Munji RN, Soung AL, Weiner GA, Sohet F, Semple BD, Trivedi A, Gimlin K, Kotoda M, Korai M, Aydin S, Batugal A, Cabangcala AC, Schupp PG, Oldham MC, Hashimoto T, Noble-Haeusslein LJ, Daneman R. Profiling the mouse brain endothelial transcriptome in health and disease models reveals a core blood-brain barrier dysfunction module. Nat Neurosci. 2019 Oct 14; PubMed.

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  1. This is an exciting paper from the Daneman group that studies how the unique characteristics of endothelial cells (ECs) around brain capillaries change in different types of pathology, in such a way that the ECs start to resemble ECs on peripheral capillaries.

    This is important because the blood-brain barrier (BBB) is normally maintained by special properties of CNS ECs: The presence of tight junctions between ECs and a suppression of transcytosis across the cells. BBB failure occurs in various types of brain pathology and contributes to neural dysfunction. In some circumstances, this failure may also provide a route for therapeutic agents to enter the CNS. Surprisingly, the paper shows that in different types of pathology there are similarities in how the gene-expression properties of the ECs alter (measured at the mRNA level: Future work will be needed to examine changes at the protein level, and also tease out the functional significance of the change in each of the many proteins whose expression is altered). This raises the possibility that successfully preventing (or increasing) EC-gene-expression changes that occur in one disease may lead to a potential therapy for other types of CNS disorder.

    View all comments by David Attwell

  2. This is a noteworthy study by the Daneman lab comparing the transcriptional profiles of mouse blood-brain barrier (BBB) endothelial cells (ECs) with those in other organs of the body (i.e., heart, lung, kidney, and liver) using RNA-Seq method and an inducible endothelial-specific reporter mouse model, ROSA-tdTomato;VE-Cadherin-CreERT2.

    First, they identified Wnt/β-catenin-related pathways (known to be crucial for BBB formation and maintenance), different transport mechanisms, and amino acid metabolism as key BBB-enriched pathways. The authors also pinpointed highly expressed tight-junction molecules including several that are BBB-enriched compared to the peripheral ECs such as Igsf5, Ocln, Lsr, Marveld2, Cgnl1, and Amot. In addition, they found BBB-enriched transporters including ATP-binding cassette (ABC) transporters (e.g., Abcb1a, encoding P-glycoprotein) and other transporters of energy metabolites (e.g., the famous glucose transporter GLUT1 encoded by the Slc2a1 gene), amino acids, neurotransmitters, ions, and others. I agree with the authors that BBB-enriched transcriptome data will provide a better understanding of BBB-specific functions as well as identifying targets to enhance drug delivery into the central nervous system.

    Importantly, the authors have also looked at disease models of stroke, traumatic brain injury, multiple sclerosis (MS), and seizures, each having profound BBB dysfunction. They noticed similar patterns of endothelial gene expression across all models in the acute/subacute phase, although each disease is caused by a unique trigger. Twelve common genes included Adamts4 (encoding ADAMTS4, a metalloproteinase), Atp8b1, Cd14, Ch25h, Kit, Lrg1, Pdlim1, Scgb3a1, Sele (encoding E-selectin, a leukocyte adhesion molecule), Tmem173, Trp53i11, and Upp1. These data suggest that finding a treatment limiting BBB dysfunction in one of the diseases may lead to treatment for others.

