This present study examined the relationship between cognitive measures and self-report hearing aid outcome. A sentence-final word identification and recall (SWIR) test was used to investigate how hearing aid use may relate to experienced explicit cognitive processing. A visually based cognitive test battery was also administered. To measure self-report hearing aid outcome, the International Outcome Inventory – Hearing Aids (IOI-HA) and the Speech, Spatial and Qualities of Hearing Scale (SSQ) were employed. Twenty-six experienced hearing aid users (mean age of 59 years) with symmetrical moderate-to-moderately severe sensorineural hearing loss were recruited. Free recall performance in the SWIR test correlated negatively with item 3 of IOI-HA, which measures residual difficulty in adverse listening situations. Cognitive abilities related to verbal information processing were correlated positively with self-reported hearing aid use and overall success. The present study showed that reported residual difficulty with hearing aid may relate to experienced explicit processing in difficult listening conditions, such that individuals with better cognitive capacity tended to report more remaining difficulty in challenging listening situations. The possibility of using cognitive measures to predict hearing aid outcome in real life should be explored in future research. Speech audiometry is widely used in research and clinics to objectively evaluate different hearing aid models and settings (e.g. compression and noise reduction), as well as hearing aid fittings. However, conventional speech tests may not be sensitive in predicting hearing aid outcome in daily life . TherefoClinically, there are a number of questionnaires that quantify hearing aid outcome on different dimensions. These include the Satisfaction with Amplification in Daily Life SADL; ; the AbbQuestions regarding the ability to perceive or to understand speech in different listening conditions when listening is aided and/or unaided are common to all the aforementioned questionnaires. Cognition plays a role in both speech perception and speech understanding, particularly in adverse listening conditions  test  at 84% speech intelligibility was obtained using the Swedish Hearing In Noise Test (HINT) . After the final word of the eighth sentence was reported, they were prompted to recall all the final words that they had previously reported . In this paper, we report the recall performance from the five background conditions analyzed in This battery assesses verbal information processing speed, lexical access speed, quality of phonological representations and working memory capacity. All have been found to be important for speech understanding under adverse listening conditions  . This teThe task was to judge whether a string of three letters shown was a real Swedish word  or not . This test measures lexical access speed and is scored based on reaction time for correct trials.The test was to judge whether two words of equal length rhymed or not . There wThis test consisted of two parallel tasks. First, the participants had to judge, whether three-word sentences shown at the centre of a computer screen, at a rate of 800 ms per word with an inter stimulus interval of 75 ms, were sensible or absurd  equipped with an IEC 711 ear simulator. The amplified signal was presented to a pair of ER3A insert earphones (see n = 25) and SSQ (n = 26) are generally comparable (within 1 SD) to published studies on hearing aid users with more than 6 months of experience . No significant correlations were found for any IOI-HA item with SRT. Table all background conditions, with the exception of the 4T/NoP condition, correlated negatively with item 3 of IOI-HA, which measures residual difficulty in adverse listening situations. Correlations with the rest of the items were not statistically significant. Furthermore, factor 2 of IOI-HA, which measures hearing handicap, also correlated with recall performance in the SSN/NoP and 4T/NR.The results of the free recall task in the SWIR test in the five background conditions were reported in P = 0.013), which reinforces the specificity of the pattern of results.In order to reduce the number of comparisons in the correlation analysis, the results of the memory task in all five background conditions were pooled to give a composite score. The relationship between the free recall composite score and IOI-HA was re-examined Table . Again, Table Physical matching test: the reaction time measure significantly correlated with the SSQ subscale Localization , indicating that faster processing speed was associated with better self-reported sound localization ability. The Lexical decision making test: the reaction time measure significantly correlated with item 1 and factor 1 of the IOI-HA , indicating that faster lexical processing speed was related to more frequent hearing aid use and better overall hearing aid benefit, yet greater overall reported residual difficulties with hearing aid, and less experienced listening effort. The Rhyme judgment test: the accuracy measure (percent correct) significantly correlated with items 1 and 3 of the IOI-HA , indicating that individuals who used hearing aids more frequently and with better phonological processing reported more remaining difficulties experienced in a challenging listening situation. The correlation with IOI-HA factor 1 was on the verge of significance . The Reading Span test: although not statistically significant, a tendency of correlation was observed between the reading span test and item 3 of IOI-HA .Some of the IOI-HA items and one SSQ subscale significantly correlated with performance on the following cognitive tests: The Summarizing the findings, we demonstrated that cognitive measures used in the present study were related to self-reported outcome with own hearing aids and settings. The ability to recall the final words of heard sentences amplified and processed with an experimental hearing aid in all but one of the background conditions was related to reported remaining difficulty in an adverse listening situation (IOI-HA item 3), and performance in two background conditions was related to the overall reported residual difficulty (IOI-HA factor 2). The quality of phonological representations in lexicon, which was measured by the rhyme judgment test, was also related to IOI-HA item 3. These relationships were negative, which mean that individuals with better recall performance reported more remaining difficulties in an adverse listening situation. On the other hand, better overall subjective benefit (IOI-HA factor 1) and more frequent hearing aid usage (IOI-HA item 1) were found to be associated with faster lexical access speed and better quality of phonological representations. It is also worthy of note that the overall recall performance was positively associated, though not significantly, with IOI-HA factor 1. To summarize, cognitive abilities relating to verbal information processing were inversely related to the perceived handicap or residual difficulty experienced in real life, but positively associated with the reported benefits and usage of hearing aid.Our results suggested that individuals with better recall performance in the SWIR test consistently reported more remaining difficulties in daily situations in which they wanted to hear better. Although we expected that persons having good cognitive abilities would consequently experience less (remaining) problems with hearing aids in daily situations, hence reporting less hearing handicap, our subjective outcome data did not show the expected pattern. One way of explaining the clear and specific pattern of results here is that the self-reported remaining difficulties could actually indicate the relative degree of engagement of explicit processing resources in working memory. In an adverse situation where the hearing aid user wants to hear better, listening is difficult. With the application of appropriate hearing aid amplifications and signal processing algorithms, listening, or even speech communication can be improved. However, hearing aids may partially but not entirely reduce experienced listening difficulties. Thus, the hearing aid user may still encounter or experience remaining difficulties even when listening with a hearing aid, and this is measured by item 3 of the IOI-HA. We assume that the situation described by item 3 is an adverse listening situation, such as when the target speech is degraded or presented in the presence of competing noise. High performers on the free recall task (SWIR) have better cognitive spare capacity  between item 3 of the IOI-HA and the reading span test. Item 3 was found to be significantly correlated with the recall performance of the SWIR test. For the same group of participants, the recall performance of the SWIR test was significantly correlated with the reading span test , which have been found to be associated with speech recognition thresholds in previous studies. No question pinpoints the overall difficulties or even remaining difficulties in listening in adverse conditions, in which working memory is taxed substantially. Thus, the SSQ was found to be less effective than IOI-HA in revealing the relationship between self-report and cognitive measures in the present study.In the other self-report outcome measure used in the present study (SSQ), there were correlations between speech reception threshold and five subscales  and only one signification correlation was found between general processing speed and the Since speech tests alone cannot always satisfactorily predict self-reported benefits , furtherThis study demonstrated a relationship between cognitive measures and residual difficulty reported by hearing aid users. The results showed that reported residual difficulty with hearing aids may relate to experienced explicit processing in difficult listening conditions, such that individuals with better cognitive capacity tended to report more remaining difficulty in challenging listening situations. Cognitive abilities related to verbal information processing are also associated with self-reported hearing aid use and overall success, such that individuals with better cognitive abilities reported better hearing aid use and overall success. The possibility of using cognitive measures to predict hearing aid outcome in real life should be explored in future.


