Publications by authors named "Shangting You"

17 Publications

  • Page 1 of 1

Bioprinting of dual ECM scaffolds encapsulating limbal stem/progenitor cells in active and quiescent statuses.

Biofabrication 2021 08 13;13(4). Epub 2021 Aug 13.

Department of NanoEngineering, University of California San Diego, La Jolla, CA 92093, United States of America.

Limbal stem cell deficiency and corneal disorders are among the top global threats for human vision. Emerging therapies that integrate stem cell transplantation with engineered hydrogel scaffolds for biological and mechanical support are becoming a rising trend in the field. However, methods for high-throughput fabrication of hydrogel scaffolds, as well as knowledge of the interaction between limbal stem/progenitor cells (LSCs) and the surrounding extracellular matrix (ECM) are still much needed. Here, we employed digital light processing (DLP)-based bioprinting to fabricate hydrogel scaffolds encapsulating primary LSCs and studied the ECM-dependent LSC phenotypes. The DLP-based bioprinting with gelatin methacrylate (GelMA) or hyaluronic acid glycidyl methacrylate (HAGM) generated microscale hydrogel scaffolds that could support the viability of the encapsulated primary rabbit LSCs (rbLSCs) in culture. Immunocytochemistry and transcriptional analysis showed that the encapsulated rbLSCs remained active in GelMA-based scaffolds while exhibited quiescence in the HAGM-based scaffolds. The primary human LSCs encapsulated within bioprinted scaffolds showed consistent ECM-dependent active/quiescent statuses. Based on these results, we have developed a novel bioprinted dual ECM 'Yin-Yang' model encapsulating LSCs to support both active and quiescent statues. Our findings provide valuable insights towards stem cell therapies and regenerative medicine for corneal reconstruction.
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http://dx.doi.org/10.1088/1758-5090/ac1992DOI Listing
August 2021

Rapid 3D Bioprinting of Glioblastoma Model Mimicking Native Biophysical Heterogeneity.

Small 2021 04 27;17(15):e2006050. Epub 2021 Jan 27.

Department of NanoEngineering, University of California San Diego, La Jolla, CA, 92093, USA.

Glioblastoma multiforme (GBM) is the most lethal primary brain tumor characterized by high cellular and molecular heterogeneity, hypervascularization, and innate drug resistance. Cellular components and extracellular matrix (ECM) are the two primary sources of heterogeneity in GBM. Here, biomimetic tri-regional GBM models with tumor regions, acellular ECM regions, and an endothelial region with regional stiffnesses patterned corresponding to the GBM stroma, pathological or normal brain parenchyma, and brain capillaries, are developed. Patient-derived GBM cells, human endothelial cells, and hyaluronic acid derivatives are used to generate a species-matched and biochemically relevant microenvironment. This in vitro study demonstrates that biophysical cues are involved in various tumor cell behaviors and angiogenic potentials and promote different molecular subtypes of GBM. The stiff models are enriched in the mesenchymal subtype, exhibit diffuse invasion of tumor cells, and induce protruding angiogenesis and higher drug resistance to temozolomide. Meanwhile, the soft models demonstrate enrichment in the classical subtype and support expansive cell growth. The three-dimensional bioprinting technology utilized in this study enables rapid, flexible, and reproducible patient-specific GBM modeling with biophysical heterogeneity that can be employed by future studies as a tunable system to interrogate GBM disease mechanisms and screen drug compounds.
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http://dx.doi.org/10.1002/smll.202006050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8049977PMC
April 2021

High-fidelity 3D Printing using Flashing Photopolymerization.

Addit Manuf 2019 Dec 19;30. Epub 2019 Aug 19.

Department of NanoEngineering, University of California San Diego, 9500 Gilman Drive, La Jolla, California, 92093, USA.

