KR20190045135A - Composition for preventing acute kidney injury - Google Patents

Abstract

The present invention relates to a pharmaceutical composition for preventing, alleviating, or treating kidney disease. More specifically, the present invention relates to the pharmaceutical composition and health functional food for preventing acute kidney disease which contains, as an active component, a peroxisome proliferator activated receptor γ coactivator 1α (PGC-1α) protein or drug inducing the overexpression of the protein.

Description

[Technical Field] The present invention relates to a composition for preventing acute kidney injury,

The present invention relates to a pharmaceutical composition for the prevention of kidney disease, and more particularly, to a pharmaceutical composition for prevention of acute kidney disease comprising, as an active ingredient, a PGC-1α (peroxisome proliferator activated receptor γ coactivator 1α) protein or a drug inducing over- And to a health functional food.

Kidney is an important organ that maintains homeostasis. It regulates body fluid volume, ion concentration and pH in the blood, excretes waste such as metabolic waste, toxins, drugs, and controls blood pressure and other metabolic and endocrine functions. . It also activates vitamin D to help calcium in the small intestine, and is involved in the synthesis of various hormones.

A kidney disease is a state in which the kidneys fail to perform their excretion, control, metabolism, and endocrine function normally, resulting in a decrease in function or an abnormality. Decreased function due to renal damage results in increased kidney and related structures, atrophy of the kidney, changes in body fluid volume, electrolyte imbalance, metabolic acidosis, gas exchange disorder, impaired anti-infective function and accumulation of toxic toxins.

The renal diseases are classified into acute renal disease and chronic renal disease depending on the progression or are classified into glomerulonephritis caused by deposition of vascular complex depending on the cause of the disease, diabetic kidney disease accompanying diabetes or hypertensive kidney disease accompanying hypertension, Toxic nephropathy caused by administration of drugs such as antibiotics or anticancer drugs, and bacterial infection. Regardless of the cause of the renal disease, if chronic renal failure progresses and the glomerular filtration rate decreases to less than 50%, the glomerular filtration rate will be continuously decreased in most cases, eventually reaching end stage renal failure, Complications such as nervous system complications, gastrointestinal complications, immunological complications, infection or bone dystrophy occur and, in severe cases, leads to death.

In particular, acute kidney disease, which is a sudden decrease in renal function, is closely related to the increase in the mortality rate of patients. It is caused by various causes such as kidney damage, sepsis, ischemia / reperfusion injury, . In particular, the occurrence of acute renal disease in patients with underlying diseases such as diabetes and hypertension is even more dangerous. If the acute kidney disease is not completely treated, the possibility of conversion to chronic kidney disease is high, And precise treatment is required.

Because the causes of acute renal disease are very diverse, other mechanisms are involved in the progression stage. Therefore, although it is necessary to apply a therapeutic agent according to the progress of the disease, most drugs do not have appropriate therapeutic agents because many of them do not consider the progress of the disease. Oxidative stress has been shown to be a major risk factor for cardiovascular injury as well as an important cause of kidney disease, but its effectiveness has not yet been clearly demonstrated.

Tran M, et al. &Quot; α-promotes recovery after acute kidney injury during systemic inflammation in mice. "J Clin Invest. 2011 Oct; Vol.121 (10) pp.4003-14 (2011.09.01.)

The inventors of the present invention have made efforts to develop an agent capable of preventing or treating acute kidney disease by effectively inhibiting apoptosis, which is one of the main mechanisms of acute kidney disease. As a result, by confirming the mechanism of inhibition of PGC- It is possible to prevent or treat acute renal disease caused by death, and the present invention has been completed.

Accordingly, it is an object of the present invention to identify the molecular mechanisms that PGC-1α overexpressed in renal tubular cells exert on apoptotic signaling pathway by oxidative stress in renal tubular cells and to establish a point of action as an apoptosis inhibitor, Which is capable of preventing or treating acute kidney disease by using PGC-1 alpha which inhibits acute renal disease.

Another object of the present invention is to provide a composition for preventing, ameliorating or treating acute renal disease, which can be effectively used for treating a sudden decrease in renal function, which is an adverse effect caused by administration of various anti-cancer drugs.

The objects of the present invention are not limited to the above-mentioned objects, and other objects not mentioned can be clearly understood by those skilled in the art from the following description.

