Proteoglycan 4 (PRG4) treatment improves skin wound healing in a porcine model.

Proteoglycan 4 (PRG4) is a boundary lubricant originally identified in articular cartilage and has been since shown to have immunomodulation and antifibrotic properties. Previously, we have demonstrated that recombinant human (rh)PRG4 treatment accelerates auricular cartilage injury closure through an inhibition of the fibrotic response, and promotion of tissue regeneration in mice. The purpose of the current study was to examine the effects of rhPRG4 treatment (vs. a DMSO carried control) on full-thickness skin wound healing in a preclinical porcine model. Our findings suggest that while rhPRG4 did not significantly accelerate nor impede full-thickness skin wound closure, it did improve repair quality by decreasing molecular markers of fibrosis and increasing re-vascularization. We also demonstrated that rhPRG4 treatment increased dermal adipose tissue during the healing process specifically by retaining adipocytes in the wound area but did not inhibit lipolysis. Overall, the results of the current study have demonstrated that rhPRG4 acts as antifibrotic agent and regulates dermal adipose tissue during the healing processes resulting in a tissue with a trajectory that more resembles the native skin vs. a fibrotic patch. This study provides strong rationale to examine if rhPRG4 can improve regeneration in human wounds.

Influence of microarchitecture on stressed volume and mechanical fatigue behaviour of equine subchondral bone.

Fractures of the equine metacarpophalangeal (MCP) joint are among the most common and fatal injuries experienced by racehorses. These bone injuries are a direct result of repetitive, high intensity loading of the skeleton during racing and training and there is consensus that they represent a mechanical fatigue phenomenon. Existing work has found the fatigue life of bone to be strongly determined by bone microarchitecture and the resulting stressed volume (i.e., the volume of bone stressed above assumed yield). The purpose of this study was to quantify the influence of bone microarchitecture on the mechanical fatigue behaviour of equine subchondral bone from the MCP joint across a wide variety of sample types. Forty-eight subchondral bone samples were prepared from the third metacarpal (MC3) and proximal phalanx (P1) of 8 horses and subsequently imaged using high resolution micro-computed tomography (µCT) to quantify microarchitectural features of interest, including bone volume fraction, tissue mineral density, pore size, pore spacing, and pore number. Samples were cyclically loaded in compression to a stress of 70 MPa, and fatigue life was defined as the number of cycles until failure. Finite element models were created from the µCT images and used to quantify stressed volume. Based on the expected log point-wise predictive density, stressed volume was a strong predictor of fatigue life in both the MC3 and P1. A regional analysis indicated fatigue life was more strongly associated with bone volume fraction in the superficial (r2 = 0.32, p < 0.001) and middle (r2 = 0.70, p < 0.001) regions of the subchondral bone, indicating the prominent role that the cortical plate played in the fatigue resistance of equine subchondral bone. By improving our understanding of the variance in fatigue life measurements, this research helps clarify the underlying mechanisms of the mechanical fatigue process and provides a basic understanding of subchondral bone injuries in the equine fetlock joint.

Comparing Inertial Measurement Units to Markerless Video Analysis for Movement Symmetry in Quarter Horses.

BACKGROUND: With an increasing number of systems for quantifying lameness-related movement asymmetry, between-system comparisons under non-laboratory conditions are important for multi-centre or referral-level studies. This study compares an artificial intelligence video app to a validated inertial measurement unit (IMU) gait analysis system in a specific group of horses. METHODS: Twenty-two reining Quarter horses were equipped with nine body-mounted IMUs while being videoed with a smartphone app. Both systems quantified head and pelvic movement symmetry during in-hand trot (hard/soft ground) and on the lunge (left/right rein, soft ground). Proportional limits of agreement (pLoA) were established. RESULTS: Widths of pLoA were larger for head movement (29% to 50% in-hand; 22% to 38% on lunge) than for pelvic movement (13% to 24% in-hand; 14% to 24% on lunge). CONCLUSION: The between-system pLoAs exceed current "lameness thresholds" aimed at identifying the affected limb(s) in lame horses. They also exceed published limits of agreement for stride-matched data but are similar to repeatability values and "lameness thresholds" from "non-lame" horses. This is encouraging for multi-centre studies and referral-level veterinary practice. The narrower pLoA values for pelvic movement asymmetry are particularly encouraging, given the difficulty of grading hind limb lameness "by eye".

