RESUMO
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 post-ischemia 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.
Assuntos
Edema Encefálico/genética , Encéfalo/patologia , Deleção de Genes , Infarto da Artéria Cerebral Média/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Encéfalo/metabolismo , Edema Encefálico/etiologia , Edema Encefálico/patologia , Feminino , Infarto da Artéria Cerebral Média/complicações , Infarto da Artéria Cerebral Média/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Bainha de Mielina/genética , Bainha de Mielina/metabolismo , Bainha de Mielina/patologia , Substância Branca/metabolismo , Substância Branca/patologiaRESUMO
Microglia play important roles in extracellular matrix remodeling, tumor invasion, angiogenesis, and suppression of adaptive immunity in glioma. Na(+)/H(+) exchanger isoform 1 (NHE1) regulates microglial activation and migration. However, little is known about the roles of NHE1 in intratumoral microglial activation and microglia-glioma interactions. Our study revealed up-regulation of NHE1 protein expression in both glioma cells and tumor-associated Iba1(+) microglia in glioma xenografts and glioblastoma multiforme microarrays. Moreover, we observed positive correlation of NHE1 expression with Iba1 intensity in microglia/macrophages. Glioma cells, via conditioned medium or non-contact glioma-microglia co-cultures, concurrently upregulated microglial expression of NHE1 protein and other microglial activation markers (iNOS, arginase-1, TGF-ß, IL-6, IL-10 and the matrix metalloproteinases MT1-MMP and MMP9). Interestingly, glioma-stimulated microglia reciprocally enhanced glioma proliferation and migration. Most importantly, inhibition of microglial NHE1 activity via small interfering RNA (siRNA) knockdown or the potent NHE1-specific inhibitor HOE642 significantly attenuated microglial activation and abolished microglia-stimulated glioma migration and proliferation. Taken together, our findings provide the first evidence that NHE1 function plays an important role in glioma-microglia interactions, enhancing glioma proliferation and invasion by stimulating microglial release of soluble factors. NHE1 upregulation is a novel marker of the glioma-associated microglial activation phenotype. Inhibition of NHE1 represents a novel glioma therapeutic strategy by targeting tumor-induced microglial activation.
