Tracheal Replacement Research Articles

Establishing Benchmarks for Airway Replacement: Long-Term Outcomes of Tracheal Autografts in a Large Animal Model.

Airway replacement is a challenging surgical intervention and remains an unmet clinical need. Due to the risk of airway stenosis, anastomotic separation, poor vascularization, and necrosis, it is necessary to establish the gold-standard outcomes of tracheal replacement. In this study, we use a large animal autograft model to assess long-term outcomes following tracheal replacement. Four New Zealand White rabbits underwent tracheal autograft surgery and were observed for 6 months. Clinical and radiographic surveillance were recorded, and grafts were analyzed histologically and radiographically at endpoint. All animals survived to the endpoint with minimal respiratory symptoms and normal growth rates. No complications were observed. Computed tomography scans of the post-surgical airway demonstrated graft patency at all time points. Histological sections showed no sign of stenosis or necrosis with preservation of the native structure of the trachea. We established benchmarks for airway replacement. Our findings suggest that a rabbit model of tracheal autograft with direct reimplantation is feasible and does not result in graft stenosis or airway collapse.

Microvascular Regeneration and Perfusion of Partially Decellularized Tracheal Grafts.

A critical barrier to successful tracheal transplantation is poor vascularization. Despite its importance, little is known about microvascular regeneration in tissue-engineered grafts. We have demonstrated that partially decellularized tracheal grafts (PDTG) support neotissue formation including new submucosal microvasculature (CD31+). However, the perfusion of this neovasculature is unknown. In this study, we used a mouse model of tracheal replacement to measure the microvascular regeneration and perfusion of PDTG. PDTG and syngeneic tracheal grafts (STG, surgical control) (n = 5 for each group) were orthotopically transplanted into C5BL/6 J mice. We quantified vascularity of STG and PDTG samples at 1 and 3 months with conventional histology (N = 3 ~ 10/group). At 1, 3, and 6 months, animals were injected with fluorescein isothiocyanate (FITC) tomato lectin into the left ventricle. After perfusion, tracheas were fixed, harvested, mounted, stained for CD31 expression, and imaged with resonant scanning confocal microscopy. Percent CD31+, FITC area was compared between groups and endpoints compared with native trachea. Microvascular intersections were quantified using Sholl analysis. Functional microvasculature was seen in both groups. Although percent vascularization (CD31) in PDTG was restored by 3 months, microvascular pattern in PDTG displayed a unique morphology compared with control. Surgery alone appeared to globally change microvascular pattern and perfusion. PDTG demonstrated equivalent perfusion to surgical control by 6 months. Sholl analysis revealed a reduction of microvessel intersectionality that persisted in PDTG and was not seen in surgical or native controls. PDTG exhibited microvascular regeneration. Perfusion was present in PDTG, improved, and persisted over long-term time points. NA Laryngoscope, 2024.

Tracheal Replacement: A Scoping Review

Abstract Objective To summarize patient characteristics and outcomes for the historical and current methods of long-segment tracheal replacement in humans. Materials and Methods A single reviewer screened the abstracts and full texts using Covidence for file management. Studies published in English that reported human subjects with circumferential or near-circumferential (>270 degrees) cervical tracheal replacements were included. Articles with subjects treated with primary anastomosis alone, retracted articles, abstracts, expert opinion articles, and conference presentations were excluded. Results A total of 32 articles were included in the review reporting 156 cases of long-segment tracheal replacement including synthetic (alive at 1–8 years n = 6/64), regenerative medicine (dead at 15 days–55 months n = 4, not reported n = 6), cadaveric tracheal allograft (alive at 5 months–10 years n = 32/38), aortic allograft (alive at 6–85 months n = 12/16), free tissue transfer (alive at 6–108 months n = 13/21), allotransplantation (alive at 6–24 months n = 5/8), and vascular composite allograft (VCA) (alive at 20 months n = 1/1). Conclusion Silicone and Marlex prostheses have poor long-term outcomes. The cadaveric tracheal allograft can only replace near-circumferential tracheal defects and is therefore limited to benign tracheal pathology. Inadequate structural support plagues the aortic allograft and often requires numerous invasive procedures and maintenance of an intraluminal stent. A lack of mucociliary clearance exists in all methods of tracheal replacement except cadaveric tracheal allograft and VCA and can cause fatal mucous plugging and chronic pulmonary infections. VCA and allotransplantation require long-term immunomodulation therapy.

