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  • Magnetic stimulation allows focal activation of the mouse cochlea. eLife Lee, J. I., Seist, R., McInturff, S., Lee, D. J., Brown, M. C., Stankovic, K. M., Fried, S. 2022; 11


    Cochlear implants (CIs) provide sound and speech sensations for patients with severe to profound hearing loss by electrically stimulating the auditory nerve. While most CI users achieve some degree of open set word recognition under quiet conditions, hearing that utilizes complex neural coding (e.g., appreciating music) has proved elusive, probably because of the inability of CIs to create narrow regions of spectral activation. Several novel approaches have recently shown promise for improving spatial selectivity, but substantial design differences from conventional CIs will necessitate much additional safety and efficacy testing before clinical viability is established. Outside the cochlea, magnetic stimulation from small coils (micro-coils) has been shown to confine activation more narrowly than that from conventional microelectrodes, raising the possibility that coil-based stimulation of the cochlea could improve the spectral resolution of CIs. To explore this, we delivered magnetic stimulation from micro-coils to multiple locations of the cochlea and measured the spread of activation utilizing a multielectrode array inserted into the inferior colliculus; responses to magnetic stimulation were compared to analogous experiments with conventional microelectrodes as well as to responses when presenting auditory monotones. Encouragingly, the extent of activation with micro-coils was ~60% narrower compared to electric stimulation and largely similar to the spread arising from acoustic stimulation. The dynamic range of coils was more than three times larger than that of electrodes, further supporting a smaller spread of activation. While much additional testing is required, these results support the notion that magnetic micro-coil CIs can produce a larger number of independent spectral channels and may therefore improve auditory outcomes. Further, because coil-based devices are structurally similar to existing CIs, fewer impediments to clinical translational are likely to arise.

    View details for DOI 10.7554/eLife.76682

    View details for PubMedID 35608242

  • Orally administered fluorescent nanosized polystyrene particles affect cell viability, hormonal and inflammatory profile, and behavior in treated mice. Environmental pollution (Barking, Essex : 1987) Nikolic, S., Gazdic-Jankovic, M., Rosic, G., Miletic-Kovacevic, M., Jovicic, N., Nestorovic, N., Stojkovic, P., Filipovic, N., Milosevic-Djordjevic, O., Selakovic, D., Zivanovic, M., Seklic, D., Milivojevic, N., Markovic, A., Seist, R., Vasilijic, S., Stankovic, K. M., Stojkovic, M., Ljujic, B. 2022: 119206


    Commercially manufactured or generated through environmental degradation, microplastics (MPs) and nanoplastics (NPs) considerably contribute to environmental pollution. There is a knowledge gap in how exposure to MPs/NPs changes cellular function and affects animal and human health. Here, we demonstrate that after oral uptake, fluorescent polystyrene (PS) nanoparticles pass through the mouse digestive system, accumulate and aggregate in different organs, and induce functional changes in cells and organs. Using cochlear explant as a novel in vitro system, we confirmed the consequences of PS-MP/NP interaction with inner ear cells by detecting aggregates and hetero-aggregates of PS particles in hair cells. The testes of treated males accumulated MPs/NPs in the interstitial compartment surrounding the seminiferous tubules, which was associated with a statistically significant decrease in testosterone levels. Male mice showed increased secretion of interleukins (IL-12p35 and IL-23) by splenocytes while cyto- and genotoxicity tests indicated impaired cell viability and increased DNA damage in spleen tissue. Males also showed a broad range of anxiogenic responses to PS nanoparticles while hippocampal samples from treated females showed an increased expression of Bax and Nlrp3 genes, indicating a pro-apoptotic/proinflammatory effect of PS treatment. Taken together, induced PS effects are also gender-dependent, and therefore, strongly motivate future research to mitigate the deleterious effects of nanosized plastic particles.

    View details for DOI 10.1016/j.envpol.2022.119206

    View details for PubMedID 35405220

  • Conductive hearing loss in the Hyp mouse model of X-linked hypophosphatemia is accompanied by hypomineralization of the auditory ossicles. Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research Delsmann, M. M., Seist, R., Sturznickel, J., Schmidt, F. N., Mansour, A., Kobelski, M. M., Broocks, G., Peichl, J., Oheim, R., Praetorius, M., Schinke, T., Amling, M., Demay, M. B., Stankovic, K. M., Rolvien, T. 2021


