Mesenchymal stem cells from human umbilical cord express preferentially secreted factors related to neuroprotection, neurogenesis, and angiogenesis. 8600 Rockville Pike Although glutamate is the excitatory transmitter in the brain required for proper function, post-traumatic burst release of glutamate leads to excitotoxicity and subsequent cellular injury as has been also shown after noise-induced damage of the inner ear. doi: 10.1002/stem.1129, Yamane, H., Nakai, Y., Takayama, M., Iguchi, H., Nakagawa, T., and Kojima, A. Cell. This suggests that neuronal repair and control of excitotoxicity could be key components of recovery from severe sound trauma. Delivery of MSCs did not have any effect on baseline hearing. Shearer, A. E., Hildebrand, M. S., and Smith, R. J. For control cochleae, an average of 155 ng/l of RNA and for MSC treated cochleae 182 ng/l of RNA were extracted. Human Mesenchymal Stem Cell Treatment Normalizes Cortical Gene Expression after Traumatic Brain Injury. doi: 10.1016/j.heares.2015.10.010, Yogarajah, M., Matarin, M., Vollmar, C., Thompson, P. J., Duncan, J. S., Symms, M., et al. Group 3 animals were unilaterally exposed to sound followed by MSC (n = 5) or artificial perilymph (n = 5) at 48 h post-sound trauma. JH and MD data analysis and writing of the manuscript. Semi quantitative RT-PCR showed an up-regulation of TNF gene expression after sound trauma. J. Neurosci. Clinical trials with mesenchymal stem cells: an update. Up regulated and down regulated genes were identified and organized according to their gene ontology (GO) classification. 146, 40204032. Neurosci. A mouse model for human deafness DFNB22 reveals that hearing impairment is due to a loss of inner hair cell stimulation. Figure 8. Similar is observed for the neuronal differentiation factor 2, Pax6 and homeobox protein engrailed-2 (EN2). Bethesda, MD 20894, Web Policies doi: 10.1016/s0896-6273(02)00824-3, Puel, J. L., dAldin, C., Ruel, J., Ladrech, S., and Pujol, R. (1997). Near the threshold level, 5 dB SPL steps using up to 1024 presentations were carried out at each frequency. Federal government websites often end in .gov or .mil. Biochimie 95, 22712285. Cell Rep. 19, 307320. Pre-treatment ABR measurements showed normal hearing threshold in both groups (Figure 2; control ore and MSC treated pre). At 16 kHz, treated animals showed better hearing thresholds than sound trauma and artificial perilymph treated animals, but this was not statistically significant. official website and that any information you provide is encrypted

