| Objective:Polymethyl methacrylate(PMMA)bone cement is a room temperature self-setting material consisting of polymethyl methacrylate or modified polymethyl methacrylate copolymer and methyl methacrylate monomer(MMA),containing or without X-ray blocking additives.In arthroplasty and vertebroplasty,PMMA bone cement is an important material widely used.PMMA bone cement can firmly fix the prosthesis to the bone tissue during hip and knee joint replacement such as femoral neck fracture and severe hip and knee degeneration,play a filling and anchoring role between the prosthesis and bone.In vertebroplasty,PMMA cement injection into the vertebral body can strengthen the vertebral body,achieve the purpose of supporting and stabilizing the spine,at the same time significantly reduce pain and improve function.It has the advantages of short operation time,minimally invasive,early ambulation and so on.But after cement implantation,it may cause a series of complications,such as blood pressure drop,dyspnea,deep vein thrombosis,pulmonary embolism,and cardiac arrest and sudden death,which are collectively called bone cement implantation syndrome(BCIS).In recent years,BCIS studies have found that a large number of thrombus were detected in the pulmonary artery when bone cement was implanted.The reason may be that the aggregation of PMMA bone cement releases a large amount of heat,and residual monomers may be released during the reaction,activating the body’s coagulation system,forming thrombus and blocking the pulmonary artery.Our preliminary results show that in hip replacement,because of the operation,the coagulation system is activated and the operation area is in a hypercoagulable state.At the same time,when the bone cement is injected into the medullary cavity and the prosthesis is inserted,the intramedullary pressure increases rapidly,and the Micro-thrombus is squeezed into the venule circulation system to form embolism.In vertebroplasty,the compression fracture vertebral blood sinus is destroyed.After injection of bone cement,the pressure in the vertebral cavity increases,resulting in more thromboemboli entering the blood circulation through the blood sinus to form thrombus.Therefore,the improvement of traditional PMMA bone cement and the reduction of thrombosis rate at the site of cement implantation deserve our further study.Enoxaparin sodium(ES)is a kind of low molecular weight heparin sodium obtained by chemical beta-elimination pyrolysis of ordinary heparin sodium.It is the most widely used low molecular weight heparin sodium in the world and has a market share of more than 60%in the United States.The antithrombotic and anticoagulant activities of enoxaparin sodium are separated and the main feature is that the anti-FXa activity is higher than anti-FIIa,and local application does not cause thrombocytopenia.This allows enoxaparin sodium to be antithrombotic without exacerbating the coagulation system and exacerbating bleeding.In this study,a new type of enoxaparin sodium bone cement was prepared by mixing a certain amount of enoxaparin sodium in traditional PMMA bone cement.The aim of this study was to observe whether enoxaparin sodium can be released stably and effectively in PMMA bone cement,to meet the local anti-thrombotic purpose of bone cement implantation in orthopaedics,and to lay a pharmacokinetic foundation for the clinical application of enoxaparin sodium PMMA bone cement.Methods:Preparation of enoxaparin sodium bone cement:The operating room environment was adjusted to 23±1°C,and all experimental materials were placed in this environment for 4 hours.The sodium enoxaparin was mixed with PMMA bone cement in different proportions,and seven groups of A-G were set up.The amount of sodium enoxaparin added to each 40 g Palacos~?R PMMA bone cement was 4000 AxaIU,8000 AxaIU,12000 AxaIU,16000AxaIU,20000 AxaIU and 24000 AxaIU,respectively.Group G was the blank control group,without adding sodium enoxaparin.The enoxaparin sodium lyophilized powder was first ground into a powder with a mortar and then thoroughly mixed with 40 g of PMMA bone cement powder(copolymer).The bone cement liquid(methyl methacrylate monomer)was poured into the powder mixture and stirred rapidly.After mixing period,waiting period and dough period,the bone cement is placed into a cylindrical photosensitive resin mold with a length of(12±0.1)mm and an inner diameter of(6±0.1)mm.After the cement enters the curing period,carefully remove the mold and use