Amino acid metabolism in the rat tapeworm, Hymenolepis diminuta

1. The amino acid metabolism of the rat tapeworm, Hymenolepis diminuta was investigated. 2. In addition to the characteristic end products of helminth metabolism, H. diminuta also forms substantial amounts of 14C-alanine during incubations in 14C-glucose. 3. Of 10 amino acids tested, only 14C-labelled asparate and, to a lesser extent alanine, generated substantial amounts of 14CO2 when incubated with H. diminuta. 4. 14C-aspartate was incorporated into both succinate and acetate, major products of the worms mitochondrial metabolism, but the rates were low when compared to the metabolism of exogenous glycogen. 5. These results suggest that amino acid metabolism in H. diminuta is very limited.     

Galactose utilization by the rat tapeworm, Hymenolepis diminuta.

1. Hymenolepis diminuta incorporated label from 14C-galactose into glycogen, but the sugar would not support net glycogen synthesis. Glucose stimulated the incorporation of label from 14C-galactose into glycogen, while glycerol did not. 2. During incubations in galactose, large internal pools of galactose and galactose 1-P accumulated, while the concentration of glucose 6-phosphate remained unchanged. 3. In vitro culture experiments indicated that galactose would not support worm growth. Therefore, while galactose can be metabolized to a limited extent, it cannot substitute for glucose as a nutrient source.     

Identification of ADPR-transferase activity in the rat tapeworm, Hymenolepis diminuta

1. The nuclear fraction of the rat tapeworm Hymenolepis diminuta (Cestoda) contains the enzyme adenosine diphosphoribosyl transferase (ADPR-transferase). 2. The enzyme catalyzes the postsynthetic modification of some nuclear proteins by the covalent attachment of the (ADP-ribose) moiety of NAD to such proteins. 3. The reaction is dependent on DNA which contains strand-breaks, and chain lengths equivalent to (ADP-ribose) is estimated. 4. The formation of polynucleotide products was competitively inhibited by 3-acetamidobezamide, with a Km of 125 microM. 5. The catalytic properties of ADPR-transferase in Hymenolepis diminuta are similar to those in T. brucei.     

Enzymes of galactose utilization in the rat tapeworm, Hymenolepis diminuta.

1. Crude enzyme preparations from Hymenolepis diminuta contained galactokinase, galactose 1-phosphate uridyl transferase and UDPgalactose 4-epimerase activity, although their specific activities were low. 2. Galactose 1-phosphate non-competitively inhibited galactose phosphorylation. This inhibition, together with the low specific activities of the enzymes in the pathway of galactose utilization, probably accounts for the inadequacy of galactose as a main nutritive carbohydrate for development of the worm.     

The selective uptake of cholesterol by the rat tapeworm Hymenolepis diminuta (Cestoda)

1. The sterols of Hymenolepis diminuta are almost exclusively cholesterol or similar C-27 sterols; the free sterols of its environment (the lumen of the rat intestine) are cholesterol and various phytosterols. 2. During incubation of tapeworms with mixed micelles of taurocholate, glyceryl monooleate, and equimolar [3H]cholesterol and [14C]beta-sitosterol, the uptake of cholesterol is 40 times more rapid than the uptake of sitosterol. 3. Following uptake, the desorption of labeled sitosterol is six times more rapid than that of cholesterol. 4. We did not detect the esterification of absorbed sterols or the conversion of absorbed sitosterol of cholesterol. 5. The highly selective uptake of cholesterol and the moderately selective desorption of phytosterols can account for the selective accumulation of C-27 sterol by the tapeworm.     

Cytoskeletal features of the syncytial epidermis of the tapeworm Hymenolepis diminuta

