206 EFFECT OF OVARIAN HYPERSTIMULATION ON CIRCULATORY microRNA PROFILE IN BOVINE FOLLICULAR FLUID AND BLOOD PLASMA

2013 ◽  
Vol 25 (1) ◽  
pp. 251 ◽  
Author(s):  
S. Seifi Noferesti ◽  
M. Hoelker ◽  
M. H. Sohel ◽  
D. Salilew-Wondim ◽  
F. Rings ◽  
...  
Keyword(s):  
Blood Plasma ◽  
Small Rna ◽  
Follicular Fluid ◽  
Mirna Expression ◽  
Target Genes ◽  
Axonal Guidance ◽  
Differentially Expressed ◽  
Ovarian Hyperstimulation ◽  
Pcr Array ◽  
In Blood Plasma

MicroRNAs (miRNA) are noncoding small RNA that are known to play a role in posttranscriptional regulation of genes involved in various physiological processes including disease and reproduction. The circulatory forms of miRNA, which are present in body fluids, are reported to be used as biomarkers for disease and pregnancy. This study was conducted to investigate the effect of ovarian hyperstimulation on the expression pattern of circulatory miRNA in blood plasma and follicular fluid. For this, one group of Simmental heifers (n = 6) was hyper-stimulated using a superovulation (SO) protocol of 8 consecutive FSH injections over 4 days in decreasing doses, and others (n = 6) were synchronized (SY) using a standard synchronization protocol. Following this, the blood samples were collected at Day 0 (onset of oestrus) and Day 7, and follicular fluid was collected from both groups at Day 0 of the oestrous cycle. Total RNA, including small RNA, was then isolated from follicular fluid and blood plasma using a miRNeasy mini kit (Qiagen, Valencia, CA, USA) and subsequently used for circulatory miRNA expression studies using the human miRCURY LNA™ Universal RT miRNA PCR array system (Exiqon, Woburn, MA, USA). Following the miRNA PCR array run, data analysis was performed using a delta threshold cycle (ΔCT) method. Results showed that 23 miRNAs were found to be differentially expressed in blood plasma (fold change ≥2 and P ≤ 0.05) between SO and SY groups. Among these, 8 miRNAs including miR-127-3p, miR-494, miR-147, miR-134, and miR-153 were downregulated and 15 miRNAs including miR-34a, miR-103, miR-181c, miR-24-2-3p, miR-18a-3p, miR-20b-3p, and miR-708-3p were found to be upregulated in SO groups. Similarly, in follicular fluid, 71 miRNAs were found to be differentially expressed between the SO and SY groups. Of these, 33 miRNA including miR-100, miR-877, miR-659, miR-200c, miR-29b-2-3p, miR-361-5p, and miR-145 were downregulated, whereas 38 miRNAs including miR-374a, miR-720, miR-155-3p, miR-202-3p, miR-33b, miR-12-3p, and miR-224-3p were upregulated in follicular fluid aspirated from SO cows compared with the SY animals. Ingenuity pathway analysis of predicted target genes of miRNA, which are dysregulated due to ovarian hyperstimulation, showed the dominance of pathways related to neuregulin signalling, axonal guidance signalling, GNRH signalling, and Wnt β-catenin signalling pathways. In conclusion, this study revealed alternation in circulatory miRNA expression profile both in blood plasma and follicular fluid as a result of ovarian hyperstimulation.

scholarly journals Small RNA sequencing evaluation of renal microRNA biomarkers in dogs with X-linked hereditary nephropathy

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Candice P. Chu ◽  
Shiguang Liu ◽  
Wenping Song ◽  
Ethan Y. Xu ◽  
Mary B. Nabity
Keyword(s):  
Small Rna ◽  
Target Genes ◽  
Differentially Expressed ◽  
Small Rna Sequencing ◽  
Rna Seq ◽  
Ckd Progression ◽  
Critical Pathways ◽  
Qrt Pcr ◽  
Hereditary Nephropathy ◽  
Differentially Expressed Mirnas

