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Biomedical subjects

D W Fawcett

Publications and source records attributed to D W Fawcett.

At least 19 recordsLinked to original sources

Ultrastructure of the intra-erythrocytic stage of Theileria species from cattle and waterbuck.

A transmission electron microscopic study of the intra-erythrocytic stages of a pathogenic Theileria parva from cattle and a previously uncharacterized Theileria sp. from waterbuck (Kobus defassa) in Kenya revealed several novel ultrastructural features, associated with feeding and multiplication, in these parasites. In trophozoites a connecting channel was observed between the parasite's cytostome and its intracytoplasmic food vacuole. In some cases the limiting membrane of the food vacuole was seen to be continuous with a close-meshed network of membrane-bounded, anastomosing tubules. This labyrinthine structure, which has not been described previously, may function as a digestive organelle in theilerial trophozoites. Electron micrographs also revealed the mode of intra-erythrocytic multiplication of these parasites in vivo. Prior to division, electron-dense cisternae and rhoptries appeared beneath the parasite's plasmalemmal membrane, marking the sites of merozoite formation. From a single parasite, a maximum of four merozoites were formed by schizogonous division and subsequently separated from a residual body by constriction at the base of each merozoite. In addition, observations on two double-membraned organelles seen in trophozoites and the intra-erythrocytic crystalline structures associated with Theileria sp. in waterbuck are reported.

Animals

Passive endocytosis of sporozoites of Theileria parva in macrophages at 1-2 degrees C.

Phagocytosis by macrophages is generally considered to involve two steps which can be experimentally dissociated: attachment, which is not energy-dependent, and interiorization, which does not occur below 15 degrees C. We have previously reported entry of the sporozoites of Theileria parva into bovine lymphocytes at 2 degrees C by a process of passive endocytosis which appears to depend entirely upon progressive circumferential binding of ligands on the parasite to receptors on the host cell membrane. We report here similar entry into macrophages in vitro at 1-2 degrees C. It is suggested that in vivo small (1 micron) spherical parasites bearing appropriate ligands can invade phagocytic or non-phagocytic host cells by a process of endocytosis that requires little or no expenditure of energy.

Animals

Electron microscopy of Leishmania donovani in splenic aspirates from patients with visceral leishmaniasis during treatment with sodium stibogluconate.

Studies were made of the ultrastructure of amastigotes of Leishmania donovani before and during treatment of patients with sodium stibogluconate. The most consistent effects of treatment on the amastigotes were a reduction in average size, greater irregularity of the cell outline, and a moderate increase in the electron density of the cytoplasm associated with a greater concentration of ribosomes. It is suggested that the drug affects active transport functions or permeability of the plasma membrane.

Adolescent

The entry of sporozoites of Theileria parva into bovine lymphocytes in vitro. Immunoelectron microscopic observations.

In an electron microscopic investigation of the entry of sporozoites of Theileria parva into bovine lymphocytes, the fate of the surface coat of the parasite was traced by immunocytochemical methods. A monoclonal antibody (MAbD1) raised in mice and directed against a surface antigen of sporozoites, was applied to ultrathin frozen sections of bovine lymphocytes infected in vitro. Sites of binding of MAbD1 were localized using a protein A-colloidal gold conjugate as an electron-dense label. The surface of all free sporozoites was labelled. Sporozoites in the process of entering were labelled only on that portion of the membrane not yet tightly bound to the lymphocyte membrane. No label was detected on sporozoites that had completed entry. After fixation with formaldehyde, but not with glutaraldehyde, local areas of labelling were found on lymphocytes in contact with sporozoites and on cells already invaded. The sporozoite organelles, called micronemes, occasionally appeared to contain labelled antigen. No label was found on sporozoites or lymphocytes in control preparations previously exposed to non-specific antibody or treated with protein A-colloidal gold alone. The findings support the conclusion that the sporozoite surface coat, containing the antigen recognized by MAbD1, is shed as the sporozoite enters the host cell.

