CN112656827A - Radix araliae armatae total saponin and application thereof in preparation of medicine for treating restenosis after vascular injury - Google Patents
Radix araliae armatae total saponin and application thereof in preparation of medicine for treating restenosis after vascular injury Download PDFInfo
- Publication number
- CN112656827A CN112656827A CN202110029784.0A CN202110029784A CN112656827A CN 112656827 A CN112656827 A CN 112656827A CN 202110029784 A CN202110029784 A CN 202110029784A CN 112656827 A CN112656827 A CN 112656827A
- Authority
- CN
- China
- Prior art keywords
- total saponins
- beta
- armata
- aralia
- formula
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
- A61P9/08—Vasodilators for multiple indications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/25—Araliaceae (Ginseng family), e.g. ivy, aralia, schefflera or tetrapanax
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
- A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
- A61P9/14—Vasoprotectives; Antihaemorrhoidals; Drugs for varicose therapy; Capillary stabilisers
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
Abstract
The invention provides total saponins of aralia armata and application thereof in preparing a medicine for treating restenosis after vascular injury, belonging to the technical field of vascular medicines. The invention uses the aralia armata L dry root bark coarse powder as a material to carry out reflux extraction by using 70-80% ethanol water solution, extracts are sequentially extracted by using petroleum ether, trichloromethane or dichloromethane and water saturated n-butyl alcohol, and supernatant is taken to remove a solvent and is evaporated to dryness to obtain total saponins of aralia armata L. Based on the characteristics that the total saponins of radix araliae armatae can effectively inhibit the hyperplasia of intima of blood vessels, inhibit the proliferation and/or metastasis of smooth muscle cells of blood vessels and inhibit the expression of protein molecules in a Wnt signal pathway, the invention provides the application of the total saponins of radix araliae armatae in preparing a medicine for treating restenosis after vascular injury.
Description
Technical Field
The invention belongs to the technical field of vascular drugs, and particularly relates to total saponins of radix araliae armatae and application thereof in preparation of drugs for treating restenosis after vascular injury.
Background
Since vascular injury frequently occurs in the lumen, research on restenosis of blood vessels after injury focuses on intima and media, and although a great deal of research results are obtained, the exact mechanism is not clear at present, and most scholars have a consensus: after vascular injury, proliferation and migration of intima and media are important factors in restenosis following vascular injury.
There are many kinds of restenosis after vascular injury, for example, patent publication No. CN104174010A discloses that the knockout of SHPS1 gene can significantly promote neointima and cell proliferation; transforming growth factor beta 1 plays a role in the formation of restenosis after vascular injury; the compound Stichopus japonicus mucopolysaccharide has effects of promoting proliferation and apoptosis of blood vessel intima in rabbit with restenosis after blood vessel injury; matrix metalloproteinases, for example, are also involved in the process of restenosis following arterial injury.
The regulatory role of the Wnt signaling pathway in vascular restenosis is a new area of research. The research of organized embryo development shows that the Wnt signal channel is in an activated state in the early formation stage of Vascular Smooth Muscle Cells (VSMC), and the research of Quansnichka and the like finds that the expression of CyclinD1 is reduced after inhibiting the Wnt canonical channel, so that the proliferation of VSMC is reduced; the wang et al study showed activation of the β -catenin signal following injury to the rat carotid artery. Further, researchers have found that upstream signaling molecules such as Wnts/Fzds, Dvl, GSK-3B are also involved in smooth muscle cell proliferation after vascular injury. Therefore, the Wnt/beta-catenin signaling pathway may be involved in the vascular development or proliferation after injury.
Radix araliae armatae is a common plant in Zhuang nationality, and root bark is widely used as a traditional medicine for treating cardiovascular diseases, chronic nephritis, prostatitis, edema, rheumatic arthralgia and other diseases. However, in the prior art, no report about the cardiovascular protection effect of the radix araliae armatae exists.
Disclosure of Invention
In view of the above, the present invention aims to provide total saponins of aralia armata and applications thereof in preparation of drugs for treating restenosis after vascular injury.
The invention provides aralia armata total saponin which is prepared by the following steps:
mixing the aralia armata dry root bark coarse powder with an ethanol water solution with the volume concentration of 70-80%, performing reflux extraction, collecting an extracting solution, concentrating to obtain an extract, re-dissolving the extract in water, extracting with petroleum ether, collecting a water phase, removing fat-soluble components, mixing with water-saturated n-butyl alcohol, extracting, taking supernate, removing the n-butyl alcohol, and evaporating to dryness to obtain total saponins of aralia armata.
Preferably, the mass ratio of the aralia armata dry root bark coarse powder to the volume ratio of the ethanol water solution is 1 g: 6-10 mL.
Preferably, the temperature of the reflux extraction is 60-80 ℃; the reflux extraction time is 1-1.5 h.
Preferably, the relative density of the extract at 60 ℃ is 1.15-1.20.
Preferably, the method for removing fat-soluble components is extraction by using dichloromethane or trichloromethane.
Preferably, the volume ratio of the dichloromethane or the trichloromethane to the water, the petroleum ether and the water-saturated n-butanol is 1: 1: 1: 1.
