KR20220030552A - Hepatocyte proliferation or anti-inflammation composition comprising exosomes from placenta - Google Patents

Abstract

The present invention relates to a pharmaceutical composition or a health functional food composition for alleviating, preventing or treating liver disease, and having anti-inflammatory efficacy which contains the placenta-derived exosomes to have excellent hepatocellular proliferation effect and anti-inflammatory effect. The placenta-derived exosome is a composition obtained by treating the placenta with trypsin-EDTA.

Description

Hepatocyte proliferation or anti-inflammatory efficacy composition comprising placental-derived exosomes {HEPATOCYTE PROLIFERATION OR ANTI-INFLAMMATION COMPOSITION COMPRISING EXOSOMES FROM PLACENTA}

The present invention relates to a composition for hepatocellular proliferation or anti-inflammatory effect comprising a placenta-derived exosome, specifically, a pharmaceutical composition for improving liver function, preventing or treating liver disease, and treating or preventing a disease related to inflammation, and It relates to a health functional food composition.

Research has been reported that the secretome contains various bioactive factors that control the behavior of cells, and in particular, 'exosome' or 'extracellular vesicle' with many functions in the cellular secretion )', so research on its ingredients and functions is being actively conducted. Cells release various membrane types of ERs to the extracellular environment, and these ERs are commonly referred to as extracellular vesicles (EVs). The extracellular vesicles are also called cell membrane-derived ERs, ectosomes, shedding vesicles, microparticles, exosomes, and the like, and in some cases, they are used separately from exosomes.

Exosomes are nano-sized bi-lipid membrane vesicles secreted from living cells that play an important role in cell-to-cell communication. During human pregnancy, the placenta plays an important role in regulating physiological homeostasis or supporting fetal development. It is known that extracellular vesicles and exosomes secreted from the placenta contribute to communication between the placenta and maternal tissues to maintain maternal-fetal resistance. Exosomes contain active biological substances including lipids, cytokines, microRNAs, mRNAs and DNA, as well as proteins that may be present on the surface of exosomes. Exosomes are known to be useful in a number of therapeutic approaches, including immune modulation, promotion of angiogenesis, and drug delivery. Therefore, the demand for more efficient methods that makes it possible to isolate large amounts of exosomes is clearly increasing recently.

The liver is one of the major human organs and is responsible for protein synthesis, metabolism, especially lipid metabolism. Fatty liver caused by alcohol or metabolic syndrome causes general abnormalities in the body. Dietary therapy is being implemented as a treatment for alleviating the symptoms of lipid or fat accumulation. However, to date, a method for fundamentally treating lipid or fat accumulation in the liver has not been developed.

Since inflammatory diseases can be acute, chronic, ulcerative, allergic or necrotic, whether any disease is acute, chronic, ulcerative, allergic or necrotic so long as it falls within the definition of an inflammatory disease as above. regardless of gender

Hepatitis is a disease caused by viruses, alcohol, drugs, and immune disorders. It can be interpreted as an all-inclusive term. Fatty hepatitis is diagnosed by distinguishing between non-alcoholic steatohepatitis (NASH) and alcoholic steatohepatitis (ASH), in which an excessive amount of triglyceride is accumulated in the liver regardless of drinking. Fat Liver Disease) and non-alcoholic fatty liver (NAFLD, Non-alcoholic Fat Liver Disease) are being diagnosed.

Int. J. Mol. Sci. 2019, 20, 2434; doi:10.3390/ijms20102434 Extra Purified Exosomes from Human Placenta Contain an Unpredictable Small Number of Different Major Proteins 2010 Fall Conference Proceedings of the Korean Liver Society Invasive/noninvasive diagnosis in nonalcoholic fatty liver disease/nonalcoholic steatohepatitis

It is an object of the present invention to provide a hepatocyte proliferation or anti-inflammatory effect pharmaceutical composition or food composition comprising a placenta-derived exosome.

