Application of narrow band imaging and Lugol's iodine staining in screening for nasopharyngeal carcinoma.

BACKGROUND: To investigate the diagnostic value of conventional white light endoscopy (WLE), narrow band imaging (NBI) endoscopy, and Lugol's iodine staining under WLE (endoscopic iodine staining) in the screening and early diagnosis of nasopharyngeal carcinoma. METHODS: Patients with nasopharyngeal lesions requiring biopsy attending the Department of Otolaryngology Head and Neck Surgery in our hospital between January 2021 and April 2023 were included in this study. Before biopsy, all subjects underwent conventional WLE, NBI endoscopy, and endoscopic iodine staining. On WLE, according to nasopharyngeal lesion morphology and color, patients were diagnosed with nasopharyngeal carcinoma ( +) or chronic hyperplastic nasopharyngitis (-). On NBI endoscopy, according to nasopharyngeal lesion vascular morphology, patients with type V manifestations (nasopharyngeal carcinoma) were categorized as NBI ( +) and patients with type I-IV manifestations (chronic hyperplastic nasopharyngitis) were categorized as NBI (-). Endoscopic iodine staining (1.6% Lugol's iodine solution) was positive ( +) if the mucosal surface was brown with no white patches, or negative (-) if there was no or light brown staining of the mucosal surface. Patients were divided into 2 groups based on histopathological diagnosis: nasopharyngeal carcinoma or chronic hyperplastic nasopharyngitis. Endoscopic diagnoses were compared with histopathological findings. The diagnostic performance of WLE, NBI endoscopy and endoscopic iodine staining for nasopharyngeal carcinoma were determined. RESULTS: This study included 159 patients. On histopathology, 29 patients were diagnosed with nasopharyngeal carcinoma, and 130 patients were diagnosed with chronic hyperplastic nasopharyngitis. There were no significant differences in the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), accuracy, and area under the receiver operating characteristic (ROC) curve (AUC) of conventional WLE, NBI endoscopy or endoscopic iodine staining for differentiating nasopharyngeal carcinoma and chronic hyperplastic nasopharyngitis. The diagnostic performance of the combination of conventional WLE, NBI endoscopy and endoscopic iodine staining was significantly improved compared to any procedure alone. CONCLUSIONS: Conventional WLE, NBI endoscopy or endoscopic iodine staining had good diagnostic performance for differentiating nasopharyngeal carcinoma and chronic hyperplastic nasopharyngitis. In particular, NBI endoscopy and endoscopic iodine staining alone or combined had clinical utility for identifying patients with nasopharyngeal lesions that are eligible for a watch-and-wait strategy.

Horizontal middle partial laryngectomy with cricothyroidopexy: an effective treatment for stage T1b glottic carcinoma.

BACKGROUND: The current treatment options for T1b glottic carcinoma often lead to poor treatment outcomes or voice quality. OBJECTIVES: This study evaluates the therapeutic efficacy of horizontal middle partial laryngectomy with cricothyroidopexy (HMPL-CTP) for stage T1b glottic carcinoma. MATERIAL AND METHODS: A retrospective analysis was conducted on 73 patients with T1b glottic carcinoma. The patients were categorized into three groups: Group A (n = 22) underwent transoral laser microsurgery (TLMS), Group B (n = 21) received frontolateral vertical partial laryngectomy (FVPL), and Group C (n = 30) underwent HMPL-CTP. The study analyzed the 5-year overall survival rate (OS), recurrence rate, phonatory status, and incidence of laryngeal stenosis. RESULTS: Voice quality scores varied significantly in the three groups, while the 5-year OS were similar. The local recurrence rate is higher in Group A than in the other two groups. The laryngeal stenosis rate in Group B is higher than in Groups A and C. Adhesions in the anterior commissure were observed in 18 cases in Group A and nine cases in Group C. CONCLUSIONS AND SIGNIFICANCE: HMPL-CTP demonstrates efficacy as a treatment for stage T1b glottic carcinoma, offering favorable preservation of laryngeal function and minimal complications.

