Cell-Specific Transcriptome Analysis Shows That Adult Pillar and Deiters' Cells Express Genes Encoding Machinery for Specializations of Cochlear Hair Cells

The mammalian auditory sensory epithelium, the organ of Corti, is composed of hair cells and supporting cells. Hair cells contain specializations in the apical, basolateral and synaptic membranes. These specializations mediate mechanotransduction, electrical and mechanical activities and synaptic transmission. Supporting cells maintain homeostasis of the ionic and chemical environment of the cochlea and contribute to the stiffness of the cochlear partition. While spontaneous proliferation and transdifferentiation of supporting cells are the source of the regenerative response to replace lost hair cells in lower vertebrates, supporting cells in adult mammals no longer retain that capability. An important first step to revealing the basic biological properties of supporting cells is to characterize their cell-type specific transcriptomes. Using RNA-seq, we examined the transcriptomes of 1,000 pillar and 1,000 Deiters' cells, as well as the two types of hair cells, individually collected from adult CBA/J mouse cochleae using a suction pipette technique. Our goal was to determine whether pillar and Deiters' cells, the commonly targeted cells for hair cell replacement, express the genes known for encoding machinery for hair cell specializations in the apical, basolateral, and synaptic membranes. We showed that both pillar and Deiters' cells express these genes, with pillar cells being more similar to hair cells than Deiters' cells. The fact that adult pillar and Deiters' cells express the genes cognate to hair cell specializations provides a strong molecular basis for targeting these cells for mammalian hair cell replacement after hair cells are lost due to damage

Identifying microRNAs involved in aging of the lateral wall of the cochlear duct.

Age-related hearing loss is a progressive sensorineural hearing loss that occurs during aging. Degeneration of the organ of Corti and atrophy of the lateral wall of the cochlear duct (or scala media) in the inner ear are the two primary causes. MicroRNAs (miRNAs), a class of short non-coding RNAs that regulate the expression of mRNA/protein targets, are important regulators of cellular senescence and aging. We examined miRNA gene expression profiles in the lateral wall of two mouse strains, along with exploration of the potential targets of those miRNAs that showed dynamic expression during aging. We show that 95 and 60 miRNAs exhibited differential expression in C57 and CBA mice during aging, respectively. A majority of downregulated miRNAs are known to regulate pathways of cell proliferation and differentiation, while all upregulated miRNAs are known regulators in the pro-apoptotic pathways. By using apoptosis-related gene array and bioinformatic approaches to predict miRNA targets, we identify candidate miRNA-regulated genes that regulate apoptosis pathways in the lateral wall of C57 and CBA mice during aging

Changes of Soil Nutrients and Enzyme Activities in Calcareous Purple Soil Under Long-term Continuous Cropping for Tobacco Planting

【Objective】The problem of soil nutrient imbalance becomes more and more serious with the increase of tobacco planting years. The effects of continuous cropping of flue-cured tobacco on soil nutrients and enzyme activities in calcareous purple soil were systematically studied to provide data support for the improvement of soil fertility and the prevention and control of continuous cropping obstacles.【Method】In April 2021, typical plots of calcareous purple soil with different continuous cropping years (0 year, and 8, 10, 27, 33 years) in Zhaohua District, Guangyuan City were selected as the research objects, and the effects of long-term continuous cropping on soil nutrients, enzyme activities and the growth and development of flue-cured tobacco were analyzed.【Result】Long-term continuous cropping of flue-cured tobacco led to the decrease of soil organic matter, pH and alkaline nitrogen by 18.34%, 17.43% and 23.15%, while the contents of soil available phosphorus and available potassium increased by 124% and 54.55%, respectively. Long-term continuous cropping resulted in a decrease of 58.51%-91.28% in urease activity and a decrease of 29.14%-48.66% in sucrase activity in soil. The results of correlation analysis showed that soil sucrase activity was significantly negatively correlated with soil available phosphorus (P < 0.01). Soil β-glucosidase was significantly positively correlated with soil available potassium content (P < 0.01), while significantly negatively correlated with soil available phosphorus (P < 0.05). The soil urease activity was significantly positively correlated with soil pH (P < 0.01).【Conclusion】Long-term continuous cropping of flue-cured tobacco in calcareous purple soil has an inhibitory effect on soil nutrients and enzyme activities. Attention should be paid to the balanced application of nitrogen, phosphorus and potassium fertilizers, and organic fertilizer can be appropriately added to regulate the soil microenvironment to promote the transformation and absorption of nutrients

Morphology and ciliary motion of mucosa in the Eustachian tube of neonatal and adult gerbils.

