Therapeutically Targeting Tumor Necrosis Factor-α/Sphingosine-1-Phosphate Signaling Corrects Myogenic Reactivity in Subarachnoid Hemorrhage

Abstract

S ubarachnoid hemorrhage (SAH) affects ≈10 in 100 000 people/y.1 Although SAH accounts for only 5% to 10% of all strokes, it is particularly devastating: half of all SAH patients die within 1 month and half of those who survive will require life-long assistance for daily living (ie, ≈75% of patients die or are seriously debilitated).2,3 The poor clinical outcome can be attributed to the biphasic course of the disease, characterized by an initial hemorrhagic stroke that is frequently followed by delayed cerebral ischemia (DCI) within 4 to 12 days. 4,5 The latter is a primary therapeutic concern when treating SAH, because DCI causes at least half of the morbidity and mortality in SAH. 2The transition from the hemorrhagic insult to secondary ischemia is driven by prominent changes in cerebrovascular reactivity, which compromises cerebral autoregulation 6,7 and evokes angiographic vasospasm (ie, radiographically identifiable arterial narrowing in the proximal cerebrovascular circulation).2,8 Conceptually, both events originate from augmentedBackground and Purpose-Subarachnoid hemorrhage (SAH) is a complex stroke subtype characterized by an initial brain injury, followed by delayed cerebrovascular constriction and ischemia. Current therapeutic strategies nonselectively curtail exacerbated cerebrovascular constriction, which necessarily disrupts the essential and protective process of cerebral blood flow autoregulation. This study identifies a smooth muscle cell autocrine/paracrine signaling network that augments myogenic tone in a murine model of experimental SAH: it links tumor necrosis factor-α (TNFα), the cystic fibrosis transmembrane conductance regulator, and sphingosine-1-phosphate signaling. Methods-Mouse olfactory cerebral resistance arteries were isolated, cannulated, and pressurized for in vitro vascular reactivity assessments. Cerebral blood flow was measured by speckle flowmetry and magnetic resonance imaging. Standard Western blot, immunohistochemical techniques, and neurobehavioral assessments were also used. Results-We demonstrate that targeting TNFα and sphingosine-1-phosphate signaling in vivo has potential therapeutic application in SAH. Both interventions (1) eliminate the SAH-induced myogenic tone enhancement, but otherwise leave vascular reactivity intact; (2) ameliorate SAH-induced neuronal degeneration and apoptosis; and (3) improve neurobehavioral performance in mice with SAH. Furthermore, TNFα sequestration with etanercept normalizes cerebral perfusion in SAH. Conclusions-Vascular smooth muscle cell TNFα and sphingosine-1-phosphate signaling significantly enhance cerebral artery tone in SAH; anti-TNFα and anti-sphingosine-1-phosphate treatment may significantly improve clinical outcome.

Tumor Necrosis Factor/Sphingosine-1-Phosphate Signaling Augments Resistance Artery Myogenic Tone in Diabetes

“…*P < 0.05 for an unpaired comparison. myogenic tone in these arteries (10)(11)(12)16). HFD/STZ augments mesenteric and olfactory cerebral artery myogenic tone without altering PE responsiveness or passive diameter; myogenic tone in PCAs and cremaster skeletal muscle resistance arteries is not affected (Fig.…”

“…These strains include wild type C57BL/6N mice (Charles River Laboratories, Montreal, Quebec, Canada), germline TNF knockout mice (TNF 2/2 ) (Taconic Biosciences, Hudson, NY), germ line sphingosine kinase 1 (Sphk1) knockout mice (Sphk1 2/2 ) (a gift from Dr. Richard L. Proia, National Institutes of Health, Bethesda, MD) (15) and inducible, smooth muscle cell-targeted TNF knockout mice (12). All mice were housed under a standard 14 h:10 h light:dark cycle and had access to food and water ad libitum.…”

“…Mice were maintained on their respective diets for 12 additional weeks, at which point resistance arteries were isolated for assessment. Mesenteric (207 6 3 mm in diameter, n = 178), olfactory cerebral (113 6 2 mm, n = 140), cremaster skeletal muscle (79 6 2 mm, n = 15), and posterior cerebral arteries (PCAs) (147 6 3 mm, n = 28) were microsurgically isolated as previously described (10)(11)(12)16).…”

“…Resistance arteries were carefully cannulated onto micropipettes, pressurized, and warmed to 37°C in MOPS-buffered saline as previously described (10)(11)(12)16). Mouse olfactory and PCAs were pressurized to 45 mmHg; mouse mesenteric and human skeletal muscle resistance arteries were pressurized to 60 mmHg; and all treatments were diluted in MOPS-buffered saline.…”

“…Resistance arteries are the key determinants of tissue perfusion: their behavior is primarily governed by the myogenic response, an intrinsic and dynamically regulated mechanism that permits highly localized control of blood flow. We have identified a signaling mechanism that augments resistance artery myogenic responses in several etiologically distinct diseases (9)(10)(11)(12): it relies on vascular smooth muscle cell tumor necrosis factor (TNF) signaling and ultimately enhances sphingosine-1-phosphate (S1P)-mediated vasoconstriction. As a key molecular entry point into this signaling pathway, the increase in TNF activity triggered by diabetes (13) is positioned to augment resistance artery tone.…”