Renal Ischemia-Reperfusion Injury in a Diabetic Monkey Model and Therapeutic Testing of Human Bone Marrow-Derived Mesenchymal Stem Cells

Abstract

Clinically, acute kidney injury (AKI) episodes in diabetes mellitus (DM) patients are associated with a cumulative risk of developing end-stage renal disease. In this study, we asked whether the severity of AKI induced by renal ischemia-reperfusion injury (IRI) is more prominent in DM than in non-DM control using a cynomolgus monkey (Macaca fascicularis) model. We also investigated whether human bone marrow-derived mesenchymal stem cells (hBM-MSCs) infused via the renal artery could ameliorate renal IRI in DM monkeys. The experimental data, including mortality rate, histologic findings, and urinary albumin secretion indicate that the severity of AKI was greater in DM monkeys than in control animals. Moreover, histological findings and qRT-PCR analysis of Ngal mRNA in renal biopsy tissue showed that hBM-MSC promoted the recovery of tubular damage caused by AKI. Serum analysis also revealed that the level of albumin and ALT was increased 24 and 48 hours after AKI, respectively, suggesting that AKI induced acute liver injury. We suggest that this nonhuman primate model could provide essential information about the renal and nonrenal impairment related to DM and help determine the clinical usefulness of MSCs in AKI.

Figure 1

The survival rate and changes in renal function markers. DM monkeys that experienced renal ischemia-reperfusion injury (IRI) were subsequently injected with MSC (DM + MSC) or not (DM). Non-DM animals were used as the control. The serum levels of creatinine (a) and blood urea nitrogen (BUN) (b) were measured at designated study points. Data are median with range. n = 3, 3, and 2 in the non-DM, DM, and DM + MSC groups, respectively.

Figure 2

Histological examination of renal tissues. At designated time points, renal biopsy specimens were collected and subjected to H&E staining (a) to assess their tissue morphology. Scale bar = 100 μm. Magnification: ×200. The extent of tubular injury (b) and necrosis (c) were also calculated. n = 3, 3, and 2 in the non-DM, DM, and DM + MSC groups, respectively. ^∗ P < 0.05 and ^∗∗ P < 0.001.

Figure 3

Analysis of proinflammatory markers following renal IRI and MSC treatment. (a) Blood was collected every 24 hours, and the serum levels of each cytokine were measured by a multiplex bead assay. Data are median with range. (b) Analysis of urinary markers for AKI. The concentration of urinary albumin and NGAL was measured by ELISA, and Ngal mRNA in renal tissue was analyzed by qRT-PCR. The level of GST-alpha and TIMP-1 in urine was obtained by a multiplex bead assay. Data are the median with range. n = 3, 3, and 2 in the non-DM, DM, and DM + MSC groups, respectively. ^∗ P < 0.05.

Figure 4

Serum biochemical analysis of markers indicative of liver function. Blood was collected every 24 hours, and the serum levels of total protein (a), albumin (b), ALT (c), and AST (d) were measured. n = 3, 3, and 2 in the non-DM, DM, and DM + MSC groups, respectively. ^∗ P < 0.05 and ^∗∗ P < 0.01. Data are the mean with range.