CN110013476A - 6- hydroxyl -1,2,3,7- tetramethoxy xanthone promotees the application in the drug that hippocampal neural occurs in preparation - Google Patents
6- hydroxyl -1,2,3,7- tetramethoxy xanthone promotees the application in the drug that hippocampal neural occurs in preparation Download PDFInfo
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- CN110013476A CN110013476A CN201910278335.2A CN201910278335A CN110013476A CN 110013476 A CN110013476 A CN 110013476A CN 201910278335 A CN201910278335 A CN 201910278335A CN 110013476 A CN110013476 A CN 110013476A
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Abstract
The present invention relates to the medical usages of 6- hydroxyl -1,2,3,7- tetramethoxy xanthone.6- hydroxyl -1,2,3,7- tetramethoxy xanthones are one of Japanese polygala ingredients, and the chemical structural formula of compound is as shown in Equation 1.Experimental result shows that the chemical combination has the function of increasing the number of hippocampus neural stem cells, and clinical treatment depression can be used for for it and provides experimental basis.
Description
Technical field
The present invention relates to field of medicaments, and in particular to a kind of 6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote in preparation
Pathological characteristics disease caused by hippocampal neural occurs and/or prevents and/or treatment hippocampus neural stem cells are damaged or apoptosis
Application in drug, especially 6- hydroxyl -1,2,3,7- tetramethoxy xanthones are in preparation prevention and/or the medicine for the treatment of depression
Application in object.
Background technique
Depression is the fourth-largest disease in the whole world, the big disease of neuropsychiatric disease first, about 3.5 hundred million people's illness.WHO
Tissue expects the year two thousand twenty, and depression can exceed that cancer, become the second largest illness of the mankind for being only second to cardiovascular and cerebrovascular diseases.At me
State's incidence of depression is about the 3%-5% of total population, has estimated 61,000,000 patients 2009, it is now possible to be suffered from close to 100,000,000
Person is the most country of world's patients with depression.Depression easily recurs simultaneously, needs to take medicine throughout one's life after multiple relapse.Therefore, press down
The disease that strongly fragrant disease drug burden will rank first place as Chinese Disease Spectrum.Although there are many antidepression class drugs, due to strong
Strong side effect or drug effect is very poor, these medicines do not have therapeutic effect for about 30% patients with depression.Therefore existing
On the basis of antidepressant, need to find new drug.
Depression is a kind of multifactorial disease of pathogenic mechanism complexity, existing research shows that the structure and function of hippocampus becomes
Change is closely related with the formation of depression, therefore maintains hippocampus function and be important the pass for improving Depressive behavior and Pathological Physiology
Key brain area.Hippocampal neural generation and the function of maintaining hippocampus are closely related.Hippocampal neural is abnormal in patients with depression, and is resisted
Depressed drug, which all has, significantly increases hippocampal neural, in the animal of rodent, if eliminating hippocampal neural, resists
The antidepressant effect of depressed drug disappears.It is essential in the treatment of depression that this display hippocampal neural occurs, therefore has
The significant compound for promoting hippocampal neural generation has potential antidepressant effect.
6- hydroxyl -1,2,3,7- tetramethoxy xanthones (6-hydroxy-1,2,3,7-tetramethoxyxanthone)
It is a kind of xanthene ketone compounds extracted from Japanese polygala.The present inventor by a large amount of the study found that 6- hydroxyl -1,2,3,
7- tetramethoxy xanthone have it is significant promote nerve to occur, after further investigation, it has further been found that its can raise ERK1/2 and
The expression of BCL2 inhibits the activation of caspase-3, reduces the apoptosis of newborn nerve cell, to reach the newborn nerve cord of increase
Cell maturation increases the neurogenetic effect of adult hippocampal, to achieve the effect that improve depression.
Summary of the invention
The present invention provides a kind of 6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote the medicine that hippocampal neural occurs in preparation
New application in object, which, which has, promotees nerve to occur effect, may be used as promoting neurogenic agent and is used to prepare prevention
And/or treatment hippocampus neural stem cells damage or apoptosis caused by pathological characteristics disease, in particular for preparation prevention and/or
The drug of depression is treated, provides a kind of new medicament selection approach to prevent and/or treating depression.
Specifically, 3,7- tetramethoxy xanthones promote hippocampus mind in preparation the present invention provides a kind of 6- hydroxyl -1,2
Structural formula compound shown in formula I is promoted hippocampal neural as preparation and occurs and/or prevent by the application in the drug through occurring
And/or the effective component of the drug of pathological characteristics disease caused by the damage for the treatment of hippocampus neural stem cells or apoptosis.
