Hesperidin activates Nrf2 to protect cochlear hair cells from cisplatin-induced damage.

Cisplatin is widely employed in clinical oncology as an anticancer chemotherapy drug in clinical practice and is known for its severe ototoxic side effects. Prior research indicates that the accumulation of reactive oxygen species (ROS) plays a pivotal role in cisplatin's inner ear toxicity. Hesperidin is a flavanone glycoside extracted from citrus fruits that has anti-inflammatory and antioxidant effects. Nonetheless, the specific pharmacological actions of hesperidin in alleviating cisplatin-induced ototoxicity remain elusive. The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) is a critical mediator of the cellular oxidative stress response, is influenced by hesperidin. Activation of Nrf2 was shown to have a protective effect against cisplatin-induced ototoxicity. The potential of hesperidin to stimulate Nrf2 in attenuating cisplatin's adverse effects on the inner ear warrants further investigation. This study employs both in vivo and in vitro models of cisplatin ototoxicity to explore this possibility. Our results reveal that hesperidin mitigates cisplatin-induced ototoxicity by activating the Nrf2/NQO1 pathway in sensory hair cells, thereby reducing ROS accumulation, preventing hair cell apoptosis, and alleviating hearing loss.

Gram-negative Bacteria are Associated With Sensorineural Hearing Loss in Chronic Otitis Media.

OBJECT: Chronic otitis media (COM) is an inflammatory disease that commonly presents with otorrhea and hearing loss. Bacteria-induced inflammation can cause inner ear damage, leading to sensorineural hearing loss (SNHL). This study aimed to compare the prevalence and severity of SNHL in patients with gram-negative versus gram-positive cultures and examine associations between the concentrations of circulating monocytes and neutrophils with bacteria species and SNHL. METHODS: This was a retrospective study. Cholesteatoma or chronic suppurative otitis media patients with otorrhea were enrolled. Middle ear secretions were collected using sterile swabs under an otoscope, and sent for bacterial detection within 30 min. Pure tone audiometry and circulating leukocyte counts were recorded and analyzed in patients infected with different pathogens. Logistic regression analysis was used to identify the risk factors associated with SNHL. RESULTS: A total of 137 patients were enrolled, including 45 patients infected with gram-negative bacteria, 41 with gram-positive bacteria, 20 with polymicrobial infection, and 31 with no bacterial growth. Logistic regression analysis showed that bacterial culture positive infections (OR = 7.265, 95% CI 2.219-23.786, p = 0.001) were an independent risk factor for SNHL. Patients with gram-negative bacteria had higher risks of SNHL (p < 0.0001) and more severe hearing loss (p = 0.005) than those with gram-positive bacteria. COM patients infected with gram-negative bacteria showed an increase in circulating monocytes, which correlated with the occurrence of SNHL (p = 0.0343). CONCLUSION: Gram-negative bacteria are associated with elevated circulating monocyte counts and have a higher risk of severe SNHL. LEVEL OF EVIDENCE: 4 Laryngoscope, 2024.

IGF2BP3-mediated enhanced stability of MYLK represses MSC adipogenesis and alleviates obesity and insulin resistance in HFD mice.

Mesenchymal stem cells (MSCs) hold immense potential as multipotent stem cells and serve as a primary source of adipocytes. The process of MSC adipogenesis plays a crucial role in maintaining systemic metabolic homeostasis and has garnered significant attention in tissue bioengineering. N6-methyladenosine (m6A), the most prevalent RNA modification, is known to regulate cell fate and disease. However, the precise involvement of m6A readers in MSC adipogenesis remains unclear. In this study, we investigated the impact of IGF2BP3, a prominent m6A reader, on MSC adipogenesis. Our findings revealed a decrease in IGF2BP3 expression during the natural adipogenic differentiation of MSCs. Furthermore, IGF2BP3 was found to repress MSC adipogenesis by augmenting the levels of MYLK, a calcium/calmodulin-dependent kinase. Mechanistically, IGF2BP3 interacted with MYLK mRNA in an m6A-dependent manner, extending its half-life and subsequently inhibiting the phosphorylation of the ERK1/2 pathway, thereby impeding the adipogenic differentiation of MSCs. Additionally, we successfully achieved the overexpression of IGF2BP3 through intraperitoneal injection of adeno-associated virus serotype Rec2, which specifically targeted adipose tissue. This intervention resulted in reduced body weight and improved insulin resistance in high-fat diet mice. Overall, our study provides novel insights into the role of IGF2BP3 in MSC adipogenesis, shedding light on adipocyte-related disorders and presenting potential targets for related biomedical applications.

