inhibits the MAPK/ERK signaling pathway and the PI3K/AKT signaling pathway.
6. The medicament for treating hepatic ischemia-reperfusion injury according to claim 5, wherein the medicament comprises miR-25-3 p.
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