miR-483-3p suppresses the proliferation and progression of human triple negative breast cancer cells by targeting the HDAC8>oncogene

Triple negative breast cancer (TNBC) is a heterogeneous subclass of breast cancer (BC) distinguished by lack of hormone receptor expression. It is highly aggressive and difficult to treat with traditional chemotherapeutic regimens. Targeted-therapy using microRNAs (miR) has recently been proposed to improve the treatment of TNBC in the early stages. Here, we explore the roles of miR-483-3p/HDAC8 HDAC8 premiR-vector on tumorigenicity in TNBC patients. Clinical TNBC specimens and three BC cell lines were prepared. miR-483-3p and expression levels were measured using quantitative real-time polymerase chain reaction. Cell cycle progression was assessed by a flow-cytometry method. We also investigated cell proliferation by 3-2, 5-diphenyl tetrazolium bromide assay and colony formation assay. We used a to overexpress miR-483-3p, and a HDAC8-KO-vector for knocking out the endogenous production of HDAC8. Our data showed significant downregulation of miR-483-3p expression in TNBC clinical and cell line samples. The HDAC8 was also upregulated in both tissue specimens and BC cell lines. We found that increased levels of endogenous miR-483-3p affects tumorigenecity of MDA-MB-231. Downregulation of HDAC8 using the KO-vector showed the same pattern. Our results revealed that the miR-483-3p suppresses cellular proliferation and progression in TNBC cell lines via targeting HDAC8. Overall, our outcomes demonstrated the role of miR-483-3p as a tumor suppressor in TNBC and showed the possible mechanism via HDAC8. In addition, targeted treatment of TNBC with miR-483-3p might be considered in the future.     

Tumor suppressor function of miR-483-3p on squamous cell carcinomas due to its pro-apoptotic properties

The frequent alteration of miRNA expression in many cancers, together with our recent reports showing a robust accumulation of miR-483-3p at the final stage of skin wound healing, and targeting of CDC25A leading to an arrest of keratinocyte proliferation, led us to hypothesize that miR-483-3p could also be endowed with antitumoral properties. We tested that hypothesis by documenting the in vitro and in vivo impacts of miR-483-3p in squamous cell carcinoma (SCC) cells. miR-483-3p sensitized SCC cells to serum deprivation- and drug-induced apoptosis, thus exerting potent tumor suppressor activities. Its pro-apoptotic activity was mediated by a direct targeting of several anti-apoptotic genes, such as API5, BIRC5, and RAN. Interestingly, an in vivo delivery of miR-483-3p into subcutaneous SCC xenografts significantly hampered tumor growth. This effect was explained by an inhibition of cell proliferation and an increase of apoptosis. This argues for its further use as an adjuvant in the many instances of cancers characterized by a downregulation of miR-483-3p.     

Consumption of Distinct Dietary Lipids during Early Pregnancy Differentially Modulates the Expression of microRNAs in Mothers and Offspring

Diet during pregnancy and lactation influences the offspring’s health in the long-term. Indeed, human epidemiological studies and animal experiments suggest that different type of fatty acids consumption during pregnancy affect offspring development and susceptibility to metabolic disorders. Epigenetic changes are thought to be elicited by dietary factors during critical timing of development. microRNAs (miRNAs) are versatile regulators of gene expression. Thus, we aimed to determine the influence of different fatty acids on miRNA expression in offspring when given during early pregnancy. We fed pregnant either soybean (SO), olive (OO), fish (FO), linseed (LO), or palm-oil (PO) diets from conception to day 12 of gestation; and standard diet thereafter. miRNA expression was assessed in liver an adipose tissue of pregnant rats and their virgin counterparts. While liver concentrations of fatty acids in pregnant or virgin rats replicated those of the diets consumed during early pregnancy, their pups’ liver tissue marginally reflected those of the respective experimental feeds. By contrast, the liver fatty acid profile of adult offsprings was similar, regardless of the diet fed during gestation. Different parental miRNAs were modulated by the different type of fatty acid: in adult offspring, miR-215, miR-10b, miR-26, miR-377-3p, miR-21, and miR-192 among others, were differentially modulated by the different fatty acids fed during early pregnancy. Overall, our results show that maternal consumption of different types of fatty acids during early pregnancy influences miRNA expression in both maternal and offspring tissues, which may epigenetically explain the long-term phenotypic changes of the offspring.     