It is also an object of the present invention to provide a pharmaceutical composition or food composition for improving liver function, preventing or treating liver disease, including exosomes obtained by treating the placenta with trypsin-EDTA.

It is an object of the present invention to provide a hepatocyte proliferation or anti-inflammatory effect pharmaceutical composition or food composition comprising exosomes obtained by treating dry placenta with trypsin-EDTA.

An object of the present invention is to improve the symptoms of jaundice caused by hepatitis (NASH, ASH, NAFLD or AFLD), liver cirrhosis, liver fibrosis, cirrhosis, fatty liver or liver disease, including liver tissue regeneration including placental-derived exosomes, improvement of liver function , to provide a pharmaceutical composition or food composition for prevention or treatment.

One. A pharmaceutical composition for hepatocellular proliferation or anti-inflammatory efficacy comprising placental-derived exosomes.

2. The composition of the above 1, wherein the placenta-derived exosome is obtained by treating the placenta with trypsin-EDTA.

3. The composition according to the above 1, characterized in that the exosome derived from the dried placenta dried placenta.

4. The pharmaceutical composition of claim 1, wherein the placental-derived exosome expresses one or more markers selected from the group consisting of CD63, CD81, ALIX, FLOT1, ICAM1, EpCam, ANXA5 and TSG101.

5. In the above 1, hepatitis (NASH, ASH, NAFLD or AFLD) or liver cirrhosis, liver fibrosis, cirrhosis, fatty liver or jaundice caused by liver disease, including liver tissue regeneration, liver function improvement, hepatocyte proliferation or hepatocellular protective effect A pharmaceutical composition for improving symptoms.

6. Hepatocyte proliferation or anti-inflammatory functional food composition comprising placental-derived exosomes.

7. The composition of the above 6, wherein the placenta-derived exosome is obtained by treating the placenta with trypsin-EDTA.

8. The composition according to the above 6, characterized in that it is an exosome derived from dry placenta in which the placenta is dried.

9. The food composition according to the above 6, wherein the placenta-derived exosome expresses one or more markers selected from the group consisting of CD63, CD81, ALIX, FLOT1, ICAM1, EpCam, ANXA5 and TSG101.

10. In the above 6, liver tissue regeneration, liver function improvement, hepatocyte proliferation or hepatocyte protective effect, including hepatitis (NASH, ASH, NAFLD or AFLD), cirrhosis, liver fibrosis, cirrhosis, fatty liver or jaundice caused by liver disease Symptom improvement functional food composition.

11. The method according to any one of 1 or 6 above, wherein the exosome is separated using a commercially available exosome separation reagent, TFF method, ultracentrifuge, ultrafiltration, and SEC (Size Exclusion Chromatography). One or more of the methods to separate the exosomes by a method characterized in that the composition.

The exosome according to the present invention proliferates hepatocytes and has an anti-inflammatory effect, so the effect of regenerating liver tissue is excellent, so that hepatitis (NASH, ASH, NAFLD or AFLD), liver cirrhosis, liver It can be used for the purpose of preventing, improving, or treating the symptoms of jaundice caused by fibrosis, cirrhosis, fatty liver or liver disease.

1 is a result of measuring the size distribution and concentration of exosomes derived from the placenta by NTA (Nanoparticle Tracking Analysis).
Figure 2 is a TEM (Transmission Electron Microscopy; Transmission Electron Microscopy) results of exosomes derived from the placenta.
3 is a result of confirming the surface protein marker of the exosome derived from the placenta using an antibody.
4 is a result of confirming the cell proliferation ability of exosomes derived from placenta in human liver cancer cells (HepG2).
5 is a result of confirming the anti-inflammatory efficacy in mouse macrophages (RAW264.7) of exosomes derived from the placenta.

Hereinafter, the present invention will be described in detail.

