Research Of Hydrogel Microcapsule Encapsulation In Human Red Blood Cell Cryopreservation

Research Background:As an important component of the human body,the importance of blood to people’s health cannot be overstated.According to statistics,in 2018,more than 118.5 million units of blood were donated worldwide alone.Blood transfusions are required in clinical surgery,aplastic anemia,hemorrhage in pregnancy,and some other cases of excessive blood loss.In today’s blood transfusion,whole blood is separated and prepared into single components for transfusion,of which red blood cells（RBCs）are an important part.Most of the methods used for clinical preservation of RBCs are refrigerated at 2-6°C with the addition of RBC preservation solution,which can be stored for about 42 days.However,the refrigeration process suffers from damage such as loss of ATP,2-3 DPG,and reduced membrane fluidity,which can hinder subsequent transfusion.Cell metabolism can be suspended at very low temperatures,and long-term preservation of cells can be achieved by placing them at deep cryogenic temperatures.RBCs have been widely studied as a typical application of cryopreservation.The most widely used clinical methods of RBCs cryopreservation are slow freezing with a high concentration of glycerol at 40% and fast freezing with a low concentration of glycerol at 20%.The higher concentration of glycerol can cause osmotic damage in addition and removal,so clinical staff need to go through complex and careful operations to remove it,which is time-consuming and obviously a potential problem for emergency blood use,such as in military battlefields.Therefore,it is important to investigate a method to reduce glycerol concentration and washing time for rapid freezing of red blood cells.Research Content:In this study,the penetraing cryoprotevtive agent（CPA）glycerol and the non-penetraing cryoprotevtive agent trehalose were studied on the liquid nitrogen rapid freezing of RBCs,and the recovery of RBCs under different hematocrit of the two CPA was compared..A method of hydrogel microcapsule encapsulation was developed and the cryopreservation method was improved.Tesed its freezing recovery of RBCs,and it was compared with RBCs using cryovials as a freezing carrier.The washing method was then improved to characterize the key indicators of post-freeze washed RBCs.Finally,we investigated the protection mechanism of hydrogels during freezing,and using a cryomicroscope platform,we observed the crystallization phenomenon and inferred the possible protection mechanism.Research Results:In the rapid freezing experiments of RBCs,when the hematocrit was low,the preservation effect of trehalose and glycerol was comparable,and the post-freezing recovery was similar.As the hematocrit continued to increase,the post-freezing recovery of trehalose-loaded RBCs differed significantly from that of glycerol.After using hydrogel microcapsules to encapsulate RBCs and adhering them in a stainless steel grid,their recovery after fast freezing was significantly improved compared to that of RBCs frozen in cryovials.The improved washing method was completed within 30 min,and the characterization of the washed RBCs showed no difference from that of fresh RBCs.Image analysis of the cryomicroscopy experiments of the hydrogels revealed that the alginate hydrogels could lower the crystallization point of the solution and reduce the size of the ice crystals and change the density of the ice crystals.In conclusion,hydrogel microencapsulation of RBCs for rapid freezing is a feasible cryopreservation strategy,which provides potential applications for cryopreservation of RBCs as well as clinical infusion,and has good practical value and research significance.