Innovations in the treatment of endometrial diseases: Role of human umbilical cord mesenchymal stem cells and their exosomes

Abstract

Endometrial disorders, notably intrauterine adhesions and thin endometrium, are major causes of female infertility and are often poorly managed by conventional therapies. Human umbilical cord-derived mesenchymal stem cells and their exosomes have emerged as promising regenerative alternatives, leveraging their immunomodulatory, anti-fibrotic, and pro-angiogenic properties. This review provides a critical synthesis of evidence from 125 studies (2007-2025), focusing not only on therapeutic mechanisms and clinical progress but also on the specific hurdles that impede clinical translation. Their mechanistic actions converge on key repair processes, including immune modulation, inhibition of transforming growth factor-β/Smad-mediated fibrosis, and promotion of vascularization. Early phase clinical trials have reported improved endometrial thickness and pregnancy rates, including live births in 30.8%-38.5% of severe intrauterine adhesion cases, with initially favorable safety profiles. The efficacy is further augmented by combining it with biomaterial carriers or estrogen. However, clinical translation is significantly constrained by inherent challenges, including small-scale, heterogeneous trials with short follow-ups; manufacturing complexities due to donor variability and a critical lack of potency assays linking cell phenotype to in vivo efficacy; and unresolved safety concerns, particularly regarding long-term and intergenerational risks, ectopic tissue formation, and context-dependent pro-tumorigenic potential in conditions such as endometriosis and cancer. Therefore, future advancements hinge on addressing these interconnected fronts: Conducting mechanism-informed, large-scale clinical trials, establishing manufacturing standards integrated with biologically relevant potency assays, and resolving context-dependent efficacy and safety paradoxes.

Key Words: Human umbilical cord mesenchymal stem cells; Exosomes; Endometrial regeneration; Endometrial diseases; Treatment

Core Tip: This review offers a critical analysis of human umbilical cord mesenchymal stem cell and exosome therapies for endometrial diseases. It synthesizes compelling mechanistic and clinical evidence while rigorously identifying the key translational roadblocks - standardization, clinical validation, and safety - that must be overcome to move from promising research to widespread clinical practice.

INTRODUCTION

Endometrial disorders, including intrauterine adhesions (IUA), thin endometrium, endometrial cancer, and endometriosis, pose a significant threat to women’s reproductive health. These conditions arise from diverse etiologies, such as uterine surgery, infection, and endocrine dysfunction[1]. Their prevalence underscores the scale of the problem: IUA is found in 0.8%-45.5% of infertile women, depending on the subpopulation[2], endometriosis affects an estimated 10% (190 million) of reproductive-aged females globally[3], and endometrial cancer constitutes roughly 5% of cases in women under 40 years of age, half of whom are nulliparous[4]. Clinically, these diseases are frequently associated with chronic pelvic pain and irregular menstruation, which can precipitate complications such as salpingitis and pelvic inflammatory disease, thereby increasing the risk of miscarriage[5]. Current management relies primarily on pharmacotherapy and surgery[6], but both approaches have inherent limitations: Prolonged drug use may incur toxic side effects, whereas surgery can inflict new trauma and carry recurrence risks. Furthermore, treatment-induced endometrial fibrosis is a major contributor to infertility, as a scarred endometrium disrupts embryo recognition, adhesion, and invasion[7]. Consequently, mitigating post-interventional fibrosis and restoring endometrial receptivity represent urgent unmet clinical needs in this field.

To address these challenges, regenerative medicine strategies centered on mesenchymal stem cells (MSCs) have gained considerable interest. First identified by Friedenstein et al[8], MSCs are multipotent, self-renewing stromal cells that reside in diverse tissues, including the bone marrow, adipose tissue, dental pulp, umbilical cord, and placenta[9,10]. Their therapeutic power stems from their robust proliferative and differentiation capacities, coupled with the pivotal paracrine signaling roles of MSC-derived exosomes[11]. Among various sources, umbilical cord-derived MSCs are particularly favored for their high proliferative potential, low immunogenicity, and proangiogenic capability[12,13]. Human umbilical cord MSCs (hUC-MSCs) can differentiate in vitro into endometrial epithelial and stromal cells[14], demonstrate significant reparative effects on the damaged endometrium, and mitigate fibrosis by downregulating the transforming growth factor-β (TGF-β)/Smad pathway[15]. Furthermore, encapsulating hUC-MSC-derived exosomes in hydrogels as sustained-release carriers significantly enhances the treatment efficacy of IUA[16]. This review critically evaluates these emerging hUC-MSC- and exosome–based therapeutic strategies for endometrial regeneration. We argue that the field has reached an inflection point, where the paramount task is no longer to prove efficacy but to confront the specific complexities that impede translation. Therefore, our analysis is structured around three defining challenges: Context dependency of therapeutic outcomes, standardization paradox in product manufacturing, and strategic ambiguities in clinical application.

LITERATURE SEARCH STRATEGY AND SELECTION CRITERIA

We conducted a systematic literature search to ensure a comprehensive and unbiased synthesis. We queried the PubMed and CNKI databases for publications from January 2007 to February 2025. The search strategy combined MeSH terms and free-text keywords in English and Chinese, spanning three core concepts: (1) HUC-MSCs (e.g., “human umbilical cord mesenchymal stem cells”, “hUC-MSCs”); (2) Exosomes (e.g., “exosomes”, “extracellular vesicles”); and (3) Target endometrial diseases (e.g., “intrauterine adhesions”, “Asherman syndrome”, “thin endometrium”, “endometriosis”, “endometrial cancer”). Boolean operators (“AND”, “OR”) were used to link these terms. After duplicate removal, two authors independently screened the retrieved records by title and abstract, followed by a full-text evaluation against predefined criteria. The inclusion criteria were as follows: (1) Original research (in vitro, in vivo, or clinical) or high-quality reviews focusing on the therapeutic application or mechanism of hUC-MSCs or their exosomes in IUA, thin endometrium, endometriosis, or endometrial cancer; (2) Publication in peer-reviewed English or Chinese journals; and (3) Studies that included characterization of hUC-MSCs (e.g., surface marker analysis and differentiation assays). The exclusion criteria were as follows: (1) Non-peer-reviewed literature (e.g., editorials, conference abstracts alone); (2) Articles with unavailable full text; (3) Duplicate publications; and (4) Studies not primarily focused on hUC-MSCs or their exosomes (e.g., those employing MSCs from other sources). This selection process yielded 125 articles that formed the evidence base for this review. The key findings of these studies are synthesized and analyzed in Tables 1 and 2.

