The differentiation in vitro of the neural crest cells of the mouse embryo

Development ◽  
1984 ◽  
Vol 84 (1) ◽  
pp. 49-62
Author(s):  
Kazuo Ito ◽  
Takuji Takeuchi
Keyword(s):  
Neural Crest ◽  
Mouse Embryo ◽  
Neural Crest Cells ◽  
Culture Method ◽  
Gene Control ◽  
Developmental Potential ◽  
Neural Tubes ◽  
Epithelial Sheet ◽  
Culture Phase

A culture method for neural crest cells of mouse embryo is described. Trunk neural tubes were dissected from 9-day mouse embryos and explanted in culture dishes. The developmental potential of mouse neural crest in vitro was shown to be essentially similar to that of avian neural crest. In the mouse, however, melanocytes always appeared in association with the epithelial sheet close to the explant. Neural crest cells surrounding the epithelial sheet, which probably migrated from the neural tubes in the early culture phase, never differentiated into melanocytes. The bimodal behaviour of mouse crest cells seems to be due to the heterogenous potency of the crest cells and the interaction of these cells with the surrounding microenvironment. This culture system is well suited for various experiments including the analysis of gene control on the differentiation of neural crest cells.

Mouse neural crest cells secrete both urokinase-type and tissue-type plasminogen activators in vitro

Development ◽  
1989 ◽  
Vol 106 (4) ◽  
pp. 685-690 ◽  
Author(s):  
P.A. Menoud ◽  
S. Debrot ◽  
J. Schowing
Keyword(s):  
Neural Crest ◽  
Cultured Cells ◽  
Neural Crest Cells ◽  
Plasminogen Activators ◽  
Cellular Migration ◽  
Tissue Type ◽  
Neural Tubes ◽  
Type Plasminogen Activator ◽  
Anterior Segments

Neural tubes of E8-5 day mouse embryos were dissected and cultured in serum substitute-supplemented medium to allow the emigration of neural crest cells. After 48 h of culture the neural tubes were removed. The neural crest cells were then cultured for 12 h in serum-free medium, and their culture supernatant was studied by electrophoresis and zymography. The cultured cells were shown to secrete both urokinase-type and tissue-type plasminogen activators. When the truncal neural tube was divided in four equal segments, the secretion pattern of the two types of plasminogen activators was similar for the cells from the three most anterior segments; cells having migrated from the most caudal one, i.e. consisting of the neural plate, secreted a higher level of urokinase-type plasminogen activator. The secretion in vitro of plasminogen activators by neural crest cells is in accord with the postulated importance of these proteases in cellular migration.

Migration and proliferation of cultured neural crest cells in W mutant neural crest chimeras

Development ◽  
1991 ◽  
Vol 112 (1) ◽  
pp. 131-141 ◽  
Author(s):  
D. Huszar ◽  
A. Sharpe ◽  
R. Jaenisch
Keyword(s):  
Neural Crest ◽  
Coat Color ◽  
Neural Crest Cell ◽  
Neural Crest Cells ◽  
Experimental Manipulation ◽  
Preimplantation Embryos ◽  
Host Animal ◽  
Developmental Potential ◽  
Host Embryo

