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Background Carpal tunnel syndrome is the most common compression syndrome of the peripheral nerve. Transthyretin amyloidosis and dialysis-related β2-microglobulin amyloidosis are known causes of carpal tunnel syndrome. Case Report A Japanese woman showed carpal tunnel syndrome 16 years after a domino liver transplantation (DLT) from the donor with hereditary transthyretin amyloidosis. DLT indication was congenital extrahepatic portosystemic shunt, and the patient had been put on maintenance hemodialysis because of chronic kidney disease 6 years before DLT. Moreover, the amyloid precursor protein of the patient was histologically confirmed not to be β2-microglobulin, but transthyretin. Conclusions The existence of amyloid was speculated when the patient who underwent DLT from hereditary transthyretin amyloidosis showed carpal tunnel syndrome. Additionally, elucidating the amyloid precursor protein when the patient has another cause of amyloidosis is necessary.

Amyloid fibrils are polymeric structures originating from aggregation of misfolded proteins. In vivo, proteolysis may modulate amyloidogenesis and fibril stability. In light chain (AL) amyloidosis, fragmented light chains (LCs) are abundant components of amyloid deposits; however, site and timing of proteolysis are debated. Identification of the N and C termini of LC fragments is instrumental to understanding involved processes and enzymes. We investigated the N and C terminome of the LC proteoforms in fibrils extracted from the hearts of two AL cardiomyopathy patients, using a proteomic approach based on derivatization of N- and C-terminal residues, followed by mapping of fragmentation sites on the structures of native and fibrillar relevant LCs. We provide the first high-specificity map of proteolytic cleavages in natural AL amyloid. Proteolysis occurs both on the LC variable and constant domains, generating a complex fragmentation pattern. The structural analysis indicates extensive remodeling by multiple proteases, largely taking place on poorly folded regions of the fibril surfaces. This study adds novel important knowledge on amyloid LC processing: although our data do not exclude that proteolysis of native LC dimers may destabilize their structure and favor fibril formation, the data show that LC deposition largely precedes the proteolytic events documentable in mature AL fibrils.

In light chain amyloidosis (AL), fibrillar deposition of monoclonal immunoglobulin light chains (LCs) in vital organs, such as heart, is associated with their severe dysfunction. In addition to the cellular damage caused by fibril deposition, direct toxicity of soluble prefibrillar amyloidogenic proteins has been reported, in particular, for cardiotoxicity. However, the molecular bases of proteotoxicity by soluble LCs have not been clarified. Here, to address this issue, we rationally engineered the amino acid sequence of the highly cardiotoxic LC H6 by introducing three residue mutations, designed to reduce the dynamics of its native state. The resulting mutant (mH6) is less toxic than its parent H6 to human cardiac fibroblasts and C. elegans. The high sequence and structural similarity, together with the different toxicity, make H6 and its non-toxic designed variant mH6 a test case to shed light on the molecular properties underlying soluble toxicity. Our comparative structural and biochemical study of H6 and mH6 shows closely matching crystal structures, whereas spectroscopic data and limited proteolysis indicate that H6 displays poorly cooperative fold, higher flexibility, and kinetic instability, and a higher dynamic state in its native fold. Taken together, the results of this study show a strong correlation between the overall conformational properties of the native fold and the proteotoxicity of cardiotropic LCs.

Muscle histology of sporadic inclusion body myositis (sIBM) demonstrates inflammatory findings and degenerative features including accumulation of TAR DNA-binding protein of 43 kDa (TDP-43). However, whether sarcoplasmic accumulation of TDP-43 is a primary trigger of muscle degeneration or a secondary event resulting from muscle degeneration in the pathophysiology of sIBM remained unclear. Our study aimed to discover whether muscle-dominant expression of TDP-43 is a primary cause of muscle degeneration. We generated several lines of wild-type TDP-43 transgenic mice driven by a creatine kinase 8 promoter, and analyzed the phenotypes via biochemical, histological, and proteomic techniques. The mice showed increased serum levels of myogenic enzymes. Muscle histology demonstrated myopathic changes including fiber size variation, abundant tubular aggregates, and TDP-43 aggregation with upregulation of endoplasmic reticulum (ER) stress. Proteomic analysis with aggregated materials in degenerative myofibers identified increased sarcoplasmic reticulum (SR)/ER-resident proteins that regulated calcium homeostasis, as well as cytosolic 5′-nucleotidase 1A. Muscle-dominant wild-type TDP-43 expression indeed caused myotoxicity featuring tubular aggregates and TDP-43-positive inclusions. Our observation suggested that TDP-43 aggregates might not be sufficient to trigger the pathogenesis of sIBM although myofiber sarcoplasmic aggregation of TDP-43 led to myofiber degeneration via ER stress and possibly calcium dysregulation, independently of inflammatory process.

To detect vascular Notch3 extracellular domain aggregates in CADASIL, we developed a novel dot-blot assay with both autopsy and biopsy skin samples. We obtained samples from 11 patients with CADASIL and 12 control patients, and we performed dot-blot analyses by using sequential biochemical tissue extractions with three different antibodies against specific regions of the Notch3 extracellular domain. We also analyzed clinical features and vascular accumulations of Notch3 by immunohistochemistry. Via the dot-blot assay with the antibody against the C-terminal region of the Notch3 extracellular domain, we successfully detected Notch3 extracellular domain aggregates in skin tissue homogenates obtained from patients with CADASIL. Our novel method may therefore aid the diagnosis of CADASIL.