it.The vernier caliper reviews the test die size,excludes the test mode that does not meet the production standard,and then uses the C-arm X-ray fluoroscopy to eliminate the test mode with a significantly low density.The prototype of enoxaparin sodium PMMA bone cement was completed.Scanning electron microscopy(SEM)analysis was performed on the test models of 0,8000 AxaIU,16000 AxaIU and 24000 AxaIU with the addition of enoxaheparin sodium.In the enoxaparin sodium release experiment,4 specimens were taken from each group of A to G.The standard test specimens of the cylinder were labeled one by one,placed in a 1.5 ml centrifuge tube,and 1 ml of Tris7.4buffer solution was added,and placed in a 37°C incubator.The extract is collected once every 24 hours.Each time the extract was collected,the test piece was rinsed with Tris 7.4 buffer for 3 times,and then the test piece was placed in a new centrifuge tube and 1 ml of Tris 7.4 buffer was added to continue the leaching,and sampling was continued for 10 days.The storage condition of the extract is-20°C cryopreservation.According to the parallel line method of quantitative reaction in the statistical method of biological verification(Appendix 1431 of Chinese Pharmacopoeia 2015 edition),the anti-Xa factor titers of extracts of each group were designed and calculated by4.4 method.After 10 days of extraction with 0,8000 AxaIU,16000 AxaIU and 24000 AxaIU,one of the test models was analyzed by scanning electron microscopy.In the stability experiment of enoxaparin sodium,the average anti-Xa factor titer of group B(8000 AxaIU/40g)bone cement extract was determined.On this average,the same concentration of enoxaparin sodium solution was prepared with enoxaparin sodium standard material as the test solution of group M,the same concentration of enoxaparin sodium solution was prepared with enoxaparin sodium standard material after heat treatment at 75°C for 10minutes as the test solution of group N,and Tris 7.4 buffer solution as the blank control of group R.The anticoagulant activity of the four groups B,M,N and R was evaluated by the rabbit plasma recalcification method(heparin bioassay in Appendix 1208 of Chinese Pharmacopoeia 2015 edition).Sixteen clean and dry tubes with uniform diameter(0.8cm*3.8cm)were taken and0.8ml anticoagulant rabbit plasma was added to each tube,and put them in a constant temperature water bath at 37°C for 5 minutes.Each tube was sequentially added with 0.1 ml of test solution B,M,N,and R and 0.1 ml of1%calcium chloride solution,and each group was measured four times in parallel.After adding 1%calcium chloride solution,mix immediately to avoid air bubbles,and start calculating time,observe and record the condensation time of each tube.Results:1.Scanning electron microscopy(SEM)of PMMA bone cement showed a large number of copolymer beads and fine zirconia particles,which are the powder components of bone cement.There are pores between beads,which can be used as space for drug delivery and release.PMMA bone cement with enoxaparin sodium was found to have syrup-like substances between copolymer beads.With the increase of ES loading,syrup-like substances also increased.Finally,they were coated like sugar on the surface of copolymer beads.Comparing the SEM photos before and after the release of the samples,we can find that the syrup-like substances are greatly reduced.2.Cement release systems with different ES loads were released rapidly at the initial stage,with a sudden release effect.On the first day,the peak of ES release was observed,and then decreased rapidly.On the fourth day,the release rate entered a slow period,and tended to be stable.3.The coagulation time of plasma in the three groups containing ES was significantly longer than that in the blank control group,with statistical difference.There was no statistically significant difference in plasma clotting time between the ES-PMMA group and the ES 75°C treatment group and the ES standard group.Conclusions:Enoxaparin sodium can be physically embedded in the pore structure of PMMA cement.ES-PMMA bone cement can effectively release enoxaparin sodium,which has a high release rate in the first three days,and then rapidly declines into a slow release phase.Enoxaparin sodium does not participate in the PMMA bone cement polymerization,and can withstand the peak temperature of the polymerization reaction,keeping the anticoagulant activity substantially unchanged. |
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