Rapid Prototyping of Multilayer Microphysiological Systems.

Microfluidic organs-on-chips aim to realize more biorelevant in vitro experiments compared to traditional two-dimensional (2D) static cell culture. Often such devices are fabricated via poly(dimethylsiloxane) (PDMS) soft lithography, which offers benefits (e.g., high feature resolution) along with drawbacks (e.g., prototyping time/costs). Here, we report benchtop fabrication of multilayer, PDMS-free, thermoplastic organs-on-chips via laser cut and assembly with double-sided adhesives that overcome some limitations of traditional PDMS lithography. Cut and assembled chips are economical to prototype ($2 per chip), can be fabricated in parallel within hours, and are Luer compatible. Biocompatibility was demonstrated with epithelial line Caco-2 cells and primary human small intestinal organoids. Comparable to control static Transwell cultures, Caco-2 and organoids cultured on chips formed confluent monolayers expressing tight junctions with low permeability. Caco-2 cells-on-chip differentiated ∼4 times faster, including increased mucus, compared to controls. To demonstrate the robustness of cut and assemble, we fabricated a dual membrane, trilayer chip integrating 2D and 3D compartments with accessible apical and basolateral flow chambers. As proof of concept, we cocultured a human, differentiated monolayer and intact 3D organoids within multilayered contacting compartments. The epithelium exhibited 3D tissue structure and organoids expanded close to the adjacent monolayer, retaining proliferative stem cells over 10 days. Taken together, cut and assemble offers the capability to rapidly and economically manufacture microfluidic devices, thereby presenting a compelling fabrication technique for developing organs-on-chips of various geometries to study multicellular tissues.

Bioactive Organic Rosette Nanotubes Support Sensory Neurite Outgrowth.

Regardless of the intervention for peripheral nerve repair, slow rates of axonal regeneration often result in poor clinical outcomes. Thus, using new materials such as biologically inspired, biocompatible, organic rosette nanotubes (RNTs) could provide a tailorable scaffold to modulate neurite extension and attachment for improved nerve repair. RNTs are obtained through the spontaneous self-assembly of a synthetic DNA base analogue featuring the hydrogen bond triads of both guanine and cytosine, the G∧C base. Here, we investigated the potential of RNTs functionalized with lysine and Arg-Gly-Asp-Ser-Lys (RGDSK) peptide to support neural growth. We hypothesized that (a) due to their dimensions, the RNTs would support neuron attachment, and (b) their conjugation to the integrin-binding peptide RGDSK would further enhance neurite outgrowth compared to unfunctionalized RNT. Neurite extension was examined on a variety of RNT structures, including RNT with a lysine side chain (K1), a mixture of the K1 and a free RGDS peptide, RNT alone, an RGDSK-functionalized RNT, in addition to poly-d-lysine and laminin controls. Both whole dorsal root ganglion (DRG) and single dissociated DRG neurons were seeded onto RNT-coated substrates containing various ratios of peptides. Analysis of neuron morphometrics showed that RNT blends support DRG neuron attachment and neurite extension, with RGDS presentation increasing neurite outgrowth from whole DRG by up to 47% over a 7-day period compared to K1 alone (p < 0.013). In addition, while RNTs increased the sprouting of primary neurites extending from dissociated DRG neurons, the total neurite outgrowth per neuron remained the same. These results show that functionalized biomimetic RNTs provide a support for neurite growth and extension and have the ability to modulate neuronal morphology. These results also pave the way for the design of injectable RNT-based nanomaterials that support guided neural regeneration following traumatic injury.