CCR2 Positive Exosome Released by Mesenchymal Stem Cells Suppresses Macrophage Functions and Alleviates Ischemia/Reperfusion-Induced Renal Injury

Figure 2

CCR2 knockdown MSC-derived exosomes failed to bind CCL2. (a) CCR2 knockdown efficacy in the MSCs. A CCR2-specific cocktail of siRNAs (si-CCR2) was transfected into the MSCs for 72 hrs, and a scramble RNA was used as a negative control RNA (NC RNA). Quantitative RT-PCR was used to analyze the changes of mRNA levels in these cells and the change folders for each group compared to untreated MSCs were shown. (b) CCR2 expression levels of the exosomes from the conditional mediums of treated MSCs. The CCR2 protein concentrations were determined by ELISA and the change folders for each group compared to untreated MSCs were shown. (c) CCL2 binding efficiency of exosomes. Recombinant mouse CCL2 protein (100 ng per well) was incubated with the exosomes for 1 hour, as indicated by the groups. Then, the exosome-free (exo-free) medium and exosomes were separated by ultracentrifugation. Then, the free CCL2 protein levels in the medium were determined by ELISA analysis and the ratios of free CCL2/total CCL2 were shown. Bars represent standard error of the mean ± S.D from three repeats. All the significance was determined by ANOVA and Student’s -test. .

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