Methodology is described for the culture of avian and mammalian chondrocytes in ionotrophically gelled "semi-solid" and "hollow" alginate beads. Chondrocytes grown in "semi-solid" gels exhibited a spherical shape as opposed to a fibroblastic morphology observed in monolayer culture. In the "semi-solid" beads, the cells grew as small clumps and as large aggregates. The aggregates were round or elliptical in appearance and surrounded by a dense Alcian Blue positive halo. Preliminary studies with collagen and chitosan matrixes encapsulated in "hollow" beads suggest that cell growth and morphology are profoundly influenced by the composition of the cellular environment. Chondrocyte structure and function in the "semi-solid" and "hollow" beads were partially characterized by light microscopy, histochemical and biochemical means. The encapsulation methodology is readily applicable for the culture of chondrocytes in single beads, in multiwell dishes, or mass culture.

A receptor that binds the phosphomannosyl recognition marker of bovine testicular P-galactosidase (13-D-galactoside galactohydrolase, EC 3.2.1.23) was isolated from bovine liver membranes. The receptor was extracted from crude plasma membrane preparations with Triton X-100 and immunoprecipitated as a receptor-f-galactosidase complex with anti-,B-galactosidase. The receptor was dissociated from the precipitate with mannose 6-phosphate, labeled with`RI, and purified on a (3-galactosidase-Sepharose 4B affinity matrix. A quantitative binding assay employing anti-(-galactosidase and IgGsorb (formalin-fixed Staphylococcus aureus) was devised to study the binding of`5I-labeled receptor to (3-galactosidase. Maximal binding of receptor to enzyme occurred at pH values between 5.7 and 6.5. Divalent cations were not required for binding. The values of the dissociation constant obtained for 83-galactosidase varied between 200 nM observed with "lower uptake" forms and 20 nM for "higher uptake" forms of the enzyme. A number of phosphorylated monosaccharides were tested as inhibitors of binding of enzyme to receptor; mannose 6-phosphate and fructose 1-phosphate served as inhibitors and exhibited 1I values of 0.064 mM and 0.24 mM, respectively. The receptor has a subunit molecular weight of 215,000. Similar receptors were also demonstrated in Triton X-100 extracts of human skin fibroblasts, Chinese hamster ovary cells, and rat hepatocytes. These cell types are known to assimilate lysosomal enzymes containing covalently bound mannose 6-phosphate residues.Certain lysosomal enzymes are selectively and efficiently taken up by cultured human fibroblasts (1). This process is thought to be mediated by a specific cell surface receptor that recognizes phosphomannosyl residues on oligosaccharide chains of the enzymes (2-5). Recognition of lysosomal enzymes by a phosphomannosyl receptor has been proposed as an essential step for the delivery of newly synthesized lysosomal enzymes to lysosomes (6-8). Direct evidence for the existence of phosphomannosyl receptors has been obtained by demonstration of the reversible binding of a-L-iduronidase to the cell surface ofhuman skin fibroblasts (9) and by the binding of P3-glucuronidase to fibroblast cell membranes (10).Phosphomannosyl receptors also occur in other mammalian cell types and tissues. Phosphomannosyl-dependent uptake or binding of lysosomal enzymes has been observed in Chinese hamster ovary cells (11), normal rat kidney cells (12), a rat liver epithelial cell line (13), rat hepatocytes (12,14), and Swarm rat chondrosarcoma (15). Binding studies using f-hexosaminidase suggest the presence of the phosphomannosyl receptor in all major tissues of the rat and in several rat liver subcellular fractions (16).In previous studies we demonstrated the presence of mannose 6-phosphate residues in f3-galactosidase (P-D-galactoside galactohydrolase, EC 3.2.1.23) (17,18) and showed that the enzyme is subject to endocytosis by the phosphomannosyl uptake system in human skin fibroblasts (5). We ...

Vaccines are among the most efficacious and cost-effective tools for reducing morbidity and mortality caused by infectious diseases. The vaccine investigation and online information network (VIOLIN) is a web-based central resource, allowing easy curation, comparison and analysis of vaccine-related research data across various human pathogens (e.g. Haemophilus influenzae, human immunodeficiency virus (HIV) and Plasmodium falciparum) of medical importance and across humans, other natural hosts and laboratory animals. Vaccine-related peer-reviewed literature data have been downloaded into the database from PubMed and are searchable through various literature search programs. Vaccine data are also annotated, edited and submitted to the database through a web-based interactive system that integrates efficient computational literature mining and accurate manual curation. Curated information includes general microbial pathogenesis and host protective immunity, vaccine preparation and characteristics, stimulated host responses after vaccination and protection efficacy after challenge. Vaccine-related pathogen and host genes are also annotated and available for searching through customized BLAST programs. All VIOLIN data are available for download in an eXtensible Markup Language (XML)-based data exchange format. VIOLIN is expected to become a centralized source of vaccine information and to provide investigators in basic and clinical sciences with curated data and bioinformatics tools for vaccine research and development. VIOLIN is publicly available at http://www.violinet.org

The integrative Vaccine Investigation and Online Information Network (VIOLIN) vaccine research database and analysis system (http://www.violinet.org) curates, stores, analyses and integrates various vaccine-associated research data. Since its first publication in NAR in 2008, significant updates have been made. Starting from 211 vaccines annotated at the end of 2007, VIOLIN now includes over 3240 vaccines for 192 infectious diseases and eight noninfectious diseases (e.g. cancers and allergies). Under the umbrella of VIOLIN, >10 relatively independent programs are developed. For example, Protegen stores over 800 protective antigens experimentally proven valid for vaccine development. VirmugenDB annotated over 200 ‘virmugens’, a term coined by us to represent those virulence factor genes that can be mutated to generate successful live attenuated vaccines. Specific patterns were identified from the genes collected in Protegen and VirmugenDB. VIOLIN also includes Vaxign, the first web-based vaccine candidate prediction program based on reverse vaccinology. VIOLIN collects and analyzes different vaccine components including vaccine adjuvants (Vaxjo) and DNA vaccine plasmids (DNAVaxDB). VIOLIN includes licensed human vaccines (Huvax) and veterinary vaccines (Vevax). The Vaccine Ontology is applied to standardize and integrate various data in VIOLIN. VIOLIN also hosts the Ontology of Vaccine Adverse Events (OVAE) that logically represents adverse events associated with licensed human vaccines.

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