Neural crest ablation does not alter pulmonary vein development in the chick embryo

Cranial neural crest, which extends from the mid-diencephalon to somite five, plays an integral role in development of pharyngeal arch derivatives and supplies mesenchyme to the aortic arch arteries. Neural crest cells in pharyngeal arches three, four, and six migrate to the heart and are involved in aorticopulmonary and conotruncal septation. Ablation of the "cardiac" neural crest cells in chick embryos results in a variety of outflow tract anomalies, including persistent truncus arteriosus. Although other studies have shown the importance of the neural crest in the development of the cardiac outflow tract, the role of neural crest in venous development has not been established. This investigation evaluates the effect of cardiac neural crest ablation on the morphological development of the pulmonary vein. The presence of the pulmonary vein was confirmed initially at early stage 15 using histological sections and computer reconstructions of serially sectioned, normal embryos. India ink injections demonstrated a complete, patent pulmonary circuit at stage 18. Cardiac neural crest was ablated at stages 8-10. Operated, sham-operated, and control embryos were sacrificed at incubation day 11, and acrylic plastic casts prepared of the intravascular compartment. In experimental embryos with persistent truncus arteriosus, there were no morphological differences in the pulmonary veins, compared with shams and controls. These data indicate that the lesions of the cardiac neural crest have little morphological impact on pulmonary vein development. It is concluded that alterations in the cardiac neural crest are not involved in venous anomalies such as cor triatriatum and total or partial anomalous pulmonary venous return.     

Dual role for neural crest cells during outflow tract septation in the neural crest-deficient mutant Splotch2H

Splotch^2H ( Sp^2H ) is a well-recognized mouse model of neural crest cell (NCC) deficiency that develops a spectrum of cardiac outflow tract malformations including common arterial trunk, double outlet right ventricle, ventricular septal defects and pharyngeal arch artery patterning defects, as well as defects in other neural-crest derived organ systems. These defects have been ascribed to reduced NCC in the pharyngeal and outflow regions. Here we provide a detailed map of NCC within the pharyngeal arches and outflow tract of Sp^2H/Sp^2H embryos and fetuses, relating this to the development of the abnormal anatomy of these structures. In the majority of Sp^2H/Sp^2H embryos we show that deficiency of NCC in the pharyngeal region results in a failure to stabilize, and early loss of, posterior pharyngeal arch arteries. Furthermore, marked reduction in the NCC-derived mesenchyme in the dorsal wall of the aortic sac disrupts fusion with the distal outflow tract cushions, preventing the initiation of outflow tract septation and resulting in common arterial trunk. In around 25% of Sp^2H/Sp^2H embryos, posterior arch arteries are stabilized and fusion occurs between the dorsal wall of the aortic sac and the outflow cushions, initiating outflow tract septation; these embryos develop double outlet right ventricle. Thus, NCC are required in the pharyngeal region both for stabilization of posterior arch arteries and initiation of outflow tract septation. Loss of NCC also disrupts the distribution of second heart field cells in the pharyngeal and outflow regions. These secondary effects of NCC deficiency likely contribute to the overall outflow phenotype, suggesting that disrupted interactions between these two cell types may underlie many common outflow defects.     

Spatiotemporally separated cardiac neural crest subpopulations that target the outflow tract septum and pharyngeal arch arteries

We used lacZ-retrovirus labeling combined with neural crest ablation in chick embryos to determine whether the cardiac neural crest cells constitute one group of multipotent cells, or they emigrate from the neural tube in time-dependent groups with different fates in the developing cardiovascular system. We demonstrated that early-migrating cardiac neural crest cells (HH9-10) massively target the aorticopulmonary septum and pharyngeal arch arteries, while the late-migrating cardiac neural crest cells (HH12) are restricted to the proximal part of the pharyngeal arch arteries. These results suggest a prominent role for early-migrating cells in outflow tract septation, and a function for late-migrating cells in pharyngeal arch artery remodeling. We demonstrated in cultures of neural tube explants an intrinsic difference between the early and late populations. However, by performing heterochronic transplantations we showed that the late-migrating cardiac neural crest cells were not developmentally restricted, and could contribute to the condensed mesenchyme of the aorticopulmonary septum when transplanted to a younger environment. Our findings on the exact timing and migratory behavior of cardiac neural crest cells will help narrow the range of factors and genes that are involved in neural crest-related congenital heart diseases.     

