Development Of Pituitary Gland Research Articles

“Hot Spot” in the PROP1 Gene Responsible for Combined Pituitary Hormone Deficiency1

As pituitary function depends on the integrity of the hypothalamic-pituitary axis, any defect in the development and organogenesis of this gland may account for a form of combined pituitary hormone deficiency (CPHD). Although pit-1 was 1 of the first factors identified as a cause of CPHD in mice, many other homeodomain and transcription factors have been characterized as being involved in different developmental stages of pituitary gland development, such as prophet of pit-1 (prop-1), P-Lim, ETS-1, and Brn 4. The aims of the present study were first to screen families and patients suffering from different forms of CPHD for PROP1 gene alterations, and second to define possible hot spots and the frequency of the different gene alterations found. Of 73 subjects (36 families) analyzed, we found 35 patients, belonging to 18 unrelated families, with CPHD caused by a PROP1 gene defect. The PROP1 gene alterations included 3 missense mutations, 2 frameshift mutations, and 1 splice site mutation. The 2 reported frameshift mutations could be caused by any 2-bp GA or AG deletion at either the 148-GGA-GGG-153 or 295-CGA-GAG-AGT-303 position. As any combination of a GA or AG deletion yields the same sequencing data, the frameshift mutations were called 149delGA and 296delGA, respectively. All but 1 mutation were located in the PROP1 gene encoding the homeodomain. Importantly, 3 tandem repeats of the dinucleotides GA at location 296-302 in the PROP1 gene represent a hot spot for CPHD. In conclusion, the PROP1 gene seems to be a major candidate gene for CPHD; however, further studies are needed to evaluate other genetic defects involved in pituitary development.

Fetal development of the pituitary gland in the beagle

Fetal development of the pituitary gland in the beagle

Three-dimensional analysis of the developing pituitary gland in the mouse.

Prenatal development of the mouse pituitary gland was analyzed in three-dimensions by using confocal laser-scanning microscopy. In any wholemount preparation of the fetal pituitary gland from day 10 to day 18, immunofluorescence of laminin was observed in the deeper regions of the organ and demarcated the boundary of the epithelial tissue from the surrounding mesenchyme. Three-dimensional capillary networks in the fetal pituitary gland at day 13, day 15, and day 18 were visualized clearly by perfusing the blood vessels with an fluorescein isothiocyanate-labelled gelatin solution. During days 12-14, cellular plates protruded from the anterior wall of the pituitary anlage and extended toward the base of the infundibulum. At the same time, the mesenchyme situated among the cellular plates separated from the surrounding mesenchyme except at the anteroventral portion of the anlage. In a posterior wall of the anlage, intercellular depositions of laminin were found among the epithelial cells, some of which reached the laminin sheet in the basement membrane of the anlage. From days 15-18, the pituitary gland enlarged, and the mesenchyme spread radially from the median anteroventral portion to the lateral, posterior, and dorsal regions and finally expanded throughout the entire organ. At the periphery, the mesenchyme in the pituitary gland connected with that surrounding the capsule. Laminin was still found in the intercellular spaces between the epithelial cells, and most of the intercellular depositions of laminin were combined with the laminin sheet in the epithelial basement membrane. An abundance of vessels entered the organ from its anteroventral portion, spread, and connected with one another throughout the gland, similar to the distribution of mesenchyme. Intercellular laminin would induce the polarization of the contacting epithelial cells, resulting in rearrangement of the epithelial cells. The mesenchyme might spread in the pituitary gland, providing the intercellular laminin a foothold, in order to cleave the epithelial cell mass into lobules. The development of capillary nets in the adenohypophysis corresponded well with that of the mesenchyme. Because laminin localizes mainly in the basement membrane in the fetal organs, immunostaining of laminin clearly indicates the boundary between epithelial tissue and the mesenchyme. Three-dimensional observation of laminin in wholemount preparation is very useful for studying morphogenesis.

Morphogenesis of the hypothalamus and hypophysis: their association, dissociation and reassociation before and after "Rathke".

