Foreign Body Granuloma Originating From the Lateral Process of Malleus: A Case Series of 19 Children.

OBJECTIVE: A special presentation of foreign body granuloma originating from the lateral process of the malleus (FBGLP) was noted in the absence of a history of foreign body entry into the external auditory canal (EAC). This study reported the clinical features, pathology, and prognosis of patients with FBGLP. DESIGN: Retrospective study. SETTING: Shandong Provincial ENT Hospital. PATIENTS: Nineteen pediatric patients (age, 1-10 yr) with FBGLP. INTERVENTIONS: Clinical data were collected from January 2018 to January 2022. MAIN OUTCOME MEASURES: Clinicopathologic characteristics of the patients were analyzed. RESULTS: All patients had an acute course, and were within 3 months of ineffective medical treatment. The most common symptoms were suppurative (57.9%) and hemorrhagic (42.1%) otorrhea. FBGLP imaging examinations demonstrated a soft mass blocking the EAC without bone destruction and occasionally concomitant effusion in the middle ear. The most common pathologic findings were foreign body granuloma (94.7%,18/19), granulation tissue (73.7%, 14/19), keratotic precipitate (73.7%, 14/19), calcium deposition (63.2%, 12/19), hair shafts (47.4%, 9/19), cholesterol crystals (5, 26.3%), and hemosiderin (15.8%, 3/19). Foreign body granuloma and granulation tissue showed higher expression levels of CD68 and cleaved caspase-3 than did the normal tympanic mucosa, whereas Ki-67 levels were similarly low in all tissues. The patients were followed up for 3 months to 4 years without recurrence. CONCLUSION: FBGLP is caused by endogenous foreign particles in the ear. We recommend the trans-external auditory meatus approach for FBGLP surgical excision, as this shows promising outcomes.

Single-cell HER2 quantification via instant signal amplification in microdroplets.

Accurate and ultrasensitive evaluation of human epidermal growth factor receptor 2 (HER2) protein is key to early diagnosis and subtype differentiation of breast cancer. Single-cell analyses to reduce ineffective targeted therapies due to breast cancer heterogeneity and improve patient survival remain challenging. Herein, we reported a novel droplet microfluidic combined with an instant cation exchange signal amplification strategy for quantitative analysis of HER2 protein expression on single cells. In the 160 µm droplets produced by a tapered capillary bundle, abundant Immuno-CdS labeled on HER2-positive cells were replaced by Ag + to obtain Cd2+ that stimulated Rhod-5N fluorescence. This uniformly distributed and instantaneous fluorescence amplification strategy in droplets improves sensitivity and reduces signal fluctuation. Using HER2 modified PS microsphere to simulate single cells, we obtained a linear fitting of HER2-modified concentration and fluorescence intensity in microdroplets with the limit detection of 11.372 pg mL-1. Moreover, the relative standard deviation (RSD) was 4.2-fold lower than the traditional immunofluorescence technique (2.89% vs 12.21%). The HER2 protein on SK-BR-3 cells encapsulated in droplets was subsequently quantified, ranging from 9862.954 pg mL-1 and 205.26 pg mL-1, equivalent to 9.795 × 106 and 2.038 × 105 protein molecules. This detection system provides a universal platform for single-cell sensitive quantitative analysis and contributes to the evaluation of HER2-positive tumors.

Decellularized tympanic membrane scaffold with bone marrow mesenchymal stem cells for repairing tympanic membrane perforation.

BACKGROUND: Tympanic membrane perforation (TMP) is a common disease in otology, and few acellular techniques have been reported for repairing this condition. Decellularized extracellular matrix (ECM) scaffolds have been used in organ reconstruction. OBJECTIVE: This study on tissue engineering aimed to develop a tympanic membrane (TM) scaffold prepared using detergent immersion and bone marrow mesenchymal stem cells (BMSCs) as repair materials to reconstruct the TM. RESULTS: General structure was observed that the decellularized TM scaffold with BMSCs retained the original intact anatomical ECM structure, with no cell residue, as observed using scanning electron microscopy (SEM), and exhibited low immunogenicity. Therefore, we seeded the decellularized TM scaffold with BMSCs for recellularization. Histology and eosin staining, SEM and immunofluorescence in vivo showed that the recellularized TM patch had a natural ultrastructure and was suitable for the migration and proliferation of BMSCs. The auditory brainstem response (ABR) evaluated after recellularized TM patch repair was slightly higher than that of the normal TM, but the difference was not significant. CONCLUSION: The synthetic ECM scaffold provides temporary physical support for the three-dimensional growth of cells during the tissue developmental stage. The scaffold stimulates cells to secrete their own ECM required for tissue regeneration. The recellularized TM patch shows potential as a natural, ultrastructure biological material for TM reconstruction.

