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ATTR amyloidosis is caused by deposition of large, insoluble aggregates (amyloid fibrils) of cross-β-sheet TTR protein molecules on the intercellular surfaces of tissues. The process of amyloid formation from monomeric TTR protein molecules to amyloid deposits has not been fully characterized and is therefore modeled in this paper. Two models are considered: 1) TTR monomers in the blood spontaneously fold into a β-sheet conformation, aggregate into short proto-fibrils that then circulate in the blood until they find a complementary tissue where the proto-fibrils accumulate to form the large, insoluble amyloid fibrils found in affected tissues. 2) TTR monomers in the native or β-sheet conformation circulate in the blood until they find a tissue binding site and deposit in the tissue or tissues forming amyloid deposits in situ. These models only differ on where the selection for β-sheet complementarity occurs, in the blood where wt-wt, wt-v, and v-v interactions determine selectivity, or on the tissue surface where tissue-wt and tissure-v interactions also determine selectivity. Statistical modeling in both cases thus involves selectivity in fibril aggregation and tissue binding. Because binding of protein molecules into fibrils and binding of fibrils to tissues occurs through multiple weak non-covalent bonds, strong complementarity between β-sheet molecules and between fibrils and tissues is required to explain the insolubility and tissue selectivity of ATTR amyloidosis. Observation of differing tissue selectivity and thence disease phenotypes from either pure wildtype TTR protein or a mix of wildtype and variant molecules in amyloid fibrils evidences the requirement for fibril-tissue complementarity. Understanding the process that forms fibrils and binds fibrils to tissues may lead to new possibilities for interrupting the process and preventing or curing ATTR amyloidosis., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Criddle et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: We have previously shown that transthyretin (TTR) amyloidosis patients have amyloid fibrils of either of two compositions; type A fibrils consisting of large amounts of C-terminal TTR fragments in addition to full-length TTR, or type B fibrils consisting of only full-length TTR. Since type A fibrils are associated with an older age in ATTRVal30Met (p.Val50Met) amyloidosis patients, it has been discussed if the TTR fragments are derived from degradation of the amyloid deposits as the patients are aging. The present study aimed to investigate if the fibril composition type changes over time, especially if type B fibrils can shift to type A fibrils as the disease progresses., Material and Methods: Abdominal adipose tissue biopsies from 29 Swedish ATTRVal30Met amyloidosis patients were investigated. The fibril type in the patients´ initial biopsy taken for diagnostic purposes was compared to a biopsy taken several years later (ranging between 2 and 13 years). The fibril composition type was determined by western blot., Results: All 29 patients had the same fibril composition type in both the initial and the follow-up biopsy (8 type A and 21 type B). Even patients with a disease duration of more than 12 years and an age over 75 years at the time of the follow-up biopsy had type B fibrils in both biopsies., Discussion: The result clearly shows that the amyloid fibril composition containing large amounts of C-terminal fragments (fibril type A) is a consequence of other factors than a slow degradation process occurring over time., Competing Interests: The authors have declared that no competing interests exist.

Background: The amyloid fibril in hereditary transthyretin (TTR) Val30Met (pVal50Met) amyloid (ATTR Val30Met) amyloidosis is composed of either a mixture of full-length and TTR fragments (Type A) or of only full-length TTR (Type B). The type of amyloid fibril exerts an impact on the phenotype of the disease, and on the outcome of diagnostic procedures and therapy. The aim of the present study was to investigate if the type of amyloid fibril remains the same within ATTR Val30Met amyloidosis families., Methods: Fifteen families were identified in whom at least two first-degree relatives had their amyloid fibril composition determined. The type of ATTR was determined by Western blot in all but two patients. For these two patients a positive 99mTc-3,3-diphosphono-1,2-propanodicarboxylic acid scintigraphy indicated ATTR Type A., Results: In 14 of the 15 families, the same amyloid fibril composition was noted irrespective of differences in age at onset. In the one family, different ATTR fibril types was found in two brothers with similar ages at onset., Conclusions: Family predisposition appears to have an impact on amyloid fibril composition in members of the family irrespective of their age at onset of disease, but if genetically determined, the gene/genes are likely to be situated at another location than the TTR gene in the genome., Competing Interests: The authors have declared that no competing interests exist.

