CN110478430A - Application of the civil and military soup of a thousand pieces of gold in the drug that retinal pigment epithelium ARPE-19 fibrosis is intervened in preparation - Google Patents
Application of the civil and military soup of a thousand pieces of gold in the drug that retinal pigment epithelium ARPE-19 fibrosis is intervened in preparation Download PDFInfo
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Abstract
The present invention relates to application of the civil and military soup of a thousand pieces of gold in the drug that retinal pigment epithelium ARPE-19 fibrosis is intervened in preparation, belong to technical field of traditional Chinese medicines.The present invention provides application of the civil and military soup of a thousand pieces of gold in the drug that retinal pigment epithelium ARPE-19 fibrosis is intervened in preparation.Compared with model group, the civil and military soup of a thousand pieces of gold can raise E-cad protein expression with dosage-dependent manner, lower α-SMA and COL-I protein expression.
Description
Technical field
The present invention relates to technical field of traditional Chinese medicines, and in particular to the civil and military soup of a thousand pieces of gold intervenes retinal pigment epithelium in preparation
Application in the drug of ARPE-19 fibrosis.
Background technique
The civil and military soup of a thousand pieces of gold (Qian-Jin-Wen-Wu Decoction, QWD) come from towards medical literature " east doctor four-quadrant it is new
Compile ", it is made of pueraria lobata, radix scutellariae, rhizoma ligustici, Chinese yam, Schisandra chinensis, campanulaceae, cimicifugae foetidae, the root of Dahurain angelica and nine taste medicine of Radix Ophiopogonis, cures mainly lunar people and disappear
Thirsty nephrosis, is clinically used for diabetes and its complication is significant in efficacy.Early period, pharmaceutical research showed it to RBP ELISA
Expression have inhibiting effect, can slow down or inhibit diabetic nephropathy (Diabetic kidney disease, DKD) rat view
Film lesion disease incidence and the course of disease, but its specific mechanism of action needs to be probed into.
Summary of the invention
The purpose of the present invention is to provide the civil and military soup of a thousand pieces of gold to intervene retinal pigment epithelium ARPE-19 fiber in preparation
Application in the drug of change.
The present invention provides the civil and military soup of a thousand pieces of gold to prepare the drug for intervening retinal pigment epithelium ARPE-19 fibrosis
In application.
The present invention also provides the civil and military soup of a thousand pieces of gold to prepare the medicine for inhibiting retinal pigment epithelium ARPE-19 fibrosis
Application in object.
The present invention also provides the civil and military soup of a thousand pieces of gold to inhibit high glucose induction retinal pigment epithelium ARPE-19 fine in preparation
Application in the drug of dimensionization.
Preferably, the standby raw material processed of the civil and military soup of described a thousand pieces of gold by pueraria lobata, radix scutellariae, rhizoma ligustici, Chinese yam, Schisandra chinensis, campanulaceae,
Cimicifugae foetidae, the root of Dahurain angelica and Radix Ophiopogonis composition.
Preferably, the standby raw material processed of the civil and military soup of described a thousand pieces of gold by pueraria lobata 10g, radix scutellariae 10g, rhizoma ligustici 10g, Chinese yam 10g,
Schisandra chinensis 5g, campanulaceae 5g, cimicifugae foetidae 5g, root of Dahurain angelica 5g and Radix Ophiopogonis 5g composition.
The present invention provides the civil and military soup of a thousand pieces of gold to prepare the drug for intervening retinal pigment epithelium ARPE-19 fibrosis
In application.The present invention can intervene high glucose induction retinal pigment epithelium ARPE-19's towards hospital " the civil and military soup of a thousand pieces of gold "
Progression of fibrosis.After test result shows high glucose induction, compared with normal group, model group cell is obvious elongated in spindle, and
E-cad protein expression is lowered, α-SMA and the up-regulation of COL-I protein expression；After (the civil and military soup of a thousand pieces of gold) is administered, compared with model group, with
Dosage-dependent manner raises E-cad protein expression, lowers α-SMA and COL-I protein expression.
