miR-200a Levels Research Articles

Down Regulated Expression Levels of miR-27b and miR-451a as a Potential Biomarker for Triple Negative Breast Cancer Patients Undergoing TAC Chemotherapy.

Background:Triple negative breast cancer (TNBC) is associated with poor prognosis, aggressive phenotype(s) of tumours, partial chemotherapy response, and lack of clinically proven therapies. MicroRNAs (miRNAs) can target and modulate key genes that are involved in TNBC chemotherapy. Deregulated miRNA expression is highly involved in anti-cancer drug resistance phenotype and thus, miRNAs tend to be promising candidates for prediction of chemotherapy response and recurrence. Aim:This study aimed to investigate the expression levels of selected miRNAs (miR-21, miR-27b, miR-34a, miR-182, miR-200c and miR-451a) in cancerous and normal adjacent tissues of TNBC patients and to correlate with the clinicopathological data. Methods:Forty-one (41) FFPE tissue block of histopathologically confirmed TNBC patients was collected. Total RNA from the cancerous and adjacent non-cancerous tissues were isolated, transcribed, and pre-amplified. The relative expression level of miRNAs in tumour and normal adjacent tissues of TNBC patients was analysed using qRT-PCR. Results: Out of six miRNAs studied, the relative expression of miR-27b and miR-451a were found to be significantly lower in cancerous as compared to normal adjacent tissues of TNBC patients. In addition, a significant down regulation of miR-451a was also observed in infiltrating ductal carcinoma subtype, stages I and II, in both grade II and III, premenopausal and postmenopausal as well as in those with positive axillary lymph node metastases.Conclusion:The results suggest the possible utilization of miR-27b and miR-451a expression levels as potential predictive risk markers for TNBC patients undergoing TAC chemotherapy.

Abstract P5-10-06: Arginine- depleting enzyme, pegylated arginase, isolated from beef-liver tissue induces growth inhibition, apoptosis, cell cycle arrest and inhibits induced mammary tumors: A promising strategy for human breast cancer treatment in vitro and in vivo

Abstract Arginine auxotrophy occurs in certain tumor types, renders tumors vulnerable to treatment with arginine-degrading enzymes, and leads to their rapid demise. Arginine is a semi-essential amino acid but is essential for rapidly proliferating cells. Thus, Arginine deprivation can be exploited as a potential targeted therapy for the treatment of various cancers. Arginase, an enzyme of the urea cycle, converts L-arginine to L-ornithine, which is the precursor of polyamines that are essential components of cell proliferation. However, Arginine deprivation and Arginase clinical implications in human breast cancer have not been fully elucidated. The objective of the current study was to investigate the potential role of the Arginine degrading enzyme, Arginase on two breast cell lines that have different differentiation patterns, Luminal A breast cancer (T-47D), and TNBC (MDA-MB-231), respectively. Herein, Arginase was extracted and purified from the beef liver and its specific activity was 0.2 IU/mg protein yielded 49% compared to the crude beef liver Arginase. The purified Arginase is covalently bound via a succinamide propionic acid (SPA) linker to polyethylene glycol (PEG) of molecular weight 5,000 to enhance the half-life time compared to the native enzyme with a maximum specific activity between 37~55°C and pH 7~9. The potential influence of extracted Peg-Arginase was examined on cell proliferation, clonogenic assays, morphological alteration, and oncogenic signaling pathways of STAT3, β-catenin, and miR-23a. Our data showed that Peg-Arginase treatment displayed a significant inhibition in cell proliferation and viability in a dose-dependent manner in both breast cancer cell lines compared to control after 48h. Furthermore, mechanistic studies showed that Peg-Arginase induced apoptosis that was associated with cell morphological changes, histone release from fragmented DNA, caspase-3 activation, and cell cycle arrest at S and/or G2/M phases in a dose-. dependent manner after 48h. Moreover, Peg-Arginase down-regulated miR-23a, STAT3, β-catenin, cyclin D1, and mutant p53 levels as well as increased p21and Bax, pro-apoptotic signal, in both breast cancer cells as determined by qRT-PCR and western blot analysis. In vivo, the effects of native- and Peg-Arginase enzymes were examined on MNU-induced mammary tumors in Albino-Wister rats. Our data showed that injecting 400 IU of native- or peg- Arginase twice per week for two weeks significantly inhibited the growth of MNU-induced mammary tumors by 58.8±4.1 % for peg-Arginase compared to 19.7±3.5 % for native-Arginase. Furthermore, the results also revealed a significant reduction in activities of ALT, AST, and ALP in the Peg-Arginase group compared to MNU untreated or native-Arginase groups, suggesting that Peg-Arginase may eliminate the hepatotoxic effect of the MNU. Furthermore, Peg-Arginase treatment reduced the localization and expression of oncogenic β-catenin in the nucleus compared to MNU-untreated and native-Arginase groups. All in all, our results suggested that the arginine-depleting enzyme, Peg-Arginase, exerted an anticancer effect against both luminal A and triple-negative breast cancer cell lines, through cell proliferation inhibition and apoptosis induction, which might provide a promising alternative strategy targeting oncometabolite in human breast cancer. Citation Format: Islam Yahiya, Asmaa Ali, Omnia Ali, Ahmed Sultan. Arginine- depleting enzyme, pegylated arginase, isolated from beef-liver tissue induces growth inhibition, apoptosis, cell cycle arrest and inhibits induced mammary tumors: A promising strategy for human breast cancer treatment in vitro and in vivo [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-10-06.

