CN105982881A - Application of bavachalcone and analogues thereof - Google Patents
Application of bavachalcone and analogues thereof Download PDFInfo
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- CN105982881A CN105982881A CN201510064923.8A CN201510064923A CN105982881A CN 105982881 A CN105982881 A CN 105982881A CN 201510064923 A CN201510064923 A CN 201510064923A CN 105982881 A CN105982881 A CN 105982881A
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Abstract
The invention discloses an application of bavachalcone and analogues thereof. As for the application, the bavachalcone or/and the analogues thereof, as active ingredients, are used for preparing a composition for activating a retinoic acid-related orphan receptor-[alpha]. Research results show that the bavachalcone and the analogues thereof can directly activate the activity of the ROR[alpha] (retinoic acid-related orphan receptor-[alpha]), present dose dependency, and can activate the expression of ROR[alpha] protein and ROR[alpha]mRNA in a concentration-dependent manner and effectively regulate and control the expression of a downstream circadian clock gene thereof, namely BMAL1, showing that the bavachalcone and the analogues thereof, as a good ROR[alpha] positive activator, can be used for preventing or treating ROR[alpha]-mediated related diseases; and the bavachalcone and the analogues thereof have a broad application prospect.
Description
Technical field
The present invention relates to the purposes of a kind of Bavachalcone and the like, belongs to pharmaceutical technology field.
Background technology
Lonely nuclear receptor (Orphan nuclear receptor) is that a big class is currently without the nuclear receptor finding and determining native ligand.
The relevant lonely nuclear receptor-α (Retinoic acid-related orphan receptor α, ROR α) of retinoic acid is distributed widely in each of body
Individual tissue, is a kind of transcription regulatory factor belonging to steroid hormone receptor superfamily member, and it can be directly entered nucleus
Transcribing of regulation target gene, thus participate in the regulation of multiple important physiological process, at morphological development, cell proliferation and differentiation, machine
The aspects such as body stable state maintains, biological metabolism regulation and control, the control of higher nerve function and immunomodulating play a significant role.
Research finds, ROR α is relevant to multiple disease, can be as the target spot of Drug therapy.Such as:
1) with the dependency of cardiovascular disease, research is pointed out: the rs10519096 of ROR α and downstream signal BMAL1's
The single nucleotide polymorphism (SNPs) of rs3816358 and the non-Wood-scoop type phenotype hypertension significant correlation of the young patient of 372 examples
(Hypertens Res.2014 Nov 20.);Melatonin can reduce male's constitutional height blood as internal ROR alpha activators
Nighttime blood pressures (the J Biol Chem.1994 of pressure patient;269:28531-28534;Hypertension.2004;43:192-197);
ROR α mutant mouse suffers from serious atherosclerosis (Circulation.1998;98:2738-2743.);RORα
In the expression of component protein ApoAI and V that receptor modulators has the high density lipoprotein (HDL) of suppression tremulous pulse medicated porridge sample effect
Rise (J Biol Chem.1997;272:22401-22404;Arterioscler Thromb Vasc Biol.2005;25:1186-1192.).
2) with the dependency of metabolic disease, research shows: in pancreas and islets of langerhans, to present day-night cycle poor for ROR family gene
Express (Mol Cell Endocrinol.2013 different in naturely;365:129-138.), its insulin secretion is regulated and controled by ROR α
(FEBS Lett.2014;Mar 18;588(6):1071-1079.);ROR α also changes hepatocellular glucose utilization rate
(Hepatology.2014;Oct 25.doi:10.1002/hep.27577.);ROR α is the fat core with the energy balance of regulation
Receptor (FEBS Lett.2009;583:2031-2036.);Selective activation ROR α may be fat and atherosclerotic
Effective (Lau P, Nixon SJ, Parton RG, et al.J Biol Chem.2004 in treatment;279:36828-36840.);Retinoic acid
Relevant lonely nuclear receptor α channel genes promote the circadian rhythm of CYP8B1 and expression on an empty stomach, increase hepatic cholesterol amount,
Increase serum and 12 Alpha-hydroxy bile acid levels (the J Biol Chem.2013 in bile acid pond;288:37154-65.);7 Alpha-hydroxies are solid
Alcohol (7 α-OHC) is the key intermediate of bile acid biosynthesis, and 7 α-OHC can stimulate the expression of ROR alpha target genes, such as Fructus Vitis viniferae
Expression (the J Biol Chem.2010 of sugar-6-phosphatase and phosphoenolpy ruvate carboxy kinase;285:5013-5025.).
3) with cerebral atrophy and the dependency of neurodegenerative diseases, research shows: spinocebellar ataxia is adult
The poly glumine neurodegenerative diseases of morbidity.In adult spinal cord cerebellar ataxia mice, ROR is alpha mediated
The development of Purkinje cell determines the order of severity of disease, and the partial loss of ROR α strengthens ataxin 1 (ATXN1)
Pathogenic (the Cell.2006 of sudden change;127:697-708.);ROR α controls the early stage step (J of Purkinje cell dendron shape differentiation
Neurosci.2006;26:1531-1538.);ROR α process LAN is possible to prevent the apoptosis (J of the neuron that oxidative stress induces
Neurochem.2006;96:1778-1789.);Under anaerobic condition, ROR α also has neuron and astrocyte
Neuroprotective function (J Neurosci.2011;31:14314-14323.), this improve nerve retrograde affection protective effect be
(the Antioxid Redox Signal.2014 realized is expressed by ROR α induction SOD2 and GPx1;21:2083-2094.).
