CN107929286A - Application of the oxymatrine in the medicine for preparing anti-cartilage degeneration - Google Patents
Application of the oxymatrine in the medicine for preparing anti-cartilage degeneration Download PDFInfo
- Publication number
- CN107929286A CN107929286A CN201711352558.6A CN201711352558A CN107929286A CN 107929286 A CN107929286 A CN 107929286A CN 201711352558 A CN201711352558 A CN 201711352558A CN 107929286 A CN107929286 A CN 107929286A
- Authority
- CN
- China
- Prior art keywords
- oxymatrine
- cartilage
- cell
- medicine
- apoptosis
- 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.)
- Pending
Links
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
- A61K31/4375—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a six-membered ring having nitrogen as a ring heteroatom, e.g. quinolizines, naphthyridines, berberine, vincamine
Abstract
The invention discloses application of the oxymatrine in the medicine for preparing anti-cartilage degeneration.Test result indicates that：For the inhibitory action of cartilage cell's inflammatory reaction and catabolic reaction, oxymatrine protects cartilage cell in cellular level, delays its regression oxymatrine.At the same time, by establishing arthritis mouse model, oxymatrine is injected intraperitoneally, further demonstrates protective effect of the oxymatrine for cartilage in vivo.
Description
Technical field
The present invention relates to biomedicine technical field, specifically, it is related to oxymatrine and moves back preparing anti-articular cartilage
Application in the medicine of change.
Background technology
Osteoarthritis (OA) be it is a kind of common with the relevant degenerative joint disease of aging, using bone articular cartilage regression as
Main feature, clinical manifestation are mainly arthralgia, deformity and moving obstacle that progressive aggravates etc..Articular cartilage is thin by cartilage
Born of the same parents form, the denaturation of cartilage cell with it is dead play the role of in the occurrence and development of OA it is key.According to WHO Report,
The whole world have exceed 200,000,000 people endure to the fullest extent OA torment, seriously endanger patients ' life quality.The domestic report on OA illness rates has been up to
8.3%, the joint that middle-aged and elderly people bears a heavy burden greatly, more than activity is mainly in, is common in hip, knee etc., it is the most common with knee joint.In recent years
Come with world population ages, the incidence of OA shows the trend increased year by year in the world, and China is aging
Arrive, it is contemplated that incidence of the OA in China will be higher and higher.Modern medicine it is not immediately clear the lesion mechanism of OA.At present
The overall therapeutic principle of OA is that non-drug is combined with drug therapy, oral drugs such as Meloxicam etc., if necessary operative treatment,
Wherein non-operative treatment is occupied an leading position, but curative effect is not definite enough, and operative treatment removes the joint peeled off with debridement arthroscopy
Based on cartilage, equating articular surface, but offer limited effectiveness, and it is costly.
Matrine be the dry root by legume kuh-seng, plant, fruit through the organic solvents such as ethanol extract made of, be
A kind of alkaloid.Banlangen is the general designation of all alkaloids of kuh-seng, its main component with matrine, oxymatrine concentration most
It is high.Other sources are Sophora alopecuroide fruit, subprostrate sophora and subprostrate sophora aerial part, and sterling appearance is off-white color to white powder.Oxygen
Change matrine, also known as kushenin, transparent grain crystallization, 207 DEG C -208 DEG C of fusing point, soluble easily in water, methanol, chloroform, benzene, are insoluble in
Ether.Its molecular formula is C15H24N2O2, structural formula is shown in formula I.Chinese patent 201610000529.2 discloses Oxymatrine
Application of the alkali in antitumor drug sensitizer is prepared, oxymatrine have increase tumor multi-medicine drug-resistant cells against neoplastic medicine
The effect of thing sensitiveness, can use as chemotherapeutic sensitizer.Chinese patent 201410396882.8 discloses oxymatrine
The application of parenteral solution antianginal drug, Oxymatrine Injection are with 5% glucose 100ml by 2mg/ml oxymatrines
Dilution, the intravenous fluid being configured to；And take seat to be administered, its curative effect is than contrast groups nitroglycerin, isosorbidi dinitras
Improve 10%.Chinese patent 201510128920.6 discloses oxymatrine as treatment neonatal rat with hypoxic-ischemic brain damage
The purposes of vulnerary thing, test result indicates that oxymatrine dosage can reduce ischemic area cerebral infarction when being 120mg/kg weight
Volume and Neuron Apoptosis rate, mitigate cerebral tissue pathology injury, improve activities of antioxidant enzymes in brain tissue, reduce malonaldehyde and contain
Amount, oxymatrine cause injury after having the function that prevention neonates with HIBD and die and promote neurological functional recovery.