Initial migration and distribution of the cardiac neural crest in the avian embryo: An introduction to the concept of the circumpharyngeal crest

The distribution and migration of the cardiac neural crest was studied in chick embryos from stages 11 to 17 that were immunochemically stained in whole-mount and sectioned specimens with a monoclonal antibody, HNK-1. The following results were obtained: (1) The first phase of the migration in the cardiac crest follows the dorsolateral pathway beneath the ectoderm. (2) In the first site of arrest, the cardiac crest forms a longitudinal mass of neural-crest cells, called in the present study, the circumpharyngeal crest; this mass is located dorsolateral to the dorsal edge of the pericardium (pericardial dorsal horn) where splanchic and somatic lateral mesoderm meet. (3) A distinctive strand of neural-crest cells, called the anterior tract, arises from the mid-otic level and ends in the circumpharyngeal crest. (4) By stage 16, after the degeneration of the first somite, another strand of neural-crest cells, called the posterior tract, appears dorsal to the circumpharyngeal crest. It forms an arch-like pathway along the anterior border of the second somite. (5) The seeding of the pharyngeal ectomesenchyme takes place before the formation of pharyngeal arches in the postotic area, i.e., the crest cells are seeded into the lateral body wall ventrally from the circumpharyngeal crest; and, by the ventralward regression of the pericardial dorsal horn, lateral expansion of pharyngeal pouch, and caudal regression of the pericardium, the crest cell population is pushed away by the pharyngeal pouch. Thus the pharyngeal arch ectomesenchyme is segregated. (6) By stage 14, at the occipital somite level, ventrolateral migration of the neural crest is observed within the anterior half of each somite. Some of these crest cells are continuos with the caudal portion of the circumpharyngeal crest. An early contribution to the enteric neuroblasts is apparent in this area.     

Effect of neural crest ablation on development of the heart and arch arteries in the chick

Mesenchymal derivatives of the neural crest contribute to the connective tissues and blood vessels of the pharyngeal arches, and participate in the septation of the outflow tract of the heart. The present study was designed to determine the nature and timing of alterations in the development of the heart and arch arteries subsequent to diminished neural crest contributions. The neural crest contributing to the three caudalmost pharyngeal arches was ablated bilaterally in chick embryos and compared with sham or unoper-ated controls. Heart development was studied by scanning electron microscopy. Arch artery development was studied microscopically after intravascular injection of India ink and clearing of the specimen. Neural crest ablation caused morphological changes in most hearts. Hearts in experimental animals commonly were elongate and were subject to inappropriate development of ventricular and atrial areas. A surgical effect delayed the disappearance of arch arteries one and two, and removal of neural crest produced an additional delay. Neural crest ablation caused failure of arch arteries three, four (right), and six to develop to the proper size in some animals. Survival of those whose sixth arch arteries achieved the proper size caused group measurements to reach normal values again by stage 32. Closure of arch arteries in some animals and maintenance in others produced greater variability in experimental animals than in controls. It is significant that heart morphology was altered before septation of the outflow tract normally occurs. This indicates at the least that another factor, such as altered blood flow, contributes to the abnormal development. Altered flow may result from changes in pharyngeal arch mesenchyme and arch artery endothelium.     

