CN105106184A - Application of astaxanthin in preparation of medicine curing decreased insulin sensitivity - Google Patents

Abstract

The invention discloses the application of astaxanthin in the preparation of a pharmaceutical composition for promoting the expression of IRS-1 and IRS-2 in the treatment of decreased insulin sensitivity; and a pharmaceutical composition for treating insulin resistance, which is characterized in that it contains astaxanthin white.

Description

Application of astaxanthin in the preparation of drugs for the treatment of decreased insulin sensitivity

technical field

The present invention relates to the application of astaxanthin in the preparation of drugs for treating insulin sensitivity reduction, more specifically, the present invention relates to the application of astaxanthin in the preparation of drugs for promoting the expression of IRS-1 and IRS-2 in the preparation of drugs for treating insulin sensitivity reduction Applications.

Background technique

Insulin resistance refers to the decline in the efficiency of insulin in promoting glucose uptake and utilization due to various reasons, and the body compensatoryly secretes excessive insulin to produce hyperinsulinemia in order to maintain the stability of blood sugar. Insulin resistance predisposes to metabolic syndrome and type 2 diabetes. In the 1950s, Yallow et al. used radioimmunoassay techniques to measure plasma insulin concentrations and found that patients with lower plasma insulin levels had higher insulin sensitivity, while those with higher plasma insulin levels were insensitive to insulin, thus proposing the concept of insulin resistance. As far as its cause is concerned, there are the following factors, 1. genetic factors, such as abnormal structure of insulin, the presence of insulin antibodies, insulin receptors or gene mutations behind insulin receptors in the body (such as Glut4 gene mutation, glucokinase gene mutation and insulin receptor body substrate gene mutation, etc.); 2. Obesity, obesity is the main cause of insulin resistance, especially central obesity. Obesity is mainly related to long-term lack of exercise and excessive intake of dietary energy. 80% of patients with type 2 diabetes are diagnosed with obesity; 3. Other related diseases, such as long-term hyperglycemia, hyperlipidemia, certain drugs such as Hormone, certain trace element deficiency such as chromium and vanadium deficiency, pregnancy and increased insulin resistance hormones in the body, etc.; 4. Increased tumor necrosis factor a (TNF-a), enhanced TNF-a activity can promote lipolysis and cause increased plasma free fatty acid levels , leading to insulin resistance and hyperinsulinemia; 5. Other factors, such as leptin resistance and adiponectin level reduction or activity weakening, increased triglyceride content in skeletal muscle cells, excessive accumulation of cholesterol in B cells caused its Decreased function. In recent years, it has been found that resistin secreted by adipocytes can reduce glucose uptake after insulin stimulation, and tissue uptake of glucose will rise after neutralizing resistin. According to the insulin dose-response curve, it can be seen that there are three forms of insulin resistance: 1. The simple curve shifts to the right, indicating that the effector organs of insulin are less sensitive to insulin, and it is necessary to increase the dose of insulin to achieve the maximum response; 2. The height of the simple curve decreases , increasing the dose of insulin can not reach the maximum response height, which suggests that the target organ's response to insulin decreased; 3. At the same time, the curve shifted to the right and the maximum height of the curve decreased, indicating that both insulin sensitivity and responsiveness decreased. Insulin resistance checks currently include: 1. Fasting insulin, which is a good indicator of insulin resistance in the population; 2. Euglycemic insulin clamp technique, which is currently recognized as a method for detecting insulin resistance and is considered to be the The gold standard for evaluating other methods for detecting insulin resistance, 3. Insulin inhibition test, is a simple and easy method, but the results are not as accurate as the clamp method. 4. Miniature model method, miniature model technology (MMT) is to use computer to simulate the relationship between blood glucose and insulin dynamic metabolism, and simultaneously calculate the insulin sensitivity index (ISI) indicating the degree of insulin resistance and the self-metabolism of glucose independent of insulin action Efficacy (SG); 5. Simultaneous measurement of insulin release curve in glucose tolerance test, the common advantage of this method is that compared with blocking glucose-insulin feedback method, it does not interfere with the physiological mechanism of glucose-insulin feedback; The glucose tolerance test is a more physiological experiment.

