Interleukin-10-engineered mesenchymal stem/stromal cells exhibit robust immunomodulatory effects in vitro and in vivo

Abstract

Introduction  A dysregulated inflammatory response to infection can lead to sepsis, a leading cause of mortality worldwide, and effective anti-inflammatory therapies remain limited. Mesenchymal stem/stromal cells (MSCs) are attractive candidates as immunomodulatory agents. This study evaluated whether genetic modification of MSCs to express interleukin-10 (IL-10), a key anti-inflammatory cytokine, enhances their immunomodulatory effects. Methods  Bone marrow-derived MSCs from C57Bl/6 mice were genetically engineered by lentiviral transduction to express mouse IL-10 (MSC-IL-10). The immunomodulatory activity in vitro was assessed by co-cultures with macrophages stimulated with LPS and IFN-γ, as well as in Con A–stimulated splenocytes. BALB/c mice subjected to lipopolysaccharide (LPS)-induced endotoxemia were treated with vehicle, dexamethasone, wild-type MSCs (MSC-WT), or MSC-IL-10. Survival, plasma cytokines, leukocyte profiles, CD11b⁺ inflammatory cells, and organ histopathology and biodistribution were evaluated in vivo. Results  MSC-IL-10 maintained the mesenchymal phenotype and multipotent characteristics while exhibiting robust IL-10 expression. In in vitro assays, MSC-IL-10 significantly decreased the production of the cytokines TNF-α, IL-1β, IL-6, IL-12 or Nos2 expression by stimulated macrophages or splenocytes, demonstrating superior immunomodulatory effects compared to MSC-WT. In in vivo mice models, MSC-IL-10 significantly reduced systemic pro-inflammatory cytokines, restored circulating leukocyte counts, and attenuated CD11b⁺ (Mac-1 integrin) inflammatory cell recruitment, surpassing MSC-WT-treated groups. Importantly, MSC-IL-10 mitigated tissue damage mainly to lungs and exhibited biodistribution to liver, lungs and spleen in LPS-challenged mice. Conclusions  These results support an enhanced immunomodulatory effect of IL-10-expressing MSCs as a promising cell-based therapeutic approach for sepsis and other inflammatory and immune mediated disorders.

1. Introduction

Inflammation plays a key role in the protection against infections, but also in restoring the body’s homeostasis, as well as repairing, reshaping, and renewing tissues in a range of challenging medical conditions. Infections from Gram-negative bacteria can trigger inflammation by lipopolysaccharides (LPS) activation of toll like receptor 4 (TLR4), which initiates a central inflammatory response mediated by MyD88-dependent and TRIF-dependent intracellular signaling, leading to the activation of the transcription factors NF-κB and IRF3 [1, 20]. Activation of TLR4 results in the production of pro-inflammatory cytokines, chemokines, and vasoactive mediators that promote vasodilation, increased vascular permeability, and the recruitment of leukocytes to the site of infection [5].

When the inflammatory response becomes excessive or poorly regulated, it can culminate in sepsis, which is a complex clinical syndrome characterized by a dysregulated systemic inflammatory response to infection, resulting in potentially fatal organ dysfunction. When sepsis progresses to persistent hypotension and tissue hypoperfusion despite adequate fluid resuscitation, it is defined as septic shock. Endotoxic shock refers specifically to sepsis caused by Gram-negative bacteria, whose cell walls contain LPS, also known as endotoxins [9, 30].

The immune response in sepsis triggers intracellular signaling pathways and production of pro-inflammatory cytokines, especially tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6) [1]. Massive cytokine release causes systemic inflammation, vasodilation, endothelial activation, increased vascular permeability, and leukocyte recruitment, leading to hypotension, tissue injury, and microthrombosis [11, 16]. Microvascular dysfunction and hypotension impair oxygen delivery, resulting in cellular hypoxia, mitochondrial dysfunction, and multi-organ failure, particularly affecting the lungs, liver, and kidneys.