preferably, the total saponins of radix araliae armatae comprise the following components: 1-O- [ (2 alpha, 3 beta, 5 xi, 9 xi, 18 xi, 19 alpha) -2,3,19, 23-Tetrahydroxy-28-oxolean-12-en-28-yl ] -beta-D-glucopyranose with the structural formula I, 1-O- [ (2 alpha, 3 beta, 5 xi, 9 xi, 18 xi, 19 alpha) -2,3,19,23, 24-Pentahydroxy-28-oxolean-12-en-28-yl ] -beta-D-glucopyranose with the structural formula II, 1-O- [ (3 beta, 5 xi, 9 xi, 18 xi, 19 alpha) -3,19, 24-Tetrahydroxy-24, 28-dioxirs-12-en-28-yl ] hexopyranose with the structural formula III, 3 beta with the structural formula IV, 5 ξ,9 ξ) -3- { [2-O- (. beta. -D-Glucopyranosyl) -. beta. -D-Glucopyranosyl ] oxy } -23-hydroxyazulene-12-en-28-oic acid, 1-O- [ (2. beta., 3. beta., 5. zeta., 9. zeta., 18. zeta.) 3- (. beta. -D-glucopyranosyloxy) -2, 23-dihydroxy-28-oxolene-12-en-28-yl ] -P-D-glucopyranose of the formula V, 6-Deoxy-. alpha. -L-mannopyranosyl- (1- >3) - [ beta. -D-Glucopyranosyl- (1- >6) ] -1-O- [3,27-dihydroxy-27,28-dioxolup-20(29) -en-28-yl ] -beta-D-glucopyranose, the structural formula of which is (3 beta, 5 xi, 9 xi, 18 xi) -28-Hydroxy-28-oxole-12-en-3-yl 6-deoxy-alpha-L-mannopyranosyl- (1- >3) - [ beta-D-glucopyranosyl- (1- >2) ] -beta-D-glucopyranoside acid, the structural formula of which is 1-O- (1,3,23,24,25-Pentahydroxy-28-oxo-9, 19-cyclolactostan-28-yl) heptapyranose, the structural formula of which is Dixigon;
the invention provides application of total saponins of radix araliae armatae in preparation of a medicine for treating restenosis after vascular injury.
The invention provides application of total saponins of radix araliae armatae in preparing a medicament for inhibiting vascular intimal hyperplasia.
The invention provides application of total saponins of radix araliae armatae in preparing a medicine for inhibiting proliferation and/or metastasis of vascular smooth muscle cells.
The invention provides application of total saponins of aralia armata in preparing an inhibitor for inhibiting expression of protein molecules in a Wnt signal path.
The invention provides aralia armata total saponin which is prepared by the following steps: extracting total saponins from the dry radix araliae armatae bark coarse powder serving as an extraction material by using an ethanol water solution with the volume concentration of 70-80%, performing reflux extraction, concentrating a collected extracting solution to obtain an extract, re-dissolving the extract in water, extracting the extract by using petroleum ether to remove impurities such as pigments and the like, retaining a water phase, mixing and extracting by using water-saturated n-butyl alcohol to obtain saponin components with higher polarity, taking supernate to remove the n-butyl alcohol, and evaporating to dryness to obtain the total saponins of radix araliae armatae. The total saponins of radix araliae armatae prepared by the method have high extraction rate and high content of effective components (6-7 percent), and have the characteristics of simple and rapid extraction method.
The invention provides application of total saponins of radix araliae armatae in preparation of a medicine for treating restenosis after vascular injury. The invention proves that the total saponins of radix araliae armatae can relieve neointimal hyperplasia after vascular injury for the first time; meanwhile, the total saponins of radix araliae armatae can remarkably reduce the neointima formation after the artery injury, and the ratio of the vascular infarction is obviously lower than that of a model. In order to prove that the total saponins of radix araliae armatae have effective inhibition effect on the growth of VSMC, the expression conditions of alpha-SMA and PCNA at the damaged blood vessel are detected by immunohistochemistry, VSMCs cells are cultured in vitro to carry out CCK-8 detection and a single-layer cell wound healing experiment, and the influence of the total saponins of radix araliae armatae on the proliferation and migration of VSMCs induced by serum is evaluated. The result shows that the total saponins of radix araliae armatae have obvious inhibition effect on the proliferation and migration of VSMCs induced by serum. Therefore, in vivo and in vitro experimental results prove that the total saponins of radix araliae armatae have the effect of improving restenosis after vascular injury. Meanwhile, in order to deeply research the mechanism of inhibiting VSMC proliferation and migration by the total saponins of radix araliae armatae, and simultaneously detect the expression conditions of signal molecules on a wnt signal path and downstream, the results show that when VSMCs proliferate under serum induction, the mRNA and protein expressions of genes wnt3a, GSK-3B, Dvl-1 and beta-catenin are obviously increased, and when the total saponins of radix araliae armatae inhibit cell proliferation induced by FBS, the mRNA and protein expressions of wnt3a, GSK-3B, Dvl-1 and catenin are also reduced. The effect of the total saponins of aralia armata on the down-regulation of the expression levels of beta-catenin and Gsk-3 beta is also proved in animal experiments. In addition, the content of Cyclin is rapidly changed along with the circulation of a cell cycle, the cell in a static state does not express Cyclin D1, when the cell is stimulated by growth factors and the like, Cyclin D1 is firstly expressed, Cyclin D1 is then combined with Cyclin-dependent kinase to phosphorylate the Cyclin-dependent kinase, and the Cyclin D1 is a downstream target gene of Dvl-1, so that the cell enters a cell proliferation state. In an in vitro cell experiment, the olecranon total saponins inhibit the high expression of a serum-induced wnt signal molecule and also down-regulate the expression of Cyclin D1, so that the change of the expression level of Cyclin D1 further supports the conclusion that the olecranon total saponins can inhibit the hyperplasia of intima after vascular injury by down-regulating a wnt signal pathway.
Drawings
FIG. 1 shows the compound and structural formula contained in the total saponins extracted from radix araliae armatae of the present invention;
FIG. 2 is the results of in vivo experiments with total saponins of Artabotrys hexapetalus in administration treatment, wherein FIG. 2a is a morphology chart of HE stained and immunohistochemical cell sections; FIG. 2b is the statistics of the infarction rates of the blood vessels in each group; FIG. 2c is a statistical result of positive expression conditions of α -SMA, PCNA, β -catenin, and Gsk-3 β;
FIG. 3 shows the results of in vitro smooth muscle cell assays; FIG. 3a shows the toxicity of total saponins of radix araliae armatae to normally growing smooth muscle cells at each dose; FIG. 3b is a graph of the effect of each dose of total saponins of Artabotrys on FBS-induced smooth muscle cell proliferation; figure 3c is a graph of the effect of total saponins of aralia armata on serum-induced VSMC wound healing at 24 hours post-injury; fig. 3d is a statistical result of the healing degree of total saponins of aralia armata on serum-induced VSMC; FIG. 3e is a diagram showing RT-qPCR detection of expression of critical protein molecules of Wnt signaling pathway of vascular smooth muscle cells induced by FBS in the high-dose group of total saponins of aralia armata;
FIG. 4 shows the result of detecting Wnt signaling pathway molecular protein in cells by Western blot; FIG. 4a is an electrophoresis diagram for detecting Wnt signaling pathway molecular proteins of cells in an administration group of total saponins of radix araliae armatae; fig. 4b is a statistical graph of the expression of Wnt signaling pathway molecular proteins induced by inhibition of 15% FBS by total saponins of aralia armata.