Photopolymerization-based 3D printing has emerged as a promising technique to fabricate 3D structures. However, during the printing process, polymerized materials such as hydrogels often become highly light-scattering, thus perturbing incident light distribution and thereby deteriorating the final print resolution. To overcome this scattering-induced resolution deterioration, we developed a novel method termed flashing photopolymerization (FPP). Our FPP approach is informed by the fundamental kinetics of photopolymerization reactions, where light exposure is delivered in millisecond-scale 'flashes', as opposed to continuous light exposure. During the period of flash exposure, the prepolymer material negligibly scatters light. The material then polymerizes and opacifies in absence of light, therefore the exposure pattern is not perturbed by scattering. Compared to the conventional use of a continuous wave (CW) light source, the FPP fabrication resolution is improved. FPP also shows little dependency on the exposure, thus minimizing trial-and-error type optimization. Using FPP, we demonstrate its use in generating high-fidelity 3D printed constructs.
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http://dx.doi.org/10.1016/j.addma.2019.100834DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7442265PMC
December 2019

A sequential 3D bioprinting and orthogonal bioconjugation approach for precision tissue engineering.

Biomaterials 2020 11 9;258:120294. Epub 2020 Aug 9.

Department of NanoEngineering, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA; Materials Science and Engineering Program, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA; Department of Bioengineering, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA; Chemical Engineering Program, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA. Electronic address:

Recent advances in 3D bioprinting have transformed the tissue engineering landscape by enabling the controlled placement of cells, biomaterials, and bioactive agents for the biofabrication of living tissues and organs. However, the application of 3D bioprinting is limited by the availability of cytocompatible and printable biomaterials that recapitulate properties of native tissues. Here, we developed an integrated 3D projection bioprinting and orthogonal photoconjugation platform for precision tissue engineering of tailored microenvironments. By using a photoreactive thiol-ene gelatin bioink, soft hydrogels can be bioprinted into complex geometries and photopatterned with bioactive moieties in a rapid and scalable manner via digital light projection (DLP) technology. This enables localized modulation of biophysical properties such as stiffness and microarchitecture as well as precise control over spatial distribution and concentration of immobilized functional groups. As such, well-defined properties can be directly incorporated using a single platform to produce desired tissue-specific functions within bioprinted constructs. We demonstrated high viability of encapsulated endothelial cells and human cardiomyocytes using our dual process and fabricated tissue constructs functionalized with VEGF peptide mimics to induce guided endothelial cell growth for programmable vascularization. This work represents a pivotal step in engineering multifunctional constructs with unprecedented control, precision, and versatility for the rational design of biomimetic tissues.
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http://dx.doi.org/10.1016/j.biomaterials.2020.120294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7489302PMC
November 2020

Photopolymerizable Biomaterials and Light-Based 3D Printing Strategies for Biomedical Applications.

Chem Rev 2020 10 23;120(19):10695-10743. Epub 2020 Apr 23.

Department of NanoEngineering, University of California San Diego, 9500 Gilman Drive, La Jolla, California 92093, United States.

Since the advent of additive manufacturing, known commonly as 3D printing, this technology has revolutionized the biofabrication landscape and driven numerous pivotal advancements in tissue engineering and regenerative medicine. Many 3D printing methods were developed in short course after Charles Hull first introduced the power of stereolithography to the world. However, materials development was not met with the same enthusiasm and remained the bottleneck in the field for some time. Only in the past decade has there been deliberate development to expand the materials toolbox for 3D printing applications to meet the true potential of 3D printing technologies. Herein, we review the development of biomaterials suited for light-based 3D printing modalities with an emphasis on bioprinting applications. We discuss the chemical mechanisms that govern photopolymerization and highlight the application of natural, synthetic, and composite biomaterials as 3D printed hydrogels. Because the quality of a 3D printed construct is highly dependent on both the material properties and processing technique, we included a final section on the theoretical and practical aspects behind light-based 3D printing as well as ways to employ that knowledge to troubleshoot and standardize the optimization of printing parameters.
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http://dx.doi.org/10.1021/acs.chemrev.9b00810DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7572843PMC
October 2020

High throughput direct 3D bioprinting in multiwell plates.

Biofabrication 2020 Apr 16. Epub 2020 Apr 16.

Nanoengineering, University of California San Diego, La Jolla, California, UNITED STATES.