Drosophila melanogaster. We monitored the grooming responses of decapitated flies to compounds known to activate the immune system, e.g., dead Escherichia coli (Ec) and lipopolysaccharides (LPS), and to tastants such as quinine, sucrose, and salt. LPS, quinine, and Ec were quite effective in triggering grooming movements when touching the distal border of the wings and the legs, while sucrose had no effect. Contact chemoreceptors are necessary and sufficient to elicit such responses, as grooming could not be elicited by LPS in poxn mutants deprived of external taste sensilla, and as grooming was elicited by light when a channel rhodopsin receptor was expressed in bitter-sensitive cells expressing Gr33a. Contact chemoreceptors distributed along the distal border of the wings respond to these tastants by an increased spiking activity, in response to quinine, Ec, LPS, sucrose, and KCl. These results demonstrate for the first time that bacterial compounds trigger grooming activities in D. melanogaster, and indicate that contact chemoreceptors located on the wings participate in the detection of such chemicals.In social insects, grooming is considered as a behavioral defense against pathogen and parasite infections since it contributes to remove microbes from their cuticle. However, stimuli which trigger this behavior are not well characterized yet. We examined if activating contact chemoreceptive sensilla could trigger grooming activities in  Insects, especially Diptera, devote a considerable amount of time to self-grooming. Grooming involves brushing the body and the wings with the legs, and cleaning the legs and the antenna with the mouthparts. In cockroaches, grooming helps cleaning external chemosensory receptors  and lipopolysaccharides (LPS), and with non-volatile chemicals known to be either aversive (quinine), appetitive (sucrose) or both depending on the concentration (NaCl and KCl)  flies. Flies devoid of external taste sensilla were Poxn70/Cyo; MKRS, Sb/TM6B, Tb (Poxn70).Gr33a-Gal4 construction (generously given by J. Carlson) with flies carrying UAS-channel rhodopsin (UAS-CHR2) . Since the balancer chromosome of this construction carried the defect curly (CyO), we could select in the progeny individuals which did not express the construction  and others (siblings) which expressed the phenotype. For Gr33a, the genotype was Gr33a-Gal4[1] /CyO; Dr/TM3, Sb, Ser × UAS-H134R-CHR2, where the siblings expressed the curly wing phenotype and the mutants had normal wings. During development, the larvae were fed on normal medium added with 1 mM trans-retinal . This suspension was subsequently diluted 100, 102, 104, and 106 fold.The TOP 10 strain of Escherichia coli 0111:B4, Sigma), sucrose, quinine, NaCl, and KCl were provided by Sigma-Aldrich and dissolved in distilled water.LPS . 4-day old flies were used for all tests.For optogenetic experiments, flies were exposed to a continuous pulse of blue light for 3 min, over the whole body. Headless flies were placed on a filter paper as usual, but the experiment was conducted in a dark condition. The intensity of the grooming response was scaled from 0 to 5  placed over the abdomen. Recordings were performed on the wing part near LV2 . Taste bristles of the wing margin  were stimulated by covering their tip with a glass electrode containing an electrolyte (1 mM KCl) and the stimulus during 2 s. To avoid adaptation, consecutive stimulations were applied at least 2 min apart. The stimulus tested were E. coli, LPS, sucrose, quinine, and NaCl at the same concentrations as in the grooming tests (see above). As a control, we used increasing concentrations of KCl . Recordings were performed on 10 females and 10 males for each stimulus category.Flies (4-day old) were secured to a support with tape and electrically grounded via a silver electrode contacting a drop of electrocardiogram gel   was applied. For the analysis of fly visits, in the 2-choices assay, a Dunnett test was applied. Optogenetic induction of grooming Decapitated flies are capable of self-grooming movements following a stimulation. In order to induce this behavior, we gently brushed their wing margin or one of their leg with the tip of a toothpick. Flies which never exhibited grooming in response to stimulation with a toothpick dipped into a chemical stimulus never showed grooming after stimulation with water alone. These movements mostly involved the meta-thoracic legs which were raised and moved independently in a succession of strokes, brushing the wings, the abdomen and the dorsum or which were extended under the abdomen and touched each other in a series of reciprocal sliding movements  and LPS induced grooming responses in 4-day old adults , the front legs (p < 0.001), and the hind legs (p < 0.001). Responses to LPS were obtained from the wings (p < 0.001) but not from the legs .Dead s Figure . Responsp = 0.692) did not trigger any grooming response at any concentration tested . Quinine elicited a more intense grooming response than NaCl, and also triggered a dose-dependent response of the frontlegs (p = 0.002) but not of the hindlegs (p = 0.696). NaCl was not effective as quinine, but it induced a dose-dependent response from the hindleg (p = 0.008). The other tastants did not trigger responses when brought in contact with the legs (p > 0.1). There were no obvious differences between genders , a bitter (quinine), and a salty one (NaCl). As in the previous experiments, water did not induce grooming responses. Also, sucrose  or of the legs  and females (p = 0.044) in the absence of any chemosensory stimulus. We found a sexual dimorphism in the flies carrying the construction as photoactivation induced more grooming in males than in females . With LPS, flies avoided walking on side treated with LPS  , to the concentration (p < 0.001), to sex (p < 0.001), and according to the sensillum position in the row . However, we did not find any difference between dorsal and ventral sensilla . These observations confirm that wing contact chemoreceptive sensilla are functional, that they respond to classical taste stimuli. These sensilla show dose-dependent responses to LPS and E. coli suspensions. They also respond in a dose-dependent way to NaCl, to KCl, to quinine, but apparently not to sugar.We thus counted the total number of spikes occurring within the first 2 s of the stimulation Figure . When anp < 0.001 for all chemicals except KCl, p = 0.002 in males and p < 0.001 in females and NaCl, p = 0.254 in males and p = 0.420 in females, analysis of covariance). However, with sucrose, the initial burst peaked at 30–35 spikes/s which may indicate that 1 mM sucrose already elicits a response. As the concentration of each stimulus increased, the firing rate globally increased. For salts, it was mostly the tonic part of the response which was affected, while with quinine, E. coli and LPS both the phasic and tonic parts of the responses increased with the dose tested. On the contrary, the spiking activities were markedly depressed in both sexes when the concentration of sucrose was raised from 1 mM to 1 M.We further analyzed the temporal course of the responses by computing the number of spikes occurring during consecutive 100 ms bins Figures , S2. At D. melanogaster adults are triggered by bacterial suspensions of E. coli, its surface compound, LPS, and by aversive chemicals like quinine and NaCl at a high concentration. Mechanosensation is not required to elicit this response as sucrose and water did not induce grooming, and Poxn70 mutants deprived of their external taste receptors were insensitive to LPS. Furthermore, grooming activities could be elicited without physical contact with a substrate by optogenetic activation of cells expressing the bitter receptor Gr33a. Since our behavioral observations indicated that grooming reactions were elicited by stimulating the costal outer vein of the wings, we examined the contact chemoreceptors located there using tip-recording. These sensilla were found to house contact chemoreceptive neurons increasing their firing activity in response bacterial extracts, LPS, quinine, and to high salt concentrations thus supporting the hypothesis that these wing contact chemoreceptors contribute to triggering grooming responses in decapitated flies.We demonstrate for the first time that grooming activities in In this work, we recorded grooming by scoring the intensity of the behavior in relation to the number of strokes made by the legs and by the duration of this reaction. A more detailed analysis of this behavior was not relevant here as we focused on finding chemical stimuli which could trigger this behavior. In order to simplify the experimental procedures, we also observed the reactions of decapitated insects. In such insects, the responses induced bypass the downstream control normally exerted by higher order nervous centers in intact animals. Our observations are thus by no mean representative of the normal grooming behavior but they give us a good indication of the strength of the sensory stimuli sent to the local ganglia.While self-grooming has been described to occur in response to touch  to drive the expression of a calcium-sensitive probe, G-cAMP (Nakai et al., Gr family, thus confirming that contact chemoreceptive sensilla of the wings of Drosophila are fully functional and respond at least to some sapid molecules detected by contact chemoreceptors located on the legs and on the proboscis.Raad  introducIn our tests, sucrose is the only stimulus which did not elicit grooming in our panel of stimuli. The electrophysiological responses obtained from wing taste sensilla did not show any increase of the firing activity with the dose, but on the contrary revealed an inhibition of the basal firing activity. This observation is consistent with recent results obtained by Raad  on fly wD. melanogaster. Our study shows that LPS induces vigorous grooming activities from flies and also elicits consistent spiking activities in contact chemoreceptors of the wing. LPS has been used repeatedly in multiple organisms as an inducer of the immune system. LPS and bacterial peptidoglycans are strong inducers of the immune system in Drosophila (Charroux et al., So far, our findings support that contact chemicals play a decisive role in triggering hygiene grooming in The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


