RESUMEN
Triple-negative breast cancer (TNBC) is an aggressive breast tumor subtype characterized by poor clinical outcome. In recent years, numerous advancements have been made to better understand the biological landscape of TNBC, though appropriate targets still remain to be determined. In the present study, we have determined that the expression levels of FGF2 and S100A4 are higher in TNBC with respect to non-TNBC patients when analyzing "The Invasive Breast Cancer Cohort of The Cancer Genome Atlas" (TCGA) dataset. In addition, we have found that the gene expression of FGF2 is positively correlated with S100A4 in TNBC samples. Performing quantitative PCR, Western blot, CRISPR/Cas9 genome editing, promoter studies, immunofluorescence analysis, subcellular fractionation studies, and ChIP assays, we have also demonstrated that FGF2 induces in TNBC cells the upregulation and secretion of S100A4 via FGFR1, along with the ERK1/2-AKT-c-Rel transduction signaling. Using conditioned medium from TNBC cells stimulated with FGF2, we have also ascertained that the paracrine activation of the S100A4/RAGE pathway triggers angiogenic effects in vascular endothelial cells (HUVECs) and promotes the migration of cancer-associated fibroblasts (CAFs). Collectively, our data provide novel insights into the action of the FGF2/FGFR1 axis through S100A4 toward stimulatory effects elicited in TNBC cells.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/fisiología , Proteínas de Neoplasias/fisiología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/fisiología , Proteína de Unión al Calcio S100A4/fisiología , Transducción de Señal/fisiología , Neoplasias de la Mama Triple Negativas/fisiopatología , Antígenos de Neoplasias/fisiología , Movimiento Celular/efectos de los fármacos , Medios de Cultivo Condicionados/farmacología , Femenino , Factor 2 de Crecimiento de Fibroblastos/farmacología , Fibroblastos/patología , Regulación Neoplásica de la Expresión Génica/fisiología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Proteínas Quinasas Activadas por Mitógenos/fisiología , Neovascularización Patológica/fisiopatología , Comunicación Paracrina , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-rel/fisiología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/antagonistas & inhibidores , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Transducción de Señal/efectos de los fármacos , Neoplasias de la Mama Triple Negativas/irrigación sanguínea , Células Tumorales CultivadasRESUMEN
Background Carotid plaques with expansive arterial remodeling are closely related to cerebral ischemic events. Although S100A4 (S100 calcium-binding protein A4) is expressed in atherosclerotic lesions, its role in atherosclerotic plaque progression remains unknown. In this study, we examined the association between carotid arterial expansive remodeling and S100A4 expression. Methods and Results Preoperative high-resolution magnetic resonance imaging was used to assess luminal stenosis and vascular remodeling in patients undergoing carotid endarterectomy. To examine murine carotid atherosclerosis, we induced experimental lesions by flow cessation in apolipoprotein E-deficient mice fed a high-fat diet. The role of S100A4 in plaque formation and smooth muscle cell proliferation was investigated in vivo and in vitro, respectively. Human carotid arterial expansive remodeling showed positive correlations with the expression of S100A4, MMP2, and MMP9. S100A4 mRNA levels were positively correlated with those of MMP2, MMP9, and MMP13. S100A4 was expressed in vascular smooth muscle cells (VSMCs) and VSMC-derived foam cells in the plaque shoulder and marginal areas. S100A4 expression increased concomitantly with plaque formation in our animal model. Exogenous recombinant S100A4 protein enhanced the levels of Mmp2, Mmp9, and Mmp13 and the cell proliferation ability in VSMCs. A chemotaxis assay indicated that extracellular S100A4 functions as a chemoattractant for VSMCs. Conclusions S100A4 expression was elevated in human carotid plaques and showed a positive correlation with the degree of expansive remodeling. S100A4-positive VSMC-derived cells are considered to play an important role in carotid expansive remodeling.
