Does a Rare Fibronectin Variant Protect Against APOE4?

APOE4 strongly increases the risk of sporadic Alzheimer’s disease. Yet some carriers evade the disease entirely. Now, scientists led by Richard Mayeux, Caghan Kizil, and Badri Vardarajan of Columbia University in New York peg some of that protection to a missense mutation in the gene for fibronectin. In the April 10 Acta Neuropathologica, they reported that APOE4 carriers with a glycine-to-glutamic-acid mutation in fibronectin were 71 percent less likely to get AD. If they did get the disease, it was 3.5 years later than expected. In a neuropathology cohort, APOE4 carriers who had been cognitively healthy had less fibronectin surrounding blood vessels in the brain than did those who had died having AD. Though these resilient cases did not carry the protective fibronectin mutation, the data supports the authors’ contention that the variant limits pathological accumulation of the extracellular matrix protein.

  • Some APOE4 carriers are resilient to Alzheimer’s.
  • A glycine-to-glutamic-acid substitution at amino acid 357 of fibronectin reduces their risk of AD.
  • Resilient E4 carriers had limited fibronectin surrounding blood vessels and minimal gliosis in the brain.

Joel Blanchard and Braxton Schuldt at the Icahn School of Medicine at Mount Sinai, New York, called the work exciting. “While previous literature strongly supports a link between the extracellular matrix and AD, Bhattarai and colleagues uniquely introduce APOE4 to this discussion and highlight potential protective variants,” they wrote (comment below).

One copy of APOE4 increases AD risk two- to threefold, while having two alleles ups risk 10 to 15 times. Still, some E4 carriers stay sharp into their 70s and 80s. In search of protective gene variants that might explain this resiliency, co-first authors Prabesh Bhattarai and Tamil Iniyan Gunasekaran at Columbia analyzed whole-genome sequencing data on 2,430 people 60 years or older from three cohorts: the National Institute on Aging Alzheimer's Disease Family Based Study (NIA-AD FBS), the Washington Heights/Inwood Columbia Aging Project (WHICAP), and the Estudio Familiar de Influencia Genetica en Alzheimer (EFIGA) cohort. Of these, 379 people had two copies of APOE4 and 1,162 people had one. Half of the participants had AD.

The authors searched for rare coding variants, i.e., frequencies below 1 percent. They identified 510 mutations in 476 genes among cognitively healthy APOE4/4 carriers that were not found in the E4 carriers with AD, or in the E4 noncarriers, hinting that the variants protect. Of these, 56 lay in proteins found in the extracellular matrix (ECM), especially the basement membrane of the blood-brain barrier. The ECM surrounding BBB vessels thickens during AD (Lepelletier et al., 2015; reviewed by Sun et al., 2021).

Bhattarai and colleagues focused on the rs140926439 missense coding variant in the fibronectin gene, FN1, because it stuck out as a top hit in all three cohorts. The variant also caught the attention of co-first author Michael Belloy, at the Washington University School of Medicine in St. Louis, because he had independently found that the mutation was protective in three additional cohorts: the Alzheimer’s Disease Genetic Consortium (ADGC), the Alzheimer’s Disease Sequencing Project (ADSP), and the UK Biobank. Among 7,185 people 60 or older in those cohorts who carried two copies of APOE4, rs140926439 carriers had 71 percent less chance of getting dementia. In rs140926439 carriers who did have AD, they developed symptoms 3.5 years later than noncarriers, on average. The researchers decided to combine their results.

How does the variant protect? Three pathogenicity algorithms, SIFT, REVEL, and MetaL R, predicted the glycine-to-glutamic-acid mutation would alter the structure of the protein sufficiently to affect function. To test this idea, the scientists turned to zebrafish because they had extensively characterized the effect of amyloidosis on the BBB in these animals and because the barrier is highly conserved among vertebrates. After injecting Aβ42 into the cerebral ventricle, amyloid accumulates in neurons throughout the zebrafish brain, microglia become activated, synapses degenerate, and astroglia detach from blood vessels (Bhattarai et al., 2016; Lee et al., 2022). Brain vascular smooth muscle also churns out 20 percent more fibronectin.

Thin Fibronectin Protects. Around blood vessels in APOE3 carriers (left), microglia (blue) clear amyloid plaques (black chevrons) with the help of astrocytes (green) that communicate with the vessels. In APOE4 carriers (middle), the extracellular matrix (gray) surrounding blood vessels thickens, limiting plaque clearance and hindering astrocyte-blood vessel connections. In E4 carriers with the fibronectin variant (right), the extracellular matrix remains thin, allowing amyloid clearance and astrocyte contacts. [Courtesy of Bhattarai et al., Acta Neuropathologica, 2024.]

