Clarifying the role of exosomal miR-137-3p in endometrial regeneration: Mechanistic gaps and future directions

Abstract

This article comments on the study by Zhang et al, which proposed that exosomes derived from hypoxia-injured endometrial epithelial cells promote human umbilical cord mesenchymal stem cell migration and differentiation into endometrial epithelial cells via exosomal miR-137-3p. The authors demonstrated that miR-137-3p targets ubiquitin protein ligase E3C and activates signal transducer and activator of transcription 3 signaling, thereby driving epithelial lineage transition. While this study expands our understanding of exosome-mediated intercellular communication in endometrial repair, several key gaps remain. Notably, microRNA (miRNA) profiling was performed in human umbilical cord mesenchymal stem cells post-exosome treatment, not in the exosomes derived from hypoxia-injured endometrial epithelial cell themselves, leaving open whether miR-137-3p is directly transferred or indirectly induced. In addition, data on exosome characterization were unavailable, and the rationale for selecting miR-137-3p over other differentially expressed miRNAs was not well justified. Future studies should include direct exosomal miRNA content analysis, in vivo validation, and deeper mechanistic exploration of the ubiquitin protein ligase E3C-signal transducer and activator of transcription 3 ubiquitination axis to establish the clinical and biological relevance of this pathway.

Key Words: Mesenchymal stem cells; Exosomal miR-137-3p; Ubiquitin protein ligase E3C; Signal transducer and activator of transcription 3; Endometrial repair; Hypoxia

Core Tip: This commentary examines the study by Zhang et al on exosomal miR-137-3p-mediated endometrial regeneration. While the proposed mechanism involving ubiquitin protein ligase E3C inhibition and signal transducer and activator of transcription 3 activation is compelling, several issues remain unresolved, including the unclear source of miR-137-3p, lack of exosome characterization, and absence of in vivo validation. We highlight the need for direct evidence of exosomal microRNA transfer, mechanistic exploration of the ubiquitin protein ligase E3C-signal transducer and activator of transcription 3 ubiquitination axis, and functional evaluation in animal models to substantiate the therapeutic relevance of this approach.

TO THE EDITOR

We read with interest the recent article by Zhang et al[1], which reports that exosomal miR-137-3p derived from hypoxia-injured endometrial epithelial cells (EECD) enhances human umbilical cord mesenchymal stem cell (hUCMSC) migration and epithelial differentiation by targeting ubiquitin protein ligase E3C (UBE3C) and activating the signal transducer and activator of transcription 3 (STAT3) pathway. This work provides important insights into microRNA (miRNA)-mediated communication under hypoxic stress and highlights a potentially novel molecular axis (miR-137-3p/UBE3C/STAT3) in endometrial regeneration. The findings contribute to the expanding field of exosome-based regenerative strategies; however, several scientific and conceptual ambiguities merit deeper discussion.

Unclear origin of miR-137-3p

A central claim of the study is that miR-137-3p from EECD-exosomes acts as a key effector. However, the miRNA sequencing was conducted on hUCMSCs treated with exosomes, not on the exosomes themselves. Therefore, it remains unclear whether the observed upregulation of miR-137-3p originates from direct exosomal transfer or is a secondary response of hUCMSCs to other EECD-exosomal signals. This distinction is critical because if miR-137-3p is endogenously induced rather than exogenously delivered, the mechanism of action shifts from cargo-mediated signaling to cellular adaptation. The schematic in Figure 7 also suggests direct transfer of miR-137-3p from EECD-exosomes, which lacks direct experimental support and might therefore misrepresent the actual biological mechanism(s). Furthermore, while the authors identified miR-137-3p as a differentially expressed miRNA in mesenchymal stem cells (MSCs) after EECD-exosome exposure, all subsequent functional validations were performed using synthetic mimics or inhibitors of miR-137-3p in MSCs. These experiments confirmed that miR-137-3p modulated MSC migration and differentiation, but they did not establish that miR-137-3p is a major functional component of the EECD-exosomes themselves.