Asunto(s)
Estenosis Carotídea/metabolismo , Proteína de Unión al Calcio S100A4/metabolismo , Remodelación Vascular , Animales , Estenosis Carotídea/diagnóstico por imagen , Estenosis Carotídea/patología , Estenosis Carotídea/fisiopatología , Humanos , Angiografía por Resonancia Magnética , Masculino , Ratones , Músculo Liso Vascular/fisiopatología , Reacción en Cadena de la Polimerasa , Proteína de Unión al Calcio S100A4/fisiología , Regulación hacia ArribaRESUMEN
BACKGROUND: Fibroblast dysfunction is the main pathogenic mechanism of idiopathic pulmonary fibrosis (IPF). S100 calcium-binding protein A4 (S100A4) plays critical roles in the proliferation of fibroblasts and in the development of pulmonary, hepatic, and renal fibrosis. However, the clinical implications of S100A4 in IPF have not been evaluated. METHODS AND MATERIALS: The S100A4 mRNA and protein levels were measured by real-time PCR and immunoblotting in fibroblasts from IPF patients and controls. The S100A4 level was measured by enzyme-linked immunosorbent assay in bronchoalveolar lavage fluid (BALF) from the normal controls (NCs; n = 33) and from patients with IPF (n = 87), non-specific interstitial pneumonia (NSIP; n = 22), hypersensitivity pneumonitis (HP; n = 19), and sarcoidosis (n = 9). S100A4 localization was evaluated by immunofluorescence staining. RESULTS: The S100A4 mRNA and protein levels were significantly higher in fibroblasts from IPF patients (n = 14) than in those from controls (n = 10, p < 0.001). The S100A4 protein level in BALF was significantly higher in the IPF (89.25 [49.92-203.02 pg/mL]), NSIP (74.53 [41.88-131.45 pg/mL]), HP (222.36 [104.92-436.92 pg/mL]) and sarcoidosis (101.62 [59.36-300.62 pg/mL]) patients than in the NCs (7.57 [1.31-14.04 pg/mL], p < 0.01, respectively). Cutoff S100A4 levels of 18.85 and 28.88 pg/mL had 87.4% and 87.8% accuracy, respectively, for discriminating IPF and other lung diseases from NCs. CONCLUSIONS: S100A4 is expressed by α-SMA-positive cells in the interstitium of the IPF patients. S100A4 may participate in the development of IPF, and its protein level may be a candidate diagnostic and therapeutic marker for IPF.
Asunto(s)
Fibrosis Pulmonar Idiopática/diagnóstico , Fibrosis Pulmonar Idiopática/genética , Pulmón/metabolismo , Proteína de Unión al Calcio S100A4/genética , Proteína de Unión al Calcio S100A4/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/metabolismo , Femenino , Expresión Génica , Humanos , Fibrosis Pulmonar Idiopática/metabolismo , Masculino , Persona de Mediana Edad , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteína de Unión al Calcio S100A4/fisiologíaRESUMEN
Extensive studies on metastasis-associated proteins, S100A4 and MTA1, have been carried out for over two decades, but correlation of both proteins remains obscure. Here we show evidence for the correlation in angiogenesis. First, silencing of each protein by siRNA-mediated knockdown in mouse endothelial MSS31 cells resulted in the inhibition of tube formation. Unexpectedly, the knockdown of MTA1 affected not only its own expression but also the expression of S100A4, whereas silencing of S100A4 did not affect the MTA1 expression. Additionally, non-muscle myosin IIA (NMIIA) phosphorylation, which was partly controlled by S100A4, was found to be upregulated by knockdown of both proteins in MSS31 cells. Moreover, cycloheximide treatment of MSS31 cells revealed that the rate of S100A4 degradation was accelerated by MTA1 knockdown. This finding, together with our observation that cytoplasmic MTA1, but not nuclear MTA1, was colocalized with S100A4, suggested the involvement of MTA1 in S100A4 stability. The direct in vivo angiogenesis assay showed that both protein siRNAs provoked a significant inhibition of new blood vessel formation induced by angiogenic factors, indicating their anti-angiogenic activities. Treatment of human pancreatic tumor (PANC-1) xenograft in mice with mMTA1 siRNA resulted in tumor regression via suppression of angiogenesis in vivo, as also observed in the case of human prostate cancer xenograft treated with mS100A4 siRNA. Taken together, these data led us to conclude that the MTA1-S100A4-NMIIA axis exists in endothelial cells as a novel pathway in promoting tumor vascular formation and could be a target for suppressing tumor growth and metastasis.
