Association of Glutathione Peroxidase 3 (GPx3) and miR-196a with Carbohydrate Metabolism Disorders in the Elderly.

The escalating prevalence of carbohydrate metabolism disorders (CMDs) prompts the need for early diagnosis and effective markers for their prediction. Hyperglycemia, the primary indicator of CMDs including prediabetes and type 2 diabetes mellitus (T2DM), leads to overproduction of reactive oxygen species (ROS) and oxidative stress (OxS). This condition, resulting from chronic hyperglycemia and insufficient antioxidant defense, causes damage to biomolecules, triggering diabetes complications. Additionally, aging itself can serve as a source of OxS due to the weakening of antioxidant defense mechanisms. Notably, previous research indicates that miR-196a, by downregulating glutathione peroxidase 3 (GPx3), contributes to insulin resistance (IR). Additionally, a GPx3 decrease is observed in overweight/obese and insulin-resistant individuals and in the elderly population. This study investigates plasma GPx3 levels and miR-196a expression as potential CMD risk indicators. We used ELISA to measure GPx3 and qRT-PCR for miR-196a expression, supplemented by multivariate linear regression and receiver operating characteristic (ROC) analysis. Our findings included a significant GPx3 reduction in the CMD patients (n = 126), especially in the T2DM patients (n = 51), and a decreasing trend in the prediabetes group (n = 37). miR-196a expression, although higher in the CMD and T2DM groups than in the controls, was not statistically significant, potentially due to the small sample size. In the individuals with CMD, GPx3 levels exhibited a negative correlation with the mass of adipose tissue, muscle, and total body water, while miR-196a positively correlated with fat mass. In the CMD group, the analysis revealed a weak negative correlation between glucose and GPx3 levels. ROC analysis indicated a 5.2-fold increased CMD risk with GPx3 below 419.501 ng/mL. Logistic regression suggested that each 100 ng/mL GPx3 increase corresponded to a roughly 20% lower CMD risk (OR = 0.998; 95% CI: 0.996-0.999; p = 0.031). These results support the potential of GPx3 as a biomarker for CMD, particularly in T2DM, and the lack of a significant decline in GPx3 levels in prediabetic individuals suggests that it may not serve reliably as an early indicator of CMDs, warranting further large-scale validation.

Overexpressed miR-196a accelerates osteogenic differentiation in osteoporotic mice via GNAS-dependent Hedgehog signaling pathway.

Osteoporosis (OP), a common metabolic bone disease, is accompanied by reduced bone mass, bone mineral density (BMD), as well as microstructure destruction of bone. Previously, microRNA-196a-2 (miR-196a-2) and miR-196a-3p were reported for its involvement in BMD. Herein, this study set out to identify the functional relevance of miR-196a in osteogenic differentiation in osteoporotic mice and explore the associated mechanism by establishing an OP mouse model. Guanine nucleotide binding protein, alpha stimulating (GNAS) was verified as a target gene of miR-196a, which was decreased in OP mice. Furthermore, the bone marrow stromal cells (BMSCs) were then extracted from OP mice and treated with miR-196 mimic/inhibitor or small interfering RNA against GNAS to investigate miR-196a interaction with GNAS and the Hedgehog signaling pathway. BMSCs in OP mice transfected with miR-196a mimic or si-GNAS displayed the elevated expression of Smo, ALP, Runx2, and OPN, as well as bone gla protein and tartrate-resistant acid phosphatase, elevated ALP vitality and bone formation ability as well as reduced expression of GNAS and PTCH. Taken conjointly, overexpression of miR-196a repressed GNAS expression by activating the Hedgehog signaling pathway, thus promoting osteogenic differentiation in mice with OP.

Upregulation of microRNA-196a improves cognitive impairment and alleviates neuronal damage in hippocampus tissues of Alzheimer's disease through downregulating LRIG3 expression.

