Interactions of Matrix Proteins from Mineralized Tissues with Octacalcium Phosphate

Dental cementum contains unique molecules that regulate the mineralization process in vitro and in vivo, such as cementum protein 1 (CEMP1). This protein possesses amino acid sequence motifs like the human recombinant CEMP1 with biological activity. This novel cementum protein 1-derived peptide (CEMP1-p3, from the CEMP1’s N-terminal domain: (QPLPKGCAAVKAEVGIPAPH), consists of 20 amino acids. Hydroxyapatite (HA) crystals could be obtained through the combination of the amorphous precursor phase and macromolecules such as proteins and peptides. We used a simple method to synthesize peptide/hydroxyapatite nanocomposites using OCP and CEMP1-p3. The characterization of the crystals through scanning electron microscopy (SEM), powder X-ray diffraction (XRD), high--resolution transmission electron microscopy (HRTEM), and Raman spectroscopy revealed that CEMP1-p3 transformed OCP into hydroxyapatite (HA) under constant ionic strength and in a buffered solution. CEMP1-p3 binds and highly adsorbs to OCP and is a potent growth stimulator of OCP crystals. CEMP1-p3 fosters the transformation of OCP into HA crystals with crystalline planes (300) and (004) that correspond to the cell of hexagonal HA. Octacalcium phosphate crystals treated with CEMP1-p3 grown in simulated physiological buffer acquired hexagonal arrangement corresponding to HA. These findings provide new insights into the potential application of CEMP1-p3 on possible biomimetic approaches to generate materials for the repair and regeneration of mineralized tissues, or restorative materials in the orthopedic field.

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Cellular and Chemical Events During Enamel Maturation

Critical Reviews in Oral Biology & Medicine ◽

10.1177/10454411980090020101 ◽

1998 ◽

Vol 9 (2) ◽

pp. 128-161 ◽

Cited By ~ 489

Author(s):

C.E. Smith

Keyword(s):

Crystal Growth ◽

Driving Forces ◽

Morphological Changes ◽

Matrix Proteins ◽

Secretory Product ◽

Water Matrix ◽

Mineralized Tissues ◽

Progressive Growth ◽

Chemical Events ◽

Protein Rich

This review focuses on the process of enamel maturation, a series of events associated with slow, progressive growth in the width and thickness of apatitic crystals. This developmental step causes gradual physical hardening and transformation of soft, newly formed enamel into one of the most durable mineralized tissues produced biologically. Enamel is the secretory product of specialized epithelial cells, the ameloblasts, which make this covering on the crowns of teeth in two steps. First, they roughly "map out" the location and limits (overall thickness) of the entire extracellular layer as a protein-rich, acellular, and avascular matrix filled with thin, ribbon-like crystals of carbonated hydroxyapatite. These initial crystals are organized spatially into rod and interrod territories as they form, and rod crystals are lengthened by Tomes' processes in tandem with appositional movement of ameloblasts away from the dentin surface. Once the full thickness of enamel has been formed, ameloblasts initiate a series of repetitive morphological changes at the enamel surface in which tight junctions and deep membrane infoldings periodically appear (ruffle-ended), then disappear for short intervals (smooth-ended), from the apical ends of the cells. As this happens, the enamel covered by these cells changes rhythmically in net pH from mildly acidic (ruffle-ended) to near-physiologic (smooth-ended) as mineral crystals slowly expand into the "spaces" (volume) formerly occupied by matrix proteins and water. Matrix proteins are processed and degraded by proteinases throughout amelogenesis, but they undergo more rapid destruction once ameloblast modulation begins. Ruffle-ended ameloblasts appear to function primarily as a regulatory and transport epithelium for controlling the movement of calcium and other ions such as bicarbonate into enamel to maintain buffering capacity and driving forces optimized for surface crystal growth. The reason ruffle-ended ameloblasts become smooth-ended periodically is unknown, although this event seems to be crucial for sustaining long-term crystal growth.

