1. BAFF and Autoimmune Disease
Samia Ragheb, Ph.D.
Wayne State University
Department of Neurology
Department of Immunology & Microbiology

2. MG is a B-cell Mediated Disease
1. AChR-specific antibodies are present in the serum of 85% of all patients with generalized MG; these antibodies are responsible for the pathology of MG, leading to impaired neuromuscular transmission and subsequent muscle weakness
2. In vitro, lymphocyte cultures from MG patients produce AChR-specific antibodies
3. EAMG can be induced in lab animals; serum anti-AChR antibodies are present in these animals
4. EAMG can be passively transferred, via antibodies, from immune animals to naïve animals and from man to mouse

3. Overexpression of CXCL13, CD23, and Bcl-2 in the Myasthenic Thymus
CXCL13: promotes B-cell migration and homing of B-cells to lymphoid follicles
CD23: promotes the survival and differentiation of germinal center B-cells through a mechanism that involves upregulation of Bcl-2
Bcl-2: pro-survival molecule that inhibits apoptosis

4. BAFF = TNFSF13b
Myeloid cells produce and secrete Baff (B-cell activating factor of the tumor necrosis factor superfamily)
A membrane-bound form of Baff is also expressed on the surface of myeloid cells; these include monocytes, macrophages, and dendritic cells
Baff transgenic animals exhibit hypergammaglobulinemia, lymphoproliferation, and B-cell hyperplasia; and they develop autoimmune disease
In Baff deficient animals, there are defects in peripheral B-cell maturation, and there are decreased levels of circulating immunoglobulins

5. Serum BAFF Levels

6. Effect of Clinical Extent and Severity on BAFF Levels

7. Effect of Presence or Absence of Serum Anti-AChR Antibody on BAFF Levels

8. Receptors for BAFF
Baff-R deficient mice lack marginal zone and follicular B-cells
TACI deficient mice have an increased number of peripheral B-cells, they develop autoimmune disease, and they exhibit fatal lymphoproliferation
The pro survival signals of Baff are mediated by the Baff-R, whereas the interaction of Baff with TACI delivers inhibitory signals

9. Immunofluorescent Staining of Peripheral Blood Mononuclear Cells
CD3: mouse IgG1-FITC
CD20: mouse IgG2a-FITC
Baff-R: goat IgG anti human Baff-R + rabbit F(ab)’2 anti goat IgG-Cy5
TACI: biotinylated goat IgG anti human TACI + streptavidin-Cy5

10. Cell-Surface Expression of BAFF-R and TACI

11. Thangarajh et al.:
MG thymus with hyperplasia: macrophages express Baff
Germinal center: B-cells express Baff-R
Plasma levels of Baff were not different
No differences in the percentages of BCMA, Baff-R, or TACI-positive B-cells
Li et al.:
Increased percentage of CD19+ Baff-R+ B-cells
Kim et al.:
Serum Baff levels are higher in MG patients

12. Myeloid Cells Produce and Secrete BAFF
Monocyte
Dendritic cell
B-cell
Baff

13. Immunofluorescent Staining of Peripheral Blood Monocytes
CD14: mouse IgM-FITC
Baff: mouse IgG1 anti human Baff + goat F(ab)’2 anti mouse IgG-PE

14. BAFF Production by Adherent Monocytes

15. BAFF Production by Adherent Monocytes

16. BAFF Production by THP-1 Cells
THP-1: human acute monocytic leukemia
1 year old male
HLA: A2, A9, B5, DRw1, DRw2

17. BAFF Production by THP-1 Cells

18. Enrichment of Monocytes by Immunomagnetic Separation
PBMC are treated with a mixture of mouse anti CD2, CD7, CD16, CD19, CD56, and CD235a followed by anti mouse IgG-coated magnetic beads

19. BAFF Production by Monocytes

21. Dendritic Cell Lineages
Banchereau et al., 2000

22. Differentiation of Monocytes Into Dendritic Cells

23. Maturation of Dendritic Cells With sCD40L (CD154)

24. Cytokine Production by Dendritic Cells
sCD40L
Immature DC Mature DC
254.0 pg/ml IL pg/ml IL IL IL

25. BAFF Production by Dendritic Cells

26. Ueno et al., 2007

27. Contributors Basic Science Clinical
Samia Ragheb, Ph.D. Robert Lisak, M.D.
Felicitas Gonzales, B.S. Richard Lewis, M.D.
Kirk Simon, B.S. Gregory Van Stavern, M.D.
Yanfeng Li, M.S.
Wayne State University School of Medicine
Supported by the Muscular Dystrophy Association (MDA)