Endoplasmic reticulum stress-induced necroptosis promotes cochlear inflammation: Implications for age-related hearing loss.

Age-related hearing loss (ARHL) is the most common sensory disorder associated with human aging. Chronic inflammation is supposed to be an important contributor to ARHL. Yet, the underlying mechanisms of developing cochlear inflammation are still not well understood. In this study, we found that the inflammation, endoplasmic reticulum (ER) stress and necroptosis signalings are activated in the cochlea of aged C57BL/6 mice. ER stress activator tunicamycin (TM) induced necroptosis in cochlear HEI-OC1 cells and cochlear explants, while necroptosis inhibitors protected cochlear cells from ER stress-induced cell death. The antioxidants inhibited necroptosis and protected HEI-OC1 cells from TM insults. Necroptotic HEI-OC1 cells promoted the activation of the co-cultured macrophages via Myd88 signaling. Moreover, necroptosis inhibitor protected from TM-induced hearing loss, and inhibited inflammation in C57BL/6 mice. These findings suggest that ER stress-induced necroptosis promotes cochlear inflammation and hearing loss. Targeting necroptosis serves as a potential approach for the treatment of cochlear inflammation and ARHL.

Autophagy-dependent ferroptosis contributes to cisplatin-induced hearing loss.

Cisplatin-induced hearing loss is a common side effect of cisplatin chemotherapy, for which clinical therapy remains unavailable. Apoptosis of hair cells is considered the primary cause of cisplatin-induced ototoxicity; however, inhibiting apoptosis can only partially restore cisplatin-induced hearing loss. Therefore, auditory cell death caused by cisplatin damage requires further study. Ferroptosis, a novel form of regulated cell death, has been shown to play a role in the mechanism of cisplatin toxicity. In this study, we observed proferroptotic alterations (lipid peroxidation and impaired antioxidant capacity) in the cochleae of C57BL/6 mice after cisplatin damage, verifying the induction of ferroptosis. Using the HEI-OC1 cell line, we observed that cisplatin induced proferroptotic alterations and activated ferritinophagy (specific autophagy pathway). Employing chloroquine, we confirmed that the blockage of autophagy remarkably alleviated cisplatin-induced ferroptosis in HEI-OC1 cells; therefore, the induction of ferroptosis in cisplatin-treated auditory cells was dependent on the activation of autophagy. In addition, the ferroptosis inhibitor ferrostatin-1 and iron chelator deferoxamine significantly attenuated cisplatin-induced cytotoxicity in HEI-OC1 cells and cochlear explants. Moreover, pharmacologically inhibiting ferroptosis using ferrostatin-1 significantly decreased the auditory cell loss and, notably, attenuated hearing loss in C57BL/6 mice after cisplatin damage. Collectively, these findings indicate that autophagy-dependent ferroptosis plays an integrated role in the mechanism of cisplatin-induced hearing loss.

Transcriptomic analysis highlights cochlear inflammation associated with age-related hearing loss in C57BL/6 mice using next generation sequencing.

BACKGROUND: In our aging society, age-related hearing loss (AHL) is the most common sensory disorder in old people. Much progress has been made in understanding the pathological process of AHL over the past few decades. However, the mechanism of cochlear degeneration during aging is still not fully understood. METHODS: Next generation sequencing technique was used to sequence the whole transcriptome of the cochlea of C57BL/6 mice, a mouse model of AHL. Differentially expressed genes (DEGs) were identified using the Cuffdiff software. GO and KEGG pathway enrichment analyses of the DEGs were implemented by using the GOseq R package and KOBAS software, respectively. RESULTS: A total of 731 genes (379 up- and 352 down-regulated) were revealed to be differentially expressed in the cochlea of aged mice compared to the young. Many genes associated with aging, apoptosis, necroptosis and particularly, inflammation were identified as being significantly modulated in the aged cochlea. GO and KEGG analyses of the upregulated DEGs revealed that the most enriched terms were associated with immune responses and inflammatory pathways, whereas many of the downregulated genes are involved in ion channel function and neuronal signaling. Real-time qPCR showed that H2O2 treatment significantly induced the expression of multiple inflammation and necroptosis-related genes in HEI-OC1 cells. CONCLUSION: Using next generation sequencing, our transcriptomic analysis revealed the differences of gene expression pattern with age in the cochlea of C57BL/6 mice. Our study also revealed multiple immune and inflammatory transcriptomic changes during cochlear aging and provides new insights into the molecular mechanisms underlying cochlear inflammation in AHL.