Table 1 Animal studies of human umbilical cord-derived mesenchymal stem cells and their derived exosomes in endometrial diseases.

Species	Disease	Treatment	Administration	Key findings (↑ increase; ↓ decrease)	Ref.
Rabbit	IUA	hUC-MSCs	Uterine wall injection of 1 × 106 cells	↑ Gland number; ↑ ER & Ki-67 expression; ↓ fibrosis	[22]
Rat	IUA	hUC-MSCs	Intraperitoneal injection of 2 × 106 cells	↓ TGF-β1/Smad3 expression; ↑ CD31 (angiogenesis)	[24]
Mouse	IUA	Cytokine-preconditioned hUC-MSCs	Uterine wall injection of 2 × 104 cells	↓ TNF-α, IL-6, CD301+ macrophages, α-SMA, collagen I; ↑ IL-10; JAK-STAT pathway modulated	[25]
Mouse	IUA	hUC-MSCs	Tail vein injection of 1 × 106 cells	↓ Fibrosis; ↓ collagen deposition	[30]
Mouse	IUA	hUC-MSCs	Tail vein injection of 2 × 105 cells	Improved endometrial architecture; ↑ visible glands	[38]
Mouse	IUA	hUC-MSC-derived exosomes	Tail vein injection (equivalent to 2 × 106 cells)	↓ IL-1β & IL-6; ↑ M2 macrophage polarization; JAK-STAT pathway activated	[32]
Monkey	IUA	hUC-MSCs/HA-GEL	Intrauterine injection of 1-2 × 107 cells in 200 μL HA-GEL	↑ Endometrial thickness; ↑ gland number; ↓ fibrotic area	[80]
Rat	IUA	hUC-MSCs/HA-GEL	Intrauterine injection of 1 × 1010 cells with 300 μL HA-GEL	↑ Number of pregnancy sacs; ↓ inflammatory & fibrotic factors	[79]
Rat	IUA	hUC-MSCs/HA-GEL	Intrauterine injection of 3 × 105 cells in 1 mL HA-GEL	Uterine cavity expansion; ↑ endometrial thickness & glandular cavity; ↑ gland number	[92]
Mouse	IUA	hUC-MSCs/scaffold	Intrauterine injection of 1 × 106 cells with silk fibroin-submucosa scaffold	↑ Gland number; ↓ fibrotic area	[106]
Rat	IUA	hUC-MSC-exosomes/scaffold	Intrauterine injection of 3 × 1010 exosomes with collagen scaffold	Enhanced endometrial regeneration; ↑ fertility rate	[81]
Rat	IUA	hUC-MSC-exosomes + estrogen	Intraperitoneal injection of exosomes + oral estrogen	↓ Fibrosis; uterine cavity expansion; ↑ blood vessels & glands; ↓ TGF-β; ↑ VEGF	[82]
Rat	Thin endometrium	hUC-MSCs	Tail vein injection of 1 × 107 cells	Preserved endometrial structure; ↑ embryo implantation rate; ↓ fibrosis & inflammation; ↑ cell proliferation & vascularization	[26]
Rat	Thin endometrium	hUC-MSCs	Intrauterine injection of 1 × 107 cells	↑ Endometrial thickness, area & gland number; ↑ cell proliferation & angiogenesis	[29]
Rat	Thin endometrium	hUC-MSCs/hydrogel	Tail vein injection of 5 × 106 cells with pluronic F-127 hydrogel	↑ Endometrial thickness & gland number; ↑ neovascularization	[97]
Rat	Thin endometrium	hUC-MSCs/matrigel microspheres	Intrauterine injection of cell-laden microspheres	↑ Endometrial thickness; ↑ fertility rate (25% to 75%)	[98]
Rat	IUA	hUC-MSCs	Sublingual vein injection of 5 × 106 cells	↓ Endometrial fibrosis; ↑ gland quantity	[100]
Mouse	Thin endometrium	hUC-MSCs/hydrogel	Intrauterine injection of 1 × 106 cells in GelMA/SerMA hydrogel	↑ Endometrial thickness; ↓ fibrosis; improved endometrial repair	[101]
Mouse	Thin endometrium	hUC-MSCs/hydrogel	Intrauterine injection of 5 × 105 cells in alginate-rCo III hydrogel	Restored endometrial function; induced mesenchymal-epithelial transition; ↑ endometrial regeneration & fertility	[102]
Rat	Endometriosis	hUC-MSCs	Tail vein injection of 1 × 105 cells (3 doses)	↓ Nerve fiber density in lesions; ↓ pain symptoms	[43]

IUA: Intrauterine adhesion; hUC-MSCs: Human umbilical cord-derived mesenchymal stem cells; ER: Estrogen receptor; TGF-β: Transforming growth factor β; CD31: Cluster of differentiation 31; TNF-α: Tumor necrosis factor alpha; IL: Interleukin; α-SMA: Alpha-smooth muscle actin; JAK: Janus kinase; STAT: Signal transducer and activator of transcription; M2: Macrophage type 2; HA-GEL: Hyaluronic acid gel; VEGF: Vascular endothelial growth factor; GelMA: Gelatin methacryloyl; SerMA: Sericin methacryloyl; Alginate-rCo III: Alginate-recombinant collagen III.

Table 2 Human trials of human umbilical cord-derived mesenchymal stem cells in endometrial diseases.