A Clinical-Grade Partially Decellularized Matrix for Tracheal Replacement: Validation In Vitro and In Vivo in a Porcine Model.

The management of extensive tracheal resection followed by circumferential replacement remains a surgical challenge. Numerous techniques are proposed with mixed results. Partial decellularization of the trachea with the removal of the mucosal and submucosal cells is a promising method, reducing immunogenicity while preserving the biomechanical properties of the final matrix. Despite many research protocols and proofs of concept, no standardized clinical grade protocol is described. Furthermore, local and systemic biointegration mechanisms of decellularized trachea are not well known. Therefore, in a translational research perspective, this work set up a partial tracheal decellularization protocol in line with Cell and Tissue Products regulations. Extensive characterization of the final product is performed in vitro and in vivo. The results show that the Partially Decellularized Trachea (PDT) is cell-free in the mucosa and submucosa, while the cartilage structure is preserved, maintaining the biomechanical properties of the trachea. When implanted in the muscle in vivo for 28 days, no systemic inflammation is observed, and locally, the PDT shows an excellent biointegration and vascularization. No signs of graft rejection are observed. These encouraging results confirmed the efficacy of the clinical grade PDT production protocol, which is an important step for future clinical applications.

Cartilage regeneration after tracheal replacement with aortic allograft: myth or fact?

Cartilage regeneration after tracheal replacement with aortic allograft: myth or fact?

Feasibility of airway replacement using stented aortic matrices: Bench to TRITON‐01 study

AbstractPatients with long‐segment tracheal defects, deemed technically inoperable, constitute a population in critical need of airway replacement. Regardless of the underlying cause—be it benign or malignant processes—this patient category requires either a tracheal transplant or admission to a palliative care facility. Despite over 50 years of exploration in thoracic surgery and regenerative medicine, airway transplantation remains a significant challenge. Various tracheobronchial substitutes, such as synthetic prostheses, bioprostheses, allografts, autografts, and bioengineered conduits, have been experimentally tested, yet none have provided a standardized method for airway replacement. Aortic grafts were suggested by Aortic grafts have been suggested as a biological matrix for extensive airway reconstruction as a biological matrix for extensive airway reconstruction. Cryopreserved aortic allografts, because of their availability in tissue banks and the lack of need for immunosuppressive therapy, were first used in clinical applications. The TRITON‐01 study (NCT04263129) focuses on patients who received airway replacement to determine the routine applicability of this novel approach. The primary objective of this review is to provide information on advancements in the use of aortic allografts as tracheal replacements.

Full-circumferential tracheal replacement with cartilage-reinforced forearm free flaps in the real world.

Full-circumferential tracheal replacement with cartilage-reinforced forearm free flaps in the real world.

Confirmation of de novo cartilage generation on aortic matrices after tracheal replacement.

Confirmation of de novo cartilage generation on aortic matrices after tracheal replacement.

Feasibility of tracheal reconstruction using silicone-stented aortic allografts.

Tracheal reconstruction post-extensive resection remains an unresolved challenge in thoracic surgery. This study evaluates the use of aortic allografts (AAs) for tracheal replacement and reconstruction in a rat model, aiming to elucidate the underlying mechanisms of tracheal regeneration. AAs from female rats were employed for tracheal reconstruction in 36 male rats, with the replacement exceeding half of the tracheal length. To avert collapse, silicone stents were inserted into the AA lumens. No immunosuppressive therapy was administered. The rats were euthanized biweekly, and the AAs were examined for neovascularization, cartilage formation, respiratory epithelial ingrowth, submucosal gland regeneration and the presence of the Sex-determining region of Y-chromosome (SRY) gene. All procedures were successfully completed without severe complications. The AA segments were effectively integrated into the tracheal framework, with seamless distinction at suture lines. Histological analysis indicated an initial inflammatory response, followed by the development of squamous and mucociliary epithelia, new cartilage ring formation and gland regeneration. In situ hybridization identified the presence of the SRY gene in newly formed cartilage rings, confirming that regeneration was driven by recipient cells. This study demonstrates the feasibility of AAs transforming into functional tracheal conduits, replicating the main structural and functional characteristics of the native trachea. The findings indicate that this approach offers a novel pathway for tissue regeneration and holds potential for treating extensive tracheal injuries.

