A narrative review of the distribution and role of multipotent fibroblasts in penile tissue: implications for regenerative medicine and erectile dysfunction

Abstract

Background and Objective

Erectile dysfunction (ED) is a common male sexual disorder with multifactorial etiologies, including diabetes mellitus and cavernous nerve injury. Recent developments in single-cell RNA sequencing (scRNA-seq) have revealed substantial cellular heterogeneity within the corpus cavernosum, especially among fibroblast (FB) subclusters. This review aims to delineate the roles of these FB subclusters and their extracellular vesicles (EVs) in the pathogenesis and potential treatment of ED.

Methods

A comprehensive literature review was conducted focusing on studies utilizing scRNA-seq to analyze human and murine corpus cavernosum tissues under physiological and ED-associated conditions. Key findings regarding FB subcluster classification, transcriptomic landscapes, and intercellular communication were synthesized, with particular attention to the fibrotic and regenerative roles of FB-derived EVs.

Key Content and Findings

Distinct FB subclusters were identified, exhibiting profibrotic, regenerative, and signaling-related phenotypes. In ED models, including diabetes- and cavernous nerve injury-induced conditions, an increased prevalence of fibrotic FB phenotypes was observed, contributing to structural disruption and functional decline. FB-derived EVs were highlighted as pivotal mediators of paracrine signaling, influencing endothelial and smooth muscle cell function, modulating immune responses, and regulating extracellular matrix remodeling. Therapeutic administration of EVs demonstrated promising regenerative effects in preclinical ED models.

Conclusions

FB subclusters and their EVs play central roles in the pathophysiology of ED, functioning in both deleterious and reparative capacities. Selective targeting of specific FB subpopulations or leveraging the therapeutic properties of FB-derived EVs may represent innovative strategies for managing ED, particularly in treatment-resistant cases.

Introduction

Erectile dysfunction (ED) is a prevalent and increasingly widespread male sexual health disorder, characterized by the persistent inability to attain or sustain an erection adequate for satisfactory sexual activity (1). Globally, it affects more than 150 million men—a figure projected to rise due to population aging and increasing prevalence of comorbid conditions such as diabetes, cardiovascular disease, and metabolic syndrome (2). Beyond its physical manifestations, ED substantially impairs emotional well-being, interpersonal relationships, and overall quality of life (1). The etiology of ED is multifactorial, involving vascular, neural, hormonal, psychological, and structural components that disrupt the coordinated hemodynamic events required for normal erectile function (1). Current therapies largely emphasize symptomatic relief without addressing the underlying pathophysiological mechanisms (3). Phosphodiesterase type 5 inhibitors remain the first-line therapeutic option, enhancing nitric oxide-mediated vasodilation to facilitate erectile response (4). However, their effectiveness is often limited in individuals with severe endothelial dysfunction or neurogenic impairment. Second-line interventions—such as intracavernous injections and vacuum erection devices—and third-line options, including penile prostheses, offer mechanical assistance but are invasive, variably effective, and often compromise sexual spontaneity (5). These therapeutic limitations have fueled growing interest in regenerative medicine approaches aimed at restoring the structural and functional integrity of erectile tissue.

