Littérature scientifique sur le sujet « COCHLEAR PROGENITORS »

Cochlear supporting cells (SCs) are glia-like cells critical for hearing function. In the neonatal cochlea, the greater epithelial ridge (GER) is a mitotically quiescent and transient organ, which has been shown to nonmitotically regenerate SCs. Here, we ablated Lgr5+ SCs using Lgr5-DTR mice and found mitotic regeneration of SCs by GER cells in vivo. With lineage tracing, we show that the GER houses progenitor cells that robustly divide and migrate into the organ of Corti to replenish ablated SCs. Regenerated SCs display coordinated calcium transients, markers of the SC subtype inner phalangeal cells, and survive in the mature cochlea. Via RiboTag, RNA-sequencing, and gene clustering algorithms, we reveal 11 distinct gene clusters comprising markers of the quiescent and damaged GER, and damage-responsive genes driving cell migration and mitotic regeneration. Together, our study characterizes GER cells as mitotic progenitors with regenerative potential and unveils their quiescent and damaged translatomes.

Mechano-sensory hair cells within the inner ear cochlea are essential for the detection of sound. In mammals, cochlear hair cells are only produced during development and their loss, due to disease or trauma, is a leading cause of deafness. In the immature cochlea, prior to the onset of hearing, hair cell loss stimulates neighboring supporting cells to act as hair cell progenitors and produce new hair cells. However, for reasons unknown, such regenerative capacity (plasticity) is lost once supporting cells undergo maturation. Here, we demonstrate that the RNA binding protein LIN28B plays an important role in the production of hair cells by supporting cells and provide evidence that the developmental drop in supporting cell plasticity in the mammalian cochlea is, at least in part, a product of declining LIN28B-mammalian target of rapamycin (mTOR) activity. Employing murine cochlear organoid and explant cultures to model mitotic and nonmitotic mechanisms of hair cell generation, we show that loss of LIN28B function, due to its conditional deletion, or due to overexpression of the antagonistic miRNAlet-7g, suppressed Akt-mTOR complex 1 (mTORC1) activity and renders young, immature supporting cells incapable of generating hair cells. Conversely, we found that LIN28B overexpression increased Akt-mTORC1 activity and allowed supporting cells that were undergoing maturation to de-differentiate into progenitor-like cells and to produce hair cells via mitotic and nonmitotic mechanisms. Finally, using the mTORC1 inhibitor rapamycin, we demonstrate that LIN28B promotes supporting cell plasticity in an mTORC1-dependent manner.

Congenital Zika Syndrome (CZS) is caused by vertical transmission of Zika virus (ZIKV) to the gestating human fetus. A subset of CZS microcephalic infants present with reduced otoacoustic emissions; this test screens for hearing loss originating in the cochlea. This observation leads to the question of whether mammalian cochlear tissues are susceptible to infection by ZIKV during development. To address this question using a mouse model, the sensory cochlea was explanted at proliferative, newly post-mitotic or maturing stages. ZIKV was added for the first 24 h and organs cultured for up to 6 days to allow for cell differentiation. Results showed that ZIKV can robustly infect proliferating sensory progenitors, as well as post-mitotic hair cells and supporting cells. Virus neutralization using ZIKV-117 antibody blocked cochlear infection. AXL is a cell surface molecule known to enhance the attachment of flavivirus to host cells. While Axl mRNA is widely expressed in embryonic cochlear tissues susceptible to ZIKV infection, it is selectively downregulated in the post-mitotic sensory organ by E15.5, even though these cells remain infectible. These findings may offer insights into which target cells could potentially contribute to hearing loss resulting from fetal exposure to ZIKV in humans.

Multipotent cochlear neural progenitors (CNPs) in the organ of Corti hold the promise for cell replacement in degenerative hearing disorders. However, not much is known about the CNPs and the specific conditions for their differentiation. Here we isolate the CNPs from the postnatal day 1 organ of Corti in mice and demonstrate their capability to self-renew and to differentiate into hair cell-like and neuronal cell-like phenotypes under the guidance of sonic hedgehog (SHH), epidermal growth factor (EGF), retinoic acid (RA), and brain-derived neurotrophic factor (BDNF), herein termed SERB (abbreviation of SHH, EGF, RA, and BDNF) in an asymmetric or symmetric manner from clonal isolates. Differentiation of CNPs into hair cells by SERB was dependent on the ERK signaling pathway, whereas the differentiation of CNPs into neurons by SERB was not. This work develops a new in vitro methodology for the maintenance and self-regeneration of CNPs for future design of regenerative strategies for hearing disorders.

Emerging evidence indicates that perinatal infection and inflammation can influence the developing immune system and may ultimately affect long-term health and disease outcomes in offspring by perturbing tissue and immune homeostasis. We posit that perinatal inflammation influences immune outcomes in offspring by perturbing (1) the development and function of fetal-derived immune cells that regulate tissue development and homeostasis, and (2) the establishment and function of developing hematopoietic stem cells (HSCs) that continually generate immune cells across the lifespan. To disentangle the complexities of these interlinked systems, we propose the cochlea as an ideal model tissue to investigate how perinatal infection affects immune, tissue, and stem cell development. The cochlea contains complex tissue architecture and a rich immune milieu that is established during early life. A wide range of congenital infections cause cochlea dysfunction and sensorineural hearing loss (SNHL), likely attributable to early life inflammation. Furthermore, we show that both immune cells and bone marrow hematopoietic progenitors can be simultaneously analyzed within neonatal cochlear samples. Future work investigating the pathogenesis of SNHL in the context of congenital infection will therefore provide critical information on how perinatal inflammation drives disease susceptibility in offspring.

