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Biomedical subjects

B Christ

Publications and source records attributed to B Christ.

At least 55 records · Page 3Linked to original sources
PubMed2000

Dual origin and segmental organisation of the avian scapula.

Bones of the postcranial skeleton of higher vertebrates originate from either somitic mesoderm or somatopleural layer of the lateral plate mesoderm. Controversy surrounds the origin of the scapula, a major component of the shoulder girdle, with both somitic and lateral plate origins being proposed. Abnormal scapular development has been described in the naturally occurring undulated series of mouse mutants, which has implicated Pax1 in the formation of this bone. Here we addressed the development of the scapula, firstly, by analysing the relationship between Pax1 expression and chondrogenesis and, secondly, by determining the developmental origin of the scapula using chick quail chimeric analysis. We show the following. (1) The scapula develops in a rostral-to-caudal direction and overt chondrification is preceded by an accumulation of Pax1-expressing cells. (2) The scapular head and neck are of lateral plate mesodermal origin. (3) In contrast, the scapular blade is composed of somitic cells. (4) Unlike the Pax1-positive cells of the vertebral column, which are of sclerotomal origin, the Pax1-positive cells of the scapular blade originate from the dermomyotome. (5) Finally, we show that cells of the scapular blade are organised into spatially restricted domains along its rostrocaudal axis in the same order as the somites from which they originated. Our results imply that the scapular blade is an ossifying muscular insertion rather than an original skeletal element, and that the scapular head and neck are homologous to the 'true coracoid' of higher vertebrates.

Animals↗
PubMed2000

Sclerotomal origin of the ribs.

The somites of vertebrate embryos give rise to sclerotomes and dermomyotomes. The sclerotomes form the axial skeleton, whereas the dermomyotomes give rise to all trunk muscles and the dermis of the back. The ribs were thought to be ventral processes of the axial skeleton and therefore to be derived from the sclerotomes; however, recently a dermomyotomal origin of the distal rib (the costal shaft) was suggested, with only the proximal parts (head and neck of the rib) being of sclerotomal origin. We have re-investigated the development of the ribs in quail-chick chimeras and carried out three experimental series. (1) Single dermomyotomes and (2) single sclerotomes were grafted homotopically, and (3) the ectoderm overlying the unsegmented paraxial mesoderm was removed in the prospective thoracic region. We found that the cells of the dermomyotome gave rise to epaxial and hypaxial trunk muscles, dermis of the back and endothelial cells, but not to ribs. Cells of the sclerotome formed the axial skeleton and all parts of the ribs. Ablation of the ectoderm, which affects dermomyotome development, results in severe malformations of the ribs, probably due to disturbed interactions between dermomyotome and sclerotome. Our results strongly confirm the traditional view of the sclerotomal origin of the ribs.

Animals↗
PubMed2000

Elevated expression of hormone-regulated rat hepatocyte functions in a new serum-free hepatocyte-stromal cell coculture model.

The specific performance of the adult hepatic parenchymal cell is maintained and controlled by factors deriving from the stromal bed; the chemical nature of these factors is unknown. This study aimed to develop a serum-free hierarchical hepatocyte-nonparenchymal (stromal) cell coculture system. Hepatic stromal cells proliferated on crosslinked collagen in serum-free medium with epidermal growth factor, basic fibroblast growth factor, and hepatocyte-conditioned medium; cell type composition changed during the 2-wk culture period. During the first wk, the culture consisted of proliferating sinusoidal endothelial cells with well-preserved sieve plates, proliferating hepatic stellate cells, and partially activated Kupffer cells. The number of endothelial cells declined thereafter; stellate cells and Kupffer cells became the prominent cell types after 8 d. Hepatocytes were seeded onto stromal cells precultured for 4-14 d; they adhered to stellate and Kupffer cells, but spared the islands of endothelial cells. Stellate cells spread out on top of the hepatocytes; Kupffer cell extensions established multiple contacts to hepatocytes and stellate cells. Hepatocyte viability was maintained by coculture; the positive influence of stromal cell signals on hepatocyte differentiation became evident after 48 h; a strong improvement of cell responsiveness toward hormones could be observed in cocultured hepatocytes. Hierarchial hepatocyte coculture enhanced the glucagon-dependent increases in phosphoenolpyruvate carboxykinase activity and messenger ribonucleic acid (mRNA) content three- and twofold, respectively; glucagon-activated urea production was elevated twofold. Coculturing also stimulated glycogen deposition; basal synthesis was increased by 30% and the responsiveness toward insulin and glucose was elevated by 100 and 55%, respectively. The insulin-dependent rise in the glucokinase mRNA content was increased twofold in cocultured hepatocytes. It can be concluded that long-term signals from stromal cells maintain hepatocyte differentiation. This coculture model should, therefore, provide the technical basis for the investigation of stroma-derived differentiation factors.

