Table 1.
Summary of the studies in female reproductive cells and tissues: (A) ovary and extracellular vesicles (exosomes and microvesicles), and (B) oocytes.
| Reference | Location | Age (years) | Population Studied | Study Design | Outcome and Aim | Analysis and Normalization Methods | Principal Conclusion | Quality Score |
|---|---|---|---|---|---|---|---|---|
| (A) Ovary and extracellular vesicles (exosomes and microvesicles) | ||||||||
| [9] (Liang et al., 2007) | USA | ND | 1 ovary | Descriptive | To identify miRNA expression patterns in human tissues (including ovary). | Analysis: qRT-PCR. Normalization: Average of miR-30e, miR-92, miR-92N, and miR-423. | A total of 216 miRNAs are expressed in human ovaries. | 4/9 |
| [22] (Sang et al., 2013) | China | Desciptive: ND. Cases: 29.09 ± 0.70 Control: 30.83 ± 0.90 | Follicular fluid and microvesicles. Descriptive: 20 women undergoing ICSI. Cases: 22 PCOS patients Control: 22. | Descriptive | To identify cell-free miRNAs in human follicular fluid and to investigate the function of these miRNAs in vitro and any roles they play in PCOS. | Analysis: qRT-PCR. Normalization: U6 RNA. | Identification of hsa-miRNA-19b, 24, and 222 in human follicular fluid and another 538 known miRNAs. The expression levels of hsa-miRNA-132 and 320 were significantly lower in controls than in PCOS patients. | 4/9 |
| [23] (Santonocito et al., 2014) | Italy | <35 | FF exosomes from 15 women undergoing ICSI | Descriptive | To characterize well-represented miRNAs in human FF and exosomes. | Analysis: qRT-PCR. Normalization: Average of miR-25, miR-28-3p, and miR-145. | 37 miRNAs upregulated in human FF compared with in plasma (15 miRNAs were found to be upregulated in total FF compared with in plasma; 10 miRNAs were carried by exosomes, while five were not). 22 miRNAs were present exclusively in exosomes. | 6/9 |
| [24] (Tong et al., 2014) | China | 29.16 ± 2.7 | 5 women (CRC and COC) | Descriptive | To determine the miRNA expression profiles, via NGS technology, of CRCs and COCs. | Analysis: RNA seq and qRT-PCR validation. Normalization: U6. | A total of 785 and 799 annotated miRNAs were identified in CRCs and COCs respectively. Different expression patterns in CRCs and COCs were detected in 72 annotated miRNAs. | 6/9 |
| [16] (Cai et al., 2017) | China | 29 ± 3.5 | GC. Cases: 25 women with PCOS. Controls 20 women wihtout PCOS. | Case-control | To investigate the effect of miR-145 on cell proliferation and the underlying mechanism of miR-145 in isolated human GCs from the aspirated follicular fluid in women with PCOS. | Analysis: qRT-PCR. Normalization: GAPDH or U6. | miR-145 is downregulated in human GCs from PCOS when compared with control subjects. Insulin receptor substrate 1 (IRS1) gene is a direct target of miR-145. | 6/12 |
| [25] (Chen et al., 2017) | China | NOR: 30.57 ± 2.69. DOR: 33.74 ± 3.25 | COC. Cases: 10 women with DOR. Controls: 10 women with NOR | Case-control | To comprehensively characterize miRNA expression profiles in cumulus cells of DOR patients. | Analysis: RNA seq and qRT-PCR validation. Normalization: U6. | 79 annotated miRNAs, and 5 novel miRNAs were identified differentially expressed between DOR and NOR. mTOR pathway and meiosis-associated biological processes were enriched in the DE-miRNAs | 5/12 |
| [27] (Eisenberg et al., 2017) | Israel | <35. PCOS: 26.9 ± 4.3 MIF: 26.8 ± 4.7 | GC. Cases: 15 normally ovulating with pure male infertility factor, and 18 with PCOS. Controls: 7 normally ovulating. | Case-control | To study the role of micro-RNA (miRNA)-200b and miRNA-429 in human ovulation and to measure their expression levels in ovulatory and anovulatory patients. | Analysis: qRT-PCR. Normalization: U6. | Very low expression levels were detected for the two miRNAs in granulosa cells, with hsa-miRNA-200b expression relatively higher than miRNA-429. No significant expression difference of any of these miRNAs was identified in the GLCs of the two groups analyzed. | 5/12 |
| [28] (Fu et al., 2018) | China | <40. Control: 29.89 ± 3.16. Cases: 30.94 ± 3.82 | 91 individual FF. Cases: 53 no blastocysts. Controls: 38 viable blastocysts | Case-control | To screen miRNAs in human follicular fluid and to explore the relationship between miRNA expression and blastocyst formation | Analysis: qRT-PCR screening and qRT-PCR validation. Normalization: U6. | 13 miRNAs (hsa-miR-185, 202, 22, 224, 29c, 30a-3p, 378, 382, 424, 432, 497, 520c-3p and 663B) were up-regulated, and three miRNAs (hsa-miR-1274a, 139-5p and 150) were down-regulated in cases compared with controls. In the validation, the expression level of miR-663B was found to be significantly higher in cases than in controls. | 5/12 |
