Mitogenic Activity of Cell‐Wall Components in Mouse Spleen Cells

Abstract

In the previous paper (5), the authors examined the mitogenic activity of the cell walls of mycobacteria, nocardia and corynebacteria, and showed that these cell walls were active as mitogens in normal spleen cells, anti-0 sera-treated spleen cells, macrophage-depleted spleen cells and cortisone-resistant thymocytes of C57BL/6J mice. It was also demonstrated that these cell walls were mitogenic in the spleen cells and macrophage-depleted spleen cells of congenitally athymic nude (BALB/c -nu/nu) mice. From the above results, we have concluded that the cell walls of mycobacteria, nocardia and corynebacteria act as mitogens on both thymus-derived lymphocytes (T cells) and bone marrow-derived lymphocytes (B cells). This note deals with the chemical constituents which are responsible for mitogenic activity of the mycobacterial cell wall. It has been established that the chemical structure of mycobacterial cell wall is a acid-arabinogalactan-mucopeptide complex and the detailed chemical properties of each component were described previously (2, 3). The chemical components of mycobacterial cell walls were prepared by the following procedure and their mitogenic activity in the spleen cells of C57BL/6J and congenitally athymic nude mice, and cortisone-resistant thymocytes of C57BL/6J mice was examined. Mycolic acid, known to be a-branched i3-hydroxy fatty (1), was prepared from the cell wall of Mycobacterium tuberculosis train Aoyama B by alkaline hydrolysis using 2.5% KOH in benzene-methanol (1 :1), and mycolic acid methyl ester was prepared by refluxing mycolic acid with 3% HC1-methanol for 3 hr. D-Arabinose5-mycolate was purified from the firmly bounded lipids of M. tuberculosis Aoyama B as described previously (6). Arabinogalactan, the neutral polysaccharide of mycobacterial cell walls, was purified from the cell wall of Mycobacterium bovis strain BCG according to the method of Misaki et al (11). Mycoloyl-arabinogalactan (D-CMI-A) was prepared by further fractionation of so-called wax D which was extracted by the same procedure as described previously (3, 7) from the cells of M. bovis strain Ushi 10 which were cultured on Sauton's synthetic medium at 37 C for 8 weeks. The mycoloyl-arabinogalactan designated as D-CMI-A of M. bovis Ushi 10 is composed of arabinogalactan (46%) and mycolic acid (51 %) but lacks mucopeptide moiety. The purification and porperties of the cell wall of M. bovis BCG were described pre-