Study of Hypoxia on Embryogenesis, Pharmaceutical Testing and Stem Cell Regulation Using Drosophila Model

摘要

Drosophila melanogaster has been used to study human disease as a model organism for many years. Many basic biological, physiological, and neurological properties are conserved between mammals and fly. We investigated its applications in the study of the impact of environmental stress on embryogenesis and its compensating mechanism, its uniqueness and powerfulness in modern pharmaceutical testing and screening, and its application in gene function identification.;First we directly investigated Drosophila embryo development in vivo inside a customized microfluidic device with an established local oxygen gradient on a micrometer scale. When the embryos were placed in various conditions, two of the key developmental activities, the germ band shortening and the tail retraction, were examined during the embryogenesis. The time-lapse live cell imaging technique was used to monitor the cell morphology changes and pattern migration with the help of green fluorescence protein markers. Our results show that the examined activities during the Drosophila embryogenesis are highly sensitive to oxygen concentrations. Using this information, we presented a model to estimate the oxygen permeability across the Drosophila embryonic layers for the first time. Secondly, the Drosophila testis was used to evaluate the basic therapeutic mechanisms of the active components of several traditional Chinese medicines (TCM) that is known related to animal genital system and sexual function. Specifically, we investigated the effect of the compounds that were extracted from above-mentioned Chinese medicines on Drosophila germline stem cells (GSCs) by quantifying the GSCs mitotic activity and GSC number. Our results showed that, flies have a significantly higher cell cycle index when fed at certain concentration of icariin and Tanshinone IIA, the primary active component of YYH and DS, respectively. Other tested concentrations of extract produced either toxicity or insignificant effects on the mitotic activity. This indicates their function of promoting the GSCs mitosis. At last, we analyzed the expression and localization of two polarity genes throughout the cell cycle, and investigated how they affected mitotic spindle dynamics in asymmetric stem cell divisions. In stem cell divisions, it is critical to maintain tissue homeostasis by balancing the number of stem cells and progenitor cells. Improperly balancing may result in tumorigenesis due to tissue hyper-proliferation or tissue ageing due to tissue degeneration. Previous studies show that cyst stem cells (CySCs) in Drosophila testis divide asymmetrically. This behavior is ensured by the stem cell mitotic spindle repositioning, during which one of the spindle poles always moves close to the stem cell niche (a.k.a. hub cells) near the onsite of anaphase. Known as polarity proteins, the apically localized Par polarity complex, containing Bazooka (Baz; homolog of Par-3 in D. melanogaster), its binding target atypical Protein Kinase C (aPKC), and Par-6, are widely reported to be crucial in polarized cell epithelium and asymmetric cell division in multiple stem cell systems. We found that Baz and aPKC are required in Drosophila CySC asymmetric cell division.