In addition, the exosome derived from the placenta of the present invention prevents the symptoms of jaundice caused by hepatitis (NASH, ASH, NAFLD or AFLD), cirrhosis, liver fibrosis, cirrhosis, fatty liver or liver disease, including liver tissue regeneration and improvement of liver function , was found to have an effect of improving or treating, thereby completing the present invention.

The placenta-derived exosome according to the present invention may be obtained from, for example, drying or culturing the placenta, separating and purifying it, and the remaining culture medium or supernatant, but does not contain placental cells and does not contain culture supernatant.

In certain embodiments, purified exosomes are formulated into pharmaceutical compositions suitable for administration to a subject in need thereof. In certain embodiments, the subject is a human. The placental-derived exosome-containing pharmaceutical composition provided herein is administered to a subject in need thereof locally, subcutaneously, systemically, parenterally, intravenously, intramuscularly, externally (topical), orally, intradermally, It may be formulated for transdermal or intranasal administration. In certain embodiments, a pharmaceutical composition containing placental derived exosomes provided herein is formulated for local administration. In certain embodiments, the pharmaceutical compositions containing placental derived exosomes provided herein are formulated for systemic subcutaneous administration. In certain embodiments, a pharmaceutical composition containing placental derived exosomes provided herein is formulated for parenteral administration. In certain embodiments, a pharmaceutical composition containing placental derived exosomes provided herein is formulated for intramuscular administration. In certain embodiments, a pharmaceutical composition containing placental-derived exosomes provided herein is formulated for topical administration. In certain embodiments, the placental-derived exosome-containing pharmaceutical compositions provided herein are formulated for oral administration. In certain embodiments, a pharmaceutical composition containing placental-derived exosomes provided herein is formulated for intradermal administration. In certain embodiments, a pharmaceutical composition containing placental-derived exosomes provided herein is formulated for transdermal administration. In certain embodiments, a pharmaceutical composition containing placental derived exosomes provided herein is formulated for intranasal administration. In a specific embodiment, the placental-derived exosome-containing pharmaceutical composition provided herein is formulated for intravenous administration.

The placenta-derived exosome according to the present invention is obtained by using a commercially available exosome separation reagent (eg, ExoQuick-TC™) or by using the TFF (Tangential Flow Filtration) method. In addition to this method, exosomes can be obtained from the placenta, a culture medium of the placenta, or a dry placenta using ultracentrifuge, ultrafiltration, or SEC (Size Exclusion Chromatography) methods. It is not limited only to the exosome obtaining method described in.

Also provided is a composition comprising exosomes derived from human placenta, wherein the exosomes are CD1c, CD20, CD24, CD25, CD29, CD2, CD3, CD8, CD9, CD11c, CD14, CD19, CD31, CD40, CD41b, CD42a, CD44, CD45, CD49e, CD4, CD56, CD62P, CD63, CD69, CD81, CD86, CD105, CD133-1, CD142, CD146, CD209, CD326, HLA-ABC, HLA-DRDPDQ, MCSP, ROR1, SSEA- 4 or a combination thereof.

The exosomes described herein contain specific markers. Such markers may be useful, for example, in the identification of exosomes, and in differentiating them from other exosomes, eg, exosomes not derived from the placenta. In certain embodiments, such exosomes are positive for one or more markers determinable by flow cytometry, such as fluorescence-activated cell sorting (FACS). In addition, the exosomes provided herein can be identified based on the absence of specific markers. Determination of the presence or absence of such markers can be determined using methods known in the art, for example, fluorescence-activated cell sorting (FACS).

In some embodiments, the exosomes are CD3-, CD11b-, CD14-, CD19-, CD33-, CD192-, HLA-A-, HLA-B-, HLA-C-, HLA-DR-, CD11c - or CD34-. In some embodiments, the exosomes are CD3-, CD11b-, CD14-, CD19-, CD33-, CD192-, HLA-A-, HLA-B-, HLA-C-, HLA-DR-, CD11c- and CD34 -am.

