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Abstract Cancer cells recruit neutrophils from the bloodstream into the tumor tissue, where these immune cells promote the progression of numerous solid tumors. Studies in mice suggest that blocking neutrophil recruitment to tumors by inhibition of neutrophil chemokine receptor CXCR2 could be a potential immunotherapy for pancreatic cancer. Yet, the mechanisms by which neutrophils promote tumor progression in humans, as well as how CXCR2 inhibition could potentially serve as a cancer therapy, remain elusive. In this study, we developed a human cell-based microphysiological system to quantify neutrophil-tumor spheroid interactions in both “separated” and “contact” scenarios. We found that neutrophils promote the invasion of tumor spheroids through the secretion of soluble factors and direct contact with cancer cells. However, they promote the proliferation of tumor spheroids solely through direct contact. Interestingly, treatment with AZD-5069, a CXCR2 inhibitor, attenuates invasion and proliferation of tumor spheroids by blocking direct contact with neutrophils. Our findings also show that CXCR2 inhibition reduces neutrophil migration toward tumor spheroids. These results shed new light on the tumor-promoting mechanisms of human neutrophils and the tumor-suppressive mechanisms of CXCR2 inhibition in pancreatic cancer and may aid in the design and optimization of novel immunotherapeutic strategies based on neutrophils. 

Neuroinflammation is characterized by the elevation of cytokines and adenosine triphosphate (ATP), which in turn activates microglia. These immunoregulatory molecules typically form gradients in vivo, which significantly influence microglial behaviors such as increasing calcium signaling, migration, phagocytosis, and cytokine secretion. Quantifying microglial calcium signaling in the context of inflammation holds the potential for developing precise therapeutic strategies for neurological diseases. However, the current calcium imaging systems are technically challenging to operate, necessitate large volumes of expensive reagents and cells, and model immunoregulatory molecules as uniform concentrations, failing to accurately replicate the in vivo microenvironment. In this study, we introduce a novel calcium monitoring micro-total analysis system (CAM-μTAS) designed to quantify calcium dynamics in microglia (BV2 cells) within defined cytokine gradients. Leveraging programmable pneumatically actuated lifting gate microvalve arrays and a Quake valve, CAM-μTAS delivers cytokine gradients to microglia, mimicking neuroinflammation. Our device automates sample handling and cell culture, enabling rapid media changes in just 1.5 s, thus streamlining the experimental workflow. By analyzing BV2 calcium transient latency to peak, we demonstrate location-dependent microglial activation patterns based on cytokine and ATP gradients, offering insights contrasting those of non-gradient-based perfusion systems. By harnessing advancements in microsystem technology to quantify calcium dynamics, we can construct simplified human models of neurological disorders, unravel the intricate mechanisms of cell-cell signaling, and conduct robust evaluations of novel therapeutics.