Enhanced Maturation of 3D Bioprinted Skeletal Muscle Tissue Constructs Encapsulating Soluble Factor-Releasing Microparticles.

Several microfabrication technologies have been used to engineer native-like skeletal muscle tissues. However, the successful development of muscle remains a significant challenge in the tissue engineering field. Muscle tissue engineering aims to combine muscle precursor cells aligned within a highly organized 3D structure and biological factors crucial to support cell differentiation and maturation into functional myotubes and myofibers. In this study, the use of 3D bioprinting is proposed for the fabrication of muscle tissues using gelatin methacryloyl (GelMA) incorporating sustained insulin-like growth factor-1 (IGF-1)-releasing microparticles and myoblast cells. This study hypothesizes that functional and mature myotubes will be obtained more efficiently using a bioink that can release IGF-1 sustainably for in vitro muscle engineering. Synthesized microfluidic-assisted polymeric microparticles demonstrate successful adsorption of IGF-1 and sustained release of IGF-1 at physiological pH for at least 21 days. Incorporating the IGF-1-releasing microparticles in the GelMA bioink assisted in promoting the alignment of myoblasts and differentiation into myotubes. Furthermore, the myotubes show spontaneous contraction in the muscle constructs bioprinted with IGF-1-releasing bioink. The proposed bioprinting strategy aims to improve the development of new therapies applied to the regeneration and maturation of muscle tissues.

An All-In-One Transient Theranostic Platform for Intelligent Management of Hemorrhage.

Developing theranostic devices to detect bleeding and effectively control hemorrhage in the prehospital setting is an unmet medical need. Herein, an all-in-one theranostic platform is presented, which is constructed by sandwiching silk fibroin (SF) between two silver nanowire (AgNW) based conductive electrodes to non-enzymatically diagnose local bleeding and stop the hemorrhage at the wound site. Taking advantage of the hemostatic property of natural SF, the device is composed of a shape-memory SF sponge, facilitating blood clotting, with ≈82% reduction in hemostatic time in vitro as compared with untreated blood. Furthermore, this sandwiched platform serves as a capacitive sensor that can detect bleeding and differentiate between blood and other body fluids (i.e., serum and water) via capacitance change. In addition, the AgNW electrode endows anti-infection efficiency against Escherichia coli and Staphylococcus aureus. Also, the device shows excellent biocompatibility and gradually biodegrades in vivo with no major local or systemic inflammatory responses. More importantly, the theranostic platform presents considerable hemostatic efficacy comparable with a commercial hemostat, Dengen, in rat liver bleeding models. The theranostic platform provides an unexplored strategy for the intelligent management of hemorrhage, with the potential to significantly improve patients' well-being through the integration of diagnostic and therapeutic capabilities.

Tunable hybrid hydrogels with multicellular spheroids for modeling desmoplastic pancreatic cancer.

The tumor microenvironment consists of diverse, complex etiological factors. The matrix component of pancreatic ductal adenocarcinoma (PDAC) plays an important role not only in physical properties such as tissue rigidity but also in cancer progression and therapeutic responsiveness. Although significant efforts have been made to model desmoplastic PDAC, existing models could not fully recapitulate the etiology to mimic and understand the progression of PDAC. Here, two major components in desmoplastic pancreatic matrices, hyaluronic acid- and gelatin-based hydrogels, are engineered to provide matrices for tumor spheroids composed of PDAC and cancer-associated fibroblasts (CAF). Shape analysis profiles reveals that incorporating CAF contributes to a more compact tissue formation. Higher expression levels of markers associated with proliferation, epithelial to mesenchymal transition, mechanotransduction, and progression are observed for cancer-CAF spheroids cultured in hyper desmoplastic matrix-mimicking hydrogels, while the trend can be observed when those are cultured in desmoplastic matrix-mimicking hydrogels with the presence of transforming growth factor-ß1 (TGF-ß1). The proposed multicellular pancreatic tumor model, in combination with proper mechanical properties and TGF-ß1 supplement, makes strides in developing advanced pancreatic models for resembling and monitoring the progression of pancreatic tumors, which could be potentially applicable for realizing personalized medicine and drug testing applications.

Oxygen-generating microparticles downregulate HIF-1α expression, increase cardiac contractility, and mitigate ischemic injury.

