Clonal Expansion of Lgr5-Positive Cells from Mammalian Cochlea and High-Purity Generation of Sensory Hair Cells

Death of cochlear hair cells, which do not regenerate, is a cause of hearing loss in a high percentage of the population. Currently, no approach exists to obtain large numbers of cochlear hair cells. Here, using a small-molecule approach, we show significant expansion (>2,000-fold) of cochlear supporting cells expressing and maintaining Lgr5, an epithelial stem cell marker, in response to stimulation of Wnt signaling by a GSK3β inhibitor and transcriptional activation by a histone deacetylase inhibitor. The Lgr5-expressing cells differentiate into hair cells in high yield. From a single mouse cochlea, we obtained over 11,500 hair cells, compared to less than 200 in the absence of induction. The newly generated hair cells have bundles and molecular machinery for transduction, synapse formation, and specialized hair cell activity. Targeting supporting cells capable of proliferation and cochlear hair cell replacement could lead to the discovery of hearing loss treatments.United States. National Institutes of Health (DE-013023)United States. National Institutes of Health (DC-007174)United States. National Institutes of Health (DC-013909)United States. National Institutes of Health (RR-00168

Applications of Lgr5-Positive Cochlear Progenitors (LCPs) to the Study of Hair Cell Differentiation

The mouse cochlea contains approximately 15,000 hair cells. Its dimensions and location, and the small number of hair cells, make mechanistic, developmental and cellular replacement studies difficult. We recently published a protocol to expand and differentiate murine neonatal cochlear progenitor cells into 3D organoids that recapitulate developmental pathways and can generate large numbers of hair cells with intact stereociliary bundles, molecular markers of the native cells and mechanotransduction channel activity, as indicated by FM1-43 uptake. Here, we elaborate on the method and application of these Lgr5-positive cochlear progenitors, termed LCPs, to the study of inner ear development and differentiation. We demonstrate the use of these cells for testing several drug candidates, gene silencing and overexpression, as well as genomic modification using CRISPR/Cas9. We thus establish LCPs as a valuable in vitro tool for the analysis of progenitor cell manipulation and hair cell differentiation

Right-Wing Politicians Prefer the Emotional Left

Physiological research suggests that social attitudes, such as political beliefs, may be partly hard-wired in the brain. Conservatives have heightened sensitivity for detecting emotional faces and use emotion more effectively when campaigning. As the left face displays emotion more prominently, we examined 1538 official photographs of conservative and liberal politicians from Australia, Canada, the United Kingdom and the United States for an asymmetry in posing. Across nations, conservatives were more likely than liberals to display the left cheek. In contrast, liberals were more likely to face forward than were conservatives. Emotion is important in political campaigning and as portraits influence voting decisions, conservative politicians may intuitively display the left face to convey emotion to voters

Integration of Transcriptomics, Proteomics, and MicroRNA Analyses Reveals Novel MicroRNA Regulation of Targets in the Mammalian Inner Ear

We have employed a novel approach for the identification of functionally important microRNA (miRNA)-target interactions, integrating miRNA, transcriptome and proteome profiles and advanced in silico analysis using the FAME algorithm. Since miRNAs play a crucial role in the inner ear, demonstrated by the discovery of mutations in a miRNA leading to human and mouse deafness, we applied this approach to microdissected auditory and vestibular sensory epithelia. We detected the expression of 157 miRNAs in the inner ear sensory epithelia, with 53 miRNAs differentially expressed between the cochlea and vestibule. Functionally important miRNAs were determined by searching for enriched or depleted targets in the transcript and protein datasets with an expression consistent with the dogma of miRNA regulation. Importantly, quite a few of the targets were detected only in the protein datasets, attributable to regulation by translational suppression. We identified and experimentally validated the regulation of PSIP1-P75, a transcriptional co-activator previously unknown in the inner ear, by miR-135b, in vestibular hair cells. Our findings suggest that miR-135b serves as a cellular effector, involved in regulating some of the differences between the cochlear and vestibular hair cells

