Serum microRNA-501-3p is a potential diagnostic tool for detecting mild cognitive impairment: Ehime genome study.

Tight junction disruption and dysfunction are involved in the progression of blood-brain barrier (BBB) breakdown. Recent investigations have revealed BBB disruption in patients with vascular cognitive decline. Our previous studies showed that miR-501-3p negatively regulates cerebral endothelial tight junction protein-1, resulting in the disruption of the BBB, and playing an important role in the development of vascular cognitive impairment. BBB breakdown in white matter lesions is often seen in the patients with vascular mild cognitive impairment (MCI). We therefore hypothesize that most early-phase MCI patients may demonstrate elevated expression of miR-501-3p and sought to investigate whether serum exosome miR-501-3p levels could be a clinical indicator for detecting mild cognitive impairment. One hundred and seventy-eight subjects (aged 73 [68-75] years, 53% male) were recruited for this study. The Japanese version of the Montreal Cognitive Assessment (MoCA-J) was used for detecting MCI. Serum exosome miR-501-3p expression levels were measured by qPCR methods. Patients were divided into two groups depending on whether their miR-501-3p ∆Ct values were above ("High"; n = 74) or below ("Low"; n = 104) cutoff levels determined by ROC curve. MCI was detected significantly more often in the miR-501-3p-High group (vs. -Low group, 63.5% vs. 47.1%, respectively; p < 0.05). Multivariate logistic regression analysis showed a significant association between MCI status and High miR-501-3p (odds ratio 2.662; p < 0.01), improved vs. known risk factors. In non-diabetic patients, High miR-501-3p was positively associated with MCI status (odds ratio 3.633; p < 0.01) and also positively associated with MCI status in those with atherosclerosis (odds ratio 3.219; p < 0.01). The present study demonstrates that elevated expression of blood exosomal miR-501-3p can indicate the presence of MCI in human patients. Early detection of vascular injuries may allow a reduction in progressive dementia through the management of vascular risk factors.

Procyanidin C1 inhibits tumor growth and metastasis in colon cancer via modulating miR-501-3p/HIGD1A axis.

INTRODUCTION: Although colon (COAD) and rectal adenocarcinoma (READ) combined to refer to colorectal cancer (CRC), substantial clinical evidence urged that CRC should be treated as two different cancers due to compared with READ, COAD showed higher morbidity and worse 5-year survival. OBJECTIVES: This study has tried to screen for the crucial gene that caused the worse prognosis and investigate its mechanism for mediating tumor growth and metastases in COAD. Meanwhile, the potential anti-COAD compound implicated in this mechanism was identified and testified from 1,855 food-borne chemical kits. This study aims to bring a new perspective to the development of new anti-COAD drugs and personalized medicine for patients with COAD. METHODS AND RESULTS: The survival-related hub genes in COAD and READ were screened out from The Cancer Genome Atlas (TCGA) database and the results showed that HIGD1A, lower expressed in COAD than in READ, was associated with poor prognosis in COAD patients, but not in READ. Over-expressed HIGD1A suppressed CRC cell proliferation, invasion, and migration in vitro and in vivo. Meanwhile, the different expressed microRNA profiles between COAD and READ showed that miR-501-3p was highly expressed in COAD and inhibited HIGD1A expression by targeting 3'UTR of HIGD1A. MiR-501-3p mimics promoted cell proliferation and metastasis in CRC cells. In addition, Procyanidin C1 (PCC1), a kind of natural polyphenol has been verified as a potential miR-501-3p inhibitor. In vitro and in vivo, PCC1 promoted HIGD1A expression by suppressing miR-501-3p and resulted in inhibited tumor growth and metastasis. CONCLUSION: The present study verified that miR-501-3p/HIGD1A axis mediated tumor growth and metastasis in COAD. PCC1, a flavonoid that riched in food exerts anti-COAD effects by inhibiting miR-501-3p and results in the latter losing the ability to suppress HIGD1A expression. Subsequently, unfettered HIGD1A inhibited tumor growth and metastasis in COAD.