In order to accomplish the object of the present invention described above, the present invention provides a method for inducing overexpression of mRNA, PGC-1α protein and PGC-1α protein encoding a gene encoding PGC-1α (Peroxisome proliferator activated receptor γ coactivator 1α) And the drug that induces overexpression of the PGC-1 alpha protein is at least one selected from the group consisting of metformin, phenformin HCl, AICAR, A-769662, GSK621, Rosiglitazone, Rosiglitazone maleate and Pioglitazone And a pharmaceutical composition for preventing acute kidney disease.

In a preferred embodiment, the active ingredient increases phosphorylation of p38, and when phosphorylation of p38 is increased, inactivation of GSK3? (Glycogen synthase kinase 3 beta) is increased, and when inactivation of GSK3? Is increased, Nrf-2 (Nuclear factor-erythroid-2-related factor 2) protein expression is increased.

The present invention also provides a method for preventing acute renal disease comprising administering the above-described pharmaceutical composition to a mammal other than a human.

Also, the present invention includes at least one of mRNA of a gene coding for PGC-1α (peroxisome proliferator activated receptor γ coactivator 1α), PGC-1α protein and a drug inducing overexpression of PGC-1α protein as an active ingredient, Wherein the drug inducing overexpression of PGC-1 alpha protein is at least one selected from the group consisting of metformin, phenformin HCl, AICAR, A-769662, GSK621, Rosiglitazone, Rosiglitazone maleate and Pioglitazone. Provide food.

In a preferred embodiment, the active ingredient increases phosphorylation of p38, and when phosphorylation of p38 is increased, inactivation of GSK3? (Glycogen synthase kinase 3 beta) is increased, and when inactivation of GSK3? Is increased, Nrf-2 (Nuclear factor-erythroid-2-related factor 2) protein expression is increased.

The composition for preventing, ameliorating or treating acute renal disease according to the present invention is involved in an increase of phosphorylation of p38 in renal tubular cells of the kidney, due to an increase in GSK3? Inactivation, leads to increase of Nrf-2 protein, Inducing gene expression and inhibiting renal cell death. Therefore, the present invention can be effectively utilized as a pharmaceutical composition and a functional food material for preventing, alleviating, or treating various acute renal diseases.

FIG. 1 schematically shows a mechanism of PGC-1α protein involved in the signaling pathway of the 38 / GSK3β / Nrf-2 axis in inhibiting apoptosis induced by oxidative stress.
FIG. 2 shows data on survival rate and degree of apoptosis induced by induction of apoptosis in PGC-1α overexpressed renal tubular cells.
FIG. 3 shows data obtained by confirming the level of ROS in cytoplasm or mitochondria between PGC-1α-overexpressing cell line and control (Mock) cell line.
FIGS. 4A to 4E show data obtained by comparing changes in expression of proteins involved in cell death between PGC-1α overexpressing cell line and control (Mock) cell line. FIG.
FIG. 5 shows data obtained by confirming whether antioxidative and anti-apoptotic effects of PGC-1α-overexpressing cell lines are related to the expression of Nrf-2.
FIGS. 6A to 6D show data obtained by stepwise confirming that the effect of increasing PGC-1α overexpression on the expression of Nrf-2 is mediated by the p38 / GSK3β / Nrf-2 axis.

Although the terms used in the present invention have been selected as general terms that are widely used at present, there are some terms selected arbitrarily by the applicant in a specific case. In this case, the meaning described or used in the detailed description part of the invention The meaning must be grasped.

Hereinafter, the technical structure of the present invention will be described in detail with reference to the accompanying drawings and preferred embodiments.

However, the present invention is not limited to the embodiments described herein but may be embodied in other forms. Like reference numerals used to describe the present invention throughout the specification denote like elements.

The technical feature of the present invention is that a PGC-1α overexpressed in renal tubular cells of the kidney leads to an increase in phosphorylation of p38, an increase in GSK3β inactivation and an increase in protein of Nrf-2, 1α, PGC-1α, and PGC-1α, which has been shown to be effective in inhibiting apoptosis induced by oxidative stress through a signal transduction pathway. And to provide a pharmaceutical composition and a health functional food which can prevent, improve or treat kidney disease.

