MicroRNA 340 Is Involved in UVB-Induced Dendrite Formation through the Regulation of RhoA Expression in Melanocytes

The influence of UV irradiation on pigmentation is well established, but the molecular and cellular mechanisms controlling dendrite formation remain incompletely understood. MicroRNAs (miRNAs) are a class of small RNAs that participate in various cellular processes by suppressing the expression of target mRNAs. In this study, we investigated the expression of miRNAs in response to UVB irradiation using a microarray screen and then identified potential mRNA targets for differentially expressed miRNAs among the genes governing dendrite formation. We subsequently determined the ability of miRNA 340 (miR-340) to suppress the expression of RhoA, which is a predicted miR-340 target gene that regulates dendrite formation. The overexpression of miR-340 promoted dendrite formation and melanosome transport, and the downregulation of miR-340 inhibited UVB-induced dendrite formation and melanosome transport. Moreover, a luciferase reporter assay demonstrated direct targeting of RhoA by miR-340 in the immortalized human melanocyte cell line Pig1. In conclusion, this study has established an miRNA associated with UVB irradiation. The significant downregulation of RhoA protein and mRNA expression after UVB irradiation and the modulation of miR-340 expression suggest a key role for miR-340 in regulating UVB-induced dendrite formation and melanosome transport.     

Changes in microRNA expression profile in hippocampus during the acquisition and extinction of cocaine-induced conditioned place preference in rats

Background

MicroRNA (miRNA) emerges as important player in drug abuse. Yet, their expression profile in neurological disorder of cocaine abuse has not been well characterized. Here, we explored the changes of miRNA expression in rat hippocampus following repeated cocaine exposure and subsequent abstinence from cocaine treatment.

Results

Conditioned place preference (CPP) procedure was used to assess the acquisition and extinction of cocaine-seeking behavior in rats. MiRNA microarray was performed to examine miRNAs levels in rat hippocampus. Quantitative RT-PCR was conducted to further confirm results in microarray study. Finally, bioinformatic predictions were made to suggest potential target genes of cocaine-responsive miRNA in this study. MiRNA array found that 34 miRNA levels were changed in rat hippocampus while acquiring cocaine CPP and 42 miRNAs levels were altered after the cocaine-induced CPP were extinguished, as compared to normal controls. The findings from qRT-PCR study support results from microarray analysis.

Conclusions

The current study demonstrated dynamic changes in miRNA expression in rat hippocampus during the acquisition and extinction of cocaine-induced CPP. Some miRNAs which have been previously reported to be involved in brain disorders and drug abuse, including miR-133b, miR-134, miR-181c, miR-191, miR-22, miR-26b, miR-382, miR-409-3p and miR-504, were found to be changed in their expression following repeated cocaine exposure and subsequent abstinence from cocaine treatment. These findings may extend our understanding of the regulatory network underlying cocaine abuse and may provide new targets for the future treatment of drug abuse.     

大鼠颅脑损伤后差异表达基因及miRNA研究

研究背景创伤性脑损伤(Traumatic brain injury, TBI)是致死率和致残率极高的外科疾患,我国在对于TBI的判断、治疗等方面还处于薄弱阶段,因此我们需要在分子层面了解大鼠颅脑损伤后基因及miRNA表达差异,以便更好地对症治疗。目的了解大鼠颅脑损伤后基因及miRNA表达差异,为临床治疗TBI提供新的思路。方法利用GEO2R筛选基因,然后用MiRwalk软件对筛选的miRNA的靶基因进行预测,再用DAVID做基因本体论功能富集分析,最后利用cytoscape做网络关系图。结果发现247个相对明显的差异表达的基因,包括150个上调表达基因和97个下调表达基因;7个差异表达的miRNA,包括2个上调表达miRNA和5个下调表达miRNA。这些差异表达基因在细胞内和细胞外都起作用,并且在炎症反应,药物应答等生物过程中起作用。将差异基因与靶基因对比后,可得到48个重合基因,同时发现这些重合基因与差异表达的miRNA有着一定的联系。     

Grouping Pentylenetetrazol-Induced Epileptic Rats According to Memory Impairment and MicroRNA Expression Profiles in the Hippocampus

