Human umbilical cord mesenchymal stem cell-derived exosomes attenuate neuroinflammation and oxidative stress through the NRF2/NF-κB/NLRP3 pathway.

We investigated whether human umbilical cord mesenchymal stem cell (hUC-MSC)-derived exosomes bear therapeutic potential against lipopolysaccharide (LPS)-induced neuroinflammation. Exosomes were isolated from hUC-MSC supernatant by ultra-high-speed centrifugation and characterized by transmission electron microscopy and western blotting. Inflammatory responses were induced by LPS in BV-2 cells, primary microglial cultures, and C57BL/6J mice. H2 O2 was also used to induce inflammation and oxidative stress in BV-2 cells. The effects of hUC-MSC-derived exosomes on inflammatory cytokine expression, oxidative stress, and microglia polarization were studied by immunofluorescence and western blotting. Treatment with hUC-MSC-derived exosomes significantly decreased the LPS- or H2 O2 -induced oxidative stress and expression of pro-inflammatory cytokines (IL-6 and TNF-α) in vitro, while promoting an anti-inflammatory (classical M2) phenotype in an LPS-treated mouse model. Mechanistically, the exosomes increased the NRF2 levels and inhibited the LPS-induced NF-κB p65 phosphorylation and NLRP3 inflammasome activation. In contrast, the reactive oxygen species scavenger NAC and NF-κB inhibitor BAY 11-7082 also inhibited the LPS-induced NLRP3 inflammasome activation and switched to the classical M2 phenotype. Treatment with the NRF2 inhibitor ML385 abolished the anti-inflammatory and anti-oxidative effects of the exosomes. hUC-MSC-derived exosomes ameliorated LPS/H2 O2 -induced neuroinflammation and oxidative stress by inhibiting the microglial NRF2/NF-κB/NLRP3 signaling pathway.

Che J ，Wang H ，Dong J ，Wu Y ，Zhang H ，Fu L ，Zhang J ... -  《-》

Exosomes derived from human umbilical cord mesenchymal stem cells ameliorate experimental non-alcoholic steatohepatitis via Nrf2/NQO-1 pathway.

No approved effective therapy for non-alcoholic steatohepatitis (NASH) is currently available. Exosomes derived from mesenchymal stem cells (MSCs) perform the functions such as inhibiting inflammation, anti-oxidative stress, regulating immunity, but it is not clear whether human umbilical cord mesenchymal stem cells (hUC-MSCs) exosomes protect against NASH through Nrf2/NQO-1 pathway. Therefore, this study was conducted to investigate the effects of hUC-MSCs exosomes on NASH through Nrf2/NQO-1 pathway in vivo and in vitro. C57BL/6J male mice were fed with high fat and high cholesterol diet (HFHC) and methionine choline deficiency diet (MCD). Mice were treated with or without hUC-MSCs exosomes by tail intravenous injection. The liver histology, lipid metabolism and oxidative stress were evaluated. HepG2 and AML12 cells were incubated with palmitic acid (PA) and MCD conditioned medium, respectively. Then the therapeutic effect of hUC-MSCs exosomes in steatotic cells was evaluated. To elucidate the signaling pathways, the Nrf2-specific blocker ML385 was applied to intervene in vitro. In NASH models, hUC-MSCs exosomes attenuated steatosis in hepatocytes, altered the abnormal expression of lipid-related genes including SREBP-1c, PPAR-α, Fabp5, CPT1α, ACOX and FAS, suppressed the hepatic inflammatory responses by decreasing the expression of F4/80+ macrophages, CD11c+ macrophages as well as the content of TNF-α and IL-6. hUC-MSCs exosomes also inhibited oxidative stress by reducing the level of MDA, CYP2E1 and ROS, increasing the activity of SOD and GSH in hepatocytes. Notably, hUC-MSCs exosomes enhanced the protein ratio of p-Nrf2/Nrf2 and the protein expression of NQO-1. Moreover, in vitro, the therapeutic effects of hUC-MSCs exosomes on lipid deposition and ROS were reversed by ML385. Also, ML385 reduced the protein expression of p-Nrf2 and NQO-1 in vitro. Nrf2/NQO-1 antioxidant signaling pathway may play a key role in the treatment of NASH by hUC-MSCs exosomes.

Kang Y ，Song Y ，Luo Y ，Song J ，Li C ，Yang S ，Guo J ，Yu J ，Zhang X ... -  《-》

Extracellular vesicles derived from mesenchymal stem cells alleviate neuroinflammation and mechanical allodynia in interstitial cystitis rats by inhibiting NLRP3 inflammasome activation.

