Regeneration Of Tracheal Epithelium Research Articles

A novel 3D-printed silk fibroin-based scaffold facilitates tracheal epithelium proliferation in vitro.

The functional epithelial regeneration is important for repairing tracheal defects. However, the potential of 3D-printed SF-based scaffolds for tracheal epithelial regeneration is still unknown. In this study, we developed a novel silk fibroin-based scaffold prepared by 3D printing of silk fibroin/hydroxypropyl methyl cellulose (SF/HPMC) thixotropic hydrogel and evaluated the tracheal epithelium proliferation on this scaffold in vitro. Combined with the freeze-dried technology, the 3D-printed SF/HPMC scaffolds had porous structures in the printed bars. After evaluation of their pore sizes, porosities, water contents and mechanical properties, the scaffolds were co-cultured with the normal human bronchial epithelial cell line (BEAS-2B) for seven days. We detected the BEAS-2B cells proliferation on the scaffolds using a CCK-8 assay, determined their mucin secretion and intercellular tight junction formation by immunofluorescence, as well as observed their cell viability and morphology by live/dead staining and scanning electronic microscopy. The results showed that the SF/HPMC scaffolds had good porosity, water content and mechanical properties. In addition, the BEAS-2B cells proliferated well on SF/HPMC scaffolds, during the seven-day co-culture, with high viability, mucin expression, and intercellular tight junction formation. In summary, these results demonstrated that the BEAS-2B cells could attach and proliferate on the 3D-printed SF/HPMC scaffolds, which were expected to have potential for facilitating tracheal epithelial regeneration.

Collagen sponge scaffolds containing growth factors for the functional regeneration of tracheal epithelium

Tracheal epithelia have barrier and mucociliary clearance functions that prevent invasion of extraneous particles and infectious materials. Hence, following tracheal reconstructions, functional and morphological regeneration of epithelia is required to prevent respiratory declines and infectious diseases. Although growth factors (GFs) promote the regeneration of tracheal epithelial morphologies, it remains unclear whether tracheal grafts containing GFs are beneficial for regeneration of tracheal epithelial functions. Thus, we fabricated collagen sponge scaffolds containing insulin-like GF-1 (IGF-1) and the basic fibroblast, hepatocyte, and epidermal GFs (bFGFs, HGFs, and EGFs, respectively), and we evaluated the effects of the grafts on the functional regeneration of tracheal epithelia. Partial tracheal defects were imposed surgically, and collagen sponges containing IGF-1, bFGF, HGF, or EGF were then transplanted to defect sites. Subsequent immunofluorescence studies suggested that EGF and bFGF contribute to regular distributions of tight junction molecules, and tracer permeability assays suggested that EGF and bFGF promote regeneration of barrier function. Increased ciliogenesis was also observed using scanning electron microscopy in reconstructed regions treated with EGF- and bFGF-supplemented collagen sponges. However, bFGF-supplemented collagen sponges led to greater microsphere transport than did EGF-supplemented sponges. The present data suggested that collagen sponge scaffold containing bFGF promotes functional regeneration of tracheal epithelial tissues.

Evaluation of Cilia Function in Rat Trachea Reconstructed Using Collagen Sponge Scaffold Seeded with Adipose Tissue-Derived Stem Cells.

The tracheal lumen is essential for conducting air to the lung alveoli and for voice production. However, patients with severe tracheal stenosis and malignant tumors invading the trachea often require tracheal resection. Recently, various reported tissue engineering methods for tracheal reconstruction show that regeneration of ciliated epithelium in the reconstructed areas, as well as preservation of the luminal structure is possible. However, only few studies report on the mucociliary transport function in reconstructed tracheae. We investigated mucociliary transport function within rat tracheal epithelium, reorganized after autologous adipose tissue-derived stem cell (ASC) transplantation. Rat ASCs were expanded in culture, and then seeded in a collagen sponge, which was physically supported with a polypropylene framework. The ASC-seeded collagen sponge was transplanted into the rat tracheal defect. We then examined the motility and transport function of cilia generated in the transplanted area using ciliary beat frequency (CBF) and microsphere movement analyses. Our data suggested that autologous ASC transplantation promoted ciliogenesis, consistent with previous reports. The CBF analysis revealed that motility of the cilia generated in the ASC group was comparable to that observed in the normal rat tracheal epithelium. Transport function in the ASC group was higher than that in the control group. These data suggested that autologous ASC transplantation increased ciliated cells in the reconstructed area without significantly disrupting cilia motility, thereby promoting transport function regeneration. Autologous ASC transplantation is expected to be beneficial in morphological and functional regeneration of tracheal epithelium. Anat Rec, 303:471-477, 2020. © 2019 American Association for Anatomy.