Distinct microbiota dysbiosis in patients with laryngopharynx reflux disease compared to healthy controls.

OBJECTIVE: To compare the differences in the laryngopharynx microbiome between patients with laryngopharyngeal reflux disease (LPRD) and healthy people and further explore the influence of related risk factors pharyngeal microbiome. METHODS: This was a case-control study. Patients with a reflux symptom index (RSI) score > 13 or reflux finding score (RFS) score > 7 were diagnosed with suspected LPRD at the Department of Otolaryngology-Head and Neck Surgery of The 900th Hospital of Joint Logistic Support Force. Patients were assessed using a related risk factors questionnaire survey and examined by electronic naso-laryngoscopy. Simultaneously, laryngopharynx secretions were collected from the patients. The patients received at least eight weeks of proton pump inhibitor therapy, and those who responded were enrolled in the final experimental group. In parallel, laryngopharynx secretions were collected from healthy volunteers as the control group, and the laryngopharynx microbiota were analyzed using second-generation high-throughput sequencing. RESULTS: A total of 23 cases each in the experimental and control group were included in this study. The experimental group microbiota were composed of Streptococcus, Prevotella, Haemophilus, Neisseria, Actinobacillus, Fusobacterium, and Porphyromonas. There was no significant difference in microbial alpha and beta-diversity analysis between the two groups. However, some advantageous bacterium groups were significantly different. The abundance of Prevotella in the experimental group was significantly higher than that of the control group (U = 117, P < 0.05), while the abundance of Fusobacterium (U = 140, P = 0.006) and Porphyromonas (U = 120, P = 0.002) was significantly lower than the control group. Smoking was positively correlated with Pectin (r = 0.46, P = 0.037), Lactobacillus (r = 0.48, P = 0.027), and Clostridium (r = 0.46, P = 0.037), while alcohol was negatively correlated with Streptococcus (r = - 0.5539, P = 0.0092). CONCLUSION: The dominant microflora in the laryngopharynx of LPRD patients was significantly different from that of healthy people, suggesting that the change of laryngopharynx microflora may play an important role in the pathogenesis of LPRD. Smoking, drinking, eating habits, and age correlated with different genus levels of the laryngopharynx microbiota.

Clinicopathological factors affecting the prognosis of massive hemorrhage after radiotherapy for patients having nasopharyngeal carcinoma.

AIMS: The aim of the study is to investigate the clinicopathological factors that determine prognosis of nasopharyngeal hemorrhage after radiotherapy in patients with nasopharyngeal carcinoma (NPC). PATIENTS AND METHODS: The clinicopathological data of 539 patients with NPC, who received radiotherapy, were analyzed retrospectively. Parameters included gender; age; T-stage; N-stage; pathological type; type of radiotherapy; synchronous chemotherapy; secondary-course radiotherapy; radiation-induced skull base osteonecrosis; diabetes, hypertension, or other systemic diseases; results of nasopharyngeal bacterial culture; and nasopharyngeal tumor recurrence. Univariate and multivariate analyses were performed using the Chi-square test and logistic regression. Afterward, the Kaplan-Meier's method was applied to analyze the survival of patients with nasopharyngeal hemorrhage. RESULTS: Among all patients examined, 64 (11.9%) had nasopharyngeal hemorrhage after radiotherapy. The univariate analysis showed that T-stage (P < 0.01), secondary-course radiotherapy (P < 0.01), radiation-induced skull base osteonecrosis (P < 0.01), nasopharyngeal bacterial culture results (P < 0.01), and nasopharyngeal tumor recurrence (P < 0.01) were associated with nasopharyngeal hemorrhage. Multivariate analysis showed that only radiation-induced skull base osteonecrosis was significantly associated with nasopharyngeal hemorrhage after radiotherapy (odds ratio = 41.83, P = 0.0001). Nevertheless, in patients with internal carotid artery hemorrhage, the survival rate was much lower than that in patients with external carotid artery bleeding. The main cause of death during follow-up was rebleeding. CONCLUSION: The rate of mortality in patients with nasopharyngeal hemorrhage after radiotherapy was high. The presence of radiation-induced skull base osteonecrosis was a decisive factor in these patients. However, after successful rescue, arterial embolization or stent implantation is proposed to prolong survival.