Animals

Seasonal changes in fine structure of the ductuli efferentes of the ground squirrel, Citellus lateralis (Say).

The morphological changes that the ductuli efferentes undergo during the seasonal breeding cycle of the ground squirrel Citellus lateralis were examined by means of electron microscopy. At the time of spermatogenetic activity the epithelium of the ductuli was composed of highly differentiated principal and ciliated cells. Distinctive cytological features of these cells during this period were the presence of a heterogeneous collection of numerous membrane-bound granules in principal cells and large accumulations of glycogen in ciliated cells. Structurally these cells were specialized for movement of luminal contents and its modification by absorption and possible secretion. With the onset of testicular regression, profound changes occurred in both cell types. Initially the lumen of the ductuli became occluded by masses of apical cytoplasm protruded from principal and ciliated cells as well as by degenerating cells which had been sloughed from the epithelium. This leads eventually, by the time of complete testicular regression, to reduced ductuli containing cells smaller in size with fewer organelles than those present during the period of spermatogenesis. The membrane-bound granules in principal cells and the accretions of glycogen in ciliated cells had now virtually disappeared. There was, however, a dramatic increase in dense inclusions representing deposits of lipofuchsin. As yet the cellular mechanisms controlling and effecting these dramatic changes in morphological appearance are unknown.

Animals

Sertoli cells of the golden-mantled ground squirrel (Spermophilus lateralis): a model system for the study of shape change.

Sertoli cells of the ground squirrel (Spermophilus lateralis), a seasonal breeder, were examined by light and electron microscopy and their structure, particularly the organization of the cytoskeleton, was related to events that occur in the seminiferous epithelium during spermatogenesis. Among the events considered and described are the apical movement of elongate spermatids, withdrawal of residual cytoplasm from germ cells, transport of smooth endoplasmic reticulum (SER) between the base and apex of the Sertoli cells, and sperm release. These events are dramatically evident in this species because the seminiferous epithelium is thin, i.e., there are few germ cells, and both the germ cells and Sertoli cells are large. Sertoli cells of the ground squirrel have a remarkably well developed cytoskeleton. Microfilaments occur throughout the cell but are most evident in ectoplasmic specializations associated with junctions. Intermediate filaments occur around the nucleus, as a layer at the base of the cell, and adjacent to desmosome-like junctions with germ cells. Intermediate filaments, together with microtubules, are also abundant in regions of the cell involved with the transport of SER, in cytoplasm associated with elongate spermatids, and in processes that extend into the residual cytoplasm of germ cells. Our observations of ultrastructure are consistent with the hypothesis that Sertoli cell microtubules are involved with the movement of germ cells within the seminiferous epithelium, and further implicate these structures as possibly playing a role in the retraction of residual cytoplasm from germ cells and the intracellular transport of SER. The abundance and organization of intermediate filaments suggest that these cytoskeletal elements may also be involved with events that occur during spermatogenesis.

Animals

Salivary gland of the tick vector of East Coast fever. III. The ultrastructure of sporogony in Theileria parva.

Sporogony of the sporozoan Theileria parva in the salivary gland of the tick vector of East Coast fever was studied in electron micrographs. The findings differ in several respects from previous interpretations based upon light microscopy. Cytokinesis of the primary sporoblast to form secondary and tertiary sporoblasts is not substantiated. Instead it is suggested that the parasite develops as a ramifying, multinucleate syncytium rapidly increasing in size and complexity until it gives rise to myriad sporozoites in a terminal episode of cytoplasmic fission. The proliferating nuclei initially occupy peripheral lobules that are continuous with a central labyrinth of branching and anastomosing processes which present a very large surface area for interchange of metabolites with the host cell cytoplasm. The membrane of the labyrinth is rich in cytostomes, but no evidence if found to bulk uptake of host cytoplasmic matrix or organelles into food vacuoles. Rhoptries are the first of the polar organelles of the parasite to develop and are associated with dense plaques irregularly distributed on the inner aspect of the parasite membrane. Micronemes form independently of the rhoptries at a later stage. After 3-4 days of tick feeding, sporogeny is complete and the infected salivary gland cell contains up to 50, 000 spherical or ovoid sporozoites about 1 micrometer in diameter. These are limited by a simple plasma membrane. The inner layer of the 'pellicle', the polar ring, and the conoid described for zoites of other Apicomplexa are lacking. Maturational changes are noted in sporozoites after sporogony is completed. Micronemes appear to increase in size, and possibly in number, from days 3-5 and the majority take up positions immediately subjacent to the plasmalemma.

