| BackgroundTranscatheter arterial embolization(TAE)is an effective treatment for hemostasis.However,there is a risk of rebleeding.Rebleeding can be caused by a variety of factors,one of which is the embolic agent feature.The necrosis of tumor tissue in the treatment of tumors with transcatheter arterial chemoembolization(TACE)is caused not only by the high concentration of chemotherapy drugs in the local area but also by the embolization effect of TACE,which blocks or reduces the blood flow of the feeding artery.Embolic agents currently in use,such as gelatin sponge and polyvinyl alcohol(PVA)particles,have flaws.Thrombin is currently mostly used as an oral medication to treat gastrointestinal bleeding,with minimal intravascular use.The intravascular usefulness of thrombin can be increased by combining it with embolic agents.Thrombin promotes thrombus formation and,when combined with an embolic agent,can improve embolization and prevent vascular recanalization.Purposes1.To create a microsphere that can load and release thrombin efficiently.2.To test the embolic and hemostatic effects of thrombin microspheres in a rabbit hemorrhage model,observe short-term vascular recanalization,and estimate the value of thrombin microspheres in hemorrhage treatment.3.To test the effect of thrombin microspheres in treating tumors in rabbits,observe recanalization of tumor feeding arteries and tumor growth,and calculate the value of thrombin microspheres in tumor therapy.Materials and methods1.The following methods were used to prepare thrombin microspheres.(a)In synthesis,thrombin microspheres were prepared using the water-in-oil emulsification method.Sodium alginate and thrombin were used in the water phase,liquid paraffin with Span-80 was used in the oil phase,and calcium chloride was used as the cross-linking agent.(b)The morphological properties of the thrombin microspheres were evaluated using optical and scanning electron microscopes as well as a laser particle size analyzer to determine the microsphere size distribution.(c)Calculating the loading efficiency and encapsulation percentage of thrombin microspheres,testing the pharmacokinetic characteristics of thrombin microspheres,testing the in vitro degradation characteristics of thrombin microspheres,and testing the effect of contrast agents on thrombin microspheres were all part of the pharmaceutical characteristic evaluation.(d)The cytocompatibility and hemolysis tests were performed to assess the safety of thrombin microspheres.2.The following procedures were used in the study of thrombin microspheres for bleeding treatment.(a)In the modeling approach,the rabbit renal hemorrhage and stomach artery hemorrhage models were constructed using the puncture method in the investigation of thrombin microspheres for hemorrhage treatment.(b)In the group of experimenters,45 New Zealand white rabbits were randomly assigned to one of three groups based on the embolic agent: thrombin microspheres(300–500 μm),blank microspheres(calcium alginate microspheres without thrombin,300–500 μm),and gelatin sponge(350–560 μm).Five New Zealand rabbits were randomly used for TAE treatment of the rabbit renal hemorrhage model and observed for 7 days.Five rabbits were used for TAE treatment of the rabbit renal hemorrhage model and observed for 14 days.Five rabbits were used for the rabbit gastric artery hemorrhage model and observed for 7 days among the 15 New Zealand rabbits in each group.(c)Angiography was used to compare the success rate of hemostasis and vascular recanalization in each group after TAE and 7/14 days after the surgery.Blood was drawn before TAE and before the last angiography to examine the blood routine,blood biochemistry,and coagulation function.The samples were sacrificed and subjected to pathological evaluation after the last angiography.3.The following procedures were used in the study of thrombin microspheres for tumor treatment.(a)A rabbit ear tumor model was created by injecting a substance into the back of the ear.(b)In the experimental group,15 New Zealand white rabbits were divided into three groups based on the embolic agent used: thrombin microspheres(300–500 μm),PVA(300–500 μm),and lipiodol.Each group had five rabbits undergoing TAE with an appropriate embolic agent.(c)For the observation,before and after TAE,angiography and Dyna CT were performed.Angiography was used to identify tumor staining and feeding arteries,and the greatest diameter of the axis pictures in Dyna CT was used to estimate the tumor size.Blood was collected before TAE and before the last angiography to assess routine blood and blood biochemistry.The samples were sacrificed and subjected to pathological analysis after the last angiography.4.For statistics,continuous data were compared using one-way ANOVA,and categorical data were compared using the chi-square test or Fisher’s exact test.Statistical significance was defined as P<0.05.Results1.The thrombin microsphere synthesis outcomes included the following.