CN100434072C - Medication of possessing function of anti inflammation, for improving rheological property of blood, and restraining thrombosis - Google Patents
Medication of possessing function of anti inflammation, for improving rheological property of blood, and restraining thrombosis Download PDFInfo
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- CN100434072C CN100434072C CNB200610042992XA CN200610042992A CN100434072C CN 100434072 C CN100434072 C CN 100434072C CN B200610042992X A CNB200610042992X A CN B200610042992XA CN 200610042992 A CN200610042992 A CN 200610042992A CN 100434072 C CN100434072 C CN 100434072C
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Abstract
The present invention discloses medicine for cooperatively improving blood rheological property and inhibiting thrombosis with an antiinflammatory action. Four kinds of compound formulas are formed from two kinds or three kinds among celecoxib, sodium selenite and dipyridamole for the medicine. A research for the medicine compound formulas by an applicant proves that all of celecoxib, sodium selenite and dipyridamole have the function of resisting adjuvant-induced arthritis, and the three compound formulas of celecoxib and the compound formulas of dipyridamole also have the function of resisting inflammatery arthritis. The function of resisting early inflammation of compound formula celecoxib is more obvious than that of using celecoxib singly. The compound formula celecoxib and dipyridamole selenium of the present invention have the advantage of reasonable prescription. The defect of promoting thrombosis of celecoxib is overcome. When the cooperative antiinflammatory action is exerted, the present invention also has a favorable action of cooperatively improving blood rheological property and inhibiting thrombosis.
Description
Technical field
The present invention relates to a class compound chemical medicine (new drug I (5) class) developmental research, be specifically related to have the anti-inflammatory effect and improve hemorheological property and the thrombotic compound medicine of inhibition.
Background technology
Former times dry goods (--coxib) medicine is selectivity NSAID (non-steroidal anti-inflammatory drug) (NSAIDs), it also is high selectivity Cycloxygenase-2 (Cox-2) inhibitor, a little less than the inhibitory action to Cycloxygenase-1 (Cox-1), therefore, former times dry goods and reduced the incidence rate of the common gastrointestinal side effect of NSAIDs, it is obvious to have antiinflammatory action, to advantage such as the GI irritation effect is slight, very big at global sales.2004, one piece of senior summary is reached a conclusion from clinical research and pharmacology's foundation: the application of Cox-2 inhibitor might increase the danger of cardiovascular disease
[1]Because rofecoxib (rofecoxib) causes patient's cardiovascular adverse events incidence rate increase, JIUYUE in 2004 U.S. Merck ﹠ Co., Inc. on the 30th announces that rofecoxib is carried out the whole world removes the city, former times the drug safety of dry goods appear suddenly out.With rofecoxib and celecoxib respectively with placebo, experiment finds that two medicines all can increase the sickness rate of cardiovascular event
[2]Find that in recent experiment the celecoxib group has 86 patients the thrombosis cardiovascular disease to occur, wherein 70% is myocardial ischemia and myocardial infarction, and 30% apoplexy occurred.But there is 56% patient that cardiovascular diseases's history is arranged among these patients, illustrates that celecoxib has the possibility that makes it to worsen
[3]Studies show that the Cox-2 selective depressant can strengthen hematoblastic activity, thereby start the generation of thrombotic episodes
[4]Experiment and clinical research show, and Cox-2 is and is expressed in kidney, blood vessel endothelium etc. some have the tissue of important physiological function, and the prostanoid that relies on the Cox-2 generation not only participates in the adjusting of pain or inflammation signal, but also keeps the integrity of blood vessel.Especially prostacyclin (the PGI that has vasodilation and anti-platelet activity
2) generate PGI when Cox-2 is suppressed by Cox-2
2Level will be reduced to below 50% of normal level
[5]There are some researches show that celecoxib can reduce PGI in healthy human urine and the blood
2Level, increased thrombotic danger
[6]Show in the experiment of delivering in February, 2005 (APC) about usefulness celecoxib (celecoxib) prevention adenoma, contrast with placebo, celecoxib (pfizer inc product) causes that the dangerous of lethal or non-lethality cardiovascular event obviously increases, there are 16 patients that take these two dosage cardiovascular event to occur, the danger of its lethal or non-lethality is 2.5 times of placebo group, simultaneously, in thrombosis incident patient occurring, mostly suffers from cardiovascular disease, the prompting celecoxib may increase the weight of original cardiovascular disease, American National cancer mechanism has ended the dosage of the celecoxib that adopted in the APC experiment: 400,800 mg/day.
Many researchs are thought, mainly are because this type of medicine has suppressed PGI
2Generation, cause TXA
2Raise, thus the PGI of adjusting platelet aggregation
2/ TXA
2The balance disorder, in addition, amount of literature data shows in recent years, ischemic cardio cerebrovascular diseases blood samples of patients viscosity abnormality increases, and exceeds 5 to 6 times than the normal person.Therefore most scholars think that it also is one of main hazard factor that causes ischemic cardio cerebrovascular diseases that blood viscosity raises, thus thrombotic dangerous the increasing of cardiovascular and cerebrovascular vessel.
