Impact of PEG sensitization on the efficacy of PEG hydrogel-mediated tissue engineering.

While poly(ethylene glycol) (PEG) hydrogels are generally regarded as biologically inert blank slates, concerns over PEG immunogenicity are growing, and the implications for tissue engineering are unknown. Here, we investigate these implications by immunizing mice against PEG to stimulate anti-PEG antibody production and evaluating bone defect regeneration after treatment with bone morphogenetic protein-2-loaded PEG hydrogels. Quantitative analysis reveals that PEG sensitization increases bone formation compared to naive controls, whereas histological analysis shows that PEG sensitization induces an abnormally porous bone morphology at the defect site, particularly in males. Furthermore, immune cell recruitment is higher in PEG-sensitized mice administered the PEG-based treatment than their naive counterparts. Interestingly, naive controls that were administered a PEG-based treatment also develop anti-PEG antibodies. Sex differences in bone formation and immune cell recruitment are also apparent. Overall, these findings indicate that anti-PEG immune responses can impact tissue engineering efficacy and highlight the need for further investigation.

Rescuing Photoreceptors in RPE Dysfunction-Driven Retinal Degeneration: The Role of Small Extracellular Vesicles Secreted from Retinal Pigment Epithelium.

Dysfunction of the retinal pigment epithelium (RPE) is a common shared pathology in major degenerative retinal diseases despite variations in the primary etiologies of each disease. Due to their demanding and indispensable functional roles throughout the lifetime, RPE cells are vulnerable to genetic predisposition, external stress, and aging processes. Building upon recent advancements in stem cell technology for differentiating healthy RPE cells and recognizing the significant roles of small extracellular vesicles (sEV) in cellular paracrine and autocrine actions, we investigated the hypothesis that the RPE-secreted sEV alone can restore essential RPE functions and rescue photoreceptors in RPE dysfunction-driven retinal degeneration. Our findings support the rationale for developing intravitreal treatment of sEV. We demonstrate that intravitreally delivered sEV effectively penetrate the full thickness of the retina. Xenogenic intraocular administration of human-derived EVs did not induce acute immune reactions in rodents. sEV derived from human embryonic stem cell (hESC)-derived fully differentiated RPE cells, but not sEV-depleted conditioned cell culture media (CCM minus sEV), rescued photoreceptors and their function in a Royal College of Surgeons (RCS) rat model. This model is characterized by photoreceptor death and retinal degeneration resulting from a mutation in the MerTK gene in RPE cells. From the bulk RNA sequencing study, we identified 447 differently expressed genes in the retina after hESC-RPE-sEV treatment compared with the untreated control. Furthermore, 394 out of 447 genes (88%) showed a reversal in expression toward the healthy state in Long-Evans (LE) rats after treatment compared to the diseased state. Particularly, detrimental alterations in gene expression in RCS rats, including essential RPE functions such as phototransduction, vitamin A metabolism, and lipid metabolism were partially reversed. Defective photoreceptor outer segment engulfment due to intrinsic MerTK mutation was partially ameliorated. These findings suggest that RPE-secreted sEV may play a functional role similar to that of RPE cells. Our study justifies further exploration to fully unlock future therapeutic interventions with sEV in a broad array of degenerative retinal diseases.

Serum Extracellular Vesicle Protein Profiling for Prediction of Corneal Transplant Rejection.

BACKGROUND: Corneal transplantation is the most common transplant procedure worldwide. Despite immune and angiogenic privilege of the cornea, 50% to 70% of corneal transplants fail in high-risk recipients, primarily because of immune rejection. Therefore, it is crucial to identify predictive biomarkers of rejection to improve transplant survival. METHODS: In search for predictive biomarkers, we performed proteomics analysis of serum extracellular vesicles (EVs) in a fully major histocompatibility complex-mismatched (C57BL/6-to-BALB/c) murine corneal transplantation model, wherein 50% of transplants undergo rejection by day 28 following transplantation. RESULTS: Our time course study revealed a decrease in the number of serum EVs on day 1, followed by a gradual increase by day 7. A comparative analysis of proteomics profiles of EVs from transplant recipients with rejection (rejectors) and without rejection (nonrejectors) found a distinct enrichment of histocompatibility 2, Q region locus 2, which is a part of major histocompatibility complex-class I of donor C57BL/6 mice, in day 7 EVs of rejectors, compared with nonrejectors, syngeneic controls, or naïve mice. In contrast, serum amyloid A2, a protein induced in response to injury, was increased in day 7 EVs of nonrejectors. CONCLUSIONS: Our findings offer noninvasive EV-based potential biomarkers for predicting corneal allograft rejection or tolerance.

