Shi Z, Li HY, Lv BK, Li D, Lu PY, Zhou XY, Xue ME, Wang RB. GrpE-like 1-engineered synovial mesenchymal stromal cell exosomes: Mechanistic and translational priorities in osteoarthritis. World J Stem Cells 2026; 18(5): 119231 [DOI: 10.4252/wjsc.v18.i5.119231]
Corresponding Author of This Article
Rui-Bo Wang, MD, Associate Chief Physician, Professor, Second Department of Orthopedics, The Fifth Affiliated Hospital of Zhengzhou University, No. 3 Kangfu Qian Street, Erqi District, Zhengzhou 450052, Henan Province, China. 18336397518@163.com
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Shi Z, Li HY, Lv BK, Li D, Lu PY, Zhou XY, Xue ME, Wang RB. GrpE-like 1-engineered synovial mesenchymal stromal cell exosomes: Mechanistic and translational priorities in osteoarthritis. World J Stem Cells 2026; 18(5): 119231 [DOI: 10.4252/wjsc.v18.i5.119231]
Zhen Shi, Hao-Yu Li, Bo-Kang Lv, Dong Li, Rui-Bo Wang, Second Department of Orthopedics, The Fifth Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, Henan Province, China
Zhen Shi, Hao-Yu Li, Bo-Kang Lv, Dong Li, Rui-Bo Wang, Henan Provincial Key Discipline of Orthopedics, The Fifth Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, Henan Province, China
Peng-Yu Lu, First Department of Orthopedics, The Fifth Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, Henan Province, China
Xin-Yu Zhou, School of Stomatology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 451191, Henan Province, China
Meng-En Xue, Spine and Spinal Cord Surgery Ward I, Zhengzhou University People’s Hospital, Zhengzhou 462000, Henan Province, China
Author contributions: Shi Z and Li HY contributed equally to this work and share first authorship. Shi Z and Li HY contributed to writing of the original draft; Shi Z contributed to conceptualization; Shi Z, Li HY, and Lu PY contributed to methodology; Shi Z and Xue ME contributed to formal analysis; Shi Z and Zhou XY contributed to data curation; Shi Z, Li D, and Zhou XY contributed to investigation; Lv BK contributed to software, visualization, and validation; Li D and Lu PY contributed to resources; Zhou XY contributed to data curation; Xue ME contributed to figure preparation; Wang RB contributed to supervision, manuscript review & editing, and project administration. All authors participated in drafting the manuscript and have read, contributed to, and approved the final version of the manuscript.
Supported by Key Scientific Research Projects of Colleges and Universities in Henan Province, No. 26A320038; Henan Province Medical Science and Technology Research Plan Project (Joint Construction), No. LHGJ20250403, No. LHGJ20220566, and No. LHGJ20240365; Key Research and Development Program of Henan Province, No. 231111311000; and the Medical Education Research Project in Henan Province, No. WJLX2023079.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Corresponding author: Rui-Bo Wang, MD, Associate Chief Physician, Professor, Second Department of Orthopedics, The Fifth Affiliated Hospital of Zhengzhou University, No. 3 Kangfu Qian Street, Erqi District, Zhengzhou 450052, Henan Province, China. 18336397518@163.com
Received: January 22, 2026 Revised: February 9, 2026 Accepted: April 22, 2026 Published online: May 26, 2026 Processing time: 123 Days and 17.7 Hours
Abstract
Osteoarthritis (OA) remains a highly prevalent degenerative joint disorder for which truly disease-modifying therapies are still lacking. Accumulating evidence positions mitochondrial dysfunction and impaired mitochondrial quality control as central drivers of chondrocyte failure, extracellular matrix breakdown, and inflammation amplification. Mitophagy—particularly the phosphatase and tensin homolog-induced kinase 1 (PINK1)/Parkin axis—has therefore emerged as an attractive, mechanistically grounded intervention point. In this context, synovial mesenchymal stem cell-derived exosomes (SMSC-Exos) represent a compelling cell-free platform capable of delivering functional biomolecules into inflamed cartilage microenvironments. Recent experimental work demonstrates that engineering SMSC-Exos to deliver the mitochondrial co-chaperone GrpE-like 1 (GRPEL1) restores chondrocyte proliferative and migratory capacity under interleukin-1β stress, preserves anabolic extracellular matrix markers (collagen type II alpha 1/aggrecan), suppresses catabolic mediators (matrix metalloproteinase 13/A disintegrin and metalloproteinase with thrombospondin motifs 5), and mitigates oxidative damage while enhancing mitophagy signatures. Mechanistically, GRPEL1 directly associates with PINK1, and PINK1 knockdown attenuates the protective phenotype, supporting a GRPEL1-PINK1 coupling model. In vivo, intra-articular administration of GRPEL1-enriched SMSC-Exos improves histological cartilage integrity and mitophagy-related readouts in a rat OA model. Here, we synthesize mechanistic implications, highlight interpretive nuances (e.g., mitophagy activation concurrent with membrane potential recovery), and outline translational priorities, including cargo quantification, mitophagy flux validation, dosing/retention kinetics, manufacturing standardization, and biomarker-driven patient stratification.
