Gene expression analysis of oxidative stress-related genes in the apical, middle, and basal turns of the cochlea.

It can be observed from aminoglycoside-induced hair cell damage that the cochlea basal turn is more susceptible to trauma than the apex. Drug-induced hearing loss is closely related to oxidative damage. The basilar membrane directly exposed to these ototoxic drugs exhibits differences in damage, indicating that there is an inherent difference in the sensitivity to oxidative damage from the apex to the base of the cochlea. It has been reported that the morphology and characteristics of the cochlea vary from the apex to the base. Therefore, we investigated oxidative stress-related gene expression profiles in the apical, middle, and basal turns of the cochlea. The Oxidative Stress RT2 Profiler™ PCR Array revealed that three of the 84 genes (Mb, Mpo, and Ncf1) were upregulated in the middle turn compared to their level in the apical turn. Moreover, eight genes (Mb, Duox1, Ncf1, Ngb, Fmo2, Gpx3, Mpo, and Gstk1) were upregulated in the basal turn compared to their level in the apical turn. The qPCR verification data were similar to that of the PCR Array. We found that MPO was expressed in the rat cochlea and protected against gentamicin-induced hair cell death. This study summarized the data for the gradient of expression of oxidative stress-related genes in the cochlea and found potential candidate targets for prevention of ototoxic deafness, which may provide new insights for cochlear pathology.

Translocation of telomerase reverse transcriptase coincided with ATP release in postnatal cochlear supporting cells.

The spontaneous bursts of electrical activity in the developing auditory system are derived from the periodic release of adenosine triphosphate (ATP) by supporting cells in the Kölliker's organ. However, the mechanisms responsible for initiating spontaneous ATP release have not been determined. Our previous study revealed that telomerase reverse transcriptase (TERT) is expressed in the basilar membrane during the first postnatal week. Its role in cochlear development remains unclear. In this study, we investigated the expression and role of TERT in postnatal cochlea supporting cells. Our results revealed that in postnatal cochlear Kölliker's organ supporting cells, TERT shifts from the nucleus into the cytoplasm over time. We found that the TERT translocation tendency in postnatal cochlear supporting cells in vitro coincided with that observed in vivo. Further analysis showed that TERT in the cytoplasm was mainly located in mitochondria in the absence of oxidative stress or apoptosis, suggesting that TERT in mitochondria plays roles other than antioxidant or anti-apoptotic functions. We observed increased ATP synthesis, release and activation of purine signaling systems in supporting cells during the first 10 postnatal days. The phenomenon that TERT translocation coincided with changes in ATP synthesis, release and activation of the purine signaling system in postnatal cochlear supporting cells suggested that TERT may be involved in regulating ATP release and activation of the purine signaling system. Our study provides a new research direction for exploring the spontaneous electrical activity of the cochlea during the early postnatal period.

[Comparison between macroscopic identification and DNA barcoding identification of Amomi Fructus].

This study aims to explore the consistency between macroscopic identification and DNA barcoding identification of Amomi Fructus. With the DNA barcoding identification results, we evaluated the reliability of identifying Amomi Fructus quality by combining macroscopic traits with main volatile chemical components. Thirteen batches of Amomi Fructus samples were collected for identification. Firstly, the morphological and sensory characteristics of each sample were observed and recorded according to the standard in Chinese Pharmacopoeia(2020 edition). The 100-fruit weight, longitudinal diameter, transverse diameter, and longitudinal diameter-to-transverse diameter ratio were measured, which correspond to large, solid, and full kernel representing good quality in the sensory evaluation. The odor value detected by electronic nose and major volatile components(borneol, camphor, limonene, and borneol acetate) correspond to the sensory evaluation of strong odor representing good quality. Secondly, DNA barcoding was employed to identify the 13 batches of samples. Finally, clustering analysis was performed for the main volatile components and macroscopic traits, and the identification results were compared with those of DNA barcoding. Except two batches of samples(No.6 and No.10), the macroscopic identification showed the results consistent with those of DNA barcoding, with an identification rate of 84.62%. The clustering results of the content of four volatile chemical components and macroscopic traits were also consistent with the DNA barcoding identification results. DNA barcoding can verify the results of macroscopic identification and provide a scientific basis for the inheritance and development of macroscopic identification. Moreover, the combination of macroscopic traits and chemical components demonstrates higher accuracy in the quality evaluation of Chinese medicinal materials.