Development of cranial nerves in the chick embryo with special reference to the alterations of cardiac branches after ablation of the cardiac neural crest

Summary Development of cranial nerve branches in the cardiac region was observed in whole-mount specimens which were stained with a monoclonal antibody, E/C8, after the ablation of the cardiac neural crest. In early embryos, nerve trunks of IX and X were lacking or only poorly developed, while the early development of pharyngeal branch primordia was normal. In day 5 embryos, the nerve trunks of IX–X were present in all the embryos, however; extensive communication was observed between X and XII. On day 6 and later, the spiral pattern of superior cardiac branches was disturbed, as were the blood vessels. Furthermore, the distal branches of XII passed within the superficial layer of cardiac outflow mesenchyme. Vagal branches passed within the deeper layer. There was no apparent change in the development of the sinal branch. Using quail — chick chimeras, it was found that the cardiac neural crest cells formed the Schwann cells of XII, and that they were also associated with the hypobranchial muscle primordium, suggesting that the absence of the cardiac neural crest not only disturbs the development of the cardiac outflow septation, but also affects the normal morphogenesis of the hypobranchial musculature and its innervation. Embryologically, the tongue is located close to the cardiac outflow tract, which is the migration pathway of the cardiac neural crest-derived cells.     

Experimental study on the significance of abnormal cardiac looping for the development of cardiovascular anomalies in neural crest-ablated chick embryos

During normal development, ectomesenchyme from the cardiac neural crest migrates to pharyngeal arches 3, 4, 6 and the developing heart. It participates in the formation of the aorticopulmonary septum and the wall of the great arteries. Removal of the cardiac neural crest resulted in anomalies of the great arteries and in two categories of severe heart defects: (1) outflow septation defects of the persistent truncus arteriosus (PTA) type, (2) alignment defects. It has been hypothesized that PTA occurs if the number of cardiac neural crest cells is reduced below a level critical for complete formation of the aorticopulmonary septum. Alignment defects would be indirect consequences of neural crest defects, possibly caused by altered blood flow in the pharyngeal arch region. We found that these concepts were not in agreement with some experimental facts reported previously, so we considered whether there could be other mechanisms responsible for the heart defects described. To investigate whether mechanical interference with cardiac looping could possibly contribute to the pathogenesis of these anomalies, we removed the entire cardiac neural crest in chick embryos with microneedles. Postoperative development was checked during cardiac looping and after normal completion of cardiac septation. Our data suggested that abnormal cardiac looping did not contribute to the pathogenesis of the aortic arch artery anomalies and PTA. With respect to the alignment heart defects, we could not elucidate the role of looping anomalies because we did not observe such heart defects. Moreover, PTA occurred only in 28% of survivors. This finding conflicts with previous studies where extensive ablation of the cardiac neural crest has led to a high incidence of PTA (73–100% of survivors). The possible reasons for this discrepancy are discussed. It is shown that the use of different microsurgical techniques (mechanical cutting/microcautery) may be responsible for the different incidence of PTA. We speculate that microcautery hampers a normal complete repair of neural crest defects, possibly by release of abnormally high levels of growth factors.     

Alteration of early vascular development after ablation of cranial neural crest

A previous study has shown that, subsequent to ablation of cranial neural crest, heart morphology and pharyngeal arch vessels (aortic arches) are altered before septation of the outflow tract normally occurs. In the present study, we concentrated on very early development of the aortic arch apparatus in the chick (incubation days 3-5). The three-dimensional organization of the arch vessel apparatus was studied by scanning electron microscopy after intravascular injection of Mercox, and by serial sections of embryos embedded in plastic. Alterations in the arch vessel apparatus were already present by day three in embryos with neural crest ablation at stage 9-10. Bilateral symmetry frequently was lost. Arch vessels sometimes were enlarged and occupied most of the arch, with little surrounding mesenchyme. Some arch vessels were small or occluded. Mesenchyme was significantly reduced in quantity in the arches, and was not condensed and symmetrical as in controls. There was a significant increase in the proportion of direct apposition of vessel endothelium with epithelium, without the intervening mesenchyme typical of controls. The surgical manipulation used in this study leads to distinct alterations in the arches of components and relationships which are important in development. Altered blood flow likely affects the development of the heart.     

Expression of Hox 2.1 protein in restricted populations of neural crest cells and pharyngeal ectoderm.