Scalable manufacture of therapeutic mesenchymal stromal cell products on customizable microcarriers in vertical wheel bioreactors that improve direct visualization, product harvest, and cost.

BACKGROUND AIMS: Human mesenchymal stromal cells (hMSCs) and their secreted products show great promise for treatment of musculoskeletal injury and inflammatory or immune diseases. However, the path to clinical utilization is hampered by donor-tissue variation and the inability to manufacture clinically relevant yields of cells or their products in a cost-effective manner. Previously we described a method to produce chemically and mechanically customizable gelatin methacryloyl (GelMA) microcarriers for culture of hMSCs. Herein, we demonstrate scalable GelMA microcarrier-mediated expansion of induced pluripotent stem cell (iPSC)-derived hMSCs (ihMSCs) in 500 mL and 3L vertical wheel bioreactors, offering several advantages over conventional microcarrier and monolayer-based expansion strategies. METHODS: Human mesenchymal stromal cells derived from induced pluripotent cells were cultured on custom-made spherical gelatin methacryloyl microcarriers in single-use vertical wheel bioreactors (PBS Biotech). Cell-laden microcarriers were visualized using confocal microscopy and elastic light scattering methodologies. Cells were assayed for viability and differentiation potential in vitro by standard methods. Osteogenic cell matrix derived from cells was tested in vitro for osteogenic healing using a rodent calvarial defect assay. Immune modulation was assayed with an in vivo peritonitis model using Zymozan A. RESULTS: The optical properties of GelMA microcarriers permit noninvasive visualization of cells with elastic light scattering modalities, and harvest of product is streamlined by microcarrier digestion. At volumes above 500 mL, the process is significantly more cost-effective than monolayer culture. Osteogenic cell matrix derived from ihMSCs expanded on GelMA microcarriers exhibited enhanced in vivo bone regenerative capacity when compared to bone morphogenic protein 2, and the ihMSCs exhibited superior immunosuppressive properties in vivo when compared to monolayer-generated ihMSCs. CONCLUSIONS: These results indicate that the cell expansion strategy described here represents a superior approach for efficient generation, monitoring and harvest of therapeutic MSCs and their products.

TWIST1 and TSG6 are coordinately regulated and function as potency biomarkers in human MSCs.

Mesenchymal stem/stromal cells (MSCs) have been evaluated in >1500 clinical trials, but outcomes remain suboptimal because of knowledge gaps in quality attributes that confer potency. We show that TWIST1 directly represses TSG6 expression that TWIST1 and TSG6 are inversely correlated across bone marrow-derived MSC (BM-MSC) donor cohorts and predict interdonor differences in their proangiogenic, anti-inflammatory, and immune suppressive activity in vitro and in sterile inflammation and autoimmune type 1 diabetes preclinical models. Transcript profiling of TWIST1HiTSG6Low versus TWISTLowTSG6Hi BM-MSCs revealed previously unidentified roles for TWIST1/TSG6 in regulating cellular oxidative stress and TGF-ß2 in modulating TSG6 expression and anti-inflammatory activity. TWIST1 and TSG6 levels also correlate to donor stature and predict differences in iPSC-derived MSC quality attributes. These results validate TWIST1 and TSG6 as biomarkers that predict interdonor differences in potency across laboratories and assay platforms, thereby providing a means to manufacture MSC products tailored to specific diseases.

Inhibitory Effects of Extracellular Vesicles from iPS-Cell-Derived Mesenchymal Stem Cells on the Onset of Sialadenitis in Sjögren's Syndrome Are Mediated by Immunomodulatory Splenocytes and Improved by Inhibiting miR-125b.

