The retention and ultrastructural appearances of various extracellular matrix molecules incorporated into three-dimensional hydrated collagen lattices

E. A. Turley, C. A. Erickson, Richard P Tucker

Research output: Contribution to journalArticle

53 Citations (Scopus)

Abstract

Artificial extracellular matrices composed of collagen, glycosaminoglycans (GAG), proteoglycans (PG), plasma fibronectin (FN), and a hyaluronate-binding protein (HABP) have been prepared that morphologically resemble embryonic extracellular matrices in vivo at the light and electron microscope level. The effect of each of the above matrix molecules on the structure and "self-assembly" of these artificial matrices was delineated. (1) Matrix components assembled in vitro morphologically resemble their counterparts in vivo, for the most part. Scanning and transmission electron microscopy indicate that under our assembly and fixation conditions, collagen forms striated fibrils that are 125 nm in diameter, FN forms 30- to 60-nm granules, chondroitin sulfate proteoglycan (CSPG) forms 27- to 37-nm granules, chondroitin sulfate (CS) assembles into 100- to 250-nm spheres, and hyaluronate (HA) appears either as granular mats when fixed with cetylpyridinium chloride (CPC) or as 1.5- to 3-nm microfibrils when preserved with ruthenium red plus tannic acid. These molecules are known to assume the same configurations in embryonic matrices when the same preservation techniques are used with the exception of FN, which generally forms fibrillar arrays. (2) Addition of various matrix molecules can radically change the appearance of the collagen gels. HA greatly expands the volume of the gel and increases the space between collagen fibrils. CSPG at low concentrations (<1 mg/ml) and CS at high concentrations (>20 mg/ml) bundle the collagen fibrils into twisted ropes. (3) A variety of assays were used to examine binding between various matrix components and retention of these components in the hydrated collagen lattices. These assays included solid-phase binding assays, negative staining of spread mixtures of matrix components, cryostat sections of unfixed mixtures of matrix components, and retention of radiolabeled matrix molecules in fixed and washed gels. A number of these binding interactions may play a role in the assembly and stabilization of the matrix. (a) HA, CSPG, and FN bind to collagen. CS appears to only weakly bind to collagen, if at all. (b) FN promotes the increased retention of HA, CSPG, and to a very small degree, CS, in collagen gels. Conversely, the GAG increase the retention of 3H-FN in the gels. Furthermore, FN binds to HA, CS, and CSPG as demonstrated by solid surface binding assays and morphological criteria. The increased retention of GAG and CSPG by the addition of FN may be due to both stabilization of binding to the collagen and trapping of matrix complexes within the gel. (c) HA binds to both CS and CSPG. Its association with CSPG monomers produces the typical "bead-on-a-string" appearance that has been well characterized in vivo. HA association with CS suggests yet another way that matrix can be stabilized. (d) A 70-kDa protein that has been shown to bind to both HA and collagen increases the retention of HA in the collagen gels and promotes HA fibril formation. Fibronectin, GAG, and CSPG spontaneously organize in the collagen matrices in a manner that is consistent both with binding characteristics that have been previously published and those that are reported here.

Original languageEnglish (US)
Pages (from-to)347-369
Number of pages23
JournalDevelopmental Biology
Volume109
Issue number2
DOIs
StatePublished - 1985

Fingerprint

Chondroitin Sulfate Proteoglycans
Extracellular Matrix
Collagen
Fibronectins
Chondroitin Sulfates
Gels
Glycosaminoglycans
Cetylpyridinium
Microfibrils
Ruthenium Red
Negative Staining
Scanning Transmission Electron Microscopy
Tannins
Proteoglycans
Carrier Proteins
Electrons
Light

