DsbA-L deletion attenuates LPS-induced acute kidney injury by modulating macrophage polarization.

Disulfide bond A oxidoreductase-like protein (DsbA-L) drives acute kidney injury (AKI) by directly upregulating the expression of voltage-dependent anion-selective channels in proximal tubular cells. However, the role of DsbA-L in immune cells remains unclear. In this study, we used an LPS-induced AKI mouse model to assess the hypothesis that DsbA-L deletion attenuates LPS-induced AKI and explore the potential mechanism of DsbA-L action. After 24 hours of LPS exposure, the DsbA-L knockout group exhibited lower serum creatinine levels compared to the WT group. Furthermore, peripheral levels of the inflammatory cytokine IL-6 were decreased. Transcriptomic data analysis revealed a significant down-regulation in the IL-17 and tumor necrosis factor pathways in DsbA-L knockout mice following LPS induction. Metabolomic analysis suggested that arginine metabolism was significantly different between the WT and DsbA-L knockout groups after LPS treatment. Notably, the M1 polarization of macrophages in the kidneys of DsbA-L knockout AKI mice was significantly reduced. Expression of the transcription factors NF-κB and AP-1 was downregulated after DsbA-L knockout. Our results suggest that DsbA-L regulates LPS-mediated oxidative stress, promotes M1 polarization of macrophages, and induces expression of inflammatory factors via the NF-κB/AP-1 pathway.

Novel immunosuppressive effect of FK506 by upregulation of PD-L1 via FKBP51 in heart transplantation.

The calcineurin inhibitor-FK506-is a first-line immunosuppressant that regulates T cell secretion of IL-2 and other cytokines. However, the mechanism of its protective effect on target cells and its role on tumour recurrence and interaction with anti-tumour immune checkpoint inhibitors, such as PD-L1 blocking, are still unclear. Here, in a murine heart transplantation model, we observed the upregulation of programmed death-ligand 1 (PD-L1) expression by FK506 in both dendritic cells (DCs) and allografts. Blocking PD-L1 during FK506 treatment increased IFN-γ and TNF-α expression, enhanced CD4+ and CD8+ T cell proliferation, and suppressed Treg differentiation. Moreover, PD-L1 decreased T cell infiltration and induced T cell apoptosis in both the spleen and graft. PD-L1 was not only required in FK506-mediated immunosuppression but also upregulated by FK506. Treatment with SAFit2, a FKBP51 selective inhibitor, reduced the expression of PD-L1 on DCs and the grafts and interfered with the immunosuppressive effect of FK506, suggesting that the mechanism depends on FK506-binding protein (FKBP) 51 expression. Overall, our results add new insights into the role of FK506, not only on T cell cytokine secretion but also on co-inhibitory molecular regulation and target cell immune privilege.

Enantioselective total synthesis, divergent optimization and preliminary biological evaluation of (indole-N-alkyl)-diketopiperazines.

The first enantioselective total synthesis of the antifungal natural product (indole-N-isoprenyl)-tryptophan-valine diketopiperazine 5 was accomplished. Four stereoisomers of 5 were intentionally prepared, and the (R, R)-isomer is more favorable in enhancing the antifungal bioactivity. Divergent structural optimization of this attractive model was conducted from the chiral pool amino acids. Fine-tuning of the structure protruded the broad-spectrum antifungal 6b, which also showed good preventative efficacy against Sclerotinia scleotiorum. Compound 5d could accelerate both hypocotyl elongation and root growth of Eclipta prostrata even at the concentration of <2.5 ppm. This unique and easily accessible scaffold will be of prime importance in achieving agrochemical candidates with the novel scaffold.

Management of Severe and Complex Hypopharyngeal and/or Laryngotracheal Stenoses by Various Open Surgical Procedures: A Retrospective Study of Seventeen Patients.

