CN1919225A - Application of cultured saussurea medusa and extract thereof in preparation of medicine for treating II type diabetes and its complications - Google Patents
Application of cultured saussurea medusa and extract thereof in preparation of medicine for treating II type diabetes and its complications Download PDFInfo
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Abstract
The invention discloses jellyfish saussurea involucrate cell and extract application in the II-typed diabetes and complication prevention drug, which is characterized by the following: (1) possessing little side-effect, reducing the density of blood sugar and cholesterol, triglyceride and LDL effectively, (2) improving sensitivity of insulin acceptor, (3) protecting valued saussurea involucrate due to culturing plant cell.
Description
Technical field
Saussurea medusa that the present invention relates to cultivate and extract thereof the purposes in preparing the medicine for the treatment of type ii diabetes and preventing its complication belongs to biological engineering and modern Chinese traditional medicine field.
Background technology
Diabetes are incretion metabolism diseases that one group of cause of disease and pathogeny are not illustrated as yet fully, are its common sign with hyperglycemia.Water, the metabolic disturbance of electrolyte of or sugar, protein, fat and secondary that relative hyposecretion caused absolute because of insulin.It can relate to each system of whole body, even bring out many mortality complication, as infect complication, retinopathy, hypertension, nephropathy, coronary heart disease, lower limb vascular pathological changes and cerebrovascular disease etc., have a strong impact on people's work capacity, and threaten people's life security.Diabetes have several types: 1) insulin dependent diabetes mellitus (IDDM) (type i diabetes IDDM); 2) non-insulin-dependent diabetes mellitus (type ii diabetes NIDDM); 3) malnutrition type diabetes; 4) other type diabetes.Wherein the type ii diabetes patient accounts for 95% of the total number of patients of diabetes.
Along with the aging of world population, diabetes have become a kind of commonly encountered diseases, frequently-occurring disease, are the diseases that becomes the third-largest serious harm human health after tumor and cardiovascular and cerebrovascular disease.According to The World Health Organization's statistics in 2005, the whole world has 200,000,000 diabeticss approximately, China is one of maximum three countries (India, China, the U.S.) of diabetics, and diabetics quantity has surpassed 4,000 ten thousand people (" Chinese diabetes control guide ") at present.In addition, tens million of sugared regulatory function sufferers (IGR) is arranged still, this type of crowd becomes huge " the reserve legion " of diabetes.Preventing and treating diabetes has become one of important clinically, urgent health care problem, is paid attention to by people day by day.
The medicine of clinical treatment diabetes can be divided into oral antidiabetic drug, insulin, immunosuppressant and other medicines, and wherein oral antidiabetic drug and insulin are topmost classifications.Still do not have at present the effective ways of curing diabetes, control patient blood sugar level, complication prevention are the keys of treatment diabetes.Clinical oral antidiabetic drug commonly used mainly contains sulphanylureas, biguanides, alpha-glucosidase inhibitor, euglycemic agent and meals regulator five classes.Wherein, sulphanylureas, biguanide drug belong to older generation's product, and alpha-glucosidase inhibitor, euglycemic agent belong to product in Mesozoic Era, and the meals regulator is a product of new generation.The market trend of five class Western medicine is: sulphanylureas and biguanides still are line medications of present stage of china clinical prescription, the successful running of acarbose makes the whole alpha-glucosidase inhibitor market share enlarge rapidly, the consumption sum of sensitizer and blood sugar regulator used during user having meals and quantity all are tangible ascendant trend, and increasing degree is also very approaching.
Until the nineties in 20th century, oral antidiabetic drug has only sulfonylureas (SU) class and biguanide (BG) class 2 classes.Recent years, (class of α-GI), novel blood sugar lowing medicines such as thiazolidinedione (TZD) class and quick-acting insulin secretion accelerating medicines are developed listing to alpha-glucosidase inhibitor in succession.The third generation SU class medicine listing that in addition has insulin resistant improvement effect reaches the center of gravity assessment to BG class medicine, and the range of choice of type 2 diabetes mellitus medicine becomes more and more wideer.
(1) sulfonylureas (SU) class medicine SU class medicine mainly act as the promotion insulin secretion.The three generations has been experienced in its development: the first generation mainly is tolbutamide (tolbutamide) and chlorpropamide (ehloromide); The second filial generation has glibenclamide (glibenclamide, glyburide), gliclazide (gliclazide, diamicron), glipizide (glipizide, glipizide) and gliquidone (gliquidone, gliquidone) etc.; The third generation has glimepiride (glimepiride) etc.What China was used clinically at present mainly is the second filial generation, and the first generation still has a small amount of use, and the third generation is not seen the clinical practice report as yet.