Myocardial hypoxia is the low oxygen tension in the heart tissue implicated in many diseases, including ischemia, cardiac dysfunction, or after heart procurement for transplantation. Oxygen-generating microparticles have recently emerged as a potential strategy for supplying oxygen to sustain cell survival, growth, and tissue functionality in hypoxia. Here, we prepared oxygen-generating microparticles with poly D,L-lactic-co-glycolic acid, and calcium peroxide (CPO), which yielded a continuous morphology capable of sustained oxygen release for up to 24 h. We demonstrated that CPO microparticles increased primary rat cardiomyocyte metabolic activity while not affecting cell viability during hypoxia. Moreover, hypoxia-inducible factor (HIF)-1α, which is upregulated during hypoxia, can be downregulated by delivering oxygen using CPO microparticles. Single-cell traction force microscopy data demonstrated that the reduced energy generated by hypoxic cells could be restored using CPO microparticles. We engineered cardiac tissues that showed higher contractility in the presence of CPO microparticles compared to hypoxic cells. Finally, we observed reduced myocardial injuries in ex vivo rabbit hearts treated with CPO microparticles. In contrast, an acute early myocardial injury was observed for the hearts treated with control saline solution in hypoxia. In conclusion, CPO microparticles improved cell and tissue contractility and gene expression while reducing hypoxia-induced myocardial injuries in the heart. STATEMENT OF SIGNIFICANCE: Oxygen-releasing microparticles can reduce myocardial ischemia, allograft rejection, or irregular heartbeats after heart transplantation. Here we present biodegradable oxygen-releasing microparticles that are capable of sustained oxygen release for more than 24 hrs. We then studied the impact of sustained oxygen release from microparticles on gene expresseion and cardiac cell and tissue function. Previous studies have not measured cardiac tissue or cell mechanics during hypoxia, which is important for understanding proper cardiac function and beating. Using traction force microscopy and an engineered tissue-on-a-chip, we demonstrated that our oxygen-releasing microparticles improve cell and tissue contractility during hypoxia while downregulating the HIF-1α expression level. Finally, using the microparticles, we showed reduced myocardial injuries in rabbit heart tissue, confirming the potential of the particles to be used for organ transplantation or tissue engineering.

A Microfluidic Contact Lens to Address Contact Lens-Induced Dry Eye.

The contact lens (CL) industry has made great strides in improving CL-wearing experiences. However, a large amount of CL wearers continue to experience ocular dryness, known as contact lens-induced dry eye (CLIDE), stemming from the reduction in tear volume, tear film instability, increased tear osmolarity followed by inflammation and resulting in ocular discomfort and visual disturbances. In this article, to address tear film thinning between the CL and the ocular surface, the concept of using a CL with microchannels to deliver the tears from the pre-lens tear film (PrLTF) to the post-lens ocular surface using in vitro eye-blink motion is investigated. This study reports an eye-blink mimicking system with microfluidic poly(2-hydroxyethyl methacrylate) (poly(HEMA)) hydrogel with integrated microchannels to demonstrate eye-blink assisted flow through microchannels. This in vitro experimental study provides a proof-of-concept result that tear transport from PrLTF to post-lens tear film can be enhanced by an artificial eyelid motion in a pressure range of 0.1-5 kPa (similar to human eyelid pressure) through poly(HEMA) microchannels. Simulation is conducted to support the hypothesis. This work demonstrates the feasibility of developing microfluidic CLs with the potential to help prevent or minimize CLIDE and discomfort by the enhanced transport of pre-lens tears to the post-lens ocular surface.

Recent Advances in Bioinspired Hydrogels: Materials, Devices, and Biosignal Computing.

The remarkable ability of biological systems to sense and adapt to complex environmental conditions has inspired new materials and novel designs for next-generation wearable devices. Hydrogels are being intensively investigated for their versatile functions in wearable devices due to their superior softness, biocompatibility, and rapid stimulus response. This review focuses on recent strategies for developing bioinspired hydrogel wearable devices that can accommodate mechanical strain and integrate seamlessly with biological systems. We will provide an overview of different types of bioinspired hydrogels tailored for wearable devices. Next, we will discuss the recent progress of bioinspired hydrogel wearable devices such as electronic skin and smart contact lenses. Also, we will comprehensively summarize biosignal readout methods for hydrogel wearable devices as well as advances in powering and wireless data transmission technologies. Finally, current challenges facing these wearable devices are discussed, and future directions are proposed.