Extracellular vesicles (EVs) from allogeneic-tissue-derived mesenchymal stem cells (MSCs) are promising to improve Sjögren's syndrome (SS) treatment, but their application is hindered by high variations in and limited expandability of tissue MSCs. We derived standardized and scalable MSCs from iPS cells (iMSCs) and reported that EVs from young but not aging iMSCs (iEVs) inhibited sialadenitis onset in SS mouse models. Here, we aim to determine cellular mechanisms and optimization approaches of SS-inhibitory effects of iEVs. In NOD.B10.H2b mice at the pre-disease stage of SS, we examined the biodistribution and recipient cells of iEVs with imaging, flow cytometry, and qRT-PCR. Intravenously infused iEVs accumulated in the spleen but not salivary glands or cervical lymph nodes and were mainly taken up by macrophages. In the spleen, young but not aging iEVs increased M2 macrophages, decreased Th17 cells, and changed expression of related immunomodulatory molecules. Loading miR-125b inhibitors into aging iEVs significantly improved their effects on repressing sialadenitis onset and regulating immunomodulatory splenocytes. These data indicated that young but not aging iEVs suppress SS onset by regulating immunomodulatory splenocytes, and inhibiting miR-125b in aging iEVs restores such effects, which is promising to maximize production of effective iEVs from highly expanded iMSCs for future clinical application.

MHC Class I Enables MSCs to Evade NK-Cell-Mediated Cytotoxicity and Exert Immunosuppressive Activity.

Allogeneic mesenchymal stem/stromal cells (MSCs) are frequently used in clinical trials due to their low expression of major histocompatibility complex (MHC) class I and lack of MHC class II. However, the levels of MHC classes I and II in MSCs are increased by inflammatory stimuli, raising concerns over potential adverse effects associated with allogeneic cell therapy. Also, it is unclear how the host immune response to MHC-mismatched MSCs affects the therapeutic efficacy of the cells. Herein, using strategies to manipulate MHC genes in human bone marrow-derived MSCs via the CRISPR-Cas9 system, plasmids, or siRNAs, we found that inhibition of MHC class I-not MHC class II-in MSCs lowered the survival rate of MSCs and their immunosuppressive potency in mice with experimental autoimmune uveoretinitis, specifically by increasing MSC vulnerability to natural killer (NK)-cell-mediated cytotoxicity. A subsequent survey of MSC batches derived from 6 human donors confirmed a significant correlation between MSC survival rate and susceptibility to NK cells with the potency of MSCs to increase MHC class I level upon stimulation. Our overall results demonstrate that MHC class I enables MSCs to evade NK-cell-mediated cytotoxicity and exert immunosuppressive activity.

Identification of Molecules Responsible for Therapeutic Effects of Extracellular Vesicles Produced from iPSC-Derived MSCs on Sjo¨gren's Syndrome.

Recent research indicated that extracellular vesicles (EVs) derived from mesenchymal stem/stromal cells (MSCs) are a promising alternative to MSCs for immunomodulatory therapy. However, the contents of MSC-EVs would change as their parent MSCs change, hence the therapeutic efficacy of MSC-derived EVs (MSC-EVs) would largely depend on donors, tissue sources and culture conditions of MSCs. To overcome limitations of tissue-derived MSCs, we previously used MSCs derived from human induced pluripotent stem cells (iMSCs) to produce EVs and demonstrated their therapeutic potential in a mouse model of secondary Sjo¨gren's Syndrome. Here, we further found that EVs from early-passage iMSCs had better immunomodulatory potency than EVs from late-passage iMSCs in TLR4-stimulated splenocytes and in a mouse model of primary Sjögren's syndrome. Comparative molecular profiling using proteomics and microRNA sequencing revealed distinctive molecular profiles of iMSC-EVs with or without immunomodulation capacity. Amongst them, manipulation of TGF-ß1, miR-21 and miR-125b levels in iMSC-EVs significantly affected their immunosuppressive effects. These findings would help improve our understanding of the molecular mechanism underlying iMSC-EV-mediated immunomodulation and further provide strategies to improve regulatory function of EVs for the treatment of immune-mediated diseases.

A scalable system for generation of mesenchymal stem cells derived from induced pluripotent cells employing bioreactors and degradable microcarriers.