Previous studies have demonstrated a close relationship between abnormal regulation of microRNA (miRNA) and various types of diseases, including epilepsy and other neurological disorders of memory. However, the role of miRNA in the memory impairment observed in epilepsy remains unknown. In this study, a model of temporal lobe epilepsy (TLE) was induced via pentylenetetrazol (PTZ) kindling in Sprague-Dawley rats. First, the TLE rats were subjected to Morris water maze to identify those with memory impairment (TLE-MI) compared with TLE control rats (TLE-C), which presented normal memory. Both groups were analyzed to detect dysregulated miRNAs in the hippocampus; four up-regulated miRNAs (miR-34c, miR-374, miR-181a, and miR-let-7c-1) and seven down-regulated miRNAs (miR-1188, miR-770-5p, miR-127-5p, miR-375, miR-331, miR-873-5p, and miR-328a) were found. Some of the dysregulated miRNAs (miR-34c, miR-1188a, miR-328a, and miR-331) were confirmed using qRT-PCR, and their blood expression patterns were identical to those of their counterparts in the rat hippocampus. The targets of these dysregulated miRNAs and other potentially enriched biological signaling pathways were analyzed using bioinformatics. Following these results, the MAPK, apoptosis and hippocampal signaling pathways might be involved in the molecular mechanisms underlying the memory disorders of TLE.     

microRNA层面上癌症的公共机制

microRNAs(miRNA)是一类内源性的非编码小RNA。已有研究表明miRNAs的靶基因中有不少癌症的相关基因。为了全面研究miRNA与癌症的关系,作者将19种癌症的相关基因集合分别富集到494个miRNAs靶基因集合上,得到各类癌症所富集的miRNAs。结果发现19种癌症仅集中地富集在144个miRNAs上,由此验证了癌症在miRNAs上的公共机制。在此基础上,作者对癌症富集较多的8个miRNAs做了进一步研究,结果发现这8个miRNAs均为高度保守的miRNAs,且它们的靶基因集合一致富集在基因本体论(gene ontology,GO)的基本生物学过程上,并与转录因子活性以及蛋白激酶活性相关。另一方面,在基于miRNA构建的癌症网络中,前列腺癌与乳腺癌,结肠癌与乳腺癌之间共享较多的miRNAs,表明了这些癌症在miRNA层面上存在密切的关系。     

Identification of Differential MicroRNAs in Cerebrospinal Fluid and Serum of Patients with Major Depressive Disorder

Major depression is a debilitating disease. To date, the development of biomarkers of major depressive disorder (MDD) remains a challenge. Recently, alterations in the expression of microRNAs (miRNAs) from post-mortem brain tissue and peripheral blood have been linked to MDD. The goals of this study were to detect the differential miRNAs in cerebrospinal fluid (CSF) and serum of MDD patients. First, the relative expression levels of 179 miRNAs (relative high levels in serum) were analyzed by miRNA PCR Panel in the CSF of MDD patients. Then, the differentially altered miRNAs from CSF were further assessed by qRT-PCR in the serum of the same patients. Finally, the serum differentially altered miRNAs were further validated by qRT-PCR in the serum of another MDD patients. The CSF-results indicated that 11 miRNAs in MDD patients were significantly higher than these in control subjects, and 5 miRNAs were significantly lower than these in control subjects. The serum-results from the same patients showed that 3 miRNAs (miR-221-3p, miR-34a-5p, and let-7d-3p) of the 11 miRNAs were significantly higher than these in control subjects, and 1 miRNA (miR-451a) of 5 miRNAs was significantly lower than these in control subjects. The up-regulation of miR-221-3p, miR-34a-5p, let-7d-3p and down-regulation of miR-451a was further validated in another 32 MDD patients. ROC analysis showed that the area under curve of let-7d-3p, miR-34a-5p, miR-221-3p and miR-451a was 0.94, 0.98, 0.97 and 0.94, with specificity of 90.48%, 95.24%, 90.48% and 90.48%, and sensitivity of 93.75%, 96.88%, 90.63% and 84.85%, respectively. In addition, target gene prediction found that the altered miRNAs are involved in affecting some important genes and pathway related to MDD. Our results suggested that differentially altered miRNAs in CSF might be involved in MDD, and serum miR-221-3p, miR-34a-5p, let-7d-3p, and miR-451a might be able to serve as biomarkers for MDD.     

MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in type 2 diabetes mellitus

Background

Dysregulation of microRNA (miRNA) expression in various tissues and body fluids has been demonstrated to be associated with several diseases, including Type 2 Diabetes mellitus (T2D). Here, we compare miRNA expression profiles in different tissues (pancreas, liver, adipose and skeletal muscle) as well as in blood samples from T2D rat model and highlight the potential of circulating miRNAs as biomarkers of T2D. In parallel, we have examined the expression profiles of miRNAs in blood samples from Impaired Fasting Glucose (IFG) and T2D male patients.

Methodology/Principal Findings

Employing miRNA microarray and stem-loop real-time RT-PCR, we identify four novel miRNAs, miR-144, miR-146a, miR-150 and miR-182 in addition to four previously reported diabetes-related miRNAs, miR-192, miR-29a, miR-30d and miR-320a, as potential signature miRNAs that distinguished IFG and T2D. Of these microRNAs, miR-144 that promotes erythropoiesis has been found to be highly up-regulated. Increased circulating level of miR-144 has been found to correlate with down-regulation of its predicted target, insulin receptor substrate 1 (IRS1) at both mRNA and protein levels. We could also experimentally demonstrate that IRS1 is indeed the target of miR-144.