Experimental animal model for assessment of tracheal epithelium regeneration.

To develop an experimental model in rabbits for assessment of tracheal epithelium regeneration through application of either natural or artificial polymer scaffolds. First, we identified the size of full-thickness mucosal defect, which does not allow self-healing (a "critical defect"), thus representing an adequate experimental model for regenerative therapy of tracheal epithelium damage. Then, two methods of polymer scaffold fixation at the site of the epithelium defect were compared: suturing and fixation with a stent. This was done through: 1) formation of a full-thickness anterolateral mucosal defect by tracheal mucosa excision; and 2) fixation of the scaffold at the site of the tracheal epithelium defect using sutures (through a tracheal wall "window") or a vascular stent (through a small tracheal incision). The dimension of a critical anterolateral mucosal defect of the trachea for rabbits was found to be 1.5 cm in length and more than 50% of the tracheal circumference. Fixation of the scaffold with a stent proved to be more efficient due to a uniform distribution of the pressure over the entire surface of the scaffold, whereas the suturing of the scaffold provided unsatisfactory results. In addition, fixation of the scaffold by suturing required formation of a large "window" in the tracheal wall. Thus, using the stent appeared to be technically less complicated and much less traumatic as compared to suturing. We present an experimental in vivo animal model of tracheal epithelium injury and recovery. It can be effectively used with certain further modifications as a basis for routine testing of bioengineered constructs. NA Laryngoscope, 129:E213-E219, 2019.

Niche-mediated BMP/SMAD signaling regulates lung alveolar stem cell proliferation and differentiation.

ABSTRACTThe bone morphogenetic protein (BMP) signaling pathway, including antagonists, functions in lung development and regeneration of tracheal epithelium from basal stem cells. Here, we explore its role in the alveolar region, where type 2 epithelial cells (AT2s) and Pdgfrα+ type 2-associated stromal cells (TASCs) are components of the stem cell niche. We use organoids and in vivo alveolar regrowth after pneumonectomy (PNX) – a process that requires proliferation of AT2s and differentiation into type 1 cells (AT1s). BMP signaling is active in AT2s and TASCs, transiently declines post-PNX in association with upregulation of antagonists, and is restored during differentiation of AT2s to AT1s. In organoids, BMP4 inhibits AT2 proliferation, whereas antagonists (follistatin, noggin) promote AT2 self-renewal at the expense of differentiation. Gain- and loss-of-function genetic manipulation reveals that reduced BMP signaling in AT2s after PNX allows self-renewal but reduces differentiation; conversely, increased BMP signaling promotes AT1 formation. Constitutive BMP signaling in Pdgfrα+ cells reduces their AT2 support function, both after PNX and in organoid culture. Our data reveal multiple cell-type-specific roles for BMP signaling during alveolar regeneration.

Heparin cross-linked collagen sponge scaffolds improve functional regeneration of rat tracheal epithelium.