| [29] (Karakaya et al., 2015) | Turkey | 4 groups: <35, 35–37, 38–40, >40. | COC in 189 women undergoing IVF–ICSI. Cases: 24 poor-responders. Controls: 32 non-poor responders. | Case-control | To analyze the association of miRNA expression with the number of oocytes retrieved, in women undergoing in vitro fertilization (IVF). | Analysis: Microarray screening and qRT-PCR validation. Normalization: RNU43. | MicroRNA microarray analysis showed up-regulation of 16 miRNAs and down-regulation of 88 miRNAs in poor responders. qRT-PCR confirmed that miR-21-5p expression was significantly up-regulated in poor responders, whereas miR-21-3p expression was significantly lower. | 5/12 |
| [30] (Luo et al., 2019) | China | Poor ovarian response (37 ± 3.16), PCOS (27 ± 3.26), normal patients (29 ± 3.22). | GC. Cases: poor ovarian response (n = 7), PCOS (n = 20). Controls: normal patients (n = 18). | Case-control | To determine the microRNA (miRNA) profiles in GCs from the FF of patients with varying levels of ovarian reserve function. | Analysis: RNA seq. | Identified 20 conserved and 3 novel miRNAs that were upregulated in the poor ovarian response group and 30 conserved miRNAs and 1 novel miRNA that were upregulated in the PCOS group. | 7/12 |
| [17] (Roth et al., 2014) | USA | PCOS: 33.1 ± 4.4. Oocyte donors: 27.1 ± 3.6 | FF. Cases: PCOS (n = 10). Controls: Oocyte donors (n = 10). | Case-control | To determine if miRNAs are differentially expressed in the FF of women with PCOS compared to fertile oocyte donors | Analysis: qRT-PCR screening and qRT-PCR validation. Normalization: U6snRNA. | 29 miRNAs are differentially expressed between PCOS and OD samples. In the validation step only five of these upregulated miRNAs (hsa-miR-9, 18b, 32, 34c, and 135a) displayed a significant increase in expression in the PCOS group compared to OD controls. | 7/12 |
| [18] (Scalici et al., 2016) | France | 19–43 | FF. Cases: 30 women with PCOS. Controls: 91 women with normal ovarian reserve. | Case-control | To investigate the expression profiles of five circulating miRNAs (let-7b, miR-29a, miR-30a, miR-140 and miR-320a) in human FF | Analysis: qRT-PCR. Normalization: miR-16. | Hsa-miR-30a was significantly up-regulated, while miR-140 and let-7b were significantly down-regulated in FF pools from patients with PCOS (n = 30) compared to women with normal ovarian reserve. | 6/12 |
| [19] (Shi et al., 2015) | China | Non-PCOS (28.5 ±3.6). PCOS (28.3 ± 3.3). | COC. Cases: PCOS (n = 24). Controls: Non-PCOS (n = 24). | Case-control | To compare the expression of miRNAs in COC from PCOS and non-PCOS women. | Analysis: qRT-PCR screening and qRT-PCR validation. Normalization: ND. | Hsa-miR-483-5p and 486-5p are significantly decreased in COC of PCOS patients compared with non-PCOS. Four predicted genes, SOCS3, SRF, PTEN and FOXO1, were significantly increased in PCOS COC, and IGF2 was significantly decreased in PCOS COC. | 7/12 |
| [20] (Wang et al., 2018) | China | Non-PCOS (30.00 ± 0.74). PCOS (28.67 ± 0.675) | GC. Cases: 25 PCOS. Controls: 17 non-PCOS healthy women. | Case-control | To characterize the function of microRNA-27a-3p (miR-27a-3p) in PCOS | Analysis: qRT-PCR. Normalization: U6 RNA. | 21 miRNAs were upregulated and 38 were downregulated in PCOS GCs. Hsa-miR-27a-3p was significantly increased in both excised GCs and the ovaries of patients with PCOS compared with the controls. | 6/12 |
| [21] (Xu et al., 2015) | China | Non-PCOS (29.43 ± 3.92). PCOS (28.76 ± 3.51). | GC. Cases: 21 women with PCOS. Controls: 20 women without PCOS. | Case-control | To describe the altered miRNA expression profiles and miRNA targeted signaling pathways in PCOS. | Analysis: RNAseq screening and qRT-PCR validation. Normalization: U6. | A total of 59 known miRNA were identified that are differentially expressed in PCOS cumulus granulosa cells, including 21 miRNAs increased and 38 miRNAs decreased. Notch signaling, regulation of hormone, and energy metabolism of the DE-miRNA target genes. | 6/12 |