Patients (n)	Disease	Treatment & administration	Study design & follow-up	Primary outcomes	Limitations	Ref.
10	IUA (n = 6), cesarean scar diverticulum (n = 4)	hUC-MSCs, 2 × 107 cells, intrauterine infusion (× 2 cycles)	Phase I, open-label, 6 months	(1) Improved menstrual volume (4/10); (2) Increased endometrial thickness (6/10); and (3) Increased uterine cavity volume (6/10)	(1) Very small sample size (n = 10); (2) No control group; (3) Short follow-up (6 months); (4) Open-label, non-randomized; and (5) Heterogeneous patient population & endpoints	[64]
26	Recurrent, moderate-severe IUA	hUC-MSCs/collagen scaffold, 1 × 107 cells on scaffold, intrauterine	Phase I, open-label, 30 months	(1) Improved menstrual parameters; (2) Reduced IUA score; (3) Increased endometrial thickness & blood flow; (4) Pregnancy: 10/26 (38.5%); and (5) Live birth: 8/26 (30.8%)	(1) Small sample size; (2) Single-center, open-label; (3) No placebo control; and (4) Follow-up ended at delivery (no long-term offspring data)	[65]
18	Thin endometrium (Asherman’s)	hUC-MSCs/collagen scaffold, 1 × 107 cells on scaffold, intrauterine (× 2 cycles)	Pilot study, open-label. Until the pregnancy outcome	(1) Increased endometrial thickness, microvascular density, Ki67 index, estrogen receptor α, progesterone receptor; pregnancy (5/18); (2) Delivering healthy babies (3/18); (3) Pregnancy: 5/18 (27.8%); and (4) Live birth: 3/18 (16.7%)	(1) Small sample size; (2) Non-randomized, open-label; (3) Short follow-up (pregnancy-defined); and (4) Included only the scaffold-treated arm in this analysis	[66]
25	Refractory thin endometrium	hUC-MSCs/collagen scaffold (hUC-MSC/CS) vs saline/CS (control). Intrauterine implantation post-hysteroscopy	Single-center, randomized, double-blind, controlled trial. Follow-up for cLBR	(1) cLBR: 3/11 (27.3%) vs 1/13 (7.7%) (P = 0.30); (2) Clinical pregnancy: 5/11 (45.5%) vs 1/13 (7.7%) (P = 0.06); and (3) Trend toward improved outcomes; mechanism linked to cytokine pathways	(1) Small sample size per group (n = 11, 13); (2) Single-center design; (3) Primary outcome (cLBR) did not reach statistical significance; and (4) Follow-up focused on pregnancy (1-year safety reported)	[67]

IUA: Intrauterine adhesion; hUC-MSC: Human umbilical cord-derived mesenchymal stem cell; CS: Collagen scaffold; cLBR: Cumulative live-birth rate.

PREPARATION AND ADMINISTRATION OF HUC-MSCS AND THEIR EXOSOMES

Preparation of hUC-MSCs

HUC-MSCs are typically isolated from Wharton’s jelly of full-term umbilical cord. Briefly, the tissue undergoes enzymatic digestion and is then cultured in standard medium to facilitate the adherence and expansion of fibroblast-like MSCs. HUC-MSC identity is confirmed by the positive expression of characteristic surface markers (e.g., CD73, CD90, and CD105), the absence of hematopoietic markers (e.g., CD34, CD45, and human leukocyte antigen-DR), and the verification of multipotent differentiation potential into adipogenic and osteogenic lineages[17]. For therapeutic applications, hUC-MSCs at early passages (e.g., passages 3-5) are commonly used.

Preparation of hUC-MSC-derived exosomes

Exosomes are nanoscale extracellular vesicles secreted by hUC-MSCs that mediate key paracrine effects in the recipient cells. The most widely used isolation method is the differential centrifugation of conditioned cell-culture medium. The harvested vesicles are subsequently characterized to confirm their identity. Standard characterization comprises: (1) Analysis of particle size distribution and concentration via nanoparticle tracking analysis; (2) Morphological assessment using transmission electron microscopy; and (3) Immunoblotting to detect exosome-enriched surface marker proteins, such as tumor susceptibility gene 101, CD63, and CD81[18,19].

THERAPEUTIC MECHANISMS OF HUC-MSCS AND THEIR EXOSOMES IN ENDOMETRIAL DISEASES

The following sections detail the mechanisms by which hUC-MSCs and their exosomes mediate endometrial repair. However, a critical synthesis of the literature reveals that their therapeutic profiles are not monolithic. This review is structured to analyze not only efficacy but also two critical, intertwined dimensions that define the translational landscape: (1) Contingency: How the disease-specific microenvironment (e.g., inflammation in IUA vs estrogen-rich milieu in endometriosis) critically shapes and can even reverse therapeutic outcomes; and (2) Challenge: How do these contingent responses translate into concrete hurdles for clinical development, such as safety concerns in oncological contexts or unpredictable efficacy? This framework aims to move beyond a catalog of mechanisms towards a nuanced understanding of the factors that will ultimately determine clinical success or failure.

Integrated core mechanisms: A foundation for endometrial repair

The therapeutic efficacy of hUC-MSCs and their exosomes in various endometrial diseases stems from a convergent set of core biological actions that target the common pathological hallmarks of tissue injury: Inflammation, fibrosis, impaired regeneration, and dysfunctional vasculature (Figure 1).

Figure 1 Schematic figure illustrating the mechanisms of human umbilical cord mesenchymal stem cells and their exosomes in the treatment of endometrial diseases. HUC-MSCs: Human umbilical cord mesenchymal stem cells; HUC-MSCs-exo: Human umbilical cord mesenchymal stem cells-derived exosomes; ZEB1: Zinc finger E-box binding homeobox 1; IL-1β: Interleukin 1β; TNF-α: Tumor necrosis factor alpha; IL-6: Interleukin 6; TGF-β1: Transforming growth factor β1; Smad3: Mothers against decapentaplegic homolog 3; α-SMA: Alpha smooth muscle actin; VEGF: Vascular endothelial growth factor; PTEN: Phosphatase and tensin homolog; PI3K: Phosphoinositide 3-kinase; AKT: Protein kinase B.

