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Mammalian auditory hair cells transduce sound-evoked traveling waves in the cochlea into nerve stimuli, which are essential for hearing function. Pillar cells located between the inner and outer hair cells are involved in the formation of the tunnel of Corti, which incorporates outer-hair-cell-driven fluid oscillation and basilar membrane movement, leading to the fine-tuned frequency-specific perception of sounds by the inner hair cells. However, the detailed molecular mechanism underlying the development and maintenance of pillar cells remains to be elucidated. In this study, we examined the expression and function of brain-specific angiogenesis inhibitor 3 (Bai3), an adhesion G-protein-coupled receptor, in the cochlea. We found that Bai3 was expressed in hair cells in neonatal mice and pillar cells in adult mice, and, interestingly, Bai3 knockout mice revealed the abnormal formation of pillar cells, with the elevation of the hearing threshold in a frequency-dependent manner. Furthermore, old Bai3 knockout mice showed the degeneration of hair cells and spiral ganglion neurons in the basal turn. The results suggest that Bai3 plays a crucial role in the development and/or maintenance of pillar cells, which, in turn, are necessary for normal hearing function. Our results may contribute to understanding the mechanisms of hearing loss in human patients. [ABSTRACT FROM AUTHOR]

A number of transcriptional pathways regulating fetal lung development are active during repair of the injured lung. We hypothesized that C/EBP[alpha], a transcription factor critical for lung maturation, plays a role in protection of the alveolar epithelium following hyperoxic injury of the mature lung. Transgenic [Cebp[alpha].sup.[DELTA]/[DELTA]] mice, in which Cebp[alpha] was conditionally deleted from Clara cells and type II cells after birth, were developed. While no pulmonary abnormalities were observed in the [Cebp[alpha].sup.[DELTA]/[DELTA]] mice (7-8 wk old) under normal conditions, the mice were highly susceptible to hyperoxia. [Cebp[alpha].sup.[DELTA]/[DELTA]] mice died within 4 days of exposure to 95% oxygen in association with severe lung inflammation, altered maturation of surfactant protein B and C, decreased surfactant lipid secretion, and abnormal lung mechanics at a time when all control mice survived, mRNA microarray analysis of isolated type II cells at 0, 2, and 24 h of hyperoxia demonstrated the reduced expression of number of genes regulating surfactant lipid and protein homeostasis, including Srebf, Scap, Lpcat1, Abca3, Sftpb, and Napsa. Genes influencing cell signaling or immune responses were induced in the lungs of [Cebp[alpha].sup.[DELTA]/[DELTA]] mice. C/EBP[alpha] was required for the regulation of genes associated with surfactant lipid homeostasis, surfactant protein biosynthesis, processing and transport, defense response to stress, and cell redox homeostasis during exposure to hyperoxia. While C/EBP[alpha] did not play a critical role in postnatal pulmonary function under normal conditions, C/EBP[alpha] mediated protection of the lung during acute lung injury induced by hyperoxia. Vnn1; GSH; microarray; Napsa; suffactant

Xu Y, Saegusa C, Schehr A, Grant S, Whitsett JA, Ikegami M. C/EBPa is required for pulmonary cytoprotection during hyperoxia. Am J Physiol Lung Cell Mo! Physiol 297: L286-L298, 2009. First published May 22, 2009; doi:10.1152/ajplung.00094.2009.- A number of transcriptional pathways regulating fetal lung development are active during repair of the injured lung. We hypothesized that Cebpα, a transcription factor critical for lung maturation, plays a role in protection of the alveolar epithelium following hyperoxic injury of the mature lung. Transgenic Cebpα[supΔ/Δ], in which Cebpα was conditionally deleted from Clara cells and type II cells after birth, were developed. While no pulmonary abnormalities were observed in the Cebpα[supΔ/Δ]mice (7-8 wk old) under normal conditions, the mice were highly susceptible to hyperoxia. Cebpα[supΔ/Δ] mice died within 4 days of exposure to 95% oxygen in association with severe lung inflammation, altered maturation of surfactant protein B and C, decreased surfactant lipid secretion, and abnormal lung mechanics at a time when all control mice survived. mRNA microarray analysis of isolated type II cells at 0, 2, and 24 h of hyperoxia demonstrated the reduced expression of number of genes regulating surfactant lipid and protein homeostasis, including Srebf, Scap, Lpcar1 , Abca3, Sftpb, and Napsa. Genes influencing cell signaling or immune responses were induced in the lungs of Cebpα[supΔ/Δ] mice. C/EBPα was required for the regulation of genes associated with surfactant lipid homeostasis, surfactant protein biosynthesis, processing and transport, defense response to stress, and cell redox homeostasis during exposure to hyperoxia. While C/EBPα did not play a critical role in postnatal pulmonary function under normal conditions, C/EBPα mediated protection of the lung during acute lung injury induced by hyperoxia. [ABSTRACT FROM AUTHOR]