The present invention also provides a kind of 6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote what hippocampal neural occurred in preparation
The compound of the acceptable salt of structural formula compound shown in formula I is promoted hippocampal neural hair by application in drug
The drug of pathological characteristics disease caused by raw and/or prevention and/or the damage for the treatment of hippocampus neural stem cells or apoptosis it is effective
Ingredient.
The present invention also provides a kind of 6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote what hippocampal neural occurred in preparation
Application in drug, being will be using a effective amount of structural formula compound shown in formula I to be subjected on active constituent, with pharmacy
Carrier and/or auxiliary material be made any one dosage form in pharmacy and occur and/or prevention and/or control for promoting hippocampal neural
Treat the pharmaceutical composition of pathological characteristics disease caused by hippocampus neural stem cells damage or apoptosis.
Further, the 6- hydroxyl -1,2,3,7- tetramethoxy xanthones are making each promote in neurogenetic drug
In, pathological characteristics disease caused by the rush hippocampal neural occurs and/or hippocampus neural stem cells are damaged or apoptosis is
Depression.
The principle of the present invention is:
6- hydroxyl -1,2,3,7- tetramethoxy xanthones can increase the neurogenetic effect of adult hippocampal, change to reach
It is apt to the effect of depression.
The present invention compared to the prior art, the beneficial effect is that:
The present invention provides 6- hydroxyl -1,2, the novel medical use of 3,7- tetramethoxy xanthones has and promotees hippocampal neural
The effect of generation can be used as and promote hippocampal neural generation drug and prevention and/or the damage for the treatment of hippocampus neural stem cells or apoptosis
The drug of caused pathological characteristics disease especially provides a kind of new medicament selection for the treatment of depression.
Detailed description of the invention
Fig. 1 is the hippocampus BrdU cell dyeing figure in embodiment 1 (cell of yellow arrows meaning is BrdU positive cell).
Wherein, figure A is normal mice hippocampus BrdU cell dyeing figure, and figure B is depression model mouse BrdU cell dyeing figure, and figure C is PMT mouse
BrdU cell dyeing figure, figure D are the statistical graph of hippocampus BrdU positive cell.
Fig. 2 is the DCX immunohistochemical staining figure in embodiment 2.Wherein, figure A is DCX immunohistochemical staining representative diagram
Piece, figure B are DCX positive cell statistical graph.
Fig. 3 is the hippocampal dentate representative diagram in embodiment 3.Wherein, figure A is control mouse hippocampal dentate representative diagram
Piece, figure B are depressed mouse hippocampal dentate representativeness picture, and figure C is PMT mouse hippocampal dentate representativeness picture, and figure D is dentate fascia
Volume statistical graph.
Fig. 4 is the mouse Depressive behavior statistical graph in embodiment 4.Wherein, figure A is the deadlock of each processing group mouse in tail-suspention test
Straight time statistical form, figure B are the stiff time statistical form of each processing group mouse in forced swim test, and figure C is the overhead experiment of cross
In each processing group mouse open arms time statistical form.
It illustrates: * * p < 0.01 in figure.
Specific embodiment
Combined with specific embodiments below, the present invention is further described.
The therapeutic agent Prozac that the present invention implements to select clinical use effect relatively good is as positive control.
One 6- hydroxyl -1,2 of embodiment, 3,7- tetramethoxy xanthones promote hippocampal neural and have an effect
Normal C57BL/6 mouse is randomly divided into three groups, depression model mouse+physiological saline as a control group (stress group),
Depression model mouse+6- hydroxyl -1,2,3,7- tetramethoxy xanthones are as administration group (PMT group), normal mice+physiological saline conduct
Normal group (control group).By fetter for a long time building depression model mice (depression model group mouse fetters 4 hours daily,
Continuous constraint 21 days can detect depressive state by forced swimming and tail-suspention test at the 21st day, if depressed shape is presented in mouse
State can determine whether experiment modeling success.) start within PMT group mouse the 8th day after modeling that 6- hydroxyl -1,2,3,7- tetra- methoxies are injected intraperitoneally
Base xanthone 100ug/kg, x14 days 1 time a day, stress group and control group gave same volume physiology in same time point
Salt water intraperitoneal injection.Last day, all mouse peritoneals inject 5- bromodeoxyuridine nucleosides (BrdU) (50mg/Kg), take brain.
First by taking brain after 4% paraformaldehyde perfusion, then it is dehydrated with 10%, 20% and 30% sucrose solution.Finally by frost
It cuts machine and obtains mouse brain slice according to every 30um offspring, brain piece is placed in phosphate buffer (PBS), is put into 4 DEG C of refrigerators
It can long-term preservation.