Yap-lin28a axis targets let7-Wnt pathway to restore progenitors for initiating regeneration.

The sox2 expressing (sox2+) progenitors in adult mammalian inner ear lose the capacity to regenerate while progenitors in the zebrafish lateral line are able to proliferate and regenerate damaged HCs throughout lifetime. To mimic the HC damage in mammals, we have established a zebrafish severe injury model to eliminate both progenitors and HCs. The atoh1a expressing (atoh1a+) HC precursors were the main population that survived post severe injury, and gained sox2 expression to initiate progenitor regeneration. In response to severe injury, yap was activated to upregulate lin28a transcription. Severe-injury-induced progenitor regeneration was disabled in lin28a or yap mutants. In contrary, overexpression of lin28a initiated the recovery of sox2+ progenitors. Mechanistically, microRNA let7 acted downstream of lin28a to activate Wnt pathway for promoting regeneration. Our findings that lin28a is necessary and sufficient to regenerate the exhausted sox2+ progenitors shed light on restoration of progenitors to initiate HC regeneration in mammals.

Nrf2 activation protects auditory hair cells from cisplatin-induced ototoxicity independent on mitochondrial ROS production.

Cisplatin is a well-known and commonly used chemotherapeutic agent. However, cisplatin-induced ototoxicity limits its clinical use. Previous studies have shown an important role of reactive oxygen species (ROS) accumulation in the pathogenesis of cisplatin-induced ototoxicity. In many cell types, the transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2) and antioxidant response element (ARE) protect against oxidative stress by suppressing ROS. Here our results showed that cisplatin injury reduced Nrf2 expression and inhibited Nrf2 translocation in HEI-OC1 cells and Nrf2 activator tert-butylhydroquinone (TBHQ) rescued hair cells from cisplatin induced apoptosis by suppressing the total cellular ROS accumulation. Moreover, we found that decreased ROS accumulation induced by TBHQ didn't depend on mitochondrial derived ROS production, indicating that Nrf2 activation alleviated cisplatin induced oxidative stress and apoptosis through mitochondrial-independent ROS production. Therefore, we provide a potential strategy of prevention and treatment for cisplatin-induced ototoxicity by Nrf2 activation. In conclusion, Nrf2 activation protects auditory hair cells from cisplatin-induced ototoxicity through suppressing the total cellular ROS levels which arise from sources other than mitochondria.

LncRNA AW112010 Promotes Mitochondrial Biogenesis and Hair Cell Survival: Implications for Age-Related Hearing Loss.