Recent progress in the experimental morphology of the development of amphibian pituitary gland is reviewed. A series of transplantation experiments were carried out using wild-type embryos of the toad as a donor and albino embryos as a recipient. Melanin granules in the wild-type cells allowed tracing of the developmental fate of the grafts as a visible cell marker. These studies have demonstrated that the pituitary gland is not a stomodeal derivative, as it has long been believed to be under the name of "Rathke's pouch". The adenohypophysis is of neural origin. The anterior part of the neural ridge (ANR) in the neuroectoderm of the open neurula gives rise to the whole adenohypophysis, i. e., pars distatis, pars intermedia and pars tuberalis. The presumptive hypothalamus is apposed caudally to the pituitary primordium. A part of the ANR contributes neurons to the preoptic hypothalamus even after closure of the neural tube. The anlagen of the olfactory system, which include the nasal epithelia and the olfactory bulbs, are situated on both sides of the pituitary primordium in the neural ridge. In both hypothalamic-hypophyseal and olfactory systems, the peripheral and central parts derive from closely affiliated cell populations, suggesting their clonal relationships. Development of the hypophysis and hypothalamus is interdependent. On one hand, a connection with the embryonic hypothalamus is essential for the pituitary proopiomelanocortin cells to develop. On the other hand, neither the hypothalamic median eminence nor its axonal supply develops without the presence of the pituitary primordium. Novel aspects of the ontogeny and phylogeny of these organs are discussed with special reference to the role of the neural ridge in the generation of a spectrum of chemoreceptive organs.

Role of estrogen receptor-alpha in the anterior pituitary gland.

Targeted insertional disruption of the mouse estrogen receptor-alpha (ER alpha) gene has provided a genetic model in which to test hypotheses that estrogens exert important effects in development and homeostatic functions of the anterior pituitary gland, particularly in the lactotroph and gonadotroph cell types. Analysis of ER alpha gene-disrupted mice reveals a marked reduction in PRL mRNA and a decrease in lactotroph cell number, but normal specification of lactotroph cell phenotype. Gonadotropin mRNA levels in ER alpha gene-disrupted female mice are elevated, consistent with previously described transcriptional suppression of gonadotropin subunit gene expression in response to sustained administration of estrogen in wild type mice. These results provide genetic evidence that ER alpha plays a critical role in PRL and gonadotropin gene transcription and is involved in lactotroph cell growth, but is not required for specification of lactotroph cell phenotype.

Pituitary homeobox 2, a novel member of the bicoid-related family of homeobox genes, is a potential regulator of anterior structure formation.

Genetic analysis of mouse mutants has demonstrated the importance of the homeobox genes Rpx, Lhx3 and Pit1 for anterior pituitary gland development. Pit1 mutations have also been identified in several human families with multiple pituitary hormone deficiencies. To identify additional homeobox regulators of pituitary development, we screened an adult pituitary gland cDNA library for homeobox sequences. Here, we report the identification of a novel bicoid-related homeodomain gene expressing two alternatively spliced mRNA products, which encode proteins of 271 and 317 amino acids, respectively. The proteins have been named Ptx2a and Ptx2b since they are highly related to Ptx1/P-OTX. Ptx2 is expressed in both developing and adult pituitary gland, eye and brain tissues, suggesting an important role in development and maintenance of anterior structures. Ptx2 was mapped close to Egf on mouse chromosome 3, in a region having extensive synteny homology with HSA 4q. These data make the human Ptx2 homologue a candidate gene for Rieger syndrome, an autosomal-dominant disorder with variable craniofacial, dental, eye and pituitary anomalies.

Ontogeny of somatolactin mRNA in the gilthead sea bream Sparus aurata

Expression of somatolactin (SL) gene during embryonal and early stages of larval development of the teleost Sparus aurata was determined by Northern blot analysis. Poly(A+)RNA was prepared from a pool of embryos collected at early and late stages or from larvae collected on different days after hatching. When hybridized to Sparus aurata SL cDNA, SL specific mRNA was seen both in embryos and in larvae. Levels of SL increased from day 1 onwards and reached the highest levels on day 21. Re-hybridization with Sparus aurata growth hormone (GH) cDNA revealed GH specific mRNA first on day 6 post-hatching. Levels of GH increased to maximal levels on day 10 and then decreased on days 15 and 21, thereby confirming the pattern of GH expression (Funkenstein and Cohen 1996). The patterns of SL and GH gene expression are different in gilthead sea bream during embryonal and larval development. Furthermore, the presence of SL transcript in embryos, prior to pituitary gland development, suggests the possibility that SL mRNA of maternal origin may be present in the oocyte.