Nerve to occur detection, using the method for immunohistochemistry, BrdU dyeing, manual count counts hippocampus of mice dentate fascia
All BrdU positive cell numbers in granulocyte lower layer.As a result as shown in Figure 1.
It can be obtained from Fig. 1, PMT can significantly improve the growth rate of the stem cell of mouse, can achieve and normal mouse
Identical level.This show PMT can significantly improve due to stress caused by the effect that reduces of hippocampal neural.
Two 6- hydroxyl -1,2 of embodiment, 3,7- tetramethoxy xanthones increase hippocampus new life nerve cell
Analysis PMT is converted into the influence of nerve cell to neural stem cell.The reaction of hippocampal dentate BrdU positive cell
It is nerve stem cell proliferation situation, but whether to be converted into neuron also unknown for increased neural stem cell, therefore we pass through
Both adrenal glands cortin (newborn nerve cell differential protein, DCX) dyeing, DCX are newborn but immature nerve cells
Marker, the number of hippocampus of mice new life nerve cell can be reacted by counting DCX positive cell number in hippocampus of mice.
C57BL/6 mouse is handled according to the method in embodiment one, obtains mouse brain slices.
Pass through the method analysis analysis hippocampal dentate DCX positive cell number purpose variation of immunohistochemistry.As a result such as Fig. 2 institute
Show.
It can be obtained from Fig. 2, PMT significantly increases mouse neonatal nerve cell number.This display neural stem cell division generates
Neonatal cell be converted into newborn nerve cell.
Three 6- hydroxyl -1,2 of embodiment, 3,7- tetramethoxy xanthones increase the volume of depression model mouse hippocampal dentate
C57BL/6 mouse is handled according to the method in embodiment one, obtains mouse brain slices.
The calculating of hippocampal dentate (DG) volume.Brain piece passes through 4 ', 6- diamidino -2-phenylindone (4 ', 6-
Diamidino-2-phenylindole, DAPI) label.DAPI can mark all nucleus, since hippocampal dentate is thin
Born of the same parents are closeer than fine and close, and after DAPI label, hippocampal dentate and other region (figures can be easily discriminated under fluorescence microscope
3).Continuous brain piece is obtained by continuous brain section first, then is dyed by DAPI, can get after taking pictures under fluorescence microscope
The picture of continuous brain piece.The area of each brain piece hippocampal dentate is calculated by software image J.Due to each brain piece
Thickness be 30um, according to Cavaglieri principle, pass through formula
V=Tx ∑T=1,2,3 ... nA
V, hippocampal dentate volume；T, the distance between continuous two brain pieces；A, the area of hippocampal dentate cross section；I, brain
The number of piece can calculate the total volume of DG.
C57 mouse is handled according to the method in embodiment one, obtains mouse brain slices.Sea purslane can be counted after DAPI dyeing
Shape ex vivo product.Count variation of the PMT to hippocampal dentate volume.As a result as shown in Figure 3.
It can be obtained from Fig. 3, the hippocampal dentate volume of depression model mouse after PMT processing, restore by hippocampal dentate volume.
Example IV 6- hydroxyl -1,2,3,7- tetramethoxy xanthones improve the depressed shape behavior of depression model mouse
Normal c57BL/6 mouse is randomly divided into four groups, and one group is normal mouse+physiological saline (control as a control group
Group), one group is depression model mouse+physiological saline as negative control group (stress group), and one group is depression model mouse+plus 6- hydroxyl
Base -1,2, for 3,7- tetramethoxy xanthones as administration group (PMT group), one group is depression model mouse+Prozac as positive right
According to group (fluoxetine group).Depression model mouse passes through constraint building for a long time, building mode reference implementation example one.PMT group
(100ug/kg) and fluoxetine group (1mg/kg) begin through intraperitoneal administration on the 14th day constraint, and x7 days 1 time a day.
Control group and stress group give isometric physiological saline in the identical point time.
At the 21st day, by the experiment of tail-suspention test, forced swim test and cross elevated plus-maze test to assess mouse depression
State.As a result as shown in Figure 4.
It can be obtained from the A of Fig. 4, in tail-suspention test, relative to depression model mouse, PMT and Prozac handle depression model mouse
The dead time that mouse can be substantially reduced shows 6- hydroxyl -1,2, and 3,7- tetramethoxy xanthones are on improving depression
It acts on identical as Prozac.
It can be obtained from the B of Fig. 4, in forced swim test, PMT and Prozac processing depression model mouse can substantially reduce suppression
The dead time of strongly fragrant model mouse.