Long noncoding RNA (lncRNA) disorder has been found in many kinds of age-associated diseases. However, the role of lncRNA in the development of age-related hearing loss (AHL) is still largely unknown. This study sought to uncover AHL-associated lncRNAs and the function. RNA-sequencing was conducted to profile lncRNA expression in the cochlea of an early-onset AHL mouse model. RT-qPCR assay was used to validate the expression pattern of lncRNAs. ATP assay, JC-1 assay, mitochondrial probe staining, CCK-8 assay, Western blot, and immunocytochemistry were performed to detect the effects of lncRNA AW112010 in HEI-OC1 cells and the mouse cochlea. We identified 88 significantly upregulated lncRNAs and 46 significantly downregulated lncRNAs in the cochlea of aged C57BL/6 mice. We focused on the significantly upregulated AW112010. Silencing of AW112010 decreased the ATP level, mitochondrial membrane potential, and cell viability and increased mitochondrial ROS generation under oxidative stress in HEI-OC1 cells. AW112010 overexpression promoted cell survival in HEI-OC1 cells. AW112010 knockdown reduced mitochondrial mass and impaired mitochondrial biogenesis in HEI-OC1 cells. Activation of mitochondrial biogenesis by resveratrol and STR1720 promoted cell survival. The mitochondrial biogenesis process was activated in the cochlea of aged mice. Moreover, AW112010 regulated AMPK signaling in HEI-OC1 cells. Transcription factor Arid5b elevated in the aged cochlea and induced AW112010 expression and mitochondrial biogenesis in HEI-OC1 cells. Taken together, lncRNAs are dysregulated with aging in the cochlea of C57BL/6 mice. The Arid5b/AW112010 signaling was induced in the aged mouse cochlea and positively modulated the mitochondrial biogenesis to maintain mitochondrial function.

SIRT1 protects cochlear hair cell and delays age-related hearing loss via autophagy.

Age-related hearing loss (AHL) is typically caused by the irreversible death of hair cells (HCs). Autophagy is a constitutive pathway to strengthen cell survival under normal or stress condition. Our previous work suggested that impaired autophagy played an important role in the development of AHL in C57BL/6 mice, although the underlying mechanism of autophagy in AHL still needs to be investigated. SIRT1 as an important regulator involves in AHL and is also a regulator of autophagy. Thus, we hypothesized that the modulation between SIRT1 and autophagy contribute to HC death and the progressive hearing dysfunction in aging. In the auditory cell line HEI-OC1, SIRT1 modulated autophagosome induction because of SIRT1 deacetylating a core autophagy protein ATG9A. The deacetylation of ATG9A not only affects the autophagosome membrane formation but also acts as a sensor of endoplasmic reticulum (ER) stress inducing autophagy. Moreover, the silencing of SIRT1 facilitated cell death via autophagy inhibition, whereas SIRT1 and autophagy activation reversed the SIRT1 inhibition media cell death. Notably, resveratrol, the first natural agonist of SIRT1, altered the organ of Corti autophagy impairment of the 12-month-old C57BL/6 mice and delayed AHL. The activation of SIRT1 modulates the deacetylation status of ATG9A, which acts as a sensor of ER stress, providing a novel perspective in elucidating the link between ER stress and autophagy in aging. Because SIRT1 activation restores autophagy with reduced HC death and hearing loss, it could be used as a strategy to delay AHL.

Modulation of miR-34a/SIRT1 signaling protects cochlear hair cells against oxidative stress and delays age-related hearing loss through coordinated regulation of mitophagy and mitochondrial biogenesis.

Mitophagy and mitochondrial biogenesis are 2 pathways that regulate mitochondrial content and metabolism maintaining cellular homeostasis. The imbalance between these opposing processes impairs mitochondrial function and is suggested to be the pathophysiological basis of a variety of neurodegenerative diseases and aging. Here we investigated the role of mitophagy and mitochondrial biogenesis in oxidative damage to the cochlear hair cells and age-related hearing loss. In cultured mouse House Ear Institute-Organ of Corti 1 hair cells, oxidative stress activated mitophagy but inhibited mitochondrial biogenesis and impaired mitochondrial function. Pharmacological inhibition of miR-34a/SIRT1 signaling enhanced mitophagy, mitochondrial biogenesis, and attenuated House Ear Institute-Organ of Corti 1 cell death induced by oxidative stress. In the cochlea of C57BL/6 mice, mitophagy and mitochondrial biogenesis were both upregulated during aging. Long-term supplementation with resveratrol, a SIRT1 activator, not only improved the balance between mitophagy and mitochondrial biogenesis but also significantly reduced age-related cochlear hair cell loss, spiral ganglion neuron loss, stria vascularis atrophy, and hearing threshold shifts in C57BL/6 mice. Moreover, SIRT1 overexpression or miR-34a deficiency both attenuated age-related cochlear hair cell loss and hearing loss in C57BL/6 mice. Our findings reveal that imbalance between mitophagy and mitochondrial biogenesis contributes to cochlea hair cell damage caused by oxidative stress and during aging. Coordinated regulation of these 2 processes by miR-34a/SIRT1 signaling might serve as a promising approach for the treatment of age-related cochlear degeneration and hearing loss.