There is currently no therapeutic drug treatment for traumatic brain injury (TBI) despite decades of experimental clinical trials. This may be because the mechanistic pathways for improving TBI outcomes have yet to be identified and exploited. As such, there remains a need to seek out new molecular targets and their drug candidates to find new treatments for TBI. This review presents supporting evidence for cathepsin B, a cysteine protease, as a potentially important drug target for TBI. Cathepsin B expression is greatly up-regulated in TBI animal models, as well as in trauma patients. Importantly, knockout of the cathepsin B gene in TBI mice results in substantial improvements of TBI-caused deficits in behavior, pathology, and biomarkers, as well as improvements in related injury models. During the process of TBI-induced injury, cathepsin B likely escapes the lysosome, its normal subcellular location, into the cytoplasm or extracellular matrix (ECM) where the unleashed proteolytic power causes destruction via necrotic, apoptotic, autophagic, and activated glia-induced cell death, together with ECM breakdown and inflammation. Significantly, chemical inhibitors of cathepsin B are effective for improving deficits in TBI and related injuries including ischemia, cerebral bleeding, cerebral aneurysm, edema, pain, infection, rheumatoid arthritis, epilepsy, Huntington’s disease, multiple sclerosis, and Alzheimer’s disease. The inhibitor E64d is unique among cathepsin B inhibitors in being the only compound to have demonstrated oral efficacy in a TBI model and prior safe use in man and as such it is an excellent tool compound for preclinical testing and clinical compound development. These data support the conclusion that drug development of cathepsin B inhibitors for TBI treatment should be accelerated. Traumatic brain injury (TBI) occurs when an external force, such as that due to a vehicular accident, a football collision, or a bullet, causes brain dysfunction and pathology. Unfortunately, TBI is all too common with over 10 million people worldwide afflicted each year , 3. FallTBI encompasses a continuum of injuries and pathologies and is symptomatically classified into mild, moderate, or severe based on the level of patient consciousness , 7. MuchBut despite this knowledge, there is currently no pharmaceutically effective treatment for TBI even though many experimental clinical trials have been conducted over decades to find such –28. ThisStreptococcus pneumoniae meningitis , was given to the protease activity in an acidic environment. Those proteases were subsequently found to be of different catalytic types and include the serine proteases cathepsins A and G, the aspartic proteases cathepsins D and E, and the lysosomal papain-like cysteine proteases , 71.In man, cathepsin B is encoded as a single gene on chromosome 8 at position p22-23.1 , spans 2Human cathepsin B messenger ribonucleic acid (mRNA) consists of multiple messages that differ in their 5′ and 3′ untranslated regions (UTRs) and arise by alternative splicing. Most tissues express 2.3 and 4.0 kilobase (kb) transcripts at a ratio of 2:1; but the ratio of mRNAs with variant 5′ UTRs differs widely . Normal Small, non-coding microRNA (miRNA) inhibit the expression and function of endogenously encoded proteins . The miRCathepsin B post-translational processing is well understood and has been the subject of many reviews , 70, 87.Proteases are stored in the cell as inactive zymogens that require hydrolysis for activation, and allow cells to rapidly deploy active protease as needed . In the As shown in Table Table TBI causes many vascular pathologies including ischemia, aneurysms, subarachnoid hemorrhages, and edema –15. IncrBrain infections are a common TBI complication  and cathFinally, as discussed above, TBI is a risk factor for several neurodegenerative diseases. Cathepsin B mRNA is elevated 3.5-fold relative to controls in spinal cord samples from an ALS mouse model . MoreoveWe are not aware of any clinical studies on acute cathepsin B regulation due to TBI, but non-brain polytrauma patients show increases in plasma cathepsin B activity during the first day after trauma, which subsequently falls to moderately elevated levels by the third day, and remains roughly at that level for up to 2 weeks. Importantly, the increase in plasma cathepsin B activity correlates with the severity of injury. Patients with a sixfold increase in plasma cathepsin B activity 1 day post-trauma subsequently developed fatal or reversible multiple organ failure, whereas those who had only a threefold increase, at the same day, did not display organ failure , 124.Moreover, in human cerebral aneurysm tissue, cathepsin B is highly expressed in the endothelial cell layer and the media in the aneurysmal walls in contrast to control artery tissue where it is barely expressed at all . Thus, cPatients having the chronic inflammatory neurological diseases Guillain–Barre syndrome, chronic demyelinating polyneuropathy, or MS have higher cerebrospinal fluid (CSF) cathepsin B activity levels than controls , 126. CaIn the peripheral inflammatory conditions of RA and osteoarthritis (OA) synovial cells and chondrocytes have increased cathepsin B mRNA levels and cathepsin B protein secretion relative to controls , 134. FuThe health of cathepsin B gene knockout mice is maintained and generally indistinguishable from normal littermates in behavior, histology, and fertility , as showTable Deleting the cathepsin B gene produces significant beneficial outcomes in TBI-related pathological animal models and includes surgery, epilepsy, AD, inflammation, pain, and cytokine cell death models. Deletion of both cathepsin B and cathepsin S improves the outcomes in an MS model  is a cytokine that induces a cytoplasmic molecular cascade including cathepsin B release from lysosomes and apoptotic cell death  and TBI TBI has been associated with an increased risk of subsequent MS development in the Chinese population . In an eThe cathepsin B gene knockout mice show little or no adverse impact on normal functions, but have major beneficial effects in TBI, including reductions in neuromotor deficits, brain pathology, and neuronal cell death. Cathepsin B deficiency produces substantial improvement in important pathologies related to TBI, including reductions in ECM breakdown, neuroexcitatory-induced cell death, inflammation, inflammatory pain, TNFα-induced cell death, Aβ levels, and memory deficits.It is likely that the absence of developmental defects associated with cathepsin B deficiency is due to redundant protease specificity of the closely related protease, cathepsin L, which can substitute for the normal function of cathepsin B. Evidence in support of this hypothesis is that cathepsin B and L double knockouts are lethal and have profound neurological abnormalities , 163. ThThe large volume of data from the cathepsin B knockout mice indicates that this protease is central to many different pathological processes. Many benefits occur from this deficiency, leading to the compelling conclusion that cathepsin B inhibition is a promising therapeutic approach for TBI.Early studies demonstrated that cathepsin B is present in human tissues throughout the body , but thahttp://www.proteinatlas.org/). Moreover, the amount of cathepsin B expression varies among different tissues and cell types.An on-line human protein tissue atlas  also shoEarly immuno-microscopic analysis showed that human brain contains cathepsin B and it is concentrated in neuronal cell types, especially in the hippocampus , 168–171Early cathepsin B mRNA analysis of rat brain also showed cathepsin B expression concentrated in neurons  and moreTable A major destination for cathepsin B is the lysosome, the name of which is from the Greek lysis: loosen and soma: body. Christian de Duve discovered that these subcellular structures were recycling centers in which unwanted proteins are dismantled to their amino acid building blocks for recycling into new proteins for which he received the 1974 Nobel Prize in Physiology or Medicine . LysosomIn addition to digestive lysosomes, cathepsin B is also trafficked to secretory lysosomes, which release their contents to the extracellular environment in response to a triggering event. These lysosomes are primarily found in immunological cell types and brain astrocytes , 183. CoBut lysosomal cathepsin B can also cleave proteins at specific recognition sites to produce protein fragments having important biological activities. For example, cathepsin B cleaves in the lysosome non-phosphorylated myristoylated alanine-rich C kinase substrate (MARCKS), which is the primary substrate for protein kinase C (PKC), to prevent phosphorylation and thereby control cell morphology and motility –187. In Autophagosomes are structures in which unnecessary or dysfunctional cellular components are degraded. Lysosomes fuse with autophagosomes to provide the proteases needed for that degradation and cathepsin B is a key protease in autophagy catabolism .Lysosomes are not the only subcellular destination for cathepsin B . It is aFrom a TBI point of view, the important point is that cathepsin B is normally mostly contained within subcellular organelles and this compartmentalization allows the cell to safely utilize the proteolytic powers of cathepsin B without damaging the cell structure. As discussed below, that segregation is compromised after brain trauma leading to cathepsin B-induced injury and pathology.Cathepsin B zymogen is activated by cathepsins in the lysosome/endosome compartments where they have optimum efficiency in the acidic environment of about pH 5.0 and the papain-like cysteine proteases require a reducing environment . The lysIn vitro, cathepsin B exopeptidase activity predominates below pH of 5.5 and endopeptidase activity prevails above pH 5.5  when exposed to cystatin C at the concentrations in the extracellular fluids is less than a second  family pyrin domain-containing 3 (NLRP3) by cathepsin release into the cytoplasm . As discApoptotic cell death cascade occurs following trauma and cathepsin B is a key component of that cascade , 98. TNFStudies in hepatocytes from cathepsin B knockout animals that show the TNFα-induced lysosomal leakage of cathepsin B to the cytosol cleaves Bid and activates caspase 2, which facilitate mitochondrial release of cyt c to the cytosol and that triggers caspase apoptotic cell death , 161. TrCathepsin B is secreted by inflamed microglia and induces apoptosis. Trauma to the CNS causes diverse inflammatory cells expressing high levels of cathepsin B, which include microglia and macrophages, to gather at a site of injury . InflammTBI can cause massive cellular and cytokine inflammatory responses that focus on cathepsin B as the center of TBI-caused damages. As discussed above, cathepsin B is clearly involved in the production of IL-1β by activating caspase 1, which in turn cleaves pro-IL-1β to produce active IL-1β  as has bPeroxisome proliferator-activated receptor (PPAR) receptors are key regulators of neuroinflammation after CNS injury, including TBI . PPARα aPathological axonal damage is a predictor of outcome in CNS diseases  and cathTBI increased autophagy activation within 1 h and up to at least 3 days after trauma in the brain relative to non-traumatized controls as measured by lipidated microtubule-associated protein light chain 3 (LC3II) , 238–241While it has been suggested that autophagy might provide neuroprotection after TBI , adminisTable The important conclusion from Table Compounds, which inhibit cathepsin B, also produced substantial beneficial effects in TBI-related injuries in animal models. For example, neuromotor, neurological learning and cognitive defects were improved by cathepsin B inhibition in cerebral bleeding , ischemiEdema is often a major complication of TBI and compounds, which inhibit cathepsin B, reduced edema in ischemic , 106 andThe data in Table Glia fibrillary acidic protein (GFAP), which is biomarker of CNS inflammation and a potential TBI diagnostic marker , is reduThese data support the conclusion that compounds, which inhibit cathepsin B, are effective in many TBI relevant animal models at improving behavior and pathology. As such, these data provide strong motivation to explore cathepsin B inhibitors as potential TBI therapeutic agents.Most of the preclinical and essentially all the clinical studies on small molecule inhibitors of cathepsin B investigated E64 and its derivatives E64d, E64c, CA-074Me, and CA-074. Table Without additional data, it is difficult to conclude that the beneficial TBI effects of E64d are due to cathepsin B inhibition because the biologically active acid form of E64d, which is E64c, inhibits proteases in addition to cathepsin B, which are the papain-like cysteine proteases and calpains 1 and 2 . However, the relative importance of cathepsin B inhibition can be resolved by evaluating E64d in cathepsin B gene-deficient animals. Those studies show that E64d treatment produced, with one exception, similar improvement