Choriocarcinoma - a rare association with squamous cell carcinoma of esophagus.

Extragenital choriocarcinoma involving the gastrointestinal tract is rare. We report a 60-year-old woman with squamous cell carcinoma of esophagus with a choriocarcinomatous focus. She was palliated with chemotherapy and an endoprosthesis.

Immunological detection of type II collagen degradation: use in the evaluation of anti-arthritic therapies.

Propagated Swaram rat chondrosarcoma cells, rabbit chrondrocytes (from articular cartilage of knee, shoulder and hip joints), and bovine nasal cartilage explant cultures were studied. Type II collagen (CII) and its peptide fragments were quantitated in cell medium and cell layer separately, using two previously developed assays; one assay employed a monoclonal antibody, C4F6, that reacts specifically with triple helical CII and the other assay used an antibody, EIE5, that reacts specifically with a peptide of CII. A time-dependent increase in the content of CII and CII-derived peptides was observed in both rat and rabbit cultures. In both culture systems the majority of the native type II collagen is found associated with the cell layer (97% in rat cultures and 73% in rabbit cultures), while the major part of the CII peptides is found in the media (73% in rat cultures, 88% in the rabbit cultures). The concentration of peptides in the media reaches approximately 2 micrograms mL-1 in both chondrocyte monolayer cultures after 4 days. The CII peptide assay employing E1E5 was well suited to quantitate articular cartilage collagen degradation in explant culture. thus it can be used to evaluate potential therapeutic agents that can modify or inhibit cartilage degradation. The assay has the added potential that it could be used in-vivo to evaluate the effectiveness of potential metalloproteinase inhibitors in animal models of osteoarthritis or in clinical trials.

Effects of certain antiarthritic agents on the synthesis of type II collagen and glycosaminoglycans in rat chondrosarcoma cultures.

Cartilage destruction is a characteristic feature of osteoarthritis. Treatment with certain nonsteroidal anti-inflammatory drugs could exacerbate cartilage destruction by impairing the synthesis of cartilage matrix proteins, type II collagen and proteoglycan. In order to monitor the changes occurring in cartilage collagen synthesis, we developed a type II collagen specific ELISA. The effects of antiarthritic agents on type II collagen and glycosaminoglycan synthesis were examined in rat chondrosarcoma cultures. Drugs were added to the monolayer cultures and 4 days later the total type II collagen, as determined by the type II collagen ELISA, and glycosaminoglycan content, as measured by dimethyl-methylene blue dye binding assay, was measured. All drugs except tiaprofenic acid decreased type II collagen synthesis by at least 40% at 100 micrograms/ml. Tiaprofenic acid at 1 microgram/ml increased type II collagen content by 54% of the controls. Glycosaminoglycan synthesis was decreased by acetylsalicylic acid, diclofenac and tiaprofenac acid, at 50 micrograms/ml or above. Indomethacin, naproxen and dexamethasone had no effect. Interestingly, tenidap stimulated the glycosaminoglycan synthesis by 32% at 100 micrograms/ml. We show that the combination of chondrosarcoma cultures, type II collagen specific ELISA and dimethylmethylene blue dye binding assay serves as a useful model for screening the effects of agents capable of modulating type II collagen and glycosaminoglycan synthesis.

Production of type II collagen specific monoclonal antibodies.

Immunoassays are used for the specific measurement of type II collagen, a major cartilage protein, which is lost in osteoarthritic joints. Poor immunogenicity and species dependent immune response to type II collagen make it difficult to obtain specific antibodies required for immunoassay development. In addition, type II collagen antibodies exhibit reactivity to structurally dissimilar antigens such as actin, myoglobin, thyroglobulin and ssDNA, complicating the isolation of specific antibodies. It is therefore necessary to characterize the antibody reactivity against both noncollagenous antigens and different collagen types. In this study, immune response to type II collagen was improved by conjugation to carrier proteins, KLH and BSA. Hybridomas were generated by fusions of lymphocytes derived from lymph nodes or spleens with X63-653-Ag8 myeloma cells. Compared to spleens, the utilization of lymph nodes as a source of lymphocytes resulted in a 23% higher number of hybridomas secreting type II collagen antibodies. Hybridomas secreting polyreactive antibodies were identified based on their reactivity to thyroglobulin and eliminated. Extensive testing of the remaining monoclonal antibodies with other structurally dissimilar antigens and various types of collagen for reactivity, allowed us to isolate specific monoclonal antibodies to type II collagen. We emphasize the importance of characterization of the reactivity of type II collagen monoclonal antibodies before employing them for immunoassays.

Quantitative immunoassays for type II collagen and its cyanogen bromide peptides.

This paper describes the development of quantitative immunoassays utilizing mouse monoclonal antibodies which are monospecific to type II collagen. The monoclonal antibodies were characterized and tested extensively for reactivity against a panel of antigens including actin, myoglobin, thyroglobulin, ssDNA, tetanus toxin and different types of collagens including their CnBr-derived peptides. Four monoclonal antibodies having strict monospecificity to type II collagen were selected. Quantitative immunoassays developed with these antibodies can measure either type II collagen in its native conformation or type II collagen-derived cyanogen bromide peptides. The assay conditions such as coating concentration of antigen, monoclonal antibody concentration, second antibody concentration and incubation times were optimized to obtain maximum possible sensitivity. These quantitative immunoassays can be employed to measure type II collagen or type II collagen-derived peptides in low amounts ranging from 20 to 100 ng/ml. The assays can be applied to chondrocyte cultures without interference from serum components or other collagen types.

Evidence for polyreactivity seen with monoclonal antibodies produced against type II collagen.

Specific type II collagen monoclonal antibodies are needed for the quantification of articular cartilage collagen. In this study we produced and characterized 29 type II collagen monoclonal antibodies. Hybridomas were generated from mice immunized with rat type II collagen, selected for high antibody production against type II collagen using ELISA. Antibodies from selected and cloned hybridoma cells were purified by affinity chromatography and their reactivity tested by ELISA against a panel of antigens including actin, thyroglobulin, and single stranded DNA, all of which have been used to characterize the 'naturally occurring antibodies.' It was found that many of the anti-type II collagen monoclonal antibodies reacted to more than one antigen. The monospecific antibodies had higher affinity to type II collagen than the antibodies which demonstrated multireactivity. Because of the prevalence of polyreactive anti-type II collagen antibodies, it is advisable to employ highly selective methodologies to isolate high affinity monospecific antibodies to type II collagen.