Role of Integrins Involved in Mn2+-Dependent Adhesion to Fibronectin Peptide of Mastocytoma P-815 Cells and Peritoneal Mast Cells

Mn²⁺-dependent integrin-mediated adhesion to the extracellular matrix is extensively studied, however, its implication in mast cell biology remains unexplored. This study aims to investigate the role of Mn²⁺ in promoting adhesion in mouse mastocytoma P-815 cells (P-815) and peritoneal mast cells (PMC) to the Arg-Gly-Asp (RGD)-enriched fibronectin peptide (RGD matrix) within the culture medium. Our findings indicate that Mn²⁺ induces cell adhesion, with optimal results achieved when P-815 were exposed to 2 mM Mn²⁺ for 30 min at 37ºC, resulting in approximately 40% cell adhesion to the RGD matrix. The Mn²⁺-dependent P-815 adhesion was inhibited by anti-integrin α4, β1, and β3 subunit function-blocking antibodies, and by the integrin αIIbβ3 antagonist tirofiban, indicating the involvement of integrins α4β1 and αIIbβ3. Similarly, Mn²⁺-dependent PMC adhesion to the RGD matrix was inhibited by anti-integrin α4, α5, β1, β3, and β7 subunit function-blocking antibodies and tirofiban, demonstrating the involvement of integrins α4β1, α4β7, α5β1, and αIIbβ3. Integrins α4β1 and αIIbβ3 were consistently involved in Mn²⁺-induced adhesion reactions in both P-815 and PMC, while integrins α4β7 and α5β1 were specifically implicated in the response to PMC only. The addition of the actin inhibitor cytochalasin D, glycosylphosphatidylinositol-anchored protein (GPI-AP) cleaving enzyme phosphatidylinositol-specific phospholipase C, and the PKA inhibitor H-89 significantly reduced Mn²⁺-dependent P-815 adhesion to the RGD matrix. However, adding the myosin II inhibitor brebbistatin and the RhoA inhibitor Y27632 did not produce the same effect. Furthermore, cellular cholesterol removal with 6-O-α-maltosyl-β cyclodextrin significantly diminished Mn²⁺-dependent P-815 adhesion, concomitant with a decrease in the expression of integrin α4 and β1 subunits on the cell surface.

In summary, Mn²⁺ fosters adhesion to the RGD matrix through integrins α4β1 and αIIbβ3, which are common between P-815 and PMC, while integrins α4β7 and α5β1 are specifically involved in PMC adhesion. The Mn²⁺-induced adhesion reaction in P-815 closely correlates with signal expression, including cAMP/PKA, GPI-AP, cellular cholesterol, and actin cytoskeleton, demonstrating a correlation between Mn²⁺-induced P-815 adhesion and signaling pathways within lipid rafts. These results may clarify questions regarding adhesion and detachment of mast cells to the extracellular matrix involved in metal ion-induced immunity and inflammation suppression.