scholarly journals DEPENDENCE OF SALIVARY EPITHELIAL MORPHOLOGY AND BRANCHING MORPHOGENESIS UPON ACID MUCOPOLYSACCHARIDE-PROTEIN (PROTEOGLYCAN) AT THE EPITHELIAL SURFACE

1972 ◽  
Vol 52 (3) ◽  
pp. 674-689 ◽  
Author(s):  
Merton R. Bernfield ◽  
Shib D. Banerjee ◽  
Ronald H. Cohn
Keyword(s):  
Basal Lamina ◽  
Branching Morphogenesis ◽  
Culture Conditions ◽  
Acid Mucopolysaccharide ◽  
Proliferating Cells ◽  
In Vitro Morphogenesis ◽  
Epithelial Surface ◽  
Epithelial Basement Membrane ◽  
Epithelial Morphology

The morphogenetic role of the acid mucopolysaccharide (glycosaminoglycan) at the epithelial surface of mouse embryo submandibular glands has been studied by comparing the in vitro morphogenesis of epithelia from which the mucopolysaccharide was removed with that of those that retained the mucopolysaccharide. Epithelia isolated free of mesenchyme by procedures which retain the bulk of surface mucopolysaccharide maintain their lobular shape and undergo uninterrupted branching morphogenesis in culture in direct combination with fresh mesenchyme. Under identical culture conditions, epithelia from which surface mucopolysaccharide was removed lose their lobules and become spherical masses of tissue. During continued culture, the spherical epithelia produce outgrowths from which branching morphogenesis resumes. The morphogenetically active mucopolysaccharide is localized within the basal lamina of the epithelial basement membrane and appears to be bound to protein. During culture in combination with mesenchyme, epithelia undergoing uninterrupted morphogenesis show maximal accumulation of newly synthesized surface mucopolysaccharide at the distal ends of the lobules, the sites of incipient branching. In contrast, the material accumulates nearly equivalently over the surface of the spherical epithelia, with the exception that there is greater accumulation of the material at the surfaces of the budding outgrowths, the sites where morphogenesis will resume. Rapidly proliferating cells are localized within the lobules of epithelia undergoing uninterrupted morphogenesis, but are distributed uniformly in the cortex of the spherical epithelia, except for the outgrowths which show a greater localization of proliferating cells. It is concluded that normal salivary epithelial morphology and branching morphegenesis require the presence of acid mucopolysaccharide-protein within the epithelial basal lamina.

scholarly journals ACID MUCOPOLYSACCHARIDE (GLYCOSAMINOGLYCAN) AT THE EPITHELIAL-MESENCHYMAL INTERFACE OF MOUSE EMBRYO SALIVARY GLANDS

1972 ◽  
Vol 52 (3) ◽  
pp. 664-673 ◽  
Author(s):  
Merton R. Bernfield ◽  
Shib D. Banerjee
Keyword(s):  
Mouse Embryo ◽  
Uronic Acid ◽  
Branching Morphogenesis ◽  
Ruthenium Red ◽  
Acid Mucopolysaccharide ◽  
Epithelial Surface ◽  
Specific Staining ◽  
Ruthenium Red Staining ◽  
Cleft Formation ◽  
Rate Of Accumulation

Acid mucopolysaccharide (glycosaminoglycan) has been demostrated at the epithelial-mesenchymal interface of mouse embryo submandibular glands by (a) specific staining for polymeric sulfate with Alcian blue 8 GX at various magnesium concentrations, (b) specific staining for polymeric uronic acid by selective oxidation of these residues to Schiff-reactive compounds, (c) electron microscope localization of ruthenium red staining, (d) radioautographic localization of glucosamine-3H and 35SO4, and (e) by susceptibility of the glucosamine radioactivity at the interface to digestion with protease-free hyaluronidase. Moreover, material labeled with glucosamine-3H and 35SO4 and with chemical characteristics identical with those of acid mucopolysaccharide were isolated from the glands. Acid mucopolysaccharide is distributed over the entire epithelial surface. The amount of acid mucopolysaccharide, as revealed by the staining procedures, is nearly equivalent at all sites. In contrast, the rate of accumulation of glucosamine-labeled mucopolysaccharide is greater at the surface of the distal ends of the growing and branching lobules. This distribution of newly synthesized acid mucopolysaccharide at the sites of incipient cleft formation suggests that surface-associated acid mucopolysaccharide is involved in the morphogenetic process. A mechanism of branching morphogenesis is proposed which accounts for the distribution of collagen fibers and total and newly synthesized acid mucopolysaccharide at the epithelial surface.

scholarly journals Determination of the curvature of epithelial cell mass by mesenchyme in branching morphogenesis of mouse salivary gland

