bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential expression of muscle regulatory factor genes in normal and denervated adult rat hindlimb muscles
Authors: Voytik Sherry L. , Przyborski Maryjo , Badylak Stephen F. , Konieczny Stephen F.
Published: February 2005

Skeletal muscle represents an excellent model system in which to examine regulatory mechanisms that modulate gene expression in the mature adult organism. Individual muscle fibers can be categorized as fast‐ or slow‐twitch based upon several physiological and molecular criteria, including metabolic enzyme activity and contractile protein isoforms. Each property can be influenced by a variety of factors such as changes in motor neuron activity or alterations in hormone levels, although the molecular pathways by which environmental factors affect gene expression remain largely unknown. As a first step in identifying potential regulators of fiber‐type diversity, the expression patterns of four basic/helix‐loop‐helix muscle regulatory factors (MRFs), referred to as MyoD, myogenin, Myf‐5, and MRF4, were examined in normal adult rat muscles which differed in their phenotypic properties. As expected, all four MRFs were expressed at detectable levels in the muscles studied. However, different muscles accumulated different proportions and combinations of MRF transcripts. For example, myogenin expression was maximally detected in slow‐twitch muscles whereas MyoD transcripts were found predominantly in muscles exhibiting a fast‐twitch phenotype. Induced phenotypic changes in two fast‐twitch muscles via denervation lead to a large and rapid increase in transcript levels of all four MRFs as early as 24 hr following denervation, with myogenin transcripts approaching 150–200‐fold higher levels than innervated contralateral muscles within 7 days. These results suggest that myogenin, as well as the other three MRFs, may be involved in both the initial establishment as well as maintenance of fiber‐type diversity in the developing organism. © 1993 Wiley‐Liss, Inc.
Expression of gap junction proteins Cx26, Cx31.1, Cx37, and Cx43 in developing and mature rat epidermis
Authors: Goliger Jeffrey A. , Paul David L.
Published: February 2005

To elucidate mechanisms underlying gap junction‐mediated intercellular communication in epidermal keratinocytes, we have examined the expression of four connexin genes, Cx26, Cx31.1, Cx37, and Cx43, in fetal (embryonic day 17–18), newborn (post‐natal day 0), and mature rat skin. Northern analyses of total skin RNA showed that levels of Cx26, Cx37, and Cx43 mRNAs remained relatively constant throughtout the three developmental stages examined, whereas Cx31.1 mRNA was 15–30 times more abundant in mature skin than in fetal skin. Antibodies specifically recognizing these connexin proteins were then used in conjunction with a recently described amplification technique to immunohistochemically stain sections of paraffin embedded rat tail epidermis. We show that Cx26, Cx31.1, Cx37, and Cx43 display overlapping but distinct patterns of expression within the keratinocyte cell layers of developing and mature epidermis. © 1994 Wiley‐Liss, Inc.
Stage‐ and region‐dependent chondrogenesis and growth of chick wing‐bud mesenchyme in serum‐containing and defined tissue culture media
Authors: Paulsen Douglas F. , Chen Wei‐Dang , Pang Lei , Johnson Benita , Okello David
Published: February 2005

During development, limb‐bud mesenchymal cells carry out complex spatiotemporal patterns of growth and differentiation. Tissue and organ culture facilitate analysis of environmental influences on these cell behaviors, allowing their partial dissection into exogenous and endogenous components. Two factors that complicate such in vitro analyses are the heterogeneity of the cultured cells and imprecise knowledge of culture medium composition. Limb mesenchyme comprises a heterogenous cell population with important regional differences in cell type. Dividing the limb into subregions helps limit the cellular heterogeneity and using chemically defined, serum‐free medium allays concerns about medium composition. In the present study, mesenchyme from different regions along the anteroposterior and proximodistal axes of stage 21–22 and stage 23–24 chick wing buds was grown in high‐density microtiter cultures in chemically defined and in serum‐containing medium. Fourday cultures of the various regions were compared in terms of culture morphology and the accumulation of Alcian blue‐positive cartilage matrix and DNA. The results demonstrate stage‐ and region‐dependent differences in the in vitro growth, differentiation, and responsiveness of these cells. For example, mesenchyme from the distal anterior region of the wing bud exhibited lower intrinsic chondrogenic capacity and greater responsiveness to serum than other regions. Patterns of in vitro chondrogenesis also suggest that, at the stages examined, distal wing‐bud mesenchyme may be less homogeneous than has been believed. A case is made for the suitability of serum‐free medium for future in vitro studies of chick limb‐bud mesenchyme. The results are considered in relation to the process of limb development and regional expression of pattern‐related genes. © 1994 Wiley‐Liss, Inc.
Increased mesenchymal cell density accompanies induction of tropoelastin expression in developing elastic tissue
Authors: Lee Katherine A. , Pierce Richard A. , Mecham Robert P. , Parks William C.
Published: February 2005

