Human CD62Ldim neutrophils identified as a separate subset by proteome profiling and in vivo pulse-chase labeling

During acute inflammation, 3 neutrophil subsets are found in the blood: neutrophils with a conventional segmented nucleus, neutrophils with a banded nucleus, and T-cell-suppressing CD62L(dim) neutrophils with a high number of nuclear lobes. In this study, we compared the in vivo kinetics and proteomes of banded, mature, and hypersegmented neutrophils to determine whether these cell types represent truly different neutrophil subsets or reflect changes induced by lipopolysaccharide (LPS) activation. Using in vivo pulse-chase labeling of neutrophil DNA with 6,6-H-2(2)-glucose, we found that H-2-labeled banded neutrophils appeared much earlier in blood than labeled CD62L(dim) and segmented neutrophils, which shared similar label kinetics. Comparison of the proteomes by cluster analysis revealed that CD62L(dim) neutrophils were clearly separate from conventional segmented neutrophils despite having similar kinetics in peripheral blood. Interestingly, the conventional segmented cells were more related at a proteome level to banded cells despite a 2-day difference in maturation time. The differences between CD62L(dim) and mature neutrophils are unlikely to have been a direct result of LPS-induced activation, because of the extremely low transcriptional capacity of CD62L(dim) neutrophils and the fact that neutrophils do not directly respond to the low dose of LPS used in the study (2 ng/kg body weight). Therefore, we propose CD62L(dim) neutrophils are a truly separate neutrophil subset that is recruited to the bloodstream in response to acute inflammation. This trial was registered at www.clinicaltrials.gov as #NCT01766414.