Three-dimensional growth of extravillous cytotrophoblasts promotes differentiation and invasion.
Human trophoblast research relies on a combination of in vitro models, including isolated primary cultures, explant cultures, and trophoblast cell lines. In the present study, we have utilized the rotating wall vessel (RWV) bioreactor to generate a three-dimensional (3-D) model of human placentation for the study of cytotrophoblast (CTB) invasion. The RWV supported the growth of the human CTB cell line SGHPL-4 and allowed for the formation of complex, multilayered 3-D aggregates that were morphologically, phenotypically, and functionally distinct from SGHPL-4 monolayers. The cells cultured three-dimensionally differentiated into an aggressively invasive cell population characterized by the upregulation of matrix metalloproteinase-2 (MMP-2), MMP-3, MMP-9 and urokinase-type plasminogen activator (uPA) secretion and activation. Microarray analysis of the 3-D and 2-D cultured cells revealed increased expression in the 3-D cells of various genes that are known mediators of invasion, including MT1-MMP, PECAM-1 and L-selectin, as well as genes not previously associated with CTB differentiation such as MMP-13 and MT5-MMP. These results were verified by quantitative real-time PCR. These findings suggest that when cultured in 3-D, SGHPL-4 cells closely mimic differentiating in utero CTBs, providing a novel approach for the in vitro study of the molecular mechanisms that regulate CTB differentiation and invasion.
LaMarca
H L
HL
Department of Microbiology and Immunology, Tulane University Health Sciences Center, New Orleans, LA, USA.
Ott
C M
CM
Höner Zu Bentrup
K
K
Leblanc
C L
CL
Pierson
D L
DL
Nelson
A B
AB
Scandurro
A B
AB
Whitley
G St J
GS
Nickerson
C A
CA
Morris
C A
CA
D019149:Bioreactors; D015153:Blotting, Western; D002449:Cell Aggregation / Q000502:physiology; D002454:Cell Differentiation / Q000502:physiology; D048708:Cell Growth Processes / Q000502:physiology; D002460:Cell Line; D005260:Female; D006801:Humans; D019041:L-Selectin / Q000096:biosynthesis / Q000235:genetics; D020782:Matrix Metalloproteinases / Q000235:genetics / Q000378:metabolism; D008855:Microscopy, Electron, Scanning; D008856:Microscopy, Fluorescence; D010929:Placentation / Q000502:physiology*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000096:biosynthesis / Q000235:genetics; D011247:Pregnancy; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014327:Trophoblasts / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism / Q000648:ultrastructure; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism
D019408:Platelet Endothelial Cell Adhesion Molecule-1; D012333:RNA, Messenger; D019041:L-Selectin; D014568:Urokinase-Type Plasminogen Activator; D020782:Matrix Metalloproteinases