Identification of Serum miR-501-3p and miR-338-3p as Novel Diagnostic Biomarkers for Breast Cancer and Their Target Genes Associated with Immune Infiltration.

Background: MicroRNAs influence the growth and metastasis of breast cancer (BC) by regulating their target genes. Our study aims to screen and identify miRNAs that are closely related to the development of breast cancer, and explore the role of these miRNAs and their target genes in breast cancer. Methods: Bioinformatics tools were applied to screen breast cancer-associated miRNAs and predict their potential target genes. Serum miRNAs were measured using RT-PCR. The correlation between miRNA expression and different clinicopathological features of BC patients was analyzed. Receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value. GEPIA, Kaplan-Meier Plotter, TIMER, and TISIDB databases were used to validate the expression levels and their prognostic value, as well as their target gene associated with immune infiltrating cells and immune checkpoints. Results: Breast cancer-associated serum miR-338-3p and miR-501-3p were screened and verified for the first time. Serum miR-501-3p was elevated in BC and was closely linked to the ki-67 index and histological grade. CDKN2C, as a potential target gene of miR-501-3p, was enriched in the cGMP-PKG signaling pathway. Serum miR-338-3p was reduced in BC and was strongly linked to lymph node metastasis and histological grading. ACTR2, CDH1, COL1A1, RBBP5, RRM1, and TPM3, as potential target genes of miR-338-3p, were enriched in MAPK, PI3K-Akt, and RAS signaling pathways. These target genes were found to be linked to breast cancer prognosis, immune infiltrating cells, and immune checkpoint inhibitors. Analysis of ROC curve showed that serum miR-501-3p combined with serum miR-338-3p had a high diagnostic value in breast cancer (AUC: 0.89, 95% CI: 0.821-0.958). Conclusion: Serum miR-501-3p combined with serum miR-338-3p show obvious clinical significance in the diagnosis and prognosis of breast cancer, which suggests that they may act as novel diagnostic biomarkers for breast cancer.

Crosstalk Among NLRP3 Inflammasome, ETBR Signaling, and miRNAs in Stress-Induced Depression-Like Behavior: a Modulatory Role for SGLT2 Inhibitors.

Depression is an overwhelming health concern, and many patients fail to optimally respond to available standard therapies. Neuroplasticity and blood-brain barrier (BBB) integrity are the cornerstones of a well-functioning central nervous system, but they are vulnerable to an overly active NLRP3 inflammasome pathway that can also indirectly trigger the release of ET-1 and contribute to the ET system disturbance, which further damages stress resilience mechanisms. Here, the promising yet unexplored antidepressant potential of dapagliflozin (Dapa), a sodium-glucose co-transporter-2 inhibitor, was investigated by assessing its role in the modulation of the NLRP3 inflammasome pathway and ETBR signal transduction, and their impact on neuroplasticity and BBB integrity in an animal model of depression. Dapa (1 mg/kg/day; p.o.) with and without BQ-788 (1 mg/kg/day; i.p.), a specific ETBR blocker, were administered to adolescent male Wistar rats exposed to a 5-week chronic unpredictable stress protocol. The depressive animals demonstrated marked activation of the NLRP3 inflammasome pathway (NF-κB/NLRP3/caspase-1/IL/TNF-α), which was associated with both peripheral and central inflammatory responses. The ET system was disrupted, with noticeable reduction in miR-125a-5p and ETBR gene expression. Cortical ZO-1 expression was downregulated under the influence of NLRP3/TNF-α/miR-501-3p signaling, along with a prominent reduction in hippocampal BDNF and synapsin-1. With ETBR up-regulation being a cornerstone outcome, Dapa administration efficiently created an overall state of resilience, improved histopathological and behavioral variables, and preserved BBB function. These observations were further verified by the results obtained with BQ-788 co-administration. Thus, Dapa may exert its antidepressant action by reinforcing BBB integrity and promoting neuroplasticity through manipulation of the NLRP3/ET-1/ETBR/BDNF/ZO-1 axis, with a significant role for ETBR signaling. Graphical illustration for the proposed mechanisms of the anti-depressant potential of Dapa. Dapa suppressed NLRP3 inflammasome activation and assembly with subsequent inhibition of pro-inflammatory ILs. This results in attenuation of neuro-inflammation, BBB disruption, glial cell activation, TNF-α and ET-1 release, and the enhanced production of neurotrophins. The role of ETBR signaling was emphasized; Dapa possibly augmented ETBR expression, which is thought to boost neurotrophins production. The ETBR blocker, BQ-788, suppressed most of the positive outcomes of Dapa. Finally, miR-125a-5p and miR-501-3p that played major roles in these pathological pathways were modulated by Dapa. It is not yet clear whether Dapa has a direct or rather indirect effect on their expression. BBB, blood-brain barrier; Dapa, dapagliflozin; ET-1, endothelin-1; ETBR, endothelin B receptor; IL, interleukin; NF-κB, nuclear factor kappa B; NLRP3, nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing protein 3; TNF-α, tumor necrosis factor-α. Created with BioRender.com.