We studied the differentiation of elastin‐producing fetal bovine chondrocytes to understand the regulatory processes associated with induction of elastin expression. Analysis of auricular elastic cartilage development in vivo indicated that differentiation of the prechondrogenic blastema to an elastogenic phenotype was preceded and accompanied by condensation of the mesenchymal cells. In addition, induction of elastin production was temporally and spatially linked to expression of type II collagen and proteoglycans. We assessed the influence of cell density on the induction of tropoelastin expression in pre‐elastogenic cells from developing ear buds. Tropoelastin expression was induced in prechondrogenic mesenchymal cells only if the cells were maintained at a high cellular density. In addition, high density culture upregulated tropoelastin expression in fully differentiated chondrocytes. Together these data suggest that high cell density facilitates cell:cell interactions that affect cell proliferation and influence tropoelastin expression. © 1994 Wiley‐Liss, Inc.
Barrier inhibition of a temporal neuraxial influence on early chick somitic myogenesis
Authors: Borman William H. , Yorde Donald E.
Published: February 2005

Skeletal myogenesis in the chick embryo first occurs in the somite. Somites are transient, paired mesodermal structures adjacent to the neural tube. Somites form from the segmental plate mesenchyme at approximately 90‐min intervals. We identify somitic myogenic cells by using confocal microscopy to detect the muscle specific intermediate filament protein, desmin, in whole mount chick embryo preparations. The appearance of desmin in somitic cells does not occur at a constant interval after the somite has formed. The rate of chick somitic myogenic onset, as evidenced by detection of desmin, is approximately 1.5 times faster than the rate of somitogenesis (Borman and Yorde [1994] J. Histochem. Cytochem. 42:265–272). Somitic myogenesis does not appear to be directly linked to somitogenesis but instead may be regulated by some influence external to the somite. Here we have specifically addressed the issue of whether an impermeable barrier placed between the neuraxis and the somites can prevent the onset of somitic myogenesis. When tantalum foil barriers are placed medial to the caudalmost 3–5 somites of embryos having up to 20 somites total (stage 13), the predominant result is an inhibition of myogenic cells lateral to the barrier. Conversely, when the tantalum foil is placed medial to the caudal somites of an embryo having 21 somites (stage 14) or more, desmin is detected lateral to the barrier in most cases. There is a temporal influence originating in the neuraxis which plays a role in the onset of somitic myogenesis. Although the nature of this interaction between the neuraxis and the somites is not yet clear, we have defined a precise temporal location within the developing embryo at which this tissue interaction is taking place. © 1994 Wiley‐Liss, Inc.
Inhibition of retinal growth cone activity by specific metalloproteinase inhibitors in vitro
Authors: Sheffield J. B. , Krasnopolsky V. , Dehlinger E.
Published: February 2005

The developing neural retina expresses a set of extracellular proteases including plasminogen activator and gelatinases. Since neurites of retina cells cultured on fluorescent gelatin digest the substrate in their paths, we have suggested that the proteases are used by the tips of growing fibers to allow them to migrate within the mass of the tissue in vivo. In order to obtain further information about relationships between extracellular proteases and fiber growth, we have examined the effects of the specific inhibitors HS‐LFA (HS‐Leu‐Phenylala‐Ala, enantiomeric forms 1 and 2), bathophenanthroline sulfonate (BPS), phenylmethyl sulfonyl fluoride (PMSF), and relevant controls on the activity of retinal growth cones in vitro monitored by time lapse video microscopy. Of the inhibitors tested, only the two enantiomeric forms of HS‐LFA caused a reproducible cessation of both spike extensiion and filopodial processes at the growth cone ruffling, while control media had no effect. In some cases, the growth cone swelled and exhibited small protrusions. The behavior of growth cones was in sharp distinction to that of the cytoplasm of neural cells, and membrane ruffling of flat cells, which continued in activity throughout. Growth cone activity returned after several hours in the presence of the agent. BPS was toxic at concentrations above 2.5 mM. Below that, it had no effect. L‐cysteine, PMSF, and control media had no effect. The relevance of these results to the possible role of proteases in fiber outgrowth from retinal cells is discussed. © 1994 Wiley‐Liss, Inc.
Transforming growth factor‐β and bone morphogenetic protein‐2 act by distinct mechanisms to promote chick limb cartilage differentiation in vitro
Authors: Roark Eileen F. , Greer Karen
Published: February 2005

A number of studies suggest that several members of the transforming growth factor‐β (TGF‐β) family of peptide growth factors may be involved in the regulation of cartilage differentiation. It has been previously reported that TGF‐β1 and TGF‐β2 promote the chondrogenic differentiation of chick limb mesenchymal cells in high density micromass cultures (Kulyk et al. [1989a] Dev. Biol. 135:424–430). In this study we report that chick limb mesenchymal cells express mRNA for chicken TGF‐β1, TGF‐β2, and TGF‐β3 during cartilage differentiation in vitro. In addition, the time course of their expression during cartilage differentiation is consistent with their playing a role in the initiation of this differentiation process. We also report that two members of the TGF‐β family, TGF‐β3 and bone morphogenetic protein‐2 (BMP‐2), are capable of promoting the accumulation of cartilage extracellular matrix molecules by differentiating chick limb mesenchymal cells in micromass culture. Significant differences, however, were noted between the specific effects on matrix production elicited by these two growth factors which suggest that they may be acting by distinct mechanisms to regulate cartilage matrix production. TGF‐β appears to be most effective on cells which have not yet undergone cell condensation, a critical event in early cartilage differentiation, wherease BMP‐2 is most effective after cells have condensed or differentiated. These observations suggest that TGF‐β3 and BMP‐2 may be acting in a sequential manner to regulate chick limb mesenchymal cells through the different stages of cartilage differentiation. © 1994 Wiley‐Liss, Inc.
Murine pulmonary myocardium: Developmental analysis of cardiac gene expression
Authors: Jones W. Keith , Sánchez Alejandro , Robbins Jeffrey
Published: February 2005