(a)For synthesis,a water-in-oil emulsification approach was used to successfully synthesize the thrombin microspheres.(b)For the morphological characteristics,the shape of the thrombin microsphere was spherical,the surface was dense and nonporous,and the swelling degree was12.3 m L/g.The diameter parameters of the thrombin microsphere were as follows:Dmin,4.31 μm;Dmax,698.61 μm;D[4,3],361.2 μm;D[v,0.1],170.6 μm;D[v,0.5],351.6 μm;and D[v,0.9],578.3 μm.(c)For the pharmaceutical properties,the thrombin microsphere loading efficiency was 3420 IU/g,and the encapsulation percentage was 33.40%.The thrombin microspheres released thrombin faster in the first 8 h of the drug release test,followed by a slower rate over time,similar to the sustained-release mode,with a cumulative release rate of 65.54% in 72 h.The thrombin microspheres degraded in the degradation test,the rate of deterioration was essentially uniform over time,and the cumulative degradation rate followed a linear trend with time.Within 5 and 20 min,the contrast agent had no influence on the unreleased enzyme activity in the thrombin microspheres.When the contrast agent was diluted to a reasonable concentration,the thrombin microspheres may be suspended,meeting the condition of equal dispersion of the embolic material in the medium during the embolization procedure.(d)For the safety assessment,the thrombin microsphere had good cytocompatibility,grade 0 toxicity,no indications of hemolysis,and an overall hemolysis rate of <5%.2.Thrombin microsphere application in bleeding treatment yielded the following results.(a)The success rate of hemostasis in each group was 100%(15/15)in the study of thrombin microspheres for hemorrhage treatment.(b)The vascular recanalization rates in the thrombin microsphere,blank microsphere,and gelatin sponge groups were 0%(0/15),60%(9/15),and73.3%(11/15),respectively.The recanalization rate was statistically different between the groups(P=0.000).The vascular recanalization rates in the thrombin microsphere,blank microsphere,and gelatin sponge groups were 0%(0/5),40%(2/5),and 60%(3/5),respectively,on the seventh day after embolization of the renal hemorrhage model.The rate of vascular recanalization did not differ significantly between the groups(P=0.251).The vascular recanalization rates in the thrombin microsphere,blank microsphere,and gelatin sponge groups were 0%(0/5),60%(3/5),and 60%(3/5),respectively,on the 14 th day following embolization of the renal hemorrhage model.The rate of vascular recanalization did not differ significantly between groups(P=0.126).The vascular recanalization rates in the thrombin microsphere,blank microsphere,and gelatin sponge groups were 0%(0/5),80%(4/5),and 100%(5/5),respectively,7 days after embolization of the gastric artery hemorrhage model.The recanalization rate was statistically different between the groups(P=0.006).(c)Blood tests showed no statistically significant differences between the groups(P>0.05).(d)The intra-arterial mixed thrombus was more visible in the thrombin microsphere group than in the other groups according to pathological testing.3.The following were the outcomes of using thrombin microspheres in tumor treatment.(a)Regarding angiography and Dyna CT,before TAE,angiography of each group revealed tumor staining.Ten days after TAE,the thrombin microsphere group showed no recanalization of arteries and no tumor staining,the PVA group showed partial recanalization of arteries and no obvious tumor staining,and the lipiodol group showed obvious tumor staining,the feeding arteries were thickened and increased.The change ratios of tumor size were-54.34±3.67%,-28.49±21.32%,and 50.27±7.69% in the thrombin microsphere,PVA,and lipiodol groups,respectively.A statistically significant difference in the change ratio of tumor size was noted between the groups(P=0.000).(b)There was no significant difference in the blood tests before TAE and the last angiography in each group(P>0.05).(c)On pathological examination,the intra-arterial mixed thrombus was more obvious in the thrombin microsphere group than in the other groups.Conclusion1.For thrombin microsphere preparation,the emulsification method effectively loaded and continuously released thrombin.Thrombin microspheres had good thrombin-release properties and could be used with contrast agents.This seems promising as an endovascular embolization material.2.Thrombin microspheres can effectively control bleeding and prevent recanalization of arteries.It is an endovascular embolic agent that is both safe and effective for TAE hemostasis.3.There was no recanalization of the embolized arteries or the creation of collateral arteries after thrombin microsphere embolization in the tumors,and the tumor size was greatly reduced.Thrombin microspheres were found to be safe and effective embolic agents for tumor intervention. |
