Chronic rhinosinusitis with nasal polyps (CRSwNP) is likely to relapse due to aberrant eosinophil infiltration. The deficiency of Vitamin D (VD) is associated with increased eosinophil infiltration in eosinophilic oesophagitis. However, the role of VD in eosinophilic CRSwNP (ECRSwNP) remains unclear. This study aims to explore the effects of VD on eosinophil chemotaxis in ECRSwNP and the underlying mechanisms. Human nasal mucosal tissues were collected from the control group, patients with non-ECRSwNP and those with ECRSwNP. Enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of VD and CCL26 in the nasal mucosa, plasma, or human primary nasal epithelial cells (hNECs). hNECs and eosinophils from patients were cultured to investigate the effect of VD on eosinophil chemotaxis and CCL26 expression via eosinophil migration assay, Western blot, and ELISA. Transcriptome sequencing, pathway enrichment analysis, Western blot and immunohistochemical staining were used to determine the key signaling pathway involved in eosinophil chemotaxis. A significant decrease in VD levels was observed in the nasal mucosa of patients with ECRSwNP, which correlated with increased local eosinophil infiltration. Furthermore, pathway enrichment analysis suggested that glycolysis signaling was promoted in the ECRSwNP group, verified by enhanced expression of glycolytic key enzymes that were positively correlated with eosinophil infiltration in nasal mucosa from patients with ECRSwNP. VD suppressed eosinophil chemotaxis in vitro by inhibiting CCL26 expression. Glycolysis regulated CCL26 expression via the ERK pathway and lactate, which promoted the expression and stability of CCL26 protein. VD attenuated glycolysis, leading to decreased production of lactate and inactivation of the ERK pathway. The decrease in lactate production suppressed eosinophil chemotaxis. Moreover, the ERK pathway activator reversed the inhibitory effect of VD on eosinophil chemotaxis. VD impedes eosinophil chemotaxis by inhibiting glycolysis - induced CCL26 expression via attenuating the activation of the ERK pathway and reducing lactate production. VD supplementation may be a novel strategy to treat ECRSwNP.

BACKGROUD:The recurrence rate of chronic rhinosinusitis with nasal polyps (CRSwNP) is positively correlated with eosinophil infiltration. Increased interleukin (IL)-19 and eosinophil chemokine RANTES levels have been reported in patients with CRSwNP. This study aimed to clarify the role of IL-19 in mediating RANTES expression and eosinophilic infiltration in eosinophilic CRSwNP (Eos CRSwNP).METHODS:Nasal tissue samples were obtained from patients with CRSwNP and controls. The expression of IL-19, its receptors, ECP, and RANTES in tissues was investigated. Primary human nasal epithelial cells (HNECs) and nasal polyp tissue blocks were cultured, then stimulated by IL-19; ERK phosphorylation, NF-κB pathway activation, RANTES level, eosinophils migration and infiltration were detected using RT-qPCR, ELISA, western blotting, HE, immunohistochemistry, immunofluorescence staining, confocal microscopy, and transwell migration assay.RESULTS:The expression of IL-19 and its receptors (IL-20R1/IL-20R2), eosinophil cationic protein, and RANTES in nasal tissues from patients with Eos CRSwNP was significantly increased compared to that in non-Eos CRSwNP and control subjects. IL-19 co-localized with RANTES in nasal tissues and significantly elevated RANTES expression in HNECs. IL-19-blocking antibody and siRNA knockdown of IL-20R1 ameliorated the effect of IL-19 on RANTES secretion in HNECs. Moreover, IL-19-induced RANTES upregulation was associated with the activation of the ERK and NF-κB pathways. NF-κB activation was mediated by the ERK pathway in IL-19-treated HNECs, and IL-19 enhanced eosinophil infiltration in nasal polyp tissue blocks.CONCLUSIONS:Our findings indicate that IL-19 promotes RANTES expression via the ERK/NF-κB pathway in HNECs and is implicated in eosinophil infiltration in patients with Eos CRSwNP.