Animals

Salivary gland of the tick vector of East Coast fever. IV. Cell type selectivity and host cell responses to Theileria parva.

Responses of cells in the tick salivary gland to parasitism by Theileria parva were studied by electron microscopy. The gland is composed of three distinct types of acini (I, II, III) which together include ten or more different cell types. Of some 30 infected cells observed in the present study, all were E-cells of acinus III. The parasite thus exhibits a high degree of selectivity for acinus and cell type. The glandular cell invaded undergoes massive hypertrophy and accumulates glycogen deposits in its cytoplasm which may serve as an energy source for the growing intracellular parasite. As synthesis of its secretory material declines the product is packaged in progressively smaller secretory granules. The extensive arrays of endoplasmic reticulum are dismantled and eliminated in autophagic vacuoles. Excess secretory granules are also broken down by crinophagy. After 4 days, sporogony is completed and the host cell contains 30,000-50,000 sporozoites in an electron-lucent cytoplasm largely devoid of cytomembranes and secretory granules. Mitochondria are still present and normal in appearance. The loss of basophilia and secretory granules observed heretofore by light microscopy have been attributed to ingestion and destruction of host organelles by the parasite. The pallid appearance of the cytoplasm has been interpreted as a sign of impending degeneration of the host cell. In electron micrographs no ingestion of organelles by the parasite or degenerative changes were found. The host cell clearly remains viable and metabolically active throughout sporogony. The striking changes in its ultrastructure result from active elimination of organelles and inclusions by the host cell itself in response to parasitism.

Animals

Localization of tritiated 1-(2, 4-dichlorobenzyl)-1H-indazole-3-carboxylic acid ([3H] AF 1890) in rat testis using freeze-drying autoradiography.

Tritiated 1-(2, 4-dichlorobenzyl)-1H-indazole-3-carboxylic acid ([3H]AF 1890) was localized within rat testis using freeze-drying autoradiographic techniques. Ligated seminiferous tubule preparations were exposed in vitro to 80 microCi of [3H]AF 1890 in the absence or presence of excess unlabeled AF 1890 for 30 min at 31 degrees C. Autoradiographs showed a generalized distribution of silver grains over the seminiferous epithelium, in the tubular lumen, and in the interstitial spaces. In addition, clusters of grains were concentrated over the seminiferous epithelium in regions of spermatid heads and tails and in the basal portion of the Sertoli cell cytoplasm, and over Leydig cells in the interstitial space. The generalized distribution was reduced and the grain clusters eliminated by incubation in an excess of unlabeled AF 1890. [3H]inulin was used to assess the effectiveness of the tubular ligation because inulin does not normally penetrate the blood-testis barrier. This extracellular marker was not localized in the tubular lumen or in the seminiferous epithelium. Therefore, the labeling observed with [3H]AF 1890 indicates that this compound crosses the blood-testis barrier.

Animals

The entry of sporozoites of Theileria parva into bovine lymphocytes in vitro. Electron microscopic observations.

