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Introduction: Thyroid hypofunction during early development results in anatomical alterations in the cerebellum, cerebrum, hippocampus and other brain structures. The plastic organization of the nucleus basalis of Meynert (nBM) projections to the whiskers-related somatosensory (wS1) cortex in adolescent pups with maternal thyroid hypofunction and sensory deprivation was assessed through retrograde WGA-HRP labeling., Methods: Congenital hypothyroidism induced by adding PTU (25 ppm) to the drinking water from embryonic day 16 to postnatal day (PND) 60. Pregnant rats were divided to intact and congenital hypothyroid groups. In each group, the total whiskers of pups (4 of 8) were trimmed continuously from PND 0 to PND 60., Results: Following separately WGA-HRP injections into wS1, retrogradely labeled neurons were observed in nBM. The number of labeled neurons in nBM were higher in the congenital hypothyroid and whisker deprived groups compared to their controls (P < 0.05)., Conclusion: Based on our results both congenital hypothyroidism and sensory deprivation may disturb normal development of cortical circuits in of nBM afferents to the wS1 cortex., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The development, maintenance and regeneration of epithelial appendages such as hairs or vibrissae depend on reciprocal interactions between the epidermal and the dermal components of the integument. Growth factors are among a number of signaling molecules that have been identified during these developmental events. Growth factors such as fibroblast growth factors (FGFs) bind cell surface heparan sulfate proteoglycans (HSPGs) on their heparan sulfate side chains and as such these proteoglycans act as co-receptors for FGF receptors (FGFRs) by forming a ternary signaling complex of HSPG, FGFR and FGF. The syndecans make up a family (syndecan-1-4) of transmembrane HSPGs. In the present study we examined the growth response of mouse vibrissae to HSPG-binding growth factors as a function of the presence or absence of syndecan-4 in an organ culture system. Syndecan-4 is expressed on keratinocytes that make up the inner root sheath of the vibrissa. Vibrissae from wild-type mice, but not from syndecan-4 null mice, displayed a statistically significant and dose-dependent growth response to FGF-1, FGF-2 and FGF-7. In contrast, a statistically significant growth response is seen in vibrissae from both wild-type and syndecan-4 null mice when the culture medium is supplemented with either hepatocyte growth factor (HGF) that binds to HSPG, insulin that does not bind to HSPG or 5% fetal bovine serum. The syndecan-4 dependent effect of FGF-1, -2 and -7 on the transcriptional activity of IRS expressed genes and of genes involved in cell proliferation reveals a number of different response patterns. In vivo, the vibrissae of syndecan-4 null mice are shorter and have a smaller diameter than those of wild-type mice and this phenotype may result from a suboptimal response to growth factors. Syndecan-1, which is expressed in the outer root sheath of the vibrissae shaft, does not influence the response of the vibrissae to FGF-1, -2 and -7 and the length and diameter of vibrissae of syndecan-1 null mice do not differ from those of wild-type mice.

Development of skin appendages strongly depends on epithelial-mesenchymal interactions. One of the genes involved in this process is p63, a member of the p53 family of transcription factors, essential for ectodermal development, as elucidated by the phenotype of p63 knock-out mice. Surprisingly, no information on p63 expression in tooth and hair is yet available. Here, we show p63 expression during teeth and vibrissae morphogenesis in mouse embryos and we also show a correlation with the expression patterns of the epithelial marker keratin 5 and the proliferation marker Ki67. Our results show that p63 colocalizes with both K5 and Ki67 in the epithelium of developing vibrissae, while in teeth p63 is expressed, together with K5, in the undifferentiated ectoderm (enamel organ), and in ameloblasts, a subpopulation of differentiated ectodermal cells. Moreover, p63 expression in tooth seems not to be fully colocalized with nuclear Ki67 expression.

