Top

Published in:

01-01-2008 | Article

Proliferation of sorted human and rat beta cells

Authors: G. Parnaud, D. Bosco, T. Berney, F. Pattou, J. Kerr-Conte, M. Y. Donath, C. Bruun, T. Mandrup-Poulsen, N. Billestrup, P. A. Halban

Published in: Diabetologia | Issue 1/2008

Abstract

Aims/hypothesis

The aim of the study was to determine whether purified beta cells can replicate in vitro and whether this is enhanced by extracellular matrix (ECM) and growth factors.

Methods

Human beta cells were purified by FACS by virtue of their high zinc content using Newport Green, and excluding ductal and dead cells. Rat beta cells were sorted by autofluorescence or using the same method developed for human cells. Cells were plated on poly-l-lysine or ECMs from rat or human bladder carcinoma cells or bovine corneal ECM and incubated in the presence of BrdU with or without growth factors.

Results

The newly developed method for sorting human beta cells yields a population containing 91.4 ± 2.8% insulin-positive cells with a low level of spontaneous apoptosis and a robust secretory response to glucose. Beta cells from 8-week-old rats proliferated in culture and this was increased by ECM. Among growth factors, only human growth hormone (hGH) and the glucagon-like peptide-1 analogue liraglutide enhanced proliferation of rat beta cells, with a significant increase on both poly-l-lysine and ECM. By contrast, sorted adult human beta cells from 16 donors aged 48.9 ± 14.3 years (range 16–64 years) failed to replicate demonstrably in vitro regardless of the substratum or growth factors used.

Conclusions/interpretation

These findings indicate that, in our conditions, the fully differentiated human adult insulin-producing beta cell was unable to proliferate in vitro. This has important implications for any attempt to expand cells from pancreases of donors of this age group. By contrast, the rat beta cells used here were able to divide in vitro, and this was enhanced by ECM, hGH and liraglutide.

Available only for authorised users

Hisaoka M, Haratake J, Hashimoto H (1993) Pancreatic morphogenesis and extracellular matrix organization during rat development. Differentiation 53:163–172PubMedCrossRef

Lelievre S, Weaver VM, Bissell MJ (1996) Extracellular matrix signaling from the cellular membrane skeleton to the nuclear skeleton: a model of gene regulation. Recent Prog Horm Res 51:417–432PubMed

Li S, Edgar D, Fassler R, Wadsworth W, Yurchenco PD (2003) The role of laminin in embryonic cell polarization and tissue organization. Dev Cell 4:613–624PubMedCrossRef

Roskelley CD, Srebrow A, Bissell MJ (1995) A hierarchy of ECM-mediated signalling regulates tissue-specific gene expression. Curr Opin Cell Biol 7:736–747PubMedCrossRef

Bosco D, Gonelle-Gispert C, Wollheim CB, Halban PA, Rouiller DG (2001) Increased intracellular calcium is required for spreading of rat islet beta-cells on extracellular matrix. Diabetes 50:1039–1046PubMedCrossRef

Bosco D, Meda P, Halban PA, Rouiller DG (2000) Importance of cell-matrix interactions in rat islet beta-cell secretion in vitro: role of alpha6beta1 integrin. Diabetes 49:233–243PubMedCrossRef

Hammar E, Parnaud G, Bosco D et al (2004) Extracellular matrix protects pancreatic beta-cells against apoptosis: role of short-and long-term signaling pathways. Diabetes 53:2034–2041PubMedCrossRef

Beattie GM, Cirulli V, Lopez AD, Hayek A (1997) Ex vivo expansion of human pancreatic endocrine cells. J Clin Endocrinol Metab 82:1852–1856PubMedCrossRef

Beattie GM, Montgomery AM, Lopez AD et al (2002) A novel approach to increase human islet cell mass while preserving beta-cell function. Diabetes 51:3435–3439PubMedCrossRef

10.

