Fine Structure of the Intestinal Epithelium of Dirofilaria immitis and Changes Occurring after Vermicidal Treatment with Caparsolate Sodium

Abstract

The intestinal epithelium of Dirofilaria immitis from an untreated dog and dogs treated with Caparsolate Sodium was studied by electron microscopy. The intestine of D. immitis is composed of a single layer of thin, columnar epithelial cells, joined laterally by terminal complexes. Each cell is divisible into an apical, central, and basal region. The latter contains a labyrinth of cisternae formed by deep invaginations of the basal plasma membrane. The apical cell surface is modified into irregularly arranged microvilli, which lack a dense central core. No terminal web is present. The central region of the cell contains a large nucleus and a limited number of organelles. These include a few rudimentary mitochondria, poorly defined endoplasmic reticulum, scanty ribosomes, microbodylike structures, and well-developed Golgi complexes. Glycogen was not observed. Lipid and pigment granules constitute the major cell inclusions and are closely associated with each other. Pigment granules are polymorphic and probably iron-positive as determined histochemically. They possess myelin figures in a dense matrix. These granules occur abundantly in the anterior portion and are almost absent from the posterior portion of the intestine. Their probable polyphasic role during absorption of host hemoglobin is discussed. A reduction in the number of these granules was observed 24 hr following administration of Caparsolate Sodium. They also seemed to aggregate, lose most of their myelin figures, and assume varying densities. Dense deposits covering the surface of the microvilli and the apical cell membrane are assumed to be a protein-arsenical complex which alters the morphology and function of the striated border. This is considered the probable mechanism of the filaricidal action of the therapeutic agent. Carbonell and Apitz (1959) described pigment granules from the intestinal cells of Ascaris lumbricoides which they regarded as intermediary products of metabolism. Similar granules were reported from the intestine of Ancylostoma caninum and their relationship to host hemoglobin has been indicated (Lee, 1968a, b). Another blood-feeding nematode, Dirofilaria immitis, also contains intestinal pigment granules. Infections with this nematode respond well to treatment with organic arsenical compounds (Otto and Maren, 1950; Drudge, 1952; Kune and Ohishi, 1957; Bailey, 1958). Little is known, however, about the mechanism of their vermicidal action. The intestinal fine structure of filariae from an untreated dog and those treated with Caparsolate Sodium [Sodium (p-carboamyl-phenylarsylene-dithio)diacetate] was studied to ascertain the possible mechanism of action of the therapeutic agent and to elucidate further the role of intestinal pigment granules. Received for publication 29 November 1968. * This investigation was supported by PHS Research Grant AI-02347 from NIAID of the NIH. MATERIALS AND METHODS Two dogs, naturally infected with D. immitis, were treated with Caparsolate Sodium at doses of 1 mg/lb of body weight. One naturally infected dog served as a control. One treated dog received 2 consecutive doses of the drug intravenously within 24 hr. The other was given 4 consecutive injections in 2 days. Both were killed with an overdose of nembutal 24 hr after the last injection of arsenical compound. The control dog was killed at the same time. Adult D. immitis were collected from the right ventricles and washed with warm saline. Segments of about 1 inch were taken from the anterior, middle, and posterior portions of worms and fixed in cold (5 C) glutaraldehyde (Pease, 1964) for 24 hr. Each segment was then subdivided into pieces of about 1 mm and postosmicated (Millonig, 1961) overnight at 5 C. They were embedded in Araldite 502 (Luft, 1961) after dehydration with ethanol. Sections were obtained with diamond knives on a PorterBlum MT-1 microtome, stained with lead hydroxide (Watson, 1958), and viewed with a Hitachi HU-11A. Histochemical tests for iron were performed on paraffin sections using a combination of microincineration and Prussian blue technique as described by Fenton et al. (1964).