Lymphoid Enhancer Factor 1 (Lef-1) is an important developmental transcription factor required for the inductive formation of several epithelial-derived organs including hair follicles. Inductive expression of Lef-1 mRNA is tightly regulated during embryo development, suggesting the involvement of a highly regulated promoter. In vitro analysis of the Lef-1 gene has demonstrated the existence of at least two spatially distinct promoters with multiple transcriptional start sites that are responsive to the canonical Wnt/beta-catenin pathway. Regions of the Lef-1 promoter required for inductive regulation in vivo, however, have yet to be determined. To this end, we utilized LacZ-reporter transgenic mice to define segments of the human Lef-1 promoter capable of reproducing mesenchymal- or epithelial-restricted transcriptional patterns of Lef-1 expression during hair and vibrissa follicle development. These studies have revealed that a 110 bp Wnt/beta-catenin-responsive element, contained within a minimal 2.5 kb Lef-1 promoter, plays an important role in regulating mesenchymal, and potentially epithelial, expression during follicle development in mouse embryos. This 2.5 kb Lef-1 promoter also demonstrated inductive mesenchymal expression during postnatal anagen stage hair-follicle cycling. Additionally, analysis of Lef-1 promoter expression revealed previously uncharacterized regions of endogenous Lef-1 expression seen in the sebaceous glands of vibrissa and hair follicles in transgenic lines harboring the minimal Lef-1 promoter and additional intronic sequences. In summary, these studies have begun to dissect the transcriptional diversity of the human Lef-1 promoter during the hair/vibrissa follicle and sebaceous gland formation.

Merkel cells originate from the neural crest. They are located in hairy and glabrous skin and have neuroendocrine characteristics. Together with A beta afferents, Merkel cells form a slowly adapting mechanoreceptor, the Merkel nerve ending, which transduces steady skin indentation. Neurotphin-3 (NT-3) plays important roles in neural crest cell development. We thus sought to determine whether neurotrophin signaling is essential for Merkel cell development in the whisker pad of the mouse. Our data indicate that at embryonic day 16.5 (E 16.5), NT-3 and its receptors, p75 neurotrophin receptor (p75NTR) and tyrosine kinase receptor, TrkC are not expressed at detectable levels in Merkel cells. After a perinatal switch, however, Merkel cells in whiskers of newborn mice are immunoreactive for p75NTR, TrkC and NT-3. Immunoreactivity of all three markers persists into adulthood. By contrast, innervating fibers are intensely p75NTR-immunoreactive in E16.5 whiskers, but no TrkC immunoreactivity is detected. At birth, and at 6 weeks of age, afferent fibers are intensely immunoreactive for both p75NTR and TrkC. In TrkC null whiskers, numerous Merkel cells are present at E16.5, and they are innervated. We draw three major conclusions from these observations: (i) NT-3 signaling through p75NTR or TrkC is not required for the development and prenatal survival of either a major subset or of all Merkel cells, (ii) the postnatal survival of Merkel cells is supported by autocrine or paracrine NT-3, rather than by neuron-derived NT-3, and (iii) Merkel cell-derived NT-3 is not a chemoattractant for innervating A beta fibers, but is likely to be involved in maintaining Merkel cell innervation postnatally.

Fibroblast Growth Factor (FGF) signaling is known to play an important role during cutaneous development. To elucidate the role of FGF10 during whisker formation, we examined the expression of Fgf10 in normal developing whiskers and phenotypes of Fgf10-deficient whiskers. Fgf10 is first expressed in the maxillary process, lateral and medial nasal processes, then in the mesenchymal cells underneath the future whisker placodes, and in the surrounding mesenchyme of developing whiskers. Fgf10-null whiskers exhibit a significant decrease in number and their structure is disorganized as revealed by scanning electron microscopy. Hair follicle marker genes such as Sonic hedgehog, Patched, and Patched 2 are aberrantly expressed in the mutant whiskers. Thus, FGF10 is required for proper whisker development mediated by SHH signaling in the mouse.

