Cytological Aspects of Lipid Assimilation by Cestodes. Incorporation of Linoleic Acid into the Parenchyma and Eggs of Hymenolepis diminuta

Abstract

The distribution of radioactivity in adult tapeworm tissues following in vitro exposure to 14C-linoleic acid was studied by tissue section autoradiography. These observations were correlated with the chemical distribution of radioactivity in the neutral lipids fractionated by thin-layer chromatography and assayed by liquid scintillation spectrometry. The present results indicate that linoleic acid absorbed from the tapeworm's environment is rapidly esterfied within the cortical and medullary parenchyma. Radioactivity is subsequently transported to the eggs and incorporated therein. Recent studies on the origins of tapeworm lipids have indicated that the ability of the adult worms so far examined to synthesize fatty acids is limited to a chain lengthening process (Meyer et al., 1966; Ginger and Fairbairn, 1966b; Jacobsen and Fairbairn, 1967). However, when supplied with exogenous fatty acids and sterols, cestodes are capable of assembling triglycerides, sterol esters, glycoand phospholipids (Meyer et al., 1966; Ginger and Fairbairn, 1966b; Jacobsen and Fairbairn, 1967). Linoleic acid, the prevalent fatty acid of the lipids of hymenolepidid tapeworms (Harrington, 1965; Ginger and Fairbairn, 1966a) is readily incorporated into the neutral lipids and phospholipids of Hymenolepis diminuta as demonstrated by Lumsden and Harrington (1966) and Jacobsen and Fairbairn (1967). While large quantities of linoleic acid-containing lipids are concentrated in the parenchyma (Lumsden and Harrington, 1966), their role remains enigmatic. No evidence has yet been obtained that the extensive depot lipids of cestode tissues serve as an energy source or are otherwise metabolized by the adults and little or no carbon from non-lipid materials appears in fatty acid constituents (Ginger and Fairbairn, 1966b; Meyer et al., 1966; Jacobsen and Fairbairn, 1967). It is conceivable, however, that at least a portion of the tissue lipids of adult tapeworms might be incorporated into the developing eggs for utilization by some subsequent stage in the life cycle. Lipids comprise more than 30% of the total solids in Received for publication 29 October 1968. * Supported by grants from the NIH (GMN 13330, AI 08673) and NSF (GB 7276). the terminal gravid proglottids of H. diminuta, the infective eggs containing approximately 11% lipid on a dry weight basis (Fairbairn et al., 1961). Lipid droplets are visible in the eggs of this and many other tapeworm species. The possible transport of parenchymal lipids into the eggs of Hymenolepis diminuta was therefore investigated by tissue section autoradiography of worms maintained in vitro in the presence of 14C-linoleic acid. MATERIALS AND METHODS The Hymenolepis diminuta utilized in this study were obtained from 14-day-old infections of albino rats. Following removal of the worms from the excised host gut by flushing with tris maleate buffered Kreb's Ringer's saline (Read et al., 1963), the worms were incubated in the medium of Schiller (1965) containing 2 /uC/ml linoleic acid1-14C (52.9 mc/mM, Nuclear-Chicago). Radiochemical purity of the linoleic acid was ascertained before use by thin layer chromatography (see below). Incubations were carried out under a carbon dioxide-nitrogen atmosphere (5:95) in a Dubnoff shaking water bath for 5, 15, 30, 60, and 120 min at 37 C or 1 C. Some of the worms incubated with 14C-linoleic acid for 120 min were subsequently transferred to isotope-free medium and maintained in vitro for up to 6 days according to Schiller (1965). After incubation, worms were either fixed with osmic acid and processed for autoradiography or quickly rinsed in isotope-free Hank's balanced salt solution, frozen on glass plates resting atop blocks of dry ice, weighed, then stored in nitrogen purged vials at -70 C for subsequent lipid analysis. Chemical determinations Worms were homogenized in 10 ml chloroformmethanol (2:1). A 0.2 ml aliquot of the homogenate was removed and hydrolyzed in 30% potassium hydroxide for determination of total radioactivity assimilated. The remaining material was filtered, washed, and the chloroform layer isolated according to the method of Folch et al.