miRNA Primer: Gene Expression

This is the second piece in our miRNA Primer series, providing an introduction to microRNA (miRNA) and its use in Inti Labs’ tests. If you haven’t read the first post, we recommend starting there.

Gene Expression

At the end of the human genome project in 2003, researchers reached some surprising conclusions. It was discovered that only about 20,000 genes (approximately 2% of DNA) code for proteins. The function of the remaining 98% of DNA was mostly unknown, but it is now understood that much of this “dark genome” is responsible for generating non-coding regulatory material, which controls the protein-making process (1).

As more is revealed about which phenotypes are coded into which parts of our DNA, we begin to understand how different types of cells in different parts of the body control the proliferation of new gene products. While it is understood that 99.99% of DNA is virtually identical in every human being, we still experience a dramatic variance from person to person.

The 0.01% difference accounts for approximately 3 million base pairs which might differ, leaving ample room for variance in phenotype. Furthermore, the field of epigenetics describes environmental factors such as diet, stress, exercise, sleep and disease that affect gene expression and are also responsible for the variance in phenotype.

microRNA and Phenotype

MicroRNAs (miRNAs) have a significant role as regulators of gene expression, and their observed dysregulation has been associated with the establishment and progression of various diseases and disorders (2). Having tight control of the production of new cells is vital to a functioning body.

In disease as well as in traits of physical appearance such as eye and hair color, miRNA profiles are being catalogued. For example, it is feasible to study a group of some ten plus individual miRNA, including miR-25 and miR-137, and conclude whether someone is dark-haired or blonde, has vitiligo, melanoma, or suffering from a sunburn. Each of these states creates a detectable snapshot of cell activity, where up-regulation or down-regulation of certain miRNA turns protein-coding genes on or off. Where there is cell activity there is a miRNA signature that gives us information about what is happening in the body (3). 

miRNA Profiles and Endometrial Receptivity

Assisted Reproductive Therapy (ART) research is ongoing, with the goal of understanding how to increase the chances of successful IVF. Approximately 30% of the embryos transferred to the uterus lead to a successful pregnancy; therefore, having as much information about each patient is vital to increasing the likelihood of success (4).

Numerous studies have outlined miRNA profiles related to endometrium development, specifically concerning differing miRNA profiles in successful versus unsuccessful IVF (5-7). There is a significantly different expression of certain miRNAs in the window of implantation (WOI) for receptive vs non-receptive endometria. Members of the let-7, miR-200, miR-30 families, and the miR-17-92 clusters are more commonly found to be associated with endometrial receptivity (ER).

One study performed miRNA microarray on the samples from repeated implantation failure (RIF) and control groups. Differentially expressed miRNAs were found and analyzed for their role in the establishment of endometrial receptivity. Hsa-miR-145, hsa-miR-374, hsa-miR-4668-5p, hsa-miR-429, and hsa-miR-5088 may be relevant to the low endometrial receptivity of RIF patients (4).

Endometrial Health and the Menstrual Cycle

Recall that during the proliferative phase of menstruation, estrogen increases, and the uterine wall thickens. During the normal secretory phase, progesterone increases, and an egg is deposited into the uterus. The endometrium continues to develop until the abrupt decrease in hormones triggers menstruation. Timing of the biopsy for an endometrial receptivity test (ERT) and its results (pre-receptive, receptive, or post-receptive) is key to informing providers when to initiate embryo transfer.

Altmae et.al. have compared the dynamic genomic expression profiles of the endometrium from the proliferative phase, to the WOI in fertile women. Has-miR-30b, has-miR-30d, and has-miR-494 were considered to play important roles in regulating endometrial receptivity. Compared with the pre-receptive endometrium, hsa-miR-30b and hsa-miR-30d were found to be significantly upregulated and hsa-miR-494 was found to be downregulated in receptive endometria (11).

Another study highlighted a group of 12 miRNAs (miR29B, mi R29C, miR30B, miR30D, miR31, miR193A-3P, miR203, miR204, miR200C, miR210, miR582-5P, and miR345) that were significantly upregulated during the mid-secretory phase. This group is predicted to target many cell cycle genes, suppressing cell proliferation during the secretory phase (8).

It has also been suggested that different sections or cell types of the endometrium can demonstrate separate miRNA profiles. In mice two studies have suggested that implantation sites versus inter-implantation sites have differing miRNA profiles (9,10). Another human study showed endometrial gene expression differs between fertile women and women with repeated implantation failure during IVF or recurrent miscarriages, specifically at the site of implantation (12).

