ABSTRACT
A small chondroitin sulfate proteoglycan was isolated and partially characterized for core protein and glycosaminoglycan structures from the mucus surrounding embryos in the developmental pouch of Viviparus ater (Mollusca Gastropoda). The protein bearing polysaccharide nature was confirmed by gel-permeation chromatography separation of fractions positive to the uronic acid dosage, 7.5% SDS-PAGE under reducing conditions, sequential staining with alcian blue and ammoniacal silver. Its molecular mass was calculated at about 228,800. After degradation of the galactosaminoglycan components by chondroitinase ABC in the presence of proteinase inhibitors, the molecular mass of the core protein was determined at about 72,200. Treatment of the proteoglycan with keratanase did not modify its electrophoretic migration. Isoelectric focusing of the core protein demonstrated a micro-heterogeneity with the presence of two isoforms with different isoelectric point, pI=8.2 and 6.6, in a ratio of about 1:2.2. The glycosaminoglycan component of the proteoglycan was characterized as chondroitin sulfate with a molecular mass of about 30,750 composed of 5% non-sulfated unsaturated disaccharide, 94% monosulfated disaccharides (4-monosulfated to 6-monosulfated disaccharide ratio of 1.36) and 1. 5% disulfated disaccharides (in particular 1.3% 2,6-disulfated disaccharide) with a sulfate to carboxyl ratio of 0.96. Degradation of the chondroitin sulfate with chondroitinase ABC and ACII permitted to determine a percentage of glucuronic acid of about 78.4. The proteoglycan isolated from the mucus surrounding the embryos of Viviparus ater is formed by a small core protein bearing about five chondroitin sulfate chains (80% chondroitin sulfate/20% dermatan sulfate) with potential function in the developmental processes of molluscs embryos.
Subject(s)
Chondroitin Sulfate Proteoglycans/chemistry , Chondroitin Sulfate Proteoglycans/isolation & purification , Embryo, Nonmammalian/chemistry , Glycoside Hydrolases , Mollusca/chemistry , Mucus/chemistry , Animals , Chondroitin Sulfate Proteoglycans/analysis , Chondroitinases and Chondroitin Lyases/metabolism , Chromatography, High Pressure Liquid , Electrophoresis, Polyacrylamide Gel , Histocytochemistry , Hydrogen-Ion Concentration , Isoelectric Focusing , Molecular Weight , Periodic Acid-Schiff Reaction , Sodium Dodecyl Sulfate , beta-Galactosidase/metabolismABSTRACT
Cultures of isolated neuronal populations from the central ganglia of the gastropod mollusc Planorbarius corneus were used for testing the effects of inorganic lead. The examined parameters were cell survival, neurite outgrowth and cytoskeletal morphology. In large heterogeneous neuronal populations as obtained from a whole cerebral or pedal ganglion, the different sensitivity to lead is reflected mainly on the cell survival. The neurons belonging to the homogeneous E cluster population are more sensitive; in fact a higher percentage of them do not survive in the presence of lead. Moreover, in this neuronal cell type the neurite outgrowth is dramatically affected by lead only when the neurons are cultured on conditioned substrate. Possibly, membrane mechanisms activated for the neurite outgrowth represent a target for inorganic lead. The few neurites sprouted in presence of lead do not evidence changes in the cytoskeletal components.