Animals↗
PubMed2000

An avian model for studies of embryonic lymphangiogenesis.

Embryonic development of lymphatics (lymphangiogenesis) in recent years has rarely been studied experimentally. Using an avian model, we showed that both intra- and extra-embryonic blood vessels of chick and quail embryos are accompanied by lymphatics. The lymphatics of the chorioallantoic membrane (CAM) are drained by lymphatic trunks of the umbilicus and are connected to the posterior lymph hearts. Intra-embryonic lymphatics are drained via paired thoracic ducts into the jugulo-subclavian junction. The lymphatic endothelial cells are characterized by the expression of Vascular Endothelial Growth Factor Receptors (VEGFR) -2 and -3. Application of VEGF-C, the ligand of these two receptors, on the differentiated CAM, induces proliferation of lymphatic endothelial cells and formation of huge lymphatic sinuses. These lymphatics derive from pre-existing lymphatic endothelial cells, whereas, in early embryos lymphangioblasts are present in the mesenchyme. This phenomenon can be demonstrated by interspecific grafting experiments between chick and quail embryos. Together with the early lymph sacs, the lymphangioblasts form the embryonic lymphatic system. Our studies demonstrate the importance of lymphangioblasts and lymphangiogenic growth factors in embryonic lymphangiogenesis.

Animals↗
PubMed1999

Macrowell cultures identify a subpopulation of neonatal rat dorsal root ganglionic neurons displaying nerve growth factor independent survival.

Dorsal root ganglionic (DRG) neurons of the newborn rat in vitro die by apoptosis within 24-48 h unless nerve growth factor (NGF) is added. Using a novel cell culture system (macrowell), we identified a neuronal subpopulation displaying NGF-independent survival in vitro. Neurons were grown on glass coverslips at standard cell density in different volumes of defined medium (standard: 500 microl; macrowell: 10 ml). In standard culture, 40% of neurons survived in the presence of NGF whereas there was no survival under control conditions. In macrowell culture, however, about 15% of neurons survived even in the absence of NGF. Addition of NGF to these cultures increased survival up to 65%. Neurons surviving independent of NGF in macrowell culture were heterogeneous in size and were lacking the low-affinity NGF receptor.

Animals↗
PubMed1999

Lymphangioblasts in the avian wing bud.

The development of the lymphatics has not yet been studied experimentally. Descriptive studies could not answer the question whether the lymphatics are exclusively derived by sprouts of the early embryonic lymph sacs, or whether lymphangioblasts in the mesenchyme contribute to the lymphatic system. We have studied the development of the lymphatics in quail-chick chimeras. In 6.5-day-old quail embryos, the endothelium of the jugulo-axillary lymph sac can be demonstrated with the QH1 antibody. In contrast to the jugular vein and the aorta, the lymph sac is irregularly shaped and does not possess a media of smooth muscle cells, and, the lymph sac endothelium starts to express the vascular endothelial growth factor receptor-3 (VEGFR-3). Cells of the quail paraxial mesoderm grafted into chick embryos integrate into the endothelium of the jugular lymph sac, strongly indicating the existence of lymphangioblasts. In the wing of 10-day-old quail embryos, VEGFR-3-positive lymphatics are accompanying all major blood vascular routes. On day 3.5 of development, that is about one day before the first occurrence of the jugulo-axillary lymph sac, we grafted distal wing buds of chick embryos homotopically into quail embryos. The chimeric wings were analyzed on day 10. The VEGFR-3 and QH1 double staining revealed that the lymphatics were formed by both chick and quail endothelial cells. This result shows that the lymphatics of the wing do not exclusively develop from sprouts of the lymph sacs, but also by recruitment of local lymphangioblasts.

Animals↗
PubMed1999

Origin and development of the avian tongue muscles.

