Stimulation of glycosaminoglycan production in murine tumors

Abstract

Three types of murine tumors, B-16 melanoma, A-10 carcinoma, and S-180 sarcoma, were shown to contain elevated glycosaminoglycan (GAG) concentrations in vivo as compared to normal muscle or subcutaneous tissue. Hyaluronate was especially concentrated in the A-10 carcinoma, which contained approximately six times more hyaluronate than subcutaneous tissue and 18 times more than muscle. In all three tumors, chondroitin sulfates, especially chondroitin-4-sulfate, were present in higher concentrations than in the normal tissues. In culture, however, all three tumor cell lines produced less than 5% as much GAG as mouse fibroblasts, when measured by incorporation of [3H] acetate or by chemical analysis. Varying the culture passage number or the medium composition, ie, glucose, serum, and insulin concentrations, had little effect on GAG synthesis by the tumor cells. The low GAG levels in the tumor cell cultures were not due to hyaluronidase activity in their media. In an attempt to mimic possible host-tumor cell interactions that could account for the elevated GAG levels in vivo, tumor cells were cocultured with fibroblasts, but no stimulation above the amount made by the tumor cells alone plus that by the fibroblasts alone was observed. Conditioned media from the tumor cells, either dialyzed or not against fresh complete medium, had no effect on fibroblast GAG synthesis. Tumor extracts, however, were found to stimulate synthesis of hyaluronate by fibroblasts. Stimulation by extracts of A-10 carcinoma was greater than and additive to that of serum. The above results strongly suggest that GAG production in these tumors is in part regulated by host-tumor interactions.

Epidermal Growth Factor Induces CD44 Gene Expression through a Novel Regulatory Element in Mouse Fibroblasts

“…In the EGF induction CAT assays, transfected cells were equally divided, allowed to recover in complete medium for 24 h, and then treated with or without 10 nM EGF for an additional 24 h in 0.1% dialyzed fetal bovine serum containing medium. CAT activity was measured from cell extracts by the CAT Enzyme Assay System (Promega) based on liquid scintillation counting using 14 C-labeled chloramphenicol (Amersham) and n-butynyl coenzyme A. The level of CAT activity from pRVBCAT transfectants without EGF induction was set at 100% and the percent (fold) increase in the other transfectants determined and compared with the standard.…”

“…Among the regulated genes are those encoding cell surface adhesion molecules (8 -10) as well as nuclear transcriptional factors that regulate gene expression. Cell surface glycoprotein CD44, an adhesion molecule that binds to the extracellular matrix components, hyaluronic acid (11) and osteopontin (12), has been postulated to modulate tumor invasion and metastasis (13)(14)(15). Our previous studies with mouse 3T3 fibroblasts demonstrated that EGF induced CD44 gene expression on both the protein and mRNA levels and that this was accompanied by enhanced cell attachment to the extracellular matrix component hyaluronic acid (16).…”

Layilin, a Novel Integral Membrane Protein, Is a Hyaluronan Receptor

“…Because tumors are frequently characterized by local accumulation of HA in vivo (Toole et al, 1979;Knudson et al, 1984), we stained sections from pancreata of RIP1-Tag2 transgenic mice, which spontaneously develop pancreatic tumors. Incubation of frozen sections from the pancreata of RIPTag2 mice with layilin-Fc showed positive staining of the ECM of the tumors ( Figure 5B), whereas no staining was observed with the E-cadherin-Fc control ( Figure 5A).…”

“…Because tumors are frequently characterized by local accumulation of HA in vivo (Toole et al, 1979;Knudson et al, 1984), we stained sections from pancreata of RIP1-Tag2 transgenic mice, which spontaneously develop pancreatic tumors.…”

Kunitz-type Protease Inhibitor Bikunin Disrupts Phorbol Ester-induced Oligomerization of CD44 Variant Isoforms Containing Epitope v9 and Subsequently Suppresses Expression of Urokinase-type Plasminogen Activator in Human Chondrosarcoma Cells

“…Since one of the bik-binding proteins is identical to LP, which is apparently held at the cell surface by HA (19), it is reasonable to think that CD44 would be involved in this bik-mediated complex at the cell surface. It has been established that CD44 is a polymorphic integral membrane glycoprotein with a postulated role in matrix adhesion (23), lymphocyte activation (24 -26), lymph node homing (27), and tumor invasion (28,29) and metastasis (30). CD44 plays an important role in the physiology of normal and tumor cells (31,32).…”