Application of icariin in preparation of medicine for preventing and treating chronic kidney disease mineral substance and bone metabolism disorder

Abstract

The invention discloses application of icariin in preparation of drugs for preventing and treating chronic kidney disease mineral substances and bone metabolism disorders. Relates to the technical field of medicine. The invention provides an application of icariin as an active pharmaceutical ingredient in preparation of a CKD-MBD treatment drug. Pharmacological experiments show that icariin can improve renal fibrosis, inhibit hyperparathyroidism, correct calcium and phosphorus metabolism, regulate and control bone metabolism related molecular expression, inhibit transdifferentiation of vascular smooth muscle cells to bone cells, inhibit vascular calcification and correct bone metabolism disorder, so that CKD-MBD is effectively prevented and treated. When icariin is used for preventing or treating CKD-MBD, the icariin is clear in component, exact in curative effect and low in side effect, and has a wide medical application prospect.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medicine, more particularly relates to the application of icariin in the preparation of drugs for preventing and treating chronic kidney disease mineral and bone disorder. BACKGROUND

[0002] Chronic kidney disease mineral and bone disorder, namely CKD-MBD, can cause bone fracture and blood vessel calcification, and the blood vessel calcification can increase the mortality rate of cardiovascular disease and the overall mortality rate. The CKD-MBD seriously endangers the life quality and healthy life of patients and greatly consumes medical funds. Therefore, the CKD-MBD is a chronic disease that seriously threatens the health of the public.

[0003] However, there is still lack of clear and effective prevention and treatment drugs and intervention means. The main methods include adjusting high-phosphorus diet, applying phosphorus binder with meals, reducing phosphorus through dialysis, etc., and the effects are limited.

[0004] Traditional Chinese medicine is also applied to the treatment of CKD-MBD, but the traditional Chinese medicine treatment has problems such as complex components and unclear mechanism of action.

[0005] Icariin is an important active component of Epimedium, and pharmacological studies have shown that icariin can increase cardiovascular blood flow, promote hematopoietic function and immune function, and has the effects of tonifying kidney and strengthening yang, anti-aging, anti-tumor, etc.

[0006] Therefore, whether icariin can be used for preparing drugs for preventing and treating chronic kidney disease mineral and bone disorder and overcoming the above technical deficiencies is a problem that needs to be solved by those skilled in the art. SUMMARY

[0007] Therefore, the present application provides the application of icariin in the preparation of drugs for preventing and treating chronic kidney disease mineral and bone disorder. The application achieves the inhibition of hyperparathyroidism and blood vessel calcification and the correction of bone metabolic disorder.

[0008] In order to achieve the above purpose, the present application adopts the following technical solutions: The application of icariin, icariin derivatives or pharmaceutically acceptable salts thereof in the preparation of drugs for preventing or treating chronic kidney disease mineral and bone disorder.

[0009] Preferably, the chronic kidney disease mineral and bone disorder includes blood vessel calcification, secondary hyperparathyroidism and bone metabolic disorder.

[0010] Preferably, the drug improves blood vessel calcification, secondary hyperparathyroidism and bone metabolic disorder.

[0011] Preferably: the drug: regulates the expression of bone metabolism related molecules and inhibits the transdifferentiation of vascular smooth muscle cells into bone cells.

[0012] Preferably: the dosage form of the drug: oral preparation, injection preparation.

[0013] Preferably: the oral preparation is a microemulsion preparation, tablet, pill, capsule or granule; the injection preparation of icariin is an injection solution or lyophilized powder.

[0014] Further, the administration time and administration frequency for preventing or treating CKD-MBD according to the present application need to be determined according to the specific diagnosis of the condition. For example, the treatment regimen for preventing or treating CKD-MBD in rats or mice is applied to humans, and the effective dose of the drug for humans can be calculated from the effective dose of the drug for rats or mice.

