Deletion of the WNK3-SPAK kinase complex in mice improves radiographic and clinical outcomes in malignant cerebral edema after ischemic stroke

Abstract

The WNK-SPAK kinase signaling pathway controls renal NaCl reabsorption and systemic blood pressure by regulating ion transporters and channels. A WNK3-SPAK complex is highly expressed in brain, but its function in this organ remains unclear. Here, we investigated the role of this kinase complex in brain edema and white matter injury after ischemic stroke. Wild-type, WNK3 knockout, and SPAK heterozygous or knockout mice underwent transient middle cerebral artery occlusion. One cohort of mice underwent magnetic resonance imaging. Ex-vivo brains three days postischemia were imaged by slice-selective spin-echo diffusion tensor imaging magnetic resonance imaging, after which the same brain tissues were subjected to immunofluorescence staining. A second cohort of mice underwent neurological deficit analysis up to 14 days post-transient middle cerebral artery occlusion. Relative to wild-type mice, WNK3 knockout, SPAK heterozygous, and SPAK knockout mice each exhibited a >50% reduction in infarct size and associated cerebral edema, significantly less demyelination, and improved neurological outcomes. We conclude that WNK3-SPAK signaling regulates brain swelling, gray matter injury, and demyelination after ischemic stroke, and that WNK3-SPAK inhibition has therapeutic potential for treating malignant cerebral edema in the setting of middle cerebral artery stroke.

Functional kinomics establishes a critical node of volume-sensitive cation-Cl− cotransporter regulation in the mammalian brain

“…Genetic inhibition of WNK3-SPAK signaling in the MCAO model also ameliorated two cell swelling-associated components of ischemic cerebral edema: perivascular cytotoxic edema of astrocytes and endothelial cell cytotoxic edema (both contributing to BBB breakdown). These findings provide a mechanistic explanation for the improved radiographic and clinical outcomes of malignant cerebral edema after ischemic stroke in mice genetically lacking either WNK3 or SPAK46. Our in vitro data (Fig.…”

“…These collectively contribute to BBB breakdown, vasogenic edema, and brain swelling4445. WNK3 KO mice develop significantly less cerebral edema and infarct volume after MCAO, and exhibit accelerated neurobehavioral recovery, but the mechanisms of these effects remain incompletely understood46.…”

Selective role of Na+/H+ exchanger in Cx3cr1+ microglial activation, white matter demyelination, and post‐stroke function recovery

“…Neurological functional deficits in mice were also determined by the corner test, as described before (Zhao et al, 2017). Corner tests were conducted prior to tMCAO and at 1, 2, 3, 5, 7, 10, and 14 days after tMCAO.…”

“…The adhesive contact test and the adhesive removal test were used to measure somatosensory deficits as described previously (Zhao et al, 2017). Animals were tested prior to tMCAO and at 1, 2,3, 5, 7,10, and 14 days after tMCAO.…”

WNK Kinases in Development and Disease

“…Indeed, mice in which WNK3 or SPAK are knocked out have decreased edema, as well as decreased infarct size and axonal demyelination. Importantly, functional neurological outcomes after stroke are also improved, suggesting that the WNK3-SPAK/OSR1-NKCC1 pathway could be a therapeutic target in stroke (Begum et al, 2015; Zhao et al, 2016). The currently available NKCC1 inhibitor, bumetanide, has low blood-brain barrier permeability, and a poor side effect profile due to inhibition of renal NKCC2 (Donovan et al, 2016; Pressler et al, 2015).…”