Effect of circRNA_FOXO3 rs12196996 polymorphism and FOXO3 rs2232365 polymorphism on survival rate and severity of intensive care unit-acquired sepsis

ABSTRACT The expression of circRNA_FOXO3 was found to be positively associated with the expression of Forkhead Box O3 (FOXO3), which is targeted and regulated by miR-23a. Polymorphisms in rs12196996 and rs2232365 have been reported in various diseases. In this study, we recruited intensive care unit (ICU)-acquired sepsis patients and grouped them according to their genotypes of rs12196996 and rs2232365. Quantitative real-time PCR was performed to analyze the expression of circRNA_FOXO3, FOXO3 mRNA, and miR-23a. ELISA was carried out to evaluate the abundance of cytokines and luciferase assay was used to explore the inhibitory role of miR-23a on circRNA_FOXO3 and FOXO3. Accordingly, we found that rs12196996 GG and rs2232365 AA were significantly correlated with prolonged survival of ICU-acquired sepsis patients. Rs12196996 GG and rs2232365 AA were also correlated with increased level of miR-23a, IL-10 and decreased level of TNF, IL-2, IFN, IL-6 and IL-1β in the peripheral blood cell samples of patients with ICU-acquired sepsis. The luciferase activity of wild-type (WT) circRNA_FOXO3 and FOXO3 were severely reduced by miR-23a. MiR-23a precursors could effectively suppress the expression of circRNA_FOXO3 and FOXO3 in the cells. Moreover, LPS-induced cell viability loss and dysregulation of cytokines were effectively restored by the knockdown of FOXO3 or circRNA_FOXO3 siRNA in the cells. This study revealed that the minor allele of rs12196996 polymorphism and rs2232365 polymorphism collaboratively contributed to the increased survival and suppressed severity of ICU-acquired sepsis.

MicroRNAs miR-451a and Let-7i-5p Profiles in Circulating Exosomes Vary among Individuals with Different Sickle Hemoglobin Genotypes and Malaria.

Sickle cell disease (SCD) occurs when two alleles of mutated hemoglobin (HbS or HbC) are inherited (HbSS and HbSC) rather than one (HbAS or HbAC), which indicates a person carries the sickle cell trait. The high prevalence of these two alleles in Africa have been associated with reduced malaria susceptibility. Recent in vitro research has been shown that microRNAs (miRNAs) miR-451a and let-7i-5p are differentially expressed in HbSS erythrocytes compared to healthy controls (HbAA) and are overexpressed in Plasmodium-infected malaria erythrocytes. However, these miRNAs have not been fully examined in the plasma of people with different sickle hemoglobin genotypes. Plasma circulating miRNAs are commonly encapsulated in extracellular vesicles, such as exosomes, and are thought to play a role in disease development. Circulating exosomal miR-451a and let-7i-5p were quantified from individuals with various hemoglobin genotypes (HbAA, HbAS, HbAC, HbSS, HbSC, and HbCC) with (+) and without (−) malaria. The results showed a higher level of exosomal let-7i-5p and miR-451a in HbSS-. Exosomal let-7i-5p and miR-451a levels were lower in HbSS+ compared to other genotypes. Based on the area under the curve (AUC) of the Receiver Operating Characteristics (ROCs), both exosomal miRNAs may be useful disease biomarkers for SCD with malaria. Finally, miR-451a and let-7i-5p modulate genes involved in inflammation, making them potential biomarkers of pathogenesis for both diseases.

