The focus of my laboratory is the understanding of the molecular mechanisms regulating cell determination during mammalian hematopoiesis and embryonic development. Our approach to these problems extensively utilizes the mouse as a model system. In particular, we exploit the generation of transgenic and knockout mice as a method to study the differentiation and effector functions of eosinophils. My laboratory collaborates extensively with Dr. Nancy Lee (Mayo Clinic Scottsdale) to achieve our experimental goals. Eosinophils are a type of white blood cell (granulocyte) involved in host defenses to parasites and often increase in numbers as a consequence of allergic inflammation (e.g., hayfever, asthma etc.). We have created several novel mouse models of human diseases characterized by eosinophil infiltration of specific tissues. These mouse models allow us to characterize at a molecular and cellular level the specific pathways leading to pathology.
Molecular Characterization of Genes Expressed in Mouse Eosinophils
In an attempt to understand regulatory pathways used to achieve a terminally differentiated cell type during murine hematopoiesis, we are cloning genes that are expressed in the eosinophil and its progenitor cells. Eosinophils play pivotal roles in inflammatory responses to both infection and injury. These responses are mediated primarily through the expression of gene products specific to the eosinophil granule.
The eosinophil contains numerous specific granules that sequester the toxic granule proteins prior to release. The cationic property of these proteins yields the characteristic crimson staining of the eosinophil with the acidic aniline dye, eosin. In humans, four major proteins have been purified and cloned from the human eosinophil specific granule. These include the major basic protein (MBP), eosinophil derived ribonuclease (EDN), eosinophil cationic protein (ECP), and eosinophil peroxidase (EPO).
One of the goals of the laboratory is the identification, characterization, and cloning of the eosinophil granule proteins genes in the mouse. We have isolated and cloned the genes coding for murine MBP, EPO and three eosinophil associated granule ribonucleases (EARs). We are continuing studies of the transcriptional, and possibly coordinate, regulation of these genes during eosinophilopoesis.
These knock-out mice will also permit a longer-term goal of "humanizing" the mouse eosinophil. In this project, clones for the human granule proteins are used with eosinophil-specific promoters to generate transgenic mice expressing these human proteins. Selective breeding with the knock-out animals thus allows a "tit for tat" replacement of the mouse proteins with their human counterparts. The hope is that these studies will create mouse models much more characteristic of human disease.
Animal Models of Eosinophil-Mediated Inflammation
Respiratory inflammation is often accompanied by the release of immunoregulative cytokines and the recruitment of proinflammatory cells to the lung. The infiltration of the bronchial mucosa by activated cells, and the resulting inflammation, is thought to play a major role in the reduction of bronchopulmonary function associated with many diseases, including asthma and cystic fibrosis. Observations from a number of laboratories have shown that the prominent groups of cells present in the lungs of asthmatics are activated T lymphocytes, macrophages, and eosinophils. In particular, eosinophilia and eosinophil recruitment to the lung have been implicated in the pathogenesis of asthma. We are currently using transgenic and gene knockout mouse models to examine the role(s) of eosinophils during immune-mediated inflammation. In addition, we are exploiting established murine models of respiratory inflammation to investigate the exacerbation of airway inflammation by environmental stimuli. These models include ozone exposure (a major air pollutant), allergen treatment, and infection with the influenza virus, each of which exacerbates the symptoms of asthma patients.