Though third generation product glimepiride promotes that the effect of pancreatic insulin secretion is more weak, can strengthen the sensitivity of insulin.Along with the practical situation of SU receptor in the pancreatic is grasped by people, the SU class medicine that research and development have dual function should be a noticeable direction.
(2) the main effect of biguanide (BG) class medicine BG class medicine is to increase surrounding tissue to the utilization of glucose with suppress the absorption of intestinal to glucose, suppresses glycogen heteroplasia and glycogen output.This type of medicine comprises: phenformin (phenformin, insoral) and metformin (flumamine, metformin) etc.
Phenformin has the toxic and side effects of serious lactic acidosis, in the use that has been under an embargo of many countries, is domesticly clinically also reducing consumption.And metformin can be avoided lactic acidosis fully, simultaneously can also blood fat reducing, delay the generation of diabetic vascular complications, and make the incidence rate of scheming infarction reduce by 39%; Compare with sulphanylureas, the metformin blood sugar reducing function relaxes relatively, and hypoglycemia takes place few.Therefore metformin has become clinical practice biguanides the most widely, is mainly used in drug combination, treatment obese diabetic patient.
(3) (α-GI) the main effect of class medicine α-GI class medicine is by suppressing alpha-glucosidase, the absorption of slow down hydrolysis and glucogenic speed and delay glucose, thereby reduction post-prandial glycemia to alpha-glucosidase inhibitor.Because this inhibitory action is reversible, thus only be to postpone to the conversion of glucose, rather than blocking-up fully.It also has certain effect for reducing blood fat, can prevent and treat the chronic complicating diseases of diabetes.Represent medicine to have: acarbose (acarbose) and voglibose (voglibose).This type of medicine is proposed as the two wires medicine of treatment type 2 diabetes mellitus at present, and the domestic market is occupied by the import product mostly.Main side effects is the intestinal reaction, and price is more expensive.
(4) though this type of medicine of non-sulfonylurea insulin secretion stimulators and SU class medicines structure need not, mechanism of action has points of resemblance, and is to promote the pancreatic excreting insulin, difference is that the bonded position of they and beta cell is different.The medicine that has gone on the market has: repaglinide (repaglinide) and Nateglinide (nateglinide).The medicine that is in the III phase clinical research stage has the mitiglinide of Kissei company.
Nateglinide is 0.78h insulin level peaking (repaglinide is 0.92h) after the meal, insulin level similar to placebo (repaglinide is 4h) behind the administration 1.5h.Because reduced total insulin contact, the fluctuation of glucose when weakening meal is so it is littler to bring out hypoglycemic danger.
(5) insulin resistant improves this type of medicine of medicine and is euglycemic agent again, and it can increase surrounding tissue (muscle and fatty tissue) to the utilization rate of glucose and reduce the output rating of liver to glucose.Finding the earliest has the chemical compound of insulin-sensitizing effect all to have 2, and the 4-thiazolidinedione (2,4-thiazolidinedione) skeleton, so this class medicine also is called TZD class medicine.Represent medicine to have: rosiglitazone (rosiglitagone) and pioglitazone (pioglitagone).
Clinical research shows that rosiglitazone 4mg every day or 8mg are oral, and type 2 diabetes mellitus patient empty stomach, blood glucose after the meal and HbAlc are all descended, and keeping hypoglycemic effect can reach more than 30 months.Use rosiglitazone in early days at onset diabetes and help protecting the beta cell function.Rosiglitazone and statins coupling can be treated diabetes hyperlipemia disease, and the HDL cholesterol is risen, and triglyceride remains unchanged, and FFA significantly descends, and LDL significantly descends.Rosiglitazone can also reduce the off normal diethylarginine, and (asymdimethylarginene ADMA), reduces cardiovascular disease danger.In routine ND volunteer's research to one group 64, (steady state plasma glucose, SSPG) concentration and serum ADMA find that the rising of ADMA can reduce the utilization of glucose, and SSPG is risen to measure stable state blood glucose; Then the hyperpietic to wherein 7 example companion IR, serum ADMA risings gives rosiglitazone 4mg/d, in totally 4 weeks, changes 8mg/d then into, in totally 8 weeks, through checking once more after the treatment in 12 weeks, finds these patients when SSPG reduces, and serum ADMA all has decline.Even the ND to tool IR is described, rosiglitazone has the protective effect to blood vessel endothelium when improving insulin sensitivity.
