Phenotype and Genetics of Progressive Sensorineural Hearing Loss (Snhl1) in the LXS Set of Recombinant Inbred Strains of Mice

Progressive sensorineural hearing loss is the most common form of acquired hearing impairment in the human population. It is also highly prevalent in inbred strains of mice, providing an experimental avenue to systematically map genetic risk factors and to dissect the molecular pathways that orchestrate hearing in peripheral sensory hair cells. Therefore, we ascertained hearing function in the inbred long sleep (ILS) and inbred short sleep (ISS) strains. Using auditory-evoked brain stem response (ABR) and distortion product otoacoustic emission (DPOAE) measurements, we found that ISS mice developed a high-frequency hearing loss at twelve weeks of age that progressed to lower frequencies by 26 weeks of age in the presence of normal endocochlear potentials and unremarkable inner ear histology. ILS mice exhibited milder hearing loss, showing elevated thresholds and reduced DPOAEs at the higher frequencies by 26 weeks of age. To map the genetic variants that underlie this hearing loss we computed ABR thresholds of 63 recombinant inbred stains derived from the ISS and ILS founder strains. A single locus was linked to markers associated with ISS alleles on chromosome 10 with a highly significant logarithm of odds (LOD) score of 15.8. The 2-LOD confidence interval spans ∼4 Megabases located at position 54–60 Mb. This locus, termed sensorineural hearing loss 1 (Snhl1), accounts for approximately 82% of the phenotypic variation. In summary, this study identifies a novel hearing loss locus on chromosome 10 and attests to the prevalence and genetic heterogeneity of progressive hearing loss in common mouse strains

Salsalate treatment following traumatic brain injury reduces inflammation and promotes a neuroprotective and neurogenic transcriptional response with concomitant functional recovery

Neuroinflammation plays a critical role in the pathogenesis of traumatic brain injury (TBI). TBI induces rapid activation of astrocytes and microglia, infiltration of peripheral leukocytes, and secretion of inflammatory cytokines. In the context of modest or severe TBI, such inflammation contributes to tissue destruction and permanent brain damage. However, it is clear that the inflammatory response is also necessary to promote post-injury healing. To date, anti-inflammatory therapies, including the broad class of non-steroidal anti-inflammatory drugs (NSAIDs), have met with little success in treatment of TBI, perhaps because these drugs have inhibited both the tissue-damaging and repair-promoting aspects of the inflammatory response, or because inhibition of inflammation alone is insufficient to yield therapeutic benefit. Salsalate is an unacetylated salicylate with long history of use in limiting inflammation. This drug is known to block activation of NF-jB, and recent data suggest that salsalate has a number of additional biological activities, which may also contribute to its efficacy in treatment of human disease. Here, we show that salsalate potently blocks pro-inflammatory gene expression and nitrite secretion by microglia in vitro. Using the controlled cortical impact (CCI) model in mice, we find that salsalate has a broad antiinflammatory effect on in vivo TBI-induced gene expression, when administered post-injury. Interestingly, salsalate also elevates expression of genes associated with neuroprotection and neurogenesis, including the neuropeptides, oxytocin and thyrotropin releasing hormone. Histological analysis reveals salsalate-dependent decreases in numbers and activation-associated morphological changes in microglia/macrophages, proximal to the injury site. Flow cytometry data show that salsalate changes the kinetics of CCI-induced accumulation of various populations of CD11b-positive myeloid cells in the injured brain. Behavioral assays demonstrate that salsalate treatment promotes significant recovery of function following CCI. These pre-clinical data suggest that salsalate may show promise as a TBI therapy with a multifactorial mechanism of action to enhance functional recovery

<i>Snhl1</i> phenotype/genotype correlation.

<p>ABR thresholds at the 32 kHz stimulus of each of the 63 RI strains (grey) and the parental ILS (green) and ISS (red) strains as a function of their genotype at markers <i>rs3682060</i>, <i>rs13480620</i>, and <i>Cdh23⁷⁵³</i> are shown. ABR thresholds (in dB SPL; Y-axis) are given as mean ± SD (n = 4). The grey box denotes the number of strains and threshold range unexplained by <i>Snhl1</i>. The ISS allele is shown in green (C, cytosine; G, guanine) and the ILS allele is represented in red (T, thymidine; A, adenine) boxes. The number of the RI strain carrying a recombinant chromosome and the threshold at the 32 kHz is shown below the haplotypes. On the right, the physical location of the marker on chromosome 10 in base pairs (bp) is given and the most likely location of <i>Snhl1</i> is indicated.</p

ABR threshold distribution in the LXS RI set.