Inhibition of DRP-1-Dependent Mitophagy Promotes Cochlea Hair Cell Senescence and Exacerbates Age-Related Hearing Loss.

Background: Mitochondrial dysfunction is considered to contribute to the development of age-related hearing loss (AHL). The regulation of mitochondrial function requires mitochondrial quality control, which includes mitophagy and dynamics. Dynamin-related Protein 1 (DRP-1) is believed to play a central role in this regulation. However, the underlying mechanism of DRP-1 in AHL remains unclear. Here, we examined whether the decline of DRP-1-dependent mitophagy contributes to the development of AHL. Methods: We induced cellular and cochlear senescence using hydrogen peroxide (H2O2) and evaluated the level of senescence through senescence-associated ß-galactosidase staining. We evaluated mitophagy levels via fluorescence imaging and Western Blotting of LC3II and P62. Mitochondrial function was assessed by ATP assay, mtDNA assay, and JC-1. Results: We found that both the expression of DRP-1 and the mitophagy level decreased in senescent cells and aged mice. DRP-1 overexpression in HEI-OC1 cells initiated mitophagy and preserved mitochondrial function when exposed to H2O2, while cells with DRP-1 silencing displayed otherwise. Moreover, inhibition of DRP-1 by Mdivi-1 blocked mitophagy and exacerbated hearing loss in aged C57BL/6 mice. Conclusion: These results indicated that DRP-1 initiated mitophagy, eliminated mitochondrial dysfunction, and may protect against oxidative stress-induced senescence. These results provide a potential therapeutic target for AHL.

Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line.

Cisplatin-induced ototoxicity is one of the major adverse effects in cisplatin chemotherapy, and hearing protective approaches are unavailable in clinical practice. Recent work unveiled a critical role of autophagy in cell survival in various types of hearing loss. Since the excessive activation of autophagy can contribute to apoptotic cell death, whether the activation of autophagy increases or decreases the rate of cell death in CDDP ototoxicity is still being debated. In this study, we showed that CDDP induced activation of autophagy in the auditory cell HEI-OC1 at the early stage. We then used rapamycin, an autophagy activator, to increase the autophagy activity, and found that the cell death significantly decreased after CDDP injury. In contrast, treatment with the autophagy inhibitor 3-methyladenine (3-MA) significantly increased cell death. In accordance with in vitro results, rapamycin alleviated CDDP-induced death of hair cells in zebrafish lateral line and cochlear hair cells in mice. Notably, we found that CDDP-induced increase of Sirtuin 1 (SIRT1) in the HEI-OC1 cells modulated the autophagy function. The specific SIRT1 activator SRT1720 could successfully protect against CDDP-induced cell loss in HEI-OC1 cells, zebrafish lateral line, and mice cochlea. These findings suggest that SIRT1 and autophagy activation can be suggested as potential therapeutic strategies for the treatment of CDDP-induced ototoxicity.

Autophagy inhibition impairs the epithelial-mesenchymal transition and enhances cisplatin sensitivity in nasopharyngeal carcinoma.