Studies show that 2, sodium selenite (Se) can make PGI
2Raise the synthetic PGI of aorta when people such as Valentrovic find to lack selenium
2Obviously reduce
[7]Report such as Schiavon is given 6 week of healthy volunteer's selenium supplement back PGI
2Synthetic increasing discovers that further adding Se can make porcine aorta endothelium cultured cell PGI
2Generating increases
[8]Rat aorta endotheliocyte glutathion peroxidase (GSH-Px) activity increases PGI simultaneously after giving the normal rat selenium supplement
2Generation increases
[9], studies show that selenium is with the lipoxygenase metabolic pathway of GSH-Px form participation AA, catalyzing hydrogen peroxide arachidonic acid (HPETE) is converted into hydroxylation arachidonic acid (HETE)
[10], and HPETE is arterial endothelium PGI
2Synthetase inhibitors.In addition, Se can also make TXA by making the active rising of GSH-Px make the oxide level reduction suppress the epoxidase activity
2Generate and reduce
[11]Based on above two effects, Se can make PGI
2/ TXA
2Ratio increases, and produces the inhibitory action of platelet aggregation.
3, in vitro study finds that double density does not reach does not have influence to platelet adhesion reaction, but can anticoagulant.Double density reaches the not interior cAMP concentration of platelet increasing by suppressing cyclic nucleotide phosphodiesterase activity, and strengthens PGI
2Effect.Recently relevant bibliographical information double density reaches and does not have good antiinflammatory action
[12]
Below be the relevant references that the applicant retrieves:
1.Clark?DWJ，Layton?D，Shakir?SAW.Do?some?inhibitors?of?COX-2increase?the?risk?ofthromboembo-lic?events？Linking?all?of?the?data?analysed?in?this?report.Drug?Saf?2004；27(7)：427-56；
2.Drazen?JM.COX?II?inhibitors：a?lesson?in?unexpected?problems?for?her?help?andrheumatology?exper-tise?in?assessing?the[online].Available?from?URL：http://www.nejm.org[Accessed?2005Feb?15]；
3.Mira?H.W，Peter?H.，Rachael?M.，et?al.Incidence?of?Thrombotic?Cardiovascular?Eventsin?Patients?Tak-ing?Celecoxib?Compared?with?Those?Taking?Rofecoxib?Interim?Results?fromthe?New?Zealand?Inten-sive?Medicines?Monitoring?Programme.Drug?study2005；28(5)：435-442；
4.Pidgeon?GP，Tamosiuniene?R，Chen?G，et?al：Intravascular?thrombosis?afterhypoxia-induced?pulm-onary?hypertension：Regulation?by?cyclooxygenase-2.Circulation2004；110：2701-2707；
5.Florian?
，Thomas?M.，Schiele?et?al.Selective?COX-2Inhibitors?and?Risk?ofMyocardial?Infarction.J?Vasc?Res?2005；42：312-324；
6. poplar algae great mansion is edited pharmacology and pharmacotherapeutics People's Health Publisher the 59th chapter antiplatelet drug: 997-998;
7.Doni?MG，The?serum?selenium?loncentration?of?patients?with?alute?myocardialinfarction，Haemostasis，1983，13：248；
8.Schiouon?R，Arterial?walls?generate?prostaglandin?endoperoxide?a?substance，ThrombRes，1986，36：340；
9.Valentrovic?MA，Platelet?glutathione?peroxidase?activity?as?an?index?of?selenium?statusin?rats，J?Toxiol?Environ?Health，1985，15：193；
10.Bryant?RW，Prostalyclin?and?thrombox?ane?synthase?new?aspects?of?hemethiocatecatelysis，J?Biol?Chem?1982，257：14937；
11.Cuidi?G，The?relation?of?selenium?glutathione?peroxydase?and?the?release?ofthromboxane?X2，Jlabclin?Med，1984，104：574；
12.Weyrich?AS，Denis?MM，Kuhlmann-Eyre?JR，Spencer?EDDipyridamole?selectivelyinhibits?inflame-matory?gene?expression?in?platelet-monocyte?aggregates?circulation2005FEB?8111(5)：633-642。
Summary of the invention
The objective of the invention is to, provide a class to have the anti-inflammatory effect and improve hemorheological property and suppress thrombotic medicine, this medicine can be in collaborative antiinflammatory action, also has good collaboratively improve hemorheological property and suppress thrombotic effect.
In order to realize above-mentioned task, the present invention takes following technical solution:
Have the anti-inflammatory effect and improve hemorheological property and suppress thrombotic medicine, it is characterized in that, this medicine reaches not in three kinds of medicines two or more by celecoxib, sodium selenite, double density and constitutes, wherein:
Celecoxib, sodium selenite and double density reach does not form the celecoxib compound recipe, and its weight proportion compatibility is: celecoxib: sodium selenite: double density reaches not=and 1: 0.0013: 2.3; Or
Celecoxib and sodium selenite are formed the celecoxib compound recipe, and its weight proportion compatibility is: celecoxib: sodium selenite=1: 0.0013; Or
Celecoxib and double density reach does not form the celecoxib compound recipe, and its weight proportion compatibility is: celecoxib: double density reaches not=and 1: 2.3; Or
Sodium selenite and double density reach not to be formed double density and reaches not compound recipe, and its weight proportion compatibility is: sodium selenite: double density reaches not=and 0.000565: 1.
Through the applicant said medicine compound recipe be studies have shown that celecoxib, sodium selenite, double density reach and all do not have anti-adjuvant-induced arthritis effect; (celecoxib+sodium selenite+double density reaches not above-mentioned 3 compound recipe celecoxibs, celecoxib+sodium selenite; Celecoxib+double density reaches not) and double density reaches not, and compound recipe (double density reaches not+sodium selenite) also has the effect of anti-inflammatory arthritis.The anti-early stage inflammatory effect of compound recipe celecoxib (celecoxib+sodium selenite+double density reaches not, and celecoxib+double density reaches not) is single obvious with celecoxib.
The applicant will finish the related work of declaring new drug as early as possible.