Core Tip: Osteoarthritis still lacks reliable disease-modifying therapies. This review highlights an emerging mitochondria-centered strategy in which synovial mesenchymal stem cell-derived exosomes deliver GrpE-like 1 to engage the phosphatase and tensin homolog-induced kinase 1-dependent mitophagy pathway, improving mitochondrial homeostasis and cartilage outcomes across in vitro and in vivo models. We synthesize the mechanistic rationale and delineate the minimal translational checkpoints required for clinical readiness: Flux-based and specificity-anchored causal validation; chemistry, manufacturing, and controls—including a standardized product definition with mechanism-linked potency assays; and intra-articular pharmacokinetics/pharmacodynamics coupled to biomarker-driven patient stratification.
Citation: Shi Z, Li HY, Lv BK, Li D, Lu PY, Zhou XY, Xue ME, Wang RB. GrpE-like 1-engineered synovial mesenchymal stromal cell exosomes: Mechanistic and translational priorities in osteoarthritis. World J Stem Cells 2026; 18(5): 119231
Osteoarthritis (OA) is a heterogeneous whole-joint disease and a leading cause of pain, disability, and work loss worldwide[1,2]. Global burden analyses estimate that 595 million people were living with OA in 2020, with further substantial increases projected by 2050[1]. Although the conceptual framework of OA has expanded from a cartilage-centered disorder to a multi-tissue disease involving biomechanical, inflammatory, metabolic, and post-traumatic processes, current treatment remains largely symptom-directed, and no disease-modifying OA drug (DMOAD) has yet entered routine clinical us[2-4].
Among the upstream regulatory systems now attracting increasing interest, mitochondrial dysfunction is increasingly regarded as a pathogenic amplifier rather than a secondary epiphenomenon. In OA chondrocytes, impaired oxidative phosphorylation, excessive mitochondrial reactive oxygen species (ROS) production, loss of mitochondrial membrane potential, and danger-signal propagation converge to promote inflammatory activation, extracellular matrix (ECM) catabolism, and cell fate dysregulation[5,6]. Accordingly, mitochondrial quality control (MQC)—encompassing mitochondrial dynamics, proteostasis, biogenesis, and selective clearance of damaged organelles—has emerged as a mechanistically coherent therapeutic framework in OA[5,6]. Within this network, mitophagy, particularly the phosphatase and tensin homolog-induced kinase 1 (PINK1)/Parkin axis, is especially attractive because it is both biologically central and experimentally tractable[6].
Recent studies have further strengthened the therapeutic relevance of MQC-directed intervention in OA. Restoring mitochondria-endoplasmic reticulum crosstalk, activating the SIRT3-PINK1-PKM2 axis, promoting optineurin (OPTN)-mediated mitophagy, and engineering exosomal cargoes to modulate chondrocyte mitophagy have each been shown to attenuate OA-related cellular or structural injury in preclinical models[7-9]. These findings suggest that mitochondrial renewal is not merely a downstream readout of cartilage degeneration, but a targetable control layer linking stress adaptation to tissue preservation.
In parallel, cell-free biologic delivery platforms are becoming increasingly important in musculoskeletal translational research. Our previous work has highlighted the clinical promise of exosomes and gene vectors in intervertebral disc degeneration, the therapeutic relevance of TFEB-regulated autophagy in degenerative disc disease, and the pathogenic importance of upstream matrix-metabolic injury pathways in diabetic bone fragility[10-13]. Against this background, the recent study by Xiang et al[14] is of particular interest because it links an engineered exosomal cargo, GrpE-like 1 (GRPEL1), with PINK1-dependent mitophagy and cartilage repair in OA models. By integrating bioinformatic target nomination, interleukin (IL)-1β-induced chondrocyte injury experiments, and intra-articular validation in a destabilization of the medial meniscus model, this work advances the GRPEL1-PINK1-mitophagy axis from a descriptive mitochondrial phenotype toward a potentially actionable therapeutic node[14]. Nevertheless, important translational issues remain unresolved, including flux-based validation of mitophagy, quantitative definition of cargo loading, mechanism-linked potency assessment, manufacturing standardization, and intra-articular pharmacokinetic/pharmacodynamic (PD) feasibility[3,14]. Therefore, this review summarizes the mechanistic basis, emerging evidence, and translational priorities of GRPEL1-engineered synovial mesenchymal stem cell (SMSC)-derived exosomes (SMSC-Exos) in OA, with particular emphasis on MQC-centered causal inference and clinical translation. The overall mechanism-to-translation framework is summarized in Figure 1, and the current evidence-and-gap map is outlined in Table 1.