OBJECTIVE: To systematically study various surgical approaches for treating complex hypopharyngeal and/or laryngotracheal stenoses at a variety of sites and levels. PATIENTS AND METHODS: We retrospectively analyzed the treatment of 17 patients with severe and complex hypopharyngeal and/or laryngotracheal stenosis at various sites and levels of severity. All of the 17 patients initially had a tracheostomy. Thirteen had failed the previous laser lysis and/or dilation treatment. Given the high severity and complexity of stenosis, all of these patients were treated by open surgical reconstruction techniques using repairing grafts (flaps), followed by stenting. RESULTS: Thirteen of 17 patients had successful decannulation 1-8 months post-operation and had stable airway and adequate vocal and swallow function. Two patients with complex hypopharyngeal and esophageal stenosis had unsuccessful decannulation. Follow-up was lost in 1 patient with complex hypopharyngeal and esophageal stenosis and 1 patient with original hypopharyngeal stenosis and recurrent thoracotracheal stenosis. CONCLUSION: Despite the failure by the regular treatments using laser lysis and/or dilation therapy, severe and complex hypopharyngeal and/or laryngotracheal stenosis may be successfully treated by variable open surgical reconstruction techniques using different grafts (flaps) depending on the site and severity of the stenosis.

MicroRNA-24 inhibits growth, induces apoptosis, and reverses radioresistance in laryngeal squamous cell carcinoma by targeting X-linked inhibitor of apoptosis protein.

BACKGROUND: Increasing evidence indicates that dysregulation of microRNAs is involved in tumor progression and development. The aim of this study was to investigate the expression of microRNA-24 (miR-24) and its function in laryngeal squamous cell carcinoma (LSCC). METHODS: Quantitative RT-PCR (qRT-PCR) was used to detect miR-24 expression in LSCC cell lines and tissue samples. MTT, colony formation, and flow cytometry was performed to analyze the effects of miR-24 expression on growth, apoptosis, and radiosensitivity of LSCC cells. Dual-luciferase reporter assays were performed to examine regulation of putative miR-24 targets. Expression of X-linked inhibitor of apoptosis protein (XIAP) mRNA and protein, cleaved or total caspase-3, and cleaved or total PARP protein were detected by qRT-PCR and western blotting assays, respectively. RESULTS: miR-24 expression levels in LSCC cell lines or tissue were significantly lower than in a normal human keratinocyte cell line or adjacent normal tissues. Functional analyses indicated that re-expression of miR-24 inhibits growth, reduces colony formation, and enhances apoptosis in LSCC cells. In addition, miR-24 upregulation increases LSCC sensitivity to irradiation by enhancing irradiation-induced apoptosis, and luciferase activity indicated that miR-24 binds to the 3'-untranslated region (3'-UTR) of XIAP mRNA. Upregulation of miR-24 inhibits XIAP protein expression in LSCC cells, and silencing of XIAP mimics the effects of miR-24 upregulation on LSCC cells. In addition, XIAP mRNA expression significantly increases in LSCC tissues and is inversely correlated with miR-24 expression. CONCLUSIONS: Our data suggest that miR-24 inhibits growth, increases apoptosis, and enhances radiosensitivity in LSCC cells by targeting XIAP. Therefore, miR-24 may be a potential molecular target for the treatment of human LSCC.

Visible-Light-Mediated Bimetal-Catalyzed meta-Alkylation of Arenes.

A mild approach to the visible-light-mediated bimetal-catalyzed meta-alkylation of arenes has been accomplished. The regioselective meta-alkylation is realized by a bimetallic ruthenium-palladium system. Ruthenium acts as a catalyst for the directing effect and as a photosensitizer, while the cocatalyst palladium behaves as a catalyst for the generation of fluoroalkyl radicals. This reaction not only is suitable for two-component meta-fluoroalkylation of arenes but can also be extended to three-component reactions to achieve bifunctionalization of olefins.

Silver(I)-catalyzed highly para-selective phosphonation of 2-aryloxazolines.

A silver-catalyzed phosphonation of 2-aryloxazolines has been accomplished. This protocol provides highly regioselective access to para-phosphonation products with good functional group tolerance and moderate to good yields via cross-dehydrogenation coupling. Mechanistic studies have shown that para-phosphonation products are obtained via a radical pathway. Furthermore, the directing oxazoline group in the para-phosphonation products is removable and can be converted to benzoic esters.

Study of Manipulative Pore Formation upon Polymeric Coating for the Endowment of the Switchable Property between Passive Daytime Radiative Cooling and Heating.

