CN115581697A - Application of nicotine and derivatives as FXR-meglin/cublin regulator in preparation of choleretic products - Google Patents

Abstract

The invention discloses the application of nicotine and its derivatives as FXR-meglin/cublin regulator in the preparation of choleretic products, and the choleretic products are anti-cholestasis or anti-galebladder stone products. In this application, nicotine and its derivatives are used to effectively treat cholestasis and gallstones by inhibiting the expression and function of FXR-megalin/cublin in the gallbladder. It has novel targets, reliable curative effect, no toxic side effects, rich resources, Low price and other advantages.

Description

Nicotine and its derivatives as FXR-meglin/cublin regulators in the preparation of choleretic Application in products

technical field

The invention relates to the technical field of medicine, in particular to the application of nicotine and its derivatives as FXR-meglin/cublin regulators in the preparation of choleretic products.

Background technique

Gallstone disease is one of the most prevalent and costly gastrointestinal disorders worldwide. Cholesterol gallstones are a major public health problem in all developed countries. The formation of gallstones is affected by several factors, including smoking. However, some studies have shown that smoking is not a risk factor for gallstones and may even have the opposite effect. Elderly smokers who have smoked for most of their lives have a lower risk of gallstone disease than non-smokers. In view of this, the development of new and effective drugs from tobacco to change the level of gallstone-related proteins in the gallbladder or bile components, thereby affecting the formation of gallstones, has positive significance for the prevention and treatment of gallstones.

Gallstones form in the biliary system from secretions of cholesterol in the bile, which results from excess cholesterol and/or a deficiency of bile salts. Many genes have been found to be related to the formation of gallstones, and they are involved in the regulation of cholesterol supersaturation in bile, crystallization nucleation, abnormal lipid metabolism, abnormal bile components, gallbladder dysfunction, and abnormal absorption of gallbladder inner wall mucosa. Affects cholesterol/bile acid metabolism, induces supersaturation of cholesterol from bile, leading to stone formation. Therefore, regulating the expression of genes involved in cholesterol/bile acid metabolism may be related to stone formation.

Currently, clinically used oral gallstone drugs mainly include chenodeoxycholic acid (CDCA) and ursodeoxycholic acid (VDCA). These two drugs desaturate bile by reducing the secretion of bile cholesterol, and unsaturated bile has the effect of dissolving cholesterol, so that the cholesterol molecules on the surface of gallstones are continuously dissolved, and the volume of gallstones gradually shrinks until they are completely dissolved. However, studies have shown that these stone treatment drugs are only suitable for gallstones with diameters below 2 cm. In addition, the drugs for stone treatment are expensive, and have certain side effects and toxic reactions (mainly including gastrointestinal tract, liver and gallbladder dysfunction, and allergic reactions), and must be taken for life. Once it becomes supersaturated again, the calculus will recur. According to statistics, the recurrence rate in 3 years can reach 25%. Therefore, there is an urgent need to find high-efficiency and low-toxic drugs based on new targets for the clinical treatment of gallstones.

Nicotine, commonly known as nicotine, is a powerful parasympathetic alkaloid found in plants of the family Solanaceae. It is the main chemical component and main addictive component of cigarettes, and is a kind of stimulant. Nicotine is a nicotinic acetylcholine receptor agonist and acts as a receptor antagonist on nAChRα9 and nAChRα10. A number of authoritative evidence shows that nicotine can be used to treat depression, Parkinson and other diseases. At the same time, nicotine itself has anti-inflammatory effects, so it is beneficial to ulcerative colitis. However, the effect on gallstones has not been reported yet.

Contents of the invention

The technical problem to be solved by the present invention is to provide the application of nicotine and its derivatives as FXR-meglin/cublin regulator in the preparation of choleretic products. Expression and function of bile, regulate bile cholesterol metabolism, effectively inhibit the formation of gallstones.

The technical problem to be solved by the present invention is achieved through the following technical solutions:

Application of nicotine and its derivatives as FXR-megalin/cublin regulator in the preparation of choleretic products.

Preferably, the choleretic product is an anti-cholestasis or anti-gallstone product.

Preferably, the nicotine is extracted from tobacco, and the derivative is (1'S.2'S)-nicotine 1'-oxide. Of course, nicotine can also be a pure product or a product obtained by other means, and this application is not limited thereto.

Preferably, said nicotine affects gallstone formation by regulating cholesterol metabolism in gallbladder epithelial cells.

Preferably, the choleretic product is medicine or health product.

A pharmaceutical composition comprising nicotine and its derivatives.

Preferably, the composition is administered intravenously or orally.

Preferably, the pharmaceutical composition is tablet, granule, capsule, suspension or freeze-dried formulation.

Preferably, the pharmaceutical composition also includes medically acceptable excipients.