Efficient remediation of crude oil-contaminated soil using a solvent/surfactant system.

Crude oil contaminated soil has been widely recognized to constitute a major environmental issue due its adverse effects on human health and ecological safety. The main objective of this study is to explore the possibility of using an ex situ solvent/surfactant washing technique for the remediation of crude oil-contaminated soil. Three organic solvents (methanol, acetone, and toluene) and one surfactant (AES-D-OA) were employed to form three kinds of solvent/surfactant systems, and utilized to evaluate the desorption performance of crude oil from soil. Natural soil, crude oil-contaminated soil, and after-remediation soil were characterized by SEM, EDX, FT-IR, and contact angle. The ability of solvent/surfactant systems to remove crude oil from soil was determined as a function of solvent polarity, mass ratio of solvent to surfactant, temperature, and ionic strength. The removal of crude oil by the toluene/AES-D-OA system was found to be more effective than the other systems. At a high toluene ratio, more than 97% of crude oil could be removed from contaminated soil. Crude oil removal efficiency was also found to increase with rising temperature or increasing ionic strength appropriately. Experimental results suggested that, compared to conventional surfactant-aided remediation, the combined utilization of surfactant and solvent achieved superior results for crude oil removal because of their similar compositions and structures in terms of aromaticity and polarity.

Dynamic Covalent Silica Nanoparticles for pH-Switchable Pickering Emulsions.

Dynamic covalent surfactants have been recently reported for preparation of pH-switchable emulsions [ Sun , D. Langmuir , 2017 , 33 , 3040 ]. In this study, dynamic covalent silica (SiO2-B) nanoparticles of switchable wettability were fabricated by a pH-responsive dynamic (covalent) imine bond between hydrophilic amino silica (SiO2-NH2) nanoparticles and hydrophobic benzaldehyde molecules. The properties of SiO2-B were characterized by Fourier transform infrared spectroscopy, elemental analysis, contact angle measurement, and ζ potential measurement. The hydrophilicity and hydrophobicity of SiO2-B were shown to be readily switchable by adjusting pH between 7.8 and 3.5. At pH 7.8, SiO2-B was partially hydrophobic and adsorbed at oil-water interface to stabilize O/W Pickering emulsions, which were characterized by electrical conductivity, optical microscopy, and confocal laser scanning microscopy. Upon lowering the pH to 3.5, the dynamic covalent bond is dissociated to convert partially hydrophobic SiO2-B into highly hydrophilic SiO2-NH2 and surface-inactive benzaldehyde. Both of them desorb from oil-water interface, resulting in a rapid oil-water separation of the Pickering emulsions. Alternating stabilization and phase separation of the Pickering emulsions over 3 cycles were demonstrated by adjusting the pH. The pH-switchable Pickering emulsions show great potential in application to effective oil-water separation of emulsions.

Prognostic significance of residual or recurrent lymph nodes in the neck for patients with nasopharyngeal carcinoma after radiotherapy.

BACKGROUND: Nasopharyngeal carcinoma is the most frequently diagnosed primary tumor originating from the nasopharynx, and the preferred treatment modality is radiotherapy. AIMS: To identify nasopharyngeal carcinoma prognostic factors in patients with residual or recurrent cervical lymph node metastases after radiotherapy. PATIENTS AND METHODS: The clinicopathologic characteristics and prognoses of 67 nasopharyngeal carcinoma patients with residual or recurrent cervical lymph node metastases who were diagnosed and treated were analyzed retrospectively. The Chi-squared test and Cox proportional hazard regression model were used to investigate associations between survival and clinicopathological features. Cumulative survival plots were obtained using the Kaplan-Meier method. RESULTS: Data analysis using the Cox proportional hazard regression model revealed that the size of residual or recurrent lymph node metastases, level V lymph node involvement, number of involved levels, surgical procedure performed, and distant metastases were significantly associated with overall survival. Chi-squared analysis only determined a significant correlation between distant metastases and patient survival. Furthermore, the Kaplan-Meier analysis demonstrated that the 1-, 3-, and 5-year survival rates for patients were 86.6%, 52.2%, and 38.6%, respectively. Radical neck dissection resulted in substantially longer overall survival than modified neck dissection. CONCLUSION: The size of residual or recurrent lymph node metastases, level V lymph node involvement, number of involved levels, surgical procedure performed, and presence of distant metastases were prognostic factors for nasopharyngeal carcinoma patients with residual or recurrent cervical lymph node metastases after radiotherapy, with distant metastases being the most important determinant. Classical radical neck dissection is recommended for treating recurrent nodal disease in nasopharyngeal carcinoma.

