Cyclic Adenosine-3′,5′ -monophosphate Stimulates Islet B Cell Replication in Neonatal Rat Pancreatic Monolayer Cultures

Rabinovitch, Alex; Blondel, Benigna; Murray, Thomas; Mintz, Daniel H.

doi:10.1172/jci109935

Cited by 51 publications

(21 citation statements)

References 37 publications

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“…Similar results were obtained when 10 ,uM Ro.20-1724 was added instead of50 ,M IBMX (results not shown). seen that DNA synthesis is markedly stimulated after a lag of 16 A number of reports have suggested a role for cAMP in stimulating proliferation in cell types other than fibroblasts (22)(23)(24)(25)(26)(27)(28)(29)(30)(31). However, there are important differences between these studies and the present findings with 3T3 cells.…”

Section: Methodscontrasting

confidence: 82%

“…2 and 3 (15,16,18). Other studies performed with mixed cell populations (26,30,31) or in vivo (26) are difficult to interpret because changes in cAMP levels cannot be properly documented and because it cannot be ruled out that cAMP is acting indirectly by promoting the release ofa growth factor. Finally, in contrast to most previous studies, our experiments with 3T3 cells were con- of DNA synthesis (s) by cholera toxin and IBMX.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Cyclic AMP: a mitogenic signal for Swiss 3T3 cells.

Rozengurt¹,

Legg²,

Strang³

et al. 1981

Proc. Natl. Acad. Sci. U.S.A.

135

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Addition ofcholera toxin (100 ng/ml) to quiescent cultures of Swiss 3T3 cells acts synergistically with serum (2-4%), insulin, phorbol esters, epidermal growth factor, and fibroblastderived growth factor to stimulate DNA synthesis. In the presence of insulin, cholera toxin caused a dose-dependent increase in cumulative [3H]thymidine incorporation into acid-insoluble material and in the intracellular cyclic AMP (cAMP) level. The dose-response curves for the two processes were similar. Furthermore, addition of 1-methyl-3-isobutylxanthine (15-500 FLM) or of 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (5-100 gLM), both of which are potent inhibitors ofcyclic nucleotide phosphodiesterase activity, stimulated DNA synthesis and increased cAMP levels in Swiss 3T3 cells. These compounds strikingly potentiated the effect of cholera toxin on DNA synthesis and on cAMP levels. When quiescent Swiss 3T3 cells were exposed to cholera toxin (100 ng/ ml) and insulin at 10 pg/ml (4-to 7-fold increase in cAMP level) or to these agents and 1-methyl-3-isobutyl xanthine at 50 gIM (35-fold increase in cAMP level), DNA (6)(7)(8)(9)(10)). An objection to many ofthese studies has been that these effects were elicited by high concentrations of analogues of cAMP and could be regarded as nonspecific (2, 4). Furthermore, a few other studies with 3T3 cells suggested that cAMP promotes proliferation, but this contention remained unproven. Thus, addition of cAMP enhanced the initiation of DNA synthesis produced by serum, but noncyclic purine nucleotides or nucleosides were also effective (11). In addition, cholera toxin, an irreversible activator of the adenylate cyclase of eukaryote cells (12) was reported to stimulate, rather than inhibit, cell division in 3T3 cells maintained in-the presence of serum (13).However, the mitogenic effect of cholera toxin was not mimicked by other agents that elevated cAMP levels of 3T3 cells (13). All these inconsistent findings preclude any definitive conclusion concerning the effect ofcAMP on the initiation of DNA synthesis of fibroblast cell lines.A complicating factor in most early studies concerning the effect of cyclic nucleotides on the initiation of DNA synthesis is that they have been conducted in cultures maintained in nutrient medium supplemented with unfractionated serum. The complex nature ofserum and its multiplicity ofactions have frequently rendered difficult the interpretations of experiments with cultured animal cells grown in medium containing serum. In recent years, certain combinations of insulin, epidermal growth factor (EGF), vasopressin, phorbol esters, and fibroblast-derived growth factor (FDGF), a cell-derived growth factor, have been shown to completely replace serum in stimulating DNA synthesis in quiescent cultures of 3T3 cells (1, 14-16). We used such mitogenic factors and defined culture conditions to assess the effect ofcAMP elevating agents on the stimulation of DNA synthesis in cultures of Swiss 3T3 cells. In contrast to previous reports (2-10), we found that incr...

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Section: Methodscontrasting

confidence: 82%

Section: Methodsmentioning

confidence: 99%

Cyclic AMP: a mitogenic signal for Swiss 3T3 cells.

Rozengurt¹,

Legg²,

Strang³

et al. 1981

Proc. Natl. Acad. Sci. U.S.A.