Anatomically, the penis comprises fibroblasts (FB), smooth muscle cells (SMC), endothelial cells (EC), nerve fibers, and connective tissue, with EC and FB constituting the predominant cell types within the corpus cavernosum (6,7). While EC have been extensively studied for their essential role in regulating penile blood flow and nitric oxide signaling (3), FB have historically received less attention despite their numerical abundance. Traditionally viewed as passive structural cells responsible for extracellular matrix (ECM) synthesis and tissue homeostasis (8), FB are now increasingly recognized as active modulators of the local tissue milieu through the secretion of cytokines, growth factors, and extracellular vesicles (EVs) (9). Seminal work by Guimaraes et al. (10) and Ryu et al. (11) underscored the functional relevance of FB surrounding corpus cavernosum vasculature, highlighting their active contribution to penile vascular homeostasis. These findings suggest that FB are not merely structural scaffolds but are intimately involved in biological processes such as angiogenesis, neurogenesis, immunomodulation, and fibrosis—each closely linked to erectile physiology and ED pathogenesis. The regulatory influence of FB on the penile microenvironment is facilitated by their phenotypic plasticity and their capacity for dynamic interactions with endothelial and neural elements (12). Nevertheless, the molecular mechanisms through which FB exert their effects on erectile function remain incompletely understood, and their therapeutic potential has yet to be fully realized. This review aims to provide a detailed characterization of the cellular composition of penile tissue in humans and mice, with an emphasis on FB subpopulations identified through single-cell RNA sequencing (scRNA-seq). By integrating findings from Zhao et al. (7) and Bae et al. (6), the review delineates both conserved and species-specific cellular features of the corpus cavernosum. In both species, FB comprise a major, heterogeneous cell population implicated in ECM remodeling, vascular regulation, immune response, and neural support. However, interspecies differences—such as the presence of transcriptionally active chondrocytes in murine models—suggest anatomical or developmental distinctions with potential implications for translational research. A nuanced understanding of these cellular dynamics, particularly the dual regenerative and fibrotic roles of FB subclusters, is critical to the advancement of novel therapeutic strategies for ED. We present this article in accordance with the Narrative Review reporting checklist (available at https://tau.amegroups.com/article/view/10.21037/tau-2025-318/rc).

Methods

A comprehensive literature search was conducted using the PubMed database without restrictions on publication date. The search terms included “erectile dysfunction”, “erectile function”, “fibrosis, fibroblasts”, “neurovascular regeneration”, and “penile erection”. The analysis of FB distribution in penile tissue was primarily based on studies employing single-cell RNA sequencing (scRNA-seq) of human and mouse penile tissue. The analysis of FB function was informed by all relevant retrieved articles. Selection criteria emphasized both scientific quality and relevance. The search encompassed a broad scope, including both animal and human studies. An overview of the search strategy is presented in Table 1.

Table 1. The search strategy summary.

Items	Specification
Date of search	December 10, 2024 to April 22, 2025
Databases searched	PubMed
Search terms used	Erectile dysfunction, erectile function, fibrosis, fibroblast, neurovascular regeneration, penile erection
Timeframe	Up to April 2025
Inclusion criteria	Only English
Selection process	Authors conducted the selection independently

Discussion

Cellular composition of human and mouse penile tissue

Penile tissue, particularly the corpus cavernosum, plays a pivotal role in erectile function and male sexual health. A detailed understanding of its cellular composition is essential for elucidating the mechanisms underlying both normal physiology and pathophysiological conditions such as ED. Advances in scRNA-seq technology have enabled high-resolution mapping of the cellular landscape in human and mouse penile tissue, uncovering notable cellular heterogeneity and novel subpopulations. This review synthesizes findings from two key investigations: Zhao et al. (7), who examined the human corpus cavernosum, and Bae et al. (6), who profiled mouse corpus cavernosum tissue, highlighting major similarities and species-specific differences with implications for translational research.

Zhao et al. (7) applied scRNA-seq to 64,993 cells derived from the human corpus cavernosum, including samples from individuals with normal erectile function and those with organic ED. Six primary cell types were identified: FB (approximately 46%), EC (~36%), SMCs (~6%), pericytes (~5%), immune cells (macrophages and T cells, ~7%), and Schwann cells (<1%). FB represented the largest cellular component and were classified into six transcriptomically distinct subclusters (FB1–FB6), each associated with specific spatial distribution and functional annotations. FB1 was characterized by high expression of Apolipoprotein C1 (APOC1) and Patched Domain Containing 1; FB2 by APOC1 and Protein Phosphatase 1 Regulatory Inhibitor Subunit 14A; FB3 by Peptidase Inhibitor 16 (PI16) and Flavin Containing Monooxygenase 2; FB4 by PI16 and Bone Morphogenetic Protein 7; FB5 by Cartilage Oligomeric Matrix Protein (COMP) and Keratocan; and FB6 by COMP and Microfibril Associated Protein 5. In parallel, Bae et al. (6) analyzed 12,894 single cells from the corpus cavernosum of mice under both normal and diabetic conditions. They identified 15 distinct cell clusters corresponding to major cell types, including FB (~55%), EC (~6%), chondrocytes (~36%), SMCs (~1%), pericytes (~1%), immune cells (~1%), and Schwann cells (<1%). Based on marker gene expression, FB were further subdivided into four subtypes: Egr1-high-expressing FB, C-X-C motif chemokine ligand 12 (Cxcl12)-high-expressing FB, matrix Gla protein (Mgp)-high-expressing FB (reticular FB), and a ribosomal FB subcluster, characterized by high expression of ribosomal protein genes, including Ribosomal Protein S25 (Rps25) and Ribosomal Protein S17 (Rps17), indicative of elevated biosynthetic activity. The detailed cellular composition is summarized in Table 2 (6,7,13-27).