Language processing of auditory cortex revealed by functional magnetic resonance imaging in presbycusis patients.

CONCLUSION: Contralateral temporal lobe activation decreases with aging, regardless of hearing status, with elderly individuals showing reduced right ear advantage. BACKGROUND: Aging and hearing loss possibly lead to presbycusis speech discrimination decline. OBJECTIVES: To evaluate presbycusis patients' auditory cortex activation under verbal stimulation. METHOD: Thirty-six patients were enrolled: 10 presbycusis patients (mean age = 64 years, range = 60-70), 10 in the healthy aged group (mean age = 66 years, range = 60-70), and 16 young healthy volunteers (mean age = 25 years, range = 23-28). These three groups underwent simultaneous 1 kHz and 90 dB single-syllable word stimuli and (blood-oxygen-level-dependent functional magnetic resonance imaging) BOLD fMRI examinations. RESULTS: The main activation regions were superior temporal and middle temporal gyrus. For all aged subjects, the right region of interest (ROI) activation volume was decreased compared with the young group. With left ear stimulation, bilateral ROI activation intensity held. With right ear stimulation, the aged group's activation intensity was higher. Using monaural stimulation in the young group, contralateral temporal lobe activation volume and intensity were higher vs ipsilateral, while they were lower in the aged and presbycusis groups. On left and right ear auditory tasks, the young group showed right ear advantage, while the aged and presbycusis groups showed reduced right ear advantage.

RNF8 plays an important role in the radioresistance of human nasopharyngeal cancer cells in vitro.

Tumor residue or recurrence is common after radiation therapy for nasopharyngeal cancer (NPC) since the tumor cells can repair irradiation-induced DNA damage. The ubiquitination cascade mediates the assembly of repair and signaling proteins at sites of DNA double-strand breaks (DSBs). Ring finger protein 8 (RNF8) is an E3 ubiquitin ligase that triggers ubiquitination at the site of DSBs. The present study aimed to identify whether and how RNF8 small interfering RNA (siRNA) treatment enhances the radiosensitivity of irradiated human NPC cell lines. The CNE1, CNE2, and SUNE human NPC cell lines were stably transfected with a constructed RNF8-targeting siRNA expression vector. Western blotting was used to detect the effectiveness of RNF8 downregulation by RNF8 siRNA. The siRNA-transfected (RNF8-) and non-transfected (RNF8+) cells were irradiated at different doses by a linear accelerator. The growth inhibition ratio and apoptosis rate were detected by the methyl thiazolyl tetrazolium (MTT) assay and flow cytometry, respectively. The ataxia-telangiectasia mutated (ATM), DNA-PKcs, Chk1, Chk2, Nbs1 and Ku80 protein levels in each group were determined. The growth inhibition ratio and apoptotic percentage of RNF8- cells were higher than those of the RNF8+ cells in each of the three cell lines. Lower protein expression levels of Chk1, Chk2, ATM, and Nbs1 were observed in the irradiated RNF8- cells compared to the irradiated RNF8+ cells in each of the three cell lines (P<0.01). As a result, a conclusion could be drawn that RNF8 recruits and ubiquitinates many factors to repair DNA damage, including DSBs, thereby conferring radioresistance to NPC cells.

[The correlation of human papillomavirus and EB virus infection in nasopharyngeal carcinoma in Fujian].

