Dynamic matrices with DNA-encoded viscoelasticity for cell and organoid culture.

Three-dimensional cell and organoid cultures rely on the mechanical support of viscoelastic matrices. However, commonly used matrix materials lack control over key cell-instructive properties. Here we report on fully synthetic hydrogels based on DNA libraries that self-assemble with ultrahigh-molecular-weight polymers, forming a dynamic DNA-crosslinked matrix (DyNAtrix). DyNAtrix enables computationally predictable and systematic control over its viscoelasticity, thermodynamic and kinetic parameters by changing DNA sequence information. Adjustable heat activation allows homogeneous embedding of mammalian cells. Intriguingly, stress-relaxation times can be tuned over four orders of magnitude, recapitulating mechanical characteristics of living tissues. DyNAtrix is self-healing, printable, exhibits high stability, cyto- and haemocompatibility, and controllable degradation. DyNAtrix-based cultures of human mesenchymal stromal cells, pluripotent stem cells, canine kidney cysts and human trophoblast organoids show high viability, proliferation and morphogenesis. DyNAtrix thus represents a programmable and versatile precision matrix for advanced approaches to biomechanics, biophysics and tissue engineering.

An in vitro fibrotic liver lobule model through sequential cell-seeding of HSCs and HepG2 on 3D-printed poly(glycerol sebacate) acrylate scaffolds.

Cirrhosis is a major cause of global morbidity and mortality, and significantly leads to a heightened risk of liver cancer. Despite decades of efforts in seeking for cures for cirrhosis, this disease remains irreversible. To assist in the advancement of understanding toward cirrhosis as well as therapeutic options, various disease models, each with different strengths, are developed. With the development of three-dimensional (3D) cell culture in recent years, more realistic biochemical properties are observed in 3D cell models, which have gradually taken over the responsibilities of traditional 2D cell culture, and are expected to replace some of the animal models in the near future. Here, we propose a 3D fibrotic liver model inspired by liver lobules. In the model, 3D-printed poly(glycerol sebacate) acrylate (PGSA) scaffolds facilitated the formation of 3D tissues and guided the deposition of fibrotic structures. Through the sequential seeding of hepatic stellate cells (HSCs), HepG2 and HSCs, fibrotic septum-like tissues were created on PGSA scaffolds. As albumin secretion is considered a rather important function of the liver and is found only among hepatic cells, the detection of albumin secretion up to 30 days indicates the mimicking of basic liver functions. Moreover, the in vivo fibrotic tissue shows a high similarity to fibrotic septa. Finally, via complete encapsulation of HSCs, a down-regulated albumin secretion profile was observed in the capped model, which is a metabolic indicator that is important for the prognosis for liver cirrhosis. Looking forward, the incorporation of the vasculature will further upgrade the model into a sound tool for liver research and associated treatments.

Design of photocurable, biodegradable scaffolds for liver lobule regeneration via digital light process-additive manufacturing.

The regeneration of damaged or lost tissue is considered to be a critical step toward realizing full organ regeneration in modern medicine. Although surgical techniques continue to advance, treatment for missing tissues in irregular wounds remains particularly difficult. With increasing interest in the application of additive manufacturing in tissue engineering, the fabrication of customized scaffolds for the regeneration of missing tissue via three-dimensional (3D) printing has become especially promising. Amongst the work on the regeneration of many important organs, liver regeneration is particularly interesting because liver diseases are increasingly prevalent in many countries around the world, resulting in a greater need for liver transplantation. The generation of hexagonal scaffolds for the regeneration of liver lobules is highly demanding, but their 3D structure has been proved difficult to reproduce by traditional fabrication methods. In this work, various hexagonal scaffolds are developed for liver lobule regeneration via 3D printing using novel biodegradable polymeric materials, including poly(glycerol sebacate) acrylate and poly(ethylene glycol) diacrylate. Through fine-tuning of printing parameters, a series of hexagonal scaffolds were designed and printed to mimic liver lobule units. The scaffolds were printed with various structures together with varying surface areas and 3D structures to enhance cell seeding density and diffusivity of the culture medium. Analysis of cell metabolic activities showed that the high-diffusion staircase (HDS) scaffold could support potential differences in cell proliferation rate. Furthermore, the HDS scaffolds composed of different copolymers were cultured with cells for up to 16 days to investigate the relationship between physical properties and hepatocyte proliferation. The results indicate that the combination of the high flexibility 3D printing with biodegradable, photocurable copolymers shows great promise for the regeneration of 3D liver lobules.

Laser-pattern induced contact guidance in biodegradable microfluidic channels for vasculature regeneration.

The direct cell control by surface topographic patterns in the micrometer and nanometer range has been proven to be important for the maintenance of tissue structures. This study presents the application of direct laser writing to fabricate micro-gratings on the biodegradable material 1,3-diamino-2-hydroxypropane-co-polyol sebacate (APS). The 193 nm excimer laser is applied to form microgrooves with widths of 2 to 10 µm and depths of 400 to 2884 nm. Two kinds of cells, fibroblasts of the rabbit synoviocyte cell line (HIG-82) and endothelial cells of human umbilical vein endothelial cells (HUVECs), were cultured on the flat and patterned APS to evaluate the biocompatibility of APS as well as the influence of contact guidance for cellular behaviours, respectively. The results show that both HIG-82 and HUVECs grow actively on APS scaffolds with directional growth, which was observed through cell morphology and proliferation rate, indicating their applicability in tissue regeneration. HIG-82 was observed to exhibit directional growth with the highest cell spreading area and density on the scaffolds with 7 µm width and 1350-1500 nm depth of gratings. Meanwhile, high cell spreading area and cell density of HUVECs were observed on laser ablated APS with 5 µm gratings and at depths greater than 1485 nm. The proposed microgrooves on APS could significantly enhance the cell growth, adhesion and even promote selective cell proliferation, which poses potential application for further tissue engineering studies.