Table 1.
miRNAs related to postmenopausal OP.
| Study | Study design | Biomarker source | Sample handling | Quantification platform | Evaluated miRNA | Normalization strategy | Validated miRNA biomarker | Potential target gene | AUC-Sensitivity(%)-Specificity(%) | Limits |
|---|---|---|---|---|---|---|---|---|---|---|
| (Wang et al., 2012) | 20 PM Caucasian women (age 57-68 years): 10 with low BMD (hip/spine Z-score < -0.84); 10 with high BMD (hip/spine Z-score > 0.84) | Circulating monocytes | Monocytes separated by density gradients in UNI-SEP tubes (sodium metrizoate 9.6% and polysucrose 5.6% with 1.077 g/ml density), and isolated using a negative isolation kit | Screening: TaqMan Human MicroRNA Array v1.0 Validation: TaqMan RT-qPCR |
Screening: 365 miRNAs tested Validation: miR-133a and miR-382 |
RNU48 | ↑miR-133a in low vs. high BMD group | CXCR3, CXCL11, and SLC39A1 (identified for miR-133a using miRDB and TargetScan database but not validated) | / | Small sample size; no significant correlation between the expression level of miR-133a and the potential target genes; no information about the stem-loop arm of miRNA origin; no ROC analysis |
| (Cao et al., 2014) | 21 PM Caucasian women (age 57-68 years): 10 with low BMD (hip/spine Z-score < -0.84); 10 with high BMD (hip/spine Z-score > 0.84) | Circulating monocytes | Monocytes separated by density gradients in UNI-SEP tubes (sodium metrizoate 9.6% and polysucrose 5.6% with 1.077 g/ml density), and isolated using a negative isolation kit | Screening: TaqMan Human MicroRNA Array v1.0 Validation: TaqMan RT-qPCR |
Screening: 365 miRNAs tested Validation: miR-27b, miR-422a, miR-151, and miR-152 |
RNU48 | ↑miR-422a in low vs. high BMD group | CD226, CBL, IGF1, TOB2, and PAG1 (identified for miR-422a using TargetScan database but not validated) | / | Small sample size; no significant correlation between miR-422a and the evaluated target genes; no information about the stem-loop arm of miRNA origin; no ROC analysis |
| (Chen et al., 2014a) | 31 Chinese PM women with OP and 30 healthy women (age 50-59 years). | PBMCs CD14+ | Ficoll-Paque separation step and CD14 antibody-coated magnetic cell sorting MicroBeads used for buffy coat PBMCs isolation and CD14+ purification, respectively | Screening: MicroRNA microarray by LC Sciences Validation: SYBR Green RT-qPCR |
Screening: 721 miRNAs tested Validation: miR-503 |
snRNU6 | ↓miR-503 in OP group vs. non-OP group | RANK (validated as miR-503 target gene) | / | Small sample size; no information about the stem-loop arm of miRNA origin; no ROC analysis |
| (Yang et al., 2013) | 5 OP PM women (age 53-63 years) and 5 premenopausal women (age 39-45 years) | BM-MCSs | Percoll density gradient centrifugation methodology obtaining BM-MCSs from the BM | Screening: LC Sciences microarray platform Validation: RT-qPCR |
Screening: 1040 miRNAs tested Validation: miR-21 |
snRNU6 | ↓miR-21 in PM OP group vs. non-OP group | SPRY1 (identified for miR-21 using Target Scan 6.0 and Pic Tar databases and validated by in vitro experiments) | / | Small sample size; no information about the stem-loop arm of miRNA origin; no ROC analysis |
| (Li et al., 2014) | 40 PM Chinese women with normal, 40 with OP, and 40 with osteopenia range BMD (age 46-69 years) | Cell-free plasma | Plasma obtained from fasting blood samples and stored in liquid nitrogen | miRCURY LNA RT-qPCR | miR-21, miR-133a, and miR-146a | miR-16 | ↓ miR-21 and ↑ miR-133a in OP and osteopenia groups vs. control | / | / | Small sample size; no information about the used anticoagulant; small sample size; arbitrary decision of the reference gene; no evaluation of the target genes; no information about the stem-loop arm of miRNA origin; no ROC analysis |