Heme Oxygenase-1 Protects Hair Cells From Gentamicin-Induced Death.

Gentamicin ototoxicity can generate free radicals within the inner ear, leading to permanent damage to sensory hair cells (HCs) and eventually hearing loss. The following study examined the alterations of oxidative damage-related genes in the cochlea and important molecules responsible for oxidation following gentamicin injury in vitro. The RT2 Profiler polymerase chain reaction (PCR) array was used to screen candidate targets for treatment to prevent hearing loss caused by gentamicin. We found that during gentamicin-induced death in HCs, Heme oxygenase-1 (HO-1) had a high fold change in the HCs of the cochlea. Moreover, the use of CoPPIX to induce HO-1 inhibited gentamicin-induced HC death, while HO-1 inhibitors ZnPPIX after CoPPIX reversed this process. Furthermore, the inhibitors of NF-E2-related factor-2 (Nrf2) reduced the expression of HO-1 and inhibited the protective effect of HO-1 after gentamicin, thus suggesting that the Nrf2/HO-1 axis might regulate gentamicin-associated ototoxicity. We further demonstrated that induction of HO-1 up-regulated the expression of Nrf2 in both cochlear and HEI-OC1 cells. In summary, these findings indicated that HO-1 protects HCs from gentamicin by up-regulating its expression in HCs and interacting with Nrf2 to inhibit reactive oxygen species (ROS).

Hsp70/Bmi1-FoxO1-SOD Signaling Pathway Contributes to the Protective Effect of Sound Conditioning against Acute Acoustic Trauma in a Rat Model.

Sound conditioning (SC) is defined as "toughening" to lower levels of sound over time, which reduces a subsequent noise-induced threshold shift. Although the protective effect of SC in mammals is generally understood, the exact mechanisms involved have not yet been elucidated. To confirm the protective effect of SC against noise exposure (NE) and the stress-related signaling pathway of its rescue, we observed target molecule changes caused by SC of low frequency prior to NE as well as histology analysis in vivo and verified the suggested mechanisms in SGNs in vitro. Further, we investigated the potential role of Hsp70 and Bmi1 in SC by targeting SOD1 and SOD2 which are regulated by the FoxO1 signaling pathway based on mitochondrial function and reactive oxygen species (ROS) levels. Finally, we sought to identify the possible molecular mechanisms associated with the beneficial effects of SC against noise-induced trauma. Data from the rat model were evaluated by western blot, immunofluorescence, and RT-PCR. The results revealed that SC upregulated Hsp70, Bmi1, FoxO1, SOD1, and SOD2 expression in spiral ganglion neurons (SGNs). Moreover, the auditory brainstem responses (ABRs) and electron microscopy revealed that SC could protect against acute acoustic trauma (AAT) based on a significant reduction of hearing impairment and visible reduction in outer hair cell loss as well as ultrastructural changes in OHCs and SGNs. Collectively, these results suggested that the contribution of Bmi1 toward decreased sensitivity to noise-induced trauma following SC was triggered by Hsp70 induction and associated with enhancement of the antioxidant system and decreased mitochondrial superoxide accumulation. This contribution of Bmi1 was achieved by direct targeting of SOD1 and SOD2, which was regulated by FoxO1. Therefore, the Hsp70/Bmi1-FoxO1-SOD signaling pathway might contribute to the protective effect of SC against AAT in a rat model.

The Absorption Characteristics of Nonvolatile Components in a Water Extraction From Amomi fructus as Determined by In Situ Single-Pass Intestinal Perfusion and High-Performance Liquid Chromatography.