In July 1998, WHO defined insulin resistance syndrome as: ① insulin resistance; ② abnormal glucose tolerance; ③ blood pressure ≥ 160/90mmHg; ④ triglyceride ≥ 1.7mmol/L, high-density lipoprotein L; ⑤ central obesity; ⑥BMI > 30kg/m2; ⑦Waist-to-hip ratio, male > 0.9, female > 0.85; ⑧Hyperuricemia; ⑨Microalbuminuria. An individual with diabetes or impaired glucose tolerance and/or insulin resistance, and a combination of two or more at the same time, can be defined as insulin resistance syndrome. The treatment of insulin resistance, general primary or hereditary insulin resistance has no effective treatment, but for some high-risk groups of insulin resistance, such as first-degree relatives with family history of diabetes, families with hypertension, hyperlipidemia Children with a history of low birth weight or a history of intrauterine malnutrition should pay special attention to avoid obesity during their acquired life, so as to prevent the occurrence of insulin resistance as much as possible. For people who have shown insulin resistance, different methods should be adopted according to different groups of people to reduce the factors that cause or aggravate insulin resistance, and at the same time, individualize the treatment for the different components of the metabolic syndrome that the individual has. 1. Strengthen exercise and control diet. Obese people emphasize a reasonable diet plan to reduce weight. At the same time, carry out long-term scientific and regular exercise to reduce weight; 2. Insulin sensitizers, thiazolidinedione drugs are powerful insulin sensitizers, which can reduce insulin resistance in type 2 diabetes by 33%. Rosiglitazone , pioglitazone, etc.; 3. To control blood sugar, for mild, moderately obese or overweight type 2 diabetic patients, thiazolidinedione derivatives, biguanides, glucotaxime inhibitors (Biglucopine) can be selected. In addition, For type 2 seminal diabetes treated with sulfonylureas or insulin, if hemorrhage control is not ideal, the above antihyperglycemic drugs can be combined with the above-mentioned antihyperglycemic drugs according to the specific situation, which can have a synergistic hypoglycemic effect; 4. Individually select antihypertensive drugs, Many hypertensive patients are often accompanied by insulin resistance. Diuretics and β-blockers may aggravate insulin resistance and have adverse effects on glucose metabolism. Long-term high-dose use should be avoided. Calcium ion antagonists have no adverse effects on glucose metabolism; α-receptor blockers, angiotensin-converting enzyme inhibitors, and angiotensin II receptor blockers can slightly improve insulin resistance while lowering blood pressure, which may be to a certain extent Reduce the risk of diabetes in patients with high blood pressure; 5. Correct lipid metabolism disorders, lipid metabolism disorders such as hypertriglycerides and hyperfree fatty acids are closely related to insulin resistance, hypertriglyceridemia and hyperfree fatty acids further exacerbate insulin resistance. Applying lipid-lowering drugs to improve lipid metabolism can reduce insulin resistance; 6. Supplementing trace elements, the lack of trace elements such as chromium and vanadium, may be related to insulin resistance, and proper dietary supplementation of trivalent chromium ions and trace elements vanadium is beneficial to the alleviation of insulin resistance .

Insulin receptor substrate (insulin receptor substrate, IRS), a phosphorylated protein involved in insulin and other cytokine signal transduction. After IRS is phosphorylated by the insulin receptor, it binds to proteins with SH2 domains like a "magnet", and produces different effects according to the specific structure of the bound proteins, such as activating the enzymatic activity of SH2 proteins, changing the protein configuration and Bind to another protein or cause the transfer of a protein from one part of the cell to another. Four members of the IRS family, IRS-1 to IRS-4, have been found, which differ in tissue distribution, subcellular localization, expression timing during development, binding to insulin, and interaction with SH2-containing proteins. It is a key intermediary molecule in the insulin signal transduction system; it acts as an anchor protein between insulin receptors and the complex network of signaling molecules containing SH2 domains in cells, and participates in the signal transduction of various hormones and cytokines, and It plays a central role in maintaining the basic functions of cells such as growth, survival and material metabolism.