Passive daytime radiative cooling (PDRC) emerges as a promising cooling strategy with an attractive feature of no energy and refrigerant consumption. In the current study, for the purpose of achieving cost-efficient fabrication of a PDRC polymeric material, a microporous polymeric coating is prepared by a novel "inverse emulsion"-"breath figure" (Ie-BF) method using water droplets as pore-formation template, and the porous morphologies of both the surface and bulk layer can be dynamically manipulated by tuning the emulsion composition as well as environmental conditions. Therefore, the solar reflectivity of the Ie-BF coating can be efficiently tuned within a rather wide range (21-91%) by facile modulation of porosity and thickness. The Ie-BF coating with a thickness of only 125 µm exhibits a high solar reflectance of 85.4% and a long-wave infrared emissivity of 96.3%, realizing a subambient radiative cooling of 6.7 °C and a cooling power of ∼76 W m-2 in the open air. Moreover, by employing the reversible feature of in situ pore formation and erasure combined with the additional attachment of a carbon black layer, the composite film could be easily switched between cooling and heating modes by solvent post-treatment. This research establishes a cost-efficient strategy with high flexibility in the structural manipulation concerning the construction of porous polymeric PDRC coating.

De-Epithelialized Viable Tracheal Allotransplantation Without Immunosuppressants: 5-Year Follow-Up.

OBJECTIVE: Tracheal transplantation could be a better option for patients with long segmental laryngotracheal stenosis or defects, but the need for immunosuppressants limits its widespread use due to the antigenicity of the tracheal epithelium. Chemically treated or cryopreserved nonviable tracheal allografts have no immunogenicity but lead to necrosis and stenosis in long-term outcomes. The present report describes the 5-year outcomes of de-epithelialized viable tracheal allotransplantation without immunosuppressants in a patient with severe laryngotracheal stenosis. METHODS: The recipient was a 47-year-old female with relapsing polychondritis affecting the larynx and cervical trachea and producing a 5 cm long stenosis that could not be repaired using resection and anastomosis. A tracheal allograft was obtained from a 45-year-old male donor and treated with a combination of 3% sodium dodecyl sulfate (SDS) and organ preservation solution for 138 hours. The allograft was revascularized by heterotopical implantation in the infrahyoid muscles of the recipient for 3 months and then transplantation to the laryngotracheal defect with a split-thickness skin graft sutured to the lumen and a silicon T-tube. No immunosuppressants were used postoperatively. RESULTS: The allograft was de-epithelialized, and most of the cartilage rings remained viable after the treatment. The allograft was revascularized, viable, and mechanically stable after 3 months of heterotopic implantation. No apparent signs of rejection or destruction were observed. The T-tube was removed, and the internal lining of the allograft was repopulated 4 months after orthotopic transplantation, despite the skin graft necrotizing at 2 weeks. Endoscopy and computed tomography showed a patent airway 5 years after orthotopic transplantation. The patient was able to resume her usual quality of life. CONCLUSION: The present study demonstrates that transplantation of the de-epithelialized viable tracheal allograft without immunosuppressants is safe and promising for patients with long laryngotracheal stenosis or defects, especially for those with malignant tumor resections.

Swallowing rehabilitation of patients undergoing T-tube implantation for the treatment of laryngotracheal stenosis.

OBJECTIVE: Patients with laryngotracheal stenosis (LTS) often have dysphagia after laryngotracheal reconstruction with T-tube insertion, which affects the quality of life. The purpose of this study is to observe the effect of swallowing rehabilitation therapy on the improvement of quality of life in patients of otolaryngology-head and neck surgery with dysphagia undergoing T-tube implantation treatment through longitudinal study. METHODS: Thirty-eight patients with LTS who experienced dysphagia after laryngotracheal reconstruction and T-tube implantation were recruited. All patients received swallowing rehabilitation therapy. The assessment of swallowing function was performed using the 10-item Eating Assessment Tool (EAT-10), the 30 mL water swallow test (WST), and flexible endoscopic evaluation of swallow (FEES). RESULTS: After swallowing rehabilitation therapy, timing of swallowing, grade of dysphagia, performance on FEES and 30 mL WST, and EAT-10 score all improved. Thirty-eight patients successfully transitioned to oral feeding and were able to remove their nasogastric tubes without experiencing any complications, including aspiration pneumonia. CONCLUSION: For patients with LTS who experienced dysphagia after laryngotracheal reconstruction and T-tube implantation, swallowing rehabilitation therapy could improve swallowing function of the patients, so as to reduce the potential harm caused by the pain and complications of surgery experienced by patients.