MITOCHONDRIAL DYSFUNCTION IN OA: FROM PHENOTYPIC SIGNATURES TO DRUGGABLE QUALITY-CONTROL NODES
Mitochondrial stress-inflammation coupling relevant to the GRPEL1-PINK1 axis
OA-relevant inflammatory stress [e.g., IL-1β/tumor necrosis factor-alpha (TNF-α)] induces a stereotyped mitochondrial stress-inflammation program in chondrocytes: Impaired oxidative phosphorylation/respiratory reserve with ATP insufficiency, destabilized mitochondrial membrane potential (ΔΨm), and accumulation of mitochondrial ROS (mtROS), together with mtROS- and mitochondrial DNA-linked innate sensing (e.g., nuclear factor kappa B/inflammasome and Cgas-STING), reinforce catabolic gene expression (A disintegrin and metalloproteinase with thrombospondin motifs 5/matrix metalloproteinase-13) and accelerate ECM loss[15-17]. We keep this background concise because it maps directly to the primary study’s mechanism-anchored readouts: GRPEL1-enriched exosomes reduced oxidative injury and restored ΔΨm under IL-1β challenge while preserving matrix anabolism and dampening catabolism, and PINK1 loss-of-function attenuated these benefits—supporting interruption of a mitochondrial stress-inflammation feed-forward loop via the GRPEL1-PINK1-mitophagy axis[14]. This framing motivates focusing on MQC modules and flux-based validation below.
The MQC system: Dynamics, proteostasis, biogenesis, and selective clearance
MQC should be framed as a system-level, modular network rather than a single pathway, because cartilage homeostasis depends on how efficiently chondrocytes isolate damage, activate repair capacity, and remove irreversibly impaired organelles under inflammatory/mechanical stress[5,6,18]. Recent OA-focused syntheses converge on a four-module MQC architecture: (1) Dynamics (fission/fusion) to segregate dysfunctional units and remodel the network; (2) Proteostasis/UPRmt to maintain import, folding, and protease surveillance; (3) Biogenesis to replenish functional mitochondrial mass and restore respiratory reserve; and (4) Selective clearance (mitophagy) to subtract the ROS/damage-associated molecular pattern-generating fraction at its source[5]. Mechanistically, OA-relevant stressors (e.g., IL-1β) can push dynamics toward fragmentation via extracellular regulated kinase 1/2-dynamin-related protein 1 activation, with concomitant reduction in fusion capacity (e.g., mitofusin 1 downshift), creating a permissive architecture for damage propagation unless counterbalanced by repair/clearance modules[16,19]. In parallel, mitochondrial proteostasis emerges as an actionable hub: The UPRmt is inducible in OA cartilage and experimentally protective, and pharmacologic augmentation (e.g., nicotinamide riboside in the cited work) improves mitochondrial function and attenuates OA phenotypes in a manner dependent on pathway-level regulators (e.g., activating transcription factor 5), supporting proteostasis as a bona fide disease-modifying lever rather than an epiphenomenon[20,21]. Finally, the “replacement arm” of MQC—biogenesis—is commonly discussed through AMP-activated protein kinase/SIRT1-peroxisome proliferator-activated receptor-gamma coactivator-1 alpha-TFAM logic, which defines mitochondrial reserve and resilience; in OA-oriented reviews, impaired or insufficiently recruited biogenesis is repeatedly positioned as a reason why short-term compensations fail to translate into durable tissue-level benefit[22-24].
Within this system, mitophagy functions as the terminal “damage-load valve”: When clearance is inadequate (or when flux is blocked), ROS-producing mitochondria persist, amplifying inflammation and catabolism upstream of ECM failure—hence the conceptual advantage of MQC over single-node downstream inhibition[8,25,26]. Importantly, the translational claim for MQC-targeting interventions increasingly hinges on flux-aware evidence (i.e., demonstrating that mitochondria are actually being turned over, not merely that LC3/PINK1/Parkin markers change). Consistent with this logic, recent work also supports that restoring higher-order mitochondrial organization can be therapeutic—for example, re-establishing mitochondria-endoplasmic reticulum contacts has been reported to reverse cartilage degeneration and relieve OA in vivo, underscoring MQC as an integrated engineering target rather than an isolated molecular slogan[7]. Together, these data motivate a mechanistically disciplined transition from “mitochondrial dysfunction” as a descriptive phenotype to MQC modules as druggable, pathway-instrumented nodes, and they set up why the next section prioritizes PINK1-centered mitophagy as the most experimentally tractable entry point for causality and PD readouts[6].