BACKGROUND:Regulatory T (Treg) cells, which prevent inflammation-induced eosinophil infiltration, are deficient in nasal polyps (NPs) in patients with eosinophilic chronic rhinosinusitis (ECRS). It is concomitant with loss of Foxp3 after certain inflammatory stimuli. OBJECTIVE:We sought to determine the inflammatory cytokines involved in inducing the loss of Treg cells in NPs. METHODS:The abundance of cytokines in ECRS patients or mice were tested using ELISA, immunochemistry, immunofluorescence, quantitative reverse transcription PCR (qPCR), and/or flow cytometry. Expression of eosinophil cationic protein (ECP), CD4+ T cells, IL-4, and IL-17A and eosinophils in nasal mucosa of mouse model was investigated by immunochemistry, immunofluorescence, and hematoxylin and eosin staining. The percentage and death of induced Treg (iTreg) cells, source of IL-21 in NPs from ECRS and non-ECRS patients, and abundance of different systemic phenotypes of CD4+ T cells in a mouse model were studied by flow cytometry. Western blot analysis, scanning, and transmission electronic microscopy were used to detect pyroptosis of iTreg cells. RESULTS:IL-21 was highly expressed in nasal mucosa of ECRS patients and mice, causing pyroptosis and preventing development of iTreg cells in vitro. The elevated IL-21 in NPs from ECRS patients was mainly produced by CD3+ T cells, including T follicular helper, T peripheral helper, TH2, and TH17 cells and CD3+CD4- T cells. T peripheral helper cells and CD3+CD4- T cells were the predominant source of IL-21 in NPs from non-ECRS patients. Blocking IL-21/IL-21R signaling significantly reduced the number of eosinophils and CD4+ T cells along with ECP, IL-4, and IL-17A expression in the nasal mucosa of ECRS mice. It also increased Treg cell percentage and systemically decreased TH2 and TH17 ratios. Akt-mTOR inhibition prevented IL-21-induced pyroptosis in human and mouse iTreg cells. CONCLUSION:Elevated IL-21 drives pyroptosis and prevents Treg cell development in ECRS patients. IL-21 induced pyroptosis via activating Akt-mTOR-NLRP3-caspase 1 signaling.

BACKGROUND:Tissue remodeling is a prominent characteristic of chronic rhinosinusitis (CRS). Excess deposition of fibronectin (FN) and collagen (Col) I by fibroblasts is crucial for the pathologic tissue remodeling in CRS without nasal polyps (CRSsNP). Increased interleukin (IL)-19 level in patients with CRS had been demonstrated in our previous studies. Here, we aimed to evaluate the role of IL-19 in mediating FN and Col I expression in CRS.METHODS:Nasal mucosal tissue samples were collected from patients with CRS with nasal polyps (CRSwNP), CRSsNP, and controls. The expression of IL-19, vimentin, FN, and Col I were detected using immunohistochemistry and immunofluorescence. Primary human nasal fibroblasts were treated with IL-19, then the activation of Smad2/3, NF-κB and relevant pathways, and the expression of FN and Col I were measured.RESULTS:Expression levels of vimentin, FN, and Col I were significantly increased in nasal tissues from patients with CRSsNP compared with CRSwNP and control subjects. Moreover, IL-19 co-localized with FN and Col Ι in nasal tissues. IL-19-treated fibroblasts had increased production of FN and Col I, which was associated with the activated Smad2/3 and NF-κB pathways. Moreover, Smad2/3 activation was mediated by the NF-κB pathway in IL-19-treated fibroblasts.CONCLUSIONS:IL-19 promotes FN and Col I production via the activated NF-κB-Smad2/3 pathway in fibroblasts, leading to fibrosis and collagen deposition in patients with CRS.

