Identification and Characterization of α3β1 Integrin on Primary and Transformed Rat Islet Cells

Kantengwa, Salomé; Baetens, D.; Sadoul, Karin; Buck, Clayton A.; Halban, Philippe A.; Rouiller, D.

doi:10.1006/excr.1997.3803

Cited by 67 publications

(63 citation statements)

References 44 publications

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“…Expression of 6 1 integrin on isolated -cells and p a n c r e a s . I m m u n o fluorescence for the 3 integrin subunit resulted in labeling of all -cells, as expected from our previous studies (15). Because of the absence of heterogeneity in expression between -cells, and lack of stimulation by secretagogues (data not shown), it was estimated that 3 1 was not responsible, at least by itself, for the observed effects of -cell-matrix interactions.…”

Section: Spreading Of -Cells Is Influenced By Insulinsupporting

confidence: 82%

Importance of cell-matrix interactions in rat islet beta-cell secretion in vitro: role of alpha6beta1 integrin.

et al. 2000

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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...

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Section: Spreading Of -Cells Is Influenced By Insulinsupporting

confidence: 82%

Importance of cell-matrix interactions in rat islet beta-cell secretion in vitro: role of alpha6beta1 integrin.

et al. 2000

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“…The enzymatic and mechanical dissociation of the pancreatic cells may alter their behavior. Furthermore, islets have been shown to contain predominantly the ␣3␤1 integrin, which does not bind laminin-1 (57). We deduced from the data compiled by Jiang et al (56) that stem cells of the pancreas initially require laminin-1 contact to allow later endocrine differentiation when laminin-1 is removed.…”

Section: Discussionmentioning

confidence: 89%

Expression and role of laminin-1 in mouse pancreatic organogenesis.

et al. 2000

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Previous studies have suggested that basement membrane alone may induce ductal differentiation and morphogenesis in the undifferentiated embryonic pancreas. The mechanism by which this induction occurs has not been investigated. Studies of other organ systems such as the lungs and mammary glands, where differentiation has been shown to be induced by basement membrane, have suggested a major role for laminin as a mediator of ductal or tubular morphogenesis and differentiation. We first defined the ontogeny of laminin-1 in the developing mouse pancreas. To determine the specific role of basement membrane laminin in pancreatic ductal morphogenesis and differentiation, we microdissected 11-day mouse embryonic pancreatic epithelium free from its surrounding mesenchyme and then suspended the explants in a 3-dimensional organ culture to allow us to assay cell differentiation and morphogenesis. When the pancreatic epithelium buds off the foregut endoderm, the pancreatic mesenchyme diffusely expresses laminin-1. This laminin subsequently organizes to the interface between the epithelium and the mesenchyme by E12.5. As gestation progresses, epithelial cells in direct contact with laminin-1 seem to differentiate into ducts and acini, whereas those spared intimate contact with laminin-1 appeared to organize into islets. Although basement membrane gel could induce pancreatic ductal morphogenesis of embryonic pancreatic epithelium, this induction was blocked when we added neutralizing antibodies against any of the following: 1) laminin (specifically laminin-1), 2) the "cross-region" of laminin-1, and 3) the ␣6 moiety of the integrin receptor, which is known to bind laminins. Immunohistochemistry, however, showed that pancreatic duct cell-specific differentiation (carbonic anhydrase II) without ductal morphogenesis was still present, despite the blockage of duct morphogenesis by the anti-laminin-1 neutralizing antibodies. Interestingly, there appeared to be a decrease in carbonic anhydrase II expression over time when the epithelia were grown in a collagen gel, rather than in a basement membrane gel. The pattern of laminin-1 expression in the embryonic pancreas supports the conclusion that laminin-1 is important in the induction of exocrine (ducts and acini) differentiation in the pancreas. Furthermore, our data demonstrate that 1) pancreatic ductal morphogenesis appears to require basement membrane laminin-1 and an ␣6-containing integrin receptor; 2) the cross-region of basement membrane laminin is a biologically active locus of the laminin molecule necessary for pancreatic ductal morphogenesis; 3) duct-specific cytodifferentiation, in the form of carbonic anhydrase II expression, is not necessarily coupled to duct morphogenesis; and 4) the basement membrane gel may contain components (e.g., growth factors) other than laminin-1 that can sustain both carbonic anhydrase II expression and, possibly, the capacity to form ducts, despite the absence of duct structures. Diabetes 49:936-944, 2000

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“…Studies from our own group have further demonstrated the specific role of integrins α3β1 and α6β1 on rat beta cells [12,33]. Disruption of inter-islet cell contacts and purification of beta cells away from other cell types possibly contributing to the deposition of ECM components in islet cultures was considered to be a confounding factor in allowing the purified cells to thrive in culture.…”

Section: Discussionmentioning

confidence: 99%

Impact of integrin-matrix matching and inhibition of apoptosis on the survival of purified human beta-cells in vitro

et al. 2002

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Aims/hypothesis. Human islet cells survive poorly in culture and are overgrown by non-endocrine cells. The aims of this study were to sort human beta cells and to develop approaches for their improved survival in culture. Methods. Human islets were infected with recombinant adenovirus expressing green fluorescent protein (GFP) under the control of the rat insulin promoter such that only beta cells expressed GFP. GFP-positive beta cells were sorted by flow cytometry, and expression of select integrins evaluated by RT-PCR. Beta cells were cultured on different extracellular matrices for up to 15 days. Apoptosis was measured by annexin V binding and ELISA. Insulin secretion was measured by ELISA. Results. Sorted beta cells survived less well in culture than unsorted islet cells. This did not appear to be due to adenoviral infection and/or GFP expression. Purified beta cells expressed the integrins α3, α5, α6, αV, β1, but not β4. Of the various matrices tested, sorted beta cells attached and spread best on a lawn of lysed human bladder carcinoma cells (5637 cells). However, survival remained poor. Cell death was decreased but not prevented by continued presence of 10 mmol/l nicotinamide and apoptosis decreased by 24 h incubation with 20 µmol/l Z-VAD. Insulin secretion was maintained over 6 days following treatment with both agents. Conclusions/interpretation. Purification of human beta cells induces marked apoptosis limiting their function and survival in vitro. This was improved by matching the extracellular matrix to the specific expression of integrins and by addition of nicotinamide and Z-VAD. [Diabetologia (2002) 45:841-850]

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Identification and Characterization of α3β1 Integrin on Primary and Transformed Rat Islet Cells

Cited by 67 publications

References 44 publications

Importance of cell-matrix interactions in rat islet beta-cell secretion in vitro: role of alpha6beta1 integrin.

Importance of cell-matrix interactions in rat islet beta-cell secretion in vitro: role of alpha6beta1 integrin.

Expression and role of laminin-1 in mouse pancreatic organogenesis.

Impact of integrin-matrix matching and inhibition of apoptosis on the survival of purified human beta-cells in vitro

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