Biomedical subjects

H Magloire

Publications and source records attributed to H Magloire.

At least 19 recordsLinked to original sources

PubMed1992

Immunohistochemical localization of nerve growth factor (NGF) and NGF receptor (NGF-R) in the developing first molar tooth of the rat.

Nerve growth factor (NGF) is a well established target-derived trophic factor supporting sympathetic and sensory innervation in the peripheral tissues as well as cholinergic innervation in the brain. Despite its name, NGF may have broader biological functions early in development in a wide range of non-neuronal differentiating cells. The many effects of NGF are directly dependent on initial binding of NGF to specific plasma membrane receptors on target cells. Here we use immunohistochemical methods to show that NGF and its receptor (NGF-R) are localized in a variety of embryonic epithelial and mesenchymal cells in the rat developing molar tooth. Dental cells known to play important roles in morphogenesis and inductive tissue interactions show NGF-like reactivity. Thus, labelling is seen in epithelial preameloblasts and mesenchymal odontoblasts. We also show a transient expression of NGF-R in restricted parts of the dental epithelium (inner dental epithelium) and dental mesenchyme differentiating cells (post-mitotic, polarizing odontoblasts). The expression patterns of NGF are different to those of NGF-R during embryogenesis and this is illustrated in detail in the developing tooth. The histochemical findings reported here support the notion that NGF may have multiple roles during morphogenetic and cytodifferentiation events in the tooth.

Animals↗

PubMed1992

Odontoblast response under carious lesions.

The local regulation of odontoblast response to caries is viewed through initiation and elaboration of sclerotic as well as reparative dentin. Dentin tissue represents a multiple source of potent environment factors when teeth are affected by the demineralization phases of carious process. Some of them have already been identified in sound tissue (matrix glycoproteins, proteoglycans, growth factors, Bone Morphogenetic Protein) and may act on the cell through membrane receptors. Thus, the amplification in collagen synthesis and alkaline phosphatase activity previously observed during sclerotic dentin deposition can be related to the interaction between matrix signals and cell receptors such as the 165 kDa protein shown only by odontoblasts under the affected zone. Similarly, under established lesions generating cell death, the specific matrix made of odontoblasts debris and damage tissues, probably rich in active molecules, may trigger pulp cells to elaborate a cartilage-like layer (identified by type II and XI collagen) followed by odontoblast-like cells to give rise to abnormal tubular dentin. Here, odontoblast response is identical to bone-cells response to injury. What remains to be elucidated concern: The nature of signals found in carious dentin (matrix components, growth factors, bacterial products). The nature and regulation of expression of cell membrane receptors during tooth repair. How the odontoblast produces specific responses to each of these signaling molecules will be the focus of important new investigations.

Cell Differentiation↗

PubMed1991

Morphological and immunocytochemical characterization of cultured rat incisor cervical epithelial cells.

Epithelial cells from the cervical loop of the rat incisor were isolated by co-culture of apical explants with growth-arrested 3T3 fibroblasts. The epithelial phenotype of the expanding outgrowths was confirmed 10 days after the seeding of the explants by phase-contrast microscopy and immunocytochemical identification of cytokeratins. After 3 weeks in culture, the epithelial cells covered the entire surface of the coverslips and were then passaged. Subcultures gave rise to a confluent sheet within 10-12 days. Light and electron microscopy showed that confluent cervical epithelial cells generally reconstituted a bi-layered structure similar to Hertwig's epithelial sheath. Epithelial cells from the rat palate, cultured and subcultured according to the same procedure, organized themselves in 5-6 cell layers, the upper cells having generally a squamous morphology. Synthesis of extracellular matrix molecules by rat incisor cervical epithelial cells was studied with specific antibodies. These cells failed to produce type I collagen, but synthesized all the major basement membrane components (type IV collagen, laminin, heparan sulphate proteoglycan and fibronectin). These observations suggest that the culture conditions allowed the reconstitution of a typical Hertwig's epithelial sheath by rat incisor cervical epithelial cells.

Animals↗

PubMed1991

Immunoblotting and cytochemical characterization of human enamel proteins.