Table 2. Cellular composition and function of human penile tissue.

Cell type	Approximate proportion	Distribution	Function	References
Fibroblasts (human)	Around 46%	Throughout corpus cavernosum and tunica albuginea	Structural support and ECM production, paracrine signaling and intercellular communication, response to injury and fibrosis, cellular plasticity	(7,13-15)
Fibroblasts (mouse)	Around 55%
Endothelial cells (human)	Around 36%	Throughout corpus cavernosum and subtunical area	Regulation of blood flow and erection, vascular homeostasis, interaction with other cell types, role in angiogenesis	(6,16-18)
Endothelial cells (mouse)	Around 6%
Chondrocyte (human)	Around 0%	Throughout corpus cavernosum	Primarily play a role in ectopic chondrogenesis, can be linked to conditions like Peyronie’s disease	(19)
Chondrocyte (mouse)	Around 36%
Smooth muscle cells (human)	Around 6%	Predominantly in trabecular regions of corpus cavernosum	Control of erection and detumescence, vasoactive signaling mediators, structural and mechanical support, interaction with other cells	(6,20,21)
Smooth muscle cells (mouse)	Around 1%
Pericytes (human)	Around 5%	Predominantly in the subtunical area and around the dorsal nerve bundle, particularly in microvessels	Vascular stability and integrity, blood flow regulation, paracrine and immunomodulatory functions, angiogenesis	(6,22-24)
Pericytes (mouse)	Around 1%
Immune cells (macrophages and T cells) (human)	Around 7%	Throughout corpus cavernosum	Immune surveillance and homeostasis, interaction with other cells, involvement in regeneration and repair	(6,7,25)
Immune cells (macrophages and T cells) (mouse)	Around 1%
Schwann cells (human)	Less than 1%	Along nerve fibers	Supports cavernous nerve function, nerve regeneration and repair, and secretion of bioactive factors	(26,27)
Schwann cells (mouse)	Less than 1%

ECM, extracellular matrix.

Comparative analysis of human and mouse corpus cavernosum tissue reveals both conserved and divergent cellular features. In both species, FBs exhibit pronounced heterogeneity and play central roles in ECM organization, immune modulation, and neurovascular support. Nevertheless, interspecies distinctions are evident. Human FB subclusters exhibit defined neuro-supportive and fibrogenic phenotypes closely tied to their spatial microenvironment, whereas mouse FBs are characterized by stress-responsive and remodeling-related activity, particularly under diabetic conditions. A notable divergence lies in the abundance and role of chondrocytes. In mice, chondrocytes constitute a substantial proportion of the corpus cavernosum cell population (~36%) and are transcriptionally active, especially in the diabetic ED model, where ECM gene expression is impaired. In contrast, chondrocytes are not prominently represented in the human dataset by Zhao et al. (7), this may reflect the presence of a penile bone/cartilage structure, which is absent in humans. Therefore, caution is warranted when extrapolating murine findings, and validation in human penile tissue is essential to confirm translational relevance.