Detailed description of the invention
Fig. 1 is the morphological observation result figure of different glucose serum free medium provided by the invention；
Fig. 2 is that MTT experiment provided by the invention detects the civil and military soup of a thousand pieces of gold to ARPE-19 cytotoxicity result figure；
Fig. 3 is that ARPE-19 cell provided by the invention is given cell E-cad, α-SMA and COL-I albumen table after drug
Up to western blot figure；
Fig. 4 is the albumen that ARPE-19 cell provided by the invention is given cell E-cad, α-SMA and COL-I after drug
Express histogram.
Specific embodiment
The present invention provides the civil and military soup of a thousand pieces of gold to prepare the drug for intervening retinal pigment epithelium ARPE-19 fibrosis
In application.The civil and military Tang Weichao hospital of a thousand pieces of gold " the civil and military soup of a thousand pieces of gold " of the present invention, in the present invention, the civil and military soup of a thousand pieces of gold
Standby raw material processed is made of pueraria lobata, radix scutellariae, rhizoma ligustici, Chinese yam, Schisandra chinensis, campanulaceae, cimicifugae foetidae, the root of Dahurain angelica and Radix Ophiopogonis；It is furthermore preferred that described
The standby raw material processed of the civil and military soup of a thousand pieces of gold is by pueraria lobata 10g, radix scutellariae 10g, rhizoma ligustici 10g, Chinese yam 10g, Schisandra chinensis 5g, campanulaceae 5g, cimicifugae foetidae
5g, root of Dahurain angelica 5g and Radix Ophiopogonis 5g composition.The present invention is not particularly limited the source of the civil and military soup of described a thousand pieces of gold.Specifically, of the invention
The preparation of the civil and military soup of a thousand pieces of gold preferably includes: the composition of raw material is mentioned through 10 times of amounts and 8 times of 30% ethyl alcohol of amount twice respectively
After taking mixing, it is concentrated into solid-liquid ratio 1:1；Again with 0.05g.mL-1For sample concentration, adsorbed with macroporous absorbent resin (MAR)；
Medical fluid after purification is finally again concentrated to solid-liquid ratio 1:1, after being freeze-dried, obtains the civil and military soup of a thousand pieces of gold.
The present invention also provides the civil and military soup of a thousand pieces of gold to prepare the medicine for inhibiting retinal pigment epithelium ARPE-19 fibrosis
Application in object.
The present invention also provides the civil and military soup of a thousand pieces of gold to inhibit high glucose induction retinal pigment epithelium ARPE-19 fine in preparation
Application in the drug of dimensionization.After high glucose induction, compared with normal group, model group cell it is obvious it is elongated be in spindle, and E-cad
Protein expression is lowered, α-SMA and the up-regulation of COL-I protein expression；After administration, compared with model group, on dosage-dependent manner
E-cad protein expression is adjusted, α-SMA and COL-I protein expression are lowered.
It is thin that retinal pigment epithelium is intervened in preparation to the civil and military soup of a thousand pieces of gold of the present invention combined with specific embodiments below
Application in the drug of born of the same parents' ARPE-19 fibrosis is further described in detail, and technical solution of the present invention includes but is not limited to
Following embodiment.
Embodiment 1
Material is as shown in Table 1 and Table 2 with instrument.
1 medicinal material of table and reagent
2 key instrument of table and equipment
Cell culture
1, the configuration of culture medium
Fetal calf serum is taken, is inactivated at 55 DEG C.DMEM is heated under 37 DEG C of water bath conditions.Configuration is containing 10%FBS's
DMEM culture medium.
2, cell recovery
The culture medium of formulated completion is put into 37 DEG C of water-bath and is heated, human retinal pigment's epithelium ARPE-
19 cells take out from -80 DEG C of refrigerators, are placed in 37 DEG C of water-bath and melt for use.It is good in ventilation equipment, lighting apparatus
Super-clean bench in, with 75% alcohol wipe testing stand, light alcolhol burner, carry out culture medium bottleneck sterilizing, use liquid-transfering gun
5ml culture medium is sucked out, is placed into 15ml test tube, is transferred in test tube and is centrifuged using liquid-transfering gun absorption cells frozen storing liquid
3min(1000r·min-1) with aspirator the supernatant after centrifugation is sucked out, suitable culture medium piping and druming cell is taken until distribution is equal
It is even, it is finally transferred in culture dish, 10ml culture medium is added.Culture dish is put into 37 DEG C, the CO of 5% concentration2In incubator, training
Educate 48h.