PD readouts for MQC-targeting DMOAD development
In MQC-targeting DMOAD development, PD packages should be pre-specified as a translational, tiered “mechanism-function-tissue” evidence stack that can propagate from in vitro screening to in vivo dose-finding and, ultimately, early-phase clinical trials. Mechanistic PD should demonstrate target engagement within the MQC circuitry [e.g., PINK1 stabilization and phospho-Ser65 ubiquitin (pSer65-ubiquitin) accumulation, Parkin recruitment with mitochondrial substrate ubiquitination, and/or activation of compensatory MQC modules such as UPRmt and biogenesis nodes], thereby de-risking claims that phenotypic rescue is mitochondria-driven rather than pleiotropic[20,27,28]. Functional PD should quantify mitochondrial performance (ATP output, oxygen consumption rate/ECAR, ΔΨm stability, mtROS burden, Ca2+ handling and mitochondrial permeability transition pore susceptibility, plus network dynamics), ideally with flux-aware controls (lysosomal blockade or reporter-based mitophagy turnover) to distinguish true increases in selective clearance from static marker accumulation[28,29]. Disease-level PD should connect these proximal events to cartilage matrix homeostasis and joint pathology (key ECM turnover readouts, proteoglycan retention, and standardized histopathological scoring), while also considering scalable biochemical and imaging biomarkers for patient stratification and response monitoring[30,31]. Such a tiered PD strategy is increasingly viewed as essential for OA drug development, where heterogeneity and endpoint limitations have repeatedly undermined DMOAD translation, and where dose/schedule optimization hinges on demonstrating both engagement and joint-level structural benefit[32,33].
PINK1-DEPENDENT MITOPHAGY IN CARTILAGE HOMEOSTASIS: MECHANISTIC TRACTABILITY AND CONTEXT DEPENDENCY
GRPEL1 engagement of the canonical PINK1-Parkin axis: Evidence, biochemical specificity, and remaining gaps
Because the PINK1-Parkin pathway is canonical knowledge for the expert audience, we do not recapitulate its stepwise biochemistry here. Instead, we treat it as an operational “rigor scaffold” to evaluate whether the primary study demonstrates pathway-level target engagement and to define PD-ready readouts that can support translation. In the study of Xiang et al[14], the proposed linkage between exosomal GRPEL1 and PINK1-dependent mitophagy is supported by three convergent lines of evidence: (1) Co-immunoprecipitation consistent with a GRPEL1-PINK1 association; (2) Directionally coherent shifts in commonly used mitophagy/autophagy markers (increased PINK1/Parkin and LC3-II/I with reduced p62); and (3) PINK1 loss-of-function that attenuates mitochondrial rescue and ECM protection under inflammatory challenge and in vivo[14].
However, from a biochemical specificity standpoint, the current “association-plus-perturbation” package is insufficient to conclude a direct, specific GRPEL1 → PINK1 activation mechanism[34]. Co-immunoprecipitation does not discriminate direct binding from an indirect complex, and key parameters that would validate specificity—binding affinity (Kd), stoichiometry, and interaction interfaces (e.g., whether PINK1’s kinase/regulatory regions and GRPEL1’s co-chaperone motifs are required)—remain undefined. Mechanistic closure would therefore benefit from orthogonal interaction validation and mapping (e.g., domain truncation/mutagenesis with loss-of-binding tests, proximity approaches under mitochondrial stress, and purified-protein biophysics such as SPR/BLI/ITC where feasible), which would also reduce off-target interpretations[35-37].
At the pathway-event level, the study does not yet resolve the most specific and quantifiable tags of canonical PINK1-Parkin activation (e.g., pSer65-ubiquitin accumulation, Parkin recruitment and substrate ubiquitination, and OPTN/NDP52-dependent initiation), which are increasingly viewed as the mechanistically anchored PD nodes for mitophagy-targeting interventions[28,38,39]. Moreover, because static changes in PINK1/Parkin/LC3/p62 can reflect either induction or stalled turnover, flux-aware confirmation (lysosomal end-blockade designs and/or mitophagy reporters such as mito-QC/mt-Keima) would materially strengthen causal closure and improve translational interpretability[28,39,40].
A further interpretability issue—highly relevant for an engineered-cargo strategy—is that GRPEL1’s canonical biology lies in mitochondrial proteostasis and protein import chaperone networks. Altering GRPEL1 abundance/stoichiometry via engineered exosome delivery could, in principle, perturb import/proteostasis programs, induce compensatory mitochondrial stress responses, or secondarily modulate mitophagy markers, thereby confounding pathway attribution[37,41,42]. Accordingly, mechanism claims would be strengthened by import/proteostasis controls (e.g., mitochondrial import reporter/precursor processing assays and UPRmt/proteostasis stress readouts) alongside dose-response titration of GRPEL1 loading to avoid supraphysiologic stress[42,43].
Taken together, we retain the PINK1-Parkin axis here not for didactic purposes, but as a concrete framework that specifies what should be measured to substantiate a GRPEL1 → PINK1 mechanism, distinguish direct pathway engagement from proteostasis-driven confounding, and convert mechanistic plausibility into PD-ready target engagement for an engineered exosome product, as further elaborated in the following discussion.