Glimepiride, acarbose, rosiglitazone, pioglitazone and repaglinide come into the market later, but they are owing to having good and clinical curative effect and huge market potential is called as " Five Golden Flowers ", and the representative brand of its corresponding product is respectively Ya Moli, acarbose, Avandia, Ai Ting, NovoNorm.First generation sulfonylurea drugs glibenclamide (glyburide) has been in the edge that is eliminated; Glipizide (glipizide, enlightening sand, Yi Bida, Glipizide XL), gliclazide and gliquidone (gliquidone) are second filial generation sulfonylurea drugs, although it is bigger to purchase the medicine amount of money at present, all present slow downtrend.Glimepiride is a third generation sulfonylurea drugs, Time To Market is later, develop by German Hoechst Marion Roussel company, nineteen ninety-five is produced listing by the third-largest An Wante of drugmaker in the whole world first, commodity are called Amaryl, it is the oral chemical medicine of the treatment type ii diabetes of present western developed country first-selection, also is the best berculon B series products of global marketing, and annual sales amount in 2005 is 6.77 hundred million Euros (being equivalent to 800,000,000 dollars).Sulfonylurea drugs mainly by stimulating the islet cells excreting insulin, improves the level of insulin in the body.
Because treating diabetes is a secular process, various Western medicine life-time service all have certain limitation and untoward reaction, or even serious adverse effects very, as cause hypoglycemia, lactic acidosis etc.Up to the present, go back the neither one medicine and can cure diabetes, Western medicine also is a blood sugar control; Use insulin, not only cost an arm and a leg, and need drug administration by injection every day, patient to be difficult for adhering to, and administration time and dosage needs strict the grasp, otherwise easily cause hypoglycemia and dawn phenomenon at night.So the natural sugar reducing substance treatment diabetes in the appliable plant, have and be easy to accept and adhere to and advantage that toxic and side effects is little.
Show as diabetes at diabetics more,, use the method for clearing away heat-fire, supplementing QI for promoting the production of body fluid more, improve clinical symptoms as thirsty polydipsia, dysphoria with feverish sensation in the chest palms and soles, weak hyperhidrosis etc.Represent medicine such as diabetes pill (actual is the Chinese medicine and western medicine mixture), ginseng, astragalus diabetes granule etc.But Chinese medicine preparation lacks curative effect hypoglycemic medicine sure, that clinician and patient take like a shot, and seldom set up the blood sugar lowering model of pharmacy circle approval and select the positive control medicine of internationally recognized determined curative effect in zoopery and clinical trial, promptly the science of Chinese medicine is often out in the cold or suspected by the people.Western medicine generally is based on blood sugar lowering, and the new drug that exploitation has good hypoglycemic effect can prevent and treat diabetic complication simultaneously is Chinese medicine worker's the task of top priority.According to statistics, diabetics accounts for 80% by the mortality rate that cardiovascular and cerebrovascular disease causes, and makes its life expectancy reduce 1/3.Diabetics merges blood fat rising person more than 80%, and blood viscosity rising person reaches 90%.Diabetic complication focuses on prevention, and promptly seeking can blood sugar lowering, and the little medicine of life-time service toxic and side effects that can prevent and treat diabetic complication again is present optimal selection.
Saussurea medusa (Saussurea medusa Maxim) is a Compositae phoenix hair Chrysanthemum plant, is a kind of very famous and precious vegetable drug, originates between the high mountain gravel of 4800～5100 meters of height above sea level on ground such as China Gansu, Qinghai, Tibet, Xinjiang.Have effects such as dispelling cold and removing dampness, tonifying yang and enriching blood, anti-inflammatory and antalgic, rheumatic arthritis, sexual impotence, menoxenia, placenta retention, the high mountain inadaptation etc. of being used for the treatment of among the people.Clinical and pharmacological research proves recently, and Herba Saussureae Involueratae has slow down aging and treats diseases such as cerebral arteriosclerosis, cerebral infarction.Flavone compounds such as Herba Saussureae Involueratae extract acacetin, jaceosidin, hisidulin and quercetin have tangible effect to treatment cancer, antiviral, defying age (removing free radical).
Summary of the invention
The technical problem to be solved in the present invention provides the saussurea medusa and the purposes of extract in preparing the medicine for the treatment of type ii diabetes and preventing its complication thereof of cultivation.
For achieving the above object, the present invention is by the following technical solutions:
The purposes of the saussurea medusa of cultivating in preparing the medicine for the treatment of type ii diabetes and preventing its complication.
Wherein the saussurea medusa of Pei Yanging is to utilize biotechnology, makes the callus producer of Herba Saussureae Involueratae suddenly change the saussurea involucrata cell line of the stable high yield Herba Saussureae Involueratae flavone of acquisition by induced mutations.
