The Dorsal Skinfold Chamber as a New Tympanic Membrane Wound Healing Model: Intravital Insights into the Pathophysiology of Epithelialized Wounds.

BACKGROUND: Tympanic membrane perforations (TMPs) are a common complication of trauma and infection. Persisting perforations result from the unique location of the tympanic membrane. The wound is surrounded by air of the middle ear and the external auditory canal. The inadequate wound bed, growth factor, and blood supply lead to circular epithelialization of the perforation's edge and premature interruption of defect closure. Orthotopic animal models use mechanical or chemical tympanic membrane laceration to identify bioactive wound dressings and overcome premature epithelialization. However, all orthotopic models essentially lack repetitive visualization of the biomaterial-wound interface. Therefore, recent progress in 3D printing of customized wound dressings has not yet been transferred to the unique wound setup of the TMP. Here, we present a novel application for the mice dorsal skinfold chamber (DSC) with an epithelialized full-thickness defect as TMP model. METHODS: A circular 2-mm defect was cut into the extended dorsal skinfold using a biopsy punch. The skinfold was either perforated through both skin layers without prior preparation or perforated on 1 side, following resection of the opposing skin layer. In both groups, the wound was sealed with a coverslip or left unclosed (n = 4). All animals were examined for epithelialization of the edge (histology), size of the perforation (planimetry), neovascularization (repetitive intravital fluorescence microscopy), and inflammation (immunohistology). RESULTS: The edge of the perforation was overgrown by the cornified squamous epithelium in all pre-parations. Reduction in the perforation's size was enhanced by application of a coverslip. Microsurgical preparation before biopsy punch perforation and sealing with a coverslip enabled repetitive high-quality intravital fluorescence microscopy. However, spontaneous reduction of the perforation occurred frequently. Therefore, the direct biopsy punch perforation without microsurgical preparation was favorable: spontaneous reduction did not occur throughout 21 days. Moreover, the visualization of the neovascularization was sufficient in intravital microscopy. CONCLUSIONS: The DSC full-thickness defect is a valuable supplement to orthotopic TMP models. Repetitive intravital microscopy of the epithelialized edge enables investigation of the underlying pathophysiology during the transition from the inflammation to the proliferation phase of wound healing. Using established analysis procedures, the present model provides an effective platform for the screening of bioactive materials and transferring progress in tissue engineering to the special conditions of tympanic membrane wound healing.

The possible mechanism of Hippophae fructus oil applied in tympanic membrane repair identified based on network pharmacology and molecular docking.

OBJECTIVE: This study aimed to explore the mechanisms of Hippophae fructus oil (HFO) in the treatment of tympanic membrane (TM) perforation through network pharmacology-based identification. METHODS: The compounds and related targets of HFO were extracted from the TCMSP database, and disease information was obtained from the OMIM, GeneCards, PharmGkb, TTD, and DrugBank databases. A Venn diagram was generated to show the common targets of HFO and TM, and GO and KEGG analyses were performed to explore the potential biological processes and signaling pathways. The PPI network and core gene subnetwork were constructed using the STRING database and Cytoscape software. A molecular docking analysis was also conducted to simulate the combination of compounds and gene proteins. RESULTS: A total of 33 compounds and their related targets were obtained from the TCMSP database. After screening the 393 TM-related targets, 21 compounds and 22 gene proteins were selected to establish the network diagram. GO and KEGG enrichment analyses revealed that HFO may promote TM healing by influencing cellular oxidative stress and related signaling pathways. A critical subnetwork was obtained by analyzing the PPI network with nine core genes: CASP3, MMP2, IL1B, TP53, EGFR, CXCL8, ESR1, PTGS2, and IL6. In addition, a molecular docking analysis revealed that quercetin strongly binds the core proteins. CONCLUSION: According to the analysis, HFO can be utilized to repair perforations by influencing cellular oxidative stress. Quercetin is one of the active compounds that potentially plays an important role in TM regeneration by influencing 17 gene proteins.

L1CAM-ILK-YAP Mechanotransduction Drives Proliferative Activity of Epithelial Cells in Middle Ear Cholesteatoma.

