[Clinical classification, staging and treatment strategy of radionecrosis of the nasopharynx and skull base].

Objective:To establish a staging system for guiding clinical treatment and prognostic risk assessment by retrospectively analyzing the cases with radionecrosis of the nasopharynx and skull base （RNSB） after radiotherapy for nasopharyngeal carcinoma. Methods:A total of 86 cases of RNSB from January 2019 to December 2022 visited Department of Otorhinolaryngology Head and Neck, the People's Hospital of Guangxi Zhuang Autonomous Region. Seventeen patients gave up the treatment, and 69 patients who underwent treatment were included for analysis. By analyzing the results of electronic nasopharyngolaryngoscopy combined with magnetic resonance （MR）, CT, and other imaging examinations, a staging system for RNSB was proposed. The relationship between the staging system and the surgical effectiveness and clinical prognosis was further analyzed. Results:According to the severity and extent of destruction of soft tissue, bone, and the adjacent neurovascular structures, the RNSB was categorized into closed type （n=5） and open type （n=64）, of which the open type was subdivided into five types: type Ⅰ（n=4）, type Ⅱ（n=6）, type Ⅲ（n=39, of which 21 cases were type Ⅲa and 18 cases were type Ⅲb）, type Ⅳ（n=12）, and type Ⅴ（n=8）. The clinical stage of RNSB were classified based on nasopharyngolaryngoscopy and imaging examinations, receiving the second course of radiotherapy or not, the involvement of the infection site, the extent of bone destruction, the degree of internal carotid artery involvement, and the degree of brain tissue necrosis: stageⅠ（1-2 scores）, 11 cases at stageⅡ（3-4 scores）, 24 cases at stage Ⅲ（5-6 scores）, and 30 cases at stage Ⅳ（ ≥ 7 scores or more）. Twenty-two patients chose conservative treatment （2 patients at stage Ⅰ, 3 patients at stage Ⅱ, 7 patients at stage Ⅲ, and 10 patients at stage Ⅳ）. Forty-seven patients chose nasal endoscopic surgical treatment （2 patients at stage Ⅰ, 8 patients at stage Ⅱ, 17 patients at stage Ⅲ, and 20 patients at stage Ⅳ）, of which 16 cases had received free mucosal flap and/or stented septum mucosal flap repair. Patients at stages Ⅰ, Ⅱ, and Ⅲ achieved satisfactory efficacy after surgical treatment. In addition, higher clinical stage was found to correlate with the worse prognosis and higher incidence of perioperative complications, which included failure of healing because of surgical site infection, cerebrospinal fluid nasal leakage, progressive osteonecrosis, nasopharyngeal hemorrhage, and death. Conclusion:The staging system proposed in our study can be used for early detection of RNSB during regular follow-up, and is also valuable for clinical treatment guidance and prognosis assessment.

[Diagnosis of nasopharyngeal carcinoma with convolutional neural network on narrowband imaging].

Objective:To evaluate the diagnostic accuracy of the convolutional neural network（CNN） in diagnosing nasopharyngeal carcinoma using endoscopic narrowband imaging. Methods:A total of 834 cases with nasopharyngeal lesions were collected from the People's Hospital of Guangxi Zhuang Autonomous Region between 2014 and 2016. We trained the DenseNet201 model to classify the endoscopic images, evaluated its performance using the test dataset, and compared the results with those of two independent endoscopic experts. Results:The area under the ROC curve of the CNN in diagnosing nasopharyngeal carcinoma was 0.98. The sensitivity and specificity of the CNN were 91.90% and 94.69%, respectively. The sensitivity of the two expert-based assessment was 92.08% and 91.06%, respectively, and the specificity was 95.58% and 92.79%, respectively. There was no significant difference between the diagnostic accuracy of CNN and the expert-based assessment （P=0.282, P=0.085）. Moreover, there was no significant difference in the accuracy in discriminating early-stage and late-stage nasopharyngeal carcinoma（P=0.382）. The CNN model could rapidly distinguish nasopharyngeal carcinoma from benign lesions, with an image recognition time of 0.1 s/piece. Conclusion:The CNN model can quickly distinguish nasopharyngeal carcinoma from benign nasopharyngeal lesions, which can aid endoscopists in diagnosing nasopharyngeal lesions and reduce the rate of nasopharyngeal biopsy.

Epithelial-mesenchymal transition classification of circulating tumor cells predicts clinical outcomes in progressive nasopharyngeal carcinoma.