The hypothalamic-pituitary axis; co-development of two organs

Development of the anterior pituitary gland ultimately leads to the appearance of five distinct cell types that are defined by the trophic hormones which they produce, providing an instructive model system for elucidating the molecular mechanisms that underlie the determination of distinct cell phenotypes within an organ from a common precursor lineage. The recent identification of several homeodomain transcription factors expressed specifically in the anterior pituitary gland has revealed a transcriptional cascade orchestrating a developmental program that leads to the determination of the five mature cell types. Recent data from gene-targeting experiments in mice further imply that the execution of this program is dependent on inductive signals originating in the floor of the diencephalon.

ROLE OF PIT-1 IN THE GENE EXPRESSION OF GROWTH HORMONE, PROLACTIN, AND THYROTROPIN

To date, nine different mutations in the Pit-1 gene resulting in CPHD have been described in mammals. Four of these mutations alter residues important for DNA binding or alter the predicted alpha helical nature of the Pit-1 protein (A158P, R172X, E250X, and W261C). The A158P mutation, however, has minimal effects on DNA binding. Four mutations lie outside alpha helical regions (P24L, R143Q, K216E, and R271W) and do not significantly alter DNA binding either experimentally or by prediction. One mutation is a large deletion of the Pit-1 gene locus in the Jackson dwarf mouse. Mutant Pit-1 proteins that do not interfere with binding cause CPHD through interference with target gene activation and regulation. The R271W mutant acts as a dominant inhibitor of transcription of the GH and Prl genes. The A158P mutant is incapable of activating transcription from the GH-I site and has low activation of transcription of the distal enhancer and proximal promoter sites of Prl and of 320 bp of the 5' GH promoter sequence. Some mutant proteins interfere with nuclear receptors. For example, the K216E mutant has defective retinoic acid signaling on the Pit-1 gene enhancer. There is phenotypic variability in the degree of CPHD and in pituitary size in patients with Pit-1 gene mutations. Since Pit-1 has different functions in the somatotroph, lactotroph, and thyrotroph, it is not surprising that point mutations in different regions of the gene interfere in different ways with Pit-1 function. A mutant Pit-1 may be able to carry out its developmental role, but may be aberrant in GH and Prl gene activation or Pit-1 autoregulation. Study of Pit-1 mutations and their diverse pathophysiologic mechanisms should increase the understanding of anterior pituitary gland development and gene regulation in normal and disease states.

P-OTX: a PIT-1-interacting homeodomain factor expressed during anterior pituitary gland development.

A novel OTX-related homeodomain transcription factor has been identified on the basis of its ability to interact with the transactivation domain of the pituitary-specific POU domain protein, Pit-1. This factor, referred to as P-OTX (pituitary OTX-related factor), is expressed in primordial Rathke's pouch, oral epithelium, first bronchial arch, duodenum, and hindlimb. In the developing anterior pituitary, it is expressed in all regions from which cells with distinct phenotypes will emerge in the mature gland. P-OTX is able to independently activate and to synergize with Pit-1 on pituitary-specific target gene promoters. Therefore, P-OTX may subserve functions in generating both precursor and specific cell phenotypes in the anterior pituitary gland and in several other organs.

Pit-1 determines cell types during development of the anterior pituitary gland. A model for transcriptional regulation of cell phenotypes in mammalian organogenesis.

Pit-1 determines cell types during development of the anterior pituitary gland. A model for transcriptional regulation of cell phenotypes in mammalian organogenesis.

Transcriptional mechanisms in anterior pituitary cell differentiation

Development of the anterior pituitary gland involves the establishment of five distinct cell lineages which are each characterized by the expression of specific trophic hormone genes. Recent studies of the thyrotrope, somatotrope, and lactotrope cell types have investigated the molecular decisions responsible for the commitment and differentiation of these cell types and have characterized the regulatory mechanisms that govern cell-specific expression of individual hormone genes. In particular, elucidation of the molecular basis of heritable dwarf phenotypes lacking particular pituitary cell lineages, such as the Snell, Jackson, and little dwarf mice, and studies of the regulation of rans-acting factors, including Pit-1, involved in pituitary cell restricted gene activation have begun to delineate the pathways responsible for development of this organ.