    X-linked hypophosphatemia (XLH) is a hereditary musculoskeletal disorder caused by loss-of-function mutations in the PHEX gene. In XLH, increased circulating fibroblast growth factor 23 (FGF23) levels cause renal phosphate wasting and low concentrations of 1,25-dihydroxyvitamin D, leading to an early clinical manifestation of rickets. Importantly, hearing loss is commonly observed in XLH patients. We here present data from two XLH patients with marked conductive hearing loss. To decipher the underlying pathophysiology of hearing loss in XLH, we utilized the Hyp mouse model of XLH and measured auditory brain stem responses (ABRs) and distortion product otoacoustic emissions (DPOAEs) to functionally assess hearing. As evidenced by the increased ABR/DPOAE threshold shifts in the mid-frequency range, these measurements indicated a predominantly conductive hearing loss in Hyp mice compared to wild type (WT) mice. Therefore, we carried out an in-depth histomorphometric and scanning electron microscopic analysis of the auditory ossicles. Quantitative backscattered electron imaging (qBEI) indicated a severe hypomineralization of the ossicles in Hyp mice, evidenced by lower calcium content (CaMean) and higher void volume (i.e., porosity) compared to WT mice. Histologically, voids correlated with unmineralized bone (i.e., osteoid), and the osteoid volume per bone volume (OV/BV) was markedly higher in Hyp mice than WT mice. The density of osteocyte lacunae was lower in Hyp mice than in WT mice, whereas osteocyte lacunae were enlarged. Taken together, our findings highlight the importance of ossicular mineralization for hearing conduction and point towards the potential benefit of improving mineralization to prevent hearing loss in XLH. This article is protected by copyright. All rights reserved.

    View details for DOI 10.1002/jbmr.4443

    View details for PubMedID 34523743

  • Two Photon Fluorescence Microscopy of the Unstained Human Cochlea Reveals Organ of Corti Cytoarchitecture FRONTIERS IN CELLULAR NEUROSCIENCE Iyer, J. S., Seist, R., Moon, I., Stankovic, K. M. 2021; 15: 690953


    Sensorineural hearing loss (SNHL) is the most common sensory deficit worldwide, and it typically originates from the cochlea. Methods to visualize intracochlear cells in living people are currently lacking, limiting not only diagnostics but also therapies for SNHL. Two-photon fluorescence microscopy (TPFM) is a high-resolution optical imaging technique. Here we demonstrate that TPFM enables visualization of sensory cells and auditory nerve fibers in an unstained, non-decalcified adult human cochlea.

    View details for DOI 10.3389/fncel.2021.690953

    View details for Web of Science ID 000684747300004

    View details for PubMedID 34421541

    View details for PubMedCentralID PMC8376148

  • Cochlin Deficiency Protects Against Noise-Induced Hearing Loss FRONTIERS IN MOLECULAR NEUROSCIENCE Seist, R., Landegger, L. D., Robertson, N. G., Vasilijic, S., Morton, C. C., Stankovic, K. M. 2021; 14: 670013


    Cochlin is the most abundant protein in the inner ear. To study its function in response to noise trauma, we exposed adolescent wild-type (Coch +/+ ) and cochlin knock-out (Coch -/-) mice to noise (8-16 kHz, 103 dB SPL, 2 h) that causes a permanent threshold shift and hair cell loss. Two weeks after noise exposure, Coch-/- mice had substantially less elevation in noise-induced auditory thresholds and hair cell loss than Coch + / + mice, consistent with cochlin deficiency providing protection from noise trauma. Comparison of pre-noise exposure thresholds of auditory brain stem responses (ABRs) and distortion product otoacoustic emissions (DPOAEs) in Coch-/- mice and Coch + / + littermates revealed a small and significant elevation in thresholds of Coch-/- mice, overall consistent with a small conductive hearing loss in Coch-/- mice. We show quantitatively that the pro-inflammatory component of cochlin, LCCL, is upregulated after noise exposure in perilymph of wild-type mice compared to unexposed mice, as is the enzyme catalyzing LCCL release, aggrecanase1, encoded by Adamts4. We further show that upregulation of pro-inflammatory cytokines in perilymph and cochlear soft-tissue after noise exposure is lower in cochlin knock-out than wild-type mice. Taken together, our data demonstrate for the first time that cochlin deficiency results in conductive hearing loss that protects against physiologic and molecular effects of noise trauma.