Tracheal epithelial cells maintain airway homeostasis by mediating mucociliary clearance. Following tracheal reconstruction, timely epithelial regeneration is required to prevent respiratory compromise and infectious diseases. To achieve rapid tracheal epithelial regeneration, a heparin cross-linked collagen sponge containing fibroblast growth factor-2 (FGF-2) was prepared as a graft for tracheal reconstruction. The heparin cross-linked sponge exhibited a high FGF-2 retaining capacity, and tracheal epithelial and mesenchymal cells cultured in this sponge containing FGF-2 showed high proliferative capacities. Subsequently, heparin-free collagen sponge scaffolds (C/F scaffold) and collagen sponge scaffolds cross-linked with 10μg/ml heparin retained FGF-2 (C/H10/F scaffold), and were transplanted into rats with tracheal defects. Invasion of both epithelial and non-epithelial cells was greater in rats treated with the C/H10/F scaffold at 1week post-transplantation than in rats treated with the C/F scaffold. Moreover, at 2weeks after transplantation, improved cilia formation was observed in the C/H10/F scaffold group, with higher motility and more potent posterior-anterior flow generation than in the C/F scaffold group. These results suggest that heparin improves functional regeneration of tracheal epithelium. Copyright © 2017 John Wiley & Sons, Ltd.

Changes in the methylation status of the Oct3/4, Nanog, and Sox2 promoters in stem cells during regeneration of rat tracheal epithelium after injury.

We investigated the relationship between promoter methylation and tracheal stem cell activation. We developed a model of rat tracheal epithelium regeneration after 5-fluorouracil (5-FU)-induced injury. Using immunohistochemistry and Western blotting, the expression levels of the stem cell pluripotency regulator Oct3/4 and differentiation marker CK14 were measured after 5-FU treatment. The methylation status of the Oct3/4, Nanog, and Sox2 promoters was investigated using methylation-specific PCR. Additionally, the effects of 5-azacytidine (5-azaC), a demethylating agent, on Oct3/4, Nanog, and Sox2 mRNA and protein expression were evaluated. Finally, we measured the activity of the maintenance and de novo DNA methyltransferases DNMT1, DNMT3a, and DNMT3b. Our data indicate that Oct3/4, Sox2, and Nanog are transiently expressed in response to 5-FU-induced injury, and then they are gradually silenced as the cells differentiate. DNA methylation can result in silencing of gene expression, and it can determine whether tracheal stem cells are in an active or dormant state. Treatment with 5-FU reversed the methylation of the Oct3/4, Nanog, and Sox2 promoters, which corresponded to increases in Oct3/4, Nanog, and Sox2 mRNA and protein. Thus, both maintenance and de novo methyltransferases are involved in regulating tracheal stem cell dormancy and activation.

Effects of artificial tracheal fixation on tracheal epithelial regeneration and prevention of tracheal stenosis

Conclusion: Tight fixation of the artificial trachea is important for epithelialization and tracheal stenosis.Objective: The authors have developed an artificial trachea and have used it for tracheal reconstruction. Although various studies on tracheal reconstruction have been conducted, no studies have examined the effect of artificial tracheal fixation on tracheal stenosis and regeneration. Therefore, the purpose of the present study was to evaluate the effect of artificial tracheal fixation.Study design: Preliminary animal experiment.Methods: Artificial tracheae were implanted into rabbits with partial tracheal defects. Tracheal stenosis and regeneration of the tracheal epithelium on the artificial tracheae were evaluated by endoscopic examination, scanning electron microscopic analysis, and histological examination. The artificial tracheae fixed to the tracheal defects were classified into three groups (0-point, 4-point, and 8-point) by the number of fixation points.Results: At 14 and 28 days post-implantation, the luminal surface of the implantation area was mostly covered with epithelium in all fixation groups. However, a small amount of granulation tissue was observed in the 0-point fixation group at 14 days post-implantation. Moreover, tracheal stenosis did not occur in the 8-point fixation group, but stenosis was detected in the other groups.

