Marie U. Nylen

Observations on function, composition, and structure of cestode calcareous corpuscles

Abstract 1. 1. Taenia taeniaeformis contains about half as much calcareous corpuscle material as does Cysticercus fasciolaris. The corpuscles show no quantitative gradient along the strobila of Taenia taeniaeformis, but the larger individual proglottids contain more corpuscle material than the smaller ones. 2. 2. More calcareous material disappears from Cysticercus faciolaris during anaerobic than during aerobic incubation in non-nutritive media. In both cases considerably more corpuscle material disappears when the worms are incubated in slightly acid than in slightly alkaline surroundings. It is concluded that the corpuscles serve to buffer acids entering the body from the outside and, possibly, to buffer acids produced during aerobic and anaerobic fermentations. 3. 3. The corpuscles yield Ca, Mg, P, and CO2. The mineral component of the corpuscles is amorphous, but upon heating with KOH crystallization takes place and brucite and hydroxyapatite crystals are formed. The carbonates, however, do not crystallize. 4. 4. Electron microscopy indicates that the lamellae of the corpuscles may be paired rings. It reveals also an amorphous or granular background substance, but no crystalline elements. 5. 5. The inorganic material of the corpuscles can readily be demonstrated by various histochemical staining methods. It is also possible to demonstrate histochemically a glycogen-like polysaccharide, a mucopolysaccharide, lipids, and proteins as constituents of the organic base of the corpuscles.

Matrix-Mineral Relationships-A. Morphologist's Viewpoint

The literature on the ultrastructural morphology of the enamel matrix and its relationship to the crystals is reviewed. Two morphological entities of the matrix are discussed: One is the so-called stippled material which may be the initial cell product; the other, variously described as fibrillar, lamellar, tubular or helical, is thought by many to play a crucial role in nucleation and orientation of the crystals. A number of observations, however, suggest that the latter structures form secondarily to the crystals and that in reality they represent organic material adsorbed to the crystal surface and maintained as independent structures upon removal of the mineral. The need for additional studies is stressed including systematic studies of interactions between constituents of the organic matrix and the apatite crystals.

Variations in the mineralogical composition of cestode calcareous corpuscles

Abstract 1. 1. The calcareous corpuscles of Cysticercus cellulosae , larval Spirometra mansonoides , Taenia saginata , and Diphyllobothrium latum contain various proportions of calcium, magnesium, phosphorus, and CO 2 as major components. 2. 2. The corpuscles of these worms, and those of the previously studied Taenia taeniaeformis , contain in addition several minor inorganic constituents. The corpuscles of the larval worms show a smaller variety of the minor components than those of the adult worms. 3. 3. The crystallization patterns induced by heat or KOH treatment vary, especially in respect to the phosphorus-containing compounds. Three of the latter have been identified: Hydroxyapatite, whitlockite, and calcium-magnesium-orthophosphate. The crystallization variations may be related to the ratios in which the major components occur in the corpuscles of the different species.

The mineralogical composition of the calcareous corpuscles of Taenia taeniaeformis.

Abstract 1. 1. The mineral content of calcareous corpuscles of cestodes is amorphous in vivo and consists primarily of Ca, Mg, P, and CO2. 2. 2. The results of a series of heat and chemical transformations of isolated Taenia taeniaeformis corpuscles and synthetic mixtures suggest that the major inorganic components of the native corpuscles are in the form of an amorphous calcium phosphate and an amorphous MgCaCO2 complex. 3. 3. The fact that still other inorganic constituents must be present is demonstrable not only from the analytical data but also from the isolation of a thermolabile fraction.

Composition and crystallization patterns of calcareous corpuscles of cestodes grown in different classes of hosts.

The chemical composition and the crystallization patterns of the calcareous corpuscles of four and six species of cestodes, respectively, have been studied. Considerable variations were found in both areas. These seemed to be more related to the species of tapeworm than to the type of host in which the worms grew. Thus, very pronounced differences were found between the corpuscles of two larval worms (Taenia crassiceps and Mesocestoides corti), both of which developed in the peritoneal cavity of mice. On the contrary, the crystallization patterns of one fish tapeworm (larval Ligula intestinalis) corresponded upon heating to 600 or 900 C to what is found in such mammalian species as Taenia taeniaeformis, Bertiella studeri, and several others. The 300 C pattern of Ligula conformed to that found in another mammalian species, Mesocestoides corti, and a similar pattern has not yet been encountered in any other tapeworm, of mammalian or nonmammalian origin. The corpuscles of the chicken tapeworm Raillietina cesticillus, on the other hand, were characterized by the almost complete absence of magnesium and the fact that ethylenediamine-isolated corpuscles showed a definite calcite pattern before subsequent heating. In other species, corpuscles isolated by this method have been amorphous with only those of Cysticercus cellulosae showing traces of calcite. Our previous studies (von Brand et al., 1960, 1965a, b; Scott et al., 1962) on the chemical composition and crystallization of cestode calcareous corpuscles revealed quantitative and qualitative differences between the granules isolated from certain species. All tapeworms studied so far in these respects had been grown in mammals. Thus the question arose whether corpuscles from cestodes isolated from other groups of hosts would show more pronounced differences or other types of differences. The present communication is a first attempt to present material having a bearing on this ques-

Cestode calcareous corpuscles: phosphate relationships, crystallization patterns, and variations in size and shape.