Why PINK1 mitophagy is experimentally actionable: Genetic interrogation and pathway-level PD metrics
A principal advantage of the PINK1 axis, relative to many upstream “mitochondrial dysfunction” descriptors, is that it comes with a built-in causality toolbox: Necessity and sufficiency can be tested by PINK1/PRKN loss-of-function (knockdown/knockout), gain-of-function, and rescue designs, and key pathway relays (e.g., OPTN) can be perturbed to interrogate initiation vs execution[27,44]. This genetic tractability has been paired with event-based PD that can be assembled into a contiguous evidence chain—PINK1 activation/outer mitochondrial membrane accumulation, pSer65-ubiquitin formation, Parkin recruitment and substrate ubiquitination, adaptor-driven initiation, and readouts of mitochondrial turnover—where pSer65-ubiquitin has emerged as a particularly specific, quantifiable “pathway tag” enabled by ultrasensitive immunoassays and reagent standardization[28,45]. Importantly, these event PD nodes are not merely biochemical ornaments: They can be explicitly linked to functional outputs (mtROS/ΔΨm/oxygen consumption rate-ATP recovery) and inflammatory-catabolic attenuation in cartilage-relevant systems, as illustrated by recent OA studies in which OPTN deficiency impairs mitophagy and accelerates OA phenotypes, while interventions that promote PINK1/Parkin-dependent mitophagy reduce chondrocyte injury and slow structural degeneration[38,46,47]. This logic creates a clear translational bar for engineered delivery platforms: If an engineered exosomal cargo is claimed to act through PINK1-dependent mitophagy, it should demonstrate target engagement at one or more event-PD steps (e.g., pSer65-ubiquitin/Parkin-ubiquitin cascade and adaptor-linked initiation) alongside concordant mitochondrial functional rescue, rather than relying on static marker shifts alone—an expectation that directly applies to the GRPEL1-exosome paradigm positioned by Xiang et al[14].
Context-dependent mitophagy states across OA microenvironments and disease stages
A practical way to reconcile apparently discordant OA mitophagy literature is to treat mitophagy as a state variable shaped by stage, microenvironment, and clearance capacity, and to classify observations into three operational states: (1) Insufficient initiation describes settings where mitochondrial damage accumulates but PINK1-axis event signals are weak (or fail to rise appropriately), consistent with human OA data showing reduced PINK1/Parkin-dependent mitophagy in injured cartilage and experimental evidence that PINK1 deficiency aggravates cartilage degeneration, while PINK1 gain-of-function can be protective—supporting initiation as a limiting step in at least a subset of contexts[27]; (2) Compensatory activation without effective clearance captures scenarios in which upstream mitophagy signaling is induced but downstream disposal is rate-limited (typically by the autophagy-lysosome system), so damaged mitochondria persist despite “activation-like” signaling; this is mechanistically plausible in metabolic-risk OA, where dyslipidemia-relevant stressors (e.g., oxidized low-density lipoprotein) disrupt TFEB-regulated autophagy-lysosome function and autophagic flux, and in obesity-associated OA where autophagic dysfunction drives STING1 activation and worsens cartilage degeneration—both pointing to lysosomal competence as a key determinant of whether mitophagy signaling translates into organelle subtraction[48]; and (3) Blocked flux/pseudo-activation refers to cases where static markers (e.g., LC3 puncta and PINK1/Parkin accumulation) rise because mitophagy progression stalls, creating an “apparent activation” phenotype that is, in fact, consistent with impaired turnover; this risk is heightened by aging-linked declines in lysosomal capacity and by mechanical microenvironments that alter pathway kinetics[49]. Notably, mechanical context can even invert interpretation: ECM stiffening—a hallmark of aging cartilage—has been reported to hyperactivate Parkin-dependent mitophagy via mechanosensitive Parkin acetylation and to accelerate chondrocyte senescence and OA, emphasizing that “more mitophagy markers” is not automatically beneficial without flux confirmation[50]. Taken together, context dependence mandates stratification and window-period thinking (stage, inflammatory intensity, metabolic background, aging/Lysosomal reserve, and mechanical load), and it sets a methodological requirement for the remainder of this section: OA-relevant claims about PINK1 mitophagy must demonstrate flux (turnover) rather than static marker shifts, which underscores the need for flux-based validation in the following discussion[46].
