Tracheal Replacement Therapy with a Stem Cell-Seeded Graft: Lessons from Compassionate Use Application of a GMP-Compliant Tissue-Engineered Medicine.

Tracheal replacement for the treatment of end-stage airway disease remains an elusive goal. The use of tissue-engineered tracheae in compassionate use cases suggests that such an approach is a viable option. Here, a stem cell-seeded, decellularized tissue-engineered tracheal graft was used on a compassionate basis for a girl with critical tracheal stenosis after conventional reconstructive techniques failed. The graft represents the first cell-seeded tracheal graft manufactured to full good manufacturing practice (GMP) standards. We report important preclinical and clinical data from the case, which ended in the death of the recipient. Early results were encouraging, but an acute event, hypothesized to be an intrathoracic bleed, caused sudden airway obstruction 3 weeks post-transplantation, resulting in her death. We detail the clinical events and identify areas of priority to improve future grafts. In particular, we advocate the use of stents during the first few months post-implantation. The negative outcome of this case highlights the inherent difficulties in clinical translation where preclinical in vivo models cannot replicate complex clinical scenarios that are encountered. The practical difficulties in delivering GMP grafts underscore the need to refine protocols for phase I clinical trials. Stem Cells Translational Medicine 2017;6:1458-1464.

How biophysical in vivo testing techniques can be used to characterize full thickness skin equivalents.

BACKGROUND: The reliability of the biophysical properties of skin equivalents (SEs) remains a challenge for medical applications and for product efficacy tests following the European Directive 2003/15/EC2 on the prohibition of animal experiments for cosmetic products. METHODS: We propose to adapt the biophysical in vivo testing techniques to compare full thickness model growth vs. time. The interest in using such techniques lies in possible comparisons between in vivo and in vitro skin as well as monitoring samples over the culture time. RESULTS: High frequency ultrasound technique, optical coherence tomography (OCT), and laser scanning microscopy were used to analyze SEs morphology at days D42 and D60 whereas their microstructure was assessed through transmission electron microscopy and classical histology. A correlation between these observations and mechanical measurements has been proposed so as to underline the consequence of both the development of the dermis elastic fibers and the epidermis differentiation. CONCLUSION: Ultrasounds measurements show a highly homogeneous dermis whereas the OCT technique clearly distinguishes the stratum corneum and the living epidermis. The increase in the thicknesses of these layers as well as the growth in elastin and collagen fibers results in strong modifications of the samples mechanical properties.

Periprosthetic Tissue Loss in Patients With Idiopathic Vitreous Inflammation After the Boston Keratoprosthesis.

PURPOSE: Idiopathic vitritis is a poorly understood complication after Boston keratoprosthesis surgery with unclear etiology. We sought to determine whether an association exists between periprosthetic corneal tissue loss and the development of idiopathic vitritis in keratoprosthesis recipients. METHODS: Thirteen Boston type I keratoprosthesis recipient eyes with a history of idiopathic vitritis and 34 type I keratoprosthesis recipient eyes with no history of idiopathic vitritis underwent anterior segment optical coherence tomography (AS-OCT) at a median time postoperatively of 2.4 years versus 1.9 years (range, 0.5-14.2 vs. 0.1-13.6 years), respectively. Areas of corneal graft tissue loss ("gaps") around the keratoprosthesis stem were identified and analyzed by 2 masked observers. The difference in the presence, number, and size of gaps was compared between cases and controls. RESULTS: A periprosthetic gap was identified more commonly in idiopathic vitritis cases than in controls on AS-OCT (11/13, 86% vs. 11/34, 33.3%, P < 0.001). The number of gaps between cases and controls was also significantly different (2.6 ± 1.6 vs. 0.5 ± 0.8, P < 0.001), but not the estimated gap area (0.056 ± 0.049 mm² vs. 0.039 ± 0.025 mm², P = 0.22). CONCLUSIONS: A significantly higher proportion of keratoprosthesis recipient eyes with idiopathic vitritis had corneal tissue loss around the keratoprosthesis stem than did controls. Tissue loss could serve as an entry point for debris or bacterial components, triggering idiopathic vitritis. Our study underscores the utility of AS-OCT imaging in the postoperative management of keratoprosthesis patients.

Guided tissue regeneration: porcine matrix does not transmit PERV.

OBJECTIVE: For cardiovascular tissue engineering, acellularized scaffolds of porcine matrices have been successfully used. However, the possibility of porcine endogenous retrovirus (PERV) transmission remains debatable. In this study, we investigated whether acellularized porcine vascular scaffolds cause cross-species transmission of PERV in a xenogenic model. METHODS: Porcine pulmonary arteries were acellularized and implanted into sheep in orthotopic position (n=6). Cardiopulmonary bypass support was used for all operations. Blood samples were collected regularly up to 6 months after the operation, and cellular components were tested for PERV infection by PCR and RT-PCR. Grafts were explanted 6 and 12 months after implantation. Tissue samples were characterized by histology and electron microscopy and tested for PERV sequences. RESULTS: All animals survived the procedure and follow up until explantation of the grafts. PERV DNA was detectable in acellularized scaffolds of porcine matrices. Acellular porcine pulmonary arteries scaffolds were repopulated in vivo by autologous cells of the host, leading to a vessel consisting of all cellular components of the vessel wall. No PERV sequences were detectable neither in all tested peripheral blood samples nor in tissue samples of in vivo recellularized grafts up to 6 months after implantation. Electron microscopy revealed no signs of graft infection by retrovirus. CONCLUSIONS: Guided tissue regeneration of acellularized vascular porcine matrix scaffolds leads to structured vessels up to one year without risk of PERV transmisson.

Development of an ambulatory artificial lung in an ovine survival model.

We are developing an artificial lung (AL) as an eventual bridge to lung transplant or recovery. The device is rigidly housed, noncompliant, and has a very low resistance to blood flow. In eight sheep, arterial cannulae were anastomosed end-to-side to the proximal and distal main pulmonary artery, and attached to the AL. A pulmonary artery snare between anastomoses diverted full pulmonary blood flow through the AL. Eight of eight sheep survived the preparation. Mean pressure gradient across the AL was 8 mm Hg (3 Wood units; 8 mm Hg/2.8 L/min). Four of eight sheep tolerated immediate full diversion of blood flow and died at 24 and 40 hours (exsanguination) or 168 and 168 hours (elective sacrifice). Four of eight sheep were intolerant of full flow: two died of right heart failure at <8 hours with full flow through the device (full snare); the other two survived with partial device flow (partial snare), but the device clotted. These two then underwent successful closed-chest cannula thrombectomy and device change-out at 53 and 75 hours, and subsequently tolerated full flow. Long-term (up to 7 day) survival with complete diversion of pulmonary blood flow through a non-compliant, low-resistance AL is possible. Initial right heart failure in this model was 50% (4 of 8).