In some embodiments, exosomes comprise non-coding RNA molecules.

Specifically, in the present invention, inflammatory diseases include atopic dermatitis, psoriasis, dermatitis, allergy, arthritis, rhinitis, otitis media, sore throat, tonsillitis, cystitis, nephritis, pelvic inflammation, inflammatory bowel disease, ankylosing spondylitis, systemic lupus erythematodes (SLE) , atherosclerosis, asthma, arteriosclerosis, edema, rheumatoid arthritis, delayed allergy (type IV allergy), transplant rejection, graft-versus-host disease, autoimmune encephalomyelitis, multiple sclerosis, arthritis, cystic fibrosis, diabetic retinopathy, rhinitis , ischemic-reperfusion injury, vascular restenosis, glomerulonephritis, gastrointestinal allergy, etc., but are not limited thereto. Examples of diseases associated with inflammation include acne vulgaris, asthma, autoimmune diseases, celiac disease, chronic prostatitis, glomerulonephritis, hypersensitivities, inflammatory bowel disease inflammatory bowel diseases, pelvic inflammatory disease, reperfusion injury, rheumatoid arthritis, sarcoidosis, transplant rejection, vasculitis, and interstitial cystitis ) is included. Many diseases are associated with inflammation or are classified as autoimmune diseases. Some different types of inflammatory diseases include gout, lupus, asthma, pleurisy, eczema, arthritis, gastritis, splenitis, sinusitis, hepatitis ( hepatitis, nephritis, psoriasis, vasculitis, laryngitis, thyroiditis, prostatitis, pharyngitis, sarcoidosis, atherosclerosis, allergic reactions, multiple sclerosis sclerosis, some myopathies, rheumatoid arthritis, seborrheic dermatitis, Wegener's granulomatosis, irritable bowel syndrome (IBS; Crohn's disease), ulcerative colitis colitis) and diverticulitis. In addition, diseases related to inflammation include sepsis, polychondritis, scleroderma, periodontitis, gingivitis, Behcet's syndrome, vasculitis, Kawasaki disease, pancreatitis, bronchitis, inflammatory skin disease, stomatitis, peritonitis, stroke, cerebral infarction, Alzheimer's disease, Parkinson's disease, and paresis Jet's disease, conjunctivitis, pneumonia, gastric ulcer, colitis, hemorrhoids, rheumatoid lupus, parotiditis, tendinitis, myositis, Sjogren's syndrome, appendicitis, encephalitis, encephalomyelitis, biliary tract infection, mastitis, iritis, scleritis, uveitis, aphthous stomatitis, etc. It is not limited to the diseases described in

The pharmaceutical composition of the present invention may be formulated as a pharmaceutical preparation for the prevention and treatment of skin diseases or liver diseases, including pharmaceutically acceptable carriers, diluents or excipients.

In another embodiment, exosomes and/or pharmaceutical compositions comprising exosomes described herein can be used as hepatoprotective agents. In another aspect, the exosomes and/or the pharmaceutical composition comprising the exosomes described herein may be provided in the form of a kit suitable for pharmaceutical use.

The carrier, excipient and diluent include, for example, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose , microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

The pharmaceutical composition of the present invention is formulated into oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, or sterile injection solutions according to conventional methods to be prepared into pharmaceutical preparations. In general, in the case of formulation, it may be prepared using a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, a surfactant, etc. usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and such solid preparations include at least one excipient in the pharmaceutical composition, for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc. It can be prepared by mixing. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Liquid formulations for oral use include suspensions, solutions, emulsions, and syrups. In addition to commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives are included. can Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, and the like. Non-aqueous solvents and suspending agents include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin, and the like can be used.