Regeneration Process of an Autologous Tissue-Engineered Trachea (aTET) in a Rat Patch Tracheoplasty Model.

The treatment of long-tracheal lesion is difficult because there are currently no viable grafts for tracheal replacement. To solve this problem, we have developed an autologous Tissue-Engineered Trachea (aTET), which is made up of collagenous tissues and cartilage-like structures derived from rat chondrocytes. This graft induced successful long-term survival in a small-animal experiment in our previous study. In this study, we investigated the regeneration process of an aTET to attain reproducible success. We prepared an aTET by using a specially designed mold and performed patch tracheoplasty with an aTET. We assigned twenty-seven rats to three groups according to the three types of patch grafts used: aTET patches (the aTET group), fresh tracheal autograft patches (the Ag group), or polylactic acid and polycaprolactone copolymer sheets (the PPc group). In each group, gross and histological evaluations were performed at 1 month (n = 3), 3 months (n = 3), and 6 months (n = 3) after implantation. We obtained high survival rates in all groups, but only the PPc group attained thick tracheal walls with granular tissues and no tracheal regeneration. On the other hand, the aTET and Ag groups reproducibly achieved complete tracheal regeneration in 6 months. So, an aTET could be a promising candidate for tracheal regeneration grafts.

Extended autologous tracheal replacement by a novel pedicled thoraco-chondro-costal flap: a cadaveric proof of concept.

Our aim was to report an anatomic model of an autologous flap based on the internal thoracic blood supply: the pedicled thoraco-chondro-costal flap; and establish the feasibility of various types of extended tracheal replacement with this novel flap, according to a newly proposed topographic classification. In a cadaveric model, a cervicotomy combined with median sternotomy was performed. The incision was extended laterally to expose the chest wall. The internal thoracic pedicle was freed from its origin down to the upper limit of the delineated flap to be elevated. The perichondria and adjacent periostea were incised longitudinally to remove cartilages and adjacent rib segments, preserving perichondria and periostea. A full-thickness quadrangular chest wall flap pedicled on internal thoracic vessels was then elevated and shaped into a neo conduit to replace the trachea with the pleura as an inner lining. Various types of extended non-circumferential and full-circumferential tracheal replacements were achieved with this composite flap. No anastomosis tension was noticed despite the absence of release manoeuvres. This model could represent a suitable autologous tracheal substitute, which is long, longitudinally flexible and eventually transversely rigid. No microsurgical vascular anastomoses are required. The technique is reproducible. The perichondria and periostea would regenerate vascularized neo-cartilaginous rings, potentially decreasing the need for long-term stenting. The inner pleural lining could potentially transform into ciliated epithelium as shown in previous preclinical studies.

Tracheal Tissue Engineering: Principles and State of the Art.

Patients affected by long-segment tracheal defects or stenoses represent an unsolved surgical issue, since they cannot be treated with the conventional surgery of tracheal resection and consequent anastomosis. Hence, different strategies for tracheal replacement have been proposed (synthetic materials, aortic allografts, transplantation, autologous tissue composites, and tissue engineering), each with advantages and drawbacks. Tracheal tissue engineering, on the other hand, aims at recreating a fully functional tracheal substitute, without the need for the patient to receive lifelong immunosuppression or endotracheal stents. Tissue engineering approaches involve the use of a scaffold, stem cells, and humoral signals. This paper reviews the main aspects of tracheal TE, starting from the choice of the scaffold to the type of stem cells that can be used to seed the scaffold, the methods for their culture and expansion, the issue of graft revascularization at the moment of in vivo implantation, and experimental models of tracheal research. Moreover, a critical insight on the state of the art of tracheal tissue engineering is also presented.

The effect of bioactivity of airway epithelial cells using methacrylated gelatin scaffold loaded with exosomes derived from bone marrow mesenchymal stem cells.

