Nasopharyngeal carcinoma (NPC) is a serious threat to public health, and the biomarker discovery is of urgent needs. The data-independent mode (DIA) based sequential window acquisition of all theoretical fragment-ion spectra (SWATH) mass spectrometry (MS) has been proved to be precise in protein quantitation and efficient for cancer biomarker researches. In this study, we performed the first SWATH-MS analysis comparing the NPC and normal tissues. Spike-in stable isotope labeling by amino acids in cell culture (super-SILAC) MS was used as a shotgun reference. We identified and quantified 1414 proteins across all SWATH-MS analyses. We found that SWATH-MS had a unique feature to preferentially detect proteins with smaller molecular weights than either super-SILAC MS or human proteome background. With SWATH-MS, 29 significant differentially express proteins (DEPs) were identified. Among them, carbonic anhydrase 2 (CA2) was selected for further validation per novelty, MS quality and other supporting rationale. With the tissue microarray analysis, we found that CA2 had an AUC of 0.94 in differentiating NPC from normal tissue samples. In conclusion, SWATH-MS has unique features in proteome analysis, and it leads to the identification of CA2 as a potentially new diagnostic biomarker for NPC.

Objective: An abnormal state of the central auditory system (CAS) likely plays a large role in the occurrence of phantom sound of tinnitus. Various tinnitus studies using resting-state functional MRI (RS-fMRI) have reported aberrant spontaneous brain activity in the non-auditory system and altered functional connectivity between the CAS and non-auditory system. This study aimed to investigate abnormal functional connections between the aberrant spontaneous activity in the CAS and the whole brain in tinnitus patients, compared to healthy controls (HC) using RS-fMRI. Materials and Methods: RS-fMRI from 16 right-ear tinnitus patients with normal hearing (TNHs) and 15 HC individuals was collected, and the time series were extracted from different clusters of a CAS template, supplied by the Anatomy Toolbox of the Statistical Parametric Mapping software. These data were used to derive the smoothed mean amplitude of low-frequency fluctuation (smALFF) values and calculate the relationship between such values and the corresponding clinical data. In addition, clusters in the CAS identified by the smALFF maps were set as seed regions for calculating and comparing the brain-wide connectivity between TNH and HC. Results: We identified the different clusters located in the left higher auditory cortex (HAC) and the right inferior colliculus (IC) from the smALFF maps that contained increased (HAC) and decreased (IC) activity when the TNH group was compared to the HC group, respectively. The value of increased smALFF cluster in the HAC was positively correlated with the tinnitus score, but the decreased smALFF cluster in the IC was not correlated with any clinical characters of tinnitus. The TNH group displayed increased connectivity, compared to the HC group, in brain regions that encompassed the left IC, bilateral Heschl gyrus, bilateral supplementary motor area, right insula, bilateral superior temporal gyrus, right middle temporal gyrus, left hippocampus, left amygdala, and right supramarginal gyrus. Conclusion: Tinnitus may be linked to abnormal spontaneous activity in the HAC, which can arise from the neural plasticity induced from the increased functional connectivity between the auditory network, cerebellum, and limbic system.

Background: Chronic suppurative otitis media (CSOM) is a common otolaryngology disease, and cholesteatoma is the most aggressive type of CSOM. CSOM with and without cholesteatoma both result in a certain level of sensorineural damage, which can be categorized as air conduction (AC), bone conduction (BC), or air-bone gap (ABG), and AC, BC, and ABG are affected by many factors. Further analyses and comparisons of factors affecting sensorineural damage in CSOM with and without cholesteatoma were conducted in this study. Methods: A comparative study was conducted of 79 patients with CSOM (39 with cholesteatoma and 40 without cholesteatoma) whose diagnoses were mainly based on chronic middle ear infections and hearing loss (HL), typical computed tomography (CT) and surgical findings. Audiological evaluation included AC, BC and ABG, and sensorineural damage was defined as mixed and sensorineural HL (SNHL). Results: In relation to the types of HL, there were no significant differences between both groups. The CSOM with cholesteatoma group had significantly greater AC (P=0.000) and a significantly greater ABG (P>0.05) than the CSOM without cholesteatoma group, but BC did not differ significantly between both groups (P>0.05). The average AC-middle frequency (MF), AC-high frequency (HF), ABG-MF and ABG-HF of CSOM without cholesteatoma were smaller than these of CSOM with cholesteatoma (P<0.05). The degree of HL differed significantly between both groups (P=0.000). The CSOM with cholesteatoma group showed a higher level of HL than the CSOM without cholesteatoma group. The presence of cholesteatoma was presented a protective factor associated with sensorineural damage (P<0.05), while higher degrees of hearing and aging were risk factors (P<0.05), respectively. Conclusions: Our direct comparisons showed that HL progressed more rapidly in the CSOM with cholesteatoma group, which had higher frequencies in relation to AC, the ABG, and severity. However, in relation to BC, there were no significant differences between both groups, which was in line with the similar proportions of the types of HL in both groups. The logistic regression showed that the presence of cholesteatoma was a protective factor, and the degree of hearing and aging were risk factors associated with sensorineural damage.