Exosome miR-501-3p Elevation Contributes to Progression of Vascular Stiffness.

Background: Tight junction (TJ) disruption and dysfunction are involved in the progression of arteriosclerosis. miR-501-3p regulates endothelial TJ protein-1, resulting in TJ disruption. Because exosomal microRNAs can travel to distant tissues and influence cell behavior, patients with elevated miR-501-3p may experience accelerated vascular disease progression secondary to miR-501-3p-induced reductions in TJ. This study investigated whether plasma exosome miR-501-3p levels are associated with vascular stiffness, an indicator for arteriosclerotic changes. Methods and Results: Fifty-one subjects (mean [±SD] age 70±8 years, 37% male) enrolled in a medical checkup program were recruited to the study. Brachial-ankle arterial pulse wave velocity (baPWV) and plasma exosome miR-501-3p expression were measured. Patients were divided into 2 groups depending on whether their miR-501-3p ∆Ct values were above ("High"; n=24) or below ("Low"; n=27) the cut-off levels determined by receiver operating characteristic (ROC) curve analysis. Median (interquartile range) baPWV levels were significantly higher in the miR-501-3p High than Low group (1,664 [1,496-1,859] vs. 1,450 [1,353-1,686] cm/s, respectively; P<0.05). Multivariate logistic regression analysis showed a significant association between increased baPWV and High miR-501-3p expression (odds ratio 4.66). At follow-up visits (mean 62 months later), baPWV remained significantly higher in the miR-501-3p High than Low group (1,830 [1,624-2,056] vs. 1,620 [1,377-1,816] cm/s, respectively; P<0.05). Conclusions: High expression levels of exosome miR-501-3p contribute to arteriosclerotic changes.

MiR-501-3p promotes osteosarcoma cell proliferation, migration and invasion by targeting BCL7A.

Increasing numbers of evidences have demonstrated that microRNAs (miRNAs) play an important role in osteosarcoma (OS) cell functions. MiR-501-3p has been reported to play an important role in several types of tumors, including prostate cancer and hepatocellular carcinoma. However, the biological function and potential mechanism of miR-501-3p in OS have not been well investigated until now. Here, we analyzed the expression of miR-501-3p in OS tissues and cell lines and its clinical significance in OS patients. Quantitative reverse transcription PCR showed miR-501-3p was significantly up-regulated in OS tissues and cell lines. Up-regulated miR-501-3p expression was associated with TNM stage, distal metastasis and worse prognosis in OS patients. MiR-501-3p knockdown and overexpression were achieved by miR-501-3p inhibitor and mimics transfection, respectively. CCK-8, colony formation and transwell assays showed that miR-501-3p knockdown in U2OS and Saos-2 cells suppressed, while miR-501-3p overexpression in Saos-2 cells promoted cell proliferation, migration and invasion. Moreover, luciferase reporter assay supporting BCL7A was a target of miR-501-3p and its expression was increased by miR-501-3p inhibitor, but inhibited by miR-501-3p mimics. By performing rescue experiments, we further demonstrated that BCL7A was a downstream functional regulator involved in miR-501-3p promoting OS cell functions. In summary, our findings suggest that miR-501-3p targets BCL7A may provide novel therapeutic targets for the treatment of OS.

