Efficacy of balloon Eustachian tuboplasty plus tympanostomy tube insertion in postirradiation otitis media with effusion.

OBJECTIVE: This study aimed to compare the efficacy of balloon Eustachian tuboplasty (BET) plus tympanostomy tube insertion (TTI) and simple TTI for postirradiation otitis media with effusion (OME) in patients with nasopharyngeal carcinoma. METHOD: This study included 36 patients (51 ears) with OME after the first radiotherapy course for nasopharyngeal carcinoma and categorized them into the BET + TTI and simple TTI groups. Effective rates, pure tone hearing threshold, Eustachian tube function score, and complication incidences were compared. RESULTS: The effective rates of the BET+TTI and TTI groups were 93.75 % and 75 %, respectively, with no statistically significant difference (P = 0.29). The pure tone hearing threshold examination at 9 months postoperatively revealed significantly lower mean air-pure tone and air-bone gap in both the BET + TTI and TTI groups than preoperatively. Eustachian Tube Dysfunction Questionnaire-7 (ETDQ-7) scores at every postoperative visit were significantly higher than preoperative scores in the two groups (all P < 0.05); ETDQ-7 score reduction in the BET + TTI group at 3, 9, and 12 months postoperatively was significantly higher than that in the TTI group. Otorrhea and recurrence both occurred in the BET+TTI and TTI groups, but the BET+TTI group demonstrated a lower incidence. CONCLUSION: BET + TTI is an effective treatment method for postirradiation OME.

Tumor-derived exosomal microRNA-15b-5p augments laryngeal cancer by targeting TXNIP.

Tumor-derived exosomes (EXO) are information carriers of microRNA (miR) in cancer development. Here, we explored the synergism of tumor-derived EXO and miR-15b-5p in laryngeal cancer (LCa). miR-15b-5p and thioredoxin-interacting protein (TXNIP) levels were firstly measured in clinical LCa tissues. The association between miR-15b-5p and TXNIP was determined. miR-15b-5p mimic was transfected into HEP-2 cells, and the corresponding exosomes were extracted. miR-15b-5p mimic-modified EXO were co-cultured with HEP-2 cells, and TXNIP low expression/high expression vector was transfected into HEP-2 cells Finally, cell growth was observed in vitro and in vivo. miR-15b-5p level was high while TXNIP level was low in LCa, and miR-15b-5p negatively modulated TXNIP expression. HEP-2 cells-derived EXO or inhibition of TXNIP enhanced HEP-2 cell growth in vitro and in vivo. Up-regulated miR-15b-5p further strengthened the pro-tumor effect of EXO, but this effect was reversed by overexpression of TXNIP. Overall, tumor-derived exosomal miR-15b-5p augments LCa through targeting down-regulation of TXNIP.

Analysis of Prognostic Alternative Splicing Reveals the Landscape of Immune Microenvironment in Thyroid Cancer.

BACKGROUND: The incidence of thyroid cancer (THCA) continues to increase in recent decades. Accumulating evidence showed that the unbalanced alternative splicing (AS) promotes the occurrence of cancers and leads to poor prognosis of patients. However, the research on alternative splicing events in THCA is lacking, and its underlying mechanism is not fully understood. This study identifies a novel prognostic signature based on AS events to reveal the relationship of AS with tumor immune microenvironment. METHODS: Based on the AS data, transcriptional data, and clinical information, the differentially expressed alternative splicings (DEASs) were screened out. Least absolute shrinkage and selection operator (LASSO) regression and multi-Cox regression analyses were employed to identify prognostic results related to AS events and establish a prognostic signature. The predictive ability of the signature was assessed by Kaplan-Meier (K-M) survival curve, risk plots, and receiver operating characteristic (ROC) curves. Furthermore, correlations between tumor-infiltrating immune cells, immune checkpoints, immune score and prognostic signature were analyzed. RESULTS: According to the LASSO regression analysis, a total of five AS events were selected to construct the signature. K-M survival curve showed that the higher the risk score, the worse the OS of the patients. Risk plots further confirmed this result. ROC curves indicated the high predictive efficiency of the prognostic signature. As for tumor immune microenvironment, patients in the high-risk group had a higher proportion of immune cells, including plasma cell, CD8+ T cell, macrophages (M0 and M2), and activated dendritic cell. Immune checkpoint proteins, such as PDCD1LG2, HAVCR2, CD274, etc., were significantly higher in the high-risk group. We also found that the ESTIMATE score, stromal score, and immune score were lower in the high-risk group, while the result of tumor purity was the opposite. CONCLUSIONS: Collectively, a prognostic signature consisting of five AS events in THCA was established. Furthermore, there was an inextricable correlation between immune cell infiltration, immune checkpoint proteins, and AS events. This study will provide a basis for THCA immunotherapy in the future.

