Molecular and Clinical Significance of Fibroblast Growth Factor 2 in Development and Regeneration of the Auditory System

The fibroblast growth factor 2 (FGF2) is a member of the FGF family which is involved in key biological processes including development, cellular proliferation, wound healing, and angiogenesis. Although the utility of the FGF family as therapeutic agents has attracted attention, and FGF2 has been studied in several clinical contexts, there remains an incomplete understanding of the molecular and clinical function of FGF2 in the auditory system. In this review, we highlight the role of FGF2 in inner ear development and hearing protection and present relevant clinical studies for tympanic membrane (TM) repair. We conclude by discussing the future implications of FGF2 as a potential therapeutic agent.

Knockout of mafba Causes Inner-Ear Developmental Defects in Zebrafish via the Impairment of Proliferation and Differentiation of Ionocyte Progenitor Cells

Zebrafish is an excellent model for exploring the development of the inner ear. Its inner ear has similar functions to that of humans, specifically in the maintenance of hearing and balance. Mafba is a component of the Maf transcription factor family. It participates in multiple biological processes, but its role in inner-ear development remains poorly understood. In this study, we constructed a mafba knockout (mafba−/−) zebrafish model using CRISPR/Cas9 technology. The mafba−/− mutant inner ear displayed severe impairments, such as enlarged otocysts, smaller or absent otoliths, and insensitivity to sound stimulation. The proliferation of p63+ epidermal stem cells and dlc+ ionocyte progenitors was inhibited in mafba−/− mutants. Moreover, the results showed that mafba deletion induces the apoptosis of differentiated K+-ATPase-rich (NR) cells and H+-ATPase-rich (HR) cells. The activation of p53 apoptosis and G0/G1 cell cycle arrest resulted from DNA damage in the inner-ear region, providing a mechanism to account for the inner ear deficiencies. The loss of homeostasis resulting from disorders of ionocyte progenitors resulted in structural defects in the inner ear and, consequently, loss of hearing. In conclusion, the present study elucidated the function of ionic channel homeostasis and inner-ear development using a zebrafish Mafba model and clarified the possible physiological roles.

Characterization of the microRNA Transcriptomes and Proteomics of Cochlea Tissue-derived Small Extracellular Vesicles From Different Age of Mice After Birth

The cochlea is an important sensory organ for both balance and sound perception, and the formation of the cochlea is a complex developmental process. The development of the mouse cochlea begins on embryonic day (E)9 and continues until postnatal day (P)21 when the hearing system is considered mature. Small extracellular vesicles (sEVs), with a diameter ranging from 30 nm to 200 nm, have been considered as a significant medium for information communication in both the processing of physiological and pathological. However, there are no studies exploring the role of sEVs in the development of the cochlea. Here, we isolated tissue-derived sEVs from the cochleae of FVB mice at P3, P7, P14, and P21 by ultracentrifugation. These sEVs were first characterized by transmission electron microscopy, nanoparticle tracking analysis, and western blotting. Next, we used small RNA-seq and mass spectrometry to characterize the microRNA transcriptomes and proteomics of cochlear sEVs from mice at different ages. Many microRNAs and proteins were discovered to be related with inner ear development, anatomical structure development, and the auditory nervous system development. These results all suggest that sEVs exist in the cochlea and are likely to be essential for the normal development of the auditory system. Our findings provide many sEV microRNA and protein targets for future studies of the roles of cochlear sEVs.