Here, PGC-1α (peroxisome proliferator-activated receptor gamma coactivator-1 alpha) binds to PPAR-gamma and several nuclear receptors to regulate gene expression associated with energy metabolism, and increases expression of mitochondria Specific amino acid sequence and nucleotide sequence information of the gene encoding the amino acid sequence can be obtained from a known database such as NCBI's GenBank, which promotes the biogenesis and increases the content of oxidative myofibers in the skeletal muscle. However, there is no evidence that PGC-1α is involved in the suppression of apoptosis induced by oxidative stress, which is one of the causes of acute renal disease in renal tubular cells.

As a result, when the expression of PGC-1 alpha gene is promoted or enhanced, the onset of acute kidney disease can be prevented or treated. Therefore, the expression of PGC-1 alpha gene of the present invention or PGC-1 alpha protein or PGC- Compositions comprising one or more drugs can be used to prevent or treat the onset of acute renal disease by the possibility of, or administration to, an acute renal disease. The present inventors have confirmed that PGC-1α protein expression is important to suppress apoptosis caused by oxidative stress, which is one of the major causes of acute kidney disease. Therefore, Overexpression of human PGC-1α was associated with increased phosphorylation of p38, increased GSK3β inactivation, increased Nrf-2 protein expression, and led to the expression of various genes involved in cell protection The signal transmission path was established.

Accordingly, the pharmaceutical composition for preventing, ameliorating or treating acute renal disease according to the present invention is a composition for inducing overexpression of mRNA, PGC-1 alpha protein and PGC-1 alpha protein coding for PGC-1 alpha (Peroxisome proliferator activated receptor gamma coactivator 1 alpha) One or more of the drugs as an active ingredient.

In other words, PGC-1α protein overexpressed in renal tubular cells stimulates the signaling pathway of p38 / GSK3β / Nrf-2 axis step by step to increase cell survival rate, remarkably lower ROS level of cytoplasm and mitochondria, This is because the anti-apoptotic effect is shown by inhibiting the expression of the involved protein. More specifically, as shown in FIG. 1, a signal transduction system involving an increase in p38 phosphorylation and an inactivation of GSK3β by activation of p38 was induced by the overexpressed PGC-1α protein. Inactivated GSK3β reacted with Nrf- 2 protein expression and the protective effect of Nrf-2 protein on renal cell protection.

The drug that induces overexpression of the PGC-1α protein may be any one selected from the group consisting of metformin, phenformin HCl, AICAR, A-769662, GSK621, Rosiglitazone, Rosiglitazone maleate, and Pioglitazone. Metformin, phenformin HCl AICAR, A-769662, and GSK621 are drugs known as AMPK activators. Rosiglitazone, Rosiglitazone maleate, and Pioglitazone are drugs known as PPARγ agonists.

As a result, the pharmaceutical composition of the present invention can be orally or parenterally administered to prevent, ameliorate or treat acute renal diseases of mammals including humans.

The pharmaceutical composition of the present invention contains, as an active ingredient, at least one of mRNA of a gene encoding PGC-1 alpha (peroxisome proliferator activated receptor gamma coactivator 1 alpha), a drug that induces overexpression of PGC-1 alpha protein and PGC- And may further include a pharmacologically acceptable carrier or excipient depending on the other formulation, the method of use, and the intended use. When provided as a mixture, the active ingredient may be included in the pharmaceutical composition in an amount of 0.0001 to 50% by weight.

The pharmaceutical composition of the present invention comprising a pharmaceutically acceptable carrier may be in the form of tablets, pills, powders, granules, capsules, suspensions, solutions, emulsions, syrups, sterilized aqueous solutions, nonaqueous solutions, suspensions, emulsions, And a suppository, and may be various oral or parenteral formulations. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablet pills, powders, granules, capsules and the like, which may contain one or more excipients such as starch, calcium carbonate, sucrose or lactose lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate, talc, and the like are also used. Examples of the liquid preparation for oral administration include various excipients such as wetting agent, sweetening agent, fragrance, preservative, etc. in addition to water and liquid paraffin, which are simple diluents commonly used for suspension, solution, emulsion and syrup . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the non-aqueous solvent and the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate. Examples of the suppository base include witepsol, macrogol, tween 61, cacao paper, laurin, glycerogelatin and the like.