in neuromotor deficits after trauma as did vehicle controlled treatment of cathepsin B knockout animals . Also, tThe E64d-treated cathepsin B knockout animals performed significantly (20%) better in the neuromotor assay than carrier-treated cathepsin B knockout animals on day 1 post-TBI . These rPrimary benefits due to inhibiting cathepsin B and the additional benefits from inhibiting related proteases have also been demonstrated in ischemic non-human primate models. CA-074, which has absolute specificity for cathepsin B , given iv immediately after ischemia significantly inhibited brain cathepsin B by about 75% in the CA1 region of the hippocampus and caused substantial neuronal protection by saving about 67% of the neurons in that region relative to controls . StudiesAnother advantage of multiple protease inhibition by E64d is the indirect reduction in matrix metallopeptidase-9 (MM-9), which is a protease that contributes to TBI dysfunction . Up-reguThe cysteine protease inhibitor, morpholinurea-leucine-homophenylalanine-vinylsulfone-phenyl (LHVS), provides significant improvements in behavioral deficits, brain edema, and cell survival following TBI . LHVS waThus, cathepsin B inhibition is a primary means by which E64d provides beneficial outcomes in TBI and ischemic models, but additional benefits are obtained by the compound’s inhibition of other related proteases, especially calpains, cathepsin L, and cathepsin S. The ability of E64d to inhibit multiple related cysteine proteases is thus an advantage and may be required to “move the needle” and affect outcomes in the complex condition of TBI.E64 and its related compounds as well as many other cathepsin B inhibitors are irreversible inhibitors meaning that they covalently bind to the traget protease and permanently stop that protease’s activity. As a result, cathepsin B activity lost due to these inhibitors can only be restored through production of new mature cathepsin B protein. Normally, cathepsin B turnover (net synthesis and degradation) as measured by the half-life is about 14 h and results in a lysosomal content of about 330 mg/g in rat macrophages , 92. E64Aspergillus japonicas selectively inhibits the papain-like cathepsin proteases (except cathepsin C) and calpains 1 and 2. But E64 does not affect cysteine proteases belonging to the CD clan, which includes caspases, or proteases belonging to the aspartyl, serine or metalloprotease classes or thiol enzymes , which was developed by Taisho Drug Company, Japan, and the Japanese Ministry of Health in the 1980s as an experimental therapeutic agent for treating muscular dystrophy . The cliBut importantly, extensive E64d pharmaceutical data were published as a result of this effort in many peer-reviewed Japanese scientific articles showing that the compounds could be safely used in man. Some of those data are summarized below .50) of E64c against rat liver cathepsins L, B, H, and porcine kidney calpain 2 is 0.09, 3.36, 1,640, and 3,000 nM, respectively, with the substrates Z-Phe-Arg-MCA, Z-Arg-Arg-MCA, Arg-MCA, and casein, respectively , which bind to the active site of proteases and report on activities via a biotin or quencher-fluorophore tag –289. ABPin vivo , 291 andin vivo . ABPs coAn important result from a TBI translational point of view is that oral administration of E64d (one 10 mg/kg dose) has been shown to be efficacious in a TBI animal model when given up to 8 h after trauma . That isFurthermore, administration of E64d by oral gavage to guinea pigs  resulted in dose–response reductions in brain cathepsin B activity and Aβ . And feeRadioactive E64d biodistribution studies show that about 60% of an oral E64d dose is taken up in rats and hamsters and most of that occurs in the small intestine. Blood levels quickly rise and the maximum blood concentration varies among animal species with rabbits having the highest, and hamsters, dogs, and rats having 75%, 33%, and 25% that of rabbits, respectively. The excretion also varies among species, with most occuring via the urine in rabbits and hamsters whereas the feces was the primary route in dogs and rats. Biliary excretion was greater than 50% and 13% of the dose in rats and hamsters, respectively. Respiration only accounted for about 7% of the dose .The tissue distribution was similar for rats, rabbits, and hamsters, but the highest drug concentration was in the gastrointestinal track followed by the kidney and liver which had about 25- and 12-fold higher concentrations than the plasma concentration, respectively. But the brain concentration was one-tenth that of the plasma concentration. However, the liver drug concentration in rats was about twice that of hamsters .At a cellular level, subcutaneous radioactive E64c injection of rats resulted in a muscle cytoplasm to lysosome-specific activity ratio of about 5:1 . That diPharmacokinetic properties of E64d have been determined in animals and man , 297. InIn vitro E64d mutagenic studies using the reverse mutation  and chromosome aberration test (Chinese hamster lung fibroblast cells) showed no mutations or chromosome aberrations. In vivo mutagenic studies using the mouse bone marrow chromosome aberration test showed that E64d did not have any effects in that assay. The conclusion was that E64d is unlikely to have mutagenic effects ( effects .50), with the main clinical effects being depression of voluntary movements, closed eye, and loss of righting reflex, from which it was concluded that acute toxicity is not an issue  and multi-dose  studies in normal volunteers and showed no effect on observed clinical symptoms, including electrocardiograms, body temperature, blood pressure, grip strength, and physician interviews, or clinical tests, including hematological tests, blood chemistry tests, and urinalysis , 296. ThOpen trials on 73 Duchene’s muscular dystrophy patients were conducted for 3 years at four Japanese national sanatoriums and resulted in some muscle strength improvement but the results were inconclusive and final double-blind studies did not confirm the results . The facThe gross abnormalities seen in offspring of double cathepsin B and L knockout mice clearly show that some cysteine cathepsin activity is required for normal embryogenesis. Possibly, the loss of cysteine cathepsin protease protein degradation in lysosomes and autophagosomes results in nutritional deficiencies, which cause the abnormalities. As discussed above, sufficiently high E64d doses can also cause teratogenic abnormalities when administered to pregnant animals.This has consequences for the therapeutic use of E64d or its derivative, the most obvious being that very careful consideration will have to be given to its use in pregnant women, women of child-bearing age, and infants. But as most young civilian and military TBI patients are adolescent or young adult males, the teratogenic danger may not be as much of a risk for the majority of that TBI patient population. Moreover, the teratogenic risk is not likely an issue for a large and growing segment of the TBI population, which are elderly who are beyond the reproductive age.These data also suggest that minimizing inhibition of lysosomal cathepsin B and cathepsin L activities is desirable in adults. To that end, E64c is unique among cysteine protease inhibitors in that it concentrates in the cytosol and not in the lysosome (see above section on E64d/E64c biodistribution), thus preferentially spares inhibition of lysosomal cathepsin B and L activities. The fact that E64d has been safely administered to pediatric patients for many years shows that, if lysosomal protease inhibition is a toxicity issue, it is not significant with an appropriate dose.The low brain dose is the primary issue with E64d as a TBI therapeutic because a small portion of an oral dose gets into the brain as discussed above (section on biodistribution). Even though a low brain dose can effectively inhibit brain cathepsin B activity, it would be desirable to have a derivative compound, which delivers a higher brain dose than does E64d so that efficacy could be achieved with reduced systemic exposure.E64d is unique among the small molecule cathepsin B inhibitors because of the extraordinary amount of data on the compound. No other compound has the extensive information known for E64d on its efficacy, toxicology, and safety. These data greatly reduce the risk of using E64d vs. other small molecule compounds for testing in TBI and related models. Therefore, E64d is a promising tool compound for preclinical TBI testing.The extraordinary amount of data makes the E64d scaffold extremely useful for developing clinical derivatives. Only the E64d compound has data on pharmaceutical properties that provides an expectation roadmap for the behavior of a clinical compound that is derived from it. Other small molecule scaffolds lack that information and thus all the data on clinical candidates derived from them will be without information as to the likelihood of pharmaceutical success. Thus, the E64d data greatly reduce the inherently high risk of drug development.One might ask why not use a cathepsin B-specific inhibitor rather than the multi-protease inhibitor E64d since cathepsin B knockouts have no pathology? A druggable cathepsin B-specific inhibitor may be useful as a follow-on therapeutic. However, no orally bioavailable, brain effecting, cathepsin B-specific inhibitor is currently available and developing such will be difficult and will take considerable resources and time. For example, a high throughput screen of the National Institutes of Health (NIH) small molecule library for a cathepsin B-specific inhibitor did not find a druggable hit . The knoTBI is an urgent unmet need that cannot wait. Furthermore, as discussed above, the multi-cysteine protease inhibition by E64c is more efficacious than selective cathepsin B inhibition and thus E64d derivatives offer a better chance of obtaining positive outcomes in clinical trials.Cathepsin B gene deletion studies show that cathepsin B is a key player in the pathology of TBI. Cathepsin B is elevated following TBI and its removal or inhibition dramatically improves outcomes following trauma in animal models. Moreover, deleting or inhibiting cathepsin B improves outcomes in injury models related to TBI including epilepsy, aneurysm, ischemia, pain, surgical trauma, spinal cord trauma, infectious disease, and neurodegeneration. E64d is a promising compound for further preclinical testing as a candidate therapeutic agent for TBI because it has been shown to be orally effective when administered after trauma. E64d is the best scaffold upon which a TBI therapeutic can be developed because only E64d has extensive clinical and preclinical data available. It will also be beneficial to consider that inhibition of cathepsin B targets several mechanisms involved in TBI, including calpain and matrix metalloproteinases that contribute to TBI, as explained in this review. In conclusion, basic and translational research on cathepsin B for improving TBI-caused deficits should be accelerated as it is highly likely to generate an effective new TBI therapeutic.Gregory Hook is an employee and has equity in ALSP, Inc., which is developing drugs to treat neurological conditions, including TBI. Vivian Hook is the chair of ALSP’s Scientific Advisory Board and J. Steven Jacobsen and Kenneth Grabstein are members of that Board and all hold equity in ALSP. J. Steven Jacobsen is an employee of AstraZeneca, a major pharmaceutical company. Vivian Hook and J. Steven Jacobsen have disclosed their relationship with ALSP to the University of California, San Diego  and AstraZeneca, respectively. Mark Kindy has no conflict of interest to declare.


