Amyloidosis and Amyloidogenesis: One Name, Many Diseases.

Amyloidosis is a heterogenous group of disorders, caused by the deposition of insoluble fibrils derived from misfolded proteins in the extracellular space of various organs. These proteins have an unstable structure that causes them to misfold, aggregate, and deposit as amyloid fibrils with the pathognomonic histologic property of green birefringence when viewed under cross-polarized light after staining with Congo red. Amyloid fibrils are insoluble and degradation-resistant; resistance to catabolism results in progressive tissue amyloid accumulation. The outcome of this process is organ disfunction independently from the type of deposited protein, however there can be organ that are specifically targeted from certain proteins.

The Role of Scintigraphy with Bone Radiotracers in Cardiac Amyloidosis.

Cardiac amyloidosis (CA) is caused by the myocardial deposition of misfolded proteins, either amyloid transthyretin (ATTR) or immunoglobulin light chains (AL). The paradigm of this condition has transformed, since CA is increasingly recognized as a relatively prevalent cause of heart failure. Cardiac scintigraphy with bone tracers is the unique noninvasive technique able to confirm CA without performing tissue biopsy or advanced imaging tests. A moderate-to-intense myocardial uptake (Perugini grade ≥2) associated with the absence of a monoclonal component is greater than 99% specific for ATTR-CA, while AL-CA confirmation requires tissue biopsy.

Effects of SPI1-mediated transcriptome remodeling on Alzheimer's disease-related phenotypes in mouse models of Aß amyloidosis.

SPI1 was recently reported as a genetic risk factor for Alzheimer's disease (AD) in large-scale genome-wide association studies. However, it is unknown whether SPI1 should be downregulated or increased to have therapeutic benefits. To investigate the effect of modulating SPI1 levels on AD pathogenesis, we performed extensive biochemical, histological, and transcriptomic analyses using both Spi1-knockdown and Spi1-overexpression mouse models. Here, we show that the knockdown of Spi1 expression significantly exacerbates insoluble amyloid-ß (Aß) levels, amyloid plaque deposition, and gliosis. Conversely, overexpression of Spi1 significantly ameliorates these phenotypes and dystrophic neurites. Further mechanistic studies using targeted and single-cell transcriptomics approaches demonstrate that altered Spi1 expression modulates several pathways, such as immune response pathways and complement system. Our data suggest that transcriptional reprogramming by targeting transcription factors, like Spi1, might hold promise as a therapeutic strategy. This approach could potentially expand the current landscape of druggable targets for AD.

Western diet increases brain metabolism and adaptive immune responses in a mouse model of amyloidosis.

Diet-induced increase in body weight is a growing health concern worldwide. Often accompanied by a low-grade metabolic inflammation that changes systemic functions, diet-induced alterations may contribute to neurodegenerative disorder progression as well. This study aims to non-invasively investigate diet-induced metabolic and inflammatory effects in the brain of an APPPS1 mouse model of Alzheimer's disease. [18F]FDG, [18F]FTHA, and [18F]GE-180 were used for in vivo PET imaging in wild-type and APPPS1 mice. Ex vivo flow cytometry and histology in brains complemented the in vivo findings. 1H- magnetic resonance spectroscopy in the liver, plasma metabolomics and flow cytometry of the white adipose tissue were used to confirm metaflammatory condition in the periphery. We found disrupted glucose and fatty acid metabolism after Western diet consumption, with only small regional changes in glial-dependent neuroinflammation in the brains of APPPS1 mice. Further ex vivo investigations revealed cytotoxic T cell involvement in the brains of Western diet-fed mice and a disrupted plasma metabolome. 1H-magentic resonance spectroscopy and immunological results revealed diet-dependent inflammatory-like misbalance in livers and fatty tissue. Our multimodal imaging study highlights the role of the brain-liver-fat axis and the adaptive immune system in the disruption of brain homeostasis in amyloid models of Alzheimer's disease.

Amyloid fibril cytotoxicity and associated disorders.