4) with osteoporosis and the dependency of rheumatoid arthritis, research shows: the mescenchymal stem cell of derived from bone marrow is expressed
ROR α, it can promote the Osteoblast Differentiation of mescenchymal stem cell, directly activate Mouse Bone sialoprotein and express nearly 7 times.This research
Also showing, total bone mineral content, density, region and the length of the tibia of ROR α sudden change staggerer mice are less than ROR α
Heterozygote and wild-type mice (Proc Natl Acad Sci USA.2000;97:9197-9202.);As ROR alpha activators
Melatonin may have three aspects to the impact of bone function: (I) improves osteoblastic differentiation and activity;(II) skeletonization is increased
The expression of cell osteoprotegerin, prevents the differentiation of osteoclast;(III) free radical and the bone resorption of removing osteoclast generation are lived
Property (J Osteoporos.2010;2010:830231.);Thiazolidinedione (Thiazolidine diones) has good antirheumatic
Property arthritic effect, its mechanism is found to be by courier's mediation in ROR α core, and be that rheumatic arthritis etc. is treated
Provide a kind of new Therapeutic Method (J Biol Chem.1996;271:13515-13522.).
5) with the dependency of cancer, research shows: melatonin can suppress the growth (Mol of human breast carcinoma through activating ROR α
Cell Endocrinol.2001;176:111-120.);In human breast carcinoma tissue and cell line, ROR alpha expression is down regulation trend,
The mRNA level in-site of SEMA3F also declines, and Prognosis in Breast Cancer is poor, again recovers ROR alpha expression with strengthening
The expression of SEMA3F, can suppress breast cancer cell tumor growth (Cancer Res.2012 in nude mouse;72:
1728-1739.);The drug treating of the activator thiazolidine diketone derivative CGP52608 of ROR α, at mRNA and albumen
Significantly reducing the expression of 5-lipoxidase in level, CGP52608 drug treating can also completely inhibit by linoleic acid and Semen arachidis hypogaeae
Proliferation function (the Int J Cancer.2004 of the human prostate cancer cell line DU 145 of tetraenoic acid induction;112:87-93.);ROR α passes through
PKC α path weakens Wnt/ β-catenin signal (the Mol Cell.2010 of colon cancer;37:183-195.);ROR alpha activators takes off
Melanocyte also strengthens apoptosis (the Mol Cell Biochem. of people's promyelocytic leukemia HL-60 cell of hydrogen peroxide-induced
2011;353:167-176.);In hepatoma carcinoma cell, SPARC, PLG, G6PC, NR1D2 and AGRP gene are can
(the PLoS One.2011 regulated and controled by ROR α;6:e22545.);ROR alpha expression is lowered poor with liver cancer patient prognosis relevant,
Its expression is notable and closely related (the Tumour Biol. of serum alpha-fetoprotein, pathological grading, tumor recurrence rate and vessel invasion
2014;35:7603-7610.).
6) with the dependency of diseases associated with inflammation, research shows: ROR α can suppress hydroxysteroid sulfotransferase (SULT2A1)
Expression, it is also possible to strengthen AMPK activity, improve fatty liver, fat hepatitis and hepatic fibrosis (Mol Endocrinol.2013;
27:106-115;Hepatology.2012;55:1379-1388.);ROR α can also be by inducing the table of SOD2 and GPx1
Reach reduction oxidative stress, prevent non-alcoholic stellato-hepatitis from developing (Antioxid Redox Signal.2014;21:
2083-9204.);ROR α gene mutation SG/SG mice shows the degree of inflammation to LPS induction than wild-type mice to be had more
High reactivity, the inflammatory cell quantity of SG/SG mouse lung, IL-1 β, IL-6 and MIP-2 (MIP-2)
Secretory volume also exceeds wild-type mice, and the high response of this inflammation regulates and controls unrelated (Am J Physiol Lung with NF kB activity
Cell Mol Physiol.2005;289:L144-L152.);The process LAN of ROR α 1 and ROR α 4 can suppress TNF-α to stimulate
P50 and p65 to core internal shift, and then the vascular cell adhesion molecule-1 (VCAM-1) of suppression vascular endothelial cell and
Intracellular adhesion molecule-1 (ICAM-1) expresses (FEBS Lett.2004;557:269-274;J.Arterioscler Thromb Vasc
Biol.2005;25:710-716.);The melatonin of the ROR alpha activators of high concentration can suppress NF-κ B to express and and DNA
Binding ability, this effect via ERK/AKT/PKC path, and may thus reduce the propagation (J of vascular endothelial cell
Pineal Res.2008;44:107-114.).
The most visible: ROR α and cardiovascular disease (such as: atherosclerosis, hypertension), metabolic disease (such as:
Diabetes, obesity, abnormal carbohydrate metabolism, hyperlipidemia), nervous system disease (such as: cerebral atrophy, neurodegenerative diseases),
Skeleton disease (such as: osteoporosis, rheumatoid arthritis), Cancerous disease (such as: breast carcinoma, hepatocarcinoma, carcinoma of prostate,
Colorectal carcinoma), the multiple disease phase such as inflammation related disease (such as: chronic obstructive pulmonary disease, nonalcoholic steatohepatitis)
Closing, ROR alpha activators can be applied in scope the most widely, and therefore, this area is in the urgent need to developing effective ROR α
Activator.
Bavachalcone (Bavachalcone) is to belong to Rosales pulse family Fructus Psoraleae (to have another name called: Fructus Psoraleae, mother-in-law solid fat, recklessly fragrant-flowered garlic
Son) main active.2005 by Ming Hong Lee et al. first from Fructus Psoraleae (Psoralea corylifolia)
In separate, and named Bavachalcone, its No. CAS is: 28448-85-3, and molecular formula is: C20H20O4, point
Son amount is: 324.4, and chemical structural formula is:Existing research only shows Fructus Psoraleae Cha Er
Ketone and the like has good bacteriostatic activity to multiple frequently seen plants disease, so far there are no Bavachalcone and similar
Other purposes report of thing.
