Empiric broad-spectrum antimicrobials therapy is suggested to be started immediately for sepsis patients. Empiric antimicrobial therapy should be narrowed once pathogen identification and sensitivities are established. However, the detail mechanisms of de-escalation strategy are still unclear. Here we hypothesized neutrophil extracellular trap (NETs) played an essential role and de-escalation strategy might alleviate organs injury through regulation of NETs formation in sepsis. We evaluated the effect of imipenem and ceftriaxone on NETs formation in vitro and examined the role of reactive oxygen species (ROS). Next, we designed de-escalation and escalation strategy based on their effects on NETs formation in CLP model. Organ injury, inflammatory cytokines, NETs levels were compared and evaluated. The in vitro study showed that imipenem and ceftriaxone had opposite effects on NETs formation in activated neutrophils. De-escalation therapy resulted in an evaluated MPO-DNA during early stage and decreased MPO-DNA during late stage, which exerted the reverse effects in escalation therapy sepsis animal model. Inflammatory response and organ injury exacerbated when eliminated NETs with DNAseI during early stage of sepsis (p<0.01). Histopathological analysis showed decreased injury in lung, liver and intestine in de-escalation therapy compared with escalation therapy (p<0.01). De-escalation therapy results in the highest 6-day survival rate compared with the control group (p<0.01), however, no significant difference was found between de-escalation and escalation group (p=0.051). We demonstrate that de-escalation, not escalation, therapy reduces organ injury, decreases inflammatory response by promoting NETs formation in the early stage and inhibiting NETs formation in the late stage of sepsis.