Circulating microRNA signature of steroid‐induced osteonecrosis of the femoral head

Objectives

Steroid‐induced osteonecrosis of the femoral head (ONFH) is a common orthopaedic disease of which early detection remains clinically challenging. Accumulating evidences indicated that circulating microRNAs (miRNAs) plays vital roles in the development of several bone diseases. However, the association between circulating miRNAs and steroid‐induced ONFH remains elusive.

Materials and methods

miRNA microarray was performed to identify the differentially abundant miRNAs in the serums of systemic lupus erythematosus (SLE) patients with steroid‐induced ONFH as compared with SLE control and healthy control group. We predicted the potential functions of these differentially abundant miRNAs using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses and reconstructed the regulatory networks of miRNA‐mRNA interactions.

Results

Our data indicated that there were 11 differentially abundant miRNAs (2 upregulated and 9 downregulated) between SLE‐ONFH group and healthy control group and 42 differentially abundant miRNAs (14 upregulated and 28 downregulated) between SLE‐ONFH group and SLE control group. We also predicted the potential functions of these differentially abundant miRNAs using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses and reconstructed the regulatory networks of miRNA‐mRNA interactions.

Conclusions

These findings corroborated the idea that circulating miRNAs play significant roles in the development of ONFH and may serve as diagnostic markers and therapeutic targets.     

MicroRNA‐26a is a novel regulator of vascular smooth muscle cell function

Aberrant smooth muscle cell (SMC) plasticity has been implicated in a variety of vascular disorders including atherosclerosis, restenosis, and abdominal aortic aneurysm (AAA) formation. While the pathways governing this process remain unclear, epigenetic regulation by specific microRNAs (miRNAs) has been demonstrated in SMCs. We hypothesized that additional miRNAs might play an important role in determining vascular SMC phenotype. Microarray analysis of miRNAs was performed on human aortic SMCs undergoing phenotypic switching in response to serum withdrawal, and identified 31 significantly regulated entities. We chose the highly conserved candidate miRNA‐26a for additional studies. Inhibition of miRNA‐26a accelerated SMC differentiation, and also promoted apoptosis, while inhibiting proliferation and migration. Overexpression of miRNA‐26a blunted differentiation. As a potential mechanism, we investigated whether miRNA‐26a influences TGF‐β‐pathway signaling. Dual‐luciferase reporter assays demonstrated enhanced SMAD signaling with miRNA‐26a inhibition, and the opposite effect with miRNA‐26a overexpression in transfected human cells. Furthermore, inhibition of miRNA‐26a increased gene expression of SMAD‐1 and SMAD‐4, while overexpression inhibited SMAD‐1. MicroRNA‐26a was also found to be downregulated in two mouse models of AAA formation (2.5‐ to 3.8‐fold decrease, P < 0.02) in which enhanced switching from contractile to synthetic phenotype occurs. In summary, miRNA‐26a promotes vascular SMC proliferation while inhibiting cellular differentiation and apoptosis, and alters TGF‐β pathway signaling. MicroRNA‐26a represents an important new regulator of SMC biology and a potential therapeutic target in AAA disease. J. Cell. Physiol. 226: 1035–1043, 2011. © 2010 Wiley‐Liss, Inc.     

Characterization of microRNAs from sheep (<Emphasis Type="Italic">Ovis aries</Emphasis>) using computational and experimental analyses

Ovis aries is one of the most important agricultural livestock for meat production, and also is an ideal model organism for biological and comparative genomics studies. Many miRNAs have been reported for their important roles in developmental processes in various animals, but there is limited information about O. aries miRNAs. In this study, combining a computational method based on expressed sequence tag (EST) analysis with experimental identification based on small RNA cDNA library, we identified 31 miRNAs belong to 24 families in sheep, 2 of which were novel miRNAs which had never been previously identified in any species. Especially, we cloned 12 miRNAs from the sheep skeletal muscle, which were good candidate miRNAs to be studied about the miRNA-dependant regulated process of muscle development, and we identified four pairs of miRNA/miRNA* and one pair of miRNA-3p/miRNA-5p from sheep EST sequences. Expression analysis indicated that some miRNAs were expressed in a specific tissue, and the pair of miRNA-3p/miRNA-5p and one pair of miRNA/miRNA* had a similar relative expression pattern in some tissues, respectively. Further, we predicted 120 potential target genes of 31 oar-miRNAs on the 3′UTR of O. aries genes. Gene ontology analysis showed that most of these genes took part in the cellular process and metabolic process. Our results enriched the O. aries miRNA database and provided useful information for investigating biological functions of miRNAs and miRNA* in sheep.     