But in the prior art, on application of the oxymatrine in the medicine for preparing anti-cartilage degeneration, it yet there are no report
Road.
The content of the invention
First purpose of the present invention is to be directed to deficiency of the prior art, there is provided the pharmaceutical applications of oxymatrine.
Second object of the present invention is to be directed to deficiency of the prior art, there is provided a kind of medicine of anti-cartilage degeneration
Thing.
Third object of the present invention is to be directed to deficiency of the prior art, there is provided oxymatrine is in reagent preparation
Purposes.
To realize above-mentioned first purpose, the present invention adopts the technical scheme that：
The purposes of oxymatrine, is used to prepare the medicine of anti-cartilage degeneration.
As the preferred embodiment of the present invention, the cartilage degeneration refers to that osteoarthritic joint cartilage moves back
Become.
As the preferred embodiment of the present invention, the table of the Drug inhibition matrix metalloproteinase and inflammatory factor
Reach, suppress the activation of inflammation associated signal paths, reduce the apoptosis of cartilage cell.
To realize above-mentioned second purpose, the present invention adopts the technical scheme that：
A kind of medicine of anti-cartilage degeneration, the medicine is by oxymatrine and pharmaceutically acceptable carrier or tax
Shape agent forms.
The pharmaceutically acceptable carrier or excipient include but is not limited to：It is brine, buffer solution, glucose, water, sweet
Oil, ethanol, and combinations thereof.Pharmaceutical preparation matches with administering mode.The oxymatrine of the present invention can be made into injection
Form, such as the aqueous solution with physiological saline or containing glucose and other assistant agents are prepared by conventional method.
To realize above-mentioned 3rd purpose, the present invention adopts the technical scheme that：
Purposes of the oxymatrine in reagent preparation, the reagent are used for：
(1) matrix metalloproteinase MMP2, MMP9, MMP13 and inflammatory factor IL-6, IL-8, the expression of TNF-α are suppressed；
(2) activation of NF-KB signal paths and MAPK signal paths is suppressed；
(3) apoptosis of cartilage cell is reduced.
The invention has the advantages that：
Present invention firstly discovers that oxymatrine can be as the medicine of anti-cartilage degeneration.In cell experiment, I
Find：(1) under the stimulation of lipopolysaccharides, matrix metalloproteinase (MMP2, MMP9, MMP13) and inflammatory factor (IL-6, IL-
8, TNF-α) expression significantly increase, and in oxymatrine treatment group, the expressions of these genes has obtained bright
Aobvious suppression.(2) under conditions of lipopolysaccharides stimulation, a degree of apoptosis occurs for cartilage cell, and at oxymatrine
In reason group, the apoptosis number of cartilage cell substantially reduces.(3) under the stimulation of lipopolysaccharides, the proteoglycans of cartilaginous tissue is significantly lost
Lose, the expression of II Collagen Type VIs reduces；And in oxymatrine treatment group, the loss of proteoglycans and the drop of II Collagen Type VIs
Solution has obtained effective alleviation, shows under the effect of oxymatrine, and the regression of articular cartilage has obtained effective suppression.
(4) GAG content in lipopolysaccharides stimulation group, supernatant is significantly raised compared with untreated fish group, and in the intervention of oxymatrine
Under, organize the release of glycosaminoglycan to obtain effective suppression.(5) oxymatrine can be by suppressing inflammation coherent signal
The activation of path, and then suppress the inflammatory regression of cartilage cell.In mouse osteoarthritis, it has been found that：(1) in model
In group, mouse articular cartilage serious wear, and in oxymatrine injection group, the abrasion of mouse articular cartilage significantly mitigates.
(2) Showed by immune group result, in model group, mouse articular chondrocytes apoptosis is obvious, and oxymatrine can be notable
Reduce the apoptosis of articular chondrocytes.(3) in model group, NF-KB signal paths significantly activate in mouse layer of articular cartilage, phase
The catabolism expression of enzymes rise answered；And in oxymatrine injection group, layer of articular cartilage NF-KB activation is lighter, matrix gold
The expression of Proteases is relatively low.It is above-mentioned test result indicates that, oxymatrine have suppress cartilage cell's inflammatory reaction
With the effect of catabolic reaction, cartilage cell is protected, delays its regression, therefore can be used for preparing anti-cartilage degeneration
Medicine.