In situ hybridization analysis of anterior pituitary hormone gene expression during fetal mouse development.

We used 35S-labeled oligonucleotides and cRNAs (riboprobes) to detect the temporal order and spatial pattern of anterior pituitary hormone gene expression in (B6CBF1 x B6CBF1)F2 fetal mice from embryonic Day 9.5 (E9.5) to postnatal Day 1 (P1). Pro-opiomelanocortin (POMC) mRNA was expressed in the basal diencephalon on Day E10.5, in the ventromedial zone of the pars distalis on Day E12.5, and in the pars intermedia on Day E14.5. The common alpha-glycoprotein subunit (alpha-GSU) mRNA first appeared in the anterior wall of Rathke's pouch on Day E11.5 and extended to the pars tuberalis and ventromedial zone of the pars distalis on Day E12.5. Thyroid-stimulating hormone-beta (TSH beta) subunit mRNA was expressed initially in both the pas tuberalis and ventromedial pars distalis on Day E14.5, with an identical spatial distribution to alpha-GSU at the time. In contrast, luteinizing hormone-beta (LH beta) subunit and follicle-stimulating hormone beta (FSH beta) subunit mRNAs were detected initially only in the ventromedial pars distalis on Days E16.5 and E17.5, respectively, in an identical distribution to each other. POMC-, alpha-GSU-, TSH beta, LH beta-, and FSH beta-positive cells within the pars distalis all increased in number and autoradiographic signal with differing degrees of spatial expansion posteriorly, laterally, and dorsally up to Day P1. POMC expression was typically the most intense and extended circumferentially to include the entire lateral and dorsal surfaces of the pars distalis. The expression of both growth hormone (GH) and prolactin (PRL) started coincidentally on Day E15.5. However PRL cells localized in the ventromedial area similarly to POMC and the glycoprotein hormone subunits, whereas GH cells were found initially in a more lateral and central distribution within the lobes of the pars distalis. Somatotrophs increased dramatically in number and autoradiographic signal, extending throughout the pars distalis except for the most peripheral layer of cells on Day E17.5. Mammotrophs also increased in number but less abundantly than somatotrophs, and PRL expression remained more confined to central-medial and ventrolateral areas of the pars distalis up to Day P1. These data demonstrate distinctive patterns of expression for each of the major anterior pituitary hormone genes during development of the mouse pituitary gland and suggest that different groups of committed cells are the immediate precursors to the terminally differentiated hormone-secreting cell types.

Functional Differentiation of the Pituitary Gland during Development of the Domestic Ring Dove (Streptopelia roseogrisea)

The development and differentiation of the anterior pituitary gland was studied in the domestic ring dove (Streptopelia roseogrisea) and compared with the pattern of body growth. By combining histochemical staining techniques with immunohistochemical methods, cells producing prolactin (PRL), growth hormone (GH), thyrotropic hormone (TSH), and adrenocorticotropic hormone were identified in the pituitary of the developing ring dove. The distribution of the four cell types within the anterior pituitary was illustrated by three-dimensional reconstruction from immunohistochemically stained serial sections. The pattern, which was shown to change through embryonic and juvenile development, was described and compared with examples from other vertebrate classes.The analysis of body growth showed that the development of the nestlings could he divided into two distinct growth phases. The first seven days were characterized by a consistently high rate of body growth, whereas after Day seven the growth rate was markedly reduced. Although GH-immunoreactive cells formed the dominant cell type in the pituitary during the whole postembryonic development, there was no correlation between the rate of body growth and the abundance of GH-immunoreactive celIs in the pituitary. In contrast, changes in the abundance of PRL- and TSH-immunoreactive cells were observed simultaneously with the transition from the first to the second growth phase. The proportion of PRL-immunoreactive cells was relatively high during the first seven days, whereas TSH-immunoreactive cells became more abundant during the second growth phase. It is therefore postulated that PRL and TSH play an important role in the regulation of the growth pattern in the ring dove.