Flexible patch with printable and antibacterial conductive hydrogel electrodes for accelerated wound healing.

Electrical stimulation can facilitate wound healing with high efficiency and limited side effects. However, current electrical stimulation devices have poor conformability with wounds due to their bulky nature and the rigidity of electrodes utilized. Here, a flexible electrical patch (ePatch) made with conductive hydrogel as electrodes to improve wound management was reported. The conductive hydrogel was synthesized using silver nanowire (AgNW) and methacrylated alginate (MAA), with the former chosen as the electrode material considering its antibacterial properties, and the latter used due to its clinical suitability in wound healing. The composition of the hydrogel was optimized to enable printing on medical-grade patches for personalized wound treatment. The ePatch was shown to promote re-epithelization, enhance angiogenesis, mediate immune response, and prevent infection development in the wound microenvironment. In vitro studies indicated an elevated secretion of growth factors with enhanced cell proliferation and migration ability in response to electrical stimulation. An in vivo study in the Sprague-Dawley rat model revealed a rapid wound closure within 7 days compared to 20 days of usual healing process in rodents.

Co-Electrospun Silk Fibroin and Gelatin Methacryloyl Sheet Seeded with Mesenchymal Stem Cells for Tendon Regeneration.

Silk fibroin (SF) is a promising biomaterial for tendon repair, but its relatively rigid mechanical properties and low cell affinity have limited its application in regenerative medicine. Meanwhile, gelatin-based polymers have advantages in cell attachment and tissue remodeling but have insufficient mechanical strength to regenerate tough tissue such as tendons. Taking these aspects into account, in this study, gelatin methacryloyl (GelMA) is combined with SF to create a mechanically strong and bioactive nanofibrous scaffold (SG). The mechanical properties of SG nanofibers can be flexibly modulated by varying the ratio of SF and GelMA. Compared to SF nanofibers, mesenchymal stem cells (MSCs) seeded on SG fibers with optimal composition (SG7) exhibit enhanced growth, proliferation, vascular endothelial growth factor production, and tenogenic gene expression behavior. Conditioned media from MSCs cultured on SG7 scaffolds can greatly promote the migration and proliferation of tenocytes. Histological analysis and tenogenesis-related immunofluorescence staining indicate SG7 scaffolds demonstrate enhanced in vivo tendon tissue regeneration compared to other groups. Therefore, rational combinations of SF and GelMA hybrid nanofibers may help to improve therapeutic outcomes and address the challenges of tissue-engineered scaffolds for tendon regeneration.

Lab-on-a-Contact Lens: Recent Advances and Future Opportunities in Diagnostics and Therapeutics.

The eye is one of the most complex organs in the human body, containing rich and critical physiological information (e.g., intraocular pressure, corneal temperature, and pH) as well as a library of metabolite biomarkers (e.g., glucose, proteins, and specific ions). Smart contact lenses (SCLs) can serve as a wearable intelligent ocular prosthetic device capable of noninvasive and continuous monitoring of various essential physical/biochemical parameters and drug loading/delivery for the treatment of ocular diseases. Advances in SCL technologies and the growing public interest in personalized health are accelerating SCL research more than ever before. Here, the current status and potential of SCL development through a comprehensive review from fabrication to applications to commercialization are discussed. First, the material, fabrication, and platform designs of the SCLs for the diagnostic and therapeutic applications are discussed. Then, the latest advances in diagnostic and therapeutic SCLs for clinical translation are reviewed. Later, the established techniques for wearable power transfer and wireless data transmission applied to current SCL devices are summarized. An outlook, future opportunities, and challenges for developing next-generation SCL devices are also provided. With the rise in interest of SCL development, this comprehensive and essential review can serve as a new paradigm for the SCL devices.

Epidermis-Inspired Wearable Piezoresistive Pressure Sensors Using Reduced Graphene Oxide Self-Wrapped Copper Nanowire Networks.