    View details for DOI 10.3389/fnmol.2021.670013

    View details for Web of Science ID 000658161600001

    View details for PubMedID 34108864

    View details for PubMedCentralID PMC8180578

  • Altered expression of genes regulating inflammation and synaptogenesis during regrowth of afferent neurons to cochlear hair cells PLOS ONE Wu, C., Brugeaud, A., Seist, R., Lin, H., Yeh, W., Petrillo, M., Coppola, G., Edge, A. B., Stankovic, K. M. 2020; 15 (10): e0238578


    The spiral ganglion neurons constitute the primary connection between auditory hair cells and the brain. The spiral ganglion afferent fibers and their synapse with hair cells do not regenerate to any significant degree in adult mammalian ears after damage. We have investigated gene expression changes after kainate-induced disruption of the synapses in a neonatal cochlear explant model in which peripheral fibers and the afferent synapse do regenerate. We compared gene expression early after damage, during regeneration of the fibers and synapses, and after completion of in vitro regeneration. These analyses revealed a total of 2.5% differentially regulated transcripts (588 out of 24,000) based on a threshold of p<0.005. Inflammatory response genes as well as genes involved in regeneration of neural circuits were upregulated in the spiral ganglion neurons and organ of Corti, where the hair cells reside. Prominent genes upregulated at several time points included genes with roles in neurogenesis (Elavl4 and Sox21), neural outgrowth (Ntrk3 and Ppp1r1c), axonal guidance (Rgmb and Sema7a), synaptogenesis (Nlgn2 and Psd2), and synaptic vesicular function (Syt8 and Syn1). Immunohistochemical and in situ hybridization analysis of genes that had not previously been described in the cochlea confirmed their cochlear expression. The time course of expression of these genes suggests that kainate treatment resulted in a two-phase response in spiral ganglion neurons: an acute response consistent with inflammation, followed by an upregulation of neural regeneration genes. Identification of the genes activated during regeneration of these fibers suggests candidates that could be targeted to enhance regeneration in adult ears.

    View details for DOI 10.1371/journal.pone.0238578

    View details for Web of Science ID 000590270000017

    View details for PubMedID 33001981

    View details for PubMedCentralID PMC7529247

  • Regeneration of Cochlear Synapses by Systemic Administration of a Bisphosphonate FRONTIERS IN MOLECULAR NEUROSCIENCE Seist, R., Tong, M., Landegger, L. D., Vasilijic, S., Hyakusoku, H., Katsumi, S., McKenna, C. E., Edge, A. B., Stankovic, K. M. 2020; 13: 87


    Sensorineural hearing loss (SNHL) caused by noise exposure and attendant loss of glutamatergic synapses between cochlear spiral ganglion neurons (SGNs) and hair cells is the most common sensory deficit worldwide. We show here that systemic administration of a bisphosphonate to mice 24 h after synaptopathic noise exposure regenerated synapses between inner hair cells and SGNs and restored cochlear function. We further demonstrate that this effect is mediated by inhibition of the mevalonate pathway. These results are highly significant because they suggest that bisphosphonates could reverse cochlear synaptopathy for the treatment of SNHL.

    View details for DOI 10.3389/fnmol.2020.00087

    View details for Web of Science ID 000556565200001

    View details for PubMedID 32765216

    View details for PubMedCentralID PMC7381223

  • Combination therapy with mTOR kinase inhibitor and dasatinib as a novel therapeutic strategy for vestibular schwannoma SCIENTIFIC REPORTS Sagers, J. E., Beauchamp, R. L., Zhang, Y., Vasilijic, S., Wu, L., DeSouza, P., Seist, R., Zhou, W., Xu, L., Ramesh, V., Stankovic, K. M. 2020; 10 (1): 4211


    Neurofibromatosis type 2 (NF2) is an inherited disorder characterized by bilateral vestibular schwannomas (VS) that arise from neoplastic Schwann cells (SCs). NF2-associated VSs are often accompanied by meningioma (MN), and the majority of NF2 patients show loss of the NF2 tumor suppressor. mTORC1 and mTORC2-specific serum/glucocorticoid-regulated kinase 1 (SGK1) are constitutively activated in MN with loss of NF2. In a recent high-throughput kinome screen in NF2-null human arachnoidal and meningioma cells, we showed activation of EPH RTKs, c-KIT, and SFK members independent of mTORC1/2 activation. Subsequently, we demonstrated in vitro and in vivo efficacy of combination therapy with the dual mTORC1/2 inhibitor AZD2014 and the multi-kinase inhibitor dasatinib. For these reasons, we investigated activated mTORC1/2 and EPH receptor-mediated signaling in sporadic and NF2-associated VS. Using primary human VS cells and a mouse allograft model of schwannoma, we evaluated the dual mTORC1/2 inhibitor AZD2014 and the tyrosine kinase inhibitor dasatinib as monotherapies and in combination. Escalating dose-response experiments on primary VS cells grown from 15 human tumors show that combination therapy with AZD2014 and dasatinib is more effective at reducing metabolic activity than either drug alone and exhibits a therapeutic effect at a physiologically reasonable concentration (~0.1 µM). In vivo, while AZD2014 and dasatinib each inhibit tumor growth alone, the effect of combination therapy exceeds that of either drug. Co-targeting the mTOR and EPH receptor pathways with these or similar compounds may constitute a novel therapeutic strategy for VS, a condition for which there is no FDA-approved pharmacotherapy.