Conclusion

We demonstrate that peripheral blood microRNAs can be developed as unique biomarkers that are reflective and predictive of metabolic health and disorder. We have also identified signature miRNAs which could possibly explain the pathogenesis of T2D and the significance of miR-144 in insulin signaling.     

miR-200c suppression increases tau hyperphosphorylation by targeting 14-3-3γ in early stage of 5xFAD mouse model of Alzheimer's disease

Background and Purpose: Recently, several abnormally regulated microRNAs (miRNAs) have been identified in patients with Alzheimer''s disease (AD). The purpose of this study was to identify abnormally expressed miRNAs and to investigate whether they affect pathological changes in AD in the 5xFAD AD mouse model.Experimental Approach: Using microarray analysis and RT-qPCR, miRNA expression in the hippocampus of a 4-month-old 5xFAD mouse model of AD was investigated. A dual-luciferase assay was performed to determine whether the altered miR-200c regulates the translation of the target mRNA, Ywhag. Whether miR-200c modulates AD pathology was determined in primary hippocampal neurons and C57BL/6J mice transfected with miR-200c inhibitor. In addition, total miRNAs were extracted from the serums of 28 healthy age-matched controls and 22 individual participants with cognitive impairment, and RT-qPCR was performed.Key results: miR-200c expression was reduced in the hippocampus of 5xFAD mice. In primary hippocampal neurons, miR-200c regulated the translation of 14-3-3γ and increased tau phosphorylation (p-tau) by increasing p-GSK-3β (GSK-3β phosphorylation). It was also confirmed that miR-200c inhibition in the hippocampus of C57BL/6J mice induces cognitive impairment and increases tau phosphorylation through 14-3-3γ activation. Finally, aberrant expression of miR-200c was confirmed in the blood serum of human AD patients.Conclusion and Implications: Our results strongly suggest that dysregulation of miR-200c expression contributes to the pathogenesis of AD, including cognitive impairment through hyperphosphorylated tau.     

miRNA profiling of naïve, effector and memory CD8 T cells

microRNAs have recently emerged as master regulators of gene expression during development and cell differentiation. Although profound changes in gene expression also occur during antigen-induced T cell differentiation, the role of miRNAs in the process is not known. We compared the miRNA expression profiles between antigen-specific na?ve, effector and memory CD8+ T cells using 3 different methods--small RNA cloning, miRNA microarray analysis and real-time PCR. Although many miRNAs were expressed in all the T cell subsets, the frequency of 7 miRNAs (miR-16, miR-21, miR-142-3p, miR-142-5p, miR-150, miR-15b and let-7f) alone accounted for approximately 60% of all miRNAs, and their expression was several fold higher than the other expressed miRNAs. Global downregulation of miRNAs (including 6/7 dominantly expressed miRNAs) was observed in effector T cells compared to na?ve cells and the miRNA expression levels tended to come back up in memory T cells. However, a few miRNAs, notably miR-21 were higher in effector and memory T cells compared to na?ve T cells. These results suggest that concomitant with profound changes in gene expression, miRNA profile also changes dynamically during T cell differentiation. Sequence analysis of the cloned mature miRNAs revealed an extensive degree of end polymorphism. While 3'end polymorphisms dominated, heterogeneity at both ends, resembling drosha/dicer processing shift was also seen in miR-142, suggesting a possible novel mechanism to generate new miRNA and/or to diversify miRNA target selection. Overall, our results suggest that dynamic changes in the expression of miRNAs may be important for the regulation of gene expression during antigen-induced T cell differentiation. Our study also suggests possible novel mechanisms for miRNA biogenesis and function.     

Human chromosome 21-derived miRNAs are overexpressed in down syndrome brains and hearts

Down syndrome (DS), or Trisomy 21, is the most common genetic cause of cognitive impairment and congenital heart defects in the human population. To date, the contribution of microRNAs (miRNAs) in DS has not been investigated. Bioinformatic analyses demonstrate that human chromosome 21 (Hsa21) harbors five miRNA genes; miR-99a, let-7c, miR-125b-2, miR-155, and miR-802. MiRNA expression profiling, miRNA RT-PCR, and miRNA in situ hybridization experiments demonstrate that these miRNAs are overexpressed in fetal brain and heart specimens from individuals with DS when compared with age- and sex-matched controls. We hypothesize that trisomic 21 gene dosage overexpression of Hsa21-derived miRNAs results in the decreased expression of specific target proteins and contribute, in part, to features of the neuronal and cardiac DS phenotype. Importantly, Hsa21-derived miRNAs may provide novel therapeutic targets in the treatment of individuals with DS.     

