Bone Marrow Mesenchymal Stem Cell Secretome from Acute Patients Reveals a Distinct Signature

Ada I Ozcan¹, Abdul-Razak Masoud¹, Jenna R Dennis¹, Olivia Warren¹, John Hunt¹, Jonathan E Schoen¹, Jeffrey E Carter¹, Herbert A Phelan¹, Patrick Greiffenstein¹, Lance Stuke¹, Luis Marrero², Vinod Dasa², Patrick McTernan³, Alison A Smith^1*

¹Department of Surgery, Louisiana State University Health Sciences Center, 2021 Perdido Street, New Orleans, Louisiana 70112, USA
²Department of Orthopedic Surgery, Louisiana State University Health Sciences Center, 2021 Perdido Street, New Orleans, Louisiana 70112, USA
³Department of Physiology, Louisiana State University Health Sciences Center, 433 Bolivar Street, New Orleans, Louisiana 70112, USA

*Correspondence author: Alison A Smith, MD, PhD, FACS, Assistant Professor of Clinical Surgery, Louisiana State University Health Sciences Center New Orleans, Department of Surgery, 2021 Perdido Street, Room 8143, New Orleans, LA 70112, USA; Email: [email protected]

Citation: Ozcan AI, et al. Bone Marrow Mesenchymal Stem Cell Secretome from Acute Patients Reveals a Distinct Signature. J Reg Med Biol Res. 2025;6(1):1-9.

Copyright^© 2025 by Ozcan AI, et al. All rights reserved. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Table 1: Donor (patient) characteristics.

Table 2: Statistical analysis of secreted cytokines.

Figure 1: Flow cytometry plots of a representative sample obtained using a Cytek® Northern Lights (Cytek Biosciences, Fremont, CA, USA) machine, analyzed with SpectroFlo (Cytek Biosciences, Fremont, CA, USA) software. FSC-H: Forward Scatter-Height; FSC-A: Forward Scatter-Area, SSC-A: Side Scatter-Area, APC-eFluor780: viability dye, BV510: CD90, PE-Cy7: CD73, PE: CD34, APC: CD45, BB700: CD105, P1 gate represents single cells, P2 gate represents alive cells.

Figure 2: Secretion levels of human cytokines derived from the secretome of bone marrow-derived MSCs from emergency patients compared to kOA patients. Profile shows the mean concentration (pg/mL) of cytokines analyzed in triplicates. N=3, *p=0.01, **p=0.001.

Discussion

Cells in the bone marrow produce a variety of cytokines, chemokines and growth factors, collectively referred to as the ‘secretome’; to regulate vital processes such as osteogenesis, hematopoiesis and stem cell regeneration [5]. Following trauma, BMSCs have been shown to migrate to the wound site in response to chemical signals, serving as progenitors of epidermal, dermal and endothelial cells that will be used to regenerate tissue [6,7].  The improved healing outcomes reported with BMSC transplantation in acute and chronic wounds have also been attributed to the autocrine and paracrine factors that constitute the “secretome” [16].  However, adoptive stem cell transfer poses several challenges including limited efficacy due to compromised cell viability and low engraftment after implantation [7,8]. Additionally, stem cells harvested from patients with a dysfunctional trauma response for auto-transplant can harbor the same disruptions that impair wound healing, thereby reducing treatment efficacy. However, despite the variable reports on the efficacy of adoptive transfer; there is promising data suggesting that MSC conditioned media alone can improve wound healing in in-vitro assays [7].  Identifying key elements, such as paracrine factors, of the MSC response to acute stressors such as trauma can guide tailored treatment strategies to overcome these challenges. 