The entry of sporozoites of Theileria parva into bovine lymphoid cells in vitro was studied with the electron microscope. Endocytosis is completed in less than 10 min. No local mobilization of actin or other cytoskeletal elements is detected in the cytoplasm of the cell being invaded and no engulfing pseudopods are formed. At the site of initial contact, the membranes of parasite and host cell come into very close apposition. As the zippering up of the membranes spreads laterally, the sporozoite sinks into a progressively deepening recess in the surface of the host cell until the rim of the invagination closes and fuses over the parasite. The observation that sporozoites are interiorized at 1-2 degrees C as well as at 37 degrees C suggests that endocytosis depends mainly upon a ligand-receptor interaction of the parasite and host cell membranes and requires little energy. Sporozoites may enter in any orientation, unlike other sporozoan parasites in which the membrane overlying an apical complex is invariably the site of attachment. 24 h after entry, the sporozoite is located in the Golgi region and the investing host cell membrane acquired during endocytosis has disappeared. The Golgi complex has been activated to form small lysosomes which gather around the parasite but are ineffective for lack of a membrane which they can fuse. It is suggested that removal of the investing host-cell membrane permits the parasite to evade destruction by the phagolysosomal system of the host cell. Persistence of micronemes after entry of the sporozoite and their subsequent disappearances invites the speculation that these parasite organelles may play a role in dispersal of the invaginated host cell membrane.

Animals

Salivary gland of the tick vector (R. appendiculatus) of East Coast fever. I. Ultrastructure of the type III acinus.

The brown ear tick Rhipicephalus appendiculatus is the vector for East Coast fever, a disease that seriously limits livestock production in East Africa. The sporozoites of the infectious agent Theileria parva develop in the tick salivary gland. This paper describes the organization of the type III acinus of the gland and establishes unambiguous ultrastructural criteria for identification of the three secretory cell types: the d-cell, e-cell and f-cell. These observations are basic to exploration of possible cell-type specificity of the invading theileria and other aspects of host-parasite relations.

Animals

Salivary gland of the tick vector (R. appendiculatus) of East Coast fever. II. Cellular basis for fluid secretion in the type III acinus.

Fluid balance is a major physiological problem for hematophagous ticks. To maintain osmotic balance they must conserve water for prolonged periods while seeking a mammalian host, and they must eliminate a very large volume of excess fluid taken in during a relatively short period of feeding. This is accomplished in part by modification of the salivary gland during 7-10 days of feeding to secrete a copious saliva which is pumped into the bovine host. This function has previously been attributed to certain interstitial epithelial cells of the type III acinus which differentiate in the course of feeding into cells reminiscent of those of the avian salt gland. The ultrastructural changes in the type III acinus during the blood-meal were studied. In addition to the differentiation of the interstitial cells, this paper describes a remarkable sequence of changes in external form and internal organization of the e-cells. This results in their transformation from typical protein-secreting glandular cells to transport cells having myriad basolateral processes interdigitating with those of the ablumenal interstitial cells to form a basal labyrinth comparable to that of other fluid-transporting epithelia. The findings are discussed in relation to various postulated mechanisms for fluid and electrolyte transport.

Animals

Immunocytochemical localization of androgen-binding protein in the male rat reproductive tract.

The localization of androgen-binding protein (ABP) in the reproductive tract of young adult male rats was studied with the peroxidase-antiperoxidase technique using frozen sections and light microscopy. Within the seminiferous tubules, a positive reaction was noted in the apical portion of the epithelium, apparently in spermatids and/or Sertoli cells. ABP was localized in granules in the apical cytoplasm of the principal epithelial cells of the proximal part of the caput epididymis and in the epithelial cells of the ductuli efferentes. The cells in the distal part of the caput as well as the corpus and cauda of the epididymis did not contain ABP. Numerous coated vesicles and multivesicular bodies were present in the supranuclear cytoplasm of the epididymal epithelium where ABP was taken up. The results indicate that ABP is taken up from the lumen by epithelial cells of the ductuli efferentes and proximal part of the caput epididymis.

Androgen-Binding Protein

Structure and function of the undulating membrane in spermatozoan propulsion in the toad Bufo marinus.