MR imaging of the pituitary gland in infants and children: changes in size, shape, and MR signal with growth and development.

It has been reported that on MR images of the brain in neonates the entire pituitary gland is hyperintense. This is quite different from the well-established observation that portion is isointense. To better understand the development of the neonatal pituitary gland, we studied the timing and magnitude of changes in the size, shape, and MR signal of the pituitary gland in early childhood and infancy. The study included 76 infants and children ages 3 days to 4 years old. In each MR study, the signal intensity of the pituitary gland on T1-weighted coronal (n = 24) and sagittal (n = 76) images was compared with the signal intensity of the pons by using region-of-interest methods. The size and shape of the pituitary glands were recorded. Statistical comparisons were made for signal intensity and size of the pituitary gland between boys and girls and among age groups. The results showed that the MR signal of the posterior lobe remained hyperintense relative to the pons for all age groups represented in our study (through 4 years). However, the signal intensity of the anterior lobe gradually diminished and by approximately age 2 months was isointense with the pons. Thus, by approximately age 2 months the MR signal intensity of the infant's pituitary gland has changed from being entirely hyperintense (as seen in neonates) to isointense in the anterior lobe and hyperintense in the posterior lobe (as seen in adults). In the first 2 months of life the pituitary gland is bulbous in shape; later it has a flatter upper surface. We found no significant changes in the length or signal intensity of the pituitary gland; no differences among sexes in size, shape, or signal intensity were demonstrated. Findings of a bulbous pituitary gland and high MR signal of the entire pituitary are normal in neonates up to 2 months of age. After that, the gland gradually flattens and the MR signal of the anterior lobe decreases until it has characteristics similar to the adult pituitary. Lack of high signal or discovery of a small pituitary gland in a neonate should alert the radiologist to the possibility of pituitary malformation or dysfunction.

Immunohistochemical distribution of simple-epithelial-type keratins and other intermediate filament proteins in the developing human pituitary gland

An immunohistochemical study of the production of the intermediate filaments [vimentin, cytokeratin, and glial filament acidic protein (GFAP)] during development of the pituitary gland was made by use of fetal and adult human pituitary tissue. Among these intermediate filament proteins in the anterior and intermediate lobes of the pituitary, cytokeratin is the first to appear, followed by GFAP and vimentin. However, only cytokeratin is seen during the period of morphogenesis of the pituitary gland, with the type-II subfamily cytokeratin 8 being the earliest to appear. Among the simple-epithelial-type cytokeratins, cytokeratins 8 and 19 were observed within the pituitary primordium during morphogenesis. Cells immunoreactive for cytokeratins 8 and 19 showed a heterogeneous three-dimensional distribution pattern in Rathke's pouch. Both cytokeratins 8 and 19 tended to be strongly positive at sites in the pituitary primordium where cells had become more loosely arranged (i.e., areas far from the diencephalon) but were only weakly positive in areas in which the epithelial cells were densely packed (i.e., areas closely associated with the diencephalon). It is concluded that, during the period of morphogenesis, Rathke's pouch has the intermediate filaments characteristic of simple epithelium and shows different immunoreactivity for simple-epithelial-type cytokeratins from place to place according to the extent of cellular differentiation.

Alternative translation initiation site usage results in two structurally distinct forms of Pit-1

Pit-1 is a pituitary-specific transcription factor that plays a critical role in the normal development of the anterior pituitary gland. Previous analyses have shown that this protein exists in the rat pituitary gland and in rat pituitary-derived cell lines as two forms of relative molecular mass 33 and 31 kDa. This aspect of Pit-1 expression has been conserved throughout the evolution from rodents to humans. Here, we determine the origin of these structurally distinct forms of Pit-1 protein and find that these arise as a consequence of the alternative usage of translation initiation sites present in Pit-1 mRNA.

Dwarf locus mutants lacking three pituitary cell types result from mutations in the POU-domain gene pit-1

Mutations at the mouse dwarf locus (dw) interrupt the normal development of the anterior pituitary gland, resulting in the loss of expression of growth hormone, prolactin and thyroid-stimulating hormone, and hypoplasia of their respective cell types. Disruptions in the gene encoding the POU-domain transcription factor, Pit-1, occur in both characterized alleles of the dwarf locus. The data indicate that Pit-1 is necessary for the specification of the phenotype of three cell types in the anterior pituitary, and directly link a transcription factor to commitment and progression events in mammalian organogenesis.