Purpose. Congenital rubella syndrome (CRS) is a global cause of preventable hearing impairment, blindness, and intellectual impairment. The present study sought to identify ocular and auditory manifestations of CRS in school-aged children in Mbingo, Cameroon. Design. Cross sectional study. Subjects. Students at two schools, one for children with hearing impairment, were screened for cataract, congenital glaucoma, and pigmentary retinopathy. Methods. Students underwent seven-field digital fundus photography through a dilated pupil using a Topcon NW200 nonmydriatic camera. Images were assessed by retina specialists in Canada via teleophthalmology. Clinical evidence was integrated to form case definitions for CRS based on Center for Disease Control and Prevention guidelines. Serological evidence of rubella infection was obtained using standardized IgG antibody titers. Main Outcome Measure. Number of probable and suspicious cases of CRS. Results. Between September 2009 and May 2010, 320 students participated. There were 28 (10.2%) probable cases, 104 (37.8%) suspects, and 143 (52.0%) unaffected. Rubella IgG serology was positive in 79 (48.7%) of children with hearing impairment and 11 (7.4%) of children with normal hearing. Conclusions. The present study identified 28 probable cases of CRS. Furthermore, 92.6% of students with normal hearing did not possess rubella IgG antibodies making future cases of CRS likely without intervention. Worldwihe virus .Maternal rubella infection in pregnancy may lead to a spectrum of fetal outcomes: an infant may be born normally, with congenital abnormalities, or spontaneously abort. Congenital rubella syndrome (CRS) denotes the presence of any combination of classic features which may accompany rubella infection early in gestation. As many as 50% of infants with CRS may appear normal at birth but will develop features of CRS afterwards . The conNuclear cataracts occur in about one-third of all cases and are bilateral in 50% , 8. OtheThe primary purpose of this study was to identify cases of CRS with ocular or auditory involvement in children attending two schools, one for students with normal hearing and the other for students with hearing impairment in Mbingo, Cameroon, using teleophthalmology. Secondary outcomes included identifying the seroprevalence of rubella IgG antibodies and the presence of salt and pepper retinopathy.Students at two schools in Mbingo, Cameroon, were screened for clinical evidence of congenital rubella syndrome. The two schools were chosen due to their proximity to the town of Mbingo. One of the schools was for children with normal hearing and the second school was for children with hearing impairment. A teleophthalmology partnership between the University of Alberta and the Mbingo Hospital enabled transmission of fundus photographs to retina specialists in Edmonton, Alberta, for confirmation of the diagnosis of rubella retinopathy. Research ethics approval was granted through both the Cameroon Baptist Convention Institutional Review Board and the University of Alberta Research Ethics Board. This research adhered to the tenets of the Declaration of Helsinki.https://teleophthalmology.com/; Secure Diagnostic Imaging Ltd., Edmonton, Alberta, Canada) for review by Canadian-based ophthalmologists at the Royal Alexandra Hospital Teleophthalmology Reading Centre. Images were viewed on a dual-monitor system through liquid crystal display shutter goggles  using proprietary 3D viewing software  and an accompanying online ETDRS grading template.Patients underwent seven-field digital photography of the retina through a dilated pupil using a Topcon NW200 nonmydriatic retinal camera. Images of the anterior segment, optic disc, and macula were captured. Original Joint Photographic Experts Group (JPEG) images were saved  and stored onto an attached laptop computer. The images were then further JPEG compressed 22 : 1 to facilitate transmission via a satellite connection with limited bandwidth. Digital images were then linked to Extendable Markup Language (XML) metadata as a single encrypted file and transmitted via the internet to a secure, web-based server (http://www.rapidtest.com/) which offers excellent sensitivity (100%) and specificity (100%) according to the manufacturer. The positive cutoff was >15 IU/mL and antibody titers less than this were considered negative. IgM levels were not measured as the sample population would no longer demonstrate positive IgM levels from CRS due to age. Some participants were unavailable for retinal photographs or blood testing and therefore not all participants had complete testing records.Serological evidence of rubella infection was obtained using standardized rubella IgG antibody titers analyzed at a local medical laboratory in Mbingo, Cameroon. Rubella IgG antibodies were detected using the human rubella IgG ELISA Test Kit  and Prevention guidelines, 2012 –15. Syst CRS suspect was defined as an individual with one of cataract, congenital glaucoma, pigmentary retinopathy, or hearing impairment. A probable case of CRS was defined as an individual with two or more of cataract, congenital glaucoma, pigmentary retinopathy, or hearing impairment. Unaffected children demonstrated none of the above findings. Because serological testing could not be performed according to the Center for Disease Control guidelines, no cases of CRS could be confirmed.For the purposes of this study aBetween September 2009 and May 2010, all 320 students from two schools in Mbingo, Cameroon, participated in this study. The average age of students from both schools was 11.8 ± 2.8 years and 58.2% of students were male. Fundus photographs were available for 275 (85.9%) participants and rubella IgG serology was available for 310 (96.9%) patients. Complete records including fundus photographs and serology were available for 268 (83.8%) participants.P < 0.0001). Ninety students (29.0%) were positive for rubella IgG antibodies and 220 (71.0%) were negative. Hearing impaired children were seven times more likely to have positive serology (48.8%) than children with normal hearing . Conversely, children with normal hearing were almost two times more likely to have negative serology (92.6%) than hearing impaired children   see .Fundus photographs were obtained for 549 eyes of 275 (85.9%) students. 58 (10.5%) eyes of 29 patients showed clear evidence of rubella retinopathy. Twenty-six of the 29 (89.7%) patients with rubella retinopathy had bilateral retinal disease. Twelve (2.2%) eyes were suspicious for rubella retinopathy and were given the designation of “peripheral stippling,” while 473 eyes (86.0%) had no evidence of rubella retinopathy.P < 0.0001). Serologic testing suggested that positive rubella titers were associated with the presence of rubella retinopathy, with 55.1% of affected children testing positive (P = 0.001).Students with rubella retinopathy were much more likely to be hearing impaired; of the 29 students with rubella retinopathy, 28 (96.6%) had impaired hearing (Ten eyes (1.8%) showed evidence of other ocular pathology such as ocular albinism (4), toxoplasmosis (1), macular scar (1), corneal scar (1), and retinal pigment atrophy. (1). One patient had undergone previous surgery for cataracts , while no cases of congenital glaucoma were identified.n = 143; 52.0%) were unaffected. There were 104 (37.8%) suspects and 28 (10.2%) probable cases of CRS . If students with hearing impairment represented cases of CRS, then older age would be expected to underestimate the prevalence of IgG as diminution of antibodies is known to occur [This study found a significant relationship between rubella IgG seropositivity and hearing impairment in unvaccinated children. This relationship is confounded by the effect of age, as students with hearing impairment were significantly older than those without hearing impairment  were in the hearing impaired population and 26 (89.7%) cases were bilateral. Our study is consistent with estimates in the literature reporting bilateral retinopathy in 80% of patients with CRS . Only 55in utero .Cataracts were expected to be observed in approximately one-third of CRS cases as reported in the literature , 23; howWHO recommendations encourage all countries not routinely immunizing against rubella to quantify the burden of disease due to congenital rubella syndrome . InterdiThe present study provides clinical evidence of CRS in students in Northwest Cameroon using teleophthalmology. Moreover, a large population vulnerable to acquiring rubella infection later in life was identified: 92.6% of students with normal hearing were negative for rubella IgG. Other studies have estimated the rate of rubella serosusceptibility among women of reproductive age in West Africa to be 10% to 30% . These r