The liver is an important organ for glucose and lipid metabolism and the main target organ for insulin action. Numerous studies in recent years have shown that lipid metabolism disorders play an important role in the occurrence and development of type 2 diabetes. When the liver cells are damaged or dysfunctional, the synthesis of lipoproteins by the liver cells decreases, resulting in the inability of the lipoproteins to transport lipid substances and the transportation is blocked, which eventually leads to fat accumulation and severe lipid metabolism disorders. Research has shown that most patients with type 2 diabetes suffer from fatty liver, and in chronic liver disease, especially in patients with fatty liver, the incidence of type 2 diabetes increases significantly and the fasting blood sugar is higher. The two are closely related. Reciprocal causation. Clinically, in view of the coexistence of both diseases, the treatment is often coordinated. When lowering blood sugar, it is necessary to treat chronic liver diseases. Improving liver function is obviously beneficial to blood sugar control.

The main pathology of diabetic liver damage is hepatic steatosis, which can gradually evolve into non-alcoholic steatohepatitis, liver fibrosis, cirrhosis, etc. Now it is being researched and developed all over the world. , and even natural anti-lipid metabolism disorder drugs without toxic and side effects. Since the early 1960s, marine biological resources with great development potential have been the focus of attention in the medical field. Finding new marine biological resources has now become the focus of natural drug research. an important goal. The full name of astaxanthin (ASX) is 3,3′-dihydroxy-β-carotene-4,4′-one, which is a red carotenoid widely distributed in marine bacteria, algae, crustaceans and fish Among them, many literatures have reported that astaxanthin has shown strong effects in anti-oxidation, anti-inflammation, anti-cancer, anti-Helicobacter pylori (HP) infection and anti-ultraviolet light, and has few toxic side effects.

More and more research results show that astaxanthin has considerable medicinal value in protecting the function of islet β cells, improving insulin resistance and preventing diabetes. ZhangM et al. used carbon tetrachloride to establish a liver fibrosis model, and treated rats with astaxanthin at doses of 20, 40, and 80 mg/kg respectively. It was observed that astaxanthin can significantly reduce the pathological changes of liver fibrosis, and also It can inhibit the expression of NF-kb and TGF-β, thereby reducing the activation of hepatic stellate cells and the production of extracellular matrix, especially with the intervention of large doses of astaxanthin, the greater the improvement of liver fibrosis. BhuvaneswariS found that the antioxidant and anti-inflammatory effects of astaxanthin can effectively reduce the occurrence of oxidative stress in the endoplasmic reticulum of liver cells, prevent and delay the chronic symptoms caused by inflammation.

The etiology of insulin resistance is complex, and its occurrence and development are closely related to many factors such as fat metabolism disorder, obesity and oxidative stress, but the relationship between them and the molecular mechanism of insulin resistance development are still unclear. Clinically, the prevention and correction of dyslipidemia is the first step, both in the primary prevention and early treatment of diabetes.

Contents of the invention

In the prior art, there is no drug used to promote the expression of IRS-1 and IRS-2. The present inventor established a classic type 2 diabetes animal model in which lipid metabolism disorder caused by high-fat feeding was supplemented by STZ partially destroying pancreatic islet b cells, and observed the effects of astaxanthin on blood sugar and blood lipids in high-fat-fed rats, and the effect of astaxanthin on insulin Sensitivity, surprisingly found that astaxanthin can promote the expression of IRS-1 and IRS-2, and then increase the sensitivity of insulin, to achieve the purpose of lowering blood sugar, thus completing the present invention.