Detailed Description
The invention provides aralia armata total saponin which is prepared by the following steps: mixing the aralia armata dry root bark coarse powder with an ethanol water solution with the volume concentration of 70-80%, performing reflux extraction, collecting an extracting solution, concentrating to obtain an extract, re-dissolving the extract in water, extracting with petroleum ether, collecting a water phase, removing fat-soluble components, mixing with water-saturated n-butyl alcohol, extracting, taking supernate, removing the n-butyl alcohol, and evaporating to dryness to obtain total saponins of aralia armata.
The invention takes the dry root bark of radix araliae armatae as an extraction material to extract the total saponins of the radix araliae armatae, and compared with other parts of the radix araliae armatae, the dry root bark of the radix araliae armatae has obvious advantages in the total extraction rate of the total saponins of the radix araliae armatae. The particle size of the aralia armata dry root bark coarse powder is preferably 10-50 meshes, and more preferably 24 meshes. The radix araliae armatae dry root bark coarse powder is beneficial to improving the extraction rate of active ingredients in medicinal materials. The source of the eagle-leaf dried root bark is not particularly limited in the invention, and the eagle-leaf dried root bark can be obtained from commercial materials of the eagle-leaf dried root bark well known in the field. In the embodiment of the invention, the sources of the aralia armata dry root bark are purchased from Guangxi Xianzhu Chinese medicine science and technology limited company.
The invention takes ethanol water solution with volume concentration of 70-80% as extraction solvent. Compared with other extraction solvents (such as water), the method for extracting the total saponins of radix araliae armatae by using the ethanol water solution has the characteristic of high extraction rate. Meanwhile, the invention specifically limits the volume concentration of the ethanol water solution to be preferably 75-80%, and the total saponins of radix araliae armatae in the medicinal materials can be fully extracted as far as possible under the concentration. The mass ratio of the aralia armata dry root bark coarse powder to the volume ratio of the ethanol water solution is preferably 1 g: 6-10 mL, more preferably 1 g: 7-9 mL, most preferably 1 g: 8 mL.
In the invention, the temperature of the reflux extraction is preferably 60-80 ℃, and more preferably 70 ℃. The time for reflux extraction is preferably 1-1.5 h, and more preferably 1.2 h. The concentration is preferably concentration under reduced pressure. The method of concentration under reduced pressure in the present invention is not particularly limited, and a method of concentration under reduced pressure known in the art may be used. The decompression concentration can remove the redundant water in the extracting solution on one hand and is beneficial to removing the volatile components in the extracting solution on the other hand. And preferably, the reduced pressure concentration is finished when the relative density of the extract reaches 1.15-1.20 at the temperature of 60 ℃.
In the invention, after the extract is obtained, the extract is re-dissolved in water and then extracted by petroleum ether. The dosage of the water is that every 200g of extract prepared from the aralia armata dry root bark coarse powder is redissolved by 500mL of water. And extracting the re-dissolved extract by using petroleum ether. The petroleum ether extraction is beneficial to removing pigment components in the extract. The extraction is preferably performed using a separatory funnel.
In the present invention, after extraction, the lower aqueous phase collected is freed from fat-soluble components. The method for removing fat-soluble components is preferably extraction with dichloromethane or chloroform.
In the invention, fat-soluble components are removed and then mixed with water saturated n-butyl alcohol for extraction, the supernatant is taken to remove the n-butyl alcohol, and the obtained product is evaporated to dryness to obtain the total saponins of radix araliae armatae. The volume ratio of the dichloromethane or trichloromethane to the water, petroleum ether and water-saturated n-butanol is preferably 1: 1: 1: 1. the water-saturated n-butanol has the following functions: firstly, the extraction of saponin components with larger polarity is facilitated; and secondly, the n-butyl alcohol saturated by water can increase the solubility of the n-butyl alcohol, is convenient to separate and is not easy to emulsify. The extraction with water-saturated n-butanol is preferably repeated twice. And (3) combining the two obtained extraction liquids, preferably recovering the solvent by rotary evaporation, preferably evaporating the extraction liquid by a water bath to dryness, and obtaining a dry extract, namely the total saponins of radix araliae armatae. The storage of the total saponins of aralia armata is preferably kept in brown bottles. The total saponins of radix araliae armatae prepared by the method have high extraction rate which is not less than 7%. The total saponins of aralia armata are detected by UPLC Q-active Orbitrap MS technology, the obtained data are subjected to database retrieval comparison (mzCloud, mzVault, ChemSepider) after primary arrangement is finished by CD2.1(Thermo Fisher), and 299 compounds are identified in total, wherein 8 oleanolic acid type triterpenoid saponins and 1 cardiac glycoside compound have the structural formula.
The invention provides application of total saponins of radix araliae armatae in preparation of a medicine for treating restenosis after vascular injury.
In the invention, the administration group of total saponins of radix araliae armatae can obviously improve severe intimal hyperplasia of damaged blood vessels, incomplete repair of intima, discontinuous inner elastic plates, rough inner surface of a lumen, migration and proliferation of a large number of smooth muscle cells from a media to the intima, obvious thickening of the intima, accumulation of a large number of matrixes and obvious narrowing of the lumen; but also can obviously reduce the vascular embolism rate. This indicates that total saponins of radix araliae armatae can inhibit neointimal hyperplasia induced by vascular injury.
The total saponins of radix araliae armatae can inhibit transformation of VSMCs from contraction phenotype to synthesis type in neointima of damaged blood vessel, thereby inhibiting hyperplasia of blood vessel intima. Meanwhile, the detection of the expression condition of PCNA (PCNA), which is an important biological index reflecting cell proliferation, and the semi-quantitative analysis result of immunohistochemistry show that the average optical density of the radix araliae armatae total saponin group is significantly different from that of a model group (P is less than 0.05), so that the radix araliae armatae is prompted to inhibit the proliferation of intimal cells after vascular injury, and therefore, the luminal stenosis is reduced.
