BackgroudAccumulating evidences indicate that circular RNAs (circRNAs), a class of non-coding RNAs, play important roles in tumorigenesis. However, the function of circRNAs in triple negative breast cancer (TNBC) is largely unknown.MethodsWe performed circRNA microarrays to identify circRNAs that are aberrantly expressed in TNBC cell lines. Expression levels of a significantly upregulated circRNA, circGFRA1, was detected by quantitative real-time PCR (qRT-PCR) in TNBC cell lines and tissues. Kaplan-Meier survival analysis was used to explore the significance of circGFRA1 in clinical prognosis. Then, we examined the functions of circGFRA1 in TNBC by cell proliferation, apoptosis and mouse xenograft assay. In addition, luciferase assay was used to explore the miRNA sponge function of circGFRA1 in TNBC.ResultsMicroarray analysis and qRT-PCR verified a circRNA termed circGFRA1 that was upregulated in TNBC. Kaplan-Meier survival analysis showed that upregulated circGFRA1 was correlated with poorer survival. Knockdown of circGFRA1 inhibited proliferation and promoted apoptosis in TNBC. Via luciferase reporter assays, circGFRA1 and GFRA1 was observed to directly bind to miR-34a. Subsequent experiments showed that circGFRA1 and GFRA1 regulated the expression of each other by sponging miR-34a.ConclusionsTaken together, we conclude that circGFRA1 may function as a competing endogenous RNA (ceRNA) to regulate GFRA1 expression through sponging miR-34a to exert regulatory functions in TNBC. circGFRA1 may be a diagnostic biomarker and potential target for TNBC therapy.

MicroRNA-124 (miR-124), a pivotal member of the p53 network, was found to be down-regulated in multiple types of tumors and further reported as tumor suppressor microRNA. In this study, we found that miR-124 was down-regulated in gastric cancer cell lines and specimens. Restoration of miR-124 expression inhibited the proliferation and colony formation of gastric cancer cells. EZH2 (enhancer of zeste homolog 2), which has been shown to be an important transcription factor involved in the proliferation and metastasis of tumor cells, was here confirmed to be a direct target gene of miR-124. On the other hand, silencing EZH2 also inhibits cell proliferation of gastric cancer cells. Furthermore, the treatment combining miR-124 with 5-fluorouracil (5-FU) significantly showed more efficient anti-tumor effects than single treatment of miR-124 or 5-FU, and over-expression of miR-124 suppresses the tumor growth in vivo. Our study indicate that miR-124 can suppress gastric cancer cell growth by directly targeting the EZH2 gene and sensitize the treatment effect of 5-FU. Therefore, miR-124 shows tumor-suppressive activity and may be a new and useful approach of gastric cancer therapy.

Aim: To investigate the effects of the major component of high-density lipoprotein apolipoprotein A-I (apoA-I) on the development of atherosclerosis in LPS-challenged ApoE -/-mice and the underlying mechanisms. Methods: Male ApoE-KO mice were daily injected with LPS (25 μg, sc) or PBS for 4 weeks. The LPS-challenged mice were intravenously injected with rAAV-apoA-I-GFP or rAAV-GFP. After the animals were killed, blood, livers and aortas were collected for biochemical and histological analyses. For ex vivo experiments, the abdominal cavity macrophages were harvested from each treatment group of mice, and cultured with autologous serum, then treated with LPS. Results: Chronic administration of LPS in ApoE -/-mice significantly increased the expression of inflammatory cytokines (TNF-α, IL-1β, IL-6, and MCP-1), increased infiltration of inflammatory cells, and enhanced the development of atherosclerosis. In LPS-challenged mice injected with rAAV-apoA-I-GFP, viral particles and human apoA-I were detected in the livers, total plasma human apoA-I levels were grammatically increased; HDL-cholesterol level was significantly increased, TG and TC were slightly increased. Furthermore, overexpression of apoA-I significantly suppressed the expression of proinflammatory cytokines, reduced the infiltration of inflammatory cells, and decreased the extent of atherosclerotic lesions. Moreover, overexpression of apoA-I significantly increased the expression of the cytokine mRNA-destabilizing protein tristetraprolin (TTP), and phosphorylation of JAK2 and STAT3 in aortas. In ex vivo mouse macrophages, the serum from mice overexpressing apoA-I significantly increased the expression of TTP, accompanied by accelerated decay of mRNAs of the inflammatory cytokines. Conclusion: ApoA-I potently suppresses LPS-induced atherosclerosis by inhibiting the inflammatory response possibly via activation of STAT3 and upregulation of TTP.