Development ◽  
1983 ◽  
Vol 73 (1) ◽  
pp. 221-232
Author(s):  
Hiroyuki Nogawa
Keyword(s):  
Salivary Gland ◽  
Epithelial Cell ◽  
Cell Mass ◽  
Branching Morphogenesis ◽  
Epithelial Surface ◽  
In Vivo Recombination ◽  
Recombination Experiments

Mouse submaxillary epithelium undergoes branching morphogenesis with increase in the curvature of its surface in vivo. Recombination experiments in vitro of the epithelium and mesenchyme between 13- and 14-day rudiments showed (1) that the 14-day mesenchyme more actively induced the epithelium to branch than the 13-day mesenchyme, (2) that the 14-day mesenchyme could produce clefts on smaller epithelial lobes than the 13-day mesenchyme, (3) that the 14-day mesenchyme produced lobes with similar diameter to lobes of the 14-day intact rudiment, and (4) that the lobular morphology of assembled 14-day lobes became obscure in recombinates with the 13-day mesenchyme while it was well maintained in recombinates with the 14-day mesenchyme. From these results it is concluded that the mesenchyme determines the curvature of epithelial surface, and that clefts are formed on the epithelial surface as a result of increase in the epithelial curvature.

scholarly journals Cooperative effects of colony-stimulating factor 1 and recombinant interleukin 2 on proliferation and induction of cytotoxicity of macrophage precursors generated from mouse bone marrow cell cultures.

1989 ◽  
Vol 169 (3) ◽  
pp. 973-986 ◽  
Author(s):  
H Li ◽  
R Schwinzer ◽  
M Baccarini ◽  
M L Lohmann-Matthes
Keyword(s):  
Bone Marrow ◽  
Marrow Cell ◽  
Interleukin 2 ◽  
Culture Conditions ◽  
Mouse Bone Marrow ◽  
Nk Activity ◽  
Proliferating Cells ◽  
Mouse Bone Marrow Cell ◽  
Cooperative Effects

Precursor cells for NK activity, present in the light fraction of fresh mouse bone marrow, were cultivated in vitro in the presence of either CSF-1, IL-2, or a combination of both factors. In the presence of only CSF-1, strong proliferation was induced. Cells quickly passed the macrophage precursor stage and matured to typical macrophages. Neither granula formation nor NK activity were induced. Under culture conditions with only IL-2 NK activity had developed after 3 d, however, no significant proliferation occurred. In the presence of both factors strong proliferation was induced, and concomitantly, granula formation and NK activity developed. Apparently, proliferation depended on CSF-1 and granula formation, and NK cytotoxicity was induced by IL-2. When proliferating cells with strong anti-YAC-1 activity from a culture in CSF-1 plus IL-2 were further cultivated in only IL-2, the content of granula further increased, whereas proliferation gradually stopped. In contrast, when these cells from CSF-1 plus IL-2 culture were further cultivated in only CSF-1, granula disappeared and NK activity was lost, whereas sustained proliferation and differentiation to macrophages occurred. Only under culture conditions with both factors were proliferation and NK activity both maintained. More than 90% of cells from a 3-d culture in CSF-1 plus IL-2 expressed the NK 1.1. marker, whereas F4/80 was only marginally detected by FACS analysis. After two further days in culture, 70% of the cells expressed F4/80 and 60% coexpressed NK 1.1. and F4/80. By setting the size scatter in order to gate for large granular cells, a population was obtained with 100% coexpression of NK1.1. and F4/80. The data indicate that early cells of the macrophage lineage can develop into different functional and morphological directions depending on the varying influence of IL-2 and CSF-1.

scholarly journals Transcriptomic analysis of native versus cultured human and mouse dorsal root ganglia focused on pharmacological targets

2019 ◽  
Author(s):  
Andi Wangzhou ◽  
Lisa A. McIlvried ◽  
Candler Paige ◽  
Paulino Barragan-Iglesias ◽  
Carolyn A. Guzman ◽  
...  
Keyword(s):  
Dorsal Root ◽  
Culture Conditions ◽  
Proliferating Cells ◽  
Illumina Platform ◽  
Human Organ ◽  
Expression Of Genes ◽  
Pharmacological Targets ◽  
Human And Mouse

