The Expression Of TRPC1Protein In Nasopharyngeal Carcinoma And Its Clinical Significance

Background and objective:nasopharyngeal carcinoma is one of the most common Head and neck cancer in China, which is highly prevalent in Southeast Asia and southern China. the incidence rate of NPC In southern Chinese and immigrantsfrom southern China was25-30times higher than in other parts of Europe and America. Tumor formation and metastasis is a complex and continuous process that needs multi-step, multi-stage,multi-factor, such as environmental factors, genetic susceptibility and Epstein-Barr virus. owing to the deep location of nasopharynx and vague symptoms of the disease, approximately70%of patients diagnosed with NPC would present with stage Ⅲ or Ⅳ.Contrast to other head and neck cancer,nasopharyngeal carcinoma is prone to invade and metastasis, because of poorly differentiation and abundant peripheral lymphoid tissues. Radiation therapy is the preferred treatment of NPC, but the rate of local recurrence and distant metastasis was very high and five-year survival rate after radiotherapy is as low as50%. Therefore, improvement in early diagnosis, seeking new therapeutic targets and new prognosis indicators is the most promising way forward in NPC research.Transient receptor potential (TRP) channels is a non-selective cation channel superfamily, first identified in Drosophila species, which were initially identified in Drosophila melanogaster and named after their role in phototransduction:Fruit flies carrying mutations in their TRP gene exhibited a transient voltage response of their photoreceptors to continuous light. Most of these proteins display a putative topology of six transmembrane domains with a pore loop between the fifth and sixth segment. Both C-and Ntermini are presumably located intracellularly. Very similar primary structures are typical for other ion channels such as voltage-gated channels and the cyclic-nucleotide-gated channels. On the bases of homology and channel function, the TRP family is divided into seven main subfamilies:TRPA(anktmi). TRPC(canonical TRP). TRPM(melastatin, long-TRP)、TRPML(mucolipin)、TRPN(nompc)、 TRPP(polycystin)、TRPV(vanilloid).Transient receptor potential canonical1(TRPC1) is the first cloned mammalian TRP channels、which shares about34%amino acid identity with Drosophila TRP. TRPC1genes are on human chromosome3q22-q24and mouse chromosome9. The molecular massis of full-length human TRPC1is91kDa, and its amino acid sequence appears highly conserved in rat, mouse, rabbit, cow and human.TRPC1is widely expressed in brain, heart, lung, ovary, testis, intestine, kidney, skeletal muscle, placenta, prostate, spleen, salivary glands. TRPC1recognized function is involved in activation of phospholipase C(PLC)-mediated after of calcium ions to enter. TRPC1, as a core component of store-operated calcium channels, contributes to changes intracellular Ca2+concentrations which controls these cellular processes, including proliferation, apoptosis, gene transcription, and angiogenesis. Cytoplasmic Ca2+concentration [Ca2+]i (intracellular Ca2+concentration) control a variety of cellular functions, but is also a key process of various signal transduction receptor activation. Ca2+homeostasis mechanisms, in which the Ca2+entry pathways play a key role, controls these cellular processes, including proliferation, apoptosis, migration,gene transcription, angiogenesis, contraction and secretion. TRP (Transient Receptor Potential) channels contribute to changes in intracellular Ca2+concentrations, either by acting as Ca2+entry pathways in the plasma membrane or via changes in membrane polarization, modulating the driving force for Ca2+entry mediated by alternative pathways. In addition, TRP channels are expressed onthe membranes of internal Ca2+stores, where they may act as triggers for enhanced proliferation, aberrant differentiation, and impaired ability to die, leading to the uncontrolled expansion and invasion characteristic of cancer. TRPC1protein,as one member of TRP, also contribute to changes in intracellular Ca2+concentrations. suppression of TRPC1reduces Ca2+influx, on contrary, TRPC1overexpression amplifies Ca2+influx with consequent the destruction of calcium homeostasis, thus contributing to aberrant differentiation and proliferation.Several studies report overexpression of TRPC1induced the proliferation of airway smooth muscle cells, these data show TRPC1plays an important role in asthma and chronic obstructive pulmonary disease (COPD), suggesting TRPC1become the potential to prevent peripheral airway fibrosistargets. TRPC1protein expression was upregulated in ischemic brain injury. Yassine El Hiani et have reported that TRPC1channels are expressed in MCF-7cells and mediate Ca2+influx in response to high[Ca2+]o stimulation, they also have found TRPC1is required for Ca2+entry, ERK1/2phosphorylation, and CaR-proliferative effect in MCF-7cell.Treatment with2-aminoethoxydiphenyl borate (2-APB), and SKF-96365or by siTRPC1reduced the Ca2+entry induced by CaR activation. These data suggest TRPC1play an in important role in [Ca2+] i and cell proliferation. Benjamin Beck et have showed TRPC1and TRPC4channels are key elements in keratinocyte Ca2+homeostasis and differentiation and their loss was related to the failure of BCC cells to differentiate. Vanden Abeele et have showed The TRPC1and TRPV6are the most likely molecular candidates for the formation of prostate-specific endogenous SOCs and these poreins reduced when LNCaP cells differentiate to an androgen-insensitive, apoptotic-resistant neuroendocrine phenotype. Valerie C. Bomben et have reported TRPC1channel is essential for chemotactic migration of human malignant gliomas, also demonstrated Disruption of TRPC1causes incomplete cytokinesis and