The technical scheme of the present invention has the following beneficial effects:

The invention provides the application of nicotine and its derivatives in antagonizing the formation of gallbladder stones through FXR-megalin/cublin. Function, used to effectively treat cholestasis and gallstones, it has the advantages of novel action target, reliable curative effect, no toxic side effects, abundant resources, and low price.

In addition, nicotine and its derivatives have the activity of inhibiting the expression of FXR-megalin/cublin, and can be used to prepare choleretic drugs or health products, and to prepare drugs for treating cholestasis or gallstones.

This application clarifies the regulation mechanism of nicotine between gallstone formation and cholesterol/bile acid balance, and provides a theoretical basis for the development of preventive drugs for corresponding diseases.

Description of drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the principles of the invention.

Fig. 1 is the therapeutic effect of nicotine of the present application on mouse gallstones (wherein: A is the formation of gallbladder stones in mice of different groups, and B is a histogram of gallbladder weights of mice in different groups).

Fig. 2 is the regulatory effect of nicotine of the present application on bile metabolism (wherein: A is the level of the mouse cholesterol of different groups, B is the level of the phospholipids of the mice of different groups, C is the bile salt of the mice of different groups D is the level of triglyceride in different groups of mice, E is the level of total cholesterol in different groups of mice, F is the level of low-density lipoprotein in the bile of different groups of mice, G is the level of different groups of mice The level of high-density lipoprotein in mice of different groups, H is the level of total bile acid in mice of different groups).

Fig. 3 is the regulatory effect of nicotine of the present application on mouse gallbladder FXR-megalin/cublin (wherein: A is the mRNA level of bile acid metabolism-related factors detected by fluorescence quantification, and B is the protein of FXR, Megalin and Cubilin detected by western blotting Level, C is the expression of FXR and Megalin detected by immunohistochemical staining).

detailed description

Various exemplary embodiments of the present invention will now be described in detail with reference to the accompanying drawings. It should be noted that the relative arrangements of components and steps, numerical expressions and numerical values set forth in these embodiments do not limit the scope of the present invention unless specifically stated otherwise.

Therapeutic effect of embodiment 1 nicotine on mouse gallbladder stone

Experimental Materials:

Wild C57 mice, 8 weeks old, male. Randomly divided into 5 groups: normal diet group (ND), normal diet+nicotine group (ND+nicotine(H)), stone-forming diet group (LD), stone-forming diet+low-dose nicotine group (LD+nicotine(L) ), stone-forming diet+high-dose nicotine group (LD+nicotine(H)).

experimental method:

(1) Normal diet in the first week, and a high bile salt diet (1.25% cholesterol + 0.5% cholic acid) was replaced from the second week. The normal diet group was given normal saline, the normal diet + nicotine group, the stone-forming diet group, the stone-forming diet + low-dose nicotine group, and the stone-forming diet + high-dose nicotine group were given nicotine (low dose of 1.1 mg/kg) from the fifth week. and high dose of 6.6mg/kg), administered by intraperitoneal injection, once every 2 days, at the end of the 10th week.

(2) At the 10th week, the mice were sacrificed and then laparotomized to observe gallbladder stones, take pictures, ligate the common bile duct, take out the gallbladder, and weigh the stones.

Experimental results:

The results showed that after the stone-forming diet, the rate of gallstone formation in mice was higher. Under the administration of nicotine, the weight of the gallbladder was significantly lower than that of the stone-forming diet group, and the volume of bile increased, indicating that nicotine has the effect of treating gallstones and cholestasis, as shown in Figure 1.

Influence of nicotine on the metabolism of bile in the body of embodiment 2

Experimental Materials:

Wild C57 mice, 8 weeks old, male, were randomly divided into 5 groups: normal diet group (ND), normal diet+nicotine group (ND+nicotine(H)), stone-forming diet group (LD), stone-forming diet+ Low-dose nicotine group (LD+nicotine(L)), stone-forming diet+high-dose nicotine group (LD+nicotine(H)).

experimental method:

(1) Normal diet in the first week, and a high bile salt diet (1.25% cholesterol + 0.5% cholic acid) was replaced from the second week. The normal diet group was given normal saline, the normal diet + nicotine group, the stone-forming diet group, the stone-forming diet + low-dose nicotine group, and the stone-forming diet + high-dose nicotine group were given nicotine (low dose of 1.1 mg/kg) from the fifth week. and high dose of 6.6mg/kg), administered by intraperitoneal injection, once every 2 days, at the end of the 10th week.

(2) At the 10th week, mice were anesthetized with 35 mg/kg of barbital sodium, and gallbladder bile was collected, and blood was collected intravenously. Biochemical analyzer for cholesterol, phospholipids, bile salts, triglycerides (TG), total cholesterol (TC), low-density lipoprotein (LDL-C), high-density lipoprotein (HDL-C), total bile acid (TBA) to test. Liver tissues were collected, and the expression levels of bile metabolism-related mRNAs were determined by qPCR, including FXR, Cyp7A1, Cyp7B1, Cyp8B1, Cyp27A1, BSEP, NTCP, OATP1, OSTα, OSTβ, MRP2, MRP3, and MRP4.