The dosage of the pharmaceutical composition of the present invention may vary depending on the age, sex, and weight of the patient, but generally in an amount of 0.1 to 100 mg/kg, preferably in an amount of 1 to 30 mg/kg, once a day to several It can be administered in divided doses. Also, the dosage may be increased or decreased according to the route of administration, the degree of disease, sex, weight, age, health status, diet, administration time, administration method, excretion rate, etc. Accordingly, the above dosage does not limit the scope of the present invention in any way.

The pharmaceutical composition of the present invention may be administered to mammals such as rats, mice, livestock, and humans by various routes. Any mode of administration can be envisaged, for example, dermal application, oral, rectal or intravenous, intramuscular, subcutaneous, intradermal, intrauterine dural administration, intranasal (nasal), inhalation administration using a nebulizer, etc. Alternatively, it may be administered by intracerebrovascular injection or the like. For the purpose of improving or preventing liver disease, the pharmaceutical composition of the present invention may be administered directly into the portal vein or hepatic vein.

Hereinafter, in order to describe the present invention in detail, examples will be described in detail.

Example 1. Obtaining exosomes from placenta

1.1 Obtaining and pretreatment of the placenta

The placenta prior to obtaining the exosome of the present invention is derived from a human, but is not limited thereto.

In order to homogenize the dry placenta, Trypsin-EDTA reagent was used. 250 mg of dried placental tissue was mixed with 20 mL of Trypsin 0.25%-EDTA reagent and stirred vigorously. This mixture was incubated at 4 °C for 72 h. It was also possible to incubate at 37° C. when a trypsin reagent was used. Thereafter, centrifugation was performed at 6,000 rpm for 30 minutes to remove tissue and cellular impurities in the mixture, and the supernatant was purified using a 0.22 μm filter. If a 0.22㎛ filter is used, cells and cell debris present in the pretreated dry placenta are removed.

1.2 Obtaining exosomes from placenta

For the isolation of exosomes from the placenta, a commercial exosome isolation reagent ExoQuick-TC™ (System Biosciences) was used.

Mix the pretreated dry placenta solution from which cells and cellular impurities have been removed with ExoQuick-TC™ in a ratio of 5:1. This was incubated overnight at 4 °C. The next day, centrifuged at 1,500 x g at 4 °C for 30 min and the supernatant was removed. After centrifugation was performed once more at 1,500 x g at 4 °C for 5 minutes to remove all possible remaining fluid, the supernatant was completely removed. 200 μl of PBS was added to the exosome pellet and resuspended. The exosomes isolated in this process were named exosomes (EXQ) for convenience.

1.3 Isolation and purification of exosomes by TFF method

Exosomes were isolated and concentrated using a TFF (Tangential Flow Filtration) method from the placenta filtered with a 0.22㎛ filter. A cassette filter (Merck Millipore) was used for the TFF method. The TFF filter can be selected for the purpose of various molecular weight cutoff (MWCO), and in this experiment, a filter with a MWCO of 300,000 Da (Dalton) was used in consideration of the molecular weight of the exosome. Exosomes were selectively separated and concentrated by the selected MWCO filter, and particles smaller than 300,000 Da, proteins, lipids, nucleic acids, and low molecular weight compounds were removed.

The following experiments were conducted using the exosomes (EXQ) of the above Examples, but the method of obtaining the exosomes in order to achieve the object of the present invention is ultracentrifuge, ultrafiltration, or SEC (Size Exclusion). Chromatography), and is not limited to the exosome obtaining method described herein.

Example 2. Identification of exosomes

2.1 NTA (Nanoparticle Tracking Analysis)

Exosomes isolated from the placenta were appropriately diluted in PBS, and the size distribution and concentration of exosome particles were measured using Nanosight (NS300, Malvern, UK) according to the device usage method.

