RESUMEN
Breast cancer (BC), the most prevalent cancer in women, is a heterogenous disease. Despite advancements in BC diagnosis, prognosis, and therapeutics, survival rates have drastically decreased in the metastatic setting. Therefore, BC still remains a medical challenge. The evolution of high-throughput technology has highlighted gaps in the classification system of BCs. Of particular interest is the notorious triple negative BC, which was recounted as being heterogenous itself and it overlaps with distinct subtypes, namely molecular apocrine (MA) and luminal androgen (LAR) BCs. These subtypes are, even today, still misdiagnosed and poorly treated. As such, researchers and clinicians have been looking for ways through which to refine BC classification in order to properly understand the initiation, development, progression, and the responses to the treatment of BCs. One tool is biomarkers and, specifically, microRNA (miRNA), which are highly reported as associated with BC carcinogenesis. In this review, the diverse roles of miRNA in estrogen receptor negative (ER-) and androgen receptor positive (AR+) BC are depicted. While highlighting their oncogenic and tumor suppressor functions in tumor progression, we will discuss their diagnostic, prognostic, and predictive biomarker potentials, as well as their drug sensitivity/resistance activity. The association of several miRNAs in the KEGG-reported pathways that are related to ER-BC carcinogenesis is presented. The identification and verification of accurate miRNA panels is a cornerstone for tackling BC classification setbacks, as is also the deciphering of the carcinogenesis regulators of ER - AR + BC.
RESUMEN
Flow cytometry (FCM) is a state-of-the-art technique for the qualitative and quantitative assessment of cells and other particles' physical and biological properties. These cells are suspended within a high-velocity fluid stream and pass through a laser beam in single file. The main principle of the FCM instrument is the light scattering and fluorescence emission upon the interaction of the fluorescent particle with the laser beam. It also allows for the physical sorting of particles depending on different parameters. A flow cytometer comprises different components, including fluidic, optics, and electronics systems. This article briefly explains the mechanism of all components of a flow cytometer to clarify the FCM technique's general principles, provides some useful guidelines for the proper design of FCM panels, and highlights some general applications and important applications in cancer research. © 2023 Wiley Periodicals LLC.
Asunto(s)
Neoplasias , Proyectos de Investigación , Humanos , Citometría de Flujo/métodos , ColorantesRESUMEN
Gap-junction-forming connexins are exquisitely regulated by post-translational modifications (PTMs). In particular, the PTM of connexin 43 (Cx43), a tumor suppressor protein, regulates its turnover and activity. Here, we investigated the interaction of Cx43 with the ubiquitin-related modifier 1 (URM-1) protein and its impact on tumor progression in two breast cancer cell lines, highly metastatic triple-negative MDA-MB-231 and luminal breast cancer MCF-7 cell lines. To evaluate the subsequent modulation of Cx43 levels, URM-1 was downregulated in these cells. The transcriptional levels of epithelial-to-mesenchymal transition (EMT) markers and the metastatic phenotype were assessed. We demonstrated that Cx43 co-localizes and interacts with URM-1, and URMylated Cx43 was accentuated upon cellular stress. The significant upregulation of small ubiquitin-like modifier-1 (SUMO-1) was also observed. In cells with downregulated URM-1, Cx43 expression significantly decreased, and SUMOylation by SUMO-1 was affected. The concomitant expression of EMT markers increased, leading to increased proliferation, migration, and invasion potential. Inversely, the upregulation of URM-1 increased Cx43 expression and reversed EMT-induced processes, underpinning a role for this PTM in the observed phenotypes. This study proposes that the URMylation of Cx43 in breast cancer cells regulates its tumor suppression properties and contributes to breast cancer cell malignancy.