Long‐standing observations that cardiac muscle exists in the walls of the pulmonary and caval veins have recently been confirmed at the molecular level (Lyons et al. [1990] J. Cell Biol. 111:2427–2436; Springall et al. [1988] Thorax 43:44–52; Subramaniam et al. [1991] J. Biol. Chem. 266:24613–24620). Using ventricle‐ and atrial‐specific riboprobes, we determined that the pulmonary myocardium exhibits an atrial pattern of cardiac‐specific gene expression. Additionally, the developmental pattern of expression was studied using a riboprobe specific to the α‐cardiac myosin heavy chain (α‐MHC) gene transcript. We find that α‐MHC gene expression is first detectable in the lung between 13.9–14.3 days post‐coitum. Extension of the α‐MHC specific hybridization signal into the pulmonary venous bed progresses through the neonatal period. The data are consistent with the hypothesis that the extension of α‐MHC gene expression into the lung occurs via the migration of atrial myoblasts into the vein during atrial septation and remodeling of the sinus venosus and pulmonary venous trunk. © 1994 Wiley‐Liss, Inc.
Pattern formation in chick feather development: Distribution of β1‐integrin in normal and scaleless embryos
Authors: Song Hee Kyung , Carver Wayne E. , Sawyer Roger H.
Published: February 2005

We have examined the immunolocalization of β1‐integrin during feather development in the spino‐lumbar tract of the backskin from normal and scaleless chick embryos. β1‐integrin appears during early feather development in three distinct phases which correspond to important developmental events. The first phase (5–5½ days of incubation; Hamburger and Hamilton [H.H.] stage 27) represents the period prior to the formation of dermis. During this phase, β1‐integrin antiserum labels mesenchymal cells located in the central region of the spino‐lumbar tract where the initiation site for feather development is located. The second phase (5½–7½ days of incubation; H.H. stages 28–32) corresponds to the period during which dermis is formed. The cells that make up the dermis are readily distinguished by their lack of β1‐integrin immunostaining. The third phase (7½–10 days of incubation; H.H. stages 33–36) begins with the sudden appearance of β1‐integrin in the central and lateral regions of the dermis. The pattern of β1‐integrin immunostaining in scaleless backskin becomes different from that of normal backskin during this phase. In normal backskin the dermal condensations of feather germs are not labeled with the β1‐integrin antiserum. This produces a heterogeneous immunostaining pattern very similar to the pattern seen for Type I collagen (Mauger et al. [1982] Dev. Biol. 94:93–105). In contrast, homogeneous immunostaining is observed in the dermis of scaleless backskin. The initial time of appearance, manner of appearance, and pattern of integrin expression in the third phase suggest that β1‐integrin may be involved in the stabilization of the feather pattern.

We also observed the appearance of β1‐integrin on the epidermal basal cells during the time of feather follicle formation. The β1‐integrin antiserum reacts strongly with the baso‐lateral surfaces of normal basal cells, yet the basal surfaces of the scaleless basal cells are unstained. This lack of immunostaining along the basal surfaces of the scaleless basal cells may relate to the abnormal adhesion between the epidermis and dermis in scaleless backskin. © 1994 Wiley‐Liss, Inc.
Anterior endoderm is a specific effector of terminal cardiac myocyte differentiation of cells from the embryonic heart forming region
Authors: Sugi Yukiko , Lough John
Published: February 2005

The ability of anterior lateral plate mesoderm cells in the heart‐forming region (HFR) of stage 6 chicken embryos to respond to cardiogenic stimuli from cells in adjacent germ layers has been investigated using explants cultured under defined conditions. Two types of explantation were evaluated: those in which two germ layers were explanted in contiguity, and those in which germ layers were isolated and co‐cultured. Two parameters—contractility and expression of sarcomeric alpha‐actin—were monitored to evaluate the terminal differentiation of cardiac myocytes. Contiguously explanted anterior endoderm/mesoderm became multilayered and underwent terminal differentiation within 2 days. By contrast, although contiguous anterior ectoderm/mesoderm or posterior endoderm/mesoderm co‐explants also became multilayered, these explants did not differentiate, up to 5 days. To ascertain the cardiogenic potential of cells from different regions of the embryo, individual germ layers were isolated and co‐cultured by placing the explants in separate areas of the culture chamber. These determinations demonstrated that anterior, but not posterior, endoderm effected differentiation of anterior mesoderm. As before, mesoderm in both types of co‐culture survived and became multilayered; by contrast, mesoderm did not survive when cultured in isolation. These experiments provide evidence that anterior endoderm regulates the terminal differentiation, as opposed to growth, of presumptive cardiac myocytes in mesoderm cells from the anterior lateral plate. Finally, anterior endoderm was co‐cultured with mesoderm from the posterior half of the embryo, which does not contain an HFR. The failure of these co‐cultured explants to differentiate infers that pre‐cardiac myoblasts in stage 6 anterior mesoderm are previously specified to respond to the terminal cardiogenic effects of endoderm. © 1994 Wiley‐Liss, Inc.
Growth dynamics of pancreatic islet cell populations during fetal and neonatal development of the rat
Authors: Kaung Hue‐lee Cheng
Published: February 2005

