Oxygenation and adenosine deaminase support growth and proliferation of ex vivo cultured Drosophila wing imaginal discs

Abstract

The Drosophila wing imaginal disc has been an important model system over past decades for discovering novel biology related to development, signaling and epithelial morphogenesis. Novel experimental approaches have been enabled using a culture setup that allows ex vivo cultures of wing discs. Current setups, however, are not able to sustain both growth and cell-cycle progression of wing discs ex vivo. We discover here a setup that requires both oxygenation of the tissue and adenosine deaminase activity in the medium, and supports both growth and proliferation of wing discs for 9 h. Nonetheless, further work will be required to extend the duration of the culturing and to enable live imaging of the cultured discs in the future.

Cell dynamics underlying oriented growth of theDrosophilawing imaginal disc

“…Although previous studies have used insulin to sustain discs long term in culture (Zartman et al, 2013;Handke et al, 2014;Mao et al, 2011Mao et al, , 2013Legoff et al, 2013;Heller et al, 2016;Tsao et al, 2016;Strassburger et al, 2017), we show that the expression of numerous genes involved in patterning declines when discs are cultured without the steroid hormone 20E. 20E was previously thought to promote imaginal growth by repressing the expression of the translational inhibitor 4E-BP (Herboso et al, 2015); however, we show here that, in addition, 20E broadly affects many different signaling pathways, each one of which is crucial for growth.…”

“…Under well-fed conditions, insulin signaling must be able to combine with 20E signaling to further increase imaginal disc growth, but this feature is not well-reproduced in culture (Fig. 1Biv) (Handke et al, 2014;Strassburger et al, 2017). Drosophila has seven insulin-like peptides that interact not only with the single receptor, but also with other proteins that modulate their activities (Honegger et al, 2008;Arquier et al, 2008;Okamoto et al, 2013).…”

“…Insulin signaling regulates animal size in response to nutrition, and bovine insulin has been widely used to stimulate growth of explanted discs (Zartman et al, 2013;Handke et al, 2014;Mao et al, 2011Mao et al, , 2013Legoff et al, 2013;Heller et al, 2016;Tsao et al, 2016;Strassburger et al, 2017). However, proliferation in insulin-cultured discs arrests within a few hours (Handke et al, 2014;Tsao et al, 2016); thus, clearly other signals are required for long-term growth.…”

Multi-view light-sheet imaging and tracking with the MaMuT software reveals the cell lineage of a direct developing arthropod limb

“…So far, these processes have not been possible to live image and quantify in direct developing arthropod limbs. Our understanding of cell dynamics shaping arthropod limbs has relied exclusively on studies of the indirectly developing Drosophila limbs (primarily the wing disc) using clonal analysis and lineage tracing across fixed specimens ( Baena-López et al, 2005 ; González-Gaitán et al, 1994 ; Resino et al, 2002 ; Weigmann and Cohen, 1999 ; Worley et al, 2013 ) and recent improvements in imaging discs in vivo and ex vivo ( Dye et al, 2017 ; Heemskerk et al, 2014 ; Legoff et al, 2013 ; Mao et al, 2013 ; Strassburger et al, 2017 ; Tsao et al, 2016 ; Zartman et al, 2013 ).…”

Reverse-engineering organogenesis through feedback loops between model systems

“…The development of culture conditions that recapitulates long-term post-embryonic micro-organ growth remains an exciting challenge in the field. Recent years have witnessed a resurgence in efforts to the develop new media formulations [57].…”