Described induced mutations comprises the mutation of 60Co-radiation gamma, temperature mutation and ultraviolet mutagenesis.Original saussurea involucrata cell line is adopted the mutation of 60Co-radiation gamma, to obtain the higher saussurea involucrata cell line of yield of flavone; Adopt temperature mutation promptly to cultivate to the mutagenic obtained saussurea involucrata cell line of 60Co-radiation gamma, its gene is undergone mutation, adapting to the needs of wide temperature range growth, and improve the ability of the synthetic secondary metabolite flavone of cell biological with high temperature and low temperature; The Herba Saussureae Involueratae cell is adopted the ultraviolet light irradiation, its gene is undergone mutation, to improve the ability of the synthetic secondary metabolite flavone of cell biological.
Pharmacodynamics test of the present invention has used Chinese invention patent ZL02156868.5 and the described saussurea medusa of ZL200310124256.5 system, has obtained beyond thought excellent effect.Can know by inference according to result of the test of the present invention, by mutagenic obtained saussurea involucrata cell line all may have the present invention the drug effect that discloses first.Therefore, even other people use other saussurea involucrata cell line to be used to prepare the medicine for the treatment of type ii diabetes or are used to prepare the medicine that prevents the type ii diabetes complication, all belong to protection scope of the present invention.
The saussurea involucrata cell line that the present invention preferably protects is saussurea involucrata cell line TUIP-8, and its preserving number is that CGMCC No.0855 and saussurea medusa are SMXL-615, and its preserving number is CGMCC No.1072.The cultivation of this two strains Herba Saussureae Involueratae cell is not limited by natural conditions, the growth temperature range broad, and well-grown in 18～30 ℃ of scopes, growth is fast, and controllable product quality is good; This cell strain yield of flavone is big, the culture density height, and cultivation cycle is short; The cytogenetics stability of characteristics, in the continuous passage incubation, the ability of the synthetic flavone of cell is not degenerated; Cell aggregation growth, the cell mass quality is harder, can resist higher shearing force, can be in the reactor of higher rotation speed well-grown, be fit to commercial scale reactor and cultivate plant cell and realize industrialization.
The purposes of the saussurea medusa extract of cultivating in preparing the medicine for the treatment of type ii diabetes and preventing its complication.This extract is the boiling water extraction thing, can but be not limited to prepare by the following method: the fresh saussurea medusa culture of getting certain mass, squeezing is also collected the juice that obtains, residue Saussurea medusa cell culture is added in the boiling water that 8-12 doubly measures, stirred 1-3 minute, filter immediately, the juice that juice and the squeezing that obtains obtained merges promptly.
The conventional method in utilization Chinese materia medica field, the Herba Saussureae Involueratae cell extract also can obtain by other technique known, for example the Herba Saussureae Involueratae cell culture is added decoction in a certain amount of water, squeezes the juice or adds organic solvent extraction.But no matter the purposes of the Herba Saussureae Involueratae cell extract with drug effect that the present invention discloses first that obtains by which kind of mode all belongs to protection scope of the present invention.
The present invention is an experimental subject with internationally recognized type ii diabetes animal model KK-Ay diabetic mice (Chinese Academy of Medical Sciences's Experimental Animal Center); with saussurea medusa culture (China invention ZL02156868.5; China's invention 200310124256.5) boiling water leaching thing (Aqueous Extract of Saussurea Medusa; AESM) be medicine; with the best positive contrast of type ii diabetes medicine glimepiride (glimepiride) of global sales; research has also disclosed AESM at blood sugar lowering; blood lipid regulation; protecting myocardial cell and liver; improve liver function; prevent aspects such as fatty liver formation, have tangible curative effect.
Advantage of the present invention: (1) is compared with the chemicals of treatment type ii diabetes, AESM life-time service toxic and side effects is little, be not only simple blood sugar lowering concentration, and energy cholesterol reducing, triglyceride and low density lipoprotein, LDL (LDL), the prevention hyperlipidemia, the energy protecting myocardial cell can improve liver function, prevents the formation of fatty liver; (2) compare with insulinize, AESM can improve Insulin receptor INSR sensitivity, makes in the blood insulin level reduce to normal level, and takes medicine conveniently; (3) because the present invention adopts modern biotechnology, obtain medicine, protected rare Herba Saussureae Involueratae plant, also solved the limited problem of rare medicine plant origin, and can guarantee the homogeneity of product well by the method for culture plant cell.
The invention will be further described below in conjunction with the drawings and specific embodiments, all any this areas of having done according to the disclosure of invention be equal to replacement, all belong to protection scope of the present invention.
Description of drawings
Fig. 1-A is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can reduce the blood glucose of KK/Ay type ii diabetes model mice and improve its carbohydrate tolerance result 1: continuous oral administration 28d, glimepiride group and AESM group mouse blood sugar are controlled at normal level, every group of each 10 mice, compare with model group (normal saline group), significant difference (P＜0.001 is represented with * * *) is arranged.
