Towards the characterization of plasmatocyte heterogeneity in Drosophila melanogaster

• Author(s): Leitão, Alexandre Castanho Barata
• Date: 2009
• Persistent ID: http://hdl.handle.net/10451/1442
• Origin: Repositório da Universidade de Lisboa
• Subject(s): Imunologia; Sistema imunitário; Drosophila melanogaster; Teses de mestrado
• Document Type: Master thesis
• Language: English
• Advisor(s): Sucena, José Élio da Silva
• Contributor(s): Repositório da Universidade de Lisboa

Description

Tese de mestrado, Biologia (Biologia Evolutiva e do Desenvolvimento), 2009, Universidade de Lisboa, Faculdade de Ciências

Resumo alargado em português disponível no documento

Insect innate immune system can be partitioned into humoral and cellular defense mechanisms. However, both branches of immune system are interconnected, acting in a synergistic way. The study of these interconnections is an important step to comprehend the immune response as an integrated system. In Drosophila genus 95% of 'blood cells' (hemocytes) are plasmatocytes. These cells participate in numerous processes during development and immune response. Throughout embryogenesis and pupal stage plasmatocytes play an important role in tissue remodeling, phagocytizing dead cells and synthesizing extracellular matrix. During an immune response they are responsible for production of antimicrobial peptides, phagocytosis and aggregation of pathogens. In addition, it is possible to find two other types of hemocytes in Drosophila's hemolymph, crystal cells and lamellocytes, both with very specialized functions in immune response. Crystal cells constitute 5% of hemocytes in Drosophila and lamellocytes are only found in larvae upon infection with large pathogens, such as wasp eggs. Classification of these three hemocyte types is based on morphological and biochemical criteria. However, some new studies have begun to characterize RNA and protein expression in Drosophila's hemocyte classes. Here, we want to go further in the genetic characterization of plasmatocytes with the purpose off asking if there are different subpopulations of plasmatocytes performing different functions during immune response. For this propose we used flow cytometry technique to analyze gene expression in Drosophila larvae plasmatocytes. Our results show that two out of the five GAL4 lines analyzed drive expression of GFP in subpopulations of Drosophila larval plasmatocytes. This observation indicates that plasmatocytes do not form a homogeneous population of cells in Drosophila's larvae hemolymph. We then used Fluorescence Activated Cell Sorting (FACS) to sort and perform in vitro experiments with hemese positive and hemese negative plasmatocytes independently. In vitro analysis confirmed that both subpopulations correspond to previous plasmatocyte descriptions. We hypothesized that one of these two different subpopulations of plasmatocytes is responsible for modulation of melanization, an immune response of insects. However, the results obtained in our specific in vitro setting did not support this hypothesis but further work is needed to ascertain this matter in a definitive way.