PURPOSE: This study is launched to uncover the inner function of microRNA-196a (miR-196a) on cognitive dysfunction and neuronal damage in Alzheimer's disease (AD) rats through regulating the PI3K/Akt signaling pathway. METHODS: The establishment of AD rat model was performed by a microinjection of Aß25-35 . miR-196a and LRIG3 expression was detected, and the putative binding site between them was also determined. The spatial learning and memory capability, the hippocampal neurons ultrastructure as well as the survival, and apoptosis of hippocampal neurons of rats were observed. The expression of apoptosis-associated protein, oxidative stress index, and inflammatory factors as well as the PI3K/Akt pathway-related factors was determined. RESULTS: Initially, decreased miR-196a and increased LRIG3 were exhibited in hippocampus tissues of AD rats. In addition, restored miR-196a and deleted LRIG3 ameliorated spatial learning and memory capability, suppressed the pathological injury, induced the survival, and suppressed the apoptosis of hippocampal neurons, as well as inhibited oxidative stress injury together with inflammatory injury in AD rats. Furthermore, upregulation of miR-196a activated the PI3/Akt pathway in AD rats. CONCLUSION: This current study suggests that upregulation of miR-196a and downregulation of LRIG3 improve cognitive impairment and alleviate neuronal damage in hippocampus tissues in AD rats via the modulation of the PI3K/Akt pathway.

MIR196A2 rs11614913 C > T polymorphism correlates with an increased risk of hepatopulmonary syndrome in liver cirrhosis: a case-control study in China.

OBJECTIVE: This case-control study is designed to explore the relationship between microRNA-196a2 (MIR196A2) rs11614913 C > T polymorphism and the risk of hepatopulmonary syndrome (HPS) in liver cirrhosis. METHODS: From January 2013 to January 2015, 163 liver cirrhosis patients with HPS (case group), 264 liver cirrhosis patients without HPS (control group), and 195 healthy people (normal group) were selected. A DNA extraction kit was used to extract plasma DNA from peripheral blood. Polymerase chain reaction-restriction fragment length polymorphism was used to detect the allele and genotype frequencies of MIR196A2 C > T polymorphism. Real-time quantitative polymerase chain reaction was adopted to detect the relative expression of MIR196A. RESULTS: The frequencies of C allele in the case group were higher than those in the control and normal groups (all P < 0.05), whereas no significant difference was found between the control and normal groups, which indicated that MIR196A2 C > T polymorphism was closely associated with an increased risk of HPS in patients with liver cirrhosis. Compared with the normal group, the relative expression of MIR196A in the case group was significantly increased (P < 0.05), but there was no significant difference in the control group (P > 0.05). In the case group, compared with patients carrying the TT genotype, the relative expression of MIR196A of patients carrying the C allele (CT + CC) evidently increased (P < 0.05). CONCLUSIONS: The MIR196A2 rs11614913 C > T polymorphism may contribute to an increased risk of HPS in liver cirrhosis patients.

MiR-196a regulates heme oxygenase-1 by silencing Bach1 in the neonatal mouse lung.

In the lung, heme oxygenase-1 (HO-1) is developmentally regulated, with its highest expression in the first days of life. In addition, neonatal mice have limited HO-1 induction in hyperoxia compared with adults. However, few reports have addressed the functional effect of microRNAs (miRNAs) in the regulation of HO-1 in vivo. The aims of the present study were to characterize changes in lung miRNA expression during postnatal development and in response to hyperoxic exposure, and to identify miRNAs that target lung HO-1 gene expression. Neonatal (<12 h old) and adult (2 mo old) mice were exposed to room air or hyperoxia (95% oxygen) for 72 h. TaqMan low-density array rodent miRNA assays were used to calculate miRNA expression changes between control and hyperoxia groups in neonatal and adult lungs. In neonates, we identified miR-196a, which binds to the 3'-untranslated region of the transcriptional repressor BTB and CNC homology 1 (Bach1) and regulates its expression, and subsequently leads to higher levels of lung HO-1 mRNA compared with levels in adults. Despite the increase at baseline, miR-196a was degraded in hyperoxia resulting in limited HO-1 induction in neonatal mice lungs. Furthermore, the developmental differences in lung HO-1 gene expression can be explained in part by the variation in miRNA-196a and its effect on Bach1. This report is the first to show developmental differences in lung miR-196a and its effect on Bach1 and HO-1 expression at baseline and in hyperoxia.

The role of microRNA-196a in tumorigenesis, tumor progression, and prognosis.

