A kind of Chinese medicine composition and application thereof with promotion mescenchymal stem cell proliferation function
Technical field
The invention belongs to the cytobiology field, relate to a kind of purposes of Chinese medicine composition, be specifically related to Icaritin and rhodioside in the purposes that promotes aspect mescenchymal stem cell propagation.The invention still further relates to the purposes of this Chinese medicine composition in treatment cell injury disease.
Background technology
Stem cell is a kind of initiating cell with self-renewal capacity and multi-lineage potential, and under given conditions, can also breeding also, directed differentiation becomes different functioning cells.Therefore the research of stem cell is significant to renewal and the injury repairing of each histoorgan of life entity, becomes gradually unique hope of the many diseases that can't cure, particularly cell and tissue disappearance or injury disease.Stem cell comprises embryonic stem cell and adult stem cell, and embryonic stem cell is due to ethics problem, and its application is subject to great restriction; Form the soma cell owing to can being divided into functioning cell and tissue, for the widespread use of stem cell provides the foundation, for research Mammals early development provides good external model, the cell replacement therapy that can be numerous disease provides new cell derived, indicates that also the development of modern biology and the research and development of novel drugs have entered the New Times.
Up to the present, only have a small amount of adult tissue stem cell, can cultivate at the external serum-free amplification that carries out, but need to add different somatomedins, signaling molecule etc. to carry out genetic regulation.Obviously, the amplification in vitro culturing stem cells need to have safer, more effective cell culture technology.
Herba Epimedii (Herba Epimedii) is the Berberidaceae barrenwort, the effect such as have kidney invigorating and YANG supporting, dispel rheumatism.Modern pharmacological research proves, Herba Epimedii contains special chemical composition and significant biological activity, is a kind of Chinese medicinal materials with extensive pharmacologically active.The flavonols compounds such as the Icaritin in Herba Epimedii have the treatment cardiovascular and cerebrovascular diseases, prevent and treat the pharmacological action of osteoporosis etc.Icaritin is the aglycon of main effective constituent icarin in Herba Epimedii, and chemical structural formula is as follows:
Root of Kirilow Rhodiola is red-spotted stonecrop (Crassulaceae) section Rhodida plant, is perennial herb or semishrub plant, mainly is distributed in Himalayas, the northwestward, Asia and the North America height above sea level in the Northern Hemisphere in the area of 1600-4000 rice.The Root of Kirilow Rhodiola medicinal history is long, have the title of " gold ginseng ", " Radix Rhodiolae ".Pharmacopeia record Root of Kirilow Rhodiola is the dry root and rhizome of crassulaceae plants Radix Rhodiolae, and its property is sweet, bitter, put down, and benefiting QI for activating blood circulation, promotes blood circulation and relievings asthma, and can be used for qi deficiency to blood stasis, Obstruction of qi in the chest and cardialgia, stroke hemiplegia and indolent and listless asthma.Modern pharmacological research shows, the biological activity of Root of Kirilow Rhodiola is inseparable with contained number of chemical composition.
Rhodioside is the natural product of separating-purifying from Root of Kirilow Rhodiola, is a kind of monomeric compound, is one of effective constituent of Root of Kirilow Rhodiola, and partly relevant section belongs to Chinese medicine and also contains rhodioside, and chemical structural formula is as follows:
Summary of the invention
The purpose of this invention is to provide the Chinese medicine composition with stem cells hyperplasia effect.Chinese medicine composition provided by the invention can be used as stem cells hyperplasia promotor and as the medicines of a large amount of preparation stem cells, also can be used for scientific research, for the fundamental research of stem cell provides a large amount of stem cells.Chinese medicine composition provided by the invention can promote the reparation of stem cells hyperplasia and functioning cell, also can be used for the stem cells hyperplasia research on mechanism.
Chinese medicine composition provided by the invention is prepared as follows:
The icarin ultrasonic dissolution slowly adds the damping fluid of cellulase while stirring in the least possible methyl alcohol, 37 ℃ of reactions are spent the night.The structural glucose of icarin and rhamnosyl fully are hydrolyzed, the ethyl acetate extraction hydrolysate, the Rotary Evaporators volatile dry namely gets Icaritin.The research report is arranged, and in the body of Herba Epimedii extract, pharmacological evaluation shows that Icaritin has the estrogen-like pharmacologically active, can make the female mice uterus weight, and estradiol content raises.This class has the natural phant compound of estrogen activity, have the effects such as anti-oxidant, angiogenesis inhibitor, anticancer differentiation, to hormonal dependent cancer, cardiovascular disorder and postmenopausal osteoporosis and climacterium uncomfortable disease, early old dementia has preventive and therapeutic action preferably.
