Obesity Blunts the Effect of Mesenchymal Stem Cell-Derived Extracellular Vesicles

Figure 1. Effects of obese-MSC-EVs and lean-MSC-EVs on kidney volume, oxygenation, and perfusion. (a) Tracking showed that the ratios of lean- and obese-MSC-EV numbers were similar in the STK and CLK. (b) The volume, (c–d) oxygenation, and perfusion (e) assessed by MRI were all lower in the STK compared to the sham+vehicle kidneys. Lean- and obese-MSC-EVs improved the STK volume to a similar degree. Lean-MSC-EVs rescued the elevated medullary hypoxia (P < 0.05) but not cortical hypoxia. Neither lean- nor obese-MSC-EVs restored the decreased perfusion. (f) RAS surgery and MSC-EVs treatment did not have significant effects on the volume, oxygenation, or perfusion of the CLK. ∗P < 0.05; ∗∗P < 0.01. BOLD, blood oxygen level-dependent MRI; CLK, contralateral kidney; MRI, magnetic resonance imaging; MSC-EVs, mesenchymal stem/stromal cell-derived extracellular vesicles; ; ns, no significant difference; RAS, renal artery stenosis; STK, stenotic kidney.

Figure 2. Effects of obese-MSC-EVs and lean-MSC-EVs on blood pressure, plasma renin content, plasma creatinine, and histological changes in the stenotic mouse kidney. (a) Blood pressure increased 2 weeks after RAS surgery (P < 0.01); by 2 weeks after injection, systolic blood pressure (SBP) decreased by both lean- and obese-MSC-EVs (P < 0.05), and diastolic blood pressure (DBP) only by lean-MSC-EVs (P < 0.05). (b) Plasma renin was decreased by both types of EVs (P < 0.01), suggesting the regulation of the renin-angiotensin-aldosterone system. (c,f) Masson’s trichrome staining assessed renal fibrosis and only lean-MSC-EVs rescued the elevated fibrosis in STK (P < 0.05). (d, f) Only lean-MSC-EVs attenuated tubular injury score in RAS (P < 0.05), evaluated based on H&E staining. (e) Plasma creatinine was elevated by RAS (P < 0.01) and unchanged by EVs. ∗P < 0.05; ∗∗P < 0.01. H&E, hematoxylin and eosin; MSC-EVs, mesenchymal stem/stromal cell-derived extracellular vesicles; STK, stenotic kidney.

Figure 3. Effects of obese-MSC-EVs and lean-MSC-EVs on inflammation and oxidative damage. (a)The mRNA of IL-1β, IL-6, and TNF-α was quantified by quantitative polymerase chain reaction to assess the anti-inflammation effect of EVs. Both lean- and obese-MSC-EVs rescued the elevated TNF-α and IL-6 in the STK whereas only lean-MSC-EVs decreased IL-1β (P < 0.01). (b–d) The immunoreactivity of 8-OHdG and HIF-1α was detected by immunohistochemistry. 8-OHdG was downregulated by both lean and obese-MSC-EVs (P < 0.05). HIF-1α was only elevated in cortical tubules and rescued by lean-MSC-EVs (P < 0.05). ∗P < 0.05; ∗∗P < 0.01. IL, interleukin; TNF-α, tumor necrosis factor alpha; HIF-1α, hypoxia inducible factor-1α; MSC-EVs, mesenchymal stem/stromal cell-derived extracellular vesicles; STK, stenotic kidney.

Figure 4. Potency of obese-MSC-EVs and lean-MSC-EVs in promoting angiogenesis and protecting PK-1 tubular cells. (a–b) Only lean-MSC-EVs, but not obese-MSC-EVs, increased the immunoreactivity of the vascular marker CD31 in the stenotic mouse kidney. (c) Both the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (cell proliferation) and crystal violet (cell viability) assays showed that only lean-MSC-EVs protect PK-1 cells from injury by antimycin-A and TNF-α. ∗P < 0.05; ∗∗P < 0.01. TNF-α, tumor necrosis factor alpha.

Figure 5. Potency of obese and lean-MSC-EVs in suppressing macrophage infiltration. (a) Only lean-MSC-EVs downregulated the elevated MCP-1 mRNA expression in the stenotic mouse kidney (STK). (b–c) Macrophages stained by F4/80 were detected by immunofluorescence. A greater number of macrophages was observed in the STK, which was restored only by lean-MSC-EVs compared to both RAS+vehicle and RAS+obese-MSC-EVs. ∗P < 0.05; ∗∗P < 0.01.

Figure 6. Effect of obese and lean-MSC-EVs on TNF-α and MCP-1 obese-MSC-EVs increased the expression of TNF-α and MCP-1, but lean-MSC-EVs didn’t. TNF-α decreased in obese-MCS-EVs treated-macrophage when MCP-1 was inhibited. ∗P < 0.05; ∗∗P < 0.01. MCP-1, monocyte chemotactic protein-1; MSC-EVs, mesenchymal stem/stromal cell-derived extracellular vesicles; TNF-α, tumor necrosis factor alpha.

Figure 7. Effect of obese and lean-MSC-EVs on macrophage migration. The wound scratch study showed that compared to control and lean-MSC-EVs obese-MSC-EVs promoted macrophage migration, which was blunted by an MCP-1 inhibitor. ∗P < 0.05. MCP-1, monocyte chemotactic protein-1; MSC-EVs, mesenchymal stem/stromal cell-derived extracellular vesicles.