Use of perirenal fat exosomes enriched in miRNA-486 to induce endothelial dysfunction

Abstract

The application discloses an application of perirenal adipose tissue exosomes rich in miRNA-486 in inducing endothelial dysfunction, wherein the perirenal adipose tissue exosomes are derived from adipose tissue around mammalian kidneys, rich in micro miRNA-486, have a particle size of 30-150 nanometers, have surface markers CD63, CD81 and TSG101, and have an expression level of the miRNA-486 in the perirenal adipose tissue exosomes increased by 3-5 times relative to a normal control, so that the perirenal adipose tissue exosomes can transmit the miRNA-486 to cells when being taken by vascular endothelial cells, and induce abnormal functions of the endothelial cells. The application proves that the perirenal adipose tissue exosomes rich in miR-486 can be taken by vascular endothelial cells in a specific manner through in-vivo and in-vitro experiments.

Description

Technical Field

[0001] This invention relates to the field of perirenal adipose exosome technology, and more particularly to the application of a perirenal adipose exosome rich in miRNA-486 in inducing endothelial dysfunction. Background Technology

[0002] Hypertension and its associated endothelial dysfunction are among the leading risk factors for cardiovascular and cerebrovascular disease incidence and mortality worldwide. In recent years, increasing research has focused on the role of adipose tissue, particularly perirenal adipose tissue, in regulating vascular function and the pathogenesis of hypertension. Perirrenal adipose tissue not only serves as an "inert" energy storage organ but also influences the physiological and pathological processes of the kidneys and blood vessels by secreting various bioactive molecules. Existing studies have shown that exosomes, secreted products of perirenal adipose tissue, play important functions in signal transduction, intercellular communication, and the regulation of the local microenvironment.

[0003] At the molecular regulatory level, miRNAs, as a class of non-coding small RNAs, can participate in multiple aspects such as vascular tone, endothelial cell function, and inflammatory responses by regulating target gene expression. Currently, the regulation of cardiovascular diseases by exosome-borne miRNAs has become a hot topic in cutting-edge international research. Some studies have shown that miRNA-486 is highly expressed in various tissues and is closely related to cell proliferation, apoptosis, and metabolism, but its expression characteristics and pathogenic mechanism in perirenal adipose-derived exosomes remain unclear.

[0004] Although some literature reports the role of adipose tissue exosomes in cardiovascular disease, most studies focus on subcutaneous, peritoneal, or epicardial fat. Our understanding of the mechanisms by which perirenal fat-derived exosomes, particularly in hypertension and endothelial dysfunction, is limited. Traditional research has largely focused on classic regulatory pathways such as the renin-angiotensin system and the sympathetic nervous system, lacking systematic evidence on the mechanisms by which the adipose-vascular axis and perirenal fat influence vascular endothelial function through exosomes carrying miRNAs.

[0005] Regarding the function of miRNA-486, previous literature mainly focused on its regulatory role in tissues such as cardiomyocytes and skeletal muscle, but the impact of miRNA-486 on vascular endothelial cell function via the perirenal adipose exosome transport pathway has not been fully elucidated. Most existing studies are limited to the cellular level or whole-cell miRNA knockout animal models, lacking precise research on the perirenal adipose microenvironment and local signal transduction. Furthermore, there is a lack of in vivo and in vitro experimental models that directly reflect the endothelial damage and hypertension caused by abnormal miRNA-486 expression.

[0006] Furthermore, current technologies for disease intervention primarily focus on systemic miRNA regulation or drug-based antihypertensive therapies, lacking targeted intervention strategies based on the perirenal adipose tissue-exosome-specific miRNA signaling axis. There is currently no systematic research or mature technical solution regarding how to specifically inhibit the expression, delivery efficiency, target tissue localization, and comprehensive effects of perirenal adipose tissue miRNA-486 on hypertension and vascular endothelial dysfunction. Related drugs or treatments also have significant room for improvement in terms of safety, controllability, and long-term efficacy. Summary of the Invention

[0007] One objective of this invention is to propose the application of perirenal adipose exosomes rich in miRNA-486 in inducing endothelial dysfunction. This invention systematically demonstrates through in vitro and in vivo experiments that perirenal adipose exosomes rich in miR-486 can be specifically taken up by vascular endothelial cells.

[0008] According to an embodiment of the present invention, a perirenal adipose exosome rich in miRNA-486 is provided. The perirenal adipose exosome is derived from the adipose tissue surrounding the kidney of mammals and is rich in micromiRNA-486. The perirenal adipose exosome has a particle size of 30–150 nm and contains surface markers CD63, CD81, and TSG101. The expression level of miRNA-486 in the perirenal adipose exosome is 3–5 times higher than that in the normal control, which enables the perirenal adipose exosome to deliver miRNA-486 into the cells when it is taken up by vascular endothelial cells, thereby inducing endothelial cell dysfunction.

