Abstract
BackgroundEven though mesenchymal stromal cells (MSCs) mitigate lung and distal organ damage in experimental polymicrobial sepsis, mortality remains high. We investigated whether preconditioning with eicosapentaenoic acid (EPA) would potentiate MSC actions in experimental sepsis by further decreasing lung and distal organ injury, thereby improving survival.MethodsIn C57BL/6 mice, sepsis was induced by cecal hligation and puncture (CLP); sham-operated animals were used as control. Twenty-four hours after surgery, CLP mice were further randomized to receive saline, adipose tissue-derived (AD)-MSCs (105, nonpreconditioned), or AD-MSCs preconditioned with EPA for 6 h (105, EPA-preconditioned MSCs) intravenously. After 24 h, survival rate, sepsis severity score, lung mechanics and histology, protein level of selected biomarkers in lung tissue, cellularity in blood, distal organ damage, and MSC distribution (by technetium-99m tagging) were analyzed. Additionally, the effects of EPA on the secretion of resolvin-D1 (RvD1), prostaglandin E2 (PGE2), interleukin (IL)-10, and transforming growth factor (TGF)-β1 by MSCs were evaluated in vitro.ResultsNonpreconditioned and EPA-preconditioned AD-MSCs exhibited similar viability and differentiation capacity, accumulated mainly in the lungs and kidneys following systemic administration. Compared to nonpreconditioned AD-MSCs, EPA-preconditioned AD-MSCs further reduced static lung elastance, alveolar collapse, interstitial edema, alveolar septal inflammation, collagen fiber content, neutrophil cell count as well as protein levels of interleukin-1β and keratinocyte chemoattractant in lung tissue, and morphological abnormalities in the heart (cardiac myocyte architecture), liver (hepatocyte disarrangement and Kupffer cell hyperplasia), kidney (acute tubular necrosis), spleen (increased number of megakaryocytes and lymphocytes), and small bowel (villi architecture disorganization). EPA preconditioning of MSCs resulted in increased secretion of pro-resolution and anti-inflammatory mediators (RvD1, PGE2, IL-10, and TGF-β).ConclusionsCompared to nonpreconditioned cells, EPA-preconditioned AD-MSCs yielded further reductions in the lung and distal organ injury, resulting in greater improvement in sepsis severity score and higher survival rate in CLP-induced experimental sepsis. This may be a promising therapeutic approach to improve outcome in septic patients.
Highlights
Even though mesenchymal stromal cells (MSCs) mitigate lung and distal organ damage in experimental polymicrobial sepsis, mortality remains high
eicosapentaenoic acid (EPA) preconditioning of MSCs resulted in increased secretion of pro-resolution and antiinflammatory mediators (RvD1, prostaglandin E2 (PGE2), IL-10, and transforming growth factor (TGF)-β)
EPA preconditioning did not affect adipose tissue-derived (AD)-MSC morphology, cell surface markers, survival, or expression of extracellular vesicles, but increased secretion of biomarkers by AD-MSCs To evaluate whether EPA has any effect on AD-MSC, an in vitro analysis of AD-MSC morphology, cell surface markers, expression of extracellular vesicles, and biomarkers secreted by AD-MSCs was performed
Summary
Ethics statement This study was approved by the Animal Care and Use Committee (CEUA:121/18) of the Health Sciences Center, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil. A total of 152 C57BL/6 mice (144 females and 8 males, weight 25–30 g, age 8–10 weeks), obtained from the animal care facility of the Laboratory of Pulmonary Investigation, Federal University of Rio de Janeiro, were used: 32 females for lung mechanics, histology, biomarker measurement, and distal organ analysis; 32 females for the analysis of blood cellularity; 40 females for the analysis of clinical score and survival rate; 20 females for the analysis of biodistribution in tissues; 20 females to obtain alveolar macrophages; and 8 males as cell donors. Alveolar macrophages were plated with nonpreconditioned or EPApreconditioned AD-MSCs on a 96-well tissue culture plate and incubated in RPMI1640 10% FBS with 1% penicillin/streptomycin for 2 h at 37 °C, in a 5% CO2 atmosphere, for cell adherence. All mice had free access to water and food and were monitored for 7 days to evaluate survival rate During this period, all animals received tramadol hydrochloride (20 mg g−1 body weight) via subcutaneous injection every 6 h for pain management. A two-sided P value < 0.05 was considered significant
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