Functional roles of FB in penile tissue

Functional roles of FB: focus on regenerative effects

FB are mesenchymal cells found in nearly all connective tissues, with a primary and well-established role in synthesizing and organizing essential ECM components, including collagen and fibronectin (28). Beyond this structural function, FB are increasingly recognized as active participants in maintaining tissue homeostasis, modulating immune responses, and facilitating repair processes following injury or disease (29). Their phenotypic plasticity enables context-specific functional adaptation to local microenvironmental cues in both physiological and pathological settings (30,31). Recent scRNA-seq studies—most notably those by Zhao et al. (7) in human tissue and Bae et al. (6) in murine models—have revealed previously unrecognized heterogeneity among penile FB. These FB subpopulations exhibit distinct genetic profiles, spatial distributions, and functional properties that contribute to erectile function and tissue structural integrity.

In human corpus cavernosum, FB3 and FB5 subpopulations are key regulators of ECM homeostasis and tissue architecture. FB3, characterized by the expression of PI16 and folate-binding protein 2, exhibits a quiescent, ECM-producing phenotype (32,33) and is localized in the interstitial region between smooth muscle and vascular structures. This subpopulation supports tissue elasticity and mechanical resilience, which are critical for the venous occlusion mechanism during erection. FB5, marked by expression of COMP and keratocan, is enriched in ECM-organizing genes (34,35) and likely contributes to the structural stability of the corpus cavernosum. Additionally, FB4 and FB6 express genes associated with neural support and are situated adjacent to nerve bundles, suggesting specialized roles in maintaining neurovascular integrity and promoting regeneration (7). These subpopulations may secrete neurotrophic factors such as nerve growth factor and brain-derived neurotrophic factor (36,37) and remodel the ECM to enhance nerve repair and vascular stabilization, processes essential for preserving erectile function and enabling recovery following neuropathic or surgical insult.

Consistent findings in murine models by Bae et al. further elucidate functionally distinct FB subtypes. FB populations expressing high levels of Cxcl12 are implicated in angiogenesis and vascular repair (38), while reticular FB appear to support ECM maintenance and general stromal organization (28,39). These populations facilitate dynamic remodeling of vascular and connective tissues under normal physiological conditions. Even in diabetic models, despite progressive pathology, FB may initially exert protective effects by releasing angiogenic signals, stabilizing ECM structure, and modulating local immune responses. Although these regenerative functions may decline with disease progression, the early involvement of FB in tissue defense and repair underscores their critical role in maintaining penile tissue homeostasis. Table 3 summarizes the distribution and regenerative roles of principal FB subpopulations identified in human and murine penile tissues. Collectively, these findings offer a more comprehensive cellular framework for understanding the functional diversity of FB and their dual roles in promoting regeneration and mediating pathological remodeling.

Table 3. Comparative summary of FB subclusters in human and mouse penile tissue.

Species	Subcluster	Key markers	Functional role	Localization/notes
Human penile tissues	FB1	APOC1+, PTCHD1+	Fibrotic, proliferative	Increased in ED tissues
	FB2	APOC1+, PPP1R14A+	Lipid metabolism	Similar to FB1
	FB3	PI16+, FMO2+	ECM-producing, quiescent	Between SMCs and vessels
	FB4	PI16+, BMP7+, KERA+, NEFL+	Neuro-supportive, tenogenic	Near nerve bundles
	FB5	COMP+, KERA+	ECM-structural organizer	Between smooth muscle bundles
	FB6	COMP+, MFAP5+	Neuron-associative stroma	Enveloping nerves
Mouse penile tissues	Egr1-high FB	Egr1+	Stress response, remodeling	Increased in diabetic ED
	Cxcl12-high FB	Cxcl12+	Angiogenic, immune signaling	May aid in vascular repair
	Reticular FB	Mgp+	ECM maintenance	General stromal structure
	Ribosomal FB	Rps25+, Rps17+	Translationally active, stress-adaptive	May reflect reactive phenotype in diabetes

ECM, extracellular matrix; ED, erectile dysfunction; FB, fibroblasts; SMCs, smooth muscle cells.

