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Abstract BackgroundLaryngeal injury associated with traumatic or prolonged intubation may lead to voice, swallow, and airway complications. The interplay between inflammation and microbial population shifts induced by intubation may relate to clinical outcomes. The objective of this study was to investigate laryngeal mechanics, tissue inflammatory response, and local microbiome changes with laryngotracheal injury and localized delivery of therapeutics via drug-eluting endotracheal tube. MethodsA simulated traumatic intubation injury was created in Yorkshire crossbreed swine under direct laryngoscopy. Endotracheal tubes electrospun with roxadustat or valacyclovir- loaded polycaprolactone (PCL) fibers were placed in the injured airway for 3, 7, or 14 days (n = 3 per group/time and ETT type). Vocal fold stiffness was then evaluated with normal indentation and laryngeal tissue sections were histologically examined. Immunohistochemistry and inflammatory marker profiling were conducted to evaluate the inflammatory response associated with injury and ETT placement. Additionally, ETT biofilm formation was visualized using scanning electron microscopy and micro-computed tomography, while changes in the airway microbiome were profiled through 16S rRNA sequencing. ResultsLaryngeal tissue with roxadustat ETT placement had increasing localized stiffness outcomes over time and histological assessment indicated minimal epithelial ulceration and fibrosis, while inflammation remained severe across all timepoints. In contrast, vocal fold tissue with valacyclovir ETT placement showed no significant changes in stiffness over time; histological analysis presented a reduction in epithelial ulceration and inflammation scores along with increased fibrosis observed at 14 days. Immunohistochemistry revealed a decline in M1 and M2 macrophage markers over time for both ETT types. Among the cytokines, IL-8 levels differed significantly between the roxadustat and valacyclovir ETT groups, while no other cytokines showed statistically significant differences. Additionally, increased biofilm formation was observed in the coated ETTs with notable alterations in microbiota distinctive to each ETT type and across time. ConclusionThe injured and intubated airway resulted in increased laryngeal stiffness. Local inflammation and the type of therapeutic administered impacted the bacterial composition within the upper respiratory microbiome, which in turn mediated local tissue healing and recovery. 

Abstract Corticosteroid-eluting endotracheal tubes (ETTs) were developed and employed in a swine laryngotracheal injury model to maintain airway patency and provide localized drug delivery to inhibit fibrotic scarring. Polycaprolactone (PCL) fibers with or without dexamethasone were electrospun onto the ETT surface PCL-only coated ETTs and placed in native airways of 18 Yorkshire swine. Regular and dexamethasone-PCL coated ETTs were placed in airways of another 18 swine injured by inner laryngeal mucosal abrasion. All groups were evaluated after 3, 7 and 14 days (n = 3/treatment/time). Larynges were bisected and localized stiffness determined by normal indentation, then sequentially matched with histological assessment. In the native airway, tissue stiffness with PCL-only ETT placement increased significantly from 3 to 7 days (p = 0.0016) and 3 to 14 days (p < 0.0001) while dexamethasone-PCL ETT placement resulted in stiffness decreasing from 7 to 14 days (p = 0.031). In the injured airway, localized stiffness at 14 days was significantly greater after regular ETT placement (23.1 ± 0.725 N/m) versus dexamethasone-PCL ETTs (17.10 ± 0.930 N/m,p < 0.0001). Dexamethasone-loaded ETTs were found to reduce laryngotracheal tissue stiffening after simulated intubation injury compared to regular ETTs, supported by a trend of reduced collagen in the basement membrane in injured swine over time. Findings suggest localized corticosteroid delivery allows for tissue stiffness control and potential use as an approach for prevention and treatment of scarring caused by intubation injury. 

Abstract Beta-tricalcium phosphate (β-TCP)-based bioinks were developed to support direct-ink 3D printing-based manufacturing of macroporous scaffolds. Binding of the gelatin:β-TCP ink compositions was optimized by adding carboxymethylcellulose (CMC) to maximize the β-TCP content while maintaining printability. Post-sintering, the gelatin:β-TCP:CMC inks resulted in uniform grain size, uniform shrinkage of the printed structure, and included microporosity within the ceramic. The mechanical properties of the inks improved with increasing β-TCP content. The gelatin:β-TCP:CMC ink (25:75 gelatin:β-TCP and 3% CMC) optimized for mechanical strength was used to 3D print several architectures of macroporous scaffolds by varying the print nozzle tip diameter and pore spacing during the 3D printing process (compressive strength of 13.1 ± 2.51 MPa and elastic modulus of 696 ± 108 MPa was achieved). The sintered, macroporous β-TCP scaffolds demonstrated both high porosity and pore size but retained mechanical strength and stiffness compared to macroporous, calcium phosphate ceramic scaffolds manufactured using alternative methods. The high interconnected porosity (45–60%) and fluid conductance (between 1.04 ×10 −9 and 2.27 × 10 −9  m 4 s/kg) of the β-TCP scaffolds tested, and the ability to finely tune the architecture using 3D printing, resulted in the development of novel bioink formulations and made available a versatile manufacturing process with broad applicability in producing substrates suitable for biomedical applications. 