MicroRNAs are a large group of non-coding RNAs that have emerged as regulators of various biological processes, especially carcinogenesis and cancer progression. Recent evidence has shown that microRNA-196a (miR-196a) is upregulated in most types of tumors and involved in multiple biological processes via translational inhibition and mRNA cleavage, such as cell proliferation, migration, and invasion, mostly functioning as an oncogene. Dysregulation of miR-196a promotes oncogenesis and tumor progression. In this review, we summarize the upstream regulators, target genes, signaling pathways, and single nucleotide polymorphisms of miR-196a, which collectively affect cell proliferation, migration, and invasion. In addition, we review the clinical outcomes and significance of miR-196a. miR-196a may serve as a novel biomarker or target for diagnosis, prognosis, and therapy in several human cancers.

Inhibition of microRNA-196a might reverse cisplatin resistance of A549/DDP non-small-cell lung cancer cell line.

We aimed to explore the possible mechanism of microRNA-196a (miR-196a) inhibition and reversion of drug resistance to cisplatin (DDP) of the A549/DDP non-small-cell lung cancer (NSCLC) cell line. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect expression differences of miR-196a in the drug-resistant A549/DDP NLCLC cell line and the parental A549 cell line, and expressions of miR-196a in the A549/DDP NLCLC cell line transfected with miR-196a inhibitor (anti-miR-196a group) and the miR-196a negative control (miR-NC) group and blank group (without transfection). 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test was applied in examining the cell viability of A549/DDP cell line before and after transfection. Clonogenic assay was used to detect cell proliferation ability. Flow cytometry was applied in detecting apoptosis rate of assayed tumor cell and rhodamine-123 changes in cells. Western blot was applied in detecting proteins of drug-resistant related gene in A549/DDP cell line. Significantly higher expression of miR-196a was detected in the drug-resistant A549/DDP cell line than that in the parental A549 cell line (P < 0.05). However, miR-196a expression in the anti-miR-196a group decreased obviously compared to that in the blank group and the miR-NC group (both P < 0.05); The value of IC50 in the anti-miR-196a group showed remarkably lower than that in the blank group and the miR-NC group (both P < 0.05); Rh-123 absorbing ability in the anti-miR-196a group increased 2.51 times and 2.49 times respectively compared to that in the blank group and the miR-NC group (both P < 0.05). No statistical differences in the apoptosis rate of A549/DDP cell line in the early stage were found among the three groups (all P > 0.05), but the late-stage apoptosis rate in the anti-miR-196a group was significantly higher than that in the blank group and the miR-NC group (both P < 0.05); The expressions of human multidrug resistance 1 (MDR1), multidrug resistance protein 1 (MRP1), excision repair cross-complementation 1 (ERCC1), survivin, and B cell lymphoma 2 (Bcl-2) decreased significantly while RhoE increased significantly in the anti-miR-196a group than the blank group and the miR-NC group (all P < 0.05). Inhibition of miR-196a could reverse cisplatin resistance of A549/DDP cell lines, which might relate with inhibition of drug efflux, down-regulation of drug-resistant protein expression, cell apoptosis, and cell proliferation suppression.

MicroRNA-196a promotes an oncogenic effect in head and neck cancer cells by suppressing annexin A1 and enhancing radioresistance.

Radiotherapy is a major treatment modality for head and neck squamous cell carcinoma (HNSCC). Up to 50% of patients with locally advanced disease relapse after radical treatment and there is therefore a need to develop predictive bomarkers for clinical use that allow the selection of patients who are likely to respond. MicroRNA (miRNA) expression profiling of a panel of HNSCC tumours with and without recurrent disease after surgery and radiotherapy detected miR-196a as one of the highest upregulated miRNAs in the poor prognostic group. To further study the role of miR-196a, its expression was determined in eight head and neck cancer cell lines. Overexpression of miR-196a in HNSCC cells, with low endogenous miR-196a expression, significantly increased cell proliferation, migration and invasion, and induced epithelial to mesenchymal transition. Conversely, miR-196a knockdown in cells with high endogenous expression levels significantly reduced oncogenic behaviour. Importantly, overexpression of miR-196a increased radioresistance of cells as measured by gamma H2AX staining and MTT survival assay. Annexin A1 (ANXA1), a known target of miR-196a, was found to be directly modulated by miR-196a as measured by luciferase assay and confirmed by Western blot analysis. ANXA1 knockdown in HNSCC exhibited similar phenotypic effects to miR-196a overexpression, suggesting the oncogenic effect of miR-196a may at least be partly regulated through suppression of ANXA1. In conclusion, this study identifies miR-196a as a potential important biomarker of prognosis and response of HNSCC to radiotherapy. Furthermore, our data suggest that miR-196a and/or its target gene ANXA1 could represent important therapeutic targets in HNSCC.