Amyloidosis is a protein conformational disorder with the distinctive feature of extracellular accumulation of amyloid fibrils that come from different proteins. In the ligamentum flavum of the lumbar spine, amyloid deposits were frequently found in elderly patients with lumbar spinal canal stenosis and were at least partially formed by wild-type transthyretin. However, how amyloid deposits in the ligamentum flavum affect lumbar spinal canal stenosis has remained unclear. In this study, we analyzed clinical, pathologic, and radiologic findings of patients with lumbar spinal canal stenosis who had amyloid deposits in the ligamentum flavum. We studied 95 ligamentum flavum specimens obtained from 56 patients with lumbar spinal canal stenosis and 21 ligamentum flavum specimens obtained from 19 patients with lumbar disk herniation. We evaluated histopathologic findings and clinicoradiologic manifestations, such as thickness of the ligamentum flavum and lumbar spinal segmental instability. We found that all 95 ligamentum flavum specimens resected from patients with lumbar spinal canal stenosis had amyloid deposits, which we classified into two types, transthyretin-positive and transthyretin-negative, and that transthyretin amyloid formation in the ligamentum flavum of patients with lumbar spinal canal stenosis was an age-associated phenomenon. The amount of amyloid in the ligamentum flavum was related to clinical manifestations of lumbar spinal canal stenosis, such as thickness of the ligamentum flavum and lumbar spinal segmental instability, in the patients with lumbar spinal canal stenosis with transthyretin-positive amyloid deposits. To our knowledge, this report is the first to show clinicopathologic correlations in transthyretin amyloid deposits of the ligamentum flavum. In conclusion, transthyretin amyloid deposits in the ligamentum flavum may be related to the pathogenesis of lumbar spinal canal stenosis in elderly patients.

Familial amyloidotic polyneuropathy (FAP) is a hereditary amyloidosis induced by amyloidogenic transthyretin (ATTR). Because most transthyretin (TTR) in serum is synthesized by the liver, liver transplantation (LT) is today the only treatment available to halt the progression of FAP, even though LT is associated with several problems. Despite the urgent need to develop alternatives to LT, the detailed pathogenesis of FAP is still unknown; also, no model fully represents the relevant processes in patients with FAP. The induction of induced pluripotent stem (iPS) cells has allowed development of pluripotent cells specific for patients and has led to useful models of human diseases. Because of the need for a tool to elucidate the molecular pathogenesis of FAP, in this study we sought to establish heterozygous ATTR mutant iPS cells, and were successful, by using a Sendai virus vector mixture containing four transcription factors (Oct3/4, Sox2, Klf4, and c-Myc) to reprogram dermal fibroblasts derived from FAP patients. Moreover, FAP-specific iPS cells had the potential to differentiate into hepatocyte-like cells and indeed expressed ATTR. FAP-specific iPS cells demonstrated the possibility of serving as a pathological tool that will contribute to understanding the pathogenesis of FAP and development of FAP treatments.

Senile systemic amyloidosis is a common age-related amyloidosis that involves accumulation of wild-type transthyretin, with cardiac dysfunction being a predominant result. The importance of obtaining an accurate diagnosis of senile systemic amyloidosis has been increasingly recognized, so that novel treatments are being developed. However, the clinicopathological features of senile systemic amyloidosis remain to be completely understood. Here, we evaluated cardiac specimens from 181 consecutive post-mortem cases older than 40 years, including 6 cases of senile systemic amyloidosis, and 5 cases of familial amyloidotic polyneuropathy, which is a hereditary systemic amyloidosis caused by mutant forms of transthyretin. Furthermore, we studied ante-mortem clinicopathological findings of 11 senile systemic amyloidosis cases, in which 9 cases underwent gastrointestinal tract biopsy and/or subcutaneous tissue biopsy, at Kumamoto University Hospital. Of the autopsied cases of elderly Japanese (older than 80 years), 12% had senile systemic amyloidosis, with the percentage increasing with age. The occurrence of senile systemic amyloidosis in elderly Japanese patients was lower than that in previous reports, which suggests that a genetic background and/or environmental factor(s) may have important roles in the occurrence of senile systemic amyloidosis. Transthyretin amyloid deposits in familial amyloidotic polyneuropathy cases developed mainly in the pericardium and the surrounding muscle fascicles, whereas in cases with senile systemic amyloidosis the transthyretin amyloid deposits had a patchy plaque-like shape and developed mainly inside the ventricular wall. Biopsies from senile systemic amyloidosis patients evidenced amyloid deposits in 44% (4/9) of gastrointestinal tract and subcutaneous tissue samples combined. As myocardial biopsy may be dangerous for elderly people, the use of a combination of gastrointestinal tract and subcutaneous tissue biopsies may make diagnosis of senile systemic amyloidosis easier.

Summary Background & aims Early high nasogastric tube feeding (NTF) is effective for improving the nutritional status of critical illness. However, potential complications of NTF in tetanus include aspiration pneumonia because of dysphagia and gastrointestinal dysfunction induced by over-activity of autonomic nervous system. Methods We serially evaluated gastric myoelectrical activities using electrogastrogram (EGG) recorder in 4 elderly severe tetanus patients, and assessed its potential as a marker to increase volume and calorie of NTF. Results Although dominant frequencies of EGG in all patients (1.8 ± 0.6 cycle/min (cpm)) were lower than those in healthy volunteers (2.9 ± 0.2 cpm) at least until 12th hospital day, it tended to improve from 14–24th hospital day and reached the same levels in healthy volunteers at least until 28th hospital day. We transferred total NTF along their metabolic costs when the timing of dominant frequency tended to improve in each patient (17–24th hospital day). No aspiration pneumonia and malnutrition occurred in their hospital days. Conclusion EGG data may be a useful marker to know the level of over-activity of autonomic nervous system and to guess the best timing to increase volume and calorie of NTF especially in elderly severe tetanus.