Immunomodulation and anti-inflammation: The modulation of the local immune microenvironment is a cornerstone of their action. hUC-MSCs secrete interleukin-6 (IL-6), which can inhibit CD4+ T cell proliferation and promote the generation of anti-inflammatory regulatory T cells, thereby shifting the immune balance from a pro-inflammatory (e.g., T helper cell 17) to an anti-inflammatory state[20,21]. Critically, they suppress the central nuclear factor kappa B signaling pathway, leading to a downstream reduction in key pro-inflammatory cytokines, such as IL-1β, tumor necrosis factor-α, and IL-6, which are fundamental for resolving chronic endometrial inflammation[22,23].

Antifibrotic action: Fibrosis is a terminal consequence of chronic inflammation. HUC-MSCs directly counteract this process by downregulating the pivotal TGF-β1/Smad3 signaling axis, a master driver of extracellular matrix deposition and myofibroblast activation (marked by alpha smooth muscle actin)[24]. Furthermore, they secrete factors such as complement 1 inhibitor, which can modulate macrophage polarization, specifically by preventing the induction of pro-fibrotic CD301+ macrophages via the Janus kinase-signal transducer and activator of transcription pathway, thereby further ameliorating tissue scarring[25].

Promotion of angiogenesis and cell survival: Restoration of an adequate blood supply is essential for regeneration. HUC-MSCs and their exosomes upregulate pro-angiogenic factors, such as vascular endothelial growth factor (VEGF) and CD31, facilitating the formation of new microvasculature[24,26]. Concurrently, they enhance the survival and proliferation of resident endometrial cells (epithelial and stromal) by activating pro-survival pathways, such as phosphoinositide 3-kinase/protein kinase B and Wnt/β-catenin pathways, while inhibiting apoptosis[27-29].

Direct cell engagement and microenvironment reprogramming: Beyond paracrine secretion, hUC-MSCs can directly interact with and modulate key effector cells. For instance, they can mitigate macrophage ferroptosis, a proinflammatory cell death, by regulating the CD36 pathway, thereby preserving the reparative functions of macrophages[30]. Their exosomes carry specific microRNAs (miRNAs, e.g., miR-21-5p) that directly promote stromal cell proliferation and proteins like galectin-1 that drive macrophages towards the reparative M2 phenotype[31-33].

Mechanism-focused application in IUA

In IUA, where the primary pathology is physical adhesion formation and fibrosis following severe injury, hUC-MSCs exert their effects via direct tissue reconstitution and targeted anti-fibrosis effects. A unique aspect of IUA is their demonstrated ability to transdifferentiate into endometrial epithelial cells, stromal fibroblasts, and vascular endothelial cells in situ, directly replacing the lost cellular components[34]. This process is complemented by strong activation of the Wnt/β-catenin pathway, which drives the proliferation of surviving epithelial cells[27]. To address the pathological features of fibrosis, anti-TGF-β/Smad action is particularly salient[24]. Genetic modification of hUC-MSCs to overexpress hepatocyte growth factor represents a potent enhancement strategy, as hepatocyte growth factor synergistically promotes angiogenesis, inhibits apoptosis, and facilitates mesenchymal-to-epithelial transition, which is crucial for regenerating a functional lining[35-38]. Exosomes offer a cell-free alternative, wherein their cargo (e.g., miR-21-5p) specifically targets pathways that promote stromal cell proliferation and suppress fibrosis[31,39].

Mechanism-focused application in thin endometrium

For a thin endometrium, characterized by an insufficient proliferative response and poor vascularization, the therapeutic emphasis shifts to the potent stimulation of proliferation and angiogenesis. The phosphoinositide 3-kinase/protein kinase B signaling pathway has been identified as a central mediator. Its activation by hUC-MSCs critically promotes cell cycle progression in endometrial cells and enhances the expression and activity of VEGF, driving both endometrial thickening and neovascularization[29,40,41]. The upregulation of characteristic stromal (vimentin) and epithelial (cytokeratin-19) markers confirms the successful stimulation of both endometrial compartments[42]. Thus, the core proliferative and angiogenic mechanisms are hyperactivated to overcome the developmental threshold required for successful embryo implantation.

Context-dependent and complex mechanisms in endometriosis

Endometriosis presents a more complex and paradoxical scenario in which the effects of hUC-MSCs can be highly context-dependent, illustrating the nuances of cell therapy. The beneficial effects observed in some studies align with the core mechanisms: Intravenous administration has been shown to reduce lesion nerve fiber density (alleviating pain), upregulate tumor suppressor phosphatase and tensin homolog to induce ectopic cell apoptosis, and suppress pro-inflammatory cytokines[43-45]. Similarly, exosomes can inhibit ectopic stromal cell proliferation and key estrogen-synthesizing enzymes[46].

However, a critical discussion must highlight this potential dual role. The unique inflammatory and estrogen-rich microenvironment of this disease can reprogram MSCs. Notably, intraperitoneal injection of hUC-MSCs into a primate model exacerbated lesions[47]. This may be due to the aberrant activation of otherwise beneficial pathways: TGF-β, abundantly secreted by MSCs, which is pro-fibrotic in IUA, may potently induce regulatory T cells in the peritoneal cavity, potentially fostering immune escape and survival of ectopic implants[48-50]. Furthermore, factors within endometriotic lesions may prime MSCs or their exosomes to inadvertently promote epithelial-mesenchymal transition or angiogenesis, thereby supporting lesion establishment[51-53]. This stark contrast underscores a fundamental translational challenge: The therapeutic action of hUC-MSCs is context-dependent, not intrinsic. This necessitates meticulous route optimization, stringent preclinical safety evaluation in disease-specific models, and a cautious clinical approach tailored to the complex endometriotic microenvironment.

Tumor-microenvironment interactions and dual roles in endometrial cancer

The application of MSCs in endometrial cancer uses their tumor-homing property, but is fraught with the most significant “dual-effect” concerns. Homing, driven by chemokine axes such as stromal cell-derived factor-1/CXC chemokine receptor 4, is advantageous for the targeted delivery of therapeutic exosomal miRNAs (e.g., miR-503-3p), which suppress cancer cell proliferation and migration[54-57]. Furthermore, hUC-MSC-derived exosomes can modulate the tumor microenvironment and inhibit oncogenic pathways, such as Wnt/β-catenin, thereby suppressing malignant behavior[58]. This underscores the potential of exosomes as a targeted and low-toxicity therapeutic strategy.