ASJC Scopus subject areas

  • Developmental Biology

Cite this

@article{a8294f241cf24896a397b14f08120331,
title = "The retention and ultrastructural appearances of various extracellular matrix molecules incorporated into three-dimensional hydrated collagen lattices",
abstract = "Artificial extracellular matrices composed of collagen, glycosaminoglycans (GAG), proteoglycans (PG), plasma fibronectin (FN), and a hyaluronate-binding protein (HABP) have been prepared that morphologically resemble embryonic extracellular matrices in vivo at the light and electron microscope level. The effect of each of the above matrix molecules on the structure and {"}self-assembly{"} of these artificial matrices was delineated. (1) Matrix components assembled in vitro morphologically resemble their counterparts in vivo, for the most part. Scanning and transmission electron microscopy indicate that under our assembly and fixation conditions, collagen forms striated fibrils that are 125 nm in diameter, FN forms 30- to 60-nm granules, chondroitin sulfate proteoglycan (CSPG) forms 27- to 37-nm granules, chondroitin sulfate (CS) assembles into 100- to 250-nm spheres, and hyaluronate (HA) appears either as granular mats when fixed with cetylpyridinium chloride (CPC) or as 1.5- to 3-nm microfibrils when preserved with ruthenium red plus tannic acid. These molecules are known to assume the same configurations in embryonic matrices when the same preservation techniques are used with the exception of FN, which generally forms fibrillar arrays. (2) Addition of various matrix molecules can radically change the appearance of the collagen gels. HA greatly expands the volume of the gel and increases the space between collagen fibrils. CSPG at low concentrations (<1 mg/ml) and CS at high concentrations (>20 mg/ml) bundle the collagen fibrils into twisted ropes. (3) A variety of assays were used to examine binding between various matrix components and retention of these components in the hydrated collagen lattices. These assays included solid-phase binding assays, negative staining of spread mixtures of matrix components, cryostat sections of unfixed mixtures of matrix components, and retention of radiolabeled matrix molecules in fixed and washed gels. A number of these binding interactions may play a role in the assembly and stabilization of the matrix. (a) HA, CSPG, and FN bind to collagen. CS appears to only weakly bind to collagen, if at all. (b) FN promotes the increased retention of HA, CSPG, and to a very small degree, CS, in collagen gels. Conversely, the GAG increase the retention of 3H-FN in the gels. Furthermore, FN binds to HA, CS, and CSPG as demonstrated by solid surface binding assays and morphological criteria. The increased retention of GAG and CSPG by the addition of FN may be due to both stabilization of binding to the collagen and trapping of matrix complexes within the gel. (c) HA binds to both CS and CSPG. Its association with CSPG monomers produces the typical {"}bead-on-a-string{"} appearance that has been well characterized in vivo. HA association with CS suggests yet another way that matrix can be stabilized. (d) A 70-kDa protein that has been shown to bind to both HA and collagen increases the retention of HA in the collagen gels and promotes HA fibril formation. Fibronectin, GAG, and CSPG spontaneously organize in the collagen matrices in a manner that is consistent both with binding characteristics that have been previously published and those that are reported here.",
author = "Turley, {E. A.} and Erickson, {C. A.} and Tucker, {Richard P}",
year = "1985",
doi = "10.1016/0012-1606(85)90461-0",
language = "English (US)",
volume = "109",
pages = "347--369",
journal = "Developmental Biology",
issn = "0012-1606",
publisher = "Academic Press Inc.",
number = "2",

}

TY - JOUR

T1 - The retention and ultrastructural appearances of various extracellular matrix molecules incorporated into three-dimensional hydrated collagen lattices

AU - Turley, E. A.

AU - Erickson, C. A.

AU - Tucker, Richard P

PY - 1985

Y1 - 1985

N2 - Artificial extracellular matrices composed of collagen, glycosaminoglycans (GAG), proteoglycans (PG), plasma fibronectin (FN), and a hyaluronate-binding protein (HABP) have been prepared that morphologically resemble embryonic extracellular matrices in vivo at the light and electron microscope level. The effect of each of the above matrix molecules on the structure and "self-assembly" of these artificial matrices was delineated. (1) Matrix components assembled in vitro morphologically resemble their counterparts in vivo, for the most part. Scanning and transmission electron microscopy indicate that under our assembly and fixation conditions, collagen forms striated fibrils that are 125 nm in diameter, FN forms 30- to 60-nm granules, chondroitin sulfate proteoglycan (CSPG) forms 27- to 37-nm granules, chondroitin sulfate (CS) assembles into 100- to 250-nm spheres, and hyaluronate (HA) appears either as granular mats when fixed with cetylpyridinium chloride (CPC) or as 1.5- to 3-nm microfibrils when preserved with ruthenium red plus tannic acid. These molecules are known to assume the same configurations in embryonic matrices when the same preservation techniques are used with the exception of FN, which generally forms fibrillar arrays. (2) Addition of various matrix molecules can radically change the appearance of the collagen gels. HA greatly expands the volume of the gel and increases the space between collagen fibrils. CSPG at low concentrations (<1 mg/ml) and CS at high concentrations (>20 mg/ml) bundle the collagen fibrils into twisted ropes. (3) A variety of assays were used to examine binding between various matrix components and retention of these components in the hydrated collagen lattices. These assays included solid-phase binding assays, negative staining of spread mixtures of matrix components, cryostat sections of unfixed mixtures of matrix components, and retention of radiolabeled matrix molecules in fixed and washed gels. A number of these binding interactions may play a role in the assembly and stabilization of the matrix. (a) HA, CSPG, and FN bind to collagen. CS appears to only weakly bind to collagen, if at all. (b) FN promotes the increased retention of HA, CSPG, and to a very small degree, CS, in collagen gels. Conversely, the GAG increase the retention of 3H-FN in the gels. Furthermore, FN binds to HA, CS, and CSPG as demonstrated by solid surface binding assays and morphological criteria. The increased retention of GAG and CSPG by the addition of FN may be due to both stabilization of binding to the collagen and trapping of matrix complexes within the gel. (c) HA binds to both CS and CSPG. Its association with CSPG monomers produces the typical "bead-on-a-string" appearance that has been well characterized in vivo. HA association with CS suggests yet another way that matrix can be stabilized. (d) A 70-kDa protein that has been shown to bind to both HA and collagen increases the retention of HA in the collagen gels and promotes HA fibril formation. Fibronectin, GAG, and CSPG spontaneously organize in the collagen matrices in a manner that is consistent both with binding characteristics that have been previously published and those that are reported here.