Upregulation of microRNA-196a and microRNA-196b cooperatively correlate with aggressive progression and unfavorable prognosis in patients with colorectal cancer.

BACKGROUND: Both microRNA (miR)-196a and miR-196b are implicated in normal cell differentiation, proliferation, and in tumorigenesis of various cancer types. Especially, miR-196a exerts a pro-oncogenic influence in colorectal cancer (CRC) cells and miR-196b expression is upregulated in CRC tissues. The aim of this study was to evaluate the associations of miR-196a and miR-196b dysregulation with clinicopathological characteristics and prognosis in patients with CRC. METHODS: Quantitative real time-PCR (qRT-PCR) was performed to detect the expression levels of miR-196a and miR-196b in 126 pairs of fresh tumor samples matched with adjacent colorectal mucosa obtained from 126 patients with CRC. RESULTS: miR-196a and miR-196b expression levels in CRC tissues were significantly higher than those in adjacent colorectal mucosa (both P < 0.002). Interestingly, the expression levels of miR-196a in CRC tissues were positively correlated with those of miR-196b. Then, high miR-196a expression and high miR-196b expression, alone or in combination, were all statistically linked to the presence of lymph node metastasis, the poor differentiation grade, and the advanced TNM stage of CRC. Moreover, overall and disease-free survivals of CRC patients with high miR-196a expression, high miR-196b expression and miR-196a-high/miR-196b-high expression tended to be shorter than the corresponding control groups (log-rank statistic, all P < 0.001). Furthermore, multivariate analysis indicated miR-196a and/or miR-196b expression as independent prognostic indicators for CRC patients (all P < 0.05). CONCLUSIONS: Both miR-196a and miR-196b may be correlated with aggressive progression and unfavorable clinical outcome in CRC patients. Combined expression of miR-196a and miR-196b may be a promising biomarker in identifying a poor prognosis group of CRC.

Electrochemiluminescence biosensor for specific detection of pancreatic ductal carcinoma through dual targeting of MUC1 and miRNA-196a.

Pancreatic ductal adenocarcinoma (PDAC) poses significant diagnostic challenges due to its asymptomatic nature in its early stages, low specificity of conventional in vitro assays, and limited efficacy of surgical interventions. However, clinical specificity of the current serum biomarkers is suboptimal, leading to diagnostic inaccuracies and oversights. Therefore, this study introduced a novel dual-target electrochemiluminescence (ECL) biosensor to address these critical issues. The ECL biosensor synergistically employs the serum biomarker MUC1 and microRNA-196a to detect early-stage PDAC precisely. While MUC1 is a differential marker between normal and cancerous pancreatic cells, its standalone diagnostic performance is limited. However, integrating miRNA-196a as a complementary marker substantially enhances the specificity of the assay. This biosensor exhibits distinct ECL signal modulation-"on-off" in the presence of MUC1 and "off-on" upon concurrent detection of MUC1 and miRNA-196a. The biosensor achieves remarkably low limits of detection (LODs) at 0.63 fg mL-1 and 4.57 aM for MUC1 and miRNA-196a, respectively. Thus, it facilitates the real-time differentiation between human normal pancreatic (hTERT-HPNE) and pancreatic cancer (PANC-1) cells in authentic biological matrices. This innovative approach heralds a significant advancement in the early and specific detection of PDAC, offering promising prospects for clinical translation and the broader landscape of cancer diagnostics.

Upregulation of miRNA-196a and miRNA-196b correlates with Bryne's prognostic score in oral squamous cell carcinoma.