Noise-induced hearing loss has been an intractable problem for heavy industry.To report our experience in reducing the incidence of age-corrected confirmed 10 dB hearing shifts  in employees in the primary aluminium industry in Australia over the period 2006–13.We analysed annual audiometric data to determine the number of permanent hearing shifts that occurred in employees in two bauxite mines, three alumina refineries and two aluminium smelters. Annual hearing shift rates were calculated based on the number of employees tested per year. Hearing conservation initiatives undertaken during the study period are described. An assessment of similar exposure group noise exposures was also undertaken to determine the magnitude of noise exposure reduction during the study period.P < 0.001). The decline in shift rates was greater in mines and refineries, where baseline shift rates were higher, than in smelter workers. Modest reductions in noise exposure occurred during the study period.Across all operations, hearing shift rates declined from 5.5% per year in 2006 to 1.3% per year in 2013 (We observed a substantial decline in hearing shift rates during the study period. We describe the hearing conservation initiatives that were collectively associated with this decline. We suspect these initiatives could be deployed relatively easily and at modest cost in other industries with noise-exposed employees. Exposure to noise in industry is ubiquitous and noise-induced hearing loss (NIHL) has been an intractable problem for heavy industry. For example, in the USA, ~22 million workers are exposed to hazardous noise levels and occupational hearing loss is one of the most common work-related diseases . The preWe report our experience reducing the incidence of age-corrected confirmed 10 dB hearing shifts among employees in the primary aluminium industry in Australia, including bauxite mining, alumina refining and aluminium smelting operations, over the period 2006–13. During this period, the employer  introduced a number of new initiatives, including quantitative fit testing of hearing protection.At the start of the study period, the mandated elements of Alcoa’s global health standard on hearing conservation had already been deployed for several years. This standard requires annual audiometry for personnel whose noise exposure for 12 or more days per year equals or exceeds 85 dB(A)Leq8h, a 15-min short-term exposure limit of ≥100 dB(A) or a sound pressure level of 140 dB linear peak . If this ‘background’ shift rate could be attained in noise-exposed employees, occupational NIHL would have been minimized. Alcoa therefore adopted a target age-corrected confirmed 10 dB hearing shift rate of 1% per year for employees exposed to occupational noise, to be achieved by the year 2020. ‘Background’ hearing shifts are due to non-occupational noise exposures  and a variety of other causes of sensorineural hearing loss  .From 2006 to 2013, Alcoa of Australia introduced a range of initiatives in addition to the mandated elements of hearing conservation outlined above see .These included1. Holding noise summit meetings and health promotion campaigns to motivate and educate employees about the cause and consequences of noise-induced hearing loss (NIHL).2. Producing a DVD on NIHL called ‘The Road to Silence’.™ E-A-RFit™ equipment and fit testing programme was adopted in Australia  within the SEG were converted to linear metric values reflecting the ratio of the noise exposures to the OEL (expressed as percentages of the OEL). The arithmetic mean of these percentages of OEL values was taken to summarize noise exposure within each unaccept able SEG. To assess the overall noise exposure at each location, the geometric mean of all of the summary unacceptable SEG values at each location was then calculated for the beginning of the 3-year metric cycle (baseline) and for the end of the cycle. The difference between baseline and end of cycle (expressed as a percentage of baseline) was taken to be the noise reduction achieved at that location. The noise reduction results were then averaged across all locations (weighted by the baseline number of unacceptable SEGs at each location) to determine the overall reduction across the Australian organization. The chi-squared test for trend was used to determine the statistical significance of reductions in hearing shift rates over the study period . StanforIn 2011, there were 865 employees in the mines, 2994 in refineries and 1246 working in smelters. Audiometric surveillance was required for 796 mining employees (92%), 2631 refinery employees (88%) and 1091 smelter employees (88%) exposed to noise. When all of the operations were considered in aggregate, there was a statistically significant decline in the age-corrected confirmed 10 dB shift rate, from 5.5% per year in 2006 to 1.3% per year in 2013 . The decThe 2013 results for the worksites reported here approximate the ‘background’ 1% per year shift rate found previously in non-noise-exposed occupations within the aluminium industry in the USA . This suggests occupational NIHL has been minimized in the worker population under surveillance here.Given the descriptive nature of the study, we are not able to state which of the initiatives contributed to the improvement in hearing shift rates. We posit that those associated with improving the knowledge of the workforce about NIHL and particularly about the importance of correctly wearing hearing protection had the greatest impact. The noise summits organized in 2006 and 2007 were conducted across all operations and enabled a core body of employees to become champions for hearing conservation. The summits included a specially produced educational DVD, ‘The Road to Silence,’ in which employees with NIHL explained how it had affected their lives. This was followed by other campaigns, in particular, a programme to fit test all noise-exposed employees to ensure they understood correct fitting of earplugs or earmuffs. This fit testing programme found that many employees had not previously been wearing hearing protection effectively and was continued on a biannual basis for all noise-exposed employees. Annual audiometric testing for all noise-exposed employees and personal hearing conservation reviews for those with early NIHL helped to identify the most significant noise risks in the employee’s area as well as assist the worker with their own hearing protection management.Although it is likely that the initiatives described above have collectively brought about the observed decline in hearing shift rates, we also considered potential confounders. There were modest changes in turnover of the workforce during the study period. The percentage turnover of employees each year during the period 2006–13 ranged from 5 to 10% in the mines, from 8 to 12% in the refineries and from 4 to 9% in the smelters. It seems unlikely that changes of these magnitudes would materially impact hearing shift rates. There were no meaningful changes in the average hours worked per week. During the period 2008–13, these ranged from 35.7 to 36.0 in the mines, from 35.4 to 36.5 in the refineries and from 40.8 to 41.0 in the smelters. There were modest changes in production levels: bauxite production increased from 30996 kt in 2006 to 34181 kt in 2013 (10%); alumina production increased from 8224 kt in 2006 to 9182 kt in 2013 (12%) and aluminium production decreased from 530 kt in 2006 to 488 kt in 2013 (−8%). It seems unlikely that increases in production in mining and refining would lead to reduced noise exposures and contribute to the observed decline in hearing shift rates. Although it seems more likely that the decline in aluminium production might have contributed to the observed effect in smelters, the decline was modest (8%) and the exposure data for unacceptable SEGs probably give a more useful insight.We also considered the possibility that the decline in confirmed hearing shift rates might be due to a decline in compliance with recall for confirmation audiometry. As the OHM database is set to automatically confirm hearing shifts detected on a first test if the person is not re-tested within 90 days, non-compliance with re-testing could not have contributed to the figures we present.There was a significant change in the average noise levels between the two cycle periods shown in this led to an increase in the number of unacceptable SEGs, particularly in the 2006–08 cycle. This was especially so at Kwinana Refinery where reorganization of work groups led to an increased number of smaller work groups. In While the overall noise reductions appear to be modest, we believe that these efforts would have had some impact in reducing NIHL, although probably less so than the improved use of hearing protection. Apart from the reduction in exposure experienced by some SEGs, the increased focus on identifying noise sources also focused attention on the need to wear hearing protection near loud plant and equipment. Unfortunately, for some SEGs, the noise levels increased, possibly as a result of ageing equipment or reduced maintenance, countering the impact of improvements in other SEGs. The modest results indicate that even with a high degree of focus, engineering reductions in noise exposures are difficult and expensive.In summary, we report a substantial decline in the incidence of age-corrected confirmed 10 dB hearing shifts among employees in the primary aluminium industry. We have detailed the initiatives collectively associated with this decline and suspect they could be deployed easily and at modest cost in other industries. More research could define which of the initiatives are most effective.Enhanced hearing conservation initiatives in the primary aluminium industry were associated with a decline in age-corrected confirmed 10 dB hearing shift rates from 5.5% per year in 2006 to 1.3% per year in 2013.We posit the most important initiatives were improved education of the workforce and quantitative fit checking of hearing protection attenuation.Modest reductions in noise exposures occurred during the study period.Alcoa Inc.A. M. D. is employed full time by Alcoa of Australia. N. F. and B. J. C. were previously employed by Alcoa of Australia. C.D.-E. was previously employed by Alcoa Inc. M. R. C. receives salary support from an Alcoa grant.