Functional roles of FB: focus on fibrosis

FB play a critical role in maintaining tissue repair and homeostasis; however, their dysregulation—particularly under conditions of chronic stress or pathological stimuli—can transform them into central mediators of fibrosis (29), a key mechanism underlying the pathogenesis of ED. Fibrosis is characterized by excessive deposition of ECM components, which leads to tissue stiffening, impaired vascular integrity, and disruption of smooth muscle function essential for normal erectile response (8). Single-cell transcriptomic profiling of the human corpus cavernosum by Zhao et al. (7) identified two FB subclusters, FB1 and FB2, that were particularly enriched in tissue samples from patients with organic ED. These subclusters demonstrated high expression of APOC1, a gene implicated in lipid metabolism and inflammation, and exhibited transcriptional signatures indicative of proliferative and fibrotic activity (40). Specifically, FB1 and FB2 were marked by elevated expression of collagen genes, such as collagen type I alpha 1 chain (COL1A1) and collagen type III alpha 1 chain (COL3A1), fibronectin (FN1), and profibrotic cytokines, such as transforming growth factor β1 (TGFB1) and connective tissue growth factor (CTGF) (7). These transcriptional programs are associated with enhanced ECM production and remodeling, contributing to decreased compliance of the corpus cavernosum and compromised veno-occlusive function, both of which are critical determinants of erectile competence.

Fibrosis represents a pathological hallmark of structural ED, particularly in the context of diabetes and postoperative nerve injury (41). The accumulation of fibrotic tissue disrupts the balance between smooth muscle and ECM, a homeostatic relationship that is vital for efficient penile blood flow during erection (42). This results in diminished penile rigidity and an impaired hemodynamic response to sexual arousal, driven by reduced distensibility and elasticity of the corpus cavernosum (43). Underlying mechanisms include the overproduction of type I and III collagen, which increases tissue rigidity (44); upregulation of ECM-stabilizing enzymes such as lysyl oxidase (45); suppression of matrix metalloproteinases and concurrent activation of tissue inhibitors of fibrosis, which collectively hinder ECM turnover (46); and the chronic release of cytokines including interleukin-6 (IL-6) and tumor necrosis factor alpha that perpetuate FB activation (47). In a murine model, Bae et al. (6) demonstrated that under diabetic conditions, FB in the corpus cavernosum lose their reparative functions and adopt a pro-fibrotic phenotype. Among the four identified subclusters, FB expressing high levels of Egr1 were associated with stress-response pathways and tissue remodeling (48), potentially mirroring the activated FB states observed in human ED. These cells exhibited downregulation of genes involved in angiogenesis and ECM regulation, resulting in vascular rarefaction and spongiform fibrosis. Additionally, the emergence of ribosomal FB subpopulations (Rps25+, Rps17+) may signify an adaptive response to chronic metabolic stress; however, their persistent activation could exacerbate tissue dysfunction through maladaptive protein synthesis and matrix deposition. Recent scRNA-seq profiling of Peyronie’s disease (PD) tissue (49) revealed FB subpopulations enriched in fibrotic and inflammatory gene signatures, including high expression of COL1A1, actin, alpha 2, smooth muscle (ACTA2), and IL-6. While some features overlap with fibrotic FBs observed in ED (e.g., FB1/FB2), PD-associated FBs also exhibit enhanced immune cell crosstalk, particularly involving mast cells and macrophages. These differences highlight disease-specific stromal responses and underscore the need for comparative analyses across fibrotic penile disorders. Thus, selectively targeting FB subclusters—such as FB1 and FB2 in humans or Egr1-high FB in mice—may offer a novel therapeutic avenue to mitigate pathological fibrosis and preserve or restore erectile function.

In summary, these scRNA-seq studies provide a refined and functionally informative map of the penile microenvironment. They both corroborate and extend prior findings, generating new hypotheses regarding the roles of distinct FB subclusters, the significance of intercellular signaling, and the molecular underpinnings of ED. Future investigations integrating spatial transcriptomics, lineage tracing, and in vivo perturbation models will be essential for validating these FB populations and delineating their contributions to both disease progression and tissue repair.