AbstractDogs with X-linked hereditary nephropathy (XLHN) are an animal model for Alport syndrome in humans and progressive chronic kidney disease (CKD). Using mRNA sequencing (mRNA-seq), we have characterized the gene expression profile affecting the progression of XLHN; however, the microRNA (miRNA, miR) expression remains unknown. With small RNA-seq and quantitative RT-PCR (qRT-PCR), we used 3 small RNA-seq analysis tools (QIAGEN OmicSoft Studio, miRDeep2, and CPSS 2.0) to profile differentially expressed renal miRNAs, top-ranked miRNA target genes, and enriched biological processes and pathways in CKD progression. Twenty-three kidney biopsies were collected from 5 dogs with XLHN and 4 age-matched, unaffected littermates at 3 clinical time points (T1: onset of proteinuria, T2: onset of azotemia, and T3: advanced azotemia). We identified up to 23 differentially expressed miRNAs at each clinical time point. Five miRNAs (miR-21, miR-146b, miR-802, miR-142, miR-147) were consistently upregulated in affected dogs. We identified miR-186 and miR-26b as effective reference miRNAs for qRT-PCR. This study applied small RNA-seq to identify differentially expressed miRNAs that might regulate critical pathways contributing to CKD progression in dogs with XLHN.

115 Exosome-mediated microRNA expression profile in follicular fluid of metabolically divergent postpartum cows

2019 ◽  
Vol 31 (1) ◽  
pp. 183
Author(s):  
T. Hailay ◽  
M. Hoelker ◽  
S. Gebremedhn ◽  
F. Rings ◽  
M. M. Saeed-Zidane ◽  
...  
Keyword(s):  
Energy Balance ◽  
Follicular Fluid ◽  
Mirna Expression ◽  
Cell Communication ◽  
Negative Energy ◽  
Negative Energy Balance ◽  
Differentially Expressed ◽  
Metabolic Status ◽  
Lactation Period ◽  
Down Regulation

Most high-milking cows enter a state of negative energy balance during the early lactation period. This phenomenon disturbs the metabolic status of the follicular fluid microenvironment, resulting in delayed ovulation. Cell-to-cell communication between the oocyte and the surrounding cells is crucial during folliculogenesis. Exosomes, evolutionarily conserved cargo molecules (30-150nm in diameter) carrying RNA and proteins, are known to be involved in cell-to-cell communication. Here, we aimed to investigate the association between postpartum metabolic status and the expression of exosomal microRNA (miRNA) in follicular fluid of Holstein-Friesian cows. For this, follicular fluid was collected from antral follicles (>8mm in diameter) using ovum pickup procedure from cows (n=30) on a weekly basis between weeks 5 and 10 postpartum. Follicular fluid collected from heifers (n=8) was used as a control. The energy status of each cow was assessed based on the blood metabolite (nonesterified fatty acids and β-hydroxybutyrate) concentration, body weight curve, and overall energy balance determined by dry matter intake. Afterwards, cows were categorized as early negative and late positive (cows show negative energy balance at early weeks and recovered at late weeks postpartum), always negative (cows did not recover until 15 weeks postpartum), and always positive (cows did not enter in to a state of negative energy balance). Following this, exosomes were isolated from pooled samples from each animal category using ultracentrifugation, and their morphology and size was characterised using electron microscopy and nanosight, respectively. Exosomal total RNA enriched with miRNA was isolated using an exosomal RNA isolation kit. Next-generation sequencing of miRNA was performed using Illumina NextSEqn 500 (Illumina Inc., San Diego, CA, USA). MicroRNAs with a fold change ≥2, P-value <0.05, and a false discovery rate of <0.1 were considered differentially expressed. The results showed that a total of 356 known and 156 novel miRNA were identified across samples. Differential expression analysis of miRNA between always-negative cows versus always-positive cows revealed down-regulation of all 6 differentially expressed miRNA, including bta-miR-451, bta-miR-132, and bta-miR-2285. Similarly, down-regulation of 14 miRNA, including bta-miR-20b, bta-miR-363, bta-miR-132, and bta-miR-451, and up-regulation of 3 miRNA was observed in always-negative cows compared to heifers. Furthermore, the target prediction analysis of the down-regulated miRNA have been shown to be involved in regulating different pathways including transforming growth factor-β signalling, cell cycle, hippo signalling, forkhead box O signalling, and endometrial cancer, among others. In conclusion, the results revealed that although negative energy balance in postpartum dairy cows suppressed exosomal miRNA expression in follicular fluid, the opposite was observed in metabolically unstressed cows. This divergence of exosome-mediated miRNA expression in the follicular fluid of metabolically stressed cows could be associated with the reduced fertility of those cows.

