In this pilot cross-sectional study of pre-menopausal women, we found a cumulative measure of recent stressful life events and a measure of perceived social standing were significantly associated with expression of select miRNAs in the myometrium, but not in fibroid tumors. We found that one miRNA, miR-27a-5p, was both associated with our composite stressful life event measure and differentially expressed between fibroids and myometrium. These results suggest miRNAs in the myometrium may be altered in response to adverse social conditions, such as exposure to common stressors, and may help us understand the disproportionate burden of fibroids observed in Black women, who often are exposed to a greater range, duration, and intensity of psychosocial stressors [8]. Taken together, we propose that healthy myometrium may become primed to a transitionary state in fibroid development via miRNAs that are potentially dysregulated by acute stressors.
Common stressors can have short- and long-term effects on physical health, and miRNAs may provide a key regulatory mechanism by which stress may contribute to fibroid pathogenesis [9, 24, 31, 32]. In our analysis of myometrial tissue, we observed 16 miRNAs that were significantly associated with a cumulative measure of exposure to common acute stressors, wherein greater stress exposures were associated with higher levels of miRNA expression. In agreement with these results, two miRNAs were associated with one’s perceived social standing (i.e., ladder response), as an indirect measure of psychosocial stress, wherein high social standing, and potentially less stress exposure, was associated with lower miRNA expression. However, we did not observe any significant associations between psychosocial stress factors, such as a composite score or through perceived social standing, on fibroid miRNA expression. This may reflect the microenvironment of a diseased tissue (i.e., fibroids) exhibiting an already distressed state that is less sensitive to modification caused by additional stressors, which has been observed in other types of cancers as compared to healthy tissues [33].
MiRNAs play an essential role in post-transcriptional gene regulation with each miRNA having the capability to regulate multiple gene targets and genes having multiple miRNAs [34, 35]. Due to the inherent regulatory role of miRNAs, we investigated the predicted gene targets of psychosocial stress-associated miRNAs in the myometrium to allude to the functional role of miRNA via their predicted mRNA targets. The most significant GO biological process was enriched at viral processes, and recent evidence suggests reproductive tract infections and reparative mechanisms that occur in response may initiate fibroid development [36]. Additionally, of the most significant GO biological processes enriched by miRNA-mRNA targets, we found several processes related to cancer. Fibroids are benign tumors that can be characterized by a life cycle similar to cancers, which starts with aberrant cell proliferation and eventually ends in cell death, a continual process that, in addition to extracellular matrix deposition, forms a fibrotic tumor [37]. These processes are likely guided, in part, by regulatory mechanisms of miRNAs, that when dysregulated, as can occur in psychosocial stress-related exposure, may induce cancer-related pathways in the myometrium that contribute to fibroid development [38]. Corroborating these findings, we found the most significant KEGG pathways enriched by social stress-associated miRNA-mRNA targets were at pathways relevant to reproductive tumorigenesis, including endometrial cancers, which have been observed concurrently in those with fibroids [39], and p53 signaling, a fundamental signaling pathway required for fibroid growth [40], as well as pathways in the endoplasmic reticulum (ER) and proteoglycans. Indeed, fibroids exhibit a higher abundance and swelling of the ER [37] and contain different composition of proteoglycans within extracellular matrix deposits [41].
We observed that miR-27a-5p was differentially expressed between myometrium and fibroids, and significantly associated with psychosocial stress exposure, which suggests that miR-27a-5p may be dysregulated by cumulative stress exposure and potentially contribute to the transformation of healthy myometrium into fibroids. Notably, miR-27a is a novel biomarker for solid tumors and has been considered as a therapeutic target for a variety of cancers [42, 43] and may be involved in female infertility [44]. Consistent with our findings, previous studies have identified miR-27a as being differentially expressed between myometrium and adjacent fibroids [45]. Taken together, exposure to psychosocial stress may lead to long-term changes in miRNA expression within the myometrium, and dysregulation of miRNAs in myometrium may have a regulatory role for functionally relevant genes involved in pathways necessary for the transformation of healthy tissue into fibrotic states characteristic of fibroids.