Chimeric mice, generated by aggregating preimplantation embryos, have been instrumental in the study of the development of coat color patterns in mammals. This approach, however, does not allow for direct experimental manipulation of the neural crest cells, which are the precursors of melanoblasts. We have devised a system that allows assessment of the developmental potential and migration of neural crest cells in vivo following their experimental manipulation in vitro. Cultured C57Bl/6 neural crest cells were microinjected in utero into neurulating Balb/c or W embryos and shown to contribute efficiently to pigmentation in the host animal. The resulting neural crest chimeras showed, however, different coat pigmentation patterns depending on the genotype of the host embryo. Whereas Balb/c neural crest chimeras showed very limited donor cell pigment contribution, restricted largely to the head, W mutant chimeras displayed extensive pigmentation throughout, often exceeding 50% of the coat. In contrast to Balb/c chimeras, where the donor melanoblasts appeared to have migrated primarily in the characteristic dorsoventral direction, in W mutants the injected cells appeared to migrate in the longitudinal as well as the dorsoventral direction, as if the cells were spreading through an empty space. This is consistent with the absence of a functional endogenous melanoblast population in W mutants, in contrast to Balb/c mice, which contain a full complement of melanocytes. Our results suggest that the W mutation disturbs migration and/or proliferation of endogenous melanoblasts. In order to obtain information on clonal size and extent of intermingling of donor cells, two genetically marked neural crest cell populations were mixed and coinjected into W embryos. In half of the tricolored chimeras, no co-localization of donor crest cells was observed, while, in the other half, a fine intermingling of donor-derived colors had occurred. These results are consistent with the hypothesis that pigmented areas in the chimeras can be derived from extensive proliferation of a few donor clones, which were able to colonize large territories in the host embryo. We have also analyzed the development of pigmentation in neural crest cultures in vitro, and found that neural tubes explanted from embryos carrying wt or weak W alleles produced pigmented melanocytes while more severe W genotypes were associated with deficient pigment formation in vitro.

scholarly journals Maintaining trunk neural crest cells as crestospheres

2018 ◽  
Author(s):  
Sofie Mohlin ◽  
Ezgi Kunttas ◽  
Camilla U. Persson ◽  
Reem Abdel-Haq ◽  
Aldo Castillo ◽  
...  
Keyword(s):  
Neural Crest ◽  
Progenitor Cells ◽  
Neural Crest Cell ◽  
Neural Crest Cells ◽  
Culture Method ◽  
Culture Conditions ◽  
List Type ◽  
Stem And Progenitor Cells ◽  
Trunk Neural Crest

AbstractNeural crest cells have broad migratory and differentiative ability that differs according to their axial level of origin. However, their transient nature has limited understanding of their stem cell and self-renewal properties. While an in vitro culture method has made it possible to maintain cranial neural crest cells as self-renewing multipotent crestospheres (Kerosuo et al., 2015), these same conditions failed to preserve trunk neural crest in a stem-like state. Here we optimize culture conditions for maintenance of trunk crestospheres, comprised of both neural crest stem and progenitor cells. Trunk crestospheres display elevated expression of neural crest cell markers as compared to those characteristic of neural tube or mesodermal fates. Moreover, trunk crestospheres have increased expression of trunk-related markers as compared to cranial genes. Finally, we use lentiviral transduction as a tool to manipulate gene expression in trunk crestospheres. Taken together, this method enables long-term in vitro maintenance and manipulation of trunk neural crest cells in a premigratory stem or early progenitor state to probe the mechanisms underlying their stemness and lineage decisions.HighlightsTrunk-derived multipotent neural crest stem cells can be cultured as crestospheresTrunk-derived crestospheres require different conditions than cranialTrunk crestospheres consist of neural crest stem and progenitor cellsTrunk crestospheres can be efficiently transduced using lentiviral vectors

scholarly journals Developmental potential of trunk neural crest cells in the mouse

Development ◽  
1994 ◽  
Vol 120 (7) ◽  
pp. 1709-1718 ◽  
Author(s):  
G.N. Serbedzija ◽  
M. Bronner-Fraser ◽  
S.E. Fraser
Keyword(s):  
Neural Crest ◽  
Neural Tube ◽  
Mouse Embryo ◽  
Dorsal Root ◽  
Single Cells ◽  
Neural Crest Cell ◽  
Experimental System ◽  
Neural Crest Cells ◽  
Pigment Cells ◽  
Developmental Potential