Knockdown of Circular RNA hsa_circ_PVT1 Inhibited Laryngeal Cancer Progression via Preventing wnt4/ß-Catenin Signaling Pathway Activation.

AIM: To explore the function and mechanism of circular has_circ_PVT1 on laryngeal cancer (LC). METHODS: Microarray chip was performed to screen the differential expression of circRNA. Western blot and qRT-PCR was employed to detect the protein and mRNA level. CCK-8, clone formation, cell cycle, wound healing, and Transwell assay were performed to detect the cell proliferation, migration, and invasion ability. Luciferase assay and Fish were used to confirm the relationship between circ_PVT1/CBX4 and miR-21-5p. Flow cytometry and TUNEL assay were carried out to assess the apoptosis level. RESULTS: The upregulation of circ_PVT1 was found in LC tissues and cells. Silencing of circ_PVT1 inhibited LC progression via targeting miR-21-5p and indirectly controlling CBX4. Wnt4/ß-catenin signal pathway was inactivated by inhibiting the expression of circ_PVT1. CONCLUSION: Knockdown of circ_PVT1 prevented LC progression via targeting miR-21-5p/CBX4 by inhibiting wnt4/ß-catenin signal pathway, which could provide a novel therapeutic target for LC.

[The study of siRNA interference after laryngeal cancer Hep-2 cells to cisplatin sensitivity of ß-catenin gene expression].

OBJECTIVE: To investigate the changes of laryngeal cancer Hep-2 cells to cisplatin chemosensitivity after the interference of siRNA of ß-catenin gene expression. METHOD: Using a small interference RNA (siRNA) technology interfere ß-catenin gene of Hep-2 cells . The mRNA and protein levels of ß-catenin in the Hep-2 cells of different groups were detected by qPCR and Western blot. It was divided into siRNA-ß-catenin-Hep-2 siRNA group, ß-catenin-Neg negative control group and blank control group. Cell proliferation inhibition rate of different concentrations of cisplatin on three groups was detected by MTT assay. Calculate the 50% inhibitory effective concentration IC50 value. Check the change of three groups of cells' apoptosis rate by flow cytometry after the same concentrations of cisplatin stimulation. RESULT: ß-catenin-siRNA interference fragment can specifically reduce the expression levels of ß-catenin mRNA and protein. qPCR illustrated the expression of mRNA in ß-catenin-siR-NA-Hep-2 interference group decreased 70% (P < 0.05) compared with the control group, Western blot results showed that the ß-catenin protein expression of interference group (0. 545 ± 0.111) decreased significantly compared with blank control group (1.507 ± 0.139) and negative control group (1.429 ± 0.089), P < 0.05. The IC50 calculation software showed that IC50 of cisplatin on ß-catenin-siRNA IC50 interference group is (5.81 ± 0.46)µg/ml, the blank control group is (10.10 ± 1.01) µg/ml, the difference between the two groups has statistical signifi- cance (P < 0.01). Cell apoptosis rate of ß-catenin-siRNA interference group was (26.15 ± 0.60)%, significantly higher than the control group (14.16 ± 0.05)%, P < 0.05. CONCLUSION: To interfere the expression of ß-catenin can effectively enhance the sensitivity of laryngeal cancer cells to chemotherapeutic drugs cisplatin. It provides a theoretical support for the reduction of laryngeal cancer chemotherapy drug cisplatin dosage.