Abstract Calcareous corpuscles isolated from larval Echinococcus granulosus yielded similar average phosphate values when worms of Chilean, Argentinian, Australian, and Lebanese provenience were analyzed contraindicating the existence of distinct geographic strains of the parasite. Instead, the observed variations are more likely due to nutritional differences. Considerable variations in phosphate content of calcareous corpuscles were encountered in analyses of both individual larval and adult Taenia taeniaeformis , the variations being somewhat more pronounced in the latter than in the former. In general, corpuscles isolated from laboratory-infected cats contained significantly less phosphate than those taken from stray cats, again presumably indicating the influence of the nutrition of the host on the phosphate content of the calcareous corpuscles. Isolated calcareous corpuscles incorporated variable amounts of phosphate, depending upon the following factors: pH of the medium, isolation procedure, species of tapeworm, and ratio between the amount of corpuscles to the volume of incubation fluid. Crystallographic studies showed that the originally amorphous corpuscles of T. taeniaeformis gave a weak dolomite-like pattern after 180 days of uninterrupted heating to 150 °C. Definite dolomite formation was induced by heating to 180 °C. The dolomite was stable up to 400 °C, but decomposed readily at 450 °C. Hydroxyapatite appeared first upon heating to 400 °C and remained stable up to the highest temperature tested, 900 °C. The heat-induced crystallization patterns of corpuscles from ten additional tapeworms were determined. Especially remarkable was the fact that in two species ( Cysticercus bovis and Cysticercus tenuicollis ) a weak, but definite dolomite pattern was found after ethylenediamine isolation, without the necessity of applying additional heat. Considerable variations in size and shape between corpuscles of various species were found. Echinococcus multilocularis , for example, had giant corpuscles with diameters up to 34 μ, while those of Priapocephalus sp. measured usually between 2 and 4 μ. Some species had predominantly round corpuscles, others oval or irregularly shaped ones. Where comparisons between larval and adult corpuscles were possible no significant differences were found in most cases. The only exception was Taenia saginata where the corpuscles of the adult were definitely smaller than those of the larva.

Organic matrix of cestode calcareous corpuscles.

Abstract A method is described for isolating pure samples of calcareous corpuscles of Taenia taeniaeformis, suitable for analysis of the organic matrix. Electron microscopic studies of corpuscles decalcified either in situ or after isolation showed that they are surrounded by an envelope composed of two layers, the outer one consisting probably of the compressed remnants of the corpuscle-forming cell. In the interior of the corpuscles concentric layers of organic material are found which may represent the structural framework for the deposition of mineral. Chemical analysis of isolated corpuscles showed that they had lost all carbohydrates that are demonstrable histochemically in corpuscles studied in situ. The corpuscles contained lipid separable into glyceride, phospholipid, and sterol fractions. The sterol was probably cholesterol. Unsaturated fatty acids predominated both in the glycerides and in the phospholipids. The corpuscles contained relatively little protein. An amino acid analysis gave a result resembling that of cellular protein in general with no indication for the presence of collagen.

Tetracycline-induced enamel defects in the rat incisor. A microroentgenographic and fluorescence microscopic study.

Abstract The effect of single and multiple intraperitoneal injections of tetracycline hydrochloride (TC) on rat incisor enamel was investigated in a study utilizing microroentgenography and fluorescence microscopy. Thirty-four animals received single dosages ranging between 6 and 130 mg TC/kg body weight and 20 animals received 5 successive injections of 130 mg TC/kg body weight. The animals were sacrificed 10, 20, 30 or 40 days later. With the exception of the lowest dosages, both single and multiple injections caused disturbances in the enamel, the severity of which was proportional to the dosage. The enamel defects ranged from hypomineralized incremental bands to hypoplastic lesions. Mineralization disturbances were associated with all defects, internal as well as external. The localization of defects to the developing enamel surface indicates that the primary effect of TC is impairment of the ameloblasts, the youngest secreting cells being especially susceptible. The similarity between the effect of TC and that of sodium fluoride on enamel formation is discussed. The hypoplasias which were the result of multiple injections were labelled permanently by the antibiotic; those caused by a single injection were non-fluorescent. It is suggested that some special feature of the altered enamel laid down subsequent to the first injection permits retention of the fluorophor.

Observations on Calcareous Corpuscles of Larval Echinococcus granulosus of Various Geographic Origins

Summary Calcareous corpuscles isolated from larval Echinococcus granulosus collected from sheep in Chile and New Zealand and from cattle in Lebanon showed only minor differences in their chemical composition and their heat-induced crystallization patterns. The only definite differences found were that the Chilean material lacked manganese, and that it had a somewhat higher magnesium and a lower phosphorus content than the other samples. As a consequence of the relatively low phosphorus content the Chilean material yielded upon heating at most traces of apatite. Previously described much more pronounced differences in chemical composition and crystallization patterns between corpuscles from different tapeworm species are hence probably essentially species-specific.

Microstructural changes induced in mammalian cell cultures by omission and replacement of a single essential amino acid

Abstract Maintenance of a strain of HeLa cells on a culture medium deficient in a single essential amino acid, valine, gave rise to an immediate cessation of growth, a series of degenerative changes in cell structure, and subsequently a marked decrease in the cell population. When the amino acid was reintroduced, the surviving cells appeared to undergo repair, and following this the culture resumed normal growth.