Abstract Background Tissue remodeling is a crucial characteristic of chronic rhinosinusitis (CRS). Imbalance between matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) is crucial for the pathologic tissue remodeling in CRS. Elevation of interleukin (IL)‐19 or MMP‐9 levels in patients with CRS had been proven in previous studies. Here, we aimed to investigate the role of IL‐19 in mediating MMP‐9 expression in CRS. Methods Nasal tissue samples were collected from 45 individuals having chronic rhinosinusitis with nasal polyps (CRSwNP), 24 CRS without nasal polyps (CRSsNP), and 17 controls. Expression of IL‐19, its receptors (IL‐20R1/IL‐20R2), and MMP‐9 were investigated using RT‐qPCR and Immunofluorescence (IF). Human nasal epithelial cells (HNECs) were stimulated by IL‐19; ERK phosphorylation, nuclear factor‐κB (NF‐κB) pathway activation, and MMP‐9 level were detected by RT‐qPCR, enzyme‐linked immunosorbent assay, western blot, and IF. We also explored the effect of type1/2/3 cytokines on IL‐19 production by RT‐qPCR, and western blot. Results Expression levels of IL‐19, its receptors (IL‐20R1/IL‐20R2), and MMP‐9 were increased in nasal tissues from individuals with CRSwNP compared to those with CRSsNP as well as the controls. IL‐19 significantly elevated the production of MMP‐9 in HNECs. Furthermore, IL‐19 could activate the ERK and NF‐κB pathways, accompanied by increased MMP‐9 production in HNECs. Conversely, both ERK and NF‐κB inhibitors significantly attenuated the role of IL‐19 in MMP‐9 production. siRNA knockdown of IL‐20R1 suppressed ERK and NF‐κB pathway activation, thereby decreasing MMP‐9 expression. IL‐13 and IL‐17A were found to stimulate IL‐19 production in HNECs. Conclusion IL‐19, promoted by IL‐13 and IL‐17A, contributes to the upregulation of secretion of the tissue remodeling factor MMP‐9 in patients with CRS.

AllergyVolume 76, Issue 2 p. 593-596 LETTER TO THE EDITOR Treatment-emergent adverse events in dupilumab-treated patients with allergic diseases: A meta-analysis Xiaohong Chen, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorMenghui Liu, Department of Cardiology, The First Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorShuo Wu, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorZizhen Huang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorXia Li, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorXiaoping Lai, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorHongwei Bao, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorJiancong Huang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorLihong Chang, Corresponding Author changlihong19840719@126.com Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China Correspondence Lihong Chang and Gehua Zhang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China. Emails: changlihong19840719@126.com; zhanggeh@mail.sysu.edu.cnSearch for more papers by this authorGehua Zhang, Corresponding Author zhanggeh@mail.sysu.edu.cn Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China Correspondence Lihong Chang and Gehua Zhang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China. Emails: changlihong19840719@126.com; zhanggeh@mail.sysu.edu.cnSearch for more papers by this author Xiaohong Chen, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorMenghui Liu, Department of Cardiology, The First Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorShuo Wu, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorZizhen Huang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorXia Li, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorXiaoping Lai, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorHongwei Bao, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorJiancong Huang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, ChinaSearch for more papers by this authorLihong Chang, Corresponding Author changlihong19840719@126.com Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China Correspondence Lihong Chang and Gehua Zhang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China. Emails: changlihong19840719@126.com; zhanggeh@mail.sysu.edu.cnSearch for more papers by this authorGehua Zhang, Corresponding Author zhanggeh@mail.sysu.edu.cn Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China Correspondence Lihong Chang and Gehua Zhang, Department of Otorhinolaryngology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China. Emails: changlihong19840719@126.com; zhanggeh@mail.sysu.edu.cnSearch for more papers by this author First published: 05 October 2020 https://doi.org/10.1111/all.14610 Chen and Liu these authors contributed equally to this work. Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinked InRedditWechat Supporting Information Filename Description all14610-sup-0001-AppS1.docxWord document, 379.6 KB App S1 Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article. Volume76, Issue2China IssueFebruary 2021Pages 593-596 RelatedInformation