Brief description of the drawings
Toxic action of the attached drawing 1 for CCK-8 experimental studies oxymatrine to people's Primary chondrocyte.
Attached drawing 2 is the lipopolysaccharide-induced inflammatory factor of oxymatrine suppression and matrix metalloproteinase gene expression.
It is scorching that attached drawing 3 is further confirmed that oxymatrine can suppress the cartilage primary cell that lipopolysaccharides induces by ELISA
The expression of inflammation factor and matrix metalloproteinase.
Attached drawing 4 is flow cytometry analysis articular chondrocyte apoptosis situation, it was demonstrated that lipopolysaccharides can promote withering for cartilage cell
Die, and oxymatrine protection cartilage cell, reduce the articular chondrocyte apoptosis that lipopolysaccharides is induced.
Attached drawing 5 is cartilaginous tissue cultured in vitro, and oxymatrine can delay the cartilaginous tissue block that lipopolysaccharides induced
Regression.
Attached drawing 6 is that the coloration result of the cartilaginous tissue of cultured in vitro is counted.
Attached drawing 7 confirms that oxymatrine can reduce the release of cartilaginous tissue glycosaminoglycan for DMMB detections, so as to delay soft
The regression of bone.
Attached drawing 8 suppresses the activation of the NF-KB and MAPK signal paths in cartilage cell for oxymatrine.
Attached drawing 9 is that NF-KB signals in cartilage cell are led to immunofluorescence and caryoplasm Separation Research oxymatrine
The inhibitory action of road activation.
Attached drawing 10 is in zoopery, it was demonstrated that oxymatrine intraperitoneal injection can significantly inhibit moving back for mouse articular cartilage
Become, TUNEL dyeing explanations are substantially reduced in oxymatrine injection group, mouse articular cartilage apoptotic cell.
Attached drawing 11 confirms that oxymatrine injection can suppress the NF-KB signal paths in cartilaginous tissue for immunohistochemistry
Activation, suppresses the expression of matrix metalloproteinase, so as to delay cartilage degeneration.
Embodiment
The invention will be further elucidated with reference to specific embodiments.It is to be understood that these embodiments are merely to illustrate this hair
It is bright rather than limit the scope of the invention.In addition, it should also be understood that, after the content of the invention recorded has been read, art technology
Personnel can make various changes or modifications the present invention, and such equivalent forms equally fall within the application the appended claims and limited
Fixed scope.
Protective effect of 1 oxymatrine of embodiment to articular cartilage
(1) material and method
1. human articular cartilage primary cell extracts and culture
The cartilage specimens from pri of old row knee prosthesis patient is taken, under sterile working, hand is used in superclean bench
Art knife is cut into about 1mm uniform in size3Cartilage fritter, collection be placed in centrifuge tube, with PBS rinse 3 times, 3 minutes every time, fully
Clean the tissue of blood and the non-cartilaginous element of surrounding.Then, to 2.5% pancreatin of centrifuge tube plus 10mL, disappear in 37 degree of incubators
Change processing 30 minutes.Centrifuge tube is placed in a centrifuge centrifugation 5 minutes after digestion, suctions out trypsase.The 0.2%II that will be prepared
Collagenase Type is added in centrifuge tube, when digestion 6-8 is small in 37 degree of incubators, adds the complete culture containing 10% hyclone
In base and digest, centrifuge supernatant discarding after five minutes, be resuspended with the complete medium containing 10% hyclone, be inoculated in culture
In bottle, culture (37 DEG C of temperature, saturated humidity, 5% carbon dioxide) in two change carbon incubators is placed in.Daily under inverted microscope
Chondrocyte growth situation is observed, the adherent situation of visual cell replaces nutrient solution.
After growth and proliferation of cell forms cell monolayer, Microscopic observation cartilage cell aggregation is in blocks, treats that about 80% cell converges
Close, that is, carry out cell passage.Nutrient solution is absorbed, after PBS cleaning, 0.25% trypsase 2mL is added, stands digestion at room temperature
3 minutes, cytoplasm retraction is observed under inverted microscope, cell becomes round again, and space between cells increase, a small amount of cell starts to float
Trip, is neutralized rapidly with the complete medium containing 10% hyclone at this time.Centrifugation, abandons supernatant, is resuspended with complete medium, connect
Kind is in the experiment of new blake bottle or progress next step.