[0015] According to the condition and the administration site, icariin can be prepared into a suitable pharmaceutical preparation for convenient administration. The oral preparation is preferably a tablet; the sublingual preparation is a pharmaceutical preparation containing icariin and suitable for sublingual administration, preferably a sublingual tablet; the injection preparation can be an injection solution, injection microemulsion, etc., preferably an injection solution. When icariin is prepared into an injection solution, the pharmaceutically acceptable carrier can be water for injection, sodium chloride, sodium citrate, citric acid, glycerol, ethanol, propylene glycol, etc. The above-mentioned icariin injection solution can also add appropriate additives according to the properties of the drug, such as osmotic pressure regulators, pH regulators, solubilizers, bacteriostatic agents, emulsifiers, suspending agents, etc., wherein the solubilizer is any one or both of polyethylene glycol 400 and Tween-80.

[0016] In the above-mentioned pharmaceutical preparations, the content of icariin in each preparation unit is 0.001 mg to 100 mg.

[0017] Through the above technical solution, compared with the prior art, the present application provides the use of icariin in the preparation of drugs for preventing and treating mineral and bone metabolism disorders in chronic kidney disease, and achieves the following technical effects: The present application shows that icariin has a significant effect on preventing or treating CKD-MBD. It can significantly improve renal fibrosis, inhibit hyperparathyroidism, correct calcium and phosphorus metabolism, regulate the expression of bone metabolism related molecules, inhibit the transdifferentiation of vascular smooth muscle cells (VSMC) into bone cells, inhibit vascular calcification and correct bone metabolism disorders through the regulation of the kidney-parathyroid-bone-vascular axis, thereby effectively preventing and treating CKD-MBD.

[0018] Icariin is a natural traditional Chinese medicine monomer extracted from the traditional Chinese medicine Epimedium, which has low toxicity and side effects on the human body, can significantly improve the safety and compliance of patients, and thus greatly improve the treatment effect and quality of life of patients with CKD-MBD. Attached Figure Description

[0019] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0020] Figure 1 The attached figure shows the HE and Masson staining results of rat kidneys provided by this invention.

[0021] Figure 2 The attached figure shows the results of Masson staining measurement of rat kidneys provided by this invention, compared with the sham-operated group. ### p < 0.001; compared with the model group, *** p < 0.001.

[0022] Figure 3 The attached figure shows the results of HE staining, Vonkossa staining, and Alizarin Red S staining of the aorta provided by the present invention.

[0023] Figure 4 The attached figure shows the expression of vascular smooth muscle cell markers α-SMA and SM22α, calcification transcription factor Runx2, osteopontin OPN and osteocalcin OCN in the aorta provided by the present invention.

[0024] Figure 5 The attached figure shows the statistical results of α-SMA expression in the aorta provided by this invention, compared with the sham surgery group. ### p < 0.001; compared with the model group, *** p < 0.001.

[0025] Figure 6 The attached figures show the results of toluidine blue staining and Alcian blue staining of the femur provided by this invention. A: Toluidine blue staining and Alcian blue staining show the results of new bone formation in the femoral neck; B: Toluidine blue staining; C: Alcian blue staining measurement results; compared with the sham surgery group, ### p < 0.001; compared with the model group, * p < 0.05 ** p < 0.01.

[0026] Figure 7The figure is a chart of bone metabolism related molecule expression level in femoral section provided by the present application, A: expression level of bone Gla protein OCN, bone bridge protein OPN and bone matrix related Collagen I molecule; B: expression measurement result of Collagen I molecule; C: expression measurement result of OCN; D: expression measurement result of OPN; compared with the sham operation group, ### p<0.05, compared with the model group, * p<0.05, ** p<0.01, *** p<0.001.

[0027] Figure 8 The figure is a chart of hyperparathyroidism of CKD-MBD rats corrected by icariin provided by the present application, wherein, A: HE staining of rat parathyroid, B, C: measurement result of parathyroid area and perimeter; D, E, F: serum parathyroid hormone, serum phosphorus and serum calcium level of rats; compared with the sham operation group, ## p<0.01, ### p<0.001, compared with the model group, * p<0.05, *** p<0.001. DETAILED DESCRIPTION

[0028] The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0029] The embodiments of the present application disclose application of icariin in preparation of drugs for preventing and treating mineral and bone metabolism disorder of chronic kidney disease.

[0030] The specific experimental methods not mentioned in the embodiments are generally carried out according to conventional experimental methods. The reagents and biological materials, if not specially specified, can be obtained from commercial channels.