LINC00452 promotes ovarian carcinogenesis through increasing ROCK1 by sponging miR-501-3p and suppressing ubiquitin-mediated degradation.

Ovarian cancer refers to all sorts of cancerous growth that starts from the ovary. Dysregulation of long non-coding RNAs (lncRNAs) is associated with ovarian cancer development and progression. Cellular expression and localization of LINC00452 in ovarian cancer cells were detected by qPCR and FISH. The roles of LINC00452 in ovarian carcinogenesis were characterized by MTT, transwell and colony-formation assays in vitro as well as xenograft mouse model. The underlying mechanism was explored by microarray, RIP, Co-IP and luciferase reporter assays. This study identified a novel lncRNA LINC00452 being elevated in both ovarian cancer cells and tumor tissues in patients. Such aberrant expression of LINC00452 was negatively correlated with relapse-free survival of ovarian cancer patients. Overexpression of LINC00452 potentiated CaOV3 cell viability, migration and invasion in vitro as well as xenograft tumor growth in vivo. Evidence from the current study suggests that the carcinogenicity of LINC00452 is partially due to competitive sponging of miR-501-3p followed with release of repression on the ROCK1, a key effector in Rho signaling pathway. Irrespective of its miRNA sponge function, LINC00452 is capable of preventing ROCK1 protein from ubiquitin/proteasome-mediated degradation via their mutual physical interaction. Our study makes LINC00452 a potential therapeutic target for ovarian cancer.

Circ-ZNF609 Accelerates the Radioresistance of Prostate Cancer Cells by Promoting the Glycolytic Metabolism Through miR-501-3p/HK2 Axis.

BACKGROUND: The development of radioresistance remains the obstacle for prostate cancer (PCa) treatment. Here, we explored the role and potential mechanism of circular RNA zinc finger protein 609 (circ-ZNF609) in the radioresistance of PCa cells. MATERIALS AND METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of circ-ZNF609, microRNA-501-3p (miR-501-3p) and hexokinase 2 (HK2) messenger RNA (mRNA). The viability, apoptosis, metastasis and radioresistance of PCa cells were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometry, transwell assays and colony formation assay. The glycolytic rate was assessed through measuring the glucose consumption and lactate production using fluorescence-based glucose and lactate assay kits. The target interaction between miR-501-3p and circ-ZNF609 or HK2 was predicted by StarBase software and confirmed by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. The protein level of HK2 was detected by Western blot assay. In vivo tumor growth assay was used to explore the role of circ-ZNF609 in the radioresistance of PCa in vivo. RESULTS: Circ-ZNF609 was abnormally up-regulated in PCa tissues and cell lines. Circ-ZNF609 silencing hampered the viability, metastasis, radioresistance and promoted the apoptosis through suppressing cell glycolysis. MiR-501-3p was a direct target of circ-ZNF609, and si-circ-ZNF609-induced influence in PCa cells was partly alleviated by the addition of anti-miR-501-3p. MiR-501-3p functioned through directly interacting with and down-regulating HK2. HK2 was modulated by circ-ZNF609/miR-501-3p axis in PCa cells. Circ-ZNF609 silencing enhanced the radiosensitivity of PCa cells in vivo. CONCLUSION: Circ-ZNF609 promoted the progression and radioresistance of PCa cells through accelerating the glycolysis via miR-501-3p/HK2 axis, providing promising targets for improving the prognosis of PCa patients.

MiR-501-3p functions as a tumor suppressor in non-small cell lung cancer by downregulating RAP1A.