[Exploration of the role of cisplatin on transformation of larvngeal tumor cells to stem-like cancer cells].

OBJECTIVE: To explore the possibility mechanism of non-side population cells (NSP) of Hep-2 be induced into stem-like cancer cells by chemotherapy drug--cisplatin. METHOD: Hep-2 cell lines were sorted by fluorescence-actived cell sorting. The acquired NSP cells in trail group were co-cultured with cisplatin for more than 48 hours,while the control group with normal saline(NS). Then identified the percentage of the side population (SP) cells by flow cytometer. The ß-catenin, notch-1 mRNA in trial and control group were detected using quantitative realtime PCR, and the ß-catenin, notch-1 protein in two groups were compared by Western blot. RESULT: The percentage of side population cells in two groups were (17.16 ± 0.18)%, (10.05 ± 1.20)%, respectively. There was significant difference between two groups (t = 5.844, P < 0.01). The expression of ß-catenin, notch-1 was higher in trail group by qRT-PCR; the protein levels of ß- catenin, notch-1 was found to inceased in the trail group by Western blot (t = 5.155, P = 0.031; t = 5.977, P = 0.004). Statistical analysis showed significant difference between two groups (P < 0.05). CONCLUSION: NSP cells can be differentiated into stem-like cancer cells after being treating with cisplatin. The supposed mechanism is maybe through wnt/ß-catenin, notch signaling transduction pathway abnormalities.

[The optimizing conditions in sorting of side population in Hep-2].

OBJECTIVE: To investigate the optimizing conditions in isolation of the side population in laryngeal carcinoma cell line Hep-2. METHOD: Single-cell suspension cells were detached from the culture flask with trypsin EDTA, at a concentration of 1 x 10(6) cells/ml. (1) The trail Samples were incubated with Hoechst33342 at a concentration of 5 microg/ml, 9 microg/ml, 10 microg/ml, 11 microg/ml for 90 minutes. (2) They were incubated with Hoechst for 50, 70, 90, 110, 130 min in water bath individually. (3) The single-cell suspension were incubated Hoechst in water bath and in thermostat each. (4) The two different density of cells were harvested, which were 100% and 70%, and then di gest into single-cell suspension. Once incubation finished, suspended in phosphate buffered saline (PBS), then test SP% by flow cytometry. Among all groups,Verapamil hydrochloride was added to the control samples, incubated at 37 degrees C for 30 minutes, the other condition were keep the same with their trial groups. RESULT: (1) The percentage of Hoechst-negative cells in trial group was (39.96 +/- 0.24)%, (26.23 +/- 0.39)%. (18.79 +/- 0.02)%, (19.01 +/- 0.14)% at the concentration of 5 microg/ml, 9 microg/ml, 10 microg/ml, 11 microg/ml respectively, when the PI-positive cells were (30.45 +/- 0.63)%, (49.9 +/- 0.42)%, (50.12 +/- 0.68)%, (64.16 +/- 0.39)% separately. (2) Varying the duration of staining incubation showed that there was a typical FACS pattern and SP% was constant when the incubation was at least 90 min. (3) Compare to water bath, SP% was more than in thermostat, the SP% was (18.67 +/- 0.45)%, (22.6 +/- 0.50)% respectively; (4) Cell density is also responsible for SP%. The low density the cell is, the less in SP%. SPSS13.0 was used in statistical analysis, the groups were compared using t-Test. P < 0.05 was considered statistically significant. CONCLUSION: The optimum concentration and duration of incubation of Hoechst33342 in isolation of the side population cells in laryngeal carcinoma cell line Hep-2 is 10 microg/ml and 90 min. Incubated in water bath is better than in thermostat. The best staining cell density is around 80%-90%.