Fig. 1-B is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can reduce the blood glucose of KK/Ay type ii diabetes model mice and improve its carbohydrate tolerance result 2: the carbohydrate tolerance experiment shows, ((2g/kg), glimepiride group and AESM group can be reduced to normal level with blood sugar concentration rapidly behind injectable dextrose monohydrate.Every group of each 10 mice are compared with model group (normal saline group), and significant difference (* * * P＜0.001, * * P＜0.01) is arranged;
Fig. 1-C is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can reduce the blood glucose of KK/Ay type ii diabetes model mice and improve its carbohydrate tolerance result 3: behind successive administration 28d, put to death mice and get hematometry serum insulin level, glimepiride group and AESM group all maintain normal level.Every group of each 10 mice are compared with model group (normal saline group), and significant difference is arranged.(**P＜0.01)。
The blood fat that Fig. 2 can effectively regulate KK/Ay type ii diabetes model mice for the boiling water of oral saussurea medusa culture leaching thing (AESM) is figure as a result.
Fig. 3 can effectively protect the myocardial cell figure of KK/Ay type ii diabetes model mice for the boiling water leaching thing (AESM) of oral saussurea medusa culture.
Fig. 4-A is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can effectively improve the liver function of KK/Ay type ii diabetes model mice and prevent that fatty liver from forming result 1: the treatment terminal point is respectively organized KK/Ay mouse liver form.
Fig. 4-B is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can effectively improve the liver function of KK/Ay type ii diabetes model mice and prevent that fatty liver from forming result 2: the treatment terminal point is respectively organized KK/Ay mouse liver tissue slice.
Fig. 4-C is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can effectively improve the liver function of KK/Ay type ii diabetes model mice and prevent that fatty liver from forming result 3: the treatment terminal point is respectively organized KK/Ay mice serum aspartic transaminase (AST) and alanine aminotransferase (ALT) activity.
Fig. 4-D is that the boiling water leaching thing (AESM) of oral saussurea medusa culture can effectively improve the liver function of KK/Ay type ii diabetes model mice and prevent that fatty liver from forming result 4: the treatment terminal point is respectively organized the concentration of glycogen in the KK/Ay mouse liver.
The specific embodiment
The preparation of the boiling water leaching thing (AESM) of embodiment 1 saussurea medusa culture
The culture (Chinese invention patent ZL02156868.5, preserve this chamber) of getting the fresh saussurea medusa TUIP-8 of 50g shake-flask culture grinds extruding in mortar, the juice that grinds out is collected separately.Get the 500mL deionized water and boil, the saussurea medusa culture that grinds is poured in the boiling water, stirred 2 minutes, it is online to be poured on 4 layer of 300 order stainless steel silk immediately, juice is collected in extruding, and mixes with the juice that grinds out, promptly prepares the boiling water leaching thing (AESM) of saussurea medusa culture.
The AESM branch is filled in the 10mL teat glass, frozen in-20 ℃.With preceding thawing and disposable finishing using.
Embodiment 2AESM is used for the treatment of type ii diabetes animal pattern KK/Ay mice
Buy type ii diabetes animal pattern KK/Ay mice from Chinese Academy of Medical Sciences's Experimental Animal Center, continuous illumination was raised separately in 12 hours under 25 ℃ of conditions, arbitrarily the higher fatty acid high heat food of feed.Mouse blood sugar filters out the high KK/Ay mice of blood sugar concentration after measured, and is divided into 3 groups at random, and 10 every group, male and female half and half.(1) model group (negative control group), every day oral (irritate feed) normal saline, dosage is a 2.5ml/100g body weight/mice.(2) Saussurea medusa (AESM) treatment group, every day, the boiling water of 1 oral (irritate and feed) 1ml saussurea medusa culture leached thing.(3) glimepiride treatment group (positive controls), every day, 1 filling was fed, and each dosage is 0.5mg/kg/.Get tail vein weekly once, with GLUCOCARDTM Test (Arkray Inc.Japan) kit measurement blood sugar concentration.Treatment was put to death mice, the curative effect of blood sampling and tissue-estimating AESM after 28 days continuously.
Blood glucose of embodiment 3 AESM may command type ii diabetes model KK/Ay mices and insulin level successive administration 28 days, monitor the blood sugar concentration of KK/Ay mice weekly, experiment shows, oral AESM can be fine the blood sugar level (Fig. 1-A and table 1) of control KK/Ay mice, the model group mouse blood sugar maintains the hyperglycemia state of 20-28mmol/L always, treat through glimepiride, the blood sugar concentration of this group mice maintains the reduced levels about 12mmol/L always, and the mouse blood sugar of Saussurea medusa group remains on the level of 8-10mmol/L always, this blood sugar concentration is in close proximity to normal level (7-8mmol/L), with model group significance difference (P＜0.001) is arranged, the level of its blood sugar control also is better than the glimepiride group.
