tretinoin and batimastat

To raise peptide antibodies recognizing the C-terminal amino acid sequence in the G1 domain of porcine aggrecan, generated by the action of either aggrecanase or neutral metalloproteinase(s), in rabbits and to use them to investigate the release of aggrecan from porcine articular cartilage.. An explant culture system was used to investigate the release of the G1 domain of aggrecan from porcine articular cartilage treated with retinoic acid or interleukin 1beta and to study how the activity of these agents is modified by the proteinase inhibitor, batimastat (BB94).. Retinoic acid and interleukin 1beta induced both enzyme activities and the release of the G1 domain into the culture medium. Proteinase activity was significantly reduced when the tissue was incubated in the presence of BB94. The functional properties of the enzyme-generated G1 domain were studied using large-pore, agarose/polyacrylamide gel electrophoresis, and it was shown to interact with hyaluronan and link protein.. The results show that there must be a mechanism for removing a functional G1 domain from aggrecan during tissue turnover using this culture system.

Topics: Aggrecans; Animals; Antibody Specificity; Cartilage, Articular; Culture Media, Conditioned; Culture Techniques; Endopeptidases; Extracellular Matrix Proteins; Glycosaminoglycans; Hyaluronic Acid; Immune Sera; Interleukin-1; Lectins, C-Type; Metalloendopeptidases; Peptide Fragments; Phenylalanine; Protease Inhibitors; Proteoglycans; Rabbits; Swine; Thiophenes; Tretinoin

In myeloid leukemia, immature leukemic cells are able to egress into peripheral blood to infiltrate extra-medullary organs. We therefore analyzed the migrating and invasive potential of human HL-60 and NB4 cell lines, representative of acute myelogenous leukemia, their ability to express matrix metalloproteases (MMPs), tissue inhibitors of metalloproteases (TIMPs) and urokinase plasminogen activator (uPA) in response to differentiating agents. Granulocytic differentiation by all-trans-retinoic acid (ATRA) and aclacinomycin (ACLA) strongly increased HL-60 and NB4 cell migration and invasion. At mRNA and protein levels, these cell lines produced significant amounts of MMP-9 (HL-60

Topics: Aclarubicin; Antineoplastic Agents; Cell Differentiation; Cell Movement; HL-60 Cells; Humans; Leukemia; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Phenylalanine; Serine Endopeptidases; Thiophenes; Tissue Inhibitor of Metalloproteinases; Tretinoin; Tumor Cells, Cultured

All-trans-retinoic acid (RA) is a potent differentiating agent that is very effective in the treatment of patients with acute promyelocytic leukemia (APL). Since clinical response can be accompanied by extramedullary manifestations, we have investigated the influence of RA on cell adhesion to and migration through reconstituted basement membranes (Matrigel) in the APL cell line NB4. No apparent cellular differentiation was observed during a 24-h incubation with 1 microM RA, as indicated by the nitroblue tetrazolium reduction test. However, exposure to RA significantly enhanced NB4 cell adhesion to Matrigel and consecutive migration through Matrigel barriers in a dose-dependent manner. Several integrin molecules potentially involved in this process, i.e., CD29, CD18, CD11a, CD11b and CD11c, were therefore studied by fluorescence-activated cell sorting analysis. The expression of the beta subunit of the beta2 integrins (CD18), but not that of beta1 integrins (CD29), was increased during 24-h RA treatment. Among the beta2 integrins, the expression of LFA-1 (CD11a) and of Mac-1 (CD11b), but not of p150,95 (CD11c), was induced by RA. When monoclonal antibodies that specifically block the interaction of these integrins with their ligands were used, we observed that CD29 is only involved in adhesion and CD11b only in migration, whereas CD11a participates in both processes. NB4 cells constitutively secreted the matrix metalloproteinases MMP-9 and MMP-2, which are known to promote cellular invasion processes by degradation of the extracellular matrix. RA treatment had no influence on the quantity of secreted MMP-9 or MMP-2 in these cells as determined by zymography. Addition of Batimastat (BB-94), a synthetic inhibitor of matrix metalloproteinases, blocked RA-induced cell migration without affecting cellular adhesion to Matrigel. These findings indicate that adhesion molecules as well as matrix metalloproteinases are involved in RA-stimulated migration of NB4 cells through Matrigel, possibly providing some explanation of tissue infiltration by leukemic cells as observed during treatment of APL patients with RA.

Topics: Antibodies, Monoclonal; Antineoplastic Agents; Cell Adhesion; Cell Line; Cell Movement; Cell Separation; Collagen; Dose-Response Relationship, Drug; Drug Combinations; Extracellular Matrix; Flow Cytometry; Fluorescence; Gelatinases; Humans; Integrins; Laminin; Leukemia, Promyelocytic, Acute; Matrix Metalloproteinase Inhibitors; Phenylalanine; Proteoglycans; Thiophenes; Time Factors; Tretinoin

To investigate mechanisms of cartilage matrix destruction by a study of the effects of a specific inactivator of cathepsin B and an inhibitor of several matrix metalloproteinases (MMP) on cartilage proteoglycan release.. Cartilage explants were treated with either recombinant human interleukin-1 alpha (rHuIL-1 alpha) or retinoic acid in the presence or absence of the inhibitors, and proteoglycan release was quantitated. Tests for nonspecific effects of the inhibitors included reversibility, rates of protein synthesis and glycolysis, and effects on other rHuIL-1 alpha-mediated events.. The cathepsin B inactivator inhibited rHuIL-1 alpha-stimulated proteoglycan release at nanomolar concentrations, but failed to significantly inhibit retinoic acid-stimulated proteoglycan release. An inhibitor of MMP was inhibitory to both rHuIL-1 alpha-stimulated release and retinoic acid-stimulated release.. Cathepsin B is implicated in rHuIL-1 alpha-stimulated loss of cartilage proteoglycan. Its lack of involvement in retinoic acid-stimulated proteoglycan release suggests the existence of at least 2 pathways of cartilage proteoglycan breakdown, which may converge at the activation of a matrix prometalloproteinase.

Topics: Aggrecans; Amino Acid Sequence; Animals; Cartilage; Cartilage, Articular; Cathepsin B; Cattle; Dipeptides; Enzyme Activation; Extracellular Matrix Proteins; Interleukin-1; Lectins, C-Type; Metalloendopeptidases; Molecular Sequence Data; Nasal Septum; Peptide Fragments; Phenylalanine; Proteoglycans; Recombinant Proteins; Thiophenes; Tretinoin