Conversely, a substantial body of evidence warns of its potential tumor-promoting effect. Adipose-derived MSCs, particularly those from omental fat, can be recruited to tumors and accelerate endometrial cancer growth and vascularization[59]. These MSCs may enhance cancer cell proliferation, mobility, and heterogeneity through direct interactions or fusion[60,61]. This pro-tumor activity appears to be highly dependent on the MSC tissue source and donor characteristics, revealing a critical layer of complexity in MSC therapy. Therefore, while the core immunosuppressive and tropic mechanisms of MSCs offer a delivery platform, their inherent capacity to modify the tumor microenvironment can be exploited to promote malignancy. This constitutes a major safety concern that must be addressed before clinical translation. This mandates not only extreme caution and rigorous donor-source selection but also dedicated research to understand the conditions under which MSCs switch from tumor-suppressive to tumor-promoting phenotypes. Extensive long-term preclinical safety studies focused on oncogenic risk are non-negotiable prerequisites for any future clinical consideration in oncology.

APPLICATION SAFETY OF HUC-MSCS AND THEIR EXOSOMES

Quality control and preclinical safety assessment

The clinical translation of hUC-MSCs and their exosomes as a specific subset necessitates rigorous safety assessments, beginning at the manufacturing stage. Standardized quality control protocols are essential for ensuring product safety and consistency. This includes guaranteeing sterility (freedom from bacterial, fungal, and viral contamination) and confirming cellular identity. Authentic hUC-MSCs exhibit characteristic spindle-shaped morphology, express high levels of typical surface markers, and lack hematopoietic markers. Similarly, exosomes are validated by the presence of specific surface proteins[62]. Furthermore, the functional potency of hUC-MSCs must be verified through their demonstrated multipotent differentiation capacity into adipocytes, osteoblasts, and chondrocytes under defined induction conditions[63]. These foundational characterizations provide critical assurance regarding the purity, identity, and biological functionality of therapeutic products before their in vivo administration.

Clinical safety profile from early-phase trials

Early phase clinical trials have provided initial evidence supporting the relative safety of intrauterine hUC-MSC transplantation for endometrial diseases, with no reports of serious adverse events directly attributable to these cells.

IUA: In a phase I trial by Huang et al[64], intrauterine infusion of clinical-grade hUC-MSCs in ten patients (six with IUA and four with cesarean scar diverticulum) was well-tolerated. No adverse symptoms were observed. Hematological parameters (white blood cell count, hemoglobin, and platelets), kidney function, ovarian function, tumor biomarkers, and immune indicators remained within the normal range during the 3- and 6-month follow-up periods[64]. Similarly, Cao et al[65] reported no postoperative complications (e.g., fever or abnormal vaginal discharge) in 26 patients with recurrent IUA who received hUC-MSCs cultured on collagen scaffolds. Endometrial biopsies at three months post-treatment showed no signs of inflammation, and no adverse events were recorded during the 30-month follow-up[65].

Thin endometrium: Zhang et al[66] observed no postoperative complications (e.g., fever) or tumor occurrence in patients receiving hUC-MSCs on collagen scaffolds. Postoperative hematological and liver/kidney function parameters were normal, and pathological examination of the uterine tissue revealed no inflammatory reaction[66]. In another trial by Hou et al[67], transient mild symptoms, such as dizziness, nausea, and lower abdominal pain, were reported in some patients post-transplantation, but resolved within hours. Two of the eleven participants developed urticaria, which subsided spontaneously within days without treatment. Importantly, coagulation, liver and kidney functions, serum electrolyte levels, thyroid function, and tumor markers remained within normal limits following treatment[67]. Collectively, these clinical findings from phase I trials indicate that the intrauterine administration of hUC-MSCs, whether alone or combined with scaffolds, is associated with a favorable short-term safety profile, with no severe treatment-related complications or systemic toxicity.

Ethical and regulatory considerations for future translation

Despite the encouraging initial safety data, the widespread clinical application of hUC-MSCs faces significant ethical and regulatory challenges that must be addressed. Although the use of umbilical cord tissue circumvents the ethical controversies associated with embryonic stem cells, rigorous ethical standards, including informed consent and donor privacy, must govern its procurement and use. A major translational hurdle is the lack of harmonized global regulatory frameworks, which can lead to inconsistencies in product quality and clinical oversight.

The inherent differentiation potential of MSCs, which is central to their therapeutic mechanism, also presents a theoretical long-term risk, such as unintended differentiation or tumor formation in specific microenvironments. To mitigate these risks and ensure consistent therapeutic efficacy, the production of clinical-grade hUC-MSCs must adhere to stringent good manufacturing practice (GMP) standards. This requires the establishment of independent committees for quality control, ethics, scientific review, and biosafety. A comprehensive battery of quality tests, including sterility, mycoplasma, viral and endotoxin detection, cell viability, identity (morphology and surface markers), genetic stability (karyotyping), and assessments of tumorigenic potential, must be implemented. Furthermore, a system of batch release and verification testing with meticulous long-term record-keeping is essential. Only through such a robust multilayered system of quality assurance and regulatory vigilance can the therapeutic promise of hUC-MSCs be safely and reliably translated into clinical practice.

CLINICAL TRANSLATION OF HUC-MSCS AND THEIR EXOSOMES IN ENDOMETRIAL DISEASES

The evolving landscape of stem cell therapy for endometrial regeneration

The clinical exploration of stem cell therapy for endometrial repair and regeneration has advanced significantly since its initial report. Following early case studies using autologous bone marrow-derived stem cells[68], subsequent studies have employed various cell sources, including peripheral blood CD133+ cells[69], adipose-derived stromal vascular fractions[70], and menstrual blood-derived stem cells[71]. These studies, which utilized different delivery methods (intrauterine instillation, local injection, scaffold-based delivery), collectively demonstrated the feasibility and potential of stem cell-based approaches to improve endometrial thickness, restore menstrual function, and achieve pregnancy in patients with IUA and a thin endometrium[69-75]. However, the heterogeneity in cell sources, delivery methods, and outcome measures across these early studies also reflects the field’s initial, empirical stage - a stage that must now evolve to address the standardization and complexity challenges highlighted in this review.