BACKGROUND: microRNA(miRNA)-196a and miRNA-196b expression has been found to be dysregulated and involved in tumorigenesis and tumor progression in array of cancers through different targets. The role of these miRNAs together in clinical application is not always consistent and, its prognostic value in oral squamous cell carcinoma (OSCC) is still elusive. This study was performed to investigate the correlation of these miRNAs expression with histological grades of OSCC according to Bryne's histological grading system, to predict prognosis and to evaluate their relationship with clinico-pathological data. METHODS: Real-time quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR) was done to evaluate the expression levels of miRNA-196a and miRNA-196b in 75 pairs of OSCC tissue matched with adjacent normal mucosa, used as a control. RESULTS: miRNA-196a and 196b expression in OSCC was significantly higher than that in corresponding adjacent normal tissues (p > 0.001). Also, a significant differential correlation was found in between the expression levels of these two miRNAs (Pearson correlation test r = 0.676, p-value<0.0001). The increased expression of these miRNAs was more frequently observed in OSCC tissues with advanced clinical and pathological TNM stages (IVa and IVb, pIVb respectively, p-value<0.0001). Significant correlation was found between miRNA-196a upregulation and moderate prognostic score (p-value<0.0001) in comparison with good and poor prognostic score of histological grades of OSCC. Sensitivity and specificity for miRNA-196a were 95 % and 85 %, respectively (AUC = 1, 95 % CI = 0.617-0.850; p 0.001), while for miRNA-196b were 94 % and 86 %, respectively (AUC = 0.808, 95 % CI = 0.701-0.916; p0.0001). CONCLUSIONS: These findings suggest that the increased expression of miRNA-196a and 196b may play an important role in tumor progression in OSCC. miRNA-196a might be a useful marker for predicting the clinical outcome of OSCC, especially for advanced stages. In conclusion, our data demonstrate for the first time that these miRNAs may serve as a potent prognostic marker for tumor progression. We further highlight miRNA-196a and miRNA-196b as a promising predictor of prognostic assessment in OSCC.

MicroRNA-196a-5p targeting LRP1B modulates phenotype of thyroid carcinoma cells.

INTRODUCTION: Thyroid cancer (TC) is a common endocrine malignancy, comprising nearly one-third of all head and neck malignancies worldwide. MicroRNAs (miRNAs) have been implicated in the malignant progression of multiple cancers; however, their contribution to thyroid diseases has not been fully explored. MATERIAL AND METHODS: This study aimed to illustrate the regulatory mechanism of microRNA-196a-5p in TC progression and to investigate whether microRNA-196a-5p affects progression of TC cells by targeting low-density lipoprotein receptor-associated protein 1B (LRP1B). MicroRNA-196a-5p and LRP1B expression status in TC cells and normal human thyroid cells was detected by quantative reverse transcription polymerase chain reaction (qRT-PCR) and western blot. Dual-luciferase reporter assay, cell counting kit-8 (CCK-8) assay, scratch healing assay, and Transwell assay were also performed. RESULTS: The results showed that microRNA-196a-5p expression was up-regulated and LRP1B expression was down regulated in TC cells. In addition, the upregulation of microRNA-196a-5p facilitated progression of TC cells. Silencing microRNA-196a-5p led to the opposite results. Dual-luciferase reporter assay offered evidence for microRNA-196a-5p targeting LRP1B in TC. MicroRNA-196a-5p could target LRP1B to facilitate proliferation, invasion, and migration of TC cells. CONCLUSION: Overall, this study revealed that microRNA-196a-5p may be a cancer-promoting microRNA that plays an important role in TC progression.

Serum exosomal microRNA-370-3p and microRNA-196a-5p are potential biomarkers for the diagnosis and prognosis of hepatocellular carcinoma.