Electrical stimulus-responsive drug delivery from conducting polymers such as polypyrrole (PPy) has been limited by lack of versatile polymerization techniques and limitations in drug-loading strategies. In the present study, we report an in-situ chemical polymerization technique for incorporation of biotin, as the doping agent, to establish electrosensitive drug release from PPy-coated substrates. Aligned electrospun polyvinylidene fluoride (PVDF) fibers were used as a substrate for the PPy-coating and basic fibroblast growth factor and nerve growth factor were the model growth factors demonstrated for potential applications in musculoskeletal tissue regeneration. It was observed that 18-h of continuous polymerization produced an optimal coating of PPy on the surface of the PVDF electrospun fibers with significantly increased hydrophilicity and no substantial changes observed in fiber orientation or individual fiber thickness. This PPy-PVDF system was used as the platform for loading the aforementioned growth factors, using streptavidin as the drug-complex carrier. The release profile of incorporated biotinylated growth factors exhibited electrosensitive release behavior while the PPy-PVDF complex proved stable for a period of 14 days and suitable as a stimulus responsive drug delivery depot. Critically, the growth factors retained bioactivity after release. In conclusion, the present study established a systematic methodology to prepare PPy coated systems with electrosensitive drug release capabilities which can potentially be used to encourage targeted tissue regeneration and other biomedical applications. 

Objectives/Hypothesis Novel laryngotracheal wound coverage devices are limited by complex anatomy, smooth surfaces, and dynamic pressure changes and airflow during breathing. We hypothesize that a bioinspired mucoadhesive patch mimicking how geckos climb smooth surfaces will permit sutureless wound coverage and also allow drug delivery. Study Design ex‐vivo. Methods Polycaprolactone (PCL) fibers were electrospun onto a substrate and polyethylene glycol (PEG) – acrylate flocks in varying densities were deposited to create a composite patch. Sample topography was assessed with laser profilometry, material stiffness with biaxial mechanical testing, and mucoadhesive testing determined cohesive material failure on porcine tracheal tissue. Degradation rate was measured over 21 days in vitro along with dexamethasone drug release profiles. Material handleability was evaluated via suture retention and in cadaveric larynges. Results Increased flocking density was inversely related to cohesive failure in mucoadhesive testing, with a flocking density of PCL‐PEG‐2XFLK increasing failure strength to 6880 ± 1810 Pa compared to 3028 ± 791 in PCL‐PEG‐4XFLK density and 1182 ± 262 in PCL‐PEG‐6XFLK density. The PCL‐PEG‐2XFLK specimens had a higher failure strength than PCL alone (1404 ± 545 Pa) or PCL‐PEG (2732 ± 840). Flocking progressively reduced composite stiffness from 1347 ± 15 to 763 ± 21 N/m. Degradation increased from 12% at 7 days to 16% after 10 days and 20% after 21 days. Cumulative dexamethasone release at 0.4 mg/cm2 concentration was maintained over 21 days. Optimized PCL‐PEG‐2XFLK density flocked patches were easy to maneuver endoscopically in laryngeal evaluation. Conclusions This novel, sutureless, patch is a mucoadhesive platform suitable to laryngeal and tracheal anatomy with drug delivery capability. 

Abstract Skeletal muscle is a tissue that is directly involved in the progression and persistence of type 2 diabetes (T2D), a disease that is becoming increasingly common. Gaining better insight into the mechanisms that are affecting skeletal muscle dysfunction in the context of T2D has the potential to lead to novel treatments for a large number of patients. Through its ability to emulate skeletal muscle architecture while also incorporating aspects of disease, tissue-engineered skeletal muscle (TE-SkM) has the potential to provide a means for rapid high-throughput discovery of therapies to treat skeletal muscle dysfunction, to include that which occurs with T2D. Muscle precursor cells isolated from lean or obese male Zucker diabetic fatty rats were used to generate TE-SkM constructs. Some constructs were treated with adipogenic induction media to accentuate the presence of adipocytes that is a characteristic feature of T2D skeletal muscle. The maturity (compaction and creatine kinase activity), mechanical integrity (Young's modulus), organization (myotube orientation), and metabolic capacity (insulin-stimulated glucose uptake) were all reduced by diabetes. Treating constructs with adipogenic induction media increased the quantity of lipid within the diabetic TE-SkM constructs, and caused changes in construct compaction, cell orientation, and insulin-stimulated glucose uptake in both lean and diabetic samples. Collectively, the findings herein suggest that the recapitulation of structural and metabolic aspects of T2D can be accomplished by engineering skeletal muscle in vitro. Impact Statement The tissue engineering of skeletal muscle to model disease and injury has great promise to provide a tool to develop and/or improve therapeutic approaches for improved health care. A tissue-engineered skeletal muscle model of one of the most common and debilitating diseases, type 2 diabetes, has been developed in vitro as evidenced by the structural and metabolic alterations that are consistent with the disease phenotype in vivo.