Excessive growth of VSMC is thought to be a major factor in causing restenosis from vascular injury. The inhibition effect of the total saponins of radix araliae armatae on VSMCs cell proliferation is determined by detecting the content of CCK-8.
The influence of the total saponins of aralia armata on VSMC migration is evaluated through a wound healing test, and the result shows that the total saponins of aralia armata slow down the wound healing of a monolayer of cells.
The expression condition of key protein molecules of the vascular smooth muscle cell Wnt signal pathway is detected by adopting an RT-qPCR method, and the result shows that the olecranon total saponin inhibits the expression level of related genes of Wnt3 alpha/Dvl-1/beta-catenin pathway. The remarkable effect of the radix araliae armatae on treating the restenosis after the vascular injury is proved from multiple aspects and multiple angles, so that the radix araliae armatae total saponin is used as the only active substance to realize the treatment of the restenosis after the vascular injury.
Based on the experimental results, the invention provides the application of the total saponins of radix araliae armatae in preparing the medicine for inhibiting the intimal hyperplasia of blood vessels. The invention also provides application of the total saponins of aralia armata in preparing a medicament for inhibiting proliferation and/or metastasis of vascular smooth muscle cells and application of the total saponins of aralia armata in preparing an inhibitor for inhibiting expression of protein molecules in a Wnt signal pathway.
The total saponins of aralia armata and the application thereof in the preparation of the medicament for treating restenosis after vascular injury are described in detail in the following examples, but the description should not be construed as limiting the scope of the invention.
Example 1
Taking dry radix araliae armatae, crushing into coarse powder, precisely weighing 200g, adding 2000ml of 80% ethanol, performing reflux extraction, performing reduced pressure concentration to obtain an extract, adding 500ml of double distilled water for sufficient dissolution, transferring to a separating funnel, adding 500ml of petroleum ether for extraction, taking a lower-layer aqueous phase solution in the separating funnel, calculating the volume of an aqueous phase, adding trichloromethane with the same volume, standing after sufficient shaking, taking an upper-layer solution in the separating funnel, adding n-butanol with the same volume, slightly shaking, taking the upper-layer solution, repeating n-butanol extraction for 2 times, combining extraction liquids, performing rotary evaporation to recover a solvent, and performing water bath evaporation to obtain 14g of a dry extract. The extraction yield of the extracted total saponins of radix araliae armatae is 7 percent by calculation. Packaging into brown glass bottle for use.
The total saponins of aralia armata are detected by UPLC Q-active Orbitrap MS technology, the obtained data are subjected to database retrieval comparison (mzCloud, mzVault, ChemSPEder) after primary arrangement is finished by CD2.1(Thermo Fisher), and 299 compounds are identified in total, wherein the structural formulas of 8 oleanolic acid type triterpene saponins and 1 cardiac glycoside compound are shown in figure 1.
Example 2
Method for extracting total saponins of radix araliae armatae with different extraction solvents and comparison
Taking 2 parts of the coarse powder of the aralia armata nakai bark, 200g of each coarse powder, performing reflux extraction on one part of the aralia armata nakai bark by using 70% ethanol water solution, extracting the other part of the aralia armata nakai bark by using water of which the amount is 10 times that of the coarse powder, extracting the other part of the aralia armata nakai bark by using water for 2 times, 1 hour each time, performing the other steps of the method in the embodiment 1), and finally weighing and detecting the.
12.0g of total saponins are obtained in the alcohol extraction scheme, and the extraction rate is 6 percent; the total saponins extracted in the water extraction scheme is 3.0g, and the extraction rate is 1.5%.
Example 3
1. Rat femoral artery injury model and grouping
32 SD rats were housed in cages, acclimatized to 1W, free diet and water. 24 rats were randomly selected to establish a femoral artery endothelial denudation model, and the reference literature is briefly described as follows: rat isoflurane inhalation anesthesia, lower limb skin is incised, femoral artery branches are dissociated and incised, an interventional guide wire (Micro Therapeutics, Inc) with the diameter of 1.00mm is inserted into femoral artery, drawing is carried out for 3 times back and forth to cause the inner cortex of artery to drop, the arteriotomy is ligated after the operation is finished, and the skin is sutured. In the sham group, only skin and artery incisions were made, and no interventional guidewire was inserted. Rats successfully modeled were randomly divided into a model group, an olecranon total saponin low dose group (40mg/kg) and an olecranon total saponin high dose group (80 mg/kg). The drug group is infused with corresponding liquid medicine, and the normal group and the model group are infused with corresponding volume of normal saline (10mL/kg) for 1 time/d, and the total volume is 4W. General conditions including body mass, water and food, stool and urine, hair, etc. were observed in each group of rats during the intervention.
2. Measurement of vascular infarction proportion and results
After the administration, the animals are circularly perfused with neutral formalin at constant pressure for 15min under anesthesia, the femoral artery is cut, washed with normal saline, soaked in formalin for fixation for 24h, the femoral artery and surrounding tissues are cut into segments with the length of 2mm, dehydrated by conventional alcohol, embedded by paraffin, and prepared into blood vessel cross section continuous sections with the thickness of 5 μm. And (3) carrying out HE staining on the tissue section, observing the shape of the blood vessel by a microscope, measuring the perimeter of the blood vessel inner cavity by Image-Pro plus 6.0 Image analysis software, converting the equivalent circular area of the blood vessel inner cavity, measuring the area of the blood vessel inner cavity, and then calculating the blood vessel infarction proportion (inhibition ratio) according to formula I. The measurement was performed by a single blind method, and the measurement was not related to the experiment.