The musculature of the vertebrate tongue is composed of cells recruited from the somites. In this paper we have investigated the migration and organisation of the muscle cells that give rise to the tongue muscle during chick embryogenesis. At the molecular level, our data suggests that a population of Tbx-3 expressing cells migrate away from the occipital somites prior to the migration of muscle precursors that express Pax-3. Both populations take the same pathway and form the hypoglossal cord. The first signs of muscle cell differentiation were not detected until cells had migrated some distance from the somites. We have determined the contribution of single somites to the musculature of the tongue and show in contrast to previous data that somites 2-6 take part in the formation of all glossal and infrahyoid muscles to the same extent but do not contribute to suprahyoid muscle. This is particularly interesting since glossal and infrahyoid muscle differ from the suprahyoid muscles not only in their morphology, but also in their developmental origin. Furthermore we show that myocytes cross the midline and contribute to the contralateral glossal and infrahyoid muscles. This is supported from our molecular data, which showed that the migratory precursor population was maintained primarily at the rostral tip of the developing hypoglossal cord.

Animals↗
PubMed1999

Genetic and epigenetic control of muscle development in vertebrates.

The skeletal body muscle of vertebrates is derived from segmentally arranged mesodermal structures, the somites. Only the dorsal epithelial half of the somite, the dermomyotome, gives rise to muscle cells during normal development. Head muscle takes its origin from the somites, the unsegmented paraxial head mesoderm and the prechordal mesoderm. Some muscle precursor cells, for instance those for limb and tongue muscle, migrate over considerable distances before differentiating at their target sites. In recent years, our understanding of the molecular events underlying myogenesis has increased considerably. Muscle differentiation is preceded by several steps during which precursor cells are specified. Markers of myogenic specification are myf5, myoD, mrf4 and myogenin, which encode transcription factors of the basic helix-loop-helix family. These factors bind to promoters of many muscle-specific genes and interact with MEF2 (myocyte enhancer binding factor-2) belonging to the MADS (MCM1, agamous, deficiens, serum response factor) box transcription factors. Signalling events leading to myogenic precursor cell specification and to the formation of muscle fibres are being elucidated. Inductive signals emanate from the neural tube, notochord and ectoderm. Controversial findings concerning the role of the notochord and neural tube in muscle development suggest that the epigenetic events leading to myogenesis are more complex than originally anticipated. Signals from the lateral plate counteract those from the axial organs and induce the locally restricted emigration of muscle precursor cells. Future investigations will have to show how signalling molecules and their receptors interact in the process of fine-tuning muscle formation in the embryo.

Animals↗
PubMed1999

Embryonic lymphangiogenesis.

About 8-9 decades ago the development of embryonic lymphatics was studied intensively. Since then our knowledge has not considerably increased in this field, and it is still unknown whether lymphatics are derived by sprouting from veins, de novo from lymphangioblasts, or by both mechanisms. However, very recent studies have shown that the vascular endothelial growth factor-C (VEGF-C) is a highly specific lymphangiogenic growth factor. This raises new questions and perspectives. Here we will review the literature on embryonic lymphangiogenesis and lymphangiogenic growth factors. We also present a description of the pattern of the lymphatics of avian embryos with emphasis on lymph hearts. The avian embryo is highly suited for studies on lymphatics, because these can be demonstrated by injection methods, serial sectioning and in situ hybridization with VEGF-receptor-2 and -3 probes. The greatest advantage resides in the fact that the lymphatics of the chorioallantoic membrane are easily accessible for experimental studies.

Allantois↗
PubMed1999

The monoclonal antibody 23E9 defines a novel developmentally-regulated Schwann cell surface antigen.

The present study describes the identification and partial characterization of a novel Schwann cell surface molecule by means of a monoclonal antibody (23E9). The 23E9 antigen was found in association with Schwann cells of the peripheral nerve but not with sensory neurons and satellite cells of the dorsal root ganglion. The expression of the antigen in the sciatic nerve starts after birth, is high around postnatal day 8 and becomes down-regulated towards the adult stage. This suggests that it may be involved in the induction of myelin formation. On Western blots, the antibody identified two major bands of approximately 27 and 42 kDa. Treatment of cultured Schwann cells with forskolin, an agent known to mimic neuronal contact in vitro, stimulated the up-regulation of the antigen. This implies that the expression of 23E9 is induced and maintained by axon-derived signals in vivo. Comparison of the presented data with the literature suggests that we have identified a novel cell surface molecule not previously characterized in the context of Schwann cell biology. To clarify the molecular identity of the antigen and define its physiological relevance, the antibody will be used in future studies for immunoprecipitation and functional in vitro assays.

Aging↗
PubMed1999

Cytokines and the hepatic acute-phase response.