Deregulation of neuronal miRNAs induced by amyloid-β or TAU pathology

Background

Despite diverging levels of amyloid-β (Aβ) and TAU pathology, different mouse models, as well as sporadic AD patients show predictable patterns of episodic memory loss. MicroRNA (miRNA) deregulation is well established in AD brain but it is unclear whether Aβ or TAU pathology drives those alterations and whether miRNA changes contribute to cognitive decline.

Methods

miRNAseq was performed on cognitively intact (4 months) and impaired (10 months) male APPtg (APP^swe/PS1^L166P) and TAUtg (THY-Tau22) mice and their wild-type littermates (APPwt and TAUwt). We analyzed the hippocampi of 12 mice per experimental group (n =?96 in total), and employed a 2-way linear model to extract differentially expressed miRNAs. Results were confirmed by qPCR in a separate cohort of 4 M and 10 M APPtg and APPwt mice (n =?7–9 per group) and in human sporadic AD and non-demented control brain. Fluorescent in situ hybridization identified their cellular expression. Functional annotation of predicted targets was performed using GO enrichment. Behavior of wild-type mice was assessed after intracerebroventricular infusion of miRNA mimics.

Results

Six miRNAs (miR-10a-5p, miR-142a-5p, miR-146a-5p, miR-155-5p, miR-211-5p, miR-455-5p) are commonly upregulated between APPtg and TAUtg mice, and four of these (miR-142a-5p, miR-146a-5p, miR-155-5p and miR-455-5p) are altered in AD patients. All 6 miRNAs are strongly enriched in neurons. Upregulating these miRNAs in wild-type mice is however not causing AD-related cognitive disturbances.

Conclusion

Diverging AD-related neuropathologies induce common disturbances in the expression of neuronal miRNAs. 4 of these miRNAs are also upregulated in AD patients. Therefore these 4 miRNAs (miR-142a-5p, miR-146a-5p, miR-155-5p and miR-455-5p) appear part of a core pathological process in AD patients and APPtg and TAUtg mice. They are however not causing cognitive disturbances in wild-type mice. As some of these miRNA target AD relevant proteins, they may be, in contrast, part of a protective response in AD.

     

Temporal differences in microRNA expression patterns in astrocytes and neurons after ischemic injury

MicroRNAs (miRNAs) are small, non-protein-coding RNA molecules that modulate gene translation. Their expression is altered in many central nervous system (CNS) injuries suggesting a role in the cellular response to stress. Current studies in brain tissue have not yet described the cell-specific temporal miRNA expression patterns following ischemic injury. In this study, we analyzed the expression alterations of a set of miRNAs in neurons and astrocytes subjected to 60 minutes of ischemia and collected at different time-points following this injury. To mimic ischemic conditions and reperfusion in vitro, cortical primary neuronal and astrocytic cultures prepared from fetal rats were first placed in oxygen and glucose deprived (OGD) medium for 60 minutes, followed by their transfer into normoxic pre-conditioned medium. Total RNA was extracted at different time-points after the termination of the ischemic insult and the expression levels of miRNAs were measured. In neurons exposed to OGD, expression of miR-29b was upregulated 2-fold within 6 h and up to 4-fold at 24 h post-OGD, whereas induction of miR-21 was upregulated 2-fold after 24 h when compared to expression in neurons under normoxic conditions. In contrast, in astrocytes, miR-29b and miR-21 were upregulated only after 12 h. MiR-30b, 107, and 137 showed expression alteration in astrocytes, but not in neurons. Furthermore, we show that expression of miR-29b was significantly decreased in neurons exposed to Insulin-Like Growth Factor I (IGF-I), a well documented neuroprotectant in ischemic models. Our study indicates that miRNAs expression is altered in neurons and astrocytes after ischemic injury. Furthermore, we found that following OGD, specific miRNAs have unique cell-specific temporal expression patterns in CNS. Therefore the specific role of each miRNA in different intracellular processes in ischemic brain and the relevance of their temporal and spatial expression patterns warrant further investigation that may lead to novel strategies for therapeutic interventions.     

MicroRNA-432 contributes to dopamine cocktail and retinoic acid induced differentiation of human neuroblastoma cells by targeting NESTIN and RCOR1 genes

MicroRNA (miRNA) regulates expression of protein coding genes and has been implicated in diverse cellular processes including neuronal differentiation, cell growth and death. To identify the role of miRNA in neuronal differentiation, SH-SY5Y and IMR-32 cells were treated with dopamine cocktail and retinoic acid to induce differentiation. Detection of miRNAs in differentiated cells revealed that expression of many miRNAs was altered significantly. Among the altered miRNAs, human brain expressed miR-432 induced neurite projections, arrested cells in G0–G1, reduced cell proliferation and could significantly repress NESTIN/NES, RCOR1/COREST and MECP2. Our results reveal that miR-432 regulate neuronal differentiation of human neuroblastoma cells.