A Model of Waardenburg Syndrome Using Patient-Derived iPSCs With a SOX10 Mutation Displays Compromised Maturation and Function of the Neural Crest That Involves Inner Ear Development

Waardenburg syndrome (WS) is an autosomal dominant inherited disorder that is characterized by sensorineural hearing loss and abnormal pigmentation. SOX10 is one of its main pathogenicity genes. The generation of patient-specific induced pluripotent stem cells (iPSCs) is an efficient means to investigate the mechanisms of inherited human disease. In our work, we set up an iPSC line derived from a WS patient with SOX10 mutation and differentiated into neural crest cells (NCCs), a key cell type involved in inner ear development. Compared with control-derived iPSCs, the SOX10 mutant iPSCs showed significantly decreased efficiency of development and differentiation potential at the stage of NCCs. After that, we carried out high-throughput RNA-seq and evaluated the transcriptional misregulation at every stage. Transcriptome analysis of differentiated NCCs showed widespread gene expression alterations, and the differentially expressed genes (DEGs) were enriched in gene ontology terms of neuron migration, skeletal system development, and multicellular organism development, indicating that SOX10 has a pivotal part in the differentiation of NCCs. It’s worth noting that, a significant enrichment among the nominal DEGs for genes implicated in inner ear development was found, as well as several genes connected to the inner ear morphogenesis. Based on the protein-protein interaction network, we chose four candidate genes that could be regulated by SOX10 in inner ear development, namely, BMP2, LGR5, GBX2, and GATA3. In conclusion, SOX10 deficiency in this WS subject had a significant impact on the gene expression patterns throughout NCC development in the iPSC model. The DEGs most significantly enriched in inner ear development and morphogenesis may assist in identifying the underlying basis for the inner ear malformation in subjects with WS.

Inner Ear and Muscle Developmental Defects in Smpx-Deficient Zebrafish Embryos

The last decade has witnessed the identification of several families affected by hereditary non-syndromic hearing loss (NSHL) caused by mutations in the SMPX gene and the loss of function has been suggested as the underlying mechanism. In the attempt to confirm this hypothesis we generated an Smpx-deficient zebrafish model, pointing out its crucial role in proper inner ear development. Indeed, a marked decrease in the number of kinocilia together with structural alterations of the stereocilia and the kinocilium itself in the hair cells of the inner ear were observed. We also report the impairment of the mechanotransduction by the hair cells, making SMPX a potential key player in the construction of the machinery necessary for sound detection. This wealth of evidence provides the first possible explanation for hearing loss in SMPX-mutated patients. Additionally, we observed a clear muscular phenotype consisting of the defective organization and functioning of muscle fibers, strongly suggesting a potential role for the protein in the development of muscle fibers. This piece of evidence highlights the need for more in-depth analyses in search for possible correlations between SMPX mutations and muscular disorders in humans, thus potentially turning this non-syndromic hearing loss-associated gene into the genetic cause of dysfunctions characterized by more than one symptom, making SMPX a novel syndromic gene.

Ceramide Kinase Inhibition Blocks IGF-1-Mediated Survival of Otic Neurosensory Progenitors by Impairing AKT Phosphorylation

Sphingolipids are bioactive lipid components of cell membranes with important signal transduction functions in health and disease. Ceramide is the central building block for sphingolipid biosynthesis and is processed to form structurally and functionally distinct sphingolipids. Ceramide can be phosphorylated by ceramide kinase (CERK) to generate ceramide-1-phosphate, a cytoprotective signaling molecule that has been widely studied in multiple tissues and organs, including the developing otocyst. However, little is known about ceramide kinase regulation during inner ear development. Using chicken otocysts, we show that genes for CERK and other enzymes of ceramide metabolism are expressed during the early stages of inner ear development and that CERK is developmentally regulated at the otic vesicle stage. To explore its role in inner ear morphogenesis, we blocked CERK activity in organotypic cultures of otic vesicles with a specific inhibitor. Inhibition of CERK activity impaired proliferation and promoted apoptosis of epithelial otic progenitors. CERK inhibition also compromised neurogenesis of the acoustic-vestibular ganglion. Insulin-like growth factor-1 (IGF-1) is a key factor for proliferation, survival and differentiation in the chicken otocyst. CERK inhibition decreased IGF-1-induced AKT phosphorylation and blocked IGF-1-induced cell survival. Overall, our data suggest that CERK is activated as a central element in the network of anti-apoptotic pro-survival pathways elicited by IGF-1 during early inner ear development.