Regeneration of tracheal epithelium using mouse induced pluripotent stem cells

Conclusion The findings demonstrated the potential use of induced pluripotent stem cells for regeneration of tracheal epithelium.Objective Autologous tissue implantation techniques using skin or cartilage are often applied in cases of tracheal defects with laryngeal inflammatory lesions and malignant tumor invasion. However, these techniques are invasive with an unstable clinical outcome. The purpose of this study was to investigate regeneration in a tracheal defect site of nude rats after implantation of ciliated epithelium that was differentiated from induced pluripotent stem cells.Method Embryoid bodies were formed from mouse induced pluripotent stem cells. They were cultured with growth factors for 5 days, and then cultured at the air–liquid interface. The degree of differentiation achieved prior to implantation was determined by histological findings and the results of real-time polymerase chain reaction. Embryoid bodies including ciliated epithelium were embedded into collagen gel that served as an artificial scaffold, and then implanted into nude rats, creating an ‘air–liquid interface model’. Histological evaluation was performed 7 days after implantation.Results The ciliated epithelial structure survived on the lumen side of regenerated tissue. It was demonstrated histologically that the structure was composed of ciliated epithelial cells.

Patch tracheoplasty in body tissue engineering using collagenous connective tissue membranes (biosheets)

BackgroundCollagenous connective tissue membranes (biosheets) are useful for engineering cardiovascular tissue in tissue engineering. The aim was to evaluate the use of biosheets as a potential tracheal substitute material in vivo in a rabbit model. MethodsGroup 1: Rectangular-shaped Gore-Tex (4×7mm) was implanted into a 3×6mm defect created in the midventral portion of the cervical trachea. Group 2: Rectangular-shaped dermis was implanted into a tracheotomy of similar size. Group 3: Biosheets were prepared by embedding silicone moulds in dorsal subcutaneous pouches in rabbits for 1month.Rectangular-shaped biosheets were implanted into a tracheotomy of similar size in an autologous fashion. All groups (each containing 10 animals) were sacrificed 4weeks after implantation. Main ResultsAll materials maintained airway structure for up to 4weeks after implantation. Regenerative cartilage in implanted Biosheets in group 3 was confirmed by histological analysis. Tracheal epithelial regeneration occurred in the internal lumen of group 3. There were significant differences in the amounts of collagen type II and glycosaminoglycan between group 3 and group 1 or 2. ConclusionWe confirm that cartilage can self-regenerate onto an airway patch using Biosheets.

Effect of Structural Differences in Collagen Sponge Scaffolds on Tracheal Epithelium Regeneration.

We developed an in situ regeneration-inducible artificial trachea composed of a porcine collagen sponge and polypropylene framework and used it for tracheal reconstruction. In the present study, collagen sponges with different structures were prepared from various concentrations of collagen solutions, and their effect on the regeneration of tracheal epithelium was examined. Collagen sponges were prepared from type I and III collagen solutions. The structures of the sponges were analyzed using scanning electron microscopy (SEM). Artificial tracheae, which were formed using the collagen sponges with different structures, were implanted into rabbits, and regeneration of the tracheal epithelium on the artificial tracheae was evaluated by SEM analysis and histological examination. The SEM analysis showed that collagen sponges prepared from 0.5% and 1.0% collagen solutions had a porous structure. However, the sponges prepared from a 1.5% collagen solution had a nonporous structure. After implantation of artificial tracheae prepared from 0.5% and 1.0% collagen solutions, their luminal surfaces were mostly covered with epithelium within 14 days. However, epithelial reorganization occurred later on artificial tracheae prepared from the 1.5% collagen solution. Collagen sponges with a porous structure are suitable for regeneration of the tracheal epithelium in our artificial trachea.

Human turbinate mesenchymal stromal cell sheets with bellows graft for rapid tracheal epithelial regeneration