Misfolded proteins assemble into fibril structures that are called amyloids. Unlike usually folded proteins, misfolded fibrils are insoluble and deposit extracellularly or intracellularly. Misfolded proteins interrupt the function and structure of cells and cause amyloid disease. There is increasing evidence that the most pernicious species are oligomers. Misfolded proteins disrupt cell function and cause cytotoxicity by calcium imbalance, mitochondrial dysfunction, and intracellular reactive oxygen species. Despite profound impacts on health, social, and economic factors, amyloid diseases remain untreatable. To develop new therapeutics and to understand the pathological manifestations of amyloidosis, research into the origin and pathology of amyloidosis is urgently needed. This chapter describes the basic concept of amyloid disease and the function of atypical amyloid deposits in them.

Therapeutic approaches in proteinopathies.

A family of maladies known as amyloid disorders, proteinopathy, or amyloidosis, are characterized by the accumulation of abnormal protein aggregates containing cross-ß-sheet amyloid fibrils in many organs and tissues. Often, proteins that have been improperly formed or folded make up these fibrils. Nowadays, most treatments for amyloid illness focus on managing symptoms rather than curing or preventing the underlying disease process. However, recent advances in our understanding of the biology of amyloid diseases have led to the development of innovative therapies that target the emergence and accumulation of amyloid fibrils. Examples of these treatments include the use of small compounds, monoclonal antibodies, gene therapy, and others. In the end, even if the majority of therapies for amyloid diseases are symptomatic, greater research into the biology behind these disorders is identifying new targets for potential therapy and paving the way for the development of more effective treatments in the future.

Deactivation of the Unfolded Protein Response Aggravated Renal AA Amyloidosis in HSF1 Deficiency Mice.

Systemic amyloid A (AA) amyloidosis, which is considered the second most common form of systemic amyloidosis usually takes place several years prior to the occurrence of chronic inflammation, generally involving the kidney. Activated HSF1, which alleviated unfolded protein response (UPR) or enhanced HSR, is the potential therapeutic target of many diseases. However, the effect of HSF1 on AA amyloidosis remains unclear. This study focused on evaluating effect of HSF1 on AA amyloidosis based on HSF1 knockout mice. As a result, aggravated amyloid deposits and renal dysfunction have been found in HSF1 knockout mice. In progressive AA amyloidosis, HSF1 deficiency enhances serum amyloid A production might to lead to severe AA amyloid deposition in mice, which may be related to deactivated unfolded protein response as well as enhanced inflammation. Thus, HSF1 plays a significant role on UPR related pathway impacting AA amyloid deposition, which can mitigate amyloidogenic proteins from aggregation pathologically and is the possible way for intervening with the pathology of systemic amyloid disorder. In conclusion, HSF1 could not only serve as a new target for AA amyloidosis treatment in the future, but HSF1 knockout mice also can be considered as a valuable novel animal model for renal AA amyloidosis.

Thermal evaporation as sample preparation for silver-assisted laser desorption/ionization mass spectrometry imaging of cholesterol in amyloid tissues.

Cholesterol plays an important biological role in the body, and its disruption in homeostasis and synthesis has been implicated in several diseases. Mapping the locations of cholesterol is crucial for gaining a better understanding of these conditions. Silver deposition has proven to be an effective method for analyzing cholesterol using mass spectrometry imaging (MSI). We optimized and evaluated thermal evaporation as an alternative deposition technique to sputtering for silver deposition in MSI of cholesterol. A silver layer with a thickness of 6 nm provided an optimal combination of cholesterol signal intensity and mass resolution. The deposition of an ultrathin nanofilm of silver enabled high-resolution MSI with a pixel size of 10 µm. We used this optimized method to visualize the distribution of cholesterol in the senile plaques in the brains of APP/PS1 mice, a model that resembles Alzheimer's disease pathology. We found that cholesterol was evenly distributed across the frontal cortex tissue, with no evidence of plaque-like accumulation. Additionally, we investigated the presence and distribution of cholesterol in myocardial sections of a human heart affected by wild-type ATTR amyloidosis. We identified the presence of cholesterol in areas with amyloid deposition, but complete colocalization was not observed.

Light chain deposition disease presenting with gastrointestinal disorder as primary manifestation: report of two cases and literature review.