A hallmark feature of parasitic platyhelminths is a cytoarchitecturally unusual syncytial epidermis composed of a peripheral layer of continuous cytoplasm (the ectocytoplasm) connected to underlying nucleated cell bodies by small cytoplasmic bridges. The helminth epidermis, or tegument, plays important roles in protection and nutrient acquisition; cestodes, in fact, completely lack a gastrointestinal tract and absorb all nutritive material through the tegument. Perhaps not surprisingly, the cestode tegument bears certain resemblances to the mucosal epithelium of the vertebrate small intestine, including the possession of a microvillous brush border upon the surface of the ectocytoplasm. In contrast to the intestinal epithelial cell, however, very little is known concerning the nature and organization of the cytoskeleton within the helminth epidermis. Therefore, a number of different microscopical preparative techniques were used to examine the tegument of the tapeworm Hymenolepis diminuta for the presence and distribution of microfilaments, intermediate filaments, and microtubules. It was found that both actin-containing microfilaments and intermediate-sized filaments are present but are restricted to specific locations along the plasmalemmae of the ectocytoplasm. In contrast, microtubules are found throughout the tegument, and are concentrated in the supranuclear regions of the perikarya and in the cytoplasmic bridges interconnecting the perikarya and ectocytoplasm. Unlike brush borders of most other epithelia, the cestode epidermal brush border lacks a filamentous terminal web and is instead associated with microtubules. A network of fine filaments, 5-8 nm in diameter but distinct from actin-containing microfilaments, runs throughout the ectocytoplasm and appears to interlink tegumental vesicles. These fine filaments may represent the primary "skeletal" system responsible for maintaining the structure of the tegumental cytoplasm.     

Modulation of caudal intestinal permeability in the rat during infection by the tapeworm Hymenolepis diminuta.

Bidirectional movement of solutes between the intestinal lumen and systemic circulation is restricted by tissue barriers that may be altered under conditions such as intestinal infection. In a study using an in vitro everted sac preparation to assess small intestinal permeability in a lumen-to-serosa direction, 51Cr-EDTA movement was compared regionally in the jejunum and ileum of rats infected and uninfected by tapeworms. Whereas jejunal segments showed no significant differences in permeability to 51Cr-EDTA at 6, 15, or 32 days postinfection (dpi), ileal segments displayed an increased permeability on 15 and 32 dpi, but not 6 dpi. The alterations in permeability were not reversed 1 wk after removal of the tapeworm from the intestine. In conclusion, the strictly lumen-dwelling tapeworm infection allows increased movement of molecules from the lumen into ileal, but not jejunal, tissues by 15 dpi.     

Inhibition of growth of a tapeworm Hymenolepis diminuta in its normal host (rat).

The biomass of 8-day-old worms of Hymenolepis diminuta in secondary infections, administered to rats 3-10 days after chemotherapeutically expelling a primary infection, was 70-90% less, and the worms were more posteriorly distributed, than in naive controls. The strong depressive effect on growth waned rapidly over 2-5 weeks, but even in rats not challenged until 17 months later, worm growth was weakly depressed by 30%. The extent to which growth was depressed in a secondary infection was independent of the number of worms in the challenge but increased with number of worms in the immunizing infection up to four to eight worms. Further increase up to 64 worms had little effect. This suggests, as it is known that the biomass of worms in a rat reaches a maximum with infections of between five and 10 worms, that the change in the intestine is proportional to biomass, not number, of worms. It is argued that partially suppressed immuno-inflammatory changes in the intestine, which will affect secondary worms so strongly, will also have depressed growth and fecundity effects on the primary worms, that a dynamic equilibrium is reached between the strength of the intestinal response and the biomass of the tapeworm, and that it is reaching this equilibrium, not a 'crowding effect', which limits H. diminuta to a level compatible with the survival of the rat.     

Studies on the mechanism of cholesterol uptake by the rat tapeworm Hymenolepis diminuta (Cestoda)

1. With increasing cholesterol content in mixed micelles, the rate of cholesterol uptake by the tapeworm approaches a limiting, maximal value. 2. This uptake is inhibited only 32-40% by other sterols, but is not markedly dependent on medium pH or tapeworm energy metabolism. 3. The competitive exchange diffusion of absorbed [14C]cholesterol could not be demonstrated. 4. The above results partially support the hypothesis that the tapeworm absorbs cholesterol by a specific carrier-mediated process. 5. Cholesterol uptake is reduced when the capacity of the micellar phase of the medium is increased, suggesting that uptake involves the intermediate partitioning of sterol from micelles into the aqueous phase of the medium.     

