Gut Immune Cells, not Blood Pressure, Blamed for Salt’s Effect on Brain

If you believe a new paper on salt and dementia, the culprit is not hypertension but a T cell response. In the January 15 Nature Neuroscience, researchers led by Costantino Iadecola at Weill Cornell Medical College in New York reported that mice eating a high-salt diet for three months pumped less blood to their brains and underperformed on tests of memory and behavior. The deficits had nothing to do with blood pressure, since that did not change. Instead, the researchers found that salt stimulated immune cells in the gut to release an inflammatory cytokine that harmed blood vessels in the brain. In turn, these vessels produced less nitric oxide, a key signaling molecule that regulates blood flow and synaptic function. The data delineate a new axis between the gut and brain that links diet to memory, and may suggest new therapeutic targets to pursue, Iadecola said. In the meantime, the findings provide yet another reason to limit salt intake. This is true especially in aging, since memory faltered sooner in old mice on the salty diet.

  • In mice, a salty diet led to expansion of TH17 cells in the gut, which released IL-17 into the blood.
  • In the brain, IL-17 curbed nitric oxide in blood vessels, strangling blood flow and impairing memory.
  • This gut-brain axis may suggest therapeutic approaches.

Other researchers appreciated the complex connections the study uncovered. “To my knowledge, this is the first mechanistic study explaining the effect of salt on loss of endothelial-mediated vasodilation of brain vessels,” Berislav Zlokovic at the University of Southern California, Los Angeles, wrote to Alzforum. Donna Wilcock at the University of Kentucky, Lexington, agreed. “This is an elegant and fascinating set of experiments. We hadn’t really considered the gut-brain axis with respect to vascular impairment before,” Wilcock said.

Its known that excess dietary salt can harm organs such as the heart, kidneys, blood vessels, and brain, and that elevated blood pressure is not the only mediator (for review, see Farquhar et al., 2015). The pathways were unclear, though bits and pieces were known. Some studies reported high oxidative stress and low nitric oxide (NO) production in the endothelial cells lining blood vessels; others linked high salt to accumulation in the gut of TH17 T helper cells, which produce the proinflammatory cytokine IL-17 (Boegehold, 2013; Cosic et al., 2016; Kleinewietfeld et al., 2013; Wu et al., 2013). Because IL-17 can dampen NO production, Iadecola decided to see if he could connect the dots into a more complete mechanism (Nguyen et al., 2013). 

High Salt, Low Flow. Mice on a normal diet (left) have vigorous blood flow in the brain (color bar). Perfusion wanes on a high-salt diet (middle), and bounces back upon returning to a normal diet (right). [Courtesy of Faraco et al., Nature Neuroscience.]

First author Giuseppe Faraco fed healthy young mice a diet containing 8 percent NaCl,  or 3 percent sodium, which equates to 16 times more salt than in normal chow. At most, people consume about five times the recommended daily sodium, or 8 g (Powles et al., 2013). The U.S. Food and Drug Administration, the American Heart Association, and the WHO recommend people consume around 2 g sodium per day, with 1.5 g considered ideal.

Over 12 weeks on the salty diet, the animals’ blood pressure stayed the same. Even so, by eight weeks cerebral blood flow had shrunk by about a quarter. Four weeks later, endothelial cells no longer pumped out NO in response to stimulation with acetylcholine, which dilates blood vessels. The mice did poorly on tests of novel-object recognition and spatial memory, and built shoddy nests. Notably, when the mice were returned to a normal diet for four weeks, cerebral blood flow, NO production, and behavior all recovered, indicating that the effects of high salt consumption are reversible, at least in the short term.

How did salt wreak this havoc? The mice developed no inflammation in brain blood vessels, ruling that out as a mechanism. Instead, NO seemed a likely culprit, since other vasodilators that do not depend on NO, such as adenosine, still increased cerebral blood flow in the high-salt mice. To test the NO hunch, the authors added the NO precursor L-arginine to the animals’ drinking water from weeks eight to 12 of the high-salt diet. They made normal amounts of NO and maintained brain blood flow and memory, showing that restoring NO levels was sufficient to counteract the negative effects of high salt intake.

Next, the authors probed whether IL-17 caused the problem by feeding the high-salt diet to IL-17 knockout mice. Lo and behold, they maintained normal NO production and memory. Mice lacking lymphocytes, and thus TH17 cells, also fared well on the high-salt diet, as did mice treated with neutralizing antibodies against IL-17 from week 10–12 of the diet. On the other hand, injecting IL-17 into mice eating normal chow mimicked the effects of high salt and led to behavioral deficits. The authors also confirmed previous findings that in response to high salt TH17 cells proliferated in the gut, but not in the blood, lymph nodes, spleen, or meninges. These results implicated IL-17 produced by TH17 cells in the gut as a key mediator of the vascular and cognitive problems.