Flux-based validation and causal inference as prerequisites for translational claims
A central methodological prerequisite for any “PINK1-mitophagy-DMOAD” translational claim is that static markers are not evidence of enhanced mitophagy: Increases in LC3 puncta, p62 abundance, or total PINK1/Parkin expression can reflect induction, blockade, or stalled turnover, and therefore cannot, on their own, establish that mitochondrial clearance has occurred; accordingly, the field-standard position is that autophagy/mitophagy must be interpreted through flux logic and orthogonal readouts rather than single-time-point marker snapshots[46,51]. At minimum, flux-supporting evidence should fall into three categories: (1) Turnover/flux designs under lysosomal-end perturbation, where lysosome inhibition (or equivalent end-stage blockade) is used to distinguish increased delivery from impaired degradation and to quantify net turnover[51]; (2) Mitochondria-to-lysosome-specific flux reporters/readouts, such as tandem-fluorescence mito-QC-type systems that directly score mitolysosome formation (acidic quenching of GFP with retained mCherry) and enable quantitative cytometric or imaging-based enumeration of mitochondrial cargo reaching the lysosome, complemented where appropriate by biochemical “event markers” that are tightly coupled to PINK1 pathway activation (e.g., pS65-ubiquitin detection using validated ultrasensitive assays/antibodies)[28,29,44]; and (3) Time-resolved event-based PD coherence, requiring a consistent sequence from PINK1 stabilization/ubiquitin phosphorylation to adaptor recruitment, LC3 engagement, and degradation-compatible endpoints, rather than isolated upstream activation signals that may persist even when canonical autophagy machinery is not the dominant disposal route[29,45]. Critically, causal closure should be treated as a gating criterion for translational language: Pathway interception (genetic or pharmacologic) must attenuate the protective phenotype, and pathway restoration/rescue must reinstate it—standards already exemplified in OA-relevant mitophagy interventions where loss-of-function of a mitophagy receptor (e.g., OPTN) blocks mitophagy and abrogates benefit, while interventions that claim Pink1/Parkin-dependent mitophagy show concordant molecular and disease-level improvements[27,46]. These flux-and-causality requirements are not merely academic: They dictate which PD endpoints are credible for dose/frequency optimization and target engagement, and they should directly inform chemistry, manufacturing, and controls (CMC) potency assays for engineered delivery products (e.g., exosome lots), which should be anchored to mechanism-proximal, flux-sensitive endpoints rather than static marker shifts[27,52] (Table 2).
Table 2 Mechanism-to-translation checklist for GrpE-like 1-exosomes.
Domain
Minimal assay set (examples)
What counts as “engagement/Go”
Pitfall if omitted
Target engagement (event): PINK1 on mitochondria
Mito/cyto fractionation + WB; IF colocalization with TOM20/VDAC (time-course)
↑Mitochondria-localized PINK1 with temporal precedence
DELIVERY-TARGET-PATHWAY: AN EVIDENCE MAP AND MECHANISTIC FRAMEWORK FOR GRPEL1- EXOSOMES
Disease association and target prioritization
Public transcriptomic mining can nominate candidates, but “disease association” is robust only when supported by cross-cohort reproducibility, orthogonal validation in human joint tissues, and careful interpretation of immune correlations as associative rather than causal. In the study by Xiang et al[14], GRPEL1 was nominated from two public human OA cartilage transcriptome cohorts (GSE169077 and GSE114007) and linked to inflammatory features and radiographic severity, providing an initial cross-dataset consistency signal[14]. However, these in silico immune-association readouts remain correlational and may be influenced by cell-composition shifts and systemic inflammation, such that they should be interpreted as “association tags” rather than mechanistic evidence of immune causality.
Importantly, although the study reports reduced GRPEL1 in OA joint compartments (synovial fluid/cartilage) and correlation with inflammatory burden[1], a key translational question is whether the protein-level observations were confirmed in an independent validation cohort and whether assay performance was sufficiently controlled for pre-analytical and analytic variables (e.g., tissue handling/fixation, antibody specificity, and scoring reproducibility) before the work proceeds to animal efficacy claims. Methodological literature on tissue-based biomarkers and contemporary immunohistochemistry analytic-validation guidelines emphasize that fit-for-purpose protein-level validation is a prerequisite for clinically interpretable biomarker claims and for prioritizing targets based on human tissue biology[53,54].
With these caveats stated explicitly, GRPEL1 still retains mechanistic plausibility as an upstream “proteostasis-MQC interface” node. This is consistent with evidence that mitochondrial proteostasis programs (e.g., UPRmt) track with OA progression and can be therapeutically leveraged, supporting the rationale for selecting protein quality-control nodes as intervention points[20]. Independent transcriptome-based diagnostic modeling has also highlighted GRPEL1 among discriminative features across analytic pipelines[55]; nevertheless, prospective, protein-level validation in well-phenotyped human OA tissues would strengthen the robustness of the disease-association-to-prioritization step.
In vitro functional evidence
A reductionist IL-1β chondrocyte-injury setting provides a tractable bridge from “delivery” to “repair-relevant function”. In this context, GRPEL1-enriched exosomes restore repair-associated cellular phenotypes (viability/proliferation and migration) while shifting matrix balance toward protection (anabolism preserved and catabolism dampened), thereby aligning mitochondria-centered target engagement with an ECM-facing functional output that is directly relevant to DMOAD intent[14]. Notably, this phenotype-to-matrix linkage is consistent with broader SMSC-Exos OA literature, where engineered or cargo-optimized SMSC-Exos reduce Osteoarthritis Research Society International scores and suppress ECM degradation programs in destabilization of the medial meniscus models, underscoring that synovium-derived exosomes can serve as a feasible intra-articular chassis for modular cargo-to-cartilage effects[56,57].
