CN114831998B

CN114831998B - Application of tramadol acid in treating kidney disease

Info

Publication number: CN114831998B
Application number: CN202210464141.3A
Authority: CN
Inventors: 李亚峰; 李荣山; 段琦; 田玲玲; 李莉红; 冯嘉炳; 平鑫博
Original assignee: Shanxi Provincial Peoples Hospital
Current assignee: Shanxi Provincial Peoples Hospital
Priority date: 2022-04-28
Filing date: 2022-04-28
Publication date: 2023-09-22
Anticipated expiration: 2042-04-28
Also published as: CN114831998A

Abstract

The application relates to the field of disease treatment, in particular to application of tramadol acid in kidney disease treatment.

Description

Application of tramadol acid in treating kidney disease

Technical Field

The application belongs to the field of disease treatment, and particularly relates to application of tramadol acid in treatment of kidney diseases (especially diabetic nephropathy).

Background

Diabetic nephropathy (Diabetic nephropathy, DN) is a progressive decrease in proteinuria and Glomerular Filtration Rate (GFR) due to prolonged diabetes mellitus, a fatal complication of diabetes mellitus, characterized by thickening of basement membrane, glomerulosclerosis, tubular interstitial fibrosis and urinary albumin. The incidence of DN in China is on the rise, and the DN becomes the second cause of end-stage renal disease at present, and is next to various glomerulonephritis. DN has no specific treatment. Current methods of treating DN include tightly controlling blood glucose, lowering blood pressure, and reducing proteinuria using renin-aldosterone system blockers, which, while effective, many patients continue to promote progression to end stage renal disease by oral administration of hypoglycemic agents in combination with angiotensin-converting enzyme inhibitors or angiotensin receptor blockers, with limited efficacy and some side effects, and no significant improvement in patient survival. Therefore, the development of novel drugs has great significance for the treatment of DN.

The natural traditional Chinese medicine and the active ingredients thereof gradually become a new strategy for treating various diseases due to the advantages of high efficiency, small side effect, low cost and the like. The tramadol acid (Trametenolic acid, TA) is the main component of lanostane type tetracyclic triterpene compounds, and has effects of cytotoxicity including tumor cells, enzyme inhibition activity, antimicrobial activity, blood glucose lowering effect, antiviral effect, and gastric ulcer resistance. In addition, TA also has a certain scavenging effect on oxygen radicals. It has been found that TA shows potent anti-cancer and anti-inflammatory activity in human prostate and breast cancer, but TA has not found application in DN.

In view of this, the present application has been proposed.

Disclosure of Invention

In order to solve the technical problems, the application provides that the tramadol acid can be used for treating kidney diseases (especially diabetic nephropathy) for the first time. The application is proved by a series of experiments that the tramadol acid can protect kidney injury caused by kidney disease through antioxidant and anti-inflammatory ways.

It is therefore a first object of the present application to provide the use of tramadol acid in the treatment of kidney disease (especially diabetic nephropathy) and its use in the manufacture of a medicament for the treatment of diabetic nephropathy;

a second object of the present application is to provide a pharmaceutical composition for the treatment of kidney disease, in particular diabetic nephropathy.

In particular, the present application first provides the use of tramadol acid in the manufacture of a medicament for the prevention and/or treatment of a disease or condition, or for reducing the risk of a disease or condition.

Further, the disease is kidney disease;

further, the kidney disease includes diabetic nephropathy;

further, the prevention and/or treatment is manifested by an enhancement of the antioxidant capacity of the subject;

further, the preventing and/or treating is manifested as inhibiting an inflammatory response in the subject;

further, the prevention and/or treatment is manifested as enhanced protection of the foot cells of the subject from injury;

further, the prevention and/or treatment is manifested as inhibiting renal fibrosis in the subject.

The application also provides a pharmaceutical composition comprising the active ingredient tramadol acid.

Further, the pharmaceutical composition further comprises optionally a pharmaceutically acceptable excipient, carrier and/or diluent.