As a result of nanoparticle tracking analysis (NTA) to measure the size distribution and concentration of the exosomes, it was confirmed that the particle size distribution of the placental-derived exosomes (EXQ) was approximately 50-200 nm. The average particle size of placental-derived exosomes (EXQ) is about 177 nm. (Fig. 1)

2.2 TEM (Transmission Electron Microscopy)

For morphological analysis of the exosomes, the exosomes isolated from the placenta were fixed on a grid for 1 minute and washed with sterile water. After staining with 2% uranyl acetate for 20 seconds, the completely dried grid was photographed with a TEM (Transmission electron microscope, Talos L120C, FEI, Czech).

As a result of TEM imaging, it was confirmed that the exosomes of the present invention were spherical particles of various sizes and shapes. (Fig. 2)

2.3 Antibody analysis of exosome surface markers (Exosome Ab array)

Exo-Check Exosome Antibody Arrays (EXORAY200A-4, System Bioscience, USA) were used to identify 8 types of surface protein markers (CD63, CD81, ALIX, FLOT1, ICAM1, EpCam, ANXA5, TSG101) of exosomes isolated from placenta. was used. Exosomes separated from the placenta and 1 X blocking buffer were added to the membrane provided by the Exo-Check Exosome Antibody Arrays kit and reacted at 2-8 °C for 24 hours. After the reaction, it was washed twice with 1 X Washing buffer at room temperature for 5 minutes each. Then, after adding 5ml Detection buffer and reacting at room temperature for 30 minutes, luminescence signals were obtained and the expression level of each protein was analyzed by chemiluminescence (FUSION_SOLO.6S, Vilber, France).

As a result of analyzing the protein markers of placental-derived exosomes (EXQ) with 8 types of antibodies, the presence of all 8 types of antibodies including CD63, CD81, ALIX, FLOT1, ICAM1, EpCam, ANXA5, and TSG101 was confirmed. In particular, it was confirmed that the expression levels of CD81, a type of tetraspanin, a membrane protein, and ICAM or ANXA5 markers were higher than those of other protein markers. (Fig. 3)

Experimental Example 1. Evaluation of hepatocyte proliferation ability

To evaluate the liver function improvement efficacy of the placental-derived exosomes of the present invention, more specifically, the hepatocellular proliferation and hepatocellular protective efficacy, human hepatocellular carcinoma HepG2 cells were dispensed into a 96-well plate at 1 x 10 ⁴ cells per well, and then Cell culture conditions were cultured for 24 hours. The hepatocyte proliferation efficacy was confirmed by removing the existing medium, treating the test substance, and culturing alone for 24 hours.

In addition, after pretreatment of the test substance for 24 hours, 20 mM of D-Galactosamine (D-GalN), which is known to induce hepatotoxicity, was treated. For the test substance in this experiment, the negative control group (-), the FBS-treated group, and the placental-derived exosome (EXQ) were diluted 2-fold from 10% to 2.5%, respectively, and CCK-8 kit assay was performed. The FBS-treated group is a MEM medium containing 10% FBS corresponding to the medium conditions in which the HepG2 cells used in this experiment proliferate smoothly, and was used for the purpose of a positive control.

After washing with DPBS, in order to measure the cell viability, 1/10 of the Cell Counting Kit-8 reaction solution (Dojindo) was diluted in a medium excluding FBS and treated with 100 μl per well for 30 minutes to 1 hour. After reacting in an incubator for a while, absorbance was measured at 450 nm using a spectrophotometer.

As a result of the experiment, hepatocyte proliferative capacity was increased in a concentration-dependent manner by placental-derived exosomes, and in particular, the 10% treatment group of placental-derived exosomes exhibited up to 179% of the proliferative capacity promotion compared to the control group.