The specific embodiment
One, compound recipe celecoxib prescription research
1. research purpose
The most important untoward reaction of celecoxib is: promote thrombosis, cause cardiovascular and cerebrovascularinfarction.This research is purpose to alleviate the short thrombotic untoward reaction of celecoxib, with healthy rat thrombosis is observation index, research celecoxib and sodium selenite and double density reach reciprocal action not, have good inhibiting celecoxib, sodium selenite, double density to reach not compound recipe prescription and prescription ratio thereof in the hope of searching to thrombosis.
2. experiment material
2.1 medicine and reagent
Celecoxib by pfizer inc production (lot number: BK050609), sodium selenite purchase in Zhengzhou Fu Run moral industry and trade company limited (lot number: 041121), double density reach not by Linfen, Shanxi pharmaceutical factory produce (lot number: 0512031),
2.2 instrument
1. extracorporeal thrombosis forming device (9103-B) is produced by Sensitive Technology Co. of the Chinese Academy of Sciences.
2. torsion balance, microscope, slide gauge, filter paper etc.
2.3 animal
Healthy adult ♂ SD rat, body weight 250 ± 25g, animal and feedstuff provide (the secondary animal quality certification number: the moving word of Shan doctor: 08-005) by Xi'an Jiaotong University Medical College experimental animal center.
3. experimental technique
3.1 weight recipe design
Utilize external thrombus to form experiment, the weight recipe design by DAS software does not reach celecoxib, sodium selenite, double density carries out the compatibility design according to three factors, six levels, shown in table 1-1;
Table 1-1. weight recipe design table
| Component medicine sequence number | 1 | 2 | 3 | 4 |
| Select row for use | A | C | B | F |
| Row number | A | B | C | D |
| Compatibility group 1 | Dosage 1 | Dosage 2 | Dosage 3 | Dosage 4 |
| Compatibility group 2 | Dosage 2 | Dosage 4 | Dosage 6 | Dosage 1 |
| Compatibility group 3 | Dosage 3 | Dosage 6 | Dosage 2 | Dosage 5 |
| Compatibility group 4 | Dosage 4 | Dosage 1 | Dosage 5 | Dosage 2 |
| Compatibility group 5 | Dosage 5 | Dosage 3 | Dosage 1 | Dosage 6 |
| Compatibility group 6 | Dosage 6 | Dosage 5 | Dosage 4 | Dosage 3 |
3.1 divide into groups and external thrombus forms and thrombosis assessment of indices method
(body weight: 200 ± 25g) are divided into 7 groups at random by the body weight equilibrium, and 6 every group, wherein 6 groups is the medication group of different proportionings, and 1 group is the blank group in addition to get 42 healthy adult ♂ SD rats.Each is organized the medication capacity and is 1ml/100g.Dosage (seeing Table 1-2) gastric infusion according to the arrangement of weight recipe design table, irritate stomach every day once, continuous 10 days, after administration in the 10th day one hour, get blood from rat carotid artery, with 1.8ml not anticoagulation inject the silication plastic hoop of extracorporeal thrombosis forming device fast, to be located at once on the thrombosis instrument rotating disk, with the rotating speed rotation of (17 ± 1) rpm (linear velocity 437mm/s), rotating ring and ground angle are 74 °, take off behind the rotation 15min, thrombosis is poured on the filter paper in will encircling, and inhales and removes surplus blood, surveys thrombosis length L (mm) with slide gauge, claim its weight in wet base W (mg) with torsion balance, and according to formula thrombosis index
Calculate Q-value.
Table 1-2. celecoxib, sodium selenite and double density reach recipe design table not
| The compatibility group | Celecoxib (D1) (mg/kg) | Sodium selenite (D2) (μ g/kg) | Double density reaches not (D3) (mg/kg) |
| 1 | 9(1) | 7(2) | 11(3) |
| 2 | 12(2) | 11(4) | 28(6) |
| 3 | 16(3) | 20(6) | 8(2) |
| 4 | 21(4) | 5(1) | 20(5) |
| 5 | 27(5) | 9(3) | 6(1) |
| 6 | 36(6) | 15(5) | 15(4) |
3.3 data analysis
Gained weight formulation data is all used DAS software, and (DAS, ver1.0) the drug interaction module is analyzed.
4. experimental result
Because what compare during DAS software drug interaction analysis is ceiling effect, and the purpose of this experiment is to thrombotic depression effect when observing celecoxib, sodium selenite and double density and reaching not three medicine compatibilities, so now thrombosis index Q is carried out inverse conversion back (1/Q) reuse DAS software analysis, draw the optimization dosage of three medicine compatibilities, the result is shown in table 1-3,1-4:
Table 1-3. three medicine compatibilities are to the influence (mean of E:1/Q, the standard deviation of SD:1/Q) of thrombosis index Q-value
Table 1-4. three medicines are the prescription injectivity optimizing of index with the inverse (1/Q) of thrombosis index Q
Effect and the interaction qualitative analysis thereof of each component in prescription
5, interpretation of result
The DAS The results of data processing shows, reach with celecoxib, sodium selenite, double density not three medicine prescriptions to thrombotic influence in, it is not principal agent (weight index b=1.853 that double density reaches, and P==0.011), and the also apparent in view (b=1.811 of the effect of sodium selenite, and P=0.037), also can be used as principal agent; Celecoxib (weight index b=-0.067, and P=0.071) has short thrombosis trend.
In addition, in 6 compatibility groups, the mean of second group of thrombosis inverse exponential (1/Q) is maximum (the mean X=0.078 of 1/Q), otherwise, the mean minimum of Q, promptly drug effect is best.Remove this, the result shows that sodium selenite and double density do not reach and to suppressing thrombosis concertedness arranged in the compatibility, thus at the purpose that reduces the short thrombosis side effect of celecoxib, select for use double density reach not with (or) sodium selenite and celecoxib compatibility.