Middle-ear cholesteatoma (cholesteatoma) is a chronic otitis media with an enhanced proliferation of epithelial cells. Negative pressure in the middle ear is thought to be important for the etiology of cholesteatoma. However, the mechanism of cholesteatoma formation remains unclear. Integrin-linked protein kinase (ILK), an important modulator of actin cytoskeletal dynamics, interacts with extracellular matrix and results in the up-regulation of mechanotransduction effector Yes-associated protein (YAP). The L1 cell adhesion molecule (L1CAM) has recently been reported as an activator of the mechanotransduction effectors related to cell proliferation and migration. In this study, we demonstrated a stretch assay for middle-ear cultured cells and performed immunohistochemistry using antibodies against Ilk, Yap, and L1cam. The tympanic membrane was also analyzed within a new middle-ear negative-pressure animal model and human cholesteatoma tissues, using immunohistochemistry with antibodies against ILK, YAP, Ki-67, and L1CAM. The expression of cytoplasmic ILK and nuclear shift of YAP increased in the thickened epithelium of the tympanic membrane under a negative-pressure load and the cholesteatoma. The expression of L1CAM was detected in the stromal cells, which enhanced epithelial cell proliferation depending on ILK signaling events. In conclusion, we demonstrated the possibility that the stromal L1CAM and epithelial ILK-YAP signaling played an important role in epithelial growth under mechanotransduction in cholesteatoma formation.

The Possible Role of Sclerostin in the Pathogenesis of Tympanosclerosis.

OBJECTIVE: The aim of this study was to investigate sclerostin (SOST) expression in a rat model of experimental tympanosclerosis (TS) and its possible role in the formation of TS. MATERIALS AND METHODS: Thirty-four SD rats were randomly divided into 2 groups: experimental group (n = 17) and normal group (n = 17). The left tympanic cavities in the experimental group were inoculated with methicillin-resistant Staphylococcus aureus. The changes of tympanic membranes were examined and recorded under otoendoscope. Haematoxylin-eosin staining was adopted to detect the morphological changes in the tympanic membrane and middle ear mucosa. Immunohistochemistry and Western blot analysis were used to observe the expression of SOST, Wnt3a, ß-catenin, and P-ERK1/2. RESULTS: In the experimental group, sclerotic lesions were observed in 54.5% ears in the end of 6 weeks. Morphological changes such as mucosa incrassation, inflammatory cells infiltration, fibrous tissue proliferation, and interstitial tissue incrassation prominently appeared in the tympanic membrane and middle ear mucosa. SOST protein was mainly distributed in the cytoplasm of epithelial cells and gland cells, the expression of which increased significantly in the calcified experimental ears. In addition, expression levels of Wnt3a, ß-catenin, and P-ERK1/2 increased significantly in the calcified group too. CONCLUSION: The upregulated expression level of SOST may be involved in the formation of TS, first, through the pro-phosphorylation of ERK1/2 in the inflammatory stage, and then through the enhancement of Wnt3a in the osteogenic stage.

Differing contributions of the first and second pharyngeal arches to tympanic membrane formation in the mouse and chick.

We have proposed that independent origins of the tympanic membrane (TM), consisting of the external auditory meatus (EAM) and first pharyngeal pouch, are linked with distinctive middle ear structures in terms of dorsal-ventral patterning of the pharyngeal arches during amniote evolution. However, previous studies have suggested that the first pharyngeal arch (PA1) is crucial for TM formation in both mouse and chick. In this study, we compare TM formation along the anterior-posterior axis in these animals using Hoxa2 expression as a marker of the second pharyngeal arch (PA2). In chick, the EAM begins to invaginate at the surface ectoderm of PA2, not at the first pharyngeal cleft, and the entire TM forms in PA2. Chick-quail chimera that have lost PA2 and duplicated PA1 suggest that TM formation is achieved by developmental interaction between a portion of the EAM and the columella auris in PA2, and that PA1 also contributes to formation of the remaining part of the EAM. By contrast, in mouse, TM formation is highly associated with an interdependent relationship between the EAM and tympanic ring in PA1.