 The pharmaceutical composition of the present invention can be administered orally or parenterally in a pharmaceutically effective amount. When administered parenterally, the composition can be administered topically to the skin, intravenously, subcutaneously, intramuscularly, intraperitoneally, ≪ / RTI > A pharmaceutical composition of the invention may be administered, wherein a " pharmaceutically effective amount " means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, The type and severity of the disease, age, sex, drug activity, drug sensitivity, time of administration, route of administration and rate of release, duration of treatment, factors including co-administered drugs and other well known factors in the medical field. The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. And can be administered singly or multiply. It is important to take into account all of the above factors and administer an amount that will achieve the maximum effect in the least amount without side effects.

The dose of the pharmaceutical composition for preventing, ameliorating or treating acute renal disease of the present invention may be appropriately selected depending on the purpose of use, the degree of addiction of the disease, the age, body weight, sex, history, Lt; / RTI > For example, the composition of the present invention comprising an active ingredient such as the mRNA of the PGC-1 alpha gene can be administered at about 0.1 ng to about 100 mg / kg, especially 1 ng to about 10 mg / kg per adult, It is preferable to administer the pharmaceutical composition of the present invention at a dose of about 0.0001 to 100 mg / kg, especially 0.001 to 10 mg / kg, more particularly 0.01 to 1 mg / kg, per adult, It is not limited, but it can be administered once a day or divided into several doses.

Next, the health functional food for preventing and ameliorating acute kidney disease according to the present invention comprises a mRNA of a gene coding for PGC-1 alpha (Peroxisome proliferator activated receptor gamma coactivator 1?), A PGC-1 alpha protein and a PGC- 1α protein as an active ingredient, it can be used for the purpose of preventing and improving symptoms of acute kidney disease.

In the present invention, the term 'health functional food' means a natural product or a processed product containing one or more nutrients. Preferably, the function of the food is determined by physical, biochemical, biotechnological, Means a food group that has been imparted with added value to function and express, and a food which is designed and manufactured so that the body control function related to regulation of bio-defense rhythm of food composition, prevention and recovery of disease, etc. is sufficiently expressed in living body. The health functional food may include a pharmaceutically acceptable food supplementary additive and may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of health functional foods.

The health functional food of the present invention includes, for example, various foods, beverages, gums, tea, and vitamin complex. In addition, it is also possible to use nutritional products such as special nutritious foods (eg crude oil, milk, baby food), health supplements, confectionery (eg snacks), dairy products (eg fermented milk, Beverages, two-oil, fermented beverages, etc.), but are not limited thereto. The food, beverage or food additives described above can be produced by a conventional production method.

The health functional food of the present invention can be used as a nutritional supplement, a vitamin, a mineral (electrolyte), a flavoring agent such as a synthetic flavor agent and a natural flavor agent, a colorant and an aging agent (cheese, chocolate etc.), a pectic acid and its salt, Salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, water, carbonating agents used in carbonated beverages and the like. These components can be used independently or in combination.

Thus, the health functional food of the present invention has various formulations as described above, and may have any one of powder, granule, tablet, capsule and beverage.

In one embodiment, when the health functional food of the present invention is implemented as a beverage, the overexpression of mRNA, PGC-1α protein and PGC-1α protein encoding a gene encoding PGC-1α (Peroxisome proliferator activated receptor γ coactivator 1α) There are no particular restrictions on the other ingredients other than those containing a drug that induces the drug, and may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary beverages. Examples of natural carbohydrates include monosaccharides such as disaccharides such as glucose and fructose such as maltose, sucrose and the like and polysaccharides such as dextrins, cyclodextrins and the like, and xylitol, Sorbitol, and erythritol. Natural flavors (tau martin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.) can be advantageously used as flavors other than those described above In the health functional food for the prevention and improvement of acute kidney disease, overexpression of mRNA, PGC-1α protein and PGC-1α protein encoding the active ingredient PGC-1α (Peroxisome proliferator activated receptor γ coactivator 1α) The content of the inducing drug can be suitably determined according to its intended use (for prevention or improvement).

In addition, the method for treating acute kidney disease of the present invention can treat acute kidney disease, including oral or parenteral administration of the above-mentioned pharmaceutical composition to treat acute renal diseases of mammals other than humans.

Example 1: Establishment of human renal tubular cell line overexpressing PGC-1?