Mature enamel proteins (tuft proteins) and fetal enamel proteins were extracted by an homogenizing buffer method, subjected to SDS-PAGE and immunoblotted with a polyclonal antibody raised against the mature enamel proteins. Both fetal and tuft proteins were recognized by this immunoblotting. With the same antibody, immunolocalization of the developing enamel proteins was done on semi-thin-sections of human fetal tissue at the secretory stage, using an immunoperoxidase technique. Specific labelling of the enamel protein matrix was observed. It is concluded that a polyclonal antibody against mature enamel proteins (anti-tuft) can recognize the developing protein matrix at the secretory stage. This suggests that a common antigenic determinant is maintained throughout the course of amelogenesis in human enamel.

Adult↗

PubMed1991

Odontoblast-like cytodifferentiation of human dental pulp cells in vitro in the presence of a calcium hydroxide-containing cement.

The cement produced microcrystals of calcite by reaction with culture medium supplemented with calf serum. Human dental pulp cells seeded on such a substrate preferentially adhered and aggregated around the microcrystals. Immunofluorescence and immunogold labelling revealed a high affinity of serum fibronectin molecules for the calcite crystals. At 4 weeks in culture, the cells had various features of differentiated odontoblasts, notably nuclear polarization, typical appearance of the Golgi apparatus, synthesis of type I collagen and absence of type III, and apical accumulation of actin and vimentin. These cells also elaborated a collagenous extracellular matrix which did not mineralize.

Adolescent↗

PubMed1991

Differential expression of type I and type III collagen genes during tooth development.

Collagen gene expression during mouse molar tooth development was studied by quantitative in situ hybridization techniques. Different expression patterns of type I and type III collagen mRNAs were observed in the various mesenchymal tissues that constitute the tooth germ. High concentration for pro-alpha 1(I) and pro-alpha 2(I) collagen mRNAs were found within the osteoblasts. We found that the cellular content of type I collagen mRNAs in the odontoblasts varies throughout the tooth formation: whereas mRNA concentration for pro-alpha 1(I) collagen decreases and that of pro-alpha 2(I) increases, during postnatal development. Moreover, different amounts of pro-alpha 1(I) and pro-alpha 2(I) collagen mRNAs were observed in crown and root odontoblasts, respectively. Type III collagen mRNAs were detected in most of the mesenchymal cells, codistributed with type I collagen mRNAs, except in odontoblasts and osteoblasts. Finally, this study reports differential accumulation of collagen mRNAs during mouse tooth development and points out that type I collagen gene expression is regulated by distinct mechanisms during odontoblast differentiation process. These results support the independent expression of the collagen genes under developmental tissue-specific control.

Animals↗

PubMed1991

Isolation and characterization of rat alveolar bone cells.

Samples of rat alveolar bone were first treated by collagenase digestion and then used as explants for cell culture. The cells obtained were subcultured and characterized by morphological and functional criteria. Their alkaline phosphatase activity was increased after incubation in 1,25-(OH)2 vitD3 10(-8) M whereas with gingival cells it did not change. The bone derived-cells organized nodular structures, synthesized type I collagen, Gla-protein, few type III collagen, and fibronectin. In the defined culture conditions no mineralization was observed. However, the method used allows to obtain cells from rat alveolar bone displaying some features of the osteoblastic phenotype.

Alveolar Process↗

PubMed1991

Extracellular matrix and intermediate filaments in the first stages and repair of experimental gingivitis in man.

By the indirect immunoperoxidase labelling procedure the expressions of type I and III collagens, laminin and fibronectin and of KL1 cytokeratin and vimentin were examined in the first stages and repair of experimental gingivitis in young subjects. A two month longitudinal study was performed using the tissue from buccal marginal gingival biopsies of four subjects taken sequentially at five specific times: before and during plaque accumulation, and during plaque elimination. The sites examined microscopically were the coronal half of the junctional epithelium and the underlining infiltrated connective tissue fraction. No clinical change could be observed during the study period. Histological examination showed reversible cellular changes during the accumulation of plaque. There were increases in vascularization and cellularity and loss of collagen. They recovered 56 days after plaque elimination their baseline level. Electron microscopic examination showed myofibroblastic aspects in some fibroblasts. The changes in the expression of laminin, fibronectin and KL1 in the J.E. might be due to a proliferation rate enhancement, and suggest an adaptation to the alterations brought about by the inflammatory process. They also reinforce the hypothesis that this epithelium resembles a developmental tissue. Type I collagen demonstrated the "collagen loss-repair" cycle shown in connective tissue by the histological study. The rise in type III collagen and the vimentin fall, both at the initial stage, suggest that these protein profiles may yield information for clinical research purposes during the very early inflammatory process. The variations in fibronectin indicate its key role in the early inflammatory and repair processes. Finally, the variations in matrix and cytoskeletal proteins variations, as well as the morphological modifications observed, were nearly all reversible.