The Eustachian tube is a small canal that connects the tympanic cavity with the nasal part of the pharynx. The epithelial lining of the Eustachian tube contains a ciliated columnar epithelium at the tympanic cavity and a pseudostratified, ciliated columnar epithelium with goblet cells near the pharynx. The tube serves to equalize air pressure across the eardrum and drains mucus away from the middle ear into the nasopharynx. Blockage of the Eustachian tube is the most common cause of all forms of otitis media, which is common in children. In the present study, we examined the epithelial lining of the Eustachian tube in neonatal and adult gerbils, with a focus on the morphological and functional development of ciliated cells in the mucosa. The length of the tube is ∼8.8 mm in adult gerbils. Scanning electron microscopy showed that the mucosal member near the pharyngeal side contains a higher density of ciliated cells and goblet cells than that near the tympanic side. The cilia beat frequency is 11 Hz. During development, the length of the Eustachian tube increased significantly between postnatal day 1 (P1) and P18. Scanning electron microscopy showed that the mucosa contained a high density of ciliated cells with a few goblet cells at P1. The density of ciliated cells decreased while the density of goblet cells increased during development. At P18, the mucosa appeared to be adult-like. Interestingly, the ciliary beat frequency measured from ciliated cells at P1 was not statistically different from that measured from adult animals. Our study suggests that the Eustachian tube undergoes significant anatomical and histological changes between P1 and P18. The tube is morphologically and functionally mature at P18, when the auditory function (sensitivity and frequency selectivity) is mature in this species

miRNA expression detected by q-PCR and <i>in situ</i> hybridization.

<p>A: Comparison of changes in miRNA expression detected by q-PCR versus microarray analyses for four miRNAs in the SV of C57 and CBA mice. Asterisks indicate statistically significant differences (p<0.05) compared to P21. Each q-PCR plot represents means from three repeats. B: Expression of four miRNAs in the LW of C57 mice using <i>in situ</i> hybridization technique. SV: Stria vascularis. SL: Spiral ligament. 1, 2, and 3 mark the three cell types in the SV (marginal, intermediate, and basal cells), respectively. Bar: 20 µm.</p

Differentially expressed miRNAs in the LW of CBA mice during aging.

<p>The X-axis relates Log2 transformed fold change in miRNA expression at 9 m (in blue) and 16 m (in red) compared to P21. Only those miRNAs whose levels were statistically different (p<0.05) from P21 were included.</p

Comparative analysis of sigma factors RpoS, FliA and RpoN in Edwardsiella tarda

Sigma factors are important regulators that bacteria employ to cope with environmental changes. Studies on the functions of sigma factors have uncovered their roles in many important cellular activities, such as growth, stress tolerance, motility, biofilm formation, and virulence. However, comparative analyses of sigma factors, which examine their common and unique features or elucidate their cross-regulatory relationships, have rarely been conducted for Edwardsiella tarda. Here, we characterized and compared motility and resistance to oxidative stress of E. tarda strains complemented with rpoS, fliA, and rpoN mutants. The results suggest that the sigma factors, FliA and RpoN, regulated motility, whereas RpoS exhibited no such function. RpoS and RpoN were essential for oxidative stress resistance, whereas FliA had no obvious impact under oxidative stress conditions. Furthermore, 2-dimensional gel electrophoresis (2-DE) based proteomics analysis, combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS/MS), revealed 12 differentially expressed protein spots that represented 11 proteins between the mutant and wild-type strains. Quantification of the expression of target genes by quantitative reverse transcription PCR (qRT-PCR) confirmed the results of our proteomics analysis. Collectively, these results suggest that these sigma factors are multi-functional mediators involved in controlling the expression of many metabolic pathway genes.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Characterization of quinoxaline derivatives for protection against iatrogenically induced hearing loss

Hair cell loss is the leading cause of hearing and balance disorders in humans. It can be caused by many factors, including noise, aging, and therapeutic agents. Previous studies have shown the therapeutic potential of quinoxaline against drug-induced ototoxicity. Here, we screened a library of 68 quinoxaline derivatives for protection against aminoglycoside-induced damage of hair cells from the zebrafish lateral line. We identified quinoxaline-5-carboxylic acid (Qx28) as the best quinoxaline derivative that provides robust protection against both aminoglycosides and cisplatin in zebrafish and mouse cochlear explants. FM1-43 and aminoglycoside uptake, as well as antibiotic efficacy studies, revealed that Qx28 is neither blocking the mechanotransduction channels nor interfering with aminoglycoside antibacterial activity, suggesting that it may be protecting the hair cells by directly counteracting the ototoxin’s mechanism of action. Only when animals were incubated with higher doses of Qx28 did we observe a partial blockage of the mechanotransduction channels. Finally, we assessed the regulation of the NF-κB pathway in vitro in mouse embryonic fibroblasts and in vivo in zebrafish larvae. Those studies showed that Qx28 protects hair cells by blocking NF-κB canonical pathway activation. Thus, Qx28 is a promising and versatile otoprotectant that can act across different species and toxins

Network analysis of the relationship between differentially expressed miRNAs and mRNAs in the LW.

<p>A: Upregulated apoptosis-related genes predicted to be targeted by downregulated miRNAs. B: Downregulated apoptosis-related genes predicted to be targeted by upregulated miRNAs. Small circles represent downregulated miRNAs or mRNAs while large circles represent upregulated miRNAs or mRNAs. Purple and orange circles denote differentially expressed miRNAs in C57 and CBA mice, respectively. Red and blue circles represent pro-apoptotic and anti-apoptotic genes, respectively.</p