MicroRNA-501-3p (miR-501-3p) has been reported to play tumor-suppressive roles in different cancers; however, its expression pattern and biological function in non-small cell lung cancer (NSCLC) remain unknown. In this study, we noted downregulation of miR-501-3p in NSCLC tissues and cell lines. Functional assays showed that overexpression of miR-501-3p suppressed NSCLC cell proliferation, clonogenicity, migration, and invasion. Moreover, miR-501-3p overexpression attenuated in vivo tumor growth in a nude mouse model. In terms of the mechanism, RAP1A was identified as a novel target of miR-501-3p. Overexpression of RAP1A strongly attenuated the inhibitory effects of miR-501-3p on the capacity of NSCLC cells for proliferation and motility. In the clinical samples of NSCLC, miR-501-3p levels negatively correlated with RAP1A expression, which was upregulated in NSCLC. Collectively, these results indicate that miR-501-3p acts as a tumor suppressor in NSCLC by directly targeting RAP1A mRNA and may serve as a theranostic biomarker for patients with NSCLC.

MicroRNA-501-3p restricts prostate cancer growth through regulating cell cycle-related and expression-elevated protein in tumor/cyclin D1 signaling.

MicroRNA-501-3p (miR-501-3p) has been reported as a novel cancer-related miRNA in many types of cancer. However, the precise biological function of miR-501-3p in prostate cancer remains unknown. In this study, we aimed to investigate the regulatory effect and mechanism of miR-501-3p on cell growth of prostate cancer cells. We found that miR-501-3p expression was significantly downregulated in prostate cancer tissues and cell lines. Gain-of-function experiments showed that upregulation of miR-501-3p expression significantly decreased cell proliferation and colony formation, and induced cell cycle arrest in the G0/G1 phase. Bioinformatics analysis predicted that cell cycle-related and expression-elevated protein in tumor (CREPT) was a potential target gene of miR-501-3p., and the results of our luciferase reporter assay confirmed that miR-501-3p bound to the 3'-untranslated region of CREPT at the predicted binding site. Moreover, miR-501-3p was shown to negatively regulate CREPT expression in prostate cancer cells. Correlation analysis showed that miR-501-3p was inversely correlated with CREPT expression in prostate cancer tissues. Knockdown studies revealed that miR-501-3p regulated the expression of cyclin D1 by targeting CREPT. Additionally, the inhibitory effect of miR-501-3p on prostate cancer cell growth was partially reversed by CREPT overexpression. Overall, these results suggest that miR-501-3p restricts prostate cancer cell growth by targeting CREPT to inhibit the expression of cyclin D1. These findings indicate that the miR-501-3p/CREPT/cyclin D1 axis plays a crucial role in the progression of prostate cancer and may serve as potential therapeutic target.

Serum microRNA miR-501-3p as a potential biomarker related to the progression of Alzheimer's disease.

MicroRNAs (miRNAs) are attractive molecules to utilize as one of the blood-based biomarkers for neurodegenerative disorders such as Alzheimer's disease (AD) because miRNAs are relatively stable in biofluid, including serum or plasma. To determine blood miRNA biomarkers for AD with next-generation sequencing genome-wide, we first surveyed 45 serum samples. These came from 27 AD patients and 18 controls (discovery set) that underwent autopsy within two weeks after their serum sampling and were neuropathologically diagnosed. We found that three miRNAs, hsa-miR-501-3p, hsa-let-7f-5p, and hsa-miR-26b-5p, were significantly deregulated between the AD samples and the controls. The deregulation for hsa-miR-501-3p was further confirmed by quantitative reverse transcription polymerase chain reaction (PCR) in a validation set composed of 36 clinically diagnosed AD patients and 22 age-matched cognitively normal controls with a sensitivity and specificity of 53% and 100%, respectively (area under the curve = 0.82). Serum hsa-miR-501-3p levels were downregulated in AD patients, and its lower levels significantly correlated with lower Mini-Mental State Examination scores. Contrary to its serum levels, we found that hsa-miR-501-3p was remarkably upregulated in the same donors' AD brains obtained at autopsy from the discovery set. The hsa-miR-501-3p overexpression in cultured cells, which mimicked the hsa-miR-501-3p upregulation in the AD brains, induced significant downregulation of 128 genes that overrepresented the Gene Ontology terms, DNA replication, and the mitotic cell cycle. Our results suggest that hsa-miR-501-3p is a novel serum biomarker that presumably corresponds to pathological events occurring in AD brains.