2.CCK-8 detects cell viability
Ex vivo cartilage primary cell is inoculated in 96 orifice plates, culture plate is stayed overnight in incubator preculture, treats that cell pastes
After wall, the oxymatrine of various concentrations is added to culture plate, concentration gradient is respectively 0.5,1,2,4mg/ml, and culture plate is existed
After incubator is incubated a period of time when small (12,24 and 48), adds 10 μ L CCK-8 solution to every hole and (be careful not in the mistake added
Bubble is generated in journey, otherwise can influence the reading of O.D values).Culture plate is incubated in incubator 1-4 it is small when after surveyed with microplate reader
The absorbance being scheduled at 450nm.
Vigor calculates：Cell viability (%)=[A (dosing)-A (blank)]/[A (0 dosing)-A (blank)] × 100%.Tool
Body：A (dosing)：The absorbance in the hole with cell, CCK-8 solution and drug solution；A (blank)：With culture medium and CCK-8
Absorbance of the solution without the hole of cell；A (0 dosing)：With cell, CCK-8 solution without the suction in the hole of drug solution
Luminosity.
3. when fluorescence quantitative PCR detection inflammatory factor and matrix metalloproteinase gene expression it is horizontal
The cartilage cell's inflammatory reaction and the influence of catabolic reaction that research oxymatrine is induced for lipopolysaccharides.
First cartilage cell is inoculated in 6 orifice plates, when cell attachment converges to 80%, first with the oxymatrine of various concentrations gradient
(0.5,1,2mg/ml) pretreatment 2 it is small when, then with lipopolysaccharides (1 μ g/ml) stimulate 24 it is small when after, Trizol methods extracting sample it is total
RNA.Culture medium in 6 orifice plates is washed off, PBS is washed one time, and Trizol lysates are added per hole, EP pipes are fully sucked after cracking
In, it is placed in -80 degree refrigerators and preserves, during follow-up extracting, takes out the cell for freezing and having cracked, room temperature, which is placed 5 minutes, makes its complete
Dissolving.The chloroform of 0.2ml is added in the sample that TRIZOL reagents per 1ml crack, covers tightly tube cover.Tube body 15 is acutely vibrated manually
After second, 15 to 30 DEG C are incubated 2 to 3 minutes.12000rpm is centrifuged 15 minutes at 4 DEG C.Mixing liquid is classified into lower floor after centrifugation
Red phenol chloroform phase, intermediate layer and colourless aqueous phase upper strata.RNA is all distributed in water phase.Carefully upper strata aqueous phase is shifted
Into the one totally centrifuge tube without RNase.Add isometric isopropanol, sightless RNA precipitate will be in bottom of the tube before centrifuging at this time
With formation gelatinous precipitate block on side wall.Supernatant is removed, adds at least 1ml's in the sample per the cracking of 1ml Trizol reagents
75% ethanol (75% ethanol is prepared with DEPCH2O), cleans RNA precipitate.After mixing, 7500rpm is centrifuged 5 minutes at 4 DEG C.Carefully
Most of ethanol solution is sucked, makes RNA precipitate 5 minutes dry in air at room temperature.Add the 40 μ l of water without RNase with rifle repeatedly
Piping and druming several times, is completely dissolved it, the RNA solution of acquisition be stored in -80 DEG C it is stand-by.
Take 1ug total serum IgEs to carry out reverse transcription, add 2ul MIX, remaining uses DEPC water polishings.Total system is put into PCR instrument progress
Reverse transcription, negates the cDNA after transcription and carries out follow-up quantitative analysis, reaction system cDNA：2ul；SYBR Green：10ul；
Upstream and downstream primer is respectively 0.8ul, ROX：0.4ul；Distilled water：6ul.After reaction, with 2-ΔΔctMethod analyzes gene expression
Difference.
4.ELISA detects inflammatory factor and matrix metalloprotease protein expression
The cartilage cell's inflammatory reaction and the influence of catabolic reaction that research oxymatrine is induced for lipopolysaccharides.