Drug resistance restricts the efficacy of cisplatin in the treatment of nasopharyngeal carcinoma (NPC). Increasing evidence indicates that autophagy and the epithelial-mesenchymal transition (EMT) participate in cancer progression and drug sensitivity. The aim of the present study was to investigate the function of autophagy and EMT in cisplatin treatment, and to reveal the underlying impact of autophagy on the EMT process in NPC. Transmission electron microscopy assays and western blot analyses confirmed that cisplatin activates autophagy in NPC cells. Alterations in cell morphology and biomolecular markers confirmed that cisplatin induces the EMT phenotype in NPC cells. Cell viability assays showed that the combination of the autophagy inhibitor chloroquine (CQ) increased the cytotoxicity of cisplatin in NPC cells and that the EMT inducer transforming growth factor ß1 promoted the resistance to cisplatin in NPC cells. Moreover, autophagy inhibition by CQ and microtubule-associated protein 1 light chain 3B-knockdown reversed the EMT phenotype in NPC cells. In conclusion, autophagy and the EMT process promote cisplatin resistance in NPC cells, while the inhibition of autophagy impairs the EMT process.

miR-185-3p regulates the invasion and metastasis of nasopharyngeal carcinoma by targeting WNT2B in vitro.

MicroRNAs (miRs) have been recognised as important regulators of malignant behaviour in different types of human cancer, including nasopharyngeal carcinoma (NPC). A previous study by our group revealed that miR-185-3p regulates the radioresistance of NPC cells. The present study aimed to investigate the effect of miR-185-3p on NPC invasion and metastasis. Human NPC CNE-2 and 5-8F cell lines were transfected with a miR-185-3p mimic and miR-185-3p inhibitor, respectively, and their effects on the invasion and metastasis of these cells was assessed using a wound healing assay and Matrigel invasion assay. The target gene of miR-185-3p, Wnt family member 2B (WNT2B) was silenced in 5-8F cells using siRNA in order to investigate its function in NPC. Data from the present study demonstrated that the expression of miR-185-3p was the highest in 5-8F and lowest in CNE-2 cells out of a range of NPC cell lines. Following the transfection of miR-185-3p mimic into CNE-2 cells, the wound healing and Matrigel invasion assays indicated that the migration and invasion ability of CNE-2 cells was significantly reduced compared with the negative control group. In addition, the inhibition of miR-185-3p in 5-8F cells significantly increased the capacity for migration and invasion. Furthermore, silencing WNT2B expression resulted in a significant reduction in the invasion and metastasis in 5-8F cells. The inhibition of miR-185-3p, which promotes invasion and metastasis, could be reversed through the silencing of WNT2B in 5-8F cells. The results of the present study indicate that miR-185-3p mediates the invasion and metastasis of NPC by targeting WNT2B in vitro.

miR-324-3p suppresses migration and invasion by targeting WNT2B in nasopharyngeal carcinoma.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is a malignant epithelial carcinoma of the head and neck with strong ability of invasion and metastasis. Our previous study indicated that miR-324-3p, as a tumor-suppressive factor, could regulate radioresistance of NPC cells by targeting WNT2B. The purpose of this study is to investigate the role of miR-324-3p on migration and invasion in NPC cells. METHODS: Quantitative real time PCR was applied to measure the expression level of miR-324-3p and WNT2B mRNA in both cells and tissues, and the expression level of WNT2B protein was determined by western blotting. The capacity of migration and invasion were tested by using wound healing and transwell invasion assay. RESULTS: Ectopic expression of miR-324-3p or silencing its target gene WNT2B could dramatically suppress migration and invasion capacity of NPC cells. Meanwhile, the alterations of miR-324-3p in NPC cells could influence the expression level of the biomarkers of epithelial-mesenchymal transition (EMT), including E-cadherin and Vimentin. Moreover, the expression of miR-324-3p was obviously downregulated and WNT2B was significantly upregulated in NPC tissues. The expression levels of miR-324-3p and WNT2B were closely correlated with T stage, clinic stage and cervical lymph node metastasis of NPC (P < 0.05). CONCLUSION: miR-324-3p could suppress the migration and invasion of NPC by targeting WNT2B and the miR-324-3p/WNT2B pathway possibly provide new potential therapeutic clues for NPC.