OBJECTIVE: To investigate the prevalence of human papillomavirus in nasopharyngeal carcinomas of Fujian province in China. METHOD: Samples from 70 patients with NPC and 25 controls. All samples were detected HPV DNA by polymerase chain reaction (PCR) suing GP5+/6+ and MY09/11 primers and genotyped by surface plasmon resonance (SPR) and HPV 16/18 E6 and LMP-1 using immunohistochemistry and EBER using in situ hybridization. RESULT: Only 2 cases of 70 patients were showed evidence of HPV DNA by PCR, the 2 HPV positive cases subtype HPV-70 and HPV-18 were genotyped by SPR, both the 2 HPV positive cases are non-keratinizing carcinomas (the HPV-70 positive one is differentiated and the HPV-18 positive one is undifferentiated), both the 2 HPV positive cases do not show any evidence of EBV. Data showed that 57 of 70 NPC detected EBER positive, but only 25 out of 70 NPC samples were detected LMP-1 positive. CONCLUSION: Our study showed a low prevalence of human papillomavirus in NPC patients of Fujian province in Southern China, there is no evidence about HPV and EBV co-infection.

Retrospective study of selective neck dissection versus radical neck dissection as salvage therapy for patients with recurrent nasopharyngeal squamous cell carcinoma.

PURPOSE: While salvage surgery has been shown to improve the survival of patients with recurrent nasopharyngeal carcinoma, it is not known whether selective neck dissection has a beneficial or harmful effect on the survival of patients compared with modified neck dissection. The present study was aimed to compare the outcomes among patients receiving either selective neck dissection or modified neck dissection as salvage therapy for recurrent nasopharyngeal carcinoma. PATIENTS AND METHODS: In the present retrospective study, a total of 67 patients with recurrent nasopharyngeal squamous cell carcinoma were included. 23 patients received selective neck dissection and 44 patients received modified neck dissection. The clinical parameters and Kaplan-Meier 1-, 3-, 5-year overall survivals were compared for the two groups. RESULTS: The clinical parameters were comparable between the two groups. The 1-, 3-, 5-year survivals were 91.3%, 62.0% and 55.1% respectively for selective neck dissection and 93.1%, 82.5% and 77.9% respectively for modified neck dissection. Patients receiving modified neck dissection had a significant better overall survival than patients receiving selective neck dissection (χ(2) = 4.079, P = 0.043). CONCLUSION: Although selective neck dissection was associated with fewer complications, it was associated with poor over all survival compared with modified neck dissection. Further prospective, large-scale and long-term study is needed to confirm this conclusion.

Hypoxia promotes radioresistance of CD133-positive Hep-2 human laryngeal squamous carcinoma cells in vitro.

Hypoxia promotes the radioresistance of laryngeal carcinomas and CD133 is one of the markers expressed by tumor-initiating, human laryngeal carcinoma cells. In order to investigate whether CD133-positive Hep-2 cells exhibit a radioresistant phenotype and to determine whether hypoxia promotes this phenotype, we performed a series of experiments. Hep-2 cells, and Hep-2 cells stably expressing hypoxia-inducible factor (HIF)-targeted small interfering RNA (siRNA) were cultured under hypoxic and normoxic conditions and were treated with varying doses of γ-rays (0, 5, 10, 15 and 20 Gy). MTT and cell cycle assays were subsequently performed. Using fluorescence-activated cell sorting (FACS), CD133-positive Hep-2 cells and CD133-positive HIF-siRNA Hep-2 cells were isolated. These cells were grown as spheres under hypoxic and normoxic conditions for MTT and soft agar colony formation assays. The expression levels of DNA-dependent protein kinase catalytic subunit (DNA-PKcs), survivin, p53 and ataxia-telangiectasia mutated (ATM) were also assayed using flow cytometry. The data showed that the growth of Hep-2 cells exposed to hypoxic conditions and treated with 10 Gy radiation (group A) was less compared to that of groups B-D (P<0.05). In addition, more cells in group A were arrested in the G1 phase of the cell cycle compared to groups B-D (P<0.05). The percentage of CD133+ cells detected after radiation increased and was the highest for group A (P<0.05). In sphere formation assays, significantly more CD133+ cells grew in spheres than CD133- cells (P<0.001). Moreover, sphere formation was the highest for CD133+ Hep-2 cells grown under hypoxic conditions and exposed to irradiation (group E) (P<0.05). Lastly, expression of DNA-PKcs and survivin for group E was the highest (P<0.05), while ATM and p53 levels remained largely unchanged (P>0.05). In conclusion, CD133-positive Hep-2 cells exhibited a radioresistant phenotype that was enhanced with hypoxia. Furthermore, an increase in DNA-PK activity was associated with this enhancement.