Rapid functional epithelial regeneration on the luminal surface is essential when using artificial tracheal grafts to repair tracheal defects. In this study, we imposed human turbinate mesenchymal stromal cell (hTMSC) sheets for tracheal epithelial regeneration, and then assessed their potential as a new clinical cell source. In vitro, hTMSCs sheets showed high capacity to differentiate into tracheal epithelium. We fabricated a poly(ε-caprolactone) (PCL) tracheal graft by indirect three-dimensional (3D) printing technique and created a composite construct by transplanting the hTMSC sheets to its luminal surface of the tracheal graft, then applied this tissue-engineered tracheal graft to non-circumferential tracheal reconstruction in a rabbit model. 4weeks after implantation, the luminal surface of tissue-engineered tracheal graft was covered by a mature and highly-ciliated epithelium, whereas tracheal grafts without hTMSC sheets were covered by only a thin, immature epithelium. Therefore, hTMSC sheets on the luminal surface of a tissue-engineered tracheal graft can accelerate the tracheal epithelial regeneration, and the tissue-engineered tracheal graft with hTMSC sheets provides a useful clinical alternative for tracheal epithelial regeneration.

Synergistic effect of laminin and mesenchymal stem cells on tracheal mucosal regeneration

Although several studies have been successfully undertaken of tracheal reconstruction in terms of the maintaining the framework of the graft, most cases of reconstruction failure have resulted from delayed mucosal regeneration. The purposes of this study were to evaluate whether laminin-coated asymmetrically porous membrane (APM) scaffold enhances mucosal regeneration, to compare the mucosalization capability with mesenchymal stem cell (MSC) seeded APM, and to determine whether laminin coating and MSC seeding has a synergistic effect on mucosal regeneration. We reconstructed the full-thickness anterior tracheal defect of 36 New Zealand White rabbits with the APM scaffold. MSCs were isolated from the rabbit's inguinal fat. The animals were divided into 4 groups by the presence of laminin coating on APM and application of MSC [Group I, −/− (laminin/MSC); Group II, −/+; Group III, +/−; Group IV, +/+]. Endoscopy and histologic evaluation were performed and the results were compared among the groups. The results showed that ciliated columnar epithelium was regenerated earlier in groups II and III than in group I. Furthermore, the application of laminin and MSC had synergistic effects on tracheal epithelial regeneration. These results demonstrate that tracheal reconstruction by laminin-coated APM seeded with MSCs is most effective in enhancing tracheal mucosalization, and appears to be promising strategy in the regenerative treatment of tracheal defects.

Tracheoplasty with cartilage-engineered esophagus environments

PurposeOur objective was to investigate the feasibility of engineering cartilage on the esophagus layer and outside the esophagus. Moreover, we investigated the feasibility of tracheoplasty with cartilage engineered on the esophagus in rabbits. MethodsChondrocytes were isolated from auricular cartilages. 1. Engineered cartilage formation by histological findings on/into the esophageal layer was compared with that of injectable scaffold and preformed scaffold with chondrocytes. 2. Chondrocytes adhered to gelatin+vicryl mesh™ and b-FGF, were implanted on the outer esophageal surface. Four weeks after seeding, we found that cartilage was implanted in the midposterior portion of the cervical trachea (n=5), and it was retrieved 8weeks after seeding. Results1. A gelatin sponge incorporating β-TCP with vicryl mesh™ showed the best performance for fabricating engineered cartilage on the outer side of the esophagus. 2. Two of 5 rabbits died due to obstructed esophagus. Cartilage engineered outside the esophagus by a composite scaffold as the main material in the gelatin sponge, maintained the airway structure for up to 1month after implantation. Tracheal epithelial regeneration occurred in the internal lumen of this engineered cartilage. ConclusionTracheoplasty with cartilage engineered outside the esophagus may be useful for reconstructing airways.

