Renal ischemia/reperfusion (I/R) injury is the main reason for acute kidney injury (AKI) and is closely related to high morbidity and mortality. In this study, we found that exosomes from human-bone-marrow-derived mesenchymal stem cells (hBMSC-Exos) play a protective role in hypoxia/reoxygenation (H/R) injury.hBMSC-Exos were enriched in miR-199a-3p, and hBMSC-Exo treatment increased the expression level of miR-199a-3p in renal cells. We further explored the function of miR-199a-3p on H/R injury. miR-199a-3p was knocked down in hBMSCs with a miR-199a-3p inhibitor. HK-2 cells cocultured with miR-199a-3p-knockdown hBMSCs were more susceptible to H/R injury and showed more apoptosis than those cocultured with hBMSCs or miR-199a-3poverexpressing hBMSCs. Meanwhile, we found that HK-2 cells exposed to H/R treatment incubated with hBMSC-Exos decreased semaphorin 3A (Sema3A) and activated the protein kinase B (AKT) and extracellular-signal-regulated kinase (ERK) pathways. However, HK-2 cells cocultured with miR-199a-3p-knockdown hBMSCs restored Sema3A expression and blocked the activation of the AKT and ERK pathways. Moreover, knocking down Sema3A could reactivate the AKT and ERK pathways suppressed by a miR-199a-3p inhibitor. In vivo, we injected hBMSC-Exos into mice suffering from I/R injury; this treatment induced functional recovery and histologic protection and reduced cleaved caspase-3 and Sema3A expression levels, as shown by immunohistochemistry. On the whole, this study demonstrated an antiapoptotic effect of hBMSC-Exos, which protected against I/R injury, via delivering miR-199a-3p to renal cells, downregulating Sema3A expression and thereby activating the AKT and ERK pathways. These findings reveal a novel mechanism of AKI treated with hBMSC-Exos and provide a therapeutic method for kidney diseases. K E Y W O R D S apoptosis, exosomes, human-bone-marrow-derived mesenchymal stem cells, ischemia/reperfusion injury, microRNA SUPPORTING INFORMATION Additional supporting information may be found online in the Supporting Information section. How to cite this article: Zhu G, Pei L, Lin F, et al. Exosomes from human-bone-marrow-derived mesenchymal stem cells protect against renal ischemia/reperfusion injury via transferring miR-199a-3p.

Tumour-associated macrophages (TAMs) compose a major component of the tumour microenvironment and form in this microenvironment prior to cancer metastasis. However, the detailed mechanisms of TAM remodelling in the context of bladder cancer have not been clearly defined. The present study collected exosomes from the conditioned medium of human bladder T24 cancer cells. The effects of macrophages treated with exosomes derived from T24 cells on bladder cancer cell migration and invasion were analysed by Transwell assays. The expression levels of endogenous and exosomal microRNA-21 (miR-21) were examined by reverse transcription-quantitative PCR, while the expression level of the target protein was analysed by western blot analysis. Luciferase reporter plasmids and mutants were used to confirm direct targeting. The effects of miR-21 on bladder cancer cell migration and invasion were analysed by Transwell and Matrigel assays following miR-21 transfection. It was identified that exosomes derived from bladder cancer cells polarized THP-1 cell-derived macrophages into the M2 phenotype, and TAM-mediated pro-migratory and pro-invasive activity was determined. Moreover, it was found that miR-21 was highly expressed in exosomes derived from bladder cancer cells as well as in macrophages treated with exosomes. In addition, macrophages transfected with miR-21 exhibited M2 polarization and promoted T24 cell migratory and invasive ability. Mechanistically, exosomal miR-21 derived from bladder cancer cells inhibited phosphatase and tensin homolog activation of the PI3K/AKT signalling pathway in macrophages and enhanced STAT3 expression to promote M2 phenotypic polarization. The present results suggest that exosomal miR-21 can promote cancer progression by polarizing TAMs.