Bioinspired Bouligand-Structured Cellulose Nanocrystals/Poly(vinyl alcohol) Composite Hydrogel for Enhanced Impact Resistance.

Impact-protective materials are gaining importance because of the widespread occurrence of impact damage. Hydrogels have emerged as promising candidates owing to their lightweight and flexible nature. However, achieving soft impact-resistant hydrogels with exceptional stiffness, strength, and toughness remains a challenge. Inspired by the Bouligand structure found in the smasher dactyl club of stomatopods, we propose a straightforward multiscale hierarchical structural design strategy. This strategy integrates self-assembly and salting-out techniques to enhance the impact resistance of soft hydrogels. Rigid cellulose nanocrystals (CNCs) self-assemble into Bouligand-like structures within soft poly(vinyl alcohol) (PVA) matrix via supramolecular interactions. This rational structural design combines the CNC Bouligand structure with a cross-linked network of soft PVA crystalline domains, resulting in a composite hydrogel with impressive mechanical properties: high tensile fracture strength (30.2 MPa), elastic modulus (62.7 MPa), and fracture energy (75.6 kJ m-2), surpassing those of other tough hydrogels. Moreover, the multiscale hierarchical structure facilitates various energy dissipation mechanisms, including crack twisting, tortuous crack paths, and PVA chain orientation, resulting in notable force attenuation (80.4%) in the composite hydrogel. This biomimetic design strategy opens new avenues for developing soft and lightweight impact-resistant materials.

Inhibition of interferon regulatory factor 4 orchestrates T cell dysfunction, extending mouse cardiac allograft survival.

BACKGROUND: T cell dysfunction, which includes exhaustion, anergy, and senescence, is a distinct T cell differentiation state that occurs after antigen exposure. Although T cell dysfunction has been a cornerstone of cancer immunotherapy, its potential in transplant research, while not yet as extensively explored, is attracting growing interest. Interferon regulatory factor 4 (IRF4) has been shown to play a pivotal role in inducing T cell dysfunction. METHODS: A novel ultra-low-dose combination of Trametinib and Rapamycin, targeting IRF4 inhibition, was employed to investigate T cell proliferation, apoptosis, cytokine secretion, expression of T-cell dysfunction-associated molecules, effects of MAPK and mammalian target of Rapamycin (mTOR) signaling pathways, and allograft survival in both in vitro and BALB/c to C57BL/6 mouse cardiac transplantation models. RESULTS: In vitro, blockade of IRF4 in T cells effectively inhibited T cell proliferation, increased apoptosis, and significantly upregulated the expression of programmed cell death protein 1 (PD-1), Helios, CD160, and cytotoxic T lymphocyte-associated antigen (CTLA-4), markers of T cell dysfunction. Furthermore, it suppressed the secretion of pro-inflammatory cytokines interferon (IFN)-γ and interleukin (IL)-17. Combining ultra-low-dose Trametinib (0.1 mg·kg-1·day-1) and Rapamycin (0.1 mg·kg-1·day-1) demonstrably extended graft survival, with 4 out of 5 mice exceeding 100 days post-transplantation. Moreover, analysis of grafts at day 7 confirmed sustained IFN regulatory factor 4 (IRF4) inhibition, enhanced PD-1 expression, and suppressed IFN-γ secretion, reinforcing the in vivo efficacy of this IRF4-targeting approach. The combination of Trametinib and Rapamycin synergistically inhibited the MAPK and mTOR signaling network, leading to a more pronounced suppression of IRF4 expression. CONCLUSIONS: Targeting IRF4, a key regulator of T cell dysfunction, presents a promising avenue for inducing transplant immune tolerance. In this study, we demonstrate that a novel ultra-low-dose combination of Trametinib and Rapamycin synergistically suppresses the MAPK and mTOR signaling network, leading to profound IRF4 inhibition, promoting allograft acceptance, and offering a potential new therapeutic strategy for improved transplant outcomes. However, further research is necessary to elucidate the underlying pharmacological mechanisms and facilitate translation to clinical practice.