BACKGROUND: Amomi fructus is a famous traditional Chinese medicine (TCM) that can exert beneficial effects during the treatment of gastrointestinal diseases and is used widely in China and other countries in Southeast Asia. However, the nonvolatile active ingredients that are present in the water extractions from A. fructus used to treat gastrointestinal diseases have yet to be elucidated. The goal of this study was to identify the nonvolatile active ingredients of A. fructus. METHODS: We used an in situ single-pass intestinal perfusion (SPIP) model to identify the active ingredients of A. fructus that play significant roles in gastrointestinal absorption. In addition, we developed a high-performance liquid chromatography (HPLC) method to identify key fractions in intestinal outflow perfusate. RESULTS: Nineteen components were identified in a water extraction from A. fructus; these exhibited different absorption capabilities in different intestinal segments. Of these, six components were determined by the newly developed HPLC method: catechin, vanillic acid, epicatechin, polydatin, isoquercitrin, and quercitrin. CONCLUSIONS: The current study aimed to identify the active ingredients present in water extractions prepared from A. fructus in a single-intestinal perfusate from rats. Our findings provide an experimental basis to explain the pharmacodynamic actions of A. fructus.

miR-182 prevented ototoxic deafness induced by co-administration of kanamycin and furosemide in rats.

Ototoxic drugs may induce auditory sensory hair cell loss and permanent deafness; however, there is still no effective treatments or prevention strategies for this side effect. A recent study found that microRNA182 (miR-182) protected cochlear hair cells from ototoxic drug-induced apoptosis in vitro. However, it remains unclear whether miR-182 can protect drug-induced deafness in vivo. In this study, we overexpressed cochlear miR-182 in Sprague-Dawley rats by trans-round window niche delivery of miR-182 mimics. The rats subsequently received intraperitoneal injections of kanamycin and furosemide to induce acute cochlear outer hair cell death and permanent deafness. Auditory brainstem response tests showed that miR-182 attenuated permanent threshold shifts. Consistent with this result, miR-182 reduced the loss of outer hair cells and missing stereocilia. miR-182 treatment also increased the level of phosphoinositide-3 kinase regulatory subunit p85α in the outer hair cells after co-administration of kanamycin and furosemide. Our findings suggest that miR-182 has powerful protective potential against ototoxic drug-induced acute auditory sensory hair cell loss and permanent deafness.

In vivo construction of lymphoid node by implantation of adipose-derived stromal cells with hydroxypropyl methyl cellulose hydrogel in BALB/c nude mice.

Adipose-derived stromal cells have multilineage potential to differentiate into several specialized tissue types. Herein, we investigated whether ADSCs could differentiate into lymphoid node in vivo. Human ADSCs from routine liposuction were cultured in differentiation medium and were supplemented with transforming growth factor ß1 (TGF)-ß1 and basic fibroblast growth factor (bFGF). The induced hADSCs mixed with 13% (w/v) hydroxypropyl methylcellulose (HPMC) were injected into BALB/c nude mice subcutaneously. Eight weeks later, nodules were found under the injected sites. Histology, immunohistochemistry, and species identification analysis confirmed that the nodules were lymphoid nodes that were derived from the injected hADSCs. Our experiment demonstrated that the hADSCs could differentiate into lymphocyte-like cells and form lymphoid nodes in vivo. TGF-ß1 and bFGF might play important roles in the differentiation of hADSCs into lymphocyte-like cells. Our study might present an alternative approach for engineering immune organs and thus offer potential treatment for immunodeficiency diseases.

Decreased expression of TERT correlated with postnatal cochlear development and proliferation reduction of cochlear progenitor cells.