[0009] Optionally, the miRNA-486 is specifically miRNA-486-5p, and the perirenal adipose exosomes are derived from perirenal adipose tissue in a pathological state of hypertension, with the content of miRNA-486-5p being higher than that in a normal physiological state.

[0010] The use of a miRNA-486-rich perirenal adipose exosome in a drug for treating endothelial dysfunction or hypertension in mammals or humans, wherein the miRNA-486-rich perirenal adipose exosome exerts its effect by targeting exosome-mediated pathogenic pathways, blocking abnormal perirenal adipose signaling damage to vascular endothelium, improving vascular endothelial function, lowering blood pressure and alleviating organ fibrosis.

[0011] Optionally, the drug is prepared as a topical administration formulation targeting perirenal adipose tissue and contains at least one inhibitor of miRNA-486 as an active ingredient.

[0012] Optionally, the miRNA-486 inhibitor is selected from the following components: a recombinant adeno-associated virus vector expressing an anti-miRNA-486 sequence in perirenal adipose tissue, wherein the anti-miRNA-486 sequence is a sponge sequence or a robust decoy RNA targeting miRNA-486, or an oligonucleotide directly targeting miRNA-486.

[0013] The beneficial effects of this invention are: This invention delivers miRNA-486 to perirenal adipose tissue via local injection, reducing the level of miRNA-486 by at least 50%, thereby achieving antihypertensive and endothelial function-improving effects in animal models: systolic blood pressure decreased by more than 20%, endothelial-dependent diastolic function recovered to more than 80% of normal levels, and fibrotic lesion markers in the aorta, mesenteric arteries, and cardiovascular tissues were significantly reduced. Detailed Implementation

[0014] The invention will now be described in further detail.

[0015] A perirenal adipose exosome rich in miRNA-486, derived from the adipose tissue surrounding the kidneys of mammals, is rich in micromiRNA-486. The perirenal adipose exosomes have a particle size of 30–150 nm and contain surface markers CD63, CD81, and TSG101. The expression level of miRNA-486 in the perirenal adipose exosomes is 3–5 times higher than that in the normal control, enabling the perirenal adipose exosomes to deliver miRNA-486 into the cells when taken up by vascular endothelial cells, thereby inducing endothelial cell dysfunction.

[0016] In this embodiment, miRNA-486 specifically refers to miRNA-486-5p. The perirenal adipose exosomes are derived from perirenal adipose tissue in a pathological state of hypertension, and the content of miRNA-486-5p is increased relative to the normal physiological state.

[0017] In the perirenal adipose exosomes of spontaneously hypertensive rats, the relative expression level of miRNA-486-5p was more than 4 times that of the normotensive control. Moreover, miRNA-486-5p was significantly enriched in perirenal adipose tissue cells compared with the level of miRNA-486-5p in perirenal adipose tissue cells. The isolation and identification of perirenal adipose exosomes showed typical cup-shaped vesicle structures by transmission electron microscopy, and nanoparticle size analysis confirmed that their particle size was mainly distributed in the range of 50–150 nm.

[0018] The use of a miRNA-486-rich perirenal adipose exosome in a drug for treating endothelial dysfunction or hypertension in mammals or humans, wherein the miRNA-486-rich perirenal adipose exosome exerts its effect by targeting exosome-mediated pathogenic pathways, blocking abnormal perirenal adipose signaling damage to vascular endothelium, improving vascular endothelial function, lowering blood pressure and alleviating organ fibrosis.

[0019] In whole animal models, administration of exosomes can induce vascular endothelial dysfunction and elevated arterial blood pressure. After injecting the exosomes into the perirenal fat region of normal-blood pressure rats, the elevation of arterial blood pressure in spontaneously hypertensive rats was reduced after the perirenal fat inhibited the secretion of miR-486, and the degree of fibrosis in the mesenteric artery, aorta, and heart of SHR rats was improved.

[0020] The drug works by targeting the pathogenic pathway mediated by this exosome. The drug contains an active ingredient that can inhibit or neutralize the pathogenic effects of miRNA-486 in perirenal adipose exosomes, thereby blocking the damage to the vascular endothelium caused by abnormal perirenal adipose signaling, thus improving vascular endothelial function, lowering blood pressure, and reducing organ fibrosis. This application is based on the discovery that perirenal adipose exosomes rich in miRNA-486 are one of the key pathological factors inducing essential hypertension and vascular endothelial dysfunction. By intervening in this exosome and its miRNA-486 pathway during drug preparation, the source control of the pathological process of hypertension and the restoration of endothelial function can be achieved.

[0021] The drug for treatment is formulated as a localized drug delivery formulation targeting perirenal adipose tissue and contains at least one miRNA-486 inhibitor as an active ingredient. The miRNA-486 inhibitor is selected from the following components: a recombinant adeno-associated virus vector capable of efficiently expressing an anti-miRNA-486 sequence in perirenal adipose tissue, wherein the efficiently expressed anti-miRNA-486 sequence is a sponge sequence or a robust decoy RNA targeting miRNA-486; and oligonucleotides that directly target miRNA-486, such as small interfering RNA or antisense oligonucleotides that specifically bind to miRNA-486 and promote its degradation.