TREM2 deficiency aggravates renal injury by promoting macrophage apoptosis and polarization via the JAK-STAT pathway in mice.

The triggering receptor expressed on myeloid cells 2 (TREM2) is an immune receptor that affects cellular phenotypes by modulating phagocytosis and metabolism, promoting cell survival, and counteracting inflammation. Its role in renal injury, in particular, unilateral ureteral obstruction (UUO) or ischemia-reperfusion injury (IRI)-induced renal injury remains unclear. In our study, WT and Trem2-/- mice were employed to evaluate the role of TREM2 in renal macrophage infiltration and tissue injury after UUO. Bone marrow-derived macrophages (BMDM) from both mouse genotypes were cultured and polarized for in vitro experiments. Next, the effects of TREM2 on renal injury and macrophage polarization in IRI mice were also explored. We found that TREM2 expression was upregulated in the obstructed kidneys. TREM2 deficiency exacerbated renal inflammation and fibrosis 3 and 7 days after UUO, in association with reduced macrophage infiltration. Trem2-/- BMDM exhibited increased apoptosis and poorer survival compared with WT BMDM. Meanwhile, TREM2 deficiency augmented M1 and M2 polarization after UUO. Consistent with the in vivo observations, TREM2 deficiency led to increased polarization of BMDM towards the M1 proinflammatory phenotype. Mechanistically, TREM2 deficiency promoted M1 and M2 polarization via the JAK-STAT pathway in the presence of TGF-ß1, thereby affecting cell survival by regulating mTOR signaling. Furthermore, cyclocreatine supplementation alleviated cell death caused by TREM2 deficiency. Additionally, we found that TREM2 deficiency promoted renal injury, fibrosis, and macrophage polarization in IRI mice. The current data suggest that TREM2 deficiency aggravates renal injury by promoting macrophage apoptosis and polarization via the JAK-STAT pathway. These findings have implications for the role of TREM2 in the regulation of renal injury that justify further evaluation.

Visible-Light-Promoted C4-Selective Phosphorylation of Pyridine and Quinoline Derivatives.

The visible-light-promoted C4-selective phosphorylation of unprefunctionalized pyridine and quinoline derivatives has been accomplished. This Minisci-type protocol provides highly regioselective access to C4-phosphorylation products with good functional group tolerance and moderate to good yields via cross-dehydrogenation coupling under mild conditions. Mechanistic studies have shown that the C4-phosphorylation products are obtained via a radical pathway.

Homoharringtonine promotes heart allograft acceptance by enhancing regulatory T cells induction in a mouse model.

BACKGROUND: Homoharringtonine (HHT) is an effective anti-inflammatory, anti-viral, and anti-tumor protein synthesis inhibitor that has been applied clinically. Here, we explored the therapeutic effects of HHT in a mouse heart transplant model. METHODS: Healthy C57BL/6 mice were used to observe the toxicity of HHT in the liver, kidney, and hematology. A mouse heart transplantation model was constructed, and the potential mechanism of HHT prolonging allograft survival was evaluated using Kaplan-Meier analysis, immunostaining, and bulk RNA sequencing analysis. The HHT-T cell crosstalk was modeled ex vivo to further verify the molecular mechanism of HHT-induced regulatory T cells (Tregs) differentiation. RESULTS: HHT inhibited the activation and proliferation of T cells and promoted their apoptosis ex vivo. Treatment of 0.5 mg/kg HHT for 10 days significantly prolonged the mean graft survival time of the allografts from 7 days to 48 days (P <0.001) without non-immune toxicity. The allografts had long-term survival after continuous HHT treatment for 28 days. HHT significantly reduced lymphocyte infiltration in the graft, and interferon-γ-secreting CD4+ and CD8+ T cells in the spleen (P <0.01). HHT significantly increased the number of peripheral Tregs (about 20%, P <0.001) and serum interleukin (IL)-10 levels. HHT downregulated the expression of T cell receptor (TCR) signaling pathway-related genes (CD4, H2-Eb1, TRAT1, and CD74) and upregulated the expression of IL-10 and transforming growth factor (TGF)-ß pathway-related genes and Treg signature genes (CTLA4, Foxp3, CD74, and ICOS). HHT increased CD4+ Foxp3+ cells and Foxp3 expression ex vivo, and it enhanced the inhibitory function of inducible Tregs. CONCLUSIONS: HHT promotes Treg cell differentiation and enhances Treg suppressive function by attenuating the TCR signaling pathway and upregulating the expression of Treg signature genes and IL-10 levels, thereby promoting mouse heart allograft acceptance. These findings may have therapeutic implications for organ transplant recipients, particularly those with viral infections and malignancies, which require a more suitable anti-rejection medication.