The acute-phase reaction comprises a variety of systemic changes in response to tissue injury and infection. Synthesis of several plasma proteins in the liver undergoes dramatic changes during the acute-phase reaction. These changes are mediated by the action of cytokines, mainly interleukin 6-(IL6-) and interleukin 1-(IL1-) type cytokines. The role of these cytokines is reviewed in detail. Attention is payed to the differences recently detected between the locally and systemically induced acute-phase response--in particular, the newest findings concerning the meaning of single cytokines. Also, changes in liver metabolic enzyme expression under acute-phase conditions are discussed. The impact of cytokine and cytokine-receptor knockout or transgenic mouse models on current knowledge is provided. The role of cytokines together with that of soluble cytokine receptors in the resolution of the acute-phase response is discussed.

Acute-Phase Proteins↗
PubMed1999

SF/HGF is a mediator between limb patterning and muscle development.

Scatter factor/hepatocyte growth factor (SF/HGF) is known to be involved in the detachment of myogenic precursor cells from the lateral dermomyotomes and their subsequent migration into the newly formed limb buds. As yet, however, nothing has been known about the role of the persistent expression of SF/HGF in the limb bud mesenchyme during later stages of limb bud development. To test for a potential role of SF/HGF in early limb muscle patterning, we examined the regulation of SF/HGF expression in the limb bud as well as the influence of SF/HGF on direction control of myogenic precursor cells in limb bud mesenchyme. We demonstrate that SF/HGF expression is controlled by signals involved in limb bud patterning. In the absence of an apical ectodermal ridge (AER), no expression of SF/HGF in the limb bud is observed. However, FGF-2 application can rescue SF/HGF expression. Excision of the zone of polarizing activity (ZPA) results in ectopic and enhanced SF/HGF expression in the posterior limb bud mesenchyme. We could identify BMP-2 as a potential inhibitor of SF/HGF expression in the posterior limb bud mesenchyme. We further demonstrate that ZPA excision results in a shift of Pax-3-positive cells towards the posterior limb bud mesenchyme, indicating a role of the ZPA in positioning of the premuscle masses. Moreover, we present evidence that, in the limb bud mesenchyme, SF/HGF increases the motility of myogenic precursor cells and has a role in maintaining their undifferentiated state during migration. We present a model for a crucial role of SF/HGF during migration and early patterning of muscle precursor cells in the vertebrate limb.

Animals↗
PubMed1999

A molecular mechanism enabling continuous embryonic muscle growth - a balance between proliferation and differentiation.

Embryonic muscle growth requires a fine balance between proliferation and differentiation. In this study we have investigated how this balance is achieved during chick development. Removal of ectoderm from trunk somites results in the down-regulation of Pax-3 expression and cell division of myogenic precursors is halted. This initially leads to an up-regulation of MyoD expression and to a burst in terminal differentiation but further muscle growth is arrested. Locally applied bone morphogenetic protein-4 (BMP-4) to somites mimics the effect of the ectoderm and stimulates Pax-3 expression which eventually results in excessive muscle growth in somites. Surprisingly, BMP-4 up-regulates expression of noggin which encodes a BMP-4 antagonist. This suggests that the proliferation enhancing activity of BMP-4 can be limited via up-regulation of noggin and that myogenic cells differentiate, as an intrinsic property, when deprived of BMP-4 influence. In contrast to BMP-4, Sonic hedgehog (Shh) locally applied to somites arrests muscle growth by down-regulation of Pax-3 and immediate up-regulation of MyoD expression. Such premature muscle differentiation in somites at tongue and limb levels prevents myogenic migration and thus tongue and limb muscle are not formed. Therefore, precise limitation of differentiation, executed by proliferative and Pax-3 promoting signals, is indispensable for continuous embryonic muscle growth.

Animals↗
PubMed1998

Experimental induction of BMP-4 expression leads to apoptosis in the paraxial and lateral plate mesoderm.