scholarly journals Identification of MicroRNAs That Respond to Soybean Cyst Nematode Infection in Early Stages in Resistant and Susceptible Soybean Cultivars

2019 ◽  
Vol 20 (22) ◽  
pp. 5634 ◽  
Author(s):  
Piao Lei ◽  
Bing Han ◽  
Yuanyuan Wang ◽  
Xiaofeng Zhu ◽  
Yuanhu Xuan ◽  
...  
Keyword(s):  
Small Rna ◽  
Soybean Cyst Nematode ◽  
High Throughput Sequencing ◽  
Target Genes ◽  
Cyst Nematode ◽  
Differentially Expressed ◽  
Early Stages ◽  
Differentially Expressed Mirnas ◽  
Plant Pathogen Interactions ◽  
Insight Into

Soybean cyst nematode (SCN) causes heavy losses to soybean yield. In order to investigate the roles of soybean miRNAs during the early stages of infection (1 and 5 dpi), 24 small RNA libraries were constructed from SCN resistant cultivar Huipizhi (HPZ) and the susceptible Williams 82 (W82) cultivar for high-throughput sequencing. By sequencing the small RNA libraries, a total of 634 known miRNAs were identified, and 252 novel miRNAs were predicted. Altogether, 14 known miRNAs belonging to 13 families, and 26 novel miRNAs were differentially expressed and may respond to SCN infection in HPZ and W82. Similar expression results were also confirmed by qRT-PCR. Further analysis of the biological processes that these potential target genes of differentially expressed miRNAs regulate found that they may be strongly related to plant–pathogen interactions. Overall, soybean miRNAs experience profound changes in early stages of SCN infection in both HPZ and W82. The findings of this study can provide insight into miRNAome changes in both HPZ and W82 at the early stages of infection, and may provide a stepping stone for future SCN management.

scholarly journals Small RNA Sequencing Reveals Differentially Expressed miRNAs in Necrotizing Enterocolitis in Rats

2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Ren-qiang Yu ◽  
Min Wang ◽  
Shan-yu Jiang ◽  
Ying-hui Zhang ◽  
Xiao-yu Zhou ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Signaling Pathway ◽  
Rna Sequencing ◽  
Necrotizing Enterocolitis ◽  
Small Rna ◽  
Mirna Expression ◽  
Expression Patterns ◽  
Wnt Signaling Pathway ◽  
Differentially Expressed ◽  
Small Rna Sequencing

Necrotizing enterocolitis (NEC) is the leading cause of death due to gastrointestinal disease in preterm infants. The role of miRNAs in NEC is still unknown. The objective of this study was to identify differentially expressed (DE) miRNAs in rats with NEC and analyze their possible roles. In this study, a NEC rat model was established using Sprague-Dawley rat pups. Small RNA sequencing was used to analyze the miRNA expression profiles in the NEC and control rats. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were carried out to identify target mRNAs for the DE miRNAs and to explore their potential roles. The DE miRNAs were verified by real-time quantitative PCR (RT-qPCR). The status of intestinal injury and the elevated levels of inflammatory cytokines in the NEC group confirmed that the NEC model was successfully established. The 16 miRNAs were found to be differentially expressed between the NEC group and the control group of rats. Bioinformatics analysis indicated that the parental genes of the DE miRNAs were predominantly implicated in the phosphorylation, cell migration, and protein phosphorylation processes. Moreover, the DE miRNAs were mainly found to be involved in the pathways of axon guidance, endocytosis, and focal adhesion, as well as in the Wnt signaling pathway, which is related to colitis. The expression patterns of the candidate miRNAs (rno-miR-27a-5p and rno-miR-187-3p), as assessed by RT-qPCR, were in accordance with the expression patterns obtained by miRNA-sequencing. The miRNA/mRNA/pathway network revealed that rno-miR-27a-5p and rno-miR-187-3p might be involved in NEC via the Wnt signaling pathway. We found an altered miRNA expression pattern in rats with NEC. We hypothesize that rno-miR-27a-5p and rno-miR-187-3p might mediate the NEC pathophysiological processes via the Wnt signaling pathway.