Pituitary cell phenotypes involve cell-specific Pit-1 mRNA translation and synergistic interactions with other classes of transcription factors.

Development of the anterior pituitary gland involves proliferation and differentiation of ectodermal cells in Rathke's pouch to generate five distinct cell types that are defined by the trophic hormones they produce. A detailed ontogenetic analysis of specific gene expression has revealed novel aspects of organogenesis in this model system. The expression of transcripts encoding the alpha-subunit common to three pituitary glycoprotein hormones in the single layer of somatic ectoderm on embryonic day 11 established that primordial pituitary cell commitment occurs prior to formation of a definitive Rathke's pouch. Activation of Pit-1 gene expression occurs as an organ-specific event, with Pit-1 transcripts initially detected in anterior pituitary cells on embryonic day 15. Levels of Pit-1 protein closely parallel those of Pit-1 transcripts without a significant lag. Unexpectedly, Pit-1 transcripts remain highly expressed in all five cell types of the mature pituitary gland, but the Pit-1 protein is detected in only three cell types--lactotrophs, somatotrophs, and thyrotrophs and not in gonadotrophs or corticotrophs. The presence of Pit-1 protein in thyrotrophs suggests that combinatorial actions of specific activating and restricting factors act to confine prolactin and growth hormone gene expression to lactotrophs and somatotrophs, respectively. A linkage between the initial appearance of Pit-1 protein and the surprising coactivation of prolactin and growth hormone gene expression is consistent with the model that Pit-1 is responsible for the initial transcriptional activation of both genes. The estrogen receptor, which has been reported to be activated in a stereotypic fashion subsequent to the appearance of Pit-1, appears to be capable, in part, of mediating the progressive increase in prolactin gene expression characteristic of the mature lactotroph phenotype. This is a consequence of synergistic transcriptional effects with Pit-1, on the basis of binding of the estrogen receptor to a response element in the prolactin gene distal enhancer. These data imply that both transcriptional and post-transcriptional regulation of Pit-1 gene expression and combinatorial actions with other classes of transcription factors activated in distinct temporal patterns, are required for the mature physiological patterns of gene expression that define distinct cell types within the anterior pituitary gland.

Analysis of Proopiomelanocortin Gene Expression during Prenatal Development of the Rat Pituitary Gland

The POMC gene is transcribed in 5-10% of adult rat anterior pituitary cells and nearly all intermediate lobe cells, but the end product peptide hormones produced by these two cell populations are distinct. We have here used in situ hybridization with 3H-POMC cRNA probes to investigate the temporal and spatial distribution of POMC mRNA containing cells during development of both lobes of the adenohypophysis. These hybridization patterns were, in addition, compared to the immunocytochemical distribution of POMC-related peptides on the same or adjacent sections. In both POMC populations, POMC-mRNA detection preceded POMC-derived peptide accumulation by at least 0.5 days. POMC-mRNA was first detected in the ventral aspect of Rathke's pouch (which will give rise to the anterior lobe) at e 13, which was two days earlier than POMC transcripts were first detected in the intermediate lobe; this result provides strong evidence that factors eliciting POMC transcription in these two populations act at distinct times during development. Further, since POMC mRNA containing cells were restricted to the anterior lobe during stages when intermediate lobe-like POMC peptides have been characterized biochemically, these results confirm immunocytochemical results and demonstrate that developmentally regulated alterations in the extent of POMC posttranslational processing occur during prenatal differentiation of anterior lobe POMC cells. POMC expression in the intermediate lobe was first observed in the midcaudal region in the midsagittal plane and subsequently extended in a gradient-like pattern to the rest of the intermediate lobe. Finally, semiquantitation of in situ hybridization signals documented that increases in the relative levels of POMC mRNA/cell occur at specific ages in both anterior and intermediate lobe. POMC cell populations and demonstrated that adult POMC mRNA/cell levels are not achieved before birth in either the anterior or intermediate lobe POMC cell populations.