Mechanistic anchoring and in vivo relevance
A key strength of the GRPEL1-exosomes proposition is the attempt to build a complete “cargo-target-pathway-tissue endpoint” chain rather than relying on marker-level associations. Mechanistically, direct GRPEL1-PINK1 interaction evidence (e.g., Co-immunoprecipitation) is coupled with concordant mitochondrial functional readouts (ROS, membrane potential, and lipid peroxidation/oxidative injury indices) and mitophagy-linked molecular shifts, forming a pathway-anchored rationale for how exosomal cargo could re-balance chondrocyte MQC under inflammatory stress[14]. Causality is further reinforced when pathway interception (PINK1 knockdown) attenuates the pro-survival/pro-migratory and matrix-protective effects, establishing a minimal causal scaffold that is frequently missing in exosome-based OA claims[14]. In vivo, intra-articular dosing in the destabilization of the medial meniscus rat model improves proteoglycan preservation and cartilage structural integrity, providing a tissue-level efficacy endpoint that aligns with the DMOAD ambition of structural benefit rather than symptomatic modulation[14]. This mechanistic framing is also coherent with emerging, high-impact OA studies that nominate PINK1/Parkin-mitophagy as a druggable, pathway-level lever through diverse modalities (e.g., nanoengineered gene-editing cargoes, metabolic/mitochondrial renewal axes, and exosome-mediated programs), collectively strengthening the rationale that PINK1-centered mitophagy is not only mechanistically interpretable but also therapeutically engineerable in the joint microenvironment[8,9,57]. This integrated delivery-to-mechanism-to-translation workflow is summarized in Figure 2.
Figure 2 Integrated mechanism-to-translation schematic for GrpE-like 1-engineered synovial mesenchymal stem cell-derived exosomes targeting phosphatase and tensin homolog-induced kinase 1-dependent mitophagy in osteoarthritis.
A left-to-right workflow summarizes the proposed therapeutic logic. Osteoarthritis-relevant inflammatory stress (interleukin-1β/tumor necrosis factor-alpha) and oxidative pressure (ROS) precipitate chondrocyte mitochondrial injury, characterized by decreased mitochondrial membrane potential (ΔΨm) and ATP production with increased mitochondrial ROS. Synovial mesenchymal stem cell-derived exosomes engineered to carry GrpE-like 1 are delivered intra-articularly and internalized by chondrocytes. GrpE-like 1 associates with phosphatase and tensin homolog-induced kinase 1 at the outer mitochondrial membrane, promoting phosphatase and tensin homolog-induced kinase 1 signaling (phospho-Ser65 ubiquitin), Parkin recruitment, and LC3-positive autophagosome formation followed by lysosomal degradation, thereby activating mitophagy. Enhanced mitochondrial renewal (mitochondrial ROS↓, ΔΨm↑, ATP↑) is linked to restoration of extracellular matrix homeostasis (collagen type II alpha 1/aggrecan↑; matrix metalloproteinase-13/A disintegrin and metalloproteinase with thrombospondin motifs 5↓) and improved cartilage integrity. The bottom ribbon highlights the minimal translational priorities required to move from proof-of-concept to an actionable disease-modifying osteoarthritis drug pathway: Flux validation, chemistry, manufacturing, and controls and mechanism-linked potency, and intra-articular pharmacokinetics/pharmacodynamics with endotype-aware stratification. OA: Osteoarthritis; IL-1β: Interleukin-1β; TNF-α: Tumor necrosis factor-alpha; ROS: Reactive oxygen species; ΔΨm: Mitochondrial membrane potential; mtROS: Mitochondrial reactive oxygen species; SMSC: Synovial mesenchymal stem cell; GRPEL1: GrpE-like 1; PINK1: Phosphatase and tensin homolog-induced kinase 1; pS65-Ub: Phospho-Ser65 ubiquitin; OMM: Outer mitochondrial membrane; ECM: Extracellular matrix; COL2A1: Collagen type II alpha 1; MMP-13: Matrix metalloproteinase-13; ADAMTS5: A disintegrin and metalloproteinase with thrombospondin motifs 5; CMC: Chemistry, manufacturing, and controls; IA: Intra-articular; PK/PD: Pharmacokinetics/pharmacodynamics.
FROM PROOF-OF-CONCEPT TO AN ACTIONABLE DMOAD: A MINIMAL TRANSLATIONAL CHECKLIST
A GRPEL1-enriched SMSC-Exos strategy will only become “DMOAD-ready” if it is specified and controlled as a product rather than a preclinical condition. At minimum, this requires a CMC package that fixes: (1) Source and upstream process (donor eligibility or master cell bank strategy, passage window, medium/exosome-depletion strategy, and culture format and scale); (2) Separation and impurity control (defined isolation workflow with quantifiable removal of protein/Lipoprotein contaminants and process residuals); and (3) Critical quality attributes aligned to community standards for extracellular vesicle (EV) identity and characterization (particle/marker profiling, orthogonal sizing/quantitation, and transparency on method limitations per MISEV2023)[58] (Table 2).