INTRODUCTION: Evidence has shown that some microRNAs (miRNAs) play a role in tumorigenesis of hepatocellular carcinoma (HCC). Herein, we aimed to evaluate the diagnostic and prognostic values of serum exosomal miR-370-3p and miR-196a-5p in patients with HCC. MATERIAL AND METHODS: Serum exosomes in 90 HCC patients were extracted and identified. Serum exosomal miR-370-3p and miR-196a-5p expression in HCC patients were detected. The diagnostic value of miR-370-3p and miR-196a- 5p, relationship between miR-370-3p and miR-196a-5p expression and clinicopathological features and prognosis of patients with HCC were analyzed. Relationship between miR-370-3p and miR-196a-5p expression and liver function indices such as alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin (TBIL) in HCC patients were analyzed. The effects of miR-370-3p and miR-196a-5p on Huh-7 HCC cells' proliferation, invasion and migration were determined. RESULTS: Lower expression of miR-370-3p and higher expression of miR-196a-5p were found in serum exosomes of HCC patients. Serum exosomal miR-370-3p and miR-196a-5p were associated with tumor size, tumor grade and TNM stage as well as prognosis and liver function indices of HCC patients. Overexpressed miR-370-3p or silenced miR-196a-5p suppressed proliferation, invasion and migration of Huh-7 HCC cells. CONCLUSIONS: We suggest that miR-370-3p/miR-196a-5p in serum exosomes of HCC patients could be potential biomarkers for the diagnosis and prognosis of HCC.

Differential impact of the angiotensin-converting enzyme-2 (ACE2 rs4343 G>A) and miR-196a2 rs11614913 C>T gene alterations in COVID-19 disease severity and mortality.

The recent coronavirus outbreak from Wuhan China in late 2019 caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) resulted in a global pandemic of coronavirus-19 disease (COVID-19). Understating the underlying mechanism of the pathogenesis of coronavirus infection is important not only because it will help in accurate diagnosis and treatment of the infection but also in the production of effective vaccines. The infection begins when SARS-CoV-2 enters the cells through binding of its envelope glycoprotein to angiotensin-converting enzyme2 (ACE2). Gene variations of ACE2 and microRNA (miR)-196 are associated with viral infection and other diseases. The present study investigated the association of the ACE2 rs4343 G>A and miR-196a2 rs11614913 C>T gene polymorphisms with severity and mortality of COVID-19 using amplification refractory mutation system PCR in 117 COVID-19 patients and 103 healthy controls from three regions of Saudi Arabia. The results showed that ACE2 rs4343 GA genotype was associated with severity of COVID-19 (OR=2.10, P-value 0.0028) and ACE2 rs4343 GA was associated with increased mortality with OR=3.44, P-value 0.0028. A strong correlation between the ACE2 rs4343 G>A genotype distribution among COVID-19 patients was reported with respect to their comorbid conditions including sex (P<0.023), coronary artery disease (P<0.0001), oxygen saturation <60 mm Hg (P<0.0009) and antiviral therapy (0.003). The results also showed that the CT genotype and T allele of the miR-196a2 rs11614913 C>T were associated with decreased risk to COVID-19 with OR=0.76, P=0.006 and OR=0.54, P=0.005, respectively. These results need to be validated with future molecular genetic studies in a larger sample size and different populations.

Graphene-Assisted Electrochemical Sensor for Detection of Pancreatic Cancer Markers.

Pancreatic cancer is a highly lethal gastrointestinal malignancy. Most patients are already in the middle to advanced stages of pancreatic cancer at the time of diagnosis and cannot be treated completely. As a single-atom planar two-dimensional crystal, graphene's unusual electronic structure, specific electronic properties and excellent electron transport capacity make it uniquely advantageous in the field of electrochemical sensing. In this mini-review, we summarize the potential application of graphene in pancreatic cancer detection. K-Ras gene, CEA and MicroRNA are important in the early diagnosis of pancreatic cancer.

MicroRNA-196a promotes renal cancer cell migration and invasion by targeting BRAM1 to regulate SMAD and MAPK signaling pathways.

