ABSTRACT
INTRODUCTION: Coagulopathy plays an important role in sepsis. The aim of this study was to determine whether bone marrow stromal cell (BMSC) administration could attenuate coagulopathy in sepsis. MATERIALS AND METHODS: In vitro: endothelial cells were cultured with/without BMSCs for 6 h following LPS stimulation and were collected for thrombomodulin (TM) and endothelial protein C receptor (EPCR) measurements. In vivo: Thirty-six mice were randomized into sham, sepsis, and sepsis + BMSC groups (n = 12 each group). Sepsis was induced through cecal ligation and puncture (CLP). BMSC infusion was started at 6 h after CLP. Lung tissues and plasma samples were collected at 24 h after CLP for enzyme-linked immunosorbent assay (ELISA), quantitative real-time RT-PCR, western blot, and immunohistochemistry analysis. RESULTS: In vitro: BMSCs attenuated the decrease in TM and EPCR mRNA and protein expression levels in LPS-stimulated endothelial cells. In vivo: BMSC treatment decreased lung injury and mesenteric perfusion impairment, and ameliorated coagulopathy, as suggested by the reduction in elevated TF, vWF, and TAT circulation levels. BMSC infusion decreased TF mRNA transcription and protein expression levels in lung tissues, and increased TM and EPCR mRNA transcription and expression levels. DISCUSSION: BMSC administration attenuated coagulopathy, and decreased lung injury and mesenteric perfusion impairment in sepsis. Key messages BMSCs increased the expression of TM and EPCR from endothelium cells exposed to LPS in vitro. BMSC treatment attenuated lung injury and coagulopathy in the mice cecal ligation and puncture (CLP) model. BMSC administration-attenuated coagulopathy is related to the reduced expression of TF and increased expression of TM and EPCR.
Subject(s)
Blood Coagulation Disorders/prevention & control , Lung Injury/prevention & control , Mesenchymal Stem Cells/cytology , Sepsis/complications , Animals , Blood Coagulation Disorders/etiology , Blotting, Western , Cells, Cultured , Disease Models, Animal , Endothelial Protein C Receptor , Enzyme-Linked Immunosorbent Assay , Gene Expression Regulation , Lung Injury/etiology , Male , Mice , Mice, Inbred C57BL , Random Allocation , Real-Time Polymerase Chain Reaction , Receptors, Cell Surface/genetics , Thrombomodulin/geneticsABSTRACT
Endothelial cell dysfunction plays an important role in the occurrence and development of sepsis, which is a consequence of the interaction between coagulation and inflammation. Kynurenine (KYN) is an endothelium-derived relaxing factor that makes a large contribution to sepsis pathophysiology. In this study, we investigated the influence of bone marrow mesenchymal stem cells (BMSCs) on KYN production by cultured endothelial cells. KYN and tryptophan (TRP) concentrations in cell supernatants were simultaneously measured with a high-performance liquid chromatography (HPLC) system equipped with a fluorescence detector (FLD) and an ultraviolet detector (UVD). Our results revealed that lipopolysaccharide-stimulated endothelial cells produced more KYN, which was accompanied by a parallel decrease in TRP. When co-cultured with BMSCs, KYN and TRP production were significantly decreased compared to lipopolysaccharide (LPS)-induction alone. Our results suggest that BMSCs can attenuate endothelial cell damage by decreasing KYN as detected with HPLC. This method is the first to be capable of capturing functional changes in the cells and is simple, fast, and suitable for cellular level research purposes.
