Dynamic changes of natural killer cell immunophenotypes and receptors according to the mortality in the intra-abdominal murine sepsis model

Sepsis is a life-threatening infectious syndrome that lacks targeted pharmacological therapies and poses major challenges in reducing mortality and long-term complications such as disability and frailty. Early and intensive intervention is critical to improving prognosis and preventing multiorgan dysfunction. However, alternative treatment strategies are urgently needed for patients who do not respond to guideline-based resuscitation, such as those outlined in the Surviving Sepsis Campaign. Natural killer (NK) cells are key effectors of the innate immune system, and their balanced activity may be crucial in preventing the progression of sepsis. Given conflicting evidence on whether NK cell activity (NKA) is protective or harmful, we investigated NKA in a murine model of intra-abdominal sepsis, assessing activating and inhibitory NK receptors (NKRs), as well as NK cell subsets in whole blood, bone marrow, lymph nodes, spleen, and liver.

C57BL/6 mice underwent cecal ligation and puncture (CLP) to induce mid-grade (MGS, 30% 7-day survival) or high-grade sepsis (HGS, 0% 7-day survival), with sham-operated mice as controls. Blood and immune-related organs were collected on days 1, 3, and 7 post-surgery (MGS: days 1, 3, 7; HGS: days 1, 3; Sham: day 7). Flow cytometry was used to analyze CD11b and CD27 expression to define maturation-associated cytolytic and cytokine-producing NK cell phenotypes. CD3⁻NK1.1⁺ NK cells were purified by FACS for RT-PCR of activating (Ly49D, Ly49H) and inhibitory (Ly49C, Ly49G2) NKRs, and ELISA was performed for granzyme B and IFN-γ.

Our experiments consistently showed that in MGS, NKA-initially suppressed-was significantly restored by day 7 after CLP. This recovery was characterized by increased expression of activating NKRs, decreased inhibitory NKRs, expansion of terminally differentiated cytotoxic NK subsets (CD11b⁺/CD27^-), higher total NK cell counts, and elevated granzyme B levels. In contrast, HGS, associated with high lethality, was marked by persistent suppression of NKA.

The sustained impairment of NK cell phenotype is associated with lethal outcomes in sepsis.