Background: Exosomes derived from mesenchymal stem cells (MSC-exos) have been demonstrated with great potential in the treatment of multiple human diseases including acute kidney injury (AKI) by virtue of their intrinsic cargoes. However, there are major challenges of low yield and the lack of an established biomanufacturing platform to efficiently produce MSC-exos, thereby limiting their therapeutic application. Here, we aimed to establish a novel strategy to produce MSC-exos with a hollow fiber bioreactor-based three-dimensional (3D) culture system and evaluate the therapeutic efficacy of 3D-exosomes (3D-exos) on AKI. Methods: Mesenchymal stem cells (MSCs) were isolated from fresh human umbilical cord and cultured in twodimensional (2D) flasks. 2 × 10 8 MSCs were inoculated into the hollow fiber bioreactor for 3D culture. The culture supernatants were collected every 1 or 2 days for isolating exosomes. Exosomes from 2D (2D-exos) and 3D cultures were characterized by transmission electron microscopy, nanoparticle tracking analysis, and western blotting analysis of exosome markers. The yield of exosomes from 2 × 10 8 MSCs seeded in 2D and 3D culture system was compared, based on protein quantification. The therapeutic efficacy of 2D-exos and 3D-exos was investigated in a murine model of cisplatin-induced AKI in vivo and in vitro. Results: 3D culture did not significantly change the surface markers of MSCs, as well as the morphology, size, and exosomal markers of 3D-exos when compared to those of 2D-exos. Compared with conventional 2D culture, the 3D culture system increased total exosome production up to 19.4-fold. 3D-exos were more concentrated in the harvested supernatants (15.5-fold) than 2D-exos, which led to a higher exosome collection efficiency of 3D culture system. In vivo, both 2D-exos and 3D-exos significantly alleviated cisplatin-induced murine AKI evidenced by improved renal function, attenuated pathological changes of renal tubules, reduced inflammatory factors, and repressed T cell and macrophage infiltration. Impressively, 3D-exos were more effective than 2D-exos. Moreover, 3D-exos were taken up by tubular epithelial cells (TECs) with improved efficiency, thereby exhibiting superior antiinflammatory effect and improved viability of TECs in vitro.

Transplanted EPCs directly incorporated into the injured pulmonary vessels and maintain the integrity of pulmonary endothelium. Erythropoietin improved the survival and proliferation of transplanted EPCs and recruited them to the injured sites to exert their repairing functions. PI3K/Akt pathway mediated EPO's functions on EPCs. The combination of EPO and EPC treatment may be a promising cell-based therapy for ALI patients.

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The aim of the present study was to investigate the effect and the underlying mechanism of activated protein C (APC) in lipopolysaccharide (LPS) induced lung injury, as well as the potential mechanism. According to the treatment, 50 rats were randomly divided into 5 groups: Control, model (LPS), low-dose group [LPS + 0.1 mg/kg recombined human activated protein C (rhAPC)], median-dose group (LPS + 0.3 mg/kg rhAPC) and high-dose group (LPS + 0.5 mg/kg rhAPC). Then, inflammation in the lung was assessed using hematoxylin and eosin (H&E) staining. Following the collection of bronchoalveolar lavage fluid (BALF), the number of leukocytes and neutrophils in BALF was counted, and superoxide dismutase (SOD) activity was assessed, as well as the expression levels of interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α using ELISA. Subsequently, the expression and phosphorylation of P-38, extracellular signal-regulated kinase (Erk)-1/2, and c-Jun N-terminal kinase (JNK) were estimated using western blotting. Based on H&E staining, rhAPC markedly suppressed inflammatory infiltration in the lung induced by LPS in a dose-dependent manner. In addition, rhAPC also significantly attenuated the accumulation of leptocytes and neutrophils, and the reduction of SOD in BALF induced by LPS in a dose-dependent manner. rhAPC also significantly attenuated the elevation of IL-1β, IL-6 and TNF-α in BALF induced by LPS in a dose-dependent manner. Further mechanistic analysis revealed that rhAPC treatment could evidently attenuate the phosphorylation levels of P-38, Erk1/2 and JNK in the lung induced by LPS in a dose-dependent manner. In conclusion, APC significantly alleviated the lung inflammation induced by LPS by downregulating the phosphorylation of P-38, ERK1/2 and JNK.