The ratio of the vascular infarction to the equivalent circular area of the endovascular membrane area/the vascular lumen is multiplied by 100 percent by the formula I
Classical histological HE staining analysis showed that the model group had significant structural changes in the vessels and vessel lumen narrowing significantly compared to the sham group. As shown in fig. 2a, in the model group, severe intimal hyperplasia, incomplete intimal repair, discontinuous inner elastic plates, rough inner surface of the lumen, a large amount of smooth muscle cells migrating and proliferating from the media to the intima, obvious intimal thickening, a large amount of matrix accumulation and obvious narrowing of the lumen were observed. The pathological changes of the administration group are obviously reduced, and the internal and external elastic plates are relatively completely preserved. The result of the blood vessel morphometric analysis by using Image Pro plus 6.0 software shows that the proportion of the blood vessel infarction in the sham operation group is very small and is about 12.37 percent, the model group is obviously increased by 97.14 percent, and the model group has a very significant difference (P <0.01) compared with the sham operation group. The ratios of vascular infarction of the high-dose and low-dose olecranon groups are 71.48% and 81.75%, respectively, and the high-dose group has a significant difference compared with the model group (P < 0.05). The radix araliae armatae total saponin can inhibit neointimal hyperplasia induced by vascular injury. The total saponins of radix araliae armatae can improve intimal hyperplasia caused by a femoral artery mechanical injury model.
Example 4
Immunohistochemical detection of expression of alpha-SMA, PCNA, GSK-3 beta, beta-catenin protein
The tissue sections prepared in example 3 were subjected to immunohistochemical staining using the SABC method. Respectively taking anti-alpha-SMA, PCNA, GSK-3 beta and beta-catenin antibodies as primary antibodies, and carrying out other operations according to the kit instructions. The new DAB is used for developing color, and the dyeing degree is controlled under a microscope. PBS was used as a negative control instead of primary antibody. Each section was randomly selected for 5 different fields, the AOI area (area of AOI) and the integrated optical density of positive staining (IOD) were measured using Image-Pro plus 6.0 Image analysis software, and the average optical density (IOD/AOI) was calculated to evaluate the degree of staining in the area.
Numerous studies have shown that VSMCs differentiated from contractile to synthetic types lead to hyperplasia of the intima of vessels by promoting the development of vascular remodeling through cell migration and proliferation, and this experiment examined the expression of the VSMCs contractile phenotype marker protein α -SMA in the neointima of damaged vessels (as shown in FIG. 2 a). Compared with a sham operation group, the expression of the alpha-SMA in the neointima of the rat blood vessel of the model group is obviously reduced, which indicates that VSMCs in the neointima of the rat blood vessel of the model group are subjected to phenotypic transformation from a contraction type to a synthesis type. Compared with the model group, the expression of the alpha-SMA of the radix araliae armatae total saponin group is increased, and the radix araliae armatae total saponin can inhibit the transformation of VSMCs in the neointima of the damaged blood vessel from a contraction phenotype to a synthesis type.
PCNA is an important biological index reflecting cell proliferation, and in order to discuss the anti-proliferation effect of total saponins of radix araliae armatae, the experiment detects the expression condition of PCNA in damaged blood vessels. The immunohistochemical result shows (as shown in fig. 2 a), no PCNA staining positive cells are seen in the sham operation group, the smooth muscle cells of the neointima of the model group are disorderly and dispersedly thickened, the PCNA positive cells are massively expressed and are deeply stained, the thickening degree of the vascular intima of the olecranon total saponin group is obviously reduced compared with that of the model group, and a small amount of PCNA positive cells are seen only in the neointima close to the lumen, and the staining degree is lighter. The semiquantitative analysis of immunohistochemistry shows that the average optical density of the radix araliae armatae total saponin group is significantly different from that of a model group (P is less than 0.05), and the radix araliae armatae is prompted to inhibit the proliferation of intimal cells after vascular injury, so that the luminal stenosis is reduced.
Example 5
1. Vascular smooth muscle cell culture and drug treatment
The culture was isolated from rat aorta by enzyme digestion. Anaesthetizing animals, opening the thoracic cavity, flushing the blood in the aorta from the left ventricle by perfusion, removing perivascular fat and connective tissue, taking out the thoracic aorta, placing into a mixed enzyme digestive fluid (HBSS, containing collagen II 1mg/ml, Elastase 0.744U/ml, Soybean Trypsin Inhibitor 1mg/ml and BSA 2mg/ml) prepared with Hanks balanced salt, incubating at 37 ℃ for 15min, carefully tearing off the outer membrane with forceps, gently scraping off the inner membrane with a cotton swab, cutting the obtained middle membrane tissue into pieces, adding fresh mixed enzyme digestive fluid, incubating at 37 ℃ for 2-3 h, gently blowing the digested tissue into a cell suspension, centrifuging at 1000rpm for 5min to collect cells, adding a culture solution containing 10% of cell to resuspend the cells, inoculating into an FBS culture bottle, after the cells are fully grown, digesting the EDTA with 0.125% Trypsin/FBS. The cultured smooth muscle cells were morphologically observed under an inverted phase contrast microscope, immunocytochemically stained with a monoclonal antibody against α -SMA and a secondary antibody labeled with Alexa Fluor 594, and the number of positive cells was observed under a fluorescence microscope. The 5 th to 9 th generation cells were used for the experiment.
2. Cell proliferation assay
Taking logarithmic growth phase VSMCs at 1.0 × 10 per well4The cells were inoculated in 96-well culture plates at a density, starved for 24h, and grouped as follows: normal control group (control); the total saponins of radix araliae armatae are prepared into various dosage groups (1.875, 3.75, 7.5, 15 and 30 mu g/ml); 5% FBS group; dosage groups of total saponins of radix araliae armatae (1.875, 3.75, 7.5 and 15 mu g/ml) + 5% FBS group. And processing the data according to the grouping for 12h, 24h and 36h respectively. And after the treatment is finished, adding 10 mu l of CCK-8 solution into each hole, placing the well in an incubator for incubation for 2h after shaking of a shaking table, measuring the absorbance value (OD) of each group at the wavelength of 450nm by using an enzyme labeling instrument, and simultaneously setting a blank control hole, wherein the OD value is measured hole-blank hole. The experiment was repeated at least three times.