Asunto(s)
Neoplasias de la Mama , Conexina 43 , Femenino , Humanos , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Conexina 43/genética , Conexina 43/metabolismo , Conexinas/metabolismo , Transición Epitelial-Mesenquimal/genética , Uniones Comunicantes/metabolismo , Células MCF-7 , Ubiquitina/metabolismoRESUMEN
Background: Cardiovascular disease that is caused by atherosclerosis is the leading cause of death worldwide. Atherosclerosis is primarily triggered by endothelial dysfunction and the accumulation of modified low-density lipoprotein (LDL) particles in the subendothelial space of blood vessels. Early reports have associated oxidized LDL with altered fibrinolysis and atherogenesis. It has been suggested that myeloperoxidase oxidized LDL (Mox-LDL) is involved in atherosclerosis because of its significant pathophysiological role in the modification of LDL in vivo. It has been equally demonstrated that Mox-LDL binds to the lectin-like oxidized low-density lipoprotein receptor-1 (lox-1) scavenger receptor which leads to the upregulation of inflammatory mediators in endothelial cells and the progression of cardiovascular disease. It has been also shown that neuroserpin, a member of the serine proteinase inhibitor (serpin) superfamily, has an important role at the level of fibrinolysis in the nervous tissue. Methods: Since little is known about the effects of Mox-LDL on endothelial cell fibrinolytic activity and the involvement of lox-1 in this process, our study aimed at evaluating the in vitro effects of Mox-LDL on neuroserpin release from human aortic endothelial cells (HAECs) and the role of lox-1 scavenger receptor in this context by relying on lox-1 gene silencing in HAECs, culturing the cells in the presence of Mox-LDL, measuring their neuroserpin expression and release by quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively, and assessing their fibrinolytic activity using the Euglobulin Clot Lysis Time (ECLT) method. Results: Our data show that Mox-LDL decreases endothelial cell fibrinolytic capacity by upregulating neuroserpin in lox-1 knockdown cells. Conclusions: Lox-1 protects the endothelial cells from a Mox-LDL-induced decrease in pro-fibrinolytic capacity, which has important consequences in the context of stroke.
RESUMEN
BACKGROUND: Inflammatory bowel disease (IBD) constitutes a substantial risk factor for colorectal cancer. Connexin 43 (Cx43) is a protein that forms gap junction (GJ) complexes involved in intercellular communication, and its expression is altered under pathological conditions, such as IBD and cancer. Recent studies have implicated epigenetic processes modulating DNA methylation in the pathogenesis of diverse inflammatory and malignant diseases. The ten-eleven translocation-2 (TET-2) enzyme catalyzes the demethylation, hence, regulating the activity of various cancer-promoting and tumor-suppressor genes. AIM: To investigate Cx43 and TET-2 expression levels and presence of 5-hydroxymethylcytosine (5-hmC) marks under inflammatory conditions both in vitro and in vivo. METHODS: TET-2 expression was evaluated in parental HT-29 cells and in HT-29 cells expressing low or high levels of Cx43, a putative tumor-suppressor gene whose expression varies in IBD and colorectal cancer, and which has been implicated in the inflammatory process and in tumor onset. The dextran sulfate sodium-induced colitis model was reproduced in BALB/c mice to evaluate the expression of TET-2 and Cx43 under inflammatory conditions in vivo. In addition, archived colon tissue sections from normal, IBD (ulcerative colitis), and sporadic colon adenocarcinoma patients were obtained and evaluated for the expression of TET-2 and Cx43. Expression levels were reported at the transcriptional level by quantitative real-time polymerase chain reaction, and at the translational level by Western blotting and immunofluorescence. RESULTS: Under inflammatory conditions, Cx43 and TET-2 expression levels increased compared to non-inflammatory conditions. TET-2 upregulation was more pronounced in Cx43-deficient cells. Moreover, colon tissue sections from normal, ulcerative colitis, and sporadic colon adenocarcinoma patients corroborated that Cx43 expression increased in IBD and decreased in adenocarcinoma, compared to tissues from non-IBD subjects. However, TET-2 expression and 5-hmC mark levels decreased in samples from patients with ulcerative colitis or cancer. Cx43 and TET-2 expression levels were also investigated in an experimental colitis mouse model. Interestingly, mice exposed to carbenoxolone (CBX), a GJ inhibitor, had upregulated TET-2 levels. Collectively, these results show that TET-2 levels and activity increased under inflammatory conditions, in cells downregulating gap junctional protein Cx43, and in colon tissues from mice exposed to CBX. CONCLUSION: These results suggest that TET-2 expression levels, as well as Cx43 expression levels, are modulated in models of intestinal inflammation. We hypothesize that TET-2 may demethylate genes involved in inflammation and tumorigenesis, such as Cx43, potentially contributing to intestinal inflammation and associated carcinogenesis.