Cell. Forty-eight hours post-cell delivery, total RNA was extracted from the cochlea and RNAseq performed to evaluate the gene expression induced by the cell therapy. Revuelta M, Santaolalla F, Arteaga O, Alvarez A, Snchez-Del-Rey A, Hilario E. Ageing Res Rev. 117, 214218. 91, 883889. 15, 425437. Sections underwent antigen retrieval in a microwave using 1:10 Dako Target Retrieval Solution (DAKO Corporation, Carpinteria, CA, United States) for 30 s. Immunohistochemistry was carried out with anti-Bcl-2 rabbit polyclonal antibody diluted 1:200 (BD Biosciences, Inc., San Jose, CA, United States). Recent advances in cochlear hair cell regeneration-A promising opportunity for the treatment of age-related hearing loss. ABR recording prior to treatment at 4, 8, 16, and 32 kHz showed an average hearing threshold of 28.3 7, 35 5, 30 5, and 30 17, respectively. doi: 10.1111/cge.12915. MS electrophysiology and manuscript approval. These were also visualized as a Voronoi tree plots to facilitate identification of key pathways. Expression of bcl-2 (brown AC) and semi quantitative RT PCR of bcl-2 mRNA (D). This study also documented functional recovery induced by dosing TBI animals with thyroxine (T4) which down-regulates transthyretin. doi: 10.1371/journal.pone.0163645, Kanzaki, S., Toyoda, M., Umezawa, A., and Ogawa, K. (2020). 7:220. doi: 10.3389/fncel.2013.00220, Cunningham, C. J., Redondo-Castro, E., and Allan, S. M. (2018). The same animals underwent re-evaluation of hearing with ABR 7 days post-MSC delivery. miR-9: a versatile regulator of neurogenesis. 109, 1935119356. Treatment with MSCs after severe sound trauma induced a moderate hearing protective effect. Front. The biological effects of MSCs have been widely studied. The role of mitochondrial oxidative stress in hearing loss. Keywords: noise trauma, hearing loss, mesenchymal stroma cells, Whartons jelly, cochlear transcriptome, hearing protection, Citation: Warnecke A, Harre J, Shew M, Mellott AJ, Majewski I, Durisin M and Staecker H (2021) Successful Treatment of Noise-Induced Hearing Loss by Mesenchymal Stromal Cells: An RNAseq Analysis of Protective/Repair Pathways. (2020)]. The secondary antibody was incubated for 6 h at room temperature in a humid chamber. doi: 10.1016/j.chembiol.2016.09.001, Chen, L., Trautwein, P. G., Powers, N., and Salvi, R. J. Down-regulated genes included miRNAs such as miR-9, miR-124a, miR-124-2, and miR-133a. ABR measurements were acquired using the Smart EP program from Intelligent Hearing Systems. The size of the category corresponds to a lower FDR with the more deeply red groups representing a higher enrichment score. Mazurek B, Knipper M, Biesinger E, Schulze H. HNO. Rajguru, R. (2013). 76, 31413156. The Sound Level Meter was calibrated using a 1000 Hz Bruel & Kjaer 4230 Sound Level Calibrator. Despite these limitations, main cell signaling pathways involved in MSC-mediated inner ear protection after noise trauma have been identified by applying pathway enrichment analysis. Threshold was defined as the SPL at which at least one of the waves could be identified in 2 or more repetitions of the recording. MSCs have been demonstrated to have a wide range of immunomodulatory and protective effects after tissue injury and have been tested in diverse disorders such as stroke, acute kidney injury and myocardial infarction (Squillaro et al., 2016; Fu et al., 2017; Chang et al., 2018; Cunningham et al., 2018; Harrell et al., 2019; Tsintou et al., 2020). Cells were cultured in medium consisting of 1% penicillin-streptomycin (Life Technologies), 10% fetal bovine serum (FBS) mesenchymal stem cell qualified (MSCq) (Life Technologies), and fibroblast basal medium (Lonza Group Ltd., Basel, Switzerland), and expanded to passage 5 for all experiments. Looking at the top down-regulated genes, mainly genes coding for transcription factors or for proteins and receptors related to synaptic function are among the most regulated. (2003). B 10, 619624. One of the most down-regulated genes identified in our study was transthyretin (5.377 fold decrease in expression after MSC treatment). Additionally, severe sound trauma results in degradation of the blood labyrinthine barrier. Within the inner ear, MSCs have been shown protective against tissue damage induced by sound and a variety of ototoxins. (2020). Walters, B. J., Coak, E., Dearman, J., Bailey, G., Yamashita, T., Kuo, B., et al. German version]. Figure 2. (2014). Evaluation of extracellular vesicles produced by WJCs for the mitigation of sound trauma has demonstrated that similar protective effects can be achieved using only the secretome alone (Warnecke et al., 2020). To fully understand the relevance of the changes induced by MSC in the inner ear, investigation of further time points after damage and treatment or even transcriptomics at single cell resolution will be needed. The Genomic Basis of Noise-induced Hearing Loss: a Literature Review Organized by Cellular Pathways. Rep. 10:15167. (2016). Res. By evaluating the diverse pathways activated in this recovering system, we may be able to select new targets for the pharmacological mitigation of sound trauma. For delivery, cells were harvested, washed in phosphate buffered saline (PBS) and resuspended to a concentration of 1 106 cells/ml in sterile PBS. A separate cohort of animals was treated in a similar fashion and allowed to survive for 2 weeks post-cell therapy and hearing outcomes determined. -. Several miRNAs (miR-9, miR-124a, miR-124-2, and miR-133a) are also down-regulated after treatment with MSCs. Exosomes and Stem Cells in Degenerative Disease Diagnosis and Therapy.

