Application of dexfoserine in preparation of medicine for preventing and / or treating renal injury

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to application of dexfoserine in preparation of a medicine for preventing and / or treating renal injury, the dexfoserine can relieve a glycerol-induced acute renal injury mouse model, obviously reduce the creatinine and urea nitrogen level in serum of the acute renal injury mouse, and improve the renal injury prevention and / or treatment effect. And the traditional Chinese medicine composition has the effect of treating the acute kidney injury and has a wide application prospect.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and in particular relates to the application of dextrophosphorylation in the preparation of a drug for preventing and / or treating kidney damage. Background Art

[0002] Acute kidney injury (AKI) refers to a clinical syndrome caused by a rapid decline in renal function due to a variety of causes. It can occur in people without previous kidney disease or in patients with chronic kidney disease. About 5% of hospitalized patients may suffer from AKI, and its incidence in intensive care units is as high as 30%. Although the nephrology community has paid more and more attention to AKI, there is still no specific treatment strategy, and the mortality rate is high. It is an acute and critical disease in kidney disease. Therefore, it is of great clinical significance to find potential drugs for the treatment of AKI. Glycerol, also known as propylene glycol, is a fat-soluble small molecule. After injection of glycerol, it can cause muscle lysis and hemolysis in the body, releasing a large amount of hemoglobin and myoglobin. Myoglobin can cause renal vasoconstriction, and the glomerular afferent arterioles are all contracted, resulting in a decrease in renal blood flow and glomerular filtration rate, causing renal ischemia and damage to renal tubular function. Because the renal tubules cannot reabsorb, casts are formed to block the renal tubules, causing pathological damage to the renal tubules and interstitium. At the same time, myoglobin and hemoglobin can be decomposed into heme, which has a direct toxic effect on the renal tubules. There are various drugs for acute kidney injury. For example, the invention patent CN113663082B discloses that gold nanoclusters stabilized by acetylcysteine ​​can be used to treat acute kidney injury, but the preparation method of gold nanoclusters is complicated; the invention patent CN109602755A discloses that eleutheroside B has the effect of treating acute kidney injury, but it is used to treat drug-induced acute kidney injury.

[0003] Dexfosfoserine (O-Phospho-L-serine) is a direct precursor in the synthesis of L-serine, an agonist of the third group of mGluR (mGluR4, mGluR6, mGluR7 and mGluR8), and an antagonist of mGluR1 and mGluR2, and is used for nutritional supplementation in the state of excessive fatigue. There are currently no reports on the use of dexfosfoserine in the treatment of acute kidney injury caused by glycerol.

[0004] The inventor unexpectedly discovered during the research process that dextrophosphorylation can significantly reduce the levels of creatinine and urea nitrogen in the serum of mice with acute kidney injury, and can also improve kidney pathological damage. It has the effect of treating acute kidney injury and has broad application prospects. Summary of the invention

[0005] The present invention aims to provide the use of dextrophosphorylation in the preparation of a drug for preventing and / or treating kidney damage, wherein the structural formula of dextrophosphorylation is as shown in formula (I):

[0006]

[0007] Preferably, the renal injury is acute renal injury.

[0008] Preferably, the acute kidney injury is induced by glycerol.

[0009] Preferably, dextrophosphorylation is used as the active compound, and excipients acceptable to pharmaceutical preparations are added to prepare the pharmaceutical preparation.

[0010] Preferably, the auxiliary materials include one or more functional carriers selected from the group consisting of excipients, stabilizers, antioxidants, colorants, diluents, and sustained-release agents.

[0011] Preferably, the pharmaceutical preparation is in the form of any one of tablets, capsules, suspensions, emulsions, powder injections and injections.

[0012] Preferably, the drug for preventing and / or treating kidney injury has one or more of the following functions:

[0013] (1) Reduce serum creatinine levels in mice with acute kidney injury;

[0014] (2) Reduce the level of serum urea nitrogen in mice with acute kidney injury.

[0015] The beneficial effects of the present invention are as follows: the present invention provides the use of dextrophosphorylation in the preparation of a drug for preventing and / or treating kidney injury, wherein the dextrophosphorylation can alleviate a mouse model of acute kidney injury induced by glycerol, significantly reduce the levels of creatinine and urea nitrogen in the serum of mice with acute kidney injury, and can also improve kidney pathological damage, has the effect of treating acute kidney injury, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 Schematic diagram of serum creatinine levels of mice in each group;

[0017] Figure 2 Schematic diagram of serum urea nitrogen levels in mice in each group;

[0018] Figure 3 HE staining results of kidneys of mice in each group;

[0019] Figure 4 The renal tubular injury scores of mice in each group. DETAILED DESCRIPTION

[0020] In order to make the purpose, technical scheme and advantages of the present invention clearer, the following examples further describe the present invention in detail. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention. The experimental methods in the following examples are conventional methods unless otherwise specified. The reagents and materials used in the following examples are commercially available unless otherwise specified.