3, cell passes on
DMEM culture medium and trypsase are taken out from refrigerator first, 37 DEG C are preheated in water-bath.In incubator
The culture dish for having covered with ARPE-19 cell is taken out, whether reaches paving with its cell quantity of micro- sem observation and cellular morphology
Full culture dish bottom 80% requires.Be up to requirement culture dish be transferred to 75% wipe across ventilation illumination it is good ultra-clean
Platform takes the culture medium of culture dish at the middle and upper levels away with aspirator, then rinses culture dish with culture medium, washes away the dead cell of floating.
Take 3ml trypsase in culture dish, uniformly rocking about 1min up and down comes into full contact with cell with trypsase, will cultivate
Ware is placed in incubator and is incubated for, and takes out after timing 3min.The culture dish in incubator is taken out, culture medium is added.Take out two
Completely new culture medium is labeled as ARPE-19 cell record experiment date and experimenter's name with marking pen, and immigration is matched in advance in right amount
Good culture medium.The culture dish in incubator is taken out, beating slides cell, observes whether cell is in flowing shape under light
State is then transferred in test tube using optical microphotograph sem observation cell dispersion.By the test tube for filling cell be put into high speed from
3min (1000rmin in scheming-1), to the time after take out.Supernatant liquor is siphoned away with aspirator, culture medium is added with liquid-transfering gun,
The cell of bottom is blown afloat, upper and lower 30 times.Cell complete after piping and druming is transferred to new culture dish, cross is then carried out and shakes
Pendulum is placed and is cultivated in incubator.37 DEG C, 5%CO2Middle cultivation 48h.
4, cell cryopreservation
When the cell in culture dish it is long to 80% or so when frozen.DMEM culture medium and tryptose are taken out from refrigerator
Enzyme is arranged in 37 DEG C of water-baths ahead of time and preheats.New disposable sterilized gloves are put on, the ultraviolet lamp closed in super-clean bench is clicked
Button, open exhaust system and fluorescent lamp, open alcolhol burner, contain 75% alcohol wipe super-clean bench.About 37 DEG C of DMEM is trained
Base, trypsase progress bottleneck sterilizing are supported, the same bottleneck sterilizing of new 15mltube is transferred to.New culture is added with liquid-transfering gun
Base washes away dead cell, then siphons away.2ml trypsase is moved into liquid-transfering gun again, is waved about 20 times in cross.Culture dish is placed in
Digest 3min in incubator, then take out and clap scattered cell, move into 5-6mlDMEM culture medium, piping and druming cell until uniformly, with from
Scheming is with 1000rmin-1, it is centrifuged 3min.Cell cryopreservation tube is taken out, (ARPE-19 is thin for the mark table note good cell category of flag
Born of the same parents), freeze the information such as date.Take out freezing liquid and with alcolhol burner sterilize bottleneck.Cell after taking out centrifugation, pumps supernatant,
1.5ml freezing liquid is moved into, piping and druming is uniformly dispersed to cell, and cell is moved into cryopreservation tube, tightens cryopreservation tube bottleneck, sealed membrane package
Cryopreservation tube after sterilizing is put in -80 DEG C of refrigerator, waits 48h, moves into liquid nitrogen container.
Mtt assay detects the civil and military soup of a thousand pieces of gold to the toxicity of ARPE-19 cell
The toxicity that drug concentration versus cell is respectively given using mtt assay measurement, so that it is determined that best administration concentration.
1, plate is planted
By DMEM culture medium, pancreatin is put into 37 DEG C of water-bath and heats.Close super-clean bench ultraviolet lamp, open illumination with
Ventilation, puts on disposable glove, sprays 75% alcohol and carries out disinfection, continue by the alcohol of the table top sprinkling 75% of super-clean bench into
Row wiping, disinfection treatment.The culture medium for covering with ARPE-19 cell in incubator is taken out, microscopically observation ARPE-19 is put into
Growing state, the ARPE-19 cell grown is taken on super-clean bench.Light alcolhol burner, culture medium, at pancreatin bottleneck disinfection
It sets.Dead cell first is washed away with PBS, siphons away PBS with aspirator.3ml pancreatin is added into culture dish, under cross rocks 30, is put into
3min in incubator.Culture dish is taken out, DMEM culture medium is added and terminates digestion, cell is transferred in test tube.It is put into superelevation
3min (1000rmin is centrifuged in fast centrifuge-1) test tube is taken out, supernatant is siphoned away, culture medium is added and dispels bottom cell, takes
15 μ l liquid are put into tally, count under the microscope, start to dilute according to the kind plate density set in advance.What is diluted
Cell suspending liquid is squeezed into 96 orifice plates, every 200 μ l of pore volume.96 orifice plates are put into incubator and cultivate 18h.
