Retinoid signalling and skeletal development.

Metabolites of vitamin A, including retinoic acid (RA), comprise a class of molecules known to be important in development and homeostasis. RA functions through a class of nuclear hormone receptors, the RA receptors (RARs), to regulate gene transcription. In the developing mammalian limb, RA affects the differentiation of many cell lineages, including those of the chondrogenic lineage. In excess, RA is a potent teratogen, causing characteristic skeletal defects in a stage- and dose-dependent manner. Genetic analysis has shown that the absence of RARs leads to severe deficiencies in cartilage formation at certain anatomical locations while promoting ectopic cartilage formation at other sites. Expression of either a dominant-negative or a weak constitutively active RAR in the developing limbs of transgenic mice adversely affects chondrogenesis leading to skeletal malformations. Together, these results show that RAR-mediated signalling plays a fundamental role in skeletogenesis. This chapter will focus on the function of RARs in regulating chondroblast differentiation and the contribution of RA signalling to appositional and longitudinal growth of the skeletal primordia.

Analysis of Nedd4 expression during skeletal development in the mouse limb.

Nedd4, a ubiquitin-protein ligase, was originally identified as being down-regulated during development of the mouse brain (Nedd denotes neural precursor cell expressed developmentally down-regulated) (Kumar, S., Tomooka, Y., Noda, M., 1992. Identification of a set of genes with developmentally down-regulated expression in the mouse brain. Biochem. Biophys. Res. Commun. 185, 1155-1161). Subtractive hybridization was used in an attempt to identify genes that are preferentially expressed early in skeletogenesis. Using this technique Nedd4 was identified multiple times. Northern blot analysis confirmed that Nedd4 is down-regulated in the forelimb and hind limb. In situ hybridization was carried out to identify regions of the limb bud expressing Nedd4. Nedd4 is expressed weakly in condensing mesenchyme, and abundantly in proliferating and prehypertrophic chondrocytes, but is undetectable in hypertrophic chondrocytes. Primary cultures, which closely mimic in vivo chondrogenesis, were also used to demonstrate the stage-specific expression of Nedd4 during early skeletal development.

Regulation of skeletal progenitor differentiation by the BMP and retinoid signaling pathways.

The generation of the paraxial skeleton requires that commitment and differentiation of skeletal progenitors is precisely coordinated during limb outgrowth. Several signaling molecules have been identified that are important in specifying the pattern of these skeletal primordia. Very little is known, however, about the mechanisms regulating the differentiation of limb mesenchyme into chondrocytes. Overexpression of RARalpha in transgenic animals interferes with chondrogenesis and leads to appendicular skeletal defects (Cash, D.E., C.B. Bock, K. Schughart, E. Linney, and T.M. Underhill. 1997. J. Cell Biol. 136:445-457). Further analysis of these animals shows that expression of the transgene in chondroprogenitors maintains a prechondrogenic phenotype and prevents chondroblast differentiation even in the presence of BMPs, which are known stimulators of cartilage formation. Moreover, an RAR antagonist accelerates chondroblast differentiation as demonstrated by the emergence of collagen type II-expressing cells much earlier than in control or BMP-treated cultures. Addition of Noggin to limb mesenchyme cultures inhibits cartilage formation and the appearance of precartilaginous condensations. In contrast, abrogation of retinoid signaling is sufficient to induce the expression of the chondroblastic phenotype in the presence of Noggin. These findings show that BMP and RAR-signaling pathways appear to operate independently to coordinate skeletal development, and that retinoid signaling can function in a BMP-independent manner to induce cartilage formation. Thus, retinoid signaling appears to play a novel and unexpected role in skeletogenesis by regulating the emergence of chondroblasts from skeletal progenitors.

Retinoids and their receptors in skeletal development.

The embryonic vertebrate limb serves as an excellent experimental model system in which to study mechanisms that regulate morphogenesis of the skeleton. The appendicular skeleton arises through the process of endochondral ossification, whereby a cartilage template is initially formed and subsequently replaced by bone. One molecule that has a dramatic effect on these processes is the vitamin-A metabolite, retinoic acid (RA). RA functions through a class of nuclear hormone receptors, the retinoic acid receptors (RARs) and retinoid-X-receptors (RXRs), to regulate gene transcription. Experimental evidence from RA teratogenesis suggests that the presence of ligand-activated RARs and/or inappropriate expression of RARs inhibits chondrogenesis. Conversely, genetic analysis has shown that the absence of the receptors can lead to deficiencies in cartilage formation while also promoting chondrogenesis at ectopic sites. Taken together, these studies suggest that the RARs play a fundamental role in the early stages of skeletal development, specifically those involved in the formation of prechondrogenic condensations and their subsequent differentiation into chondroblasts.

The influence of sentence elicitation variables on children's speech production.

This study investigated the potential influence of adult-modeled sentences on the speech production of 15 children with speech delays of unknown origin. Two comparison tokens of target words containing sounds with inconsistently realized phonemes were sampled in picture descriptions elicited with and without adult-modeled descriptive sentences. Ten listeners made forced-choice paired-comparisons to identify the children's relatively more advanced word productions. From 205 total comparisons, listeners identified 130 word pairs that included one token more advanced than the other. Significantly more of the children's advanced word productions occurred in sentences elicited with an adult model sentence. Discussion considers theoretical and clinical perspectives of an assumption that variables facilitating children's language production may benefit speech production.

Contextual and linguistic correlates of intelligibility in children with developmental phonological disorders.

Listeners' glosses of children's intended words provided data for two studies of the potential influence of selected contextual and linguistic variables on word intelligibility. Several regularities associated with the occurrence of unintelligible words were identified. In Study I, intelligibility outcomes were associated with utterance length and fluency, word position, intelligibility of adjacent words, phonological complexity, and grammatical form. In Study II, intelligibility outcomes were associated with phonological complexity, syllabic structure, and grammatical form. Discussion considers the implications of these and other regularities associated with the occurrence of unintelligible words for a comprehensive perspective on the utterance-to-utterance intelligibility deficits of children with phonological disorders of unknown origin.

Analysis of language-speech samples with salt and pepper.

Microcomputer-aided analysis of spontaneous language-speech samples offers researchers an efficient means of analyzing large amounts of data. It may be necessary, however, to format samples for more than one software program in order to obtain comprehensive morpho-syntactic and phonetic/phonologic analyses. This paper suggests a procedure for the combined use of SALT (Systematic Analysis of Language Transcripts, Miller & Chapman, 1985) and PEPPER (Programs to Examine Phonetic and Phonologic Evaluation Records, Shriberg, 1986) that is designed to minimize the duplication of effort involved in following two different formatting procedures. Results of a study undertaken to explore methodological issues in the combined use of SALT and PEPPER generally support the validity, reliability, and efficiency of the procedure. Results also raise some issues concerning the use of narrow phonetic transcription as opposed to standard orthographic transcription of continuous language-speech samples.