[Risk factors analysis of local failure following radiotherapy and chemotherapy to nasopharyngeal carcinoma].

OBJECTIVE: To analyse the risk factors involved in local failure following radiotherapy and chemotherapy to nasopharyngeal carcinoma. METHOD: A retrospective analysis was carried out to review the histopathological data from 308 NPC patients who recieved medical treatment between 2004 and 2006. The incidence and the risk factor for local treatment failure were evaluated in a model that included the following factors: sex, age, T and N staging, histological grade of primary tumor, presence of cervical lymph node metastasis, size and laterals of positive neck nodes, levels involved, ways of radiation and condition of concurrent chemotherapy. Univariate chi2 test and multivariate stepwise logistic regression model were used for the analysis. Statistical analysis of survival of patients with local residues and recurrence was performed using Kaplan-Meier method. RESULT: Ninty-three cases (30.2%) presented local residues and recurrence in 308 patients of nasopharyngeal carcinomas. In the univariate analysis, it was confirmed that the following variables correlated to local failure, i. e., T staging (P < 0.01), N staging (P < 0.01), presence of cervical lymph node metastasis (P < 0.05), size and laterals of positive neck nodes (P < 0.05, respectively). In the multivariate analysis, the most ignificant risk factors for local failure were the T staging only. Kaplan-Meier analysis showed that overall survival rates of 71 NPC patients with local residues and recurrence who received re-treatment were 77.2% at 1 year, 40.4% at 3 years, 22.4% at 5 years, respectively. CONCLUSION: T staging is the key risk factors in determining the development of local failure following radiotherapy or chemotherapy in NPC patients. Patients with primary tumor infiltrating bone have the higher risk of developing local residues and recurrence. Retreatment to the patients suffering from local failure can imrove survival rates.

Regulation of glucose transporter protein-1 and vascular endothelial growth factor by hypoxia inducible factor 1α under hypoxic conditions in Hep-2 human cells.

The present study evaluated the regulation of glucose transporter protein-1 (Glut-1) and vascular endothelial growth factor (VEGF) by hypoxia inducible factor 1α (HIF-1α) under hypoxic conditions in Hep-2 human cells to explore the feasibility of these three genes as tumor markers. Hep-2 cells were cultured under hypoxic and normoxic conditions for 6, 12, 24, 36 and 48 h. The proliferation of Hep-2 cells was evaluated using an MTT assay. The protein and mRNA expression levels of HIF-1α, Glut-1 and VEGF were detected using the S-P immunocytochemical method, western blotting and reverse transcription polymerase chain reaction (RT-PCR). The results revealed that the expression levels of HIF-1α, Glut-1 and VEGF protein in Hep-2 cells were significantly elevated under hypoxic conditions compared with those under normoxic conditions over 36 h. Under hypoxic conditions, mRNA levels of HIF-1α were stable, while mRNA levels of Glut-1 and VEGF changed over time. In conclusion, Glut-1 and VEGF were upregulated by HIF-1α under hypoxic conditions in a time-dependent manner in Hep-2 cells and their co-expression serves as a tumor marker.

[Expression of hypoxia inducible factor-1α and correlated target genes in human laryngeal carcinoma].