Genomic Duplication and Overexpression of TJP2/ZO-2 Leads to Altered Expression of Apoptosis Genes in Progressive Nonsyndromic Hearing Loss DFNA51

Age-related hearing loss is due to death over time, primarily by apoptosis, of hair cells in the inner ear. Studies of mutant genes responsible for inherited progressive hearing loss have suggested possible mechanisms for hair cell death, but critical connections between these mutations and the causes of progressive hearing loss have been elusive. In an Israeli kindred, dominant, adult-onset, progressive nonsyndromic hearing loss DFNA51 is due to a tandem inverted genomic duplication of 270 kb that includes the entire wild-type gene encoding the tight junction protein TJP2 (ZO-2). In the mammalian inner ear, TJP2 is expressed mainly in tight junctions, and also in the cytoplasm and nuclei. TJP2 expression normally decreases with age from embryonic development to adulthood. In cells of affected family members, TJP2 transcript and protein are overexpressed, leading to decreased phosphorylation of GSK-3β and to altered expression of genes that regulate apoptosis. These results suggest that TJP2- and GSK-3β-mediated increased susceptibility to apoptosis of cells of the inner ear is the mechanism for adult-onset hearing loss in this kindred and may serve as one model for age-related hearing loss in the general population

The Use and Duration of Preintubation Respiratory Support Is Associated With Increased Mortality in Immunocompromised Children With Acute Respiratory Failure

OBJECTIVESTo determine the association between preintubation respiratory support and outcomes in patients with acute respiratory failure and to determine the impact of immunocompromised (IC) diagnoses on outcomes after adjustment for illness severity. DESIGNRetrospective multicenter cohort study. SETTINGEighty-two centers in the Virtual Pediatric Systems database. PATIENTSChildren 1 month to 17 years old intubated in the PICU who received invasive mechanical ventilation (IMV) for greater than or equal to 24 hours. INTERVENTIONSNone. MEASUREMENTS AND MAIN RESULTSHigh-flow nasal cannula (HFNC) or noninvasive positive-pressure ventilation (NIPPV) or both were used prior to intubation in 1,825 (34%) of 5,348 PICU intubations across 82 centers. When stratified by IC status, 50% of patients had no IC diagnosis, whereas 41% were IC without prior hematopoietic cell transplant (HCT) and 9% had prior HCT. Compared with patients intubated without prior support, preintubation exposure to HFNC (adjusted odds ratio [aOR], 1.33; 95% CI, 1.10-1.62) or NIPPV (aOR, 1.44; 95% CI, 1.20-1.74) was associated with increased odds of PICU mortality. Within subgroups of IC status, preintubation respiratory support was associated with increased odds of PICU mortality in IC patients (HFNC: aOR, 1.50; 95% CI, 1.11-2.03; NIPPV: aOR, 1.76; 95% CI, 1.31-2.35) and HCT patients (HFNC: aOR, 1.75; 95% CI, 1.07-2.86; NIPPV: aOR, 1.85; 95% CI, 1.12-3.02) compared with IC/HCT patients intubated without prior respiratory support. Preintubation exposure to HFNC/NIPPV was not associated with mortality in patients without an IC diagnosis. Duration of HFNC/NIPPV greater than 6 hours was associated with increased mortality in IC HCT patients (HFNC: aOR, 2.41; 95% CI, 1.05-5.55; NIPPV: aOR, 2.53; 95% CI, 1.04-6.15) and patients compared HCT patients with less than 6-hour HFNC/NIPPV exposure. After adjustment for patient and center characteristics, both preintubation HFNC/NIPPV use (median, 15%; range, 0-63%) and PICU mortality varied by center. CONCLUSIONSIn IC pediatric patients, preintubation exposure to HFNC and/or NIPPV is associated with increased odds of PICU mortality, independent of illness severity. Longer duration of exposure to HFNC/NIPPV prior to IMV is associated with increased mortality in HCT patients