The separation purification method of rhodioside monomer: take the Root of Kirilow Rhodiola powder as raw material, extract rhodioside take water as solvent with microwave method, optimal processing parameter is: liquid-solid ratio 20: 1, soak time 1.5h, low power microwave is processed 60S, magnetic agitation 15min, extract 2 times, extraction yield can reach 90%.The pharmacological action of rhodioside comprises the ability of anti-hypoxia, antifatigue effect and external removing hydroxy radical qiao.Rhodioside can suppress the increase that interleukin-1 beta that Ischemia and Reperfusion in vivo in Rats causes is expressed, and reduces the expression of TNF-α in cerebral ischemia Post-per fusion Cerebral tissue, plays the cerebrovascular effect of protecting; By suppressing the expression of blood vessel ACE, can resist myocardial ischemia to a certain extent, improve body immunity.
The present inventor finds in experiment, the composition of Icaritin and rhodioside human mesenchymal stem cell to vitro culture under finite concentration and proportioning has proliferation, make these cells keep in a long time vegetative state, compare with traditional cultural method, the speed of growth of cell is faster, and cytoactive is higher.
The present invention includes following content:
Mescenchymal stem cell propagation reagent is the material that can increase mescenchymal stem cell quantity, for example, can by stimulate proliferation, Inhibited differentiation and prevent the dead mescenchymal stem cell quantity that increases of mescenchymal stem cell.
Chinese medicine composition of the present invention comprises Icaritin and rhodioside.Extract Icaritin and rhodioside according to preceding method, its preferably combination ratio is 5: 1.
Chinese medicine composition consumption of the present invention: the ratio according to 1% joins in basic medium.Basic medium comprises DMEM/F12,1640, α MEM etc.
Data of the present invention shows, uses drug regimen of the present invention to play promoter action to the multiplication culture of autologous bone marrow mesenchymal stem cells, umbilical cord mesenchymal stem cells.Microscopic observation uses drug regimen of the present invention can make mescenchymal stem cell keep greater activity, and rate of propagation increases substantially, and prevents its differentiation.The present invention is to the clinical application of mesenchymal stem cell transplantation and to use mescenchymal stem cell to carry out drug screening all significant.
The short multiplication agent of stem cell of the present invention can be used for the propagation of stem cell, it is characterized in that, contacts with stem cell by described Chinese medicine composition, promotes the propagation of stem cell.In the occasion of external use stem cells hyperplasia agent provided by the invention, micromolecular compound provided by the invention is dissolved in suitable solution uses afterwards.Described suitable solution comprises the substratum such as DMEM.
Mescenchymal stem cell of the present invention comprises umbilical cord mesenchymal stem cells and autologous bone marrow mesenchymal stem cells.
Chinese medicine composition stem cell provided by the present invention is urged directly administration of multiplication agent, uses as conventional dose.In such use, described stem cells hyperplasia agent is take Chinese medical extract provided by the invention as activeconstituents.The short multiplication agent of stem cell provided by the invention can contain separately Chinese medical extract provided by the invention, or Chinese medical extract provided by the invention is according to the mixture of specified proportion composition.
The present invention also provides a kind of method of culturing stem cells, is included in the situation of Chinese medicine composition existence of the present invention culturing stem cells.
Description of drawings
Fig. 1 is the promoter action that Icaritin and rhodioside are bred umbilical cord mesenchymal stem cells
Embodiment
In following embodiment, in the situation that the short multiplication agent of stem cell of the present invention exists, cultivate and obtain stem cell.The below enumerates embodiment, and the present invention will be described in more detail, but the present invention is not limited to these embodiment.
Embodiment 1
With Icaritin and rhodioside, mix according to the ratios of 5: 1, be dissolved in that in the DMEM/F12 substratum, (ultimate density is: Icaritin: 5uM; Rhodioside: 1uM).With this culture medium culturing umbilical cord mesenchymal stem cells.Mescenchymal stem cell (obtaining from umbilical cord according to a conventional method) is incubated in serum-free conventional medium (conventional medium comprises DMEM/F12,1640, α MEM etc.).Experiment is grouped into: medium controls (not adding Chinese medicine composition); Substratum+Chinese medicine composition.Cultivate and add MTT (5mg/ml) 10ul after four days, add lysate (10%SDS, 0.1%NH after hatching 4h
4Cl) 100ul, overnight incubation is in 490nm photometry absorption value.
As shown in Figure 1, the height of histogram represents chemiluminescence intensity, is directly proportional to cell count.This shows, under the existence of the short multiplication agent of Chinese medicine composition, umbilical cord mesenchymal stem cells propagation has 30% increase, confirms that this Chinese medicine composition has the effect that promotes propagation to mescenchymal stem cell.