AbstractDorsal root ganglion (DRG) neurons detect sensory inputs and are crucial for pain processing. They are often studied in vitro as dissociated cell cultures with the assumption that this reasonably represents in vivo conditions. However, to our knowledge, no study has ever directly compared genome-wide transcriptomes of DRG tissue in vivo versus in vitro, or between different labs and culturing protocols. We extracted bilateral lumbar DRG from C57BL6/J mice and human organ donors, and acutely froze one side and processed the other side as a dissociated cell culture, which was then maintained in vitro for 4 days. RNA was extracted and sequenced using the NextSeq Illumina platform. Comparing native to cultured human or mouse DRG, we found that the overall expression level of many ion channels and GPCRs specifically expressed in neurons is markedly lower in culture, but still expressed. This suggests that most pharmacological targets expressed in vivo are present in culture conditions. However, there are changes in expression levels for these genes. The reduced relative expression for neuronal genes in human DRG cultures is likely accounted for by increased expression of genes in fibroblast-like and other proliferating cells, consistent with the mitotic status of many cells in these cultures. We did find a subset of genes that are typically neuronally expressed, increased in human and mouse DRG cultures, including genes associated with nerve injury and/or inflammation in preclinical models such as BDNF, MMP9, GAL, and ATF3. We also found a striking upregulation of a number of inflammation-associated genes in DRG cultures, although many were different between mouse and human. Our findings suggest an injury-like phenotype in DRG cultures that has important implications for the use of this model system for pain drug discovery.

scholarly journals Maintaining Hair Inductivity in Human Dermal Papilla Cells: A Review of Effective Methods

2020 ◽  
Vol 33 (5) ◽  
pp. 280-292
Author(s):  
Ehsan Taghiabadi ◽  
Mohammad Ali Nilforoushzadeh ◽  
Nasser Aghdami
Keyword(s):  
Hair Follicle ◽  
Dermal Papilla ◽  
Culture Conditions ◽  
Hair Follicles ◽  
Main Role ◽  
Follicle Growth ◽  
In Vitro Morphogenesis ◽  
Dermal Papilla Cells ◽  
Hair Follicle Growth

The dermal papilla comprises mesenchymal cells in hair follicles, which play the main role in regulating hair growth. Maintaining the potential hair inductivity of dermal papilla cells (DPCs) and dermal sheath cells during cell culture is the main factor in in vitro morphogenesis and regeneration of hair follicles. Using common methods for the cultivation of human dermal papilla reduces the maintenance requirements of the inductive capacity of the dermal papilla and the expression of specific dermal papilla biomarkers. Optimizing culture conditions is therefore crucial for DPCs. Moreover, exosomes appear to play a key role in regulating the hair follicle growth through a paracrine mechanism and provide a functional method for treating hair loss. The present review investigated the biology of DPCs, the molecular and cell signaling mechanisms contributing to hair follicle growth in humans, the properties of the dermal papilla, and the effective techniques in maintaining hair inductivity in DPC cultures in humans as well as hair follicle bioengineering.

scholarly journals A QUANTITATIVE METHOD FOR ANALYSING 3-D BRANCHING IN EMBRYONIC KIDNEYS: DEVELOPMENT OF A TECHNIQUE AND PRELIMINARY DATA

2011 ◽  
Vol 21 (1) ◽  
pp. 37 ◽  
Author(s):  
Gabriel Fricout ◽  
Luise A Cullen-McEwen ◽  
Ian S Harper ◽  
Dominique Jeulin ◽  
John F Bertram
Keyword(s):  
Preliminary Data ◽  
Quantitative Methods ◽  
Kidney Development ◽  
Automatic Segmentation ◽  
Branching Morphogenesis ◽  
Culture Conditions ◽  
Branch Points ◽  
Total Length ◽  
Fundamental Information

The normal human adult kidney contains between 300,000 and 1 million nephrons (the functional units of the kidney). Nephrons develop at the tips of the branching ureteric duct, and therefore ureteric duct branching morphogenesis is critical for normal kidney development. Current methods for analysing ureteric branching are mostly qualitative and those quantitative methods that do exist do not account for the 3- dimensional (3D) shape of the ureteric "tree". We have developed a method for measuring the total length of the ureteric tree in 3D. This method is described and preliminary data are presented. The algorithm allows for performing a semi-automatic segmentation of a set of grey level confocal images and an automatic skeletonisation of the resulting binary object. Measurements of length are automatically obtained, and numbers of branch points are manually counted. The final representation can be reconstructed by means of 3D volume rendering software, providing a fully rotating 3D perspective of the skeletonised tree, making it possible to identify and accurately measure branch lengths. Preliminary data shows the total length estimates obtained with the technique to be highly reproducible. Repeat estimates of total tree length vary by just 1-2%. We will now use this technique to further define the growth of the ureteric tree in vitro, under both normal culture conditions, and in the presence of various levels of specific molecules suspected of regulating ureteric growth. The data obtained will provide fundamental information on the development of renal architecture, as well as the regulation of nephron number.

