AN EFFICIENT METHOD FOR ISOLATION AND PURIFICATION OF THE PANCREATIC ISLETS

On the base of the one step, operator independent method which was set up by Christophe A.E., the pancreas was infused with cold University of Wisconsin(UW) solution for the preservation, digested by the collagenase P, circuited with HBSS+5%fetal calf serum(FCS)+10mmol/L Hepes solution, and separated with the stainless steel mesh. The number of the collected islets were 400000～1800000 per pancreas, i.e. about 12150/g pancreas. After purification, the recovery was 350000～1700000 per pancreas, i.e. about 10250/g pancreas, the recovery rate was above 80%, and the purity of the final preparation was above 95%. The insulin secretion in the response to the high concentration glucose (22 mmol/L) stimulation was apparently different on the 1,3,5 day of the cultural islets, which the high level of insulin was three times the low level (5.5 mmol/L) on the 5th day, and the insulin level of the double stimulation under perfusion conditions is apparently higher than low glucose. The result demonstrated that the purified islets were functionally alive. Histological studies also show that the shape of islets are complete, and the β cell was specially stained by the dithizone (DTZ). The Trypan Blue staining had shown the living cell was above 90%. In conclusion, the new method was highly practical and yielded higher concentration of active pancreatic islets.

A humanized mouse model to study human immune response in xenotransplantation

BACKGROUND: A major barrier to the clinical application of xenotransplantation as a treatment option for patients is T cell-mediated rejection. Studies based on experimental rodent models of xenograft tolerance or rejection in vivo have provided useful information about the role of T cell immune response in xenotransplantation. However not all observations seen in rodents faithfully recapitulate the human situation This study aimed to establish a humanized mouse model of xenotransplantation, which mimics xenograft rejection in the context of the human immune system. METHODS: NOD-SCID IL2rγ -/- mice were transplanted with neonatal porcine islet cell clusters (NICC) followed by reconstitution of human peripheral blood mononuclear cells (PBMC). Human leukocyte engraftment and islet xenograft rejection were confirmed by flow cytometric and histological analyses. RESULTS: In the absence of human PBMC, porcine NICC transplanted into NOD-SCID IL2rγ -/- mice revealed excellent graft integrity and endocrine function. Human PBMC demonstrated a high level of engraftment in NOD-SCID IL2rγ -/- mice. Reconstitution of NICC recipient NOD-SCID IL2rγ -/- mice with human PBMC led to the rapid destruction of NICC xenografts in a PBMC number-dependent manner. CONCLUSIONS: Human PBMC-reconstituted NOD-SCID IL2rγ -/- mice provide an ideal model to study human immune responses in xenotransplantation. Studies based on this humanized mouse model will provide insight for improving the outcomes of clinical xenotransplantation.