We observed highly aggressively proliferating immortalized (HAPI) cells growing in cultures that had been enriched for microglia. The cells were initially obtained from mixed glial cultures prepared from 3-day-old rat brains. HAPI cells are typically round with few or no processes when cultured in 10% serum containing medium. As the percentage of serum in the medium is decreased, the HAPI cells have more processes. HAPI cells stain for the isolectin B4, OX-42, and GLUT5, which are markers for microglial cells, but the cells do not immunolabel with A2B5, a marker of cells in the oligodendroglial cell lineage, or with the astrocyte-specific marker, glial fibrillary aciidic protein (GFAP). In addition, HAPI cells are capable of phagocytosis. We conclude that HAPI cells are of microglia/macrophage lineage. Exposing HAPI cells to lipopolysaccharide (LPS) induces the mRNAs for tumor necrosis factor-alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS). LPS exposure also induces secretion of TNF-alpha and production of nitric oxide (NO) in HAPI cells. Because activation of microglia is associated with an increase in iron accumulation and ferritin expression, we tested the hypothesis that iron status affects the production of TNF-alpha and NO. Our studies demonstrate that both iron chelation and iron loading diminished the LPS-induced effect of TNF-alpha and NO. The results of this study indicate that HAPI cells possess the characteristics of microglia/brain macrophages, providing an alternative cell culture model for the study of microglia. In addition, we demonstrate that the activation of microglial cells could be modified by iron.

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The long term effects of indomethacin, cyclosporin, cyclophosphamide, and placebo on collagen-induced arthritis in mice were tested under two different treatment protocols. A prophylactic experiment examined the effects of the daily drug administration for 180 days beginning one day before the first collagen injection. Under this dosage schedule, cyclophosphamide and cyclosporin decreased the severity of arthritis, while indomethacin did not. A therapeutic protocol examined the effects of these same drugs when daily administration was delayed until the animals had active disease at 78 days after the first collagen injection. Under this protocol, all three drugs reduced the progression of disease. In both protocols, the most significant suppression of arthritis was seen in animals receiving cyclophosphamide which was associated with a decrease in anti-collagen antibody levels. Collagen-induced arthritis in mice should be further investigated as a model to study the long term effects of "slow-acting" anti-rheumatic drugs.

A series of 2-alkylisoflavone derivatives 1 was prepared with the intent to study the importance of the phenyl group (at the 3-position) of the isoflavone in imparting antihypertensive activity and the substitution effects at the 2-position of isoflavone. With the exception of the 2-isopropyl analog, the antihypertensive activity of these compounds appears to have a slow onset and long duration. None of the analogs appears better than the corresponding flavone (3) and 3-phenylflavone (2) analogs. An unsuccessful attempt to correlate the relationship between antihypertensive activity and the calculated torsional angle of C2-C3-C1'-C2' is discussed. Antiinflammatory activities of these compounds along with 7-(oxypropylamine)flavones were also evaluated and found to be not very potent. The antiinflammatory activity appears to be sensitive to steric effects of the alkyl group on the nitrogen and of substituents at the 2-position of the isoflavones, while the hydroxyl group of the propanolamine side chain is not essential.