The acute respiratory distress syndrome (ARDS) is characterized by disruption of the alveolar-capillary barrier resulting in accumulation of proteinaceous edema and increased inflammatory cells in the alveolar space. We previously found that endothelial progenitor cell (EPC) exosomes prevent endothelial dysfunction and lung injury in sepsis in part due to their encapsulation of miRNA-126. However, the effects of EPC exosomes in acute lung injury (ALI) remain unknown. To determine if EPC exosomes would have beneficial effects in ALI, intratracheal administration of lipopolysaccharide (LPS) was used to induce ALI in mice. Lung permeability, inflammation, and the role of miRNA-126 in the alveolar-epithelial barrier function were examined. The intratracheal administration of EPC exosomes reduced lung injury following LPS-induced ALI at 24 and 48 h. Compared to placebo, intratracheal administration of EPC exosomes significantly reduced the cell number, protein concentration, and cytokines/chemokines in the bronchoalveolar lavage fluid (BALF), indicating a reduction in permeability and inflammation. Further, EPC exosomes reduced myeloperoxidase (MPO) activity, lung injury score, and pulmonary edema, demonstrating protection against lung injury. Murine fibroblast (NIH3T3) exosomes, which do not contain abundant miRNA-126, did not provide these beneficial effects. In human small airway epithelial cells (SAECs), we found that overexpression of miRNA-126-3p can target phosphoinositide-3-kinase regulatory subunit 2 (PIK3R2), while overexpression of miRNA-126-5p inhibits the inflammatory alarmin HMGB1 and permeability factor VEGFα. Interestingly, both miR-126-3p and 5p increase the expression of tight junction proteins suggesting a potential mechanism by which miRNA-126 may mitigate LPS-induced lung injury. Our data demonstrated that human EPC exosomes are beneficial in LPS-induced ALI mice, in part through the delivery of miRNA-126 into the injured alveolus.

Microvascular dysfunction leads to multi-organ failure and mortality in sepsis. Our previous studies demonstrated that administration of exogenous endothelial progenitor cells (EPCs) confers protection in sepsis as evidenced by reduced vascular leakage, improved organ function, and increased survival. We hypothesize that EPCs protect the microvasculature through the exosomes-mediated transfer of microRNAs (miRNAs). Mice were rendered septic by cecal ligation and puncture (CLP), and EPC exosomes were administered intravenously at 4 hr after CLP. EPC exosomes treatment improved survival, suppressing lung and renal vascular leakage, and reducing liver and kidney dysfunction in septic mice. EPC exosomes attenuated sepsis-induced increases in plasma levels of cytokines and chemokine. Moreover, we determined miRNA contents of EPC exosomes with next-generation sequencing and found abundant miR-126-3p and 5p. We demonstrated that exosomal miR-126-5p and 3p suppressed LPS-induced high mobility group box 1 (HMGB1) and vascular cell adhesion molecule 1 (VCAM1) levels, respectively, in human microvascular endothelial cells (HMVECs). Inhibition of microRNA-126-5p and 3p through transfection with microRNA-126-5p and 3p inhibitors abrogated the beneficial effect of EPC exosomes. The inhibition of exosomal microRNA-126 failed to block LPS-induced increase in HMGB1 and VCAM1 protein levels in HMVECs and negated the protective effect of exosomes on sepsis survival. Thus, EPC exosomes prevent microvascular dysfunction and improve sepsis outcomes potentially through the delivery of miR-126.

Pericytes are specialized cells located in close proximity to endothelial cells within the microvasculature. They play a crucial role in regulating blood flow, stabilizing vessel walls, and maintaining the integrity of the blood–brain barrier. The loss of pericytes has been associated with the development and progression of various diseases, such as diabetes, Alzheimer’s disease, sepsis, stroke, and traumatic brain injury. This review examines the detection of pericyte loss in different diseases, explores the methods employed to assess pericyte coverage, and elucidates the potential mechanisms contributing to pericyte loss in these pathological conditions. Additionally, current therapeutic strategies targeting pericytes are discussed, along with potential future interventions aimed at preserving pericyte function and promoting disease mitigation.

