Meprin β expression associated with decreased proximal tubule PKA Cβ levels in ischemia/reperfusion‐induced acute kidney injury
Authors: Ahmed Faihaa , Mwiza Jean-Marie , Fernander Mizpha , Abousaad Shaymaa , Ongeri Elimelda Moige 
Published: April 2020

Ischemia/ reperfusion (IR) is a leading cause of acute kidney injury (AKI). Meprins are metalloproteases that are expressed abundantly in the brush border membranes of renal proximal tubules (PTs). The levels of expression and the localization of meprins were previously shown to associate with IR‐induced AKI. Studies from our group showed that meprin B proteolytically cleaves three isoforms of the catalytic subunit of PKA, namely, PKA Cα1, Cβ1, and Cβ2. The cleavage by meprin B resulted in decreased kinase activity of these PKA C isoforms. Furthermore, the total levels of PKA C were significantly decreased in the meprin expressing PTs at 3h post‐IR when compared to distal tubules which are deficient in meprins. The objective of the current study was to determine which of the PKA C isoforms is impacted by meprin B and thus involved in the IR‐response mediated by meprin β. To this end, surgical procedures were used to induce unilateral IR (with contralateral nephrectomy) in 12‐week old wild‐type (WT) and meprin β knockout (βKO) mice on a C57BL/6 background. Kidney tissues were harvested at 6h post IR. Kidney proteins were extracted and fractionated into BBM‐, cytosolic‐, and nuclear‐enriched fractions. Western blot analysis was used to determine the levels of Cα and Cβ in the cytosolic‐ and nuclear‐enriched fractions. Immunofluorescence staining was used to determine the localization of PKA C (total), Cβ, and Cα. Statistical analysis utilized 2‐way ANOVA. The total levels of PKA C decreased in kidneys from WT mice subjected to IR but not in βKO counterparts. However, use of isoform‐specific antibodies demonstrated increases in the levels of Cβ but not Cα in the cytosolic‐ and nuclear‐enriched fractions. Two Cβ protein bands (41 and 37 kDa) were detected in cytosolic‐ and one band (41 kDa) in nuclear‐enriched fractions. The levels of the 41 kDa Cβ protein species were higher in lysates from WT kidneys at 6h post‐IR (P=0.0001). In contrast, the 37 kDa species, which was detected in nuclear‐enriched fractions only, decreased in βKO (P=0.002) but not in WT kidneys subjected to IR. A closer histological analysis of kidney sections showed that the level of damage to the brush borders was not uniform in all proximal tubules. More importantly, immunohistochemical analysis showed that the PKA Cβ levels decreased in damaged/injured meprin β expressing PTs but appeared to increase in intact PTs. These changes were not observed in PTs from meprin βKO kidneys. Individual interstitial cells (presumed to be resident macrophages) also stained positively for Cβ in WT but not in βKO kidney sections. Taken together, the data suggest that the PKA Cβ isoform is involved in the response to IR‐induced renal injury. The Cβ levels are known to be abundant in lymphoid tissue but our data indicate expression in immune cells resident in kidney tissue as well as epithelial tubular cells in response to IR‐induced AKI. More studies are needed to determine the response genes mediated by PKA Cβ signaling in IR to increase our understanding of how meprins modulate kidney injury in mice.

Support or Funding Information

This study was supported by the National Institute of Health/National Institutes of General Medical Sciences (NIGMS) Grant #s SC3GM102049 and SC1GM118271