Physiological response to trauma is characterized by a rise in IL-6, IL-10 and MCP-1 triggered by the release of Damage Associated Molecular Patterns, molecules like ATP released by dead or dying cells to trigger inflammation, within 12 hours, followed by a T-helper 17 predominant response secreting IL-17A evolving over the first 24 hours [9-11].  IL-6, initially identified as a B-cell differentiation factor, is one of the most widely studied cytokines of the acute trauma response and the rise in IL-6 following injury has been shown to correlate with the severity of the stressor [12,13].  There is a growing body of evidence to suggest that higher levels of IL-6 can be used to predict a dysregulated inflammatory response, including Multiorgan Dysfunction Syndrome (MODS) and Systemic Inflammatory Response Syndrome (SIRS) and its levels remain elevated as it regulates the transition of acute inflammation into a chronic state [14,15]. However, as the inflammatory state is sustained more local cells become sensitive to the effects of IL-6, hence less IL-6 will be needed to propagate the immune effects [16]. Studies have revealed higher synovial fluid concentrations of IL-6 in OA patients but reports on serum IL-6 levels vary between average and elevated, likely due to the variety of patient characteristics [17]. Our study demonstrated a significantly higher concentration of IL-6 secreted by the trauma cohort, which correlates with the existing literature.  

IL-6, together with TGF- β and other cytokines, modulates the IL-17A response in acute and chronic inflammation. While initially identified by its role in chronic inflammation inducing neutrophilia, recruiting T cells and stimulating IL-6, IL-8 and MMP production, emerging studies have identified critical roles for IL-17A in the acute response to trauma, hemorrhage and wound healing as well [18]. A study of 18 blunt trauma patients identified a prominent cytokine response from the pathogenic subset of Th17 cells that secrete IL-17A and inhibit IL-10 production in the non-survivor group which is in line with our data demonstrating higher levels of IL-17A and lower levels of IL-10 in trauma patients [19]. Additionally, the same study showed that the administration of an IL-17 antagonist to mice with hemorrhagic shock could dampen the organ damage. There are conflicting reports on the role of IL-17A in wound healing with some studies identifying it as a regulator of glycolysis promoting regeneration and others suggesting improved re-epithalization in mice when treated with IL-17A antagonists [20]. Considering the time and location limited functions of pleiotropic cytokines such as IL-17A, further studies are needed to identify the inflammatory stage specific roles of IL-17A and the potential benefits of its antagonists in modulating the trauma response.  

IL-17A can also stimulate angiogenesis by promoting the production of VEGF [21,22]. Additionally, VEGF and IL-6 have been implicated in the inhibition of extracellular matrix type II collagen synthesis and in the pathogenesis of OA [23,24]. As IL-6 and VEGF both serve as essential regulators of wound healing, a dysregulated response can result in wound complications and fibrosis [25].  A study comparing the bone marrow secretome via RNA expression analysis from 52 blunt trauma patients with 33 patients undergoing elective hip replacement found a significant downregulation of VEGF expression in the trauma cohort [26]. Our study has not demonstrated a significant difference in VEGF secretion between the 2 groups and this discrepancy between studies might be explained by the different timeframes and levels of fold change required to observe differences in RNA expression, reported in the aforementioned study, versus levels of secreted cytokines, measured by our study.  

IL-8 and IL-6 are signature biomarkers of traumatic injury as elevated levels of both can be used to distinguish trauma-related death from non-traumatic causes [27].  IL-8 is responsible for neutrophil, BMSC and macrophage recruitment, respectively, to the site of acute inflammation [28]. Studies have shown that IL-8 plays a pivotal role in endochondral bone formation following injury. Higher levels of IL-8 in our trauma cohort can be explained by the acute injury inducing its release to stimulate repair. While IL-8 has been implicated in OA pathogenesis via MMP regulation, only synovial concentrations, not plasma levels, have been associated with disease activity [29,30].

