Jianxin Bao

Back signaling by the Nrg-1 intracellular domain.

Transmembrane isoforms of neuregulin-1 (Nrg-1), ligands for erbB receptors, include an extracellular domain with an EGF-like sequence and a highly conserved intracellular domain (ICD) of unknown function. In this paper, we demonstrate that transmembrane isoforms of Nrg-1 are bidirectional signaling molecules in neurons. The stimuli for Nrg-1 back signaling include binding of erbB receptor dimers to the extracellular domain of Nrg-1 and neuronal depolarization. These stimuli elicit proteolytic release and translocation of the ICD of Nrg-1 to the nucleus. Once in the nucleus, the Nrg-1 ICD represses expression of several regulators of apoptosis, resulting in decreased neuronal cell death in vitro. Thus, regulated proteolytic processing of Nrg-1 results in retrograde signaling that appears to mediate contact and activity-dependent survival of Nrg-1–expressing neurons.

Activity-dependent transcription regulation of PSD-95 by neuregulin-1 and Eos

Neuregulin-1 (Nrg-1) contains an intracellular domain (Nrg-ICD) that translocates into the nucleus, where it may regulate gene expression upon neuronal depolarization. However, the identity of its target promoters and the mechanisms by which it regulates transcription have been elusive. Here we report that, in the mouse cochlea, synaptic activity increases the level of nuclear Nrg-ICD and upregulates postsynaptic density protein-95 (PSD-95), a scaffolding protein that is enriched in post-synaptic structures. Nrg-ICD enhances the transcriptional activity of the PSD-95 promoter by binding to a zinc-finger transcription factor, Eos. The Nrg-ICD–Eos complex induces endogenous PSD-95 expression in vivo through a signaling pathway that is mostly independent of γ-secretase regulation. This upregulation of PSD-95 expression by the Nrg-ICD–Eos complex provides a molecular basis for activity-dependent synaptic plasticity.

Age-Related Loss of Spiral Ganglion Neurons

Spiral ganglion neurons (SGNs) are the relay station for auditory information between hair cells and central nervous system. Age-related decline of auditory function due to SGN loss can not be ameliorated by hearing aids or cochlear implants. Recent findings clearly indicate that survival of SGNs during aging depends on genetic and environmental interactions, which can be demonstrated at the systemic, tissue, cellular, and molecular levels. At the systemic level, both insulin/insulin-like growth factor-1 and lipophilic/steroid hormone pathways influence SGN survival during aging. At the level of organ of the Corti, it is difficult to determine whether age-related SGN loss is primary or secondary degeneration. However, a late stage of SGN degeneration may be independent of age-related loss of hair cells. At the cellular and molecular level, several pathways, particularly free radical and calcium signaling pathways, can influence age-related SGN loss, and further studies should determine how these pathways contribute to SGN loss, such as whether they directly or indirectly act on SGNs. With the advancement of recent genetic and pharmacologic tools, we should not only understand how SGNs die during aging, but also find ways to delay this loss.

Requirement of Nicotinic Acetylcholine Receptor Subunit β2 in the Maintenance of Spiral Ganglion Neurons during Aging

Age-related hearing loss (presbycusis) is a major health concern for the elderly. Loss of spiral ganglion neurons (SGNs), the primary sensory relay of the auditory system, is associated consistently with presbycusis. The causative molecular events responsible for age-related loss of SGNs are unknown. Recent reports directly link age-related neuronal loss in cerebral cortex with the loss of high-affinity nicotine acetylcholine receptors (nAChRs). In cochlea, cholinergic synapses are made by olivocochlear efferent fibers on the outer hair cells that express α9 nAChR subunits and on the peripheral projections of SGNs that express α2, α4-7, and β2-3 nAChR subunits. A significantly decreased expression of the β2 nAChR subunit in SGNs was found specifically in mice susceptible to presbycusis. Furthermore, mice lacking the β2 nAChR subunit (β2-/-), but not mice lacking the α5 nAChR subunit (α5-/-), have dramatic hearing loss and significant reduction in the number of SGNs. Our findings clearly established a requirement for β2 nAChR subunit in the maintenance of SGNs during aging.