This study investigated the effect of systemic salicylate on central auditory and non-auditory structures in mice. Since cochlear hair cells are known to be one major target of salicylate, cochlear effects were reduced by using kanamycin to remove or impair hair cells. Neuronal brain activity was measured using the non-invasive manganese-enhanced magnetic resonance imaging technique. For all brain structures investigated, calcium-related neuronal activity was increased following systemic application of a sodium salicylate solution: probably due to neuronal hyperactivity. In addition, it was shown that the central effect of salicylate was not limited to the auditory system. A general alteration of calcium-related activity was indicated by an increase in manganese accumulation in the preoptic area of the anterior hypothalamus, as well as in the amygdala. The present data suggest that salicylate-induced activity changes in the auditory system differ from those shown in studies of noise trauma. Since salicylate action is reversible, central pharmacological effects of salicylate compared to those of (permanent) noise-induced hearing impairment and tinnitus might induce different pathophysiologies. These should therefore, be treated as different causes with the same symptoms. In the inner ear (the cochlea), salicylate leads to both functional and structural changes. Salicylate markedly reduces the cochlear blood flow by vasoconstriction [Salicylate is known to induce reversible loss of auditory sensitivity and tinnitus both in humans –8 and antriction . Furthertriction  with haitriction , 20. Thitriction  and the triction –24. A patriction , 25–27.Beside these cochlear changes, salicylate also modulates neuronal activity in the central auditory pathway in a dose-dependent manner. It has been demonstrated that changes in cochlear output leads to an altered spontaneous activity in the auditory nerve. Whereby salicylate studies in cats revealed an increase in nerve fibre activity, spontaneous discharge rates were shown to decrease in gerbils , 29, 30.Beside these findings, several previous studies have investigated the effects of systemic salicylate application with regard to the modulation of spontaneous or evoked activity in central auditory structures. In summary, the results demonstrated that the majority of neurons change their spontaneous activity upon salicylate application. An increase was observed in the frequency range showing tinnitus-related behaviour, whereby neurons with low spontaneous firing rates were particularly affected, even in the auditory cortex. In contrast, a salicylate-induced reduction in multi-unit activity was also detected in awake animals These changes of central neural activity could be caused by the peripheral or the central effects of salicylate. Recordings in brain slices showed a direct, dose-dependent pharmacological action of salicylate on several central auditory and non-auditory structures , 55. HowUsing manganese-enhanced magnetic resonance imaging (MEMRI), the present study investigated the effect of systemic salicylate application on calcium-related activity in central auditory structures after reducing the peripheral effects mediated by cochlear hair cells as one major target of salicylate action.Mus musculus, NMRI strain) of both sex were examined. Different animals were used for threshold testing and MEMRI measurements. The experimental protocol was approved by the governmental commission for animal studies . Experiments were carried out in accordance with the EU Directive 2010/63/EU on the protection of animals used for scientific purposes. All efforts were made to minimize the number of animals and their suffering. The condition of animals was monitored every 6 hours after the induction of hearing loss and, in addition, every hour following substance injection . Since animals did not show any abnormal behaviour, additional analgesic treatment was not necessary.In the present study, 30 normal hearing adult  mice  dissolved in phosphate-buffered saline (PBS), at 1 mg/g body weight followed 40 minutes later by an intraperitoneal injection of bumetanide  at 0.05 mg/g body weight. This procedure has been shown to elicit a significant and rapid hair cell loss in mice in-vivo . Three oAuditory thresholds were measured for 7 hearing-impaired mice  and for 8 normal hearing control mice. Different animals were used for each group. The time interval for deaf mice was chosen to avoid any additional central pathophysiological effects arising from the damaged peripheral auditory system.® measurement system . The brainstem responses (recording epoch 10 ms following stimulus presentation) were amplified (100.000 x), filtered (bandwidth 0.15–3 kHz) and averaged (300 x) by the Viking IV®. The amplitude growth function of wave IV/V was calculated from the resulting data for each frequency tested and a linear regression was fitted to the function. By extrapolating the regression line to zero, mean auditory thresholds were estimated for the controls and the kanamycin-treated animals. Results are shown as mean auditory thresholds (± S.E.) in dB SPL by group and mean relative hearing loss of the treated group compared to the normal hearing controls.Frequency-specific  auditory brainstem responses (ABR) were recorded under anaesthesia. Sub-dermal needle electrodes were placed at the vertex (active), mastoid (reference) and at one foot (ground). Tone stimuli were delivered binaurally at different SPLs with a sine-wave generator . ABR recordings were carried out using a Viking IVTo investigate salicylate-induced changes in central calcium-dependent activity after kanamycin-induced damage of cochlear hair cells, 9 treated animals received an intraperitoneal (i.p.) injection of a sodium salicylate (SS) solution (250 mg/kg). It has been shown earlier that salicylate is able to cross the blood brain barrier shortly after systemic (e.g. i.p.) application leading to an increased cerebrospinal fluid level of salicylate –59. Compin-vivo [2+ accumulates within the tissue which results in an increase in MRI-T1 signal contrast and therefore reflects the Ca2+-dependent activity [2+-dependent activity can be imaged. This provides an opportunity to integrate neuronal activity, represented by the increase in signal contrast due to manganese accumulation, over a well-defined period of time before measurements, i.e., 24 hours in the present experiments. This is of particular importance during the investigation of auditory-related activity even inside a noisy MRI scanner [MEMRI is a powerful tool to image central nervous system activity in small animals in-vivo –64. Aftein-vivo  and entein-vivo , 67. Dueactivity . Thereby scanner , 69–71.2 solution , animals of the experimental group  as well as hearing-impaired control mice (kanamycin/bumetanide-treated only) received a 0.4 mM/kg dose of MnClnce with ). Delivence with , MRI scaManganese-enhanced MRI was carried out using a 7 Tesla rodent scanner  with a 16 cm horizontal bore magnet and a 9 cm (inner diameter) shielded gradient with a H-resonance-frequency of 300 MHz and a maximum gradient strength of 300 mT/m. For imaging a 1H-RF quadrature-volume resonator with an inner diameter of 20 mm and a T1-weighted 2D turbo spin-echo sequence  was used. Data acquisition and image processing were conducted using the Bruker Paravision 4.0 software. Thirty-five axial slices with a slice thickness of 0.3 mm, a field of view of 2.85 x 2.85 cm and a matrix of 256 x 256 resulted in an in-plane resolution of 111μm. Imaging covered the brain from brainstem to forebrain. Signal intensity analysis of defined auditory brain regions was carried out with the program Analyze 5.0 . Signal intensities were quantified bilaterally in the dorsal (DCN) and ventral (VCN) cochlear nucleus, the superior olivary complex (SOC), the inferior colliculus (IC), the medial geniculate body (MGB), the auditory cortex (AC), the preoptic area of the anterior hypothalamus (PO/AH), the Amygdala (Amyg). The masseter muscle was used as an objective intensity reference, since it is located close to the related neural tissue but is not affected by changes in brain activity. PO/AH and Amyg served as non-auditory structures investigated in the present study. Regions of interest were marked in accordance with “the mouse brain atlas in stereotaxic coordinates”  Fig 1)..Fig 1). The statistical comparison was done with the software SPSS . After confirming a normal distribution using the Kolmogoroff-Smirnoff-test, the t-test for independent samples was applied. A statistically significant difference was considered for p<0.05.Hearing loss following kanamycin and bumetanide treatment was indicated by an ABR-threshold shift of up to 40 dB in the investigated frequency range between 4 and 20 kHz compared to the untreated normal hearing control group. Frequency-dependent mean auditory thresholds in normal hearing controls ranged between 17 and 31 dB SPL. In the hearing impaired animals, thresholds were detected between 53 and 60 dB SPL . AverageFollowing systemic salicylate application, MEMRI contrast increased significantly in all investigated auditory brain areas.In detail, the relative MRI signal intensities were raised in the DCN from 128.0±2.8 in hearing-impaired controls to 152.9±1.9 in the hearing-impaired salicylate group and for the VCN from 127.2±2.4 (hearing-impaired controls) to 151.2±1.9 . In the SOC, the signal increased from 133.1±2.0 (in hearing-impaired controls) to 149.2±1.8 in salicylate-treated animals. Similar observations have been made in the IC , in the MGB  as well as in the AC  .An increased MEMRI signal was also observed for the non-auditory control areas (PO/AH and Amyg) after salicylate treatment. Signal intensities were elevated from 148.5±2.0 (hearing-impaired controls) to 175.0±2.1  in PO/AH and from 143.6±3.0 in hearing-impaired controls to 167.8±2.5 in salicylate-treated animals in the amygdala region . All datThe differences between the groups were statistically significant for all investigated auditory and non-auditory structures .The results of the present study demonstrate changes in calcium-dependent brain activity after systemic salicylate application in kanamycin-treated animals using non-invasive manganese-enhanced magnetic resonance imaging (MEMRI).The ABR recordings demonstrated that a three-fold application of kanamycin and bumetanide produces a significant threshold shift in mice, possibly in particular due to OHC destruction according to previous reports. It is further assumed that the treatment induces a minor loss of IHC and spiral ganglion cells as well , 74. AltRecent studies have reported that the central auditory system largely contributes to the salicylate-induced effects on neuronal activity and auditory phantom perceptions like tinnitus, even after disconnection of the auditory periphery , 75, 76.An altered activity of the amygdala after salicylate application has already been reported by immunostaining. An elevated c-Fos- or arg3.1-related activity was demonstrated in the auditory cortex and the amygdala after treatment with a single high-dose salicylate application , wherebyOne possible mechanism for this increase in calcium-related activity could be the direct action of salicylate on neuronal and synaptic activity within those particular brain structures. Salicylate largely influences the pre- and postsynaptic conductance of several transmembrane ion channels by modulating neurotransmitter activity (e.g. modulation of GABA-mediated inhibition) , 40. In It should be mentioned that pharmacological removal of cochlear sensory cells reduces auditory nerve input towards the central auditory system. This happens particularly when inner hair cells have been affected (in addition to OHC damage), thereby inducing changes in central neuronal activity (e.g. compensatory hyperactivity), as shown by several studies in this field. For a review, see . HoweverThe present study indicates that salicylate-induced activity changes in central auditory brain areas, including possibly tinnitus, exist to a large extent. Since observations were made after reduction of peripheral effects mediated by cochlear hair cells, these effects cannot be entirely explained by salicylate’s action on peripheral tissue.2+ activity patterns between the two studies even indicate that hearing impairment related to either salicylate application or noise trauma have varying impacts on central auditory system physiology and might therefore, rely on different pathophysiological mechanisms. This may lead to the hypothesis that the appearance of tinnitus due to salicylate or noise is also based on different sources. Calcium-related activity patterns have also been observed by another study investigating activity changes in several brain structures after induction of noise- or salicylate-related tinnitus using manganese-enhanced MRI [The present results indicate that the mechanisms through which salicylate impacts on activity in several auditory and non-auditory brain structures appears different to those known from noise-induced hearing loss (and possibly tinnitus generation); particularly immediately after treatment. Acute noise-exposure is accompanied initially by cochlear changes. In a second stage, the altered afferents are probably responsible for modulating central auditory processing. However, permanent tonotopic and other pathophysiological changes appear to be largely established by compensatory neuroplasticity, acting to overcome the reduced cochlear output and damaged central auditory structures. This is in line with the finding that an increased spontaneous firing rate (SFR) in the DCN (after 140 dB SPL noise exposure) could be recorded only after a 2–3 days’ period . Once thnced MRI . Signal In essence, the present data support the idea that the impact of salicylate application or noise trauma on the auditory system are probably based on different mechanisms, even if comparable changes of electrophysiological properties could be observed in vivo: e.g. increased SFR in central auditory areas , 90, 91,S1 Table(XLS)Click here for additional data file.S2 Table(XLS)Click here for additional data file.


