AB040. Early and long-term protective effect of uncultured adipose stromal vascular fraction against renal ischemia-reperfusion injury via pre-ischemic administration

Background: Ischemia-reperfusion (IR) induced acute kidney injury (AKI) is the common clinical syndrome. Stem/progenitor cells therapy is a promising option to foster the intrinsic capacity of renal regeneration. However, several challenges still remain due to the potential risks during in vitro cell culture, low retention rate after transplantation and unclear effect on the progression of chronic kidney disease (CKD). Adipose stromal vascular fraction (SVF) is regarded as an attractive cell source for cell-based therapy without need of in vitro expansion. Preconditioning with ischemia has been suggested to be a useful method to induce a delay of lethal cell injury, enhance cellular tolerance to injurious effects of IR, and promote cell retention and survival. The purpose of this study is to investigate the effect of pre-administration of uncultured adipose SVF on IR induced AKI and progression of fibrosis.

Methods: SVF was isolated from rat epididymal adipose tissue and characterized by flow cytometric analysis. Cell proliferation and scrape wound healing assay was performed by a co-culture system to evaluate the effects of SVF pre-hypoxic administration on proliferation, migration and apoptosis of renal tubular epithelial cells in hypoxic environment. IR induced kidney injury model was established and CM-DiI labeled SVF was injected to the subcapsular pre- or post-ischemia. Renal function was measured at 12 h, 24 h, 72 h, and 6 months after surgery. SVF retention was evaluated by the expression of CM-DiI under fluorescence microscope. Renal tissue sample was retrieved and stained with hematoxylin and eosin (HE) to evaluate the degree of tubular injury. Immunohistochemical assay was performed to detect the expression of PCNA, E-cadherin, RECA-1, and α-SMA in kidney tissue. Masson’s trichrome staining was performed for the analysis of renal fibrosis. Expression of E-cadherin and α-SMA in kidney tissue was also evaluated by western blot analysis for the analysis of epithelia-mesenchymal transition (EMT). Level of transforming growth factor-beta1 (TGF-β1) in the serum and kidney tissues at 6 months after injury was determined by enzyme-linked immunosorbent assay.

Results: Fleshly isolated SVF expressed hematopoietic, mesenchymal, and endothelial antigens. Higher cell retention rate was detected in SVF pre-ischemic administration group. SVF pre-ischemic administration showed stronger functional and morphologic protection from renal IR injury than SVF post-ischemic administration by enhancing tubular epithelial cell proliferation and reducing apoptosis. Progression of chronic kidney damage was also significantly delayed by pre-ischemic administration of SVF, compare with post-ischemic administration, through inhibiting EMT, microvascular rarefaction, and the production of TGF-β1. In addition, in vitro study showed that SVF pre-hypoxic administration could also significantly promote the proliferation, migration and survival of hypoxic renal tubular epithelial cells.

Conclusions: Our study firstly demonstrated that pre-administration of uncultured adipose SVF protect the kidney from both early IR injury and long-term risk of developing CKD. SVF administration before ischemia could enhance cell retention and then contributed to the improvement of tubular epithelial cells survival and proliferation, and inhibition of CKD.

Keywords: Acute kidney injury (AKI); stromal vascular fraction (SVF); ischemia-reperfusion (IR); kidney fibrosis