MiR-23a suppression accelerates functional decline in the rNLS8 mouse model of TDP-43 proteinopathy

Skeletal muscle dysfunction may contribute to the progression and severity of amyotrophic lateral sclerosis (ALS). In the present study, we characterized the skeletal muscle pathophysiology in an inducible transgenic mouse model (rNLS8) that develops a TAR-DNA binding protein (TDP-43) proteinopathy and ALS-like neuropathology and disease progression; representative of >90% of all familial and sporadic ALS cases. As we previously observed elevated levels of miR-23a in skeletal muscle of patients with familial and sporadic ALS, we also investigated the effect of miR-23a suppression on skeletal muscle pathophysiology and disease severity in rNLS8 mice. Five weeks after disease onset TDP-43 protein accumulation was observed in tibialis anterior (TA), quadriceps (QUAD) and diaphragm muscle lysates and associated with skeletal muscle atrophy. In the TA muscle TDP-43 was detected in muscle fibres that appeared atrophied and angular in appearance and that also contained β-amyloid aggregates. These fibres were also positive for neural cell adhesion molecule (NCAM), but not embryonic myosin heavy chain (eMHC), indicating TDP-43/ β-amyloid localization in denervated muscle fibres. There was an upregulation of genes associated with myogenesis and NMJ degeneration and a decrease in the MURF1 atrophy-related protein in skeletal muscle. Suppression of miR-23a impaired rotarod performance and grip strength and accelerated body weight loss during early stages of disease progression. This was associated with increased AchRα mRNA expression and decreased protein levels of PGC-1α. The TDP-43 proteinopathy-induced impairment of whole body and skeletal muscle functional performance is associated with muscle wasting and elevated myogenic and NMJ stress markers. Suppressing miR-23a in the rNLS8 mouse model of ALS contributes to an early acceleration of disease progression as measured by decline in motor function.

Long Non-Coding RNA H19 Regulates Glioma Cell Growth and Metastasis via miR-200a-Mediated CDK6 and ZEB1 Expression.

Long non-coding RNAs (lncRNAs) serve essential roles on various biological functions. Previous studies have indicated that lncRNAs are involved in the occurrence, growth and infiltration of brain tumors. LncRNA H19 is key regulator in the pathogenesis of gliomas, but the underlying mechanisms of H19-regulated tumor progression remain unknown. Therefore, we investigated the effects and mechanism of action of lncRNA H19 on the homeostasis of glioma cells. As a novel oncogenic factor, up-regulation of H19 was able to promote the proliferation of glioma cells by targeting miR-200a. Furthermore, elevated miR-200a levels could reverse H19-induced cell growth and metastasis. Overexpression of miR-200a could significantly suppress the proliferation, migration and invasion of glioma cells. These biological behavior changes in glioma cells were dependent on the binding to potential target genes including CDK6 and ZEB1. CDK6 could promote cell proliferation and its expression was remarkably increased in glioma. In addition, up-regulation of miR-200a lead to reduction of CDK6 expression and inhibit the proliferation of glioma cells. ZEB1 could be a putative target gene of miR-200a in glioma cells. Thus, miR-200a might suppress cell invasion and migration through down-regulating ZEB1. Moreover, overexpression of miR-200a resulted in down-regulation of ZEB1 and further inhibited malignant phenotype of glioma cells. In summary, our findings suggested that the expression of H19 was elevated in glioma, which could promote the growth, invasion and migration of tumor cells via H19/miR-200a/CDK6/ZEB1 axis. This novel signaling pathway may be a promising candidate for the diagnosis and targeted treatment of glioma.

MiR-26a enhances colorectal cancer cell growth by targeting RREB1 deacetylation to activate AKT-mediated glycolysis.

We previously reported the inhibitory effects of microRNA-26a (miR-26a) on the conversion of pyruvate to acetyl coenzyme A in glucose metabolism by directly targeting pyruvate dehydrogenase protein X component in colorectal cancer (CRC) cells (Chen B et al., BMC Cancer 2014). Here, using microRNA in situ hybridization, we confirmed that miR-26a levels were elevated in 77 human CRC tissue samples and further investigated the key miR-26a-mediated metabolic regulation elements and signaling pathways in CRC cells through quantitative proteomic dissection combined with cancer cell biology and biochemical loss-of-function analysis. We found that AKT transcription signaling was a target pathway via miR-26a-mediated deacetylation modification of Ras-responsive element-binding protein 1 (RREB1) at the Lys-60 residue. miR-26a improved the deacetylation level of RREB1, thus contributing to RREB1 binding to the AKT1 promoter to activate AKT transcription and its related signaling pathway in glycolysis. Moreover, miR-26a promoted CRC tumorigenesis in CRC cells and subcutaneous xenograft mice. Thus, miR-26a is a key regulator of CRC tumorigenesis that mediates the deacetylation modification of RREB1 to enhance AKT1 transcription and downstream target gene expression in glycolysis for CRC growth.