Identification of early inflammatory changes in the tympanic membrane with Raman spectroscopy.

The tympanic membrane (TM) is a dynamic structure that separates the middle ear from the external auditory canal. It is also integral for the transmission of sound waves. In this study, we demonstrate the feasibility of using Raman spectroscopy to identify early chemical changes resulting from inflammation in the TM that can serve as an indicator of acute otitis media. Bacterial lipopolysaccharide (LPS) was injected trans-tympanicaly in a murine model. Presence of inflammatory response was assessed with binocular microscopy, confirmed with histopathology and immunofluorescence staining. Successful discrimination suggesting spectral differences among the control and LPS treated groups was achieved using principal component analysis. Raman imaging revealed major differences in collagen distribution and nucleic acid content. Image segmentation analysis on the trichrome stained tissue sections was performed to corroborate the Raman spectra. The spectral co-localization study suggests changes in the expression of collagen IV specific signals in LPS treated samples. The overall findings of the study support prospective application of RS in the diagnosis and therapeutic monitoring of otitis media.

Sustained release of triamcinolone acetonide from an intratympanically applied hydrogel designed for the delivery of high glucocorticoid doses.

The pharmacokinetic properties and tolerability of a triamcinolone acetonide poloxamer 407 hydrogel for intratympanic application were investigated in a guinea pig model. Evaluation of in vivo release kinetics showed very high initial perilymph drug levels, with clinically relevant levels present for a minimum of 10 days. Assessment of auditory brainstem response thresholds showed a minimal, delayed and transient threshold shift, which was apparent on day 3 and resolved by day 10. No relevant histological changes of the middle and inner ear structures were noted, and hair cell counts showed no significant differences between treated and untreated ears. Thus, the triamcinolone-acetonide-loaded poloxamer 407 hydrogel is an effective vehicle for sustained high-dose inner ear glucocorticoid delivery.

TGF-α/HA complex promotes tympanic membrane keratinocyte migration and proliferation via ErbB1 receptor.

Tympanic membrane perforations are common and represent a management challenge to clinicians. Current treatments for chronic perforations involve a graft surgery and require general anaesthesia, including associated costs and morbidities. Bioactive molecules (e.g. growth factors, cytokines) play an important role in promoting TM wound healing following perforation and the use of growth factors as a topical treatment for tympanic membrane perforations has been suggested as an alternative to surgery. However, the choice of bioactive molecules best suited to promote wound healing has yet to be identified. We investigated the effects of hyaluronic acid, vitronectin, TGF-α, IL-24 and their combinations on migration, proliferation and adhesion of cultured human tympanic membrane-derived keratinocytes (hTM), in addition to their possible mechanisms of action. We found that TGF-α, TGF-α/HA and TGF-α/IL-24 promoted wound healing by significantly increasing both migration and proliferation. TGF-α and/or HA treated cells showed comparable cell-cell adhesion whilst maintaining an epithelial cell phenotype. With the use of receptor binding inhibitors for ErbB1 (AG1478) and CD44 (BRIC235), we revealed that the activation of ErbB1 is required for TGF-α/HA-mediated migration and proliferation. These results suggest factors that may be incorporated into a tissue-engineered membrane or directly as topical treatment for tympanic membrane perforations and hence reduce the need for a surgery.

[Mastoidectomy with reconstruction of the posterior canal wall and obliteration preventing postoperative morbidity].