AB - Artificial extracellular matrices composed of collagen, glycosaminoglycans (GAG), proteoglycans (PG), plasma fibronectin (FN), and a hyaluronate-binding protein (HABP) have been prepared that morphologically resemble embryonic extracellular matrices in vivo at the light and electron microscope level. The effect of each of the above matrix molecules on the structure and "self-assembly" of these artificial matrices was delineated. (1) Matrix components assembled in vitro morphologically resemble their counterparts in vivo, for the most part. Scanning and transmission electron microscopy indicate that under our assembly and fixation conditions, collagen forms striated fibrils that are 125 nm in diameter, FN forms 30- to 60-nm granules, chondroitin sulfate proteoglycan (CSPG) forms 27- to 37-nm granules, chondroitin sulfate (CS) assembles into 100- to 250-nm spheres, and hyaluronate (HA) appears either as granular mats when fixed with cetylpyridinium chloride (CPC) or as 1.5- to 3-nm microfibrils when preserved with ruthenium red plus tannic acid. These molecules are known to assume the same configurations in embryonic matrices when the same preservation techniques are used with the exception of FN, which generally forms fibrillar arrays. (2) Addition of various matrix molecules can radically change the appearance of the collagen gels. HA greatly expands the volume of the gel and increases the space between collagen fibrils. CSPG at low concentrations (<1 mg/ml) and CS at high concentrations (>20 mg/ml) bundle the collagen fibrils into twisted ropes. (3) A variety of assays were used to examine binding between various matrix components and retention of these components in the hydrated collagen lattices. These assays included solid-phase binding assays, negative staining of spread mixtures of matrix components, cryostat sections of unfixed mixtures of matrix components, and retention of radiolabeled matrix molecules in fixed and washed gels. A number of these binding interactions may play a role in the assembly and stabilization of the matrix. (a) HA, CSPG, and FN bind to collagen. CS appears to only weakly bind to collagen, if at all. (b) FN promotes the increased retention of HA, CSPG, and to a very small degree, CS, in collagen gels. Conversely, the GAG increase the retention of 3H-FN in the gels. Furthermore, FN binds to HA, CS, and CSPG as demonstrated by solid surface binding assays and morphological criteria. The increased retention of GAG and CSPG by the addition of FN may be due to both stabilization of binding to the collagen and trapping of matrix complexes within the gel. (c) HA binds to both CS and CSPG. Its association with CSPG monomers produces the typical "bead-on-a-string" appearance that has been well characterized in vivo. HA association with CS suggests yet another way that matrix can be stabilized. (d) A 70-kDa protein that has been shown to bind to both HA and collagen increases the retention of HA in the collagen gels and promotes HA fibril formation. Fibronectin, GAG, and CSPG spontaneously organize in the collagen matrices in a manner that is consistent both with binding characteristics that have been previously published and those that are reported here.

UR - http://www.scopus.com/inward/record.url?scp=0022081392&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0022081392&partnerID=8YFLogxK

U2 - 10.1016/0012-1606(85)90461-0

DO - 10.1016/0012-1606(85)90461-0

M3 - Article

VL - 109

SP - 347

EP - 369

JO - Developmental Biology

JF - Developmental Biology

SN - 0012-1606

IS - 2

ER -