Pilot Testing of a Lightweight, Pulmonary Assist System in an Ambulatory Sheep Model of Destination Therapy Respiratory Support.

A new, lightweight (2.3 kg), ambulatory pulmonary assist system (PAS) underwent preliminary evaluation in ambulatory sheep. The PAS was purposefully designed for long-term extracorporeal respiratory support for chronic lung disease and utilizes a novel, small (0.9 m 2 surface area) gas exchanger, the pulmonary assist device, with a modified Heart Assist 5 pump fitting in a small wearable pack. Prototype PAS were attached to two sheep in venovenous configuration for 7 and 14 days, evaluating ability to remain thrombus free; maintain gas exchange and blood flow resistance; avoid biocompatibility-related complications while allowing safe ambulation. The PAS achieved 1.56 L/min of flow at 10.8 kRPM with a 24 Fr cannula in sheep one and 2.0 L/min at 10.5 kRPM with a 28 Fr cannula in sheep 2 without significant change. Both sheep walked freely, demonstrating the first application of truly ambulatory ECMO in sheep. While in vitro testing evaluated PAS oxygen transfer rates of 104.6 ml/min at 2 L/min blood flow, oxygen transfer rates averaged 60.6 ml/min and 70.6 ml/min in studies 1 and 2, due to average hemoglobin concentrations lower than humans (8.9 and 10.5 g/dl, respectively). The presented cases support uncomplicated ambulation using the PAS.

Generation and morphological quantification of large scale, three-dimensional, self-assembled vascular networks.

One of the largest issues facing the field of tissue engineering is scaling due to tissue necrosis as a result of a lack of vascularization. We have developed an accessible method for generating large scale vascular networks of arbitrary geometries through the self-assembly of endothelial cells in a collagen gel, similar to vasculogenesis that occurs in the developing embryo. This system can be applied to a wide range of collagen concentrations and seeding densities, resulting in networks of varying phenotypes, lending itself to the recapitulation of vascular networks that mimic those found across different tissues. Methods are thus described for the generation and imaging of these self-assembled three-dimensional networks in addition to image processing methods for rigorous quantitative measurement of various morphological parameters. There are several advantages to the system described herein. •Varied molding procedures allow for irregular geometries, similar to those that would be required for tissue grafts.•Robust network formation translates into centimeter scale constructs.•Whereas similar processes suffer from a high degree of variability and inconsistent characterization, our method employs image analysis techniques to stringently characterize each network based on several objective characteristics.

De novo lung biofabrication: clinical need, construction methods, and design strategy.

Chronic lung disease is the 4th leading cause of death in the United States. Due to a shortage of donor lungs, alternative approaches to support failing, native lungs have been attempted, including mechanical ventilation and various forms of artificial lungs. However, each of these support methods causes significant complications when used for longer than a few days and are thus not capable of long-term support. For artificial lungs, complications arise due to interactions between the artificial materials of the device and the blood of the recipient. A potential new approach is the fabrication of lungs from biological materials, such that the gas exchange membranes provide a more biomimetic blood-contacting interface. Recent advancements with three-dimensional, soft-tissue biofabrication methods and the engineering of thin, basement membranes demonstrate the potential of fabricating a lung scaffold from extracellular matrix materials. This scaffold could then be seeded with endothelial and epithelial cells, matured within a bioreactor, and transplanted. In theory, this fully biological lung could provide improved, long-term biocompatibility relative to artificial lungs, but significant work is needed to perfect the organ design and construction methods. Like artificial lungs, biofabricated lungs do not need to follow the shape and structure of a native lung, allowing for simpler manufacture. However, various functional requirements must still be met, including stable, efficient gas exchange for a period of years. Design decisions depend on the disease state, how the organ is implanted, and the latest biofabrication methods available in a rapidly evolving field.

Fabrication of centimeter-scale and geometrically arbitrary vascular networks using in vitro self-assembly.

One of the largest challenges facing the field of tissue engineering is the incorporation of a functional vasculature, allowing effective nourishment of graft tissue beyond diffusion length scales. Here, we demonstrate a methodology for inducing the robust self-assembly of endothelial cells into stable three-dimensional perfusable networks on millimeter and centimeter length scales. Utilizing broadly accessible cell strains and reagents, we have rigorously tested a state space of cell densities (0.5-2.0 × 106 cell/mL) and collagen gel densities (2-6 mg/mL) that result in robust vascular network formation. Further, over the range of culture conditions with which we observed robust network formation, we advanced image processing algorithms and quantitative metrics to assess network connectivity, coverage, tortuosity, lumenization, and vessel diameter. These data demonstrate that decreasing collagen density produced more connected networks with higher coverage. Finally, we demonstrated that this methodology results in the formation of perfusable networks, is extensible to arbitrary geometries and centimeter scales, and results in networks that remain stable for 21 days without the need for the co-culture of supporting cells. Given the robustness and accessibility, this system is ideal for studies of tissue-scale biology, as well as future studies on the formation and remodeling of larger engineered graft tissues.