Human mesenchymal stem cells (hMSCs) are effective in treating disorders resulting from an inflammatory or heightened immune response. The hMSCs derived from induced pluripotent stem cells (ihMSCs) share the characteristics of tissue derived hMSCs but lack challenges associated with limited tissue sources and donor variation. To meet the expected future demand for ihMSCs, there is a need to develop scalable methods for their production at clinical yields while retaining immunomodulatory efficacy. Herein, we describe a platform for the scalable expansion and rapid harvest of ihMSCs with robust immunomodulatory activity using degradable gelatin methacryloyl (GelMA) microcarriers. GelMA microcarriers were rapidly and reproducibly fabricated using a custom microfluidic step emulsification device at relatively low cost. Using vertical wheel bioreactors, 8.8 to 16.3-fold expansion of ihMSCs was achieved over 8 days. Complete recovery by 5-minute digestion of the microcarriers with standard cell dissociation reagents resulted in >95% viability. The ihMSCs matched or exceeded immunomodulatory potential in vitro when compared with ihMSCs expanded on monolayers. This is the first description of a robust, scalable, and cost-effective method for generation of immunomodulatory ihMSCs, representing a significant contribution to their translational potential.

Mesenchymal stromal cells for the treatment of ocular autoimmune diseases.

Mesenchymal stromal cells, commonly referred to as MSCs, have emerged as a promising cell-based therapy for a range of autoimmune diseases thanks to several therapeutic advantages. Key among these are: 1) the ability to modulate innate and adaptive immune responses and to promote tissue regeneration, 2) the ease of their isolation from readily accessible tissues and expansion at scale in culture, 3) their low immunogenicity enabling use as an allogeneic "off-the-shelf" product, and 4) MSC therapy's safety and feasibility in humans, as demonstrated in more than one thousand clinical trials. Evidence from preclinical studies and early clinical trials indicate the therapeutic potential of MSCs and their derivatives for efficacy in ocular autoimmune diseases such as autoimmune uveoretinitis and Sjögren's syndrome-related dry eye disease. In this review, we provide an overview of the current understanding of the therapeutic mechanisms of MSCs, and summarize the results from preclinical and clinical studies that have used MSCs or their derivatives for the treatment of ocular autoimmune diseases. We also discuss the challenges to the successful clinical application of MSC therapy, and suggest strategies for overcoming them.

Mesenchymal stromal cells for corneal transplantation: Literature review and suggestions for successful clinical trials.

Corneal transplantation is a routine procedure for patients with corneal blindness. Despite the streamlining of surgical techniques and deeper understanding of the cellular and molecular pathways mediating rejection, corticosteroids are still the main immunosuppressive regimen in corneal transplantation, and the 15-year survival of corneal transplants remains as low as 50%, which is poorer than that for most solid organ transplants. Recently, mesenchymal stromal cells (MSCs) with unique regenerative and immune-modulating properties have emerged as a promising cell therapy to promote transplant tolerance, minimize the use of immunosuppressants, and prevent chronic rejection. Here, we review the literature on preclinical studies of MSCs for corneal transplantation and summarize the key findings from clinical trials with MSCs in solid organ transplantation. Finally, we highlight current issues and challenges regarding MSC therapies and suggest strategies for safe and effective MSC-based therapies in clinical transplantation.

Comprehensive Molecular Profiles of Functionally Effective MSC-Derived Extracellular Vesicles in Immunomodulation.

Accumulating evidence indicates that mesenchymal stem/stromal cell-derived extracellular vesicles (MSC-EVs) exhibit immunomodulatory effects by delivering therapeutic RNAs and proteins; however, the molecular mechanism underlying the EV-mediated immunomodulation is not fully understood. In this study, we found that EVs from early-passage MSCs had better immunomodulatory potency than did EVs from late-passage MSCs in T cell receptor (TCR)- or Toll-like receptor 4 (TLR4)-stimulated splenocytes and in mice with ocular Sjögren's syndrome. Moreover, MSC-EVs were more effective when produced from 3D culture of the cells than from the conventional 2D culture. Comparative molecular profiling using proteomics and microRNA sequencing revealed the enriched factors in MSC-EVs that were functionally effective in immunomodulation. Among them, manipulation of transforming growth factor ß1 (TGF-ß1), pentraxin 3 (PTX3), let-7b-5p, or miR-21-5p levels in MSCs significantly affected the immunosuppressive effects of their EVs. Furthermore, there was a strong correlation between the expression levels of TGF-ß1, PTX3, let-7b-5p, or miR-21-5p in MSC-EVs and their suppressive function. Therefore, our comparative strategy identified TGF-ß1, PTX3, let-7b-5p, or miR-21-5p as key molecules mediating the therapeutic effects of MSC-EVs in autoimmune disease. These findings would help understand the molecular mechanism underlying EV-mediated immunomodulation and provide functional biomarkers of EVs for the development of robust EV-based therapies.