Development of a decellularized hypopharynx with vascular pedicle scaffold for use in reconstructing hypopharynx.

BACKGROUND: Hypopharynx reconstruction after hypopharyngectomy is still a great challenge. Perfusion decellularization is for extracellular matrix (ECM) scaffolding and had been used in organ reconstruction. Our study aimed to prepare an acellular, natural, three-dimensional biological hypopharynx with vascular pedicle scaffold as the substitute materials to reconstruct hypopharynx. RESULT: Scanning electron microscope and histology staining showed that the decellularized hypopharynx with vascular pedicle scaffold retained intact native anatomical ECM structure. Myoblasts were observed on the recellularized scaffolds with bone marrow mesenchymal stem cells induced by 5-azacytidine implanted in the rabbit greater omentum by immunohistochemical analysis. CONCLUSION: The decellularized hypopharynx with vascular pedicle scaffold prepared by detergent perfusion in our study has a potential to be an alternative material to pharynx reconstruction.

The effect of balloon dilatation eustachian tuboplasty combined with grommet insertion on the structure and function of the eustachian tube in patients with refractory otitis media with effusion.

BACKGROUND: The treatments for refractory secretory otitis media with effusion usually choose long-term grommet insertion. This study evaluated the effect of balloon eustachian tuboplasty combined with grommet insertion on the function and the opening length of the eustachian tube in patients with refractory otitis media with effusion. METHODS: Fifty-seven patients with refractory otitis media with effusion were enrolled. A three-dimensional reconstruction of an iohexol-enhanced computed tomography image was applied to evaluate the structural and length changes of the eustachian tube at both resting and Valsalva maneuver states. The grommet was removed 3 months after the operation and postoperative follow-up was carried out from 3 to 12 months. We performed pre- and post-operative observation of the following: appearance of the tympanic membrane, pure-tone audiometry threshold, eustachian tube score, seven-item Eustachian Tube Dysfunction Questionnaire scores (ETDQ-7), quantitative examination of eustachian tube function dynamic observation of tympanogram peak pressure point, and computed tomography examination of the eustachian tube. RESULTS: The pure-tone audiometry at 1, 3, 6, 9, and 12 months postoperatively were all significantly lower compared to the preoperative value (all P<0.05). There was no significant difference between the pure-tone audiometry at 6 and 9 months postoperatively, neither was for the air-bone conduction gap at these time points. The quantitative examination peak pressure deviation was markedly increased at 6 months postoperatively compared with that before the operation (all P<0.05). The peak pressure deviation of tympanometry at 6 and 9 months postoperatively were both higher than the value at 12 months after surgery (P<0.05). The eustachian tube score at 1, 3, 6, 9, and 12 months postoperatively were notably higher than that before the operation (all P<0.05). A significant difference was also observed between the 6- and 12-month postoperative eustachian tube score (P<0.05). There was a significant difference in the ETDQ-7 scores at 6- and 12-month postoperatively (P<0.05). The quantitative examination peak pressure deviation and eustachian tube score were both correlated with development length of the eustachian tube after three-dimensional computed tomography reconstruction (P<0.05). CONCLUSIONS: Eustachian tube balloon dilatation combined with grommet insertion is an effective treatment for refractory otitis media with effusion.

Tissue-engineered esophagus: recellular esophageal extracellular matrix based on perfusion-decellularized technique and mesenchymal stem cells.

Perfusion-decellularization was an interesting technique to generate a natural extracellular matrix (ECM) with the complete three-dimensional anatomical structure and vascular system. In this study, the esophageal ECM (E-ECM) scaffold was successfully constructed by perfusion-decellularized technique through the vascular system for the first time. And the physicochemical and biological properties of the E-ECM scaffolds were evaluated. The bone marrow mesenchymal stem cells (BMSCs) were induced to differentiate into myocytesin vitro. E-ECM scaffolds reseeded with myocytes were implanted into the greater omenta to obtain recellular esophageal ECM (RE-ECM), a tissue-engineered esophagus. The results showed that the cells of the esophagi were completely and uniformly removed after perfusion. E-ECM scaffolds retained the original four-layer organizational structure and vascular system with excellent biocompatibility. And the E-ECM scaffolds had no significant difference in mechanical properties comparing with fresh esophagi,p> 0.05. Immunocytochemistry showed positive expression ofα-sarcomeric actin, suggesting that BMSCs had successfully differentiated into myocytes. Most importantly, we found that in the RE-ECM muscularis, the myocytes regenerated linearly and continuously and migrated to the deep, and the tissue vascularization was obvious. The cell survival rates at 1 week and 2 weeks were 98.5 ± 3.0% and 96.4 ± 4.6%, respectively. It was demonstrated that myocytes maintained the ability for proliferation and differentiation for at least 2 weeks, and the cell activity was satisfactory in the RE-ECM. It follows that the tissue-engineered esophagus based on perfusion-decellularized technique and mesenchymal stem cells has great potential in esophageal repair. It is proposed as a promising alternative for reconstruction of esophageal defects in the future.

