Hyperglycemia abolishes the protective effect of ischemic preconditioning in glomerular endothelial cells in vitro

Abstract

In preclinical investigations, ischemic preconditioning (IPC) protects kidneys from ischemia/reperfusion injury. The direct effects of IPC on glomerular endothelial cells have not been studied in detail. Most investigations of IPC have focused on healthy cells and animals, and it remains unknown whether IPC is renoprotective in the setting of medical comorbidities such as diabetes. In this study, we determined the preventive potential of IPC in healthy glomerular endothelial cell monolayers, and compared these results to monolayers cultured under hyperglycemic conditions. We exposed glomerular endothelial monolayers to 1 h of IPC 24 h prior to oxygen-glucose deprivation (OGD), an in vitro model of ischemia/reperfusion injury. Glomerular endothelial monolayer integrity was assessed by measuring transendothelial electrical resistance, albumin flux, and cell survival. We found that IPC protected healthy but not hyperglycemic glomerular endothelial monolayers from ischemia/reperfusion injury. Furthermore, not only was the protective effect of IPC lost in the setting of hyperglycemia, but IPC was actually deleterious to the integrity of hyperglycemic glomerular endothelial cell monolayers.

Keywords: Glomerular endothelial cells; hyperglycemia; ischemic preconditioning; renal ischemia–reperfusion injury.

Figure 1

(A) Graph showing transendothelial electrical resistance (TEER) recordings of GenC monolayers under normal glucose conditions. TEER (y-axis) is shown as a percentage of baseline recordings over time (x-axis). Results show that Oxygen–Glucose Deprivation (OGD) causes GenC monolayer dysfunction that was maximal in the first 2 h of OGD and remained rapid for the first 8 h (n = 5, P < 0.0001). (B) Graph showing percentage of surviving GenC normalized to normoxic controls (y-axis) following exposure to OGD over time (x-axis). Results show that OGD does not cause an increase in GenC cell death compared to normoxic controls (n = 3, P = 0.13).

Figure 2

(A) Graph of TEER of GenC monolayers cultured in either normal glucose or high glucose conditions. Results show that high glucose causes a decrease in TEER (n = 5, P < 0.0001). (B) Graph of albumin flux across GenC monolayers cultured in normal or high glucose conditions. Results show that high glucose causes an increase in albumin flux (n = 3, P = 0.02).

Figure 3

Following 8 h of OGD and 12 h of RGR, there was (A). decreased TEER in hyperglycemic compared to normoglycemic GenC monolayers (n = 5, P = 0.02), and (B). increased albumin flux across hyperglycemic compared to normoglycemic GenC monolayers (n = 3, P = 0.04).

Figure 4

Graph showing TEER of normoglycemic and hyperglycemic GenC exposed to ischemic preconditioning (IPC) prior to OGD/RGR. TEER values are represented as percentage of control GenC that were exposed to OGD/RGR but not exposed to IPC. Results show that IPC protects normoglycemic GenC function (n = 4, P = 0.01) but makes hyperglycemic GenC more susceptible to OGD/RGR-induced injury (n = 4, P = 0.03).