<p><b>A–D.</b> Histograms showing threshold distributions at the click (<b>A</b>), 8- (<b>B</b>), 16- (<b>C</b>), and 32 kHz (<b>D</b>) stimuli. The Y-axis represents the number of RI strains, and the X-axis denotes the threshold level. Histograms were fitted with a normal Gaussian distribution. r²  =  goodness-of-fit.</p

Inner ear histology in ILS and ISS mice.

<p>Tolouidin blue-stained plastic sections through the cochlear duct in twelve-week-old ILS and ISS and eight-week-old C3HeB/FeJ (C3H) mice. <b>A,F,K,</b> Cross section through the organ of Corti at the mid-apical region; tm, tectorial membrane; oh, outer hair cell; ih, inner hair cell; sc, supporting cell; tC, tunnel of Corti; sM, scala media; scale bar  =  50 µm. <b>B,G,L,</b> Cross section through the spiral ligament at the mid-apical region of the cochlear duct. White arrow indicates a fibrocyte. sL, spiral ligament; scale bar  =  50 µm. <b>C,H,M,</b> Cross section through the stria vascularis at the mid-apical region of the cochlear duct. Red arrow, basal cell; white arrow, intermediate cell; green arrow, marginal cell; sV, stria vascularis; scale bar  =  10 µm. <b>D,I,N,</b> Cross section through the spiral ganglion near the base of the cochlear duct. Red arrow points to areas of degeneration. sG, spiral ganglion; scale bar  =  50 µm. <b>E,J,O,</b> Cross section through the spiral ganglion at the mid-apical region of the cochlear duct. White arrow points to a neuron and the red arrow indicates a Schwann cell. sG, spiral ganglion; scale bar  =  50 µm.</p

Auditory characteristics of ILS and ISS mice.

<p><b>A.</b> ABR thresholds at click (c) and pure tone pips at 8-, 16-, and 32 kHz at twelve and 26 weeks (wks) of age. Data are given as mean ± SD. <b>B.</b> Wave I ABR amplitudes at 8- and 16 kHz stimuli at 60 dB SPL input levels. Data are given as mean ± SD. µV, microvolt. <b>C.</b> Latencies of ABR waves I – V at a 16 kHz stimulus of 60 dB SPL. Data are given as mean ± SD; msec, milliseconds. <b>D.</b> DPOAE output levels at 2f1-f2 in dB SPL over f2 frequency range 6–56 kHz (left panel). The right panel shows I/O function at f2 = 16 kHz. Data are given as mean ± SEM. <b>E.</b> Endocochlear potentials. Data are given as mean ± SD; mV, milliVolt. For all panels: C3HeB/FeJ (blue), ILS (green) and ISS (red); **<i>p</i><0.01; ***<i>p</i><0.001.</p

Results of genome-wide regression analysis.

<p>kHz, kiloHertz; QTL, Quantitative trait locus; Chr, chromosome;</p

ABR thresholds in ILS and ISS strains.

<p>C3H, C3HeB/FeJ; wks, weeks of age; kHz, kiloHertz; SD, standard deviation; <i>p</i>, ANONVA; n.s., not significant; n, number of animals tested;</p

Controlled cortical impact and craniotomy induce strikingly similar profiles of inflammatory gene expression, but with distinct kinetics

An immediate consequence of traumatic brain injury (TBI) is the induction of an inflammatory response. Mounting data suggest that inflammation is a major contributor to TBI-induced brain damage. However, much remains unknown regarding the induction and regulation of the inflammatory response to TBI. In this study we compared the TBI-induced inflammatory response to severe parenchymal injury (controlled cortical impact) vs. mild brain injury (craniotomy) over a 21 day period. Our data show that both severe and mild brain injury induce a qualitatively similar inflammatory response, involving highly-overlapping sets of effector molecules. However, kinetic analysis revealed that the inflammatory response to mild brain injury is of much shorter duration than the response to severe TBI. Specifically, the inflammatory response to severe brain injury persists for at least 21 days, whereas the response to mild brain injury returns to near baseline values within 10 days post-injury. Our data therefore imply that the development of accurate diagnostic tests of TBI severity that are based on imaging or biomarker analysis of the inflammatory response may require repeated measures over at least a 10 day period, post-injury