Summary of the invention
It is an object of the invention to provide the purposes of a kind of Bavachalcone and the like, with widen Bavachalcone and
The range of application of analog.
The purposes of Bavachalcone of the present invention and the like, refers to Bavachalcone or/and its analog is made
For active component be used for preparing the relevant lonely nuclear receptor-α of activation retinoic acid (Retinoic acid-related orphan receptor α,
ROR α) compositions.
Furtherly, the purposes of Bavachalcone of the present invention and the like, refer to Bavachalcone or/and
Its analog is used for preparing relevant lonely nuclear receptor-α (the Retinoic acid-related orphan receptor of retinoic acid as active component
α, ROR α) activator.
Furtherly, the purposes of Bavachalcone of the present invention and the like, refer to Bavachalcone or/and
Its analog is used for preparing prevention or treatment by relevant lonely nuclear receptor-α (the Retinoic acid-related of retinoic acid as active component
Orphan receptor α, ROR α) compositions of relevant disease that mediates.
Above-mentioned composition is pharmaceutical composition, Halth-care composition or food compositions.
Described relevant disease is cardiovascular and cerebrovascular disease, metabolic disease, nervous system disease, skeleton disease, Cancerous disease
Or at least one in inflammation disease.
Described cardiovascular and cerebrovascular disease is at least one in hypertension, atherosclerosis;Described metabolic disease is sugar
At least one during urine disease, abnormal carbohydrate metabolism, obesity, hyperlipidemia, Bile metabolism are abnormal;Described nervous system disease is
At least one in cerebral atrophy, neurodegenerative diseases;Described skeleton disease is osteoporosis, rheumatoid pass
At least one in joint inflammation;Described Cancerous disease is that breast carcinoma, hepatocarcinoma, colorectal carcinoma, carcinoma of prostate, early children's grain are white
At least one in disorders of blood;Described inflammation disease be aseptic inflammation, chronic obstructive pulmonary disease, nonalcoholic steatohepatitis,
At least one in pneumonia.
Described Bavachalcone and the like is for have the compound of Formulas I structure or pharmaceutically can connecing of described compound
Salt, tautomer, stereoisomer or the precursor compound being subject to:
Wherein: R1、R2、R3、R4、R5、R6、R7、R8、R9Be each independently selected from hydrogen, ORa, OCOORa,
OCONRaRb、OSO2Ra、OSO3Any one in Ra;Described Ra and Rb be each independently selected from hydrogen, replacement or
Unsubstituted C1~C12 alkyl, substituted or unsubstituted C2~C12 thiazolinyl, substituted or unsubstituted C2~C12 alkynyl,
Any one in substituted or unsubstituted C6~C30 aryl, substituted or unsubstituted C1~C30 heteroaryl.
Preferably, R therein1Selected from hydrogen, hydroxyl or alkoxyl, R3Selected from hydroxyl or alkoxyl, R4、R8、R9
It is each independently selected from hydrogen or hydroxyl, R2、R5、R6It is each independently selected from hydrogen, R7Selected from hydroxyl.
As further preferred scheme, described Bavachalcone and the like be have following structural formula compound or
The pharmaceutically acceptable salt of described compound, tautomer, stereoisomer or precursor compound:
The dosage form of compositions of the present invention can be diversified, as long as it is internal that active component can be made effectively to arrive
Dosage form be all possible.Such as be selected from: tablet, capsule, powder, granule, syrup, solution, suspension,
Injection, tincture, oral liquid, aerosol, to suck common dosage forms or the nanometer formulations etc. such as agent, electuary, pill, powder slow
Release dosage form.
Effective application dosage of active component of the present invention can be with compositions used, the pattern of administration and disease to be treated
The order of severity and change.But, generally give with the dosage of about 0.5-20mg/kg body weight every day when the reactive compound of the present invention
When giving, gratifying effect can be obtained, preferably give with 1-3 separate dosage every day, or be administered with sustained release forms.
It is applicable to the dosage form taken orally, comprises reactive compound intimately mixed with solid-state or liquid carrier.This dosage side of scalable
Case is to provide optimal treatment, such as: by an urgent demand for the treatment of situation, can give the most separate dosage every day, or will
Dosage reduces pari passu.
It addition, those skilled in the art should be understood that after knowing the structure of the compounds of this invention, also by multiple this area
Known to method, utilize known raw material, obtain the analog of the present invention, such as chemosynthesis or extraction etc. from plant.
Heretofore described term is defined as follows:
Term " Bavachalcone " is used interchangeably with " Bavachalcone ", and its English synonym also has " Roussochalcone
B ", " Broussochalcone B ", refer both to compound:
(E)-1-[2,4-Dihydroxy-5-(3-methyl-2-butenyl)phenyl]-3-(4-hydroxyphenyl)-2-propen-1-one
[(E)-1-[2,4-dihydroxy-5-(3-methyl-2-butene base) phenyl]-3-(4-hydroxy phenyl)-2-propylene-1-ketone].
Term " C1-C12 alkyl " refers to the straight or branched alkyl or cycloalkyl with 1-12 carbon atom, such as: methyl,
Methylene, ethyl, propyl group, isopropyl, butyl, isobutyl group, sec-butyl, the tert-butyl group, cyclopropyl, cyclobutyl, ring penta
Base, cyclohexyl, suberyl, or similar group.