Neuroinflammation in spinal dorsal horn (SDH) plays an important role in the pathogenesis of interstitial cystitis/bladder pain syndrome (IC/BPS). Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) exert potent anti-inflammatory activities in the treatment of various diseases. This study aimed to determine the therapeutic effects of MSC-EVs on IC and furtherly investigate the potential mechanism to attenuate neuroinflammation. Female IC rat model was established by intraperitoneal injection of cyclophosphamide (50 mg/kg, every 3 days for 3 doses). Inhibition of NLRP3 inflammasome was performed by intraperitoneal injection of MCC950 (10 mg/kg). MSC-EVs were isolated from the culture supernatants of human umbilical cord derived MSCs using ultracentrifugation, and then injected intrathecally into IC rats (20 μg in 10 μl PBS, every other day for 3 doses). Suprapubic mechanical allodynia was assessed using up-down method with von Frey filaments, and micturition frequency was examined by urodynamics. The expression of NLRP3 inflammasome components (NLRP3 and Caspase-1), glial cell markers (IBA-1 and GFAP), proinflammatory cytokines (TNF-α, IL-1β, IL-6 and IL-18) and TLR4/NF-κB signal pathway (TLR4, p65 NK-κB and phospho-p65 NK-κB) in L6-S1 SDH was measured by Western blot analysis. The cellular localization of NLRP3 in SDH was detected using immunofluorescence co-staining. NLRP3 inflammasome was activated in neurons in SDH of IC rats. NLRP3 inflammasome activation contributed to activation of glial cells and process of spinal neuroinflammation in IC rats, and was related to suprapubic mechanical allodynia and frequent micturition. Intrathecal injection of MSC-EVs alleviated suprapubic mechanical allodynia and frequent micturition in IC rats, restrained activation of glial cells and attenuated neuroinflammation in SDH. In addition, MSC-EV treatment significantly inhibited activation of both NLRP3 inflammasomes and TLR4/NF-κB signal pathway. NLRP3 inflammasome activation is involved in the neuroinflammation of IC. Intrathecal injection of MSC-EVs alleviates neuroinflammation and mechanical allodynia in IC by inhibiting the activation of NLRP3 inflammasome, and TLR4/NF-κB signal pathway may be the potential regulatory target.

Zhang C ，Huang Y ，Ouyang F ，Su M ，Li W ，Chen J ，Xiao H ，Zhou X ，Liu B ... -  《Journal of Neuroinflammation》

Mesenchymal Stem Cells-derived Exosomes Ameliorate Lupus by Inducing M2 Macrophage Polarization and Regulatory T Cell Expansion in MRL/lpr Mice.

Sun W ，Yan S ，Yang C ，Yang J ，Wang H ，Li C ，Zhang L ，Zhao L ，Zhang J ，Cheng M ，Li X ，Xu D ... -  《-》

Huc-MSCs-derived exosomes attenuate neuropathic pain by inhibiting activation of the TLR2/MyD88/NF-κB signaling pathway in the spinal microglia by targeting Rsad2.

Mesenchymal stem cells (MSCs)-derived exosomes have shown promise as a cell-free therapeutic strategy for neuropathic pain. This study was conducted to explore the potential mechanisms underlying the analgesic effects of MSC-derived exosomes in treating neuropathic pain. Human umbilical cord MSCs (huc-MSCs)-derived exosomes were isolated and identified. BV-2 microglia were stimulated with lipopolysaccharide (LPS) in the presence or absence of exosomes. Differentially expressed proteins were identified by tandem mass tag (TMT)-based proteomic analysis. The analgesic effects of huc-MSCs-derived exosomes were evaluated in a rat model of chronic constriction injury (CCI). The underlying mechanism was investigated by flow cytometry, RT-qPCR, Western blotting, immunofluorescent staining, and small interfering RNA transfection. In vitro, huc-MSCs-derived exosomes suppressed LPS-induced microglial activation and inhibited activation of the TLR2/MyD88/NF-κB signaling pathway. Based on the proteomic analysis, Rsad2 was identified and confirmed to be down-regulated by huc-MSCs-derived exosomes. Importantly, knockdown of Rsad2 also inhibited microglial activation and restrained activation of the TLR2/MyD88/NF-κB signaling pathway. In vivo, intrathecal injection of exosomes ameliorated CCI-induced mechanical allodynia, down-regulated Rsad2 expression and restrained TLR2/MyD88/NF-κB signaling activation in the spinal microglia. Huc-MSCs-derived exosomes exerted analgesic effects on neuropathic pain by inhibiting activation of the TLR2/MyD88/NF-κB signaling pathway in the spinal microglia. The mechanism underlying these antinociceptive effects involved exosome-mediated interference with Rsad2 expression, thereby inhibiting microglial activation.

Gao X ，Gao LF ，Zhang YN ，Kong XQ ，Jia S ，Meng CY ... -  《-》