6, conclusion
The theoretical optimization prescription is: 1. celecoxib 12mg/kg+ sodium selenite 16 μ g/kg+ double densities reach not 28mg/kg, and three's theoretical proportion compatibility is 1: 0.0013: 2.3; 2. celecoxib 12mg/kg+ sodium selenite 16 μ g/kg, the theoretical proportion compatibility of the two is 1: 0.0013; 3. celecoxib 12mg/kg+ double density reaches not 28mg/kg, and the theoretical proportion compatibility of the two is 1: 2.3; 4. sodium selenite 16 μ g/kg+ double densities reach not 28mg/kg, and the theoretical proportion compatibility of the two is 0.000565: 1.
Two, the Mus adjuvant-induced arthritis effect of the compound recipe celecoxib Chinese People's Anti-Japanese Military and Political College and the side's of tearing open research thereof
1. research purpose
Based on the front experimental result, celecoxib, sodium selenite and double density are reached not to become 4 compound recipes in 1: 0.0013: 2.3 ratio compatibility (celecoxib+sodium selenite+double density reaches not; Celecoxib+sodium selenite; Celecoxib+double density reaches not; Double density reaches not+sodium selenite), study compound medicine influence and reciprocal action thereof to rat assist agent arthritis behind the side of tearing open.
2. experiment material
2.1 medicine and reagent
Celecoxib is by pfizer inc production (lot number: BK050609), sodium selenite is purchased in Zhengzhou Fu Run moral industry and trade company limited (lot number: 041121), double density reaches is produced by Linfen, Shanxi pharmaceutical factory that (lot number: 0512031), Freund's complete adjuvant is not purchased the (lot number: R051124) in Sigma company.
2.2 experimental apparatus 20ml syringe, tripod, tee T, volumetric pipette;
2.3 laboratory animal SD rat is male, body weight 175～225g, and animal and feedstuff provide (the secondary animal quality certification number: the moving word of Shan doctor: 08-005) by Xi'an Jiaotong University Medical College experimental animal center.
3. experimental technique and result
3.1 method
Analyze the experiment needs according to the drug influence side of tearing open, the healthy adult male SD rat be divided into 11 groups at random, 10 every group, be respectively:
(1) blank group: NS 20ml/kg;
(2) model group: NS 20ml/kg;
(3) celecoxib+sodium selenite+double density reaches not (cele.+Se+Dm) heavy dose of group: 24mg/kg+32 μ g/kg+56mg/kg;
(4) celecoxib+sodium selenite+double density reaches not dosage group in (cele.+Se+Dm): 12mg/kg+16 μ g/kg+28mg/kg;
(5) celecoxib+sodium selenite+double density reaches not (cele.+Se+Dm) small dose group: 6mg/kg+8 μ g/kg+14mg/kg;
(6) celecoxib+sodium selenite (cele.+Se). middle dosage group: 12mg/kg+16 μ g/kg;
(7) celecoxib+double density reaches not dosage group: 12mg/kg+28mg/kg in (cele.+Dm);
(8) sodium selenite+double density reaches not (se+Dm). middle dosage group: 16 μ g/kg+28mg/kg;
(9) plug comes dosage group: 12mg/kg in former times (cele);
(10) dosage group in the sodium selenite (se): 16 μ g/kg;
(11) double density reaches not dosage group: 28mg/kg in (Dm).
The rat adaptability was raised after 5 days, every Mus causes inflammation in right back sufficient plantar subcutaneous injection 0.1mL Freund's complete adjuvant, administration group rat was pressed above-mentioned grouping and dosed administration on the 19th day in adjuvant injection back, once a day, successive administration d14, d1, d3, d5, d7, d14 are measured the left and right back of each Mus ankle volume according to volumetric method before administration and after the administration.
The volumetrical difference of the left and right ankle of rat is the swelling degree, calculates the average of swelling degree, calculates inhibitory rate of intumesce as follows.
3.2 data statistics
The gained data with the DAS statistical software (DAS, ver1.0) in the module between the sequential data set t-test compare.
3.3 experimental result
Observe adjuvant type rats with arthritis ankle swelling degree by above-mentioned 11 component groups, and be contrast groups with the inflammatory model group, the variation of statistics swollen joint expansibility, and then (celecoxib+sodium selenite+double density reaches not to analyze compound medicine; Celecoxib+sodium selenite; Celecoxib+double density reaches not; Double density reaches not+sodium selenite) and single compound medicine (celecoxib, sodium selenite, double density reach not) to the influence of inflammation; Be the analysis of the contrast groups side of tearing open with the celecoxib group again.
The table 2-1 compound recipe celecoxib side of tearing open afterwards different time to the therapeutical effect of rat paw edema (x ± S, n=10)
: the expression contrast groups ,-P＞0.05, * P＜0.05, * * P＜0.01 is added up with rate of change.
(continuous table 2-1)
The table 2-2 compound recipe celecoxib side of tearing open afterwards different time to the therapeutical effect of rat paw edema (x ± S, n=10)
: the expression contrast groups ,-P＞0.05, * P＜0.05, * * P＜0.01 is added up with rate of change.
Discover that pedal swelling has taken place the model group rat, and the swelling degree is obvious gradually subsequently, peaks at d5, still obviously has (P＜0.01) at the d14 pedal swelling, the success of adjuvant arthritis model copy is described.