It can be obtained from the C of Fig. 4, in the experiment of cross elevated plus-maze test, PMT and Prozac processing depression model mouse can increase suppression
Residence time of the strongly fragrant model mouse in open arms.
The above results show that 6- hydroxyl -1,2,3,7- tetramethoxy xanthones can significantly improve depression model mouse
Depressed shape behavior, therefore, the xanthone have significant antidepressant effect.
Claims (3)
1.6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote the application in the drug that hippocampal neural occurs, feature in preparation
It is: compound shown in formula I is promoted into hippocampal neural generation and/or prevention as preparation and/or treatment hippocampal neural is dry thin
Effective component in the drug of pathological characteristics disease caused by cellular damage or apoptosis.
2.6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote the application in the drug that hippocampal neural occurs, feature in preparation
Be: using the compound of the pharmaceutical salts formula of compound shown in formula I as preparation promote hippocampal neural occur and/or prevention and/
Or the effective component in the drug of pathological characteristics disease caused by the damage for the treatment of hippocampus neural stem cells or apoptosis.
3.6- hydroxyl -1,2,3,7- tetramethoxy xanthones promote the application in the drug that hippocampal neural occurs, feature in preparation
It is, using such as compound of I compound represented or its acceptable salt as preparation rush hippocampal neural generation drug and/or in advance
Effective component in anti-and/or treatment depression drug.
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| CN201910278335.2A CN110013476A (en) | 2019-04-08 | 2019-04-08 | 6- hydroxyl -1,2,3,7- tetramethoxy xanthone promotees the application in the drug that hippocampal neural occurs in preparation |
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| CN201910278335.2A CN110013476A (en) | 2019-04-08 | 2019-04-08 | 6- hydroxyl -1,2,3,7- tetramethoxy xanthone promotees the application in the drug that hippocampal neural occurs in preparation |
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Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060287297A1 (en) * | 2005-06-16 | 2006-12-21 | Decorte Bart | Tricyclic opioid modulators |
| US20080139573A1 (en) * | 2006-12-08 | 2008-06-12 | Copharm | Treatment of resistant Schizophrenia and other CNS disorders |
| CN102160873A (en) * | 2011-03-28 | 2011-08-24 | 南京泽朗农业发展有限公司 | Method for preparing total polygala xanthone |
| CN102229594A (en) * | 2011-05-06 | 2011-11-02 | 南京泽朗农业发展有限公司 | Method of dual-aqueous phase system for purifying polygala root xanthonoid compound |
| CN103191141A (en) * | 2012-01-05 | 2013-07-10 | 昆明制药集团股份有限公司 | Application of 3-methoxyl mangiferin in preparing medicine used for treating post-stroke depression |
| CN104177324A (en) * | 2013-05-20 | 2014-12-03 | 南京工业大学 | Xanthone compounds and their use in depression resistance |
| CN105218507A (en) * | 2015-09-30 | 2016-01-06 | 中国药科大学 | A kind of Norathyriol derivative and its production and use |
-
2019
- 2019-04-08 CN CN201910278335.2A patent/CN110013476A/en active Pending
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060287297A1 (en) * | 2005-06-16 | 2006-12-21 | Decorte Bart | Tricyclic opioid modulators |
| US20080139573A1 (en) * | 2006-12-08 | 2008-06-12 | Copharm | Treatment of resistant Schizophrenia and other CNS disorders |
| CN102160873A (en) * | 2011-03-28 | 2011-08-24 | 南京泽朗农业发展有限公司 | Method for preparing total polygala xanthone |
| CN102229594A (en) * | 2011-05-06 | 2011-11-02 | 南京泽朗农业发展有限公司 | Method of dual-aqueous phase system for purifying polygala root xanthonoid compound |
| CN103191141A (en) * | 2012-01-05 | 2013-07-10 | 昆明制药集团股份有限公司 | Application of 3-methoxyl mangiferin in preparing medicine used for treating post-stroke depression |
| CN104177324A (en) * | 2013-05-20 | 2014-12-03 | 南京工业大学 | Xanthone compounds and their use in depression resistance |
| CN105218507A (en) * | 2015-09-30 | 2016-01-06 | 中国药科大学 | A kind of Norathyriol derivative and its production and use |
Non-Patent Citations (2)
| Title |
|---|
| HÖLZL: "贯叶金丝桃抗抑郁作用和对情绪变化的影响", 《国外医药 植物药分册》 * |
| 夏忠庭等: "基于UPLC-ESI-IT-TOF/MS方法分析郁舒片中脂溶性成分", 《药学学报》 * |
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