Asunto(s)
Histona Desacetilasas/fisiología , Neoplasias/irrigación sanguínea , Neoplasias/patología , Neovascularización Patológica/genética , Proteínas Represoras/fisiología , Proteína de Unión al Calcio S100A4/fisiología , Animales , Proliferación Celular/genética , Células Cultivadas , Histona Desacetilasas/genética , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Ratones Desnudos , Metástasis de la Neoplasia , Neoplasias/genética , Proteínas Represoras/genética , Proteína de Unión al Calcio S100A4/genética , TransactivadoresRESUMEN
BACKGROUND: Pulmonary arterial hypertension (PAH) is a complex pulmonary vasculature disease characterized by remodeling of the pulmonary vessels and a persistent increase in the pulmonary vascular resistance (PVR) with a poor prognosis. Serotonin increases the expression of S100A4/Mts1, which in turn stimulates the proliferation and migration of human pulmonary artery smooth muscle cells through the interaction with RAGE (receptor for advanced glycation end products) and thus S100A4/Mts1 has been implicated in the development of PAH in vitro. Fluoxetine, a selective serotonin re-uptake inhibitor has been shown to protect against PAH. The current study was designed to test whether S100A4 and its associated proteins connected in the development of PAH in vivo as well as to investigate the involvement of those proteins in the protective effect of fluoxetine against PAH. METHODS: MCT-induced PAH models were established in Wistar rats by a single intraperitoneal injection of MCT (60 mg/kg). Fluoxetine (2 and 10 mg/kg/day) was intragastrically administered once a day for 3 weeks along with controls. The detection methods followed include Hematoxylin and Eosin (H&E) staining, immunohistochemistry, western blotting and real-time reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: MCT induced pulmonary hypertension, pulmonary vascular remodeling, and right ventricular hypertrophy significantly increased the expressions of S100A4 and RAGE in the pulmonary arteries, lungs and right ventricle (RV). Fluoxetine dose-dependently inhibited MCT-induced pulmonary arterial hypertension, pulmonary vascular remodeling, and right ventricular hypertrophy and reduced the S100A4 and RAGE. Further analysis revealed that fluoxetine alleviated both the increase of p53, MMP13, MMP2 and MMP9 and the decrease of pp53Ser15 and MDM2 in lungs and RV tissues of MCT-induced PAH rats. CONCLUSION: From the present investigation it could be concluded that S100A4/Mts1 and its associated proteins are involved in the evolution of MCT-induced PAH in rats and fluoxetine inhibits MCT-induced PAH in rats mainly through S100A4/RAGE signaling axis and involved factors.
Asunto(s)
Fluoxetina/uso terapéutico , Hipertensión Pulmonar/prevención & control , Proteína de Unión al Calcio S100A4/fisiología , Animales , Relación Dosis-Respuesta a Droga , Hipertensión Pulmonar/inducido químicamente , Masculino , Monocrotalina , Proteínas Proto-Oncogénicas c-mdm2/análisis , Ratas , Ratas Wistar , Receptor para Productos Finales de Glicación Avanzada/análisis , Proteína de Unión al Calcio S100A4/análisis , Proteína p53 Supresora de Tumor/análisisRESUMEN
Schwann cells (SCs) have been reported as a possible source of neurotrophic support for spiral ganglion neurons (SGNs). This study was aimed to investigate whether S100A4 was contributed in the functional effects of SCs on SGNs. SCs were transfected with S100A4 vector or small interfering RNA (siRNA) against S100A4, and the transfection efficiency was verified by quantitative PCR (qPCR) and Western blot. The migration of transfected SCs was determined by Transwell assay, and the expression levels of vascular endothelial growth factor precursor (VEGF) and matrix metallopeptidase 9 (MMP-9) were measured by Western blot. Co-culture of either S100A4 overexpressed or suppressed SCs with SGNs, and the growth associated protein 43 (GAP43) expression in SGNs was detected by immunofluorescence (IF), qPCR and Western blot. The migration of SCs was significantly enhanced by S100A4 overexpression (P < 0.001), while was suppressed by S100A4 knockdown (P < 0.01). Further, the expressions of VEGF and MMP-9 were notably up-regulated by S100A4 overexpression, while were down-regulated by S100A4 knockdown. Moreover, co-culture with the S100A4 overexpressed SCs significantly increased the expression of GAP43 in SGNs (P < 0.01). As expected, co-culture with S100A4 knockdown SCs decreased GAP43 level (P < 0.05). S100A4 enhanced the migratory ability of SCs. SCs genetically modified to overexpress the S100A4 could up-regulate the GAP43 expression in SGNs.
