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Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease that progressively and irreversibly alters the lung parenchyma, eventually leading to respiratory failure. The study of this disease has been historically challenging due to the myriad of complex processes that contribute to fibrogenesis and the inherent difficulty in accurately recreating the human pulmonary environment

Maintaining the three-dimensional architecture and cellular complexity of lung tissue ex vivo can enable elucidation of the cellular and molecular pathways underlying chronic pulmonary diseases. Precision-cut lung slices (PCLS) are one human-lung model with the potential to support critical mechanistic studies and early drug discovery. However, many studies report short culture times of 7–10 days. Here, we systematically evaluated poly(ethylene glycol)-based hydrogel platforms for the encapsulation of PCLS. We demonstrated the ability to support ex vivo culture of embedded PCLS for at least 21 days compared with control PCLS floating in media. These customized hydrogels maintained PCLS architecture (no difference), viability (4.7-fold increase, P

Pulmonary arterial hypertension (PAH) is a progressive disease of the lung vasculature, characterized by elevated pulmonary blood pressure, remodeling of the pulmonary arteries, and ultimately right ventricular failure. Therapeutic interventions for PAH are limited in part by the lack of in vitro screening platforms that accurately reproduce dynamic arterial wall mechanical properties. Here we present a 3D-bioprinted model of the pulmonary arterial adventitia comprised of a phototunable poly(ethylene glycol) alpha methacrylate (PEG-αMA)-based hydrogel and primary human pulmonary artery adventitia fibroblasts (HPAAFs). This unique biomaterial emulates PAH pathogenesis in vitro through a two-step polymerization reaction. First, PEG-αMA macromer was crosslinked off-stoichiometry by 3D bioprinting an acidic bioink solution into a basic gelatin support bath initiating a base-catalyzed thiol-ene reaction with synthetic and biodegradable crosslinkers. Then, matrix stiffening was induced by photoinitiated homopolymerization of unreacted αMA end groups. A design of experiments approach produced a hydrogel platform that exhibited an initial elastic modulus (E) within the range of healthy pulmonary arterial tissue (E = 4.7 ± 0.09 kPa) that was stiffened to the pathologic range of hypertensive tissue (E = 12.8 ± 0.47 kPa) and supported cellular proliferation over time. A higher percentage of HPAAFs cultured in stiffened hydrogels expressed the fibrotic marker alpha-smooth muscle actin than cells in soft hydrogels (88 ± 2% versus 65 ± 4%). Likewise, a greater percentage of HPAAFs were positive for the proliferation marker 5-ethynyl-2ʹ-deoxyuridine (EdU) in stiffened models (66 ± 6%) compared to soft (39 ± 6%). These results demonstrate that 3D-bioprinted, phototunable models of pulmonary artery adventitia are a tool that enable investigation of fibrotic pathogenesis in vitro.

The global COVID-19 pandemic disrupted supply chains across the world, resulting in a critical shortage of personal protective equipment (PPE) for frontline healthcare workers. To preserve PPE for healthcare providers treating COVID-19 positive patients and to reduce asymptomatic transmission, the Department of Bioengineering at the University of Colorado, Denver | Anschutz Medical Campus collaborated with National Jewish Health to design and test patterns for cloth face coverings. A public campaign to sew and donate the final pattern was launched and over 2500 face coverings have been donated as a result. Now that nearly three million cases of COVID-19 have been confirmed in the United States, many state and local governments are requiring cloth face coverings be worn in public. Here, we present the collaborative design and testing process, as well as the final pattern for non-patient facing hospital workers and community members alike.

Fibrotic disorders account for over one third of mortalities worldwide. Despite great efforts to study the cellular and molecular processes underlying fibrosis, there are currently few effective therapies. Dual-stage polymerization reactions are an innovative tool for recreating heterogeneous increases in extracellular matrix (ECM) modulus, a hallmark of fibrotic diseases in vivo. Here, we present a clickable decellularized ECM (dECM) crosslinker incorporated into a dynamically responsive poly(ethylene glycol)-α-methacrylate (PEGαMA) hybrid-hydrogel to recreate ECM remodeling in vitro. An off-stoichiometry thiol-ene Michael addition between PEGαMA (8-arm, 10 kg mol-1) and the clickable dECM resulted in hydrogels with an elastic modulus of E = 3.6 ± 0.24 kPa, approximating healthy lung tissue (1-5 kPa). Next, residual αMA groups were reacted via a photo-initiated homopolymerization to increase modulus values to fibrotic levels (E = 13.4 ± 0.82 kPa) in situ. Hydrogels with increased elastic moduli, mimicking fibrotic ECM, induced a significant increase in the expression of myofibroblast transgenes. The proportion of primary fibroblasts from dual-reporter mouse lungs expressing collagen 1a1 and alpha-smooth muscle actin increased by approximately 60% when cultured on stiff and dynamically stiffened hybrid-hydrogels compared to soft. Likewise, fibroblasts expressed significantly increased levels of the collagen 1a1 transgene on stiff regions of spatially patterned hybrid-hydrogels compared to the soft areas. Collectively, these results indicate that hybrid-hydrogels are a new tool that can be implemented to spatiotemporally induce a phenotypic transition in primary murine fibroblasts in vitro.