Nasopharyngeal carcinoma (NPC) is a head and neck tumor with high degree of malignancy and with high incidence especially in southern China. AIMP2-DX2, one isoform of the aminoacyl-tRNA synthetase interacting multifunctional proteins (AIMPs), is shown to be a potential target in many cancers. However, the detailed mechanisms of AIMP2-DX2 in NPC development remain to be elucidated. Here, we found that the mRNA expression level of AIMP2-DX2 was significantly increased in NPC specimens, compared with normal nasopharyngeal tissues. Microarray immunohistochemical analysis of NPC specimens and Kaplan–Meier analysis showed that patients with high AIMP2-DX2 protein expression had shorter overall survival than those with low AIMP2-DX2 level. Furthermore, mRNA and protein expression levels of AIMP2-DX2 were both increased in cultured NPC cell lines (5-8F, CNE-2Z, and CNE-1), by being compared with normal nasopharyngeal cell line NP69. Overexpression of AIMP2-DX2 remarkably promoted the cell viability, cell migration, and invasion of cultured NPC cells. Genetic knockdown of AIMP2-DX2 by shRNA lentiviruses significantly suppressed the proliferation, migration, and invasion and induced apoptosis of NPC cells. Inhibition of AIMP2-DX2 decreased the highly expressed level of matrix metalloproteinase- (MMP-) 2 and MMP-9, further suppressed proliferation, migration, and invasion in cultured NPC cells in vitro, and inhibited tumor growth in a xenograft mouse model in vivo. Taken together, these results suggest that AIMP2-DX2 plays an important role in the regulation of NPC and could be a potential therapeutic target and prognostic indicator for the treatment of NPC.

Background: We sought to determine the differences with respect to the proteome of nasopharyngeal tissues between patients with nasopharyngeal carcinoma (NPC) and healthy controls by using sequential windowed acquisition of all theoretical fragment ion mass spectra (SWATH^TM-MS) and ingenuity pathway analysis (IPA). Our primary purpose was to identify specific protein markers that can be applied for diagnosis or treatment of NPC. Methods: The CNE-1, CNE-2 and H1299 cell lines were cultured in stable isotope labeling of amino acids in cell culture (SILAC) medium for 10 generations to obtain labeled proteins. Thirty samples of NPC and 30 healthy control nasopharyngeal tissues were collected from the Department of Otolaryngology of the First Affiliated Hospital of Jinan University. Proteome of the nasopharyngeal tissues were analyzed and compared by SWATH-MS to identify differently expressed proteins. Further, extraction of target proteins and biological pathways was performed by IPA. Super-SILAC technique and liquid chromatography-tandem mass spectrometry were used to verify the reliability of the data obtained using SWATH-MS. Results: We identified 1,415 differentially expressed proteins between NPC patients and healthy controls. On IPA analysis, EIF2AK2 and MAPK1 proteins were found to be enriched in multiple biological pathways and functional networks. Conclusions: The differentially expressed proteins EIF2AK2 and MAPK seem to play an important role in the biological network of NPC or may help discover the specific functional proteins of NPC. Further studies are required to identify the pathways and molecular mechanisms that underlie NPC.