research
We are interested in the signaling mechanisms underlying two important and highly dynamic cellular responses:
- Neutrophil adhesion, polarity and chemotaxis
Cell migration is a highly integrated multi-step process that mediates embryonic development, contributes to tissue repair and regeneration, and drives disease progression in cancer, mental retardation, atherosclerosis, and arthritis. Although the understanding of cell-extracellular matrix (ECM) adhesion in slow-migrating mesenchymal cells has been significantly improved in the past two decades, this key step of cell migration remains poorly defined in highly polarized and rapidly moving amoeboid cells, such as leukocytes, hematopoietic stem cells, and tumor cells (Fig. 1). Because of the rapid membrane turnover rates during amoeboid movement, cell adhesion to ECM substrate is viewed as very limited.
Our recent studies contradict this notion and suggest a complex and highly dynamic pattern of adhesion during neutrophil polarization and chemotaxis. We found that cell adhesion is required for neutrophils to establish and maintain polarity. In addition, we have begun to uncover the potential key regulatory components of adhesion. We will employ the tools of cell biology, biochemistry, genetics, proteomics, and biomechanics to dissect the molecular basis of cell adhesion in neutrophils during chemotaxis. These experiments will yield a new level of understanding of the mechanisms that control cell adhesion in neutrophils and in other amoeboid cells. The results from this study will also have direct relevance to the treatment of multiple human diseases, including autoimmune disorders and cancer.
Fig. 1. Human neutrophils expressing actin-YFP were exposed to a gradient of chemoattractant delivered by a micropipette. Cells responded to the stimulation by accumulating actin at the front and migrating all the way to the pipet tip. The fluorescence and DIC images show morphology of cells 5 min after attractant stimulation. This is one of the assays used in the lab to assess the signaling mechanism underlying neutrophil polarity and chemotaxis. Bar, 10 μm
Fate decisions of human embryonic stem cells
Nearly 20 years after murine embryonic stem cells (mESC) were isolated, the first report of the derivation of human embryonic stem cells (hESCs) in 1998 spawned the field of hESC research. Although this field is only in its infancy, hESCs have already been shown to be capable of long-term self-renewal in culture and have remarkable potential to develop into many different cell types in the body (known as ˇ°pluripotencyˇ±). They therefore represent a theoretically inexhaustible source of precursor cells to treat degenerative, malignant, or genetic diseases, or injury due to inflammation, infection, and trauma. Meanwhile, hESCs are an invaluable research tool to study human development, both normal and abnormal, and can serve as a platform to develop and test new drugs.
Our long-term goal is to define new conditions and molecular programs that govern fate decisions of hESCs. The knowledge is essential if we are ultimately to use these cells for therapy. To dissect the mechanism underlying hESC long-term self-renewal, we screened a small collection of pharmacological inhibitors and identified both potential positive and negative regulators of hESC pluripotency (Fig. 2). This pilot study provided proof-of-concept for applying large-scale library screening to the study of hESCs. Accordingly, we will develop cell-based high-throughput assays by establishing hESCs containing enhanced green fluorescence protein (EGFP) reporters for pluripotency and directed differentiation. We will then use these assays to conduct large-scale library-screening to uncover more regulators of hESC long-term self-renewal. The results of these studies will markedly improve our knowledge of the molecular mechanisms underlying hESC fate determination and may contribute to effective strategies for tissue repair and regeneration.
Fig. 2. Immunofluorescence studies of Oct-4 (green) in hESCs (H9 line) treated with a specific inhibitor of a protein kinase (6 days). Because inhibition of the kinase leads to the downregulation of pluripotency genes, it is a potential positive regulator of hESC pluripotency. The fluorescence images were merged with the phase-contrast images. Bar, 500 μm