The current evidence provides support for the involvement of bone marrow mesenchymal stem cells (BMSCs) in the regulation of airway epithelial cells. However, a comprehensive understanding of the underlying biological mechanisms remains elusive. This study aimed to isolate and characterize BMSC-derived exosomes (BMSC-Exos) and epithelial cells (ECs) through primary culture. Subsequently, the impact of BMSC-Exos on ECs was assessed in vitro, and sequencing analysis was conducted to identify potential molecular mechanisms involved in these interactions. Finally, the efficacy of BMSC-Exos was evaluated in animal models in vivo. In this study, primary BMSCs and ECs were efficiently isolated and cultured, and high-purity Exos were obtained. Upon uptake of BMSC-Exos, ECs exhibited enhanced proliferation (p < .05), while migration showed no difference (p > .05). Notably, invasion demonstrated significant difference (p < .05). Sequencing analysis suggested that miR-21-5p may be the key molecule responsible for the effects of BMSC-Exos, potentially mediated through the MAPK or PI3k-Akt signaling pathway. The in vivo experiments showed that the presence of methacrylated gelatin (GelMA) loaded with BMSC-Exos in composite scaffold significantly enhanced epithelial crawling in the patches in comparison to the pure decellularized group. In conclusion, this scheme provides a solid theoretical foundation and novel insights for the research and clinical application of tracheal replacement in the field of tissue engineering.

Preparation of Decellularized Tissue as Dual Cell Carrier Systems: A Step Towards Facilitating Re-epithelization and Cell Encapsulation for Tracheal Reconstruction.

Surgical treatment of tracheal diseases, trauma, and congenital stenosis has shown success through tracheal reconstruction coupled with palliative care. However, challenges in surgical-based tracheal repairs have prompted the exploration of alternative approaches for tracheal replacement. Tissue-based treatments, involving the cultivation of patient cells on a network of extracellular matrix (ECM) from donor tissue, hold promise for restoring tracheal structure and function without eliciting an immune reaction. In this study, we utilized decellularized canine tracheas as tissue models to develop two types of cell carriers: a decellularized scaffold and a hydrogel. Our hypothesis posits that both carriers, containing essential biochemical niches provided by ECM components, facilitate cell attachment without inducing cytotoxicity. Canine tracheas underwent vacuum-assisted decellularization (VAD), and the ECM-rich hydrogel was prepared through peptic digestion of the decellularized trachea. The decellularized canine trachea exhibited a significant reduction in DNA content and major histocompatibility complex class II, while preserving crucial ECM components such as collagen, glycosaminoglycan, laminin, and fibronectin. Scanning electron microscope and fluorescent microscope images revealed a fibrous ECM network on the luminal side of the cell-free trachea, supporting epithelial cell attachment. Moreover, the ECM-rich hydrogel exhibited excellent viability for human mesenchymal stem cells encapsulated for 3 days, indicating the potential of cell-laden hydrogel in promoting the development of cartilage rings of the trachea. This study underscores the versatility of the trachea in producing two distinct cell carriers-decellularized scaffold and hydrogel-both containing the native biochemical niche essential for tracheal tissue engineering applications.

The Efficacy of Daily Administration of Nebulized Heparin on the Prevention of Endotracheal Tube Blockage in Patients With Pneumonia.

Nosocomial infections pose a significant public health concern, impacting over 100 million people worldwide annually. Within this research, we investigated heparin nebulization through the endotracheal tube and its effect on preventing blockage due to clots and mucus plugs compared to normal saline. A double-blind clinical experiment was done on a cohort of 40 pneumonia patients who were intubated and hospitalized in theintensive care unit (ICU) at Imam Khomeini Hospital in Ahvaz, Iran. The individuals were randomly assigned to two groups of 20 patients using a random allocation technique. The initial cohort was administered 5000 IU of heparin diluted in 4 ccs of 0.9% normal saline every eight hours via a nebulizer through a tracheal tube. In contrast, the second cohort was given 5 ccs of normal saline as a nebulizer through a tracheal tube. The study compared the incidence of tracheal tube obstruction caused by mucus plug or clot, the occurrence of patient hypoxia resulting in emergency tracheal tube replacement, and the frequency of emergency tracheal tube suction due to partial obstruction caused by mucus plug in both the heparin and saline groups. According to our data, the number of patients in the heparin group who could avoid the need for emergency tracheal tube replacement owing to blockage was more significant than in the ordinary saline group (P=0.013). Heparin was significantly correlated with the number of times emergency suction was required to remove a tracheal tube occlusion (P=0.01). Heparin had no significant effect on coagulation factors (international normalized ratio [INR], platelet[PLT], and partial thromboplastin time [PTT]), Acute Physiology and Chronic Health Evaluation (APACHE) score, pneumonia severity index (PSI), saturation of patients, or tracheal tube secretions. There was no statistically significant difference in total time spent in the intensive care unit (P=0.91). Further studies are suggested to determine the effect of heparin nebulization on preventing endotracheal tube obstruction due to clots and mucus plugs in intubated ICU patients.