Background: Liquid biopsy facilitates the enrichment and isolation of circulating tumor cells (CTCs) in various human cancers, including nasopharyngeal carcinoma (NPC). Characterizing CTCs allows observation of the evolutionary process of single tumor cells undergoing blood-borne dissemination, such as epithelial-mesenchymal transition. However, the prognostic value of phenotypic classification of CTCs in predicting the clinical outcomes of NPC remains poorly understood. Patients and methods: A total of 92 patients who met the inclusion criteria were enrolled in the present study. The CanPatrol™ CTC technology platform was employed to isolate CTCs, and an RNA in situ hybridization-based system was used for phenotypic classification. Kaplan-Meier survival curves were used for univariate survival analysis, and the log-rank test was performed for between-group comparisons of the survival curves. Results: CTCs were detected in 88.0% (81/92) of the enrolled patients with NPC. The total CTC number did not vary between the T and N stages or between Epstein-Barr virus DNA-positive and -negative cases. The numbers of total CTCs and epithelial/mesenchymal (E/M) hybrid CTCs decreased significantly at 3 months post concurrent chemoradiotherapy (P=0.008 and P=0.023, respectively), whereas the numbers of epithelial or mesenchymal CTCs did not decrease. E/M hybrid-predominant cases had lower disease-free survival (P=0.043) and distant metastasis-free survival (P=0.046) rates than non-E/M hybrid-predominant cases. Conclusion: CTC classification enables a better understanding of the cellular phenotypic alterations responsible for locoregional invasion and distant metastasis in NPC. E/M hybrid-predominant CTC distribution predicts unfavorable clinical outcomes in patients with progressive NPC.

Bleomycin A5 suppresses Drp1­mediated mitochondrial fission and induces apoptosis in human nasal polyp­derived fibroblasts.

Intralesional injection of bleomycin­A5 (BLE­A5) is a novel treatment for nasal polyps. Our previous study clarified that BLE­A5 could induce nasal polyp­derived fibroblast (NPDF) apoptosis in nasal polyps. However, the detailed mechanisms are still unclear. The present study aimed to determine the effects of BLE­A5 on NPDF mitochondrial dynamics and provide a theoretical basis for the local application of BLE­A5 to treat nasal polyps. In the present study, an in vitro nasal polyp tissue culture model was used to define the BLE­A5 target cell type in nasal polyps. NPDF primary cell culture was used to study the effects of BLE­A5 on the mitochondrial dynamic­related mechanism. The results showed that BLE­A5 treatment of NPDFs caused mitochondrial­mediated apoptosis. Dynamin­related protein 1 (Drp1) was shown to be altered in BLE­A5­treated NPDFs. Drp1 knockdown increased the sensitivity of NPDFs to BLE­A5 and exacerbated mitochondrial dysfunction. BLE­A5 decreased cyclin B1­CDK1 complex­mediated phosphorylation of Drp1 and inhibited Drp1­mediated mitophagy in NPDFs. Overall, the present study concluded that BLE­A5 mainly induces NPDF apoptosis in nasal polyps. BLE­A5 regulates the mitochondria by inhibiting Drp1 activation, resulting in NPDF mitochondrial dynamic disorder and apoptosis.

Reactive Oxygen Species Are Necessary for Bleomycin A5-Induced Apoptosis and Extracellular Matrix Elimination of Nasal Polyp-Derived Fibroblasts.

BACKGROUND:: The pathology of chronic rhinosinusitis with nasal polyp (CRSwNP) is characterized by the infiltration of a large number of fibroblasts, resulting in extracellular matrix (ECM) deposition. Intralesional bleomycin A5 (BLE) injection has proved to be effective and safe, providing a novel treatment for CRSwNP. However, the mechanism is not clearly understood. OBJECTIVES:: The aim of this study is to explore the possible mechanism of BLE-induced apoptosis in nasal polyp-derived fibroblasts (NPDFs). MATERIAL AND METHODS:: Dichloro-dihydro-fluorescein diacetate probe, cell migration assays, and cell cycle analysis were used to detect the growth characteristics and basal reactive oxygen species (ROS) traits of NPDFs. Annexin V/propidium iodide and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay was used to detect BLE-induced apoptosis. As a control, the antioxidant glutathione (GSH) was used to abrogate ROS induced by BLE. Western blot analysis was used to evaluate the effects of BLE on apoptosis and the ECM proteins of NPDFs. RESULTS:: The results showed that NPDFs had more active growth characteristics and higher basal ROS levels than normal nasal mucosa fibroblasts (NMFCs). NPDFs were more sensitive to BLE-induced apoptosis and ROS accumulation. GSH abrogation inhibits BLE-induced ECM degradation and apoptosis in NPDFs through a mitochondrial-mediated pathway. CONCLUSIONS:: BLE induced NPDF apoptosis and ECM degradation through a mitochondrial-mediated pathway and in a ROS-dependent manner.

Interleukin-4-induced posttranscriptional gene regulation of CCL26 by the RNA-binding protein HuR in primary human nasal polyp-derived epithelial cells.