Adolescent↗

PubMed1991

[A model of an in vitro biological assay controlled by immunofluorescence and scanning electron microscopy].

This study was designed to complete the cell culture test for the cytotoxicity assessment of dental material presented by Christen et al. (1989) and Regad et al. (1989). Two liner materials were selected for this experiment which had already been subjected to both in vivo and in vitro tests. After 2, 7 and 14 days of contact with the test materials, the cell responses were evaluated under the light microscope by their growth pattern and morphological aspects. Cell synthesis of fibronectin and type I and III collagen was studied by immunofluorescence. Surface appearance of the cells in contact with the materials as well as their structures were observed by scanning electron microscopy. This study confirmed the viability of the in vitro cytotoxicity assessment methods with cell cultures. The results also demonstrated the potentials of this method in the early screening of dental materials for their biological compatibility.

Acrylic Resins↗

PubMed1990

In vitro mineralization of a three-dimensional collagen matrix by human dental pulp cells in the presence of chondroitin sulphate.

These matrices were used as cell culture substrates to investigate the influence of extracellular molecules on mineralization. Pulp cells seeded in type I collagen or type I collagen-chondroitin-4-sulphate sponges were able to grow and were morphologically similar to cells responsible for reparative dentine formation in vivo. In sponges consisting of collagen only, the cells elaborated an abundant new matrix which became organized with time and consisted of collagen fibres surrounded by fibrillar material, but no mineralization was observed. In collagen-chondroitin sulphate sponges, cells deposited less and poorly organized matrix; in these, calcification occurred, increasing with time, and at the ultrastructural level, small needle-like crystals containing calcium and phosphorus were scattered throughout the sponge fibres. These observations suggest that chondroitin sulphate might influence in vitro calcification induced by pulp cells.

Cells, Cultured↗

PubMed1990

Expression of fibronectin and type I collagen by human dental pulp cells and gingiva fibroblasts grown on fibronectin substrate.

Specific antibodies and indirect immunoperoxidase labelling were used to study the intracellular production of collagen and fibronectin by cells grown on fibronectin-coated glass; the same cell populations seeded on uncoated glass were used as controls. Strong intracellular staining for type I collagen was seen in all cases, but immunostaining for fibronectin was very faint or negative in both gingival and pulp cells grown on the fibronectin substrate, in contrast to control cells. Thus, fibronectin substrate inhibited fibronectin synthesis by the cultured cells, but did not seem to influence type I collagen synthesis.

Cells, Cultured↗

PubMed1990

Cytokeratins as molecular markers in the evaluation of the precise differentiation stage of human gingival epithelium reconstituted in vitro.

Cytokeratins are considered to be molecular markers for different types of epithelial differentiation. They were used to investigate the precise differentiation stage of gingival epithelium, reconstituted in vitro, following two different culture procedures. Human trypsin-dissociated gingival keratinocytes were seeded either on a feeder layer of irradiated mouse 3T3 fibroblasts or on a connective tissue equivalent (lattice) made up of human fibroblasts in a collagen gel. The cytokeratins were extracted and analysed by two-dimensional gel electrophoresis. Although both methods showed on histological sections that cultured gingival keratinocytes formed a multilayered non-keratinizing epithelium, the cytokeratins patterns showed great differences. The gingival epithelium-like structure reconstituted on 3T3 feeder layer expressed some cytokeratins characteristic of the in situ gingival epithelium (K 5, 6, 14, 16, 17) and some which do not exist in the normal tissue (K 8, 18, 19, traces of K 13 and K 15) and are specific for embryonic, simple and tumour epithelia. However, the gingival epithelium reconstituted on connective tissue equivalent expressed all the cytokeratins present in the normal tissue (K 5, 6, 14, 16, 17), except those specific for terminal differentiation (K 1, 2, and 10/11). These findings suggest that the culture of gingival keratinocytes on connective tissue equivalents allows them to reproduce physiological stages of differentiation.

Biomarkers↗

PubMed1990

Ultrastructural and immunocytochemical study of bone-derived cells cultured in three-dimensional matrices: influence of chondroitin-4 sulfate on mineralization.