Term " C2-C12 thiazolinyl " refers to straight or branched that have 2-6 carbon atom, that have one or more double bond
Thiazolinyl or cycloalkenyl group, such as: vinyl, pi-allyl, 1-acrylic, isopropenyl, 1-butylene base, crotyl, hexamethylene
Thiazolinyl, cycloheptenyl, 1,3-cyclohexadienyl, 1,4-cyclohexadienyl or similar group.
Term " C2-C12 alkynyl " refers to straight or branched that have 2-12 carbon atom, that have one or more three keys
Alkynyl, such as: acetenyl, propinyl or similar group.
Term " C6~C30 aryl " refers to the aryl with 6~30 carbon atoms, including monocycle or aryl bicyclic, such as:
Phenyl, naphthyl or similar group.
Term " C1~C30 heteroaryl " refers to the heteroaryl with 1~30 carbon atom, such as: pyrrole radicals, pyridine radicals,
Furyl or similar group.
Term " replace " the one or more hydrogen atoms referring on group be selected from lower group substituent group replace: C1~C10 alkane
Base, C3~C10 cycloalkyl, C1~C10 alkoxyl, halogen, hydroxyl, carboxyl (-COOH), C1~C10 aldehyde radical,
C2~C10 acyl group, C2~C10 ester group, amino, phenyl;Described phenyl includes unsubstituted phenyl or has 1-3
The substituted-phenyl of substituent group, described substituent group is selected from halogen, C1-C10 alkyl, cyano group, OH, nitro, C3~C10 ring
Alkyl, C1~C10 alkoxyl or amino.
Term " pharmaceutically acceptable salt " refers to that described compound is formed with pharmaceutically acceptable mineral acid or organic acid
Salt, described mineral acid includes: hydrochloric acid, hydrobromic acid, phosphoric acid, nitric acid, sulphuric acid;Described organic acid includes: formic acid,
Acetic acid, propanoic acid, succinic acid, naphthalenedisulfonic acid (1,5), asiatic acid, oxalic acid, tartaric acid, lactic acid, salicylic acid, benzoic acid,
Valeric acid, diethacetic acid, malonic acid, succinic acid, fumaric acid, 1,5-pentanedicarboxylic acid., adipic acid, maleic acid, malic acid, amino
Sulfonic acid, benzenpropanoic acid, gluconic acid, ascorbic acid, nicotinic acid .gamma.-pyridinecarboxylic acid, methanesulfonic acid, p-methyl benzenesulfonic acid, citric acid, and
Aminoacid.
Term " tautomer " refers to the functional group's isomery produced because atom a certain in molecule moves rapidly two positions
Body, such as: enol and corresponding ketone.
Term " stereoisomer " refers to by isomer produced by molecule Atom spatially arrangement mode difference, such as:
Cis-trans-isomer, enantiomer, conformer etc..
Term " precursor compound " refers to the most inactive, but can carry out metabolism or chemical reaction conversion in vivo
For the active component of the present invention, thus play the compound of its pharmacological action.
Term " compositions " refers in the composition, in addition to containing main active, and also can be containing a small amount of and not
Affect the submember of effective ingredient and/or pharmaceutically acceptable carrier.For example, it is possible to containing sweeting agent with improve taste,
Antioxidant is in case oxidation, and adjuvant necessary to various preparation.
Term " pharmaceutically acceptable " refers to be applicable to people and without excessive bad side reaction (such as toxicity, stimulation and allergy),
I.e. there is the material of rational benefit/risk ratio.
The result of study of the present invention shows: Bavachalcone and the like can directly activate the activity of ROR α, and presents
Dose dependent, and concentration dependent ground activates ROR α albumen and ROR α mrna expression and can effectively regulate and control under it
The expression of trip clock gene BMAL1;Illustrate that Bavachalcone and the like is that a good ROR α forward swashs
Agent alive, can be used for prevention or treats by relevant disease alpha mediated for ROR, having wide application prospect.
Accompanying drawing explanation
Fig. 1 shows the activation to ROR α reporter gene of the Bavachalcone of variable concentrations;
Fig. 2 shows the Bavachalcone impact on ROR α expressing quantity;
Fig. 3 shows the Bavachalcone impact on ROR α mrna expression amount;
Fig. 4 shows the Bavachalcone impact on clock gene BMAL1 mrna expression amount;
Fig. 5 shows the isobavachalcone impact on ROR α mrna expression amount.
Detailed description of the invention
Below in conjunction with specific embodiment, the present invention is expanded on further.Should be understood that these embodiments be merely to illustrate the present invention and
It is not used in restriction the scope of the present invention.The experimental technique of unreceipted actual conditions in the following example, generally according to normal condition
Or according to the condition proposed by manufacturer.Unless otherwise indicated, otherwise percentage ratio and number are calculated by weight.
Embodiment
1. experiment material and equipment
1.1 experiment reagent
Cell strain: HUVEC cell strain is purchased from ATCC company of the U.S. (HUVEC, CRL-1730).
Antibody: HRP-conjugated Monoclonal Mouse Anti-glyceraldehyde-3-phosphate Dehydrogease
(GAPDH, KC-5G5) becomes biological company limited purchased from upper Haikang;Anti-ROR α antibody (ab60134) is purchased from abcam
Company.
Test kit: the Promega Luciferase Assay System (Promega, USA);High-Capacity cDNA
Reverse Transcription Kits is purchased from Applied Biosystems company.