Limitations of scRNA-seq and translational challenges

While scRNA-seq has provided unprecedented insights into the heterogeneity and function of FB subpopulations in the corpus cavernosum, several limitations must be acknowledged. First, sampling bias can influence results, particularly in human tissues where regional diversity or rare subtypes may be underrepresented due to limited sample availability or enzymatic dissociation artifacts. Second, interspecies extrapolation from murine models to humans is complicated by anatomical and developmental differences—for example, the notable abundance of chondrocytes in mouse penile tissue is not recapitulated in human data. Third, the translational targeting of FB subtypes remains technically challenging, as many subclusters share overlapping transcriptional markers, and surface antigens enabling selective delivery are poorly defined. These issues highlight the necessity of integrating scRNA-seq with complementary tools such as spatial transcriptomics, lineage tracing, and functional perturbation studies to validate the roles of specific FB subsets and advance their therapeutic exploitation.

Future directions for FB-based ED therapeutic strategies

Recent work by Guimaraes et al. (10) has highlighted the critical role of FB in regulating penile erection. Their findings demonstrate that FB constitute a key cell population within the corpus cavernosum, modulating vascular tone and blood flow during erectile function. One of the most notable observations from this study is the ability of FB to act as a molecular sink for phenylephrine, an α-adrenergic vasoconstrictor, thereby attenuating excessive vasoconstriction and stabilizing penile hemodynamics. Intriguingly, aging is associated with a progressive increase in FB number in the corpus cavernosum. While this might enhance adrenergic buffering capacity, it could also dysregulate finely balanced neurovascular signaling, especially if aging FB acquire senescent or fibrotic phenotypes. These age-related changes may partly explain altered erectile pharmacodynamics and reduced therapeutic efficacy of vasoactive agents in older patients. FB exhibit dynamic activity, with their number and function adapting in response to erectile activity. These cells contribute to ECM remodeling in response to diverse signals, a process fundamental to preserving the structural integrity and physiological function of penile tissue. Furthermore, the Notch signaling pathway has been identified as a pivotal regulator of FB populations within the corpus cavernosum. Notch inhibition has been shown to increase FB numbers, thereby enhancing penile perfusion. This pathway plays a central role in determining FB fate and activity, and is closely linked to fibrosis, vascular remodeling, and tissue repair—processes that are critical in the pathogenesis of ED. Although therapeutic modulation of this pathway is challenged by its complexity and the risk of off-target effects, Notch signaling remains a promising target, particularly for restoring FB function compromised by conditions such as diabetes or nerve injury. This study offers novel insights into the role of corpus cavernosum FB in erectile physiology and underscores their potential as therapeutic targets for ED. Future strategies should aim to inhibit specific Notch receptors (Notch1, Notch2) or ligands (Jagged1, Delta-like canonical notch ligand 4) using monoclonal antibodies or small molecule inhibitors (50,51); block proteolytic activation of Notch receptors through γ-secretase inhibitors such as N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT) or MK-0752 (52,53); suppress expression of Notch components via RNA interference, CRISPR/Cas9, or other gene-editing technologies (54,55); and modulate the FB microenvironment (e.g., oxygen levels, mechanical stress, immune context) to fine-tune Notch signaling activity (56-58).