Cochlear progenitor cells are considered as one of the best candidates for hair cell regeneration, thus, the regulation of cochlear progenitor cell proliferation has become a focus in this field. Several genes expressed in the inner ear during postnatal development have been demonstrated to be involved in maintaining the proliferative potential of progenitor cells, but the mechanism for regulating the proliferation and differentiation of cochlear progenitor cells remains poorly understood. Telomerase reverse transcriptase (TERT) has rate limiting telomerase activity and the overexpression of TERT has been shown to promote cell proliferation in series of cell lines. The aim of the present study was to evaluate the expression of TERT in the postnatal development of the cochlea and progenitor cells. The results demonstrated that TERT was expressed in the basilar membranes during the first postnatal week. In vitro, TERT expression in progenitor cells reached a maximum at day 4 after culture and decreased as the culture time prolonged or the cell passage number increased. These results led us to hypothesize that TERT may be involved in the development of the cochlea and in maintaining the proliferation ability of progenitor cells.

Combinatorial enzymatic digestion with thermolysin and collagenase type I improved the isolation and culture effects of hair cell progenitors from rat cochleae.

The high incidence of hearing loss in human combined with the lack of hair cell regeneration in mammalian cochleae had got the attention to manipulate stem/progenitor cells to participate in hair cell regeneration for years. Cochlear progenitor cells are considered as the best candidate for hair cell regeneration. However, there is not any effective and feasible way to separate hair cell progenitors from rat cochleae, yet. In this study, we tried to isolate single epithelial cells from rat basilar membrane by combinatorial enzymatic digestion with thermolysin and collagenase type I. The results showed that the harvested single cells gave rise to otospheres with features of stem cells and could be induced to differentiate into hair cells. Significantly, more otospheres of epithelial origin were obtained by digesting with thermolysin and collagenase type I. The combinatorial enzymatic digestion would be a potential method for hair cell progenitor isolation and culture with broad applications.

[Rancidness of Armeniacae Semen Amarum involving Bianzhuang Lunzhi].

This article aims to compare the qualities of Armeniacae Semen Amarum before and after rancidness, in order to study the rancidness of Armeniacae Semen Amarum. In the experiment, content of fatty oil, acid value and peroxide value were determined before and after rancidness,respectively. Meanwhile, HPLC, GC-MS were utilized to analyze laetrile and fatty acid components. Besides, colorimeter and e-nose were introduced to quantify and compare "color and odor". A correlation analysis was conducted on the above results. The results showed that color of post-rancidness Armeniacae Semen Amarum changed from yellow to brown, with sour and lower content of laetrile. On the contrary, acid and peroxide values increased significantly, with changes in fatty acid component. There was a considerable correlation between appearance characteristics and changes in internal quality. The "sensory analysis-quality identification system" can provide a certain scientific basis for prediction of the content of chemical components in traditional Chinese medicine, preliminary judgment of quality of traditional Chinese medicine and real-time quality monitoring, which offers us novel ideas and reference for storage principles of traditional Chinese medicines of "pre-event prediction, during-event intervention and post-event identification".

Apoptosis-inducing factor and calpain upregulation in glutamate-induced injury of rat spiral ganglion neurons.

Spiral ganglion neuron (SGN) damage and apoptosis can lead to noise-induced hearing loss, age-associated hearing loss and, in certain cases, auditory neuropathy. The apoptosis-inducing factor (AIF)-associated pathway may be important in this process. The present study aimed to investigate the expression levels of AIF and calpain in damaged SGNs. Glutamate (Glu) perfusion and cell culture in different concentrations of Glu were performed to damage the SGNs of Sprague-Dawley (SD) rats, with saline water used as a control Different concentrations (5, 10, 20 and 40 mM) of Glu were injected into the cochlear tympanic canal of 18 SD rats, and 10, 20 and 40 mM Glu were added to SGN cultures. Auditory brainstem responses (ABR) were measured prior to and 2 days following the injection of Glu. Immunofluorescent staining was used to detect the SGN damage and the expression levels of AIF and calpain in vivo and in in vitro. Transmission electron microscopy (TEM) was used to measure cell apoptosis and reverse transcription-quantitative polymerase chain reaction was used to analyse the gene expression levels of AIF and calpain in the damaged SGNs. The TEM identified mitochondrial vacuolisation, swelling of the SGN and heterochromatin formation. Injection of Glu reduced the number of SGNs and induced apoptosis. AIF was observed to translocate into the nuclei of the SGNs in the 20 and 40 mM Glu groups, and the expression levels of AIF and calpain were markedly upregulated in the modiolus of the Glu-damaged SGNs. The upregulation of AIF and calpain may be important in the process of SGN damage and apoptosis.