D1, d3, d5, d7, d14 after administration, administration 9 groups of (3 dosage of compound recipe celecoxib, celecoxib+sodium selenite, celecoxib+double density reaches not, and sodium selenite+double density reaches not, celecoxib, sodium selenite, double density reaches not) the rat paw edema degree is alleviated; It is little respectively to organize swelling degree rate of change behind the d1, has only in the compound recipe dosage group more obvious, has statistical significance (P＜0.05); Along with administration time prolongs, the effect that each administration group suppresses rat paw edema (suppression ratio) is tending towards obviously (seeing Table 2-3).Studies show that celecoxib, sodium selenite and double density reach and all do not have independently anti-inflammatory arthroncus effect; With the celecoxib is that the main compound recipe of forming (celecoxib+sodium selenite+double density reaches not, and celecoxib+sodium selenite, celecoxib+double density reach not, and sodium selenite+double density reaches not) also has antiinflammatory, anti-arthroncus effect.(table 2-1, table 2-2)
The side of tearing open the analysis showed that, greater than the effect that celecoxib uses separately, illustrates that each component has been brought into play mutual promoting action in the compound medicine in the effect of early stage (d1) compound medicine (celecoxib+sodium selenite+double density reaches not, and celecoxib+double density reaches not) of inflammation.
Each administration group is along with the time sees Table 2-3 to rat paw edema degree suppression ratio size; From Fig. 2-1,2-2,2-3 all can find out compound medicine and independent 3 medicines to the suppression ratio of rat assist agent arthritis along with the prolongation of time is in rising trend.
The table 2-3 compound recipe celecoxib side of tearing open back different time is to the suppression ratio (n=10) of rat paw edema
4. brief summary
1, successfully set up adjuvant-induced arthritis, inflammatory reaction reaches the peak at d5.
2, celecoxib, sodium selenite, double density reach and all do not have anti-adjuvant-induced arthritis effect; The compound recipe (celecoxib+sodium selenite+double density reaches not, and celecoxib+double density reaches not, and double density reaches not+sodium selenite) of 3 kinds of medicine compositions also has the effect of anti-inflammatory arthritis, and its effect with the administration time prolongation obviously.
3, early stage at inflammatory arthritis, the antiinflammatory action of compound medicine (celecoxib+sodium selenite+double density reaches not, and celecoxib+double density reaches not) is single obvious with celecoxib.
Three, the compound recipe celecoxib is to rat assist agent arthritis hemorheological property and thrombotic influence
There are some researches show that celecoxib can reduce PGI in healthy human urine and the blood
2Level, increased thrombotic danger.These studies have shown that theoretic supposition: selectivity Cox-2 inhibitor can strengthen hematoblastic activity, thereby can start the generation of thrombotic episodes.Amount of literature data shows that it also is one of main hazard factor that causes ischemic cardio cerebrovascular diseases that blood viscosity raises in recent years.In addition, hemorheology index that there are some researches show the AA rat is obviously unusual, the whole blood viscosity, reduced viscosity, plasma viscosity, hematocrit, erythrocyte sedimentation rate, the ESR equation value that show as under each shear rate all significantly increase, illustrate rat blood when arthritis be in dense, sticking, coagulate, poly-state.
1. purpose
Observing adjuvant, to induce in the rat arthritis celecoxib whether to have a platelet increasing active and hemorheology is exerted an influence, and study itself and sodium selenite, double density and reach not compatibility to hemorheological influence, in the hope of reducing the thrombosis side effect of celecoxib.
2.. material
2.1 medicine and reagent
Celecoxib is by pfizer inc production (lot number: BK050609), sodium selenite is purchased in Zhengzhou Fu Run moral industry and trade company limited (lot number: 041121), double density does not reach produces (lot number: 0512031) by Linfen, Shanxi pharmaceutical factory, sodium citrate is produced (lot number: 030608) by Xi'an chemical reagent factory, heparin sodium is produced (lot number: 0602113) by the biochemical-pharmaceutical factory, Xuzhou, ammonium oxalate by Xi'an chemical reagent factory produce (lot number: 021009), TXB
2, PGI
2Radioimmunological kit is purchased the medical college hematopathy laboratory (lot number: 060428) in University Of Suzhou.
2.2 instrument
LBY-N6K type Puli gives birth to automatic washing hemorheology instrument, is produced by Pulisheng Instruments Co., Ltd., Beijing, and shear rate is respectively 10,60,1501/S; F-3 type high speed microcentrifuge; Reading microscope is by Changchun No.1 optical Instrument Plant production; Extracorporeal thrombosis forming device (9103-B), platelet adhesion instrument (9103-A) are produced LXJ-II type centrifugal precipitation mechanism, Shanghai medical analytical instrument factory by Sensitive Technology Co. of the Chinese Academy of Sciences; γ radioimmunity calculating instrument, Shanghai 262 factories.
2.3 laboratory animal
Healthy adult ♂ SD rat, body weight 250 ± 25g, animal and feedstuff provide (the secondary animal quality certification number: the moving word of Shan doctor: 08-005) by Xi'an Jiaotong University Medical College experimental animal center
3. experimental technique:
3.1 grouping
Get 110 of healthy SD ♂ rats, body weight 250 ± 25g, by body weight at random equilibrium be divided into 11 groups, it is the same to divide into groups.With concentration is 10mg.mL
-1Adjuvant cause inflammation in the right back sufficient plantar subcutaneous injection 0.1mL of every Mus.In administration according to dosage in the 19th day, model group and blank group then awarded normal saline and irritate stomach administration group rat in adjuvant injection back, once a day, irritated gastric capacity and were the 10ml/kg body weight, successive administration 14d.In the 15th day, after stopping eating 12 hours, the empty stomach rat is carried out the mensuration of hemorheology index.