MicroRNA 483-3p suppresses the expression of DPC4/Smad4 in pancreatic cancer

Both deregulation of tumor-suppressor genes and misexpression of microRNAs (miRNAs) have been implicated in the development of pancreatic cancer, but their relationship during this process remains less clear. Here, we report that the expression of miR-483-3p is strongly enhanced in pancreatic cancer tissues compared to side normal tissues using a miRNA-array differential analysis. Furthermore, DPC4/Smad4 is identified as a target of miR-483-3p and their expression levels are inversely correlated in human clinical specimens. Ectopic expression of miR-483-3p significantly represses DPC4/Smad4 protein levels in pancreatic cancer cell lines, and simultaneously promotes cell proliferation and colony formation in vitro. Our findings identify miR-483-3p as a potent regulator of DPC4/Smad4, which may provide a novel therapeutic strategy for the treatment of DPC4/Smad4-driven pancreatic cancer.     

miR-483-3p靶向CD44抑制EGFR突变肺癌的作用研究

目的:探讨mi R-483-3p对CD44表达的调控作用及采用脂质体载药系统递送mi R-483-3p治疗表皮生长因子(epidermalgrowth factor receptor,EGFR)突变的非小细胞肺癌(non-small cell lung cancer,NSCLC)的作用。方法:通过mi R-483-3p靶基因的数据库发现CD44可能是mi R-483-3p的靶基因之一,从结构上及功能上进行验证。在EGFR酪氨酸激酶抑制剂(epidermal growth factor receptor-tyrosine kinase inhibitor, EGFR-TKI)耐药的NSCLC模型中,分别从基因及蛋白水平检测耐药模型及敏感模型中CD44的表达。在HCC827GR移植瘤模型中,采用脂质体-鱼精蛋白-DNA (liposome-polycation-DNA, LPD)载药系统递送mi R-483-3p进行治疗,观察肿瘤的生长情况。结果:双荧光素酶报告基因实验及Western blot实验结果显示CD44是mi R-483-3p的靶基因之一,且CD44在EGFR-TKI耐药模型中异常高表达(P0.05)。脂质体载药系统LPD-mi RNA-DSPE-PEG符合静脉给药要求,Size为66.93±21 nm,Zeta potential为8.7±2 m V,PDI (Polydispersity Index)为0.1,递送mi R-483-3p后能够抑制HCC827GR移植瘤的生长(P0.05)。结论:CD44为mi R-483-3p的靶基因之一,在耐药模型中高表达,脂质体载药系统给予mi R-483-3p能够抑制EGFR-TKI耐药肿瘤的生长。     

Chondrocyte mTORC1 activation stimulates miR-483-5p via HDAC4 in osteoarthritis progression

The hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1) in chondrocytes has been shown to accelerate the severity of destabilization of the medial meniscus-induced and age-related osteoarthritis (OA) phenotypes with aberrant chondrocyte hypertrophy and angiogenesis. Meanwhile, we previously reported that miR-483-5p is essential for the initiation and development of OA by stimulating chondrocyte hypertrophy and angiogenesis. The connection between mTORC1 and miR-483-5p activation in OA progression, however, remains unclear. In this study, we elucidated their relationship and identified the underlying mechanisms. The expression of miR-483-5p in the articular cartilage of cartilage-specific TSC1 knockout mice was assessed compared with control mice using the Agilent Mouse miRNA (8*60K) V19.0 array and real-time polymerase chain reaction (RT-PCR). The functional effects of the stimulation of miR-483-5p via histone deacetylase 4 (HDAC4) by mTORC1 in OA development, subsequently modulating its downstream targets matrilin 3 and tissue inhibitor of metalloproteinase 2, were examined by immunostaining, western blotting, and real-time PCR. This study revealed that miR-483-5p was responsible for mTORC1 activation-stimulated OA. Mechanistically, mTORC1 controls the HDAC4-dependent expression of miR-483-5p to stimulate chondrocyte hypertrophy, extracellular matrix degradation, and subchondral bone angiogenesis, and it consequently initiates and accelerates the development of OA. Our findings revealed a novel mTORC1-HDAC4-miR-483-5p pathway that is critical for OA development.     

Circulating microRNAs associated with liver fibrosis in chronic hepatitis C patients

A major challenge in hepatitis C research is the detection of early potential for progressive liver disease. MicroRNAs (miRNAs) are small RNAs that regulate gene expression and can be biomarkers of pathological processes. In this study, we compared circulating miRNAs identified in hepatitis C virus (HCV)-infected patients presenting two extremes of liver disease: mild/moderate fibrosis and cirrhosis. The patients in the cirrhosis group subsequently developed hepatocellular carcinoma (HCC). We identified 163 mature miRNAs in the mild/moderate fibrosis group and 171 in the cirrhosis group, with 144 in common to both groups. Differential expression analysis revealed 5 upregulated miRNAs and 2 downregulated miRNAs in the cirrhosis group relative to the mild/moderate fibrosis group. Functional analyses of regulatory networks (target gene and miRNA) identified gene categories involved in cell cycle biological processes and metabolic pathways related to cell cycle, cancer, and apoptosis. These results suggest that the differentially expressed circulating miRNAs observed in this work (miR-215-5p, miR-483-5p, miR-193b-3p, miR-34a-5p, miR-885-5p, miR-26b-5p and miR -197-3p) may be candidates for biomarkers in the prognosis of liver disease.     