Rationale: Endothelial progenitor cells (EPCs) have been associated with human sepsis but their role is incompletely understood. Stromal cell–derived factor (SDF)-1α facilitates EPC recruitment and is elevated in murine sepsis models. Previous studies have demonstrated that the SDF-1α analog CTCE-0214 (CTCE) is beneficial in polymicrobial sepsis induced by cecal ligation and puncture (CLP) in mice. Objectives: We hypothesized that exogenously administered EPCs are also beneficial in CLP sepsis and that CTCE provides synergistic benefit. Methods: Mice were subjected to CLP and administered EPCs at varying doses, CTCE, or a combination of the two. Mouse survival, plasma miRNA expression, IL-10 production, and lung vascular leakage were determined. The in vitro effect of CTCE on miRNA expression and EPC function were determined. Measurements and Main Results: Survival was improved with EPC therapy at a threshold of 106 cells. In coculture studies, EPCs augmented LPS-induced macrophage IL-10 production. In vivo EPC administration in sepsis increased plasma IL-10, suppressed lung vascular leakage, attenuated liver and kidney injury, and augmented miR-126 and -125b expression, which regulate endothelial cell function and/or inflammation. When subthreshold numbers of EPCs were coadministered with CTCE in CLP mice they synergistically improved survival. We demonstrated that CTCE recruits endogenous EPCs in septic mice. In in vitro analysis, CTCE enhanced EPC proliferation, angiogenesis, and prosurvival signaling while inhibiting EPC senescence. These cellular effects were, in part, explained by the effect of CTCE on miR-126, -125b, -34a, and -155 expression in EPCs. Conclusions: EPCs and CTCE represent important potential therapeutic strategies in sepsis.

Summary: β‐arrestins 1 and 2 are ubiquitously expressed proteins that alter signalling by G‐protein‐coupled receptors. β‐arrestin 2 plays an important role as a signalling adaptor and scaffold in regulating cellular inflammatory responses. We hypothesized that β‐arrestin 2 is a critical modulator of inflammatory response in experimental sepsis. β‐arrestin 2(−/−) and wild‐type (WT) mice were subjected to caecal ligation and puncture (CLP). The survival rate was significantly decreased (P < 0·05) in β‐arrestin 2(−/−) mice (13% survival) compared with WT mice (53% survival). A second group of mice were killed 18 hr after CLP for blood, peritoneal lavage and tissue sample collection. CLP‐induced plasma interleukin (IL)‐6 was significantly increased 25 ± 12 fold and caecal myeloperoxidase (MPO) activity was increased 2·4 ± 0·3 fold in β‐arrestin 2(−/−) compared with WT mice. β‐arrestin 2(−/−) mice exhibited more severe lung damage and higher bacterial loads compared with WT mice post CLP challenge as measured by histopathology and colony‐forming unit count. In subsequent experiments, splenocytes, peritoneal macrophages and bone marrow‐derived macrophages (BMDMs) were isolated and cultured from β‐arrestin 2(−/−) and WT mice and stimulated in vitro with lipopolysaccharide (LPS). Tumour necrosis factor (TNF)‐α, IL‐6 and IL‐10 production induced by LPS was significantly augmented (2·2 ± 0·2 fold, 1·8 ± 0·1 fold, and 2·2 ± 0·4 fold, respectively; P < 0·05) in splenocytes from β‐arrestin 2(−/−) mice compared with WT mice. The splenocyte response was different from that of peritoneal macrophages or BMDMs, which exhibited no difference in TNF‐α and IL‐6 production upon LPS stimulation between WT and β‐arrestin 2(−/−) mice. Our data demonstrate that β‐arrestin 2 functions to negatively regulate the inflammatory response in polymicrobial sepsis.