The overgrowth of VSMC is considered to be a main factor causing the restenosis of vascular injury, and therefore, the inhibition effect of total saponins of radix araliae armatae on the cell proliferation of VSMCs is determined by detecting CCK-8. Firstly, the cytotoxicity of the total saponins of aralia armata on normally grown VSMCs is detected, and as shown in a figure 3a, except that a dose group (30 mug/ml) of total saponins of aralia armata has cytotoxicity, all the other groups can not show obvious cytotoxicity. Therefore, the 4 doses of total saponins of aralia armata 1.875, 3.75, 7.5, 15 μ g/ml were selected to test the inhibitory effect on the proliferation of VSMC induced by 15% FBS, and the results are shown in fig. 3 b. The total saponins of radix araliae armatae can effectively inhibit the proliferation of VSMCs induced by serum, and the inhibition effect shows good time-effect and quantity-effect relationship.
Example 6
Scratch test
VSMCs were seeded in 6-well plates (3.0X 10)5cells/well), cells were attached and then streaked with a 100. mu.l pipette tip and washed twice with PBS. 2 olecranon total saponin doses (7.5, 15 mug/ml) which are less cytotoxic and can effectively inhibit 15% FBS-induced proliferation are selected for treatment for 24 h. Each well scratch was photographed by observing and photographing under a microscope (Olympus, Japan, 100 ×), respectively, recording the average width of the scratch of 0h and 24h, and calculating the degree of healing of the scratch according to formula II. The experiment was repeated at least three times.
The degree of healing of the scratch is equal to the initial average width of the scratch of 0h to the average width of the scratch of 24h
The effect of total saponins of aralia armata on VSMC migration was evaluated by a wound healing test, as shown in fig. 3c, significant inhibition of serum-induced VSMC wound healing at 24 hours after injury by total saponins of aralia armata, with score healing degrees of 578.85 and 420.97 for total saponins of aralia armata at 7.5 and 15 μ g/ml, respectively, with significant differences compared to the model group (fig. 3 d).
Example 7
RT-qPCR detection
VSMC total RNA was routinely extracted using TRNzol Universal reagent (available from TIANGEN Co.) and D (. lamda.TECAN) was measured using an infinite TECAN microplate reader (model: M200PRO))260/D(λ)280And if the number is 1.8-2.0, storing for later use. cDNA was synthesized from 1. mu.g of total RNA (Fast Reverse Transcription Master Mix from BIOTECHINC); the reaction system firstly removes residual DNA at 42 ℃ for 5min, then carries out reverse transcription at 42 ℃ for 15min, and inactivates reverse transcriptase at 95 ℃ for 5 min. Taking 1.2 μ l of reverse transcription reaction product to perform real-time fluorescence PCR reaction according to AB HS Green qPCR Mix kit (purchased from BIOTECH INC. Co.)) The method adopts two-step thermal cycle, and the reaction conditions are as follows: pre-denaturation at 95 ℃ for 3min for 1 cycle, denaturation at 95 ℃ for 10s, annealing at 60 ℃ for 30s, and repeating 40 cycles; all samples were loaded in 96-well PCR plates, 3 wells for each sample, and all reactions were performed in a Roche LightCycler Sequence Detection System. The primer sequences are shown in the table, and the relative quantitative method adopts a comparative Ct method, takes GAPDH as an internal reference, and adopts a delta Ct (Ct purpose-Ct internal reference) method to perform relative quantitative analysis so as to obtain the primer sequences
The relative expression level of the target RNA was determined.
TABLE 1 primer information for real-time RT-qPCR
Expression level of related genes of aralia armata total saponin inhibition wnt3 alpha/Dvl-1/beta-catenin pathway
The RT-qPCR detection result shows (figure 3e), the high-dose group (15 mug/ml) of the total saponins of aralia armata significantly inhibits the high expression of the mRNA of the Wnt signaling pathway key protein molecules (Wnt3 alpha, beta-catenin, Gsk-3 beta, Dvl-1 and Cyclin D1) of the vascular smooth muscle cells induced by 15% FBS.
Example 8
western blot
Digesting and collecting the cells treated by the medicines, extracting protein according to the instruction of a protein extraction kit, measuring the protein concentration of the obtained extract by a BCA method, and storing the extract at-80 ℃ for later use. Taking a proper amount of protein sample, carrying out electrophoresis by 10% SDS-PAGE, transferring the intracollagen protein to a PVDF membrane according to an immunoblotting method, sealing the membrane with 5% skimmed milk powder solution at 4 ℃ overnight, adding Wnt3 alpha, beta-catenin, Gsk-3 beta, Dvl-1, Cyclin D1 and GADPH antibodies on the next day respectively, incubating for 4h at room temperature, washing the membrane for 3 times by TBST, adding corresponding horseradish peroxidase-labeled secondary antibodies, incubating for 1h at room temperature, washing the membrane for 3 times by TBST, sequentially adding a luminescent substrate according to an ECL method, exposing, developing and fixing, and analyzing a band by using Image software.
Statistical analysis method
Statistical processing was performed using SPSS 19.0. For numerical values
And (4) showing. Comparing every two, and adopting Independent-Samples T test when the normality is met; and comparing the variance and the normality between multiple groups, wherein LSD-T or Dunnett-T test is adopted when the variance is uniform and the normality is satisfied, Nonparametric T test is adopted when the variance is not satisfied, and Dunnett's T3 or Tamhane's T2 test is adopted when the variance is not uniform and the variance is uniform. P.ltoreq.0.05 means statistically significant, P.ltoreq.0.01 means that the differences examined are of very significant significance.
