Yuan Cui: Single-cell oxygen metabolism: a universal, effect-based marker of chemical toxicity

Abstract

Understanding how organisms respond to oxygen (O2) fluctuations in their aquatic microenvironments and studying O2 consumption as an indicator of metabolism is crucial for advancing ecology, toxicology, tissue engineering and environmental sciences. However, despite their importance across scientific disciplines, traditional O2 measurements often lack the sensitivity to capture rapid or localized changes occurring at the microscale. This PhD thesis addresses this challenge by developing novel microfluidic techniques and systems to sense O2.

To investigate advective and diffusive O2 fluxes at the microscale, this thesis developed chemical sensing particle image velocimetry (sensPIV), a technique that combines optode microparticles and rapid microscope imaging. Specifically, I contributed by testing the performance of sensPIV particles within O2-permeable polydimethylsiloxane (PDMS) devices. This demonstrated the effectiveness of sensPIV in visualizing microscale O2 fluxes, with potential applications around complex biological structures such as coral segments.

To characterize single-cell O2 respiration rates this thesis developed a micro-respiration chamber device. This device consists of gastight microwells that isolate single-cells, allowing for measuring their O2 consumption via immobilized O2-sensitive optodes. When applied to human hepatic cells, this device revealed size-related respiration kinetics in single-cells and adaptively changing respiration rates due to O2 limitations. The micro-respiration device was then refined into SlipO2Chip, a microfluidic platform that quantifies single-cell O2 respiration rates before and after chemical exposures. This was achieved by adding a mechanism for opening and closing microwells via slipping a dedicated channel that introduces chemical solutions. SlipO2Chip demonstrated a dose-dependent decrease in diatom respiration when exposed to the bacterial infochemical 2-Heptyl-4-Quinolone (HHQ), thus enhancing our understanding of toxicological impacts by detailing cell-to-cell heterogeneity. In a final effort, the micro-respiration chamber device was combined with quantitative phase imaging (QPI) to jointly measure respiration rates and dry mass in individual cells from three unicellular diatom species of varying sizes (16 - 300 µm). Preliminary data were integrated into metabolic scaling theory that describes how metabolic rates scale with geometrical size/mass across all species. The results indicated an interspecific scaling exponent similar to published bulk measures, and three intraspecific exponents showing potential morphological and physiological relationships to metabolism. This proof-of-concept study highlighted that combined measurement of metabolism and mass can enhance the resolution of scaling theory by adding crucial information on cell-to-cell variability.

Overall, this PhD thesis contributed to ecotoxicology, ecology and bioengineering by providing detailed insights into spatiotemporal O2 dynamics and single-cell O2 metabolism in the presence and absence of chemical perturbations.