In the avian embryo, epithelialization of the segmental plate and formation of an epithelial dermomyotome depend on signals from the neural tube and the ectoderm overlying the paraxial mesoderm. In this study, we report that ectoderm removal in combination with barrier insertion between the axial organs and the segmental plate leads to an induction of BMP-4 expression in the paraxial mesoderm. In the lateral plate, ectoderm removal alone leads to an increase of BMP-4 expression. Application of BMP-4 protein results in a lack of epithelialization of the paraxial mesoderm. In order to investigate whether the loss of epithelial structures after these manipulations can be attributed to a change in cell fate, a change in cell proliferation, or the induction of apoptosis, the paraxial mesoderm was tested for expression of Msx-2, BMP-2, BMP-4, and BMP-7. Moreover, BrdU and TUNEL staining were carried out. The inhibition of epithelialization after ectoderm removal alone and after segregation of the axial organs is accompanied neither by an increase in apoptosis nor by a reduction of the proliferation rate in the paraxial mesoderm. On the other hand, an ectopic BMP-4 expression in the paraxial mesoderm after ectoderm removal in combination with barrier insertion coincides with the occurrence of apoptotic cells and reduction of proliferation rate in this tissue. Increase of apoptosis and decrease in cell proliferation are observed in the paraxial and lateral plate mesoderm also after application of BMP-4 protein.

Animals↗
PubMed1998

Human mitochondrial phosphoenolpyruvate carboxykinase 2 gene. Structure, chromosomal localization and tissue-specific expression.

The mitochodrial (mt) phosphoenolpyruvate carboxykinase 2 (PCK2) gene was isolated by screening a human genomic library with a rat cytosolic (cy) PCK1 cDNA probe comprising sequences from exons 2-9 and by PCR amplification of human genomic DNA spanning consecutive exons with known primer pairs from mtPCK2 cDNA containing sequences from two putative neighbouring exons. The mtPCK2 gene spans approx. 10 kb and consists of ten exons and nine introns. All exon-intron junction sequences match the classical GT/AG rule. Northern blot analysis of poly(A)+ and total RNA from various tissues revealed one mRNA species of approx. 2.4 kb. The gene is expressed in a variety of human tissues, mainly in liver, kidney, pancreas, intestine and fibroblasts. In contrast with the cytosolic isoenzyme, the mitochondrial form might not have a purely gluconeogenic function. The mtPCK2 gene maps to chromosome 14q11.2-q12, in contrast with the cyPCK1 gene located on 20q13.2-q13.31.

Animals↗
PubMed1998

Cell death of neonatal rat sensory neurons is prevented by culture at clonal density.

We studied the trophic requirements of neonatal rat sensory neurons in single neuron and standard culture (9400 neurons/cm2). In agreement with previous studies, we found that survival of sensory neurons in standard culture is strictly dependent on nerve growth factor (NGF). Contrary to this, however, no difference between NGF-treated and untreated cultures with respect to survival and neurite formation was evident when neurons were plated as single cells. Approximately 70% of singly-seeded sensory neurons survived for 2 days independent of supply with NGF. Survival of large, mid-sized, and small sensory neurons in single neuron culture demonstrated that this phenomenon is not confined to a neuronal subpopulation. We speculate that survival in single neuron culture is due to the absence or inactivation of yet unidentified cell death-inducing factors that in standard culture are overridden or suppressed by NGF.

Animals↗
PubMed1998

The importance of timing differentiation during limb muscle development.

BACKGROUND: Skeletal muscle of trunk, limbs and tongue develops from a small population of cells that originates from somites. Although promoters and inhibitors of muscle differentiation have been isolated, nothing is known about how the amplification of the muscle precursor pool is regulated; this amplification provides muscle mass during development. Furthermore, little is known about how cells accumulate in the pre-muscle masses in the limbs. We investigated the role of bone morphogenetic protein (BMPs) and Sonic hedgehog (Shh) during proliferation, differentiation and positioning of muscle. RESULTS: The proliferation of muscle precursors in limbs was linked to Pax-3 expression. Ectoderm removal downregulated Pax-3 expression, arrested proliferation and prematurely initiated muscle differentiation which exhausted the muscle precursor pool and prevented further muscle growth. BMP-2, BMP-4 and BMP-7 had a dose-dependent effect on pre-myogenic cells: low concentrations maintained a Pax-3-expressing proliferative population, substituting for ectoderm-derived proliferative signals and delaying differentiation, whereas high concentrations prevented muscle development, probably by inducing apoptosis. In the limb, Shh upregulated Bmp-2 and Bmp-7 expression which delayed muscle differentiation, upregulated Pax-3, amplified the muscle precursor population and stimulated excessive muscle growth. CONCLUSIONS: These data indicate that embryonic muscle growth requires muscle differentiation to be delayed. Muscle differentiation may occur through a default pathway after cells escape proliferative signals. Positioning of muscle is regulated by high concentrations of BMPs, thus a single type of signalling molecule can determine crucial steps in muscle development: when and where to proliferate, and when and where to differentiate.

Animals↗