MicroRNA-26a-5p Prevents Retinal Neuronal Cell Death in Diabetic Mice by Targeting PTEN

ABSTRACTAimTo explore the role of microRNA-26a-5p (miR-26a) in early diabetic retinal neuronal cell death and reveal the underlying mechanism(s).MethodsA streptozotocin (STZ)-induced diabetic mouse model was established using C57BL/6 J mice. Control or miR-26a mimic was intravitreally injected. Hematoxylin-eosin (H&E) and transmission electron microscopy (TEM) were used to observe the morphologic alterations in the retinal structure and ultrastructure, respectively. The expression of miR-26a and phosphatase and tensin homolog (PTEN) was assayed using qRT-PCR and western blotting, respectively. An immunofluorescence assay was used to investigate the distribution of PTEN expression in the retina. The expression of glial fibrillary acidic protein (GFAP) was measured to identify glial cell activation. The mRNA levels of IL-1β, NF-κB, and VEGF were examined to assess diabetic retinal inflammation.ResultsmiR-26a expression was decreased in retinal tissues of diabetic mice, and injection of miR-26a mimic restored the miR-26a level. Diabetic mice had significantly reduced neuroretinal thickness and ganglion cell number; miR-26a mimic delayed the thinning of neuroretinal layers and the loss of ganglion numbers. TEM showed damaged ultrastructure of retinal ganglions in diabetic mice, while miR-26a mitigated the damages. PTEN expression was increased mainly in the inner and outer nuclear layer of the retina in diabetic mice; miR-26a mimics lowered PTEN expression. GFAP, IL-1β, NF-κB, and VEGF expression were significantly increased in the diabetic mice, and intravitreal delivery of miR-26a resulted in a down-regulated expression of these factors.ConclusionmiR-26a can protect against retinal neuronal impairment in diabetic mice by down-regulating PTEN, highlighting the potential of miR-26a as a target for DR treatment.

Long non-coding RNA ZNF674-AS1 regulates miR-23a/E-cadherin axis to suppress the migration and invasion of non-small cell lung cancer cells.

BackgroundNon-small cell lung cancer (NSCLC) is one of the most common malignant tumors. The prognosis of patients with advanced NSCLC is poor due to metastasis. In recent years, the role of long non-coding RNAs (lncRNAs), a class of non-coding RNA molecules, in NSCLC has become an increasingly popular focus of studies. This study aimed to investigate the effects of ZNF674-AS1 and microRNA (miR)-23a on the migration and invasion abilities of NSCLC cells in vitro and explore the underpinning molecular mechanisms.MethodsThe expression levels of ZNF674-AS1 and miR-23a in NSCLC tissues and cells were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Scratch test and transwell test were used to detect the effects of ZNF674-AS1 and miR-23a on the migration and invasion of NSCLC cells. The luciferase reporter gene experiment was used to verify miRNA targets. Western blot experiments were used to analyze protein expression.ResultsZNF674-AS1 was downregulated in NSCLC tissues and cells, and inhibited the migration and invasion of NSCLC cells in vitro. In contrast, the expression of miR-23a, a downstream target of ZNF674-AS1, was increased in NSCLC tissues and cells. We found that miR-23a could antagonize the role of ZNF674-AS1 in NSCLC. E-cadherin was identified as a downstream target gene of miR-23a, and miR-23a could directly inhibit its expression.ConclusionsZNF674-AS1 inhibits the migration and invasion of NSCLC cells by regulating a miR-23a/E-cadherin axis. ZNF674-AS1 and miR-23a could become potential therapeutic targets for NSCLC.

Reprogrammed lung epithelial cells by decrease of miR-451a in extracellular vesicles contribute to aggravation of pulmonary fibrosis.

Extracellular vesicles (EVs) play novel roles in homeostasis through cell-to-cell communication in human airways via transferring miRNAs. However, the contribution of EV miRNAs to pulmonary phenotypic homeostasis is not clearly understood. Hence, the aim of this study was to elucidate the functional role of miRNAs obtained from epithelium-derived EVs in lung fibrogenesis. Pulmonary fibrosis was induced by exposure of polyhexamethylene guanidine phosphate (PHMG-p)-instilled mice. In histopathological changes, a clear phenotypic change was observed in bronchial epithelium. For figuring out the role of EVs derived from conditioned media of untreated cells (EV-Con) and PHMG-p-treated BEAS-2B (EV-PHMG), significant increase in EVs released from PHMG-p-treated BEAS-2B was detected. Functional analysis with targets of differentially expressed miRNAs in EVs was annotated to epithelial-mesenchymal transition (EMT). Especially, the most abundant miRNA, miR-451a, was downregulated in EV of PHMG-p-treated BEAS-2B cells. We found that odd-skipped related 1 (OSR1) was a putative target for miR-451a, which had been known as a transcription factor of several fibrosis-associated genes. Transfer of decreased miR-451a via EV-PHMG upregulated OSR1 and induced EMT compared to Con-EV-treated cells. In pulmonary fibrosis mice, miR-451a levels were significantly reduced in EV derived from bronchoalveolar lavage fluid and OSR1 expression was increased in lung tissues of mice with PHMG-p exposure. MiR-451a-transfected EVs markedly alleviated fibrogenesis in the PHMG-p-exposed lungs. Low level of miR-451a in EVs modulated EMT and fibrogenesis in recipient cells by increasing OSR1 levels in vitro and in vivo. Our results suggest that transferring EV miR-451a induces anti-fibrotic autocrine effect by downregulating its target, OSR1 maintaining pulmonary homeostasis disrupted by PHMG-p exposure, which can be a potential therapeutic target.