Rats from 18 days fetal to 28 days neonatal ages were studied for the total population sizes and cell proliferation activities of insulin secreting B cells, glucagon secreting A cells, somatostain secreting D cells, and pancreatic polypeptide secreting PP cells. Cell population sizes were assessed by morphometric quantitation of immunohistochemically stained cells by a linear scanning method and cell proliferation activities were estimated by [3H]‐thymidine labeling indices of these cell populations. There was a continuous increase in population sizes for all 4 islet cell types, with the fastest increase occurring in the last 4 days of gestation. The accelerated growth of these islet cell populations during late gestation was accomplished by a high cell proliferative activity at 20–22 days of gestation and a large influx of undifferentiated epithelial cells differentiating into the specific islet cell populations during this period. There was a reduction of population growth and cell proliferation for all islet cell types during the first 3–4 days of life. Growth activities continued for all islet cell populations after the 4th postnatal day, with a renewed acceleration in growth activities for B and A cells at this time. After the 10th neonatal day, the cell proliferation and total population growth continued at slow rates for all 4 islet cell types. The contribution from undifferentiated epithelial cells into the specific islet cell populations was negligible for B and A cells but continued at a low rate for PP and D cells during the first 10 days after birth. For B and A cell populations, there was a possibility that some cell loss occurred during the first 10 days of neonatal life. These dynamic changes of the growth characteristics provide a basis for understanding the abnormal growth of the endocrine pancreas. © 1994 Wiley‐Liss, Inc.
Control of somitic expression of tenascin in Xenopus embryos by myogenic factors and Brachyury
Authors: Umbhauer M. , Riou J.‐F. , Smith J. C. , Boucaut J. C.
Published: February 2005

Tenascin is a large glycoprotein which is expressed in a restricted pattern in the extracellular matrix (ECM) of vertebrate embryos. Tenascin interferes with cell‐fibronectin interactions in vitro, and may play a role in the control of cell migration and differentiation during development. In Xenopus, tenascin immunoreactivity is first detected at the early tailbud stage in the ECM of the most anterior somite. Thereafter, it is distributed dorsally along neural crest cell migration pathways. In this paper, we report that tenascin mRNA is most abundant in dorsal mesoderm at the neurula stage and in somites at the early tailbud stage, indicating that the initial accumulation of tenascin in the ECM is due to secretion from paraxial mesoderm. To understand how tenascin expression in somitic mesoderm is controlled, we have expressed Xbra and the myogenic factors XMyoD and XMyf5 in blastula animal cap tissue. The tenascin gene is activated by all three transcription factors. Interestingly, expression of tenascin mRNA, and accumulation of the protein in the ECM, can occur without formation of muscle. Our results suggest that tenascin regionalization in early Xenopus embryos depends on tenascin RNA expression by somitic mesoderm, where it is likely to be activated by myogenic factors. © 1994 Wiley‐Liss, Inc.
Type II collagen is transiently expressed during avian cardiac valve morphogenesis
Authors: Swiderski Ruth E. , Daniels Karla J. , Jensen Karen L. , Solursh Michael
Published: February 2005

We present new evidence of the temporal and spatial expression of type II collagen in the embryonic chick heart during the very early stages of its development. In particular, we emphasize the distribution of its mRNA and protein during valve formation. Type II collagen as well as several other fibrillar collagens (types I, III, and V) are present in stage 18 endocardial cushion mesenchymal cells. At stage 23, α1(II) collagen transcripts and the cognate polypeptide co‐localize in the atrioventricular valves. As development proceeds, the relative abundance of α1(II) collagen transcripts decreases during the stages studied (stages 22 to 45; day 3.5 to day 19) as assayed by RNA blotting of extracts of whole hearts. Type II collagen protein was immunologically undetectable in stage 38 (day 12) hearts, although collagens I, III, and V persisted and localize in the valve regions, in the endothelial lining of the heart, and in the epicardium. In keeping with other observations of type II collagen expression in non‐chondrogenic regions of a variety of vertebrate embryos, the avian heart also exhibits transient type II collagen expression. © 1994 Wiley‐Liss, Inc.
Developmental regulation of M‐cadherin in the terminal differentiation of skeletal myoblasts
Authors: Pouliot Yannick , Gravel Michel , Holland Paul C.
Published: February 2005