Fibroblast inflammatory priming determines regenerative versus fibrotic skin repair in reindeer.

Adult mammalian skin wounds heal by forming fibrotic scars. We report that full-thickness injuries of reindeer antler skin (velvet) regenerate, whereas back skin forms fibrotic scar. Single-cell multi-omics reveal that uninjured velvet fibroblasts resemble human fetal fibroblasts, whereas back skin fibroblasts express inflammatory mediators mimicking pro-fibrotic adult human and rodent fibroblasts. Consequently, injury elicits site-specific immune responses: back skin fibroblasts amplify myeloid infiltration and maturation during repair, whereas velvet fibroblasts adopt an immunosuppressive phenotype that restricts leukocyte recruitment and hastens immune resolution. Ectopic transplantation of velvet to scar-forming back skin is initially regenerative, but progressively transitions to a fibrotic phenotype akin to the scarless fetal-to-scar-forming transition reported in humans. Skin regeneration is diminished by intensifying, or enhanced by neutralizing, these pathologic fibroblast-immune interactions. Reindeer represent a powerful comparative model for interrogating divergent wound healing outcomes, and our results nominate decoupling of fibroblast-immune interactions as a promising approach to mitigate scar.

Application of an instructive hydrogel accelerates re-epithelialization of xenografted human skin wounds.

Poor quality (eg. excessive scarring) or delayed closure of skin wounds can have profound physical and pyschosocial effects on patients as well as pose an enormous economic burden on the healthcare system. An effective means of improving both the rate and quality of wound healing is needed for all patients suffering from skin injury. Despite wound care being a multi-billion-dollar industry, effective treatments aimed at rapidly restoring the skin barrier function or mitigating the severity of fibrotic scar remain elusive. Previously, a hydrogel conjugated angiopoietin-1 derived peptide (QHREDGS; Q-peptide) was shown to increase keratinocyte migration and improve wound healing in diabetic mice. Here, we evaluated the effect of this Q-Peptide Hydrogel on human skin wound healing using a mouse xenograft model. First, we confirmed that the Q-Peptide Hydrogel promoted the migration of adult human keratinocytes and modulated their cytokine profile in vitro. Next, utilizing our human to mouse split-thickness skin xenograft model, we found improved healing of wounded human epidermis following Q-Peptide Hydrogel treatment. Importantly, Q-Peptide Hydrogel treatment enhanced this wound re-epithelialization via increased keratinocyte migration and survival, rather than a sustained increase in proliferation. Overall, these data provide strong evidence that topical application of QHREDGS peptide-modified hydrogels results in accelerated wound closure that may lead to improved outcomes for patients.

Biomechanics of Wound Healing in an Equine Limb Model: Effect of Location and Treatment with a Peptide-Modified Collagen-Chitosan Hydrogel.

The equine distal limb wound healing model, characterized by delayed re-epithelialization and a fibroproliferative response to wounding similar to that observed in humans, is a valuable tool for the study of biomaterials poised for translation into both the veterinary and human medical markets. In the current study, we developed a novel method of biaxial biomechanical testing to assess the functional outcomes of healed wounds in a modified equine model and discovered significant functional and structural differences in both unwounded and injured skin at different locations on the distal limb that must be considered when using this model in future work. Namely, the medial skin was thicker and displayed earlier collagen engagement, medial wounds experienced a greater proportion of wound contraction during closure, and proximal wounds produced significantly more exuberant granulation tissue. Using this new knowledge of the equine model of aberrant wound healing, we then investigated the effect of a peptide-modified collagen-chitosan hydrogel on wound healing. Here, we found that a single treatment with the QHREDGS (glutamine-histidine-arginine-glutamic acid-aspartic acid-glycine-serine) peptide-modified hydrogel (Q-peptide hydrogel) resulted in a higher rate of wound closure and was able to modulate the biomechanical function toward a more compliant healed tissue without observable negative effects. Thus, we conclude that the use of a Q-peptide hydrogel provides a safe and effective means of improving the rate and quality of wound healing in a large animal model.

Transcriptional Profiling of the Adult Hair Follicle Mesenchyme Reveals R-spondin as a Novel Regulator of Dermal Progenitor Function.