A polyclonal antibody, alpha Hox 2.1a, was used to localize Hox 2.1 protein in presumptive neural crest cells and nodose ganglion of 8.5-10.0 day p.c. mouse embryos. The following results were obtained: (1) The nodose placode, in its epithelial state, first expresses Hox 2.1 protein at 9.0 d.p.c. By 9.5 d.p.c. presumptive migrating neuroblasts between the nodose placode and ganglion primordium also express Hox 2.1 protein. (2) At 9.5 d.p.c., presumptive crest cells lateral to the cephalic cardinal vein and within pharyngeal arches 4 and 6 are immunoreactive for alpha Hox 2.1a. In the arch 6 region, positive cells extend medially to a mesenchymal cell population on the lateral aspect of the foregut wall. (3) At 10.0 d.p.c., Hox 2.1 protein expression in putative crest cells is restricted to the arch 6 cell population. A similar staining pattern is seen using alpha Hox 2.1a with chick embryos. Comparison with the chicken embryo suggests that the Hox 2.1 positive cells in the pharyngeal arch and those on the lateral aspect of the foregut in the mouse embryo correspond to the caudalmost subpopulation of the circumpharyngeal crest (Kuratani and Kirby: Am. J. Anat. 191:215-227, 1991; Anat. Rec. 234:263-280, 1992). These results are consistent with a role for Hox 2.1 in pattern formation in the caudalmost region of the vertebrate head.     

Myocardial enlargement in defective heart development

Background: The cardiac neural crest (neural crest extending from the mid-otic placode to the caudal region of somite 3) provides ectomesenchymal cells that contribute to aortic arch development and are essential for aortico-pulmonary septation of the outflow tract. Bilateral ablation of the cardiac neural crest in the chick embryo, prior to migration, leads to aortic arch anomalies and failure of septation of the cardiac outflow tract, which produces a severe defect known as persistent truncus arteriosus (PTA). Altered hemodynamics resulting from abnormal aortic arch artery development and PTA and other unknown factors related to the absence of neural crest, are likely to alter the developmental history of the myocardium. Methods: In this study the wet and dry weights of ventricles and whole embryos, the total number of myocytes per ventricle and the myocyte density (number of myocytes per unit volume of ventricular myocardium) were compared in control (unwindowed eggs), sham-operated and cardiac neural crest ablated chick embryos at day 11 of incubation. Results: We found that the wet and dry weights of ventricles from hearts with PTA were not different from normal hearts in control and sham-operated embryos. However, the embryos with PTA weighed less than embryos with normal hearts. Thus, the ventricle to embryo weight ratios were greater in embryos with PTA compared to control and sham-operated embryos for both wet (14 and 20%, respectively) and dry (30 and 59%) weights. The data further implied that more water was present with respect to body weight in comparison with sham-operated and control embryos which indicated that the embryos with PTA were edematous. The total number of myocytes and the number of myocytes per unit volume were not different when comparing sham-operated with PTA. Further, there was no indication that the myocardium from hearts with PTA was abnormal despite the small size and edema of the embryos. Conclusions: It appears that hemodynamic stresses, resulting from the structural defects produced by neural crest ablation, are insufficient to increase heart growth, although cardiac function is depressed as evidenced by edema and failure of the embryo to thrive. © 1994 Wiley-Liss, Inc.     

Molecular determinants of neural crest migration

Normal septation of the cardiac outflow tract requires migration of neural crest cells from the posterior rhombencephalon to the branchial arches and developing conotruncal endocardial cushions. Proper migration of these cells is mediated by a variety of molecular cues. Adhesion molecules, such as integrins, are involved in the interaction of neural crest cells with the extracellular matrix, while cadherins allow neural crest cells to interact with each other during their migration. Pax3 appears to be important for proliferation of neural crest precursors, and connexin-43-mediated gap junction communication influences the rate of migration. Endothelin and its receptors are required for normal postmigratory differentiation. Platelet-derived growth factor and retinoic acid have roles in neural crest migration and differentiation as well. Finally, the similarity between the cardiovascular malformations seen in the DiGeorge and 22q11 deletion syndromes and animal models of neural crest deficiency has led to the examination of the role of genes located near or within the DiGeorge critical region in neural crest migration.     