Alterations in the visual system may underlie visual hallucinations in dementia with Lewy bodies (DLB). However, cortical excitability as measured by transcranial magnetic stimulation (TMS) and functional magnetic resonance imaging (fMRI) activation of lower visual areas (V1–3) to visual stimuli appear normal in DLB. We explored the relationship between TMS-determined phosphene threshold and fMRI-related visual activation and found a positive relationship between the two in controls but a negative one in DLB. This double dissociation suggests a loss of inhibition in the visual system in DLB, which may predispose individuals to visual dysfunction and visual hallucinations. This was supported by our finding of unaltered cortical activity in the early visual system , as measured by blood oxygen level-dependent (BOLD) functional magnetic resonance imaging (fMRI), in DLB.2 BOLD haemodynamic response has been suggested to be related to synaptic activity in terms of input and local processing,3 as well as to neuronal spiking4 arising from sensory stimulation, and thus may represent the activity in both excitatory and inhibitory neuronal populations. In contrast, the neuronal discharge consequent to TMS is likely to be dependent on the relative balance of excitatory and inhibitory activity within the area of stimulation. Therefore a positive relationship between BOLD activity in the early visual system and phosphene threshold may suggest that inhibition outweighs excitation, whereas a negative relationship may indicate the opposite. In this paper we report the interrelationship between fMRI BOLD activity in the early visual system and visual cortical excitability as measured using TMS, using previously published data from patients with DLB and hallucinations and from controls.2Dementia with Lewy bodies (DLB) is characterised by complex visual hallucinations. Previously, we used the threshold for eliciting phosphenes with occipital transcranial magnetic stimulation (TMS) as a measure of visual cortical excitability.2 In brief, 16 participants who met criteria for probable DLB5 and who experienced visual hallucinations at least once in the month before TMS/fMRI testing were recruited from a community-dwelling population of patients, along with 19 controls. The study was approved by the local ethics committee. The DLB diagnosis was made independently by two experienced senior clinicians.5 Occipital TMS and neuroimaging evaluations were conducted within 2 weeks of each other.Details of participant selection, neuropsychological and neuropsychiatric assessment, fMRI and TMS testing are described fully elsewhere. In the current analysis, we selected participants who had undergone both fMRI and occipital TMS.TMS and imaging protocols are described in the online supplementary methods.6 motor scores were significantly higher and cognitive test and visuoperceptual task scores significantly lower in the DLB group (online Table DS1).The DLB and control groups were similar in terms of age, gender and visual acuity. Unified Parkinson's Disease Rating Scale (UPDRS)P = 0.66). Across groups, phosphene thresholds were similar  = 44.0%; DLB group: median 77.2%, IQR = 36.0%, Mann–Whitney U-test 115.5, P = 0.23).Phosphenes were elicited in a similar proportion of the control  and DLB groups ; DLB group mean 0.71 (s.d. = 0.29), P= 0.83). Similarly, there were no perfusion differences in V1 ; DLB group, mean 28.0 (s.d. = 10.5), P= 0.20) or V2/V3 , DLB group, mean 27.0 (s.d. = 7.6), P= 0.15) between groups. However, there were significant positive correlations between the magnitude of the BOLD response (V1 and V2/V3) to the checkerboard stimulus and phosphene thresholds in the control group and a significant negative relationship in the DLB group in both V1 and V2/V3 . These relationships remained significant (P<0.05) even with the removal of non-responders (i.e. those participants with a phosphene threshold greater than 100% of the stimulator output) except for phosphene threshold v. V2/V3 in the DLB group. In TMS responders (i.e. those experiencing phosphenes) BOLD activity was significantly lower in the control group in V1 compared with the DLB group ; DLB group, mean 0.65 (s.d. = 0.25), P= 0.04).There were no differences in V1 or V2/V3 BOLD magnitude between the control and DLB groups for checkerboard stimulus ; DLB group, mean 0.57 (s.d. = 0.30), v. DLB group for V1 and V2/V3 were highly significant . BOLD activity was significantly lower in V1 and trending lower in V2/V3 in those in the DLB group who were on cholinesterase inhibitors (V1: mean 0.45 (s.d. = 0.24); V2/V3: mean 0.62 (s.d. = 0.10)) compared with those in the DLB group not taking these agents (V1: mean 0.82 (s.d. = 0.28), P= 0.02; V2/V3: mean 0.88 (s.d. = 0.33), P = 0.10) although phosphene thresholds were similar, as was cerebral perfusion in V1 and V2/V3 between the DLB group on and off cholinesterase inhibitors (phosphene threshold: mean 76.7% (s.d.= 20.2%) v. 78.2% (s.d.= 21.1%), P= 0.90. Perfusion (ml/100g/min): V1, mean 27.1 (s.d.= 9.7) v. 30.4 (s.d.= 13.7), P = 0.62; V2/V3, mean 27.4 (s.d.= 7.3) v. mean 25.8 (s.d. = 9.4), P = 0.73). There was no association between global cognitive function in the DLB group and phosphene threshold, BOLD activity or cerebral perfusion in V1 or V2/V3 (P>0.13).Differences in the slopes for the control 4 then the lack of difference between groups in mean BOLD activity suggests a loss of inhibition.We report a significant positive relationship between phosphene threshold and BOLD activity in early visual areas  in our controls, which suggests that the greater an individual's inherent visual cortical excitability, the smaller the visual cortical activation in response to a simple checkerboard stimulus. This observation implies that individuals with sensitive visual systems need less ‘activation’ to an external stimulus. In contrast, the opposite relationship in DLB infers a breakdown in this dynamic. Assuming similar visual cortical neuronal populations are activated by the checkerboard and TMS, then in the controls a positive relationship between BOLD activity and phosphene threshold implies inhibition must outweigh excitation, whereas in DLB this is reversed. If the BOLD signal represents spiking excitatory neuronal activity,7 and the BOLD effect reflects not just neuronal activity but also haemodynamic coupling, which may be altered with neurodegeneration; ideally cerebrovascular reactivity would be assessed via a CO2 challenge8 or similar paradigm. However, we did not observe any differences in occipital perfusion between the control and DLB group, and similarly, there was no relationship between perfusion and either BOLD response or phosphene threshold. Furthermore, cognitive function in DLB was not associated with phosphene threshold or any of the neuroimaging measures. Thus, although not providing a direct measure of cerebrovascular reactivity, these findings suggest that the early visual system in these participants with DLB was similar to controls.There needs to be critical consideration to our interpretations; our knowledge of the underlying neuronal activity of the BOLD signal is incomplete9 In addition, cholinergic function has been implicated in modulating visual attention and associated BOLD fMRI responses;10 our observation of differences in BOLD activity between those patients with DLB taking cholinesterase inhibitors v. those not, supports this and fits into the broader framework of reported cholinergic receptor changes in DLB11 as well as changes in occipital metabolism in individuals with DLB with hallucination amelioration treated with donepezil.12From the TMS perspective, the origin of phosphene perception may not be co-localised to the visual cortex and thus the relative state of the visual cortex may be less crucial to phosphene perception, although reciprocal connections between early visual and association areas are likely to be important.1Finally, a loss of inhibitory activity in patients might be expected to decrease phosphene threshold relative to the control group, something which was not observed. However, it is notable that for a range of phosphene thresholds, the overall BOLD response in the DLB group was larger than in the control group, significantly so for V1 after excluding TMS non-responders . Hence, one could speculate that compensatory mechanisms may be operating in patients with DLB that attempt to ‘normalise’ the balance of excitation/ inhibition in an underactive early visual system, in the face of ‘upstream’ pathology in the visual system thus leading to a maintenance of the phosphene threshold within a normative range and possibly improved bottom-up visual processing, but at the cost of increasing the risk of visual hallucination occurrence, as evidenced by our previous work linking phosphene threshold with visual hallucination frequency and severity.v. healthy controls. Whether this represents a specific loss of inhibitory drive in the visual cortex of people with DLB and what role cholinergic function plays remains to be clarified; neurochemical post-mortem studies or novel spectroscopic imaging assessing for example gamma-aminobutyric acid-ergic tone may be helpful in this regard.In summary, our findings of a double dissociation suggest a marked deviance in visual neuronal processing between patients with DLB who hallucinate