Toward Single Cell Tattoos: Biotransfer Printing of Lithographic Gold Nanopatterns on Live Cells.

Lithographic nanopatterning techniques such as photolithography, electron-beam lithography, and nanoimprint lithography (NIL) have revolutionized modern-day electronics and optics. Yet, their application for creating nanobio interfaces is limited by the cytotoxic and two-dimensional nature of conventional fabrication methods. Here, we present a biocompatible and cost-effective transfer process that leverages (a) NIL to define sub-300 nm gold (Au) nanopattern arrays, (b) amine functionalization of Au to transfer the NIL-arrays from a rigid substrate to a soft transfer layer, (c) alginate hydrogel as a flexible, degradable transfer layer, and (d) gelatin conjugation of the Au NIL-arrays to achieve conformal contact with live cells. We demonstrate biotransfer printing of the Au NIL-arrays on rat brains and live cells with high pattern fidelity and cell viability and observed differences in cell migration on the Au NIL-dot and NIL-wire printed hydrogels. We anticipate that this nanolithography-compatible biotransfer printing method could advance bionics, biosensing, and biohybrid tissue interfaces.

Toward single cell tattoos: Biotransfer printing of lithographic gold nanopatterns on live cells.

Lithographic nanopatterning techniques like photolithography, electron-beam lithography, and nanoimprint lithography (NIL) have revolutionized modern-day electronics and optics. Yet, their application for creating nano-bio interfaces is limited by the cytotoxic and two-dimensional nature of conventional fabrication methods. Here, we present a biocompatible and cost-effective transfer process that leverages (a) NIL to define sub-300 nm gold (Au) nanopattern arrays, (b) amine functionalization of Au to transfer the NIL-arrays from a rigid substrate to a soft transfer layer, (c) alginate hydrogel as a flexible, degradable transfer layer, and (d) gelatin conjugation of the Au NIL-arrays to achieve conformal contact with live cells. We demonstrate biotransfer printing of the Au NIL-arrays on rat brains and live cells with high pattern fidelity and cell viability and observed differences in cell migration on the Au NIL-dot and NIL-wire printed hydrogels. We anticipate that this nanolithography-compatible biotransfer printing method could advance bionics, biosensing, and biohybrid tissue interfaces.

Autonomous Untethered Microinjectors for Gastrointestinal Delivery of Insulin.

The delivery of macromolecular drugs via the gastrointestinal (GI) tract is challenging as these drugs display low stability as well as poor absorption across the intestinal epithelium. While permeation-enhancing drug delivery methods can increase the bioavailability of low molecular weight drugs, the effective delivery of high molecular weight drugs across the tight epithelial cell junctions remains a formidable challenge. Here, we describe autonomous microinjectors that are deployed in the GI tract, then efficiently penetrate the GI mucosa to deliver a macromolecular drug, insulin, to the systemic circulation. We performed in vitro studies to characterize insulin release and assess the penetration capability of microinjectors and we measured the in vivo release of insulin in live rats. We found that the microinjectors administered within the luminal GI tract could deliver insulin transmucosally to the systemic circulation at levels similar to those with intravenously administered insulin. Due to their small size, tunability in sizing and dosing, wafer-scale fabrication, and parallel, autonomous operation, we anticipate that these microinjectors will significantly advance drug delivery across the GI tract mucosa to the systemic circulation in a safe manner.

Directing Multicellular Organization by Varying the Aspect Ratio of Soft Hydrogel Microwells.

Multicellular organization with precise spatial definition is essential to various biological processes, including morphogenesis, development, and healing in vascular and other tissues. Gradients and patterns of chemoattractants are well-described guides of multicellular organization, but the influences of 3D geometry of soft hydrogels are less well defined. Here, the discovery of a new mode of endothelial cell self-organization guided by combinatorial effects of stiffness and geometry, independent of protein or chemical patterning, is described. Endothelial cells in 2 kPa microwells are found to be ≈30 times more likely to migrate to the edge to organize in ring-like patterns than in stiff 35 kPa microwells. This organization is independent of curvature and significantly more pronounced in 2 kPa microwells with aspect ratio (perimeter/depth) < 25. Physical factors of cells and substrates that drive this behavior are systematically investigated and a mathematical model that explains the organization by balancing the dynamic interaction between tangential cytoskeletal tension, cell-cell, and cell-substrate adhesion is presented. These findings demonstrate the importance of combinatorial effects of geometry and stiffness in complex cellular organization that can be leveraged to facilitate the engineering of bionics and integrated model organoid systems with customized nutrient vascular networks.

Hierarchically Curved Gelatin for 3D Biomimetic Cell Culture.

The stiffness, microcurvature, and meso-curvature of cellular microenvironments can significantly alter cell and tissue function. However, it is challenging to produce in vitro tissue models that feature tunability in shape, stiffness, and curvature simultaneously in a high-throughput and cost-effective manner. One of the significant challenges is the fragility of micropatterns in soft and biocompatible hydrogels. Here, we describe an approach that combines reflow photolithography, soft lithography, and strain engineering to create soft anatomically mimetic gelatin cell culture models. The models can be mechanically tuned to have stiffnesses as low as 400 Pa to as high as 50 kPa featuring hierarchical curvature at two length scales: the cellular length scale of 12 to 120 µm, and the mesoscale of 1-4 mm. We characterize the microstructured gels using optical microscopy and rheometry, highlighting tunability in the hierarchical curvature, modulus, and shape. Also, collagen-based gelatin offers high-level biocompatibility and bypasses the need for additional surface modification to enhance cell adhesion. We anticipate that this approach could advance anatomically accurate in vitro 3D cell culture models of relevance to biofabrication, cell biology, and drug screening.