In Vitro Macrophage Assay Predicts the In Vivo Anti-inflammatory Potential of Exosomes from Human Mesenchymal Stromal Cells.

Extracellular vesicles (EVs) play key roles in cell biology and may provide new clinical diagnostics and therapies. However, it has proven difficult to develop protocols for their purification and characterization. One of the major barriers in the field has been a lack of convenient assays for their bioactivity. Developing assays has not been a trivial matter, because of the heterogeneity of EVs, the multiple activities they demonstrate, and the uncertainty about their modes of action. Therefore, it is likely that multiple assays for their activities are needed. One important assay will be for the anti-inflammatory activity observed in mice after administration of the small EVs commonly referred to as exosomes. We developed an assay for the anti-inflammatory activity of exosomes with a line of mouse macrophages. The assay makes it possible to rank different preparations of exosomes by their anti-inflammatory activity, and their ranking predicts their efficacy in suppressing LPS-stimulated inflammation in mice. The assay is convenient for comparing multiple samples and, therefore, should be useful in developing protocols for the purification and characterization of anti-inflammatory exosomes.

Inhibitory Effects of iPSC-MSCs and Their Extracellular Vesicles on the Onset of Sialadenitis in a Mouse Model of Sjögren's Syndrome.

No effective treatment for Sjögren's syndrome (SS), a chronic autoimmune disease affecting mainly salivary and lacrimal glands, is available now. Systemic infusion of allogeneic mesenchymal stem cells (MSCs) isolated from tissues such as bone marrow (BM) alleviated SS in mouse models and a small clinical trial, but further research and application of this MSC therapy were hindered by limited expandability, significant donor variations, and safety concerns of tissue-derived MSCs. To circumvent these issues, we derived MSCs from human iPSCs using an optimized protocol that can be easily scaled up to produce a huge amount of standardized MSCs. Our iPSC-MSCs inhibited the onset of lymphocyte infiltration into salivary glands in the NOD mouse model of SS in the same way as BM-MSCs. Extracellular vesicles (EVs) carry bioactive molecules in the same way as their originating cells and are more stable and considered much safer than cells for therapies. We found that EVs derived from BM-MSCs and iPSC-MSCs suppressed activation of immune cells and expression of proinflammation factors essential for SS progression in vitro and that infusion of iPSC-MSC EVs at the predisease stage decreased the lymphocyte infiltration in salivary glands and serum autoantibody levels in the same way as infusion of BM-MSCs and iPSC-MSCs. These data suggested that iPSC-MSC EVs have the potential to prevent the progression of SS before the onset of sialadenitis.

Mesenchymal Stromal Cells Inhibit Inflammatory Lymphangiogenesis in the Cornea by Suppressing Macrophage in a TSG-6-Dependent Manner.

The cornea is a transparent tissue devoid of blood and lymphatic vessels. However, various inflammatory conditions can cause hemangiogenesis and lymphangiogenesis in the cornea, compromising transparency and visual acuity. Mesenchymal stem/stromal cells (MSCs) have therapeutic potentials in a variety of diseases because of anti-inflammatory properties. Herein, we investigated the effects of MSCs on corneal angiogenesis using a model of suture-induced inflammatory corneal neovascularization. Data demonstrated that an intravenous administration of MSCs suppressed corneal inflammation and neovascularization, inhibiting both hemangiogenesis and lymphangiogenesis. MSCs reduced the levels of vascular endothelial growth factor (VEGF)-C, VEGF-D, Tek, MRC1, and MRC2 in the cornea, which are expressed by pro-angiogenic macrophages. Moreover, the number of CD11b+ monocytes/macrophages in the cornea, spleen, peripheral blood, and draining lymph nodes was decreased by MSCs. Depletion of circulating CD11b+ monocytes by blocking antibodies replicated the effects of MSCs. Importantly, knockdown of tumor necrosis factor alpha (TNF-α)-stimulated gene/protein 6 (TSG-6) in MSCs abrogated the effects of MSCs in inhibiting corneal hemangiogenesis and lymphangiogenesis and monocyte/macrophage infiltration. Together, the results suggest that MSCs inhibit inflammatory neovascularization in the cornea by suppressing pro-angiogenic monocyte/macrophage recruitment in a TSG-6-dependent manner.

Data against a Common Assumption: Xenogeneic Mouse Models Can Be Used to Assay Suppression of Immunity by Human MSCs.