Asunto(s)
Movimiento Celular/fisiología , Proteína GAP-43/metabolismo , Ganglios Espinales/citología , Ganglios Espinales/metabolismo , Proteína de Unión al Calcio S100A4/fisiología , Células de Schwann/fisiología , Células Receptoras Sensoriales/metabolismo , Animales , Mejoramiento Genético/métodos , Fármacos Neuroprotectores/metabolismo , Ratas , Células de Schwann/citología , Células Receptoras Sensoriales/citologíaRESUMEN
OBJECTIVE: PRAME (Preferentially Expressed Antigen in Melanoma) is a tumor-associated antigen recognized by immunocytes, and it induces cytotoxic T cell-mediated responses in melanoma. PRAME is expressed in a wide variety of tumors, but in contrast with most other tumor-associated antigens, it is also expressed in leukemias. The physiologic role of PRAME remains elusive. Recently, it has found PRAME could be involved in the regulation of cell death in leukemias, but the mechanism of the function is unclear. Here, we confirm that PRAME induces leukemias cell death by regulation of S100A4/p53 signaling. MATERIALS AND METHODS: The pCDNA3-PRAME plasmid and its control were transfected with the KG-1 cells. The pCDNA3-PRAME transfected KG-1 cells were then transiently transfected with S100A4 cDNA or wt-p53 siRNA. The PRAME siRNA and its control were transfected with the K562 cells. The PRAME siRNA transfected K562 cells were then transiently transfected with S100A4 siRNA or pGMp53-Lu. PRAME, S100A4 and P53 were detected by Western blot assay in different time point. Annexin V/propidium iodide and MTT methods were used to detect apoptosis and cell survival rate. RESULTS: KG-1 cells overexpressing the PRAME gene significantly induces apoptosis and decreases proliferation in vitro, followed by down-regulation of S100A4 and up-regulation of p53. Up-regulation of S100A4 by S100A4 transfection inhibits PRAME-induced p53 up-regulation. Furthermore, up-regulation of S100A4 by S100A4 transfection or down-regulation of p53 by p53 siRNA transfection reduces apoptosis and increases proliferation in vitro. Knockdown of PRAME in K562 cells significantly increases proliferation in vitro, followed by up-regulation of S100A4 and down-regulation of p53. The downregulation of S100A4 by S100A4 siRNA transfection increased p53 expression. Furthermore, downregulation of S100A4 by S100A4 siRNA transfection or up-regulation of p53 by p53 transfection decreases proliferation in vitro. CONCLUSIONS: Our results suggest that the leukemias expressing high levels of PRAME has a favorable prognosis. PRAME promotes in vitro leukemia cells death by regulating S100A4/p53 signaling.
Asunto(s)
Antígenos de Neoplasias/biosíntesis , Genes p53/fisiología , Leucemia/metabolismo , Proteína de Unión al Calcio S100A4/fisiología , Transducción de Señal/fisiología , Antígenos de Neoplasias/genética , Apoptosis/fisiología , Muerte Celular/fisiología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Humanos , Células K562 , Leucemia/genética , Leucemia/patología , ARN Interferente Pequeño/biosíntesis , ARN Interferente Pequeño/genéticaRESUMEN
Epithelial-mesenchymal transition (EMT) is a major cause of endometrial cancer (EC) to initiate invasion and metastasis. S100A4, a calcium-binding protein, is implicated in multistage of tumorigenesis and tumor progression. The correlation between S100A4 and EMT in EC is still unclear. This study was aimed to clarify the role of S100A4 in EC and the relationship between S100A4 expression and EMT markers. S100A4, E-cadherin, and vimentin were detected in tissues of EC patients (n=50) by immunohistochemistry. The impact of S100A4 on EC cell proliferation, migration and invasion was investigated via RNA interference, and the correlation between S100A4 and EMT markers were also explored. The results showed that S100A4 was significantly increased in epithelial cells of EC compared with the normal endometrium (P<0.05), also S100A4 level was positively related to age (P=0.021), histological grade (P<0.001), and lymph node metastasis (P<0.001). Additionally, silencing of S100A4 remarkably attenuated EC cell migration and invasion. Significant morphological change accompanied with the downregulation of EMT markers, E-cadherin and vimentin were also observed. Aberrant S100A4 expression may predict EC progression and play an important role in regulating EC cell invasion through EMT regulation. Hence, S100A4 is a promising therapeutic target.
Asunto(s)
Neoplasias Endometriales/patología , Transición Epitelial-Mesenquimal , Proteína de Unión al Calcio S100A4/fisiología , Línea Celular Tumoral , Movimiento Celular , Progresión de la Enfermedad , Femenino , Humanos , Invasividad Neoplásica , Proteína de Unión al Calcio S100A4/análisisRESUMEN
Numerous studies have shown that S100A4 acquires its metastasis-promoting effects via inducing epithelial-mesenchymal transition (EMT). However, its role and mechanism in EMT in breast cancer had not been clearly elucidated. Herein, we showed that the knockdown of S100A4 expression in breast cancer cell lines, MDA-MB-231 and MDA-MB-468, inhibited not only cell invasion ability greatly, but also the occurrence of EMT significantly. In addition, S100A4 knockdown could also decrease the expression of MMP2, a promoter and a mediator of the EMT processes in cancer. Above all, restoring the expression of MMP2 in MDA-MB-231 and MDA-MB-468 could not only rescue the invasion ability inhibited by knockdown of S100A4, but also reverse the EMT suppressed by knockdown of S100A4. In summary, our results indicated that S100A4 could promote the invasion ability of breast cancer cells via EMT, more importantly, it could participate in EMT via regulating MMP2 in breast cancer. Therefore, S100A4 could be a candidate biomarker for defining breast cancer metastasis and useful target for therapy.