EFFECTS OF DIFFERENT ANTIOXIDANT ENZYMES ON THE TRACHEAL EPITHELIUM REGENERATION AFTER CHEMICAL BURN

The aim of the study was to investigate a role of different antioxidant enzymes for tracheal epithelium regeneration after chemical burn. Methods. The study was conducted in a rat model of chemical burn of the upper airways caused by inhaled hydrochloric acid. Results. According to results of histological examination, to the 2nd day after the exposure approximately 70 % of the epithelial surface remained injured, mostly due to death of the ciliated cells. The degree of the damage was unchanged to the 4th day after the exposure. Visible regeneration of the tracheal epithelium began to the 7th day. The death of the tracheal epithelial cells was generally due to necrosis though cell apoptosis also occurred. The expression of all antioxidant enzymes was greatly decreased during the 1st day after the exposure followed by its growth to maximum to the 7th day and normal level to the 15th day after the burn. Exposition of superoxide dismutase to the trachea resulted in a significant epithelium destruction. On contrary, peroxiredoxin 6 and a chimeric protein containing peroxiredoxin / superoxide dismutase significantly protected the tracheal epithelium. Conclusion. Peroxiredoxins and their derivates could be used as highly efficient therapeutic agents with potent antioxidant action in patients with burn injury of the upper airways.

Acceptable Warm Ischemia Time of Tracheal Grafts From Non–heart-beating Donors in Rats

Objectives Warm ischemia (WI)-induced airway complications are common in clinical lung transplantation. However, the acceptable WI time of tracheal grafts from non–heart-beating donors (NHBDs) is unknown. The purpose of this study was to determine the acceptable WI time by observing tracheal epithelial regeneration among NHBD. Method Forty-eight rats were randomly divided into four groups (each with 12 rats): WI-0 minutes (group A), WI-30 minutes (group B), WI-45 minutes (group C), and WI-60 minutes (group D). In each group, the tracheas from 6 rats were imbedded in the greater omentum of 6 other rats. Fourteen days later, the transplanted trachea was obtained from the recipient to evaluate epithelial thickness and regeneration. Six tracheas were obtained from living donors as a control group. Results There were no significant differences in tracheal transplantation time (mean, 17.66 ± 1.21 minutes). There were no significant differences in epithelial thickness and regeneration between the controls and groups A, B, and C ( P < .05). Group D showed no normal epithelial structure of the trachea only with monolayer cells. Conclusions The time limits of tolerance to WI of tracheal grafts from NHBDs may be 45 minutes.

Bone Marrow-Derived Mesenchymal Stem Cells Enhance Cryopreserved Trachea Allograft Epithelium Regeneration and Vascular Endothelial Growth Factor Expression

Epithelium regeneration and revascularization of tracheal implants are challenging issues to be solved in tracheal transplantation research. Bone marrow-derived mesenchymal stem cells (BMSCs) can migrate to the damaged tissue and promote functional restoration. Here, we applied intravenous transplantation of BMSCs combined with a cryopreserved allograft to investigate the role of BMSCs in enhancing implant survival, tracheal epithelium regeneration and revascularization. After transplantation with cryopreserved allografts, PKH-26 labeled 3 to 5 passage BMSCs were injected into recipient rats through the tail vein. Rats in the control groups were injected with a comparable amount of phosphate-buffered saline. We observed the histology of the tracheal allograft and measured vascular endothelial growth factor (VEGF) protein levels in the epithelium to evaluate the effect of BMSCs on epithelium regeneration and revascularization. Histologic observation of the rats from the BMSCs injection groups showed that the tracheal lumen was covered by pseudostriated ciliated columnar epithelium. The cartilage structure was intact. There were no signs of denaturation or necrosis. PKH-26 labeled BMSCs migrated to the implant site and exhibited red fluorescence, with the brightest red fluorescence at the anastomotic site. VEGF protein levels in the allograft epithelium of the BMSCs injection group were higher than the levels in the phosphate-buffered saline injection group. Our results indicate that given systemic administration, BMSCs may enhance epithelium regeneration and revascularization by upregulating VEGF expression.