Further, the pharmaceutical composition is used for preventing and/or treating kidney diseases;

further, the kidney disease includes diabetic nephropathy;

The present application also provides a method of preventing or treating a disease, disorder or syndrome, comprising administering to a subject in need thereof an effective amount of tramadol or a pharmaceutical composition as described above.

Further, the medicament or the pharmaceutical composition described above is administered to the subject subcutaneously, intravenously, orally, rectally or intraperitoneally.

The application also provides a method for enhancing antioxidant capacity, inhibiting inflammatory response, enhancing kidney foot cell damage protection and/or inhibiting fibrosis in a cell, tissue or subject comprising administering to a kidney cell, tissue or subject with a kidney disease an effective amount of tramadol acid or a pharmaceutical composition as described above.

Further, the subject is a subject suffering from kidney disease, preferably a subject suffering from diabetic nephropathy.

Further, the oxidative stress, inflammatory reaction, injury to renal foot cells, fibrosis, etc. are all caused by kidney disease.

Further, the cells are kidney cells and the tissue is kidney tissue; preferably, the cells and tissues are ex vivo.

The application also provides application of the tramadol acid in preparing a kidney disease animal model, in particular application in a diabetic kidney disease animal model.

The application also provides the use of tramadol acid in the manufacture of a medicament or agent for enhancing antioxidant capacity, inhibiting inflammatory response, enhancing kidney foot cell damage protection, and/or inhibiting fibrosis in a cell, tissue or subject.

Further, the subject includes human or non-human animals (including mammals), such as humans, non-human primates (apes, gibbons, gorillas, chimpanzees, orangutans, macaques), domestic animals (dogs and cats), farm animals (poultry such as chickens and ducks, horses, cows, goats, sheep, pigs), and laboratory animals (mice, rats, rabbits, guinea pigs). Human subjects include fetal, neonatal, infant, adolescent and adult subjects. Further, the subject also includes an animal disease model.

The beneficial technical effects of the application are as follows:

the application provides the therapeutic application of the tramadol acid to kidney diseases (especially diabetic nephropathy) for the first time, and illustrates the potential mechanism of the tramadol acid in treating the diabetic nephropathy.

Drawings

In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings that are needed in the description of the embodiments or the prior art will be briefly described, and it is obvious that the drawings in the description below are some embodiments of the present application, and other drawings can be obtained according to the drawings without inventive effort for a person skilled in the art.

FIG. 1, right kidney weight/body weight, BUN, scr and urinary albumin level test results comparison chart; data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group and #p < 0.05 compared to DN group;

fig. 2, TA-mitigating DN kidney histopathological change profile: (A) HE, PAS, masson representative micrograph of staining at 400× magnification. (B) Glomerular membrane matrix expansion and collagen deposition quantitative analysis; data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group and #p < 0.05 compared to DN group.

Fig. 3, comparison of TA oxidation stress results for DN reduction: after TA treatment, GSH (A) content is increased, SOD (C) and CAT (D) activities are increased, and MDA (B) content is reduced. Data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group and #p < 0.05 compared to DN group.

FIGS. 4, TA graphs of oxidative stress results for alleviating DN by activating the Nrf2/HO-1 signal path: (A) Representative western blot plots of Nrf2, HO-1 and NQO-1 protein expression; (B) quantitative analysis of Nrf2, HO-1 and NQO-1 protein expression. Data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group; the group #P < 0.05 was compared with the DN group.

FIG. 5A is a graph of the results of reducing the inflammatory response of DN by inhibiting NF- κB signaling: (A) Representative western blot results of NF- κ B, P-NF- κ B, TNF- α, IL-1β and IL-6 protein expression; (B) Quantitative analysis of P-NF- κB/NF- κ B, TNF- α, IL-1β and IL-6 protein expression. Data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group and #p < 0.05 compared to DN group.

Fig. 6, TA vs DN podocyte injury protection results chart: (A) Representative western blot results of Nephrin and Podocin protein expression; (B) quantitative analysis of Nephrin and Podocin protein expression. Data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group and #p < 0.05 compared to DN group.