Cardiac neural crest stem cells

Whereas the heart itself is of mesodermal origin, components of the cardiac outflow tract are formed by the neural crest, an ectodermal derivative that gives rise to the peripheral nervous system, endocrine cells, melanocytes of the skin and internal organs, and connective tissue, bone, and cartilage of the face and ventral neck, among other tissues. Cardiac neural crest cells participate in the septation of the cardiac outflow tract into aorta and pulmonary artery. The migratory cardiac neural crest consists of stem cells, fate-restricted cells, and cells that are committed to the smooth muscle cell lineage. During their migration within the posterior branchial arches, the developmental potentials of pluripotent neural crest cells become restricted. Conversely, neural crest stem cells persist at many locations, including in the cardiac outflow tract. Many aspects of neural crest cell differentiation are driven by growth factor action. Neurotrophin-3 (NT-3) and its preferred receptor, TrkC, play important roles not only in nervous system development and function, but also in cardiac development as deletion of these genes causes outflow tract malformations. In vitro clonal analysis has shown a premature commitment of cardiac neural crest stem cells in TrkC null mice and a perturbed morphology of the endothelial tube. Norepinephrine transporter (NET) function promotes the differentiation of neural crest stem cells into noradrenergic neurons. Surprisingly, many diverse nonneuronal embryonic tissues, in particular in the cardiovascular system, express NET also. It will be of interest to determine whether norepinephrine transport plays a role also in cardiovascular development.     

Immunolocalization of N-CAM in the heart of the early developing rat embryo

Background: Neural cell adhesion molecule (N-CAM) is important in the migration of neural crest cells and is expressed in the developing heart. The pattern of expression of N-CAM in the heart of early rat embryos was investigated to shed light on the potential role of N-CAM in cardiac neural crest cell migration. Methods: N-CAM expression was studied by immunohistochemistry in Sprague-Dawley rat hearts between embryonic days 11.5 and 15.5 HNK-1 immunoreactivity was also investigated for comparison with that of N-CAM. Results: A continuity of N-CAM immunoreactivity was transiently detected from the outflow tract to the recurrent nerve. N-CAM was also expressed around the sinus venosus, inferior vena cava, sinotrial septum, and coronary sinus, as well as on mesenchymal cells in the atrioventricular endocardial cushion tissues. Conclusions: The continuous N-CAM immunoreactivity from the outflow tract to the recurrent nerve appeared to represent the pathway along which cardiac neural crest cells migrate. N-CAM-immunoreactive sites around the sinus venosus may correspond to migrating neural crest cells that differentiate into nerve fibers or cardiac ganglia. Results indicate that N-CAM may play an important role in the migration, proliferation, and transformation of neural crest cells, thereby contributing to cardiac morphogenesis and to innervation around the heart and great arteries. © 1995 Wiley-Liss, Inc.     

Conotruncal anomalies in the trisomy 16 mouse: an immunohistochemical analysis with emphasis on the involvement of the neural crest

The trisomy 16 (Ts16) mouse is generally considered a model for human Down's syndrome (trisomy 21). However, many of the cardiac defects in the Ts16 mouse do not reflect the heart malformations seen in patients suffering from this chromosomal disorder. In this study we describe the conotruncal malformations in mice with trisomy 16. The development of the outflow tract was immunohistochemically studied in serially sectioned hearts from 34 normal and 26 Ts16 mouse embryos ranging from 8.5 to 14.5 embryonic days. Conotruncal malformations observed in the Ts 16 embryos included double outlet right ventricle, persistent truncus arteriosus, Tetralogy of Fallot, and right-sided aortic arch. This spectrum of malformations is remarkably similar to that seen in humans suffering from DiGeorge syndrome (DGS). As perturbation of neural crest development has been proposed in the pathogenesis of DGS we specifically focussed on the fate of neural crest derived cells during outflow tract development of the Ts16 mouse using an antibody that enabled us to trace these cells during development. Severe perturbation of the neural crest-derived cell population was observed in each trisomic specimen. The abnormalities pertained to: 1) the size of the columns of neural crest-derived cells (or prongs); 2) the spatial orientation of these prongs within the mesenchymal tissues of the outflow tract; and 3) the location in which the neural crest cells interact with the myocardium. The latter abnormality appeared to be responsible for ectopic myocardialization found in trisomic embryos. Our observations strongly suggest that abnormal neural crest cell behavior is involved in the pathogenesis of the conotruncal malformations in the Ts16 mouse.     