Serum MiRNA-23a as a Diagnostic and Prognostic Biomarker of Hepatitis C Related Hepatocellular Carcinoma

Background: Micro-ribonucleic acids (MiRNAs) are small, non-coding RNA molecules which regulate gene expression. Several miRNAs including miR-23a were found to be frequently deregulated in hepatocellular carcinoma (HCC). Objective: This study aimed to evaluate serum miR-23a as a biomarker of hepatitis C related HCC. Methods: This study was conducted on 60 hepatitis C virus (HCV) infected patients (group I: without cirrhosis, group II with cirrhosis and group III with HCV associated HCC) and a control group of 20 healthy volunteers. All patients were submitted to history taking, clinical examination in addition to categorization and staging of HCC patients. Following extraction of RNA from serum samples, quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed. Calculation of serum miR-23a was done using the comparative cycle threshold (Ct) method (2–ΔΔCT). Results: Serum miR-23a levels (2-ΔΔCT) were significantly higher in cirrhotic and HCC patients compared to chronic hepatitis C patients (CHC). However, no significant difference was noted between cirrhotic and HCC patients. The sensitivity and specificity of miR-23a levels for discriminating HCC patients from cirrhotic patients were 55% and 65%, respectively. MiR-23a levels had sensitivity of 90% and specificity of 70% for discriminating metastatic from non-metastatic HCC patients. Conclusion: Higher miR-23a levels were detected among metastatic HCC patients than among those without metastasis. The sensitivity and specificity of miR-23a levels for discriminating HCC patients from cirrhotic patients were lower than those of alpha fetoprotein (AFP).

Circulating skeletal muscle related microRNAs profile in Piedmontese cattle during different age

Piedmontese cattle is known for double-muscle phenotype. MicroRNAs (miRNAs) play important role as regulators in skeletal muscle physiological processes, and we hypothesize that plasma miRNAs expression profiles could be affected by skeletal muscle growth status related to age. Plasma samples of cattle were collected during four different ages from first week of life until the time of commercial end of the fattening period before slaughter. Small-RNA sequencing data analysis revealed the presence of 40% of muscle-related miRNAs among the top 25 highly expressed miRNAs and, 19 miRNAs showed differential expression too. Using qRT-PCR, we validated in a larger bovine population, miRNAs involved in skeletal muscle physiology pathways. Comparing new-born with the other age groups, miR-10b, miR-126-5p, miR-143 and miR-146b were significantly up-regulated, whereas miR-21-5p, miR-221, miR-223 and miR-30b-5p were significantly down-regulated. High expression levels of miR-23a in all the groups were found. Myostatin, a negative regulator of skeletal muscle hypertrophy, was predicted as the target gene for miR-23a and miR-126-5p and we demonstrated their direct binding. Correlation analysis revealed association between miRNAs expression profiles and animals’ weights along the age. Circulating miRNAs could be promising for future studies on their biomarker potentialities to beef cattle selection.

Correlation between serum level of miRNA-106a expression with clinicopathological characteristics and prognosis of patients with renal cell carcinoma

