In vivo two-photon microscopy reveals the contribution of Sox9+ cell to kidney regeneration in a mouse model with extracellular vesicle treatment

Abstract

<jats:p>Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) have been shown to stimulate regeneration in the treatment of kidney injury. Renal regeneration is also thought to be stimulated by the activation of Sox9+ cells. However, whether and how the activation mechanisms underlying EV treatment and Sox9+ cell-dependent regeneration intersect is unclear. We reasoned that a high-resolution imaging platform in living animals could help to untangle this system. To test this idea, we first applied EVs derived from human placenta-derived MSCs (hP-MSCs) to a Sox9-Cre<jats:sup>ERT2</jats:sup>; R26<jats:sup>mTmG</jats:sup> transgenic mouse model of acute kidney injury (AKI). Then, we developed an abdominal imaging window (AIW) in the mouse and tracked the Sox9+ cells in the inducible Sox9 Cre transgenic mice via in vivo lineage tracing with two-photon intravital microscopy. Our results demonstrated that EVs can travel to the injured kidneys post intravenous injection as visualized by Gaussia luciferase imaging and markedly increased the activation of Sox9+ cells. Moreover, the two-photon living imaging of lineage-labeled Sox9+ cells showed that the EVs promoted the expansion of Sox9+ cells in kidneys post AKI. Histological staining results confirmed that the descendants of Sox9+ cells contributed to nephric tubule regeneration which significantly ameliorated the renal function after AKI. In summary, intravital lineage tracing with two-photon microscopy through an embedded AIW provides a practical strategy to investigate the beneficial functions and to clarify the mechanisms of regenerative therapies in AKI.</jats:p>

Mesenchymal stem cells and extracellular vesicles in therapy against kidney diseases

“…Researches have shown that MSCs locate to injured areas via direct interaction and the paracrine effect [12,13], while being less dependent on the differentiation function [14]. Through marking, it is found that, after being injected into the body, MSCs can be specifically located in the injured zones of the kidney [15][16][17]. The molecular mechanisms of MSC homing are based on a multistep model, including initial tethering by selectins, activation by cytokines, arrest by integrins, diapedesis or transmigration, and migration toward chemokines [18].…”

“…Chen et al proposed that the glial-derived neurotrophic factor-modified ADMSC-Exos stimulate the perivascular capillaries in tubulointerstitial fibrosis by activating the SIRT1/eNOS pathway [ 76 ]. In addition, previous research also suggested that ADMSC-Exos upregulate the expression of the transcription factor Sox9 of TECs, and the offspring of Sox9 + cells facilitate regeneration of renal tubules rather than fibrotic transformation, thus slowing the AKI-CKD transition [ 17 , 77 ].…”

Stem/progenitor cell in kidney: characteristics, homing, coordination, and maintenance

“…There are stem/progenitor cells in tubules, especially proximal tubules [ 5 , 11 , 21 , 24 , 64 , 68 ]. Most of them are capable to differentiate into tubular epithelial cells and even could differentiate into mesodermal lineages such as adipogenic, osteogenic, and chondrogenic lineages.…”

“…It has been found that Sox9 + cells are in adult kidney, which own the high capacity of proliferation and mesodermal lineage differentiation [ 70 ]. These stem/progenitor cells are primarily located in proximal tubules, and they have epithelial polarity and brush border [ 68 ]. These cells express CD133 and Lgr4, the markers of progenitor cells, but have a negative expression of Pax-2 or common MSC markers.…”

“…They think that although most descendants of Sox9 + cells no longer express Sox9 gene in normal kidneys, it is activated after kidney injury. The researchers think that de novo activation of Sox9 rather than the expansion of the resident Sox9 + population contributes more to the recovery of kidney [ 68 , 70 , 71 ].…”