Effect of Ball Burnishing Pressure on Surface Roughness by Low Plasticity Burnishing Inconel 718 Pre-Turned Surface.

The low plasticity burnished surface roughness is significantly affected by the low plasticity burnishing (LPB) parameters. This research proposed the analytical prediction model to predict the LPBed surface roughness and optimal LPB pressure based on Hertz contact mechanics and the slip-line field theory. In this study, the surface formatted process was divided into the smoothing stage (SS) and the indentation stage (IS). The smoothing mechanism of SS and the deterioration mechanism of IS were analyzed theoretically. The analytical prediction model for the LPBed surface roughness was proposed based on Hertz contact mechanics and slip-line field theory. The proposed analytical prediction model was validated by the LPBed surface roughness of AISI 1042, and the error of the analytically predicted results was less than 13.3%. After validation, the proposed model was applied to predict the LPBed surface roughness of Inconel 718. The single-factor experiments were conducted. The error between the proposed model prediction results and experimental results was less than 7% for the LPBed surface roughness of Inconel 718. The optimal LPB pressure interval was calculated to be [12.2 MPa, 17.5 MPa], corresponding to the experimental one as [12 MPa, 18 MPa]. It indicated that the proposed model could accurately predict the LPBed surface roughness and conduct the LPB processing.

Reconstruction of tracheal wall defect with a mesh patch of nickel-titanium shape-memory alloy.

OBJECTIVES: We explored the feasibility of reconstructing tracheal wall defects with a mesh patch fashioned from a nickel-titanium shape-memory alloy. METHODS: A tracheal wall defect was first constructed surgically by resecting the anterior half of the tracheal wall between the second and sixth tracheal rings. The defect was reconstructed in 8 experimental animals by replacing the resected tracheal mucosa and tracheal cartilage with a pedicle skin flap, which was then enclosed in the mesh patch. In 4 control animals, only a pedicle skin flap with strap muscles was used in the reconstruction procedure. The performance of the animals was observed after surgery. At the end of the experiments, the reconstructed segment was harvested for anatomic evaluation. RESULTS: In the experimental group, 1 animal died 5 days after the operation. Endoscopic and anatomic examination of the 7 animals that survived the observation period showed that the reconstructed trachea was stable, with sufficient airway space for breathing. All 4 control animals died after the operation. After observing successful completion of this operation in animals, we successfully used this method to repair a tracheal wall defect in a human victim of a traffic accident. CONCLUSIONS: Tracheal defects can be successfully reconstructed by use of a mesh patch of nickel-titanium shape-memory alloy as an extraluminal stent--a method that avoids complications associated with intraluminal stents.

Effects of Machined Surface Integrity on High-Temperature Low-Cycle Fatigue Life and Process Parameters Optimization of Turning Superalloy Inconel 718.

Machined surface integrity characteristics, including surface stresses, physical-mechanical properties and metallographic structures, play important roles in the fatigue performance of machined components. This work aimed at investigating the effects of machined surface integrity on high-temperature low-cycle fatigue life. The process parameters were optimized to obtain required surface integrity and fatigue life of the turning superalloy Inconel 718. The relationships between low-cycle fatigue life and machined surface integrity characterization parameters were established based on the low-cycle fatigue tests at a high temperature (650 °C). The sensitivities of turning process parameters to high-temperature low-cycle fatigue life were analyzed, and the optimization parameters were proposed with the goal of antifatigue manufacturing. Experimental results indicated that the impact order of the characterization parameters of machined surface integrity on the high-temperature low-cycle fatigue life were the degree of work hardening RHV, the residual stress in the cutting speed direction S22, the fatigue stress concentration factor Kf, the degree of grain refinement RD and the residual stress in the feed direction S33. In the range of turning parameters of the experiments in this research, the cutting speeds could be 80~110 m/min, and the feed rate could be 0.10~0.12 mm/rev to achieve a longer high-temperature low-cycle fatigue life. The results can be used for guiding the fatigue-resistant manufacturing research of aeroengine superalloy turbine disks.