MMP-2 expression during early avian cardiac and neural crest morphogenesis

Matrix metalloproteinase-type 2 (MMP-2) degrades extracellular matrix, mediates cell migration and tissue remodeling, and is implicated in mediating neural crest (NC) and cardiac development. However, there is little information regarding the expression and distribution of MMP-2 during cardiogenesis and NC morphogenesis. To elucidate the role of MMP-2, we performed a comprehensive study on the temporal and spatial distribution of MMP-2 mRNA and protein during critical stages of early avian NC and cardiac development. We found that ectodermally derived NC cells did not express MMP-2 mRNA during their initial formation and early emigration but encountered MMP-2 protein in basement membranes deposited by mesodermal cells. While NC cells did not synthesize MMP-2 mRNA early in migration, MMP-2 expression was seen in NC cells within the cranial paraxial and pharyngeal arch mesenchyme at later stages but was never detected in NC-derived neural structures. This suggested NC MMP-2 expression was temporally and spatially dependent on tissue interactions or differed within the various NC subpopulations. MMP-2 was first expressed within cardiogenic splanchnic mesoderm before and during the formation of the early heart tube, at sites of active pharyngeal arch and cardiac remodeling, and during cardiac cushion cell migration. Collectively, these results support the postulate that MMP-2 has an important functional role in early cardiogenesis, NC cell and cardiac cushion migration, and remodeling of the pharyngeal arches and cardiac heart tube.     

Frizzled 2 is transiently expressed in neural crest-containing areas during development of the heart and great arteries in the mouse

Frizzled 2 acts as a 7-transmembrane receptor in the Wnt-Dishevelled signal transduction cascade. Among others, this cascade has been associated with neural crest cell proliferation and early migration during development in mammals. The genes for some components of this cascade are located in chromosomal regions that are deleted in human syndromes associated with neural crest cell defects, like DiGeorge and Velo-Cardio-Facial Syndrome. These syndromes are often accompanied by abnormalities in cardiac morphology. Furthermore, we have reported in previous studies the upregulation of the tissue polarity gene frizzled 2 in myofibroblasts during their migration into the necrotic area after myocardial infarction in the adult heart. It is known that genes that are upregulated during cardiac remodeling due to pathology often play a role during development. To investigate whether frizzled 2 can be associated with the process of cardiac morphogenesis we studied its expression in the thoracic arterial system and heart of mouse embryo's of 10, 12, 14, 16 and 18 days after conception by means of in situ hybridization. At day 10 after conception signal could be found in the pharyngeal arches and arch arteries. The outflow tract, the ascending aorta and the pulmonary trunk were positive for frizzled 2 from day 12 on. This expression decreased with time and at day 18 only some signal could be detected in the aorta and pulmonary trunk. In contrast, in coronary and pulmonary arteries no expression was observed at any time point. Minor myocardial expression was observed in the ventricular septum at days 12 and 14. Atrial expression, although considerably lower than ventricular expression, could be detected somewhat later at days 14 and 16. Our results indicate that there is transient expression of frizzled 2 in areas that are invested by neural crest cells. This expression is downregulated upon neural crest cell differentiation. The frizzled 2 expression supports a role for the Wnt-frizzled pathway in neural crest-related disorders.     