Human renal tubular cells, HK-2 cells, were purchased from ATCC (Manassas, Va., USA) and cultured in DMEM-F12 medium supplemented with 10% fetal bovine serum (FBS), 50 U / ml penicillin and 50 μg / ml streptomycin was added, and the cells were cultured at 37 ° C and 5% CO ₂ . In order to continuously overexpress human PGC-1α in human renal tubular cells (HK-2 cells), human PGC-1α / pCDNA4, an expression plasmid purchased from Addgene (Cambridge, USA) 2 μg of pCDNA4 plasmid was transformed into HK-2 cells by mixing 1: 3 ratio of plasmid and Fugene HD reagent in HK-2 cells using Fugene HD reagent. Twenty-four hours after the mixed solution treatment, the cell line transformed into HK-2 cells was transfected with the Zeocin gene existing in the pCDNA4 vector every 3 days with the antibiotic (200 μg / ml) Cells that survived to 2 weeks after transfection were fractionated and cells were continuously cultured. The obtained Mock and human PGC-1α-overexpressed cell lines were transfected with human PGC-1α / pCDNA4 plasmid at the protein level using Western blot, and c-Myc Expression of human PGC-1α was indirectly confirmed by gene expression. A pair of primers using a sequence in the gene of genomic DNA was also prepared at the gene level and confirmed by PCR (polymerase chain reaction). The results are shown in FIG. 2A. All subsequent examples are experiments using control and HKC cells overexpressing human PGC-1 alpha.

Example 2: Comparison of survival rates according to treatment with hydrogen peroxide (H ₂ O ₂ ) treatment (MTT assay) in control and human PGC-1α overexpressing cell lines.

The decrease in survival rate by treatment with 0.5 mM hydrogen peroxide (H ₂ O ₂ ) was confirmed by MTT assay as follows, and the results are shown in FIG. 96 dispensing a 1 X 10 ^4, the two cell lines to the well plate, and the hydrogen peroxide solution of 0.5 mM after day (H ₂ O ₂₎ for 0, 2, 4, and the final concentration to each process for 8 hours induced apoptosis 0.5 mg / ml MTT, and the amount of MTT-formazan substituted in living cells only was measured with an ELISA reader at an absorbance of 590 nm.

As shown in FIG. 2B, the survival rate of the PGC-1α-overexpressing cell line was increased in comparison with the control group, according to the treatment with hydrogen peroxide (H ₂ O ₂ ).

Example 3: Apoptosis assay by treatment with hydrogen peroxide (H ₂ O ₂ ) treatment in control and human PGC-1α overexpressing cell lines

The number of cells in the cell death process according to the treatment with 0.5 mM hydrogen peroxide (H ₂ O ₂ ) was quantified as follows, and the results are shown in FIG. The number of apoptosis and necrosis cells stained with Annexin V was measured using Annexin V-FITC Apoptosis Detection Kit (KOMA BIOTECH).

Example 4: Comparison of apoptosis by treatment with hydrogen peroxide (H ₂ O ₂ ) treatment (DAPI staining method) in control and human PGC-1α overexpressing cell lines

When cell apoptosis occurs, condensation and cleavage of DNA occurs in the nucleus. The change in nuclei that underwent apoptosis under fluorescence microscope is analyzed by using DAPI (4 ', 6-diaidino-2-phenylindole dihydrochloride) And the results are shown in Fig. 2D. 4 in the well slide was dispensed into ^{5 X 10 4 / well, 0.5} mM hydrogen peroxide (H ₂ O ₂₎ was observed for cell death according to the degree of processing for 4 hours in such condensation of the nucleus.

In FIG. 2, arrows indicate changes in the shape of the nucleus due to apoptosis. As a result, it was observed that the shape of the nucleus was less changed in the cell line overexpressing human PGC-1α than the control group.

Example 5: Comparison of the amount of reactive oxygen species (ROS) according to the treatment with hydrogen peroxide (H ₂ O ₂ ) in the control and human PGC-1α overexpressing cell line.

(ROS) in cells using 5,6-chloromethyl-2'7'-dicholorodihydrofluorescein diacetate (CM-H2DCFDA), which selectively reacts with hydrogen peroxide (H ₂ O ₂ ) The amount was observed through a fluorescence microscope and the results are shown in Fig. The results shown in FIG. 3 were obtained by dividing 5 × 10 ⁴ / well on a 4-well slide, treating with 0.5 mM H ₂ O ₂ for 30 minutes, reacting with 10 μM CM-H2DCFDA for 30 minutes, As a result, it was observed that the amount of reactive oxygen species (ROS) in the cell line overexpressing human PGC-1α was smaller than that in the control group.