Advances in three dimensional (3D) bioprinting have enabled the fabrication of sophisticated 3D tissue scaffolds for biological and medical applications, where high speed, high throughput production in well plates is a critical need. Here, we present an integrated 3D bioprinting platform based on microscale continuous optical printing, capable of high throughput in situ rapid fabrication of complex 3D biomedical samples in multiwell plate formats for subsequent culture and analysis. Our high throughput 3D bioprinter (HT-3DP) was used to showcase constructs of varying spatial geometries of biomimetic significance, tunable mechanical properties, as well as reproducibility. Live hepatocellular carcinoma 3D tissue scaffolds were fabricated in situ in multiwell plates, after which a functional drug response assay against the chemotherapy drug doxorubicin was performed. Dual cell-type populations involving both live hepatocellular carcinoma as well as human umbilical vein endothelial cells were also printed to demonstrate dual-tissue fabrication capability. This work demonstrates a significant advancement in that the production rate of 3D bioprinted tissue scaffolds with controllable spatial architectures and mechanical properties can now be done on a high throughput scale, enabling rapid generation of in vitro 3D tissue models within conventional multiwell cell culture plates for high throughput preclinical drug screening and disease modeling.
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http://dx.doi.org/10.1088/1758-5090/ab89caDOI Listing
April 2020

Bionic 3D printed corals.

Nat Commun 2020 04 9;11(1):1748. Epub 2020 Apr 9.

Bioinspired Photonics Group, Department of Chemistry, University of Cambridge, Cambridge, UK.

Corals have evolved as optimized photon augmentation systems, leading to space-efficient microalgal growth and outstanding photosynthetic quantum efficiencies. Light attenuation due to algal self-shading is a key limiting factor for the upscaling of microalgal cultivation. Coral-inspired light management systems could overcome this limitation and facilitate scalable bioenergy and bioproduct generation. Here, we develop 3D printed bionic corals capable of growing microalgae with high spatial cell densities of up to 10 cells mL. The hybrid photosynthetic biomaterials are produced with a 3D bioprinting platform which mimics morphological features of living coral tissue and the underlying skeleton with micron resolution, including their optical and mechanical properties. The programmable synthetic microenvironment thus allows for replicating both structural and functional traits of the coral-algal symbiosis. Our work defines a class of bionic materials that is capable of interacting with living organisms and can be exploited for applied coral reef research and photobioreactor design.
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http://dx.doi.org/10.1038/s41467-020-15486-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7145811PMC
April 2020

Rapid continuous 3D printing of customizable peripheral nerve guidance conduits.

Mater Today (Kidlington) 2018 Nov 27;21(9):951-959. Epub 2018 Apr 27.

Department of NanoEngineering, University of California San Diego, La Jolla, CA 92093, United States.

Engineered nerve guidance conduits (NGCs) have been demonstrated for repairing peripheral nerve injuries. However, there remains a need for an advanced biofabrication system to build NGCs with complex architectures, tunable material properties, and customizable geometrical control. Here, a rapid continuous 3D-printing platform was developed to print customizable NGCs with unprecedented resolution, speed, flexibility, and scalability. A variety of NGC designs varying in complexity and size were created including a life-size biomimetic branched human facial NGC. implantation of NGCs with microchannels into complete sciatic nerve transections of mouse models demonstrated the effective directional guidance of regenerating sciatic nerves via branching into the microchannels and extending toward the distal end of the injury site. Histological staining and immunostaining further confirmed the progressive directional nerve regeneration and branching behavior across the entire NGC length. Observational and functional tests, including the von Frey threshold test and thermal test, showed promising recovery of motor function and sensation in the ipsilateral limbs grafted with the 3D-printed NGCs.
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http://dx.doi.org/10.1016/j.mattod.2018.04.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6538503PMC
November 2018

Projection Printing of Ultrathin Structures with Nanoscale Thickness Control.

ACS Appl Mater Interfaces 2019 May 18;11(17):16059-16064. Epub 2019 Apr 18.

Department of NanoEngineering , University of California San Diego , La Jolla , California 92093 , United States.

Spatial control of photon energy has been a central part of many light-based manufacturing processes. We report a direct projection printing method for ultrathin structures with nanoscale thickness control by using a patterned evanescent field. The evanescent field is induced by total internal reflection at the interface between the substrate and a prepolymer solution, and it is patterned by a phase-only spatial light modulator. The ultrathin structure is printed on a high-refractive-index glass substrate through photopolymerization. An iterative algorithm is used to calculate the phase pattern for generating arbitrary holography images and making the image plane to coincide with the interface. The thickness of the pattern is limited by the penetration depth of the evanescent field. Experiment results demonstrated that polymer structures as thin as 200 nm can be patterned without significant process optimization. Such fine control in thickness could transform many techniques such as light-based 3D printing and laser direct-write manufacturing.
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http://dx.doi.org/10.1021/acsami.9b02728DOI Listing
May 2019

Nanoscale 3D printing of hydrogels for cellular tissue engineering.

