Fig. 6.

miR-328-3p contributed to apoptotic body-mediated rescue of impaired MRL/lpr MSCs. a qPCR analysis showed the levels of miR-328-3p, miR-22-3p, miR-124-3p and miR-148a-3p were increased after apoptotic body treatment in MRL/lpr MSCs. b Western blot showed that mimics of miR-328-3p, miR-22-3p, and miR-148a-3p, but not miR-124-3p, reduced the level of Axin1 in apoptotic body-treated MRL/lpr MSCs. c qPCR showed that siRNA knockdown efficiently reduced the levels of miR-328-3p, miR-22-3p and miR-148a-3p in MSCs. d Western blot showed that apoptotic bodies from miR-328-3p-treated MSCs showed reduced capacity to downregulate Axin1 as well as to upregulate active-β-catenin when compared to miR-22-3p and miR-148a-3p groups. e Immunostaining showed that Cy3-miR-328-3p-labeled apoptotic bodies (Red) could be engulfed by MRL/lpr MSCs. f BrdU labeling and continuous passage assay showed that miR-328-3p knockdown partially abolished apoptotic body-induced rescue of the decreased proliferation and population doubling rates in MRL/lpr MSCs, but not as effectively as in the RNF146 knockdown group. g miR-328-3p knockdown partially abolished apoptotic body-induced rescue of mineralized nodule formation, as assessed by alizarin red staining, and expression of Runx2 and ALP, as assessed by Western blot, in MRL/lpr MSCs, but not as effectively as in the RNF146 knockdown group. h miR-328-3p knockdown partially abolished apoptotic body-induced rescue of adipogenic differentiation, as assessed by Oil red O staining, and expression of adipogenic markers, PPARγ and LPL, assessed by Western blot, in MRL/lpr MSCs, but not as effectively as in the RNF146 knockdown group. i Immunostaining showed that MRL/lpr MSCs engulfed both EGFP-RNF146-labeled apoptotic bodies and Cy3-miR-328-3p-labeled apoptotic bodies in an in vitro co-culture system. After intravenous infusion of PKH67-labeled apoptotic bodies for 24 h, immunofluorescent staining showed that both EGFP-RNF146- and Cy3-miR-328-3p-labeled apoptotic bodies co-localized with CD105-positive cells in the femurs of MRL/lpr mice. All results are representative of data generated in three independent experiments. Error bars represent the S.D. from the mean values. **P < 0.01; *P < 0.05; N.S., no significance. Scale bar, 20 μm (e), (i). KD, knockdown