Type 2 inflammation and eosinophilic infiltration are prominent pathologic features of chronic rhinosinusitis with nasal polyps (CRSwNP). The purpose of the present study was to determine the roles of Tregs in controlling type 2 inflammation and inhibiting eosinophilic infiltration in CRSwNP. A total of 134 nasal polyps, 67 ostiomeatal complex from chronic rhinosinusitis (CRS) and 62 normal nasal tissues from controls were collected to study the enumeration and function of Tregs cells and the expressions of cytokine profiles via immunofluorescence staining, flow cytometry, qRT-PCR, ELISA, and/or H&E staining. The effects of Tregs on type2 and type3 inflammations were determined in an eosinophilic chronic sinusitis (ECRS) mice model. It was confirmed that the CRSwNP displayed the features of Th2 and Th17 cells-mediated inflammation, accompanying by an increased level of eosinophilic infiltration and the eosinophil cationic protein (ECP), with a decreased frequency of Treg cells. Furthermore, the percentages of CD4+CD25+CD127lowTreg and CD4+CD25+Foxp3+Treg were only decreased in the polyps of CRSwNP but not in the paired peripheral blood. The CRSwNP possessed the decreased Nrp1+Tregs, Helios+Treg, and low TGF-β and interleukin (IL)-10 expressions in Tregs. The ECRS mice showed similar inflammatory characteristics to CRSwNP patients. The adoptive transfer of CD4+CD25+Foxp3+ Treg cells significantly decreased the inflammatory cytokines, eosinophilic chemotactic factors in the mucosa of the ECRS mice without alteration of the immune balance in the peripheral blood and spleen. In conclusion, CRSwNP showed high type 2 and type3 inflammation and defective Tregs. The induced regulatory T cell (iTreg) may correct the imbalance between immune tolerance and effect via limiting the eosinophil recruitment of mucosa in CRSwNP.

Mice models were used to study the pathogenesis of mice and human diseases. Although some mice models of allergic rhinitis and rhinosinusitis have been reported, no detailed anatomic, histological and computed tomographic comparative data of the normal murine sinus are available in the literature for new researchers to establish mice models. The purpose of this study was to clarify the histological and computed tomographic characteristics of the normal nasal sinus in BALB/c mice. Fifteen sinonasal specimens were collected. Five mice were subjected to micro-computed tomography imaging, and then dissected to observe its anatomic landmarks, and 10 mice were subjected to haematoxylin and eosin staining. Important anatomic landmarks were clearly demonstrated, including the ethmoturbinates, nasoturbinal, maxilloturbinate, ethmoid sinus, maxillary sinus, nasopharyngeal duct, nasolacrimal duct and vomeronasal organ. Full and typical sinonasal landmarks can be visualized by gross anatomy, micro-computed tomography imaging and haematoxylin and eosin staining, which will be useful for establishing the mouse models of nasal disease.

PURPOSE:Budesonide improves the prognosis of chronic rhinosinusitis (CRS). However, few reports have examined whether its use for nasal irrigation, compared to normal saline, improves the prognosis of patients after endoscopic sinus surgery (ESS). We compared the effects of nasal irrigation with budesonide and normal saline in CRS patients after ESS.METHODS:Sixty CRS patients who had undergone ESS were randomly divided into an experimental group (30 patients), which used budesonide nasal irrigation, and a control group (30 patients), which used normal saline nasal irrigation. All patients received regular follow-up evaluations and were assessed via questionnaires, including the Lund-Kennedy endoscopic score (LKES), the symptom visual analog scale (VAS), the 22-item Sino-Nasal Outcome Test (SNOT-22), the Short-Form 36-Item Questionnaire (SF-36), the Self-Rating Anxiety Scale (SAS), the Self-Rating Depression Scale (SDS) and a side effects scale.RESULTS:Scores of polyposis, mucosal edema, secretions and total score of LKES; VAS scores of nasal blockage, hyposmia and rhinorrhea; and SNOT-22 results in both groups were significantly improved 3 months after ESS. Scores of polyposis, mucosal edema, secretions and scarring and total score of LKES in experimental group were significantly better than in control group 3 months after ESS. No significant differences were observed in SF-36, SAS or SDS before or 3 months after ESS within or between the two groups. The side effects of the two groups were not significantly different.CONCLUSIONS:Nasal irrigation improved the prognosis of CRS patients after ESS. Budesonide nasal irrigation had a better effect than normal saline nasal irrigation.