Asunto(s)
Adenocarcinoma , Colitis Ulcerosa , Colitis , Neoplasias del Colon , Dioxigenasas , Enfermedades Inflamatorias del Intestino , Animales , Ratones , Adenocarcinoma/patología , Carcinogénesis/patología , Colitis/inducido químicamente , Colitis Ulcerosa/patología , Colon/patología , Neoplasias del Colon/patología , Conexina 43/genética , Conexina 43/metabolismo , Sulfato de Dextran/toxicidad , Dioxigenasas/metabolismo , Modelos Animales de Enfermedad , Inflamación/patología , Enfermedades Inflamatorias del Intestino/patologíaRESUMEN
Cardiovascular disease as a result of atherosclerosis is a leading cause of death worldwide. Atherosclerosis is primarily caused by the dysfunction of vascular endothelial cells and the subendothelial accumulation of oxidized forms of low-density lipoprotein (LDL). Early observations have linked oxidized LDL effects in atherogenesis to the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) scavenger receptor. It was shown that LOX-1 is upregulated by many inflammatory mediators and proatherogenic stimuli including cytokines, reactive oxygen species (ROS), hemodynamic blood flow, high blood sugar levels and, most importantly, modified forms of LDL. Oxidized LDL signaling pathways in atherosclerosis were first explored using LDL that is oxidized by copper (Cuox-LDL). In our study, we used a more physiologically relevant model of LDL oxidation and showed, for the first time, that myeloperoxidase oxidized LDL (Mox-LDL) may affect human aortic endothelial cell (HAEC) function through the LOX-1 scavenger receptor. We report that Mox-LDL increases the expression of its own LOX-1 receptor in HAECs, enhancing inflammation and simultaneously decreasing tubulogenesis in the cells. We hypothesize that Mox-LDL drives endothelial dysfunction (ED) through LOX-1 which provides an initial hint to the pathways that are initiated by Mox-LDL during ED and the progression of atherosclerosis.
Asunto(s)
Aterosclerosis , Células Endoteliales , Aterosclerosis/metabolismo , Células Endoteliales/metabolismo , Humanos , Lipoproteínas LDL/metabolismo , Lipoproteínas LDL/farmacología , Peroxidasa/metabolismo , Receptores de LDL/metabolismo , Receptores Depuradores de Clase E/metabolismoRESUMEN
Prostate cancer is the second most commonly diagnosed cancer in men and one of the main leading causes of cancer deaths among men worldwide. Rapid uncontrolled growth and the ability to metastasize to other sites are key hallmarks in cancer development and progression. The Rho family of GTPases and its activators the GTPase-activating proteins (GAPs) are required for regulating cancer cell proliferation and migration. StarD13 is a GAP for Rho GTPases, specifically for RhoA and Cdc42. We have previously shown that StarD13 acts as a tumor suppressor in astrocytoma as well as breast and colorectal cancer. In this study, we performed a functional comparative analysis of StarD13 targets/and or interacting molecules to understand the general role that StarD13 plays in cancers. Our data highlight the importance of StarD13 in modulating several hallmarks of cancer. Findings from database mining and immunohistochemistry revealed that StarD13 is underexpressed in prostate cancers, in addition knocking down Stard13 increased cancer cell proliferation, consistent with its role as a tumor suppressor. Stard13 depletion, however, led to an increase in cell adhesion, which inhibited 2D cell migration. Most interestingly, StarD13 depletion increases invasion and matrix degradation, at least in part, through its regulation of Cdc42. Altogether, the data presented suggest that StarD13 acts as a tumor suppressor inhibiting prostate cancer cell invasion.