Western blot detection results (figure 4a and figure 4b) show that the total saponins of aralia armata can better inhibit the high expression of Wnt signal pathway molecular proteins (Wnt3 alpha, beta-catenin, Gsk-3 beta, Dvl-1 and Cyclin D1) induced by 15% FBS, and particularly have a remarkable inhibiting effect on the high expression of beta-catenin, Cyclin D1 and Gsk-3 beta. The same conclusion is also confirmed in immunohistochemical experiments, as shown in fig. 2c, the total saponins of aralia armata can inhibit the expression levels of beta-catenin and Gsk-3 beta at the damaged blood vessels, and have significant difference compared with the model group (p < 0.05). The total saponins of aralia armata inhibits the expression level of wnt3 alpha/Dvl-1/beta-catenin pathway related protein.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Sequence listing
<110> Guangxi International Zhuang medical Hospital
<120> total saponins of radix araliae armatae and application thereof in preparation of medicine for treating restenosis after vascular injury
<160> 12
<170> SIPOSequenceListing 1.0
<210> 1
<211> 17
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 1
ccctcggagc ccgtgta 17
<210> 2
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 2
accaccaaat cgggtagctg 20
<210> 3
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 3
ctgctgatct cggactggac 20
<210> 4
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 4
gtcggtatca aaccaggcca 20
<210> 5
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 5
gagacacacc tgccctcttc 20
<210> 6
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 6
tggggctgtt caggtagagt 20
<210> 7
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 7
gagctgggac tacctccaga 20
<210> 8
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 8
agtggtgcct ctccatgttg 20
<210> 9
<211> 19
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 9
tcaagtgtga cccggactg 19
<210> 10
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 10
cactacttgg tgactcccgc 20
<210> 11
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 11
agtgccagcc tcgtctcata 20
<210> 12
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 12
accagcttcc cattctcagc 20
Claims (10)
1. The total saponins of aralia armata is characterized by being prepared by the following steps:
mixing the aralia armata dry root bark coarse powder with an ethanol water solution with the volume concentration of 70-80%, performing reflux extraction, collecting an extracting solution, concentrating to obtain an extract, re-dissolving the extract in water, extracting with petroleum ether, collecting a water phase, removing fat-soluble components, mixing with water-saturated n-butyl alcohol, extracting, taking supernate, removing the n-butyl alcohol, and evaporating to dryness to obtain total saponins of aralia armata.
2. The total saponins of aralia armata of claim 1, wherein the mass ratio of the dry root bark meal of aralia armata to the volume ratio of the ethanol water solution is 1 g: 6-10 mL.
3. The total saponins of aralia armata as claimed in claim 1, wherein the temperature of the reflux extraction is 60-80 ℃; the reflux extraction time is 1-1.5 h.
4. The total saponins of aralia armata as claimed in claim 1, wherein the relative density of the extract at 60 ℃ is 1.15-1.20.
5. The total saponins of aralia armata according to claim 1, wherein the fat-soluble components are removed by extraction with dichloromethane or chloroform;
the volume ratio of the dichloromethane or the trichloromethane to the water, the petroleum ether and the water-saturated n-butanol is 1: 1: 1: 1.
6. the total saponins of Artabolic acid of claim 5, wherein the total saponins of Artabolic acid comprise 1-O- [ (2 α,3 β,5 ξ,9 ξ,18 ξ,19 α) -2,3,19, 23-Tetrahydroxy-28-oxolean-12-en-28-yl ] - β -D-glucopyranose having a structural formula of formula I, 1-O- [ (2 α,3 β,5 ξ,9 ξ,18 ξ,19 α) -2,3,19,23, 24-Pentahydroxy-28-oxolean-12-en-28-yl ] - β -D-glucopyranose having a structural formula of formula II, 1-O- [ (3 β,5 ξ,9 ξ, 18) -3,19,24-Trihydroxy-24 having a structural formula of formula III, 28-dioxiurs-12-en-28-yl ] hexopyranose, 3- { [2-O- (. beta. -D-Glucopyranosyl) -beta. -D-Glucopyranosyl ] oxy } -23-hydroxyolean-12-en-28-oic acid of formula IV, (3. beta., 5. delta., 9. delta.) -3- { [2-O- (. beta. -D-Glucopyranosyl) -beta. -D-Glucopyranosyl ] oxy } -23-hydroxyolean-12-en-28-oic acid of formula V, 1-O- [ (2. beta., 3. beta., 5. delta., 9. delta., 18. delta.) -3- (. beta. -D-glucopyranosyloxy) -2, 23-dihydroxy-28-oxolean-12-en-28-yl ] -beta-D-glucopyranose of formula V, 6-Deoxy-alpha-L-mannopyranosyl- (1- >3) - [ beta. -D-Glucopyranosyl- (1- > 6-O- [ 3) ], 27-dihydroxy-27,28-dioxolup-20(29) -en-28-yl ] - β -D-glucopyranose, (3 β,5 ξ,9 ξ,18 ξ) -28-Hydroxy-28-oxolan-12-en-3-yl 6-deoxy- α -L-Mannopyranosyl- (1- >3) - [ β -D-glucopyranosyl- (1- >2) ] - β -D-glucopyranoside acid of formula VII, 1-O- (1,3,23,24,25-Pentahydroxy-28-oxo-9, 19-cyclolactostan-28-yl) hexopyranose of formula VIII, dixigin of formula ix;
7. use of total saponins of aralia armata as claimed in any of claims 1-6 in the preparation of a medicament for the treatment of restenosis following vascular injury.
8. Use of total saponins of aralia armata as claimed in any of claims 1-6 in the preparation of a medicament for inhibiting intimal hyperplasia of blood vessels.
9. Use of total saponins of aralia armata as claimed in any of claims 1-6 for the preparation of a medicament for inhibiting proliferation and/or metastasis of vascular smooth muscle cells.
10. Use of total saponins of aralia armata according to any one of claims 1-6 in the preparation of an inhibitor for inhibiting the expression of protein molecules in the Wnt signaling pathway.
