Research Projects
Lymphopoiesis and Apoptosis in the Development of Autoimmune Diseases

Objective and Significance:

Rheumatoid arthritis (RA) is a common chronic autoimmune disease for which at present there is no effective treatment. A widely accepted experimental model for human rheumatoid arthritis is the collagen-induced arthritis (CIA) model in genetically susceptible DBA/1 mice. Extensive studies have focused on aspects of lymphocyte activation and autoantibody production, but the underlying mechanisms of prolonged B cell survival during the pathogenesis of RA still remain elusive. The proposed work will examine the regulation of peripheral B cell development and role of B cell apoptosis in the development of autoimmune arthritis and provide further insight in understanding B cell apoptosis and its modulation during normal development and autoimmunity.

Research Plan and Methodology:

(1) Collagen-induced arthritis mouse model (CIA)

Male DBA/1 mice 10-12 week of age will be immunized with 100 µg of bovine type II collagen in Freund's complete adjuvant intradermally at the base of the tail. The mice are observed for the development of arthritis and evaluated by histopathology.

(2) B cell development and apoptosis in CIA

Single cell suspensions from the bone marrow, spleen, mesenteric lymph nodes and peritoneal lavage will be prepared from normal and CIA mice.

The total number and frequency of defined B cell subsets at various differentiation stages will be immunolabeled with B lineage markers and analyzed by multi-color flow cytometry. B cells, especially B1 cells, from spleen and from peritoneal cavity will be phenotypically characterized. Apoptosis will be evaluated with Annexin-V binding and propidium iodide staining and analyzed by flow cytometry.

Research Plan and Methodology:

Research Plan

First, we shall analyze the effects of PA toxins on the changes in transcriptional outputs as measured by various reporter assays in airway epithelial cells. These effects will then be corelated changes in cytokine productions as measured by ELISA. Subsequently, the role of intracellular signaling pathways (PKA, PKC, MAP kinases etc.) in these effects will be examined using specific inhibitors.

Methodology

1. Reporter gene constructs will be generated and transfected with human airway epithelial cell lines (A549 and BEAS-2B). The luciferase activity of transfected lysed cells will be determined using a commercially available kit (Promega).
2. The release of inflammatory cytokines, such as IL-8 and GM-CSF, will be measured in the supernatants of PYO- or 1-HP-stimulated non-transfected and transfected cells by use of a sandwich ELISA according to the manufacturer's instructions (R & D).
3. Western blot analysis for various antibodies in the signaling pathways will be performed in total cell lysates, cytosolic and nuclear fractions. For activation, anti-phospho-Ab will be used in conjunction with the pan Ab. ECL detection will be used and quantification will be done by densitometry.
4. Immunocytochemistry will also be performed in coverslip-grown cells using EnVision system (DAKO). The reaction products will be visualized using DAB.

Professor LW Lu, Department of Pathology

Biography
liweilu@hku.hk

For more information or to express interest for this project, please email the supervisor or the specified contact point in the project description.  Interested candidates are advised to enclose with your email:

1. your CV,
2. a brief description of your research interest and experience, and
3. two reference letters (not required for HKUMed UG students seeking MRes[Med]/URIS projects).

Information on the research programme, funding support and admission documentations could be referenced online at the Research Postgraduate Admissions website. General admission enquiries should be directed to rpgmed@hku.hk.

HKUMed MBBS students interested in the Master of Research in Medicine (MRes[Med]) programme may visit the programme website for more information.  

HKUMed UG students interested in the Undergraduate Research Internship Scheme (URIS) may visit the scheme’s website for more information.