Much of what we know about immunology suggests that little is to be gained from experiments in which human cells are administered to immunocompetent mice. Multiple reports have demonstrated that this common assumption does not hold for experiments with human mesenchymal stem/stromal cells (hMSCs). The data demonstrate that hMSCs can suppress immune responses to a variety of stimuli in immunocompetent mice by a range of different mechanisms that are similar to those employed by mouse MSCs. Therefore, further experiments with hMSCs in mice will make it possible to generate preclinical data that will improve both the efficacy and safety of the clinical trials with the cells that are now in progress.

MSC-derived Extracellular Vesicles Attenuate Immune Responses in Two Autoimmune Murine Models: Type 1 Diabetes and Uveoretinitis.

Accumulating evidence shows that extracellular vesicles (EVs) produced by mesenchymal stem/stromal cells (MSCs) exert their therapeutic effects in several disease models. We previously demonstrated that MSCs suppress autoimmunity in models of type 1 diabetes (T1D) and experimental autoimmune uveoretinitis (EAU). Therefore, here, we investigated the therapeutic potential of MSC-derived EVs using our established mouse models for autoimmune diseases affecting the pancreas and the eye: T1D and EAU. The data demonstrate that MSC-derived EVs effectively prevent the onset of disease in both T1D and EAU. In addition, the mixed lymphocyte reaction assay with MSC-derived EVs indicated that EVs inhibit activation of antigen-presenting cells and suppress development of T helper 1 (Th1) and Th17 cells. These results raise the possibility that MSC-derived EVs may be an alternative to cell therapy for autoimmune disease prevention.

Mechanisms of mesenchymal stem/stromal cell function.

The past decade has seen an explosion of research directed toward better understanding of the mechanisms of mesenchymal stem/stromal cell (MSC) function during rescue and repair of injured organs and tissues. In addition to delineating cell-cell signaling and molecular controls for MSC differentiation, the field has made particular progress in defining several other mechanisms through which administered MSCs can promote tissue rescue/repair. These include: 1) paracrine activity that involves secretion of proteins/peptides and hormones; 2) transfer of mitochondria by way of tunneling nanotubes or microvesicles; and 3) transfer of exosomes or microvesicles containing RNA and other molecules. Improved understanding of MSC function holds great promise for the application of cell therapy and also for the development of powerful cell-derived therapeutics for regenerative medicine. Focusing on these three mechanisms, we discuss MSC-mediated effects on immune cell responses, cell survival, and fibrosis and review recent progress with MSC-based or MSC-derived therapeutics.

Stanniocalcin-1 attenuates ischemic cardiac injury and response of differentiating monocytes/macrophages to inflammatory stimuli.

Stanniocalcin-1 (STC-1) is a multifunctional glycoprotein with antioxidant and anti-inflammatory properties. Ischemic myocardial necrosis generates "danger" signals that perpetuate detrimental inflammatory reactions often involving monocyte recruitment and their subsequent differentiation into proinflammatory macrophages. Therefore, we evaluated the effects of recombinant STC-1 (rSTC-1) on monocyte phenotype and in a mouse model of myocardial infarction. Using an established protocol to differentiate human monocytes into macrophages, we demonstrated that rSTC-1 did not alter morphology of the differentiated cells, toll-like receptor (TLR) 4 expression, or expression of the myeloid cell marker CD11b. However, rSTC-1 treatment before differentiation attenuated the rise in the expression of CD14, a TLR4 coreceptor and pathogen sensor that propagates innate immune responses, and suppressed levels of inflammatory cytokines produced by the differentiated cells in response to the CD14-TLR4 ligand lipopolysaccharide. Moreover, rSTC-1 treatment reduced CD14 expression in monocytes stimulated with endogenous danger signals. Interestingly, the effects of rSTC-1 on CD14 expression were not reproduced by a superoxide dismutase mimetic. In mice with induced myocardial infarcts, intravenous administration of rSTC-1 decreased CD14 expression in the heart as well as levels of tumor necrosis factor alpha, C-X-C motif ligand 2, interleukin 1 beta, and myeloperoxidase. It also suppressed the formation of scar tissue while enhancing cardiac function. The data suggests that one of the beneficial effects of STC-1 might be attributed to suppression of CD14 on recruited monocytes and macrophages that limits their inflammatory response. STC-1 may be a promising therapy to protect the heart and other tissues from ischemic injury.