OBJECTIVE: To detect the expression of hypoxia inducible factor 1 alpha (HIF-1α), glucose transporter protein-1 (GLUT-1) and vascular endothelial growth factor (VEGF) in human laryngeal carcinoma tissue, and to study the relationship between hypoxia and HIF-1α, GLUT-1, VEGF in human laryngeal carcinoma Hep-2 cells and to explore the effect of HIF-1α, GLUT-1 and VEGF as endogenous hypoxic markers on laryngeal carcinoma. METHODS: The expression levels of HIF-1α, GLUT-1 and VEGF were detected in 35 cases of laryngeal carcinoma by SP immunohistochemical methods and in Hep-2 cells by SP immunocytochemical methods. The relationship between HIF-1α and GLUT-1, VEGF protein expression was analyzed. RESULTS: Of the 35 cases, 16 cases expressed HIF-1α, 16 cases expressed GLUT-1, 19 cases expressed VEGF. The expression of HIF-1α and VEGF were closely correlated with pathologic grading and lymphnode metastasis. GLUT-1 was correlated with lymphnode metastasis. The expression levels of HIF-1α, GLUT-1 and VEGF in Hep-2 cells under hypoxic condition were higher than those under normoxic condition. CONCLUSION: HIF-1α may promote the expression of GLUT-1 and VEGF in laryngeal carcinoma, furthermore promote tumor angiogenesis, invasion, and metastasis of the laryngeal carcinoma.

[Impacts of hypoxia on the features and chemoresistance of cancer stem cells in Hep-2 cells and underlying mechanism].

OBJECTIVE: To investigate the effects of hypoxia on the features and chemoresistance of cancer stem cells in Hep-2 cells and underlying mechanism. METHODS: The shRNA interference recombinant plasmid targeting HIF-1α was synthesized and transfected into Hep-2 cells. The HIF-1α knockdown Hep-2 cells were established after clonal selection and the expression of HIF-1α was measured. The cellular features including proliferation, clonal formation, cell cycle, apoptosis and CD133 phenotype were measured in Hep-2 cells cultured under hypoxic condition in vitro. CD133+ cells were sorted from Hep-2 cells with flow cytometry. Clonal formation test and cisplatin treatment were carried out, and the expressions of related genes (Oct-4, suvivin and p53) in CD133+ cells were measured. RESULTS: HIF-1α knockdown Hep-2 cells was successfully established, as evidenced by the reduced mRNA and protein expressions of HIF-1α. The Hep-2 cells cultured under hypoxic microenvironment showed higher proliferation and clonal formation activity, cell cycle arrest in G0/G1, lower apoptosis, up-regulated CD133, however the effects of hypoxia reduced in HIF-1α knockdown Hep-2 cells. CD133+ cells were successfully sorted from Hep-2 cells, and the CD133+ cells showed increased clonal formation activity and cisplatin treatment resistance in hypoxia. Also the effects of hypoxia on CD133+ cells decreased with HIF-1α knockdown, showing down-regulated Oct-4 and survivin and up-regulated p53. CONCLUSIONS: Hypoixa can induce the features of cancer stem cells in Hep-2 cells and increase proliferation, differentiation and chemoresistant ability of CD133+ cells, which might be correlated with the changes in expressions of HIF-1α and related genes regulated by HIF-1α.

[Cancer stem cells play an important role in resistance of laryngeal squamous cancer to irradiation mediated by hypoxia].