[0021] In the present invention, the structural formula of the dextrophosphorylation is as follows:

[0022]

[0023] Example 1: Effect of Dexphosphoserine in the Preparation of Drugs for Treating Kidney Injury

[0024] 1. Experimental Reagents and Materials

[0025] Glycerol (99%, G810575, Shanghai McLean Biochemical Technology Co., Ltd.); 4% paraformaldehyde fixative (P885233, Shanghai McLean Biochemical Technology Co., Ltd.); ethanol (E809061, Shanghai McLean Biochemical Technology Co., Ltd.); xylene (X820585, Shanghai McLean Biochemical Technology Co., Ltd.); hematoxylin and eosin (HE) staining kit (G1120, Beijing Solebow Technology Co., Ltd.); creatinine determination kit (C011-2-1, Nanjing Institute of Bioengineering Engineering); urea nitrogen test kit (C013-2-1, Nanjing Institute of Bioengineering Engineering); A total of 24 male C57BL / 6 mice aged 8-12 weeks were provided by the Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, with license number SCXK(Gan)-2020-0002. They were adapted to feeding for one week, given diet and free drinking water, and then divided into groups for experiments.

[0026] Before using glycerol for modeling, dilute 99% glycerol with physiological saline to a 50% glycerol solution in a centrifuge tube, place the centrifuge tube on a shaker, and shake for 15 minutes to mix the solution, which is ready for use.

[0027] 2. Experimental groups and drug administration

[0028] After 1 week of adaptive feeding, 24 male C57BL / 6 mice were randomly divided into a blank control group, a glycerol model group, a low-dose dextrophosphorylation group (Glycerol+60mg / kg PS) group, and a high-dose dextrophosphorylation group (Glycerol+120mg / kg PS), with 6 mice in each group. The specific grouping and administration methods are as follows:

[0029] 1) Control group: Intraperitoneal injection of normal saline, once a day, for 6 days; after the intraperitoneal injection on the 6th day, the mice were deprived of water but not food for 18 hours. The intraperitoneal injection of normal saline continued, and 30 minutes after the injection, half of the dose of normal saline was injected into the muscles of the bilateral hind limbs of the mice at a dose of 10mL / kg. The mice continued to be deprived of water but not food for 8 hours and then were free to eat.

[0030] 2) Glycerol group: intraperitoneal injection of normal saline, once a day, for 6 days; after the intraperitoneal injection on the 6th day, the mice were deprived of water but not food for 18 hours. The intraperitoneal injection of normal saline continued, and 30 minutes after the injection, 50% glycerol solution was injected into the muscles of the bilateral hind limbs of the mice at a dose of 10 mL / kg, half of the dose, and the mice continued to be deprived of water but not food for 8 hours, and then they were free to eat.

[0031] 3) Glycerol + 60mg / kg PS group: 60mg / kg PS was intraperitoneally injected once a day for 6 days; after the intraperitoneal injection on the 6th day, the mice were deprived of water but not food for 18h. 60mg / kg PS was injected intraperitoneally again, and 30min after the injection, half of the dose of 50% glycerol solution at a dose of 10mL / kg was injected into the muscles of the bilateral hind limbs of the mice, and the mice continued to be deprived of water but not food for 8h before being allowed to eat freely.

[0032] 4) Glycerol + 120mg / kg PS group: 120mg / kg PS was intraperitoneally injected once a day for 6 days; after the intraperitoneal injection on the 6th day, the mice were deprived of water but not food for 18h. 120mg / kg PS was continued intraperitoneally, and 30min after the injection, half of the dose of 50% glycerol solution at a dose of 10mL / kg was injected into the muscles of the bilateral hind limbs of the mice, and the mice continued to be deprived of water but not food for 8h before being allowed to eat freely.

[0033] 3. Renal function test

[0034] After the above grouping and administration were completed and the mice were fed for 48 hours, blood was collected from the inner canthus of the eye and placed in a centrifuge tube. The blood was placed at room temperature for 1 hour and centrifuged at 2000g and 4°C for 10 minutes. The supernatant was the serum sample. The creatinine and urea nitrogen content of the mouse serum was analyzed using a creatinine assay kit and a urea nitrogen test kit.

[0035] After the blood was collected, the mice were killed, and the kidneys were obtained and washed with saline 2-3 times to remove the blood. The left kidney was fixed in 4% paraformaldehyde for subsequent HE staining, and the right kidney was wrapped in tin foil and stored in a -80°C refrigerator.

