It has long been recognized that islet cell function is rapidly altered in vitro, but can be maintained, at least in part, when cells are layered on defined extracellular matrices. The present work addresses the influence of short-term cell-matrix interactions on islet -cell function and provides first insight into the molecular basis of these interactions. When primary rat -cells were allowed to attach to a matrix produced by a rat carcinoma cell line (804G), there was an increased insulin secretory response to secretagogues. This change was the result of an increase in the proportion of actively secreting -cells and in the amount of insulin secreted per active cell, as shown using the reverse hemolytic plaque assay. In turn, the spreading or flattening of -cells on this matrix was enhanced by secretagogues, and flattened cells secreted more insulin than rounded cells. Using indirect immunofluorescence, it was found that 1) 6 1 integrins are present at the surface of islet cells in situ, 2) 6 1 expression is heterogeneous among purified -cells and is upregulated by insulin secretagogues, 3) 6 1 expression is higher in spreading cells, and 4) anti-6 1 -s p e c i fic antibodies decrease spreading. These observations demonstrate that islet cell-matrix interactions can improve the sensitivity of insulin cells to glucose and are mediated, at least in part, by 6 1 integrins, suggesting that outside-in signaling through 6 1 integrin plays a major role in the regulation of -cell function. Diabetes 49:233-243, 2000 P rimary cells of rat islets of Langerhans lose glucose responsiveness and eventually die when maintained in culture for a long period of time. By contrast, the secretory capability of islet cells remains stable when dishes are first coated with components of extracellular matrices (ECMs) (1). Thus, adult rat islets maintained over several weeks on ECM respond to an acute glucose stimulation by a five-to eightfold increase in insulin secretion (2). In human -cells cultured 5-11 days on bovine corneal endothelial cell matrix, both basal and stimulated insulin release were also increased, compared with cells layered on gelatin, collagen, or Matrigel (3). Furthermore, when overlaid with collagen, monolayers of human islet cells undergo a gradual and complete reorganization into a threedimensional islet-like structure with a striking reinforcement of their secretory activity. Under these conditions, cells were able to survive more than 8 weeks (4). By contrast, standard cultures on uncoated plastic petri dishes exhibit a rapid and d e finitive decline in insulin secretion with a survival time not exceeding 14 days (4). Using a reverse hemolytic plaque a s s a y, Perfetti et al. (5) determined that -cells cultured for 6 weeks on Matrigel showed an equal number of insulinsecreting cells compared with freshly isolated islets cultured for only 3 days in the absence of Matrigel. The amount of insulin released per single -cell was nevertheless reduced by as much as 60%. Despite this evidence for a crucial role of islet...

We have shown previously that culture of ␤-cells on matrix derived from 804G cells and rich in laminin-5 improves their function. The purpose of this study was to investigate whether this matrix protects ␤-cells against apoptosis and to elucidate signaling pathways involved. Matrix protected sorted rat ␤-cells against apoptosis under standard conditions (11.2 mmol/l glucose, 10% serum), after serum deprivation (1% serum), and in response to interleukin-1␤ (IL-1␤; 2 ng/ml), compared with control (poly-L-lysine [pLL]). Caspase-8 activity was reduced in cells cultured on matrix, whereas focal adhesion kinase (FAK), protein kinase B (PKB, or Akt), and extracellular signal-regulated kinase (ERK) phosphorylation was augmented. Treatment (4 h) with an anti-␤1 integrin antibody, with the ERK pathway inhibitor PD98059, and/or with the phosphatidylinositol 3-kinase inhibitor LY294002 augmented cell death on 804G matrix but not on pLL. In long-term assays (48 h), PD98059 but not LY294002 drastically augmented cell death on 804G matrix but did so to a lesser extent on pLL. The protein inhibitor of nuclear factor-B (IB␣) was overexpressed in cells cultured 18 h on matrix with partial blockade by PD98059. In summary, this study provides evidence for activation of signaling pathways and gene expression by extracellular matrix leading to improved ␤-cell survival. Diabetes

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Bay 11-7082) or by a recombinant adenovirus expressing the nonphosphorylatable form of IB␣, 804G-ECM-induced cell spreading and actin cytoskeleton organization were reduced. GSIS from cells on 804G-ECM was inhibited 5-fold, whereas cell survival was not affected. In summary, the results indicate that 804G-ECM induces a transient and moderate NF-B activity. This study shows for the first time that ECM-induced NF-B activity is necessary in maintaining GSIS, although it does not affect survival of pancreatic beta cells. The effects of ECM-induced NF-B activity contrast with the deleterious effects of cytokine-induced NF-B activity. It is proposed that transient and moderate NF-B activity is essential for proper function of the pancreatic beta cell.