Research On The Preparation And Characterization Of Hemostatic Cryogel

The high mortality rate caused by uncontrollable bleeding in the field of military and civilian trauma remains a worldwide problem.At present,the commonly used hemostatic materials such as gelatin sponge,oxidized cellulose and chitosan have some problems,such as complex preparation process,high price and slow hemostatic speed,so the research on hemostatic materials which can effectively control bleeding is the current research focus.Silk fibroin（SF）is a kind of natural polymer fibrin extracted from silkworm cocoon.It has excellent mechanical properties,air permeability,good biocompatibility and biodegradability.In recent years,it has been widely concerned in biomedical applications.Sodium alginate（SA）,which is composed ofβ-D-manuronic acid andα-L-guluronic acid,is also widely used in tissue engineering.However,SA lacks cell binding sites.By adding appropriate amount of cell adhesion polymers to SA-based hydrogels,the specific interaction between cells and hydrogels can be regulated to make it more suitable for tissue engineering.In recent years,bionic studies have found that dopamine（DA）can be easily oxidized to polydopamine（PDA）with a structure similar to that of mussel podocyte（DOPA）in aqueous or alkaline conditions.PDA has been proved to have strong adhesion on various substrates.The phenolic hydroxyl and amino groups in its structure not only help to activate the blood coagulation system,but also prevent infection and promote wound healing.In this study,a high strength composite cryogel（SA/SF/DA）with SA,SF and PDA as main raw materials was prepared by combining the good water absorption of SA with the chemical hemostatic properties of SF and PDA for the first time,which provides a new idea for the development of hemostatic materials.In order to obtain the optimal preparation parameters,the three factors of SA mass fraction,SF mass fraction and Ca Cl₂soaking time were selected to obtain the optimal level combination,that is,2%SA,2%SF and 12h Ca Cl₂soaking time.Subsequently,the effect of DA on hemostatic properties was studied by adding different contents of DA,and the physical properties of the materials were characterized by FTIR spectra,SEM images,swelling rate,bloodsucking rate,porosity and mechanical properties.The test results show that SA/SF/DA cryogel has a hierarchical porous structure,which is beneficial to the absorption of liquid and the concentration of blood factors.With the increase of DA content,the liquid absorption capacity of SA/SF/DA cryogel decreased gradually,and the mechanical properties increased gradually.Therefore,the liquid absorption capacity and mechanical properties can be adjusted by the change of DA content.The hemostatic properties of cryogel were evaluated by dynamic coagulation index in vitro,rat liver injury bleeding model and rat femoral artery bleeding model.The dynamic coagulation index in vitro showed that the coagulation effect of SA/SF/DA3 was the best,and the BCI index at 30s（36.84±1.87%）and 300s（11.75±0.59%）was significantly lower than that of other groups.In the animal bleeding model,SA/SF/DA3 also showed the shortest bleeding time and the least amount of bleeding.The above results show that SA/SF/DA cryogel has a good ability to promote blood coagulation.Furthermore,the erythrocyte adhesion rate,platelet adhesion rate,intracellular calcium fluorescence intensity,P-selectin（CD62p）expression and thrombin production of cryogel were measured to explore its hemostatic mechanism.First of all,the SA/SF/DA cryogel with hierarchical microporous structure has a capillary effect to absorb a large amount of water in the blood,and rapidly increase the volume and weight in the wound site as a physical barrier to prevent wound bleeding.At the same time,the contact between SA/SF/DA and blood releases Ca²⁺around the skeleton,which promotes the electrostatic interaction of blood cells to achieve physical cross-linking.The platelets and coagulation factors attached to the surface and porous structure of SA/SF/DA will be activated by the porous structure,Ca²⁺and the phenolic hydroxyl and amino groups of PDA,and further trigger the formation of thrombin,which further promotes the formation of fibrin,which leads to faster blood clotting.Hemostatic materials not only need rapid hemostatic ability,but also have good biocompatibility.The biocompatibility of SA/SF/DA cryogel was evaluated by hemolysis test,cytotoxicity test and in vivo degradation test.The results show that SA/SF/DA cryogel has good biocompatibility and has the basic conditions to be used as hemostatic material.