To see if cytokine is connected to NO production, the authors examined endothelial nitric oxide synthase, the enzyme that generates the gas. eNOS is known to be regulated by phosphorylation (Harris et al., 2001). In mice on a high-salt diet, inhibitory eNOS phosphorylation was up by 50 percent, the authors found. Similarly, applying IL-17 to cultured mouse brain endothelial cells nearly doubled inhibitory phosphorylation. Several kinases, including Rho-kinase (ROCK), have been implicated in turning down eNOS activity (Sugimoto et al., 2007). The authors found that cultured cells treated with a ROCK inhibitor maintained normal eNOS activity and NO production during IL-17 treatment, while other kinase inhibitors had no effect. To confirm that ROCK mediated IL-17’s toxic effects, the authors treated mice on a high-salt diet with the ROCK inhibitor from week 10–12. The treatment normalized NO production and mouse behavior.

Notably, all these effects occurred in mice that were only two months old. Because vascular problems tend to increase with age, the authors tested how older mice responded to the salty diet. It induced drops in NO production and cerebral blood flow in middle-aged, 15-month-old mice, as well. However, the behavioral deficits appeared earlier, that is, by eight weeks into the high-salt diet. This suggests the older mice were more vulnerable to the cognitive consequences of a high-salt diet.

Implications for Human Health?
Do the findings translate to people? The study does not answer this question, but as a first step toward investigating it, the authors added IL-17 to cultured human endothelial cells. They saw the same suppression of NO production as in mice already at 10 times lower IL-17 concentration, suggesting human cells are more sensitive to the cytokine. In future work, Iadecola will put volunteers on a high-salt diet for short periods of time to see if their plasma IL-17 and brain blood flow changes at all.

Joanna Wardlaw at the University of Edinburgh, Scotland, said the mouse findings jibe with data from her studies of people. “We see associations between a higher salt intake and features of small-vessel disease, such as white-matter lesions and lacunae on brain scans,” she told Alzforum. These associations are independent of any effect on blood pressure, she confirmed. “It’s encouraging that this study focused on dietary salt. I think that’s been a neglected factor in studies of cerebrovascular damage,” Wardlaw said. At the same time, she noted that the study should be replicated in other labs as well as in additional strains of mice or rats that might have varying susceptibilities to salt and cerebrovascular disease.

If the findings are broadly validated, they would provide a rationale for trying drugs that raise NO levels in people with cognitive problems, Wardlaw suggested. She has just finished a pilot study, LACI-1, that tested two such drugs in people with small-vessel disease to find out if the treatment improved blood-vessel function. She is starting a larger trial, LACI-2, that will look for evidence of cognitive benefit. “We’ve thought for a long time that stroke and dementia have huge commonalities and are linked through dysfunction of the small blood vessels. Hopefully a paper like this will help people start to think more broadly in terms of therapeutic options for brain disease,” Wardlaw said.

For his part, Iadecola is drilling down on how lack of NO harms cognition. It may lower blood flow, thus starving the brain of oxygen and nutrients, or it may directly affect synapses, since NO plays a role in long-term potentiation (for review see Garthwaite, 2008). Regardless of the downstream mechanism, Iadecola believes the upstream factors in this pathway, IL-17 and ROCK, might make promising therapeutic targets for vascular and brain health. IL-17 is elevated in autoimmune diseases such as multiple sclerosis, psoriasis, and rheumatoid arthritis, and an anti-IL-17 antibody, secukinumab, is already in clinical use for psoriasis, with others in development (Wasilewska et al., 2016). 

In the meantime, however, Iadecola recommends that people cut back on excess salt consumption. The average American consumes about 3.4 g per day; the worldwide average is close to 4 g. While this is far less than most of the mice in the study consumed, ominously, mice that ate only half as much salt (4 percent NaCl diet) still made less NO than controls and developed learning and memory problems, Iadecola noted. The 4 percent NaCl diet is about equivalent to the high end of human consumption. “It’s like your mother told you: ‘All things in moderation,’” Iadecola quipped.—Madolyn Bowman Rogers

Comments

  1. This is a very important paper showing the mechanism underlying the link between dietary salt and cognitive function through endothelial-mediated regulation of cerebral blood flow responses. In an elegant series of experiments, Dr. Iadecola’s team has convincingly shown that salt acts by affecting TH17 cell in the small intestine causing IL17 production, which in turn blocks NO generation by brain endothelial cells, leading to dysregulated blood flow responses to brain activation that can set a stage for development of neurodegenerative changes and behavioral deficits. To my knowledge this is first mechanistic study explaining the effect of salt on loss of endothelial-mediated vasodilation of brain vessels through inhibition of the NO system via very complex peripheral mechanism involving cross-talk of peripheral gut-derived immune TH17 cells with endothelial cells of cerebral blood vessels.

    View all comments by Berislav Zlokovic

  2. This is an interesting study that investigated the effect of high-salt diet on alteration of blood-circulation parameters. The authors have shown that high-salt diet alters cerebral blood flow (CBF), increases plasma interleukin 17 (IL-17) and causes endothelial damage. Others, including us, have shown the effect of high salt diet on CBF as well as core blood pressure in animal models (Taheri et al., 2016). However, the results of such studies are controversial as they depend on the experimental settings, such as the number of animals, diet composition, diet regimen, the CBF measurement methods, and the statistical inference powers. To overcome these controversies more studies with a higher number of animals and more advanced CBF measurement methods are needed.

