Analyzing the in vivo contribution of G protein-coupled receptors and LFA-1 for naive B cell trafficking in lymph nodes

B cell motility within lymphoid tissue is precisely orchestrated by G-protein-coupled receptors (GPCRs) of the chemokine receptor family and other GPCRs, including the recently described 7,25-dihydroxycholesterol (7,25-OHC) receptor Ebi2. Here, we analyzed the contribution of the known B cell-expressed GPCRs CXCR4, CXCR5, CCR7 and Ebi2, and the LFA-1 integrin during the dynamic migration of naive B cells inside lymphoid tissue and their accumulation from the T cell area in B cell follicles, using intravital twophoton microscopy (2PM). We observed a non-redundant role for CXCR5 during dynamic B cell motility in the T cell area and B cell follicles, while the absence of CXCR4 and CCR7 signaling did not influence B cell migration parameters. Although CXCR5 was central for efficient B cell accumulation in B cell follicles, we did not find evidence for directed migration of T cell area-borne B cells towards B cell follicles, supporting a chemokinetic, and not chemotactic, role for the CXCR5 ligand CXCL13 during this process. Lack of Ebi2 confined B cells to the central part of B cell follicles, resulting in higher average migration velocities as compared to wild type B cells. Additional blocking of LFA-1 further reduced B cell migration velocities inside lymphoid tissue without affecting guidance by the lymphoid stromal network. Finally, we found that despite much lower expression levels of homeostatic chemokines and disrupted lymphoid structure in virus-infected PLNs, CXCR5 remained essential for B cell cluster formation. In summary, our data provide novel insight into the contribution of GPCRs and LFA-1 during the dynamic migration of naive B cells in non-inflamed and inflamed lymphoid tissue and shed light on the bioavailability of chemokines and chemokine gradients.