Fei Wang1, Shushang Liu2, Jinqiang Liu3, Fan Feng2, Yinghao Guo4, Wenming Zhang5, Gaozan Zheng2, Qiao Wang6, Lei Cai2, Man Guo2, Xiao Lian2, Guanghui Xu7, Hongwei Zhang8. 1. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China; Department of General Surgery, NO. 534 Hospital of PLA, Luoyang, Henan, China. 2. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China. 3. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China; Cadre' s sanitarium, 62101 Army of PLA, 67 Nahu Road, 464000, Xinyang, Henan, China. 4. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China; Health company, 92667 Army of PLA, 39 East Zaoshan Road, 266100, Qingdao, Shandong, China. 5. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China; Department of General surgery, Linxia People's Hospital, 731100, Linxia, Gansu, China. 6. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China; Department of General Surgery, NO. 91 Central Hospital of PLA, Jiaozuo, Henan, China. 7. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China. Electronic address: xuguanghui8@126.com. 8. Division of Digestive Surgery, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, Shaanxi, China. Electronic address: zhanghwfmmu@126.com.
Abstract
BACKGROUND: Substance P (SP) plays an important role in several types of cancer promotion and progression by binding to its preferential neurokinin 1 receptor (NK1R). However, the clinical significance and downstream mechanism of NK1R in esophageal squamous cell carcinoma (ESCC) have not been elucidated. The aim of this study was to investigate the role of SP/NK1R in the proliferation of ESCC and to screen related downstream molecules. METHODS: In the current investigation, the expression of NK1R was detected via immunohistochemistry (IHC), western blot (WB) analysis and real-time reverse transcription-polymerase chain reaction (RT-qPCR) in ESCC tissues and cell lines. Thereafter, the optimal concentration of SP was determined in vitro. The proliferation ability of SP/NK1R was assessed by cell counting kit-8 (CCK-8) and colony formation assays and subcutaneous tumour formation in nude mice with EC109 cells. Moreover, the related downstream molecules were screened by performing isobaric tags for relative and absolute quantitation (iTRAQ) protein spectrum analysis. RESULTS: NK1R was upregulated in ESCC, and its overexpression correlated with larger tumour size, deeper tumour invasion, more perineural invasion and eventually caused poorer overall survival (OS). Both intrinsic and SP-activated NK1R upregulation could promote the proliferation and clonogenic capacity of ESCC cells. In nude mice, tumour growth was suppressed by EC109 cells of NK1R downregulation. Further experiments demonstrated that Hairy and Enhancer of Split 1 (Hes1) was markedly reduced upon NK1R downregulation in EC109 cell lines and could regulate cell proliferation in the downstream of SP/NK1R. CONCLUSIONS: The significant role of NK1R in mediating ESCC cell proliferation depended on the activation of SP and might be related to the downstream regulation of Hes1.
BACKGROUND:Substance P (SP) plays an important role in several types of cancer promotion and progression by binding to its preferential neurokinin 1 receptor (NK1R). However, the clinical significance and downstream mechanism of NK1R in esophageal squamous cell carcinoma (ESCC) have not been elucidated. The aim of this study was to investigate the role of SP/NK1R in the proliferation of ESCC and to screen related downstream molecules. METHODS: In the current investigation, the expression of NK1R was detected via immunohistochemistry (IHC), western blot (WB) analysis and real-time reverse transcription-polymerase chain reaction (RT-qPCR) in ESCC tissues and cell lines. Thereafter, the optimal concentration of SP was determined in vitro. The proliferation ability of SP/NK1R was assessed by cell counting kit-8 (CCK-8) and colony formation assays and subcutaneous tumour formation in nude mice with EC109 cells. Moreover, the related downstream molecules were screened by performing isobaric tags for relative and absolute quantitation (iTRAQ) protein spectrum analysis. RESULTS:NK1R was upregulated in ESCC, and its overexpression correlated with larger tumour size, deeper tumour invasion, more perineural invasion and eventually caused poorer overall survival (OS). Both intrinsic and SP-activated NK1R upregulation could promote the proliferation and clonogenic capacity of ESCC cells. In nude mice, tumour growth was suppressed by EC109 cells of NK1R downregulation. Further experiments demonstrated that Hairy and Enhancer of Split 1 (Hes1) was markedly reduced upon NK1R downregulation in EC109 cell lines and could regulate cell proliferation in the downstream of SP/NK1R. CONCLUSIONS: The significant role of NK1R in mediating ESCC cell proliferation depended on the activation of SP and might be related to the downstream regulation of Hes1.