ATTRV30M amyloid neuropathy is a lethal autosomal dominant sensorimotor and autonomic neuropathy, caused by deposition of amyloid fibrils composed of aberrant transthyretin (TTR). Ages of onset and penetrance exhibit great variability and genetic factors have been implicated. Complement activation co-localizes with amyloid deposits in amyloidotic neuropathy and is possibly involved in the kinetics of amyloidogenesis. A candidate gene approach has recently identified C1q polymorphisms to correlate with disease onset in a Cypriot cohort of patients with ATTRV30M amyloid neuropathy. In the current study we use a double transgenic mouse model of ATTRV30M amyloid neuropathy in which C1q is ablated to elucidate further a possible modifier role for C1q. Amyloid deposition is found to be increased by 60% in the absence of C1q. Significant up regulation is also recorded in apoptotic and cellular stress markers reflecting extracellular toxicity of pre-fibrillar and fibrillar TTR. Our data further indicate that in the absence of C1q there is marked reduction of macrophages in association with amyloid deposits and thus less effective phagocytosis of TTR.

We hypothesized that tissue-resident macrophages in familial amyloid polyneuropathy (FAP) patients will exhibit qualitative or quantitative abnormalities, that may accelerate transthyretin (TTR)-derived amyloid deposition. To evaluate this, we examined the number and subset of tissue-resident macrophages in heart tissue from amyloid-deposited FAP and control patients. In both FAP and control patients, tissue-resident macrophages in heart tissue were all Iba+/CD163+/CD206+ macrophages. However, the number of macrophages was significantly decreased in FAP patients compared with control patients. Furthermore, the proportion of intracellular TTR in CD14+ monocytes was reduced in peripheral blood compared with healthy donors. Based on these results, we next examined degradation and endocytosis of TTR in human induced pluripotent stem (iPS) cell-derived myeloid lineage cells (MLs), which function like macrophages. iPS-MLs express CD163 and CD206, and belong to the inhibitory macrophage category. In addition, iPS-MLs degrade both native and aggregated TTR in a cell-dependent manner in vitro. Further, iPS-MLs endocytose aggregated, and especially polymerized, TTR. These results suggest that decreased tissue-localized macrophages disrupt clearance of TTR-derived amyloid deposits, leading to progression of a pathological condition in FAP patients. To improve this situation, clinical application of pluripotent stem cell-derived MLs may be useful as an approach for FAP therapy., Competing Interests: The authors have declared that no competing interests exist.

Purpose: Transthyretin V30M (ATTR V30M) amyloidosis is a phenotypically diverse disease with symptoms ranging from predominant neuropathy to exclusive cardiac manifestations. The aims of this study were to determine the dispersion of the two types of fibrils found in Swedish ATTR V30M patients -Type A consisting of a mixture of truncated and full length ATTR fibrils and type B fibrils consisting of full length fibrils, and to estimate the severity of cardiac dysfunction in relation to fibril composition and sex., Material and Methods: Echocardiographic data were analysed in 107 Swedish ATTR V30M patients with their fibril composition determined as either type A or type B. Measurements of left ventricular (LV) dimensions and evaluation of systolic and diastolic function including speckle tracking derived strain were performed. Patients were grouped according to fibril type and sex. Multivariate linear regression was utilised to determine factors of significant impact on LV thickness., Results: There was no significant difference in proportions of the two types of fibrils between men and women. In patients with type A fibrils, women had significantly lower median septal (p = 0.007) and posterior wall thicknesses (p = 0.010), lower median LV mass indexed to height (p = 0.008), and higher septal strain (p = 0.037), as compared to males. These differences were not apparent in patients with type B fibrils. Multiple linear regression analysis revealed that fibril type, sex and age all had significant impact on LV septal thickness., Conclusion: This study demonstrates a clear difference between sexes in the severity of amyloid heart disease in ATTR V30M amyloidosis patients. Even though type A fibrils were associated with more advanced amyloid heart disease compared to type B, women with type A fibrils generally developed less cardiac infiltration than men. The differences may explain the better outcome for liver transplanted late-onset female patients compared to males.