Example 6: Changes in the activity and protein amount of apoptosis-related proteins (Western blot) by treatment with hydrogen peroxide (H ₂ O ₂ ) treatment in control and human PGC-1α overexpressing cell lines.

After incubation with 0.5 mM hydrogen peroxide (H ₂ O ₂ ) for 6 hours, apoptosis was induced, and the phosphorylation of p53, a protein involved in apoptosis, and the increase rate of cleaved caspase 3 / caspase 3 were measured by Western blot And the results are shown in FIG. 4A to FIG. 4E.

As shown in FIG. 4A to FIG. 4E, it was confirmed that the increase of these proteins was prominent in the control group, but remarkably decreased in the cell line overexpressing human PGC-1α. In other words, Cytochrome C present in mitochondria migrates from mitochondria to cytoplasm when apoptotic process occurs. Western blot analysis of changes in mitochondria and cytoplasmic proteins showed that mitochondria-to-cytoplasmic migration was more frequent in the control group In the cell line overexpressing human PGC-1 ?. In addition, it was confirmed that there was no abnormality in the fractionation experiment as a protein existing only in mitochondria (complex V α) and cytoplasm (GAPDH).

Example 7

In the cell line overexpressing human PGC-1 alpha, the change in apoptosis-related protein and the amount of active oxygen according to the selective decrease of Nrf-2 was observed through the knock-down experiment of Nrf-2 as follows. Respectively.

To elucidate that the cytoprotective effect of Nrf-2 on human PGC-1α-overexpressed cell lines is due to the increase of Nrf-2 protein, Nrf-2 specific siRNAs were treated with human PGC-1α- Induced a decrease in expression. Nrf-2 specific siRNA was purchased from Santacruz, and 3 × 10 ⁵ cells were seeded in a 60-mm dish. After 24 hours, the cells were injected into the cells using a DhamaFect 1 transfection reagent. After culturing for 72 hours, hydrogen peroxide water (H ₂ O ₂ ) was treated to induce apoptosis. Protein expression was confirmed using Western blot.

As shown in FIG. 5, it was observed that the amount of Cleaved caspase 3 / Caspase 3, which was decreased in association with apoptosis, was induced by selective reduction of Nrf-2 in human PGC-1α overexpressed cell line, It was confirmed that the protective effect of kidney cells on the human PGC-1α overexpressing cell line was an increase in the expression of Nrf-2. In addition, the amount of reactive oxygen (ROS) in the cell was also measured as in Example 5. As a result, in the cell line overexpressing human PGC-1 alpha, the amount of active oxygen decreased in association with apoptosis Respectively.

Example 8

The mechanism of protein increase of Nrf-2 in human PGC-1α overexpressed cell line was confirmed as follows, and the results are shown in FIGS. 6a to 6d.

In order to confirm the mechanism of the protective function of the kidney cells according to the increase of Nrf-2 in the human PGC-1α overexpressed cell line, the control cell and the cell line overexpressing human PGC-1α were dispensed into a 60 mm dish at 3.5 × 10 ⁵ cells . After incubation for 24 hours, the cells were treated with hydrogen peroxide (H ₂ O ₂ ) at 0, 0.5, 1, 3, and 6 hours for 16 hours after the cells were exchanged with FBS-free culture medium to obtain Keap1, GSK3β Were confirmed by western blotting. As shown in FIG. 6A, the amount of phosphorylation of the 9th residue of serine, which is an inactivated form of GSK3?, Increased from 1 hour to 6 hours of hydrogen peroxide (H ₂ O ₂ ) treatment of human PGC-1α overexpressed cell line .