J Mater Chem B 2018 Apr 14;6(15):2187-2197. Epub 2018 Mar 14.

Department of NanoEngineering, University of California, San Diego, La Jolla, CA 92093-0448, USA.

Hydrogel scaffolds that mimic the native extracellular matrix (ECM) environment is a crucial part of tissue engineering. It has been demonstrated that cell behaviors can be affected by not only the hydrogel's physical and chemical properties, but also its three dimensional (3D) geometrical structures. In order to study the influence of 3D geometrical cues on cell behaviors as well as the maturation and function of engineered tissues, it is imperative to develop 3D fabrication techniques to create micro and nanoscale hydrogel constructs. Among existing techniques that can effectively pattern hydrogels, two-photon polymerization (2PP)-based femtosecond laser 3D printing technology allows one to produce hydrogel structures with 100 nm resolution. This article reviews the basics of this technique as well as some of its applications in tissue engineering.
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http://dx.doi.org/10.1039/C8TB00301GDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6178227PMC
April 2018

Three-Dimensional Printing of Bisphenol A-Free Polycarbonates.

ACS Appl Mater Interfaces 2018 Feb 31;10(6):5331-5339. Epub 2018 Jan 31.

Department of NanoEngineering, University of California, San Diego , La Jolla, California 92093, United States.

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.
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http://dx.doi.org/10.1021/acsami.7b18312DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6536128PMC
February 2018

A 3D Tissue-Printing Approach for Validation of Diffusion Tensor Imaging in Skeletal Muscle.

Tissue Eng Part A 2017 09 24;23(17-18):980-988. Epub 2017 Mar 24.

1 Department of Bioengineering, University of California San Diego , La Jolla, California.

The ability to noninvasively assess skeletal muscle microstructure, which predicts function and disease, would be of significant clinical value. One method that holds this promise is diffusion tensor magnetic resonance imaging (DT-MRI), which is sensitive to the microscopic diffusion of water within tissues and has become ubiquitous in neuroimaging as a way of assessing neuronal structure and damage. However, its application to the assessment of changes in muscle microstructure associated with injury, pathology, or age remains poorly defined, because it is difficult to precisely control muscle microstructural features in vivo. However, recent advances in additive manufacturing technologies allow precision-engineered diffusion phantoms with histology informed skeletal muscle geometry to be manufactured. Therefore, the goal of this study was to develop skeletal muscle phantoms at relevant size scales to relate microstructural features to MRI-based diffusion measurements. A digital light projection based rapid 3D printing method was used to fabricate polyethylene glycol diacrylate based diffusion phantoms with (1) idealized muscle geometry (no geometry; fiber sizes of 30, 50, or 70 μm or fiber size of 50 μm with 40% of walls randomly deleted) or (2) histology-based geometry (normal and after 30-days of denervation) containing 20% or 50% phosphate-buffered saline (PBS). Mean absolute percent error (8%) of the printed phantoms indicated high conformity to templates when "fibers" were >50 μm. A multiple spin-echo echo planar imaging diffusion sequence, capable of acquiring diffusion weighted data at several echo times, was used in an attempt to combine relaxometry and diffusion techniques with the goal of separating intracellular and extracellular diffusion signals. When fiber size increased (30-70 μm) in the 20% PBS phantom, fractional anisotropy (FA) decreased (0.32-0.26) and mean diffusivity (MD) increased (0.44 × 10 mm/s-0.70 × 10 mm/s). Similarly, when fiber size increased from 30 to 70 μm in the 50% PBS diffusion phantoms, a small change in FA was observed (0.18-0.22), but MD increased from 0.86 × 10 mm/s to 1.79 × 10 mm/s. This study demonstrates a novel application of tissue engineering to understand complex diffusion signals in skeletal muscle. Through this work, we have also demonstrated the feasibility of 3D printing for skeletal muscle with relevant matrix geometries and physiologically relevant tissue characteristics.
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http://dx.doi.org/10.1089/ten.tea.2016.0438DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5610393PMC
September 2017

Resolution criteria in double-slit microscopic imaging experiments.