Eosinophilic chronic rhinosinusitis with nasal polyps (ECRS) is a condition linked with type 2 inflammation, poor treatment outcomes, and high recurrence tendency. Although γδT cells have been reported to induce type 2 immune responses and eosinophilic infiltration in several diseases, their role in ECRS has not been fully explored. We aimed to evaluate the association of γδT cells with the type 2 inflammatory profiles in ECRS. Nasal tissue samples obtained from patients with chronic rhinosinusitis with nasal polyps (CRSwNP) (51 eosinophilic and 48 non-eosinophilic), 50 patients with chronic rhinosinusitis without nasal polyps (CRSsNP), and 58 control subjects were examined for γδT cells, inflammatory markers and eosinophils using HE, RT-qPCR, ELISA, immunofluorescence, and flow cytometry. In parallel, studies were also conducted in an ECRS murine model induced by anti-γδT cells neutralizing antibody administration. γδT cells expression was significantly increased in tissues from patients with ECRS compared with non-ECRS, CRSsNP and control subjects. Moreover, inflammatory markers including type 2 proinflammatory cytokines (IL-4, IL-5, IL-13), GATA3, eosinophil cationic protein (ECP), and eotaxin levels were also increased in nasal tissues of patients with ECRS, and Vγ1+ γδT cells mRNA expression was positively correlated with type 2 cytokines, GATA3, and ECP. In the ECRS murine model, anti-Vγ1+ γδT antibody treatment reduced the infiltration of eosinophils and expression of type 2 cytokines, GATA3, and ECP in nasal mucosae. In conclusion, the results of the present study suggest that γδT cells play a crucial role in the type 2 inflammatory profiles and nasal tissue eosinophilic infiltration in patients with ECRS.

Chronic rhinosinusitis (CRS) is a common chronic inflammatory disease of the upper airways involving nasal cavity and sinus. Deriving both from its clinical complexity with protean clinical manifestations as well its pathogenetic heterogeneity, the molecular mechanisms contributing to the pathogenesis of CRS remain unclear, and attract a wide interest in the field. Current evidences indicate that IL-17A is highly expressed in chronic rhinosinusitis with nasal polyps (CRSwNP). However, its pathogenetic role in regulation of tissue remodeling of CRSwNP remains unknown. The present study aimed to investigate the cellular origins and functions of IL-17A cytokine in CRSwNP, and further determined whether IL-17A could affect the expression of metalloproteinases (MMPs), the remodeling factors of CRSwNP. The results showed that the expression of IL-17A was upregulated in nasal tissues of patients with CRSwNP compared to those with chronic rhinosinusitis without nasal polyps (CRSsNP) and controls. CD8+ cytotoxic T lymphocytes (Tc) were major IL-17A producers in nasal tissues of CRSwNP. Interleukin (IL)-17-producing CD8+ T cells (Tc17) was significantly higher in nasal tissues of CRSwNP than CRSsNP and controls. Nonetheless, no difference was observed among the IL-17A in peripheral blood lymphocytes of these three groups. Moreover, in the same patients, IL-17A expression was negligible in lymphocytes of peripheral blood when compared with nasal tissues. Increased gene and protein expression of MMP-7 and MMP-9 in patients with CRSwNP compared with controls were observed. In CRSwNP samples, IL-17A receptor (IL-17AR) co-localized with MMP-9 and they were mainly expressed in the epithelial cells. MMP-9 expression was up-regulated both in Primary human nasal epithelial cells (PHNECs) and a nasal epithelial cell line (RPMI 2650) by IL-17A treatment, and diminished by anti-IL-17AR treatment. Furthermore, IL-17A promoted the expression of MMP-9 by activating the NF-κB signal pathway. Thus, our results have revealed a crucial role of IL-17A and Tc cells on pathogenesis and tissue remodeling of CRSwNP.