First cartilage cell is inoculated in 6 orifice plates, when cell attachment converges to 80%, first with the oxymatrine of various concentrations gradient
(1,2mg/ml) pretreatment 2 it is small when, then with lipopolysaccharides (1 μ g/ml) stimulate 24 it is small when after, Invitrogen Elisa kits inspection
Survey the matrix metalloproteinase (MMP2, MMP9 and MMP13) and inflammatory factor (IL-6 and TNF-α) in cultured chondrocytes liquid
Content.
5. flow cytomery articular chondrocyte apoptosis situation
The cartilage cell's inflammatory reaction that is induced for lipopolysaccharides of research oxymatrine and catabolic reaction apoptosis
Influence.First cartilage cell is inoculated in 6 orifice plates, when cell attachment converges to 80%, first with the oxidation of various concentrations gradient
Matrine (1mg/ml) pretreatment 2 it is small when, then with lipopolysaccharides (1 μ g/ml) co-oxidation matrine stimulate 24 it is small when after, use
The apoptosis situation of the cell apoptosis detection kit detection cartilage cell of invitrogen, analysis oxymatrine are thin for cartilage
The redemption effect of born of the same parents' apoptosis.
6. cartilage cultured in vitro detects protective effect of the oxymatrine for ex vivo cartilage tissue degeneration
Take because of Osteoarthritis and the specimens from pri of the patient of row operative treatment, blade cuts patient's cartilaginous tissue, uses knife
Piece is cut into 1mm3The cartilage block of size, is placed in 24 orifice plates and adds complete medium culture 7 days and 14 days, and experiment is divided into blank pair
According to group, the LPS stimulation groups of 10ug/ml, the oxymatrine treatment group of the LPS joints 1mg/ml of 10ug/ml, in the 7th of culture the
It and fortnight are respectively fixed cartilage block, row H＆E, Safranin O dyeing, immunohistochemistry detection cartilaginous tissue II
The expression of Collagen Type VI.After the supernatant of culture takes out at the same time, glycosaminoglycan detection kit detection cartilaginous tissue osamine gathers
Sugared release conditions.
7.Western blot
Detect influence of the oxymatrine for nf-kb and MAPK signal paths.Cartilage cell is inoculated in 6cm wares, is treated
When cell attachment converges to 80% or so, when first small with the oxymatrine pretreatment cell 2 of 1mg/ml, then stimulated with lipopolysaccharides
5,10,15,30,60 minutes, protein sample is collected at every point of time, after having collected protein sample, to ensure each albumen sample
The applied sample amount of product is consistent, and the protein concentration of each protein sample is measured with BCA detection kits.Add in the protein sample of collection
Enter the SDS-PAGE albumen sample-loading buffers of the 5X concentrated in right amount.100 DEG C or boiling water bath heat 10 minutes, to be fully denatured egg
In vain.After being cooled to room temperature, protein sample is directly loaded in SDS-PAGE glue well and carries out protein electrophoresis.Testing index bag
Include：P65, p-p65, IKB α, JNK, P-JNK, ERK, P-ERK, P38, P-P38, GAPDH etc..
8. mechanical damage makes mouse osteoarthritis, protection of the oxymatrine to articular cartilage in animal model is studied
Effect
We with the method for front fork ligament detachment carry out C57 mouse Osteoarthritis modelings for this part, after mouse anesthesia,
Preserved skin, disinfection, knee sprung insert a fine needle after front, and the anterior cruciate ligament of the detachment that swings mouse, is opened for postoperative 3 days
Begin intraperitoneal injection oxymatrine.This experiment is divided into four groups, and first group of sham-operation group, reorganization mouse is not cooked front fork ligament detachment,
Mouse is as blank control；Second group of mouse row front fork ligament detachment art makes Osteoarthritis model；3rd group of row bone joint
Scorching modeling, while low dosage oxymatrine (25mg/kg) is injected intraperitoneally；4th group of row Osteoarthritis modeling, while abdominal cavity
Inject high dose oxymatrine (50mg/kg).
Draw materials within postoperative 6th week, after mouse joint sample is fixed with 4% paraformaldehyde, decalcification, dehydration embedding, section, after going
Continuous dyeing processing.The knee joint slice row H＆E of several groups of mouse is dyed respectively, Safranin O/Fast green dyeing detections
Cartilage destruction degree, parallel OARSI joint tissues scoring, the apoptosis situation of TUNEL dyeing detection cartilage cells are same with this
When, we manage it immunohistochemical staining detection NF-KB signal paths activation situation, and matrix metalloproteinase MMP9 and
The expression of MMP13, comprehensive descision is in the case of body, and intraperitoneal injection oxymatrine is for caused by front fork ligament detachment
The retarding action of cartilage wear.