Significance of ATP-binding cassette transporter proteins in multidrug resistance of head and neck squamous cell carcinoma.

According to the cancer stem cell theory, a small subpopulation of cancer cells, known as cancer stem cells (CSCs), exist that are self-renewing and are involved in tumor invasion, metastasis and recurrence. A number of studies have reported that certain cancer cells are able to efflux the Hoechst 33342 dye. These cells are termed side population (SP) cells and share characteristic features of CSCs. The results of the present study revealed that 2.7% of primary head and neck squamous cell carcinoma (HNSCC) cells were SP cells. This was reduced to 0.7% following treatment with verapamil. The immunofluorescence and reverse transcription polymerase chain reaction analysis revealed that SP cells have an enhanced expression of the ATP-binding cassette (ABC) transporter protein ABC subfamily G, member 2 (ABCG2), which has been identified to be actively involved in drug exclusion. Similarly, the mRNA level of the oncogene B lymphoma Mo-MLV insertion region-1 and the stem cell surface proteins nestin and octamer-binding transcription factor-4 were highly expressed in the SP cells compared with the non-SP cells. In addition, it was demonstrated that HNSCC SP cells exhibited increased proliferation and were highly resistant to multiple drugs. These findings suggest that the presence of CSCs, such as SP cells, may be responsible for chemotherapy failure and tumor relapse in patients with HNSCC. Therefore, the identification of a novel therapeutic drug that could effectively target CSCs may help to eradicate refractory tumors.

Constitutive secretion of Interleukin-4 dictates CD133+ side population cells to resist drug treatment and cell death.

PURPOSE: In the present study, we made an attempt to elucidate the role of oversecretion of interleukin-4 (IL-4) in cancer stem cells (CSCs) of head and neck squamous cell carcinoma (HNSCC). METHODS: HNSCC samples were analyzed for the presence of CSCs by flow cytometry. In addition, we have performed drug and apoptosis resistance assays to determine the role of IL-4 in CSCs. RESULTS: HNSCC samples contained 3.3% of CD133+ cancer stem like side population (SP) cells in HNSCC which displayed infinite cell proliferation and they had high self-renewal capacity. These CD133+ cells displayed enhanced expression of IL-4, which promoted multidrug and apoptosis resistance. After neutralizing IL-4, the CD133+ SP cells became more sensitive to drug treatment and apoptosis. CONCLUSIONS: Our data suggest that the autocrine secretion of IL-4 is a potential target for the development of novel anticancer drugs to prevent the CSCs-mediated therapy failure and tumorigenesis.

Three New Cytotoxic ent-Kaurane Diterpenes from Isodon excisoides.

Three types of ent-kaurane diterpenoids were isolated from the aerial parts of Isodon excisoides, including three new diterpenoids, 1α,7α,14ß-trihydroxy-20-acetoxy-ent-kaur-15-one (1); 1α,7α,14ß,18-tetrahydroxy-20-acetoxy-ent-kaur-15-one (2); and 1α-acetoxy-14ß-hydroxy-7α,20-epoxy-ent-kaur-16-en-15-one (3); together with six known diterpenes henryin (4); kamebanin (5); reniformin C (6); kamebacetal A (7); kamebacetal B (8); and oridonin (9). The structures of the isolated compounds were elucidated by means of nuclear magnetic resonance spectroscopy and high-resolution mass spectrometry in conjunction with published data for their analogs, as well as their fragmentation patterns. Compounds 5 and 9 were isolated from Isodon excisoides for the first time. To explore the structure-activity relationships of the isolated compounds, they were tested for their cytotoxic effects against five human cancer cell lines: HCT-116, HepG2, A2780, NCI-H1650, and BGC-823. Most of the isolated compounds showed certain cytotoxic activity against the five cancer cell lines with IC50 values ranging from 1.09-8.53 µM. Among the tested compounds, compound 4 exhibited the strongest cytotoxic activity in the tested cell lines, with IC50 values ranging from 1.31-2.07 µM. Compounds 1, 6, and 7 exhibited selective cytotoxic activity.