scholarly journals Controlled ovarian hyperstimulation induced changes in the expression of circulatory miRNA in bovine follicular fluid and blood plasma

2015 ◽  
Vol 8 (1) ◽  
Author(s):  
Sina Seifi Noferesti ◽  
Md. Mahmodul Hasan Sohel ◽  
Michael Hoelker ◽  
Dessie Salilew-Wondim ◽  
Ernst Tholen ◽  
...  
Keyword(s):  
Blood Plasma ◽  
Follicular Fluid ◽  
Controlled Ovarian Hyperstimulation ◽  
Ovarian Hyperstimulation ◽  
Induced Changes

scholarly journals Screening and identification of miRNAs related to sexual differentiation of strobili in Ginkgo biloba by integration analysis of small RNA, mRNA, and degradome sequencing

2020 ◽  
Author(s):  
Xiao-Meng Liu ◽  
Shui-Yuan Cheng ◽  
Jia-Bao Ye ◽  
Ze-Xiong Chen ◽  
Yong-Ling Liao ◽  
...  
Keyword(s):  
Small Rna ◽  
Ginkgo Biloba ◽  
Sexual Differentiation ◽  
Target Genes ◽  
Differentially Expressed ◽  
Sequencing Data ◽  
Multiple Perspectives ◽  
Juvenile Period ◽  
Degradome Sequencing ◽  
Screening And Identification

Abstract Background: Ginkgo biloba, a typical dioecious plant, is a traditional medicinal plant widely planted. However, it has a long juvenile period, which severely affected the breeding and cultivation of superior ginkgo varieties.Results: In order to clarify the complex mechanism of sexual differentiation in G. biloba strobili. Here, a total of 3,293 miRNAs were identified in buds and strobili of G. biloba, including 1,085 conserved miRNAs and 2,208 novel miRNAs using the three sequencing approaches of transcriptome, small RNA, and degradome. Comparative transcriptome analysis screened 4,346 and 7,087 differentially expressed genes (DEGs) in MB _vs_ FB and MS _vs_ OS, respectively. A total of 6,032 target genes were predicted for differentially expressed miRNA. The combined analysis of both small RNA and transcriptome datasets identified 51 miRNA-mRNA interaction pairs that may be involved in the process of G. biloba strobili sexual differentiation, of which 15 pairs were verified in the analysis of degradome sequencing. Conclusions: The comprehensive analysis of the small RNA, RNA and degradome sequencing data in this study provided candidate genes and clarified the regulatory mechanism of sexual differentiation of G. biloba strobili from multiple perspectives.

scholarly journals The expression of small RNAs in exosomes of follicular fluid altered in human polycystic ovarian syndrome

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e8640 ◽  
Author(s):  
Junhe Hu ◽  
Tao Tang ◽  
Zhi Zeng ◽  
Juan Wu ◽  
Xiansheng Tan ◽  
...  
Keyword(s):  
Small Rna ◽  
Follicular Fluid ◽  
Target Genes ◽  
Polycystic Ovary ◽  
Micro Rna ◽  
Control Group ◽  
Rna Sequences ◽  
Human Follicular Fluid ◽  
Non Coding Rna ◽  
Pcos Group