Current insights in noise-induced hearing loss: a literature review of the underlying mechanism, pathophysiology, asymmetry, and management options. (2019). Finally, we identified molecules in clinical use that could modulate most significantly regulated genes, identifying 46 drugs that could potentially be used to study some of the effects induced by MSCs (Table 2). Statistical analysis was carried out by the NOISeq method. No use, distribution or reproduction is permitted which does not comply with these terms. doi: 10.1089/neu.2013.3019, Diaz-Horta, O., Abad, C., Cengiz, F. B., Bademci, G., Blackwelder, P., Walz, K., et al. Figure 3. After sound trauma there was an occasional increase in bcl-2 immunolabeling in isolated supporting cells (arrow) (B). -, Proc Natl Acad Sci U S A.

To more clearly identify effects of cell therapy the IPA software was used to compare only significantly up- and down-regulated genes to evaluate the relationship between expression data and potential pathways involved in injury and protection. Am. Before (2018). The most up and down regulated genes were additionally evaluated using Pharos software3 for identification of molecules in clinical use that can down or up-regulate the identified genes. There is no significant difference between the pre- and post-transplantation thresholds. TNF is a key factor that influences multiple processes and genes. To further characterize these effects, a more moderate sound trauma paradigm with multiple sample time points after MSC delivery and verification of the effects using PCR and immunohistochemistry will be needed. doi: 10.1016/j.ebiom.2017.01.046, Mittal, R., Grati, M., Sedlacek, M., Yuan, F., Chang, Q., Yan, D., et al. Another miRNA that we found to be down-regulated (miR-133) is involved in organogenesis, regulating the hedgehog pathway in myogenesis (Mok et al., 2018; Yu et al., 2019). Baumgartner LS, Moore E, Shook D, Messina S, Day MC, Green J, Nandy R, Seidman M, Baumgartner JE. Res. (2018). Up-regulated genes related to response to sound are specific to the ear, associated with development of the inner ear, associated with syndromes that include hearing loss or when mutated are associated with sensorineural hearing loss (Shearer et al., 1993). doi: 10.1016/j.biochi.2013.06.020, Fetoni, A. R., Lattanzi, W., Eramo, S. L. M., Barba, M., Paciello, F., Moriconi, C., et al. 2015 Mar;15(3):417-30 Thus, MSCs regulate not only transcription factors involved in neurogenesis, but also miRNAs involved in the regulation of transcription factors. Adv. The homeobox protein BarH-like 2 (Barhl2) acts as transcription factor and possesses an enhancer that can be activated by the basic helix-loop-helix factor Atoh1 and can drive spinal cord-specific gene expression. Cell Transplant. Res. Sheehan-Rooney, K., Swartz, M. E., Zhao, F., Liu, D., and Eberhart, J. K. (2013). 252, 504508. Histological evaluation of organ of Corti integrity in sound trauma (A,C) and sound trauma and MSC (B,D) treated cochlea. Molecular function annotations centered on a range of protein receptor signaling processes and processes related to matrix binding. Further insights could be gained by including analysis of the transcriptome from a group of mice not exposed to noise with and without treatment with MSC. Our data demonstrated an up-regulation in TNF gene expression which could not be confirmed by PCR (Figure 7D). Large-scale secretome analyses unveil the superior immunosuppressive phenotype of umbilical cord stromal cells as compared to other adult mesenchymal stromal cells. Expression of TNF protein (Red AC) and semi quantitative RT PCR of TNF mRNA (D). Naffah-Mazzacoratti Mda, G., Gouveia, T. L. F., Simes, P. S. R., and Perosa, S. R. (2014). Neuroscience 248, 116. Yu, X., Wang, D., Wang, X., Sun, S., Zhang, Y., Wang, S., et al. doi: 10.1007/s00018-019-03176-4, Tsintou, M., Dalamagkas, K., and Makris, N. (2020). (2014).

Neurosci.

Changes in gene expression were grouped together based on gene ontology classification. Delivery of MSCs caused a greater down-regulatory than up-regulatory effect on gene expression in sound traumatized animals treated with MSCs. Spaethling, J. M., Klein, D. M., Singh, P., and Meaney, D. F. (2008). doi: 10.1371/journal.pone.0072604. Protective paracrine effect of mesenchymal stem cells on cardiomyocytes. 127, 967976. Besides neurotrophins, MSCs produce a variety of immunomodulatory molecules and other growth factors. Neurobiol. Additional processes included transcription factor function, wound healing, regulation of apoptotic process and response to hypoxia.