Cells comprising biological tissues are surrounded by a structure known as the stroma, and their behavioradapts in response to stimuli generated by flow and transport phenomena. Despite its importance, ourunderstanding of how cells respond to their surrounding microenvironment remains limited, hindering thedevelopment of effective disease prevention and treatment strategies. A significant challenge has been thedifficulty in observing cellular behavior while simultaneously controlling the local culture environment.Although microfluidic devices have become increasingly prevalent in recent years, they have not fullyaddressed the need for comprehensive environmental control. To overcome this limitation, we developed the"stromal function chip," which focuses on three critical environmental factors within the stroma: oxygenconcentration, pH, and interstitial flow. This innovative platform enables precise and rapid manipulation ofthese parameters while facilitating real-time observation of both individual cellular responses and complexcell-cell interactions.

Traditionally, stage incubators mounted on microscopes have been employed to maintain culture conditionsduring time-lapse observations of cellular behavior. However, these conventional systems present significantlimitations in actively and rapidly controlling localized changes within the culture microenvironment. Whilerecent advances in microfluidic devices and organ-on-a-chip technologies have enhanced our ability toobserve cellular responses under controlled conditions, these approaches still exhibit considerable constraintsin achieving comprehensive environmental regulation. In contrast, our newly developed chip providesprecise, dynamic, and immediate control over the culture microenvironment during cellular experiments,enabling high-fidelity visualization and quantification of complex cellular dynamics in response to environmental stimuli.

The stromal function chip features sophisticated architecture comprising cell culture channels with multiplegas channels strategically positioned in vertical alignment above them. Through the controlled delivery ofprecisely mixed gases containing specific oxygen and carbon dioxide concentrations to these gas channels,the chip facilitates gas exchange that enables exquisite regulation of both oxygen concentration and pHwithin the cell culture microenvironment. This approach represents a significant advancement overconventional chemical reaction-based methods, as it eliminates potential cellular toxicity while providinghighly flexible and dynamic control over oxygen concentration and pH. Furthermore, the chip's innovativedesign allows for the precise modulation of interstitial flow—achieved by embedding hydrogel within theculture channels and establishing controlled hydrostatic pressure gradients between inlet and outlet ports. Bysimultaneously manipulating these three critical environmental factors—oxygen concentration, pH, andinterstitial flow—researchers can systematically investigate cellular response mechanisms and characterizehow cells adapt to specific stromal microenvironmental conditions, thereby advancing our understanding oftissue physiology and pathophysiology.

By precisely recapitulating the hypoxic and acidic microenvironmental conditions that characterize tumorniches and inflammatory sites, this innovative chip serves as a powerful platform for pre-clinical evaluationof therapeutic efficacy, enabling researchers to determine optimal drug candidates and dosage regimens priorto in vivo studies. Moreover, the system serves as a platform/tool for fundamental medical and biologicalinvestigations, allowing for high-resolution cellular observation and analysis under rigorously controlled andphysiologically relevant culture conditions.

• Microfluidic device
• Cell culture
• Cell experiment
• Oxygen concentration
• pH
• Interstitial flow
• Hypoxia
• In vivo microenvironment
• Organ-on-a-chip
• Biofunctional chip
• Cellular dynamics
• rug discovery