The GSDMD Knockout HL-60 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout population engineered to disrupt the GSDMD gene, which encodes gasdermin D, the central pore-forming executioner of pyroptosis. This heterogeneous pool, derived from the HL-60 promyelocytic leukemia line, provides a loss-of-function model for investigating inflammasome-driven cell death and inflammatory cytokine release.
The HL-60 cell line was originally isolated from the peripheral blood of a 36-year-old female with acute promyelocytic leukemia and represents an undifferentiated myeloblastic model capable of granulocytic or monocytic differentiation upon stimulation with DMSO or phorbol esters. These cells express key inflammasome components and undergo robust pyroptosis, making them a pertinent host for GSDMD disruption.
GSDMD acts downstream of inflammatory caspases activated by the NLRP3, AIM2, and NLRC4 inflammasomes. Cleavage by caspase-1, -4, -5, or -11 releases the N-terminal pore-forming domain, which oligomerizes and inserts into the plasma membrane, causing osmotic lysis and the release of IL-1??, IL-18, and HMGB1. This process is regulated by upstream adaptors ASC and cardiolipin, and requires cooperation with NINJ1 for membrane rupture. Furthermore, GSDMD-mediated K? efflux potentiates secondary NLRP3 inflammasome activation.
Knocking out GSDMD in HL-60 cells uncouples inflammasome assembly from pyroptotic execution, preserving upstream caspase-1 activation and cytokine processing while blocking lytic death. This model is especially useful when differentiated into macrophage-like cells, which exhibit enhanced inflammasome responses. It thus allows dissection of GSDMD-dependent and -independent signaling outcomes in both undifferentiated and differentiated myeloid states.
The cells are suited for LDH release and PI uptake assays to quantify pyroptosis, western blotting to monitor GSDMD cleavage, and ELISA to measure IL-1?? and IL-18 secretion. They enable confocal imaging of pore formation, co-immunoprecipitation of caspase-1 complexes, and drug screening for pyroptosis modulators. Transcriptomic and proteomic analyses can identify novel regulators, while stimulations with nigericin or ATP activate canonical NLRP3 pathways. For further technical details, please contact Ascent Research.
