Food ingestion by planarian intestinal phagocytic cells - a study by scanning electron microscopy

Ishii, Saburo; Sakurai, Takashige

doi:10.1007/978-94-011-2775-2_27

Cited by 6 publications

(6 citation statements)

References 15 publications

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“…The planarian intestinal epithelium is comprised of two cell types: secretory goblet cells that release digestive enzymes after food ingestion, and absorptive phagocytes that engulf food for intracellular digestion [ 90 - 92 ]. Intestinal phagocytes retain dyes, beads, and other compounds for up to several weeks after they are ingested [ 17 , 93 - 96 ]. We recently developed a protocol in which animals are fed iron beads and dissociated into single cell suspensions, enabling the purification of phagocytes by magnetic sorting (Figure 1 A) [ 23 ].…”

Section: Resultsmentioning

confidence: 99%

Generation of cell type-specific monoclonal antibodies for the planarian and optimization of sample processing for immunolabeling

2014

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BackgroundEfforts to elucidate the cellular and molecular mechanisms of regeneration have required the application of methods to detect specific cell types and tissues in a growing cohort of experimental animal models. For example, in the planarian Schmidtea mediterranea, substantial improvements to nucleic acid hybridization and electron microscopy protocols have facilitated the visualization of regenerative events at the cellular level. By contrast, immunological resources have been slower to emerge. Specifically, the repertoire of antibodies recognizing planarian antigens remains limited, and a more systematic approach is needed to evaluate the effects of processing steps required during sample preparation for immunolabeling.ResultsTo address these issues and to facilitate studies of planarian digestive system regeneration, we conducted a monoclonal antibody (mAb) screen using phagocytic intestinal cells purified from the digestive tracts of living planarians as immunogens. This approach yielded ten antibodies that recognized intestinal epitopes, as well as markers for the central nervous system, musculature, secretory cells, and epidermis. In order to improve signal intensity and reduce non-specific background for a subset of mAbs, we evaluated the effects of fixation and other steps during sample processing. We found that fixative choice, treatments to remove mucus and bleach pigment, as well as methods for tissue permeabilization and antigen retrieval profoundly influenced labeling by individual antibodies. These experiments led to the development of a step-by-step workflow for determining optimal specimen preparation for labeling whole planarians as well as unbleached histological sections.ConclusionsWe generated a collection of monoclonal antibodies recognizing the planarian intestine and other tissues; these antibodies will facilitate studies of planarian tissue morphogenesis. We also developed a protocol for optimizing specimen processing that will accelerate future efforts to generate planarian-specific antibodies, and to extend functional genetic studies of regeneration to post-transcriptional aspects of gene expression, such as protein localization or modification. Our efforts demonstrate the importance of systematically testing multiple approaches to species-specific idiosyncracies, such as mucus removal and pigment bleaching, and may serve as a template for the development of immunological resources in other emerging model organisms.Electronic supplementary materialThe online version of this article (doi:10.1186/s12861-014-0045-6) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

Generation of cell type-specific monoclonal antibodies for the planarian and optimization of sample processing for immunolabeling

2014

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“…In our results, receptor-mediated endocytosis seemed to be the only efficient process of food intake by the phagocytes. So far we have not observed in Stenostomum any evidence for phagocytosis or phase-fluid endocytosis, commonly seen in other platyhelminths (Bowen et al 1974;Ishii 1972;Pascolini & Gargiulo 1975;Gargiulo et al 1976;Hori 1985;Antoniazzi 1989;Ishii & Sakurai 1991).…”

Section: Gastrodermismentioning

confidence: 90%

“…These pits and vesicles were rapidly formed upon stimulation by the latex beads and are very likely associated with receptor-mediated endocytosis. By contrast, Ishii & Sakurai (1991) stated that in planarians, it is likely that small particles, including latex beads (< 1 pm), would be ingested through a non-specific process, not dependent on cell receptors. In our results, receptor-mediated endocytosis seemed to be the only efficient process of food intake by the phagocytes.…”

Section: Gastrodermismentioning

confidence: 99%

Studies on Stenostomum grande Child, 1902 (Platyhelminthes, Catenulida): fine structure of the digestive tract and the endocytotic activity of the gastrodermis

Antoniazzi

Silveira

1996

Acta Zoologica

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Antoniaui, M. M. & Silveira, M. 1996. Studies on Stenostomum grande Child, 1902 (Platyhelminthes, Catenulida): fine structure of the digestive tract and the endocytotic activity of the gastrodermis. Acta Zoologica (Stockholm) 77: 25-32.The digestive tract and its endocytotic activity in the catenulid Stenosromum g r a d e were studied by electron microscopy. The pharynx was typical of the simplex type. At the mouth, between the integumental epithelium and the pharyngeal epithelium proper, was a transition zone. Among the epithelial cells of this transition were monociliated sensory cells and the necks of bucco-pharyngeal secretory cells of two types. The pharyngeal epithelium proper was densely ciliated, with long ciliary rootlets and mitochondria. It was surrounded by two layers of muscles. The gastrodermis consisted of phagocytes and typical secretory Minotian cells. It was underlain by a delicate basal lamina and muscle fibers. Distinctive of the phagocytes was the presence of differentiated cilia, cup-shaped mitochondria, and vacuoles with dense inclusions. Morphological differences between pharyngeal and gastrodermal cilia suggest functional differences. Experiments using latex beads as tracers and the identification of acid phosphatase in cytoplasmic vacuoles pointed to a high level of endocytotic and digestive activity in the phagocytes. Our data demonstrate that the basic structure of the digestive tract in S. g r a d e conforms well to that of other free-living platyhelminths, but it does have ultrastructural peculiarities.

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“…Lately, planarians have become one of the model references for studying stem cell biology, but these flatworms may also be useful for studying other biological issues. More than 20 years ago, M. Morita and T. Sakurai (Ishii and Sakurai, 1991 ; Morita, 1991 ; Morita and Collins, 1995 ) noted the peculiar role of certain cells called “reticular cells” that could mediate the early immune response. These high-degree mobility cells could recognize foreign material, such as bacteria, as early as 8 h after their introduction.…”

Section: From Here To Eternity: the Planarians Experiencementioning

confidence: 99%

Unconventional animal models: a booster for new advances in hostâ€”pathogen interactions

Conti

Abnave

Ghigo

2014

Front. Cell. Infect. Microbiol.

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Food ingestion by planarian intestinal phagocytic cells - a study by scanning electron microscopy

Cited by 6 publications

References 15 publications

Generation of cell type-specific monoclonal antibodies for the planarian and optimization of sample processing for immunolabeling

Generation of cell type-specific monoclonal antibodies for the planarian and optimization of sample processing for immunolabeling

Studies on Stenostomum grande Child, 1902 (Platyhelminthes, Catenulida): fine structure of the digestive tract and the endocytotic activity of the gastrodermis

Unconventional animal models: a booster for new advances in hostâ€”pathogen interactions

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