| [26] (Diez-Fraile et al., 2014) | Belgium | Younger group (29.3–30.3), older group (38.7–42.4) | 16 women. Young age: < 31 years (n = 8); advanced age: >38 years (n = 8) | Prospective | To report the presence of miRNAs in human FF and identify a set of miRNAs that are differentially expressed in older women compared to younger women. | Analysis: qRT-PCR screening and qRT-PCR validation. Normalization: hsa-miR-483-5p. | Hsa-miR-21-5p was present at significantly higher levels in FF from young women, whereas hsa-miR-99b-3p, 134 and 190b were present at significantly higher levels in the FF from older women. | 8/14 |
| [32] (Moreno et al., 2015) | Spain | Younger group (32.93 ± 2.19), older group (38.20 ± 0.86) | Stage 1: Younger group: <35 (n = 15), older group: >37 (n = 15). Stage 2: MII vs. GV (n = 12) and MI vs. MII (n = 9), each patient acted as her own control. | Prospective | To determine whether there is any difference in the FF miRNA profiles from IVF patients according to their age and oocyte maturation stage. | Analysis: qRT-PCR screening and qRT-PCR validation. Normalization: RNU6B. | hsa-miR-424, which is present in higher proportions in FF from patients with advanced age. When we compared the FF from MII versus GV oocytes, they found 13 differentially expressed miRNAs. When we compared FF from MII versus MI, they found seven differentially expressed miRNAs in MII. | 7/14 |
| [31] (Martinez et al., 2018) | USA | 28.9–33.7 | 126 women. EV-miRNAs in FF from oocytes with normal fertilization (n = 93) and from oocytes that failed to fertilize (n = 33). | Cross-sectional | To assess whether EV miRNAs from FF can serve as biomarkers for fertilization status and day 3 embryo quality. | Analysis: qRT-PCR. Normalization: global mean. | 12 EV-miRNAs were differentially expressed between the normal and failed to fertilize groups. Hsa-miR-92a and miR-130b, were over-expressed in FF samples from oocytes that failed to fertilize compared to those that were normally fertilized. Hsa-miR-888 was over-expressed and miR-214 and miR-454 were underexpressed in samples that resulted in impaired day-3 embryo quality compared to top-quality day-3 embryos. | 8/14 |
| (B) Oocytes | ||||||||
| [33] (Barragán et al., 2017) | Spain | 1. Young women with high AFC (age 21 ± 1 years and 24 ± 3 follicles) and low AFC (age 24 ± 2 years and 8 ± 2 follicles); 2. Old women with high AFC (age 32 ± 2 years and 29 ± 7 follicles) and low AFC (age 34 ± 1 years and 7 ± 1 follicles). | 36 in vivo matured MII oocytes from 30 healthy women recruited for oocyte donation | Retrospective | To identify the coding and noncoding transcriptional profiles of in vivo matured MII human oocytes and evaluate their changes in relation to age and, independently, ovarian reserve. | Analysis: Microarray and qRT-PCR validation. Normalization: Actin B, ubiquitin C and DNA methyltransferase-1. | A set of five miRNAs (ENSG00000221162, hsa-miR-220b, ENSG00000239174, miR-4262 and 1260a) were increased in old group. | 7/14 |
| [35] (Battaglia et al., 2016) | Italy | Young women (28–35 years) and old women (38–40 years) | Six MII oocytes from young women and six from older women | Retrospective | To identify human oocyte miRNAs and demonstrate thatconditions altering oocyte quality, such as reproductive aging | Analysis: qRT-PCR screening and qRT-PCR validation. Normalization: RNU6B. | Twelve miRNAs are differentially expressed in women of advanced reproductive age | 7/14 |
| [34] (Xu et al., 2011) | China | 1. 30.65 ± 3.87 GV, and 2. 32.15 ± 4.63 MII oocytes | 392 oocytes at GV stage and 43 oocytes at MI stage during 251 ICSI cycles | Retrospective | To identify differentially expressed miRNAs and expression patterns of specific miRNAs during meiosis in human oocytes. | Analysis: Microarray and qRT-PCR validation. Normalization: U6 RNA | Compared with GV oocytes, MII oocytes exhibited up-regulation of 4 miRNAs (hsa-miR-193a-5p, 297, 625 and 602), and down-regulation of 11 miRNAs (hsa-miR-888*, 212, 662, 299-5p, 339-5p, 20a, 486-5p, 141*, 768-5p, 376a and 15a). | 7/14 |
Abbreviations: AFC, antral follicle count; COC, cumulus-oocyte complex; CRC, corona radiata cells; DE-miRNAs, differentially expressed miRNAs; DOR, diminished ovarian reserve; EV, extracellular vesicles; FF, follicular fluid; GC, granulosa cells; GLC, granulosa lutein cells; GV, germinal vesicle; ICSI, intracytoplasmic sperm injection; IVF, in-vitro fertilization; MI, metaphase I; MII, metaphase II; ND, no data; NGS, next generation sequencing; NOR, normal ovarian reserve; OD, oocyte donors; PCOS, polycystic ovary syndrome; qRT-PCR, quantitative real-time PCR; RNAseq, RNA sequencing.