(2) result
(1) Fig. 1 is toxic action of the CCK-8 experimental studies oxymatrine to people's Primary chondrocyte.By figure, we can
To see, the drug toxicity of oxymatrine is very low, and under the concentration of 2mg/ml, oxymatrine is not bright for cartilage cell
Aobvious toxic action.
(2) Fig. 2 is the lipopolysaccharide-induced inflammatory factor of oxymatrine suppression and matrix metalloproteinase gene expression.By
Figure is it will be seen that under the stimulation of lipopolysaccharides, matrix metalloproteinase (MMP2, MMP9, MMP13) and inflammatory factor (IL-
6, IL-8, TNF-α) expression significantly increase, and in oxymatrine treatment group, the expression of these genes obtains
Obvious suppression.
(3) further research, we are with the inflammatory factor and matrix gold in ELISA detection cultured chondrocytes supernatants
The secretory volume of Proteases.The results are shown in Figure 3, it has been found that its result and the result of quantitative PCR are basically identical, this explanation
Oxymatrine can effectively suppress inflammatory reaction and the catabolic reaction of cartilage cell, so as to play protection cartilage cell
Effect.
(4) Fig. 4 is flow cytometry analysis articular chondrocyte apoptosis situation, it was demonstrated that lipopolysaccharides can promote withering for cartilage cell
Die, and oxymatrine protection cartilage cell, reduce the articular chondrocyte apoptosis that lipopolysaccharides is induced.Streaming the results show that
Under conditions of lipopolysaccharides stimulates, a degree of apoptosis occurs for cartilage cell, and in oxymatrine treatment group, cartilage cell
Apoptosis number substantially reduce.
(5) Fig. 5 is cartilaginous tissue cultured in vitro, and oxymatrine can delay the cartilaginous tissue block that lipopolysaccharides induced
Regression.In the experiment of cartilaginous tissue cultured in vitro, it has been found that under the stimulation of lipopolysaccharides, the proteoglycans of cartilaginous tissue is significantly lost
Lose, show as the loss of sarranine dyeing, at the same time, the coloration result of articular cartilage specific marker thing II Collagen Type VIs is shown,
Under the stimulation of lipopolysaccharides, the expression of II Collagen Type VIs reduces.And in oxymatrine treatment group, the loss of proteoglycans
Degraded with II Collagen Type VIs has obtained effective alleviation, shows under the effect of oxymatrine, and the regression of articular cartilage obtains
Effective suppression is arrived.
(6) Fig. 6 is that the coloration result of the cartilaginous tissue of cultured in vitro is counted.We according to Safranin O and
The intensity of II Collagen Type VIs counts histological stain, it is found that comparison among groups has significant difference.
(7) Fig. 7 is that DMMB detections confirm that oxymatrine can reduce the release of cartilaginous tissue glycosaminoglycan, so as to delay soft
The regression of bone.We have carried out the detection of GAG content for the supernatant to cartilaginous tissue cultured in vitro, it turns out that, fat is more
Sugared stimulation group, the GAG content in supernatant are significantly raised compared with untreated fish group, and under the intervention of oxymatrine, tissue
The release of glycosaminoglycan has obtained effective suppression.
(8) Fig. 8 is the activation for the NF-KB and MAPK signal paths that oxymatrine suppresses in cartilage cell.We are to oxygen
The mechanism for changing inhibitory effect of matrine cartilage cell is further studied, it has been found that in cartilage cell, oxymatrine
The activation of NF-KB signal paths can substantially be suppressed, while the activation of MAPK signal paths can also be notable by oxymatrine
Suppress.These inflammation damnification coherent signals are by activating caused cascade of response of inflammation and accelerating the inflammatory reaction of cartilage cell
And catabolism, oxymatrine inhibit the activation of these signal paths, and then inhibit the inflammatory regression of cartilage cell.
(9) further, we further demonstrate P65 in NF-KB signal paths with caryoplasm separation and immunofluorescence
Enter core situation, as shown in Figure 9.Result above is consistent with western blot results, illustrates, oxymatrine can pass through suppression
The activation of inflammation associated signal paths processed, and then suppress the inflammatory regression of cartilage cell.
(10) further, we have carried out further verification in zoopery to our result, as shown in Figure 10.