Polycystic ovary syndrome (PCOS) can cause reproductive disorders that may affect oocyte quality from punctured follicles in human follicular fluid (HFF). The non-coding RNA family includes micro RNA (miRNA), piwi-interacting RNA (piRNA) and transfer RNA (tRNA); these non-coding RNA transcripts play diverse functions and are implicated in a variety of diseases and health conditions, including infertility. In this study, to explore the role of HFF exosomes in PCOS, we extracted and sequenced RNA from HFF exosomes of PCOS patients and compared the analysis results with those of non-PCOS control group. The HFF exosomes were successfully isolated and characterized in a variety of ways. The sequencing results of the HFF exosomal RNA showed that about 6.6% of valid reads in the PCOS group and 8.6% in the non-PCOS group were successfully mapped to the human RNA database. Using a hierarchical clustering method, we found there were ten small RNA sequences whose expression was significantly different between the PCOS and non-PCOS groups. We chose six of them to predict target genes of interest for further GO analysis, and pathway analysis showed that the target genes are mainly involved in biosynthesis of amino acids, glycine, serine and glycosaminoglycan, as well as threonine metabolism. Therefore, the small RNA sequences contained in HFF EXs may play a key role in the mechanism that drives PCOS pathogenesis, and thereby can act as molecular biomarkers for PCOS diagnosis in the future.

scholarly journals Determination of the MiRNAs Related to Bean Pyralid Larvae Resistance in Soybean Using Small RNA and Transcriptome Sequencing

2019 ◽  
Vol 20 (12) ◽  
pp. 2966 ◽  
Author(s):  
Weiying Zeng ◽  
Zudong Sun ◽  
Zhenguang Lai ◽  
Shouzhen Yang ◽  
Huaizhu Chen ◽  
...  
Keyword(s):  
Small Rna ◽  
Transcriptome Sequencing ◽  
Target Genes ◽  
Isoquinoline Alkaloid ◽  
Differentially Expressed ◽  
Pcr Analysis ◽  
Small Rna Sequencing ◽  
Regulatory Pathways ◽  
Oil Crops ◽  
Differentially Expressed Mirnas

Soybean is one of the most important oil crops in the world. Bean pyralid is a major leaf-feeding insect of soybean. In order to screen out the functional genes and regulatory pathways related to the resistance for bean pyralid larvae, the small RNA and transcriptome sequencing were performed based on the highly resistant material (Gantai-2-2) and highly susceptible material (Wan 82-178) of soybean. The results showed that, when comparing 48 h feeding with 0 h feeding, 55 differentially expressed miRNAs were identified in Gantai-2-2 and 58 differentially expressed miRNAs were identified in Wan82-178. When comparing Gantai-2-2 with Wan82-178, 77 differentially expressed miRNAs were identified at 0 h feeding, and 70 differentially expressed miRNAs were identified at 48 h feeding. The pathway analysis of the predicted target genes revealed that the plant hormone signal transduction, RNA transport, protein processing in the endoplasmic reticulum, zeatin biosynthesis, ubiquinone and other terpenoid-quinone biosynthesis, and isoquinoline alkaloid biosynthesis may play important roles in soybean’s defense against the stress caused by bean pyralid larvae. According to conjoint analysis of the miRNA/mRNA, a total of 20 differentially expressed miRNAs were negatively correlated with 26 differentially expressed target genes. The qRT-PCR analysis verified that the small RNA sequencing results were credible. According to the analyses of the differentially expressed miRNAs, we speculated that miRNAs are more likely to play key roles in the resistance to insects. Gma-miR156q, Gma-miR166u, Gma-miR166b, Gma-miR166j-3p, Gma-miR319d, Gma-miR394a-3p, Gma-miR396e, and so on—as well as their negatively regulated differentially expressed target genes—may be involved in the regulation of soybean resistance to bean pyralid larvae. These results laid a foundation for further in-depth research regarding the action mechanisms of insect resistance.

scholarly journals Genome-wide profiling of miRNA expression patterns in tubal endometriosis