Cargo definition and quantitative loading
GRPEL1 is better described as an engineered-enriched EV cargo rather than a fully defined, quantitatively controlled therapeutic attribute at the current stage. Although Xiang et al[14] demonstrate GRPEL1 enrichment in SMSC-derived EVs and biological activity in recipient cells, key translational parameters are not reported, including loading efficiency (e.g., GRPEL1 per EV particle and/or per μg EV protein), lot-to-lot consistency, and cargo stability under storage/handling. Current International Society for Extracellular Vesicles minimal-information guidance emphasizes that EV therapeutic claims should be supported by rigorous EV identity/purity characterization and fit-for-purpose quantification of both EV dose (particle/protein) and relevant cargo attributes[59,60].
Cargo functionality and mechanism-linked potency
Moreover, cargo presence does not guarantee cargo functionality. To confirm that GRPEL1 is properly folded and biologically active within EV preparations, mechanism-linked functional assays are needed, ideally comparing GRPEL1-enriched EVs to matched control EVs (vector/empty) using pathway-event and flux-aware readouts (e.g., mitochondrial PINK1 accumulation, pSer65-ubiquitin formation, Parkin recruitment/substrate ubiquitination, and mitophagy flux reporters/end-blockade designs). Quantitative cargo reporting can be operationalized by combining particle counting (e.g., nanoparticle tracking analysis) with orthogonal GRPEL1 measurement (enzyme-linked immunosorbent assay or targeted MS) to express GRPEL1 per particle and per μg EV protein, as recommended in emerging EV cargo-loading and quality-control discussions[52,61].
Cargo routing and EV compositional integrity
Finally, because GRPEL1 is canonically a mitochondrial proteostasis factor, it remains mechanistically unclear how it is preferentially sorted into EVs and whether engineering perturbs EV biogenesis or native cargo composition. This motivates comparative EV profiling (e.g., proteomics and EV marker/impurity panels) to demonstrate that the enhanced activity is attributable to GRPEL1 enrichment rather than broader compositional shifts induced by overexpression/engineering[62-65].
For intra-articular implementation, the pharmacology is typically the rate-limiting step. Intra-articular agents are rapidly cleared from the joint space, and repeated injections add procedural burden and infection risk, making residence time, joint distribution, and dose-frequency optimization non-negotiable components of translation. A pragmatic pharmacokinetics/PD plan should therefore couple: (1) Joint exposure/retention (label-free or labeled tracking, synovial fluid time-course); (2) Pathway PD (PINK1/Parkin event-based metrics and flux-consistent readouts in cartilage/synovium where feasible); and (3) Disease PD bridging to structural endpoints (ECM biomarkers, compositional imaging, and histologic scores in models) in a way that supports regimen selection and demonstrates mechanism-hit at clinically realistic dosing[32,66,67]. Given OA heterogeneity, trial feasibility also improves when PD is paired with patient stratification—for example, inflammatory endotype enrichment and baseline mitochondrial stress signatures—consistent with recent Osteoarthritis Research Society International clinical trials symposium guidance emphasizing disease activity-oriented biomarker logic and endotype-aware development pathways[32,33,68].
CONCLUSION
GRPEL1-loaded SMSC-Exos represent a promising mitochondria-centered strategy for OA by linking engineered cargo delivery to a pharmacodynamically tractable MQC pathway, namely, PINK1-dependent mitophagy. Current evidence supports their ability to improve mitochondrial homeostasis, preserve ECM balance, and confer structural benefit in preclinical models. However, clinical translation will depend on three key advances: Rigorous flux- and specificity-based mechanistic validation, CMC-standardized cargo and potency definition, and intra-articular pharmacokinetics/PD-guided regimen development with biomarker-informed patient stratification. If these requirements are met, GRPEL1-exosomes may provide not only a candidate DMOAD approach, but also a translational framework for MQC-targeted therapies in OA.
ACKNOWLEDGEMENTS
The authors gratefully acknowledge Zhejiang Luming Biotechnology Co., Ltd. (2nd Floor, 113-1 to 113-5 Nanliu Road, Chashan Street, Ouhai District, Wenzhou, Zhejiang Province, China) for providing technical support in scientific figure preparation, formatting optimization, and related manuscript assistance. The authors are solely responsible for the scientific content, interpretation, and conclusions of this manuscript.
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Footnotes
Peer review: Externally peer reviewed.
Peer-review model: Single blind
Specialty type: Cell and tissue engineering
Country of origin: China
Peer-review report’s classification
Scientific quality: Grade B, Grade C
Novelty: Grade B, Grade D
Creativity or innovation: Grade B, Grade C
Scientific significance: Grade B, Grade C
P-Reviewer: Chen GY, MD, Assistant Professor, China; Zhang JW, PhD, Academic Fellow, FRSC, Full Professor, Principal Investigator, Professor, China S-Editor: Wang JJ L-Editor: Wang TQ P-Editor: Zhao YQ