Deregulated microRNAs in CD4+ T cells from individuals with latent tuberculosis versus active tuberculosis

The mechanisms of latent tuberculosis (TB) infection remain elusive. Roles of microRNA (miRNA) have been highlighted in pathogen–host interactions recently. To identify miRNAs involved in the immune response to TB, expression profiles of miRNAs in CD4⁺ T cells from patients with latent TB, active TB and healthy controls were investigated by microarray assay and validated by RT‐qPCR. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were used to analyse the significant functions and involvement in signalling pathways of the differentially expressed miRNAs. To identify potential target genes for miR‐29, interferon‐γ (IFN‐γ) mRNA expression was measured by RT‐qPCR. Our results showed that 27 miRNAs were deregulated among the three groups. RT‐qPCR results were generally consistent with the microarray data. We observed an inverse correlation between miR‐29 level and IFN‐γ mRNA expression in CD4⁺ T cells. GO and KEGG pathway analysis showed that the possible target genes of deregulated miRNAs were significantly enriched in mitogen‐activated protein kinase signalling pathway, focal adhesion and extracellular matrix receptor interaction, which might be involved in the transition from latent to active TB. In all, for the first time, our study revealed that some miRNAs in CD4⁺ T cells were altered in latent and active TB. Function and pathway analysis highlighted the possible involvement of miRNA‐deregulated mRNAs in TB. The study might help to improve understanding of the relationship between miRNAs in CD4⁺ T cells and TB, and laid an important foundation for further identification of the underlying mechanisms of latent TB infection and its reactivation.     

Global investigation of the co‐evolution of MIRNA genes and microRNA targets during soybean domestication

Although the selection of coding genes during plant domestication has been well studied, the evolution of MIRNA genes (MIRs) and the interaction between microRNAs (miRNAs) and their targets in this process are poorly understood. Here, we present a genome‐wide survey of the selection of MIRs and miRNA targets during soybean domestication and improvement. Our results suggest that, overall, MIRs have higher evolutionary rates than miRNA targets. Nonetheless, they do demonstrate certain similar evolutionary patterns during soybean domestication: MIRs and miRNA targets with high expression and duplication status, and with greater numbers of partners, exhibit lower nucleotide divergence than their counterparts without these characteristics, suggesting that expression level, duplication status, and miRNA–target interaction are essential for evolution of MIRs and miRNA targets. Further investigation revealed that miRNA–target pairs that are subjected to strong purifying selection have greater similarities than those that exhibited genetic diversity. Moreover, mediated by domestication and improvement, the similarities of a large number of miRNA–target pairs in cultivated soybean populations were increased compared to those in wild soybeans, whereas a small number of miRNA–target pairs exhibited decreased similarity, which may be associated with the adoption of particular domestication traits. Taken together, our results shed light on the co‐evolution of MIRs and miRNA targets during soybean domestication.     

Genome‐wide identification and characterization of microRNAs responding to ABA and GA in maize embryos during seed germination

• MicroRNAs (miRNAs) are an important class of non‐coding small RNAs that regulate the expression of target genes through mRNA cleavage or translational inhibition. Previous studies have revealed their roles in regulating seed dormancy and germination in model plants such as Arabidopsis thaliana, rice (Oryza sativa) and maize (Zea mays). However, the miRNA response to exogenous gibberellic acid (GA) and abscisic acid (ABA) during seed germination in maize has yet to be explored.
• In this study, small RNA libraries were generated and sequenced from maize embryos treated with GA, ABA or double‐distilled water as control.
• A total of 247 miRNAs (104 known and 143 novel) were identified, of which 45 known and 53 novel miRNAs were differentially expressed in embryos in the different treatment groups. In total, 74 (37 up‐regulated and 37 down‐regulated) and 55 (23 up‐regulated and 32 down‐regulated) miRNAs were expressed in response to GA and to ABA, respectively, and a total of 18 known and 38 novel miRNAs displayed differential expression between the GA‐ and ABA‐treated groups. Using bioinformatics tools, we predicted the target genes of the differentially expressed miRNAs. Using GO enrichment and KEGG pathway analysis of these targets, we showed that miRNAs differentially expressed in our samples affect genes encoding proteins involved in the peroxisome, ribosome and plant hormonal signalling pathways.
• Our results indicate that miRNA‐mediated gene expression influences the GA and ABA signalling pathways during seed germination.

