Exosomes and Microvesicles From Adipose-Derived Mesenchymal Stem Cells Protects the Endothelial Glycocalyx From LPS Injury

Abstract

Introduction: Endothelial glycocalyx damage occurs in numerous pathological conditions and results in endotheliopathy. Extracellular vesicles, including exosomes and microvesicles, isolated from adipose-derived mesenchymal stem cells (ASCs) have therapeutic potential in multiple disease states; however, their role in preventing glycocalyx shedding has not been defined. We hypothesized that ASC-derived exosomes and microvesicles would protect the endothelial glycocalyx from damage by LPS injury in cultured endothelial cells. Methods: Exosomes and microvesicles were collected from ASC conditioned media by centrifugation (10,000g for microvesicles, 100,000g for exosomes). Human umbilical vein endothelial cells (HUVECs) were exposed to 1 μg/mL lipopolysaccharide (LPS). LPS-injured cells (n = 578) were compared with HUVECS with concomitant LPS injury plus 1.0 μg/mL of exosomes (n = 540) or microvesicles (n = 510) for 24 hours. These two cohorts were compared with control HUVECs that received phosphate-buffered saline only (n = 786) and HUVECs exposed to exosomes (n = 505) or microvesicles (n = 500) alone. Cells were fixed and stained with FITC-labeled wheat germ agglutinin to quantify EGX. Real-time quantitative reverse-transcription polymerase chain reaction was used on HUVECs cell lystate to quantify hyaluron synthase-1 (HAS1) expression. Results: Exosomes alone decreased endothelial glycocalyx staining intensity when compared with control (4.94 vs. 6.41 AU, P < 0.001), while microvesicles did not cause a change glycocalyx staining intensity (6.39 vs. 6.41, P = 0.99). LPS injury resulted in decreased glycocalyx intensity as compared with control (5.60 vs. 6.41, P < 0.001). Exosomes (6.85 vs. 5.60, P < 0.001) and microvesicles (6.35 vs. 5.60, P < 0.001) preserved endothelial glycocalyx staining intensity after LPS injury. HAS1 levels were found to be higher in the exosome (1.14 vs. 3.67 RE, P = 0.02) and microvesicle groups (1.14 vs. 3.59 RE, P = 0.02) when compared with LPS injury. Hyaluron synthase-2 and synthase-3 expressions were not different in the various experimental groups. Conclusions: Exosomes alone can damage the endothelial glycocalyx. However, in the presence of LPS injury, both exosomes and microvesicles protect the glycocalyx layer. This effect seems to be mediated by HAS1.

Roles of adipose-derived stem cells and derived exosomes in therapeutic applications to testicular injury caused by cisplatin

“…ADSC-Ex plays a crucial role in intercellular communication, as they can transfer bioactive molecules, such as proteins, lipids, and nucleic acids, to recipient cells [ 24 ]. In recent years, there has been a growing interest in the therapeutic potential of ADSC-Ex in various diseases, including tissue injury, inflammation, and cancer [ 25 ].…”

Dimethyl sulfoxide as a novel therapy in a murine model of acute lung injury

“…Matric metalloproteinases play an important role in the degradation of the glycocalyx during injury. 18,24 The glycocalyx sheddases found to have been affected by DMSO in a hypoxia-reoxygenation injury include MMP24 and MMP25. 4 In the present study, an LPS injury was examined, as HUVECs were injured with LPS.…”

“…Prior research has shown that DMSO can prevent the membrane organization that leads to endothelial glycocalyx damage in hypoxia-reperfusion. Matric metalloproteinases play an important role in the degradation of the glycocalyx during injury 18,24 . The glycocalyx sheddases found to have been affected by DMSO in a hypoxia-reoxygenation injury include MMP24 and MMP25 4 .…”

“…With a significance level ( α ) of 0.05 and power (1 − β ) value of 0.8, a sample number of eight animals per group was required to achieve the 0.8 power value. For cell culture glycocalyx experiments, prior work demonstrating a mean difference of 25% and an SD of 83% of the mean value of glycocalyx staining intensity, 18 a power analysis and sample size determination were performed. With a significance level ( α ) of 0.05 and power (1 − β ) of 0.8, a sample number of 172 cells was required to achieve the 0.8 power value.…”

Destruction of vascular endothelial glycocalyx during formation of pre-metastatic niches

“…Alternatively, the treated cells were stained for 1 h with FITC-WGA (2 μg/mL, 100 μL) to label vascular endothelial glycocalyx, washed twice with PBS, fixed with 4% paraformaldehyde for 15 min, washed twice with PBS, and analyzed under a fluorescence microscope. As a positive control, the cells were stimulated with lipopolysaccharide, which is known to degrade the vascular endothelial glycocalyx [ 40 ].…”