Stress signals can reshape the miRNA landscape of MSCs and their exosomes. For instance, exosomes derived from lipopolysaccharide-preconditioned MSCs were shown to have enhanced anti-inflammatory effects in sepsis models by enriching miR-150-5p, which targeted insulin receptor substrate 1 and suppressed the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of the rapamycin (PI3K/Akt/mTOR) pathway to promote M2 macrophage polarization and improve survival outcomes[2]. Hypoxia-preconditioned hair follicle MSC-derived exosomes exhibit enhanced anti-inflammatory effects in ulcerative colitis models by upregulating miR-214-3p, which mediates suppression of the PI3K/AKT/mTOR pathway and promotes mitophagy and mitochondrial homeostasis[3]. Exosomes derived from MSCs exposed to combined hypoxic and inflammatory stress alleviated intervertebral disc degeneration by delivering miR-221-3p to suppress DNA damage induced transcript 4 and inhibit nuclear factor kappa B activation, thus reducing the senescence of nucleus pulposus cells through epigenetic modification[4]. Similarly, a study using equine adipose-derived MSCs found that various stimuli, including interleukin-1β, hypoxia, shockwave stimulation, and senescence, significantly altered the miRNA composition of secreted extracellular vesicles, with several anti-inflammatory miRNAs (e.g.,miR-146a, miR-29b, and miR-221) being upregulated and identified as potential therapeutic candidates in osteoarthritis[5]. These studies collectively indicate that stress-conditioned exosomes do not simply act as static cargo carriers, but may trigger complex transcriptional or epigenetic responses in recipient cells. Therefore, without direct quantification of miR-137-3p in purified EECD-exosomes (e.g., via exosomal RNA isolation and reverse transcription-quantitative polymerase chain reaction), the hypothesis of exosomal transfer remains speculative.

One major regulatory mechanism underlying these stress-induced miRNA changes involves hypoxia-inducible factors (HIFs), particularly HIF-1α, which orchestrate transcriptional responses to low oxygen conditions. A subset of miRNAs, termed hypoxia-associated miRNAs, are transcriptionally regulated by HIFs and mediate key adaptive responses under hypoxic stress[6]. These include well-characterized miRNAs such as miR-210, miR-21, and miR-214, which are upregulated in MSCs and their exosomes under hypoxia and contribute to inflammation resolution, mitochondrial protection, or metabolic reprogramming[7-10]. Based on the data presented by Zhang et al[1], miR-137-3p may also represent a hypoxia-responsive miRNA, potentially expanding the repertoire of known hypoxia-associated miRNAs. However, this proposed protective role appears to contrast with previously published studies, in which miR-137-3p was upregulated in ischemia/reperfusion-injured cardiomyocytes, and its inhibition attenuated hypoxia/reoxygenation-induced apoptosis, oxidative stress, and pyroptosis in H9c2 and HL-1 cells[11,12]. This discrepancy could be attributed to several context-dependent factors. First, cell type specificity may play a critical role. The behavior of miR-137-3p in cardiomyocytes under ischemic stress may differ markedly from its function in mesenchymal or epithelial cells under hypoxia. Second, whether miR-137-3p is upregulated endogenously in stressed cells or introduced through exosomal transfer may influence its intracellular concentration, subcellular localization, and ability to access target genes. Third, the functional outcome of miR-137-3p may depend on the surrounding microenvironment, including factors such as inflammation, oxidative stress levels, and mitochondrial status, which can collectively influence whether its effect is harmful or beneficial. Despite these uncertainties, the identification of miR-137-3p as a potentially functional exosomal cargo involved in endometrial regeneration represents a novel and valuable contribution by Zhang et al[1]. These insights reinforce the concept that stress-conditioned exosomes can trigger complex adaptive responses in recipient cells, rather than acting as passive carriers. Further studies involving direct quantification of miR-137-3p in EECD-exosomes, along with loss-of-function experiments, are essential to substantiate the proposed mechanism and clarify whether the observed effects truly result from exosomal transfer or reflect an endogenous hypoxic response.

Lack of exosome characterization and uptake validation

Another concern lies in the insufficient characterization of the EECD-exosomes. No data were provided regarding their size distribution (e.g., via nanoparticle tracking analysis), morphology (e.g., transmission electron microscopy), or surface marker expression (e.g., CD9, CD63, and CD81), which are considered standard for exosome identification. This lack of definition undermines the interpretability of the proposed exosome-mediated signaling.