Secondly, FB-derived EVs have demonstrated significant therapeutic potential across a range of diseases, owing to their regenerative, immunomodulatory, and tissue-repair properties (59-63). For instance, FB-derived EVs enriched with growth factors and pro-survival proteins, such as vascular endothelial growth factor, have been shown to facilitate cardiac tissue repair and enhance recovery following myocardial infarction or heart failure (61). In models of idiopathic pulmonary fibrosis, these EVs exhibit anti-fibrotic properties and promote epithelial regeneration, leading to improved pulmonary function (60). In the context of diabetic foot ulcers and other chronic wounds, FB-derived EVs accelerate tissue repair by promoting cellular proliferation, migration, and angiogenesis (63). In preclinical models of Parkinson’s disease, FB-derived EVs have demonstrated neuroprotective effects by attenuating inflammation and supporting neuronal survival, suggesting potential applications in neurodegenerative disease therapy (62). Similarly, EVs derived from renal FB have been found to mitigate renal fibrosis and improve kidney function in models of both chronic kidney disease and acute kidney injury (64). In experimental liver fibrosis, FB-derived EVs have reduced fibrotic matrix deposition and stimulated hepatocyte regeneration, indicating promise for chronic liver disease treatment (59). Despite the extensive evidence supporting the therapeutic utility of FB-derived EVs in various organ systems, their application in ED remains an emerging field. This is largely due to the anatomical and physiological complexity of penile tissue, which comprises intricately interacting vascular, smooth muscle, and neural components (7). These interactions are uniquely affected in ED, making it challenging to directly extrapolate findings from other tissues. However, it is essential to emphasize that the current evidence for FB-derived EV efficacy in erectile tissue remains limited. Most mechanistic and therapeutic studies have been conducted in non-urologic organs with markedly different structural, vascular, and neurogenic characteristics. Therefore, direct extrapolation to the corpus cavernosum should be approached with caution. Functional studies specifically evaluating FB-derived EVs in penile models are urgently needed to validate their therapeutic relevance in ED. Importantly, distinct FB subclusters exhibit specialized transcriptional and functional phenotypes—some subsets (e.g., FB1, FB2, and high Egr1-expressing FB) promote fibrosis, whereas others (e.g., FB3, FB5, and high Cxcl12-expressing FB) support angiogenesis and tissue regeneration. This heterogeneity presents an opportunity to isolate specific subclusters and harness their EVs for targeted ED therapy, aligning with current advances in precision medicine and cell type–specific interventions. Nonetheless, as with all emerging cell-derived therapies, issues such as potential immunogenicity, off-target effects, and long-term safety must be rigorously evaluated prior to clinical translation. In summary, although the use of FB-derived EVs in ED treatment is currently limited, their regenerative and anti-inflammatory capacities offer promising prospects for novel therapeutic strategies. Overcoming challenges related to tissue specificity, vesicle purification, and targeted delivery will be essential for optimizing their clinical utility in ED. In a recent study of PD, FB-derived EVs were shown to exert potent pro-fibrotic effects through the delivery of TGF-β1, fibronectin, and other matrix-regulating proteins (65). These EVs induced myofibroblast differentiation and enhanced collagen deposition via activation of SMAD2/3 signaling in recipient cells, suggesting that FB-derived EVs actively modulate fibrotic cascades in the penile microenvironment. This mechanism may have relevance to other fibrotic penile disorders such as ED, especially in diabetes or cavernous nerve injury models, where FB subclusters adopt fibrogenic phenotypes. While direct evidence of FB-derived EVs fibrotic signaling in ED remains limited, the PD data emphasize the need to investigate whether similar EV-mediated intercellular pathways drive ECM accumulation and vascular impairment in ED. Such insights could inform EV-targeted antifibrotic therapies or diagnostic biomarker development.

Thirdly, the classification of FB subtypes based on scRNA-seq provides a framework for stratifying ED patients by underlying cellular mechanisms. For instance, diabetic ED is predominantly associated with fibrotic FBs (human FB1/FB2, murine Egr1-high FBs), which overexpress collagen and profibrotic mediators, leading to ECM accumulation and tissue stiffening. A similar fibroplastic response is observed in radical prostatectomy ED due to nerve injury. In contrast, regenerative FBs (e.g., FB3/FB5 in humans, Cxcl12-high FBs in mice) exhibit pro-angiogenic and neurotrophic profiles, making them attractive targets in neurogenic or aging-related ED. This understanding enables the development of subtype-specific therapeutic strategies: antifibrotic agents or Notch inhibitors may be used to reverse ECM accumulation in fibrotic phenotypes, whereas EV-based or pro-regenerative signaling interventions may support tissue repair in degenerative settings. In aging-associated ED, a combination of both approaches may be necessary due to the coexistence of vascular dysfunction and stromal fibrosis. To synthesize these insights, Table 4 summarizes the major clinical phenotypes of ED, the corresponding FB subtypes and molecular pathways, and their respective therapeutic strategies. We also provide a feasibility estimate for each approach over the next 5–10 years, based on current translational progress. This feasibility assessment is derived from a composite evaluation of preclinical maturity, target specificity, delivery compatibility, anticipated safety and immunogenicity, and regulatory considerations. Strategies supported by robust in vivo evidence, well-defined molecular targets, and practical delivery systems are rated as having higher translational potential within this timeframe.