OBJECTIVE: To study whether laryngeal cancer stem cells in hypoxia have the characteristic of resistance to irradiation and underlying mechanism. METHODS: CD133(+) cells were separated from Hep-2 cells with flow cytometry (FCM) and the purity was 92.8%. The separated CD133(+) cells were cultured in serum-free medium in hypoxia in normoxia environment respectively, and hypoxia-inducible factor 1 alpha (HIF-1α) expression was detected by FCM. The cells were exposed respectively to X-rays emitted by linear accelerator with a dose of 0, 5, 10, 15 or 20 Gy for 24 hours, with additional time points of 12, 36, and 48 hours for the cells exposed to 10 Gy. Then the growth inhibition ratios of cells in hypoxia and normoxia groups were detected with MTT assay at different time points. Soft agar colony formation assay was used to detect colony formation ratios of cells in hypoxia and normoxia groups. DNA dependent protein kinase catalytic subunit (DNA-PKcs), ataxia telangiectasia mutate (ATM), Survivin and P53 were detected by FCM. RESULTS: Growth inhibition ratio of CD133(+) cells in hypoxia group was lower than that in normoxia group (P < 0.05). Colony formation ratio of CD133(+) cells was higher than that of CD133(-) cells (P < 0.01) and the ratio of CD133(+) cells in hypoxia group was higher than that in normoxia group (P < 0.05). The ratio of hypoxia group was not affected by irradiation, while the ratio of normoxia group decreased significantly after irradiation (P < 0.05). The expressions of DNA-PKcs, ATM, Survivin and P53 in CD133(+) cells were higher than those in CD133(-) cells respectively (P < 0.01). In CD133(+) cells with radiation, the expressions of DNA-PKcs and Survivin of hypoxia group were higher those of normoxia group (P < 0.05), but no difference in the expression of ATM or P53 between the two groups. CONCLUSIONS: Laryngeal cancer stem cells play an important role in radioresistance mediated by hypoxia.

[Cancer stem cells promotes resistance of laryngeal squamous cancer to irradiation mediated by hypoxia].

OBJECTIVE: To study whether cancer stem cells promotes resistance of laryngeal squamous cancer to irradiation mediated by hypoxia. METHOD: Hep-2 cells were respectively cultured in hypoxia and normoxia environment, and the express of HIF-la was detected by western blot. Then they were radiated with different doses of gamma-rays. After that we detected growth inhibition ratio with MTT assay, cell circle and ratio of CD133+ cells with Flow cytometry at different times. RESULT: MTT assay showed that inhibition ratio of the hypoxia group was lower than that of the normoxia group after different doses of gamma-rays at each time point, and the difference was significant 24 h after 10 Gy irradiation (P < 0.05). The results of Flow cytometry demonstrated that cells of the two groups were arrested at G1 phase, and cells ratio in G1 phase of the hypoxia group was higher than that of he normoxia group after 10 Gy irradiation. The ratio of CD133-positive cells was higher in the hypoxia group than in the normoxia group after radiation, and difference was significant 24 h after 10 Gy irradiation (P < 0.05). In each group, the ratio of CD133-positive cells became higher after radiation than that before radiation (P < 0. 05). CONCLUSION: We can conclude that cancer stem cells play an important role in radioresistance mediated by hypoxia.

[Application of reversed pedicled submental island flap].

OBJECTIVE: To explore the clinical applications of reversed pedicled submental island flap in the face and oropharynx. METHODS: The clinical data of ten cases of reconstruction of defect in the area of face or oropharynx following resection of tumors with reversed pedicled submental island flaps between January of 2004 to December of 2006 were retrospectively studied. The cases included six males and four females, aged from 24 to 76 (median 55 years). One of the cases suffered from upper lid melanoma, two hard palate myoepithelioma, one maxillary ameloblastoma, four tonsil cancer and two lingual carcinoma. The submental island flaps were dissected according to the area of the defection, the distal facial vessel was used as the pedicle of the flaps, and the flaps were transferred through under the skin of the face or the mandible to the area of the defection. Radical neck dissection was performed in the four tonsil cancers and two lingual carcinomas simultaneously. RESULTS: All the flaps showed pale, edema and/or congestion after the surgical treatment, but swelling and congestion disappeared gradually 5 days later. One of the cases suffered from severe congestion of the flap, but the flap survived by stabbing with needle and draining. The flap of another case separated from the hard palate was sutured again, and healed. Lower lip palsy occurred in another case, and recovered 3 months later by conservative therapy. None of the flaps necrosed. CONCLUSIONS: Because of the upper pedicle, reversed pedicled submental island flap can be transferred to a long distance to reconstruct the defect in the upper face or around the orbit, and radical neck dissection can be performed simultaneously, it is a reasonable candidate in repairing the defect of the face and oropharynx.