The availability of naturally occurring and engineered mutations in mice which affect the neural crest makes the mouse embryo an important experimental system for studying neural crest cell differentiation. Here, we determine the normal developmental potential of neural crest cells by performing in situ cell lineage analysis in the mouse by microinjecting lysinated rhodamine dextran (LRD) into individual dorsal neural tube cells in the trunk. Labeled progeny derived from single cells were found in the neural tube, dorsal root ganglia, sympathoadrenal derivatives, presumptive Schwann cells and/or pigment cells. Most embryos contained labeled cells both in the neural tube and at least one neural crest derivative, and numerous clones contributed to multiple neural crest derivatives. The time of injection influenced the derivatives populated by the labeled cells. Injections at early stages of migration yielded labeled progeny in both dorsal and ventral neural crest derivatives, whereas those performed at later stages had labeled cells only in more dorsal neural crest derivatives, such as dorsal root ganglion and presumptive pigment cells. The results suggest that in the mouse embryo: (1) there is a common precursor for neural crest and neural tube cells; (2) some neural crest cells are multipotent; and (3) the timing of emigration influences the range of possible neural crest derivatives.

Induction of melanocyte precursors from neural crest cells surrounding the neural tube-like structures developed in vitro using mouse ES cell culture

2007 ◽  
Vol 27 (1) ◽  
pp. 45-52
Author(s):  
Koh-ichi Atoh ◽  
Manae S. Kurokawa ◽  
Hideshi Yoshikawa ◽  
Chieko Masuda ◽  
Erika Takada ◽  
...  
Keyword(s):  
Cell Culture ◽  
Neural Crest ◽  
Neural Tube ◽  
Neural Crest Cells ◽  
Es Cell ◽  
Mouse Es Cell

Cellular fibronectin promotes adrenergic differentiation of quail neural crest cells in vitro

1981 ◽  
Vol 133 (2) ◽  
pp. 285-295 ◽  
Author(s):  
Maya Sieber-Blum ◽  
Fritz Sieber ◽  
Kenneth M. Yamada
Keyword(s):  
Neural Crest ◽  
Neural Crest Cells ◽  
Cellular Fibronectin

The neural crest population responding to endothelin-3 in vitro includes multipotent cells

1997 ◽  
Vol 110 (14) ◽  
pp. 1673-1682 ◽  
Author(s):  
J.G. Stone ◽  
L.I. Spirling ◽  
M.K. Richardson
Keyword(s):  
Neural Crest ◽  
Schwann Cells ◽  
Cell Types ◽  
Developmental Potential ◽  
Colony Assay ◽  
Cellular Targets ◽  
Multipotent Cells ◽  
Endothelin 3

The peptide endothelin 3 (EDN3) is essential for normal neural crest development in vivo, and is a potent mitogen for quail truncal crest cells in vitro. It is not known which subpopulations of crest cells are targets for this response, although it has been suggested that EDN3 is selective for melanoblasts. In the absence of cell markers for different precursor types in the quail crest, we have characterised EDN3-responsive cell types using in vitro colony assay and clonal analysis. Colonies were analysed for the presence of Schwann cells, melanocytes, adrenergic cells or sensory-like cells. We provide for the first time a description of the temporal pattern of lineage segregation in neural crest cultures. In the absence of exogenous EDN3, crest cells proliferate and then differentiate. Colony assay indicates that in these differentiated cultures few undifferentiated precursors remain and there is a low replating efficiency. By contrast, in the presence of 100 ng/ml EDN3 differentiation is inhibited and most of the cells maintain the ability to give rise to mixed colonies and clones containing neural crest derivatives. A high replating efficiency is maintained. In secondary culture there was a progressive decline in the number of cell types per colony in control medium. This loss of developmental potential was not seen when exogenous EDN3 was present. Cell type analysis suggests two novel cellular targets for EDN3 under these conditions. Contrary to expectations, one is a multipotent precursor whose descendants include melanocytes, adrenergic cells and sensory-like cells; the other can give rise to melanocytes and Schwann cells. Our data do not support previous claims that the action of EDN3 in neural crest culture is selective for cells in the melanocyte lineage.