Reactive Oxygen Species-Related Disruptions to Cochlear Hair Cell and Stria Vascularis Consequently Leading to Radiation-Induced Sensorineural Hearing Loss.

Aims: Radiation-induced sensorineural hearing loss (RISNHL) is one of the major side effects of radiotherapy for head and neck cancers. At present, no effective clinical treatment or prevention is available for RISNHL. This study thus aimed to investigate the cochlear pathology so that the underlying mechanisms of RISNHL may be elucidated, consequently paving the way for potential protective strategies to be developed. Results: Functional and morphological impairment in the stria vascularis (SV) was observed after irradiation (IR), as indicated by endocochlear potential (EP) reduction, hyperpermeability, and SV atrophy. The expression of zonulae occludins-1 was found to have decreased after IR. The loss of outer hair cells (OHCs) occurred later than SV damage. The disruption to the SV and OHCs could be attributed to reactive oxygen species (ROS)-related damage. In addition, EP shifts and the loss of OHCs were reduced when ROS was reduced by N-acetylcysteine (NAC) in C57BL/6 mice, attenuating auditory threshold shifts. Innovation: The damage to the SV was found to occur before OHC loss. ROS-related damage accounted for SV damage and OHC loss. The incidences of SV damage and OHC loss were decreased through ROS modulation by NAC, subsequently preventing RISNHL, suggesting the possible role of NAC as a possible protective agent against RISNHL. Conclusion: The findings from this study suggest oxidative stress-induced early SV injury and late OHC loss to be the key factors leading to RISNHL. NAC prevents IR-induced OHC loss, and attenuates auditory brainstem response and EP shifts by regulating the level of oxidative stress. Antioxid. Redox Signal. 40, 470-491.

Higher degree of keratinization correlated with severe bone destruction in acquired Cholesteatoma.

BACKGROUND: Acquired cholesteatoma is characterized by hyper-keratinized squamous epithelium and bone destruction. However, direct evidence for hyper-keratinized epidermis promoting bone destruction is lacking. AIMS/OBJECTIVES: To determine whether higher degree of keratinization correlated with severe bone destruction and further offer direct evidence for keratinocyte-inducing osteoclastogenesis. MATERIALS AND METHODS: Histological changes and clinical relevance were analyzed in human-acquired cholesteatoma. Animal models were established by implanting autologous epidermis with different degrees of keratinization. The severity of bone resorption and the number of osteoclasts were compared in different keratinized groups. An in vitro coculture system was developed to mimic the progress of keratinocyte-inducing osteoclastogenesis. RESULTS: The matrix of cholesteatoma was composed of a thicker stratum corneum than normal skin. The stratum corneum thickness and the expression of Keratin 10 positively correlated to the severity of bone destruction. Animal models revealed that the bone destruction induced by a higher keratinized epidermis was more severe. Osteoclasts were detected in bone erosion areas, and the number of osteoclasts increased with the keratinization degrees of the graft. In vitro studies showed that keratinocytes directly promoted monocytes differentiating into osteoclasts. CONCLUSIONS AND SIGNIFICANCE: In acquired cholesteatoma, the degree of keratinization correlated with disease severity, and keratinocytes directly promote osteoclastogenesis.

Promoting TFEB nuclear localization with curcumin analog C1 attenuates sensory hair cell injury and delays age-related hearing loss in C57BL/6 mice.

Sensory hair cell (HC) injuries, especially outer hair cell (OHC) loss, are well-documented to be the primary pathology of age-related hearing loss (AHL). Recent studies have demonstrated that autophagy plays an important role in HC injury in the inner ear. In our previous works, a decline in autophagy levels and HC loss were found to occur simultaneously in the inner ears of aged C57BL/6 mice, and the administration of rapamycin promoted autophagy levels, which reduced OHC loss and delayed AHL, but the underlying mechanism of autophagy in AHL has not been well elucidated. Transcription factor EB (TFEB), an autophagy regulator and the downstream target of mammalian target of rapamycin (mTOR), is involved in the pathological development of neurodegenerative disease. This study would address the link between autophagy and TFEB in aged C57BL/6 mouse cochleae and clarify the effect of the TFEB activator curcumin analog C1 (C1) in aged cochleae. Decreased TFEB nuclear localization (p = 0.0371) and autophagy dysfunction (p = 0.0273) were observed in the cochleae of aged C57BL/6 mice that exhibited AHL, HCs loss and HCs senescence. Treatment with C1 promoted TFEB nuclear localization and restored autophagy, subsequently alleviating HC injury and delaying AHL. The protective effect of C1 on HEI-OC1 cells against autophagy disorder and aging induced by D-galactose was abolished by chloroquine, which is one of the commonly used autophagy inhibitors. Overall, our results demonstrated that the capacity to perform autophagy is mediated by the nuclear localization of TFEB in aged C57BL/6 mouse cochleae. C1 promotes the nuclear localization of TFEB, subsequently alleviating HC injury and delaying AHL by restoring the impaired autophagy function. TFEB may serve as a new therapeutic target for AHL treatment.