Synergistic Effects of SDS and H2O2 Combinations on Tracheal Scaffold Development: An In Vitro Study Using Goat Trachea.

Currently, a tissue-engineered trachea has been popularly used as a biological graft for tracheal replacement in severe respiratory diseases. In the development of tissue-engineered tracheal scaffolds, in vitro studies play a crucial role in allowing researchers to evaluate the efficacy and safety of scaffold designs and fabrication techniques before progressing to in vivo or clinical trials. This research involved the decellularization of goat trachea using SDS, H2O2, and their combinations. Various quantitative and qualitative assessments were performed, including histological analysis, immunohistochemistry, and biomechanical testing. Hematoxylin and eosin staining evaluated the cellular content, while safranin O-fast green and Masson's trichrome staining assessed glycosaminoglycan content and collagen distribution, respectively. The immunohistochemical analysis focused on detecting MHC-1 antigen presence. Tensile strength measurements were conducted to evaluate the biomechanical properties of the decellularized scaffolds. The results demonstrated that the combination of SDS and H2O2 for goat tracheal decellularization yielded scaffolds with minimal cellular remnants, low toxicity, preserved ECM, and high tensile strength and elasticity. This method holds promise for developing functional tracheal scaffolds to address severe respiratory diseases effectively.

Evaluation of a decellularized bronchial patch transplant in a porcine model

Biological scaffolds for airway reconstruction are an important clinical need and have been extensively investigated experimentally and clinically, but without uniform success. In this study, we evaluated the use of a decellularized bronchus graft for airway reconstruction. Decellularized left bronchi were procured from decellularized porcine lungs and utilized as grafts for airway patch transplantation. A tracheal window was created and the decellularized bronchus was transplanted into the defect in a porcine model. Animals were euthanized at 7 days, 1 month, and 2 months post-operatively. Histological analysis, immunohistochemistry, scanning electron microscopy, and strength tests were conducted in order to evaluate epithelialization, inflammation, and physical strength of the graft. All pigs recovered from general anesthesia and survived without airway obstruction until the planned euthanasia timepoint. Histological and electron microscopy analyses revealed that the decellularized bronchus graft was well integrated with native tissue and covered by an epithelial layer at 1 month. Immunostaining of the decellularized bronchus graft was positive for CD31 and no difference was observed with immune markers (CD3, CD11b, myeloperoxidase) at two months. Although not significant, tensile strength was decreased after one month, but recovered by two months. Decellularized bronchial grafts show promising results for airway patch reconstruction in a porcine model. Revascularization and re-epithelialization were observed and the immunological reaction was comparable with the autografts. This approach is clinically relevant and could potentially be utilized for future applications for tracheal replacement.

Reconstruction of tracheal window-shape defect by 3D printed polycaprolatone scaffold coated with Silk Fibroin Methacryloyl.