Reagent: Trizol Reagent is purchased from American I nvitrogen company;15596-026 is public purchased from Applied Biosystems
Department;RIPA lysate (weak, P0013D), BCA determination of protein concentration test kit (enhancement mode, P0010), Triton X-100/
Triton x-100 (ST795), Phenylmethanesulfonyl fluoride PMSF (100mM, ST506) are purchased from green skies biotechnology research institute;
Dodecyl sodium sulfate (sodium dodecyl sulfate, SDS), beta-mercaptoethanol, glycine (Glycine), Tris alkali, third
Dilute amide, methene double acrylamide, Ammonium Persulfate 98.5 (ammonium persulfate, APS), tetramethylethylenediamine
(tetramethylenediamine, TEMED), bovine serum albumin (Bovine Serum Albumin, BSA), Tween-20
It is purchased from Sigma company;ECL Immobilon Western Chemiluminescent HRP Substrate(WBKLS0500)
Purchased from Millipore company;Protein transfer film (nitrocellulose filter, NC film) is purchased from Pall company of the U.S.;HD Transfection
Reagent (E231A) is purchased from U.S. Promega;Cell culture medium DMEM height sugar, DMEM low sugar, RPMI1640,
FCS, 0.25% trypsin is purchased from U.S. GIBCO;Ampicillin (A6140), DMSO are purchased from Sigma;EDTA
Purchased from Shanghai biological engineering;Tryptone (LP0042) is purchased from OXOID.
1.2 experimental apparatus
CO2Cell culture incubator (HEPA CLASS 100, Thermo company of the U.S.), and Biohazard Safety Equipment (Delta serAes, beautiful
State Labconco), Tissue Culture Dish, 96 porocyte culture plates (CornAng company of the U.S.), the multi-functional reading of full wavelength scanner formula
Number instrument (VarAoskan Flash, Thermo scAentAfAc, the U.S.), GSP-9050MBE water isolation type constant incubator,
BJ-1CD antibacterial super-clean bench (Medical Equipment Plant of Shanghai Boxun Industrial Co., Ltd.), PureYAeldTM PlasmAd MAdAprep
System (U.S. Promega A2492), inverted phase contrast microscope (AX71, Japan Olympus), PCR amplification instrument
(thermocycler, BAometra company of Germany), protein electrophorese and transferring system (BAORAD company of the U.S.), QL-901
The mini centrifuge of Vortex, LX-100 Palm type centrifugal machine, LX-200, TS-1 type decolorization swinging table, TS-8 transfer decolouring is shaken
Bed, TS-8 Transference DecolorAng Shaker are Jiangsu Haimen City its woods Bel instrument manufacturing company limited and produce,
Plastic film sealing machine (FS-200 type, Wenzhou Zheng Xiong Machinery Co., Ltd.), (Ningbo is new for JY92-2D ultrasonic cell disruptor
Sesame biotech inc), 2600C automatic X-ray film developing machine, Ancubator shaker constant-temperature table
(ZHWY-211B, ZHWY-100B, Shanghai intelligence is sincere).
2. experimental technique
2.1 cells are cultivated
HUVEC with DMEM (low sugar) culture medium culturing containing 10%FCS in 10cm culture dish, in saturated humidity
Containing 5%CO237 DEG C of constant temperature culture of cell culture incubator, grow to pass on during 80%~90% fusion, and PBS washes 2 times,
0.25% pancreatin (containing 0.02%EDTA) digests about 4min, adds the DMEM culture medium containing serum and terminates digestion, collects cell,
Room temperature 800 rpm/min, 3min are centrifugal, abandon supernatant, add the culture medium containing 10%FCS, 1:3 Secondary Culture.Take
4-8 is used for testing for cell.
2.2 reporter gene
The previous day of cell inoculation, spread 24 orifice plates, transfect when cell grows to 80%.24 orifice plates are being transfected
On the same day, dilute in the serum-free medium of DNA to 50 μ L of 2 μ g, then take 2 μ L FuGENE HD transfection reagents
(Promega company, the U.S.) adds DNA in the serum-free medium of 50 μ L to and mixes with transfection reagent and hatch 15 minutes
After, add in each hole.After transfecting 4-6 hour, sop up old culture medium, add fresh culture, after one hour, add
Enter Bavachalcone or its analog so that it is final concentration of 2.5 μMs, 5 μMs, 10 μMs and 20 μMs.After 16 hours,
Sop up old culture medium, wash twice with PBS, add the lysate cell lysis of 70 μ L, be put in shaking table and sway 5min, 12000rpm,
4 DEG C, 5min is centrifuged, and adds double fluoroscopic examination liquid detection.
2.3 Western blot analysis
2.3.1 the extraction of total protein
1) discarding culture medium, every ware cell adds the PBS washed cell of 4 DEG C of pre-coolings of 3mL, discards PBS and is placed on ice;
2) melting RAPA lysate, add PMSF before using, the ultimate density making PMSF is 1mM, shakes up as ice
On;
3) every 100mm ware adds 200 μ L lysates, cracks 5 minutes on ice;
4) with cell scraper, cell is scraped, in the centrifuge tube of the 1.5mL collecting sterilizing;
5) ultrasonic cell disruption instrument ultrasonication, until fully cracking;
6) 4 DEG C, 12000rpm/mAn is centrifuged 10min, takes supernatant.
2.3.2 BCA method measures protein concentration
1) preparation BCA working solution: according to sample size, BCA reagent A is prepared by the ratio of 50:1 with B by volume
For appropriate working solution, fully mix;It is completely dissolved protein standard substance, is diluted to the concentration of 0.5mg/mL with PBS;
2) being added in the standard sample wells of 96 orifice plates by 0,1,2,4,8,12,16,20 μ L by standard substance, PBS supplies
20μL;
3) add after protein sample suitably being diluted in 20 μ L sample to 96 orifice bores;
4) each hole adds 200 μ L BCA working solutions, 37 DEG C of reaction 30min on constant temperature microwell plate concussion instrument;
5) microplate reader measures 562nm light absorption value, calculates protein concentration according to standard curve.