[Objective]To construct miR-18a overexpression and inhibition lentivirus vectors and to determine their effects on human nasopharyngeal cancer(NPC)cell line CNE1 and CNE2.[Methods]Designed the primers for Real-time polymerase chain(PCR)reaction to obtain the miR-18a premature gene.The premature gene and the siRNA oligo-nucleutides of miR-18a were connected to the lentivirus vector GV369 and GV280,respectively.The construction vectors were confirmed by DNA sequencing.Then,293T cell was infected with the vectors plus Helper 1.0 and pHelper 2.0 vec-tors to obtain recombinant lentivirus vector for miR-18a overexpression and inhibition. The NPC cell line CNE1 and CNE2 were infected with the successful recombinant lentivirus vectors.Puromycin was added to select the positive infect-ed cells. PCR method was used to detect the miR-18a expression level after infecting the recombinant lentivirus vector into the NPC cell line.[Results]A recombinant lentivirus vector expressing miR-18a interference oligonucleutides was obtained and confirmed by DNA sequencing.The virus titer was 3×108TU/mL,and the expression of its target gene ATM was downregulated in CNE1 and CNE2.A recombinant lentivirus vector expressing miR-18a premature gene was obtained and confirmed by DNA sequencing. The virus titer was 3×108TU/mL,and the miR-18a was overexpressed in CNE1 (20.3 fold upregulation,P<0.01)and CNE2(122.5 fold upregulation,P<0.01),and its target gene ATM was downregu-lated.[Conclusions]The miR-18a overexpression and suppression lentivirus vectors are successfully constructed.These vec-tors could alter the expression level of miR-18a in NPC cell line significantly,and provide a stable cell line for functional studies in the future.

The increasing resistance of nasopharyngeal carcinoma to irradiation makes the exploration of effective radiosensitizers necessary. Tetrandrine is known to be an antitumor drug, but little is known regarding its radiosensitization effect on nasopharyngeal carcinoma. We investigated the effect of combined treatment of irradiation and maximum non-cytotoxic doses of tetrandrine on the nasopharyngeal carcinoma cell lines CNE1 and CNE2. The maximum non-cytotoxic doses of tetrandrine in CNE1 and CNE2 cells were assessed using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. The radiosensitization of cells receiving the maximum non-cytotoxic doses of tetrandrine was assessed by evaluating cell proliferation and DNA damage repair using MTT, clonogenic, comet assays and detection of caspase-3 and phosphorylated histone H2AX (γ-H2AX). The cell cycle was assessed by flow cytometry, and protein expression was detected by western blot analysis. The maximum non-cytotoxic doses of tetrandrine in CNE1 and CNE2 cells were 1.5 μmol/L and 1.8 μmol/L, respectively. When cells were exposed to irradiation and the maximum non-cytotoxic doses of tetrandrine, the survival fraction was decreased. DNA damage and γ-H2AX levels markedly increased. Moreover, tetrandrine abrogated the G2/M phase arrest caused by irradiation. Combined treatment with the maximum non-cytotoxic dose of tetrandrine and irradiation caused suppression of the phosphorylation of CDK1 and CDC25C and increase in the expression of cyclin B1. The study in vivo also showed that the maximum non-cytotoxic dose of tetrandrine could reduce tumor growth in xenograft tumor model. Our results suggest that the maximum non-cytotoxic dose of tetrandrine can enhance the radiosensitivity of CNE1 and CNE2 cells and that the underlying mechanism could be associated with abrogation of radiation-induced G2/M arrest via activation of the CDC25C/CDK1/Cyclin B1 pathway.

Background: This study aimed to establish an effective prognostic nomogram with or without plasma Epstein-Barr virus DNA (EBV DNA) for nondisseminated nasopharyngeal carcinoma (NPC).Methods: The nomogram was based on a retrospective study of 4630 patients who underwent radiotherapy with or without chemotherapy at Sun Yat-sen University Cancer Center from 2007 to 2009. The predictive accuracy and discriminative ability of the nomogram were determined by a concordance index (C-index) and calibration curve and were compared with EBV DNA and the current staging system. The results were validated using bootstrap resampling and a prospective cohort study on 1819 patients consecutively enrolled from 2011 to 2012 at the same institution. All statistical tests were two-sided.Results: Independent factors derived from multivariable analysis of the primary cohort to predict recurrence were age, sex, body mass index (BMI), T stage, N stage, plasma EBV DNA, pretreatment high sensitivity C-reactive protein (hs-CRP), lactate dehydrogenase (LDH), and hemoglobin level (HGB), which were all assembled into the nomogram with (nomogram B) or without EBV DNA (nomogram A). The calibration curve for the probability of recurrence showed that the nomogram-based predictions were in good agreement with actual observations. The C-index of nomogram B for predicting recurrence was 0.728 (P < .001), which was statistically higher than the C-index values for nomogram A (0.690), EBV DNA (0.680), and the current staging system (0.609). The C-index of nomogram B (0.730) and nomogram A (0.681) remained higher for predicting recurrence among patients treated with intensity-modulated radiotherapy (P < .001). The results were confirmed in the validation cohort.Conclusions: The proposed nomogram with or without plasma EBV DNA resulted in more accurate prognostic prediction for NPC patients.

