Progression and Classification of Granular Osmiophilic Material (GOM) Deposits in Functionally Characterized Human NOTCH3 Transgenic Mice

Abstract

CADASIL is a NOTCH3-associated cerebral small vessel disease. A pathological ultrastructural disease hallmark is the presence of NOTCH3-protein containing deposits called granular osmiophilic material (GOM), in small arteries. How these GOM deposits develop over time and what their role is in disease progression is largely unknown. Here, we studied the progression of GOM deposits in humanized transgenic NOTCH3 Arg182Cys mice, compared them to GOM deposits in patient material, and determined whether GOM deposits in mice are associated with a functional CADASIL phenotype. We found that GOM deposits are not static, but rather progress in ageing mice, both in terms of size and aspect. We devised a GOM classification system, reflecting size, morphology and electron density. Six-month-old mice showed mostly early stage GOM, whereas older mice and patient vessels showed predominantly advanced stage GOM, but also early stage GOM. Mutant mice did not develop the most severe GOM stage seen in patient material. This absence of end-stage GOM in mice was associated with an overall lack of histological vascular pathology, which may explain why the mice did not reveal functional deficits in cerebral blood flow, cognition and motor function. Taken together, our data indicate that GOM progress over time, and that new GOM deposits are continuously being formed. The GOM staging system we introduce here allows for uniform GOM deposit classification in future mouse and human studies, which may lead to more insight into a potential association between GOM stage and CADASIL disease severity, and the role of GOM in disease progression.

Notch3 Signaling and Aggregation as Targets for the Treatment of CADASIL and Other NOTCH3-Associated Small-Vessel Diseases

“…45,47,51,52 In agreement with this notion, the number, size, and morphology of GOM deposits evolve through the course of the disease. 53 In vitro studies have demonstrated reduced clearance and decreased solubility of mutant Notch3 protein complexes compared with wild Notch3 in Small-Vessel Disease…”

“…66 Conversely, a different CADASIL mouse model overexpressing high levels of mutant Notch3, the tgN3 MUT 350 mouse, presented important GOM deposition but failed to develop other central disease features, including mural cell degeneration, basement membrane thickening, or changes in cerebrovascular reactivity and cognition. 53,58 The apparent lack of robust association between GOM and vessel structural or functional integrity in animal studies might be secondary to limitations of current animal models in accurately replicating the full spectrum of CADASIL pathophysiological mechanisms or of methods to detect and quantify GOM in these models.…”

Pathophysiological Mechanisms and Potential Therapeutic Targets in Cerebral Autosomal Dominant Arteriopathy With Subcortical Infarcts and Leukoencephalopathy (CADASIL)

“…The authors found a progression from initially small round GOMs, located to the abluminal side of mural cells, to large amorphous deposits which induced basement membrane bulging and mural cell indentation, seen in the last stages of the disease, and proposed a GOM classification system divided into five stages of progression. Notably, both in aged mice and in post-mortem CADASIL brain microvessels, large and amorphic GOM deposits coexisted with small and recent ones, suggesting that GOMs are continuously formed during disease progression (Gravesteijn et al, 2019).…”

“…However, GOM formation and development over time and their role in disease progression remain largely unknown. A recent study found that GOM deposits are not static, but rather change in terms of size, morphology, and number during disease course in CADASIL transgenic mice (Gravesteijn et al, 2019). The authors found a progression from initially small round GOMs, located to the abluminal side of mural cells, to large amorphous deposits which induced basement membrane bulging and mural cell indentation, seen in the last stages of the disease, and proposed a GOM classification system divided into five stages of progression.…”

CADASIL from Bench to Bedside: Disease Models and Novel Therapeutic Approaches

“…TgNotch3 R182C mice, developed by using a BAC clone containing the human NOTCH3 gene and subsequently introducing the R182C variant by two-step Red-mediated recombination, showed progressively increasing age- and NOTCH3 RNA expression level-dependent vascular accumulation of NOTCH3 and GOM deposits, which progress over time and are continuously being formed, without brain parenchyma lesions occurrence [ 65 ]. This finding led authors to realize the “NOTCH3 score” as quantitative biomarker for CADASIL, attempting to transform transgenic CADASIL mice in a proper model for pre-clinical testing of therapeutic approaches [ 66 ].…”