Fgfr1 regulates patterning of the pharyngeal region

Development of the pharyngeal region depends on the interaction and integration of different cell populations, including surface ectoderm, foregut endoderm, paraxial mesoderm, and neural crest. Mice homozygous for a hypomorphic allele of Fgfr1 have craniofacial defects, some of which appeared to result from a failure in the early development of the second branchial arch. A stream of neural crest cells was found to originate from the rhombomere 4 region and migrate toward the second branchial arch in the mutants. Neural crest cells mostly failed to enter the second arch, however, but accumulated in a region proximal to it. Both rescue of the hypomorphic Fgfr1 allele and inactivation of a conditional Fgfr1 allele specifically in neural crest cells indicated that Fgfr1 regulates the entry of neural crest cells into the second branchial arch non-cell-autonomously. Gene expression in the pharyngeal ectoderm overlying the developing second branchial arch was affected in the hypomorphic Fgfr1 mutants at a stage prior to neural crest entry. Our results indicate that Fgfr1 patterns the pharyngeal region to create a permissive environment for neural crest cell migration.     

小鼠胚胎细胞视黄酸结合蛋白1阳性神经嵴细胞的时空分布与功能

目的 探讨小鼠胚胎心神经嵴细胞的形成、分布模式及其在心血管系统发育过程中的作用。方法 选用抗细胞视黄酸结合蛋白1（CRABP1）、抗α-平滑肌肌动蛋白（α-SMA）、抗心肌肌球蛋白重链（MHC）、抗胰岛因子1（Isl-1）抗体,对45只胚龄8～12d小鼠胚胎连续切片进行免疫
组织化学染色。结果 胚龄8d,CRABP1在神经褶的外胚层未见阳性表达。胚龄8.5～9d,在心管与鳃弓水平,神经褶开始出现CRABP1阳性细胞,且有部分细胞从神经褶背侧分离进入邻近间充质。胚龄10d,神经管两侧间充质内的CRABP1阳性细胞迁移至鳃弓、弓动脉壁内皮周围以及流出道
心胶质内。胚龄11～12d,弓动脉内皮周围、流出道心内膜垫内CRABP1表达明显下降,但弓动脉管壁α-SMA阳性平滑肌细胞数量增加。主肺动脉隔及其分隔形成的升主动脉和肺动脉干管壁内均可见Isl-1阳性细胞,但未见CRABP1表达。结论 小鼠胚胎CRABP1阳性神经嵴细胞形成的时间窗
限定在胚龄8.5～9d。胚龄10d后,CRABP1阳性神经嵴细胞经过迁移,参与弓动脉中膜平滑肌和流出道心内膜垫的形成。CRABP1不能用于标记迁移后的神经嵴细胞。     

Suppression of the melanogenic potential of migrating neural crest‐derived cells by the branchial arches

The development of melanocytes from neural crest‐derived precursors that migrate along the dorsolateral pathway has been attributed to the selection of this route by cells that are fate‐restricted to the melanocyte lineage. Alternatively, melanocytes could arise from nonspecified cells that develop in response to signals encountered while these cells migrate, or at their final destinations. In most animals, the bowel, which is colonized by crest‐derived cells that migrate through the caudal branchial arches, contains no melanocytes; however, the enteric microenvironment does not prevent melanocytes from developing from crest‐derived precursors placed experimentally into the bowel wall. To test the hypothesis that the branchial arches remove the melanogenic potential from the crest‐derived population that colonizes the gut, the Silky fowl (in which the viscera are pigmented) was studied. Sources of crest included Silky fowl and quail vagal and truncal neural folds/tubes, which were cultured or explanted to chorioallantoic membranes alone or together with branchial arches or limb buds from Silky fowl, White Leghorn, or quail embryos. Crest and mesenchyme‐derived cells were distinguished by using the quail nuclear marker. Melanocytes developed from Silky fowl and quail crest‐derived cells. Melanocyte development from both sources was inhibited by quail and White Leghorn branchial arches (and limb buds), but melanocyte development was unaffected by branchial arch (and limb buds) from Silky fowl. These observations suggest that a factor(s) that is normally expressed in the branchial arches, and is lacking in animals with the Silky mutation, prevents cells with a melanogenic potential from colonizing the bowel. Anat Rec 268:16–26, 2002. © 2002 Wiley‐Liss, Inc.