Furthermore, the study does not include any direct evidence showing that hUCMSCs internalized the EECD-derived exosomes. As highlighted in the MISEV2018 guidelines, uptake validation is a necessary step in functional extracellular vesicle studies, particularly when biological effects are attributed to specific RNA cargos[13]. This is particularly critical given that the central mechanism of the current study involves the transfer of miR-137-3p via exosomes to recipient MSCs.

Rationale for selecting miR-137-3p

Among the 53 differentially expressed miRNAs identified, the selection of miR-137-3p as the study’s focus is not well substantiated. The authors did not provide comparative expression levels, fold changes, or literature-based rationale supporting its prioritization over other candidates. A deeper analysis of miRNA-mRNA regulatory networks or competing endogenous RNA interactions may have yielded a more robust mechanistic foundation. Additionally, no loss-of-function experiments (e.g., miR-137-3p inhibitors) were conducted, which would be necessary to confirm its necessity in driving the observed effects.

Mechanistic insights into UBE3C and STAT3

The study establishes that miR-137-3p downregulates UBE3C and upregulates phosphorylated STAT3, suggesting a novel post-translational regulatory axis. However, the exact molecular mechanism by which UBE3C modulates STAT3 phosphorylation remains speculative. The authors hypothesize that inhibiting UBE3C may prevent STAT3 ubiquitination, thereby stabilizing or enhancing its phosphorylation status. This intriguing possibility deserves further validation via co-immunoprecipitation or ubiquitination-specific assays. Moreover, while total STAT3 protein was elevated, STAT3 mRNA remained unchanged, reinforcing the need to investigate ubiquitin-proteasome system dynamics.

In vivo validation

While Zhang et al’s study provides meaningful in vitro data, its therapeutic potential remains to be fully understood[1]. In vivo studies using animal models of thin endometrium would be a valuable next step to determine whether the observed effects can be replicated in a physiologically relevant context. Such studies would also help address the complexity of endometrial regeneration under hypoxia, which cannot be fully modeled in vitro.

Strengths and limitations

The study is commendable for identifying a novel miRNA target (UBE3C) in the context of hUCMSC differentiation and for proposing a link to STAT3 signaling. Functional assays (e.g., wound healing, transwell, Western blot) are methodologically sound and reproducible. However, the most critical limitation lies in the unverified claim that miR-137-3p is transferred via exosomes rather than endogenously expressed in recipient cells. This key mechanistic assertion is not supported by direct experimental evidence and significantly undermines the strength of the proposed pathway. In addition, the study has other omissions, including insufficient exosome characterization, lack of in vivo validation, and a speculative explanation regarding ubiquitination, all of which reduce the overall impact and translational relevance of the findings. We summarize the proposed mechanism and critical evidential gaps in Figure 1.

Figure 1 Schematic summary of the proposed mechanism by Zhang et al[1] and the associated evidence gaps highlighted in this commentary. EEC: Endometrial epithelial cell; EEC-Exo: Exosomes derived from endometrial epithelial cell; MSC: Mesenchymal stem cell; UBE3C: Ubiquitin protein ligase E3C; STAT3: Signal transducer and activator of transcription 3; MAPK: Mitogen-activated protein kinases; AKT: Protein kinase B. The figure was created in https://BioRender.com.

Future directions

To fully clarify the role of EECD-exosomal miR-137-3p in endometrial regeneration, several follow-up studies are warranted: (1) Direct quantification of miR-137-3p in EECD-exosomes (e.g., via exosomal RNA isolation and reverse transcription-quantitative polymerase chain reaction); (2) Loss-of-function studies using miR-137-3p inhibitors and CRISPR-based knockout of UBE3C in hUCMSCs; (3) Ubiquitination assays to confirm UBE3C-mediated regulation of STAT3 stability; (4) Animal models of Asherman syndrome or thin endometrium to evaluate functional repair; and (5) Exosome biodistribution and safety studies to assess translational potential.

Conclusion

Zhang et al[1] present an interesting molecular framework linking EECD-derived miR-137-3p to enhanced epithelial differentiation of hUCMSCs via UBE3C and STAT3. However, uncertainties surrounding the actual source of miR-137-3p, the definition of the exosome preparation, and the speculative nature of the ubiquitination mechanism weaken the translational value of the findings. Future studies with direct exosome cargo analysis, robust mechanistic dissection, and in vivo validation will be essential to elevate this work from a conceptual hypothesis to a clinically relevant therapeutic strategy.