Cadherins form a large family of membrane glycoproteins which mediate homophilic calcium‐dependent cell adhesion. They are thought to mediate the initial calcium‐dependent cell adhesion which precedes the plasma membrane fusion of skeletal myoblasts. Two cadherin subtypes are known to be expressed in mammalian skeletal myoblasts: muscle cadherin (M‐cadherin) and neural cadherin (N‐cadherin). In the present study we demonstrate that (1) the expression of M‐ and N‐cadherin is differentially regulated during myoblast differentiation in vitro, (2) the expression of M‐cadherin but not N‐cadherin is inhibited by 5‐bromo‐2′‐deoxyuridine (BUdR), an agent which selectively inhibits skeletal myoblast differentiation, and (3) fusion and differentiation‐competent rat L6 myoblasts do not express detectable levels of N‐cadherin mRNA. In vivo, M‐cadherin mRNA was detectable exclusively in skeletal muscle. M‐cadherin mRNA levels peaked during the secondary myogenic wave in rat hindlimb muscle, becoming barely detectable in 1‐week‐old and adult rats. These observations indicate that M‐cadherin is unique in two ways: It is the first cadherin to be included in the family of skeletal muscle‐specific genes, and its shows peak leels of expression in developing skeletal muscle tissue. Taken together, these results suggest that M‐cadherin plays an important role in skeletal myogenesis. © 1994 Wiley‐Liss, Inc.
Developmental regulation of fatty acid binding protein in neural tissue
Authors: Sellner Peggy
Published: February 2005

Fatty acid binding proteins (FABP) constitute a family of small, cytosolic carriers of hydrophobic ligands. These proteins are thought to be important for lipid trafficking toward specific metabolic pathways, and are potentially important for the establishment of characteristic lipid compositions of neural tissue. In the embryonic chick retina and brain, FABP resembles the heart subtype, as determined by protein characterization and immunoblot studies. In this paper, the developmental expression and cellular localization of chick retinal FABP were examined. Results of immunoblot analysis suggest that FABP is maximally expressed around embryonic day 9 (E9) and declines thereafter. In adult retinas, FABP is barely detectable on a Western blot. Immunohistochemical staining of the retina shows light labeling on day E6 and a more intense staining throughout the retina on day E9. As the retina differentiates, labeling becomes increasingly localized. By day E18 subpopulations of ganglion cells and photoreceptor inner segments are stained, as are all photoreceptor cell bodies, most of the inner nuclear layer, and the nerve fiber layer. Staining is decreased in older retinas such that in adult animals, only light staining of the photoreceptor cell bodies is visible. The decrease in relative amount of FABP in the retina after day E9 suggests a role for FABP in the early stages of retinal differentiation. Localization in the retina is consistent with this hypothesis, as label becomes more restricted to those cells undergoing maturation at a particular developmental age. Thus, in young embryos (E6–E9), FABP immunolabeling is apparent throughout the retina, and transiently localizes at different ages (E12–E15) to plexiform and nuclear layers. Near hatching (E18–E21), the photoreceptors are in the final stages of maturation, and are the principal cells immunoreactive for FABP. In the adult retina, FABP lightly labels only the photoreceptor cells bodies; thus, we conclude that chick retinal FABP is not involved in outer segment membrane homeostasis. Instead, FABP may serve to sequester fatty acids needed for the period of neurite outgrowth which occurs as the retina ends its mitotic cycles and begins the process of synaptogenesis. © 1994 Wiley‐Liss, Inc.
Demethylation in the 5′‐flanking region of mouse cellular retinoic acid binding protein‐I gene is associated with its high level of expression in mouse embryos and facilitates its induction by retinoic acid in P19 embryonal carcinoma cells
Authors: Wei Li‐Na , Lee Chih‐Hao
Published: February 2005

The mouse cellular retinoic acid binding protein‐I (CRABP‐I) gene is specifically up‐regulated by retinoic acid (RA) in P19 mouse embryonal carcinoma cells, and its expression in animals is spatially and temporally restricted to RA‐sensitive tissues during embryonic development. This study demonstrates that, in adult mouse tissues and P19 cells where the expression of CRABP‐I is detected at the basal level, the 5′‐ flanking region of the CRABP‐I gene is hypermethylated at the C residues of all the Hpa II sites. Conversely, in mouse embryos during early stages of development when the expression of CRABP‐I gene is detected at a much higher level, this region is demethylated at these Hpa II sites. In P19, enhancement on the RA‐induced up‐regulation of CRABP‐I can be observed in cells treated with 5‐azacytidine (5‐AzaC) in conjunction with RA, where partial demethylation in the 5′‐flanking region of CRABP‐I gene is observed. Nuclear run‐on experiments indicate that increased message levels of CRABP‐I in P19 cells can be accounted for, at least partially, by increases in its transcription rates. The induction of retinoic acid receptor (RAR) β by RA can also be enhanced by 5‐AzaC, but to a much lesser degree. In contrast, all the Hpa II sites in the structural gene portion, at least in the first two exons, are fully demethylated at the C residues. © 1994 Wiley‐Liss, Inc.
Abnormal development of embryonic and extraembryonic cell lineages in parthenogenetic mouse embryos
Authors: Sturm Karin S. , Flannery Margaret L. , Pedersen Roger A.
Published: February 2005