In this study, we aimed to utilize autologous tracheal epithelia and BMSCs as the seeding cells, utilize PCL coated with SilMA as the hybrid scaffold to carry the cells and KGN, which can selectively stimulate chondrogenic differentiation of BMSCs. This hybrid tracheal substitution was carried out to repair the tracheal partial window-shape defect. Firstly, SilMA with the concentration of 10%, 15% and 20% was prepared, and the experiment of swelling and degradation was performed. With the increase of the concentration, the swelling ratio of SilMA decreased, and the degradation progress slowed down. Upon the result of CCK-8 test and HE staining of 3D co-culture, the SilMA with concentration of 20% was selected. Next, SilMA and the cells attached to SilMA were characterized by SEM. Furthermore, in vitro cytotoxicity test shows that 20% SilMA has good cytocompatibility. The hybrid scaffold was then made by PCL coated with 20% SilMA. The mechanical test shows this hybrid scaffold has better biomechanical properties than native trachea. In vivo tracheal defect repair assays were conducted to evaluate the effect of the hybrid substitution. H&E staining, IHC staining and IF staining showed that this hybrid substitution ensured the viability, proliferation and migration of epithelium. However, it is sad that the results of chondrogenesis were not obvious. This study is expected to provide new strategies for the fields of tracheal replacement therapy needing mechanical properties and epithelization.

Case Report: Human Tracheal Transplantation Undergoes Progressive Reepithelialization Over Time.

Tracheal transplantation is an ideal option for the reconstruction of long-segment circumferential tracheal defects. Our group performed the first successful vascularized single-staged tracheal transplantation in January 2021. Although a rigid biocompatible structure is necessary for a functioning tracheal replacement, the importance of ciliated epithelium, which allows for critical mucociliary clearance, is now being appreciated. Here, we examined the histological changes of the first single-staged human tracheal transplant from serial endoscopic biopsies. Biopsies of the tracheal mucosa were serially obtained since the time of the tracheal transplantation. Samples were examined via hematoxylin and eosin, electron microscopy, and immunohistochemistry. One week after transplantation, there is loss of ciliated epithelium and seromucinous cells, with only a basal layer of epithelium remaining. By 2 weeks, however, the epithelium begins to recover, albeit differently depending on the location of the biopsy. Near the site of tracheal anastomosis, there is epithelial proliferation, with the appearance of early ciliated cells. However, in the midgraft, there appears to be evidence of squamous metaplasia. Over time, however, normal ciliated epithelium and mucous cells appear without signs of chronic inflammation. Critically, the tracheal allograft regained normal appearing respiratory epithelium after initial ischemic injury. The histologic differences at the midgraft versus anastomosis may suggest unique mechanisms of reepithelialization. At the recipient-donor interface, there may be a faster direct migration of recipient-derived epithelial cells, in line with preclinical studies. The midgraft, in contrast, responds with epithelial proliferation from the donor basal cells or dedifferentiated mucous cells. N/A Laryngoscope, 134:2664-2671, 2024.

Preliminary results in tracheal replacement using stented aortic matrices for primary extensive tracheal cancer.

Recent studies have demonstrated the feasibility and favorable long-term results of tracheobronchial replacement using stented cryopreserved aortic allografts. We propose to investigate the outcomes of this emerging technique in the subgroup of patients with extensive tracheal cancer. This study was based on 13 patients with primary extensive tracheal cancer extracted from the prospective registry TRITON-01 (ClinicalTrials.gov Identifier: NCT04263129), which included 40 patients in total. We analyzed early and late outcomes in this subset of patients. From March 2019 to September 2022, 13 patients were included in the study. There were 9 female and 4 male patients, with a mean age of 53.9years [36-71 years]. They had tracheal replacement for extended adenoid cystic carcinoma (n=11), squamous cell carcinoma (n=1), and mucoepidermoid carcinoma (n=1). A venovenous extracorporeal membrane oxygenation was used in the 6 last cases. The mean length of resection was 81 mm [50-120 mm]. There was no 30-day postoperative mortality. A complete resection (R0) was achieved in 11 patients. The main late complications consisted of tracheal granulomas related to the stent and requiring repeated bronchoscopies (n=9), pneumonia (n=3), airway infection (n=1), bronchoesophageal fistula (n=1), mechanical stent obstruction requiring change (n=2), and mediastinitis treated by antibiotics, drainage, and omentoplasty (n=1). With a maximal follow-up of 3years and 7months, cancer recurrence was observed in 2 patients. All patients were alive at last follow-up except 2 (84.6%). Airway replacement using stented CAA represents a feasible and promising solution for extensive tracheal cancer.