    Exploring the mechanism by which a high-salt diet regimen affects endothelial cells function is a strong part of this report. Many pathways have been suggested by which high-salt diet can affect blood circulation and endothelial dysfunction, including Na/Ca channels and NO. Alteration of NO synthesis was long thought to be the main culprit of cerebral endothelial dysfunction and cerebral hypoperfusion. However, its connection to high salt diet and gut cells is very interesting.

    References:

    Taheri S, Yu J, Zhu H, Kindy MS. High-Sodium Diet Has Opposing Effects on Mean Arterial Blood Pressure and Cerebral Perfusion in a Transgenic Mouse Model of Alzheimer's Disease. J Alzheimers Dis. 2016 Oct 4;54(3):1061-1072. PubMed.

    View all comments by Saeid Taheri

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References

Paper Citations

  1. Farquhar WB, Edwards DG, Jurkovitz CT, Weintraub WS. Dietary sodium and health: more than just blood pressure. J Am Coll Cardiol. 2015 Mar 17;65(10):1042-50. PubMed.
  2. Boegehold MA. The effect of high salt intake on endothelial function: reduced vascular nitric oxide in the absence of hypertension. J Vasc Res. 2013;50(6):458-67. Epub 2013 Oct 26 PubMed.
  3. Cosic A, Jukic I, Stupin A, Mihalj M, Mihaljevic Z, Novak S, Vukovic R, Drenjancevic I. Attenuated flow-induced dilatation of middle cerebral arteries is related to increased vascular oxidative stress in rats on a short-term high salt diet. J Physiol. 2016 Sep 1;594(17):4917-31. Epub 2016 Jun 16 PubMed.
  4. Kleinewietfeld M, Manzel A, Titze J, Kvakan H, Yosef N, Linker RA, Muller DN, Hafler DA. Sodium chloride drives autoimmune disease by the induction of pathogenic TH17 cells. Nature. 2013 Apr 25;496(7446):518-22. Epub 2013 Mar 6 PubMed.
  5. Wu C, Yosef N, Thalhamer T, Zhu C, Xiao S, Kishi Y, Regev A, Kuchroo VK. Induction of pathogenic TH17 cells by inducible salt-sensing kinase SGK1. Nature. 2013 Apr 25;496(7446):513-7. Epub 2013 Mar 6 PubMed.
  6. Nguyen H, Chiasson VL, Chatterjee P, Kopriva SE, Young KJ, Mitchell BM. Interleukin-17 causes Rho-kinase-mediated endothelial dysfunction and hypertension. Cardiovasc Res. 2013 Mar 15;97(4):696-704. Epub 2012 Dec 21 PubMed.
  7. Powles J, Fahimi S, Micha R, Khatibzadeh S, Shi P, Ezzati M, Engell RE, Lim SS, Danaei G, Mozaffarian D, Global Burden of Diseases Nutrition and Chronic Diseases Expert Group (NutriCoDE). Global, regional and national sodium intakes in 1990 and 2010: a systematic analysis of 24 h urinary sodium excretion and dietary surveys worldwide. BMJ Open. 2013 Dec 23;3(12):e003733. PubMed.
  8. Harris MB, Ju H, Venema VJ, Liang H, Zou R, Michell BJ, Chen ZP, Kemp BE, Venema RC. Reciprocal phosphorylation and regulation of endothelial nitric-oxide synthase in response to bradykinin stimulation. J Biol Chem. 2001 May 11;276(19):16587-91. Epub 2001 Feb 28 PubMed.
  9. Sugimoto M, Nakayama M, Goto TM, Amano M, Komori K, Kaibuchi K. Rho-kinase phosphorylates eNOS at threonine 495 in endothelial cells. Biochem Biophys Res Commun. 2007 Sep 21;361(2):462-7. Epub 2007 Jul 18 PubMed.
  10. Garthwaite J. Concepts of neural nitric oxide-mediated transmission. Eur J Neurosci. 2008 Jun;27(11):2783-802. PubMed.
  11. Wasilewska A, Winiarska M, Olszewska M, Rudnicka L. Interleukin-17 inhibitors. A new era in treatment of psoriasis and other skin diseases. Postepy Dermatol Alergol. 2016 Aug;33(4):247-52. Epub 2016 Aug 16 PubMed.

External Citations

  1. recommend
  2. LACI-1

Further Reading

Primary Papers

  1. Faraco G, Brea D, Garcia-Bonilla L, Wang G, Racchumi G, Chang H, Buendia I, Santisteban MM, Segarra SG, Koizumi K, Sugiyama Y, Murphy M, Voss H, Anrather J, Iadecola C. Dietary salt promotes neurovascular and cognitive dysfunction through a gut-initiated TH17 response. Nat Neurosci. 2018 Feb;21(2):240-249. Epub 2018 Jan 15 PubMed.

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