Parthenogetically activated, diploid mouse oocytes can develop to midgestation stages in utero. However, even these advanced parthenogenones appear to die because of much reduced trophoblast and yolk sac development. Previous studies have compared the general features of parthenogenetic and androgenetic development and determined the fate of uniparental cells in chimeras with normal embryos. These studies led to the concept of genomic imprinting as the cause for developmental failure when either the maternal or the paternal genome is duplicated, with the corresponding deficiency of the other. Genomic imprinting appears to arise during gametogenesis and to act through dosage effects in a set of imprinted genes, whose expression depends on their parental origin. In this study we undertook a more detailed morphological analysis of parthenogenetic development in the mouse and established a classification system to quantify the developmental extent of parthenogenones. We found that the failure of parthenogenones occurred at different times during early postimplantation development, generating a spectrum of concepti which had developed to different extents, with only a small fraction of the embryos reaching advanced somite stages. In all parthenogenones differentiation and proliferation of the trophectoderm and primitive endoderm lineages (both extraembryonic) was abnormal, and in all, even the best‐developed parthenogenones, we observed similar deficiencies in the embryonic lineages, especially the mesoderm. Common to all abnormally developed lineages was that the proportion of undifferentiated precursor cells was much reduced, while their differentiated descendants were relatively abundant. We propose, therefore, that the failure of parthenogenones to develop to term is due to abnormal regulation of differentiation and proliferation in both embryonic and extraembryonic lineages. In this hypothesis, the apparent tissue specific defects observed in parthenogenones arise as a consequence of the functional importance of certain tissues (like the trophoblast) early in development. The spectrum of parthenogenones thus appears to reflect critical events in early development, whose regulation are affected by genomic imprinting. © 1994 Wiley‐Liss, Inc.
In vitro analysis of the spatial organization of chondrogenic regions of avian mandibular mesenchyme
Authors: Langille Robert M.
Published: February 2005

The mechanism(s) which control patterning in the face remain elusive, due in large part to the absence of morphologically identifiable controlling regions such as the AER of the limb bud. In order to identify the controlling region(s) and timing of patterning in the face, an investigation was launched to determine the spatial organization of tissues within this region, beginning with the chondrogenic zones of the avian (chick and quail) mandible. The mandibles from HH stage 23/24 chick and equivalent stage quail embryos were initially bisected in three planes giving rostral or caudal, proximal or distal, and medial or lateral halves. The mesenchyme from these various regions was isolated, plated out in high density micromass cultures, and grown for 4 days. Additionally, further cultures were grown, consisting of mandibular mesenchyme subdivided into quarters along the long axis of the mandible (e.g., rostro‐proximal quarter) as well as the bisecting of medial or lateral halves (e.g., medialrostral quarter). Nodule number and area were determined by morphometric analysis for each culture as well as whole mandible controls. The demarcation between chondrogenic and non‐chondrogenic regions was dramatic. Of the bisected halves, proximal and lateral were the most chondrogenic with the lateral subdivision displaying much more cartilage than whole mandible. The nodules of the lateral cultures fused into a sheet of cartilage. In constrast mesenchyme from the medial half was virtually non‐chondrogenic. When ranked by the amount of chondrogenesis, the order was, lateral > proximal = whole = core > distal > caudal > rostral > periphery ≫ medial. Interestingly, when subdivided further an altered pattern appeared. For example, the rostromedial quarter displayed a sheet of cartilage; more than the disto‐lateral or even the proximolateral which, based on the bisected‐mandible data, should have yielded the most cartilage. One possible explanation for the variance in the cartilage produced by the quartered mandibles from the amount predicted by the bisected‐mandible data is that further isolating portions of the mesenchyme changed the ratios of other cell types in the cultures and that cell‐cell interactions affect chondrogenic differentiation as suggested for limb mesenchyme. © 1994 Wiley‐Liss, Inc.
Differential expression of laminin isoforms and α6‐β4 integrin subunits in the developing human and mouse intestine
Authors: Simon‐Assmann Patricia , Duclos Brigitte , Orian‐Rousseau Véronique , Arnold Christiane , Mathelin Carole , Engvall Eva , Kedinger Michèle
Published: February 2005

The intestinal tissue is characterized by important morphogenetic movements during development as well as by a continuous dynamic crypt to villus epithelial cell migration leading to differentiation of specialized cells. In this study, we have examined the spatio‐temporal distribution of laminin A and M chains as well as of α6 and β4 integrin subunits in adult and developing human and mouse intestine by indirect immunofluorescence. Selective expression of the constituent polypeptides of laminin isoforms (A and M chains) was demonstrated. In the mature human intestine, A and M chains were found to be complementary, the M chain being restricted to the base of crypts and the A chain lining the villus basement membrane. In the developing human intestine, M chain expression was delayed as compared to that of A chain; as soon as the M chain was visualized, it exhibited the typical localization in the crypt basement membrane. A somewhat different situation was found in the adult mouse intestine, since both M and A chains were found in the crypts. During mouse intestinal development the delayed expression of the M chain as compared to that of the A chain was also obvious. The absence of M chain expression in mutant dy mouse did not impair intestinal morphogenesis nor cell differentiation. The expression of α6 and β4 subunits was not coordinated. In both species the α6 expression preceded that of β4. Furthermore, while β4 staining in adult mouse intestine was detected at the basal surface of all cells lining the cryptvillus, that of α6 was mainly confined to the crypt cell compartment. An overall similarity of location between α6 integrin subunit and laminin A chain at the epithelial/stromal interface was noted. These data indicate that the spatial and temporal distribution of laminin variants in the developing intestine may be characteristic for each species and that interactions of laminin variants with particular receptors may be important for induction and/or maintenance of differentiated cells. © 1994 Wiley‐Liss, Inc.
αv Integrin subunit is predominantly located in nervous tissue and skeletal muscle during mouse development
Authors: Hirsch Emilio , Gullberg Donald , Balzac Fiorella , Altruda Fiorella , Silengo Lorenzo , Tarone Guido
Published: February 2005