A comparison of phlorizin and phloretin adsorption by the tapeworm Hymenolepis diminuta

Phloretin and phlorizin adsorb to the tegument surface of Hymenolepis diminuta, with KDs of 2.39 mM and 14.7 microM, respectively, and Vmaxs of 1446 and 12.54 nmoles/g tissue per 2 min, respectively. Phloretin adsorption is not inhibited by phlorizin or glucose. Glucose partially inhibits phlorizin adsorption. Phlorizin, but not phloretin, adsorption to isolated tegument brush border membrane preparations is partially inhibited by N-ethylmaleimide. No indications of phlorizin hydrolysis to phloretin during incubation with H. diminuta were obtained. The data are supportive of spacially separate and distinct binding sites for phloretin and phlorizin in the tegument brush border.     

Allometric growth of the proglottids and strobila of the tapeworm, Hymenolepis diminuta

HYMENOLEPIS DIMINUTA: adults were killed and fixed in fully extended positions, and the total lengths of the strobilae, numbers of proglottids, and distances of proglottids along the strobilae were determined. The relationship of proglottid number to distance along the strobila was exponential. Beginning at proglottid 100 (P100), the lengths and widths of proglottids at 100 proglottid intervals were determined, and the surfaces areas were calculated. The relationships of proglottid length and width to proglottid number were linear, but the relationship of proglottid number to surface area (SA) was exponential. The volumes of proglottids were calculated, and the relationship of volume (V) to proglottid number was exponential. The relationship of surface area to volume ratio (SAVR) to proglottid number was exponential; at the anterior end of the worm (P100), the SAVR was 14.6, while at the posterior end of the worm (P1300) the ratio was 4.2. A single exponential equation describing the relationships among proglottid number, SA, and V was derived.     

Biochemical characterisation of a hydrophobic ligand binding protein from the tapeworm Hymenolepis diminuta

The cestode Hymenolepis diminuta contains an abundant, cytoplasmic, hydrophobic ligand, binding protein (H-HLBP). Studies with polarity sensitive probes suggest a single hydrophobic binding site, the results also indicate that the single tryptophan in the molecule (Trp41) is involved in ligand binding. Of the possible physiological ligands tested, only haematin and retinoids (retinol and retinoic acid) show appreciable binding in addition to fatty acids. H-HLBP also binds a range of anthelmintics, again with K(D) values in the nM range. The interaction of anthelmintics with hydrophobic binding proteins may be important in determining drug specificity and site of action and could have a role in the development of drug resistance.     

Characterization of the immuno-regulatory response to the tapeworm Hymenolepis diminuta in the non-permissive mouse host

Hymenolepis diminuta is spontaneously expelled from mice; concomitant with worm expulsion was protection against colitis induced by dinitrobenzene sulphonic acid (DNBS). Here we examined the immune response mobilized by Balb/c and C57Bl/6 male mice in response to H. diminuta and assessed the requirement for CD4+ cells (predominantly T cells) in worm expulsion and the anti-colitic effect. Wild-type (CD4+) or CD4 knock-out (CD4-/-) mice received five H. diminuta cysticercoids and segments of jejunum and mesenteric lymph nodes (MLNs), or spleen, were excised 5, 8 and 1l days later for mRNA analysis and cytokine production, respectively. In separate experiments uninfected and infected mice received DNBS by intra-rectal infusion and indices of inflammation were assessed 3 days later (i.e. 11 days p.i.). Infection of Balb/c mice resulted in a time-dependent increase in intestinal mRNA for Foxp3, a marker of natural regulatory T cells, and markers of alternatively activated macrophages (arginase-1, FIZZ1), while concanavalin-A activation of MLN cells revealed a significant increase in T helper 2 (TH2) type cytokines: IL-4, -5, -9, -10, -13. MLN cells showed a reduced ability to induce Foxp3 expression upon stimulation. CD4-/- mice did not display this response to infection, but surprisingly did expel H. diminuta. Moreover, DNBS-induced colitis in CD4-/- mice (wasting, tissue damage, elevated myeloperoxidase) was not reduced by H. diminuta infection, whereas time-matched infected CD4+ C57Bl/6 mice had significantly less DNBS-induced inflammation. In conclusion: (i) in addition to stereotypical TH2 events, H. diminuta-infected Balb/c mice develop a local immuno-regulatory response; and (ii) CD4+ cells are not essential for H. diminuta expulsion from mice but are critical in mediating the anti-colitic effect that accompanies infection in this model.