[0036] 4. HE staining of renal tissue

[0037] 1) Fixation: The left kidney of the mouse was placed in 4% paraformaldehyde fixative and fixed for 24 hours.

[0038] 2) Water washing: Rinse the fixed tissue blocks with running water for 4-6 hours until there is no formaldehyde smell.

[0039] 3) Dehydration: Place the washed samples in 50%, 70%, and 80% ethanol solutions for 90 min each, then in 95% ethanol solution for 90 min × 2 times, and finally in 100% ethanol solution for 60-90 min × 3 times.

[0040] 4) Transparency: Place in ethanol:xylene (volume ratio 1:1) solution for 20 minutes; then dewax in xylene I and II solutions for 20 minutes each.

[0041] 5) Wax dipping: It needs to be carried out in a constant temperature box, and the temperature of the constant temperature box is adjusted to 3°C higher than the melting point of paraffin. Paraffin I (45-50°C) 60min; Paraffin II (56-58°C) 60min; Paraffin III (56-58°C) 60min;

[0042] 6) Embedding: Place in paraffin embedding machine and embed for later use.

[0043] 7) Sectioning: Place the embedded wax block in a -20℃ refrigerator for 1 hour and slice it to a thickness of about 5μm.

[0044] 8) Slide spreading: Paraffin sections are heat-scalded in a 45-50℃ water bath and mounted on glass slides pre-treated with poly-lysine.

[0045] 9) Baking: Place the prepared slices in a 45°C oven and bake for 2 to 4 hours.

[0046] 10) Staining: Stain according to the instructions of the hematoxylin and eosin (HE) staining kit and observe the HE staining results. Figure 3 As shown, from left to right are the Control group, Glycerol group, Glycerol + 60 mg / kg PS group, and Glycerol + 120 mg / kg PS group, and from top to bottom are the HE staining results magnified 200 and 400 times.

[0047] 11) The scoring rules for renal tubules are as follows: 0 points for no abnormality; 1 point for obvious dilatation and flattened cells; 1 point for brush border damage; 1 point for cell shedding and necrosis in the tubular lumen but without tubular forms or cell fragments; 2 points for tubular forms; and 2 points for brush border shedding.

[0048] 5. Statistical Analysis

[0049] The experimental data were statistically analyzed using SPSS23.0 software, and the data were expressed as (x±s). One-way ANOVA and LSD-t method were used for pairwise comparisons between groups. P<0.05 was considered statistically significant. All statistical results were plotted using GraphPad Prism 8 software.

[0050] 6. Experimental Results

[0051] Creatinine and urea nitrogen in serum are important indicators for evaluating renal function in the body and are used to detect whether there is kidney damage and the severity of the damage. Figure 1 , Figure 2 The results showed that 60 mg / kg and 120 mg / kg of dextrophosphorylation significantly reduced the levels of creatinine and urea nitrogen in the serum of mice with acute kidney injury, indicating that 60 mg / kg and 120 mg / kg of dextrophosphorylation can alleviate glycerol-induced acute kidney injury.

[0052] The results of HE staining of kidneys of mice in each group are shown in Figure 3 The renal tubular injury scores of mice in each group were shown in Figure 4 As shown, it can be seen that 60 mg / kg of dextrophosphorylation can also effectively improve renal pathological damage, but the effect of 120 mg / kg of dextrophosphorylation is not obvious.

[0053] In summary, the present invention provides the use of dextrophosphorylation in the treatment of renal injury. The dextrophosphorylation can significantly reduce the levels of creatinine and urea nitrogen in the serum of mice with acute renal injury, and can also improve renal pathological damage, has the effect of treating acute renal injury, and has broad application prospects.

Claims

1. Use of dextrophosphorylation in the preparation of a drug for preventing and / or treating kidney damage, wherein the structural formula of dextrophosphorylation is as shown in formula (I):

2. The use according to claim 1, characterized in that The renal injury is acute renal injury.

3. The use according to claim 1, characterized in that The acute kidney injury was induced by glycerol.

4. The use according to claim 3, characterized in that Dextrophosphorylation is used as an active compound, and excipients acceptable to pharmaceutical preparations are added to prepare the pharmaceutical preparations.

5. The use according to claim 4, characterized in that The auxiliary materials include one or more functional carriers of excipients, stabilizers, antioxidants, colorants, diluents, and sustained-release agents.

6. The use according to claim 5, characterized in that The pharmaceutical preparation is in any dosage form of tablets, capsules, suspensions, emulsions, powder injections and injections.

7. The use according to any one of claims 1 to 6, characterized in that: The drug for preventing and / or treating kidney injury has one or more of the following functions: (1) Reduce serum creatinine levels in mice with acute kidney injury; (2) Reduce the level of serum urea nitrogen in mice with acute kidney injury.

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