Tracheal intubation disrupts physiological homeostasis of secretion production and clearance, resulting in secretion accumulation within endotracheal tubes (ETTs). Novel in vitro and in vivo models were developed to specifically recapitulate the clinical manifestations of ETT occlusion. The novel Sharklet™ micropatterned ETT was evaluated, using these models, for the ability to reduce the accumulation of both bacterial biofilm and airway mucus compared to a standard care ETT. Novel ETTs with micropattern on the inner and outer surfaces were placed adjacent to standard care ETTs in in vitro biofilm and airway patency (AP) models. The primary outcome for the biofilm model was to compare commercially-available ETTs (standard care and silver-coated) to micropatterned for quantity of biofilm accumulation. The AP model's primary outcome was to evaluate accumulation of artificial airway mucus. A 24-h ovine mechanical ventilation model evaluated the primary outcome of relative quantity of airway secretion accumulation in the ETTs tested. The secondary outcome was measuring the effect of secretion accumulation in the ETTs on airway resistance. Micropatterned ETTs significantly reduced biofilm by 71% (p = 0.016) compared to smooth ETTs. Moreover, micropatterned ETTs reduced lumen occlusion, in the AP model, as measured by cross-sectional area, in distal (85%, p = 0.005), middle (84%, p = 0.001) and proximal (81%, p = 0.002) sections compared to standard care ETTs. Micropatterned ETTs reduced the volume of secretion accumulation in a sheep model of occlusion by 61% (p

The extracellular matrix (ECM) controls keratinocyte proliferation, migration, and differentiation through β-integrin signaling. Wound-healing research requires expanding cells in vitro while maintaining replicative capacity; however, early terminal differentiation under traditional culture conditions limits expansion. Here, a design of experiments approach identifies poly(ethylene glycol)-based hydrogel formulations with mechanical properties (elastic modulus, E = 20.9 ± 0.56 kPa) and bioactive peptide sequences that mimic the epidermal ECM. These hydrogels enable systematic investigation of the influence of cell-binding domains from fibronectin (RGDS), laminin (YIGSR), and collagen IV (HepIII) on keratinocyte stemness and β1 integrin expression. Quantification of 14-day keratin protein expression shows four hydrogels improve stemness compared to standard techniques. Three hydrogels increase β1 integrin expression, demonstrating a positive linear relationship between stemness and β1 integrin expression. Multifactorial statistical analysis predicts an optimal peptide combination ([RGDS] = 0.67 mm, [YIGSR] = 0.13 mm, and [HepIII] = 0.02 mm) for maintaining stemness in vitro. Best-performing hydrogels exhibit no decrease in Ki-67-positive cells compared to standards (15% decrease, day 7 to 14; p < 0.05, Tukey Test). These data demonstrate that precisely designed hydrogel biomaterials direct integrin expression and promote proliferation, improving the regenerative capability of cultured keratinocytes for basic science and translational work.

Biomaterial scaffolds have been a foundational element of the tissue engineering paradigm since the inception of the field. Over the years there has been a progressive move toward the rational design and fabrication of bio-inspired materials that mimic the composition as well as the architecture and 3D structure of tissues. In this review, we chronicle advances in the field that address key challenges in tissue engineering as well as some emerging applications. Specifically, a summary of the materials and chemistries used to engineer bio-inspired 3D matrices that mimic numerous aspects of the extracellular matrix is provided, along with an overview of bioprinting, an additive manufacturing approach, for the fabrication of engineered tissues with precisely controlled 3D structures and architectures. To emphasize the potential clinical impact of the bio-inspired paradigm in biomaterials engineering, some applications of bio-inspired matrices are discussed in the context of translational tissue engineering. However, focus is also given to recent advances in the use of engineered 3D cellular microenvironments for fundamental studies in cell biology, including photoresponsive systems that are shedding new light on how matrix properties influence cell phenotype and function. In an outlook for future work, the need for high-throughput methods both for screening and fabrication is highlighted. Finally, microscale organ-on-a-chip technologies are highlighted as a promising area for future investment in the application of bio-inspired microenvironments.

Purpose To evaluate the ability of Sharklet (SK) micropatterns to inhibit lens epithelial cell (LEC) migration. Sharklet Technologies, Inc. (STI) and InSight Innovations, LLC have proposed to develop a Sharklet-patterned protective membrane (PM) to be implanted in combination with a posterior chamber intraocular lens (IOL) to inhibit cellular migration across the posterior capsule, and thereby reduce rates of posterior capsular opacification (PCO). Methods A variety of STI micropatterns were evaluated versus smooth (SM) controls in a modified scratch wound assay for the ability to reduce or inhibit LEC migration. The best performing topography was selected, translated to a radial design, and applied to PM prototypes. The PM prototypes were tested in an in vitro PCO model for reduction of cell migration behind an IOL versus unpatterned prototypes and IOLs with no PM. In both assays, cell migration was analyzed with fluorescent microscopy. Results All SK micropatterns significantly reduced LEC migration compared with SM controls. Micropatterns that protruded from the surface reduced migration more than recessed features. The best performing micropattern reduced LEC coverage by 80%, P = 0.0001 (ANOVA, Tukey Test). Micropatterned PMs reduced LEC migration in a PCO model by 50%, P = 0.0005 (ANOVA, Tukey Test) compared with both IOLs with no PM and IOLs with SM PMs. Conclusions Collectively, in vitro results indicate the implantation of micropatterned PMs in combination with posterior chamber IOLs could significantly reduce rates of clinically relevant PCO. This innovative technology is a globally accessible solution to high PCO rates. Translational relevance A novel IOL incorporating the SK micropattern in a membrane design surrounding the optic may help increase the success of cataract surgery by reducing secondary cataract, or PCO.