Table 1 Herba Saussureae Involueratae cell culture water extract is to the influence of KK/Ay mouse blood sugar
| Group | Blood sugar concentration (mmol/L) | |||||
| Time (my god) | 0 | 7 | 14 | 21 | 28 | |
| Normal saline group glimepiride group Herba Saussureae Involueratae group | 20.56±8.84 21.00±8.10 19.63±8.57 | 19.36±4.58 13.50±4.16 11.29±4.59 | 23.96±4.60 12.97±3.42 10.75±4.24 | 26.85±3.51 12.90±2.15 12.02±3.47 | 21.61±3.30 13.03±2.87 9.84±2.72 | |
The carbohydrate tolerance experiment is (Glucose Tolerance Test) result show: AESM is blood sugar lowering level (Fig. 1-B and table 2) rapidly.The method of carbohydrate tolerance experiment: the laboratory animal fasting is 16 hours before the experiment, intravenous injection glucose (2g/kg) afterwards, respectively from the tail vein 0,30,60 and 120 minutes measuring blood sugar of blood extracting concentration.Fig. 1-B shows that the intravenous injection glucose is after 30 minutes, and it is the highest that blood sugar concentration reaches, just descend afterwards, still greater than 16mmol/L, and glimepiride and AESM group is in the time of 120 minutes in the time of 120 minutes for model group, and blood sugar concentration all drops to the normal level of 7-8mmol/L.
Table 2 Herba Saussureae Involueratae cell culture water extract is to the influence of KK/Ay mice oral glucose tolerance
| Group | Blood sugar concentration (mmol/L) | ||||
| Time (minute) | 0 | 30 | 60 | 120 | |
| Normal saline group glimepiride group Herba Saussureae Involueratae group | 11.45±3.47 8.71±2.02 6.57±1.50 | 23.48±3.92 14.89±5.67 14.67±6.52 | 16.31±4.13 10.13±4.25 8.96±2.82 | 13.71±3.29 7.57±2.63 6.49±1.42 | |
The level that mices are got the hematometry serum insulin was put to death in treatment back in 28 days, adopted radioimmunity test kit (radioimmuno-assay kit) to measure, determination step by shop instruction carry out (Linco Research, St.Charles, MI).The result shows, the insulin level of glimepiride group and AESM group significantly is lower than model group (Fig. 1-C and table 3), serum insulin level height is the type ii diabetes typical characteristic, be that type ii diabetes is a non-insulin-dependent, the blood sugar concentration height is not that hypoinsulinism causes, but, relevant cell causes that this result also points out glimepiride and AESM to have insulin-sensitizing effect because descending to the sensitivity of insulin.
Table 3 Herba Saussureae Involueratae cell culture water extract is to the influence of insulin concentration and free fatty acid in the KK/Ay mouse blood
| The normal saline group | The glimepiride group | The Herba Saussureae Involueratae group | Pvalue | |
| Insulin concentration (μ U/ml) free fatty acid (μ mol/l) | 21.60±4.67 739.72±361.19 | 7.65±3.20 430.11±224.54 | 8.69±2.10 444.58±174.23 | 0.0003 0.03 |
28 days treatment backs are put to death mice and are got triglyceride in the hematometry serum (TG) (TG E-test, Wako), cholesterol (CHO), high density lipoprotein (HDL), low density lipoprotein, LDL (LDL) and the free fatty acid of fatization (NEFA) (NEFA C-test not, Wako) concentration, assay method are all undertaken by the method that test kit manufacturer provides.Glimepiride group and AESM group all can significantly reduce TG, CHO, LDL and NEFA concentration (table 3), improve the concentration of HDL, compare with model group (normal saline group), significant difference (* P＜0.05 is arranged, * P＜0.01, * * * P＜0.001), see shown in Fig. 2 and the table 4.
High density lipoprotein (HDL) concentration height can effectively prevent arteriosclerotic formation, and TG, CHO, LDL and NEFA concentration height all can promote arteriosclerosis, cause cardiovascular and cerebrovascular disease.According to statistics, the type ii diabetes patient generally is an improper diet, and Excessive Intake fat and heat food cause that the ill initial stage is fat mostly, and the later stage is understood concurrent cardiovascular and cerebrovascular disease, hyperlipidemia.In fact the mortality rate that causes of the concurrent cardiovascular and cerebrovascular disease of type ii diabetes accounts for 80%, and makes its life expectancy reduce 1/3; Diabetics merges blood fat rising person more than 80%, and blood viscosity rising person reaches 90%.Therefore, can glucose level control, can significantly improve blood lipid metabolism again and make the saussurea medusa culture become the Chinese medicine of a kind of very potential treatment type ii diabetes and blood fat reducing.