Sci Rep 2016 09 19;6:33764. Epub 2016 Sep 19.

Division of Physics and Applied Physics, School of Physical and Mathematical Sciences, Nanyang Technological University, 637371, Singapore.

Double-slit imaging is widely used for verifying the resolution of high-resolution and super-resolution microscopies. However, due to the fabrication limits, the slit width is generally non-negligible, which can affect the claimed resolution. In this paper we theoretically calculate the electromagnetic field distribution inside and near the metallic double slit using waveguide mode expansion method, and acquire the far-field image by vectorial Fourier optics. We find that the slit width has minimal influence when the illuminating light is polarized parallel to the slits. In this case, the claimed resolution should be based on the center-to-center distance of the double-slit.
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http://dx.doi.org/10.1038/srep33764DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5027385PMC
September 2016

Iterative phase-retrieval method for generating stereo array of polarization-controlled focal spots.

Opt Lett 2015 Aug;40(15):3532-5

This Letter introduces an iterative phase-retrieval method based on the Gerchberg-Saxton (G-S) algorithm for generating any arbitrary 3D pattern in image space, while simultaneously controlling the polarization orientation at each pixel. For proof-of-principle, we generate a stereo focal spot array with distinct polarization orientation for each spot. This method is universal for controlling the output polarization; the only requirement is that the input polarization should be spatially inhomogeneous. This work has the potential to impact coherent imaging techniques and spectroscopy.
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http://dx.doi.org/10.1364/OL.40.003532DOI Listing
August 2015

Resolution-enhanced surface plasmon-coupled emission microscopy.

Opt Express 2015 May;23(10):13159-71

A novel fluorescence emission difference technique is proposed for further enhancements of the lateral resolution in surface plasmon-coupled emission microscopy (SPCEM). In the proposed method, the difference between the image with phase modulation by using a 0-2π vortex phase plate (VPP) along with a diaphragm and the original image obtained from SPCEM is used to estimate the spatial distribution of the analyzed sample. By optimizing the size of the diaphragm and the subtractive factor, the lateral resolution can be enhanced by about 20% and 33%, compared with that in SPCEM with a single 0-2π VPP and conventional wide-field fluorescence microscopy, respectively. Related simulation results are presented to verify the capability of the proposed method for improving lateral resolution and reducing imaging distortion. It is believed that the proposed method has potentials to improve the performance of SPCEM, thus facilitating biological observation and research.
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http://dx.doi.org/10.1364/OE.23.013159DOI Listing
May 2015

Isotropic superresolution imaging for fluorescence emission difference microscopy.

Appl Opt 2014 Nov;53(33):7838-44

Fluorescence emission diffraction microscopy (FED) has proven to be an effective sub-diffraction-limited imaging method. In this paper, we theoretically propose a method to further enhance the resolving capability of FED. Using a coated mirror and only one objective lens, this method achieves not only the same axial resolution as 4Pi microscopy but also a higher lateral resolution. The point spread function (PSF) of our method is isotropic. According to calculations, the full width at half-maximum (FWHM) of the isotropic FED's PSF is 0.17λ along all three spatial directions. Compared with confocal microscopy, the lateral resolution is improved 0.7-fold, and the axial resolution is improved 3.1-fold. Simulation tests also demonstrate this method's advantage over traditional microscopy techniques.
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http://dx.doi.org/10.1364/AO.53.007838DOI Listing
November 2014

Eliminating deformations in fluorescence emission difference microscopy.

Opt Express 2014 Oct;22(21):26375-85

We propose a method for eliminating the deformations in fluorescence emission difference microscopy (FED). Due to excessive subtraction, negative values are inevitable in the original FED method, giving rise to deformations. We propose modulating the beam to generate an extended solid focal spot and a hollow focal spot. Negative image values can be avoided by using these two types of excitation spots in FED imaging. Hence, deformations are eliminated, and the signal-to-noise ratio is improved. In deformation-free imaging, the resolution is higher than that of confocal imaging by 32%. Compared to standard FED imaging with the same level of deformations, our method provides superior resolution.
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http://dx.doi.org/10.1364/OE.22.026375DOI Listing
October 2014
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