| Background: Islet transplantation is one of the best ways to treat diabetes.It has the advantages of permanent replacement of insulin injection,safety,and fewer complications.However,donors are scarce and the long-term survival of transplanted islets is not the main factor restricting the development of islet transplantation.Massive islet cell death after transplantation is the main cause of the above-mentioned deficiency.Islets are highly vascularized cells,and a large number of islet cells died early after transplantation due to ischemic hypoxia.However,direct contact with blood can cause severe inflammatory reactions.This experiment mainly designed a new type of scaffold,which artificially created suitable conditions to improve the ischemia and hypoxia environment and reduce the early mortality of transplanted islets.Purpose: 1.A new type of medical 3D printing scaffold was made by using 3D printing technology and medical PLA(Di Kan PLDL 7039)material,and the effects of adding VEGF and heparin on the revascularization of the scaffold were compared.2.The effect of pre-implantation of new medical PLA 3D printing scaffold into the skin on the early survival rate of transplanted islets.Method: 1.Independently designed a new type of medical PLA3 D printed scaffold which can suitable for inclusion in transplanted islets.There are three groups according to the different ingredients: group N1 uses ordinary medical PLA3 D printed scaffolds;group N2 uses heparin added medical PLA3 D printed scaffolds;group N3 uses heparin and VEGF medical PLA3 D printed scaffolds.The experimental animals were using healthy c57 male mice,which were randomly divided into 3 groups of 7 mice each.The scaffold was implanted subcutaneously in the back of the mouse.After 14 days,all the scaffolds were removed,Making the HE staining and CD31 immunohistochemistry and to calculate the microvascular density(MVD)to compare the angiogenesis.2.The experimental animals were c57bl/6 mice.The donor islets were digested by collagenase and purified by discontinuous gradient density centrifugation,Implanted marginal number of islets(300 equivalents).The diabetic rat model was made by stz(180mg/kg)and divided into 4 groups:(1)islet scaffold group: the medical PLA 3D printing scaffold was implanted subcutaneously on the back of c57bl/6 mice in advance,and the diabetic rat model was made after 9 days of implantation.14 days later,the islets were allogeneic and the islets were implanted into the subcutaneous scaffold.The scaffold was the one with the best effect in the first part of the experiment.(2)Renal dorsal membrane group: allogeneic subcapsular islet transplantation was performed in rats.(3)Subcutaneous group: subcutaneous islet transplantation was performed in allogeneic rats.(4)blank control group: normal saline was injected subcutaneously.The vital signs and the changes of blood glucose were observed after islet transplantation.Abdominal glucose tolerance test((IPGTT))was performed 14 days after operation.At14 days after implantation,the specimens were taken and the angiogenesis was observed.The function of transplanted islets were observed by insulin immunohistochemical staining.Result: 1.N3 group had the most abundant angiogenesis and the least inflammatory cells.The endothelial cells in group N1 failed to crawl along the pore diameter,and there were more inflammatory cells in group N1 than in group N3,mainly forming abundant blood vessels in the periphery.The number of N2 inflammatory cells and angiogenesis was between the two.Compared with the three groups,the average MVD of N1 was 25.6 ±4.3,the MVD of N2 was 44.4 ±3.3,the MVD of N3 was 71.6 ±3.8.N3,and there was significant difference between N3 and N2;N2and N1;N3 and N1(P<0.001).2.The subcutaneous scaffold group(using the N3 group scaffold)decreased to normal blood glucose at 4.16 ± 0.98 days after transplantation and renal capsule group decreased to normal at 4.33 ± 0.82 days,the two groups have not much different(P>0.05).Blood glucose did not return to normal in the subcutaneous group and the blank control group.The area under Intraperitoneal glucose tolerance test(IPGTT)of islet scaffold group,renal capsule group,subcutaneous group and blank control group were 1663 ±105.9,1730 ±55.33,3734 ±81.75 and 3694 ±125.4,respectively.The islet function in the islet scaffold group and renal capsule group was good,and there was no significant difference(P>0.05);The fuction of subcutaneous group ang blank control group was almost lost,There was significant difference between islet scaffold and them(p < 0.0001).Insulin immunohistochemistry showed that the islet scaffold group and renal capsule group had good islet activity without obvious inflammatory cell involvement;the subcutaneous group was sparse and infiltrated with inflammatory cells.Conclusion: 1.A new medical PLA 3D printed scaffold with heparin and VEGF added at the same time has good histocompatibility.After 14 days of subcutaneous implantation,abundant blood vessels are formed around the stent,and the revascularization effect is good.2.Pre-implantation of new medical PLA 3D printed scaffolds can create a vascular-rich environment in advance under the skin,improve islet transplantation survival,and provide a new direction for clinical islet transplantation surgery. |
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