Calpain inhibitor PD150606 attenuates glutamate induced spiral ganglion neuron apoptosis through apoptosis inducing factor pathway in vitro.

OBJECTIVE: This research aimed to investigate whether glutamate induced spiral ganglion neurons (SGNs) apoptosis through apoptosis inducing factor (AIF) pathway. And verify whether PD150606, a calpain inhibitor could prevent apoptosis by inhibiting cleaving and releasing AIF in mitochondrion. METHODS: SGNs of postnatal days 0-3 were harvested and cultured in dishes. 20 mM Glu, the caspase inhibitor Z-VAD-FMK and calpain inhibitor PD150606 were added into cultured dishes separately. We used optical microscope and immunofluoresence staining to observe cell morphology and AIF distribution, RT-PCR and Westernblot to analyse AIF and calpain expression in SGNs. TUNEL assay was used to test cell apoptosis. RESULTS: Cell morphology and nuclear translocation of AIF were altered in SGNs by 20 mM Glu treated in vitro. The axon of SGN shortened, more apoptosis SGN were observed and the expression of AIF and calpain were up-regulated in Glu-treated group than the normal one (P<0.05). The same experiments were conducted in 20 mM+PD150606 treated group and 20 mM+Z-VAD-FMK group. Obviously AIF were located from cytoplasm to the nuclear and the expressions of AIF and calpain were down-regulated by PD150606 (P<0.05). Positive cells in TUNEL staining decreased after PD150606 treating. However, Z-VAD-FMK had no influence on AIF, calpain expression or cell apoptosis. CONCLUSION: The AIF-related apoptosis pathway is involved in the process of Glu-induced SGN injury. Furthermore, the inhibition of calpain can prevent AIF from releasing the nuclear or inducing SGN apoptosis.

Viral-mediated expression of c-Myc and cyclin A2 induces cochlear progenitor cell proliferation.

Cochlear progenitor cells have a limited proliferative capability, which prevents their application in treating sensorineural hearing loss. In this study, we showed that the expression of c-Myc and cyclin A2 was down-regulated during the development of cochlear tissue and CPC differentiation. Over-expression of these two genes using adenovirus transduction, significantly affected the CPC cell cycle and promoted the CPC proliferation. We further demonstrated that this promotion involves the classic CKI-cyclin-CDK pathway. Our study suggests that genetically modified CPCs may be a promising cell source for cochlear stem cell transplantation that improves the efficacy of cell therapy.

MicroRNA-195 chemosensitizes colon cancer cells to the chemotherapeutic drug doxorubicin by targeting the first binding site of BCL2L2 mRNA.

The mechanisms underlying doxorubicin (Dox) resistance in colon cancer cells are not fully understood. MicroRNA (miRNA) play important roles in tumorigenesis and drug resistance. However, the relationship between miRNA and Dox resistance in colon cancer cells has not been previously explored. In this study, we utilized microRNA array and real-time PCR to verify that miR-127, miR-195, miR-22, miR-137 were significantly down-regulated, while miR-21, miR-592 were up-regulated in both HT29/DOX and LOVO/DOX cell lines. In vitro cell viability assay showed that knockdown of miR-195 in HT29 and LOVO cells caused a marked inhibition of Dox-induced cytotoxicity. Moreover, we explored that miR-195 is involved in repression of BCL2L2 expression through targeting its 3'-untranslated region, especially the first binding site within its mRNA. Furthermore, down-regulation of miR-195 conferred DOX resistance in parental cells and reduced cell apoptosis activity, while over-expression of miR-195 sensitized resistant cells to DOX and enhanced cell apoptosis activity, all of which can be partly rescued by BCL2L2 siRNA and cDNA expression. These results may have implications for therapeutic strategies aiming to overcome colon cancer cell resistance to Dox.