Abnormal Wnt signaling and overexpression of ABCG2 contributes to drug efflux properties of side population cells in nasopharyngeal carcinoma.

The presence of cancer stem cells (CSCs) has major implications in the choice of cancer treatment strategy and is responsible for tumor relapse. CSCs have been isolated and characterized in several types of cancer; however, studies concerning the CSCs from nasopharyngeal carcinoma (NPC) are limited. Thus, the present study was designed to isolate and characterize the cancer stem-like side population (SP) cells from NPC samples. The fluorescence-activated cell sorting (FACS)-based Hoechst 33342 dye exclusion technique identified that 3.9% of cells from NPC samples were cancer stem-like SP cells. Upon treatment with verapamil (ABC transporter inhibitor), the percentage of SP cells was significantly reduced to 0.7%, which confirms that the ABC transporter protein exhibits a significant role in drug exclusion. Fluorescence microscopy analysis revealed that the FACS purified SP cells showed increased expression of ABCG2 (ATP transporter protein), Oct-4 and CD44 (stem cell surface protein). Furthermore, these SP cells exhibited increased mRNA expression of ABCG2 and anti-apoptotic factor Bmi-1, which contribute to multi-drug resistance and increased cell survival rate. Notably, the Wnt/ß-catenin signaling pathways are altered in SP cells. In addition, using reverse transcription­quantitative polymerase chain reaction analysis it was observed that the cells exhibited increased expression of DKK1 and AXIN2. In conclusion, data from the present study clearly demonstrated that the presence of cancer stem-like SP cells from NPC may be responsible for chemotherapeutic drug resistance, tumor recurrence and invasion.

Gene expression profiling via bioinformatics analysis reveals biomarkers in laryngeal squamous cell carcinoma.

The present study aimed to identify key genes and relevant microRNAs (miRNAs) involved in laryngeal squamous cell carcinoma (LSCC). The gene expression profiles of LSCC tissue samples were analyzed with various bioinformatics tools. A gene expression data set (GSE51985), including ten laryngeal squamous cell carcinoma (LSCC) tissue samples and ten adjacent non-neoplastic tissue samples, was downloaded from the Gene Expression Omnibus. Differential analysis was performed using software package limma of R. Functional enrichment analysis was applied to the differentially expressed genes (DEGs) using the Database for Annotation, Visualization and Integrated Discovery. Protein-protein interaction (PPI) networks were constructed for the protein products using information from the Search Tool for the Retrieval of Interacting Genes/Proteins. Module analysis was performed using ClusterONE (a software plugin from Cytoscape). MicroRNAs (miRNAs) regulating the DEGs were predicted using WebGestalt. A total of 461 DEGs were identified in LSCC, 297 of which were upregulated and 164 of which were downregulated. Cell cycle, proteasome and DNA replication were significantly over-represented in the upregulated genes, while the ribosome was significantly over-represented in the downregulated genes. Two PPI networks were constructed for the up- and downregulated genes. One module from the upregulated gene network was associated with protein kinase. Numerous miRNAs associated with LSCC were predicted, including miRNA (miR)-25, miR-32, miR-92 and miR-29. In conclusion, numerous key genes and pathways involved in LSCC were revealed, which may aid the advancement of current knowledge regarding the pathogenesis of LSCC. In addition, relevant miRNAs were also identified, which may represent potential biomarkers for use in the diagnosis or treatment of the disease.

microRNA-423-3p promotes tumor progression via modulation of AdipoR2 in laryngeal carcinoma.

Despite of the variety of combined modality treatments for laryngeal carcinoma have been introduced, the distance recurrence rate and 5-year overall survival rate over the past decades are still the major issues, underlining the importance to better understand the biological bases that contribute to disease progression. Here, we reported that miR-423-3p overexpressed in primary laryngeal carcinoma cell line where it plays a critical role in tumor progression. Suppression of miR-423-3p expression resulted in decreasing cell proliferation, clonogenicity, cell migration and invasion. By using in silico prediction algorithms for target identification, AdipoR2 (adiponectin receptor 2) and DUSP4 (MAP kinase phosphatase 2) were identified to be potential targets of miR-423-3p. Overexpression of miR-423-3p was associated with epigenetic silencing of AdipoR2 in human laryngeal carcinoma samples, which have been previously implicated in suppression of tumor proliferation and angiogenesis. Luciferase reporter assays and western blot further confirmed the direct interaction of miR-423-3p with AdipoR2. Our findings have demonstrated that miR-423-3p plays an important oncogenic role in laryngeal carcinoma progression, and further suggest that suppression of miR-423-3p expression might be useful for its clinical management.