Experimental results:

The results showed that compared with the LD group, the cholesterol level in gallbladder bile was significantly decreased in the LD+nicotine(H) group. Compared with the LD group, the cholesterol level of the LD+nicotine(L) group was not significantly reduced. Compared with the LD group, there was no significant difference in the phospholipids and bile salts in the bile of the experimental group. Compared with the ND group, the serum TG, TC, and LDL-C levels in the LD group were significantly increased, while HDL-C and TBA were significantly decreased. Levels of these indicators did not change after treatment with low concentrations of nicotine. Compared with the LD group, the serum TG, TC, LDL-C of the mice in the LD+nicotine(H) group were significantly decreased, while the levels of HDL-C and TBA were increased. Compared with the LD group, the Cyp7A1, NTCP, and OSTβ mRNAs in the nicotine treatment group were significantly up-regulated; FXR and Cyp7B1 mRNAs were down-regulated. It shows that nicotine has the effect of regulating bile cholesterol, as shown in Figure 2.

Example 3 Inhibitory activity of nicotine on FXR-megalin/cublin and its influence on metabolic genes

Experimental Materials:

Wild C57 mice, 8 weeks old, male, were randomly divided into 5 groups: normal diet group (ND), normal diet+nicotine group (ND+nicotine(H)), stone-forming diet group (LD), stone-forming diet+ Low-dose nicotine group (LD+nicotine(L)), stone-forming diet+high-dose nicotine group (LD+nicotine(H)). Normal diet in the first week, and high bile salt diet (1.25% cholesterol + 0.5% cholic acid) was replaced from the second week.

experimental method:

(1) The normal diet group was given normal saline, the normal diet + nicotine group, the stone-forming diet group, the stone-forming diet + low-dose nicotine group, and the stone-forming diet + high-dose nicotine group were given nicotine (1.1 mg/kg) from the fifth week Low dose of 6.6mg/kg and high dose of 6.6mg/kg), intraperitoneal injection, once every 2 days, the end of the 10th week.

(2) At the 10th week, mice were anesthetized with 35 mg/kg sodium pentobarbital and sacrificed. The gallbladder tissue was collected, and the expression level of mRNA related to FXR-megalin/cublin signaling pathway was determined by qPCR; the protein expression level of FXR-megalin/cublin was detected by Western blotting; the expression of FXR-megalin was detected by immunohistochemistry4.

Experimental results:

The results showed that, compared with the ND group, the expression levels of NPC1L1 and megalin were significantly decreased, while the expression levels of ABCG5/G8 and SR-BI were increased in the LD group. Low and high concentrations of nicotine can attenuate their expression levels. Western blot and immunohistochemistry results showed that nicotine did not alter cubilin protein levels, but restored LD-downregulated FXR and megalin levels. This shows that nicotine can affect the formation of gallbladder stones by regulating cholesterol metabolism in gallbladder epithelial cells, as shown in Figure 3.

In this application, nicotine and its derivatives, as new FXR regulators, can regulate bile cholesterol metabolism and effectively inhibit the formation of gallstones by inhibiting the expression of FXR-meglin/cublin signaling pathway; they have the advantages of reliable curative effect and low price.

Although the present invention has been disclosed as above with the embodiments, it is not intended to limit the present invention, and any person skilled in the art can make various choices and modifications without departing from the spirit and scope of the present invention. Therefore, the present invention The scope of protection is defined by the claims and their equivalents.

Claims (9)

1. The application of nicotine and derivatives as FXR-megalin/cublin regulator in the preparation of choleretic products. 2. The application according to claim 1, wherein the choleretic product is an anti-cholestasis or anti-gallstone product. 3. The application according to claim 1, wherein the nicotine is extracted from tobacco, and the derivative is (1'S.2'S)-nicotine 1'-oxide. 4. The application according to claim 1, said nicotine affects the formation of gallbladder stones by regulating cholesterol metabolism in gallbladder epithelial cells. 5. The application according to claim 1, the choleretic product is medicine or health product. 6. A pharmaceutical composition comprising the nicotine and its derivatives according to any one of claims 1-5. 7. The pharmaceutical composition according to claim 6, characterized in that, the composition is administered intravenously or orally. 8. The pharmaceutical composition according to claim 6, wherein the pharmaceutical composition is tablet, granule, capsule, suspension or freeze-dried formulation. 9. The pharmaceutical composition according to claim 6, characterized in that, the pharmaceutical composition further comprises medically acceptable adjuvants.

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