Fig. 7, TA reduced DN kidney fibrosis result graph: (A) CoL-III and FN immunohistochemical staining, magnification: 400×; (B) semi-quantitative analysis of COL-III and FN expression. Data are expressed as mean ± standard deviation (n=10), P < 0.05 compared to NC group and #p < 0.05 compared to DN group.

Detailed Description

The following description of the embodiments of the present application will be made apparent and fully in view of the accompanying drawings, in which some, but not all embodiments of the application are shown. All other embodiments, which can be made by those skilled in the art based on the embodiments of the application without making any inventive effort, are intended to be within the scope of the application.

Partial term definition

Unless defined otherwise hereinafter, all technical and scientific terms used in the detailed description of the application are intended to be identical to what is commonly understood by one of ordinary skill in the art. While the following terms are believed to be well understood by those skilled in the art, the following definitions are set forth to better explain the present application.

As used herein, the terms "comprising," "including," "having," "containing," or "involving" are inclusive or open-ended and do not exclude additional unrecited elements or method steps. The term "consisting of …" is considered to be a preferred embodiment of the term "comprising". If a certain group is defined below to contain at least a certain number of embodiments, this should also be understood to disclose a group that preferably consists of only these embodiments.

The indefinite or definite article "a" or "an" when used in reference to a singular noun includes a plural of that noun.

The terms "about" and "substantially" in this application mean the range of accuracy that one skilled in the art can understand yet still guarantee the technical effect of the features in question. The term generally means a deviation of + -10%, preferably + -5%, from the indicated value.

Furthermore, the terms first, second, third, (a), (b), (c), and the like in the description and in the claims, are used for distinguishing between similar elements and not necessarily for describing a sequential or chronological order. It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the application described herein are capable of operation in other sequences than described or illustrated herein.

Use of tramadol of the application in the manufacture of a medicament for the prevention and/or treatment of a disease or condition, or for reducing the risk of a disease or condition.

In some embodiments, the disease described herein is kidney disease.

The Tramadol Acid (TA), "English name: trametenolic acid, chinese alias: 3-hydroxy lanosta-8, 24-diene-21-acid, is the main component of lanostane type tetracyclic triterpene compound, and has effects of cytotoxic activity including tumor cells, enzyme inhibition activity, antimicrobial activity, hypoglycemic effect, antivirus, anti-gastric ulcer and the like.

In the present application, the term "disease" or "disorder" is used interchangeably and generally refers to any deviation of a subject from a normal state, such as any change in the state of the body or certain organs, which impedes or disrupts performance of the function and/or causes symptoms such as discomfort, dysfunction, pain or even death in a person suffering from or in contact with the disease. The disease or disorder may also be referred to as a disorder (disorder), an malaise (ailment), a malady (ailment), a disorder (disorder), a disease (hickness), an illness (illness), a physical malaise (compatibility), an index of disorder, or an affection.

In the present application, the term "preventing and/or treating" includes not only preventing and/or treating a disease, but also generally includes preventing the onset of a disease, slowing or reversing the progression of a disease, preventing or slowing the onset of one or more symptoms associated with a disease, reducing and/or alleviating one or more symptoms associated with a disease, reducing the severity and/or duration of a disease and/or any symptoms associated therewith and/or preventing further increases in the severity of a disease and/or any symptoms associated therewith, preventing, reducing or reversing any physiological damage caused by a disease, and any pharmacological effects that would normally be beneficial to the patient being treated. The RNA inhibitors or pharmaceutical compositions of the application form viable therapeutic agents without the need to achieve complete cure or eradication of any symptoms or manifestations of the disease. As recognized in the relevant art, drugs used as therapeutic agents may reduce the severity of a given disease state, but need not eliminate every manifestation of the disease to be considered useful therapeutic agents. Similarly, a prophylactically administered treatment constitutes a viable prophylactic agent and need not be completely effective in preventing the onset of the condition. It may be sufficient to simply reduce the impact of the disease in the subject (e.g., by reducing the number or severity of its symptoms, or by increasing the effectiveness of another treatment, or by producing another beneficial effect), or to reduce the likelihood of disease occurrence or exacerbation.