Overexpression of miR‐483‐5p/3p cooperate to inhibit mouse liver fibrosis by suppressing the TGF‐β stimulated HSCs in transgenic mice

The transition from liver fibrosis to hepatocellular carcinoma (HCC) has been suggested to be a continuous and developmental pathological process. MicroRNAs (miRNAs) are recently discovered molecules that regulate the expression of genes involved in liver disease. Many reports demonstrate that miR‐483‐5p and miR‐483‐3p, which originate from miR‐483, are up‐regulated in HCC, and their oncogenic targets have been identified. However, recent studies have suggested that miR‐483‐5p/3p is partially down‐regulated in HCC samples and is down‐regulated in rat liver fibrosis. Therefore, the aberrant expression and function of miR‐483 in liver fibrosis remains elusive. In this study, we demonstrate that overexpression of miR‐483 in vivo inhibits mouse liver fibrosis induced by CCl₄. We demonstrate that miR‐483‐5p/3p acts together to target two pro‐fibrosis factors, platelet‐derived growth factor‐β and tissue inhibitor of metalloproteinase 2, which suppress the activation of hepatic stellate cells (HSC) LX‐2. Our work identifies the pathway that regulates liver fibrosis by inhibiting the activation of HSCs.     

MiR-125a-5p抑制3T3-L1前体脂肪细胞分化

MicroRNAs(miRNAs) 是一类在脂肪组织发育中发挥重要作用的小非编码RNA. 为探明miR-125a-5p在3T3-L1前体脂肪细胞中的作用,采用实时qPCR检测了miR-125a-5p在小鼠各组织及3T3-L1前体脂肪细胞分化过程中的表达|使用经化学修饰的miR-125a-5p模拟物agomir及抑制剂antagomir转染3T3-L1前体脂肪细胞,采用实时qPCR 和 Western印迹检测成脂标志基因Pparγ和aP2的表达,油红O染色观察脂肪细胞脂质积累. 结果显示,miR-125-5p在小鼠脂肪组织中高丰度表达,在3T3-L1前体脂肪细胞分化过程中表达下降.过表达miR-125a-5p,与对照组相比,成脂标志基因Pparγ和aP2在mRNA和蛋白质水平均明显下降|油红O染色及定量结果显示脂质积累减少. 抑制剂处理结果显示,Pparγ和aP2在mRNA和蛋白质水平均有不同程度上升,但油红O染色及定量结果差异不显著. 以上结果表明,miR-125a-5p在脂肪细胞分化中发挥负调控作用.     

The early life environment and the epigenome

Several lines of evidence point to the early origin of adult onset disease. A key question is: what are the mechanisms that mediate the effects of the early environment on our health? Another important question is: what is the impact of the environment during adulthood and how reversible are the effects of early life later in life? The genome is programmed by the epigenome, which is comprised of chromatin, a covalent modification of DNA by methylation and noncoding RNAs. The epigenome is sculpted during gestation, resulting in the diversity of gene expression programs in the distinct cell types of the organism. Recent data suggest that epigenetic programming of gene expression profiles is sensitive to the early-life environment and that both the chemical and social environment early in life could affect the manner by which the genome is programmed by the epigenome. We propose that epigenetic alterations early in life can have a life-long lasting impact on gene expression and thus on the phenotype, including susceptibility to disease. We will discuss data from animal models as well as recent data from human studies supporting the hypothesis that early life social-adversity leaves its marks on our epigenome and affects stress responsivity, health, and mental health later in life.     

miR-324-5p promotes adipocyte differentiation and lipid droplet accumulation by targeting Krueppel-like factor 3 (KLF3)

miRNAs, a kind of noncoding small RNA, play a significant role in adipose differentiation. In this study, we explored the effect of miR-324-5p in adipose differentiation, and found that miR-324-5p could promote adipocytes differentiation and increase body weight in mice. We overexpressed miR-324-5p during adipocytes differentiation, by oil red O and bodipy staining found that lipid accumulation was increased, and the expression level of adipogenic related genes were significantly increased. And the opposite experimental results were obtained after inhibiting miR-324-5p. In vivo, we injected miR-324-5p agomiR in obese mice and found that body weight, adipocyte area, and adipogenic-related gene expression level were significantly increased but lipolytic genes were decreased. To further explore the mechanism of miR-324-5p regulation in lipid accumulation, we constructed Krueppel-like factor 3 (KLF3) 3′-untranslated region luciferase reporter vector and KLF3 pcDNA 3.1 overexpression vector, and found that miR-324-5p was able to directly target KLF3. Overall, in this study we found that miR-324-5p could promote mice preadipoytes differentiation and increase mice fat accumulation by targeting KLF3.     