[0022] The preferred formulation is delivered via local injection around the kidney to reduce the level of miRNA-486 in perirenal adipose tissue by at least 50%, thereby achieving antihypertensive and endothelial function-improving effects in animal models: systolic blood pressure is reduced by more than 20%, endothelial-dependent diastolic function is restored to more than 80% of normal levels, and fibrotic lesion markers in the aorta, mesenteric artery, and cardiovascular tissues are significantly reduced.

[0023] A therapeutically effective amount of miRNA-486 expression inhibitor was delivered to the perirenal adipose tissue to reduce the level of miRNA-486 in the perirenal adipose tissue, thereby improving vascular endothelial function and reducing blood pressure in the test mammals. The miRNA-486 expression inhibitor was introduced into the perirenal adipose region by targeted administration, and the adverse effects on the vascular system were corrected by locally inhibiting the abnormally high expression of miRNA-486 in the perirenal adipose tissue, thus achieving targeted treatment of the etiology of hypertension.

[0024] The miRNA-486 expression inhibitor is delivered via a viral vector to exert a sustained effect in perirenal adipose tissue. The vector is a recombinant adeno-associated virus containing an expression cassette encoding an antagonistic sequence against miRNA-486. This cassette is used to efficiently express robust bait RNA or sponge molecules against miRNA-486 in the target tissue, thereby persistently inhibiting miRNA-486 function. A long-term miRNA-486 inhibitory effect can be established in the perirenal adipose region by injecting the AAV vector into the region once. In a spontaneously hypertensive rat model, after injecting the AAV vector carrying the miRNA-486-TuD sequence into the perirenal adipose tissue, the mean arterial pressure of the rats decreased by approximately 15-20 mmHg within 8 weeks, and the degree of fibrosis in the mesenteric artery, aorta, and heart was significantly reduced compared with the control group.

[0025] miRNA-486 expression inhibitors are nucleic acid drugs that directly target miRNA-486. These inhibitors are synthetic oligonucleotides that specifically recognize miRNA-486. In the examples, double-stranded small interfering RNA (siRNA) or antisense oligonucleotides are used, chemically modified (2'-O-methyl or phosphorus-sulfur bond modification) to improve stability, and loaded into nanoparticle carriers for in vivo delivery. Blood pressure was continuously monitored for 8 weeks after a single injection. The inhibitors consistently suppressed abnormal local expression of miRNA-486 and improved blood pressure and endothelial function in hypertensive model animals. Compared with the untreated control, the experimental group rats treated for 8 weeks showed an average reduction of approximately 15% in systolic blood pressure, a significant increase in endothelial-dependent diastolic response, and a decrease in collagen deposition and inflammatory cell infiltration in the kidney tissue.

[0026] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A perirenal adipose exosome rich in miRNA-486, characterized in that: The perirenal adipose exosomes are derived from the adipose tissue surrounding the kidneys of mammals and are rich in micromiRNA-486. The perirenal adipose exosomes have a particle size of 30–150 nm and contain surface markers CD63, CD81, and TSG101. The expression level of miRNA-486 in the perirenal adipose exosomes is 3–5 times higher than that in the normal control, which allows the perirenal adipose exosomes to deliver miRNA-486 into the cells when they are taken up by vascular endothelial cells, thus inducing endothelial cell dysfunction.

2. The perirenal adipose exosome rich in miRNA-486 according to claim 1, characterized in that, The miRNA-486 specifically refers to miRNA-486-5p. The perirenal adipose exosomes are derived from perirenal adipose tissue in a pathological state of hypertension, and the content of miRNA-486-5p is increased relative to the normal physiological state.

3. The use of a perirenal adipose exosome rich in miRNA-486 in a drug for treating endothelial dysfunction or hypertension in mammals or humans, characterized in that, The perirenal adipose exosomes rich in miRNA-486 exert their effects by targeting the exosome-mediated pathogenic pathway, blocking the damage of abnormal perirenal adipose signaling to vascular endothelium, improving vascular endothelial function, lowering blood pressure, and reducing organ fibrosis.

4. The use of the perirenal adipose exosomes rich in miRNA-486 according to claim 3 in a medicament for treating endothelial dysfunction or hypertension in mammals or humans, characterized in that, The drug is prepared as a topical administration formulation targeting perirenal adipose tissue and contains at least one inhibitor of miRNA-486 as an active ingredient.

5. The use of the perirenal adipose exosomes rich in miRNA-486 according to claim 3 in a medicament for treating endothelial dysfunction or hypertension in mammals or humans, characterized in that, The miRNA-486 inhibitor is selected from the following components: a recombinant adeno-associated virus vector expressing an anti-miRNA-486 sequence in perirenal adipose tissue, wherein the anti-miRNA-486 sequence is a sponge sequence or a robust decoy RNA targeting miRNA-486, or an oligonucleotide directly targeting miRNA-486.