2, influence of a certain concentration drug to ARPE-19 cell is investigated
The civil and military decoction extract of a thousand pieces of gold is added in 96 orifice plates, every hole is separately added into 100 μ l drugs, (every group is successively administered 10 μ g,
50 μ g, 100 μ g, 200 μ g, 300 μ g, 500 μ g, 1000 μ g, 2000 μ g) then patted around orifice plate with hand, place into incubator
Middle processing 18h.MTT reagent (5mgml is added after 18h-1) 20 μ l, it is kept in dark place.Active cell can be MTT
The blue for being reduced to water-insoluble ties brilliant formazan, is then placed in incubator and continues to be incubated for 4h.After 4h, 100 μ l lysates are added
The crystallization dissolution of Shi formazan, continues to be put into incubator after 4h, be wrapped with tinfoil.
3 Data Detections
Spectrophotometer is opened to and set parameter, detects the absorbance of 570nm.
After a thousand pieces of gold that APRE-19 cell is given various concentration civil and military soup, as a result as shown in Fig. 2, the cell of 500 μ g groups
Survival rate is 84.76%, illustrates that (concentration is 5 μ g μ l to 500 μ g of administration-1) when without overt toxicity, and cell well-grown.
Cell modeling
By DMEM culture medium, pancreatin is put into 37 DEG C of water-bath and heats.With trypsin digestion ARPE-19 cell, put
Enter and digest 3min in incubator, 3ml culture medium is added in taking-up observation cell dissociation situation after the time arrives, what piping and druming did not digested also
The ARPE-19 cell suspension of stopping digestion in culture dish is added in test tube, test tube is put into centrifuge, is set by cell
1000r·min-1, time 3min.Test tube is taken out after to the time, siphons away supernatant liquor, it is thin that addition 3ml culture medium blows afloat bottom
Then portion enters cell kind in 6 orifice plates, after placement, then put 37 DEG C into, 5%CO2It is cultivated in incubator；In each culture
Glucose Medium without serum (the final concentration of 25 μm of olL of glucose containing various concentration are added in ware-1、5μmol/L-1、30μ
mol/L-1、40μmol·L-1、50μmol·L-1) processing 72h.Microscopically observation cellular morphology tests to obtain most by modeling
Good administration concentration is used for drug-treated cell.
APRE-19 cell gives 5 μm of olL respectively-1With 50 μm of olL-1Glucose Medium without serum.With 5 μ
mol·L-1Group is compared, 50 μm of olL-1Significant spindle is presented in group cell, and space between cells becomes larger, and degrees of fusion reduces, and shows
ARPE-19 cell is 50 μm of olL in concentration of glucose-1When model index become apparent from, therefore determine 50 μm of olL-1As modeling
Concentration of glucose.Cell modeling the result is shown in Figure 1 is observed under inverted phase contrast microscope, wherein A: give 5 μm of olL-1Grape
Sugar；B: 50 μm of olL are given-1Glucose.
Cell grouping and administration
Cell is moved in 6 orifice plates, after growing 18h, observes the growing state of cell.It is added in cell modeling in every hole
Determine the serum free medium processing 72h of concentration of glucose.Cell be divided into blank group, model group, low dose group, middle dose group and
High dose group, blank group, model group do not give drug, and low dose group administration concentration is 0.1 μ g μ l-1, middle dose group is administered dense
Degree is 2 μ g μ l-1, high dose group administration concentration is 5 μ g μ l-1.Culture observation 72h after administration.The drug in every hole is drawn, so
Every hole gives lysate 130 μ l afterwards, freeze overnight.