αv integrin subunit can dimerize with different β subunits to form receptors for several matrix proteins. The function of these receptors in vivo is still largely unknown. We examined the localization of αv integrin during mouse development and showed that its distribution is dynamically regulated in the glia of the central nervous system and in skeletal muscle. Immunoreactivity in the neural tube was firstly localized at embryonic day 10.5 (E10.5) around cell bodies lining the lumen and along tiny fibers extending towards the outer margin. At E12.5 αv distribution follows the highly defined pattern of the radial glia: fascicles of immunoreactive fibres form parallel palisades, in particular along the hindbrain and the spinal cord. At E15.5, although with weaker intensity, αv was still detectable in radial glia fibres, and it codistributed with glial fibrillary acidic protein positive fascicles. After birth (P8) αv immunoreactivity in the brain and spinal cord decreased dramatically, but remained high in the radial glia of the cerebellum. In adult mice αv reactivity in the central nervous system disappeared. During myogenesis αv appears at E10.5 in myotomal cells and from E12.5 αv was evident in myoblasts and in myotubes. In the developing skeletal muscle of E15.5 embryos, immunoreactivity became more concentrated in the apical portion of the myotubes. In adult striated muscle the amount of αv subunit dramatically declined and immunostaining was no longer detectable. During development, αv was weakly evident in other sites including heart and endothelia of blood vessels, mesonephric tubula, smooth muscle of the digestive tract, and bronchia. Comparative analysis of the localization of αv, α3, and α5 integrin subunits indicated that αv has a unique and highly regulated distribution pattern. The distribution in the nervous system is consistent with a role of αv in neuron‐glia interaction during the organization of the neuronal layers in the brain cortex and in the cerebellum. Moreover, αv is likely to be involved in the myotendinous junction during embryonic life, suggesting a dual functional role of this integrin in muscle and nervous tissue. © 1994 Wiley‐Liss, Inc.
Structure and distribution of N‐cadherin in developing zebrafish embryos: Morphogenetic effects of ectopic over‐expression
Authors: Bitzur Sigal , Kam Zvi , Geiger Benjamin
Published: February 2005

N‐cadherin cDNA was cloned from a zebrafish embryonic cDNA library. Analysis of the deduced amino acid sequence of this moleoule (ZN‐eadherin) revealed a high degree of homology to N‐cadherins of other species, except that its pre‐sequence is considerably shorter. Nevertheless, following transfection into chinese hamster ovary (CHO) cells, the expressed protein was functionally active, namely participated in calcium‐dependent intercellular interactions. Moreover, ectopic over‐expression of ZN‐cad‐herin, following mRNA microinjection into 2–4 cell embryos, caused microaggregation and uneven segregation of deep cells, resulting in distorted embryos. Developmental Northern and Western blot analyses indicated that both the mRNA and the protein first appear at gastrulation. In‐situ hybridization showed that ZN‐cadherin mRNA was initially present in all deep cells, and later became restricted to various epithelial and neural tissues. Whole‐mount immunostaining indicated that while ZN‐cadherin was already present at 50% epiboly, it became associated with cell junctions only 4–5 h later. In developing somites ZN‐cadherin expression was prominent but transient. High levels of the protein were detected in epithelial somites and its expression was apparently down regulated concomitantly with the onset of myogenesis. © 1994 Wiley‐Liss, Inc.
Patterns of cell behaviour underlying somitogenesis and notochord formation in intact vertebrate embryos
Authors: Wood Andrew , Thorogood Peter
Published: February 2005

We have made a detailed analysis of cell behaviour using high resolution timelapse microscopy of the earliest cellular interactions taking place during morphogenesis of the notochord and somites in intact teleost embryos. Notochord formation is typified by active intercalation of paraxial mesenchyme cells into the lateral surfaces of the primordium. Following this recruitment phase, complete immiscibility develops between cells of the notochord and the presomitic mesenchyme. Dorso‐ventral and rostro‐caudal expansion of the notochord is characterised by translocation of cells within dorsoventral planes of section and is supported by elongation of the remaining cells and reduction in width across its latero‐medial axis. A lateral pallisading of paraxial mesenchyme against the lateral aspects of the notochord precedes overt segmentation. Intersomitic furrows form by localised de‐adhesion at small foci at the nascent intersomitic planes, which are consolidated by coalescence of such areas by de‐adhesion to produce the interface. It is not possible to predict precisely where cells would initiate de‐adhesion since there is a stochastic element to the phenomenon. Once formed, boundaries between somites are stable and provide no opportunity for mixing, except across the first formed furrow, which disintegrates at the 4‐6 somite stage. The first ten somites form at a constant rate of 2.3 somites/hr, during which time we recorded constant relative displacement of the segmental plate against the rostro‐caudally elongating notochord. Unlike teleost epiboly and gastrulation, no large‐scale movements of individual cells can be detected during elaboration of the embryonic axis. © 1994 Wiley‐Liss, Inc.
Cellular invasion and collagen type IX in the primary corneal stroma in vitro
Authors: Cai Cindy X. , Fitch John M. , Svoboda Kathy K. H. , Birk David E. , Linsenmayer Thomas F.
Published: February 2005