High-mobility group box 1 (HMGB1) has been found to mediate autophagy during chemotherapy in several cancers. However, whether HMGB1plays a role in autophagy and chemoresistance in bladder cancer is elusive. In this report, HMGB1 expression was found to be increased in 30 primary bladder cancer tissue specimens compared to their matched adjacent non-tumor tissues. While gemcitabine induced apoptotic cell death, it also induced HMGB1 expression and autophagy in bladder cancer T24 and BIU-87 cells. Suppressing HMGB1 expression with siRNA strongly potentiated gemcitabine-induced apoptosis. HMGB1 siRNA or autophagy inhibitors suppressed gemcitabine-induced autophagy. Further, gemcitabine activated c-Jun N-terminal kinase (JNK) and extracellular regulated protein kinase (ERK) and Bcl-2 phosphorylation, and blocking ERK and JNK inhibited autophagy and increased apoptosis in gemcitabine-treated cells. Interestingly, suppressing HMGB1 expression attenuated gemcitabine-induced ERK and JNK activation and Bcl-2 phosphorylation. Thus, our results suggest that while gemcitabine kills bladder cancer cells through apoptosis, a cytoprotective autophagy is also induced involving HMGB1-mediated JNK and ERK to counteract the cytotoxicity of gemcitabine, and intervention targeting this pathway may improve the anticancer efficacy of gemcitabine against bladder cancer.

Bone marrow–derived mesenchymal stem cells (BMSCs) have been recently reported to play a variety of vital roles in organ and tissue damage repair, mainly via potent paracrine activity, including secreting extracellular vesicles, such as exosomes, that serve as mediators facilitating intercellular communication and reprogramming recipient cells by delivering their contents to target cells. However, the underlying mechanisms are diverse and complex, and the influencing characteristics have rarely been studied. Accordingly, we designed this study to explore the time dependence of the effects of exosomes derived from BMSCs (BMexos) on renal ischemia‐reperfusion (I/R) injury and the underlying mechanisms associated with the reperfusion time. Impressively, our study is the first to find that BMexos protected against renal I/R injury in vitro and in vivo at the very early reperfusion stages, especially 4–8 h after reperfusion in vitro and 8–16 h after reperfusion in vivo. Interestingly, we simultaneously found that endoplasmic reticulum (ER) stress was significantly suppressed following the administration of BMexos in vitro and in vivo with a similar time dependence. Additionally, we discovered that miR‐199a‐5p, which was abundant in the BMSCs, was transferred into renal tubular epithelial cells (NRK‐52E) in a time‐dependent manner and significantly inhibited I/R‐induced ER stress by targeting binding immunoglobulin protein (BIP). Cocultivation with miR‐199a‐5p‐overexpressing BMSCs amplified the suppression of ER stress and further protected against I/R injury. However, coculture with miR‐199a‐5p‐knockdown BMSCs obviously increased ER stress and reversed the BMexos‐induced protection, and silencing BIP by small interfering RNA–1098 in NRK‐52E inhibited these effects. This study provides evidence that administering BMexos at the very early reperfusion stages significantly protects against renal I/R injury, and ER stress is closely linked to this protection. These results suggest a novel therapeutic strategy during the very early reperfusion stages of renal I/R injury.—Wang, C., Zhu, G., He, W., Yin, H., Lin, F., Gou, X., Li, X. BMSCs protect against renal ischemia‐reperfusion injury by secreting exosomes loaded with miR‐199a‐5p that target BIP to inhibit endoplasmic reticulum stress at the very early reperfusion stages. FASEB J. 33, 5440–5456 (2019). http://www.fasebj.org

The biological characteristics of bladder cancer include enhanced invasion and migration, which are the main causes of death in patients. Starvation is a typical feature of the bladder cancer microenvironment and can induce autophagy. Autophagy has an important relationship with the invasion and migration of tumors. However, the role of autophagy in the invasion and migration of bladder cancer cells remains unclear. Hence, the aim of the current study was to clarify this role and underlying mechanism. In this study, we found that starvation enhanced the epithelial‐mesenchymal transition (EMT)‐mediated invasion and migration of T24 and 5637 cells while inducing autophagy. The inhibition of autophagy with chloroquine (CQ) or 3‐methyladenine (3MA) decreased EMT‐mediated invasion and migration. In addition, the expression of transforming growth factor 1 (TGF‐β1) and phosphorylated Smad3 (p‐Smad3) increased after starvation. The inhibition of autophagy with CQ or 3MA also decreased the expression of TGF‐β1 and p‐Smad3. The inhibitor of TGF‐β receptor sb431542 also inhibited the invasion, migration, and EMT of T24 and 5637 cells during starvation. Furthermore, recombinant TGF‐β1 induced autophagy and inhibition of the TGF‐β/Smad signaling pathway with sb431542 suppressed autophagy. In summary, our results suggested that autophagy promotes the invasion and migration of bladder cancer cells by inducing EMT through the TGF‐β1/Smad3 signaling pathway. Moreover, autophagy and TGF‐β1 can form a positive feedback loop to synergistically promote invasion and migration. Thus, our findings may provide a theoretical basis for the prevention of invasion and migration in bladder cancer.