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Cardiopulmonary bypass activates classical monocytes via shear-mediated activation of Store-Operated Calcium Entry [scRNA-seq]

ABSTRACT: Patients undergoing cardiac surgery face significant inflammation induced by exposure to cardiopulmonary bypass (CPB), contributing to heightened morbidity and mortality. The molecular and cellular mechanisms that underpin this inflammatory process remain unknown. To address this knowledge gap, we conducted snRNA/ATAC sequencing to profile transcriptional and chromatin changes on circulating leukocytes from neonatal CPB patients. Classical monocytes become more prevalent after CPB, showing dysregulation of numerous inflammatory genes and significant changes in chromatin accessibility, highlighting their role in CPB-associated inflammation. The proinflammatory cytokine Interleukin 8 (IL8 or CXCL8), is significantly induced by CPB, with its promoter accessibility to the AP-1 transcription factor significantly increased. A genome-wide CRISPR screen using IL8 gene as a readout in non-adherent monocytic cells identified two genes, non-erythroid spectrin (SPTAN1) and RAF1, as novel effectors of hemodynamic stress. We further found that SPTAN1 and RAF1 activate store-operated calcium entry (SOCE) under CPB conditions, leading to elevated IL8 expression. We propose a novel shear stress-responsive pathway consisting of SPTAN1/RAF1/SOCE signaling. Drug treatment experiments offer a potential therapeutic approach to alleviate CPB-induced inflammation. Together, these findings provide novel insights into the pathogenesis of CPB-associated inflammation, enhancing our understanding of sterile inflammation and how non-adherent cells sense shear stress at molecular level.

ORGANISM(S): Homo sapiens

PROVIDER: GSE291575 | GEO | 2025/11/28

REPOSITORIES: GEO

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Project description:Cardiopulmonary bypass (CPB), the mechanical support required during the majority of cardiac surgeries, causes significant systemic inflammation and multi-organ dysfunction that is especially severe in neonatal patients. Currently, the limited understanding of the molecular mechanisms that underlie CPB-associated inflammation presents a significant barrier to the development of therapeutic interventions. To address this knowledge gap, we performed mRNA sequencing of total circulating leukocytes from neonatal patients undergoing CPB. The transcriptional signatures across 7 time points ranging from pre-CPB, during CPB, to 24 hours post-CPB were found to be dominated by myeloid cells, particularly monocytes. INTERLEUKIN-8 (IL8) and TUMOR NECROSIS FACTOR-α (TNFα) were two inflammatory cytokines specifically and robustly upregulated in the leukocytes from both CPB patients and piglets exposed to CPB. To delineate the mechanism how CPB activates cytokine production, we exposed THP-1 cells, a human monocyte cell line, in vitro to CPB-like conditions including artificial surfaces, high shear stress and cooling/rewarming. The in vitro CPB data recapitulated the in vivo CPB condition in that IL8 and TNFα were strongly upregulated. Shear stress was the major driver of cytokine expression as compared to the temperature or plastic exposure. Extracellular calcium influx was found essential for the shear-stress mediated upregulation of cytokines via the MEK/ERK/AP-1 and Calcineurin/NFAT pathways. After being exposed to CPB conditions, a subpopulation of THP-1 cells died by apoptosis and TNFα-mediated necroptosis, the latter form of which potentially contribute to the post-CPB tissue injuries. Together, our study is the first to elucidate the shear-stress modulated molecular mechanisms leading to systemic inflammation in pediatric CPB and the link between inflammation and organ damage. Calcium signaling pathways and TNFα signaling are novel targets for therapeutic development to improve clinical outcomes. To our knowledge, these are also the first data to implicate necroptosis in CPB and show that shear stress can activate necroptosis.

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Cardiopulmonary bypass activates classical monocytes via shear-mediated activation of Store-Operated Calcium Entry [scRNA-seq]

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