Salmonella typhimurium mediated delivery of Apoptin in human laryngeal cancer.

An effective cancer therapeutic should target tumours specifically with limited systemic toxicity. Here, we transformed an attenuated Salmonella typhimurium (S. typhimurium) with an Apoptin expressing plasmid into a human laryngeal carcinoma cell line. The expression of the inserted gene was measured using fluorescence and immunoblotting assays. The attenuated S. typhimurium-mediated Apoptin significantly decreased cytotoxicity and strongly increased cell apoptosis through the activation of caspase-3. The process was mediated by Bax, cytochrome c and caspase-9. A syngeneic nude murine tumour model was used to determine the anti-tumour effects of the recombinant bacteria in vivo. Systemic injection of the recombinant bacteria with and without re-dosing caused significant tumour growth delay and reduced tumour microvessel density, thereby extending host survival. Our findings indicated that the use of recombinant Salmonella typhimurium as an Apoptin expression vector has potential cancer therapeutic benefits.

[Removal of cadmium from extracts of Gentianae Radix et Rhizoma by gamma-mercaptopropyl-modified silica gel].

OBJECTIVE: To study the optimum process of removing cadmium irons from extracts of Gentianae Radix et Rhizoma with gamma-mercaptopropyl-modified silica gel (MPS) and assess its cadmium ion-removing property. METHOD: Static and dynamic adsorptions were adopted to detect the cadmium-removing rate. MPS' cadmium ion-removing property was assessed with such indicators as the cadmium-removing rate, the solid content and the HPLC fingerprint. RESULT: The process parameters of the static adsorption were as follows: 0.20 g x mL(-1) of concentration of extracts, 120 minutes of adsorption time and 15:1 between raw materials and MPS. The process parameters of the dynamic adsorption were as follows: 1:3.5 times between diameter and height, 0.20 g x mL(-1) of concentration of extracts, 0.9 mL x min(-1) of flow rate of the extracts and 50:1 between raw materials and MPS. Before and after the cadmium ion-removing process, the extracts showed no notable difference in solid content and HPLC fingerprint. CONCLUSION: gamma-mercaptopropyl-modified silica gel (MPS) can effectively remove cadmium ion from the extracts of Gentianae Radix et Rhizoma with an excellent cadmium ion-removing property.

Temperature sensor based on an isopropanol-sealed photonic crystal fiber in-line interferometer with enhanced refractive index sensitivity.

We fabricate a simple, compact, and stable temperature sensor based on a liquid-sealed photonic crystal fiber (PCF) in-line nonpolarimetric modal interferometer. Different from other reported PCF devices, it does not need expensive polarimetric devices, and the liquid is sealed in one fiber. The device consists of a stub of isopropanol-filled PCF spliced between standard single-mode fibers. The temperature sensitivity (-166 pm/°C) increases over an order of magnitude compared with those of the previous sensors based on air-sealed PCF interferometers built via fusion splicing with the same mechanism. In addition, the refractive index sensitivity also increases. Higher temperature sensitivity can be realized by infiltrating some liquid having a higher thermo-optic coefficient into the microholes of the PCF.

Changes of NF-kB, p53, Bcl-2 and caspase in apoptosis induced by JTE-522 in human gastric adenocarcinoma cell line AGS cells: role of reactive oxygen species.

AIM: To identify whether JTE-522 can induce apoptosis in AGS cells and ROS also involved in the process, and to investigate the changes in NF-kB, p53, bcl-2 and caspase in the apoptosis process. METHODS: Cell culture, MTT, Electromicroscopy, agarose gel electrophoresis, lucigenin, Western blot and electrophoretic mobility shift assay (EMSA) analysis were employed to investigate the effect of JTE-522 on cell proliferation and apoptosis in AGS cells and related molecular mechanisms. RESULTS: JTE-522 inhibited the growth of AGS cells and induced the apoptosis. Lucigenin assay showed the generation of ROS in cells under incubation with JTE-522. The increased ROS generation might contribute to the induction of AGS cells to apoptosis. EMSA and Western blot revealed that NF-kB activity was almost completely inhibited by preventing the degradation of IkBalpha. Additionally, by using Western blot we confirmed that the level of bcl-2 was decreased, whereas p53 showed a great increase following JTE-522 treatment. Their changes were in a dose-dependent manner. CONCLUSION: These findings suggest that reactive oxygen species, NF-kB, p53, bcl-2 and caspase-3 may play an important role in the induction of apoptosis in AGS cells after treatment with JTE-522.