In vitro growth of neonatal rat islet cells is stimulated by adhesion to matrix.
We examined DNA synthesis in non-enzymatically isolated neonatal rat pancreatic islets sub-cultured to eliminate fibroblast contamination, which was excluded both by demonstrating no effect of fibroblast growth factor (FGF) on 3H-thymidine incorporation and by immunofluorescence of attached islets using a mouse anti-fibroblast monoclonal antibody. 3H-thymidine incorporation in islets increased with increasing glucose up to a concentration of 21.1 mM in both free-floating islets (11,789 cpm/micrograms DNA +/- 1,610 SEM) and islets attached to fibronectin coated plastic (43,043 cpm/micrograms DNA +/- 9,203 SEM). These values were significantly higher when compared to 3H-thymidine incorporation in medium containing 11.1 mM glucose (p < 0.007, and p < 0.0001 for free-floating and attached islets respectively). 3H-thymidine incorporation was significantly higher in attached islets than in free-floating islets at all glucose concentrations tested (p < 0.005 at 11.1 mM, 16.1 mM, and 21.1 mM; and p < 0.01 at 26.1 mM). 5-bromo-2'-deoxyuridine (BrDU) staining of islets showed an increased number of positive nuclei in cells localised within attached islets (37.6 nuclei per islet +/- 5.1 SEM) compared to free-floating islets (7.62 nuclei per islet +/- 1.04 SEM, p < 0.001), indicating that attachment influenced proliferation of islet cells not physically in contact with the matrix. No difference in glucose-stimulated insulin release was observed between attached and free-floating islets. In conclusion, a fibroblast free islet culture was used to document the stimulatory effect of islet attachment on DNA synthesis, which was greater than the stimulation exerted by glucose alone.(ABSTRACT TRUNCATED AT 250 WORDS)
Hulinsky
I
I
R. Williams Institute of Paediatric Endocrinology, Royal Alexandra Hospital, Camperdown, NSW, Australia.
Cooney
S
S
Harrington
J
J
Silink
M
M
D000818:Animals; D002448:Cell Adhesion; D002455:Cell Division; D046508:Culture Techniques; D005947:Glucose / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013936:Thymidine / Q000378:metabolism
D007328:Insulin; D005947:Glucose; D013936:Thymidine