INTRODUCTION: A cholesteatoma in the mastoid or in the middle ear presents a hazard to the well-being of patients. Commonly used surgical interventions are not an ideal solution as they bear with them postoperative morbidity such as the need for water precautions, a high rate of cholesteatoma recurrence and the inability to undergo hearing rehabilitation. METHODS: Forty-five patients underwent an innovative surgical procedure that enables complete removal of the cholesteatoma, preservation of ear anatomy and hearing restoration. Our series was divided into two groups. The first group comprised those in whom this innovative procedure was the first one and the posterior bony canal was preserved (primary surgery). The second group comprised those in whom the bony wall had been removed previously during surgery (secondary surgery). RESULTS: In the first group, which included 29 patients, the middle ear cavity was found to be aerated in 69% of the patients, the tympanic membrane was intact in 93% and the rate of cholesteatoma recurrence was 10.3%. In the second group, which included 16 patients, the middle ear cavity was found to be aerated in 56.2% of cases, the tympanic membrane was intact in 75% and the rate of cholesteatoma recurrence was 25%. DISCUSSION: Mastoidectomy reconstruction of the posterior wall and obliteration (MAPRO) was found to be an effective approach for completely removing a choLesteatoma and preventing cholesteatoma recurrence. It is water-safe and provides an excellent basis for hearing restoration. The use of the original posterior bony canal for middle ear reconstruction was found to be beneficial. The authors advise an MRI study 18 months after surgery for cholesteatoma detection.

Localization of progenitor/stem cells in the human tympanic membrane.

BACKGROUND: The outer epithelium of the tympanic membrane (TM) initiates the closure of a perforation. Embryonic stem cells have been used in attempts to enhance the healing capacity of induced perforations in experimental animals. More knowledge about epithelial cell proliferation and cell migration is needed for a better understanding of the TM healing process. This includes the identification of regenerative zones within the outer epithelial layer of the TM where progenitor cells may be present. METHODS: Normal human TMs from translabyrinthine surgery were investigated using immunohistochemistry and immunofluorescence to detect the progenitor/stem cell markers α6-integrin, ß1-integrin and cytokeratin 19 (CK19). RESULTS: α6-Integrin was detected in the basal layer of the keratinizing epithelium in the umbo, in the annular region and along the malleus but not in the intermediate portion of the pars tensa. ß1-Integrin and CK19 were found in the same locations not only in the basal layer but also in the suprabasal layers of the keratinizing epithelium. CONCLUSIONS: Possible progenitor cells are found in the umbo, the annular region and along the malleus. Further studies are needed to identify the source of these cells.

CO(2) gas exchange across the human tympanic membrane is not appreciably affected by pathology.

Past in vivo studies in humans showed that the tympanic membrane (TM) is permeable to physiological gases. Animal studies show that transTM CO(2) conductance is increased by TM pathology. The objective of the study was to determine if transTM CO(2) exchange in humans is affected by atrophic and sclerotic pathologies. The study used an ear canal (EC) probe (ECP) constructed from a custom-fitted acrylic body, a glass capillary tube enclosing an oil meniscus to maintain ambient ECP + EC pressure and a silica glass microtube linked to a mass spectrometer (MS) for measuring gas composition that was hermetically sealed within the ear canal of the test ear. ECP + EC volume was measured and gas samples taken at 10 min intervals for 1 h. The fractional CO(2) pressure measured in the ECP + EC for each sample was regressed on time and the slope of the function multiplied by the ECP + EC volume and divided by the estimated transTM CO(2) gradient at the start of the experiment to yield transTM CO(2) conductance (microL/min/Pa). Data were complete for 15 normal, 13 sclerotic and 9 atrophic TMs. The average (+std) transTM CO(2) conductances were 1.76 × 10(-4) + 7.27 × 10(-5), 2.26 × 10(-4) + 1.5 × 10(-4) and 2.36 × 10(-4) + 1.14 × 10(-4) microL/min/Pa/TM for the normal, sclerotic and atrophic TMs, respectively. A pairwise comparison of data for the normal and atrophic TMs under the directional hypothesis of a greater CO(2) exchange rate for thinner TMs approached statistical significance (P = 0.07). A similar pairwise comparison for the sclerotic and normal TMs did not approach statistical significance (P = 0.28). The effect of TM pathologies on CO(2) conductance was limited.

Development of thermosensitive poloxamer 407-based microbubble gel with ultrasound mediation for inner ear drug delivery.

