Study Of Platelets In Immune Regulation

Part I Platelets promote allergic Asthma through CD154Platelet is the second largest population within blood. Except its well-accepted roles in coagulation, thrombosis and wound healing, the emerging role of platelets in immune regulation is gradually appreciated by immunologists. Platelets release a variety of immune modulating factors upon activation, including both soluble factors and membrane-bound ligands, of which CD 154 attracts much interest. CD 154 is a membrane bound protein belonging to TNF superfamily.It plays important role in immune regulation, including B cell isotype switching, germinal center formation, T helper function to B cells and CTL activation. By using different animal models, it has been characterized the important role of platelet-derived CD 154 in initiation of varieties of autoimmune diseases, including rheumatoid arthritis, virus induced hepatitis, systemic lupus erythetomasus. However, the role of platelets in allergic asthma remains unclear. Previous studies show the association of platelet activation with asthma progression, yet the exact role of platelet activation in progression of allergic asthma remains elusive. In this part of our study, we found allergen can directly activate platelets and allergen activated platelets have high expression of CD154.By using platelet transfusion or depletion technique, we demonstrated that platelet promote asthma progression. Finally, by comparing the ability of wildtype and CD154 deficient platelets in asthma promotion, we conclude that platelets promote asthma progrssion through CD 154.Part II platelet protects mice from endotoxin shockRecent studies show various TLR agonists can direct activate platelets. Under sepsis shock, multiple TLR agonists accumulate in blood which might cause massive activation of platelets, indeed platelet activation has been well documented in sepsis shock induced by different pathogens. Platelet activation is the direct consequence of entotoxin accumulation in blood, meanwhile, it can also initiate downstream cascades to amplify or attenuate the responses induced by endotoxin, however, the role of platelets in such progress remains mysterious. IL-1β is a crutial cytokine in sepsis shock and there exists different mechanisms to attenuate responses initiated by IL-1β, including IL-1Ra and sIL-1R2. In this part of our study, by using platelet transfusion and depletion technique, we found platelets protect mice from endotoxin shock, yet platelet trasfusion or depletion didn’t cause significant change in IL-1β production. We then found high expression of IL-1R1 on platelets and upon activation IL-1R1 rapidly disappeared probably through cleavage of membrane-bound form to soluble form. We tested our hypothesis in a bone marrow chimera model and concluded that IL-1R1 derived from bone marrow cells protects mouse from endotoxin shock.Part III anti-tumor anto-antibodies and vwf mediate breast cancer cell conjugation with plateletsIt has been well documented that platelets promote metastasis of various tumor cells. Meanwhile large amounts of platelet-tumor cell conjugates have been found in blood during tumor progression. Platelets help tumor cell trans-blood vessel migration through P-selectin expression on platelets. Recently another study show platelets promote tumor metastasis also through release of TGF-β,TGF-β induces EMT transition of tumor cells and facilitates tumor cell migration yet the mechanism of platelet binding to tumor cell remains mysterious.In breast cancer patients, large amounts of anti-tumor cell antibodies have been found in peripheral blood and it has been tested in various models that anti-tumor cell autoantibodies in breast cancer patients promote tumor metastasis, yet the detailed mechanism underlying the functions of these large amounts of anti-tumor antibodies is poorly understood.Our hypothesis comes from stuies of platelet association with Gram-positive bacteria, i.e Listeria Monocytogene. One study shows blockage of complement C3 in blood, or GPIb on platelets totally abrogates platelet association with Listeria Monocytogene, indicating complement C3 and GPIb on platelets are key molecules mediating platelet binding to Listeria Monocytogenes. Another study focusing on Listeria Monocytogenes binding to von Willebrand factor (vwf) demonstrates that poly saccharides on cell wall of Listeria Monocytogenes direct binds to vwf and GPIb on platelets direct binds to D’D3 domain of vwf. Thus vwf bridges platelet binding to Listeria monocytogenes. Further studies of other domains indicate a potential Ig binding site within A1 domain of vwf. Based on above findings, we hypothesize that anti-tumor cell auto-antibodies can form immune complex with tumor cells and bind to A1 domain of vwf, on the other hand, GPIb on platelet binds to D’D3 domain of vwf and completes conjugation with tumor cells.To test this hypothesis, we over-express A1 or D’D3 domain of vwf in breast cancer cell line 4T1 and observe tumor metastasis in vivo. Meanwhile, by using recombinant A1 or D’D3 we demonstrated that A1 and D’D3 inhibit anti-tumor auto-antibody mediated tumor binding to platelets.