To analyze the expression of microRNA-106a(miR-106a) in renal cell carcinoma (RCC) and its correlation with clinicopathological characteristics and prognosis of patients. Serum samples of 64 patients with newly diagnosed RCC were collected as the study group, and serum samples of 40 healthy individuals were used as the control group. Real-time fluorescence quantitative PCR was used to determine the expression level of miR-106a in each group. The correlation between miR-106a expression and clinicopathological characteristics of the patients was studied with single factor analysis and multiple Logistic regression model. Kaplan-Meier survival curve was used to analyze its correlation with the prognosis of patients. Before surgery, compared with the control group (1.17± 0.58), RCC patients with high- (9.15± 0.96) and low-expression(3.45± 0.37) had increased expression of miR-106a. Postoperatively, the expression level of miR-106a in both groups of patients decreased to 1.53± 0.18 and 1.75± 0.21, respectively. The area under the curve (AUC) of the diagnostic value of serum miR-106a for RCC was 0.782 (95% CI: 0.661-0.902). With an optimal cutoff value of 0.531, the sensitivity was 78.10% and the specificity was 75.00%. Serum miR-106a level of RCC patients with TNM stage T3 or T4, clinical stage II or III, lymph node metastasis, and recurrence were significantly increased. The high expression of serum miR-106a in RCC patients has an independent relationship with the tumor TNM stage and lymph node metastasis. Of the 64 follow-up patients, 4 were lost and 30 had died. Among them, the median survival time of patients in the miR-106a high expression group was 30 months, which was significantly shorter than that of the low expression group (52 months). The serum level of miR-106a is elevated in RCC patients, and may be used as a molecular marker for the diagnosis of RCC. High serum expression of miR-106a is an independent predictor for tumor TNM stage and lymph node metastasis, as well as an independent predictor for poor prognosis of RCC patients.

Urinary exosomal microRNA profiling in type 2 diabetes patients taking dipeptidyl peptidase-4 inhibitor compared with sulfonylurea.

BackgroundDipeptidyl peptidase-4 (DPP-4) inhibitor has been reported to have kidney-protective benefits. To elucidate how antidiabetic agents prevent diabetic kidney disease progression, it is important to investigate their effect on the kidney environment in type 2 diabetes mellitus (DM) patients. Herein, we investigated the expression pattern of urinary exosome-derived microRNA (miRNA) in patients taking a combination of DPP-4 inhibitor and metformin (DPP-4 inhibitor group) and compared them with patients taking a combination of sulfonylurea and metformin (sulfonylurea group).MethodsThis was a prospective study involving 57 patients with type 2 DM (DPP-4 inhibitor group, n = 34; sulfonylurea group, n = 23) and healthy volunteers (n = 7). We measured urinary exosomal miRNA using the NanoString nCounter miRNA array (NanoString Technologies) across the three groups (n = 4 per each group) and validated findings using real-time polymerase chain reaction.ResultsTwenty-one differentially expressed candidate miRNAs were identified, and six (let-7c-5p, miR-23a-3p, miR-26a-3p, miR-30d, miR-205, and miR-200a) were selected for validation. Validation showed no significant difference in miRNA expression between the DPP-4 inhibitor and sulfonylurea groups. Only miR-23a-3p was significantly overexpressed in the diabetes group compared with the control group (DPP-4 inhibitor vs. control, p = 0.013; sulfonylurea vs. control, p = 0.007). This trend was consistent even after adjusting for age, sex, and body mass index.ConclusionThere was no significant difference in urine exosome miRNA expression between diabetic participants taking DPP-4 inhibitor and those taking sulfonylurea. The miR-23a levels were higher in diabetic participants than in nondiabetic controls.

MicroRNA-200a inhibits TGF-β-induced epithelial-mesenchymal transition in human ovarian carcinoma cells by downregulating SOX4 expression

IntroductionThe microRNA-200 (miR-200) family and sex-determining region Y-box 4 (SOX4) could regulate EMT phenotypes, which is important to the process of tumor pathological. This study explored the association of miR-200a with SOX4 in transforming growth factor (TGF)-β-induced EMT of OC cells.Material and methodsFor the in vitro experiments, hunam CO cells subjected to TGF-β was used to induce EMT; the activity of miR-200a was selectively inhibited or overexpressioned by miR-200a inhibitor and mimics, respectively. Small interfering RNAs against SOX4 (si-SOX4) were used to inhibit SOX4 expression in human OC cell lines.ResultsDecreased miR-200a and increased SOX4 levels were detected in patients with OC and these changes were closely related to the International Federation of Gynaecology and Obstetrics stage, ovarian tumor biomarker CA125 level, lymph node status and tumor size. The TGF-β-treated cells increased the miR-200a level, decreased the SOX4 level and prompted EMT properties, including a reduction in epithelial marker (e-cadherin), induction in interstitial markers (vimentin and n-cadherin), and enhancement of proliferation, migration and invasion. The OC cells were transducted with miR-200a mimic and the overexpression cells were subsequently treated with TGF-β, decreased SOX4 expression and EMT properties were detected. Also, in miR-200a inhibited cells, TGF-β increased SOX4 expression and EMT properties. Moreover, SOX4 silencing weakened the effect of the miR-200a inhibitor.ConclusionsOverall, these results provide a link between miR-200a and SOX4 in OC pathogenesis and indicate that miRNA-200a inhibits EMT by downregulating SOX4 expression in human OC cells.