Regulation of ACSL4-Catalyzed Lipid Peroxidation Process Resists Cisplatin Ototoxicity.

Cisplatin-induced ototoxicity is one of the common side effects during its treatment and there are no effective measures to prevent it. Our study aimed to investigate the effect of ACSL4-catalyzed lipid peroxidation on cisplatin-induced hearing loss and its possible protective mechanisms. We used a variety of cisplatin ototoxicity models, including HEI-OC1 cell line, cochlear explants, and ET4 GFP+ zebrafish. After measuring the experimental concentrations of cisplatin by CCK8 assay and immunofluorescence, respectively, we examined the levels of lipid peroxidation by MDA content, 4-HNE content, and C11-BODIPY (581/591) probe. Then, we used two ferroptosis inhibitors, FER-1, and Vit-E to protect hair cells. We found that cisplatin significantly increased the levels of lipid peroxidation and that this process can be resisted by the ferroptosis inhibitors. Afterwards, we performed metabolomic assays on the cisplatin-treated hair cells. The metabolite levels were significantly altered in the experimental group compared to the control group, and the highest degree of change was observed in the glutathione metabolic pathway and the arachidonic acid metabolic pathway. Therefore, we screened the key enzymes on the arachidonic acid metabolic pathway in the hair cells after cisplatin treatment and found that ACSL4 had the greatest regulatory value. Further, we reduced the level of lipid peroxide in hair cells by specifically inhibiting the expression of ACSL4, which protected hair cells from cisplatin damage at source. In conclusion, the lipid peroxidation process regulated by ACSL4 may be an important factor contributing to the sensitivity of hair cells to cisplatin. Inhibition of ACSL4 expression may be an effective preventive measure against cisplatin ototoxicity.

Autophagy-dependent ferroptosis contributes to cisplatin-induced hearing loss.

Cisplatin-induced hearing loss is a common side effect of cisplatin chemotherapy, for which clinical therapy remains unavailable. Apoptosis of hair cells is considered the primary cause of cisplatin-induced ototoxicity; however, inhibiting apoptosis can only partially restore cisplatin-induced hearing loss. Therefore, auditory cell death caused by cisplatin damage requires further study. Ferroptosis, a novel form of regulated cell death, has been shown to play a role in the mechanism of cisplatin toxicity. In this study, we observed proferroptotic alterations (lipid peroxidation and impaired antioxidant capacity) in the cochleae of C57BL/6 mice after cisplatin damage, verifying the induction of ferroptosis. Using the HEI-OC1 cell line, we observed that cisplatin induced proferroptotic alterations and activated ferritinophagy (specific autophagy pathway). Employing chloroquine, we confirmed that the blockage of autophagy remarkably alleviated cisplatin-induced ferroptosis in HEI-OC1 cells; therefore, the induction of ferroptosis in cisplatin-treated auditory cells was dependent on the activation of autophagy. In addition, the ferroptosis inhibitor ferrostatin-1 and iron chelator deferoxamine significantly attenuated cisplatin-induced cytotoxicity in HEI-OC1 cells and cochlear explants. Moreover, pharmacologically inhibiting ferroptosis using ferrostatin-1 significantly decreased the auditory cell loss and, notably, attenuated hearing loss in C57BL/6 mice after cisplatin damage. Collectively, these findings indicate that autophagy-dependent ferroptosis plays an integrated role in the mechanism of cisplatin-induced hearing loss.

Modulation of NAD+ biosynthesis activates SIRT1 and resists cisplatin-induced ototoxicity.