2.3.3 PAGE gel electrophoresis protein sample is prepared
According to protein concentration, adjust loading volume, loading total amount with PBS consistent, with 4 × loading buffer according to 4:
1 volume adds sample-loading buffer, mixing;In thermostat water bath 95 DEG C, 5min degeneration, carry out follow-up PAGE gel
It is separated by electrophoresis or is stored in-80 DEG C of refrigerators.
2.3.4 Western Blot
1) glue (PAGE gel): prepare the resolving polyacrylamide gel of 12% according to the molecular weight of destination protein, will
The gel liquid implantation glass plate prepared, dehydrated alcohol 200 μ L sealing, prepare after gelling to be separated is solid and inject 4% dense
Contracting glue, inserts 10 hole combs of 1.5mm thickness;
2) electrophoresis: install gel slab, adds 1 × electrophoretic buffer, adds albumen pre-dyed Marker and each group in gel lane
Sample, with BAORAD mini protein electrophorese transferring system separation albumen, starts electrophoresis from 80V, transfers to after 50min
120V continue electrophoresis to dyestuff close to gel bottom, stop electrophoresis;
3) transferring film (wet robin): NC film and 4 filter paper and 4 fiber mats are immersed in and have transfering buffering liquid (methanol 20%)
Dish in, from the bottom to top according to the order of " 2 fiber mat-2 metafiltration paper-gel-NC film-2-2 layers of fiber mats of metafiltration paper ",
Noting getting rid of bubble, be placed in transferring film groove, side is on the rocks, is embedded in ice basin by whole transferring film groove, and constant current 400mA, 1h turn
Film;
4) close: film is immersed in the confining liquid containing 5% defatted milk powder or 5%BSA (using TBS-Tween buffer solution), room
Temperature shakes 1h slowly;
5) Western Immuno detection: by membrane closure in the hybridization bag containing one anti-(dilution ratio is 1:1000), be placed in eluting
On shaking table, 4 DEG C of overnight incubation;Second day, quickly wash film 8min × 3 time with TBS-T buffer;Again film is enclosed HRP
In anti-(dilution ratio the is 1:1000) hybridization bag of the two of coupling, room temperature shakes 1h slowly;Film 8min × 3 are quickly washed with TBS-T buffer
Secondary;
6) substrate chemiluminescence and development: paper using blots the moisture on film, lies against on preservative film, adds ECL (A:B 1:1 preparation)
Reaction 2min, paper using sucks unnecessary ECL, wraps with preservative film and lies against in magazine, and protein powder upwards, will turn in darkroom
Photographic film is exposed by die, and automatic film developer is developed a film;
7) result and analysis: by film scanning, calculate each band optical density value with gel analysis software Quantity One, carry out
Statistical analysis.
2.4 RNA extract and quantitative fluorescent PCR
1) sopping up cell waste liquid, PBS washes twice, and 60mm ware adds 1mL trizol, and room temperature places 5min;
2) every 1mL Trizol adds 0.2mL chloroform, cover tightly sample lid, exert oneself shaking test tube 15 seconds with hands so that it is fully
Mixing, room temperature stands 5 minutes;4 DEG C, 12000rmp is centrifuged 15 minutes;
3) upper strata aqueous phase is proceeded to (about 400-500 μ L) in new 1.5mL EP pipe, adds 0.5mL isopropanol, after mixing
Room temperature places 10min;4 DEG C, 12000rmp is centrifuged 10 minutes;
4) carefully outwell supernatant, leave and take precipitation;Add ethanol (pre-cooling) the vibration washing RNA precipitate of 1mL now joins 75%
Once, 4 DEG C, 7500rmp be centrifuged 5 minutes;
5) outwell supernatant, leave and take precipitation;Add 1mL ethanol (pre-cooling) vibration washing RNA precipitate more once, 4 DEG C, 7500rmp
Centrifugal 5 minutes;
6) carefully outwell supernatant, take precipitation and put the superclean bench machine that blows in and dry up that (about 10 minutes, now RNA precipitate became thoroughly
Bright);Note allowing RNA precipitate be completely dried (its ground solubility can be greatly reduced);In pipe, add 20 μ L's again
DEPC water dissolution, hatches 10 minutes hydrotropies at 55-60 DEG C;
7) RNA is surveyed concentration, unified furnishing 0.5 μ g/ μ L, carry out reverse transcription;CDNA is put in-80 DEG C standby;
8) quantitative fluorescent PCR is carried out to specifications.
The extraction of 2.5 biological clock RNA
1) plating cells the previous day, 1,200,000/ware;After cell grows to 80%, thin with the culture medium Nature enemy of 0.3% serum
Born of the same parents 16 hours;
2), after 16 hours, with the medium treatment two hours of 50% serum, change 0.5% blood serum medium the most again into and cultivate,
A portion cell adds the Bavachalcone of 20 μMs and carries out identical process;
3) to be zero moment after hungry 16 hours, collected cell RNA every four hours, collect 48 hours altogether;
4) after collection terminates, unified extracting RNA, reverse transcription, PCR.
2.6 data process and statistical analysis
Using GraphPad PrAsm 5 software or SPSS15.0 software to carry out data process, data are with mean ± standard deviation
(mean ± SD) represents, compares employing pairing t-inspection and carries out statistical analysis, compare employing one factor analysis of variance in group between group
Method (one-way ANOVA) analyzes and processes, and p 0.05 thinks there is significant difference, has statistical significance.