De-Epithelialized Heterotopic Tracheal Allografts without Immunosuppressants in Dogs: Long-Term Results for Cartilage Viability and Structural Integrity.

OBJECTIVES: Reconstruction of long segmental tracheal defects is difficult because no ideal tracheal substitutes are currently available. Tracheal allotransplantation maintains cartilage and epithelium viability but requires immunosuppression because of epithelial immunogenicity. We aimed to obtain an epithelium-decellularized allograft that maintains cartilage viability and to evaluate long-term outcomes of such allografts implanted on dog backs without immunosuppressants. METHODS: Twenty-five tracheas harvested from mongrel dogs were used to explore the period of epithelium decellularization by combined use of 1% sodium dodecyl sulfate and an organ preservation solution and to assess the chondrocyte viability and immunogenicity of the tracheas after decellularization. Sixteen epithelium-decellularized tracheal allografts and 10 fresh tracheal segments (6 cm long) were implanted in 26 beagles for durations of 10 days and 1, 3, 6, and 12 months. Macroscopic and microscopic examinations were used to evaluate the morphology, viability, and immune rejection of the allografts. Safranin-O staining was used to detect glycosaminoglycans. RESULTS: The epithelium disappeared after 24 hours of decellularization. At 72 hours, almost no nuclei remained in the mucosa, while the mean survival rate of chondrocytes was 88.1%. Histological analysis demonstrated that the allograft retained intact tracheal rings and viable cartilage after heterotopic implantation for 1 year, with no immunological rejection. There were no significant differences in the glycosaminoglycan contents among the implanted epithelium-decellularized allografts. CONCLUSIONS: Epithelium-decellularized tracheal allografts with chondrocyte viability can be achieved by combined use of a detergent and organ preservation solution, which showed satisfactory cartilage viability and structural integrity after long-term heterotopic transplantation. Further studies on orthotopic transplantation are needed to assess the feasibility of allografts in reconstructing long segmental tracheal defects.

Mechanical properties of de-epithelialized tracheal allografts.

BACKGROUND: Many studies have shown that mucosa-treated tracheas can be used for allotransplantation without the use of immunosuppressants. However, no literature has reported whether the transplanted trachea has normal biomechanics. In this study, fresh dog trachea was used as a control to determine the mechanical properties of allograft trachea after mucous membrane removal. METHODS: Cervical tracheas (~10 cm in length, consisting of the cartilage rings down to the tracheal carina) and the surrounding connective tissues were removed from 20 mixed-breed dogs and then divided into 4 groups with 5 samples in each group. The groups were defined as follows: group A, fresh canine tracheas without any treatment; group B, canine tracheas after sodium dodecyl sulphate (SDS) de-epithelialization; group C, canine tracheas implanted into the dorsal fascia of beagle dogs for 1 month after SDS de-epithelialization; group D, canine tracheas implanted into the dorsal fascia of beagle dogs for 6 months after SDS de-epithelialization. All heterotopically implanted beagle dogs were free of any immunosuppressive treatment. Uniaxial tensile (n=20) and compressive (n=20) experiments were performed on the 4 groups of trachea using a universal testing machine; the tensile load was recorded at 150% of the original tracheal length (50% displacement), and the compressive load was recorded at 50% of the initial anteroposterior diameter (50% displacement). The average difference in tensile and compressive loads between groups was compared by analysis of variance. RESULTS: No significant difference was found in the mean compressive strength among groups (group A =2.38±0.87 N, group B =3.33±1.03 N, group C =5.39±2.44 N, group D =3.04±2.12 N; P=0.07 for all groups). The mean tensile strength of group C was 9.68±0.86 N, resulting in a significant difference in tensile strength compared to group A (22.51±3.91 N) and B (24.24±7.46 N) (P<0.001). No significant difference in tensile strength was observed among groups A, B, and D. CONCLUSIONS: The de-epithelialized trachea showed no significant change in mechanical properties after 6 months of implantation.