    There is no doubt that this exciting paper will lead to additional studies which should consider the following:

    1. The link between endothelial-gene profile and functional protein expression is missing here. Future studies should perform these experiments that are critical to better understand the BBB properties in health and disease.
    2. The newly discovered concept of “vascular zonation” or “BBB zonation” which refers to the heterogeneity of molecular and phenotypic changes of endothelial and mural—i.e., pericytes and vascular smooth muscle cells (VSMCs)—cells along the cerebrovascular tree (Vanlandewijck et al., 2018). In fact, there is increasing evidence for genetic and functional heterogeneity of these cells depending upon their location on the cerebrovascular bed, but also their geolocation in the brain (e.g., white vs. gray matter regions, hippocampus vs. cortical mantle, etc.).
    3. The transcriptional profiles of other vascular cell types, including pericytes, VSMCs, and astrocytes. Each vascular cell type can be FACS-sorted, and their respective transcriptional profiles investigated. One would think that they have distinct spatiotemporal patterns of gene expression in both health and disease. It would be interesting to dissect all these gene profiles in the proposed disease models and see whether a certain cell type has specific gene-expression changes prior to others.
    4. AD mouse models, since vascular dysfunction has become an important etiological factor (Montagne et al., 2017). Profiling the mouse brain vascular cell transcriptome in AD mice would be of interest, not only for the endothelial gene profile, but also for pericytes and VSMCs profiles as well. Indeed, damage to brain capillary pericytes has been shown to contribute to AD pathophysiology in mice (Montagne et al., 2017), and to develop early in older adults with cognitive impairment (van de Haar et al., 2016; Nation et al., 2019). 

    Finally, the authors have found that the Vcam1 gene (encoding for an inflammatory leukocyte adhesion molecule called VCAM-1) was mostly expressed in peripheral vessels, except in the experimental autoimmune encephalomyelitis mouse model of MS where Vcam1 was detected in brain vessels. Given that VCAM-1 is not constitutively expressed at the BBB, it would be interesting to investigate whether brain endothelium activation (e.g., increased Vcam1 at the BBB) occurs early in AD using Daneman’s BBB-enriched RNA-Seq method, and whether it precedes amyloid buildup, neurodegeneration, and memory deficits. Recent clinical studies revealed that soluble VCAM-1 was the top protein among 31 that increased with age (Yousef et al., 2019) and that higher levels of plasma sVCAM-1 are found in AD cases compared to age-matched controls (Huang et al., 2015). 

    References:

    van de Haar HJ, Burgmans S, Jansen JF, van Osch MJ, van Buchem MA, Muller M, Hofman PA, Verhey FR, Backes WH. Blood-Brain Barrier Leakage in Patients with Early Alzheimer Disease. Radiology. 2016 Nov;281(2):527-535. Epub 2016 May 31 PubMed.

    Huang CW, Tsai MH, Chen NC, Chen WH, Lu YT, Lui CC, Chang YT, Chang WN, Chang AY, Chang CC. Clinical significance of circulating vascular cell adhesion molecule-1 to white matter disintegrity in Alzheimer's dementia. Thromb Haemost. 2015 Nov 25;114(6):1230-40. Epub 2015 Aug 20 PubMed.

    Montagne A, Zhao Z, Zlokovic BV. Alzheimer's disease: A matter of blood-brain barrier dysfunction?. J Exp Med. 2017 Nov 6;214(11):3151-3169. Epub 2017 Oct 23 PubMed.

    Nation DA, Sweeney MD, Montagne A, Sagare AP, D'Orazio LM, Pachicano M, Sepehrband F, Nelson AR, Buennagel DP, Harrington MG, Benzinger TL, Fagan AM, Ringman JM, Schneider LS, Morris JC, Chui HC, Law M, Toga AW, Zlokovic BV. Blood-brain barrier breakdown is an early biomarker of human cognitive dysfunction. Nat Med. 2019 Feb;25(2):270-276. Epub 2019 Jan 14 PubMed.

    Vanlandewijck M, He L, Mäe MA, Andrae J, Ando K, Del Gaudio F, Nahar K, Lebouvier T, Laviña B, Gouveia L, Sun Y, Raschperger E, Räsänen M, Zarb Y, Mochizuki N, Keller A, Lendahl U, Betsholtz C. A molecular atlas of cell types and zonation in the brain vasculature. Nature. 2018 Feb 14; PubMed.