Light chain deposition disease (LCDD) is an under-recognized condition characterized by deposition of abnormal monoclonal light chains in tissues, leading to organ dysfunction. LCDD involving the gastrointestinal tract is very uncommon, and its diagnosis is challenging. We herein report two cases of LCDD that manifested as inflammatory bowel disease-like symptoms and protein-losing gastroenteropathy. Both patients were women in their early 60s. Tissue biopsies from the gastrointestinal mucosa demonstrated extracellular deposits, which were negative by Congo red staining but positive for κ-light chain by immunohistochemistry. The recent literature on LCDD was reviewed. When patients unexpectedly show extracellular deposits in gastrointestinal biopsy specimens, evaluation of immunoglobulin chains is recommended for diagnosis of LCDD after systemic amyloidosis has been excluded.

Modulation of hippocampal protein expression by a brain penetrant biologic TNF-α inhibitor in the 3xTg Alzheimer's disease mice.

BACKGROUND: Biologic TNF-α inhibitors (bTNFIs) can block cerebral TNF-α in Alzheimer's disease (AD) if these macromolecules can cross the blood-brain barrier (BBB). Thus, a model bTNFI, the extracellular domain of type II TNF-α receptor (TNFR), which can bind to and sequester TNF-α, was fused with a mouse transferrin receptor antibody (TfRMAb) to enable brain delivery via BBB TfR-mediated transcytosis. Previously, we found TfRMAb-TNFR to be protective in a mouse model of amyloidosis (APP/PS1) and tauopathy (PS19), and herein we investigated its effects in mice that combine both amyloidosis and tauopathy (3xTg-AD). METHODS: Eight-month-old female 3xTg-AD mice were injected intraperitoneally with saline (n = 11) or TfRMAb-TNFR (3 mg/kg; n = 11) three days per week for 12 weeks. Age-matched wild-type (WT) mice (n = 9) were treated similarly with saline. Brains were processed for immunostaining and high-resolution multiplex NanoString GeoMx spatial proteomics. RESULTS: We observed regional differences in proteins relevant to Aß, tau, and neuroinflammation in the hippocampus of 3xTg-AD mice compared with WT mice. From 64 target proteins studied using spatial proteomics, a comparison of the Aß-plaque bearing vs. plaque-free regions in the 3xTg-AD mice yielded 39 differentially expressed proteins (DEP) largely related to neuroinflammation (39% of DEP) and Aß and tau pathology combined (31% of DEP). Hippocampal spatial proteomics revealed that the majority of the proteins modulated by TfRMAb-TNFR in the 3xTg-AD mice were relevant to microglial function (⁓ 33%). TfRMAb-TNFR significantly reduced mature Aß plaques and increased Aß-associated microglia around larger Aß deposits in the 3xTg-AD mice. Further, TfRMAb-TNFR increased mature Aß plaque-associated microglial TREM2 in 3xTg-AD mice. CONCLUSION: Overall, despite the low visual Aß load in the 11-month-old female 3xTg-AD mice, our results highlight region-specific AD-relevant DEP in the hippocampus of these mice. Chronic TfRMAb-TNFR dosing modulated several DEP involved in AD pathology and showed a largely microglia-centric mechanism of action in the 3xTg-AD mice.

Evaluation of abdominal fat pad aspiration highlighting challenges to interpretation.