PurposeThis study aimed to clarify the prognostic utility of high-sensitivity C-reactive protein (hs-CRP) in nasopharyngeal carcinoma (NPC) patients in the Intensity-Modulated Radiotherapy (IMRT) era.Patients and MethodsIn this observational study, 1,589 non-metastatic NPC patients treated with IMRT were recruited. Blood samples were collected before treatment for examination of hs-CRP levels. We evaluated the association of pretreatment hs-CRP levels with overall survival rate (OS), progression free survival rate (PFS), locoregional relapse free survival rate (LRFS) and distant metastasis free survival rate (DMFS).ResultsBaseline hs-CRP levels were correlated with sex, clinical stage, body mass index, smoking status, and EBV DNA level. Multivariate analysis showed that hs-CRP had significant association with OS (HR:1.723; 95% CI:1.238-2.398; p = 0.001), PFS (HR:1.621; 95% CI:1.273-2.064; p < 0.001) and DMFS (HR: 1.879; 95% CI: 1.394-2.531; p < 0.001). In subgroups such as advanced-stage group, low EBV DNA group and high EBV DNA group, elevated hs-CRP levels still predicted poor clinical outcomes. Furthermore, in patients with chronic HBV infection, decreased 4-year survival was observed in the cohort of high hs-CRP levels, with 87.4% vs. 94.9% (p = 0.023) for OS, 65.2% vs. 90.8% (p < 0.001) for PFS, and 67.6% vs. 95.0% (p < 0.001) for DMFS. A similar finding was observed for patients with cardiovascular disease, with 79.1% vs. 90.2% (p = 0.020) for PFS, and 71.4% vs. 97.6% (p = 0.002) for DMFS.ConclusionElevated serum hs-CRP levels were correlated with poor survival for NPC patients in the IMRT era, playing a complementary role to TNM stage and EBV DNA. In addition, elevated hs-CRP level was still an effective indicator for patients with chronic HBV infection and cardiovascular disease.

Purpose: To evaluate the effects of combining the assessment of circulating high-sensitivity C-reactive protein (hs-CRP) with that of Epstein-Barr virus DNA (EBV DNA) in the pretherapy prognostication of nasopharyngeal carcinoma (NPC).Patients and Methods: Three independent cohorts of NPC patients (training set of n=3113, internal validation set of n=1556, and prospective validation set of n=1668) were studied. Determinants of disease-free survival, distant metastasis -free survival, and overall survival were assessed by multivariate analysis. Hazard ratios and survival probabilities of the patient groups, segregated by clinical stage (T1-2N0-1M0, T3-4N0-1M0, T1-2N2-3M0, and T3-4N2-3M0) and EBV DNA load (low or high) alone, and also according to hs-CRP level (low or high), were compared.Results: Elevated hs-CRP and EBV DNA levels were significantly correlated with poor disease-free survival, distant metastasisefree survival, and overall survival in both the training and validation sets. Associations were similar and remained significant after excluding patients with cardiovascular disease, diabetes, and chronic hepatitis B. Patients with advanced-stage disease were segregated by high EBV DNA levels and high hs-CRP level into a poorest-risk group, and participants with either high EBV DNA but low hs-CRP level or high hs-CRP but low EBV DNA values had poorer survival compared with the bottom values for both biomarkers. These findings demonstrate a significant improvement in the prognostic ability of conventional advanced NPC staging.Conclusion: Baseline plasma EBV DNA and serum hs-CRP levels were significantly correlated with survival in NPC patients. The combined interpretation of EBV DNA with hs-CRP levels led to refinement of the risks for the patient subsets, with improved risk discrimination in patients with advanced-stage disease. (C) 2015 Elsevier Inc.