The astaxanthin involved in the present invention, also known as astaxanthin and lobster shell pigment, is a carotenoid and the highest grade product of carotenoid synthesis. It is dark pink in color and has a chemical structure similar to β-carotene. β-carotene, lutein, canthaxanthin, lycopene, etc. are all intermediate products of carotenoid synthesis, so in nature, astaxanthin has the strongest antioxidant activity. Widely present in the biological world, especially in shrimp, crab, fish, algae, yeast and bird feathers, it is one of the main carotenoids in marine organisms.

Astaxanthin chemical name: 3,3'-dihydroxy-4,4'-diketo-β-carotene, pigment Aj067-69CASNo: 472-61-7, molecular formula C40H52O4, molecular weight 596.86. Due to the optical activity of the hydroxyl groups (-OH) at both ends, astaxanthin has three isotropic species: 3S-3'S, 3R-3'S, and 3R-3'R (also known as left-handed, meso-, and right-handed). Configuration state, in which artificial astaxanthin is a mixture of 3 kinds of structure astaxanthin (levorotatory accounted for 25%, dextrorotatory accounted for 25%, mesorotatory accounted for 50%), very little antioxidant activity, and salmon and other aquaculture organisms The astaxanthin in the body (mainly in the trans structure - 3S-3S type) is quite different. The astaxanthin derived from yeast is 100% dextrorotatory (3R-3′R), and has partial antioxidant activity; the above two This source of astaxanthin is mainly used in the coloring of non-edible animals and materials. Only astaxanthin from algal sources has a 100% left-handed (3S-3'S) structure and has the strongest biological activity.

Description of drawings

Fig. 1: Experimental flowchart of the embodiment of the present invention.

Figure 2: Effect of astaxanthin on blood sugar in type 2 diabetic rats

Figure 3: Effect of astaxanthin on blood insulin in type 2 diabetic rats

Figure 4: Effect of astaxanthin on OGTT in type 2 diabetic rats

Figure 5: Effect of astaxanthin on the area under the OGTT curve in type 2 diabetic rats

Figure 6: The effect of astaxanthin on the expression of IRS-1 in the liver of type 2 diabetic rats

Figure 7: The effect of astaxanthin on the expression of IRS-2 in the liver of type 2 diabetic rats

Detailed ways

Materials and Methods

1.1 Experimental animals

Healthy male Sprague-Dawley (SD) rats, aged one month and weighing 100-120 g, were provided by the Experimental Animal Center of Zhejiang Province. The animals were kept in separate cages, and the experimental animals were adapted to the environment in the animal center for a week before starting the experiment. They were fed with standard rat chow with free drinking water, and the cages were kept clean. The room temperature was controlled at 24°C, with natural light sources, good ventilation, and a relative humidity of 60%. Animal experiments were performed at the Experimental Animal Center of Ningbo University.

1.2 Experimental Instruments and Reagents

1.2.1 Main experimental instruments

Table 1 Names and manufacturers of experimental instruments

1.2.2 Experimental drugs and reagents

Table 2 Names and manufacturers of experimental drugs

experimental method

1.2.3 Criteria for the success of the premorbid state model of type 2 diabetes

The experimental rats were first fed with a high-fat diet (the ratio of each component per 100g was crude protein: fat: carbohydrate = 20:40:40) for 6 weeks, and the model was built, and the tail was cut to take blood to measure random blood sugar, so as to ensure that the experiment was large. The rats were in a state before the onset of diabetes, and the mental state of the rats was observed every day to ensure sufficient food and drinking water.

1.2.4 Drug intervention

The administration time of the prevention group was 4 weeks and 6 weeks, and the intragastric administration dose was 25mg/kg, and the other groups were simultaneously given the same amount of normal saline, free to eat and drink.

1.2.5 Random blood glucose measurement and oral glucose tolerance test

During the drug prevention treatment, each rat in each group had a random blood glucose test at the same time every 3 days, and used an automatic blood glucose meter to take blood from the tail vein of the rats for measurement, and made relevant records immediately, and counted the blood glucose values after the experiment was over. Variety.