Monocyte Chemoattractant Protein-1 (MCP-1), also referred to as Chemokine (CC-motif) Ligand-2(CCL-2), promotes the migration of monocytes and memory T-cells to the site of injury [30]. It is known to stimulate a Th2 response, signified by the production of IL-4 and IL-10 and triggers the release of active substances locally such as histamine from basophils and mast cells [27].  Therefore, MCP-1 plays a critical role in controlling local inflammation and it has been identified as one of the key factors driving synovial disruption in OA [31]. However, it is important to note that baseline levels of MCP-1 can vary due to a wide range of genetic polymorphisms [32]. Additionally, MCP-1 is known to promote endothelial regeneration and angiogenesis after injury while attracting leukocytes and enhancing the migration of monocytes to the subendothelium, contributing to an overall pro-thrombotic effect [32].  Notably, studies have shown an increased risk of venous thromboembolism in trauma patients with higher levels of IL6, IL8 and MCP-1, however it is not yet clear whether this could be the inciting factor or a confounder effect [33].Our results are in support of the view from the literature that MCP-1 levels can be a useful tool in cases of traumatic injury, but further research is needed to clarify its role [34].  

As a cytokine with mostly anti-inflammatory effects that can inhibit IL-1, IL-6 and TNF- α; IL-10 plays a pivotal role in refining the immune response to trauma [27]. IL-6 is an important promoter of its synthesis, therefore, levels of IL-10 are elevated after acute injuries [35,36]. In contrast, serum IL-10 levels were found to be compromised in patients with osteoarthritis, possibly reflecting an inability to counter chronic inflammation [37]. Our findings are in line with data from the literature highlighting the critical role of IL-10 in the acute response to trauma.  

IL-1 β can cause vasodilation, attract granulocytes, cause fever and stimulate a Th17 response [38]. Studies on IL-1 β deficient mice have failed to produce an acute phase response with a paucity of IL-6 production, nausea and fever [39,40]. These findings, in addition to our data, seem to highlight the critical role of IL-1 β as one of the initiators of the acute response to injury.  

The cytokine response to tissue damage results in a highly dynamic serum cytokine profile with various cytokines such as IL-1 β demonstrating transient peaks due to low stability [33]. Therefore, it is important to evaluate the secretome over time to derive data that can guide clinical decisions. Future studies could assess how these cytokines, in addition to TGF- β and TNF- α, which are known to play critical roles in injury response and tissue regeneration, change over time and differ according to patient characteristics including obesity and alcohol exposure, factors associated with a dysregulated inflammatory response [3,14,41]. Additionally, it is important to measure the changes in cytokine levels over time as baseline levels will vary according to patient characteristics; for example, older age and obesity are both associated with higher baseline levels of IL-6 [42]. 

Conclusion

In conclusion, Bone Marrow Stem Cells (BMSCs) are responsible for a majority of the chemical signals that regulate the acute response to injury. Our results underscore the significance of IL-1 β, IL-6, IL-8, IL-10, IL-17A and MCP-1 as key components of the cytokine signature associated with trauma. Additionally, our data suggests that there is a strong correlation between increased MSC pro-inflammatory cytokines levels and trauma, specifically IL-6: involved in acute phase-phase immune response, IL-1 beta: a potent stimulator for the release of other cytokines and immune cells, IL-8: a chemoattractant for neutrophils and IL-17A. Similarly, a correlation between decreased IL-10 – a potent anti-inflammatory cytokine that promotes tissue repair via suppression of pro-inflammatory cytokine expression and trauma was established. However, the specific role of these cytokines may vary depending on the presence of other factors. Further studies will be conducted to establish possible cytokine-cytokine interactions in both kOA and acute injuries. Interventions to modulate the activity of these cytokines can be used to prevent and treat complications associated with dysregulated trauma response. Further studies are needed to understand how these chemical signals can be manipulated to improve treatment outcomes.

Conflict of Interest

The authors report no conflicts of interest. AAS is a paid consultant for Aroa Biologics and on the advisory board for Prytime Medical. JEC is a stockholder for Permaderm, Inc. and SpectralMD and serves as a consultant for Avita and Polynovo Ltd. HAP serves as a consultant for Avita and SpectralMD. PG is a Consultant and paid speaker for Zimmer Biomet and MedExpert. The other authors declare no conflicts of interest.

Funding

This research did not receive any specific grant from funding agencies in the public, commercial or not-for-profit sectors.

Consent for Publication

This research appeared as a poster presentation at the 2024 Annual Shock Society Meeting. 

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