Table 4 Herba Saussureae Involueratae cell culture water extract is to the regulating action of KK/Ay mice blood fat
| The normal saline group | The glimepiride group | The Herba Saussureae Involueratae group | Pvalue | |
| CHO(mmol/L) TG(mmol/L) LDL(mmol/L) HDL(mmol/L) | 5.32±1.64 2.06±0.70 8.66±2.80 0.30±0.07 | 4.21±1.47 1.03±0.40 3.28±1.47 0.48±0.13 | 3.49±0.92 1.04±0.39 3.38±1.35 0.38±0.09 | 0.02 0.003 0.0003 0.07 |
The concentration that mice is got lactic acid dehydrogenase in the hematometry serum (LDH) and creatine kinase (CK) was put to death in the treatment back in 28 days, assay method all carries out (LDH by the method that test kit manufacturer provides, the CK external diagnosis reagent case, Zhongsheng Beikong Biological Science ﹠ Technology Co., Ltd.).Glimepiride group and AESM group all can significantly reduce the concentration (* * P＜0.01, * * * P＜0.001) of LDH and CK in the serum, sees shown in Fig. 3 and the table 5.LDH and CK are the important indicators that characterizes the myocardial cell function.The type ii diabetes patient generally can concurrent cardiovascular and cerebrovascular disease, and blood supply insufficiency can cause myocardium cell necrosis, and myocardium cell necrosis can discharge a large amount of LDH and CK, and especially CK is to make a definite diagnosis the topmost biochemical indicator of acute myocardial infarction.AESM can significantly reduce CK concentration, reflects that AESM has good protective action to myocardial cell.
Table 5 Herba Saussureae Involueratae cell culture water extract is to the myofunctional influence of KK/Ay mouse core
| The normal saline group | The glimepiride group | The Herba Saussureae Involueratae group | Pvalue | |
| Lactic acid dehydrogenase (U/L) creatine kinase (U/L) | 1224.38±239.30 780.00±467.16 | 39.13±12.03 183.00±18.21 | 36.75±10.73 171.38±21.48 | 0.01 0.002 |
28 days treatment backs are put to death mice and are got hematometry serum Mid-Heaven Gate winter propylhomoserin transaminase (AST) and alanine aminotransferase (ALT) activity, assay method all carries out (AST by the method that test kit manufacturer provides, the ALT external diagnosis reagent case, Zhongsheng Beikong Biological Science ﹠ Technology Co., Ltd.).Get liver, a part of homogenate, the content of glycogen and triglyceride in the mensuration liver.Another part hepatic tissue is fixed with 10% neutral formalin, uses paraffin embedding, and section is with h and E (hematoxylinand eosin, H﹠amp; E) its form is examined under a microscope in dyeing.
During treatment in 28 days, transaminase's's---aspartic transaminase (AST) and alanine aminotransferase (ALT)---of two kinds of reflection liver functions activity is all at normal level (table 6) in glimepiride group and the AESM group serum, and two kinds of transaminases' of model mice level is all higher, exceed range of normal value (Fig. 4-C), show that AESM has the effect that improves liver function.
Table 6 Herba Saussureae Involueratae cell culture water extract is to the influence of KK/Ay Mouse Liver function
| The normal saline group | The glimepiride group | The Herba Saussureae Involueratae group | Pvalue | |
| GPT(U/L) GOT(U/L) | 76.88±20.90 287.00±70.87 | 15.67±4.46 864.00±124.89 | 12.34±3.67 871.88±245.78 | 0.0003 0.0006 |
Dissect mice and show, the obvious enlargement of the liver of model group mice, and color and luster turns white visible fibrosis of naked eyes and fatization.As seen hepatic tissue dyes, and the model group mouse liver has very tangible fat to drip, and shows that the liver of KK/Ay type ii diabetes model mice has the tendency that forms fatty liver.The liver volume of glimepiride treatment group and weight are less than model group, but organize greater than AESM, the liver color and luster of AESM group is dark red, be normal liver tissue, hepatic tissue dyeing shows that also the hepatic tissue of AESM treatment group is normal, does not have fat to drip, the phenomenon that does not also have fatization, its effect obviously are better than glimepiride treatment group (Fig. 4-A, Fig. 4-B).Measure glycogen content in the liver organization, AESM group content is the highest, shows that liver obviously strengthens (* P＜0.05, * * P＜0.01, * * * P＜0.001) with the ability that unnecessary sugar part is converted into glycogen, sees Fig. 4-D and table 7.Therefore, AESM has the liver function of improving KK/Ay type ii diabetes model mice of highly significant and prevents the curative effect that fatty liver forms.