The term "kidney disease" as used herein refers to damage to kidney structure and function caused by various causes, and may include, but is not limited to, primary glomerular diseases such as acute nephritis, chronic nephritis, latent nephritis, nephrotic syndrome, etc., secondary glomerular diseases such as lupus nephritis, purpura nephritis, hepatitis B virus associated nephritis, diabetic nephropathy, etc., hereditary kidney diseases such as polycystic kidney, thin basement membrane kidney disease, etc., interstitial nephritis such as acute interstitial nephritis, chronic interstitial nephritis, etc.

In some embodiments, the kidney disease of the present application includes, inter alia, diabetic Nephropathy (DN).

The "diabetic nephropathy" described in the present application is progressive decrease in proteinuria and Glomerular Filtration Rate (GFR) due to prolonged diabetes mellitus, and is a fatal complication of diabetes mellitus, characterized by thickening of basement membrane, glomerulosclerosis, tubular interstitial fibrosis and urinary albumin.

In some embodiments, the prophylaxis and/or treatment of the application is manifested as an enhancement of the antioxidant capacity of a subject;

in some embodiments, the prevention and/or treatment of the application is manifested by inhibiting an inflammatory response in a subject, particularly an inflammatory response caused by kidney disease;

in some embodiments, the prevention and/or treatment of the application is manifested by enhanced protection of the subject from podocyte injury, particularly from kidney disease;

in some embodiments, the prevention and/or treatment of the application is manifested as inhibiting renal fibrosis in a subject, particularly renal fibrosis caused by kidney disease.

The application also discloses a pharmaceutical composition which comprises the active ingredient of the tramadol acid, and the pharmaceutical composition comprising the tramadol acid is used for preventing and/or treating kidney diseases.

In some embodiments, the kidney disease of the present application includes, inter alia, diabetic nephropathy.

In the present application, the term "subject" generally refers to a human or non-human animal (including mammals) in need of diagnosis, prognosis, amelioration, prevention and/or treatment of a disease, such as humans, non-human primates (apes, gibbons, gorillas, chimpanzees, gorillas, macaques), domestic animals (dogs and cats), farm animals (poultry such as chickens and ducks, horses, cattle, goats, sheep, pigs) and laboratory animals (mice, rats, rabbits, guinea pigs). Human subjects include fetal, neonatal, infant, adolescent and adult subjects. The subject includes an animal disease model. In some embodiments, the subject of the application is particularly a kidney disease subject, and in other specific embodiments, the subject of the application is particularly a diabetic kidney disease subject.

In some embodiments, the subject of the application is a human or a mouse.

In some embodiments, the pharmaceutical compositions of the present application further comprise optionally a pharmaceutically acceptable excipient, carrier, and/or diluent.

In the present application, the term "pharmaceutically acceptable" generally refers to one or more non-toxic substances that do not interfere with the effectiveness of the biological activity of the active ingredient. Such formulations may generally contain salts, excipients, buffers, preservatives, compatible carriers and optionally other therapeutic agents. Such pharmaceutically acceptable formulations may also generally comprise compatible solid or liquid fillers, diluents or encapsulating materials suitable for administration to humans. For pharmaceutical use, the salt should be a pharmaceutically acceptable salt, but non-pharmaceutically acceptable salts may be conveniently used to prepare pharmaceutically acceptable salts, and they are not excluded from the scope of the present application. Such pharmacologically and pharmaceutically acceptable salts include, but are not limited to, salts prepared from the following acids: hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, maleic acid, acetic acid, salicylic acid, citric acid, boric acid, formic acid, malonic acid, succinic acid, and the like. Pharmaceutically acceptable salts may also be prepared as alkali metal salts or alkaline earth metal salts, such as sodium, potassium or calcium salts.

The term "acceptable excipients, carriers, and/or diluents" refers to excipients, carriers, and/or diluents that are pharmacologically and/or physiologically compatible with the subject and active agent, are well known in the art (see, e.g., remington's Pharmaceutical sciences. Mediated by Gennaro AR,19th ed.Pennsylvania:Mack Publishing Company,1995), and include, but are not limited to, pH adjusters, surfactants, adjuvants, and ionic strength enhancers, and the like. For example, pH modifiers include, but are not limited to, phosphate buffers; surfactants include, but are not limited to, cationic, anionic or nonionic surfactants, such as Tween-80; ionic strength enhancers include, but are not limited to, sodium chloride.