Developmental potential of neural crest-derived cells migrating from segments of developing quail bowel back-grafted into younger chick host embryos

Development ◽  
1990 ◽  
Vol 109 (2) ◽  
pp. 411-423 ◽  
Author(s):  
T.P. Rothman ◽  
N.M. Le Douarin ◽  
J.C. Fontaine-Perus ◽  
M.D. Gershon
Keyword(s):  
Neural Crest ◽  
Neural Tube ◽  
Peripheral Nerves ◽  
Sympathetic Ganglia ◽  
Limb Bud ◽  
Pigment Cells ◽  
Developmental Potential ◽  
Migratory Experience ◽  
Host Embryo

The technique of back-transplantation was used to investigate the developmental potential of neural crest-derived cells that have migrated to and colonized the avian bowel. Segments of quail bowel (removed at E4) were grafted between the somites and neural tube of younger (E2) chick host embryos. Grafts were placed at a truncal level, adjacent to somites 14–24. Initial experiments, done in vitro, confirmed that crest-derived cells are capable of migrating out of segments of foregut explanted at E4. The foregut, which at E4 has been colonized by cells derived from the vagal crest, served as the donor tissue. Comparative observations were made following grafts of control tissues, which included hindgut, lung primordia, mesonephros and limb bud. Additional experiments were done with chimeric bowel in which only the crest-derived cells were of quail origin. Targets in the host embryos colonized by crest-derived cells from the foregut grafts included the neural tube, spinal roots and ganglia, peripheral nerves, sympathetic ganglia and the adrenals, but not the gut. Donor cells in these target organs were immunostained by the monoclonal antibody, NC-1, indicating that they were crest-derived and developing along neural or glial lineages. Some of the crest-derived cells (NC-1-immunoreactive) that left the bowel and reached sympathetic ganglia, but not peripheral nerves or dorsal root ganglia, co-expressed tyrosine hydroxylase immunoreactivity, a neural characteristic never expressed by crest-derived cells in the avian gut. None of the cells leaving enteric back-grafts produced pigment. Cells of mesodermal origin were also found to leave donor explants and aggregate in dermis and feather germs near the grafts. These observations indicate that crest-derived cells, having previously migrated to the bowel, retain the ability to migrate to distant sites in a younger embryo. The routes taken by these cells appear to reflect, not their previous migratory experience, but the level of the host embryo into which the graft is placed. Some of the population of crest-derived cells that leave the back-transplanted gut remain capable of expressing phenotypes that they do not express within the bowel in situ, but which are appropriate for the site in the host embryo to which they migrate.

scholarly journals Lineage-specific requirements of β-catenin in neural crest development

2002 ◽  
Vol 159 (5) ◽  
pp. 867-880 ◽  
Author(s):  
Lisette Hari ◽  
Véronique Brault ◽  
Maurice Kléber ◽  
Hye-Youn Lee ◽  
Fabian Ille ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Neural Crest ◽  
Neural Crest Cells ◽  
Neural Crest Stem Cells ◽  
Downstream Signaling ◽  
Neural Crest Development ◽  
Sensory Neurogenesis

β-Catenin plays a pivotal role in cadherin-mediated cell adhesion. Moreover, it is a downstream signaling component of Wnt that controls multiple developmental processes such as cell proliferation, apoptosis, and fate decisions. To study the role of β-catenin in neural crest development, we used the Cre/loxP system to ablate β-catenin specifically in neural crest stem cells. Although several neural crest–derived structures develop normally, mutant animals lack melanocytes and dorsal root ganglia (DRG). In vivo and in vitro analyses revealed that mutant neural crest cells emigrate but fail to generate an early wave of sensory neurogenesis that is normally marked by the transcription factor neurogenin (ngn) 2. This indicates a role of β-catenin in premigratory or early migratory neural crest and points to heterogeneity of neural crest cells at the earliest stages of crest development. In addition, migratory neural crest cells lateral to the neural tube do not aggregate to form DRG and are unable to produce a later wave of sensory neurogenesis usually marked by the transcription factor ngn1. We propose that the requirement of β-catenin for the specification of melanocytes and sensory neuronal lineages reflects roles of β-catenin both in Wnt signaling and in mediating cell–cell interactions.

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