3. experimental result
Fig. 1 shows the activation to ROR α reporter gene of the Bavachalcone of variable concentrations.As seen from Figure 1: transfection ROR α
After reporter gene 5 hours, add after normal incubation medium hatches for 1 hour, then with 2.5 μMs, 5 μMs, 10 μMs, 20 μMs four
Concentration process ROR α reporter gene, after 6 hours, is illustrated by the testing result of double fluorescent reporter gene: Bavachalcone is dense
The expression of ROR α in degree dependency ground active cell.Data therein are by three independent experiment mean+SD;* P < 0.05,
Compare with CTRL group;* P < 0.01, compares with CTRL group.
Fig. 2 show the Bavachalcone impact on ROR α expressing quantity, A figure therein show ROR α and
GAPDH aggregate level changes, and B figure shows ROR α band statistical result;As seen from Figure 2: huve cell (HUVEC)
After the Bavachalcone of various dose processes 24 hours, the evaluation result of immunoblotting illustrate: Bavachalcone
ROR alpha active in concentration dependent ground active cell.Data therein are by three independent experiment mean+SD;* P < 0.05,
Compare with CTRL group;* P < 0.01, compares with CTRL group.
Fig. 3 shows the Bavachalcone impact on ROR α mrna expression amount, as seen from Figure 3: umbilical vein endothelium is thin
Born of the same parents (HUVEC) are after the Bavachalcone of various dose processes 24 hours, by the testing result of fluorescence quantifying PCR method
Illustrate: ROR alpha active in Bavachalcone concentration dependent ground active cell.Data therein are put down by three independent experiments
Means standard deviation;* P < 0.05, compares with CTRL group;* P < 0.01, compares with CTRL group.
Fig. 4 shows the Bavachalcone impact on clock gene BMAL1 mrna expression amount, as seen from Figure 4: umbilicus is quiet
Arteries and veins endotheliocyte (HUVEC) first processes 16 hours with 0.3% serum, then with 50% blood serum medium and 50% serum+20 μMs
The medium treatment of Bavachalcone 2 hours, finally processes thin with 0.5% serum or+20 μMs of Bavachalcones of 0.5% serum
Born of the same parents, the RNA result explanation by collecting every 4 hours: Bavachalcone can regulate and control the expression of clock gene BMAL1.
Fig. 5 shows the isobavachalcone impact on ROR α mrna expression amount, as seen from Figure 5: huve cell
(HUVEC) after the isobavachalcone at 20 μMs of dosage processes 24 hours, the testing result of fluorescence quantifying PCR method say
Bright: isobavachalcone has ROR alpha active in active cell.Data therein are by three independent experiment meansigma methods ± standards
Difference;* P < 0.05, compares with CTRL group.
The full name of the abbreviation described in embodiment is as shown in the table:
Abbreviation | English full name | Chinese full name |
PBS | Phosphate buffered saline | Phosphate buffer solution |
PMSF | Phenylmethyl sulfonylfluoide | Benzylsulfonyl chloride compound |
RORα | Retinoid acid receptor related orphan receptorα | The relevant lonely nuclear receptor α of retinoic acid |
DMSO | Dimethyl sulfoxide | Dimethyl sulfoxide |
HRP | Horseradish peroxidase | Athomin peroxidase |
GAPDH | Glyceraldehyde 3 phosphate dehydrogenase | Glyceraldehyde 3-phosphate dehydro-genase |
HUVECs | Human Umbilical Vein Endothelial Cells | Human umbilical vein endothelial cells |
HEK293 | Human Embryonic Kidney Cells 293 | HEKC 293 |
To sum up result of study is visible: Bavachalcone and the like can directly activate the activity of ROR α, and presents dosage and depend on
Lai Xing, and concentration dependent ground activates ROR α albumen and ROR α mrna expression and can effectively regulate and control biological clock downstream
The expression of gene BMAL1;Illustrate that Bavachalcone and the like is a good ROR α forward activator, available
In preventing or treating by relevant disease alpha mediated for ROR, there is wide application prospect.
Finally it is necessary described herein: those skilled in the art can carry out Fructus Psoraleae on the basis of above-described embodiment
The above-mentioned activity experiments of other analog of chalcone, will not enumerate at this.In addition, it is to be understood that reading this
After bright foregoing, the present invention can be made various changes or modifications by those skilled in the art, but the equivalent form of value done
Fall within the protection domain required by the application claims equally.
Claims (9)
1. the purposes of Bavachalcone and the like, it is characterised in that: with Bavachalcone or/and its analog
It is used for preparing the compositions of the relevant lonely nuclear receptor-α (ROR α) of activation retinoic acid as active component.
Purposes the most according to claim 1, it is characterised in that: using Bavachalcone or/and its analog is as activity
Composition is used for preparing the activator of the relevant lonely nuclear receptor-α (ROR α) of retinoic acid.
Purposes the most according to claim 1, it is characterised in that: using Bavachalcone or/and its analog is as activity
Composition is used for preparing the compositions of the relevant disease that prevention or treatment are mediated by the relevant lonely nuclear receptor-α (ROR α) of retinoic acid.
4. according to the purposes described in claim 1 or 3, it is characterised in that: described compositions is pharmaceutical composition, health product
Compositions or food compositions.
Purposes the most according to claim 3, it is characterised in that: described relevant disease is cardiovascular and cerebrovascular disease, metabolism
At least one in property disease, nervous system disease, skeleton disease, Cancerous disease or inflammation disease.