Transthyretin amyloidosis is a conformational pathology characterized by the extracellular formation of amyloid deposits and the progressive impairment of the peripheral nervous system. Point mutations in this tetrameric plasma protein decrease its stability and are linked to disease onset and progression. Since non-mutated transthyretin also forms amyloid in systemic senile amyloidosis and some mutation bearers are asymptomatic throughout their lives, non-genetic factors must also be involved in transthyretin amyloidosis. We discovered, using a differential proteomics approach, that extracellular chaperones such as fibrinogen, clusterin, haptoglobin, alpha-1-anti-trypsin and 2-macroglobulin are overrepresented in transthyretin amyloidosis. Our data shows that a complex network of extracellular chaperones are over represented in human plasma and we speculate that they act synergistically to cope with amyloid prone proteins. Proteostasis may thus be as important as point mutations in transthyretin amyloidosis.

Background: Serum amyloid P component (SAP) is a glycoprotein that is universally found associated with different types of amyloid deposits. It has been suggested that it stabilizes amyloid fibrils and therefore protects them from proteolytic degradation., Methodology/principal Findings: In this paper, we show that SAP binds not only to mature amyloid fibrils but also to early aggregates of amyloidogenic mutants of the plasma protein transthyretin (TTR). It does not inhibit fibril formation of TTR mutants, which spontaneously form amyloid in vitro at physiological pH. We found that SAP prevents cell death induced by mutant TTR, while several other molecules that are also known to decorate amyloid fibrils do not have such effect. Using a Drosophila model for TTR-associated amyloidosis, we found a new role for SAP as a protective factor in inhibition of TTR-induced toxicity. Overexpression of mutated TTR leads to a neurological phenotype with changes in wing posture. SAP-transgenic flies were crossed with mutated TTR-expressing flies and the results clearly confirmed a protective effect of SAP on TTR-induced phenotype, with an almost complete reduction in abnormal wing posture. Furthermore, we found in vivo that binding of SAP to mutated TTR counteracts the otherwise detrimental effects of aggregation of amyloidogenic TTR on retinal structure., Conclusions/significance: Together, these two approaches firmly establish the protective effect of SAP on TTR-induced cell death and degenerative phenotypes, and suggest a novel role for SAP through which the toxicity of early amyloidogenic aggregates is attenuated.

Background: Familial amyloidotic polyneuropathy (FAP) is a neurodegenerative disease caused by the extracellular deposition of mutant transthyretin (TTR), with special involvement of the peripheral nervous system (PNS). Currently, hepatic transplantation is considered the most efficient therapy to halt the progression of clinical symptoms in FAP since more than 95% of TTR is produced by the liver. However, less invasive and more reliable therapeutic approaches have been proposed for FAP therapy, namely based on drugs acting as inhibitors of amyloid formation or as amyloid disruptors. We have recently reported that epigallocatechin-3-gallate (EGCG), the most abundant catechin in green tea, is able to inhibit TTR aggregation and fibril formation, "in vitro" and in a cellular system, and is also able to disrupt pre-formed amyloid fibrils "in vitro"., Methodology and Principal Findings: In the present study, we assessed the effect of EGCG subchronic administration on TTR amyloidogenesis "in vivo", using well characterized animal models for FAP. Semiquantitative immunohistochemistry (SQ-IHC) and Western blot analysis of mice tissues after treatment demonstrated that EGCG inhibits TTR toxic aggregates deposition in about 50% along the gastrointestinal tract (GI) and peripheral nervous system (PNS). Moreover EGCG treatment considerably lowered levels of several biomarkers associated with non-fibrillar TTR deposition, namely endoplasmic reticulum (ER)-stress, protein oxidation and apoptosis markers. Treatment of old FAP mice with EGCG resulted not only in the decrease of non-fibrillar TTR deposition but also in disaggregation of amyloid deposits. Consistently, matrix metalloproteinase (MMP)-9 and serum amyloid P component (SAP), both markers of amyloid deposition, were also found reduced in treated old FAP mice., Conclusions and Significance: The dual effect of EGCG both as TTR aggregation inhibitor and amyloid fibril disruptor together with the high tolerability and low toxicity of EGCG in humans, point towards the potential use of this compound, or optimized derivatives, in the treatment of TTR-related amyloidoses.