The molecular mechanisms underlying triple-negative breast cancer (TNBC) pathology are not fully understood. Here, we reviewed the SOX8 transcript level in 24 types of cancer and normal tissues and the SOX8 expression pattern in breast cancer from the TCGA and METABRIC data sets and found that SOX8 was highly expressed in TNBC. We investigated the effect of SOX8 on tumorigenicity, migration and apoptosis in TNBC cell lines and xenografts models. We identified SOX8 as a functional oncogene that involved in the maintenance of stem-like capacities in TNBC cells. Through a promoter truncation experiment and ChIP experiment, we verified zinc finger E-box binding homeobox 1 (ZEB1) as a transcriptional activator of SOX8 that enhanced SOX8 expression by binding to its promoter. We evaluated the ZEB1 and the SOX8 levels in 240 TNBC patients and high expression of ZEB1 and SOX8 were significantly associated with poor prognosis. We demonstrated the significance of the ZEB1–SOX8 axis in regulating TNBC cancer stem-like cells (CSCs) and its connection with poor prognosis. Due to its vital role in TNBC CSCs, the ZEB1–SOX8 regulatory axis could be a promising therapeutic target for TNBC.

Fibroblast proliferation and migration are central in atrial fibrillation (AF) promoting structure remodeling, which is strongly associated with aging and hypertension. Transient receptor potential canonical-3 channel (TRPC3) is a key mediator of cardiac fibrosis and the pathogenesis of AF. Here, we have observed the increased TRPC3 expression that induced atrial fibrosis which possibly is either mediated by the aging process or related to hypertensive progression. In this study, we measured the pathological structure remodeling by H&E staining, Masson staining, and transmission electron microscope (TEM). The protein expression levels of fibrotic biomarkers and TRPC3 were measured by Western blotting with atrial tissues from normotensive Wistar Kyoto rats (WKY 4m-o (4 months old)), old WKY (WKY 24m-o (24 months old)), spontaneously hypertensive rat (SHR 4m-o (4 months old)), and old SHR (SHR 24m-o (24 months old)). To illuminate the molecular mechanism of TRPC3 in atrial fibrosis of aging rats and SHR, we detected the inhibited role of TRPC3 selective blocker ethyl-1-(4-(2,3,3-trichloroacrylamide) phenyl)-5-(trifluoromethyl)-1H-pyrazole-4-carboxylate,pyrazole-3 (Pyr3) on angiotensin II (Ang II) induced fibrosis in neonatal rat atrial fibroblasts. The pathological examination showed that the extracellular matrix (ECM) and collagen fibrils were markedly increased in atrial tissues from aged and hypertensive rats. The protein expressions of fibrotic biomarkers (collagen I, collagen III, and transforming growth factor-β1 (TGF-β1)) were significantly upregulated in atrial tissues from the WKY 24m-o group, SHR 4m-o group, and SHR 24m-o group compared with the WKY 4m-o group. Meanwhile, the expression level of TRPC3 was significantly upregulated in WKY 24m-o and SHR 4m-o atrial tissues compared to WKY 4m-o rats. In isolated and cultured neonatal rat atrial fibroblasts, Ang II induced the atrial fibroblast migration and proliferation and upregulated the expression levels of TRPC3 and fibrotic biomarkers. TRPC3 selected blocker Pyr3 attenuated the migration and proliferation in neonatal rat atrial fibroblasts. Furthermore, Pyr3 significantly alleviated Ang II-induced upregulation of TRPC3, collagen I, collagen III, and TGF-β1 through the molecular mechanism of the TGF-β/Smad2/3 signaling pathway. Similarly, knocking down TRPC3 using short hairpin RNA (shTRPC3) also attenuated Ang II-induced upregulation of TGF-β1. Pyr3 preconditioning decreased Ang II-induced intracellular Ca2+ transient amplitude elevation. Furthermore, AT1 receptor was involved in Ang II-induced TRPC3 upregulation. Hence, upregulation of TRPC3 in aging and hypertension is involved in an atrial fibrosis process. Inhibition of TRPC3 contributes to reverse Ang II-induced fibrosis. TRPC3 may be a potential therapeutic target for preventing fibrosis in aging and hypertension.

