Justin Liu

Deterministically patterned biomimetic human iPSC-derived hepatic model via rapid 3D bioprinting.

Significance The great challenge to developing an in vitro liver model lies in the limitation of current approaches to recapitulate the sophisticated liver microenvironment contributed by the complex microarchitecture and diverse cell combination. We demonstrate an innovative advancement toward simulating natural complexity by integrating a rapid 3D bioprinting technology with tissue engineering to develop a microscale hepatic construct consisting of physiologically relevant hexagonal units of liver cells and supporting cells. The entire construct is fabricated within several seconds on minimal UV illumination. The model enables the structural and functional improvements of human induced pluripotent stem cell-derived hepatic progenitor cells and therefore can be used in early personalized drug screening and liver pathophysiology studies in vitro. The functional maturation and preservation of hepatic cells derived from human induced pluripotent stem cells (hiPSCs) are essential to personalized in vitro drug screening and disease study. Major liver functions are tightly linked to the 3D assembly of hepatocytes, with the supporting cell types from both endodermal and mesodermal origins in a hexagonal lobule unit. Although there are many reports on functional 2D cell differentiation, few studies have demonstrated the in vitro maturation of hiPSC-derived hepatic progenitor cells (hiPSC-HPCs) in a 3D environment that depicts the physiologically relevant cell combination and microarchitecture. The application of rapid, digital 3D bioprinting to tissue engineering has allowed 3D patterning of multiple cell types in a predefined biomimetic manner. Here we present a 3D hydrogel-based triculture model that embeds hiPSC-HPCs with human umbilical vein endothelial cells and adipose-derived stem cells in a microscale hexagonal architecture. In comparison with 2D monolayer culture and a 3D HPC-only model, our 3D triculture model shows both phenotypic and functional enhancements in the hiPSC-HPCs over weeks of in vitro culture. Specifically, we find improved morphological organization, higher liver-specific gene expression levels, increased metabolic product secretion, and enhanced cytochrome P450 induction. The application of bioprinting technology in tissue engineering enables the development of a 3D biomimetic liver model that recapitulates the native liver module architecture and could be used for various applications such as early drug screening and disease modeling.

Three-dimensional direct cell patterning in collagen hydrogels with near-infrared femtosecond laser

We report a methodology for three-dimensional (3D) cell patterning in a hydrogel in situ. Gold nanorods within a cell-encapsulating collagen hydrogel absorb a focused near-infrared femtosecond laser beam, locally denaturing the collagen and forming channels, into which cells migrate, proliferate, and align in 3D. Importantly, pattern resolution is tunable based on writing speed and laser power, and high cell viability (>90%) is achieved using higher writing speeds and lower laser intensities. Overall, this patterning technique presents a flexible direct-write method that is applicable in tissue engineering systems where 3D alignment is critical (such as vascular, neural, cardiac, and muscle tissue).

Differential role of all-trans retinoic acid in promoting the development of CD4 and CD8 regulatory T cells

It is known that ATRA promotes the development of TGF‐β‐induced CD4+Foxp3+ iTregs, which play a vital role in the prevention of autoimmune diseases; however, the role of ATRA in facilitating the differentiation and function of CD8+Foxp3+ iTregs remains elusive. Using a head‐to‐head comparison, we found that ATRA promoted expression of Foxp3 and development of CD4+ iTregs, but it did not promote Foxp3 expression on CD8+ cells. Using a standard in vitro assay, we demonstrated that CD8+ iTregs induced by TGF‐β and ATRA were not superior to CD8+ iTregs induced by TGF‐β alone. In cGVHD, in a typical lupus syndrome model where DBA2 spleen cells were transferred to DBA2xC57BL/6 F1 mice, we observed that both CD8+ iTregs induced by TGF‐β and ATRA and those induced by TGF‐β alone had similar therapeutic effects. ATRA did not boost but, conversely, impaired the differentiation and function of human CD8+ iTregs. CD8+ cells expressed the ATRA receptor RAR and responded to ATRA, similar to CD4+ cells. We have identified the differential role of ATRA in promoting Foxp3+ Tregs in CD4+ and CD8+ cell populations. These results will help to determine a protocol for developing different Treg cell populations and may provide novel insights into clinical cell therapy for patients with autoimmune diseases and those needing organ transplantation.

