Correlative morphometric and biochemical analysis of purified extracellular matrix vesicles from rat alveolar bone

Abstract

Extracellular matrix vesicles were isolated from rat alveolar bone and purified by either gel filtration or discontinuous sucrose density gradient. Morphometric evaluation of electron-micrographs of pellets of purified and nonpurified vesicle fractions was correlated with the activity of vesicular enzymes. A high correlation was found between the percentage of area occupied by vesicles with electron-dense content (electron-dense vesicle fractional area) and the enzymatic activity. Highest enzymatic specific activities and electron-dense vesicle fractional area were recorded in the "light" vesicle-enriched fraction obtained after equilibrium density centrifugation. These parameters revealed lowest values in the "heavy" vesicle-enriched fraction resolved by the same methods. The combined electron-dense and electron-lucent fractional area (vesicular fractional area) was similar in the different purified fractions. It is therefore suggested that the fraction obtained by gel filtration contains both "light" and "heavy" vesicles. Morphometric study is proposed as an additional criterion for the degree of purification of matrix vesicle preparations.

Role of lipids in calcification of cartilage

“…, growth cartilage (Anderson and Sajdera, 19761, and dentin (Biggerstaff et al, 1983), and during osseous induction and hard tissue repair (Bab et al, 1983;Boskey et al, 1980a). The classic morphologic demonstration of this close association of membrane and mineral is in electron micrographs of growth cartilage.…”

Primary Mineralization At the Surfaces of Implants

“…Later, hyaluronidase treatment was abandoned, and the amount of collagenase reduced with increased incubation time (Wuthier, 1982;Ali, 1983). In addition, tissue homogenization and centrifugation in a variety of density gradient materials were introduced (Deutsch et al, 1981;Wuthier, 1982;Bab et al, 1983). At present, the one method consisting of collagenase digestion and differential centrifugation of the collagenase-digest supernatant after pelleting of the cells (Ali and Evans, 1973;Ali, 1976) is the one that is most commonly used.…”

Calcifying Matrices: Cartilage

“…Similar results have been found in dentin, where matrix vesicles show no direct contact with odontoblasts or their processes (Katchburian 1973a). The fact that matrix vesicles can be isolated by matrix collagenase digestion, cell pellet removal, and differential centrifugation, or else by matrix homogenization and density gradient fractionation, both from cartilage (Ali 1976;Ali and Evans 1973;Ali et al 1970Ali et al , 1971Wuthier et al 1978Wuthier et al , 1985Watkins et al 1980;Warner et al 1983;Kakuta et al 1985) and from bone (Deutsch D et al 1981;Bab et al 1983) and dentin (Slavkin et al 1972), further confirms that they are true matrix bodies, without any direct link with cells.…”

“…This could explain the often reported heterogeneity of matrix vesicles. Two fractions of matrix vesicles have been isolated by means of the equilibrium density fractionation method: a heavy fraction, rich in enzymatic activities, and a light fraction (Bab et al 1983). Two types of matrix vesicles have been described too, on the basis of their ultrastructure and enzymatic properties: a rare type I, comprising vesicles with lysosome-like properties, and a frequent type II, showing alkaline phosphatase activity (Thyberg 1972;Thyberg et al 1975).…”