In addition, hepatocellular protection efficacy was confirmed by constructing a cytotoxicity model by treating D-GalN, which is known to induce hepatotoxicity by inhibiting RNA and protein synthesis in hepatocytes. As a result, it was confirmed that the cell viability decreased by D-GalN treatment was effectively recovered by the dry placenta-derived exosomes, which can be inferred to have hepatocellular protective efficacy against liver damage diseases. It can be inferred that the hepatocyte proliferation efficacy of placental-derived exosomes is due to growth factors or physiologically active substances present in the placenta. (Fig. 4)

Experimental Example 2. Evaluation of anti-inflammatory efficacy in macrophages

Mouse macrophages (RAW264.7 cells) were seeded in a 96-well plate at 1 x 10 ⁵ cells/well and cultured for 24 hours in cell culture conditions. The old medium was removed, and lipopolysaccharide (LPS) and test substance (EXQ) were simultaneously treated and incubated for 24 hours. The LPS used at this time is one of the components of the cell wall of Gram-negative bacteria and is a representative pathogenic substance that induces an inflammatory response in cells. Then, the cell culture solution and the Griess reagent were mixed in a 1:1 ratio and reacted at room temperature for 15 minutes. And the absorbance was measured at 540 nm using a spectrophotometer, and through this, the ability to inhibit nitric oxide (NO) production was measured. As a positive control group, diclofenac, a nonsteroidal anti-inflammatory drug used for the treatment of inflammation, was used, and the inhibition rate of NO production in each group was calculated as a percentage (%) compared to that of the control group.

To evaluate the anti-inflammatory efficacy of exosomes derived from dry placenta, the test substance (EXQ) was diluted two-fold from 10% to 1.25% and treated, followed by NO assay.

As a result of the experiment, NO production was decreased in a concentration-dependent manner by exosomes derived from dry placenta. That is, it was demonstrated that the NO production inhibitory ability of the dry placenta-derived exosomes showed a concentration dependence. The dry placenta-derived exosomes decreased from 5% and 10% to 37.5% and 22.5%, respectively, which was lower than that of diclofenac (12.5 ug/mL), a positive control, 63%. Therefore, it can be seen from this study that dry placenta-derived exosomes have the potential to be used as a treatment for inflammation-related diseases. (Fig. 5)

Claims (11)

A pharmaceutical composition for hepatocellular proliferation or anti-inflammatory efficacy comprising placental-derived exosomes.
The composition according to claim 1, wherein the placenta-derived exosome is obtained by treating the placenta with trypsin-EDTA.
The composition according to claim 1, wherein the composition is an exosome derived from dry placenta obtained by drying the placenta.
The pharmaceutical composition according to claim 1, wherein the placental-derived exosome expresses one or more markers selected from the group consisting of CD63, CD81, ALIX, FLOT1, ICAM1, EpCam, ANXA5 and TSG101.
The method according to claim 1, wherein the liver tissue regeneration, liver function improvement, hepatocyte proliferation or hepatocyte protective effect, including hepatitis (NASH, ASH, NAFLD or AFLD), cirrhosis, liver fibrosis, cirrhosis, fatty liver or jaundice caused by liver disease A pharmaceutical composition for improving symptoms.
Hepatocyte proliferation or anti-inflammatory functional food composition comprising a placenta-derived exosome.
The composition according to claim 6, wherein the placenta-derived exosome is obtained by treating the placenta with trypsin-EDTA.
The composition of claim 6, wherein the composition is an exosome derived from dry placenta obtained by drying the placenta.
The food composition according to claim 6, wherein the placental-derived exosome expresses one or more markers selected from the group consisting of CD63, CD81, ALIX, FLOT1, ICAM1, EpCam, ANXA5 and TSG101.
The method according to claim 6, wherein the liver tissue regeneration, liver function improvement, hepatocyte proliferation or hepatocyte protective effect, including hepatitis (NASH, ASH, NAFLD or AFLD), cirrhosis, liver fibrosis, cirrhosis, fatty liver or jaundice caused by liver disease Symptom improvement functional food composition.
The method according to any one of claims 1 to 6, wherein the exosome is separated using a commercially available exosome separation reagent, TFF method, ultracentrifuge, ultrafiltration, and SEC (Size Exclusion Chromatography) A composition characterized in that it is separated by one or more exosome separation methods of a method comprising a.

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