[Inhibition of Th2 reaction by thymic stromal lymphopoietin blockade in vitro].

OBJECTIVE: To explore the effect of thymic stromal lymphopoietin (TSLP) on transformation of dendritic cell (DC) and T cell in vitro. METHODS: Mouse-derived immature dendritic cells and T lymphocytes were co-cultured in vitro, which were divided into 4 groups (TSLP stimulation group, TSLP stimulation and its receptor blocking group, ovalbumin stimulation group and ovalbumin stimulation and TSLP receptor blocking group). IL-4, IL-8 and IFN-ß in cell culture supernatant were detected after 2 days by ELISA. SPSS 13.0 software was used to analyze the data. RESULTS: IL-4 levels of TSLP receptor blocking groups [(48.84 ± 1.56) pg/ml, (52.53 ± 2.36) pg/ml]were significantly lower than those of corresponding TSLP stimulation group and ovalbumin stimulation group [(72.55 ± 7.76) pg/ml, (80.47 ± 21.93) pg/ml;t = 5.994, P < 0.05;t = 2.534, P < 0.05]. However, there were not significant differences of IL-8 and IFN-ß expression between corresponding two groups of whether or not TSLP receptor blocking (all P > 0.05). CONCLUSION: TSLP receptor blockade in vitro can inhibit T lymphocyte transformation to Th2, which may provide a new therapeutic strategy for clinical Th2 dominant diseases such as allergic rhinitis and asthma.

The role of RIP3 mediated necroptosis in ouabain-induced spiral ganglion neurons injuries.

Spiral ganglion neuron (SGN) injury is a generally accepted precursor of auditory neuropathy. Receptor-interacting protein 3 (RIP3) has been reported as an important necroptosis pathway mediator that can be blocked by necrostatin-1 (Nec-1). In our study, we sought to identify whether necroptosis participated in SGN injury. Ouabain was applied to establish an SGN injury model. We measured the auditory brain-stem response (ABR) threshold shift as an indicator of the auditory conditions. Positive ß3-tubulin immunofluorescence staining indicated the surviving SGNs. RIP3 expression was evaluated using immunofluorescence, quantitative real-time polymerase chain reaction and western blot. SGN injury promoted an increase in RIP3 expression that could be suppressed by application of the necroptosis inhibitor Nec-1. A decreased ABR threshold shift and increased SGN density were observed when Nec-1 was administered with apoptosis inhibitor N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD). These results demonstrated that necroptosis is an indispensable pathway separately from apoptosis leading to SGN death pathway, in which RIP3 plays an important role.

Acoustical stimulus changes the expression of stromal cell-derived factor-1 in the spiral ganglion neurons of the rat cochlea.

Neural stem cell (NSC) transplantation into the cochlea has been tested as a treatment for spiral ganglion neuron (SGN) degenerative disease and injury in various animal models. A recent study has shown evidence of functional recovery after transplantation of the stem cells into a degenerated-SGN model. Chemokine stromal cell-derived factor-1 (SDF-1, or known as CXC chemokine ligand-12, CXCL-12) signaling through CXCR4 has previously been identified as a key step in the homing of the stem cells within the injury areas; meanwhile, studies have revealed that the SDF-1/CXCR4 axis is also involved in axon guidance and pathfinding. A study found that transplanted neural precursor cells can migrate to the root of the auditory nerve when animals are subjected to an augmented acoustic environment (AAE). In accordance with these studies, we hypothesize that AAE will up-regulate the expression of SDF-1 in acoustic nerves. We tested our hypothesis by examining the expression of SDF-1 in different acoustic environments, and the results were confirmed by the auditory brainstem response (ABR), immunohistochemical and RT-PCR analyses. The results showed that SDF-1 was expressed at a relatively low level in the SGNs under normal animal unit acoustic conditions (40-50 dB). Moreover, it was significantly up-regulated in the SGNs under the 75 dB (augmented physiological process without hearing loss) and 90 dB AAE (pathological process with light hearing loss) conditions; however, under the 115 dB AAE (pathological process with severe hearing loss) condition, the expression of SDF-1 was not up-regulated. The results confirmed that appropriately augmented acoustical stimuli lead to the up-regulation of SDF-1, which may assist in the migration of the transplanted cells and the subsequent establishment of essential synaptic contacts between the exogenous cells and the host auditory pathway.