Reproduction ◽  
2019 ◽  
Vol 157 (6) ◽  
pp. 525-534 ◽  
Author(s):  
Hang Qi ◽  
Guiling Liang ◽  
Jin Yu ◽  
Xiaofeng Wang ◽  
Yan Liang ◽  
...  
Keyword(s):  
Pathway Analysis ◽  
Mirna Expression ◽  
Gene Networks ◽  
Target Genes ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Mirna Gene ◽  
Differentially Expressed ◽  
Signal Pathways ◽  
Differentially Expressed Mirnas

MicroRNA (miRNA) expression profiles in tubal endometriosis (EM) are still poorly understood. In this study, we analyzed the differential expression of miRNAs and the related gene networks and signaling pathways in tubal EM. Four tubal epithelium samples from tubal EM patients and five normal tubal epithelium samples from uterine leiomyoma patients were collected for miRNA microarray. Bioinformatics analyses, including Ingenuity Pathway Analysis (IPA), Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, were performed. Quantitative real-time polymerase chain reaction (qRT-PCR) validation of five miRNAs was performed in six tubal epithelium samples from tubal EM and six from control. A total of 17 significantly differentially expressed miRNAs and 4343 potential miRNA-target genes involved in tubal EM were identified (fold change >1.5 and FDR-adjustedPvalue <0.05). IPA indicated connections between miRNAs, target genes and other gynecological diseases like endometrial carcinoma. GO and KEGG analysis revealed that most of the identified genes were involved in the mTOR signaling pathway, SNARE interactions in vesicular transport and endocytosis. We constructed an miRNA-gene-disease network using target gene prediction. Functional analysis showed that the mTOR pathway was connected closely to tubal EM. Our results demonstrate for the first time the differentially expressed miRNAs and the related signal pathways involved in the pathogenesis of tubal EM which contribute to elucidating the pathogenic mechanism of tubal EM-related infertility.

scholarly journals Transcriptome and Small RNA Combined Sequencing Analysis of Cold Tolerance in Non-heading Chinese Cabbage

2021 ◽  
Vol 12 ◽  
Author(s):  
Jin Wang ◽  
Qinxue Zhang ◽  
Xiong You ◽  
Xilin Hou
Keyword(s):  
Gene Expression ◽  
Brassica Rapa ◽  
Small Rna ◽  
Target Genes ◽  
Differentially Expressed ◽  
Sequencing Analysis ◽  
Pak Choi ◽  
Expression Levels ◽  
Heading Chinese Cabbage ◽  
Gene Expression Levels

BackgroundNon-heading Chinese cabbage (Brassica rapa ssp. chinensis) is an important leaf vegetable grown worldwide. However, there has currently been not enough transcriptome and small RNA combined sequencing analysis of cold tolerance, which hinders further functional genomics research.ResultsIn this study, 63.43 Gb of clean data was obtained from the transcriptome analysis. The clean data of each sample reached 6.99 Gb, and the basic percentage of Q30 was 93.68% and above. The clean reads of each sample were sequence aligned with the designated reference genome (Brassica rapa, IVFCAASv1), and the efficiency of the alignment varied from 81.54 to 87.24%. According to the comparison results, 1,860 new genes were discovered in Pak-choi, of which 1,613 were functionally annotated. Among them, 13 common differentially expressed genes were detected in all materials, including seven upregulated and six downregulated. At the same time, we used quantitative real-time PCR to confirm the changes of these gene expression levels. In addition, we sequenced miRNA of the same material. Our findings revealed a total of 34,182,333 small RNA reads, 88,604,604 kinds of small RNAs, among which the most common size was 24 nt. In all materials, the number of common differential miRNAs is eight. According to the corresponding relationship between miRNA and its target genes, we carried out Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis on the set of target genes on each group of differentially expressed miRNAs. Through the analysis, it is found that the distributions of candidate target genes in different materials are different. We not only used transcriptome sequencing and small RNA sequencing but also used experiments to prove the expression levels of differentially expressed genes that were obtained by sequencing. Sequencing combined with experiments proved the mechanism of some differential gene expression levels after low-temperature treatment.ConclusionIn all, this study provides a resource for genetic and genomic research under abiotic stress in Pak-choi.

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