Multi-Functional Microsystems for Cellular Engineering and Molecular Diagnostics


Student thesis: Doctoral Thesis

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Award date5 May 2022


The integrative multifunctional microsystems are widely used in scientific research of genomics and proteomics, clinical disease diagnosis and therapeutic, new drug development, judicial appraisal, food safety, etc. Nowadays, the convergence of new fabrication technologies expands our arsenal for making more effective microsystems to understand molecular diagnostics and cellular behaviors. However, there are still many demands related to cancer screening and novel cancer therapies to be satisfied in the molecular diagnostics and cellular engineering areas. In the molecular diagnostics area, the diagnosis of tumor recurrence among patients is a severe problem during the cancer treatment, making the lagging therapy. However, limited therapeutic microsystems can provide personality healthcare for tumor patients. Microsystems not only play an essential role in cancer screening and novel cancer therapies in the molecular diagnostics area but also can help build disease models for exploring pathogenetic mechanisms and drug screening in cellular engineering.

In the cellular engineering area, our understanding of the cellular mechanism is still in the early stage due to the absence of microsystems that can mimic the developmental context of organogenesis in vitro. At the same time, the situation is complex in vivo.

In this dissertation, we proposed three kinds of microsystems that focus on building three meaningful models in molecular diagnostics and cellular engineering to address these bottlenecks and promote the development of new functional microsystems in other situations. In chapter 2, a microsystem called “iMethy” was developed to address continuously tumor monitoring in the interstitial fluid (ISF). The “iMethy,” a self-healable electronic patch for active monitoring methylated circular tumor DNA (ctDNA), is designed with two core parts, an eGaIn immunosensor for real-time in vitro analysis and a hydrogel bridge for continuously macromolecular extraction in ISF. A self-healing eutectic Gallium-Indium(eGaIn) circuit was constructed with a specifically designed multilayer structure (structural layer, affinity layer, and constrained eGaIn), which ensures the robust connection even after severe damage. Our iMethy can be sensitively responsive to the weak methylation state change, with a 10-16M detection limit, and shows good work performed in both in vitro and in vivo tests.

Secondly, in the cellular engineering area, to develop a microsystem that can rebuild the complex chemical gradient in vitro to explore the chemical cues in cellular behavior, a high-throughput chemotaxis analysis platform (HT-ChemoChip) was developed in chapter 3. The microsystem that combines Matrigel cylinder arrays allows high-throughput generation of large-scale molecular gradients with significant steepness in vitro. Hippocampal neurons were grown in the platform, which can form thousands of different gradients of a three-dimensional microcolumn of neurons and explore how neurons migrated and projected. We systematically studied neurons’ steepness sensitivity with two classic guide molecules (Sema3A, and Netrin-1) and revealed the incredible diversity and complexity in the associated chemotactic regulation of neuronal development. These results provide new insights into the role of gradient steepness in neuronal chemotaxis.

Thirdly, in the cellular engineering area, to establish a microsystem that can rebuild the cellular features of spatial constraints that shape tissue growth, an active thermosensitive-hydrogel-based culture system (NeurGel platform) was proposed in chapter 4. This microsystem can mimic the shape of the neural groove and the change from the board shape to groove shape in vitro. The embryo stem cells were seeded onto the system in the environment of Matrigel. We found different protein expressions in different curvature areas across the groove. And then, mechanical factors related to marker (F-actin) are squeezed at the bottom of the groove shape. The RNA sequencing data indicated that the groove-shape change activated some signal pathways regulating the neural tube closure. Our system will enable the attainment of more shaping change processes during embryo development in vitro.

In summary, the presented studies explore three different functional microsystems, including an “iMethy,” a “HT-ChemoChip,” and a “NeurGel platform” to improve the functionality and efficiency of the microsystems, which would essentially optimize disease diagnosis/treatment processes and deepen our understanding of cellular behavior related to disease occurrence. The quantitative results established in these studies may contribute to developing novel investigative and therapeutic microsystems in molecular diagnostics and cellular engineering.