Beta-catenin has a dual role in the cell. At the membrane, it connects E-cadherin to the actin cytoskeleton, while in the nucleus, it controls gene expression in concert with Tcf-like transcription factors. Nuclear translocation of beta-catenin is induced by the Wnt signal transduction pathway. Control of this process is essential since elevated beta-catenin levels interfere with differentiation and development, and can initiate cancer in many tissues. An important role for beta-catenin during hair follicle related development and tumorigenesis has recently been established, though little is known of its endogenous expression during the development of these structures. Here, we have investigated the expression of beta-catenin in relation to markers for proliferation, differentiation and Wnt signaling during the development of three hair follicle related structures, i.e. whiskers, normal body hair and the preputial gland, and a hair follicle-derived tumor, the epidermal cyst. We observed nuclear accumulation of beta-catenin, the hallmark of Wnt signaling, in the upper matrix, the dermal papilla, the developing ringwulst of the whisker and in the tumor, though it was never in association with proliferation or terminal differentiation. Co-localization of nuclear beta-catenin with Tcf-3/4 was found only in the dermal papilla and the developing ringwulst of the whisker, but not in the upper matrix or in the tumor. These results further elucidate the role of the Wnt signal transduction pathway during hair follicle related development and tumorigenesis and illustrate the dynamic role of beta-catenin in signal transduction and cell-adhesion.

Versican, a proteoglycan recently implicated in hair follicle induction, has been shown to influence axon outgrowth in vitro and in vivo. We used immunohistochemistry to study the relationship between versican expression and innervation, during rat vibrissa follicle development and the adult hair cycle. During development, nerve fibres were commonly associated with areas of weak versican expression, and the path of axons appeared to be delineated by sharp boundaries of versican expression. Versican expression changed in the lower follicle dermis during the adult hair follicle cycle but remained strong around the follicle neck reflecting the constant innervation. Our observations show a correlation between versican expression and peripheral innervation indicating that versican may have a dual role in hair follicle biology.

About 80% of all cases of Fanconi anemia (FA) can be accounted for by complementation groups A and C. To understand the relationship between these groups, we analyzed the expression pattern of the mouse FA group-A gene (Fanca) during embryogenesis and compared it with the known pattern of the group-C gene (Fancc). Northern analysis of RNA from mouse embryos at embryonic days 7, 11, 15, and 17 showed a predominant 4.5 kb band in all stages. By in situ hybridization, Fanca transcripts were found in the whisker follicles, teeth, brain, retina, kidney, liver, and limbs. There was also stage-specific variation in Fanca expression, particularly within the developing whiskers and the brain. Some tissues known to express Fancc (eg, gut) failed to show Fanca expression. These observations show that (1) Fanca is under both tissue- and stage-specific regulation in several tissues; (2) the expression pattern of Fanca is consistent with the phenotype of the human disease; and (3) Fanca expression is not necessarily coupled to that of Fancc. The presence of distinct tissue targets for FA genes suggests that some of the variability in the clinical phenotype can be attributed to the complementation group assignment.

The hair follicle is a source of epithelial stem cells and site of origin for several types of skin tumors. Although it is clear that follicles arise by way of a series of inductive tissue interactions, identification of the signaling molecules driving this process remains a major challenge in skin biology. In this study we report an obligatory role for the secreted morphogen Sonic hedgehog (Shh) during hair follicle development. Hair germs comprising epidermal placodes and associated dermal condensates were detected in both control and Shh -/- embryos, but progression through subsequent stages of follicle development was blocked in mutant skin. The expression of Gli1 and Ptc1 was reduced in Shh -/- dermal condensates and they failed to evolve into hair follicle papillae, suggesting that the adjacent mesenchyme is a critical target for placode-derived Shh. Despite the profound inhibition of hair follicle morphogenesis, late-stage follicle differentiation markers were detected in Shh -/- skin grafts, as well as cultured vibrissa explants treated with cyclopamine to block Shh signaling. Our findings reveal an essential role for Shh during hair follicle morphogenesis, where it is required for normal advancement beyond the hair germ stage of development., (Copyright 1999 Academic Press.)