3.2 hemorheology experiment
Each group rat with urethane (20%) 5ml/kg intraperitoneal injection of anesthesia, is separated carotid artery and gets blood 8ml by the common carotid artery intubate.Wherein 3.5ml blood injects the anticoagulant heparin pipe rapidly (the anticoagulant tube preparation method: heparin sodium is mixed with solution with distilled water, concentration is 31.25 units/ml blood, adds the test tube bottom with micro sample adding appliance, and it is standby to put into drying in oven), after shaking up, the constant water bath box of inserting 35 ℃ is standby; (preparation method: it is 3.8% solution that structure rafter acid sodium is mixed with concentration with normal saline 1.5ml blood injects the sodium citrate anticoagulant tube rapidly, getting 0.13ml adds in the silication test tube), shake up standbyly, 1.1ml is used to survey external thrombus and forms index, and 1.5ml then is used to detect blood plasma TXB
2, PGI
2Content.
1) mensuration of whole blood and plasma viscosity: get the 1ml heparin anti-coagulating and inject in the viscosity counting tube, change high, medium and low shear rate switch, from computer, directly read high, medium and low shear rate viscosity viscosity number.Viscosity apparatus cleans automatically after finishing whole blood viscosity, and remaining 2.5ml heparin anti-coagulating is used centrifugal 15 minutes of centrifuge, and rotating speed is 3000rpm, gets 1ml blood plasma and injects a survey plasma viscosity in the viscosity counting tube.
2) mensuration of erythrocyte sedimentation rate and packed cell volume: get the smooth capillary tube in bottom, insert in the heparin anti-coagulating of firm mixing, treat that blood enters 4/5 place in the capillary tube, seal lower port, insert in the plasticine lower end, stands vertically, and timing, use erythrocyte sedimentation rate after 1 hour, the hematocrit plate is read erythrocyte sedimentation rate value (wynn's method).To measure the capillary centrifuge tube of erythrocyte sedimentation rate, and use centrifugal 5 minutes of 3F-3 type high speed microcentrifuge, rotating speed is 12000rpm, and reuse erythrocyte sedimentation rate, hematocrit plate are read the long-pending value of erythrocyte.
3) platelet adhesion rate is measured: with 1ml sodium citrate anticoagulation adding capacity is in the long-neck spheroidal vial of 5ml, be placed on the platelet adhesion instrument rotating disk, with the rotation of the rotating speed of 5rpm after 15 minutes, reuse is quantitatively got blood vessel and is drawn before the 20 μ l rotation respectively and the ammonium oxalate diluent 0.4ml of postrotational anticoagulation injection 1%, measure the platelet plate number in the blood of rotation front and back, be calculated as follows then:
4) mensuration of red cell deformability:
5) external thrombus forms and thrombosis assessment of indices method: with 1.1ml not anticoagulation inject (extracorporeal thrombosis forming device adnexa) in the silication plastic hoop fast, to be located at once on the thrombosis instrument rotating disk, rotating speed rotation with (17 ± 1) rpm (linear velocity 437mm/s), rotating ring and ground angle are 74 °, take off behind the rotation 15min, and thrombosis is poured on the filter paper in will encircling, surplus blood is removed in suction, survey its length with vernier cursor, torsion balance claims its weight in wet base, and according to formula thrombosis index
Calculate Q-value, L is thrombosis length (mm) in the formula, and W is wet weight of thrombus (mg).
6) TXB in the blood plasma
2, PGF
1 αMeasure
The preparation specimen: with aforementioned 1.5ml blood with 2%EDTA solution in ratio anticoagulant in 1: 9, with 3000 rev/mins centrifugal 30 minutes, draw supernatant and place the cryogenic refrigerator preservation.
TXB
2, PGF
12Measure: operating procedure:
1. TXB
2Standard substance become 7 pipes of 2.5～160pg/0.1ml with the buffer doubling dilution, with TXB2 antiserum, TXB
2Tracer adds buffer 10ml dilution.The testing sample for preparing is taken out redissolution.
2. operate shown in the according to the form below:
From the 3rd pipe centrifugal 3000rpm30 minute, inhale and to abandon the sedimentary radioactivity of mensuration behind the supernatant.
3. respectively managing of will preparing send behind the labelling Isotope Lab to measure TXB in the sample
2, PGF
1 αContent.
4. result of calculation
3.3 statistical analysis
Data are analyzed with SPSS, and experimental result is represented with x ± s, relatively checks with t between group.
4. experimental result
4.1 the adjuvant arthritis rats hemorheology sexually revises
Table 3-1 data declaration, compare with the normal control group, the whole blood height of adjuvant arthritis rats cut viscosity, in cut, low viscosity, erythrocyte IR value, erythrocyte aggregation index, the thrombosis index Q of cutting, the platelet adhesion reaction rate all increases and has a height statistical significance (P＜0.01); The packed cell volume of adjuvant arthritis rats, erythrocyte sedimentation rate then do not have obvious change.Above result is consistent with bibliographical information, and the success of adjuvant-induced arthritis Blood stasis model copy is described.
Table 3-1. Blood stasis model copy situation (x ± S, n=10)
Compare with the normal control group
*: P＜0.05,
*: P＜0.01
4.2 celecoxib, sodium selenite, double density reach not and compound recipe is cut the influence of viscosity to the adjuvant arthritis rats whole blood
Table 3-2 shows, with model group relatively, except that celecoxib did not have the explicitly effect, sodium selenite, double density reached not and the compound recipe celecoxib (Cele+Se+Dm) (high, medium and low dosage) formed and the compound recipe (Cele.+Se of other 2 medicines composition; Cele.+Dm; Se.+Dm) the high, medium and low shear rate whole blood viscosity of adjuvant-induced arthritis Blood stasis model is all reduced, and the statistical significance of tool highly significant (P＜0.01), illustrate that they have good should kindly acting on to hemorheological property.
