Abstract:
INTRODUCTION: Dysregulation of the inflammatory response can trigger a state of systemic hyperinflammation, as observed in sepsis—a severe clinical condition with high lethality, characterized by an uncontrolled inflammatory response to infection, with challenging diagnosis and treatment. Given the limitations of available therapies, innovative approaches such as the use of genetically modified mesenchymal stromal cells (MSCs) aimed at enhancing their biological activities are being explored in the context of sepsis. Among the molecular targets is the leukemia inhibitory factor (LIF), a pleiotropic cytokine with anti-inflammatory and regenerative properties. Although its mechanisms are not yet fully elucidated, the overexpression of LIF in MSCs emerges as a promising strategy to amplify their immunomodulatory effects in systemic inflammation. OBJECTIVE: To evaluate the therapeutic effect of MSCs overexpressing leukemia inhibitory factor (MSC-LIF) in a model of endotoxemic shock induced by lipopolysaccharide (LPS). MATERIAL AND METHODS: Initially, in vitro assays were conducted using murine peritoneal macrophages stimulated with LPS and interferon-gamma (IFN-γ) and treated with unmodified MSCs (MSC-WT) or MSC-LIF. Immunomodulatory activity was assessed by quantifying nitric oxide (via the Griess method) and cytokines TNF-α, IL-6, IL-12, and IL-10 in the supernatant using ELISA. For in vivo experiments, male C57BL/6 mice were subjected to an endotoxemic shock model via intraperitoneal injection of LPS. Thirty minutes after the challenge, animals were treated with MSC-WT, MSC-LIF, dexamethasone, or glucose solution (control). Survival was monitored for four days. Hematological and biochemical analyses, as well as serum cytokine quantification (TNF-α, IL-1β, IL-6, IL-12, and IL-10), were performed to evaluate the systemic inflammatory response. RESULTS: In vitro assays, MSC-LIF inhibited nitric oxide production by 95% at a concentration of 2×10⁵ cells and modulated the inflammatory profile by reducing TNF-α and IL-12 and increasing IL-6 and IL-10 levels compared to the positive control. In the endotoxemic shock model, the dose of 1×10⁶ MSC-LIF ensured 100% survival, while MSC-WT resulted in 50% survival. The lower dose of MSC-LIF (2.5×10⁵) also maintained survival above 50%, suggesting a potential protective effect. In the evaluation of serum cytokines, the MSC-LIF treated group showed increased IL-10 levels. A reduction in the inflammatory cytokines IL-1β, IL-12, and TNF-α was also observed compared to controls. Regarding hematological parameters, thrombocytopenia was attenuated in animals treated with MSC-LIF and dexamethasone, with platelet counts approaching those of the naive group (p < 0.05). Additionally, biochemical parameters such as creatinine and urea were normalized in the treated groups, suggesting preservation of renal function in the face of systemic inflammatory insult. CONCLUSION: LIF overexpression enhanced the immunomodulatory action of MSCs, contributing to improved survival and inflammation modulation in an endotoxemia model.