Asunto(s)
Movimiento Celular/genética , Proteínas Activadoras de GTPasa/genética , Proteínas Activadoras de GTPasa/fisiología , Expresión Génica/genética , Invasividad Neoplásica/genética , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/fisiología , Línea Celular Tumoral , Progresión de la Enfermedad , Proteínas Activadoras de GTPasa/metabolismo , Humanos , Masculino , Proteínas Supresoras de Tumor/metabolismo , Proteína de Unión al GTP cdc42 , Proteínas de Unión al GTP rho , Proteína de Unión al GTP rhoARESUMEN
Primary effusion lymphoma (PEL) is a rare aggressive subset of non-Hodgkin B cell lymphoma. PEL is secondary to Kaposi sarcoma herpes virus (KSHV) and predominantly develops in serous cavities. Conventional chemotherapy remains the treatment of choice for PEL and yields high response rates with no significant comorbidities. Yet, chemotherapy often fails in achieving or maintaining long-term remission. Lenalidomide (Lena), an immunomodulatory drug, displayed some efficacy in the treatment of PEL. On the other hand, arsenic trioxide (ATO) in combination with other agents effectively treated a number of blood malignancies, including PEL. In this study, we present evidence that the combination of ATO/Lena significantly enhanced survival of PEL mice, decreased the volume of exacerbated ascites in the peritoneum, and reduced tumor infiltration in organs of treated animals. In ex vivo treated PEL cells, ATO/Lena decreased the proliferation and downregulated the expression of KSHV latent viral proteins. This was associated with decreased NF-κB activation, resulting in reactivation of viral replication, downregulation of interleukin-6 (IL-6) and IL-10, inhibition of vascular endothelial growth factor, and apoptosis. Our results elucidate the mechanism of action of ATO/Lena and present it as a promising targeted therapeutic modality in PEL management, which warrants further clinical investigation.
RESUMEN
Loss of connexin-mediated cell-cell communication is a hallmark of breast cancer progression. Pannexin1 (PANX1), a glycoprotein that shares structural and functional features with connexins and engages in cell communication with its environment, is highly expressed in breast cancer metastatic foci; however, PANX1 contribution to metastatic progression is still obscure. Here we report elevated expression of PANX1 in different breast cancer (BRCA) subtypes using RNA-seq data from The Cancer Genome Atlas (TCGA). The elevated PANX1 expression correlated with poorer outcomes in TCGA BRCA patients. In addition, gene set enrichment analysis (GSEA) revealed that epithelial-to-mesenchymal transition (EMT) pathway genes correlated positively with PANX1 expression. Pharmacological inhibition of PANX1, in MDA-MB-231 and MCF-7 breast cancer cells, or genetic ablation of PANX1, in MDA-MB-231 cells, reverted the EMT phenotype, as evidenced by decreased expression of EMT markers. In addition, PANX1 inhibition or genetic ablation decreased the invasiveness of MDA-MB-231 cells. Our results suggest PANX1 overexpression in breast cancer is associated with a shift towards an EMT phenotype, in silico and in vitro, attributing to it a tumor-promoting effect, with poorer clinical outcomes in breast cancer patients. This association offers a novel target for breast cancer therapy.