Tracheal Replacement with Fresh and Cryopreserved Aortic Allograft in Adult Dog

Many reports have described tracheal replacement using aortic allografts, with varying results and minimal understanding of the mechanism of tracheal regeneration. The present study attempted tracheal regeneration in adult dogs using fresh aortic allografts (FAA) and cryopreserved aortic allografts (CAA). Twelve adult beagles underwent tracheal resection and were transplanted with FAA (n = 5) or CAA (n = 7). Animals were followed-up with serial radiography and magnetic resonance imaging, and were euthanized at predetermined times up to 16 mo post-surgery. There were no procedural deaths, but two animals died due to stent migration. Stent migration occurred in seven of the 12 animals. Evidence of regeneration of tracheal epithelium was observed in the surviving animals, with the transformation of squamous metaplasia to mucociliary epithelium being time-dependent. Islet of cartilage were observed in animals after 6 mo, but ring-like cartilage structures were absent, even after 16 mo. During autopsy, axial graft contractions up to 68% were observed. Serial radiographs show that most of the contraction occurred within 1 mo. The results of the MRI showed that the graft area was strongly enhanced for up to 2 mo, but was clearly reduced after 3 mo. Tracheal replacement in adult dogs using FAA or CAA is feasible. However, immaturity of the neotracheal cartilage did not allow the tissue to function as native tracheal tissue. Prolonged stenting should be considered in adult if the procedure is to be clinically contemplated.

A tissue-engineered trachea derived from a framed collagen scaffold, gingival fibroblasts and adipose-derived stem cells

In some types of tracheal disease, tracheal resection is required. For patients with tracheal resection, artificial grafts, made from collagen sponge with a spiral polypropylene stent and mesh, have been clinically used by our group. However, epithelial regeneration was confirmed to be slow. In the present study, we investigated the potential of gingival fibroblasts (GFBs) and adipose-derived stem cells (ASCs) as autologous transplanted cells in combination with artificial graft for tracheal epithelial regeneration. In in vitro co-culturing with tracheal epithelial cells, GFBs stimulated epithelial cell differentiation and reconstruction of a pseudostratified epithelium. ASCs stimulated epithelial cell proliferation and reconstruction of a multi-layered epithelium. Subsequently, we prepared three kinds of bioengineered scaffolds from GFBs and/or ASCs and implanted them into rat tracheal defects. The bioengineered scaffolds containing GFBs were covered with tracheal epithelial cells after 1 week, and highly ciliated epithelium was formed after 2 weeks of transplantation. The bioengineered scaffold containing ASCs induced thick epithelium, and then pseudostratified epithelium containing goblet cells was formed. Furthermore, the application of both GFBs and ASCs had synergistic effects on tracheal epithelial regeneration, suggesting that bioengineered scaffolds containing GFBs and ASCs are useful for hastening tracheal epithelial regeneration.

Tissue‐engineered allograft tracheal cartilage using fibrin/hyaluronan composite gel and its in vivo implantation

Treatment and management of tracheal defects remain challenging in head and neck surgery. Various reconstruction techniques have been used, with no consensus on the best approach. The purpose of this study was to explore a novel strategy to fabricate tissue-engineered trachea by using fibrin/hyaluronic acid (HA) composite gel and evaluate the feasibility of creating tracheal cartilage. A preliminary animal experiment. Chondrocytes from rabbit cartilage were expanded and seeded into a culture dish at high density to form mechanically stable allograft tracheal cartilage using fibrin/HA composite gel. After a longitudinal cervical skin incision, the trachea was exposed and a rectangular defect (1 x 0.5 cm) was created on the cervical trachea by scalpel on six rabbits. Tissue-engineered cartilage using fibrin/HA composite was trimmed and fixed to defect boundaries with Tissucol (Baxter International, Deerfield, IL). Postoperatively, the site was evaluated endoscopically, histologically, radiologically, and functionally. Postoperatively, rigid telescopic examination showed that the implanted scaffolds in all cases were completely covered with regenerated mucosa without granulation or stenosis. Histologic data showed ciliated epithelium regenerated at the operated site from 2 months postoperatively. Ciliary beat frequency of ciliated epithelium on implants was very similar to normal respiratory mucosa. Computed tomography images revealed fine luminal contour of the regenerated site. However, allograft cartilage implanted was found to be partially preserved on the postoperative specimen. The tracheal luminal contour and functional epithelial regeneration without graft rejection and inflammation were observed after repair of a tracheal resection using allogeneic implants with chondrocytes cultured with fibrin/HA.