Genome-wide analyses of long noncoding RNA expression profiles correlated with radioresistance in nasopharyngeal carcinoma via next-generation deep sequencing.

BACKGROUND: Radioresistance is one of the major factors limiting the therapeutic efficacy and prognosis of patients with nasopharyngeal carcinoma (NPC). Accumulating evidence has suggested that aberrant expression of long noncoding RNAs (lncRNAs) contributes to cancer progression. Therefore, here we identified lncRNAs associated with radioresistance in NPC. METHODS: The differential expression profiles of lncRNAs associated with NPC radioresistance were constructed by next-generation deep sequencing by comparing radioresistant NPC cells with their parental cells. LncRNA-related mRNAs were predicted and analyzed using bioinformatics algorithms compared with the mRNA profiles related to radioresistance obtained in our previous study. Several lncRNAs and associated mRNAs were validated in established NPC radioresistant cell models and NPC tissues. RESULTS: By comparison between radioresistant CNE-2-Rs and parental CNE-2 cells by next-generation deep sequencing, a total of 781 known lncRNAs and 2054 novel lncRNAs were annotated. The top five upregulated and downregulated known/novel lncRNAs were detected using quantitative real-time reverse transcription-polymerase chain reaction, and 7/10 known lncRNAs and 3/10 novel lncRNAs were demonstrated to have significant differential expression trends that were the same as those predicted by deep sequencing. From the prediction process, 13 pairs of lncRNAs and their associated genes were acquired, and the prediction trends of three pairs were validated in both radioresistant CNE-2-Rs and 6-10B-Rs cell lines, including lncRNA n373932 and SLITRK5, n409627 and PRSS12, and n386034 and RIMKLB. LncRNA n373932 and its related SLITRK5 showed dramatic expression changes in post-irradiation radioresistant cells and a negative expression correlation in NPC tissues (R = -0.595, p < 0.05). CONCLUSIONS: Our study provides an overview of the expression profiles of radioresistant lncRNAs and potentially related mRNAs, which will facilitate future investigations into the function of lncRNAs in NPC radioresistance.

Ionizing radiation promotes advanced malignant traits in nasopharyngeal carcinoma via activation of epithelial-mesenchymal transition and the cancer stem cell phenotype.

Post-irradiation residual mass and recurrence always suggest a worse prognosis for nasopharyngeal carcinoma (NPC). Our study aimed to investigate the malignant behaviors of post-irradiation residual NPC cells, to identify the potential underlying mechanisms and to search for appropriate bio-targets to overcome this malignancy. Two NPC cell lines were firstly exposed to 60 Gy irradiation, and residual cells were collected. In our previous study, colony formation assay detected the radioresistance of these cells. Here, the CCK-8 assay examined the cell sensitivity to paclitaxel and cisplatin. Wound-healing and Transwell assays were performed to investigate cell motility and invasion capabilities. Inverted phase-contrast microscopy was used to observe and photograph the morphology of cells. Expression levels of epithelial-mesenchymal transition (EMT)-related proteins were detected by western blot assay in NPC cells and tissues. The mRNA levels of cancer stem cell (CSC)-related genes were detected via qRT-PCR. The results revealed that residual NPC cells exhibited enhanced radioresistance and cross-resistance to paclitaxel and cisplatin. Higher capacities of invasion and migration were also observed. An elongated morphology with pseudopodia formation and broadening in the intercellular space was observed in the residual cells. Downregulation of E-cadherin and upregulation of vimentin were detected in the residual NPC cells and tissues. CSC-related Lgr5 and c-myc were significantly upregulated in the CNE-2-Rs and 6-10B-Rs radioresistance cells. Higher proportions of Lgr5+ cells were observed in radioresistant cells via immunofluorescent staining and flow cytometry. In conclusion, our study demonstrated that residual NPC cells had an advanced malignant transition and presented with both EMT and a CSC phenotype. This provides a possible clue and treatment strategy for advanced and residual NPC.