Given that fibroids arise from the myometrium, we propose that miRNAs, in addition to other epigenetic modifications, may prime healthy myometrial tissue towards fibroids, a process that is affected by accumulating exposure to adverse environmental influences, including psychosocial stressors. This is especially important for minoritized populations, where exposure to adverse psychosocial conditions is quite common throughout the life-course. Indeed, previous evidence by our group demonstrated that adverse childhood experiences (ACEs) were associated with greater severity of these fibroids, and ACEs were more prevalent among Black women than non-Black women, further supporting the relevance of our current work to understanding racial disparities in fibroids [17]. We posit that as an individual acquires specific adverse “hits”, or risk factors (i.e., race, obesity, diet, age, parity, etc.), throughout the life-course, these hits can drive pathological processes within the myometrium that may initiate the development of fibroids [46], a process that may be guided by epigenetic mechanisms that can affect the myometrial stem cell pool, their self-renewal states, and their final differentiated state in myometrium tissues [47]. These epigenetic changes in myometrial tissues are thus primed to a pro-fibroid phenotype that may readily respond to adverse environmental conditions that contribute to fibroid development. Recent evidence found epigenetic mechanisms may underlie the racial differences in fibroid disparities and primed myometrial stem cells are expanded in Black women [48, 49]. Taken together, it is plausible that the disproportionate prevalence of fibroids in underserved communities, including Black women, may arise from epigenetic priming of progenitor myometrial cells and otherwise healthy myometrium to a pro-fibroid state early during development that is amenable to the effects of environmental exposures (i.e., psychosocial stressors), that, when accumulating over time, may readily transition to fibroids.
This study provides evidence for miRNAs as a potential epigenetic mechanism involved in psychosocial stress-associated fibroid development. However, as this is a pilot study, results should be interpreted with caution. In our analyses, we had a modest sample size of 20 participants and higher likelihood of spurious results. Our analyses were primarily focused on two types of psychosocial stressors, acute life events and stress associated with social standing (i.e., ladder response), although there are many types of psychosocial stressors (e.g., structural racism, ACEs, etc.), that were not included in this study. Thus, there could be residual confounding from these unmeasured confounders. Likewise, all participants were enrolled based on the criteria of seeking surgical treatment for fibroids, and our prior work has shown that the process of seeking medical treatment for fibroids can be a unique and important source of stress [50, 51]. As this study included participants who underwent surgical treatment for fibroids, these results may not be generalizable to women with asymptomatic fibroids. Because myometrium and fibroid tissue contain heterogenous cell populations, we were unable to ascertain the cell type-specific expression patterns of miRNAs; however, myometrium and fibroids predominantly contain smooth muscle cells. Our in-silico approach sought to determine mRNA targets of social stress-associated miRNAs to infer biological functionality, although individual miRNAs can target multiple genes and mRNA targets differ by cell type and tissue type [52]. The enrichment of biological functions and pathways relevant to tumorigenesis at an FDR p-value < 0.05 provide confidence in our analysis. Future studies should seek to expand on our findings in larger, longitudinal cohorts of diverse populations of women and include a diversity of psychosocial stressors in independent and composite analyses. Nonetheless, our preliminary study provides compelling evidence for the effects of psychosocial stress exposure from recent life events and social status-associated stress on miRNA signatures in otherwise healthy myometrium tissue that may contribute to fibroid development that can be used as a framework for future research.
In conclusion, our study suggests cumulative exposure to typical acute stressors may contribute to fibroid progression and pathogenesis through dysregulation of miRNAs. Our findings may have important implications for understanding and ultimately mitigating the disproportionate burden of fibroids in Black women since this work suggests the priming of myometrial tissues to pro-fibroid states may be susceptible to social and environmental exposures, including psychosocial stress.