Preclinical foundation: Evidence from animal models

The robust therapeutic potential of hUC-MSCs and their exosomes for endometrial regeneration is firmly supported by extensive preclinical research. As systematically summarized in Table 1, studies across diverse animal models (e.g., rodents, rabbits, and non-human primates) have consistently demonstrated their efficacy in ameliorating the defining pathologies of IUA, thin endometrium, and endometriosis. These investigations have not only validated core therapeutic mechanisms, such as the attenuation of fibrosis (via TGF-β/Smad inhibition), promotion of angiogenesis (via VEGF upregulation), and immunomodulation, but have also served as critical proof-of-concept platforms for optimizing delivery strategies. Notably, combination approaches with biomaterial carriers, such as hyaluronic acid gels (HA-GEL) or collagen scaffolds, which were later translated into clinical trials, were first shown to significantly enhance cell retention and therapeutic outcomes in these animal studies (Table 1).

Clinical evidence for hUC-MSCs

Among the various stem cell sources, hUC-MSCs offer distinct advantages for allogeneic therapy, including noninvasive procurement, low immunogenicity, robust proliferative capacity, and relative ease of standardization under GMP conditions[65,76,77]. Despite their widespread investigation in other fields (e.g., phase III trials for knee osteoarthritis), their application in gynecology remains limited to early-phase clinical trials (phase I/II). Key clinical studies on hUC-MSCs for endometrial diseases are summarized in Table 2.

Although the evidence is preliminary, it remains promising. In a landmark phase I trial, Cao et al[65] reported that transplantation of a hUC-MSC/collagen scaffold complex in 26 patients with severe IUA significantly improved endometrial parameters and resulted in 10 pregnancies within a 30-month follow-up, demonstrating both safety and potential efficacy. Subsequent studies have focused on optimizing delivery. Zhang et al[66] and Hou et al[67] further supported the scaffold-based strategy, showing significant increases in endometrial thickness and promising pregnancy rates in patients with a thin endometrium or refractory IUA. Notably, Huang et al[64] conducted a phase I trial using intrauterine perfusion of clinical-grade hUC-MSCs without a scaffold and observed improvements in menstrual volume and endometrial metrics, albeit in a small cohort, where statistical significance was limited. Collectively, these trials suggest that hUC-MSC-based therapy, particularly when combined with a scaffold for enhanced retention, can promote endometrial regeneration and improve reproductive outcomes.

Critical analysis of study designs, endpoints, and limitations

A critical synthesis of the current clinical data reveals several important insights and underscores the need for more robust evidence.

Study design and control groups: Most existing trials, including those summarized in Table 2, were open-label, single-arm, or small-scale pilot studies. The general lack of randomized, placebo-controlled, and blinded designs is a major limitation, introducing potential bias and making it difficult to definitively attribute outcomes to the intervention vs natural history or concomitant procedures (e.g., hysteroscopic adhesiolysis).

Heterogeneity in delivery strategies: Current research strongly indicates a preference for combining hUC-MSCs with biomaterial scaffolds (e.g., collagen) over simple cell suspension. This strategy is rationally grounded in improving local cell retention, prolonging paracrine action, and providing a protective physical barrier[65-67]. However, direct comparative studies evaluating different scaffold materials (e.g., collagen and synthetic polymers) for their degradation kinetics, biocompatibility, and ultimate therapeutic efficacy are conspicuously absent, leaving the optimal delivery platform undefined.

Timing and frequency of administration: Beyond dosage and delivery method, the optimal timing and frequency of hUC-MSC administration are undefined and contribute to the outcome variability. The therapeutic window may be limited because cell activity diminishes over time[69,72]. Consequently, treatment schedules vary across studies, with some employing single transplants and others employing multiple courses[69,72,78]. Decisions on repeat administration are often based on individual patient responses, such as endometrial thickness after the first procedure[78]. This lack of a standardized protocol further complicates the comparison of efficacy data across clinical trials.

Endpoints and sample size: Primary endpoints varied across studies, including morphological improvements (endometrial thickness and uterine cavity volume), functional recovery (restoration of menstruation), and reproductive outcomes (clinical pregnancy and live birth rates). Although live birth is the most clinically relevant endpoint, most trials are underpowered to detect significant differences in this outcome because of their small sample sizes (typically n < 30). This limitation affects the statistical strength and generalizability of the findings.

Safety and long-term follow-up: Although short-term safety profiles from early phase trials appear favorable[64,65], systematic long-term follow-up data are lacking. Most studies conclude follow-up at the time of live birth, with no extended monitoring of maternal health or the long-term development and health of offspring. This represents a significant gap in the safety assessment required for full clinical translation of this approach.

IUA: From empirical delivery to strategic synthesis

The clinical management of IUA with hUC-MSCs has evolved significantly, moving beyond simple cell infusion towards rationally designed combination strategies aimed at overcoming the central challenge of therapeutic retention within the injured uterine cavity. Table 3 provides a comparative synthesis of these major interventional approaches, integrating pivotal clinical trial outcomes with the supporting preclinical evidence.

Table 3 Comparative analysis and critical appraisal of therapeutic strategies for intrauterine adhesions.

