| Laryngeal cancer is a common head and neck cancer,accounting for 1-5% of systemic malignancies.The incidence rate of Laryngeal cancer is low in the body malignancy,but high in the respiratory tract and head and neck cancer.In the respiratory tract cancer,it accounts for the second place after lung cancer,and accounts for 2-5 in the head and neck tumors.Laryngeal cancer often happens at 40 or older age groups,of which the male to female ratio is about 10:1 in the worldwide.Men and women incidence difference was not significant in some areas,such as northeast China.Although the current epidemiological survey shows that smoking and drinking both are high risk factors for the incidence of laryngeal cancer,the specific pathogenesis is a complex biological process.Like most tumors,laryngeal cancer maybe caused by oncogene activation and suppressor gene inactivation,and many internal factors can promote the occurrence and development of laryngeal cancer,such as inflammation,the body ion channel mutation.There are some common clinical manifestations of laryngeal manifestations,such as hoarseness,difficulty breathing or swallowing difficulties.Neck mass can be found caused by cervical lymph node metastasis.Squamous cell carcinoma is the most of the pathological types of laryngeal.At present,the treatment of laryngeal cancer is based on surgery,combined with the comprehensive treatment of radiotherapy and chemotherapy.Although the treatment of laryngeal cancer has experienced more than 100 years,with non-stop progress of basic research forlaryngeal cancer incidence and treatment,the five-year survival rate and postoperative quality of life of patients with laryngeal cancer have not improved significantly.As a Ca2+-permeable non-selective cation channel,TRP(transient receptor potential)plays a significant role in Ca2+ signaling.TRP channel is generally divided into two categories,the first category including TRPA,TRPC,TRPN,TRPV.the second category including TRPP and TRPML.They have a similar structure which has an embedded card channel.TRP channel is widely distributed,regulating the body's mechanical sensation,pain,taste,muscle contraction cell proliferation,differentiation,apoptosis and many other life activities mainly through participation in calcium entry channels.However,the role of various TRP channels is not exactly the same,and some TRP channel subclasses can coordinate life activities with each other or synergistically.TRPP2 channel as a member of the TRP superfamily,also known as PKD2(polycystin-2),the current study shows indeed evidence that TRPP2 mutation and the incidence of polycystic kidney disease is closely related.TRPP2 is widely expressed in a variety of tissues,such as vascular endothelial smooth muscle cells,epithelial cells,cardiomyocytes,adrenal and ovarian cells.The present study suggests that subcellular localization of TRPP2 is controversial and is thought to be expressed in the cytoplasmic membrane,Golgi apparatus,endoplasmic reticulum and fibril,and the localization of endoplasmic reticulum is the most recognized.In the endoplasmic reticulum,TRPP2 acts as a channel for intracellular calcium release,which combines the inositol triphosphate inositol receptor and the sarcoside receptor to mediate intracellular calcium balance,involved in the SOCE(store-operated calcium entry channels,SOCE),playinga variety of physiological functions,such as cell proliferation,apoptosis,selfphagocytosis and other processes.Tumor necrosis factor ?(TNF-?)is one of the strongest anti-tumor factor,which was originally found.TNF-? is a multidirectional cytokine that plays an important role in inflammatory response,cell differentiation,immunoregulation,and anti-infectivity and anti-tumor.However,studies have shown that the effect of TNF-? on the tumor can produce different results due to different conditions,although the strongest anti-tumor factor,in some cases,it can also be expressed as the promotion of tumor development.Bioinformatics is an interdisciplinary subject widely used in clinical and basic research in recent years.It combines a variety of techniques such as informatics,computer science,statistics and so on.It can analyze large number of biological data from molecular level through sequence comparison,pathway analysis,Clustering,statistical analysis and other analysis to found that the abnormal information of the disease.It can be used as a first study for further confirmation study to avoid blindness of the study.In the present study,we used bioinformatics,western blot,flow cytometry,proliferation assay,Ca2+ image and small interfering(si)RNA technique to demonstrate that TNF-? enhanced the proliferation of Hep-2 cells(a cell line called human laryngeal carcinoma)through suppressing TRPP2 expression and TRPP2-mediated Ca2+ release from the ER.To further elucidate the role of TRPP2 and TNF-? in the proliferation of Hep-2 cells and to provide a new basis for the mechanism,it is helpful to elucidate the development and progression of laryngeal carcinoma,develop a new therapeutic direction for laryngeal cancer and provide laboratory evidence for laryngeal cancer.1.Construction and analysis of m RNA expression profile of TNF-? on Hep-2 cells Previous studies have also found that the role of TNF-? on the tumor is twofold,because the different conditions can be expressed as the promotion or inhibition.To investigate the effect of tumor necrosis factor-? on human laryngeal squamous cell carcinoma Hep-2 cells,we used m RNA sequencing technology and bioinformatics analysis to reveal the effect of TNF-? on human laryngeal squamous cell carcinoma cells treated with TNF-?.It was revealed that PKD2 m RNA was down-regulated in Hep-2 cells treated with TNF-?(compared with the control group).It is speculated that the TNF-? maybe involve in regulating Hep-2 cell proliferation by down-regulate TRPP2.the conclusion need further study to confirm.2.TNF-? accelerates Hep-2 cells proliferation via suppressing TRPP-2 expression TNF-? and TRPP2 are involved in a variety of functions of the body cells,and studies have shown that calcium channels play an important role in the development of tumors.TRPP2 as a member of the calcium channel,we speculate that it plays an important role in proliferation and development of laryngeal carcinoma.In the first part of the study,we found that the expression of TRPP2-related m RNA was significantly reduced in human laryngeal squamous cell carcinoma cells treated with TNF-?compared with the untreated human laryngeal squamous cell carcinoma cells in the m RNA expression profile.To further confirm whether TRPP2 is involved in the process of TNF-? on human laryngeal squamous cell carcinoma Hep-2 cells,we have conducted further studies.In the present study,we demonstrated that TNF-? treatment enhanced Hep-2 cell proliferation via PERK-e IF2? signaling pathway and possibly by suppressing TRPP2 expression and TRPP2-mediated Ca2+ signaling.The major findings are as follows:(1)TNF-? treatment significantly decreased TRPP2 expression and ATP-induced Ca2+ release in Hep-2 cells.(2)Knockdown of TRPP2 by TRPP2 specific si RNA significantly suppressed ATP-induced Ca2+ release and removed TNF-?-caused reduction of ATP-induced Ca2+ release.(3)TNF-? treatment and/or knockdown of TRPP2 did not affect IP3R-I expression level and basal [Ca2+]i of Hep-2cells.But TNF-? treatment and knockdown of TRPP2 had a synergistic effect to suppress TRPP2 expression.(4)TNF-? treatment enhanced the proliferation and the percentage of Hep-2 cells in G2/M phase,but decreased the percentage of Hep-2 cells in G0/G1 phase.TRPP2 si RNA transfection also increased the proliferation the percentage of Hep-2 cells in S and G2/M phases,but decreased the percentage of Hep-2 cells in G0/G1 phase.(5)TNF-? significantly down-regulated p-PERK and p-e IF2? expression levels,but did not affect PERK and e IF2? expression levels. |
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