When the scientists knocked out FN1B, the zebrafish ortholog, then injected Aβ42, the peptide roused 25 percent fewer astroglia, especially around the brain vasculature, and 60 percent more phagocytic microglia were retained. The zebrafish also had as many synapses as wild-types. To the authors, this suggested that fibronectin loss-of-function improves glial responses to amyloid to protect neurons, possibly by thinning the ECM and allowing the cells to clear more amyloid (image above).

Protected Vessels? Astrocytes (red) and fibronectin (green) surround blood vessels in the brains of APOE3/3 (left) and E4/4 (right) carriers who are cognitively healthy. A thicker ECM and a greater number of astrocytes surround vessels in APOE4/4 carriers who had had AD (middle). [Courtesy of Bhattarai et al., Acta Neuropathologica, 2024.]

There is some evidence that fibronectin might hamper amyloid clearance in people, too. In prefrontal cortex tissue from eight APOE4/4 carriers who had had AD, immunostaining showed 27 percent more fibronectin surrounding BBB basement membrane than in 11 APOE3/3 AD cases. Further, in six APOE4 homozygotes who did not have AD, vessels looked like those from cognitively normal E3s. Together, the findings hint that maintaining a thin ECM around BBB vessels protects against AD (image at right). Notably, none of the brain tissue donors carried the rare FN1 rs140926439 variant.

The findings tie in with prior work that fibronectin thickening around BBB vessels exacerbates cerebral amyloid angiopathy and vascular damage (Wyss-Coray et al., 2000). “Together, these [studies] suggest that vascular fibronectin accumulation and basement membrane thickening may promote pathological vascular Aβ, and that mutations preventing this could be protective,” wrote Andrew Yang of the University of California, San Francisco.—Chelsea Weidman Burke

Comments

  1. APOE4 is the strongest risk factor for Alzheimer’s disease. APOE4 homozygosity was recently identified as a distinct, highly penetrant form of AD. However, a small percentage of APOE4 homozygotes live cognitively normal lives into their 80s and beyond (Fortea et al., 2024). 

    The mechanisms that protect against APOE4 and other forms of AD are poorly understood, limiting our ability to develop therapeutics and lifestyle interventions that promote healthy cognitive aging in at-risk populations. Bhattarai et al. further our understanding of APOE4-resilience biology by exploring the genetic signatures of the AD-resilient population—APOE4 carriers who did not develop the disease. Identifying genetic factors protecting the highest-risk population will provide insight into the molecular mechanisms underlying risk and resilience to AD.

    Intriguingly, the authors found that many variants in the APOE4-resilient population are in genes associated with the extracellular matrix (ECM). They then focus on a particular loss-of-function variant in the gene encoding the ECM protein fibronectin, finding that this variant reduces the odds ratio of developing AD by approximately 70 percent, and delays disease onset by 3.37 years in APOE4 carriers. Seeking to validate their genetic findings, Bhattarai and colleagues turned to the postmortem human brain, where they found that APOE4 carriers had significantly higher fibronectin deposition along the blood-brain barrier compared to APOE3 individuals. Interestingly, when accounting for AD status, APOE4 carriers with AD had significantly more fibronectin deposition compared to APOE4 individuals who are cognitively unaffected. Hypothesizing that fibronectin may be a driver of AD pathology, the authors found that inducing a loss-of-function mutation in fibronectin in zebrafish had a protective effect on amyloid toxicity, specifically via decreased gliosis, increased microglial activity, and enhanced gliovascular remodeling. In summary, the work presented by Bhattarai et al. suggests that fibronectin is linked to AD progression in APOE4 carriers, thus highlighting the therapeutic potential of targeting this protein.

    The results presented here are particularly exciting given what is already known about the role of ECM dysregulation in AD. Studies show that the deposition of several ECM components, including fibronectin, increases with AD, and that fibronectin deposition in the postmortem human brain has a significant positive correlation with amyloid pathology (Bogdan et al., 2022; Lepelletier et al., 2017; Végh et al., 2014). Furthermore, fibronectin directly binds to amyloid and induces amyloid accumulation along the vasculature when injected into mice (Howe et al., 2018). While previous literature strongly supports a link between the ECM and AD, Bhattarai and colleagues uniquely introduce APOE4 to this discussion and highlight potential protective variants.

    From this work, several important questions emerge. Research should aim to characterize the mechanism through which APOE4 mediates an increase in fibronectin deposition and its subsequent contribution to the development of AD pathology in human brain tissue. Additionally, further studies are needed to dissect the molecular mechanism underlying the protective effect of this fibronectin variant in the APOE4 population. Overall, it is clear from this study that ECM dysregulation is emerging as a potential driver of APOE4-driven AD progression, and this field of study will likely expand in the future.

