Dataset Information

MiRNA Analysis by Quantitative PCR in Preterm Human Breast Milk Reveals Daily Fluctuations of hsa-miR-16-5p.

ABSTRACT:

Background and aims

Human breast milk is an extremely dynamic fluid containing many biologically-active components which change throughout the feeding period and throughout the day. We designed a miRNA assay on minimized amounts of raw milk obtained from mothers of preterm infants. We investigated changes in miRNA expression within month 2 of lactation and then over the course of 24 hours.

Materials and methods

Analyses were performed on pooled breast milk, made by combining samples collected at different clock times from the same mother donor, along with time series collected over 24 hours from four unsynchronized mothers. Whole milk, lipids or skim milk fractions were processed and analyzed by qPCR. We measured hsa-miR-16-5p, hsa-miR-21-5p, hsa-miR-146-5p, and hsa-let-7a, d and g (all -5p). Stability of miRNA endogenous controls was evaluated using RefFinder, a web tool integrating geNorm, Normfinder, BestKeeper and the comparative ??Ct method.

Results

MiR-21 and miR-16 were stably expressed in whole milk collected within month 2 of lactation from four mothers. Analysis of lipids and skim milk revealed that miR-146b and let-7d were better references in both fractions. Time series (5H-23H) allowed the identification of a set of three endogenous reference genes (hsa-let-7d, hsa-let-7g and miR-146b) to normalize raw quantification cycle (Cq) data. We identified a daily oscillation of miR-16-5p.

Perspectives

Our assay allows exploring miRNA levels of breast milk from mother with preterm baby collected in time series over 48-72 hours.

SUBMITTER: Floris I

PROVIDER: S-EPMC4608744 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Publications

MiRNA Analysis by Quantitative PCR in Preterm Human Breast Milk Reveals Daily Fluctuations of hsa-miR-16-5p.

Floris Ilaria I Billard Hélène H Boquien Clair-Yves CY Joram-Gauvard Evelyne E Simon Laure L Legrand Arnaud A Boscher Cécile C Rozé Jean-Christophe JC Bolaños-Jiménez Francisco F Kaeffer Bertrand B

PloS one 20151016 10

<h4>Background and aims</h4>Human breast milk is an extremely dynamic fluid containing many biologically-active components which change throughout the feeding period and throughout the day. We designed a miRNA assay on minimized amounts of raw milk obtained from mothers of preterm infants. We investigated changes in miRNA expression within month 2 of lactation and then over the course of 24 hours.<h4>Materials and methods</h4>Analyses were performed on pooled breast milk, made by combining sampl ...[more]

PMID: 26474056

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Project description:ObjectiveSkin cutaneous melanoma(SKCM) is the most severe, and complex disease of all skin cancers. The molecular mechanisms of this cancer progression are not well understood.MethodsGEPIA online database was used to validate the differentially expressed genes from two GEO datasets. The prognostic value was calculated by the Kaplan-Meier method. RT-qPCR verified the expression of TUBB4A in SKCM cell line, and the immunohistochemistry of TUBB4A in SKCM and normal skin tissues were gained from Human Protein Atlas. Seven target prediction databases predicted potential microRNAs(miRNAs), and upstream long non-coding RNAs(lncRNAs) were predicted by starBase. The co-expressed gene of TUBB4A was obtained using the two online analysis sites UALCAN and starBase. These co-expressed genes were performed by enrichment analysis, and immune infiltration result was obtained by the TIMER2 online database. The receiver operating characteristic( ROC) curve was applied to evaluate the diagnostic value of TUBB4A in the SKCM and normal skin group. A new nomogram about TUBB4A was constructed to forecast the survival rate of SKCM patients at 1, 3, and 5 years.ResultsFirstly, we found that DLL3 and TUBB4A were significantly higher expressed in skin cutaneous melanoma than normal skin. Subsequently, by analyzing progress-free interval(PFI), diseasespecific survival(DSS), and disease-free survival(DFS), only TUBB4A was the most potent gene for inhibiting shin cutaneous melanoma progression. In gene ontology(GO)/ kyoto encyclopedia of genes and genomes(KEGG) analysis, TUBB4A may play a key role in the progression of skin cutaneous melanoma by regulating mitochondrial function and affecting cellular metabolism, possibly related to the immune infiltration of CD4+Th1 cells and NK cells. The upstream non-coding RNA(ncRNA) acts through the SNHG16-hsa-let-7b-5p-TUBB4A axis.ConclusionIn conclusion, we elucidated the regulatory role of the SNHG16-hsa-let-7b-5p-TUBB4A axis in the progression of skin cutaneous melanoma by modulating mitochondrial function to affect cellular metabolism. TUBB4A may be a promising diagnostic biomarker and therapeutic target for cutaneous skin melanoma.

Dataset Information

MiRNA Analysis by Quantitative PCR in Preterm Human Breast Milk Reveals Daily Fluctuations of hsa-miR-16-5p.

Background and aims

Materials and methods

Results

Perspectives

Publications

MiRNA Analysis by Quantitative PCR in Preterm Human Breast Milk Reveals Daily Fluctuations of hsa-miR-16-5p.

Similar Datasets

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