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Postembedding colloidal-gold immunocytochemistry of noncollagenous extracellular matrix proteins in mineralized tissues

Microscopy Research and Technique ◽

10.1002/jemt.1070310105 ◽

1995 ◽

Vol 31 (1) ◽

pp. 44-62 ◽

Cited By ~ 86

Author(s):

M. D. McKee ◽

A. Nanci

Keyword(s):

Extracellular Matrix ◽

Colloidal Gold ◽

Extracellular Matrix Proteins ◽

Matrix Proteins ◽

Mineralized Tissues

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Comparative Immunochemical Analyses of the Developmental Expression and Distribution of Ameloblastin and Amelogenin in Rat Incisors

Journal of Histochemistry & Cytochemistry ◽

10.1177/002215549804600806 ◽

1998 ◽

Vol 46 (8) ◽

pp. 911-934 ◽

Cited By ~ 104

Author(s):

A. Nanci ◽

S. Zalzal ◽

P. Lavoie ◽

M. Kunikata ◽

W.-Y. Chen ◽

...

Keyword(s):

Protein Secretion ◽

Brefeldin A ◽

Matrix Proteins ◽

Developmental Expression ◽

Enamel Matrix Proteins ◽

Precise Knowledge ◽

Mineralized Tissues ◽

Maturation Stages ◽

Alternatively Spliced ◽

Enamel Layer

SUMMARY Mineralized tissues are unique in using proteins to attract and organize calcium and phosphate ions into a structured mineral phase. A precise knowledge of the expression and extracellular distribution of matrix proteins is therefore very important in understanding their function. The purpose of this investigation was to obtain comparative information on the expression, intracellular and extracellular distribution, and dynamics of proteins representative of the two main classes of enamel matrix proteins. Amelogenins were visualized using an antibody and an mRNA probe prepared against the major alternatively spliced isoform in rodents, and nonamelogenins by antibodies and mRNA probes specific to one enamel protein referred to by three names: ameloblastin, amelin, and sheathlin. Qualitative and quantitative immunocytochemistry, in combination with immunoblotting and in situ hybridization, indicated a correlation between mRNA signal and sites of protein secretion for amelogenin, but not for ameloblastin, during the early presecretory and mid-to late maturation stages, during which mRNA signals were detected but no proteins appeared to be secreted. Extracellular amelogenin immunoreactivity was generally weak near secretory surfaces, increasing over a distance of about 1.25 μm to reach a level slightly above an amount expected if the protein were being deposited evenly across the enamel layer. Immunolabeling for ameloblastin showed an inverse pattern, with relatively more gold particles near secretory surfaces and much fewer deeper into the enamel layer. Administration of brefeldin A and cycloheximide to stop protein secretion revealed that the immunoblotting pattern of amelogenin was relatively stable, whereas ameloblastin broke down rapidly into lower molecular weight fragments. The distance from the cell surface at which immunolabeling for amelogenin stabilized generally corresponded to the point at which that for ameloblastin started to show a net reduction. These data suggest a correlation between the distribution of amelogenin and ameloblastin and that intact ameloblastin has a transient role in promoting/stabilizing crystal elongation.

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Expression of matrix proteins during the development of mineralized tissues

Bone ◽

10.1016/s8756-3282(96)00218-9 ◽

1996 ◽

Vol 19 (4) ◽

pp. 371-380 ◽

Cited By ~ 91

Author(s):

B. Sommer ◽

M. Bickel ◽

W. Hofstetter ◽

A. Wetterwald

Keyword(s):

Matrix Proteins ◽

Mineralized Tissues

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Contribution of biomimetic collagen-ligand interaction to intrafibrillar mineralization

Science Advances ◽

10.1126/sciadv.aav9075 ◽

2019 ◽

Vol 5 (3) ◽

pp. eaav9075 ◽

Cited By ~ 10

Author(s):

Q. Song ◽

K. Jiao ◽

L. Tonggu ◽

L. G. Wang ◽

S. L. Zhang ◽

...

Keyword(s):

Biomechanical Properties ◽

Matrix Proteins ◽

Dynamics Simulation ◽

Cross Linking ◽

Ectopic Calcification ◽

Polycarboxylic Acid ◽

Ligand Interaction ◽

Collagen Matrices ◽

Body Tissues ◽

Mineralized Tissues

Contemporary models of intrafibrillar mineralization mechanisms are established using collagen fibrils as templates without considering the contribution from collagen-bound apatite nucleation inhibitors. However, collagen matrices destined for mineralization in vertebrates contain bound matrix proteins for intrafibrillar mineralization. Negatively charged, high–molecular weight polycarboxylic acid is cross-linked to reconstituted collagen to create a model for examining the contribution of collagen-ligand interaction to intrafibrillar mineralization. Cryogenic electron microscopy and molecular dynamics simulation show that, after cross-linking to collagen, the bound polyelectrolyte caches prenucleation cluster singlets into chain-like aggregates along the fibrillar surface to increase the pool of mineralization precursors available for intrafibrillar mineralization. Higher-quality mineralized scaffolds with better biomechanical properties are achieved compared with mineralization of unmodified scaffolds in polyelectrolyte-stabilized mineralization solution. Collagen-ligand interaction provides insights on the genesis of heterogeneously mineralized tissues and the potential causes of ectopic calcification in nonmineralized body tissues.