ABSTRACTSOur previous study first investigated feasibility of applying ultrasound (US) and microbubbles (MBs) via external auditory canal to facilitate drug delivery into inner ear. However, most drugs are in aqueous formulae and eliminated via Eustachian tubes after drug application. In this study, feasibility of sustained release of thermosensitive poloxamer 407 (P407)-based MB gel for US mediation-enhanced inner ear drug (dexamethasone, DEX) delivery was investigated. The sol-to-gel transition temperature showed that mixture of DEX and only 10% and 12.5% P407 in MBs can be used for in vitro and in vivo drug delivery experiments. In in vitro Franz diffusion experiments, the release rates of 12.5% P407-MBs + US groups in the model using DEX as the delivered reagent at 3 h resulted in values 1.52 times greater than those of 12.5% P407-MBs groups. In guinea pigs, by filling tympanic bulla with DEX in 12.5% P407-MBs (DEX-P407-MBs), USMB applied at post-treatment days 1 and 7 induced 109.13% and 66.67% increases in DEX delivery efficiencies, respectively, compared to the group without US. On the 28th day after US-mediated P407-MB treatment, the safety assessment showed no significant changes in the hearing thresholds and no damage to the integrity of cochlea or middle ear. These are the first results to demonstrate feasibility of US-modified liquid form DEX-P407-MB cavitation for enhancing permeability of round window membrane. Then, a gel form of DEX-P407-MBs was generated and thus prolonged the release of DEX in middle ear to maintain the therapeutic DEX level in inner ear for at least 7 days.

The importance of vascular endothelial growth factor in the healing of acute tympanic membrane perforation.

PURPOSE: The purpose of this study was to determine the more important growth factor expression between basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) in the healing of acute tympanic membrane (TM) perforation. MATERIALS AND METHODS: Bilateral perforations of the TM were created in 12 rats. The TM perforations in the right ears were treated with dexamethasone, and left ears were designated as the control group. The TM was examined for the growth factor expression immunohistochemically in the epithelial and fibrous layers according to the rate of TM perforation healing. RESULTS: The mean spontaneous healing time of the TM perforations was 11.0 +/- 2.0 days. However, dexamethasone-treated group showed no evidence of closure. The bFGF and VEGF expression were significantly correlated with the rate of healing of acute TM perforations. The VEGF expression was decreased both in the epithelial and fibrous layers, but bFGF expression was decreased only in the epithelial layer in the dexamethasone-treated group. The VEGF was expressed to a lesser degree than bFGF in the dexamethasone-treated group. CONCLUSION: Vascular endothelial growth factor is the more specific and important growth factor than bFGF in the healing of acute TM perforation.

Congenital cholesteatoma associated with blue eardrum.

The classic presentation of congenital cholesteatoma is a pearl behind the anterior-superior quadrant of an intact tympanic membrane. Idiopathic hemotympanum is characterized by a dark blue eardrum, the most prominent feature of which is the presence of cholesterol granulomas. Blue eardrum is associated with eustachian tube dysfunction. Despite the well-established relationship between eustachian tube dysfunction and the development of pediatric cholesteatoma, little has been written concerning the appropriate timing of tympanostomy tube placement. To date, there are no reports of congenital cholesteatoma associated with blue eardrum. A recent case of advanced congenital cholesteatoma (stage IV) associated with blue eardrum was treated using preoperative tympanostomy tube insertion. Tympanostomy tubes were helpful in preventing recurrence of the cholesteatoma after surgery. The case is presented along with a review of the literature.

[Use of forced-air to prevent intraoperative hypothermia]. / Uso da manta térmica na prevenção da hipotermia intraoperatória.

OBJECTIVE: Hypothermia is a life-threatening event during the perioperative period. No consensus has been reached about the best active warming approach for such cases. Furthermore there is no consensus on the most appropriate time to warm a hypothermic patient. This study aimed to assess the efficacy of a forced-air blanket to warm patients at 38 degrees C before and during surgery. Following utilization of the forced-air blanket, adverse effects were evaluated. METHODS: Patients submitted to orthopedic surgeries were divided into four groups of 15 patients. In the control group (Gcont), patients were not warmed with a forced-air blanket. In the preoperative group (Gpre), intraoperative group (Gintra), and total group (Gtotal), patients were warmed at 38 degrees C, during 30 minutes before anesthetic induction, after anesthetic induction up to 120 minutes and before and after the induction, respectively. Parameters evaluated were central (tympanic) temperature, peripheral (skin) temperature, operating room temperature, variations in the hemodynamic conditions and warming-induced adverse effects. RESULTS: Only Gtotal did not show significant variation in central temperature. Central temperatures of Gtotal patients were significantly higher (p <0.05) than those of other groups at 60 and 120 min after induction. In Gcont, Gpre and Gintra, patients were hypothermic at 60 min. CONCLUSION: The forced-air blanket is effective to prevent intraoperative hypothermia when applied for a period ranging from 30 min before anesthetic induction to 120 min after anesthetic induction. In the conditions of this study, adverse effects were not observed.