Accessory fibers in most sperm surround the axoneme so that their function in propulsion is difficult to assess. In the sperm of the toad Bufo marinus, an accessory fiber is displaced from the axoneme, being connected to it by the thin undulating membrane in such a way that the movement of axoneme and accessory fiber can be viewed independently. The axoneme is highly convoluted in whole mounts, and the axial fiber is straight. Cinemicrographic analysis shows that it is the longer, flexuous fiber, the presumed axoneme, that move actively. The accessory fiber follows it passively with a lower amplitude of movement. The accessory fiber does not move independent of the axoneme, even after demembranation and reactivation of the sperm. On the basis of anatomical relations in the neck region, it appears that the accessory fibers of amphibians are analogous to the dense fibers of mammalian sperm. SDS polyacrylamide gel electrophoresis of demembranated toad sperm tails reveals two principal proteins in addition to the tubulins, the former probably arising from the accessory fibers and the matrix of the undulating membrane. The function of displacing an accessory fiber into an undulating membrane may be to provide stiffness for the tail without incurring an energy deficit large enough to require a long middle piece. A long middle piece is not present in toad sperm, in contrast to those sperm that have accessory fibers around the axoneme. However, the toad sperm suffers a reduction in speed of about one-third, compared with the speed expected for a sperm without an undulating membrane.

Animals

Morphology of the epithelium of the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the adult male rabbit.

The fine structure of the epithelium lining the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the rabbit has been investigated. In the ductuli efferentes the epithelium is composed of two cell types, principal cells and ciliated cells. The latter type is distinguished from principal cells by the presence of cilia projecting into the lumen and the position of the nucleus in the apical half of the cell. Principal cells in this segment are characterized by micropinocytotic vesicles on the surface plasma membrane and a variety of small dense bodies scattered throughout the cytoplasm. In the ductus epididymidis basal cells replace ciliated cells as the second cell type, but differences between various segments of the epididymis are related to the fine structure of the principal cells. In the proximal caput epididymidis (Nicander's region 1) the principal cells are tall with long microvilli. They typically contain a small Golgi apparatus and a cluster of dense bodies adjacent to the nucleus. In the distal caput epididymidis (Nicander's regions 2-5) the apical cytoplasm of principal cells is filled with numerous micropinocytotic vesicles and large multivesicular bodies; these features are interpreted as signs of absorptive activity. The multivesicular bodies are absent from the cytoplasm of principal cells in the corpus epididymidis (Nicander's region 6) and, instead, numerous elements of smooth endoplasmic reticulum, a large Golgi apparatus, lipid droplets and dense bodies characterize principal cells in this segment. Towards the proximal cauda epididymidis (Nicander's region 7), the number of dense bodies (lysosomes) in the cytoplasm increases considerably. In the globose cauda (Nicander's region 8), the principal cells are reduced in height, and in addition to the features described in region 7, are characterized by a concentric array of rough endoplasmic reticulum in the basal cytoplasm. These observations are discussed in relation to the role of the epididymis in promoting the maturation and survival of spermatozoa.

Animals

Changes in distribution of nuclear pores during differentiation of the male germ cells.

Changes in number of nuclear pores in different states of physiologica activity have been reported, but little is known about changing patterns of distribution in the course of cell differentiation. Pore distribution in male germ cells was studied in freeze fracture preparations of immature and mature rodent testis. As in other somatic cells, pores were uniformly and apparently randomly distributed in Sertoli cell nuclei. The nucleus of gonocytes and spermatogonia showed varying degrees of pore clustering. Spermatocytes invariably exhibited very striking pore aggregation with close hexagonal packing in pore-rich areas, and large pore-free areas. In early spermatids, pores appeared to be randomly distributed. As the acrosome formed and spread over the apical pole of the nucleus, pores disappeared ahead of its advancing margin and became more concentrated in the post-acrosomal region. The relationship of pore complexes to the chromosomes and the role of the fibrous lamina are discussed. The question as to whether the changing patterns observed involve movement of pores within fluid nuclear membranes, or a dissolution and reformation of new pores remains unanswered.

Animals