We first use H＆E, Safranin O/Fast green to dye the regression situation that have detected mouse articular cartilage.It turns out that
Compared with blank control group, in front fork ligament cut-out group, articular cartilage serious wear, partially visible cartilage strips off completely, dew
Go out subchondral bone, and in oxymatrine injection group, the abrasion of mouse articular cartilage significantly mitigates.Meanwhile we are to each group
TUNEL dyeing has been done in section, detects the apoptosis situation of mouse articular chondrocytes, the results show that in front fork ligament detachment group,
Articular chondrocytes apoptosis is obvious, and oxymatrine can substantially reduce the apoptosis of articular chondrocytes.Based on the above results,
Intraperitoneal injection oxymatrine can mitigate mouse articular cartilage damage regression.
(11) Figure 11 is that immunohistochemistry confirms that oxymatrine injection can suppress the NF-KB signal paths in cartilaginous tissue
Activation, suppress the expression of matrix metalloproteinase, so as to delay cartilage degeneration.We have detected respectively with immunohistochemical staining
The NF-KB signal paths activation situation of kind mouse articular cartilage, while have detected the table of matrix metalloproteinase MMP9 and MMP13
Up to level, the results show that in front fork ligament detachment group, NF-KB signal paths significantly activate in mouse layer of articular cartilage, accordingly
Catabolism expression of enzymes rise；And in oxymatrine injection group, layer of articular cartilage NF-KB activates lighter, matrix metal
The expression of protease is relatively low.The result shows that under concrete conditions in the establishment of a specific crime, oxymatrine shows satisfied cartilage protection and makees
Use effect.
Conclusion：The oxymatrine of suppression work we have studied to(for) cartilage cell's inflammatory reaction and catabolic reaction
With oxymatrine protects cartilage cell in cellular level, delays its regression.At the same time, we establish mouse arthritis
Model, is injected intraperitoneally oxymatrine, further demonstrates protective effect of the oxymatrine for cartilage in vivo.
The above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
Member, on the premise of the method for the present invention is not departed from, can also make some improvement and supplement, these are improved and supplement also should be regarded as
Protection scope of the present invention.
Claims (5)
1. the purposes of oxymatrine, it is characterised in that be used to prepare the medicine of anti-cartilage degeneration.
2. purposes according to claim 1, it is characterised in that the cartilage degeneration refers to osteoarthritic joint cartilage
Regression.
3. purposes according to claim 1, it is characterised in that the Drug inhibition matrix metalloproteinase and inflammatory factor
Expression, suppress the activation of inflammation associated signal paths, reduce the apoptosis of cartilage cell.
4. a kind of medicine of anti-cartilage degeneration, it is characterised in that the medicine is by oxymatrine and pharmaceutically acceptable
Carrier or excipient composition.
5. purposes of the oxymatrine in reagent preparation, it is characterised in that the reagent is used for：
(1) matrix metalloproteinase MMP2, MMP9, MMP13 and inflammatory factor IL-6, IL-8, the expression of TNF-α are suppressed；
(2) activation of NF-KB signal paths and MAPK signal paths is suppressed；
(3) apoptosis of cartilage cell is reduced.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711352558.6A CN107929286A (en) | 2017-12-15 | 2017-12-15 | Application of the oxymatrine in the medicine for preparing anti-cartilage degeneration |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711352558.6A CN107929286A (en) | 2017-12-15 | 2017-12-15 | Application of the oxymatrine in the medicine for preparing anti-cartilage degeneration |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107929286A true CN107929286A (en) | 2018-04-20 |
Family
ID=61943512
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201711352558.6A Pending CN107929286A (en) | 2017-12-15 | 2017-12-15 | Application of the oxymatrine in the medicine for preparing anti-cartilage degeneration |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107929286A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110638817A (en) * | 2019-09-29 | 2020-01-03 | 谢岩 | Application of oxymatrine in promoting scar repair |