    References:

    Fortea J, Pegueroles J, Alcolea D, Belbin O, Dols-Icardo O, Vaqué-Alcázar L, Videla L, Gispert JD, Suárez-Calvet M, Johnson SC, Sperling R, Bejanin A, Lleó A, Montal V. APOE4 homozygozity represents a distinct genetic form of Alzheimer's disease. Nat Med. 2024 May;30(5):1284-1291. Epub 2024 May 6 PubMed. Correction.

    Bogdan S, Puścion-Jakubik A, Klimiuk K, Socha K, Kochanowicz J, Gorodkiewicz E. The Concentration of Fibronectin and MMP-1 in Patients with Alzheimer's Disease in Relation to the Selected Antioxidant Elements and Eating Habits. J Clin Med. 2022 Oct 27;11(21) PubMed.

    Howe MD, Atadja LA, Furr JW, Maniskas ME, Zhu L, McCullough LD, Urayama A. Fibronectin induces the perivascular deposition of cerebrospinal fluid-derived amyloid-β in aging and after stroke. Neurobiol Aging. 2018 Dec;72:1-13. Epub 2018 Aug 9 PubMed.

    Lepelletier FX, Mann DM, Robinson AC, Pinteaux E, Boutin H. Early changes in extracellular matrix in Alzheimer's disease. Neuropathol Appl Neurobiol. 2015 Nov 6; PubMed.

    Végh MJ, Rausell A, Loos M, Heldring CM, Jurkowski W, van Nierop P, Paliukhovich I, Li KW, del Sol A, Smit AB, Spijker S, van Kesteren RE. Hippocampal extracellular matrix levels and stochasticity in synaptic protein expression increase with age and are associated with age-dependent cognitive decline. Mol Cell Proteomics. 2014 Nov;13(11):2975-85. Epub 2014 Jul 20 PubMed.

    View all comments by Joel Blanchard View all comments by Braxton Schuldt
    • User Profile Image Steve Barger
      University of Arkansas for Medical Sciences
    • Posted:

    TGFꞵ1 is well-established as a contributor to cerebral amyloid angiopathy (CAA) (Wyss-Coray et al., 1997), and one of the earliest actions noted for TGFꞵ was induction of fibronectin expression (Ignotz and Massagué, 1986). It seems quite likely that this explains the TGFꞵ effect on CAA. Perhaps this relationship links to ApoE through its receptors, which generally suppress TGFꞵ actions. Cells deficient in ApoER2 show a dramatically exaggerated induction of fibronectin in response to TGFꞵ (Komaravolu et al., 2019), and deficiencies in LRP1 (Boucher et al., 2007) or VLDLR (Ma et al., 2021) are similarly permissive for vascular fibrosis. TGFꞵ1 (Huang et al., 2003) and -2 (Muratoglu et al., 2011) are both ligands for LRP1; indeed, LRP1 is also known as “TGFβ receptor V.”

    A loss-of-function phenotype for ApoE4 at one or more of these receptors is perhaps the simplest explanation. But there are considerable data indicating that the relationship of APOE ε4 genotype to AD risk is mediated by a gain of (toxic) function. Joachim Herz and colleagues have emphasized the tendency of ApoE4 to “distract” its receptors within the cell’s interior, reducing the levels of functional receptor on the cell surface (Chen et al., 2010). Like the complicated impact of mutations on presenilin/γ-secretase, this is a loss masquerading as a gain; it seems to be a plausible explanation for the apparent ability of ApoE4 to suppress its active receptors and thus promote TGFꞵ actions on fibrosis.

    References:

    Wyss-Coray T, Masliah E, Mallory M, McConlogue L, Johnson-Wood K, Lin C, Mucke L. Amyloidogenic role of cytokine TGF-beta1 in transgenic mice and in Alzheimer's disease. Nature. 1997 Oct 9;389(6651):603-6. PubMed.

    Ignotz RA, Massagué J. Transforming growth factor-beta stimulates the expression of fibronectin and collagen and their incorporation into the extracellular matrix. J Biol Chem. 1986 Mar 25;261(9):4337-45. PubMed.

    Komaravolu RK, Waltmann MD, Konaniah E, Jaeschke A, Hui DY. ApoER2 (Apolipoprotein E Receptor-2) Deficiency Accelerates Smooth Muscle Cell Senescence via Cytokinesis Impairment and Promotes Fibrotic Neointima After Vascular Injury. Arterioscler Thromb Vasc Biol. 2019 Oct;39(10):2132-2144. Epub 2019 Aug 15 PubMed.