Expression of the protein zero myelin gene in axon-related Schwann cells is linked to basal lamina formation

Development ◽  
1993 ◽  
Vol 119 (3) ◽  
pp. 867-880 ◽  
Author(s):  
C. Fernandez-Valle ◽  
N. Fregien ◽  
P.M. Wood ◽  
M.B. Bunge
Keyword(s):  
Gene Expression ◽  
Schwann Cell ◽  
Schwann Cells ◽  
Basal Lamina ◽  
Culture Conditions ◽  
Structural Protein ◽  
Gene Encoding ◽  
Serum Free ◽  
Protein Zero

A Schwann cell has the potential to differentiate into either a myelinating or ensheathing cell depending upon signals received from the axon that it contacts. Studies focusing on the pathway leading to myelination demonstrated that Schwann cells must form a basal lamina in order to myelinate an axon. In this report, we describe studies that indicate that initiation of basal lamina synthesis is required for Schwann cells to distinguish between myelination-inducing axons and axons that do not induce myelination, and to respond by undergoing the appropriate genetic and cellular changes. We have used high resolution in situ hybridization, immunocytochemistry and electron microscopy to examine changes in gene expression and morphology of Schwann cells differentiating into myelin-forming cells in vitro. These experiments were carried out in dorsal root ganglion neuron/Schwann cell co-cultures maintained in either serum-free, serum-only or serum-plus-ascorbate-containing medium. We have made four novel observations that contribute significantly to our understanding of how basal lamina and myelination are linked. (1) The addition of ascorbate (in the presence of serum), which promotes basal lamina production, appears to induce expression of the protein zero gene encoding the major structural protein of myelin. Moreover, expression of protein zero mRNA and protein, and its insertion into myelin membranes, occurs only in the subset of Schwann cells contacting myelination-inducing axons. Schwann cells in contact with axons that do not induce myelination, or Schwann cells that have not established a unitary relationship with an axon, do not express protein zero mRNA although they produce basal lamina components. (2) In serum-free conditions, a majority of Schwann cells express protein zero mRNA and protein, but this change in gene expression is not associated with basal lamina formation or with elongation of the Schwann cell along the axon and elaboration of myelin. (3) In the presence of serum (and the absence of ascorbate), Schwann cells again fail to form basal lamina or elongate but no longer express protein zero mRNA or protein. (4) Myelin-associated glycoprotein and galactocerebroside, two additional myelin-specific components, can be expressed by Schwann cells under any of the three culture conditions. Therefore, we have demonstrated that axonal induction of protein zero gene expression in Schwann cells is subject to regulation by both serum- and ascorbate-dependent pathways and that not all myelin-specific proteins are regulated in the same manner.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Generation of proliferating human adult hepatocytes using optimized 3D culture conditions

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Sophie Rose ◽  
Frédéric Ezan ◽  
Marie Cuvellier ◽  
Arnaud Bruyère ◽  
Vincent Legagneux ◽  
...  
Keyword(s):  
Cell Cycle ◽  
3D Culture ◽  
Culture Conditions ◽  
Human Hepatocytes ◽  
Cellular Interactions ◽  
Matrix Stiffness ◽  
Hepatic Differentiation ◽  
Proliferating Cells ◽  
Promising Tool

AbstractGenerating the proliferation of differentiated normal adult human hepatocytes is a major challenge and an expected central step in understanding the microenvironmental conditions that regulate the phenotype of human hepatocytes in vitro. In this work, we described optimized 3D culture conditions of primary human hepatocytes (PHH) to trigger two waves of proliferation and we identified matrix stiffness and cell–cell interactions as the main actors necessary for this proliferation. We demonstrated that DNA replication and overexpression of cell cycle markers are modulate by the matrix stiffness while PHH cultured in 3D without prior cellular interactions did not proliferate. Besides, we showed that PHH carry out an additional cell cycle after transient inhibition of MAPK MER1/2-ERK1/2 signaling pathway. Collagen cultured hepatocytes are organized as characteristic hollow spheroids able to maintain survival, cell polarity and hepatic differentiation for long-term culture periods of at least 28 days. Remarkably, we demonstrated by transcriptomic analysis and functional experiments that proliferating cells are mature hepatocytes with high detoxication capacities. In conclusion, the advanced 3D model described here, named Hepoid, is particularly relevant for obtaining normal human proliferating hepatocytes. By allowing concomitant proliferation and differentiation, it constitutes a promising tool for many pharmacological and biotechnological applications.

Induction and Differentiation of Dental Epithelium in Vivo and in Vitro

1982 ◽  
Vol 40 ◽  
pp. 142-145
Author(s):  
E. J. Kollar
Keyword(s):  
Connective Tissue ◽  
Oral Mucosa ◽  
Basal Lamina ◽  
Functional Derivatives ◽  
Specific Source ◽  
Dental Epithelium ◽  
Connective Tissue Stroma

The differentiation and maintenance of many specialized epithelial structures are dependent on the underlying connective tissue stroma and on an intact basal lamina. These requirements are especially stringent in the development and maintenance of the skin and oral mucosa. The keratinization patterns of thin or thick cornified layers as well as the appearance of specialized functional derivatives such as hair and teeth can be correlated with the specific source of stroma which supports these differentiated expressions.

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