Recent Advances in the Study of Age-Related Hearing Loss: A Mini-Review

Hearing loss is a common age-associated affliction that can result from the loss of hair cells and spiral ganglion neurons (SGNs) in the cochlea. Although hair cells and SGNs are typically lost in the same cochlea, recent analysis suggests that they can occur independently, via unique mechanisms. Research has identified both environmental and genetic factors that contribute to degeneration of cochlear cells. Additionally, molecular analysis has identified multiple cell-signaling mechanisms that likely contribute to pathological changes that result in hearing deficiencies. These analyses should serve as useful primers for future work, including genomic and proteomic analysis, to elucidate the mechanisms driving cell loss in the aging cochlea. Significant progress in this field has occurred in the past decade. As our understanding of aging-induced cochlear changes continues to improve, our ability to offer medical intervention will surely benefit the growing elderly population.

Temporal and genetic influences on protection against noise-induced hearing loss by hypoxic preconditioning in mice

The protective benefits of hypoxic preconditioning (HPC) against permanent noise-induced hearing loss (NIHL) were investigated in mice. Hypoxia induced by exposure to 8% O2 for 4 h conferred significant protection against damaging broadband noise delivered 24-48 h later in male and female CBA/J (CBA) and CBA/CaJ mice. No protection was found in C57BL/6 (B6) mice, their B6.CAST-Cdh23(CAST) (B6.CAST) congenics, or in CBAxB6 F1 hybrid mice over the same interval, suggesting that the potential for HPC depends on one or a few autosomal recessive alleles carried by CBA-related strains, and is not influenced by the Cdh23 locus. Protection against NIHL in CBA mice was associated with significant up-regulation of hypoxia-inducible factor-1alpha (HIF-1alpha) within the organ of Corti, not found in B6.CAST. In both CBA and B6.CAST mice, some hypoxia-noise intervals shorter than 24 h were associated with exacerbation of NIHL. Cellular cascades underlying the early exacerbation of NIHL by hypoxia are therefore common to both strains, and not mechanistically linked to later protection. Elucidation of the events that underlie HPC, and how these are impacted by genetics, may lead to pharmacologic approaches to mimic HPC, and may help identify individuals with elevated risk of NIHL.

Neuroprotective effects of blockers for T-type calcium channels.

Cognitive and functional decline with age is correlated with deregulation of intracellular calcium, which can lead to neuronal death in the brain. Previous studies have found protective effects of various calcium channel blockers in pathological conditions. However, little has been done to explore possible protective effects of blockers for T-type calcium channels, which forms a family of FDA approved anti-epileptic drugs. In this study, we found that neurons showed an increase in viability after treatment with either L-type or T-type calcium channel antagonists. The family of low-voltage activated, or T-type calcium channels, comprise of three members (Cav3.1, Cav3.2, and Cav3.3) based on their respective main pore-forming alpha subunits: α1G, α1H, and α1I. Among these three subunits, α1H is highly expressed in hippocampus and certain cortical regions. However, T-type calcium channel blockers can protect neurons derived from α1H-/- mice, suggesting that neuroprotection demonstrated by these drugs is not through the α1H subunit. In addition, blockers for T-type calcium channels were not able to confer any protection to neurons in long-term cultures, while blockers of L-type calcium channels could protect neurons. These data indicate a new function of blockers for T-type calcium channels, and also suggest different mechanisms to regulate neuronal survival by calcium signaling pathways. Thus, our findings have important implications in the development of new treatment for age-related neurodegenerative disorders.

Anti-epileptic drugs delay age-related loss of spiral ganglion neurons via T-type calcium channel.