Wnt1 from cochlear schwann cells enhances neuronal differentiation of transplanted neural stem cells in a rat spiral ganglion neuron degeneration model.

Although neural stem cell (NSC) transplantation is widely expected to become a therapy for nervous system degenerative diseases and injuries, the low neuronal differentiation rate of NSCs transplanted into the inner ear is a major obstacle for the successful treatment of spiral ganglion neuron (SGN) degeneration. In this study, we validated whether the local microenvironment influences the neuronal differentiation of transplanted NSCs in the inner ear. Using a rat SGN degeneration model, we demonstrated that transplanted NSCs were more likely to differentiate into microtubule-associated protein 2 (MAP2)-positive neurons in SGN-degenerated cochleae than in control cochleae. Using real-time quantitative PCR and an immunofluorescence assay, we also proved that the expression of Wnt1 (a ligand of Wnt signaling) increases significantly in Schwann cells in the SGN-degenerated cochlea. We further verified that NSC cultures express receptors and signaling components for Wnts. Based on these expression patterns, we hypothesized that Schwann cell-derived Wnt1 and Wnt signaling might be involved in the regulation of the neuronal differentiation of transplanted NSCs. We verified our hypothesis in vitro using a coculture system. We transduced a lentiviral vector expressing Wnt1 into cochlear Schwann cell cultures and cocultured them with NSC cultures. The coculture with Wnt1-expressing Schwann cells resulted in a significant increase in the percentage of NSCs that differentiated into MAP2-positive neurons, whereas this differentiation-enhancing effect was prevented by Dkk1 (an inhibitor of the Wnt signaling pathway). These results suggested that Wnt1 derived from cochlear Schwann cells enhanced the neuronal differentiation of transplanted NSCs through Wnt signaling pathway activation. Alterations of the microenvironment deserve detailed investigation because they may help us to conceive effective strategies to overcome the barrier of the low differentiation rate of transplanted NSCs.

G-CSF attenuates noise-induced hearing loss.

In this study, we investigated the effects of granulocyte colony-stimulating factor (G-CSF) for the treatment of noise-induced hearing loss (NIHL) in a guinea pig model. Forty guinea pigs were randomly divided into four groups: control, noise (white noise, 3 h/d for 2 days at 115 dB), noise+G-CSF (350 µg/kg/d for 5 days), and noise+saline. Auditory brainstem response (ABR) and distortion product otoacoustic emission (DPOAE) were used to determine the hearing threshold and outer hair cell function, respectively, in each group. Cochlear morphology was examined to evaluate hair cell injury induced by intense noise exposure. Fourteen days after noise exposure, the noise+G-CSF group had a lower ABR value than the noise group (P<0.05) or the noise+saline group (P<0.01). At most frequencies, the DPOAE value of the noise+G-CSF group showed a significant rise (P<0.05) compared to the noise group or the noise+saline group. Neither the ABR value nor the DPOAE value differed between the noise group and the noise+saline group. The morphology of the phalloidin-stained organ of Corti was consistent with the functional measurements. In conclusion, G-CSF can preserve hearing in an experimental model of NIHL in guinea pigs, by preserving hair cells after intense noise exposure.