Well-organized spheroids as a new platform to examine cell interaction and behaviour during organ development.
We present an experimental method allowing the production of three-dimensional organ-like structures, namely microtissues (MTs), in vitro without the need for exogenous extracellular matrix (ECM) or growth factors. Submandibular salivary glands (embryonic day ED14), kidneys (ED13) and lungs (ED13) were harvested from mouse embryos and dissociated into single cells by enzyme treatment. Single cells were seeded into special hanging drop culture plates (InSphero) and cultured for up to 14 days to obtain MTs. This strategy permitted full control of the quantity of seeded cells. The development of the MTs into organs was followed histologically and immunohistochemically. Well-organized epithelial structures surrounded by a basal lamina were formed, as confirmed by transmission electron microscopy. Expression of E-cadherin, vimentin, fibronectin and α-SMA was compared in organs and corresponding MTs by real-time quantitative polymerase chain reaction. Branching morphogenesis was induced in MTs (as shown by histology and immunostaining for fibronectin and perlecan) and was conserved even after 14 days of culture. MTs continued their development and their epithelial structures were comparable with those of the physiological organ at postnatal day 2 (PN2). Expression of aquaporins was investigated to obtain better support for the functional differentiation of epithelial cells. Histogenesis proceeded and led to the start of organogenesis. This experimental model might improve our knowledge of epithelial-mesenchymal histogenesis and can be employed to study development or cellular organization during the embryonic formation of organs.
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France.|Faculté de Chirurgie Dentaire, Université de Lille, Lille, France.
Thibault
T
Bécavin
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France. kuchler@unistra.fr.|Faculté de Chirurgie Dentaire, Université de Strasbourg, 1 Place de l'Hôpital, Strasbourg, F-67000, France. kuchler@unistra.fr.
Sabine
S
Kuchler-Bopp
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France.|Faculté de Chirurgie Dentaire, Université de Strasbourg, 1 Place de l'Hôpital, Strasbourg, F-67000, France.|INSERM U954-NEGRE, Université de Lorraine, Vandoeuvre-les-Nancy, France.
Tunay
T
Kökten
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France.|Faculté de Chirurgie Dentaire, Université de Strasbourg, 1 Place de l'Hôpital, Strasbourg, F-67000, France.
Olivier
O
Huck
Service de Microscopie Electronique, Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U 964, CNRS UMR 1704, UDS, Illkirch, France.
Nadia
N
Messaddeq
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France.|Faculté de Chirurgie Dentaire, Université de Strasbourg, 1 Place de l'Hôpital, Strasbourg, F-67000, France.
Hervé
H
Lesot
Faculté de Chirurgie Dentaire, Université de Lille, Lille, France.
Etienne
E
Deveaux
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France.|Faculté de Chirurgie Dentaire, Université de Strasbourg, 1 Place de l'Hôpital, Strasbourg, F-67000, France.
Nadia
N
Benkirane-Jessel
INSERM (French National Institute of Health and Medical Research), "Osteoarticular and Dental Regenerative Nanomedicine" laboratory, UMR 1109, Faculté de Médecine, FMTS, F-67085, Strasbourg Cedex, France.|Faculté de Chirurgie Dentaire, Université de Strasbourg, 1 Place de l'Hôpital, Strasbourg, F-67000, France.
Keller
K
Laetitia
D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002450:Cell Communication; D002478:Cells, Cultured; D004848:Epithelium / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation; D008648:Mesoderm / Q000378:metabolism; D008813:Mice, Inbred ICR; D038081:Organogenesis; D012469:Salivary Glands / Q000378:metabolism / Q000648:ultrastructure; D018874:Spheroids, Cellular / Q000166:cytology* / Q000378:metabolism*
D015820:Cadherins
Development; Epithelial-mesenchymal interaction; Microtissue; Spheroid; Three-dimensional cell culture