Filamin-A regulates neutrophil uropod retraction through RhoA during chemotaxis.

Filamin-A (FLNa) has been shown to be a key cross-linker of actin filaments in the leading edge of a motile melanoma cell line, however its role in neutrophils undergoing chemotaxis is unknown. Using a murine transgenic model in which FLNa is selectively deleted in granulocytes, we report that, while neutrophils lacking FLNa show normal polarization and pseudopod extension, they exhibit obvious defects in uropod retraction. This uropod retraction defect was found to be a direct result of reduced FLNa mediated activation of the small GTPase RhoA and myosin mediated actin contraction in the FLNa null cells. This results in a neutrophil recruitment defect in FLNa null mice. The compensatory increase in FLNb levels that was observed in the FLNa null neutrophils may be sufficient to compensate for the lack of FLNa at the leading edge allowing for normal polarization, however this compensation is unable to regulate RhoA activated tail retraction at the rear of the cell.

A non-invasive oral rinse assay predicts bone marrow engraftment and 6 months prognosis following allogeneic hematopoietic stem cell transplantation.

BACKGROUND: We have previously shown in a pediatric Hematopoietic stem cell transplant (HSCT) population that a non-invasive oral rinse can be used to monitor engraftment, neutrophil tissue delivery and susceptibility to infection post-HSCT. METHODS: Using the same oral rinse protocol, we studied neutrophil tissue delivery kinetics and its relationship to clinical parameters and outcomes following HSCT in 29 adult patients. Oral neutrophil counts were compared to circulating neutrophil levels, oral mucositis scores and patient health status at 6 months post-HSCT. RESULTS: Neutrophils were detected on average 8.4 ± 3.4 SD days earlier in the oral tissues than in the blood circulation, enabling us to confirm successful engraftment more than one week earlier than when using blood neutrophil counts alone. As well, in this population the time-span between oral engraftment (OE) and blood engraftment (BE) was a consistent predictor of treatment outcome at 6 months following HSCT where a BE-OE of <6 days resulted in 100% of patients having a negative outcome. CONCLUSION: We conclude that monitoring the timing of neutrophil delivery to the oral tissues with a non-invasive oral rinse has the potential to allow the physician to identify those patients who are at a high risk of HSCT failure within just a few weeks of the initiation of treatment.

Rac1 links leading edge and uropod events through Rho and myosin activation during chemotaxis.

Chemotactic responsiveness is crucial to neutrophil recruitment to sites of infection. During chemotaxis, highly divergent cytoskeletal programs are executed at the leading and trailing edge of motile neutrophils. The Rho family of small GTPases plays a critical role in cell migration, and recent work has focused on elucidating the specific roles played by Rac1, Rac2, Cdc42, and Rho during cellular chemotaxis. Rac GTPases regulate actin polymerization and extension of the leading edge, whereas Rho GTPases control myosin-based contraction of the trailing edge. Rac and Rho signaling are thought to crosstalk with one another, and previous research has focused on mutual inhibition of Rac and Rho signaling during chemotaxis. Indeed, polarization of neutrophils has been proposed to involve the activity of a negative feedback system where Rac activation at the front of the cell inhibits local Rho activation, and vice versa. Using primary human neutrophils and neutrophils derived from a Rac1/Rac2-null transgenic mouse model, we demonstrate here that Rac1 (and not Rac2) is essential for Rho and myosin activation at the trailing edge to regulate uropod function. We conclude that Rac plays both positive and negative roles in the organization of the Rhomyosin "backness" program, thereby promoting stable polarity in chemotaxing neutrophils.