[0031] Embodiment 1 Protective effect of icariin on vascular calcification, bone metabolism disorder and hyperparathyroidism of CKD-MBD rats induced by 5 / 6 nephrectomy plus high phosphorus drinking water 1 Experimental method 1.15 / 6 nephrectomy plus high phosphorus drinking water induced CKD-MBD rat model and administration scheme SPF level male Wistar rats, body weight (160-180) g, in the room temperature 24±2℃ environment to give the general standard pellet feed adaptability feeding 1 weeks after random selection of 10 as the sham operation group, the rest for modeling. Intraperitoneal injection of pentobarbital 45mg / kg anesthesia rats, except for the sham operation group, the remaining 3 groups of 5 / 6 nephrectomy (first operation to remove about 2 / 3 of the left kidney, a week after the second operation to remove the right kidney), sham operation group (two-step operation at the same period, the separation of kidney capsule, method same as above, but not any resection). After the second operation a week later the activity of the animals were randomly divided into groups, 10 in each group, respectively, model group, icariin low dose group, icariin high dose group. Icariin group 5 / 6 kidney surgery began to give icariin (low dose group 50 mg·kg–1; high dose group 100 mg·kg–1), once a day; sham operation group was given ordinary drinking water, the rest of the group was given high phosphorus drinking water (0.5 g / one, 2.5 g dissolved in 500 mL drinking water), for 14 weeks.

[0032] 1.2 animal sampling The body weight of rats was measured daily, and the death time of rats was recorded. At the 14th week after modeling, the rats were anesthetized by intraperitoneal injection of sodium pentobarbital, and blood was taken from the abdominal aorta. The kidneys, aorta, femur, thyroid and parathyroid of rats were removed, and a part of the organ tissue was fixed with 4% paraformaldehyde for pathological section staining observation. The remaining tissue was stored at -80℃ for subsequent homogenate index and protein determination.

[0033] 1.3 detection index The kidney, vascular tissue and decalcified femur were routinely paraffin-embedded to prepare sections, and the morphology of kidney, aorta, femur and parathyroid tissue was observed by routine HE staining. Masson staining was used to observe renal fibrosis, and alizarin red S staining and Von kassa staining were used to detect vascular calcification. The expression of vascular smooth muscle cell marker molecules α-SMA and SM22α, and bone metabolism related index molecules Runx2, osteocalcin OCN and osteopontin OPN in aorta were detected by routine immunohistochemistry. The expression of bone metabolism related index molecules osteocalcin OCN, osteopontin OPN and bone matrix related molecule Collagen I in femur tissue was detected by immunohistochemistry. The levels of calcium, phosphorus and parathyroid hormone were detected by blood biochemistry. 2 experimental results 2.1 Icariin significantly improves renal interstitial fibrosis in CKD-MBD rats The results of kidney HE staining and Masson staining are as follows Figure 1 and Figure 2The results showed that the kidney tissue in the sham-operated group was neatly arranged with no obvious abnormalities. In the model group, HE staining revealed partial renal tubular atrophy or compensatory dilation, scattered mononuclear and lymphocyte infiltration in the renal interstitium, and extensive myofibroblast proliferation, leading to widespread interstitial fibrosis; Masson staining showed abundant blue-stained collagen deposition in the renal interstitium. In contrast, the icariin treatment groups (low and high doses of ICA) showed reduced renal tubular dilation, significantly decreased interstitial inflammatory cell infiltration and fibrosis; Masson staining showed a significant reduction in blue-stained collagen deposition. These results indicate that icariin can alleviate pathological damage to the kidneys and has an anti-renal fibrosis effect.

[0034] 2.2 Icariin significantly inhibited vascular calcification in CKD-MBD rats Aortic sections were stained with HE, Alizarin Red S, and Von Kossa, respectively. All results showed the presence of calcifications in the arterial media in the model group, while icariin significantly inhibited aortic vascular calcification. Figure 3 Further immunohistochemical analysis was used to analyze the expression levels of molecules related to aortic vascular calcification. The results showed that the expression of vascular smooth muscle cell markers α-SMA and SM22α was significantly reduced in the aorta of the model group rats, while the expression of bone metabolism-related markers Runx2, OCN, and OPN was significantly increased. Icariin intervention significantly inhibited Runx2, OCN, and OPN, while increasing the expression of α-SMA and SM22α. Figure 4 and Figure 5 The above results indicate that icariin can inhibit the transdifferentiation of VSMCs into osteoblasts and prevent the occurrence and development of vascular calcification.