     

Identification of microRNAs and their corresponding targets involved in the susceptibility interaction of wheat response to Puccinia striiformis f. sp. tritici

MicroRNAs (miRNAs) play very important roles in plant defense responses. However, little is known about their roles in the susceptibility interaction between wheat and Puccinia striiformis f. sp. tritici (Pst). In this study, two miRNA libraries were constructed from the leaves of the cultivar Xingzi 9104 inoculated with the virulent Pst race CYR32 and sterile water, respectively. A total of 1316 miRNA candidates, including 173 known miRNAs that were generated from 98 pre‐miRNAs, were obtained. The remaining 1143 miRNA candidates included 145 conserved and 998 wheat‐specific miRNAs that were generated from 87 and 1088 pre‐miRNAs, respectively. The 173 known and 145 conserved miRNAs were sub‐classified into 63 miRNA families. The target genes of wheat miRNAs were also confirmed using degradome sequencing technology. Most of the annotated target genes were related to signal transduction or energy metabolism. Additionally, we found that miRNAs and their target genes form complicated regulation networks. The expression profiles of miRNAs and their corresponding target genes were further analyzed by quantitative real‐time polymerase chain reaction (qRT‐PCR), and the results indicate that some miRNAs are involved in the compatible wheat‐Pst susceptibility interaction. Importantly, tae‐miR1432 was highly expressed when wheat was challenged with CYR32, and the corresponding target gene, predicted to be a calcium ion‐binding protein, also exhibited upregulated expression but a divergent expression trend. PC‐3P‐7484, a specific wheat miRNA, was highly expressed in the wheat response to Pst infection, while the expression of the corresponding target gene ubiquillin was dramatically downregulated. These data provide the foundation for evaluating the important regulatory roles of miRNAs in wheat‐Pst susceptibility interaction.     

Three vital RNA functions and interactions in the process of silk gland apoptosis in silkworm Bombyx mori

The Silkworm Bombyx mori is an important insect in terms of economics and a model organism with a complete metamorphosis. The economic importance of silkworms is dependent on the functions of the silkgland, a specialized organ that synthesizes silk proteins. The silk gland undergoes massive degeneration during the larval to pupal stage, which involves in cell apoptosis. In this paper, high throughput sequencing was used to detect the expression of messenger RNA (mRNA), long noncoding RNA (lncRNA), and microRNA (miRNA) from silk glands of Day 3 in the fifth instar larvae (L5D3) and the spinning 36h (sp36h). We analyzed the Gene Ontology (GO) functions of target genes of the differentially expressed lncRNAs and miRNAs. We investigated the regulations of mRNA, lncRNA, and miRNA on silk gland apoptosis in L5D3 and sp36h. In total, 10,947 lncRNAs were detected in the silk gland and the index number TCONS‐00021360 lncRNA may be involved in the process of apoptosis. In addition, 344 miRNAs targeted 285 mRNAs were related to the death process under GO entry. The results indicated that miRNAs play an important role in the molecular regulation of the silk gland apoptosis compared with that of lncRNAs. Finally, we screened 746 lncRNAs and 20 miRNAs that might interact with BmDredd, and drew an interaction network among them.     

Analyses of a Glycine max Degradome Library Identify microRNA Targets and MicroRNAs that Trigger Secondary SiRNA Biogenesis

Plant microRNAs (miRNAs) regulate gene expression mainly by guiding cleavage of target mRNAs. In this study, a degradome library constructed from different soybean (Glycine max (L.) Merr.) tissues was deep-sequenced. 428 potential targets of small interfering RNAs and 25 novel miRNA families were identified. A total of 211 potential miRNA targets, including 174 conserved miRNA targets and 37 soybean-specific miRNA targets, were identified. Among them, 121 targets were first discovered in soybean. The signature distribution of soybean primary miRNAs (pri-miRNAs) showed that most pri-miRNAs had the characteristic pattern of Dicer processing. The biogenesis of TAS3 small interfering RNAs (siRNAs) was conserved in soybean, and nine Auxin Response Factors were identified as TAS3 siRNA targets. Twenty-three miRNA targets produced secondary small interfering RNAs (siRNAs) in soybean. These targets were guided by five miRNAs: gma-miR393, gma-miR1508, gma-miR1510, gma-miR1514, and novel-11. Multiple targets of these secondary siRNAs were detected. These 23 miRNA targets may be the putative novel TAS genes in soybean. Global identification of miRNA targets and potential novel TAS genes will contribute to research on the functions of miRNAs in soybean.