The adult hair follicle (HF) undergoes successive regeneration driven by resident epithelial stem cells and neighboring mesenchyme. Recent work described the existence of HF dermal stem cells (hfDSCs), but the genetic regulation of hfDSCs and their daughter cell lineages in HF regeneration remains unknown. Here we prospectively isolate functionally distinct mesenchymal compartment in the HF (dermal cup [DC; includes hfDSCs] and dermal papilla) and define the transcriptional programs involved in hfDSC function and acquisition of divergent mesenchymal fates. From this, we demonstrate cross-compartment mesenchymal signaling within the HF niche, whereby DP-derived R-spondins act to stimulate proliferation of both hfDSCs and epithelial progenitors during HF regeneration. Our findings describe unique transcriptional programs that underlie the functional heterogeneity among specialized fibroblasts within the adult HF and identify a novel regulator of mesenchymal progenitor function during tissue regeneration.

Dysfunction of Hair Follicle Mesenchymal Progenitors Contributes to Age-Associated Hair Loss.

Skin aging is accompanied by hair loss due to impairments in hair follicle (HF) epithelial progenitor cells and their mesenchymal niche. This inductive mesenchyme, called dermal papilla (DP), undergoes progressive cell loss and eventual miniaturization that contributes to HF pathogenesis. Using laser ablation and fate mapping, we show that HF dermal stem cells (hfDSCs) reconstitute the damaged DP and maintain hair growth, suggesting that hfDSC dysfunction may trigger degeneration of the inductive niche. Fate mapping over 24 months revealed progressive hfDSC depletion, and in vivo clonal analysis of aged hfDSCs showed impaired self-renewal and biased differentiation. Single-cell RNA-seq confirmed hfDSCs as a central precursor, giving rise to divergent mesenchymal trajectories. In aged skin, hfDSCs exhibited senescent-like characteristics, and senescence-associated secretory phenotypes were identified in the aging HF mesenchyme. These results clarify fibroblast dynamics within the HF and suggest that progressive dysfunction within the mesenchymal progenitor pool contributes to age-related hair loss.

Adult Human Dermal Progenitor Cell Transplantation Modulates the Functional Outcome of Split-Thickness Skin Xenografts.

Following full-thickness skin injuries, epithelialization of the wound is essential. The standard of care to achieve this wound "closure" in patients is autologous split-thickness skin grafting (STSG). However, patients living with STSGs report significant chronic impairments leading to functional deficiencies such as itch, altered sensation, fragility, hypertrophic scarring, and contractures. These features are attributable to the absence of functional dermis combined with the formation of disorganized fibrotic extracellular matrix. Recent work has demonstrated the existence of dermal progenitor cells (DPCs) residing within hair follicles that function to continuously regenerate mesenchymal tissue. The present work examines whether cultured DPCs could regenerate dermis within an STSG and improve overall graft function. Adult human DPCs were transplanted into a full-thickness skin wound in immune-compromised mice and closed with a human STSG. At 3 months, human DPCs (hDPCs) had successfully integrated into the xenograft and differentiated into various regionally specified phenotypes, improving both viscoelastic properties of the graft and mitigating pruritus.

Flowable Polyethylene Glycol Hydrogels Support the in Vitro Survival and Proliferation of Dermal Progenitor Cells in a Mechanically Dependent Manner.

Cell-based therapies have garnered considerable interest largely because of their potential utility for tissue regeneration in a variety of organs, including skin. Designing vehicles that enable optimal delivery and purposeful integration of donor cells within tissues will be critical for their success. Here, we investigate the utility of an injectable, self-polymerizing, fully synthetic hydrogel in supporting the survival, proliferation, and function of cultured adult dermal progenitor cells (DPCs) which may serve as a source of renewable cells to repair severe skin injuries or restore hair growth. We show that modifying the stiffness of these transglutaminase cross-linked poly(ethylene glycol) (TG-PEG) hydrogels significantly alters DPC behavior and phenotype; increasing stiffness promotes their differentiation and migration whereas softer gels maintained them in a proliferative state. We found that 2-3% TG-PEG was optimal to promote cell expansion and survival. Unexpectedly, DPCs grown in all conditions maintained their inductive function and thus generated de novo hair follicles. Our data suggests that TG-PEG hydrogels may be a versatile platform for stem and progenitor cell transplantation and fate specification while maintaining functional competence.