Precise control of organ growth and patterning is executed through a balanced regulation of progenitor self-renewal and differentiation. In the auditory sensory epithelium—the organ of Corti—progenitor cells exit the cell cycle in a coordinated wave between E12.5 and E14.5 before the initiation of sensory receptor cell differentiation, making it a unique system for studying the molecular mechanisms controlling the switch between proliferation and differentiation. Here we identify the Yap/Tead complex as a key regulator of the self-renewal gene network in organ of Corti progenitor cells. We show that Tead transcription factors bind directly to the putative regulatory elements of many stemness- and cell cycle-related genes. We also show that the Tead coactivator protein, Yap, is degraded specifically in the Sox2-positive domain of the cochlear duct, resulting in down-regulation of Tead gene targets. Further, conditional loss of theYapgene in the inner ear results in the formation of significantly smaller auditory and vestibular sensory epithelia, while conditional overexpression of a constitutively active version ofYap,Yap5SA, is sufficient to prevent cell cycle exit and to prolong sensory tissue growth. We also show that viral gene delivery ofYap5SAin the postnatal inner ear sensory epithelia in vivo drives cell cycle reentry after hair cell loss. Taken together, these data highlight the key role of the Yap/Tead transcription factor complex in maintaining inner ear progenitors during development, and suggest new strategies to induce sensory cell regeneration.

Sphingolipids are a class of complex lipids known to be not only structural components of biological membranes, but also bioactive molecules involved in several processes, such as cell differentiation, proliferation, motility and cell survival. Between them, we focused on sphingosine 1-phosphate (S1P) and ceramide 1-phosphate (C1P). S1P is intracellularly produced by sphingosine kinase (SK) 1 and SK2 and exerts many of its action consequently to its ligation to S1P specific receptors (S1PR), S1P1–5, whereas C1P is generated by the action of ceramide kinase and it is able to via activation of different signalling pathways. Recent experimental findings demonstrate an emerging role for S1P signalling axis in the maintenance of auditory function. WHO reported that age-related sensorineural hearing loss (SNHL) affects more than 360 million people worldwide, and the current unique available treatment is cochlear implant, which has important use limitations. Our main aim was to investigate S1P signalling axis role in this biological context: in the first paper, we demonstrated that the fibroblast growth factor 2 (FGF2)-induced proliferative action in US/VOT-N33 auditory neuroblasts was dependent on SK1, SK2 as well as S1P1 and S1P2. Moreover, the pro-survival effect of FGF2 from apoptotic cell death induced by staurosporine treatment was dependent on SK but not on S1PR. In addition, ERK1/2 and Akt signaling pathways were found to mediate the mitogenic and survival action of FGF2, respectively. In the second paper, we focused on the emerging role of bioactive sphingolipids as regulators of ERM (ezrin-radixin-moesin) proteins. ERM are a family of cross-linker adaptors between plasma membrane and actin cytoskeleton, playing a crucial role in cell morphology and signal transduction. S1P was found to activate ERM in a S1P2-dependent manner in US/VOT-E36 auditory epithelial progenitors and S1P-induced ERM activation potently contributed to actin cytoskeletal remodeling and to the appearance of electrophysiological changes typical of more differentiated cells. Moreover, PKC and Akt activation was found to mediate S1P-induced ERM phosphorylation. Taken together, our findings demonstrate a crucial role for S1P signalling axis in inner ear biology and disclose potential innovative therapeutic approaches for hearing loss prevention and treatment. In order to develop new methodologies to solve the difficulties of getting drugs inside the inner ear, we studied solid lipid nanoparticles (SLN) as attractive biocompatible nanocarriers for the delivery of drugs with low solubility in aqueous media. In our third paper, we showed that SLN based on stearic acid are efficiently incorporated by HEI-OC1 cells and are not ototoxic at the doses studied. The SLN loaded with glucocorticoids were more effective in protecting cells by the cisplatin-induced damage than glucocorticoids alone. Preliminary in vivo studies also indicate that intratympanic SLN are able to reach the inner ear. These results indicate that SLN are highly efficient vehicles for inner ear drug-delivery and specifically for the administration of glucocorticoids. During my Ph.D. course I have also contributed to a parallel research focused on the role of sphingolipid signalling in skeletal muscle biology. Skeletal muscle is able to regenerate thanks to the presence of satellite cells that upon trauma enter into the cell cycle and start proliferating. Starting from the previously demonstrated positive role of C1P in myoblast proliferation, in the fourth paper we showed that C1P stimulates C2C12 myoblast proliferation via LPA signalling axis. Moreover, C1P via phospholipase A2 activation leads to LPA1 and LPA3 engagement, which in turn drive Akt and ERK1/2 activation, thus stimulating DNA synthesis. The present findings highlight a new key role for bioactive sphingolipids in skeletal muscle and provide further support to the possibility of using them as therapeutic targets for its regeneration.