It has recently been proposed that synaptotagmin (Syt) VII functions as a plasma membrane Ca2+ sensor for dense-core vesicle exocytosis in PC12 cells based on the results of transient overexpression studies using green fluorescent protein (GFP)-tagged Syt VII; however, the precise subcellular localization of Syt VII is still a matter of controversy (plasma membrane versus secretory granules). In this study we established a PC12 cell line "stably expressing" the Syt VII-GFP molecule and demonstrated by immunocytochemical and immunoelectron microscopic analyses that the Syt VII-GFP protein is localized on dense-core vesicles as well as in other intracellular membranous structures, such as the trans-Golgi network and lysosomes. Syt VII-GFP forms a complex with endogenous Syts I and IX, but not with Syt IV, and it colocalize well with Syts I and IX in the cellular processes (where dense-core vesicles are accumulated) in the PC12 cell line. We further demonstrated by an N-terminal antibody-uptake experiment that Syt VIIGFP-containing dense-core vesicles undergo Ca2+dependent exocytosis, the same as endogenous Syt IX-containing vesicles. Moreover, silencing of Syt VII-GFP with specific small interfering RNA dramatically reduced high KCl-dependent neuropeptide Y secretion from the stable PC12 cell line (∼60% of the control cells), whereas the same small interfering RNA had little effect on neuropeptide Y secretion from the wild-type PC12 cells (∼85-90% of the control cells), indicating that the level of endogenous expression of Syt VII molecules must be low. Our results indicate that the targeting of Syt VII-GFP molecules to specific membrane compartment(s) is affected by the transfection method (transient expression versus stable expression) and suggested that Syt VII molecule on dense-core vesicles functions as a vesicular Ca2+ sensor for exocytosis in endocrine cells. [ABSTRACT FROM AUTHOR]

Introduction: Pendred syndrome is an autosomal-recessive disease characterized by congenital hearing loss and thyroid goiter. Previously, cell stress susceptibilities were shown to increase in patient-derived cells with intracellular aggregation using an in vitro acute cochlear cell model derived from patient-specific pluripotent stem (iPS) cells. Moreover, we showed that rapamycin can relieve cell death. However, studies regarding long-term cell survival without cell stressors that mimic the natural course of disease or the rational minimum concentration of rapamycin that prevents cell death are missing., Methods: In this report, we first investigated the rational minimum concentration of rapamycin using patient-specific iPS cells derived-cochlear cells with three different conditions of acute stress. We next confirmed the effects of rapamycin in long-term cell survival and phenotypes by using cochlear cells derived from three different patient-derived iPS cells., Results: We found that inner ear cells derived from Pendred syndrome patients are more vulnerable than those from healthy individuals during long-term culturing; however, this susceptibility was relieved via treatment with low-dose rapamycin. The slow progression of hearing loss in patients may be explained, in part, by the vulnerability observed in patient cells during long-term culturing. We successfully evaluated the rational minimum concentration of rapamycin for treatment of Pendred syndrome., Conclusion: Our results suggest that low-dose rapamycin not only decreases acute symptoms but may prevent progression of hearing loss in Pendred syndrome patients.