Rationale: MicroRNAs (miRNAs) are endogenous ~22nt RNAs that play critical regulatory roles in various biological and pathological processes, including various cancers. Their function in renal cancer has not been fully elucidated. It has been reported that miR-196a can act as oncogenes or as tumor suppressors depending on their target genes. However, the molecular target for miR-196a and the underlying mechanism in miR-196a promoted cell migration and invasion in renal cancer is still not clear. Methods: The expression, survival and correlation between miR-196a and BRAM1 were investigated using TCGA analysis and validated by RT-PCR and western blot. To visualize the effect of Bram1 on tumor metastasis in vivo, NOD-SCID gamma (NSG) mice were intravenously injected with RCC4 cells (106 cells/mouse) or RCC4 overexpressing Bram1. In addition, cell proliferation assays, migration and invasion assays were performed to examine the role of miR-196a in renal cells in vitro. Furthermore, immunoprecipitation was done to explore the binding targets of Bram1. Results: TCGA gene expression data from renal clear cell carcinoma patients showed a lower level of Bram1 expression in patients' specimens compared to adjacent normal tissues. Moreover, Kaplan­Meier survival data clearly show that high expression of Bram1correlates to poor prognosis in renal carcinoma patients. Our mouse metastasis model confirmed that Bram1 overexpression resulted in an inhibition in tumor metastasis. Target-prediction analysis and dual-luciferase reporter assay demonstrated that Bram1 is a direct target of miR-196a in renal cells. Further, our in vitro functional assays revealed that miR-196a promotes renal cell proliferation, migration, and invasion. Rescue of Bram1 expression reversed miR-196a-induced cell migration. MiR-196a promotes renal cancer cell migration by directly targeting Bram1 and inhibits Smad1/5/8 phosphorylation and MAPK pathways through BMPR1A and EGFR. Conclusions: Our findings thus provide a new mechanism on the oncogenic role of miR-196a and the tumor-suppressive role of Bram1 in renal cancer cells. Dysregulated miR-196a and Bram1 represent potential prognostic biomarkers and may have therapeutic applications in renal cancer.

Knockdown of lncRNA TUG1 protects lens epithelial cells from oxidative stress-induced injury by regulating miR-196a-5p expression in age-related cataracts.

Oxidative stress plays an important role in the pathogenesis of cataracts. Under oxidative stress, apoptosis of lens epithelial cells (LECs) is activated, which may cause lens opacity and accelerate the development of cataracts. Long non-coding RNA (lncRNA) and microRNA (miRNA/miR) are involved in cataracts. Previous studies have demonstrated that lncRNA taurine upregulated 1 (TUG1) promotes cell apoptosis induced by ultraviolet radiation by downregulating the expression of miR-421. However, the mechanism underlying TUG1 in age-related cataract remains to be elucidated. The present study aimed to investigate the effect of TUG1 in age-related cataracts and to determine the related underlying molecular mechanism. In the present study, the association between TUG1 and microRNA (miR)-196a-5p was predicted using StarBase and verified using a dual luciferase reporter assay in 293 cells. The LEC line SRA01/04 was exposed to 200 µM hydrogen peroxide (H2O2) for 24 h to establish an in vitro oxidative stress model. The mRNA expression levels of TUG1 and miR-196a-5p were analyzed using reverse transcription-quantitative PCR, whilst cell viability and apoptosis were determined using MTT and flow cytometry assays, respectively. The protein expression levels of cleaved caspase-3 and caspase-3 in SRA01/04 cells were determined using western blotting. The results of the present study revealed that TUG1 directly targeted miR-196a-5p expression. In addition, the expression levels of miR-196a-5p were downregulated in SRA01/04 cells following oxidative stress, whilst TUG1 expression was upregulated. Cell transfection with TUG1-small interfering RNA (siRNA) upregulated miR-196a-5p expression levels in SRA01/04 cells, which was reversed following co-transfection with the miR-196a-5p inhibitor. Transfection with TUG1-siRNA also reduced the levels of H2O2-induced oxidative damage in SRA01/04 cells, which was demonstrated by increased cell viability, reduced levels of apoptosis and downregulated cleaved caspase-3 levels. Conversely, transfection with the miR-196a-5p inhibitor reversed these effects aforementioned. Overexpression of miR-196a-5p reduced H2O2-induced oxidative damage in SRA01/04 cells. In conclusion, findings from the present study suggested that knocking down TUG1 expression may protect LECs from oxidative stress-induced apoptosis by upregulating the expression of miR-196a-5p.

Investigating the role and mechanism of microRNA-196a in oral squamous cell carcinoma by targeting FOXO1.