Table 7 Herba Saussureae Involueratae cell culture water extract is to the influence of glycogen and triglyceride content in the KK/Ay mouse liver tissue
| The normal saline group | The glimepiride group | The Herba Saussureae Involueratae group | Pvalue | |
| Hepatic tissue glycogen content (mg/g hepatic tissue) hepatic tissue triglyceride content (mg/g liver) | 4.33±1.60 7.41±1.09 | 6.91±1.54 4.24±1.22 | 9.99±1.02 3.78±0.99 | 0.0004 0.0002 |
Claims (10)
1. the saussurea medusa of Pei Yanging is at preparation treatment type ii diabetes and prevent purposes in the medicine of its complication.
2. purposes according to claim 1 is characterized in that: the saussurea medusa of described cultivation is to utilize biotechnology, makes the callus producer of Herba Saussureae Involueratae suddenly change the saussurea involucrata cell line of the stable high yield Herba Saussureae Involueratae flavone of acquisition by induced mutations.
3. purposes according to claim 2 is characterized in that: described induced mutations comprises the mutation of 60Co-radiation gamma, temperature mutation and ultraviolet mutagenesis.
4. purposes according to claim 1 is characterized in that: described saussurea medusa is saussurea involucrata cell line TUIP-8, and it protects minus sign is that CGMCC No.0855 and saussurea medusa are SMXL-615, and its preserving number is CGMCC No.1072.
5. the saussurea medusa extract of Pei Yanging is at preparation treatment type ii diabetes and prevent purposes in the medicine of its complication.
6. purposes according to claim 5 is characterized in that: the saussurea medusa of described cultivation is to utilize biotechnology, makes the callus producer of Herba Saussureae Involueratae suddenly change the saussurea involucrata cell line of the stable high yield Herba Saussureae Involueratae flavone of acquisition by induced mutations.
7. purposes according to claim 6 is characterized in that: described induced mutations comprises the mutation of 60Co-radiation gamma, temperature mutation and ultraviolet mutagenesis.
8. purposes according to claim 5 is characterized in that: described saussurea medusa is saussurea involucrata cell line TUIP-8, and it protects minus sign is that CGMCC No.0855 and saussurea medusa are SMXL-615, and its preserving number is CGMCC No.1072.
9. purposes according to claim 5, it is characterized in that: the saussurea medusa extract of described cultivation makes by the following method: the fresh saussurea medusa culture of getting certain mass, squeezing is also collected the juice that obtains, residue Saussurea medusa cell culture is added in the boiling water that 8-12 doubly measures, stirred 1-3 minute, filter immediately, the juice that juice and the squeezing that obtains obtained merges promptly.
10. according to any one described purposes among the claim 1-9, it is characterized in that: described treatment type ii diabetes and prevent its complication to be meant blood sugar lowering, blood lipid regulation, protecting myocardial cell and liver, improve liver function and prevent that fatty liver from forming.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101904887A (en) * | 2010-07-13 | 2010-12-08 | 大连普瑞康生物技术有限公司 | Application of Saussurea involucrata culture in food and anti-inflammatory analgesic medicine |
| CN102579818A (en) * | 2012-03-26 | 2012-07-18 | 次旺 | Traditional Tibetan medicine composition, preparation method thereof and application |
| CN108245553A (en) * | 2016-12-28 | 2018-07-06 | 兰州奇正生态健康品有限公司 | A kind of hypoglycemic composition and preparation method thereof and purposes |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1212057C (en) * | 2004-02-10 | 2005-07-27 | 北京智创信达科技发展有限公司 | Saussurea medusa Maxim cell line for stable high yielding of snow lotus flavone and its uses |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101904887A (en) * | 2010-07-13 | 2010-12-08 | 大连普瑞康生物技术有限公司 | Application of Saussurea involucrata culture in food and anti-inflammatory analgesic medicine |
| CN102579818A (en) * | 2012-03-26 | 2012-07-18 | 次旺 | Traditional Tibetan medicine composition, preparation method thereof and application |
| CN102579818B (en) * | 2012-03-26 | 2014-01-08 | 次旺 | Traditional Tibetan medicine composition, preparation method thereof and application |
| CN108245553A (en) * | 2016-12-28 | 2018-07-06 | 兰州奇正生态健康品有限公司 | A kind of hypoglycemic composition and preparation method thereof and purposes |
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