    Boucher P, Li WP, Matz RL, Takayama Y, Auwerx J, Anderson RG, Herz J. LRP1 functions as an atheroprotective integrator of TGFbeta and PDFG signals in the vascular wall: implications for Marfan syndrome. PLoS One. 2007 May 16;2(5):e448. PubMed.

    Ma X, Takahashi Y, Wu W, Chen J, Dehdarani M, Liang W, Shin YH, Benyajati S, Ma JX. Soluble very low-density lipoprotein receptor (sVLDLR) inhibits fibrosis in neovascular age-related macular degeneration. FASEB J. 2021 Dec;35(12):e22058. PubMed.

    Huang SS, Ling TY, Tseng WF, Huang YH, Tang FM, Leal SM, Huang JS. Cellular growth inhibition by IGFBP-3 and TGF-beta1 requires LRP-1. FASEB J. 2003 Nov;17(14):2068-81. PubMed.

    Muratoglu SC, Belgrave S, Lillis AP, Migliorini M, Robinson S, Smith E, Zhang L, Strickland DK. Macrophage LRP1 suppresses neo-intima formation during vascular remodeling by modulating the TGF-β signaling pathway. PLoS One. 2011;6(12):e28846. Epub 2011 Dec 9 PubMed.

    Chen Y, Durakoglugil MS, Xian X, Herz J. ApoE4 reduces glutamate receptor function and synaptic plasticity by selectively impairing ApoE receptor recycling. Proc Natl Acad Sci U S A. 2010 Jun 29;107(26):12011-6. Epub 2010 Jun 14 PubMed.

    View all comments by Steve Barger

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References

Mutations Citations

  1. APOE C130R (ApoE4)

Paper Citations

  1. Lepelletier FX, Mann DM, Robinson AC, Pinteaux E, Boutin H. Early changes in extracellular matrix in Alzheimer's disease. Neuropathol Appl Neurobiol. 2015 Nov 6; PubMed.
  2. Sun Y, Xu S, Jiang M, Liu X, Yang L, Bai Z, Yang Q. Role of the Extracellular Matrix in Alzheimer's Disease. Front Aging Neurosci. 2021;13:707466. Epub 2021 Aug 27 PubMed.
  3. Bhattarai P, Thomas AK, Cosacak MI, Papadimitriou C, Mashkaryan V, Froc C, Reinhardt S, Kurth T, Dahl A, Zhang Y, Kizil C. IL4/STAT6 Signaling Activates Neural Stem Cell Proliferation and Neurogenesis upon Amyloid-β42 Aggregation in Adult Zebrafish Brain. Cell Rep. 2016 Oct 18;17(4):941-948. PubMed.
  4. Lee AJ, Raghavan NS, Bhattarai P, Siddiqui T, Sariya S, Reyes-Dumeyer D, Flowers XE, Cardoso SA, De Jager PL, Bennett DA, Schneider JA, Menon V, Wang Y, Lantigua RA, Medrano M, Rivera D, Jiménez-Velázquez IZ, Kukull WA, Brickman AM, Manly JJ, Tosto G, Kizil C, Vardarajan BN, Mayeux R. FMNL2 regulates gliovascular interactions and is associated with vascular risk factors and cerebrovascular pathology in Alzheimer's disease. Acta Neuropathol. 2022 Jul;144(1):59-79. Epub 2022 May 24 PubMed.
  5. Wyss-Coray T, Lin C, Sanan DA, Mucke L, Masliah E. Chronic overproduction of transforming growth factor-beta1 by astrocytes promotes Alzheimer's disease-like microvascular degeneration in transgenic mice. Am J Pathol. 2000 Jan;156(1):139-50. PubMed.

Further Reading

No Available Further Reading

Primary Papers

  1. Bhattarai P, Gunasekaran TI, Belloy ME, Reyes-Dumeyer D, Jülich D, Tayran H, Yilmaz E, Flaherty D, Turgutalp B, Sukumar G, Alba C, McGrath EM, Hupalo DN, Bacikova D, Le Guen Y, Lantigua R, Medrano M, Rivera D, Recio P, Nuriel T, Ertekin-Taner N, Teich AF, Dickson DW, Holley S, Greicius M, Dalgard CL, Zody M, Mayeux R, Kizil C, Vardarajan BN. Rare genetic variation in fibronectin 1 (FN1) protects against APOEε4 in Alzheimer's disease. Acta Neuropathol. 2024 Apr 10;147(1):70. PubMed.

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