In order to confirm the mechanism of the protective function of the kidney cells according to the increase of Nrf-2 in the human PGC-1α overexpressed cell line, the control cell and the cell line overexpressing human PGC-1α were dispensed into a 60 mm dish at 3.5 × 10 ⁵ cells . After incubation for 24 hours, the cells were treated with hydrogen peroxide (H ₂ O ₂ ) at 0, 15, 30, 60, and 120 min intervals for 16 hours after the medium was replaced with FBS-free culture medium. MAPKs (ERK1 / 2, p-38, phosphorylation of JNK) was confirmed by Western blotting. As shown in FIG. 6B, the degree of phosphorylation of p38 during the activation of MAPKs (ERK1 / 2, p38, JNK phosphorylation) was significantly increased in Western blot analysis in human PGC-1α overexpressed cell line .

6a and 6b show the increase of Nrf-2 in the human PGC-1α overexpressed cell line, which increases GSK3β inactivation (p-GSK3β Ser9; phosphorylation of the 9th residue of serine) and phosphorylation of p38 (5 μM) to inhibit phosphorylation of p38, which is considered to be the highest signal, in human PGC-1α-overexpressed cell line to confirm whether the two signals are sequential through one pathway, Lt; / RTI > Western blot analysis confirmed the p38-specific response by treating the MAPKs with the ERK1 / 2 inhibitor (PD98059, 50 μM). As shown in FIG. 6C, the inactivation of GSK3β was decreased and the amount of Nrf-2 decreased, as the phosphorylation inhibitor of p38, which was increased in human PGC-1α overexpressed cell line, And the amount of the protein, hemooxygenase 1 (HO-1), was also decreased. In addition, as in Example 2, the survival rate after treatment with the p38 activity inhibitor was compared in the cell line overexpressing human PGC- As a result, it was observed that the cytoprotective effect of human PGC-1α-overexpressed cell line was inhibited, as shown in FIG. 6d.

These results demonstrate that PGC-1α is overexpressed and that the increase of PGC-1α protein in renal tubular cells effectively suppresses apoptosis induced by oxidative stress in kidney cells. The inhibitory effect of p38 / GSK3β / Nrf-2 axis The molecular mechanism through a signal transduction pathway that leads to the expression of various genes involved in cell protection, which is followed by an increase in the phosphorylation of p38, an increase in GSK3? Inactivation and an increase in the protein of Nrf-2 Respectively.

Therefore, the pharmaceutical composition comprising the PGC-1 alpha gene mRNA or the PGC-1 alpha protein itself or a drug that induces overexpression of PGC-1 alpha in the body can inhibit cell death due to oxidative stress in the kidney cells , And can be used as a pharmaceutical composition and a functional food material for prevention and symptom improvement of acute kidney disease due to lowering of kidney function, which is a side effect of acute kidney disease, especially various anticancer drugs.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is clearly understood that the same is by way of illustration and example only and is not to be taken by way of limitation, Various changes and modifications will be possible.

Claims (5)

MRNA of a gene coding for PGC-1 alpha (Peroxisome proliferator activated receptor gamma coactivator 1 alpha), a drug inducing overexpression of PGC-1 alpha protein and PGC-1 alpha protein,
The drug for inducing overexpression of PGC-1 alpha protein is at least one selected from the group consisting of metformin, phenformin HCl, AICAR, A-769662, GSK621, Rosiglitazone, Rosiglitazone maleate and Pioglitazone. Composition.
The method according to claim 1,
The active ingredient increases phosphorylation of p38 and increases the phosphorylation of p38 to increase the inactivation of GSK3? (Glycogen synthase kinase 3 beta). When the inactivation of GSK3? Is increased, Nrf-2 (Nuclear factor-erythroid- 2-related factor 2) protein expression is increased.
A method for preventing acute renal disease comprising administering the pharmaceutical composition of claim 1 or 2 to a mammal other than a human.
1 & cir & and a drug that induces overexpression of PGC-1 alpha protein (Peroxisome proliferator activated receptor gamma coactivator 1 alpha) mRNA, PGC-1 alpha protein and PGC- Wherein the drug inducing overexpression is at least one selected from the group consisting of metformin, phenformin HCl, AICAR, A-769662, GSK621, Rosiglitazone, Rosiglitazone maleate, and Pioglitazone.
5. The method of claim 4,
The active ingredient increases phosphorylation of p38 and increases the phosphorylation of p38 to increase the inactivation of GSK3? (Glycogen synthase kinase 3 beta). When the inactivation of GSK3? Is increased, Nrf-2 (Nuclear factor-erythroid- 2-related factor 2) Protein expression is increased, and a health functional food for acute kidney disease prevention.

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