Table 4. Clinical phenotypes of ED and associated FB subtypes, pathways, and therapeutic implications.

ED phenotype	Associated FB subtypes	Key pathways/features	Potential therapeutic strategies	Feasibility (5–10 years)
Diabetes-related ED	Human FB1/FB2; mouse Egr1-high FB	Fibrosis, chronic inflammation, ECM overproduction	Anti-fibrotic agents (e.g., TGF-β inhibitors); FB-EVs from regenerative subtypes; Notch pathway modulation	Moderate: early-stage EV studies; fibrosis-targeting in progress
RP (neurogenic ED)	Human FB4/FB6; mouse Cxcl12-high FB	Neurotrophic factor secretion, vascular remodeling	Neuroregenerative FB-EVs; neurotrophin-loaded vesicles; spatially guided cell therapies	Moderate–high: neuro-EV tools emerging; models available
Aging-related ED	Mixed (FB1–FB5); mouse reticular FB	ECM disorganization, oxidative stress, SMC rarefaction	ECM-modulating FB-EVs; Notch signaling fine-tuning; senescence-targeted approaches	Low–moderate: complex etiology; requires combination therapies
Vasculogenic ED	FB3, FB5 (ECM-organizing FBs)	Reduced vascular compliance, endothelial dysfunction	Angiogenic EVs; co-delivery with endothelial cells	Moderate: existing angiogenic tools adaptable
Peyronie’s disease	Activated FB1-like (myofibroblastic)	Fibrosis, plaque formation, myofibroblast transition	Anti-fibrotic EVs; matrix remodeling enzymes; FB-specific gene silencing	Moderate: localized delivery feasible in near future

ECM, extracellular matrix; ED, erectile dysfunction; EV, extracellular vesicles; FB, fibroblasts; RP, radical prostatectomy; SMC, smooth muscle cell; TGF-β, transforming growth factor β.

Lastly, FB possess stem cell–like characteristics, particularly under defined conditions or within specific subpopulations (28). Their inherent plasticity enables them to undergo transdifferentiation or reprogramming into multiple cell lineages relevant to erectile tissue regeneration (66). For example, FB have been shown to differentiate into osteoblasts (67), chondrocytes (68), and adipocytes (69)—lineages typically associated with mesenchymal stem cells. Furthermore, FB can be reprogrammed into induced pluripotent stem cells (iPSCs) through the introduction of transcription factors such as octamer-binding transcription factor 4, SRY-box transcription factor 2, Kruppel-like factor 4, and cellular myelocytomatosis oncogene (70). These iPSCs can subsequently be differentiated into EC, SMCs, and even neuron-like cells, offering a flexible and autologous source for cell-based therapies aimed at restoring erectile function.

Conclusions

The cellular diversity and functional heterogeneity of FB in human and murine penile tissues underscore their central role not only in regulating ECM architecture, but also in maintaining vascular–neural homeostasis, facilitating tissue repair, and contributing to pathological fibrosis. Under physiological conditions, specific FB subclusters contribute to tissue integrity and vascular regeneration. Conversely, in pathological contexts such as diabetes or nerve injury, other FB subpopulations are associated with enhanced fibrogenesis and vascular rarefaction. Emerging evidence also supports a dynamic role of FB in modulating penile erection, with the Notch signaling pathway serving as a critical regulator of FB abundance and activity. Additionally, FB-derived EVs have shown regenerative and anti-inflammatory potential across multiple disease models, though their application in ED remains in its early stages. Owing to the specialized structure and complex microenvironment of penile tissue, further research is required to refine strategies for the isolation, targeting, and therapeutic delivery of FB-derived vesicles. In conclusion, FB represent a compelling therapeutic target for ED. Future investigations incorporating spatial transcriptomics, lineage tracing, and in vivo functional studies will be vital for delineating the specific roles of FB subclusters and advancing the clinical translation of FB-based therapies.

Supplementary

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Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.