Protein immunoblot experiment
1, protein is extracted
Prepare cell scraping, PBS buffer solution scrapes six orifice plates with what cell scraping was exerted oneself, wipes the egg being sticked on six orifice plates off
White matter has often scraped one group and has rinsed cell scraping with PBS buffer solution.With liquid-transfering gun by the protein suspension in six orifice plates, it is transferred to
In the tube of 2ml.Tube is put into refrigerated centrifuge, 4 DEG C of centrifugation 10min, revolving speed 13000.It will be upper after centrifugation with liquid-transfering gun
Layer clear liquid is transferred in the tube of 1.5ml, and tube is put into -20 DEG C of preservations.13000r·min-1.Supernatant is collected, is put into -20
DEG C save.
2, glue
It chooses 1cm and matches glue glass plate, clean up, test leakage with tri-distilled water after being clipped with clip；Taken out from refrigerator AA,
APS, according to the separation gel and concentration glue (TEMED is first not added) for preparing 8% and 12% with glue table, concussion is mixed, and determines glass plate
It does not leak, pour out tri-distilled water and water is blotted with paper, TEMED is added in separation gel and shakes mixing, holding is sufficiently covered with, is ensuring
It under the premise of separation gel height, is injected into glass plate with the liquid-transfering gun of 1ml, space needed for reserving concentration glue, again with methanol is filled out
Full, double swerve keeps glue surface smooth, stands.After gelling to be separated is solid (about 1h), methanol is poured out, is washed three times with tri-distilled water, uses paper
It blots, tries comb, it is ensured that comb is bonded in glass plate.Concentration glue is added TEMED and shakes mixing, the implantation glass being swift in motion
Then plate is rapidly inserted into comb, it should avoid the generation of bubble；It stands after glue to be concentrated solidifies completely and extracts comb, clean
Sample hole, it is ensured that without concentration glue in sample hole, it is stand-by to dry water.If the offset plate prepared does not use immediately, tinfoil or preservative film can be used
It wraps to be put into 4 DEG C of refrigerators and refrigerate.
3, electrophoresis
Offset plate is clipped with clip and is put into electrophoresis tank, small glass plate side is inwardly；It is added new 1 between two plates ×
Runningbuffer, does not cross the small glass plate in inside, and the runningbuffer of recycling is added in surrounding；First existed with the liquid-transfering gun of 10 μ l
1 μ lmark is added in first glue hole, sequentially adds sample；Glue is run in electrophoresis, constant current (30mA), when protein example reaches gel
When the 1cm of bottom (about 1h), power supply is closed.
4, it transfers
After electrophoresis, prepares two pvdf membranes and be immersed in proper amount of methanol；It is poured into suitable container
Transbuffer is sequentially placed into clip, cotton, pvdf membrane are used in transfer.Offset plate is pried open, concentration glue is scraped off, scratches separation gel four
Week glue is placed on pvdf membrane rapidly, covers cotton, clips clip；Fixing clamp is put into transfer tank, is poured into conversion buffered
Liquid, and transfer tank is put into ice.Start to transfer, constant pressure 65V transfers 1h, and constant pressure 35V transfers 1h.
5, film is washed, antibody is pasted
Measure 100ml0.05%PBS-T20, then weigh 5g wash film with milk be dissolved in wherein, be uniformly mixed, by what is transferred
Pvdf membrane, which is put into milk, closes 1h；Pvdf membrane is then taken out to be cleaned four times with 0.05%PBS-T20, be followed successively by 15min,
5min,5min,5min；Being incubated for corresponding primary antibody, (antibody ratios are detailed in specification, with containing 5%wv-1Skimmed milk power or BSA's
The dilution of PBS-T20 buffer), 4h or 4 DEG C of incubation 12h is rocked at room temperature on shaking table；After having pasted primary antibody, then use 0.05%PBS-
T20 successively cleans 15min, 5min, 5min, 5min；Patch secondary antibody: the corresponding secondary antibody of peroxidase label is added (using containing
2.5%wv-1The PBS-T20 buffer of skimmed milk power), 1h is incubated at room temperature on shaking table；Secondary antibody takes out pvdf membrane after having shaken,
15min, 15min, 15min are successively cleaned with 0.05%PBS-T20.