Strategy	Theoretical rationale/mechanism of action	Current best efficacy signal (source)	Key methodological limitations (source)	Unresolved translational challenges
hUC-MSCs alone (intrauterine perfusion)	Minimally invasive; direct delivery of viable cells and paracrine factors	Improved menstrual volume and endometrial morphology in a small, mixed cohort[64]	Very small, heterogeneous sample; no control group; IUA-specific pregnancy data not reported[64]	Low cell retention; optimal dose and timing undefined; durability of effect unproven
hUC-MSCs + collagen scaffold	Enhanced cell retention and survival; provides 3D structural support and a physical barrier against adhesion reformation	Live birth rate of 30.8% in patients with severe IUA[65]	Single-arm, open-label design; small sample size; follow-up ended at delivery (no long-term offspring data)[65]	Risk of fibrotic recurrence; necessity for long-term tumorigenicity monitoring; scaffold standardization (e.g., degradation kinetics, porosity)
hUC-MSCs + HA-GEL	Biocompatible anti-adhesion barrier; serves as a hydrogel carrier for sustained factor release; fosters an anti-inflammatory microenvironment	Significant endometrial regeneration and reduced fibrosis in a primate IUA model[79,80]	Efficacy in humans pending validation in large-scale trials with standardized reproductive endpoints	Defining the therapeutic window of gel residence and bioactivity; clinical cost-effectiveness analysis
hUC-MSC-exosomes + scaffold	Cell-free approach mitigates risks of live-cell transplantation; scaffold enables controlled release; potent immunomodulatory cargo	Restoration of fertility and induction of M2 macrophage polarization in a rat IUA model[81]	Awaiting clinical translation; challenges in exosome standardization and scalable GMP manufacturing	Potential immunogenicity of allogeneic exosomes; long-term biodistribution and safety profile; development of validated potency assays
hUC-MSC-exosomes + estrogen	Synergistic action: Estrogen priming optimizes endometrial receptivity, while exosomes deliver targeted regenerative signals	Superior anti-fibrotic and pro-angiogenic effects compared to monotherapy in a rat IUA model[82]	No clinical data available; long-term safety of combined therapy unknown	Mechanistic understanding of synergy; criteria for patient stratification; optimization of the combined dosing regimen

hUC-MSCs: Human umbilical cord-derived mesenchymal stem cells; IUA: Intrauterine adhesions; 3D: Three-dimensional; HA-GEL: Hyaluronic acid gel; GMP: Good manufacturing practice.

The intrauterine perfusion of hUC-MSCs alone, as investigated by Huang et al[64], demonstrated initial safety and feasibility, with improvements in menstrual volume (4/10 patients) and endometrial thickness (6/10 patients) reported in a small, mixed cohort over a 6-month follow-up. However, the absence of reported IUA-specific pregnancy data in this trial hints at the potential limitations of cell suspension delivery, likely related to suboptimal local retention. In contrast, the transplantation of hUC-MSCs seeded on a collagen scaffold (Cao et al[65]) yielded the most robust reproductive outcomes to date, with a live birth rate of 30.8% (8/26 patients) during a 30-month follow-up in severe IUA, strongly supporting the scaffold’s role in enhancing cell engraftment and paracrine activity. The strategy of combining hUC-MSCs with HA-GEL presents a distinct approach that uses the gel’s dual function as a biocompatible anti-adhesion barrier and supportive hydrogel carrier[16]. Preclinical studies in primate and rat models have shown that this combination can significantly increase endometrial thickness and glandular number while reducing fibrosis and inflammatory markers[79,80]. These promising results position HA-GEL as a viable alternative to solid scaffolds, although its therapeutic efficacy and optimal parameters for clinical use (e.g., gel residence time and cell-gel ratio) await confirmation in human trials with standardized reproductive endpoints. Additionally, preclinical studies have further illuminated the path forward, showing high promise for cell-free strategies employing hUC-MSC-derived exosomes. These exosomes, particularly when delivered via a scaffold for sustained release or combined with estrogen to prime the regenerative microenvironment, have demonstrated potent efficacy in restoring endometrial architecture and fertility in animal models[81,82], although they await clinical validation.

This comparative synthesis highlights the paramount barriers that must be addressed before widespread clinical adoption: (1) Risk of fibrotic recurrence and long-term durability: It remains unknown whether endometrial regeneration is functionally durable beyond a single pregnancy cycle, particularly within a microenvironment that may retain a pro-fibrotic predisposition; (2) Long-term and intergenerational safety: The follow-up periods in current trials are inadequate to assess theoretical long-term risks, including ectopic differentiation or tumorigenicity. The establishment of mandated long-term safety registries for treated women and their offspring is a non-negotiable future requirement; and (3) Immune rejection potential: Although acute rejection is rarely reported, the long-term fate of allogeneic cells and the immunological consequences of potential repeat administrations in a chronically inflamed cavity are not systematically monitored and warrant dedicated studies.

Synthesis and implications for future clinical translation

Current clinical evidence positions hUC-MSCs and their exosomes as a highly promising, yet investigational, therapy for endometrial regeneration. The encouraging outcomes of early phase trials strongly justify their advancement to more definitive clinical studies. To translate this considerable potential into proven therapeutic benefits, future research must prioritize several key objectives. First, large-scale, multicenter, randomized, placebo-controlled trials with sufficient statistical power are urgently needed to robustly assess clinically relevant endpoints, particularly the live birth rate. Second, the field must move towards standardizing therapeutic products and administration protocols. This includes conducting rigorous head-to-head comparisons of different cell delivery combinations (e.g., various cell-scaffold constructs) to identify the most efficacious and practical strategies. Third, establishing a consensus-based core outcome set is essential to ensure consistent and comparable reporting of results across future trials. Finally, implementing mandated long-term safety registries is imperative to systematically track the health of treated women and, of paramount importance, the long-term development and well-being of their offspring over an extended period. Addressing these interconnected challenges is crucial for steering hUC-MSC therapy through a necessary paradigm shift: From demonstrating feasibility in exploratory studies to systematically addressing the complexities of standardization, validation, and safety required for reliable clinical practice.

STRATEGIES TO ENHANCE THE EFFICACY OF HUC-MSCS AND THEIR EXOSOMES

The challenges of context dependency and suboptimal delivery have spurred innovative strategies to enhance the efficacy of hUC-MSCs and their exosomes. These approaches are, in essence, direct responses to the translational complexities outlined above. They primarily focus on engineering therapeutic presentation through a combination of biomaterials or drugs, aiming to override negative microenvironmental cues, extend local bioactivity, and achieve spatiotemporal control, thereby attempting to convert unpredictable biological responses into more reliable clinical outcomes.

Combination with biomaterial carriers: Gels and hydrogels

Biomaterial gels serve as versatile carriers that can localize cells or exosomes at the injury site, prolong their retention, and provide a protective three-dimensional (3D) microenvironment.