Cisplatin, the most widely used platinum-based anticancer drug, often causes progressive and irreversible sensorineural hearing loss in cancer patients. However, the precise mechanism underlying cisplatin-associated ototoxicity is still unclear. Nicotinamide adenine dinucleotide (NAD+), a co-substrate for the sirtuin family and PARPs, has emerged as a potent therapeutic molecular target in various diseases. In our investigates, we observed that NAD+ level was changed in the cochlear explants of mice treated with cisplatin. Supplementation of a specific inhibitor (TES-1025) of α-amino-ß-carboxymuconate-ε-semialdehyde decarboxylase (ACMSD), a rate-limiting enzyme of NAD+de novo synthesis pathway, promoted SIRT1 activity, increased mtDNA contents and enhanced AMPK expression, thus significantly reducing hair cells loss and deformation. The protection was blocked by EX527, a specific SIRT1 inhibitor. Meanwhile, the use of NMN, a precursor of NAD+ salvage synthesis pathway, had shown beneficial effect on hair cell under cisplatin administration, effectively suppressing PARP1. In vivo experiments confirmed the hair cell protection of NAD+ modulators in cisplatin treated mice and zebrafish. In conclusion, we demonstrated that modulation of NAD+ biosynthesis via the de novo synthesis pathway and the salvage synthesis pathway could both prevent ototoxicity of cisplatin. These results suggested that direct modulation of cellular NAD+ levels could be a promising therapeutic approach for protection of hearing from cisplatin-induced ototoxicity.

Nrf2 activation protects auditory hair cells from cisplatin-induced ototoxicity independent on mitochondrial ROS production.

Cisplatin is a well-known and commonly used chemotherapeutic agent. However, cisplatin-induced ototoxicity limits its clinical use. Previous studies have shown an important role of reactive oxygen species (ROS) accumulation in the pathogenesis of cisplatin-induced ototoxicity. In many cell types, the transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2) and antioxidant response element (ARE) protect against oxidative stress by suppressing ROS. Here our results showed that cisplatin injury reduced Nrf2 expression and inhibited Nrf2 translocation in HEI-OC1 cells and Nrf2 activator tert-butylhydroquinone (TBHQ) rescued hair cells from cisplatin induced apoptosis by suppressing the total cellular ROS accumulation. Moreover, we found that decreased ROS accumulation induced by TBHQ didn't depend on mitochondrial derived ROS production, indicating that Nrf2 activation alleviated cisplatin induced oxidative stress and apoptosis through mitochondrial-independent ROS production. Therefore, we provide a potential strategy of prevention and treatment for cisplatin-induced ototoxicity by Nrf2 activation. In conclusion, Nrf2 activation protects auditory hair cells from cisplatin-induced ototoxicity through suppressing the total cellular ROS levels which arise from sources other than mitochondria.

Cartilage tympanoplasty combined with eustachian tube balloon dilatation in the treatment of adhesive otitis media.

OBJECTIVES/HYPOTHESIS: To investigate the simultaneous application of cartilage tympanoplasty combined with eustachian tube (ET) balloon dilatation in the treatment of adhesive otitis media (AdOM). STUDY DESIGN: Multicenter, prospective, double-blind, randomized, controlled clinical trial. METHODS: Patients with AdOM were randomly divided into four groups: control group (conservative treatment), ET balloon dilatation (ETBD) group, cartilage tympanoplasty (CT) group, and cartilage tympanoplasty combined with ET balloon dilatation (ETBD+CT) group. Patients were followed up at 3 months, 6 months, 1 year, and 2 years after treatment, receiving otoendoscopy and pure-tone audiometry, and were evaluated using the Tinnitus Handicap Inventory (THI), visual analogue scale (VAS) for the symptom of ear stuffiness, Chronic Otitis Media Outcome Test (COMOT-15), and eustachian tube scores (ETS). RESULTS: There was no improvement in tympanic membrane (TM) morphology and mean pure-tone air-bone gap (ABG) after treatment in the control and ETBD groups. The postoperative TM morphology was improved in the CT group and ETBD+CT group, although retraction pockets reoccurred in two cases of CT group. Reduced ABG and improvements in ETS, THI, VAS, and COMOT-15 were all achieved in these two groups, but the difference was not statistically significant. CONCLUSIONS: Cartilage tympanoplasty combined with ET balloon dilatation could be used as an appropriate surgical technique for AdOM, which could relieve the symptoms of tinnitus and ear stuffiness, and improve postoperative TM morphology, hearing level, ET functions, and quality of life, with low incidence of complications. LEVEL OF EVIDENCE: 2 Laryngoscope, 129:1462-1467, 2019.