Retinoic acid (RA) has marked effects on mouse upper-lip skin morphogenesis, leading to the development of glomerular gland instead of hair vibrissa follicle, but does not apparently change the dorsal pelage hair developmental program. In order to test the hypothesis that an up-regulation of the beta retinoic acid nuclear receptor (RAR beta) may be implicated in the alteration of the dermal-epidermal interactions which occur during cutaneous appendage development, RA-treated and untreated skin explants, controls as well as heterotopic recombinants, were made among nasal, upper-lip, and dorsal mouse embryonic tissues. They were analyzed by in situ hybridization with RAR beta 35S-labeled probe after 48 hr of in vitro culture as well as by identification of the morphological phenotype of cutaneous appendages after 6 additional days of culture on the chick chorioallantoic membrane. The results show that only mesenchyme from the facial region can express the RAR beta gene either normally or after RA treatment, depending on its nasal or upper-lip origin. However, the RAR beta up-regulation is unrelated to hair glandular metaplasia, which depends both on a glandular bias of the upper-lip epidermis and on the weakening of hair follicle-inducing dermal properties. The latter occurs in both the upper-lip and dorsal dermis as a consequence of RA treatment.

The spatial and temporal expression of the nuclear retinoic acid receptors alpha, beta, and gamma (RAR-alpha, beta, and gamma) was compared by in situ hybridization during hair vibrissa follicle and nasal and plantar eccrine gland morphogenesis in mouse embryo. The RAR-alpha and RAR-gamma transcripts are abundant in the dermal papilla cells of the hair vibrissa when these cells elicit epidermal hair placode (12.5-d embryos) and hair follicle (13.5-d embryos) formation. Both these transcripts are also abundant in the dermal cells of the plantar foot pad at the initiation stage (17.5-d embryos) of glandular morphogenesis. In epidermal cells, the distribution of RAR-gamma transcripts increases in parallel with hair vibrissa follicle and sweat gland differentiation, and thus may be part of the epidermal response to the dermal instructions. The RAR-beta signal is barely above control level during both hair vibrissa and plantar gland morphogenesis. By contrast, during nasal gland formation (12.5- to 15.5-d embryos), the RAR-beta signal reaches a high level in mesenchymal cells, whereas the RAR-alpha-transcripts are present in both epithelial and mesenchymal cells. These results suggest a role for RAR-alpha and RAR-gamma in the epidermal-dermal interactions that lead to hair follicle and plantar gland morphogenesis, whereas the nasal gland development implies RAR-alpha and RAR-beta gene expression. This should be correlated with the expression of the RAR-beta gene that was previously shown to be linked to the RA-induced glandular metaplasia of hair vibrissa follicles.

Retinoic acid is an exception amongst teratogenic agents: indeed, it not only interferes with morphogenesis, but it also alters positional values, which leads to the formation of additional structures. The latter effect, similar to that of homeotic mutations, which have been characterized in drosophila, is the most instructive way to study morphogenetic processes. Intramniotic injection of retinoic acid into 10-day-old chick embryos causes feather formation on the scales of the anterior side of tarsometatarsus. Likewise, when the upper-lip skin of 13.5-day mouse embryos is treated in vitro with retinoic acid, there is a development of mouse vibrissa hair buds into glands. The expression of retinoic acid nuclear receptors (RARs) has been studied by in situ hybridization during normal morphogenesis of hair vibrissa follicles as well as during retinoic acid-induced glandular metaplasia. During the normal development of hair follicle, only RAR alpha and RAR gamma genes are transcribed, whereas RAR beta gene expression remains undetected by the techniques used. Retinoic acid skin treatment brings about RAR beta expression in the dermis, leading to the formation of glomerular glands. Now, hair, gland, scale or feather differentiation implies dermal-epidermal interactions which have been extensively studied. However, little is known about the language of communication used by the two tissues. Even if the inductive role of the dermis seems dominant, the competence of the epidermis must not be neglected: the latter determines in some cases the type of cutaneous appendage which is formed.(ABSTRACT TRUNCATED AT 250 WORDS)