In the present application, the term "administration" generally refers to the introduction of the pharmaceutical formulation of the application into the body of a subject by any route of introduction or delivery. Any method known to those skilled in the art for contacting cells, organs or tissues with the drug may be used. The administration may include, but is not limited to, intravenous, intra-arterial, intranasal, intra-abdominal, intramuscular, subcutaneous transdermal, or oral. The daily dose may be divided into one, two or more doses of a suitable form to be administered at one, two or more times during a certain period of time. In some embodiments, the medicament or the pharmaceutical composition described above is administered to a subject subcutaneously, intravenously, orally, or intraperitoneally.

In the present application, the term "effective amount" or "effective dose" generally refers to an amount sufficient to achieve or at least partially achieve the desired effect. A "therapeutically effective amount" or "therapeutically effective dose" of a drug or therapeutic agent is generally any amount of drug that, when used alone or in combination with another therapeutic agent, promotes regression of the disease (as evidenced by a decrease in severity of symptoms of the disease, an increase in the frequency and duration of disease asymptomatic periods, or prevention of damage or disability due to the disease). "prophylactically effective amount" or "prophylactically effective dose" of a drug generally refers to an amount of a drug that inhibits the progression or recurrence of a disease when administered alone or in combination with another therapeutic agent to a subject at risk of disease progression or recurrence. The ability of a therapeutic or prophylactic agent to promote regression of a disease or inhibit the progression or recurrence of a disease can be assessed using a variety of methods known to those of skill in the art, such as in a human subject during a clinical trial, in an animal model system to predict efficacy in humans, or by assaying the activity of the agent in an in vitro assay. In certain embodiments, an "effective amount" refers to an amount of tramadol that produces a desired pharmacological, therapeutic or prophylactic result.

The application also provides a method for enhancing antioxidant capacity, inhibiting inflammatory response, enhancing kidney foot cell damage protection and/or inhibiting fibrosis in a cell, tissue or subject comprising administering to a kidney cell, tissue or subject with a kidney disease an effective amount of tramadol acid or a pharmaceutical composition as described above. It will be appreciated that the procedure may be performed in vivo or in vitro with respect to cells or tissues.

In some embodiments, the cells of the application are kidney cells and the tissue is kidney tissue; in some preferred embodiments, the kidney cells or kidney tissue are in an ex vivo state.

In some embodiments, the subject of the application is a subject with kidney disease, preferably a subject with diabetic nephropathy.

The application also provides application of the tramadol acid in preparing a kidney disease animal model, in particular application in a diabetic kidney disease animal model. It is understood that upon determining that tramadol can be used to treat kidney disease, those skilled in the art can prepare or assist in preparing animal models of kidney disease accordingly.

Examples

The application is further described by the accompanying drawings and the following examples, which are provided to illustrate specific embodiments of the application and are not to be construed as limiting the scope of the application in any way.

Experimental materials and packet processing

Experimental materials: SPF grade 8 week old male C57BLKS/db (db/db) mice 20 and C57BL/6 mice 10 purchased from Guangdong gold and Biotech Co., ltd; the Trametes Acid (TA) is extracted from Inonotus obliquus with purity of above 99%.

The experiments were divided into three groups: :

group 1 same week-old C57BL/6 mice as normal control group (n=10);

group 2 diabetic nephropathy DN group (n=10);

group 3 diabetic nephropathy dn+tramadol (n=10);

wherein groups 2 and 3 were randomly divided into two groups by db/db mice, group 3 was intraperitoneally injected with tramadol (10 mg/kg/d) from dn+tramadol mice for 4 weeks; normal control and DN mice were intraperitoneally injected with normal saline for 4 weeks.