HA-GEL for IUA: The combination of hUC-MSCs with self-crosslinking HA-GEL is a prominent strategy for IUA treatment. HA-GEL acts as a biocompatible physical barrier, minimizing postsurgical adhesion formation and maintaining a moist, anti-inflammatory environment conducive to healing[83-87]. Its viscoelastic properties allow sustained residence in the uterine cavity, supporting hUC-MSC survival and the gradual release of paracrine factors[80,88]. Preclinical studies in primate models have confirmed that the hUC-MSC/HA-GEL complex significantly increases endometrial thickness, reduces fibrosis, and restores glandular density[89,90]. The gel also modulates local inflammation by reducing the levels of TGF-β1, IL-6, and IL-8[39], while hUC-MSCs promote tissue remodeling via factors such as matrix metallopeptidase-9[91], collectively improving pregnancy outcomes[79]. Compatibility studies have confirmed that HA-GEL does not adversely affect the hUC-MSC phenotype or safety profile[92].

Advanced hydrogel systems for thin endometrium: Various thermosensitive and biomimetic hydrogels have been developed to enhance hUC-MSC delivery and efficacy in patients with a thin endometrium. Pluronic F-127, an Food and Drug Administration-approved thermosensitive hydrogel, transitions to a solid gel at body temperature, encapsulating hUC-MSCs and facilitating the localized release of angiogenic factors such as VEGF, leading to increased endometrial thickness and vascularization[93-97]. Matrigel microspheres provide an extracellular matrix-like environment, and when loaded with hUC-MSCs, have been shown to significantly promote endometrial regeneration[98]. Engineered hydrogels, such as gelatin methacrylate/sericin methacryloyl and alginate-recombinant collagen III, form highly biocompatible 3D networks that support hUC-MSC adhesion and proliferation. These systems not only increase endometrial thickness but also improve markers of angiogenesis (CD31) and epithelial proliferation (Ki-67) and reduce fibrosis, facilitating functional endometrial repair and creating a receptive environment for embryo implantation[99-102].

Combination with 3D scaffolds

Scaffolds provide a structural framework that guides tissue organization and offers sustained and localized delivery of therapeutic agents.

Cell-seeded scaffolds for IUA: Combining hUC-MSCs with porous, biodegradable scaffolds (e.g., collagen and silk fibroin composites) creates a construct that maintains uterine cavity architecture and supports stem cell engraftment[103-105]. When transplanted into IUA models, these scaffolds facilitate the delivery of hUC-MSCs, which in turn secrete factors that upregulate endometrial estrogen and progesterone receptors, promote stromal cell proliferation, and inhibit apoptosis, thereby driving structured tissue regeneration and collagen remodeling[106-108].

Exosome-laden scaffolds: To harness the benefits of exosomes while ensuring localized and sustained delivery, exosomes derived from hUC-MSCs were integrated into collagen scaffolds. This combination allows for the controlled release of exosomal cargo (e.g., miRNA-223-3p) over an extended period (up to 14 days). The exosomes modulate macrophage polarization towards the reparative M2 phenotype and promote the expression of hormonal receptors (e.g., estrogen receptors and progesterone receptors), effectively enhancing endometrial regeneration and restoring fertility in IUA models[81,109-111].

Pharmacological synergy: Combination with estrogen

Estrogen therapy, a standard treatment for endometrial hypoplasia, can be synergistically combined with hUC-MSCs or their exosomes.

With hUC-MSCs: Estrogen not only directly stimulates endometrial cell proliferation but may also enhance the survival, migration, and differentiation potential of co-administered hUC-MSCs. Combined treatment in IUA models results in superior restoration of endometrial morphology, glandular architecture, and reduction of fibrosis compared to either agent alone[112,113]. The mechanism may involve estrogen priming the endometrial microenvironment to better support stem cell activity[114].

With exosomes: The combination of exosomes and estrogen presents a cell-free therapeutic strategy. Exosomes can augment estrogen’s effects by delivering anti-inflammatory and pro-angiogenic signals directly to the endometrial milieu. This dual approach significantly downregulates pro-fibrotic and inflammatory mediators (tumor necrosis factor-α, TGF-β, IL-1, IL-6, runt-related transcription factor 2, collagen I) while upregulating VEGF, leading to more effective inhibition of fibrosis, promotion of angiogenesis, and overall functional restoration of the endometrium[82,115,116].

SUMMARY AND OUTLOOK

This review has critically appraised the journey of hUC-MSC and exosome therapies from bench to bedside. The accumulated evidence robustly supports their biological potential for endometrial regeneration. The central conclusion of this analysis, however, is that the field’s immediate challenge is a transition in focus: From demonstrating technical feasibility to systematically managing the complexities that hinder robust clinical translation. Three interconnected issues epitomize this challenge.

First, the paradox of context-dependency must be resolved. The same cellular agents that promote regeneration in a fibrotic IUA microenvironment may exhibit null or adverse effects in the inflammatory, angiogenic landscape of endometriosis or near-malignant tissue. This is not a mere curiosity but a fundamental translational constraint that demands a new generation of mechanistic studies to predict patient-specific responses and delineate safe therapeutic windows.

Second, the product “potency” must be meaningfully defined. Current manufacturing practices, focused on identity and sterility, overlook a critical gap: The disconnect between in vitro cell characteristics and in vivo repair efficacy. The development of standardized, biologically relevant potency assays - linking specific secretory profiles or exosomal cargo to functional outcomes - is the indispensable next step for achieving true GMP standardization and batch consistency.

Third, clinical translation requires strategic clarity, not just empirical optimization. Choices regarding delivery route (e.g., scaffold vs perfusion), treatment timing, and patient selection are currently ambiguous. These must evolve into data-driven, pathology-stratified strategies. Future clinical trials should be designed as comparative effectiveness studies to answer specific strategic questions, rather than as further proofs of concept.

CONCLUSION

In conclusion, realizing the promise of this regenerative platform will require a dual commitment: To deepen basic understanding of its contingent biology, and to execute targeted translational work that addresses these defined complexities. By framing the problem in this way, this review provides a distinct roadmap for transforming a promising therapeutic modality into a reliable clinical reality.