Matrix-Induced Autologous Chondrocyte Implantation (MACI) Using a Cell-Seeded Collagen Membrane Improves Cartilage Healing in the Equine Model.

BACKGROUND: Autologous chondrocyte implantation (ACI) using a collagen scaffold (matrix-induced ACI; MACI) is a next-generation approach to traditional ACI that provides the benefit of autologous cells and guided tissue regeneration using a biocompatible collagen scaffold. The MACI implant also has inherent advantages including surgical implantation via arthroscopy or miniarthrotomy, the elimination of periosteal harvest, and the use of tissue adhesive in lieu of sutures. This study evaluated the efficacy of the MACI implant in an equine full-thickness cartilage defect model at 1 year. METHODS: Autologous chondrocytes were seeded onto a collagen type-I/III membrane and implanted into one of two 15-mm defects in the femoral trochlear ridge of 24 horses. Control defects either were implanted with cell-free collagen type-I/III membrane (12 horses) or were left ungrafted as empty defects (12 horses). An additional 3 horses had both 15-mm defects remain empty as nonimplanted joints. The repair was scored by second-look arthroscopy (12 weeks), and necropsy examination (53 weeks). Healing was assessed by arthroscopic scoring, gross assessment, histology and immunohistology, cartilage matrix component assay, and gene expression determination. Toxicity was examined by prostaglandin E2 formation in joint fluid, and lymph node morphology combined with histologic screening of organs. RESULTS: MACI-implanted defects had improved gross healing and composite histologic scores, as well as increases in chondrocyte predominance, toluidine blue-stained matrix, and collagen type-II content compared with scaffold-only implanted or empty defects. There was minimal evidence of reaction to the implant in the synovial membrane (minor perivascular cuffing), subchondral bone, or cartilage. There were no adverse clinical effects, signs of organ toxicity, or evidence of chondrocytes or collagen type-I/III membrane in draining lymph nodes. CONCLUSIONS: The MACI implant appeared to improve cartilage healing in a critical-sized defect in the equine model compared with collagen matrix alone. CLINICAL RELEVANCE: These results indicate that the MACI implant is quick to insert, provides chondrocyte security in the defect, and improves cartilage healing compared with ACI.

Biocomposite nanofiber matrices to support ECM remodeling by human dermal progenitors and enhanced wound closure.

Cell-based therapies have recently been the focus of much research to enhance skin wound healing. An important challenge will be to develop vehicles for cell delivery that promote survival and uniform distribution of cells across the wound bed. These systems should be stiff enough to facilitate handling, whilst soft enough to limit damage to newly synthesized wound tissue and minimize patient discomfort. Herein, we developed several novel modifiable nanofibre scaffolds comprised of Poly (ε-caprolactone) (PCL) and gelatin (GE). We asked whether they could be used as a functional receptacle for adult human Skin-derived Precursor Cells (hSKPs) and how naked scaffolds impact endogenous skin wound healing. PCL and GE were electrospun in a single facile solvent to create composite scaffolds and displayed unique morphological and mechanical properties. After seeding with adult hSKPs, deposition of extracellular matrix proteins and sulphated glycosaminoglycans was found to be enhanced in composite grafts. Moreover, composite scaffolds exhibited significantly higher cell proliferation, greater cell spreading and integration within the nanofiber mats. Transplantation of acellular scaffolds into wounds revealed scaffolds exhibited improvement in dermal-epidermal thickness, axonal density and collagen deposition. These results demonstrate that PCL-based nanofiber scaffolds show promise as a cell delivery system for wound healing.

Enhanced Expansion and Sustained Inductive Function of Skin-Derived Precursor Cells in Computer-Controlled Stirred Suspension Bioreactors.

Endogenous dermal stem cells (DSCs) reside in the adult hair follicle mesenchyme and can be isolated and grown in vitro as self-renewing colonies called skin-derived precursors (SKPs). Following transplantation into skin, SKPs can generate new dermis and reconstitute the dermal papilla and connective tissue sheath, suggesting they could have important therapeutic value for the treatment of skin disease (alopecia) or injury. Controlled cell culture processes must be developed to efficiently and safely generate sufficient stem cell numbers for clinical use. Compared with static culture, stirred-suspension bioreactors generated fivefold greater expansion of viable SKPs. SKPs from each condition were able to repopulate the dermal stem cell niche within established hair follicles. Both conditions were also capable of inducing de novo hair follicle formation and exhibited bipotency, reconstituting the dermal papilla and connective tissue sheath, although the efficiency was significantly reduced in bioreactor-expanded SKPs compared with static conditions. We conclude that automated bioreactor processing could be used to efficiently generate large numbers of autologous DSCs while maintaining their inherent regenerative function. Stem Cells Translational Medicine 2017;6:434-443.