Wearable piezoresistive sensors are being developed as electronic skins (E-skin) for broad applications in human physiological monitoring and soft robotics. Tactile sensors with sufficient sensitivities, durability, and large dynamic ranges are required to replicate this critical component of the somatosensory system. Multiple micro/nanostructures, materials, and sensing modalities have been reported to address this need. However, a trade-off arises between device performance and device complexity. Inspired by the microstructure of the spinosum at the dermo epidermal junction in skin, a low-cost, scalable, and high-performance piezoresistive sensor is developed with high sensitivity (0.144 kPa-1 ), extensive sensing range ( 0.1-15 kPa), fast response time (less than 150 ms), and excellent long-term stability (over 1000 cycles). Furthermore, the piezoresistive functionality of the device is realized via a flexible transparent electrode (FTE) using a highly stable reduced graphene oxide self-wrapped copper nanowire network. The developed nanowire-based spinosum microstructured FTEs are amenable to wearable electronics applications.

pH-Responsive doxorubicin delivery using shear-thinning biomaterials for localized melanoma treatment.

Injectable shear-thinning biomaterials (STBs) have attracted significant attention because of their efficient and localized delivery of cells as well as various molecules ranging from growth factors to drugs. Recently, electrostatic interaction-based STBs, including gelatin/LAPONITE® nanocomposites, have been developed through a simple assembly process and show outstanding shear-thinning properties and injectability. However, the ability of different compositions of gelatin and LAPONITE® to modulate doxorubicin (DOX) delivery at different pH values to enhance the effectiveness of topical skin cancer treatment is still unclear. Here, we fabricated injectable STBs using gelatin and LAPONITE® to investigate the influence of LAPONITE®/gelatin ratio on mechanical characteristics, capacity for DOX release in response to different pH values, and cytotoxicity toward malignant melanoma. The release profile analysis of various compositions of DOX-loaded STBs under different pH conditions revealed that lower amounts of LAPONITE® (6NC25) led to higher pH-responsiveness capable of achieving a localized, controlled, and sustained release of DOX in an acidic tumor microenvironment. Moreover, we showed that 6NC25 had a lower storage modulus and required lower injection forces compared to those with higher LAPONITE® ratios. Furthermore, DOX delivery analysis in vitro and in vivo demonstrated that DOX-loaded 6NC25 could efficiently target subcutaneous malignant tumors via DOX-induced cell death and growth restriction.

Droplet-based microfluidics in biomedical applications.

Droplet-based microfluidic systems have been employed to manipulate discrete fluid volumes with immiscible phases. Creating the fluid droplets at microscale has led to a paradigm shift in mixing, sorting, encapsulation, sensing, and designing high throughput devices for biomedical applications. Droplet microfluidics has opened many opportunities in microparticle synthesis, molecular detection, diagnostics, drug delivery, and cell biology. In the present review, we first introduce standard methods for droplet generation (i.e. passive and active methods) and discuss the latest examples of emulsification and particle synthesis approaches enabled by microfluidic platforms. Then, the applications of droplet-based microfluidics in different biomedical applications are detailed. Finally, a general overview of the latest trends along with the perspectives and future potentials in the field are provided.

Engineering liver microtissues to study the fusion of HepG2 with mesenchymal stem cells and invasive potential of fused cells.

Increasing evidence from cancer cell fusion with different cell types in the tumor microenvironment has suggested a probable mechanism for how metastasis-initiating cells could be generated in tumors. Although human mesenchymal stem cells (hMSCs) have been known as promising candidates to create hybrid cells with cancer cells, the role of hMSCs in fusion with cancer cells is still controversial. Here, we fabricated a liver-on-a-chip platform to monitor the fusion of liver hepatocellular cells (HepG2) with hMSCs and study their invasive potential. We demonstrated that hMSCs might play dual roles in HepG2 spheroids. The analysis of tumor growth with different fractions of hMSCs in HepG2 spheroids revealed hMSCs' role in preventing HepG2 growth and proliferation, while the hMSCs presented in the HepG2 spheroids led to the generation of HepG2-hMSC hybrid cells with much higher invasiveness compared to HepG2. These invasive HepG2-hMSC hybrid cells expressed high levels of markers associated with stemness, proliferation, epithelial to mesenchymal transition, and matrix deposition, which corresponded to the expression of these markers for hMSCs escaping from hMSC spheroids. In addition, these fused cells were responsible for collective invasion following HepG2 by depositing Collagen I and Fibronectin in their surrounding microenvironment. Furthermore, we showed that hepatic stellate cells (HSCs) could also be fused with HepG2, and the HepG2-HSC hybrid cells possessed similar features to those from HepG2-hMSC fusion. This fusion of HepG2 with liver-resident HSCs may propose a new potential mechanism of hepatic cancer metastasis.