Metadherin regulation of vascular endothelial growth factor expression is dependent upon the PI3K/Akt pathway in squamous cell carcinoma of the head and neck.

Our previous study indicated overexpression of metadherin (MTDH) is an adverse prognostic factor in squamous cell carcinoma of the head and neck (SCCHN) and promotes SCCHN cell proliferation and invasion. However, its mechanism remains unclear. Recent studies have indicated that MTDH is a cancer-metastasis-associated molecule that participates in the process of angiogenesis. Therefore, the study is aimed to investigate that whether vascular endothelial growth factor (VEGF), as one of the most potent proangiogenic cytokines, is regulated by MTDH and the role of the phosphatidylinositide 3-kinases/Protein Kinase B (PI3K/Akt) pathway in this process of regulation and the clinical significance of both MTDH and VEGF in SCCHN.Immunohistochemistry was used to assay the expression of MTDH and VEGF in a cohort of 189 SCCHN patients with intact follow-up information. The expression of MTDH was then upregulated or inhibited by lentivirus-mediated MTDH Complementary deoxyribonucleic acid or MTDH short hairpin ribonucleic acid (shRNA) to observe the resulting alterations in VEGF expression and the PI3K/Akt signaling pathway in SCCHN cell lines. In addition, the PI3K/Akt pathway was modulated to observe the resulting changes in the MTDH-mediated expression of VEGF.The immunohistochemistry data showed that MTDH expression is positively correlated with VEGF expression in SCCHN tissues. Moreover, the overexpression of MTDH in SCCHN Tu686 and 5-8F cells led to increases in the expression of VEGF, and this effect was accompanied by activation of the PI3K/Akt pathway. Conversely, shRNA-mediated knockdown of MTDH led to decreased VEGF expression. In addition, inhibition of the Akt signaling pathway reversed the upregulation of VEGF resulting from MTDH overexpression. Moreover, the survival analysis revealed that VEGF is an independent prognostic factor, and a combined survival analysis based on both MTDH and VEGF showed synergistic effects in the prognosis evaluation of SCCHN patients.The findings of the present study demonstrate that MTDH regulates the expression of VEGF via the PI3K/Akt signaling pathway, indicating the potential role of the MTDH-mediated activation of VEGF signaling pathway in SCCHN angiogenesis and metastasis.

Increased expression of miR-93 is associated with poor prognosis in head and neck squamous cell carcinoma.

MicroRNA-93-5p (miR-93) is a novel oncogenic microRNA (miRNA) and is elevated in diverse human malignancies. Aberrant expression and dysfunction of miR-93 are involved in many types of human tumours. However, the exact role of miR-93 remains unclear in head and neck squamous cell carcinoma (HNSCC). The objective of this study is to determine the expression pattern and clinical significance of miR-93 in HNSCC. MiR-93 expression levels in 103 primary HNSCC tissues and 16 corresponding non-cancerous epithelia were analysed by miRNA in situ hybridisation and correlated with the clinicopathological parameters and patient outcomes. Moreover, the expression of miR-93 was examined in four HNSCC cell lines and 17 pairs of HNSCC tissues and their corresponding adjacent tissues using quantitative real-time PCR (qRT-PCR). The miR-93 levels in HNSCC tissues and cell lines were significantly higher than those in the non-cancerous tissues. Notably, high miR-93 expression was significantly associated with T classification, lymph node metastasis and clinical stage. Kaplan-Meier survival analysis demonstrated that patients with high miR-93 expression had poorer overall survival than patients with low miR-93 expression. Multivariate Cox regression analysis revealed that miR-93 overexpression and lymph node metastasis were independent prognostic factors in patients with HNSCC. This study demonstrated that miR-93 expression was significantly increased in HNSCC tissue samples and cell lines and that miR-93 overexpression was associated with tumour progression, metastasis and poor prognosis in HNSCC patients. These results suggest that miR-93 may play a critical role in the initiation and progression of HNSCC, indicating that miR-93 may be a valuable marker for the prediction of metastasis and prognosis in HNSCC.