Mechanical characterization of matrix-induced autologous chondrocyte implantation (MACI®) grafts in an equine model at 53 weeks.

There has been much interest in using autologous chondrocytes in combination with scaffold materials to aid in cartilage repair. In the present study, a total of 27 animals were used to compare the performance of matrix-assisted chondrocyte implantation (MACI®) using a collagen sponge as a chondrocyte delivery vehicle, the sponge membrane alone, and empty controls. A total of three distinct types of mechanical analyses were performed on repaired cartilage harvested from horses after 53 weeks of implantation: (1) compressive behavior of samples to measure aggregate modulus (HA) and hydraulic permeability (k) in confined compression; (2) local and global shear modulus using confocal strain mapping; and (3) boundary friction coefficient using a custom-built tribometer. Cartilage defects receiving MACI® implants had equilibrium modulus values that were 70% of normal cartilage, and were not statistically different than normal tissue. Defects filled with Maix™ membrane alone or left empty were only 46% and 51-63% of control, respectively. The shear modulus of tissue from all groups of cartilage defects were between 4 and 10 times lower than control tissue, and range from 0.2 to 0.4 MPa. The average values of boundary mode friction coefficients of control tissue from all groups ranged from 0.42 to 0.52. This study represents an extensive characterization of the mechanical performance of the MACI® grafts implant in a large animal model at 53 weeks. Collectively, these data demonstrate a range of implant performance, revealing similar compressive and frictional properties to native tissue, with inferior shear properties.

Second carpal bone slab fracture and subluxation of the middle carpal joint in a horse subsequent to arthrodesis of the carpometacarpal joint.

OBJECTIVE: To report complications of arthrodesis of the carpometacarpal (CMC) joint using a drilling technique in an adult horse. STUDY DESIGN: Case report. ANIMALS: Horse (n = 1). METHOD: A 12-year-old Quarter Horse mare with CMC osteoarthritis (CMC-OA) had arthrodesis under general anesthesia in right lateral recumbency. Under fluoroscopic guidance, a 4.5 mm drill bit was inserted at 3 drilling sites 5-6 cm into the CMC joint and was fanned 30-45° in the plane of the joint and 5-10° in the long axis of the limb to destroy articular cartilage and expose the subchondral bone. RESULTS: The horse presented 2 weeks after surgery for severe lameness of the operated limb. A slab fracture of the 2nd carpal bone (C2) and subluxation of the middle carpal (MC) joint was diagnosed. The horse was humanely euthanatized due to poor prognosis. CONCLUSION: The fanning technique of arthrodesis of the CMC joint may lead to fracture of carpal bones, joint instability, and MC joint subluxation. A balance between articular surface destruction and maintenance of joint stability should be achieved when using this technique.

Mandibular condylectomy in a cow with a chronic luxation of the temporomandibular joint.

A cow, presented after being struck by a motor vehicle, continued to have difficulty eating after mandibular fracture repair. Imaging showed a temporomandibular luxation and a mandibular condylectomy was performed. Mastication improved greatly but the cow was euthanized due to infection. This is the first report of mandibular condylectomy in cattle.

Condylectomie mandibulaire chez une vache avec une luxation chronique de l'articulation temporomandibulaire. Une vache présentée après avoir été heurtée par un véhicule automobile, continuait d'éprouver de la difficulté à manger après la réparation d'une fracture mandibulaire. L'imagerie a indiqué une luxation temporo-mandibulaire et une condylectomie mandibulaire a été réalisée. La mastication s'est améliorée considérablement mais la vache a été euthanasiée en raison d'une infection. Il s'agit du premier rapport d'une condylectomie mandibulaire chez les bovins.(Traduit par Isabelle Vallières).

In vitro elution of amikacin and ticarcillin from a resorbable, self-setting, fiber reinforced calcium phosphate cement.