Compare with celecoxib group (17mg/kg), itself and sodium selenite, double density reach the effect of the various compound recipes of not forming all greater than the celecoxib group, particularly the low dose group of compound recipe celecoxib (Cele+Se+Dm) (8.5mg 11.5 μ g; 20mg/kg) effect that reduces high, medium and low shear rate whole blood viscosity illustrates that still greater than celecoxib group (17mg/kg) medicine has produced good interaction.
Table 3-2 celecoxib, sodium selenite and double density reaches not and compound recipe to the influence of whole blood viscosity (x ± S, n=10)
: expression contrast groups ,-P＞0.05, * P＜0.05, * * P＜0.01.
4.3 celecoxib, sodium selenite and double density reach not and compound recipe to the influence of erythrocyte rigidity index IR:
The erythrocytic fragility of erythrocyte rigidity index IR value representation is inverse relation with erythrocyte deformability.Erythrocyte rigidity index IR value sees Table 3-3.
Compare with model group, reach any compound recipe (Cele+Se+Dm, the Cele.+Se that does not form by celecoxib, sodium selenite and double density; Cele.+Dm; Se.+Dm) the erythrocyte rigidity index IR value of adjuvant arthritis rat is all obviously reduced, and have height statistical significance (P＜0.01); In addition, the independent effect of sodium selenite also can make the IR value obviously reduce, and has height statistical significance (P＜0.01); Group, celecoxib group IR value change not statistically significant and double density reaches not.
Table 3-3. celecoxib, sodium selenite and double density reach not and compound recipe to the influence of IR value (x ± S, n=10)
: expression contrast groups ,-P＞0.05, * P＜0.05, * * P＜0.01.
The side's of tearing open analysis: compare with the celecoxib group, the celecoxib+double density of dosage reaches not the effect significance raising that (Cele.+Dm) reduces the IR value on year-on-year basis, illustrates that celecoxib and double density reach the collaborative erythrocyte deformability that increases; Compare with sodium selenite, the sodium selenite+double density of dosage reaches not the effect significance raising that (Se.+Dm) reduces the IR value on year-on-year basis, illustrates that sodium selenite and double density reach the collaborative erythrocyte deformability that increases; Do not reach with double density and to compare, on year-on-year basis sodium selenite+the double density of dosage reach not (Se.+Dm), celecoxib+double density and reach not+sodium selenite (Cele+Se+Dm) reduces the effect significance raising of IR value, illustrate that celecoxib, sodium selenite and double density reach the not collaborative erythrocyte deformability that increases, wherein selenium may play a key effect.
4.4 celecoxib, sodium selenite and double density reach not compound recipe to the influence of platelet adhesion reaction rate and thrombosis index Q
Find that with the model group comparative study celecoxib (Cele), sodium selenite (Se), double density reach not (Dm) and compound recipe thereof (Cele+Se+Dm, Cele.+Se, Cele.+Dm, Se.+Dm) all can reduce platelet adhesion reaction rate and thrombosis index Q-value (P＜0.01).See Table 3-4.
Table 3-4. celecoxib, sodium selenite and double density reaches not and compound recipe to platelet adhesion reaction rate and the exponential influence of thrombosis
X ± S, n=10: expression contrast groups ,-P＞0.05, * P＜0.05, * * P＜0.01
Find that with the comparative study of celecoxib group the inhibitory action of platelet adhesion reaction rate and thrombosis index Q is weaker than sodium selenite to celecoxib and double density reaches not: the compound recipe of being made up of 3 medicines (Cele+Se+Dm) antiplatelet effect of sticking when dosage on year-on-year basis obviously is better than celecoxib when using separately; The compound recipe of being made up of 3 medicines reduces when dosage on year-on-year basis when also obviously being better than celecoxib aspect the thrombosis index Q and using separately, (Cele.+Se is when Cele.+Dm) also obviously being better than celecoxib when dosage and low dosage and using separately on year-on-year basis for the compound recipe of being made up of 2 medicines.Studies show that compound medicine has been brought into play good interaction to the inhibitory action of platelet adhesion reaction rate and thrombosis index Q the time, wherein Wei Liang sodium selenite may play an important role.See Table 3-4.
Back, the 4-5 compound recipe celecoxib side of tearing open is to blood plasma TXA
2/ PGI
2The influence of ratio
Table 3-5 discloses to such an extent that quantity of information is very big, compares PGI when inflammation with model group
2Content and relative quantity (TXB
2/ PGI
2) all increase; Celecoxib can make inflammation P of Rats GI
2Reduce and TXA
2/ PGI
2Ratio increases, the antagonism inflammatory activity; Compound recipe celecoxib (Cele+Se+Dm) (high, middle dosage) when heavy dose of makes TXB2 and PGI
2All reduce, but TXA
2/ PGI
2Ratio is but increasing.
The table 3-5. compound recipe celecoxib side of tearing open is to blood plasma TXA
2, PGI
2Level influence x ± S, n=10
: expression contrast groups ,-P＞0.05, * P＜0.05, * * P＜0.01
Studies show that celecoxib can obviously increase TXA
2/ PGI
2Ratio might increase the danger that the thrombosis cardiovascular adverse effects takes place; With the celecoxib list of dosage on year-on-year basis with comparing, compound recipe celecoxib (Cele+Se+Dm) has stronger reduction TXB
2Therefore effect increases TXA
2/ PGI
2A little less than the effect of ratio; The celecoxib of dosage+sodium selenite compound recipe (Cele+Se) is also increasing TXB2 and PGI on year-on-year basis
2The time, also have increase TXA
2/ PGI
2The characteristics that the effect of ratio is more weak; Do not observe the celecoxib of dosage on year-on-year basis and double density and reach not that compound recipe (Cele+Dm) has such characteristics.Use relatively with the celecoxib list, compound recipe celecoxib (Cele+Se+Dm or Cele+Se) increases TXA
2/ PGI
2A little less than the effect of ratio, thereby reduced the danger of celecoxib cardiovascular adverse effects.