During different stages in the development of the avian cornea, various collagen types have been shown to participate in matrix formation and have been implicated in morphogenesis. One of these is the fibril‐associated collagen type IX. This molecule is present when the primary corneal stroma is in a compact state, but rapidly disappears just prior to stromal swelling and its invasion by mesenchymal cells. The temporospatial pattern of the disappearance of type IX collagen in the developing cornea suggests that this molecule may be involved in stabilizing the primary corneal stromal matrix by interacting either with other type IX collagen molecules or with other matrix components. To explore further whether the removal of type IX collagen is involved in stromal swelling, we have employed an in vitro culture system in which swelling of the primary stroma and mesenchymal cell invasion can be experimentally manipulated by culturing chick corneal explants on a Nuclepore filter support in the presence or absence of an associated lens. We have also examined the effect of exogenously added human recombinant tissue inhibitor of metalloproteinases (TIMP‐1) on the presence of type IX collagen and cellular invasion. When stage 25—26+ corneal explants were cultured with an associated lens, the primary stroma did not swell; immunohistochemically detectable type IX collagen was still present, and mesenchymal cell invasion failed to occur. Conversely, when the same stages of corneal explants were cultured without an associated lens, the primary stroma swelled; type IX collagen disappeared, and mesenchymal cell migration occurred. Under both conditions, however, the type II collagen of the stroma, which is known to be a component of the striated fibrils, remained clearly detectable and with time even seemed to increase in amount. This result is consistent with the proposition that type IX collagen is one factor involved in maintaining the primary stroma as a compact matrix, possibly by functioning as a bridging/stabilizing factor. When TIMP was added to cultures of corneal explants, type IX collagen remained detectable in focal regions, suggesting that one or more metalloproteinases are involved in the removal of the type IX collagen. In addition, some of these type IX‐containing regions contained mesenchymal cells, suggesting that in addition to type IX collagen other factors are likely to be involved in regulating mesenchymal cell migration. © 1994 Wiley‐Liss, Inc.
Expression of epidermal growth factor family gene members in early mouse development
Authors: Johnson Susan E. , Rothstein Jay L. , Knowles Barbara B.
Published: February 2005

Transcription of four members of the epidermal growth factor (EGF) family, EGF, TGF‐α, Amphiregulin, and Cripto, was investigated in the ovulated unfertilized egg and preimplantation embryo using cDNA libraries. EGF was present as a maternal message only, TGF‐α was present at low levels in each library, Amphiregulin was not detected, and Cripto transcripts were first detected in the blastocyst cDNA library. In situ hybridization studies of the implanting embryo revealed Cripto expression localized to the entire ectoderm and then to the rapidly growing ectoplacental cone. At gastrulation, Cripto was detected in the primitive streak and developing mesoderm. During organogenesis, Cripto localized to the developing heart. Two Cripto transcripts were detected: one is confined to the early embryo and teratocarcinoma cells, and the other, first found in the fetus, is the major form detected in adult organs. © 1994 Wiley‐Liss, Inc.
Stage‐specific expression of alkaline phosphatase during neural development in the mouse
Authors: Narisawa Sonoko , Hasegawa Hideaki , Watanabe Keiichi , Millán José Luis
Published: February 2005

The expression pattern of tissue nonspecific alkaline phosphatase (TNAP) in the developing neural tube of mouse is reported. Homogeneous AP activity in the neuroepithelium becomes prominent at E8.5. AT E9.5, distinctly AP‐positive cells appear in the brain and spinal cord area. At stages E10.5 to E12.5, AP positivity is observed between the mesencephalon and the rhombencephalon, along the entire spinal cord and cranial nerves emerging from the myelencephalon. At E13.5, strongly AP positive fibers become prominent in the pons. At E14.5, AP expression in brain tissue is considerably reduced and there is a complete absence of AP activity in the nerve cells and glial cells of adult brain. The choroid plexus remains distinctly positive for AP expression until the adult stage. Northern blot analysis and reverse‐transcriptase polymerase chain reaction amplification of RNA indicate that this AP activity results from the expression of the Akp‐2 locus. This AP expression pattern is distinct from those reported for the expression of GD3, nestin, Hox 2.3, and Wnt‐1 during brain development. We conclude that AP is a useful marker of a subpopulation of neuroectodermal cells present in the neural tube as early as E8.5, at which stages there are no other AP positive intraembryonic cells except PGCs. © 1994 Wiley‐Liss, Inc.