Splenectomy abrogates the induction of oral tolerance in experimental autoimmune uveoretinitis.

Oral administration of uveitogenic antigens inhibits the development of experimental autoimmune uveoretinitis (EAU) and the cellular immune response initiated by these antigens. The mechanism of oral tolerance is not completely clear, but accumulating data indicate that suppressor cells are actively involved in this process. The spleen is known to harbor suppressor cells and their precursors and the present study was aimed at testing the role of this organ in the induction of oral tolerance by S-antigen (S-AG). We report here that: (a) splenectomy abrogated the induction of oral tolerance; unlike in sham operated controls, feeding with S-Ag did not inhibit the development of EAU in splenectomized rats; (b) splenectomized rats responded with higher cellular immune responses than did sham operated controls, but feeding with S-Ag inhibited these responses in both groups of animals; (c) splenectomy also abrogated the adoptive transfer of tolerance: EAU induction was inhibited in sham operated recipients of splenocytes from S-Ag fed donors but not in the splenectomized recipients. The data thus indicate that the spleen plays an important role in the induction of oral tolerance, perhaps by acting as the site for induction and/or amplification of cells with suppressor activity.

Isolation of single cell suspensions from the rat mammary gland: separation, characterization, and primary culture of various cell populations.

Isolation and characterization of a single cell suspension from the rat mammary gland was achieved by combining selective enzymatic digestion and the mechanical agitation of a Stomacher laboratory blender with immunohistological identification of cell-specific markers. Utilizing this procedure we were able to isolate single cell suspensions of high yield (10 to 15 X 10(6) cells/rat) and viability (greater than 98%) with a concurrent decrease in isolation time and the amount of proteolytic enzymes required. Five distinct cell fractions were isolated from the mammary gland cell suspension after banding on discontinuous Percoll gradients. These populations were characterized both before and after primary cell culture by a combination of histological, immunohistological, and autoradiographic techniques. Fractions two and three were found to be enriched for mammary epithelial cells, as identified by their high binding of antikeratin antibodies. These populations also exhibited a minimal degree of binding to actin, myosin, and fibronectin antibodies. Fraction three also exhibited a high labeling index as measured by autoradiography following in vivo administration of [methyl-3H]thymidine. The remaining fractions were found to contain higher percentages of myoepithelial cells or other mammary cell types. Inasmuch as there is a direct correlation between mammary gland cell types and susceptibility to mammary gland carcinomas, further studies of these cell populations may provide new insights into the mechanisms underlying mammary gland carcinogenesis.