Bone-derived cells were cultured in three-dimensional reconstituted matrices made of type I collagen or type I collagen chondroitin-4-sulfate. As observed by microscope, their characteristics were as follows: The cells deposited a faint extracellular matrix mainly composed of type I collagen. In the collagen-chondroitin-sulfate sponge fibers, a calcification process, which involved the deposition of hydroxyapatite crystals, was demonstrated. Mineralization occurred only in collagen chondroitin sulfate sponge fibers when seeded with bone-derived cells and was not seen with nonosteogenic cells, such as gingival fibroblasts. Gla protein was intracellularly visualized in both types of sponges seeded with bone-derived cells while an extracellular secretion was seen only in the collagen chondroitin sulfate sponge fibers where calcification occurred. These results suggest that collagen chondroitin sulfate promotes in vitro mineralization of three-dimensional collagen matrices when seeded with bone-derived cells.

Animals↗

PubMed1990

[Epidermal growth factor in tooth development].

The epidermic growth factor (EGF) inhibits the morphogenesis and cellular differentiation of dental tissues during embryogenesis. These effects are modulated according to the stage of tooth development. Thus, the cupula stage seems markedly affected contrary to the bell stage which continues to evolve eve in the presence of EGF. Distribution of EGF receptors is altered during the first stages or the morphogenesis and this is specific for each stage of dental development. Finally, some rare pathological processes, related to the EGF level, are particularly characterized by a very early growth of the teeth.

Epidermal Growth Factor↗

PubMed1990

Collagen gene expression and tooth development. An overview.

The regulation aspects of type I and type III collagen gene expression are examined and the relationships with tooth morphogenesis and differentiation are discussed. Type I and III collagens constitute the major molecular proteins of the dental tissues. In addition the collagen gene expression in the mesenchyme derived odontoblasts represents an important step in the cytodifferentiation at the mesenchymal level. Furthermore, odontoblasts seem to synthesize only type I and type I trimer collagens, but not type III collagen. Therefore, the aim of this overview is to describe the molecular mechanisms that control the expression of specific collagen genes during the process of odontoblast differentiation. The available data support the main transcriptional control and argue for the existence of an independent and developmental regulation during collagen gene expression in odontoblast cells.

Animals↗

PubMed1989

Immunoelectron microscopic localization of dentin gamma-carboxyglutamic acid-containing proteins in differentiating rat odontoblasts.

The intracellular synthesis of the dentin-gamma-carboxyglutamic acid-containing proteins (DGPs) by rat odontoblasts was investigated at the electron microscopic level using a sensitive pre-embedding immunoperoxidase technique. The DGPs were detected in the rough endoplasmic reticulum and secretory vesicles, but not in the Golgi apparatus of the odontoblasts, while dentin matrix is not yet reactive. These results suggest that the DGPs synthesis is independent of mineral deposition.

1-Carboxyglutamic Acid↗

PubMed1989

Immunolocalization of cathepsin D in dental tissues.

Cathepsin D antigenicity was localized at the light and electron microscopic levels within dental cells, but not in extracellular matrix. Different intracellular sites for cathepsin D were found depending on the cell type: the enzyme was detected in secretory vesicles of the odontoblasts and in the lysosome-like structures of the ameloblasts. Otherwise, these results suggest that the secretory vesicles of the odontoblasts may contain both cathepsin D and type I collagen. These data might implicate cathepsin D in the enamel and the dentin formations.

Animals↗

PubMed1989

Human recombinant gamma-interferon stimulates proliferation and inhibits collagen and fibronectin production by human dental pulp fibroblasts.

Human recombinant-gamma-interferon was tested on human dental pulp fibroblast activity in vitro. Fibroblast proliferation was estimated by a colorimetric test. Type I and type III collagens and fibronectin were quantified by radioimmunoassay in culture supernatant from confluent fibroblasts. A dose dependent stimulation of the proliferation was observed when fibroblasts were treated with recombinant-gamma-interferon. In contrast, an inhibition of the synthesis of soluble types I and III collagen and fibronectin by confluent cell cultures treated with recombinant-gamma-interferon occurred without apparent modification of the insoluble collagen level in the cell layer. Quantimetric analysis of type I collagen immunoperoxidase labelling have demonstrated that there was no intracellular storage of type I collagen in these cultured fibroblasts. These data support the view that human recombinant-gamma-interferon can affect human dental pulp fibroblast functions and thus may play an important part in the regulation of fibrosis.

Cell Division↗