6, it images
Washed transfer film is put into darkroom, A liquid and B liquid 1ml are respectively taken, is uniformly mixed with liquid-transfering gun；Prepare developer
And clear water, it opens infrared lamp and closes fluorescent lamp, pay attention to expose, clip film；After taking out film with tweezers, with paper on film
Mixing liquid siphons away, and film is put in the mixture, is rinsed with liquid-transfering gun；The film rinsed is put into clamping plate in plastic film,
Bubble removing and fixation are removed with paper, has seen whether fluorescence；If fluorescence is more apparent, the diaphragm sheared is placed on film, closes folder
In a moment, taking out film immediately, (attention cannot misplace plate, and the band to prevent showing obscures；If fluorescence is unobvious or does not observe
To fluorescence, then shut after clamping plate is placed in dark place for a period of time and carry out subsequent operation again (closing clip prevents film from drying out)；Developing
Be put into film from bottommost in liquid, and be completely soaked, when see there is band more visible spot when, that is, can be taken off film
It cleans into the water, places into fixing solution and be fixed after cleaning, as shown in Figure 3.
ARPE-19 cell is given expression result such as Fig. 4 of cell E-cad, α-SMA and COL-I albumen after drug
Shown in (###:P < 0.001vs blank group, *: P < 0.001**:P of * * < 0.01vs model group), as can be seen from figs. 3 and 4, with blank
Group is compared, and the expression of model group E-cad is substantially reduced, and the expression of COL-I, α-SMA are significantly raised, shows that model is built
It is vertical.After giving each acute drug, drug lowers the expression of α-SMA and COL-I with dosage-dependent manner, raises E-cad's
Expression.These are the result shows that the civil and military soup of a thousand pieces of gold can intervene the progression of fibrosis of high glucose induction ARPE-19 cell.
The present invention the experimental results showed that, after high glucose induction, compared with normal group, model group cell it is obvious it is elongated be in spindle
Shape, and E-cad protein expression is lowered, α-SMA and the up-regulation of COL-I protein expression；After administration, compared with model group, with dosage according to
Rely property mode to raise E-cad protein expression, lowers α-SMA and COL-I protein expression.
Towards hospital's " civil and military soup of a thousand pieces of gold " can intervene the fibrosis of high glucose induction retinal pigment epithelium ARPE-19 into
Journey.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (5)
1. application of the civil and military soup of a thousand pieces of gold in the drug that retinal pigment epithelium ARPE-19 fibrosis is intervened in preparation.
2. application of the civil and military soup of a thousand pieces of gold in the drug that preparation inhibits retinal pigment epithelium ARPE-19 fibrosis.
3. the civil and military soup of a thousand pieces of gold answering in the drug that preparation inhibits high glucose induction retinal pigment epithelium ARPE-19 fibrosis
With.
4. described in any item applications according to claim 1~3, which is characterized in that the standby raw material processed of the civil and military soup of a thousand pieces of gold
It is made of pueraria lobata, radix scutellariae, rhizoma ligustici, Chinese yam, Schisandra chinensis, campanulaceae, cimicifugae foetidae, the root of Dahurain angelica and Radix Ophiopogonis.
5. application according to claim 4, which is characterized in that the standby raw material processed of the civil and military soup of a thousand pieces of gold by pueraria lobata 10g,
Radix scutellariae 10g, rhizoma ligustici 10g, Chinese yam 10g, Schisandra chinensis 5g, campanulaceae 5g, cimicifugae foetidae 5g, root of Dahurain angelica 5g and Radix Ophiopogonis 5g composition.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000053204A1 (en) * | 1999-03-09 | 2000-09-14 | China Pharmaceutical University | A pharmaceutical composition for treating angiocardiopathy and the method of producing thereof |
| CN108743795A (en) * | 2018-08-30 | 2018-11-06 | 延吉朝耀生物科技有限公司 | A kind of prevention diabetic nephropathy towards medicament extract and its preparation method and application |
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2019
- 2019-09-30 CN CN201910942490.XA patent/CN110478430A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000053204A1 (en) * | 1999-03-09 | 2000-09-14 | China Pharmaceutical University | A pharmaceutical composition for treating angiocardiopathy and the method of producing thereof |
| CN108743795A (en) * | 2018-08-30 | 2018-11-06 | 延吉朝耀生物科技有限公司 | A kind of prevention diabetic nephropathy towards medicament extract and its preparation method and application |
Non-Patent Citations (1)
| Title |
|---|
| 谷李影等: "下调miR-25对高糖诱导视网膜色素上皮细胞系ARPE-19细胞Smad7和VEGF表达的影响", 《临床和实验医学杂志》 * |
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Application publication date: 20191122 |