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Distribution of VLA-5 and VLA-4 fibronectin receptors in human monocytes demonstrated by immunofluorescence, immunocytochemistry, and autoradiography

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100147375 ◽

1993 ◽

Vol 51 ◽

pp. 306-307

Author(s):

Robert Williams ◽

Che-Hung Lee ◽

Sara E. Quella ◽

David M. Harlan ◽

Yuan-Hsu Kang

Keyword(s):

Present Report ◽

Matrix Proteins ◽

Extracellular Matrices ◽

Human Monocytes ◽

Integrin Receptors ◽

Morphological Evaluation ◽

The Matrix ◽

Specific Receptors ◽

Cytokine Stimulation ◽

Monocyte Adherence

Monocyte adherence to endothelial or extracellular matrices plays an important role in triggering monocyte activation in extravascular sites of infection, chronic inflammatory disorders, and tissue damage. Migration of monocytes in the tissues involves the response to a chemoattractant and movement by a series of attachments and detachments to the extracellular matrices which are regulated by expression and distribution of specific receptors for the matrix proteins such as fibronectin (FN). The VSAs (very late antigens or beta integrins), a subfamily of the transmembrane heterodimeric integrin receptors, have been thought to play a major role in monocyte adherence to the extracellular matrices and cells. In this subfamily, VLA-5 and VLA-4 are believed to be the most essential integrins mediating monocyte adherence to FN. In the present report, we have established and compared different procedures for morphological evaluation of the expression and distribution of the FN receptors on human monocytes in order to investigate their response to endotoxin or cytokine stimulation.

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Development of an in vitro Model for the Study of the Response of Equine Tendon Fibroblasts to Injury and Medication

Veterinary and Comparative Orthopaedics and Traumatology ◽

10.1055/s-0038-1632561 ◽

1997 ◽

Vol 10 (01) ◽

pp. 6-11 ◽

Cited By ~ 3

Author(s):

R. F. Rosenbusch ◽

L. C. Booth ◽

L. A. Dahlgren

Keyword(s):

Electron Microscopy ◽

Extracellular Matrix ◽

Light Microscopy ◽

Light And Electron Microscopy ◽

Tendon Healing ◽

Matrix Proteins ◽

Isolated Cells ◽

Superficial Digital Flexor Tendon ◽

Vitro Model

SummaryEquine tendon fibroblasts were isolated from explants of superficial digital flexor tendon, subcultured and maintained in monolayers. The cells were characterized by light microscopy, electron microscopy and radiolabel studies for proteoglycan production. Two predominant cell morphologies were identified. The cells dedifferentiated toward a more spindle shape with repeated subcultures. Equine tendon fibroblasts were successfully cryopreserved and subsequently subcultured. The ability to produce proteoglycan was preserved.The isolated cells were identified as fibroblasts, based on their characteristic shape by light microscopy and ultrastructure and the active production of extracellular matrix proteins. Abundant rough endoplasmic reticulum and the production of extracellular matrix products demonstrated active protein production and export. Proteoglycans were measurable via liquid scintillation counting in both the cell-associated fraction and free in the supernatant. This model is currently being utilized to study the effects of polysulfated glycosaminoglycan on tendon healing. Future uses include studying the effects of other pharmaceuticals, such as hyaluronic acid, on tendon healing.A model was developed for in vitro investigations into tendon healing. Fibroblasts were isolated from equine superficial digital flexor tendons and maintained in monolayer culture. The tenocytes were characterized via light and electron microscopy. Proteoglycan production was measured, using radio-label techniques. The fibroblasts were cryopreserved and subsequently subcultured. The cells maintained their capacity for proteoglycan production, following repeated subculturing and cryopreservation.

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