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110029784.0A CN112656827B (en) | 2021-01-11 | 2021-01-11 | Radix araliae armatae total saponin and application thereof in preparation of medicine for treating restenosis after vascular injury |
| NL2030464A NL2030464B1 (en) | 2021-01-11 | 2022-01-07 | Aralia armata (wall.) seem-derived total saponin and use thereof in preparation of drug for treating post-vascular injury restenosis |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110029784.0A CN112656827B (en) | 2021-01-11 | 2021-01-11 | Radix araliae armatae total saponin and application thereof in preparation of medicine for treating restenosis after vascular injury |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN112656827A true CN112656827A (en) | 2021-04-16 |
| CN112656827B CN112656827B (en) | 2022-02-11 |
Family
ID=75414117
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110029784.0A Active CN112656827B (en) | 2021-01-11 | 2021-01-11 | Radix araliae armatae total saponin and application thereof in preparation of medicine for treating restenosis after vascular injury |
Country Status (2)
| Country | Link |
|---|---|
| CN (1) | CN112656827B (en) |
| NL (1) | NL2030464B1 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114470019A (en) * | 2022-03-16 | 2022-05-13 | 广西国际壮医医院 | Application of radix araliae armatae extract in preparation of medicine for preventing and/or treating proteinuria and related diseases |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101040884A (en) * | 2007-04-25 | 2007-09-26 | 孙晓波 | Method for controlling the quality of Chinese traditional medicine araloside |
| CN111249295A (en) * | 2020-03-23 | 2020-06-09 | 济宁医学院附属医院 | Application of astragaloside IV in inhibiting local inflammatory reaction and treating arterial restenosis |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1355171B (en) * | 2000-11-26 | 2011-01-19 | 吉林省中医中药研究院 | Extracting process and usage of general aralic chinensis saponin |
| CN104174010B (en) | 2014-08-20 | 2016-04-13 | 武汉大学 | The function and application of SHPS1 in treatment angiostenosis after damage |
| CN108992478B (en) * | 2017-06-07 | 2022-01-04 | 浙江华润三九众益制药有限公司 | Application of Chinese Aralia extract in preparing medicine |
-
2021
- 2021-01-11 CN CN202110029784.0A patent/CN112656827B/en active Active
-
2022
- 2022-01-07 NL NL2030464A patent/NL2030464B1/en active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101040884A (en) * | 2007-04-25 | 2007-09-26 | 孙晓波 | Method for controlling the quality of Chinese traditional medicine araloside |
| CN111249295A (en) * | 2020-03-23 | 2020-06-09 | 济宁医学院附属医院 | Application of astragaloside IV in inhibiting local inflammatory reaction and treating arterial restenosis |
Non-Patent Citations (1)
| Title |
|---|
| MEI HU等: "TRITERPENOID GLUCURONIDE SAPONINS FROM ROOT BARK OF ARALIA ARMATA", 《PHYTOCHEMISTRY》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114470019A (en) * | 2022-03-16 | 2022-05-13 | 广西国际壮医医院 | Application of radix araliae armatae extract in preparation of medicine for preventing and/or treating proteinuria and related diseases |
Also Published As
| Publication number | Publication date |
|---|---|
| NL2030464A (en) | 2022-07-25 |
| CN112656827B (en) | 2022-02-11 |
| NL2030464B1 (en) | 2023-02-15 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Qi et al. | Studies on the toxicity of gambogic acid in rats | |
| CN103222988B (en) | A kind of American-cockroach-extract and its preparation method and application | |
| Piao et al. | Cucurbitacin B inhibits tumor angiogenesis by triggering the mitochondrial signaling pathway in endothelial cells | |
| Guo et al. | S-petasin induces apoptosis and inhibits cell migration through activation of p53 pathway signaling in melanoma B16F10 cells and A375 cells | |
| Lee et al. | (2S)-naringenin from Typha angustata inhibits vascular smooth muscle cell proliferation via a G0/G1 arrest | |
| CN112656827B (en) | Radix araliae armatae total saponin and application thereof in preparation of medicine for treating restenosis after vascular injury | |
| Abdelhamid et al. | The hepatoprotective effect of piperine against thioacetamide-induced liver fibrosis in mice: The involvement of miR-17 and TGF-β/smads pathways | |
| Wang et al. | Fucoxanthin prevents breast cancer metastasis by interrupting circulating tumor cells adhesion and transendothelial migration | |
| Ju et al. | Curcumin-pretreated adipose-derived stem cells enhance the neuroprotective ability to repair rheumatoid arthritis-induced damage in the rat brain | |
| Zhang et al. | Kinetin inhibits proliferation of hepatic stellate cells by interrupting cell cycle and induces apoptosis by down-regulating ratio of Bcl-2/Bax | |
| CN109097358A (en) | A kind of lncRNA is preventing or is treating the application in hypertension | |
| CN112358527B (en) | Triptolide acrylate, preparation method and application thereof | |
| CN108714149A (en) | Purposes of the garden burnet active constituent in preparing anti-tumor drug | |
| Kim et al. | Antiproliferative action of cudraflavone B, isolated from Cudrania tricuspidata, through the downregulation of pRb phosphorylation in aortic smooth muscle cell proliferation signaling | |
| CN112716940A (en) | Application of canagliflozin in preparation of medicine for treating STAT6 protein-related diseases | |
| CN111297849B (en) | Pharmaceutical composition for treating laryngeal cancer, preparation method and application thereof | |
| CN109646669A (en) | The use in conjunction and combination medicine of 9R-P201 polypeptide and 5FU | |
| Zhao et al. | Aralia armata (Wall.) Seem Improves Intimal Hyperplasia after Vascular Injury by Downregulating the Wnt3α/Dvl-1/β-Catenin Pathway | |
| Wu et al. | The preventive effects of G115 on balloon injury-induced neointima formation in rats | |
| CN111870638B (en) | Application of theabrownin in preparation of medicine for treating liver cancer | |
| CN109355389B (en) | B4GALNT2 gene as biomarker for liver cancer detection and application thereof | |
| KE et al. | Preparation of total triterpenoids from Antrodia cinnamomea fermentation mycelium and their in vitro inhibitory effects on hepatocellular carcinoma | |
| CN108066349A (en) | Applications of IV a of panax japonicus saponin in treatment ephritis or kidney fibrosis drug is prepared | |
| CN109939103B (en) | Application and preparation method of coriander lactone-1 | |
| KR20180093396A (en) | Mesenchymal stem cell line useful for developing fibrosis therapeutic agent |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| CB03 | Change of inventor or designer information | ||
| CB03 | Change of inventor or designer information |
Inventor after: Zhao Xiangpei Inventor after: Teng Hongli Inventor after: Zhong Chuanmei Inventor after: Chen Shaofeng Inventor after: Zhong Wen Inventor before: Zhao Xiangpei Inventor before: Zhong Wen Inventor before: Chen Shaofeng Inventor before: Zhong Chuanmei |
|
| GR01 | Patent grant | ||
| GR01 | Patent grant |