| (Meng et al., 2015) | Discovery cohort: 25 PM women with OP and 23 PM Chinese women with osteopenia (age 59-70 years)Validation cohort: 24 PM Chinese women with normal, 32 with OP and 30 with osteopenia range BMD (age 59-70 years) | Whole blood | Blood samples lysed using RBC lysis solution and centrifuged for 10 min at 450g | Discovery: Agilent Human miRNA microarray followed by SYBR Green RT-qPCR Validation: SYBR Green RT-qPCR |
Discovery cohort: comprehensive miRNA expression analysis (Microarray); miR-130b-3p, miR-151a-3p, miR-151b, miR-194-5p, miR-590-5p, and miR-660-5p (RT-qPCR) Validation cohort: miR-194-5p |
snRNU6 | ↑ miR-130b-3p, miR-151a-3p, miR-151b, miR-194-5p, and miR-590-5p in OP vs. osteopenia (Discovery cohort)↑ miR-194-5p in OP and osteopenia vs. control (Validation cohort) | / | / | Small sample size; no evaluation of the target genes; no ROC analysis |
| (You et al., 2016) | 155 PM Chinese women with PM OP (n = 81, age 51-62 years) or healthy (n = 74, age 40-46 years) | Cell-free serum and BM-MSCs | / | Screening: Agilent Human miRNA Microarray Validation: TaqMan RT-qPCR |
Screening: 851 miRNAs Validation: miR-27a |
snRNU6 | ↓ miR-27a in OP vs. control | Mef2c (predicted for miR-27a using TargetScan and PicTar database and validated by in vitro studies) | / | The mean age of OP and healthy women is significantly different; no information about the stem-loop arm of miRNA origin; no ROC analysis |
| (Bedene et al., 2016) | 74 PM women (age 55-65 years): 57 controls and 17 OP based on femoral neck/lumbar spine/total hip T-score ≤–2.5 SD | Cell-free plasma | Blood samples collected in EDTA tubes, centrifuged at 2800 rpm and 4°C for 10 min, then further centrifuged at 9600g and 4°C for 15 min. Plasma samples stored at -80°C | SYBR Green RT-qPCR | miR-7d-5p, miR-7e-5p, miR-30 d-5p, miR-30e-5p, miR-126-3p, miR-148a-3p, miR-199a-3p, miR-423-5p, and miR-574-5p | Combination of let-7a-5p and miR-16-5p as identified by Normfinder | ↑ miR-148a-3p in OP vs. control | / | / | Small sample size of the control group; no evaluation of the target genes; no ROC analysis |
| (Chen et al., 2016) | 36 PM women: 19 HC, 7 osteopenic, 10 OP | Cell-free serum | Serum obtained by centrifuging blood samples in two steps: for 10 min at 2000g and 4°C and for 20 min at 12000g and 4°C. Serum stored at -80°C | SYBR RT-qPCR | miR-30a-5p, miR-30e-5p, miR-425-5p, miR-142-3p, miR-191a-3p, miR-215, miR-29b-3p, miR-30b-5p, miR-26a-5p, miR-345-5p, miR-361-5p, miR-185-5p, and miR-103-3p | NormFinder and GeNorm identified miR-25-3p as the most stable reference gene in mice models of OP | ↓ miR-30b-5p in both osteopenic and OP vs. HC↓ miR-103-3p, miR-328-3p, and miR-142-3p OP vs. HC | / | 0.793 (miR-30b-5p) for both OP and osteopenia vs. HC0.793-70.6-79.0 (miR-30b-5p), 0.800-80-72.2 (miR-103-3p), 0.789-70-79.0 (miR-142-3p) and 0.874-80-100 (miR-328-3p) for OP vs. HC | Different number of subjects recruited in the 3 groups; the reference gene for humans was identified in mice models; no evaluation of the target genes |
| (Jimenez-Ortega et al., 2017) | PM Mexican-Mestizo women: 6 with normal (control group) and 6 with OP hip BMD (age 63-85 years) | PBMCs | Histopaque-1077 kit used for obtaining PCMCs by density gradients. CD14+ obtained by density gradient centrifugation for 30 min at 400g and RT and magnetic bead isolation. negative isolation kit EasySep Human Monocyte Enrichment used for naive monocyte isolation | Screening: Affymetrix GeneChip Human U133 Plus 2.0 Array Validation: TaqMan RT-qPCR |