5, brief summary
5.1, on the adjuvant arthritis rats model, observe the aggravation of the bad variation of hemorheological property and thrombosis, comprise that whole blood viscosity increases, erythrocyte deformability reduces, platelet adhesion reaction strengthens, variations such as thrombosis index Q-value increase.Adjuvant arthritis rats PGI
2Content and TXB
2/ PGI
2Ratio reduces.
5.2, sodium selenite, double density reach do not have the rat assist agent arthritis of the improvement model hemorheological property effect of (high, medium and low shear whole blood viscosity reduces, and erythrocyte deformability increases); Various prescriptions of compound recipe celecoxib (Cele+Se+Dm, Cele+Se, Cele+Dm) and double density reach not, and selenium (Se+Dm) also has the effect that improves the rat blood rheological characteristic.Illustrate that the effect of compound recipe celecoxib is better than celecoxib and uses separately.
5.3, sodium selenite, double density reach and do not have the effect that reduces rat assist agent arthritis model thrombosis index Q-value and suppress the platelet adhesion reaction rate, the various prescriptions of compound recipe celecoxib (Cele+Se+Dm, Cele+Se, Cele+Dm) also have the effect that reduces thrombosis index Q-value and suppress the platelet adhesion reaction rate, illustrate that the effect of compound recipe celecoxib is better than the celecoxib list and uses.Double density reaches not, and selenium (Se+Dm) has the effect that reduces thrombosis index Q-value and suppress the platelet adhesion reaction rate equally.
5.4, celecoxib, sodium selenite, double density reaches Mo Junke makes rat assist agent arthritis model TXB
2/ PGI
2Ratio increases, wherein celecoxib effect the most obvious (consistent with bibliographical information); Compound recipe celecoxib (Cele+Se+Dm, Cele+Se) rising TXB
2/ PGI
2The effect significance of ratio is lower than celecoxib to be used separately, illustrates that the compound recipe celecoxib causes that the dangerous significance of thrombotic disease is lower than the celecoxib list and uses.
Four, research conclusion
1, compound recipe celecoxib optimization prescription is: 1. celecoxib 12mg/kg+ sodium selenite 16 μ g/kg+ double densities reach not 28mg/kg, and three's proportion compatibility is 1: 0.0013: 2.3; 2. celecoxib 12mg/kg+ sodium selenite 16 μ g/kg, the proportion compatibility of the two is 1: 0.0013; 3. celecoxib 12mg/kg+ double density reaches not 28mg/kg, and the proportion compatibility of the two is 1: 2.3; 4. double density reaches not 28mg/kg+ sodium selenite 16 μ g/kg, and the proportion compatibility of the two is 1: 0.00057.
2, celecoxib, sodium selenite, double density reach and all do not have anti-adjuvant-induced arthritis effect; (celecoxib+sodium selenite+double density reaches not 3 compound recipe celecoxib prescriptions, celecoxib+sodium selenite; Celecoxib+double density reaches not) and double density reaches not, and selenium (double density reaches not+sodium selenite) also has the effect of anti-inflammatory arthritis.The anti-early stage inflammatory effect of compound medicine (celecoxib+sodium selenite+double density reaches not, and celecoxib+double density reaches not) is single obvious with celecoxib.
3, sodium selenite, double density reach not that prescription has the rat assist agent arthritis of improvement model hemorheological property and suppresses thrombotic good action, and celecoxib does not have above-mentioned effect; 3 prescriptions of compound recipe celecoxib (Cele+Se+Dm, Cele+Se, Cele+Dm) and double density reach not, and selenium (Dm+Se) all has the rat blood of improvement rheological characteristic and suppresses thrombotic good action.
4, conclusion: (celecoxib+sodium selenite+double density reaches not the compound recipe celecoxib, celecoxib+sodium selenite; Celecoxib+double density reaches not) and double density reach not selenium (double density reaches not+sodium selenite) reasonable recipe, overcome the short thrombotic shortcoming of celecoxib, in the collaborative antiinflammatory action of performance, also have and good collaboratively improve hemorheological property and suppress thrombotic effect.
Claims (1)
1. one kind has the anti-inflammatory effect and collaborative improve hemorheological property and suppress thrombotic medicine, it is characterized in that, this medicine reaches not two or three in three kinds of medicines by celecoxib, sodium selenite, double density and constitutes, wherein:
Celecoxib, sodium selenite and double density reach does not form the celecoxib compound recipe, and its weight proportion compatibility is: celecoxib: sodium selenite: double density reaches not=and 1: 0.0013: 2.3; Or
Celecoxib and sodium selenite are formed the celecoxib compound recipe, and its weight proportion compatibility is: celecoxib: sodium selenite=1: 0.0013.
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| CN1549704A (en) * | 2001-08-06 | 2004-11-24 | Stabilized oral suspension formulation | |
| CN1561210A (en) * | 2001-10-25 | 2005-01-05 | 迪内希尚蒂拉尔·帕特尔 | Preparation of selective epoxide hydrolase II inhibitor |
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| CN1480144A (en) * | 2002-09-05 | 2004-03-10 | 成都力思特制药有限公司 | Compound preparation for treating and preventing thrombosis and apoplexia as well as its preparing method |
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