טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentBen-Shaul Shahar
SubjectLaquinimod Modulates Responsiveness, Phenotype and IL10
Production in Human B Cells from Healthy Donors
and MS Patients
DepartmentDepartment of Medicine
Supervisors Professor Elias Toubi
Professor Ariel Miller
Professor Doron Melamed
Full Thesis textFull thesis text - English Version


Abstract

The therapeutic benefit of B cell-depleting therapy in mice and humans has emphasized a role of B cells in autoimmune diseases, including the immune-mediated neurodegenerative disease, Multiple Sclerosis (MS). Moreover, the formation of structures resembling germinal centers in the MS brain implies a possible local B cell reaction to Central Nervous System (CNS)-derived antigens, reinforcing the evidence of B cells involvement in MS pathophysiology. Laquinimod, an orally administrated drug, is being evaluated as treatment of Relapsing- Remitting MS (RRMS) patients. Our research goal was to assess the immunomodulatory effects of Laquinimod invitro on human B cells from healthy and RRMS subjects. In order to examine whether Laquinimod affects B cell fate; we assessed the expression of two proteins, the antiapoptotic Bcl2 and the pro-apoptotic Bim, both regulated through the MAPK and NFκB pathways. Our results showed that 1μM Laquinimod upregulated the expression of Bcl2 in BCR-stimulated and unstimulated B cells, while Bim expression was downregulated in unstimulated B cells. We also examined the affect of Laquinimod on the NFκB pathway, linked to B cells survival, differentiation and proliferation. We demonstrated an increase in NFκB activity with 1μM Laquinimod treatment. Additionally, we investigated Laquinimod's effect on primary B cells phenotype from healthy and RRMS subjects. Purified B cells were cultured in the presence or absence of 1mM Laquinimod and with or without 1mM ODN stimulation, then stained with specific antibodies for extracellular and intracellular markers and analyzed by FACS. In our study, Laquinimod treatment decreased the CD5 B cell population in ODN-stimulated B cells in both healthy and MS subjects. CD86, IL10 and CD25 have all been associated with B cell regulatory characteristics. In B cells from healthy donors, Laquinimod treatment increased the percentages of CD86 B cells in stimulated cells, and of CD25 cells in unstimulated cells, and further increased the expression of IL10 in CD25 cells. In B cells from MS patients, Laquinimod treatment increased the expression of CD86 and IL10 in CD25 B cells. In summary, we showed that Laquinimod modulates B cell signaling and phenotype, thereby possibly effecting B cell responsiveness and function. In view of the recent evidences of regulatory/suppressive properties of IL10CD86 B cells, the Laquinimod-mediated ascent of IL10CD86 B cell sub-population, may contribute to its beneficial therapeutic properties in MS and possibly in other autoimmune diseases.