Beattie GM, Itkin-Ansari P, Cirulli V et al (1999) Sustained proliferation of PDX-1+ cells derived from human islets. Diabetes 48:1013–1019PubMedCrossRef

11.

Lefebvre VH, Otonkoski T, Ustinov J, Huotari MA, Pipeleers DG, Bouwens L (1998) Culture of adult human islet preparations with hepatocyte growth factor and 804G matrix is mitogenic for duct cells but not for beta-cells. Diabetes 47:134–137PubMedCrossRef

12.

Bai L, Meredith G, Tuch BE (2005) Glucagon-like peptide-1 enhances production of insulin in insulin-producing cells derived from mouse embryonic stem cells. J Endocrinol 186:343–352PubMedCrossRef

13.

Bernal-Mizrachi E, Wen W, Srinivasan S, Klenk A, Cohen D, Permutt MA (2001) Activation of Elk-1, an Ets transcription factor, by glucose and EGF treatment of insulinoma cells. Am J Physiol Endocrinol Metab 281:E1286–E1299PubMed

14.

Buteau J, Foisy S, Joly E, Prentki M (2003) Glucagon-like peptide 1 induces pancreatic beta-cell proliferation via transactivation of the epidermal growth factor receptor. Diabetes 52:124–132PubMedCrossRef

15.

Friedrichsen BN, Galsgaard ED, Nielsen JH, Moldrup A (2001) Growth hormone- and prolactin-induced proliferation of insulinoma cells, INS-1, depends on activation of STAT5 (signal transducer and activator of transcription 5). Mol Endocrinol 15:136–148PubMedCrossRef

16.

Huotari MA, Palgi J, Otonkoski T (1998) Growth factor-mediated proliferation and differentiation of insulin-producing INS-1 and RINm5F cells: identification of betacellulin as a novel beta-cell mitogen. Endocrinology 139:1494–1499PubMedCrossRef

17.

Sutton R, Peters M, McShane P, Gray DW, Morris PJ (1986) Isolation of rat pancreatic islets by ductal injection of collagenase. Transplantation 42:689–691PubMedCrossRef

18.

Rouiller DG, Cirulli V, Halban PA (1990) Differences in aggregation properties and levels of the neural cell adhesion molecule (NCAM) between islet cell types. Exp Cell Res 191:305–312PubMedCrossRef

19.

Van De Winkel M, Smets G, Gepts W, Pipeleers D (1982) Islet cell surface antibodies from insulin-dependent diabetics bind specifically to pancreatic B cells. J Clin Invest 70:41–49CrossRef

20.

Kato Y, Gospodarowicz D (1985) Effect of exogenous extracellular matrices on proteoglycan synthesis by cultured rabbit costal chondrocytes. J Cell Biol 100:486–495PubMedCrossRef

21.

Friedrichsen BN, Neubauer N, Lee YC et al (2006) Stimulation of pancreatic beta-cell replication by incretins involves transcriptional induction of cyclin D1 via multiple signalling pathways. J Endocrinol 188:481–492PubMedCrossRef

22.

Demeterco C, Beattie GM, Dib SA, Lopez AD, Hayek A (2000) A role for activin A and betacellulin in human fetal pancreatic cell differentiation and growth. J Clin Endocrinol Metab 85:3892–3897PubMedCrossRef

23.

Gmyr V, Belaich S, Muharram G et al (2004) Rapid purification of human ductal cells from human pancreatic fractions with surface antibody CA19-9. Biochem Biophys Res Commun 320:27–33PubMedCrossRef

24.

Ichii H, Inverardi L, Pileggi A et al (2005) A novel method for the assessment of cellular composition and beta-cell viability in human islet preparations. Am J Transplant 5:1635–1645PubMedCrossRef

25.

Lukowiak B, Vandewalle B, Riachy R et al (2001) Identification and purification of functional human beta-cells by a new specific zinc-fluorescent probe. J Histochem Cytochem 49:519–528PubMed

26.

