Immunosuppression by glucocorticoids: inhibition of production of multiple lymphokines by in vivo administration of dexamethasone.

Glucocorticoids are extremely potent immunosuppressive agents, capable of directly affecting the function of lymphocytes. We studied the effect of in vivo dexamethasone (DEX) administration on anti-CD3-induced lymphocyte proliferation and lymphokine production in mice. To characterize the kinetics and dose responsiveness of lymphocytes to DEX, splenocytes from BALB/c mice that had received a single dose of DEX in vivo were cultured in vitro with suboptimal and optimal concentrations of anti-CD3 monoclonal antibody. Cell proliferation in response to suboptimal concentrations of anti-CD3 was decreased by DEX doses of > or = 30 mg/kg; much higher doses (> or = 200 mg/kg) were required to inhibit cell proliferation in response to optimal anti-CD3 stimulation. Inhibition of suboptimal anti-CD3-stimulated proliferation was evident within 4 hr after DEX administration, was maintained for at least 24 hr, and was no longer evident at 7 and 14 days. Lymphokine secretion induced by optimal doses of anti-CD3 in vitro was differentially affected by in vivo DEX treatment. IL-1 alpha, IL-4, IL-6, IL-10, and IFN-gamma levels were decreased by treatment with low doses of DEX (30 mg/kg), whereas higher doses were required to inhibit production of IL-2, IL-3, and TNF. GM-CSF (granulocyte-macrophage-colony stimulating factor) was least susceptible to DEX inhibition. Low-dose (30 mg/kg) DEX treatment significantly reduced anti-CD3-stimulated production of most lymphokines tested at 4 and 12 hr; by 24 hr the levels of most lymphokines had begun to return to control values. Hence, our data indicate that administration of a single dose of DEX (30 mg/kg for 4 hr) results in significant suppression of lymphokine production and cell proliferation that precedes any significant cell loss and can be used as a reversible model of immunosuppression.

Malignant properties of P635 glioma are independent of glial fibrillary acidic protein expression.

To examine the relationship between the malignant behavior of rat glioma cells and expression of the differentiation antigen glial fibrillary acidic protein (GF), we assayed the tumorigenicity and metastatic ability of P635 and its GF+ or GF- clones. We injected P635 (GF+) and clone 45 (GF-) cells intramuscularly in nude mice. Both lines formed local tumors which metastasized to lungs with equal efficiency. We then injected these lines intravenously in nude mice. Both produced experimental metastases in lungs and other organs with equal efficiency. Finally we injected P635 and several GF+ and GF- clones into the chorioallantoic membrane veins of naturally immune-deficient chick embryos and measured cell growth in embryonic liver. We again found that all clones survived and grew equally well. P635 cells are capable of extracranial growth and metastasis, two important features of the malignant phenotype. We conclude that GF in P635 is a neutral marker with respect to tumorigenicity and metastatic ability.

Comparison of metastatic properties of a variety of mouse, rat, and human cells in assays in nude mice and chick embryos.

We have previously developed an assay to measure experimental metastatic ability of cells following intravenous injection into chorioallantoic membrane (CAM) veins of naturally immune deficient chick embryos. Here we compare metastatic properties of different cell types (ras-transformed and control NIH 3T3, LTA, and 10T1/2; melanoma; and glioma) from several species (mouse, rat, human), using chick embryos and the more commonly-used immune deficient host, nude mice. We found a good correlation between the two assays. Both hosts have advantages and disadvantages in assessing metastatic properties. We conclude that the chick embryo assay is a useful alternative host for experimental metastasis studies. This assay correlates well with and is less costly than assays using nude mice.

ras-transformed NIH 3T3 cell lines, selected for metastatic ability in chick embryos, have increased proportions of p21-expressing cells and are metastatic in nude mice.

We previously reported that ras-transformed NIH 3T3 cells could be selected in vivo for increased metastatic ability after intravenous injection into chick embryos, and that the metastatic cell populations expressed increased levels of ras p21 protein. We have tested the metastatic ability of a series of these cells in nude mice, to determine if their properties in the chick embryo experimental metastasis assay predict their metastatic behavior in nude mice. We report here that cells selected for metastatic ability in the chick embryo are also metastatic when assayed in nude mice. We also asked whether the selected cells uniformly expressed higher levels of p21, using an immunocytochemical procedure. We found that p21 expression among individual cells in these populations was quite heterogeneous. There was a direct relationship between the proportions of p21-expressing cells and metastatic ability in both assays, with increased proportions of p21-expressing cells in cell lines selected for metastatic ability. Our results suggest that (a) the experimental (i.v.) metastasis assay in the chick embryo offers an efficient and cost-effective procedure for the identification and selection of cells that are also experimentally metastatic in nude mice, and (b) the metastatic properties of ras-transformed NIH 3T3 cells are due to individual cells that express increased amounts of p21.