Intravital imaging of real-time endogenous actin dysregulation in proximal and distal tubules at the onset of severe ischemia-reperfusion injury

Peter R Corridon,Ali A Khraibi,Anousha A Khan,Shurooq H Karam,Mohamed A Alhashmi

doi:10.1038/s41598-021-87807-6

Abstract

Severe renal ischemia-reperfusion injury (IRI) can lead to acute and chronic kidney dysfunction. Cytoskeletal modifications are among the main effects of this condition. The majority of studies that have contributed to the current understanding of IRI have relied on histological analyses using exogenous probes after the fact. Here we report the successful real-time visualization of actin cytoskeletal alterations in live proximal and distal tubules that arise at the onset of severe IRI. To achieve this, we induced fluorescent actin expression in these segments in rats with hydrodynamic gene delivery (HGD). Using intravital two-photon microscopy we then tracked and quantified endogenous actin dysregulation that occurred by subjecting these animals to 60 min of bilateral renal ischemia. Rapid (by 1-h post-reperfusion) and significant (up to 50%) declines in actin content were observed. The decline in fluorescence within proximal tubules was significantly greater than that observed in distal tubules. Actin-based fluorescence was not recovered during the measurement period extending 24 h post-reperfusion. Such injury decimated the renal architecture, in particular, actin brush borders, and hampered the reabsorptive and filtrative capacities of these tubular compartments. Thus, for the first time, we show that the combination of HGD and intravital microscopy can serve as an experimental tool to better understand how IRI modifies the cytoskeleton in vivo and provide an extension to current histopathological techniques.

Highlights

Severe renal ischemia-reperfusion injury (IRI) can lead to acute and chronic kidney dysfunction
Severe IRI is a common cause of acute kidney injury (AKI)[2], and produces irreversible damage that supports the progression of AKI to chronic kidney disease (CKD) and, end-stage renal failure[3]
Brightfield images were obtained from cortical sections of normal rat kidneys that were counterstained with hematoxylin and eosin (H&E) (Fig. 1A)

Summary

Introduction

Severe renal ischemia-reperfusion injury (IRI) can lead to acute and chronic kidney dysfunction. Ischemia-reperfusion injury (IRI) is a complex cascade of events that support structural and functional losses in renal tubular segments. Severe IRI is a common cause of acute kidney injury (AKI)[2], and produces irreversible damage that supports the progression of AKI to chronic kidney disease (CKD) and, end-stage renal failure[3]. This disease progression is a growing global health problem with no current specific treatment that has been the focus of research for several decades[3]. The complex nature of the renal system has provided significant challenges to progress in this field

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Conclusion