Towards long term cultivation of Drosophila wing imaginal discs in vitro

Björn Handke, J. Szabad, Peter V. Lidsky, Ernst Hafen, Christian F. Lehner

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

The wing imaginal disc of Drosophila melanogaster is a prominent experimental system for research on control of cell growth, proliferation and death, as well as on pattern formation and morphogenesis during organogenesis. The precise genetic methodology applicable in this system has facilitated conceptual advances of fundamental importance for developmental biology. Experimental accessibility and versatility would gain further if long term development of wing imaginal discs could be studied also in vitro. For example, culture systems would allow live imaging with maximal temporal and spatial resolution. However, as clearly demonstrated here, standard culture methods result in a rapid cell proliferation arrest within hours of cultivation of dissected wing imaginal discs. Analysis with established markers for cells in S- and M phase, as well as with RGB cell cycle tracker, a novel reporter transgene, revealed that in vitro cultivation interferes with cell cycle progression throughout interphase and not just exclusively during G1. Moreover, quantification of EGFP expression from an inducible transgene revealed rapid adverse effects of disc culture on basic cellular functions beyond cell cycle progression. Disc transplantation experiments confirmed that these detrimental consequences do not reflect fatal damage of imaginal discs during isolation, arguing clearly for a medium insufficiency. Alternative culture media were evaluated, including hemolymph, which surrounds imaginal discs during growth in situ. But isolated larval hemolymph was found to be even less adequate than current culture media, presumably as a result of conversion processes during hemolymph isolation or disc culture. The significance of prominent growth-regulating pathways during disc culture was analyzed, as well as effects of insulin and disc co-culture with larval tissues as potential sources of endocrine factors. Based on our analyses, we developed a culture protocol that prolongs cell proliferation in cultured discs.

Original languageEnglish
Pages (from-to)e107333
JournalPLoS One
Volume9
Issue number9
DOIs
Publication statusPublished - 2014

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Imaginal Discs
imaginal discs
Cell culture
Drosophila
Cells
Cell proliferation
Hemolymph
Culture Media
hemolymph
cell cycle
Cell Cycle
Cell Proliferation
Transgenes
interphase
transgenes
Cell growth
cell proliferation
Growth
culture media
Developmental Biology

ASJC Scopus subject areas

  • Medicine(all)

Cite this

Towards long term cultivation of Drosophila wing imaginal discs in vitro. / Handke, Björn; Szabad, J.; Lidsky, Peter V.; Hafen, Ernst; Lehner, Christian F.

In: PLoS One, Vol. 9, No. 9, 2014, p. e107333.

Research output: Contribution to journalArticle

Handke, Björn ; Szabad, J. ; Lidsky, Peter V. ; Hafen, Ernst ; Lehner, Christian F. / Towards long term cultivation of Drosophila wing imaginal discs in vitro. In: PLoS One. 2014 ; Vol. 9, No. 9. pp. e107333.
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