Example 1, renal weight/body weight determination and Biochemical index detection of mice after TA administration

Taking into account that the right kidney weight/body weight, BUN, scr and urinary albumin levels of the mice can reflect the kidney function of the mice, the metabolism cage of this example collects urine 24h before mice are sacrificed and detects the urine albumin amount of the 24h mice according to the commercial kit instructions; each mouse weighed the weight and right kidney weight, and calculated right kidney weight/body weight; serum samples were collected and assayed for BUN, scr and urinary albumin levels in mouse serum following the kit instructions.

The results are shown in FIG. 1, with DN group mice, the right kidney weight/body weight, BUN, scr and urinary albumin levels were all significantly increased (P < 0.05). While the right kidney weight/body weight, BUN, scr and urinary albumin content were significantly reduced (P < 0.05) in the TA group. The above data indicate that administration of TA significantly improves kidney function in DN mice.

Example 2 histological study of mice after TA administration

This example further demonstrates the role of TA in the treatment of kidney disease by providing a number of aspects of histological changes, oxidative stress, inflammatory response, podocyte protection, and kidney fibrosis.

The present example sacrifices mice, kidney tissue was isolated, 4% paraformaldehyde fixed, paraffin embedded, and 3 μm thick kidney tissue sections were cut. Sections were subjected to hematoxylin-eosin staining (HE), periodic acid schiff staining (PAS) and Masson staining. Randomly select 20 fields, observe kidney pathology and take images of kidney tissue, which are analyzed using Image-Pro Plus 6.0 software to assess glomerulosclerosis and the extent of tubular interstitial damage.

Protein extraction and Western blot: the total protein of the kidney of the mice is extracted by taking 1/3 kidney of the mice and adding 200 mu L of RIPA lysate. Equal amounts of proteins were separated by SDS-PAGE, wet-transfer, 5% skim milk blocked for 1h, nrf2 (1:1000), HO-1 (1:1000), NQO-1 (1:1000), NF- κB (1:1000), P-NF- κB (1:1000), nephrin (1:1000), podocin (1:1000), TNF- α (1:1000), IL-1β (1:1000), IL-6 (1:1000) and GAPDH (1:1000) antibodies, overnight at 4 ℃, TBST wash (3 times, 5min each), secondary antibody incubated for 1h at room temperature, TBST wash (3 times, 5min each), electrochemiluminescence developed, and finally Image J software was used for grey value analysis after photographing with a fully automated gel imaging system.

Immunohistochemical study: kidney tissue sections were immunohistochemically stained for collagen type iii (COL-iii) and Fibronectin (FN), respectively. 20 fields were randomly selected and the relative expression levels of COL-III and FN proteins were measured using Image-Pro Plus 6.0 software.

Statistical analysis: statistical processing and analysis of the data was performed using SPSS 19.0 software. The measurement data conforming to normal distribution is expressed in terms of mean ± standard deviation, and the comparison among multiple groups adopts single factor analysis of variance. P < 0.05 is statistically significant for the differences.

The specific experimental results are as follows:

(1) TA administration can alleviate histological changes in the kidney of DN mice

As can be seen from fig. 2, (a) a representative micrograph of HE, PAS, masson staining; (B) Quantitative analysis of mesangial matrix expansion and collagen deposition were significantly enhanced (P < 0.05) in group DN mice. After TA administration, the glomerular mesangial matrix expansion and collagen deposition in mice was significantly reduced (P < 0.05). The above results indicate that TA administration can alleviate kidney histological changes in DN mice.

(2) TA administration improves DN mice oxidative stress

Oxidative stress is one of the major risk factors involved in the pathogenesis of diabetic nephropathy. To investigate the effect of TA on DN oxidative stress, GSH and MDA contents and SOD and CAT activities were examined in this example. Compared with NC group, DN mice have obviously reduced SOD activity and obviously increased MDA content (P < 0.05), which indicates that the increased lipid peroxide level of DN mice leads to oxidative damage; while GSH content and CAT activity were significantly reduced (P < 0.05). After TA administration, GSH levels, SOD and CAT activities were significantly increased and MDA content was significantly decreased (P < 0.05). These data indicate that TA administration significantly reduced lipid peroxidation levels in mice and enhanced antioxidant capacity in mice (see in particular fig. 3).