BACKGROUND: Much attention on the pathophysiology of nasal polyp (NP) has focused on eosinophils. Interleukin (IL)-4 and eotaxin-3 (C-C motif chemokine ligand 26, or CCL26) levels have been reported to be increased in eosinophilic nasal polyps. The aim of this study was to characterize CCL26 posttranscriptional regulation by the RNA-binding protein HuR in primary human nasal polyp-derived epithelial cells (hNPDECs) challenged with IL-4. METHODS: A prospective, observational study was conducted. Nasal polyp tissues were obtained from eosinophilic (n = 12) and non-eosinophilic (n = 10) NP patients, and inferior turbinate (IT) tissues were taken from control subjects (n = 9) and cultured into hNPDECs. Expression of HuR and CCL26 were measured by immunohistochemistry, Western blot analysis, enzyme-linked immunoassay, and real-time polymerase chain reaction (PCR). The nucleocytoplasmic shuttling of HuR in hNPDECs was detected by immunofluorescence. Posttranscriptional regulation of CCL26 by HuR was tested by ribonucleoprotein immunoprecipitation assay (RIP) and dual-luciferase reporter assay. CCL26 mRNA stabilization was measured by quatititative PCR after treatment with actinomycin D. Student's t test and one-way analysis of variance were used. RESULTS: Immunohistochemical data show that both HuR and CCL26 were highly expressed in NP tissues, especially eosinophilic NP tissues (p < 0.05). IL-4 stimulation increased CCL26 mRNA stability, and overexpression and knockdown of HuR affected CCL26 expression. Immunofluorescence data indicate that IL-4 altered the subcellular distribution of HuR. The RIP and dual-luciferase reporter assay results supply strong evidence for HuR binding to CCL26. CONCLUSION: Our results provide strong support for the hypothesis that IL-4-induced expression of CCL26 in hNPDECs relies partly on CCL26 mRNA stabilization mediated by the interaction of HuR with CCL26 3'UTR.

Intralesional bleomycin A5 injection for the treatment of nasal polyps through inducing apoptosis.

CONCLUSION: An intralesional bleomycin A5 (BLE) injection might be used as an alternative therapy for eosinophilic-type nasal polyps (NPs). BLE-induced apoptosis might play an important role in shrinkage of NPs. OBJECTIVES: The aim of this study is to determine the effect and explore the possible role of apoptosis in shrinkage of NPs. METHODS: Twenty-nine patients with eosinophilic-type NPs experienced repeated local injection of BLE. The recurrence rate of this group was obtained. The mechanism of BLE treatment was investigated through an in vitro experiment. Nasal polyp tissues were treated with BLE. The apoptotic activity was detected by the presence of DNA smear and test of terminal deoxynucleotidyl transferase dUTP nick end labeling. The caspase-8 and PARP were examined through immunohistochemistry and Western blotting. RESULTS: After several local injections of BLE, the nasal polyp tissues decreased and then disappeared. During follow-up of 3 years the recurrence rate of this group was significantly lower than another one treated with operation plus medicine treatment. Apoptosis in BLE-treated tissue was prominently detected in the infiltrating inflammatory cells. The expression of PARP and casp-8 were increased in BLE-treated nasal polyp tissue compared with PBS-treated tissue.

Induction of apoptosis in nasal polyp-derived fibroblasts by bleomycin A5 in vitro.

The present study aimed to evaluate the pro-apoptotic effects of bleomycin A5 on nasal polyp­derived fibroblasts (NPDFs) and the underlying molecular mechanisms. Nasal polyp tissue was acquired from 10 patients during surgery and NPDFs were isolated from surgical tissues. Fibroblasts were identified using immunohistochemistry. Bleomycin A5 was used to treat NPDFs along a concentration gradient. Cell viability was evaluated using a Cell Counting Kit­8 assay. A flow cytometric Annexin V­fluorescein isothiocyanate/propidium iodide assay was used to determine the percentage of apoptotic NPDFs. The mRNA expression levels of apoptotic genes were determined by reverse transcription­quantitative polymerase chain reaction and levels of proteins associated with apoptosis were determined by western blotting. The results indicated that bleomycin A5 was able to induce apoptosis in NPDFs in a dose­dependent manner. NPDFs treated with bleomycin A5 were identified to contain significantly high amounts of the active forms of caspase­3 and showed considerable cleavage of poly(ADP­ribose) polymerase. The mRNA and protein expression levels of the pro­apoptotic molecule Bcl­2­associated X protein were significantly higher in treated NPDFs than in untreated NPDFs. In contrast, the mRNA and protein expression of the anti­apoptotic molecule B­cell lymphoma 2 (Bcl­2) was significantly lower in treated NPDFs. These results indicated that bleomycin A5 could induce apoptosis in primary NPDFs through activation of the Bcl­2 family and caspase cascades in a time-, and concentration-dependent manner.