[0035] 2.3 Icariin significantly improved bone metabolism disorder in CKD-MBD rats. Femoral sections were stained with toluidine blue and Alcian blue, respectively. The results showed that the model group had reduced new bone formation and impaired bone metabolism and self-renewal. Icariin treatment restored the amount of new bone formation. Figure 6 Further immunohistochemical analysis was used to analyze the expression levels of bone metabolism-related molecules OCN, OPN, and bone matrix-related Collagen I. The results showed that OCN and OPN were significantly increased in the femur of the model group rats, while the expression of Collagen I was significantly decreased. This suggests that although bone activity in the model group showed a compensatory increase trend, the ability to synthesize bone matrix was reduced, indicating a metabolic disorder. This metabolic disorder leads to poor bone repair capacity and increased bone fragility. Icariin intervention significantly inhibited OCN and OPN, while increasing the expression of Collagen I. Figure 7 This study demonstrates that icariin can regulate osteogenic activity and bone matrix synthesis, thereby correcting the development and progression of bone metabolism disorders.

[0036] 2.4 Icaritin significantly inhibits parathyroid hyperfunction in CKD-MBD rats The results of parathyroid slice HE staining and area measurement showed that the parathyroid of the model group was significantly enlarged, and the cells were hypertrophic and diffusely proliferated, combined with blood biochemical indicators, the serum phosphorus and parathyroid hormone levels of the model group were significantly increased, suggesting that the model group had parathyroid hyperfunction; Icaritin significantly inhibited parathyroid hyperfunction in CKD-MBD rats, and the parathyroid showed normal state, and no hyperplasia and cell hypertrophy was observed, the serum calcium, phosphorus and parathyroid hormone (PTH) levels remained normal (P>0.05). Figure 8 The above results show that Icaritin significantly inhibits parathyroid hyperfunction in CKD-MBD rats.

[0037] In summary, Icaritin of the present application can improve renal fibrosis, inhibit parathyroid hyperfunction, correct calcium and phosphorus metabolism, inhibit vascular smooth muscle cell (VSMC) transdifferentiation into osteocyte, regulate the expression of bone matrix related Collagen I molecules and bone metabolism related index molecules OCN and OPN, inhibit vascular calcification and correct bone metabolism disorder by regulating the kidney-parathyroid-bone-vascular axis, thereby effectively preventing and treating CKD-MBD. When Icaritin is used for prevention or treatment of CKD-MBD, the composition is clear, the efficacy is exact, the side effects are low, and it has a broad medical application prospect.

[0038] The various embodiments in the specification are described in a progressive manner, and each embodiment focuses on the difference from other embodiments, and the same or similar parts between the various embodiments can be referred to each other.

[0039] Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein can be applied to other embodiments without departing from the spirit or scope of the application. Accordingly, the present application is not to be restricted to these embodiments shown in the figures, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. Use of icariin, icariin derivatives or pharmaceutically acceptable salts thereof in the preparation of a drug for preventing or treating mineral and bone metabolism disorder of chronic kidney disease.

2. Use according to claim 1, wherein The mineral and bone metabolism disorder of chronic kidney disease includes vascular calcification, secondary hyperparathyroidism and bone metabolism disorder.

3. Use according to claim 2, wherein the compound is ###0002### The drug improves vascular calcification, secondary hyperparathyroidism and bone metabolism disorder.

4. Use according to claim 3, wherein the compound is ###0002### The drug regulates the expression of bone metabolism related molecules and inhibits the transdifferentiation of vascular smooth muscle cells into bone cells.

5. The use according to claim 4, wherein the compound is ###0002### The dosage form of the drug is oral preparation or injection preparation.

6. Use according to claim 5, wherein The oral preparation is microemulsion preparation, tablet, pill, capsule or granule; the icariin injection preparation is injection solution or freeze-dried powder injection.

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