    Yousef H, Czupalla CJ, Lee D, Chen MB, Burke AN, Zera KA, Zandstra J, Berber E, Lehallier B, Mathur V, Nair RV, Bonanno LN, Yang AC, Peterson T, Hadeiba H, Merkel T, Körbelin J, Schwaninger M, Buckwalter MS, Quake SR, Butcher EC, Wyss-Coray T. Aged blood impairs hippocampal neural precursor activity and activates microglia via brain endothelial cell VCAM1. Nat Med. 2019 Jun;25(6):988-1000. Epub 2019 May 13 PubMed.

    View all comments by Axel Montagne

  3. This exciting study provides a comprehensive data resource on the molecular underpinnings of brain endothelial cell dysfunction. It builds on the group's prior work providing a healthy blood-brain barrier (BBB) transcriptome, an already incredibly useful resource. The authors describe here a core dysfunction module, a set of genes that become upregulated across four distinct and acute insults, with several implicated in human disease. It was not known a priori whether such a core dysfunction module existed, and its identification opens a variety of fascinating questions.

    For example, it will be important to determine which of, and how, the module genes are functionally linked to observed phenotypes, such as BBB leakiness. We also wonder whether this module will be found in chronic settings of neurodegeneration and aging. A hint arises in the module's de-enrichment of BBB-specific genes, suggesting a common perturbation in surrounding mural cell signals such as from a loss of pericytes. As pericyte loss has been reported in Alzheimer's disease, this module may indeed be an even more generalizable hallmark of BBB dysfunction. And with single-cell sequencing (scRNA-Seq) technologies increasingly adopted, it will be interesting to deduce vessel segment-specific changes across insults: It may be that their stroke model differentially impacts arterial cells compared to the rest of the endothelium, and so on. 

    It is becoming increasingly clear that the BBB is a transcriptionally dynamic sensor of a variety of environmental stimuli, and that its gene products may functionally affect overall brain health. Whether these “sense-and-response” capabilities decline with normal aging and neurodegenerative disease may be an important area of study.

    —Andrew Chris Yang is a co-author of this comment.

    View all comments by Tony Wyss-Coray

  4. This is an interesting study providing useful data for the understanding of brain endothelial function in both physiological and pathological conditions. The study shows that in different models of neurodegenerative diseases, the transcriptional responses of brain endothelial cells are similar. Although the acute gene expression response seems to be different in each model, the changes are similar in the subacute phase, suggesting that the response of brain endothelial cells to injury might be the same despite the nature of the insult. The data are intriguing and to a certain degree surprising since the pathophysiologies of these conditions are different.

    In addition, even if we consider that dysfunction of the blood-brain barrier (BBB) occurs in all the conditions that were studied in the paper, the time course and the mode of the BBB disruptions are different. Also, it is interesting to note that the response of brain endothelial cells to injury might involve acquiring a “peripheral” phenotype. Lastly, considering recent data showing gene-expression signatures in different sections of the cerebrovascular tree, in future studies it will be interesting to understand whether the transcriptional changes of brain endothelial cells in response to injury are common to arteries, capillaries and veins. Equally interesting will be to understand whether changes to brain endothelial function prior to disease onset and BBB disruption might play a role in the pathogenesis of neurodegenerative diseases. In this regard, it is worth mentioning that a dysfunction of endothelial cells is not necessarily associated with BBB dysfunction. For example, we have shown that a deficit in the ability of brain endothelial cells to produce nitric oxide induces cognitive dysfunction through tau aggregation, despite an intact BBB (Faraco et al., 2018; Faraco et al., 2019). 

    References:

    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.

    Faraco G, Hochrainer K, Segarra SG, Schaeffer S, Santisteban MM, Menon A, Jiang H, Holtzman DM, Anrather J, Iadecola C. Dietary salt promotes cognitive impairment through tau phosphorylation. Nature. 2019 Oct;574(7780):686-690. Epub 2019 Oct 23 PubMed.

    View all comments by Giuseppe Faraco

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