Bioengineered Multicellular Liver Microtissues for Modeling Advanced Hepatic Fibrosis Driven Through Non-Alcoholic Fatty Liver Disease.

Despite considerable efforts in modeling liver disease in vitro, it remains difficult to recapitulate the pathogenesis of the advanced phases of non-alcoholic fatty liver disease (NAFLD) with inflammation and fibrosis. Here, a liver-on-a-chip platform with bioengineered multicellular liver microtissues is developed, composed of four major types of liver cells (hepatocytes, endothelial cells, Kupffer cells, and stellate cells) to implement a human hepatic fibrosis model driven by NAFLD: i) lipid accumulation in hepatocytes (steatosis), ii) neovascularization by endothelial cells, iii) inflammation by activated Kupffer cells (steatohepatitis), and iv) extracellular matrix deposition by activated stellate cells (fibrosis). In this model, the presence of stellate cells in the liver-on-a-chip model with fat supplementation showed elevated inflammatory responses and fibrosis marker up-regulation. Compared to transforming growth factor-beta-induced hepatic fibrosis models, this model includes the native pathological and chronological steps of NAFLD which shows i) higher fibrotic phenotypes, ii) increased expression of fibrosis markers, and iii) efficient drug transport and metabolism. Taken together, the proposed platform will enable a better understanding of the mechanisms underlying fibrosis progression in NAFLD as well as the identification of new drugs for the different stages of NAFLD.

A sub-1V, microwatt power-consumption iontronic pressure sensor based on organic electrochemical transistors.

Wearable and implantable pressure sensors are in great demand for personalized health monitoring. Pressure sensors with low operation voltage and low power-consumption are desired for energy-saving devices. Organic iontronic devices, such as organic electrochemical transistors (OECTs), have demonstrated great potential for low power-consumption bioelectronic sensing applications. The ability to conduct both electrons and ions, in addition to their low-operation voltage has enabled the widespread use of OECTs in different biosensing fields. However, despite these merits, OECTs have not been demonstrated for pressure sensing applications. This is because most OECTs are gated with aqueous electrolyte, which fails to respond to external pressure. Here, a low power-consumption iontronic pressure sensor is presented based on an OECT, in which an ionic hydrogel is used as a solid gating medium. The resultant iontronic device operated at voltages less than 1 V, with a power-consumption between ~ 101-103 µW, while maintaining a tunable sensitivity between 1 ~ 10 kPa-1. This work places OECTs on the frontline for developing low power-consumption iontronic pressure sensors and for biosensing applications.

Healthy and diseased in vitro models of vascular systems.

Irregular hemodynamics affects the progression of various vascular diseases, such atherosclerosis or aneurysms. Despite the extensive hemodynamics studies on animal models, the inter-species differences between humans and animals hamper the translation of such findings. Recent advances in vascular tissue engineering and the suitability of in vitro models for interim analysis have increased the use of in vitro human vascular tissue models. Although the effect of flow on endothelial cell (EC) pathophysiology and EC-flow interactions have been vastly studied in two-dimensional systems, they cannot be used to understand the effect of other micro- and macro-environmental parameters associated with vessel wall diseases. To generate an ideal in vitro model of the vascular system, essential criteria should be included: 1) the presence of smooth muscle cells or perivascular cells underneath an EC monolayer, 2) an elastic mechanical response of tissue to pulsatile flow pressure, 3) flow conditions that accurately mimic the hemodynamics of diseases, and 4) geometrical features required for pathophysiological flow. In this paper, we review currently available in vitro models that include flow dynamics and discuss studies that have tried to address the criteria mentioned above. Finally, we critically review in vitro fluidic models of atherosclerosis, aneurysm, and thrombosis.