slows the growth of human malignant gliomas.So far, there are rarely studies about TRPC1protein expression in tumor tissues.This research intends to test TRPC1distribution and expression in human common tumor tissues and adjacent tissues by using tissue chip technology and immunohistochemical(IHC) method, investigate the differrence of TRPC1expression in NPC and non-NPC(chronic nasopharyngitis),and analyze the relation TRPC1expression in NPC patients with their clinical pathologicalfeatures and the relationship between TRPC1expression and over survival time of NPC patients retrospectively. ALLcould provide the preliminary theory basis for the diagnosis, prognosis and treatment of NPC.Methods:1. The expression of TRPC1protein in human common tumor tissues and adjacent tissuesUsing tissue chip technology and immunohistochemical method, TRPC1distribution and expression were detected in human common tumor tissues and adjacent tissues.2. The expression of TRPC1protein in NPC tissues and chronic nasopharyngitis tissues and its clinical significance in NPC Using immunohistochemical method, the expression of TRPC1protein was detected in138cases NPC tissues and24cases chronic nasopharyngitis tissues. the relation of TRPC1protein expression with NPC patients’clinical pathological parameters were analyzed. Survival analysis was used to analyze the relationship between TRPC1expression and over survival time of NPC patients retrospectively. COX analysis was used to explore the effect of variables on NPC patient’s over survival time.3. Statistical methods:All statistical analyses were performed using SPSS13.0software, the differrence of TRPC1protein expression and intensity in NPC tissues and chronic nasopharyngitis tissues were analyzed by Chi-square test and Wilcoxon rank sum test respectively. And the relationships of TRPC1protein expression with NPC patients’ clinical pathological parameters were analyzed by Chi-square test. Kaplan-Meier method was used to analyze the relationship between TRPC1expression and over survival time of NPC patients retrospectively.COX analysis was used to explore the effect of variables on NPC patient’s over survival time.were considered as significant when P value was less than0.05.Results:1. The expression of TRPC1protein in human common tumor tissues and adjacent tissuesTRPC1protein is widely expressed in the adjacent normal tissue of the breast, esophagus, stomach, liver, lung, pharynx, larynx, pancreas, and also expressed in the corresponding tumor tissue. TRPC1protein expression is strong in breast cancer, esophageal squamous cell carcinoma, rectum adenocarcinoma, hepatocellular carcinoma, renal cell carcinoma, ovarian serous adenocarcinoma. pharyngeal squamous cell carcinoma, laryngeal squamous cell carcinoma, and nasopharyngeal carcinoma, moderate in colon adenocarcinoma, lung squamous cell carcinoma, gastric cancer and prostate cancer tissue, and low in cervical cancer and pancreatic cancer.2. The difference of TRPC1protein expression in NPC tissues and chronic nasopharyngitis tissuesThe expression of TRPC1protein was found in cytoplasm. The positive rate of TRPC1protein in NPC tissues was84.1%(116/138),the high-expression rate was52.2%(72/138).The proportion of differentlevel in NPCtissues were as followed:"-"15.9%(22/138),"+"31.9%(44/138),"++"31.9%(44/138),"+++"20.3%(28/138). While The positive rate of TRPC1protein in chronic nasopharyngitis tissues was75.0%(18/24),the high-expression rate was20.8%(5/24).The proportion of different level in NPCtissues were as followed:"-"25.0%(6/24),"+"48.1%(13/24),"++"12.5%(3/24),"+++"8.3%(2/24). The positive rate and high-expression rate of TRPC1protein in NPC tissues were significantly higher than chronic nasopharyngitis tissues(P<0.05).3. The clinical significance of TRPC1protein expression in NPCThe TRPC1protein expression correlated closely to intracranial invasion,T stage, N stage and TNM stage(P value:0.001,0.004,0.049,0.013), while not correlated to gender, age, histological type and the status of EBV-IgG(P value:0.155,0.423,0.647,0.811).The median over survival time of NPC patients with high-expression TRPC1protein was50.0months, while the NPC patients with low-expression was86.0months. The median over survival time of NPC patients with high-expression TRPC1protein was significantly lower than NPC patients with low-expression (P=0.000). Kaplan-Meier method was used to identify the variables of potential prognostic significance in NPC patients. Gender, age, histological type and the status of EBV-IgG did not affect the over survival time of NPC patients(Pvalue:0.417,0.401,0.785,0.837). TRPC1protein expression, intracranial invasion,T stage, N stage and TNM stage were found affect the over survival time of NPC patients(Pvalue<0.05). M stage, intracranial invasion and TRPC1protein expression were Independent risk factors for overall survival of patients with nasopharyngeal carcinoma(Pvalue<0.05).Conclusions:1. TRPC1protein is widely expressed in normal tissues, and upregulated in tumor tissues, suggests that TRPC1may involved in the development and progress of tumor.2. The positive rate and high-expression rate of TRPC1protein in NPC tissues were significantly higher than chronic nasopharyngitis tissues. TRPC1protein were detected highly expressed in NPC tissues, while low expressed in chronic nasopharyngitis tissues. TRPC1protein may be used as a reference valuable biomarker on NPC diagnosis and prognosis.3. TRPC1protein expression correlated closely with intracranial invasion, T stage, N stage and TNM stage.The median over survival time of NPC patients with high-expression TRPC1protein was significantly lower than NPC patients with low-expression. Thus, TRPC1protein may be the potential prognosis biomarker and pharmaceutical targets of NPC.