Relationship of T lymphocytes, cytokines, immunoglobulin E and nitric oxide with otitis media with effusion in children and their clinical significances.

OBJECTIVE: To investigate the relations of T lymphocytes, cytokines, immunoglobulin E, and nitric oxide with otitis media with effusion (OME) in children and their clinical significances. METHODS: Fifty children with OME treated in our hospital were enrolled in the study (observation group). Fifty healthy children were selected as control. The percentages of CD4+ and CD8+ T lymphocyte and CD4+/CD8+ ratio in peripheral blood, and the levels of cytokine (IL)-2, IL-4, IL-6, immunoglobulin E (IgE) and nitric oxide (NO) in peripheral blood and middle ear effusion (MEE) in both groups were detected. The correlations of these indexes with OME were analyzed. RESULTS: The percentage of peripheral blood CD4+ and CD8+ levels, CD4+/CD8 ratio, IgE, and NO levels in the observation group were significantly higher than those in the control group (P < 0.01). In the observation group, the IL-2 and IL-6 levels, and IgE and NO levels in the MEE were significantly higher than those in peripheral blood (P < 0.01). In addition, in the observation group, the MEE IL-2 and IL-6 levels were positively correlated with peripheral blood CD4+/CD8+ ratio, respectively r = 0.366, P = 0.009; r = 0.334, P = 0.018. CONCLUSIONS: The levels of peripheral blood CD4+ and CD8+ lymphocytes and MEE IL-2, IL-6, IgE, and NO levels are increased in children with OME. These indexes have provided significant clues for the diagnosis of OME in children.

Immunohistochemical analysis of retraction pocket pars tensa of tympanic membrane in children.

AIMS: Immunohistochemical analysis of retraction pocket pars tensa of tympanic membrane in children. Identification of signs typical for cholesteatoma and support of retraction theory of cholesteatoma. STUDY DESIGN: a prospective study analysing 31 surgically removed retraction pockets. DEPARTMENT: University Hospital, Children's Medical Centre Methods: Retraction pockets processed by a standard process for immunohistochemical analysis. The observed findings were specified using antibodies CD45 LCA (leukocyte common antigen), CD31 (platelet endothelial cell adhesion molecule), D2-40 (marker of lymphatic endothelium), MMP9 (marker of degradation of connective tissue extracellular matrix) and Ki67 (cellular marker of proliferation). RESULTS: All observed parameters except for MMP9 had a significantly higher incidence in retraction pocket stage III compared to stage II according to Charachon. CONCLUSION: We described immunohistochemical signs of retraction pocket pars tensa of tympanic membrane in children resulting in cholesteatoma. All the observed signs occur in the structure of matrix and perimatrix of cholesteatoma. A significantly higher incidence of all observed parameters except from MMP9 was proved in retraction pocket stage III, unlike in stage II. This observation proves the fact that retraction pocket is a progressive disease and is a procholesteatoma stage.

Tympanic membrane changes in experimental acute otitis media and myringotomy.