Screening: 2.578 miRNAs tested Validation: miR-1270, miR-548x-3p, and miR-8084 |
Screening: quantile normalization Validation: RNU44 |
↑miR-1270 in OP group vs. control group | IRF8 (identified for miR-1270 using PITA v5.0, microRNA.org, miRWalk v2.0, miRDB, and TargetScan Human v7.0 database and validated in study) | / | Small sample size; no ROC analysis |
| (De-La-Cruz-Montoya et al., 2018) | PM Mexican-Mestizo women: 7 with normal (control group) and 7 with OP hip BMD (age 63-85 years) | Human PBMCs | Blood collected in CPT tubes and PBMCs obtained. CD14+ cells enriched by negative selection (EasySep kit) | Screening: Illumia NextSeq 500 Validation: TaqMan RT-qPCR |
Validation: miR-708-5p, miR-3161, miR-939-3p, and miR-4422 | Validation: RNU44 and RNU48 | ↑miR-708-5p in osteoporosis group vs. control group | AKT1, AKT2, FKBP5, PARP1, and MP2K3 (identified for miR-708-5p using miRTarBase and MiRNet and validated in study) | / | Small sample size; no ROC analysis |
| (Ramirez-Salazar et al., 2018) | Discovery cohort: 40 PM Mexican-Mestizo women: 20 with normal (controls) and 20 with OP hip BMD (age 63-85 years)Validation cohort: 22 normal, 26 OP, 28 osteopenia, 21 with hip fracture BMD | Cell-free serum | Serum obtained within 1h of collection and stored at -80°C | Discovery stage: TaqMan Array Human MicroRNA A+B Cards Set v3.0 Validation stage: TaqMan RT-qPCR |
Screening: 754 miRNAs tested Validation: miR-23b-3p miR-140-3p, and miR-885-5p |
snRNU6 | ↑ miR-23b-3p and miR-140-3p in OP, osteopenia and bone fracture group vs. control↑ miR-885-5p in osteopenia vs. control | AKT1, AKT2, AKT3, IL6R, BMP2, GSK3B, FOXO3, PRKACB, WNT5B, and RUNX2 (identified for miR-23b-3p and miR-140-3p using miRWalk v3 database) | 0.84 (miR-140-3p) for osteopenia, 0.96 (miR-140-3p) for OP, and 0.92 (miR-140-3p) for fracture vs. HC0.73 (miR-23b-3p) for osteopenia, 0.69 (miR-23b-3p) for OP, and 0.88 (miR-23b-3p) for fracture vs. HC0.69 (miR-885-5p) for osteopenia vs. HC | No validation of the identified target genes |
AKT1, AKT serine/threonine kinase 1; AKT2, AKT serine/threonine kinase 2; AKT3, AKT serine/threonine kinase 3; BMD, bone mineral density; BM-MCSs, bone marrow mesenchymal stem cells; BMP2, bone morphogenic protein 2; CBL, casitas B-lineage lymphoma proto oncogene; CD226, cluster of differentiation 226; CXCL11, chemokine (C-X-C motif) ligand 11; CXCR3, chemokine (C-X-C motif) receptor 3; FKBP5, FK506 binding protein 5; FOXO3, forkhead box O3; FZD3, frizzled-3; GSK3B, glycogen synthase kinase 3 beta; HC, healthy controls; IGF1, insulin-like growth factor 1; IL6R, interleukin 6 receptor; IRF8, interferon regulatory factor-8; Mef2c, myocyte enhancer factor 2 c; MP2K3, mitogen-activated protein kinase kinase 3; OP, osteoporosis; OSX, osterix; PAG1, phosphoprotein associated with glycosphingolipid microdomains 1; PARP1, poly(ADP-ribose) polymerase 1; PBMCs, peripheral blood mononuclear cells; PM, postmenopausal; PRKACB, protein kinase cAMP-activated catalytic subunit beta; RANK, receptor activator of nuclear factor κ B; RANKL, receptor activator of nuclear factor k B ligand; RT, room temperature; RT-qPCR, real-time quantitative polymerase chain reaction; RUNX2, runt-related transcription factor 2; SLC39A1, solute carrier family (zinc transporter), member 1; SPRY1, protein sprouty homolog 1; TOB2, transducer of ERBB2, 2; WNT5B, Wnt family member 5B.