Action potential (AP) induces presynaptic membrane depolarization and subsequent opening of Ca2+ channels, and then triggers neurotransmitter release at the active zone of presynaptic terminal. Presynaptic Ca2+ channels and SNARE proteins (SNAREs) interactions form a large signal transfer complex, which are core components for exocytosis. Ca2+ channels serve to regulate the activity of Ca2+ channels through direct binding and indirect activation of active zone proteins and SNAREs. The activation of Ca2+ channels promotes synaptic vesicle recruitment, docking, priming, fusion and neurotransmission release. Intracellular calcium increase is a key step for the initiation of vesicle fusion. Various voltage-gated calcium channel (VGCC) subtypes exert different physiological functions. Until now, it has not been clear how different subtypes of calcium channels integrally regulate the release of neurotransmitters within 200 μs of the AP arriving at the active zone of synaptic terminal. In this mini review, we provide a brief overview of the structure and physiological function of Ca2+ channel subtypes, interactions of Ca2+ channels and SNAREs in neurotransmitter release, and dynamic fine-tune Ca2+ channel activities by G proteins (Gβγ), multiple protein kinases and Ca2+ sensor (CaS) proteins.

CircRNAs (circular RNA) are reported to regulate onset and progress multiple cancers. Nonetheless, the function along with the underlying mechanisms of circRNAs in HER‐2‐positive breast cancer (BC) remains unclear. CircRNA microarrays were performed to elucidate expression profiles of HER‐2‐positive BC cells. circRNA levels were quantified using qRT‐PCR assay. Various in vitro along with in vivo assays were employed to further explore the effects of circGFRA1 in the progress of HER‐2‐positive BC and interactions of circGFRA1, miR‐1228 and AIFM2 in Her‐2‐positive BC. CircGFRA1 was remarkably upregulated in HER‐2‐positive BC. Knockdown of circGFRA1 could attenuate HER‐2‐positive BC progression by inhibiting the proliferation, infiltration and migratory ability of HER‐2‐positive BC cells. Through ceRNA mechanism, circGFRA1 could bind to miR‐1228 and alleviate inhibitory activity of miR‐1228 on targeted gene AIFM2. In summary, circGFRA1‐miR‐1228‐AIFM2 axis regulates HER‐2‐positive BC. CircGFRA1 is a novel promising treatment option for HER‐2‐positive BC.

Background: Circular RNAs (circRNAs) have been reported to play important roles in cancer progression. However, the potential involvement of circRNAs in breast cancer metastasis to the lung remains unclear. Methods: High-throughput circular RNA microarray assays of primary breast cancer tissues and lung metastatic tissues were performed. Reactome pathway analysis and GO analysis of the linear mRNA transcripts corresponding to the circRNAs were conducted. The expression of the top downregulated circRNA was confirmed by qRT-PCR in breast cancer cell lines. Kaplan-Meier survival analysis was conducted to analyze the clinical significance of the selected circRNA in breast cancer. A series of in vitro and in vivo experiments, including cell proliferation and migration, was performed to explore the functions of the selected circRNA in breast cancer progression. We further investigated the regulatory effect of the selected circRNA on a miRNA and its target genes to explore the potential mechanisms. Results: We found that circNFIC (hsa_circ_0002018) was the most downregulated circRNA in lung metastatic tissues. Kaplan-Meier survival analysis revealed that low levels of circNFIC were related to poor outcome of breast cancer. Further experiments revealed that overexpressing circNFIC suppressed breast cancer cell proliferation and migration to the lung. A mechanistic study showed that circNFIC acted as a sponge for miR-658 and competed for binding to miR-658 with UPK1A, leading to increased expression of UPK1A. Conclusion: Our study highlighted the regulatory function of the circNFIC/miR-658/UPK1A pathway in breast cancer progression, which could be a potential therapeutic target for breast cancer.