INTRODUCTION: Fine-needle aspiration (FNA) of abdominal fibroadipose tissue is a commonly utilized method for the detection of amyloidosis. While generally regarded as an accurate and specific detection method, the sensitivity is variable. The objective of this study was to investigate the performance of fat pad FNAs in detecting amyloidosis relative to other tissue biopsies. MATERIALS AND METHODS: Fat pad FNA results from January 1, 2014, to December 31, 2022, were catalogued. Clinical data including FNA indication were ascertained for each case. The results of any subsequent tissue biopsy/biopsies evaluated for amyloidosis by Congo red staining were also assessed. Challenges to diagnostic interpretation were explored. RESULTS: A total of 334 fat pad FNAs were identified. The most common indications were peripheral neuropathy (29.3%), cardiomyopathy/heart failure (28.1%), monoclonal gammopathy (27.8%), and multiple myeloma/lymphoplasmacytic lymphoma (21.0%). Cytologic interpretations were: 7 (2.1%) nondiagnostic, 284 (85.0%) negative, 18 (5.4%) indeterminate, 16 (4.8%) suspicious, and 9 (2.7%) positive for amyloid deposition. In our sample, 103 (30.8%) patients had Congo red testing performed on a subsequent surgical specimen(s) including: 3 of 7 of nondiagnostic cases, none which were positive on the subsequent surgical; 70 of 284 negative cases, 27 which were positive on the subsequent surgical; 11 of 18 indeterminate cases, 7 which were positive on the subsequent surgical; 13 of 16 suspicious cases, 2 which were positive on the subsequent surgical; and 6 of 9 positive cases, 3 which were positive on the subsequent surgical. Challenges to FNA interpretation included scant cellularity, focal staining/birefringence, and overstaining. CONCLUSIONS: It is best to view fat pad aspiration versus other tissue biopsy results as complimentary diagnostic tests that should be interpreted in the context of the clinical setting and overall clinical suspicion for amyloidosis.

Morphological findings in different subtypes of hepatic amyloid.

The classic findings have been well described for light-chain amyloid involving the liver. In addition to light chain, however, many additional proteins are now known to be amyloidogenic and can involve the liver. A total of 58 surgical pathology specimens with amyloid deposits were analyzed for patterns of amyloid deposition, including amyloid from light chain lambda (N = 17), light chain kappa (N = 15), transthyretin (N = 15), serum amyloid A (N = 4), apolipoprotein A1 (N = 4), fibrinogen alpha (N = 2), LECT2 (N = 1). Amyloid deposits predominately targeted the liver vasculature, including the walls of the hepatic arteries, portal veins, and sinusoids. While there was overlap, light chain amyloid predominately involved the sinusoids, while transthyretin amyloid predominately targeted the hepatic arteries, especially the larger ones in the hilum and larger portal tracts. Serum amyloid A formed nodular deposits that started in the portal vasculature but then extended into the portal tract stroma, leading to large, bulbous, portal-based amyloid deposits. Apolipoprotein A amyloid also formed large portal-based nodules. Fibrinogen was mild and subtle on H&E and predominately affected portal veins. Amyloid deposits in hilar nerves were prominent with amyloid light chain, transthyretin, and apolipoprotein A1. In conclusion, the histology of hepatic amyloid is diverse and shows several distinct clusters of findings that can aide in recognition in surgical pathology specimens.

Telomere length and micronuclei trajectories in APP/PS1 mouse model of Alzheimer's disease: Correlating with cognitive impairment and brain amyloidosis in a sexually dimorphic manner.

Although studies have demonstrated that genome instability is accumulated in patients with Alzheimer's disease (AD), the specific types of genome instability linked to AD pathogenesis remain poorly understood. Here, we report the first characterization of the age- and sex-related trajectories of telomere length (TL) and micronuclei in APP/PS1 mice model and wild-type (WT) controls (C57BL/6). TL was measured in brain (prefrontal cortex, cerebellum, pituitary gland, and hippocampus), colon and skin, and MN was measured in bone marrow in 6- to 14-month-old mice. Variation in TL was attributable to tissue type, age, genotype and, to a lesser extent, sex. Compared to WT, APP/PS1 had a significantly shorter baseline TL across all examined tissues. TL was inversely associated with age in both genotypes and TL shortening was accelerated in brain of APP/PS1. Age-related increase of micronuclei was observed in both genotypes but was accelerated in APP/PS1. We integrated TL and micronuclei data with data on cognition performance and brain amyloidosis. TL and micronuclei were linearly correlated with cognition performance or Aß40 and Aß42 levels in both genotypes but to a greater extent in APP/PS1. These associations in APP/PS1 mice were dominantly driven by females. Together, our findings provide foundational knowledge to infer the TL and micronuclei trajectories in APP/PS1 mice during disease progression, and strongly support that TL attrition and micronucleation are tightly associated with AD pathogenesis in a female-biased manner.

Cardiac amyloid deposition and the forensic autopsy - A review and analysis.