Hayek A, Beattie GM, Cirulli V, Lopez AD, Ricordi C, Rubin JS (1995) Growth factor/matrix-induced proliferation of human adult beta-cells. Diabetes 44:1458–1460PubMedCrossRef

27.

Gershengorn MC, Hardikar AA, Wei C, Geras-Raaka E, Marcus-Samuels B, Raaka BM (2004) Epithelial-to-mesenchymal transition generates proliferative human islet precursor cells. Science 306:2261–2264PubMedCrossRef

28.

Ouziel-Yahalom L, Zalzman M, Anker-Kitai L et al (2006) Expansion and redifferentiation of adult human pancreatic islet cells. Biochem Biophys Res Commun 341:291–298PubMedCrossRef

29.

Atouf F, Park CH, Pechhold K, Ta M, Choi Y, Lumelsky NL (2007) No evidence for mouse pancreatic beta-cell epithelial–mesenchymal transition in vitro. Diabetes 56:699–702PubMedCrossRef

30.

Chase LG, Ulloa-Montoya F, Kidder BL, Verfaillie CM (2007) Islet-derived fibroblast-like cells are not derived via epithelial–mesenchymal transition from Pdx-1 or insulin-positive cells. Diabetes 56:3–7PubMedCrossRef

31.

Morton RA, Geras-Raaka E, Wilson LM, Raaka BM, Gershengorn MC (2007) Endocrine precursor cells from mouse islets are not generated by epithelial-to-mesenchymal transition of mature beta cells. Mol Cell Endocrinol 270:87–93PubMedCrossRef

32.

Weinberg N, Ouziel-Yahalom L, Knoller S, Efrat S, Dor Y (2007) Lineage tracing evidence for in vitro dedifferentiation but rare proliferation of mouse pancreatic beta-cells. Diabetes 56:1299–1304PubMedCrossRef

33.

Gmyr V, Kerr-Conte J, Belaich S et al (2000) Adult human cytokeratin 19-positive cells reexpress insulin promoter factor 1 in vitro: further evidence for pluripotent pancreatic stem cells in humans. Diabetes 49:1671–1680PubMedCrossRef

34.

Adams JC, Watt FM (1993) Regulation of development and differentiation by the extracellular matrix. Development 117:1183–1198PubMed

35.

Parnaud G, Hammar E, Rouiller DG, Armanet M, Halban PA, Bosco D (2006) Blockade of beta1 integrin-laminin-5 interaction affects spreading and insulin secretion of rat beta-cells attached on extracellular matrix. Diabetes 55:1413–1420PubMedCrossRef

36.

Sorenson RL, Brelje TC (1997) Adaptation of islets of Langerhans to pregnancy: beta-cell growth, enhanced insulin secretion and the role of lactogenic hormones. Horm Metab Res 29:301–307PubMedCrossRef

37.

Parsons JA, Brelje TC, Sorenson RL (1992) Adaptation of islets of Langerhans to pregnancy: increased islet cell proliferation and insulin secretion correlates with the onset of placental lactogen secretion. Endocrinology 130:1459–1466PubMedCrossRef

38.

Brelje TC, Scharp DW, Lacy PE et al (1993) Effect of homologous placental lactogens, prolactins, and growth hormones on islet B-cell division and insulin secretion in rat, mouse, and human islets: implication for placental lactogen regulation of islet function during pregnancy. Endocrinology 132:879–887PubMedCrossRef

39.

Billestrup N, Nielsen JH (1991) The stimulatory effect of growth hormone, prolactin, and placental lactogen on beta-cell proliferation is not mediated by insulin-like growth factor-I. Endocrinology 129:883–888PubMedCrossRef

40.

Kieffer TJ, Habener JF (1999) The glucagon-like peptides. Endocr Rev 20:876–913PubMedCrossRef

41.

Drucker DJ (2003) Glucagon-like peptide-1 and the islet beta-cell: augmentation of cell proliferation and inhibition of apoptosis. Endocrinology 144:5145–5148PubMedCrossRef

42.