Loss of spiral ganglion neurons is a major cause of age-related hearing loss (presbycusis). Despite being the third most prevalent condition afflicting elderly persons, there are no known medications to prevent presbycusis. Because calcium signaling has long been implicated in age-related neuronal death, we investigated T-type calcium channels. This family is comprised of three members (Ca(v)3.1, Ca(v)3.2, and Ca(v)3.3), based on their respective main pore-forming alpha subunits: α1G, α1H, and α1I. In the present study, we report a significant delay of age-related loss of cochlear function and preservation of spiral ganglion neurons in α1H null and heterozygous mice, clearly demonstrating an important role for Ca(v)3.2 in age-related neuronal loss. Furthermore, we show that anticonvulsant drugs from a family of T-type calcium channel blockers can significantly preserve spiral ganglion neurons during aging. To our knowledge, this is the first report of drugs capable of diminishing age-related loss of spiral ganglion neurons.

Stem Cell Therapy for Hearing Loss: Math1 Overexpression in VOT-E36 Cells

Hypothesis: VOT-E36 cells acquire mechanosensitivity after mammalian atonal homolog 1 (Math1) overexpression. Background: VOT-E36 cells are derived from a population of epithelial cells in the ventral region of the otocyst at embryonic Day 10.5, before hair cell differentiation. These cells express a number of specific molecular markers for hair cells under both proliferation and differentiation states. Overexpression of Math1 can convert nonsensory epithelial cells into hair cells in the cochlea. Based on this information, we tested whether VOT-E36 cells can be converted into hair cells by Math1 overexpression. Methods: Using reverse transcriptase-polymerase chain reaction-based analysis, we first compared the expression patterns of various molecular markers for hair cell development in VOT-E36 cells between proliferation and differentiation states, and also before and after overexpression of Math1. Subsequently, with a standard calcium imaging method, we examined whether VOT-E36 cells overexpressing Math1 could detect mechanical vibrations and activate spiral ganglion neurons in a coculture model. In addition, using confocal and scanning electron microscopes, we examined morphologic changes of VOT-E36 cells after Math1 overexpression. Results: Consistent with previous reports, this study has shown that VOT-E36 cells express a number of specific molecular markers for hair cells in both proliferation and differentiation states. Under appropriate culture conditions, Math1 is transiently expressed in this cell line during conditional differentiation. In VOT-E36 cells overexpressing Math1, the normal expression pattern of certain molecular markers for mature hair cells is partially restored. Interestingly, after coculture with spiral ganglion neurons, VOT-E36 cells overexpressing Math1 are able to respond to mechanical vibrations and activate spiral ganglion neurons. Possible molecular mechanisms underlying this novel finding have been explored. Conclusion: Math1 overexpression can partially restore presumably downstream signaling cascades for normal hair cell differentiation in VOT-E36 cells, which are able to detect mechanical vibrations after being cocultured with spiral ganglion neurons.

Two-phase analysis of molecular pathways underlying induced pluripotent stem cell induction.

Induced pluripotent stem cells (iPSCs) can be reprogrammed from adult somatic cells by transduction with Oct4, Sox2, Klf4, and c‐Myc, but the molecular cascades initiated by these factors remain poorly understood. Impeding their elucidation is the stochastic nature of the iPS induction process, which results in heterogeneous cell populations. Here we have synchronized the reprogramming process by a two‐phase induction: an initial stable intermediate phase following transduction with Oct4, Klf4, and c‐Myc, and a final iPS phase following overexpression of Sox2. This approach has enabled us to examine temporal gene expression profiles, permitting the identification of Sox2 downstream genes critical for induction. Furthermore, we have validated the feasibility of our new approach by using it to confirm that downregulation of transforming growth factor β signaling by Sox2 proves essential to the reprogramming process. Thus, we present a novel means for dissecting the details underlying the induction of iPSCs, an approach with significant utility in this arena and the potential for wide‐ranging implications in the study of other reprogramming mechanisms. STEM Cells 2011;29:1963–1974.