135

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“…In general, increasing cytosolic cAMP levels stimulates ␤-cell mitogenesis in most of the experimental ␤-cell models (9,25,37), with the exception of fetal rat islets (38). In INS-1 cells, GLP-1 has been shown to induce an increase of intracellular cAMP levels independently of glucose and induce mitogenesis, in part via PI3K activation leading to PKC activation (24), perhaps by a direct interaction between G protein-coupled GLP-1 receptor and IGF-1 receptor itself (39).…”

Section: S6kmentioning

confidence: 99%

Differential Activation Mechanisms of Erk-1/2 and p70S6K by Glucose in Pancreatic β-Cells

et al. 2003

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“…Monolayer cultures of islet cells were prepared by a modification of a collagenase-trypsin method (6) as described (18) . Briefly, the pancreases of newborn Wistar rats were enzymatically dissociated by ten successive 10-min incubations at 37°C in a Ca"-and Mg"-free phosphate-buffered saline (PBS) containing 5.6 mM glucose, 5 mg/ml trypsin (1 :250, Difco Laboratories, Detroit, Mich.) and 0.2 mg/ml of collagenase (CLS IV, Worthington Biochemical Corp., Freehold, N.…”

Section: Islet Cell Culturesmentioning

confidence: 99%

Direct communication of homologous and heterologous endocrine islet cells in culture.

Meda¹,

Kohen²,

Kohen³

et al. 1982

The Journal of Cell Biology

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The transfer of 6-carboxyfluorescein between islet cells in monolayer culture was observed by fluorescence microscopy, and the endocrine cells involved in this transfer were identified by immunohistochemistry and electron microscopy. The results show that carboxyfluorescein was directly exchanged between homologous B-cells and also between B-and A-or D-cells . Successive microinjections of the probe into different cells of the same cluster showed the existence of separate territories, each formed by 2-8 communicating cells. Intercellular communication was not observed after every dye microinjection, and communicating and noncom municating islet cells were found to coexist within the same cluster . The data indicate that the exchange of exogenous cytoplasmic molecules occurs between different types of endocrine islet cells. However, within a single cluster, all islet cells are not metabolically coupled to one another, at a given time.Ionic and metabolic cell-to-cell coupling (1) have been recently demonstrated between B-cells of the islets of Langerhans (2,5,8,11). In view of the heterogeneous endocrine cell composition of the islet (14) and of the possible functional interactions between various islet cells (for review, see reference 22), it was important to assess whether coupling occurred between different islet cell types .In the present study we have identified by immunohistochemistry and electron microscopy the islet cells involved in the direct exchange of 6-carboxyfluorescein, an exogenous fluorescent probe (376 mol wt) to which the cell membrane is impermeable (20). Our observations show that, in monolayer culture of neonatal endocrine pancreas, B-cells directly communicate not only with other B-cells but also with neighboring A-and D-cells, indicating that both homologous and heterologous coupling occur between differentiated islet cells . The metabolically coupled cells were found to form separate territories and to coexist with uncoupled cells within the same cluster . MATERIALS AND METHODS Islet Cell CulturesMonolayer cultures of islet cells were prepared by a modification of a collagenase-trypsin method (6) as described (18) . Briefly, the pancreases of newborn Wistar rats were enzymatically dissociated by ten successive 10-min incubations at 37°C in a Ca"-and Mg"-free phosphate-buffered saline (PBS) containing 5.6 mM glucose, 5 mg/ml trypsin (1 :250, Difco Laboratories, Detroit, Mich.) and 0.2 mg/ml of collagenase (CLS IV, Worthington Biochemical Corp., Freehold, N. J.) . The dissociated cells, collected by decantation at the end of THE JOURNAL OF CELL BIOLOGY " VOLUME 92 JANUARY 1982 221-226 © The Rockefeller University Press -0021-9525/82/01/0221/06 $1 .00 RAPID COMMUNICATIONS each of the last eight incubations, were pooled, rinsed twice by centrifugation in medium 199 (Gibco Laboratories, Grand Island Biological Co ., Grand Island, N. Y.) supplemented with 10% heat-inactivated fetal calf serum and 16 .7 mM glucose, resuspended in this medium, and plated at a concentration of 7.5 x 105 ce...

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Cyclic Adenosine-3′,5′ -monophosphate Stimulates Islet B Cell Replication in Neonatal Rat Pancreatic Monolayer Cultures

Cited by 51 publications

References 37 publications

Cyclic AMP: a mitogenic signal for Swiss 3T3 cells.

Cyclic AMP: a mitogenic signal for Swiss 3T3 cells.

Differential Activation Mechanisms of Erk-1/2 and p70S6K by Glucose in Pancreatic β-Cells

Direct communication of homologous and heterologous endocrine islet cells in culture.

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