The Role Of Matrix-integrin α3 Interations In Rat Islet Beta-cell Survival And Function In Vitro

The Role of Matrix-integrin a 3 Interactions in Rat Islet Beta-cell Survival and Function in vitro Abstract Background: Pancreatic islet transplantation can be successful in reversing diabetes. In sharp contrast to the success of whole pancreas grafts, the vast majority of IDDM patients undergoing islet transplantation have exhibited only partial and transient graft function. Islet grafts experience a unique and unexplained high failure rate, with over 60% loss by 3 to 6 months. Furthermore, it has long been recognized that islet cell function and survival is rapidly altered in vitro, with a high rate of death within 48 hours of isolation as a result of apoptosis. In contrast, it appears that islet fuziction and survival are better maintained in vitro when cells are grown in the presence of extracellular matrix. It is probable that destruction of the islet microenvironment (especially the disruption of cell-matrix relationship) and loss of tropic support that occur during isolation lead to compromised survival. The purpose of this study was to examine the role of cell-matrix interactions on islet beta-cell survival and function following islet isolation and provides insight into the molecular basis of these interactions. Objective: 1. To establish an effective model for isolation, purification and long term culture of islets in vitro; 2. To determine the role of cell-matrix interactions on islet beta-cell survival and function in vitro following islet isolation; 3. To provide insights into the molecular basis of these interactions: the expression of integrins a 3 on rat islet cells and its importance in the maintenance of islet structure and function. Methods: 1. Isolation and purification of islet: SD rats (6-8 weeks old, 200-300g) were used as islets donors. Islets were isolated by intraductal -4- collagenase injection and purified by Ficoll-400 discontinuous density gradient. The morphology and function of cultured islets were dynamically examined. 2. Islet cells were cultured on plastic surface or on different extracellular matrix. The morphology and apoptosis of cultured islets were observed during the culture period. 3. The change of the integrin a 3 expression and its relationship with islet cell function and survival in vitro were observed by the methods of immunocytochemistry and flow cytometer. Results: 1. After purification on Ficoll-400 gradients, about 1016?09 islets per pancreas were obtained from one adult SD rat, with an average purity of 90.4% ?2.3%. The purified islets were responded to high concentration glucose stimulation (22.8mmol/L Glucose) with 3.4 times increase of insulin secretion over basal levels (2.3mmol/L Glucose) (P<0.05). 2. After routine islet transplantation, the distribution of the pen-insulin BM in SD rat pancreas was damaged. Glucose-induced insulin secretion declined progressively over 6 days in every group (P<0.05). Insulin secretion from islets in the control group decreased more extensively than other three groups in any time (P<0.05). When cultured 6 days, the insulin release in the control group decreased 78.6% ?5.4%. Otherwise, the COL group, FN group and LAM group decreased 32.2% ?6.7%, 35.5% ?4.9%, 41.3% ?4.5% respectively. During the period of culture, islets experience a high rate of death in all four groups. The control group suffered a gr...