Molecular mechanism of microRNA-26a regulation of phosphatase and tensin homolog gene in condyloma acuminatum and penile squamous cell carcinoma.

ObjectiveTo investigate the expression levels and mechanisms of microRNA (miRNA) 26a (miR-26a) and phosphatase and tensin homolog (PTEN) in patients with human papillomavirus (HPV)-induced condyloma acuminatum (CA) and penile squamous cell carcinoma (PSCC).MethodsThirty-one patients with HPV-positive CA and 28 with HPV-positive PSCC were included in this retrospective, cross-sectional study. PTEN mRNA and miR-26a levels in lesion tissues, blood, and urine were analyzed by quantitative reverse transcription polymerase chain reaction, and PTEN protein was detected by western blot and enzyme-linked immunosorbent assay. Cell proliferation was assessed by MTT assay. The interaction between miR-26a and PTEN was predicted by bioinformatics analysis and confirmed by dual luciferase reporter assayResultsPTEN mRNA and protein levels were significantly lower and miR-26a levels were significantly higher in all samples from patients with PSCC compared with the CA group. Bioinformatics analysis and luciferase reporter assay confirmed PTEN as a target gene of miR-26a. Up-regulation of miR-26a significantly increased the proliferation of Penl1 PSCC cells.ConclusionsPTEN expression is down-regulated and miR-26a levels are up-regulated in PSCC compared with CA. PTEN is a direct target gene of miR-26a. These results suggest that miR-26a might regulate HPV-positive progression from CA to PSCC through modulating PTEN.

P-232 Prognostic inflammatory and microRNA biomarkers in stage IV colorectal cancer patients

Colorectal cancer (CRC) is the second leading cause of cancer deaths worldwide. Stage IV CRC patients have poor prognosis with a five-year survival rate of 14%. Systemic inflammation has a key determinant role in clinicopathological outcomes among CRC patients. Furthermore, microRNA (miRNA), small RNA non-coding molecules, have been detected to be dysregulated in tissues, as well as in biological fluids taken from colorectal cancer patients. Consequently, we are interested in investigating the prognostic potential of inflammatory markers and miRNA in Stage IV colorectal cancer patients. C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), serum albumin, fibrinogen, ferritin, haptoglobin, lymphocyte/monocyte ratio, neutrophil/lymphocyte ratio and platelets/lymphocytes ratio, and Glasgow prognostic score were prospectively evaluated pre-chemotherapy in 67 Stage IV CRC patients presenting at the American University of Beirut Medical Center between May 2018 till November 2020. miRNA (miR-21, miR-19a, miR-23a, miR-29a, miR-155, miR-210, miR-145, miR-203, miR-31 and miR-345) expression was determined using reverse transcription real-time PCR in plasma samples collected prechemotherapy. Cutoff values of the biomarkers were determined using ROC curve. Statistical analysis was performed with SPSS version 25. The median overall survival for the 67 patients was not reached and the median progression-free survival was 13 months. Patients with low pre-chemotherapy WBC count (< 9600 /cu.mm), CRP (< 3.1 mg/L) or ESR (< 20.5 mm/hr) experienced significant improvements in progression-free survival (P=0.012, P=0.016, P < 0.05 respectively) compared with patients with high pre-chemotherapy CRP, ESR or WBC count. Patients with low pre-chemotherapy haptoglobin (< 3.1 g/L) had significantly better overall survival (OS) (P < 0.05) compared with patients with high pre-chemotherapy haptoglobin. Low levels of miR-23a and miR-19a were significantly associated with better PFS (P=0.021 and P=0.031 respectively). Elevated levels of miR-345 and miR-210 were correlated with better OS (P=0.029 and P=0.035 respectively). Subsequent Cox regression multivariate analysis showed that high pre-chemotherapy of CRP, WBC count, miR-19a, and miR-23a were independent favorable prognostic factors for PFS with a HR=3.8; 95%CI: 1.149-12.648; HR=2.426; 95%CI: 1.159-5.075, HR= 2.37; 95%CI: 1.092-5.15 and, HR=2.31; 95%CI: 1.029-4.414), respectively. CRP level, WBC count, miR-19a, and miR-23a are potential prognostic biomarkers for progression-free survival in Stage IV colorectal cancer that need further validation in larger cohorts.

Retraction Note: MicroRNA-23a enhances migration and invasion through PTEN in osteosarcoma.