Purposes the most according to claim 5, it is characterised in that: described cardiovascular and cerebrovascular disease is hypertension, tremulous pulse medicated porridge
At least one in sample hardening;Described metabolic disease is diabetes, abnormal carbohydrate metabolism, obesity, hyperlipidemia, bile
At least one in Developmental and Metabolic Disorder;Described nervous system disease is at least one in cerebral atrophy, neurodegenerative diseases;
Described skeleton disease is at least one in osteoporosis, rheumatoid arthritis;Described Cancerous disease be breast carcinoma,
At least one in hepatocarcinoma, colorectal carcinoma, carcinoma of prostate, early children's grain leukemia;Described inflammation disease is that aseptic is scorching
At least one in disease, chronic obstructive pulmonary disease, nonalcoholic steatohepatitis, pneumonia.
7. according to the purposes described in any one of claim 1-3, it is characterised in that described Bavachalcone and similar
Thing is compound or the pharmaceutically acceptable salt of described compound, tautomer, the stereoisomer with Formulas I structure
Or precursor compound:
Wherein: R1、R2、R3、R4、R5、R6、R7、R8、R9Be each independently selected from hydrogen, ORa, OCOORa,
OCONRaRb、OSO2Ra、OSO3Any one in Ra;Described Ra and Rb be each independently selected from hydrogen, replacement or
Unsubstituted C1~C12 alkyl, substituted or unsubstituted C2~C12 thiazolinyl, substituted or unsubstituted C2~C12 alkynyl,
Any one in substituted or unsubstituted C6~C30 aryl, substituted or unsubstituted C1~C30 heteroaryl.
Purposes the most according to claim 7, it is characterised in that: described Bavachalcone and the like is for having
The compound of Formulas I structure or the pharmaceutically acceptable salt of described compound, tautomer, stereoisomer or precursor
Compound, wherein: R1Selected from hydrogen, hydroxyl or alkoxyl, R3Selected from hydroxyl or alkoxyl, R4、R8、R9It is each independently selected from
Hydrogen or hydroxyl, R2、R5、R6It is each independently selected from hydrogen, R7Selected from hydroxyl.
Purposes the most according to claim 8, it is characterised in that: described Bavachalcone and the like is for having
The compound of following structural formula or the pharmaceutically acceptable salt of described compound, tautomer, stereoisomer or precursor
Compound:
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Cited By (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN110025603A (en) * | 2019-05-15 | 2019-07-19 | 大连天星本草生物科技有限公司 | Application of the Bavachalcone compounds in the drug of preparation treatment obesity |
WO2020116382A1 (en) * | 2018-12-06 | 2020-06-11 | サントリーホールディングス株式会社 | Composition for suppressing blood pressure elevation and method for suppressing blood pressure elevation |
WO2020116381A1 (en) * | 2018-12-06 | 2020-06-11 | サントリーホールディングス株式会社 | Blood flow improvement composition and vascular endothelium function improvement composition |
WO2021131569A1 (en) * | 2019-12-25 | 2021-07-01 | サントリーホールディングス株式会社 | Composition for improving sleep and composition for improving circadian rhythm |
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Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20030035851A1 (en) * | 2001-02-08 | 2003-02-20 | Sophie Chen | Anti-cancer agents and method of use thereof |
CN1816328A (en) * | 2003-05-02 | 2006-08-09 | 宝生物工程株式会社 | Therapeutic agent |
CN101288666A (en) * | 2008-04-16 | 2008-10-22 | 北京珅奥基医药科技有限公司 | Application of iso-bavachalcone in preparing drug for anti-tumor, anti-osteoporosis and anti-senile dementia |
CN102772393A (en) * | 2012-07-04 | 2012-11-14 | 天津中医药大学 | Application of isobavachalcone in treating nerve inflammatory diseases |
-
2015
- 2015-02-09 CN CN201510064923.8A patent/CN105982881A/en active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20030035851A1 (en) * | 2001-02-08 | 2003-02-20 | Sophie Chen | Anti-cancer agents and method of use thereof |
CN1816328A (en) * | 2003-05-02 | 2006-08-09 | 宝生物工程株式会社 | Therapeutic agent |
CN101288666A (en) * | 2008-04-16 | 2008-10-22 | 北京珅奥基医药科技有限公司 | Application of iso-bavachalcone in preparing drug for anti-tumor, anti-osteoporosis and anti-senile dementia |
CN102772393A (en) * | 2012-07-04 | 2012-11-14 | 天津中医药大学 | Application of isobavachalcone in treating nerve inflammatory diseases |
Non-Patent Citations (1)
Title |
---|
梁正等: "异补骨脂查尔酮抑制IL-4 产生及其机制研究", 《中草药》 * |
Cited By (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
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WO2020116381A1 (en) * | 2018-12-06 | 2020-06-11 | サントリーホールディングス株式会社 | Blood flow improvement composition and vascular endothelium function improvement composition |
JPWO2020116382A1 (en) * | 2018-12-06 | 2021-10-21 | サントリーホールディングス株式会社 | Composition for suppressing blood pressure increase and method for suppressing blood pressure increase |
JPWO2020116381A1 (en) * | 2018-12-06 | 2021-10-21 | サントリーホールディングス株式会社 | Composition for improving blood flow and composition for improving vascular endothelial function |
JP7352570B2 (en) | 2018-12-06 | 2023-09-28 | サントリーホールディングス株式会社 | Composition for improving blood flow and composition for improving vascular endothelial function |
CN110025603A (en) * | 2019-05-15 | 2019-07-19 | 大连天星本草生物科技有限公司 | Application of the Bavachalcone compounds in the drug of preparation treatment obesity |
WO2021131569A1 (en) * | 2019-12-25 | 2021-07-01 | サントリーホールディングス株式会社 | Composition for improving sleep and composition for improving circadian rhythm |
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CN114848795B (en) * | 2021-02-03 | 2023-04-14 | 四川大学 | Application of RORa protein and agonist thereof in preparation of anti-aging drugs |
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