Nanofibre optic force transducers with sub-piconewton resolution via near-field plasmon–dielectric interactions

Ultrasensitive nanomechanical instruments, including the atomic force microscope (AFM)1-4 and optical and magnetic tweezers5-8, have helped shed new light on the complex mechanical environments of biological processes. However, it is difficult to scale down the size of these instruments due to their feedback mechanisms9, which, if overcome, would enable high-density nanomechanical probing inside materials. A variety of molecular force probes including mechanophores10, quantum dots11, fluorescent pairs12,13 and molecular rotors14-16 have been designed to measure intracellular stresses; however, fluorescence-based techniques can have short operating times due to photo-instability and it is still challenging to quantify the forces with high spatial and mechanical resolution. Here, we develop a compact nanofibre optic force transducer (NOFT) that utilizes strong near-field plasmon-dielectric interactions to measure local forces with a sensitivity of <200 fN. The NOFT system is tested by monitoring bacterial motion and heart-cell beating as well as detecting infrasound power in solution.

Rapid 3D bioprinting of decellularized extracellular matrix with regionally varied mechanical properties and biomimetic microarchitecture

Hepatocellular carcinoma (HCC), as the fifth most common malignant cancer, develops and progresses mostly in a cirrhotic liver where stiff nodules are separated by fibrous bands. Scaffolds that can provide a 3D cirrhotic mechanical environment with complex native composition and biomimetic architecture are necessary for the development of better predictive tissue models. Here, we developed photocrosslinkable liver decellularized extracellular matrix (dECM) and a rapid light-based 3D bioprinting process to pattern liver dECM with tailorable mechanical properties to serve as a platform for HCC progression study. 3D bioprinted liver dECM scaffolds were able to stably recapitulate the clinically relevant mechanical properties of cirrhotic liver tissue. When encapsulated in dECM scaffolds with cirrhotic stiffness, HepG2 cells demonstrated reduced growth along with an upregulation of invasion markers compared to healthy controls. Moreover, an engineered cancer tissue platform possessing tissue-scale organization and distinct regional stiffness enabled the visualization of HepG2 stromal invasion from the nodule with cirrhotic stiffness. This work demonstrates a significant advancement in rapid 3D patterning of complex ECM biomaterials with biomimetic architecture and tunable mechanical properties for in vitro disease modeling.

3D bioprinting of functional tissue models for personalized drug screening and in vitro disease modeling

3D bioprinting is emerging as a promising technology for fabricating complex tissue constructs with tailored biological components and mechanical properties. Recent advances have enabled scientists to precisely position materials and cells to build functional tissue models for in vitro drug screening and disease modeling. This review presents state-of-the-art 3D bioprinting techniques and discusses the choice of cell source and biomaterials for building functional tissue models that can be used for personalized drug screening and disease modeling. In particular, we focus on 3D-bioprinted liver models, cardiac tissues, vascularized constructs, and cancer models for their promising applications in medical research, drug discovery, toxicology, and other pre-clinical studies.

Three-Dimensional Printing of Bisphenol A-Free Polycarbonates

Polycarbonates are widely used in food packages, drink bottles, and various healthcare products such as dental sealants and tooth coatings. However, bisphenol A (BPA) and phosgene used in the production of commercial polycarbonates pose major concerns to public health safety. Here, we report a green pathway to prepare BPA-free polycarbonates (BFPs) by thermal ring-opening polymerization and photopolymerization. Polycarbonates prepared from two cyclic carbonates in different mole ratios demonstrated tunable mechanical stiffness, excellent thermal stability, and high optical transparency. Three-dimensional (3D) printing of the new BFPs was demonstrated using a two-photon laser direct writing system and a rapid 3D optical projection printer to produce structures possessing complex high-resolution geometries. Seeded C3H10T1/2 cells also showed over 95% viability with potential applications in biological studies. By combining biocompatible BFPs with 3D printing, novel safe and high-performance biomedical devices and healthcare products could be developed with broad long-term benefits to society.