The Study On Physicochemical Properties Of Drug-loaded Thermosensitive Hydrogel System For Spinal Cord Injury Repair

?Objective?The aim of this study is to prepare PLGA-PEG-PLGA thermosensitive hydrogels by opening loop polymerization and discuss their physicochemical properties.The use of nafamostat combined with thermosensitive hydrogels PLGA-PEG-PLGA,produced a durg-loaded temperature sensitive gel complex.Its physicochemical properties were studied by gel temperature,rheology and drug release in vitro.In order to provide a theoretical basis for the application of drug carrying thermosensitive hydrogels to repair spinal cord injury in animals in the future,the safety of the animal in vivo was evaluated by cytotoxicity test.?Method?1.Physicochemical properties study of thermosensitive gel solutions PLGA-PEG-PLGA thermosensitive hydrogels were prepared by open loop polymerization.After freezing,the gel was added to double steam water and 20% drug-free thermosensitive gel solution was obtained at 4? for 24 hours.It is placed in a water bath at 25?,and raised the temperature by 1? each time,then inverted 180 ° to observe the flow of the liquid surface at each temperature.The advanced rotational rheological system(ARES)was used to study rheological characteristics.The morphology was observed under a transmission electron microscope.2.physicochemical properties of drug-loaded thermosensitive gel solutions Nafamostat mesilate was added to 20% thermosensitive gel solution,and was arranged into a 3mg/ml drug containing thermosensitive gel,and 20% drug-loaded thermosensitive solution was obtained at 4? for 24 hours.it is placed in a water bath at 25?,and raised the temperature by 1? each time,and then inverted 180 °to observe the flow of the liquid surface at each temperature.The advanced rotational rheological system(ARES)was used to study rheological characteristics.The distribution of drugs in thermosensitive gels was observed under a transmission electron microscope.3.Drug sustained release test 20% drug-loaded thermosensitive gel solution was taken into round-bottom centrifuge tube,placed in a 37? thermostatic shaker to obtain a gel that is a drug-loaded thermosensitive glue blocks.Then the shake was started.The solution was taken 3 ml at 15 min,30 min,45 min,1 h,3 h,6 h,12 h,24 h,72 h,120 h,168 h and 240 h respectively.The removed solution was diluted ten times and its UV absorption spectrum was tested.The absorbance at the highest peak is taken respectively into a standard curve to obtain the nafamostat concentration,and the cumulative release rate can be calculated.4.The safety of the drug-loaded thermosensitive gel PC12 cell culture,and drug addition methods.CCK8 was used to detect cell viability to study safety,and Graphpad prism 7.0 was used for histogram.?Result? 1.The drug-free thermosensitive gel solution is a clear solution state.The gel temperature range of the 20% drug-free thermosensitive gel solution is 32-42?.Through dynamic rheology analysis,the intersecting point of the storage modulus G' and loss modulus G” of drug-free thermosensitive gel is about 31?,and the maximum viscosity of the drug-free thermosensitive gel is about 36?.Under the transmission electron microscope,you can see a ball-like micelle or micelle group,ranging from 10-30 mm in diameter.2.The drug-loaded thermosensitive gel is a more turbid solution state.The gel temperature range of the 20% drug-loaded thermosensitive gel solution is 35-56?.The intersecting point of storage modulus G' and the loss modulus G” of the drug-loaded thermosensitive gel is about 34?,and the maximum viscosity of the drug-loaded thermosensitive gel is about 40?.The drug-loaded thermosensitive gel can see drug particles that uniformly disperse in gel micelles or micelles under the transmission microscope.3.In drug-loaded thermosensitive hydrogels,the cumulative release rate of nafamostat was about 58% at first days,83% at fifth days,and 92% at tenth days.4.In the drug-loaded thermosensitive gel group,the survival rate of PC12 cells was not significantly different from that of the simple drug group,drug-free thermosensitive gel group and control group.?Conclusion? 1.The study successfully prepared NM-PLGA-PEG-PLGA loaded thermosensitive hydrogels,and proved that both of them had temperature sensitive properties.temperature measurement and dynamic rheology analysis showed that the unloaded and loaded drug thermosensitive hydrogels can form gel state under normal body temperature.By electron microscopy,the nafamostat in the thermosensitive hydrogels can be evenly distributed.2.Drug release tests showed that the release of nafamostat is slow and sustained in ten days in the drug-loaded thermosensitive hydrogels.3.The cytotoxicity analysis showed that the drug-loaded thermosensitive hydrogel is safe to cells in vivo and could be further used experiment in vivo.