A Study On Activity And Function Of Porcine Islet Culture In Vitro By Different Method Isolation Purification Porcine Islet

Type 1 is a disease currently affect about 500million people worldwide.Death on account of the complications of diabetes will seem to elevate by more than 60%.The method currently used to islet separation method of collagenase digestion,mechanical shear broken and freezing thawing method.Selective osmotic shock pressure method is found in recent years,a new method of islet cells in young mice,purification of effect obviously,its purification effect also confirmed in young porcine islet cells.But in adult porcine islet cells whether still have the same effect was not reported.During xenotransplantation,Hemeoxygenase-1(HO-1)has been identified as a ubiquitous stress protein induced in many cell types by various stimulants,such as hemolysis,inflammatory cytokines,oxidative stress,heatshock,heavy metals,and endotoxin.So the aim of the paper was to build a novel porcine islet purification method to increase the purity of islet cells for implantation as well as whether HO-lup-regulationina beta-cell and in freshly isolated porcine islets could result in protection from apoptosis and improve in vivo function performance.The main work of this dissertation is summarized as follows:Firstly,a novel islet purification means comprising a hypertonic-hypotonic treatment conducted to increase the purity of adult porcie islet was established and based on the islet function and activity,optimal conditions for islet purity compared with control significant difference.Secondly,The cobaltprotoporphyrin(CoPP)applied to the isolation of islet cell short-time incubation,Found that induced the expression of Hemeoxygenase-1 quantity by CoPP was higher than the control group,determined Hemeoxygenase-1 affected the protection to porcine islet cells.Purified islet cells were randomly divided into five groups,CoPP designed five drug concentration:0 ?mol/L,20 ? mol/L,40 ? mol/L,80 ? mol/L,160 ? mol/L,and islet cell culture together,than by DTZ staining,ELISA insulin release kit and PCR molecular detection.The results showed that the induction of Hemeoxygenase-1 expression can improve the activity of islet cells.In this paper,the results showed that:By using selective osmotic shock(SOS)method separation of porcine islet equivalents quantity(IEQ)was 589104.80 ± 82581.40 significantly higher than the collagenase method islet equivalent quantity(IEQ)267432 ± 31512.27(p<0.05);The result of 7213.47 ± 432.27 per gram tissue islet equivalents quantity(IEQ/g)is significantly higher than collagenase method per gram tissue islet equivalents 3045 ± 986(IEQ/g)(P<0.05);Islet purity(87 ± 2%)of SOS was significantly higher than the collagenase method(73 ± 3%),selective osmotic shock of islet of glucose stimulated insulin secretion increased about three times in the control group(3.06 ± 0.22vs.0.97 ± 0.085).the hypertonic glucose-low hypotonic-glucose can realize higher purity higher reactive extraction islet,and cobaltprotoporphyrin(CoPP)can be induced the expression of Hemeoxygenase-1,increased insulin release quantity and improved the protection of islet cell apoptosis,implement of islet highly active,lay the foundation of successful islet transplantation.