Increasing IVF success using MIRA™

Timing and location are proving vital to successful IVF and utilizing this breadth of research is at the heart of how MIRA operates. It is advantageous to test for miRNA expression to determine the stage of development of the endometrium at the location of intended embryo implantation. [1] [2] MIRA employs a novel group of miRNA which has an accuracy of over 90% in identifying a displaced window of implantation as the cause for repeated implantation failure (13). The high accuracy and low failure rate of miRNA detection with MIRA will mean less stress and more reliability for patients and providers, and allow providers to employ a specific strategy for each individual patient.

References

1 The ENCODE Project Consortium. An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57–74 (2012). https://doi.org/10.1038/nature11247

2 Barbarotto, E., Schmittgen, T. D., & Calin, G. A. (2008). MicroRNAs and cancer: Profile, profile, profile. International Journal of Cancer, 122(5), 969– 977.

3 Hushcha Y, Blo I, Oton-Gonzalez L, Mauro GD, Martini F, Tognon M, Mattei M. microRNAs in the Regulation of Melanogenesis. Int J Mol Sci. 2021 Jun 5;22(11):6104. doi: 10.3390/ijms22116104. PMID: 34198907; PMCID: PMC8201055.

4 Stern JE, Cedars MI, Jain T, Klein NA, Beaird CM, Grainger DA, et al. Assisted reproductive technology practice patterns and the impact of embryo transfer guidelines in the United States. Fertil Steril 2007;88:275-82. doi: https://doi.org/10.1016/j.fertnstert.2006.09.016.

5 Chen CH, Lu F, Yang WJ, Yang PE, Chen WM, Kang ST, Huang YS, Kao YC, Feng CT, Chang PC, Wang T, Hsieh CA, Lin YC, Jen Huang JY, Wang LH. A novel platform for discovery of differentially expressed microRNAs in patients with repeated implantation failure. Fertil Steril. 2021 Apr 3:S0015-0282(21)00088-1. doi:10.1016/j.fertnstert.2021.01.055. Epub ahead of print. PMID: 33823989.

6 Reza, A. M., Choi, Y., Han, S. G., Song, H., Park, C., Hong, K., & Kim, J. (2018). Roles of microRNAs in mammalian reproduction: From the commitment of germ cells to peri-implantation embryos. Biological Reviews, 94(2), 415-438. doi:10.1111/brv.12459

7 Liang, J., Wang, S., & Wang, Z. (2017). Role of microRNAs in embryo implantation. Reproductive Biology and Endocrinology, 15(1). doi:10.1186/s12958-017-0309-7

8 Satu Kuokkanen, Bo Chen, Laureen Ojalvo, Lumie Benard, Nanette Santoro, Jeffrey W. Pollard, Genomic Profiling of MicroRNAs and Messenger RNAs Reveals Hormonal Regulation in MicroRNA Expression in Human Endometrium, Biology of Reproduction, Volume 82, Issue 4, 1 April 2010, Pages 791–801, https://doi.org/10.1095/biolreprod.109.081059

9 Geng, Y., He, J., Ding, Y. et al. The Differential Expression of MicroRNAs Between Implantation Sites and Interimplantation Sites in Early Pregnancy in Mice and Their Potential Functions. Reprod. Sci. 21, 1296–1306 (2014). https://doi.org/10.1177/1933719114525273

10 Hu SJ, Ren G, Liu JL, Zhao ZA, Yu YS, Su RW, Ma XH, Ni H, Lei W, Yang ZM: MicroRNA expression and regulation in mouse uterus during embryo implantation. J Biol Chem 2008; 283:23473–23484.

11 Altmäe, S., Martinez-Conejero, J. A., Esteban, F. J., Ruiz-Alonso, M., Stavreus-Evers A., Horcajadas, J. A., & Salumets, A. (2012). MicroRNAs miR-30b, miR-30d, and miR-494 Regulate Human Endometrial Receptivity. Reproductive Sciences, 20(3), 308-317. doi:10.1177/1933719112453507

12 Zhao Y, Zacur H, Cheadle C, Ning N, Fan J and Vlahos NF: Effect of luteal-phase support on endometrial microRNA expression following controlled ovarian stimulation. Reprod Biol Endocrinol 10: 72, 2012.

13 Chen CH, Lu F, Yang WJ, Yang PE, Chen WM, Kang ST, Huang YS, Kao YC, Feng CT, Chang PC, Wang T, Hsieh CA, Lin YC, Jen Huang JY, Wang LH. A novel platform for discovery of differentially expressed microRNAs in patients with repeated implantation failure. Fertil Steril. 2021 Apr 3:S0015-0282(21)00088-1. doi:10.1016/j.fertnstert.2021.01.055. Epub ahead of print. PMID: 33823989.