Low-Dose Gamma Radiation Sterilization for Decellularized Tracheal Grafts.

One of the main key aspects in ensuring that a transplant evolves correctly is the sterility of the medium. Decellularized tracheal transplantation involves implanting an organ that was originally in contact with the environment, thus not being sterile from the outset. While the decellularization protocol (through detergent exposition [2% sodium dodecyl sulfate], continuous stirring, and osmotic shocks) is conducted in line with aseptic measures, it does not provide sterilization. Therefore, one of the main challenges is ensuring sterility prior to in vivo implantation. Although there are established gamma radiation sterilization protocols for inorganic materials, there are no such measures for organic materials. Additionally, the protocols in place for inorganic materials cannot be applied to organic materials, as the established radiation dose (25 kGy) would completely destroy the implant. This paper studies the effect of an escalated radiation dose in a decellularized rabbit trachea. We maintained the dose range (kGy) and tested escalated doses until finding the minimal dose at which sterilization is achieved. After determining the dose, we studied effects of it on the organ, both histologically and biomechanically. We determined that while 0.5 kGy did not achieve sterility, doses of both 1 kGy and 2 kGy did, with 1 kGy, therefore, being the minimal dose necessary to achieve sterilization. Microscopic studies showed no relevant changes compared to non-sterilized organs. Axial biomechanical characteristics were not altered at all, and only a slight reduction in the force per unit of length that the organ can radially tolerate was observed. We can therefore conclude that 1 kGy achieves complete sterilization of decellularized rabbit trachea with a minimal, if any, effects on the organ.

A Standardised Approach to the Biomechanical Evaluation of Tracheal Grafts.

The ideal tracheal substitute must have biomechanical properties comparable to the native trachea, but currently there is no standardised approach to evaluating these properties. Here we propose a novel method for evaluating and comparing the properties of tracheal substitutes, thus systematising both measurement and data curation. This system was tested by comparing native rabbit tracheas to frozen and decellularised specimens and determining the histological characteristics of those specimens. We performed radial compression tests on the anteroposterior tracheal axis and longitudinal axial tensile tests with the specimens anastomosed to the jaw connected to a measuring system. All calculations and results were adjusted according to tracheal size, always using variables relative to the tracheal dimensions, thus permitting comparison of different sized organs. The biomechanical properties of the decellularised specimens were only slightly reduced compared to controls and significant in regard to the maximum stress withstood in the longitudinal axis (-0.246 MPa CI [-0.248, -0.145] MPa) and the energy stored per volume unit (-0.124 mJ·mm-3 CI [-0.195, -0.055] mJ·mm-3). The proposed method is suitable for the systematic characterisation of the biomechanical properties of different tracheal substitutes, regardless of the size or nature of the substitute, thus allowing for direct comparisons.

Decellularized tracheal prelamination implant: A proposed bilateral double organ technique.

In tracheal replacement transplantation, prelamination is a critical stage. Nowadays, the most widely used prelamination technique is the prethoracic fascia flap with lateral thoracic artery. We propose a flap based on the internal thoracic artery, which allows a relatively non-aggressive double organ implant, and we have tested its efficacy in decellularized tracheas. Tracheas of albino New Zealand rabbits were decellularized following a protocol that uses detergents and cryogenization, sterilized with 1kGy gamma radiation, and tutorized with a stent. Bilateral pedicled flaps made of pectoral fascia and a muscular component were harvested through a longitudinal 3-cm central thoracic incision, wrapping the tracheas with them in 16 rabbits, remaining them implanted for 2, 4, 8, and 12 weeks. The tracheas were then studied histologically using standard stainings plus immunohistochemistry (CD31). The models were adjusted with Bayesian statistics using ordinal regression; results as odds ratios and credibility intervals. All analysis were performed using R software. Acute inflammatory cell invasion was observed at 2 weeks, which almost disappeared at week 8 after implant. Only macrophages and giant cells increased between Weeks 8 and 12 (OR 10.487, CI [1.603-97.327]). The cartilage maintained its structure, with slight signs of ischemia in a few cases. New CD31-positive vessels were observed from Week 2 and increasing thereafter, reaching a maximum peak at Week 8. We propose a bilateral implant technique that is viable and effective as a prelamination option for two concurrent tracheas, achieving perfect vascularization and integration of the organ with hardly any inflammatory response in the medium or long term.