Although amyloid material in the heart is not infrequently encountered at autopsy it may on occasion be difficult to determine the significance in terms of possible contributions to the terminal mechanisms of death. A review was undertaken of the literature and of autopsy cases at Forensic Science SA over a 20-year-period (2003-2022) for all cases where significant amyloid material had been encountered on microscopy of the heart. Sixteen cases were found consisting of 11 cases where cardiac amyloid was involved in the lethal episode, and five where it was considered an incidental feature. Of the 11 lethal cases, there were three where cardiac amyloidosis was the cause of death, and eight where it was a contributing factor, along with ischaemic heart disease (N = 7) and bronchopneumonia (N = 1). The age range was 47-92 years, average 78.6 years, with a male to female ratio of 10:1. The weights of the hearts ranged from 496 to 1059 g - average 648 g. Of the five cases where it was considered an incidental finding, the causes of death were blunt head trauma (N = 2), small intestinal ischaemia (N = 2) and small intestinal obstruction (N = 1). The weights of the hearts ranged from 299 to 487 g, average 369 g. The most relevant types of amyloidosis in forensic cases tend to be light chain amyloidosis, senile cardiac amyloidosis and familial amyloid cardiomyopathy. Other forms of amyloidosis that affect the heart, which include reactive amyloidosis, haemodialysis-related amyloidosis and isolated atrial amyloidosis, either have minimal or no clinical significance, or are of uncertain significance. While it may be difficult to determine the prognostic significance of amyloid material at autopsy clinicopathological correlation may provide useful supportive information.

Mimicking kidney flow shear efficiently induces aggregation of LECT2, a protein involved in renal amyloidosis.

Aggregation of leukocyte cell-derived chemotaxin 2 (LECT2) causes ALECT2, a systemic amyloidosis that affects the kidney and liver. Previous studies established that LECT2 fibrillogenesis is accelerated by the loss of its bound zinc ion and stirring/shaking. These forms of agitation create heterogeneous shear conditions, including air-liquid interfaces that denature proteins, that are not present in the body. Here, we determined the extent to which a more physiological form of mechanical stress-shear generated by fluid flow through a network of narrow channels-drives LECT2 fibrillogenesis. To mimic blood flow through the kidney, where LECT2 and other proteins form amyloid deposits, we developed a microfluidic device consisting of progressively branched channels narrowing from 5 mm to 20 µm in width. Shear was particularly pronounced at the branch points and in the smallest capillaries. Aggregation was induced within 24 h by shear levels that were in the physiological range and well below those required to unfold globular proteins such as LECT2. EM images suggested the resulting fibril ultrastructures were different when generated by laminar flow shear versus shaking/stirring. Importantly, results from the microfluidic device showed the first evidence that the I40V mutation accelerated fibril formation and increased both the size and the density of the aggregates. These findings suggest that kidney-like flow shear, in combination with zinc loss, acts in combination with the I40V mutation to trigger LECT2 amyloidogenesis. These microfluidic devices may be of general use for uncovering mechanisms by which blood flow induces misfolding and amyloidosis of circulating proteins.

Gastroduodenal Involvement in AL Amyloidosis: Case Report and Literature Review.

Gastrointestinal amyloidosis is a rare condition commonly found in the setting of systemic AL amyloidosis. Amyloid can deposit throughout the gastrointestinal tract and the resulting symptoms vary depending on the site of deposition. Gastrointestinal (GI) manifestations can range from weight loss or abdominal pain, to more serious complications like gastrointestinal bleeding, malabsorption, dysmotility, and obstruction. This case describes a patient with known history of IgG lambda AL amyloidosis, presenting with epigastric pain and unintentional weight loss found to have gastroduodenal amyloidosis. The definitive diagnosis of GI amyloidosis requires endoscopic biopsy with Congo red staining and visualization under polarized light microscopy. There are currently no specific guidelines for the management of GI amyloidosis. Generally, the goal is to treat the underlying cause of the amyloidosis along with symptom management. Our patient is being treated with cyclophosphamide, bortezomib, and dexamethasone (CyBorD) and started on hemodialysis due to progression of renal disease.