OBJECTIVE: To determine in vitro elution characteristics of amikacin and ticarcillin from fiber reinforced calcium phosphate beads (FRCP). SAMPLE POPULATION: Experimental. METHODS: FRCP beads with water (A), amikacin (B), ticarcillin/clavulanate (C), or both amikacin and ticarcillin/clavulanate (D) were bathed in mL phosphate-buffered saline (PBS) at 37°C, 5% CO(2) and 95% room air. PBS was sampled (eluent) and beads were placed in fresh PBS at time points 1 and 8 hours and 1, 2, 3, 4, 5, 6, 7, 10, 12, 14, 18, 21, 25, 28, 35, 42, 49, and 56 days. Antibiotic concentration and antimicrobial activity of eluent against Escherichia coli, Staphylococcus aureus, and Klebsiella pneumoniae were determined. RESULTS: Both antibiotics eluted in a bimodal pattern. Beads with a single antibiotic eluted 20.8 ± 2.5% of amikacin and 29.5 ± 0.8% of ticarcillin over 56 days. Coelution of the antibiotics resulted in a lower proportion (AUC(0-∞) ) of antibiotics eluted for both amikacin (9.5 ± 0.2%) and ticarcillin (21.7 ± 0.09%). Bioassay of antimicrobial activity of the eluent (t = 1, 8, and 24 hours) established reduced antimicrobial activity of amikacin from combination beads (D). CONCLUSIONS: FRCP beads with amikacin or ticarcillin/clavulanate, but not the combination, are suitable carriers for wound implantation. CLINICAL RELEVANCE: Duration before complete resorption of FRCP beads in vivo should be determined before clinical use as a resorbable depot. The results of this study underscore the importance of testing drug combinations, despite success of the combination systemically, before their use in local applications.

Reattachment of the articular cartilage component of type 1 subchondral cystic lesions of the medial femoral condyle with polydioxanone pins in 3 horses.

CASE DESCRIPTION: 3 horses were referred for treatment of subchondral cystic lesions of 1 or both medial femoral condyles. CLINICAL FINDINGS: All horses had clinically apparent lameness confirmed to be due to a radiographically evident subchondral cystic lesion of the medial femoral condyle with a large articular component (> 15 mm) and shallow subchondral depth (< 10 mm). Arthroscopic assessment of affected cartilage revealed undulating cartilage with a relatively smooth surface and extensive residual perimeter attachment. TREATMENT AND OUTCOME: Resorbable polydioxanone pins were used arthroscopically to reattach the cartilage overlying the subchondral cystic lesions. A biologic graft (bone marrow aspirate concentrate or allogeneic chondrocytes) was injected into the depths of the cystic cavity following cartilage reattachment. Follow-up examination confirmed radiographic resolution of the lesion and elimination of clinical signs within the treated femorotibial joint. CLINICAL RELEVANCE: Lesions with a large area of affected articular cartilage have been associated with a decreased rate of return to athletic function following arthroscopic debridement, likely secondary to the loss of subchondral architecture and the production of imperfect fibrocartilage repair. Salvage of the affected cartilage in a select population of horses with progressively expanding but shallow subchondral cystic lesions of the medial femoral condyle is possible and may improve radiographic and clinical outcome.

Partial resection of the plica salpingopharyngeus for the treatment of three foals with bilateral tympany of the auditory tube diverticulum (guttural pouch).

CASE DESCRIPTION: 3 foals were referred for treatment of tympany of the auditory tube diverticulum (guttural pouch). CLINICAL FINDINGS: Bilateral guttural pouch tympany was diagnosed in all 3 foals on the basis of clinical signs and results of radiographic and endoscopic evaluations and gutturocentesis. In each foal, previous medical or surgical interventions or both had failed to correct the problem. TREATMENT AND OUTCOME: Bilateral surgical resection of the plica salpingopharyngeus was performed. Follow-up telephone interviews with owners 8 months to 3 years after surgery revealed that long-term resolution of the tympany had been achieved in each foal, with few to no postsurgical complications. CLINICAL RELEVANCE: Surgical correction of refractory bilateral guttural pouch tympany can be accomplished by resection of the plica salpingopharyngeus within the pharyngeal opening of the Eustachian tube. In the foals of this report, permanent resolution of tympany was apparently achieved without interfering with upper airway function, providing support for the use of surgical resection of the plica salpingopharyngeus in the treatment of bilateral guttural pouch tympany.