ABSTRACT
Aberrant substance P/neurokinin-1 receptor (SP/NK-1R) system activation plays a critical role in various disorders, however, little is known about the expression and the detailed molecular mechanism of the SP and NK-1R in gallbladder cancer (GBC). In this study, we firstly analyzed the expression and clinical significance of them in patients with GBC. Then, cellular assays were performed to clarify their biological role in GBC cells. Moreover, we investigated the molecular mechanisms regulated by SP/NK-1R. Meanwhile, mice xenografted with human GBC cells were analyzed regarding the effects of SP/NK1R complex in vivo. Finally, patient samples were utilized to investigate the effect of SP/NK-1R. The results showed that SP and NK-1R were highly expressed in GBC. We found that SP strongly induced GBC cell proliferation, clone formation, migration and invasion, whereas antagonizing NK-1R resulted in the opposite effects. Moreover, SP significantly enhanced the expression of NF-κB p65 and the tumor-associated cytokines, while, Akt inhibitor could reverse these effects. Further studies indicated that decreasing activation of NF-κB or Akt diminished GBC cell proliferation and migration. In consistent with results, immunohistochemical staining showed high levels of Akt, NF-κB and cytokines in tumor tissues. Most importantly, the similar conclusion was obtained in xenograft mouse model. Our findings demonstrate that NK-1R, after binding with the endogenous agonist SP, could induce GBC cell migration and spreading via modulation of Akt/NF-κB pathway.
Subject(s)
Cell Movement/drug effects , Cell Proliferation/drug effects , Gallbladder Neoplasms/metabolism , Receptors, Neurokinin-1/metabolism , Substance P/pharmacology , Animals , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Gallbladder Neoplasms/genetics , Humans , Interleukin-1beta/metabolism , Interleukin-6/metabolism , Matrix Metalloproteinase 9/metabolism , Mice , Mice, Nude , Proto-Oncogene Proteins c-akt/metabolism , RNA, Small Interfering , Receptors, Neurokinin-1/genetics , Signal Transduction/drug effects , Signal Transduction/genetics , Transcription Factor RelA/metabolism , Transplantation, Heterologous , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Transient receptor potential canonical channel 3 (TRPC3) proteins function as non-voltage-gated Ca2+ -permeable channels and play divergent roles in many processes of pathophysiology. The purpose of this study was to determine the relationship between TRPC3 expression and airway hyperresponsiveness and remodeling in ovalbumin-induced asthmatic Kunming mice. Mice were sensitized and challenged by ovalbumin to establish asthmatic model. Hematoxylin-eosin staining, hydroxyproline assay, and isometric tracheal ring force measurement were used to evaluate airway remodeling and hyperresponsiveness in asthmatic mice. Western blot was performed to detect the expression of TRPC3 proteins. MTT assay was used to measure the proliferation of airway smooth muscle cells. TRPC3 protein expression increased in airway smooth muscle of asthmatic mice. GdCl3 , a nonspecific TRPC blocker, attenuated the contractile force of airway smooth muscle. Fetal bovine serum stimulated airway smooth muscle cells proliferation and augmented TRPC3 protein expression. Both TRPC3 blockade by GdCl3 or specific TRPC3 antibodies and gene silencing by siRNA inhibited the proliferation of airway smooth muscle cells. In contrast, the current drugs treatment for asthma such as Dexamethasone and Aminophylline had no effects on TRPC3 protein overexpression. Therefore, TRPC3 protein overexpression may be involved in airway smooth muscle hyperresponsiveness and remodeling in asthmatic mice, providing evidence for a new direction of asthma pathogenesis research and a new target for drug intervention.
Subject(s)
Airway Remodeling , Asthma/etiology , Ovalbumin/immunology , TRPC Cation Channels/metabolism , Acetylcholine/pharmacology , Airway Remodeling/drug effects , Aminophylline/pharmacology , Aminophylline/therapeutic use , Animals , Antibodies/immunology , Asthma/drug therapy , Asthma/veterinary , Calcium/metabolism , Cell Proliferation/drug effects , Cells, Cultured , Dexamethasone/pharmacology , Dexamethasone/therapeutic use , Gadolinium/pharmacology , Hydroxyproline/metabolism , Mice , Muscle Contraction/drug effects , Muscle, Smooth/cytology , Muscle, Smooth/metabolism , Protein Biosynthesis/drug effects , RNA Interference , RNA, Small Interfering/metabolism , TRPC Cation Channels/antagonists & inhibitors , TRPC Cation Channels/geneticsABSTRACT
AIMS: The transient receptor potential canonical channel 1 (TRPC1) is a crucial component of the stretch-activated ion channels (SACs). The objective of this research was to demonstrate the contribution of TRPC1 in maintaining cardiac contractile function in the hypertrophic myocardium. METHODS: Hypertrophic rat hearts were induced by injecting isoproterenol intraperitoneally, and the expressions of TRPC1/3/6 and Na(+)/Ca(2+) exchanger 1 (NCX1) proteins were analyzed by Western blot. The intracellular calcium images, the action potential of myocardium, the length-dependent contractile force of ventricle muscle and the cardiac output of isolated heart were investigated. RESULTS: The expression of TRPC1 was increased in the hypertrophic myocardium. After being stretched, the ascendant amplitude of the increase in the intracellular calcium ion concentration ([Ca(2+)]i) in the hypertrophic myocardium was higher than that in the normal myocardium. The increase of the APD50 and the amplitude of the membrane potential depolarization were more significant in the hypertrophic myocardium after the activation of SACs. When the heart preparations were perfused with Tyrode's solution, there was no difference in the cardiac systolic function between the cardiac hypertrophy group and the control group. Gadolinium, a SACs blocker, reduced the length-dependent contractile force and suppressed the ascending limb of the Frank-Starling curves in the hypertrophic heart. CONCLUSIONS: The upregulation of TRPC1 contributes to the contractile function in the hypertrophic myocardium by increasing [Ca(2+)]i through the SACs.