Oral squamous cell carcinoma (OSCC) is one of the most prevalent malignancies worldwide. MicroRNAs (miRNAs or miRs) serve crucial roles in the development of OSCC. miR-196a is upregulated in various tumors; however, the role of miR-196a in OSCC remains unclear. This present study aimed to determine the role and underlying mechanism of miR-196a in OSCC cells. Reverse transcription-quantitative PCR (RT-qPCR) was used to measure miR-196a levels in OSCC cells. MTT assays were also performed to determine cell proliferation. Cell migration was detected using wound healing assays and transwell assays, and cell apoptosis was analyzed via flow cytometry. The results indicated that the expression of miR-196a was increased in OSCC cells compared with normal oral squamous cells. TargetScan and luciferase reporter assays also confirmed that forkhead box O1 (FOXO1) was a target gene of miR-196a. It was demonstrated that FOXO1 small interfering RNA significantly promoted SCC9 cell proliferation and migration, and inhibited cell apoptosis. Furthermore, inhibition of miR-196a suppressed SCC9 cell proliferation and migration, and induced cell apoptosis. However, all effects of the miR-196a inhibitor were reversed following FOXO1 inhibition. Western blotting and RT-qPCR were subsequently performed to determine the effect of miR-196a on the PI3K/Akt signaling pathway. In the present study, transfection of miR-196a inhibitor suppressed the expression of phosphorylated (p)-PI3K and p-Akt, and enhanced the levels of FOXO1, while inhibition of FOXO1 exerted the opposite effects. Furthermore, it was demonstrated that miR-196a mimic significantly enhanced SCC9 cell proliferation and migration, and inhibited cell apoptosis. In conclusion, the results indicated that miR-196a serve as an oncogene in OSCCs. Downregulation of miR-196a inhibited the malignant biological processes of OSCC cells by targeting FOXO1. The current results may provide a novel therapeutic strategy for the treatment of patients with OSCC.

microRNA-196a Overexpression Inhibits Apoptosis in Hemin-Induced K562 Cells.

microRNAs (miRNAs) have a crucial role in erythropoiesis. However, the understanding of the apoptosis of erythroid lineage remains poorly understood. Hence, an additional examination is required. K562 cell lines can be differentiated into early erythrocytes by hemin and the model of early erythrocytes can be established, consequently. miR-196a has been proven to take part in antiapoptosis in many cell lines. However, the role of miR-196a associated with the apoptosis in hemin-induced K562 cells remains unclear. To study the potential function of miR-196a involved in the common progenitor of erythroblasts, miR-196a mimics and microRNA-small hairpin negative control (miRNA-ShNC) were transfected into hemin-induced K562 cells with lentiviruses. After that, the viability of the transfected hemin-induced K562 cells was tested by CCK-8 assay, and the alteration of cell cycle and apoptosis rate were detected by flow cytometry. Furthermore, bioinformatics and dual-luciferase report system verified that p27kip1 is a target gene of miR-196a. Additionally, the expression of some proteins associated with cell cycle and apoptosis was tested by Western blotting assays. It was found that after overexpressing miR-196a, the proliferation of hemin-induced K562 cells was promoted while the apoptosis inhibited. Furthermore, miR-196a combines with the 3'UTR of p27kip1 directly. Additionally, the relationship between miR-196a and the protein level of p27kip1 is negative. After restoring the expression of p27kip1, the growth rate of hemin-induced K562 cells was not as high as before and the inhibition of apoptosis was alleviated. The present study validates that miR-196a overexpression inhibits apoptosis in hemin-induced K562 cells through downregulating p27kip1.

microRNA-196a attenuates ischemic brain injury in rats by directly targeting high mobility group A1.

Dysfunction of the microRNA (miR) network has been indicated as a major regulator in neurological diseases. However, there is limited understanding regarding the functional significance of miRs in ischemic brain injury. In the present study, miR-196a expression was significantly increased in rat brains and neurons following transient middle cerebral artery occlusion (MCAO) or oxygen-glucose deprivation, respectively. In addition, repression of miR-196a significantly decreased neuron cell apoptosis and the infarct size in rats subjected to MCAO (P<0.05). Furthermore, miR-196a was indicated to directly target and inhibit high mobility group A1 expression, which indicated a potential role for miR-196a in ischemic brain injury. These findings suggested that miR-196a may be involved in regulating neuronal cell death, thus offering a novel target for the development of therapeutic agents against ischemic brain injury.