During the animal experiment, rats in each group underwent oral glucose tolerance test after 4 weeks of drug intervention. The specific method is as follows: after fasting for 12 hours, rats are gavaged with 50% glucose solution (gastric gavage dose is 2 g/kg). After gavage for 0h, 0.5h, 1h and 2h, the tail was cut to measure the blood glucose value. After the experiment, the glucose tolerance curve of rats in each group was drawn, and the area under the glucose tolerance curve was calculated.

1.2.6 Blood collection, specimen fixation, paraffin section, H&E staining

1.2.6.1 Blood sample collection

Blood collection: After the treatment, each rat chooses the inner canthus vein to collect blood. The specific method is: prepare a vacuum blood collection tube in advance and mark it, fix the rat with the left hand, and use a capillary glass tube with a diameter of 1 mm and a length of about 5 cm in the right hand. Rotate and insert the inner side of the eyeball of the rat. After adjusting the depth, the venous blood flows out immediately. The blood is collected with a blood collection tube. After the blood is left to stand for 2-3 hours, it is centrifuged with a low-speed centrifuge at 2000rpm for 10min, and the supernatant is taken for detection with an automatic biochemical instrument. Blood lipid index.

1.2.7 Western blot method to analyze the expression of liver antioxidant factors Nrf2, HMOX-1, SOD-1 protein

Table 3 Westernblot operation process

【Example】

Therapeutic effect of astaxanthin on type 2 diabetes

The experiment is shown in the experimental flow chart (see Figure 1): Take 20 male healthy SD rats, first randomly select 4 as the normal control group, and feed them with common rat diet for 6 weeks. Rats in other groups were synchronously fed with high-fat diet for 6 weeks and then intraperitoneally injected with 1% STZ citrate buffer at a dose of 25 mg/kg. Afterwards, the tail was cut and the blood was taken to measure the random blood glucose value. Eight animals with random blood glucose value ≥ 11.1 mol/L were selected as experimental rats, and randomly divided into the diabetes group and the treatment group with 4 rats in each group. The dose of astaxanthin given to the treatment group was 25 mg/kg; the other groups were given the same amount of normal saline at the same time, and each rat was gavaged once a day for 6 weeks, free to eat and drink. After the treatment, the rats in each group were subjected to OGTT test. Then the rats were sacrificed, and the rat serum was taken to extract the liver samples for subsequent analysis of changes in blood sugar levels, and the RNA and protein of fresh liver tissues were extracted, and the expression levels of factors related to liver lipid metabolism and insulin signaling pathways were detected at the mRNA level by RT-qPCR .

Table 4 Comparison table of Chinese and English abbreviations in the manual

STZ Streptozotocin AUC area under the glucose tolerance curve OGTT oral glucose tolerance test HE Hematoxylin and eosin staining Nrf-2 nuclear factor E2 related factor HMOX-1 heme oxygenase-1 SOD-1 SOD-1

Claims (10)

1. The application of astaxanthin in the preparation of a pharmaceutical composition for promoting the expression of IRS-1 and IRS-2 in the treatment of decreased insulin sensitivity. 2. The application according to claim 1, wherein the astaxanthin is natural astaxanthin. 3. The use according to claim 1 or 2, wherein the decreased insulin sensitivity is selected from one or more of hepatic steatosis, obesity and diabetes. 4. The use according to claim 1 or 2, wherein said decreased insulin sensitivity can cause type 2 diabetes. 5. The use according to claim 1 or 2, wherein the decreased insulin sensitivity comes from before and during the onset of diabetes in type 2 diabetic rats. 6. A pharmaceutical composition for treating decreased insulin sensitivity, characterized by containing astaxanthin. 7. The pharmaceutical composition according to claim 7, wherein said astaxanthin is natural astaxanthin. 8. The composition according to claim 6 or 7, wherein the decreased insulin sensitivity is selected from one or more of hepatic steatosis, obesity and diabetes. 9. The removing composition according to claim 6 or 7, wherein said decreased insulin sensitivity causes type 2 diabetes. 10. The composition according to claim 6 or 7, wherein the decreased insulin sensitivity comes from before and during the onset of type 2 diabetes in rats.