Cancer-on-a-Chip for Modeling Immune Checkpoint Inhibitor and Tumor Interactions.

Cancer immunotherapies, including immune checkpoint inhibitor (ICI)-based therapies, have revolutionized cancer treatment. However, patient response to ICIs is highly variable, necessitating the development of methods to quickly assess efficacy. In this study, an array of miniaturized bioreactors has been developed to model tumor-immune interactions. This immunotherapeutic high-throughput observation chamber (iHOC) is designed to test the effect of anti-PD-1 antibodies on cancer spheroid (MDA-MB-231, PD-L1+) and T cell (Jurkat) interactions. This system facilitates facile monitoring of T cell inhibition and reactivation using metrics such as tumor infiltration and interleukin-2 (IL-2) secretion. Status of the tumor-immune interactions can be easily captured within the iHOC by measuring IL-2 concentration using a micropillar array where sensitive, quantitative detection is allowed after antibody coating on the surface of array. The iHOC is a platform that can be used to model and monitor cancer-immune interactions in response to immunotherapy in a high-throughput manner.

State of the art in integrated biosensors for organ-on-a-chip applications.

Organ-on-a-chip (OoC) models are bioengineered tissue constructs integrated with microfluidics that recapitulate the key features of the physiology of human organs and tissues with applications related to drug development and personalized medicine. The characterization of OoCs relies on conventional labor-intensive approaches despite the many years of research in the field. The physical environment of the tissue constructs, functionality, and metabolic activity of the cells must be monitored to ensure the behavior of the cells, and the cellular environments represent in vivo physiology. Current efforts focus on monitoring these parameters, particularly with in-line biosensors integrated with OoCs. In this review, we describe the recent advances in different biosensing modalities applied to monitor the environment and functionality of OoC models and offer suggestions for future directions in OoC applications.

Biofabrication of endothelial cell, dermal fibroblast, and multilayered keratinocyte layers for skin tissue engineering.

The skin serves a substantial number of physiological purposes and is exposed to numerous biological and chemical agents owing to its large surface area and accessibility. Yet, current skin models are limited in emulating the multifaceted functions of skin tissues due to a lack of effort on the optimization of biomaterials and techniques at different skin layers for building skin frameworks. Here, we use biomaterial-based approaches and bioengineered techniques to develop a 3D skin model with layers of endothelial cell networks, dermal fibroblasts, and multilayered keratinocytes. Analysis of mechanical properties of gelatin methacryloyl (GelMA)-based bioinks mixed with different portions of alginate revealed bioprinted endothelium could be better modeled to optimize endothelial cell viability with a mixture of 7.5% GelMA and 2% alginate. Matrix stiffness plays a crucial role in modulating produced levels of Pro-Collagen I alpha-1 and matrix metalloproteinase-1 in human dermal fibroblasts and affecting their viability, proliferation, and spreading. Moreover, seeding human keratinocytes with gelatin-coating multiple times proved to be helpful in reducing culture time to create multiple layers of keratinocytes while maintaining their viability. The ability to fabricate selected biomaterials for each layer of skin tissues has implications in the biofabrication of skin systems for regenerative medicine and disease modeling.

Microphysiological Systems: Next Generation Systems for Assessing Toxicity and Therapeutic Effects of Nanomaterials.

Microphysiological systems, also known as organ-on-a-chip platforms, show promise for the development of new testing methods that can be more accurate than both conventional two-dimensional cultures and costly animal studies. The development of more intricate microphysiological systems can help to better mimic the human physiology and highlight the systemic effects of different drugs and materials. Nanomaterials are among a technologically important class of materials used for diagnostic, therapeutic, and monitoring purposes; all of which and can be tested using new organ-on-a-chip systems. In addition, the toxicity of nanomaterials which have entered the body from ambient air or diet can have deleterious effects on various body systems. This in turn can be studied in newly developed microphysiological systems. While organ-on-a-chip models can be useful, they cannot pick up secondary and systemic toxicity. Thus, the utilization of multi-organ-on-a-chip systems for advancing nanotechnology will largely be reflected in the future of drug development, toxicology studies and precision medicine. Various aspects of related studies, current challenges, and future perspectives are discussed in this paper.