To investigate the biological significance of abundant microRNA-23a (miR-23a) expression in osteosarcoma and its correlation with PTEN in the pathogenesis of osteosarcoma migration and invasion. The human osteosarcoma cell lines MG63, HOS58 and SaoS-2, and the human normal osteoblasts (hFOB1.19) were grown in RPMI 1640 medium supplemented with 10% fetal bovine serum. Gene and protein levels of miR-23a and PTEN were examined to determine the molecular relationship between them in the pathogenesis of osteosarcoma. Inhibition of miR-23a effectively reduced migration and invasion of osteosarcoma cell lines. Bioinformatics and luciferase-reporter assay revealed that miR-23a specifically targeted the 3'-untranslational region of PTEN and regulated its expression. Downregulation of PTEN enhanced migration and invasion of osteosarcoma cell lines. Furthermore, in tumor tissues obtained from osteosarcoma patients, the expression of miR-23a was negatively correlated with PTEN and the high expression of miR-23a combined with low expression of PTEN might serve as a risk factor for cancer patients. Besides, miR-23a-mediated suppression of PTEN led to activation of AKT/ERK pathways and epithelial-mesenchymal transition (EMT) in osteosarcoma cells, and finally enhanced the activity of osteosarcoma cell proliferation and movement and promoted osteosarcoma xenograft tumor growth in mouse models. Our study showed that miR-23a, by downregulation of PTEN, enhanced migration and invasion in osteosarcoma cells.

The miRNA expression profile directly reflects the energy metabolic differences between slow and fast muscle with nutritional regulation of the Chinese perch (Siniperca chuatsi)

Fish skeletal muscles are composed of two distinct types, slow and fast muscles, and they play important roles in maintaining the body's movement and energy metabolism. The two types of muscle are easy to separate, so they are often used as the model system for studies on their physiological and functional characteristics. In this study, we revealed that the carbohydrate and lipid metabolic KEGG pathways are different between slow and fast muscles of Chinese perch with transcriptome analysis. In fast muscle, glucose metabolism was catabolic with higher glycolysis capacity, while in slow muscle, glucose metabolism was anabolic with more glycogen synthesis. In addition, oxidative metabolism in slow muscle was stronger than that in fast muscle. By analyzing the expression levels of 40 miRNAs involved in metabolism in the muscles of Chinese perch, 18 miRNAs were significantly upregulated and 7 were significantly downregulated in slow muscle compared with fast muscle. Based on functional enrichment analysis of their target genes, the differential expression levels of 17 miRNAs in slow and fast muscles were reflected in their carbohydrate and lipid metabolism. Among these, 15 miRNAs were associated with carbohydrate metabolism, and 6 miRNAs were associated with lipid metabolism. After 3 days of starvation, the expression levels of 15 miRNAs involved in glucose metabolism in fast and slow muscles increased. However, after 7 days of starvation, the mRNA levels of miR-22a, miR-23a, miR-133a-3p, miR-139, miR-143, miR-144, miR-181a and miR-206 decreased to basal levels. Our data suggest that the possible reason for the difference in glucose and lipid metabolism is that more miRNAs inhibit the expression of target genes in slow muscle.

Circulating Levels of MicroRNAs in Hypertrophic Cardiomyopathy: The Relationship With Left Ventricular Hypertrophy, Left Atrial Dilatation and Ventricular Depolarisation-Repolarisation Parameters

MicroRNAs are small, endogenous, non-coding RNAs that regulate the expression of many genes. It has recently been shown that circulating microRNAs may be biomarkers of hypertrophy and fibrosis in patients with hypertrophic cardiomyopathy (HCM). To determine whether circulating levels of microRNAs involved in HCM are associated with electrocardiographic and echocardiographic parameters. This study enrolled 20 patients with familial HCM and 20 blood donors. Peripheral serum levels of miR-29a-3p, miR-199a-5p and miR-451a were assessed by quantitative real-time polymerase chain reaction and compared with levels in the control group. Whether circulating levels of miRNAs in HCM patients correlated with electrocardiographic and echocardiographic parameters was also assessed. Median circulating levels of miR-29a and miR-451a were significantly higher in HCM than the control group. Median miR-199a levels did not differ between groups. However, circulating levels of miR-199a negatively correlated with corrected QT duration (Bazett formula). Median miR-29a levels positively correlated with QRS duration. In addition, circulating levels of miR-29a correlated with maximal wall thickness, left ventricular mass index and left atrial volume index. The data suggested that serum levels of miR-29a and miR-451a were significantly increased in HCM patients. As the circulating level of miR-29a correlated with QRS duration, left ventricular hypertrophy and left atrial dilatation, the serum miR-199a level negatively correlated with corrected QT duration. These miRNAs may be seen as potential biomarkers for further research in HCM pathophysiology.