Ephrin type­A receptor 2 regulates sensitivity to paclitaxel in nasopharyngeal carcinoma via the phosphoinositide 3­kinase/Akt signalling pathway.

Ephrin type­A receptor 2 (EphA2) is a receptor tyrosine kinase that is associated with cancer cell metastasis. There has been little investigation into its impact on the regulation of sensitivity to paclitaxel in nasopharyngeal carcinoma (NPC). In the present study, upregulation of EphA2 expression enhanced the survival of NPC 5­8F cells, compared with control cells exposed to the same concentrations of paclitaxel. Flow cytometry and western blot analysis demonstrated that over­expression of EphA2 decreased NPC cancer cell sensitivity to paclitaxel by regulating paclitaxel­mediated cell cycle progression but not apoptosis in vitro. This was accompanied by alterations in the expression of cyclin­dependent kinase inhibitors, p21 and p27, and of inactive phosphorylated­retinoblastoma protein. Furthermore, paclitaxel stimulation and EphA2 over­expression resulted in activation of the phosphoinositide 3­kinase (PI3K)/Akt signalling pathway in NPC cells. Inhibition of the PI3K/Akt signalling pathway restored sensitivity to paclitaxel in 5­8F cells over­expressing EphA2, which indicated that the PI3K/Akt pathway is involved in EphA2­mediated paclitaxel sensitivity. The current study demonstrated that EphA2 mediates sensitivity to paclitaxel via the regulation of the PI3K/Akt signalling pathway in NPC.

Analysis of clinical features and prognosis of malignant triton tumor: A report of two cases and literature review.

Malignant triton tumor (MTT) is a malignant peripheral nerve sheath tumor with rhabdomyoblastic differentiation. The prognosis of patients is poor, and due to its rarity, large case studies are lacking. The aim of this study is to describe the clinical features and identify potential prognostic factors. Two patients with MTT in the head and neck treated at our department are reported. A literature search revealed another 198 published cases. All of these cases then went through a retrospective analysis. The ratio of male-to-female incidence was 1.5:1, and the median age at diagnosis was 29 years. In 41.7% of cases it occurred in patients with neurofibromatosis type 1. The five-year survival of MTT was found to be just 35%. Cox proportional hazards analysis revealed that complete resection (hazard ratio, 0.396; P=0.032) and metastases (hazard ratio, 3.188; P=0.004) were associated with mortality, indicating that complete resection may lead to a longer life span, and that the existence of metastasis suggested a worse prognosis for patients with MTT.

Elevated expression of histone demethylase PHF8 associates with adverse prognosis in patients of laryngeal and hypopharyngeal squamous cell carcinoma.

AIM: Overexpression of histone demethylase PHF8 has been reported to function as an oncoprotein in many cancers; however, the implications of PHF8 involvement in laryngeal and hypopharyngeal squamous cell carcinoma (LHSCC) remain unclear. This study aims to explore the expression of PHF8 and its clinical significance in LHSCC. MATERIALS & METHODS: Western blotting and immunohistochemistry were performed to evaluate PHF8 protein expression in fresh and archived LHSCC samples. Global expressions of H3K27 and H3K9 methylation were analyzed in a cell line with PHF8 siRNA treatment. RESULTS & CONCLUSION: In our study, PHF8 was upregulated in fresh LHSCC tissues. Immunohistochemical staining revealed that the expression of PHF8 was positively associated with T classification, clinical stage, primary tumor position and tumor relapse. Survival analysis demonstrated that high PHF8 expression was significantly associated with shorter overall survival and disease-free survival. Moreover, PHF8 regulates the levels of H3K9me2 and H3K27me2 in LHSCC. Taken together, PHF8 might be a novel prognostic marker for this disease.