(3) TA can relieve oxidative stress of DN by activating Nrf2/HO-1 signal path

In order to further study the mechanism of TA administration for relieving DN oxidative stress, the application adopts a western blot method to detect the expression of Nrf2/HO-1 channel key proteins. As a result, as shown in FIG. 4, DN groups showed significantly reduced expression of Nrf2, HO-1 and NQO-1 proteins (P < 0.05) compared to NC groups. In contrast, expression of Nrf2, HO-1, NQO-1 proteins (P < 0.05) was significantly improved after TA administration. The above results suggest that TA reduces oxidative stress by activating the Nrf2/HO-1 signaling pathway in DN.

(4) TA reduces inflammatory response to DN by inhibiting NF- κB signaling pathway

In order to investigate the effect of TA on DN inflammatory response and its mechanism, protein expression of NF-kappa B, P-NF-kappa B and downstream pro-inflammatory factors TNF-alpha, IL-1 beta and IL-6 (P < 0.05) was examined by western blot. As shown in FIG. 5, TA administration significantly reduced protein expression of P-NF- κB/NF- κB, pro-inflammatory factors TNF- α, IL-1β and IL-6 (P < 0.05). These results indicate that TA is capable of reducing the inflammatory response of DN and that TA reduces the expression of pro-inflammatory factors by inhibiting NF- κb signaling pathways.

(5) TA administration has protective effect on DN podocyte injury

(1) Podocytes are an important component of the glomerular filtration barrier in kidney disease, and Nephrin and Podocin together maintain the normal morphology and function of podocytes, thereby maintaining the normal glomerular filtration barrier function. This example evaluates the effect of TA dosing on DN podocyte damage by evaluating Nephrin and Podocin protein levels. As can be seen from fig. 6, the expression levels of Nephrin and Podocin proteins were significantly down-regulated in DN group (P < 0.05) compared to NC group; whereas Nephrin and Podocin expression levels were significantly elevated (P < 0.05) in tissues of mice treated with TA. The data show that TA administration treatment has obvious protection effect on DN podocyte injury.

(2) TA administration reduces DN renal fibrosis

collagen-III (COL-III) is one of the major components of the extracellular matrix (ECM) of kidney tissue, and during the progression of fibrosis of kidney tissue, the accumulation of ECM is mainly composed of abnormally expressed type III collagen and type IV collagen; fibronectin (FN) is a non-collagenous macromolecular glycoprotein, and alterations in FN expression are closely related to a number of pathological events, including fibrosis. To investigate the effect of TA on renal fibrosis, the present application used immunohistochemical methods to detect expression levels of COL-III and FN. FIG. 7 shows that DN group COL-III and FN levels are significantly higher than NC group (P < 0.05); the expression level of COL-III and FN is obviously reduced (P is less than 0.05) after TA administration, which shows that TA administration can effectively inhibit the expression of COL-III and FN, and further has anti-fibrosis effect on DN mice.

In view of the test results, TA administration can have an effective therapeutic effect on diabetic nephropathy.

The foregoing descriptions of specific exemplary embodiments of the present application are presented for purposes of illustration and description. It is not intended to limit the application to the precise form disclosed, and obviously many modifications and variations are possible in light of the above teaching. The exemplary embodiments were chosen and described in order to explain the specific principles of the application and its practical application to thereby enable one skilled in the art to make and utilize the application in various exemplary embodiments and with various modifications as are suited to the particular use contemplated. It is intended that the scope of the application be defined by the claims and their equivalents.

Claims

1. Use of tramadol acid in the manufacture of a medicament for the prevention and/or treatment of a disease, or for reducing the risk of a disease; the disease is diabetic nephropathy.

2. The use of claim 1, wherein the prevention and/or treatment is manifested by enhancing antioxidant capacity of the subject, inhibiting inflammatory response of the subject, enhancing podocyte injury protection of the subject, and/or inhibiting renal fibrosis of the subject.

3. The use according to any one of claims 1-2, wherein the subject is a human or non-human animal.