RESUMEN
Connexins regulate multiple cellular functions and are considered tumor suppressors. Connexin43 (Cx43) is frequently down-regulated in breast tumors. However, Cx43 regulation during cancer onset and metastasis is complex and context-dependent. We investigated the effect of Cx43 over-expression or knock-down on the metastatic potential of MDA-MB-231 breast cancer cells in vitro and in vivo and in human breast cancer tissues. MDA-MB-231 cells over-expressing (Cx43D) or down-regulating Cx43 (shCx43) were generated and used in proliferation, migration, and invasion assays. The regulation of genes/proteins implicated in progression, invasion and metastasis was assessed in vitro and in immune-compromized mice injected with MDA-MB-231, Cx43D or shCx43 cells. Primary tumor onset/growth, metastasis and overall survival of these animals was monitored and evaluated. In addition, Cx43 expression in human breast carcinoma samples was assessed by qPCR. Cx43 over-expression increased protein levels of epithelial markers E-cadherin and zonula occludens 1 expression and resulted in the sequestration of ß-catenin at the cell membrane, while Cx43 knock-down induced protein expression of the mesenchymal marker N-cadherin and an increased invasive potential of shCx43 cells. In vivo, in mice xenografted with breast cancer cells, Cx43 over-expression decreased tumor volume, attenuated cell metastasis to lungs and liver and increased overall mice survival. Importantly, the expression of Cx43 in triple negative human breast cancer tissues is also down-regulated. Collectively, Cx43 over-expression induced an epithelial-like phenotype in MDA-MB-231 cells and suppressed tumor growth and metastasis to secondary organs in vivo. In contrast, Cx43 knock-down in MDA-MB-231 cells induced a mesenchymal phenotype with increased cell invasion leading to an enhanced metastatic phenotype. These data provide evidence for a pivotal role of Cx43 in breast cancer metastasis and support the potential targeting of connexins in breast cancer therapy.
RESUMEN
Bevacizumab or Avastin® (Av), the recombinant antibody targeting VEGF, improves progression-free but not overall survival of metastatic breast cancer patients due to development of Av resistance. We showed that Av-therapy-induced inflammatory microenvironment contributes to the refractoriness to Av treatment. Here we present data regarding the effect of Av treatment on migration of a non-invasive breast cancer cell line, MCF-7. The data presented hereis related to the research article "Bevacizumab induces inflammation in MDA-MB-231 breast cancer cell line and in a mouse model" (Hajjar et al., 2018).
RESUMEN
BACKGROUND: Bevacizumab or Avastin® (Av) is an anti-vascular endothelial growth factor agent. It does not improve survival of breast cancer patients due to development of refractoriness. Av treatment was shown to increase inflammation in a diabetic mouse model, and also to induce epithelial-to-mesenchymal transition of non-transformed breast epithelia. This study aimed to understand if the Av-induced inflammatory microenvironment could be a mechanism of Av refractoriness. Expression profiles of inflammatory mediators, in vitro in MDA-MB-231 cells, in vivo in a mouse model xenografted with MDA-MB-231 cells and from archived cases of human breast carcinoma tissues were evaluated. Gap junctions are also crucial for angiogenesis and tumor cell extravasation. The effect of connexin 43 (Cx43) overexpression on the expression of inflammatory markers in MDA-MB-231 cells treated with Av was assessed. METHODS: MDA-MB-231 cells, control or overexpressing Cx43, were used in this study. Proliferation and invasion assays were performed. Quantitative PCR, ELISA and western blotting were performed to assess the regulation of inflammatory mediators and other factors upon Av treatment. Immunofluorescence was performed to document the translocation of Nuclear Factor-kappa B p65. RESULTS: Breast cancer tissues had elevated transcriptional levels of inflammatory mediators. Av treatment increased expression levels of inflammatory mediators and metastatic factors in vitro and in vivo. Interestingly, overexpressing Cx43 in MDA-MB-231 cells alleviated the inflammatory effects induced by Av treatment. CONCLUSION: This study attributes Av refractoriness to the Av therapy-induced inflammatory microenvironment.