Drucker DJ (2003) Glucagon-like peptides: regulators of cell proliferation, differentiation, and apoptosis. Mol Endocrinol 17:161–171PubMedCrossRef

43.

Brubaker PL, Drucker DJ (2004) Minireview: glucagon-like peptides regulate cell proliferation and apoptosis in the pancreas, gut, and central nervous system. Endocrinology 145:2653–2659PubMedCrossRef

44.

Buteau J, Roduit R, Susini S, Prentki M (1999) Glucagon-like peptide-1 promotes DNA synthesis, activates phosphatidylinositol 3-kinase and increases transcription factor pancreatic and duodenal homeobox gene 1 (PDX-1) DNA binding activity in beta (INS-1)-cells. Diabetologia 42:856–864PubMedCrossRef

45.

Teta M, Long SY, Wartschow LM, Rankin MM, Kushner JA (2005) Very slow turnover of beta-cells in aged adult mice. Diabetes 54:2557–2567PubMedCrossRef

46.

Maedler K, Schumann DM, Schulthess F et al (2006) Aging correlates with decreased beta-cell proliferative capacity and enhanced sensitivity to apoptosis: a potential role for Fas and pancreatic duodenal homeobox-1. Diabetes 55:2455–2462PubMedCrossRef

Title: Proliferation of sorted human and rat beta cells
Authors: G. Parnaud
D. Bosco
T. Berney
F. Pattou
J. Kerr-Conte
M. Y. Donath
C. Bruun
T. Mandrup-Poulsen
N. Billestrup
P. A. Halban
Publication date: 01-01-2008
Publisher: Springer-Verlag
Published in: Diabetologia / Issue 1/2008
Print ISSN: 0012-186X
Electronic ISSN: 1432-0428
DOI: https://doi.org/10.1007/s00125-007-0855-1

Live Webinar | 27-06-2024 | 18:00 (CEST)

Keynote webinar | Spotlight on medication adherence

Reserve your place

Live: Thursday 27th June 2024, 18:00-19:30 (CEST)

WHO estimates that half of all patients worldwide are non-adherent to their prescribed medication. The consequences of poor adherence can be catastrophic, on both the individual and population level.

Join our expert panel to discover why you need to understand the drivers of non-adherence in your patients, and how you can optimize medication adherence in your clinics to drastically improve patient outcomes.

Prof. Kevin Dolgin

Prof. Florian Limbourg

Prof. Anoop Chauhan

Developed by: Springer Medicine

Obesity Clinical Trial Summary

At a glance: The STEP trials

A round-up of the STEP phase 3 clinical trials evaluating semaglutide for weight loss in people with overweight or obesity.

Developed by: Springer Medicine

Keynote webinar | Spotlight on medication adherence

Springer Medicine

Abstract

Aims/hypothesis

Methods

Results

Conclusions/interpretation

Please log in to get access to this content

Other articles of this Issue 1/2008

Oral taurine but not N-acetylcysteine ameliorates NEFA-induced impairment in insulin sensitivity and beta cell function in obese and overweight, non-diabetic men

Fat depot-related differences in gene expression, adiponectin secretion, and insulin action and signalling in human adipocytes differentiated in vitro from precursor stromal cells

Association analysis of podocyte slit diaphragm genes as candidates for diabetic nephropathy

Metabolic aspects of pig-to-monkey (Macaca fascicularis) islet transplantation: implications for translation into clinical practice

The GCKR rs780094 polymorphism is associated with elevated fasting serum triacylglycerol, reduced fasting and OGTT-related insulinaemia, and reduced risk of type 2 diabetes

Diabetic polyneuropathy is associated with respiratory muscle impairment in type 2 diabetes

Keynote webinar | Spotlight on medication adherence

Live: Thursday 27th June 2024, 18:00-19:30 (CEST)

At a glance: The STEP trials