-
2017
- 2017-12-15 CN CN201711352558.6A patent/CN107929286A/en active Pending
Non-Patent Citations (6)
| Title |
|---|
| JIANG LIANG ET AL.: "Oxymatrine prevents synovial inflammation and migration via blocking NF-κB activation in rheumatoid fibroblast-like synoviocytes", 《INTERNATIONAL IMMUNOPHARMACOLOGY》 * |
| YAFEI JIANG ET AL.: "Oxymatrine exerts protective effects on osteoarthritis via modulating chondrocyte homoeostasis and suppressing osteoclastogenesis", 《J CELL MOL MED.》 * |
| 任广慧等: "氧化苦参碱对胶原蛋白诱导大鼠关节炎症及T细胞亚群的影响", 《中国药学杂志》 * |
| 詹新堂等: "苦参碱对1L-１β诱导的小鼠体外骨关节炎的作用研究", 《蛇志》 * |
| 赵薇等: "氧化苦参碱对家兔激素性股骨头坏死的影响", 《宁夏医科大学学报》 * |
| 郭伟雄: "氧化苦参碱通过NF-κB通路抑制滑膜细胞炎症因子表达的实验研究", 《万方学位论文全文数据库》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110638817A (en) * | 2019-09-29 | 2020-01-03 | 谢岩 | Application of oxymatrine in promoting scar repair |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| KR101706642B1 (en) | Composition comprising exosomes extracted from stem cells which differentiate into chondrocytes for inducing chondrogenic differentiation or regenerating cartilage tissue | |
| JP2015164967A (en) | Compositions and methods for increasing lifespan and health span | |
| KR102159791B1 (en) | Composition for preventing or treating of liver fibrosis comprising exosomes derived from adipose stem cells as an active ingredient | |
| EP2191836A1 (en) | Prophylactic or alleviating agent for peripheral nerve disorder induced by anti-cancer agent | |
| RU2649129C2 (en) | USE OF RECOMBINANT GANODERMA LUCIDUM IMMUNOMODULATORY PROTEIN (rLZ-8) IN PREPARATION OF MEDICINE FOR TREATING MELANOMA | |
| US20190048054A1 (en) | Mesenchymal Stem Cells Expressing Biomarkers that Predict the Effectiveness of Mesenchymal Stem Cells for Treating Diseases and Disorders | |
| Fadladdin | Antischistosomal activity of Origanum majorana, Ziziphus spina-christi, and Salvia fruticosa plant extracts on Hamster Infected with Schistosoma haematobium | |
| CN107929286A (en) | Application of the oxymatrine in the medicine for preparing anti-cartilage degeneration | |
| KR20180119790A (en) | Composition for preventing or treating erectile dysfunction comprising embryonic stem cells derived exosome | |
| CN1897932A (en) | Composition for treatment of osteoarthritis containing apigenin as chindroregenerative agent | |
| US20130236445A1 (en) | Bromelain proteinases (Bromelainases) as chemotherapeutic agents in treating and/or preventing various types of cancer in a mammal | |
| US20130302279A1 (en) | Identification of natural plant extracts harboring anti-hepatitis c virus ns5b polymerase activity | |
| US20130171263A1 (en) | Snake Powder Extract For Treatment Of Cancer | |
| KR100456089B1 (en) | The method for preparing purified extract showing anti-cancer activity from wild ginseng and the composition comprising the same | |
| CN113648306A (en) | Application of bergamottin in preventing or treating osteoporosis and/or bone loss | |
| CN107898785B (en) | Application of oxymatrine in preparation of medicine for resisting osteoclast-mediated bone loss | |
| TOULAH et al. | In vitro antihelminthic activity of ethanol Zingiber officinale extract on fasciola gigantica in comparison to triclabendazole | |
| KR101802668B1 (en) | Treatment use of liver failure and recovery use of liver function by liver toxin-induced adipose tissue-derived stem cell secretome | |
| CN113768956A (en) | Effect of cell-free fat extract on macrophage polarization regulation and disease treatment | |
| KR102622408B1 (en) | Composition for inducing bone formation through osteoblast activation by Opa1 inhibition or for treating bone diseases | |
| KR20200132550A (en) | A composition for preventing or treating of blood vessel disease comprising tonsil-derived mesenchymal stem cell or conditioned medium thereof | |
| CN112569338B (en) | Application of TDFA in preparation of medicine for preventing and/or treating ocular surface inflammatory diseases | |
| KR102340457B1 (en) | Novel compositions for the treatment of cancer | |
| KR102526447B1 (en) | A composition for preventing or treating of liver disease comprising conditioned medium of tonsil-derived mesenchymal stem cell | |
| US20220193006A1 (en) | Compositions for effective management of fibroblast-like synoviocytes mediated rheumatoid arthritis |
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 | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20180420 |
|
| RJ01 | Rejection of invention patent application after publication |