    View details for DOI 10.1038/s41598-020-60156-6

    View details for Web of Science ID 000560099700001

    View details for PubMedID 32144278

    View details for PubMedCentralID PMC7060236

  • Cytokine Levels in Inner Ear Fluid of Young and Aged Mice as Molecular Biomarkers of Noise-Induced Hearing Loss FRONTIERS IN NEUROLOGY Landegger, L. D., Vasilijic, S., Fujita, T., Soares, V. Y., Seist, R., Xu, L., Stankovic, K. M. 2019; 10: 977


    Sensorineural hearing loss (SNHL) is the most common sensory deficit worldwide, frequently caused by noise trauma and aging, with inflammation being implicated in both pathologies. Here, we provide the first direct measurements of proinflammatory cytokines in inner ear fluid, perilymph, of adolescent and 2-year-old mice. The perilymph of adolescent mice exposed to the noise intensity resulting in permanent auditory threshold elevations had significantly increased levels of IL-6, TNF-α, and CXCL1 6 h after exposure, with CXCL1 levels being most elevated (19.3 ± 6.2 fold). We next provide the first immunohistochemical localization of CXCL1 in specific cochlear supporting cells, and its presumed receptor, Duffy antigen receptor for chemokines (DARC), in hair cells and spiral ganglion neurons. Our results demonstrate the feasibility of molecular diagnostics of SNHL using only 0.5 μL of perilymph, and motivate future sub-μL based diagnostics of human SNHL based on liquid biopsy of the inner ear to guide therapy, promote hearing protection, and monitor response to treatment.

    View details for DOI 10.3389/fneur.2019.00977

    View details for Web of Science ID 000485185600001

    View details for PubMedID 31632328

    View details for PubMedCentralID PMC6749100

  • Cutting off the power: inhibition of leukemia cell growth by pausing basal ATP release and P2X receptor signaling? PURINERGIC SIGNALLING Ledderose, C., Woehrle, T., Ledderose, S., Strasser, K., Seist, R., Bao, Y., Zhang, J., Junger, W. G. 2016; 12 (3): 439-451


    T cells respond to antigen stimulation with the rapid release of cellular ATP, which stimulates an autocrine feedback mechanism that regulates calcium influx through P2X receptors. This autocrine purinergic feedback mechanism plays an essential role in the activation of T cells resulting in cell proliferation and clonal expansion. We recently reported that increases in mitochondrial ATP production drive this stimulation-induced purinergic signaling mechanism but that low-level mitochondrial ATP production fuels basal T cell functions required to maintain vigilance of unstimulated T cells. Here we studied whether defects in these purinergic signaling mechanisms are involved in the unwanted proliferation of leukemia T cells. We found that acute leukemia T cells (Jurkat) possess a larger number and more active mitochondria than their healthy counterparts. Jurkat cells have higher intracellular ATP concentrations and generat more extracellular ATP than unstimulated T cells from healthy donors. As a result, increased purinergic signaling through P2X1 and P2X7 receptors elevates baseline levels of cytosolic Ca(2+) in Jurkat cells. We found that pharmacological inhibition of this basal purinergic signaling mechanism decreases mitochondrial activity, Ca(2+) signaling, and cell proliferation. Similar results were seen in the leukemic cell lines THP-1, U-937, and HL-60. Combined treatment with inhibitors of P2X1 or P2X7 receptors and the chemotherapeutic agent 6-mercaptopurine completely blocked Jurkat cell proliferation. Our results demonstrate that increased mitochondrial metabolism promotes autocrine purinergic signaling and uncontrolled proliferation of leukemia cells. These findings suggest that deranged purinergic signaling can result in T cell malignancy and that therapeutic targeting aimed at purinergic signaling is a potential strategy to combat T cell leukemia.

    View details for DOI 10.1007/s11302-016-9510-y

    View details for Web of Science ID 000383603600003

    View details for PubMedID 27020575

    View details for PubMedCentralID PMC5023625