Adipose Rheb deficiency promotes miR-182-5p expression via the cAMP/PPARγ signaling pathway

Dysregulation of microRNAs (miRNAs) in adipocytes plays a critical role in the pathogenesis of obesity. However, the signaling mechanisms regulating miRNAs production in adipose tissue remain largely unclear. Here, we show that adipose tissue-specific knockout of Ras homolog enriched in brain (Rheb), a direct upstream activator of mTOR, increases miR-182-5p level in mouse subcutaneous white adipose tissues. Interestingly, the inhibition of mTOR signaling by rapamycin has no effect on miR-182-5p level in primary subcutaneous white adipocytes, suggesting the presence of a mTOR-independent mechanism regulating Rheb-mediated miR-182-5p expression. Consistent with this view, Rheb-ablation activates the cAMP/PPARγ signaling pathway. In addition, treatment of white adipocytes with pioglitazone, a PPARγ agonist, dramatically upregulates miR-182-5p levels. Our study reveals a unique mechanism by which Rheb regulates miR-182-5p in adipocytes. Given that increasing miR-182-5p in adipose tissue promotes beige fat development, our study also suggests a unique mechanism by which Rheb promotes thermogenesis and energy expenditure.     

MiR-483-5p suppresses the proliferation of glioma cells via directly targeting ERK1

MicroRNAs (miRNAs) exhibit tumor-specific expression signatures and play crucial roles in tumorigenesis by targeting oncogenes. Here, through analyzing the miRNA-array profiles of human glioblastoma tissues and the adjacent normal brain tissues, we found miR-483-5p was significantly down-regulated in gliomas, which was confirmed in both human glioma specimens and cell lines. The overexpression of miR-483-5p suppressed glioma cell proliferation and induced a G0/G1 arrest. In contrast, miR-483-5p inhibition promoted cell proliferation. Furthermore, by a dual-luciferase reporter assay and expression analysis, we identified extracellular signal-regulated kinase 1 (ERK1) as a direct target of miR-483-5p. ERK1 knockdown can block cell proliferation induced by miR-483-5p inhibition. Thus, our findings provide the first evidence that miR-483-5p can serve as a tumor suppressor in gliomas.     

Profiling of Serum and Urinary MicroRNAs in Children with Atopic Dermatitis

Background

Atopic dermatitis (AD) is the most prevalent chronic inflammatory skin disease in children characterized by dermatitis and pruritus. MicroRNAs (miRNAs) have been shown as great potential biomarkers for disease fingerprints to predict prognostics. We aimed to identify miRNA signature from serum and urine for the prognosis of AD patient by genome-wide miRNA profiling analysis.

Methods

Serum and urine from 30 children with AD and 28 healthy children were collected and their genome-wide miRNA expression profiles were measured by TaqMan-based array and confirmed by quantitative real-time PCR. Inflammatory factors in serum were detected by Antibody Array System.

Results

miR-203 and miR-483-5p were significantly up-regulated in serum of children with AD compared with healthy children. The level of miR-483-5p in serum was significantly associated with other atopic conditions, such as rhinitis and/or asthma. However, miR-203 was markedly decreased in urine of children with AD compared with healthy children. Down-regulated miR-203 in urine was significant associated with abnormal level of serum IgE in AD patients. 7 inflammatory factors in serum were altered in children with AD compared with healthy children. Up-regulated miR-203 in serum was significantly associated with increased sTNFRI and sTNFRII.

Conclusions

Up-regulated miR-483-5p in serum may be indicative of other atopic conditions in children with AD. Down-regulated miR-203 in urine may serve as a biomarker for the severity of inflammation in children with AD.     

miR-148a-3p promotes rabbit preadipocyte differentiation by targeting PTEN

Although emerging data support crucial roles for microRNAs (miRNAs) during adipogenesis, the detailed mechanisms remain largely unknown. In this study, it was shown that in rabbits, levels of miR-148a-3p not only increased in white adipose tissue during early stages of growth but also during in vitro cultured preadipocyte differentiation. Furthermore, overexpression of miR-148a-3p significantly upregulated the mRNA levels of PPARγ, C/EBPα, and FABP4, as well as the protein levels of PPARγ, as indicated by qPCR and western blotting analyses. Overexpression of miR-148a-3p also promoted intracellular triglyceride accumulation. In contrast, downregulation of miR-148a-3p inhibited the differentiation of rabbit preadipocytes. Next, based on target gene prediction and a luciferase reporter assay, we further demonstrated that miR-148a-3p directly targeted one of the 3′ untranslated regions of PTEN. Finally, it was observed inhibition of PTEN by siRNA promoted rabbit preadipocyte differentiation. Taken together, our results suggested that miR-148a-3p could be involved in regulating rabbit preadipocyte differentiation through inhibiting expression of PTEN, which further highlighted the importance of miRNAs during adipogenesis.