OBJECTIVE: The present experimental study explored pathomorphological changes and calcium depositions in the tympanic membrane during experimental acute otitis media caused by nontypeable Haemophilus influenzae in myringotomized and nonmyringotomized ears. MATERIAL AND METHODS: A rat model of experimental acute otitis media caused by nontypeable Haemophilus influenzae was employed. Sixteen Sprague-Dawley rats were used. Four days following middle ear inoculation, a bilateral myringotomy was performed in six randomly selected animals. Another group of 10 animals was inoculated only. On days 4, 7, 14, and 28 after inoculation, two animals from each group were sacrificed. The temporal bones were removed and the tympanic membranes were dissected, followed by paraffin embedding. Adjacent sections were stained with PAS-alcian blue for basic histopathological observations and by von Kossa method for determination of calcium phosphate depositions. RESULTS: Particularly intense invasion of polymorphonuclear neutrophil leukocytes was seen on day 4 after inoculation. The highest infiltration of macrophages was observed on day 7. The peak number of lymphocytes was seen on day 14. No difference occurred in the number of polymorphonuclear leukocytes in myringotomized and nonmyringotomized tympanic membranes. The infiltration with lymphocytes and activated macrophages in all parts of the myringotomized tympanic membranes was statistically significantly higher than in the nonmyringotomized animals. The total amount of interstitial calcium phosphate depositions during days 7, 14, and 28 of study was statistically higher in the sections of pars tensa from myringotomized membranes compared to the nonmyringotomized membranes. CONCLUSION: Nontypeable Haemophilus influenzae-induced acute otitis media and myringotomy provoke more extensive inflammatory reaction with microcalcification in the tympanic membranes.

A new minimally invasive method for the transtubal, microendoscopic application of fluids to the middle ear.

The Eustachian (or auditory) tube is of central importance for the regulation of ambient air pressure changes within the middle ear spaces. Dysfunction of the Eustachian tube usually leads to chronic inflammatory changes of the middle ear. The aim of the present feasibility study was to investigate an alternative, minimally invasive approach for the application of fluids to the middle ear via the Eustachian tube. This so-called transtubal application (TTA) was conducted in a prospective, non-randomized study with a total of ten subjects. The TTA approach consisted of placing a microendoscope within the Eustachian tube under local anaesthesia via its epipharyngeal opening. Subsequently, fluids were applied through an additional working channel after microendoscopic evaluation. Therefore the subjects were positioned supine-laterally and had to swallow actively. The successful fluid application into the middle ear was evidenced by microendoscopy of the tympanic membrane (visualization of the fluid level). In all cases, a successful application could be evidenced. Side effects (e.g. pain, mucosal injuries, microbleedings) were not observed. This new technique (TTA) offers the opportunity of a minimally invasive approach to treat tubal dysfunction and possibly other middle ear diseases by local fluid and/or drug application.

Permeation Enhancers for Intratympanically-applied Drugs Studied Using Fluorescent Dexamethasone as a Marker.

HYPOTHESIS: Entry of locally applied drugs into the inner ear can be enhanced by chemical manipulations. BACKGROUND: Perilymph drug concentrations achieved by intratympanic applications are well below the applied concentration due to limited entry through the round window (RW) membrane and stapes. Chemical manipulations to increase entry permeability could increase the effectiveness of drug therapy with local applications. METHODS: Dexamethasone-fluorescein (F-dex) was used as an entry marker. F-dex was applied to the RW niche of guinea pigs as a 20 µL bolus of 1 mM solution. After a 1 hour application, 10 samples of perilymph were collected sequentially from the lateral semicircular canal, allowing F-dex distribution throughout the perilymph to be quantified. Entry was also measured with the applied solution additionally containing dimethyl sulfoxide (DMSO), N-methylpyrrolidone (NMP), saponin, caprate, benzyl alcohol (BA) or poloxamer 407 (P407). Combinations of saponin or BA with P407 were also compared. RESULTS: In control experiments, F-dex entered the inner ear slowly at both the RW and stapes. The total F-dex recovered in all 10 samples from each animal averaged 2.1 pMoles for controls, 1.71 pMoles for 17% P407, 3.70 pMoles for caprate, 8.04 pMoles for DMSO, 16.32 pMoles for NMP, 31.0 pMoles for saponin, and 67.3 pMoles for 4% BA. Entry with DMSO, NMP, saponin and 4% BA were all significantly higher than the controls (one-way ANOVA). CONCLUSION: These studies confirm that entry of drugs into the ear can be markedly enhanced with the use of chemical permeation-enhancing agents.