👤 Hiroyuki Ichida

Juvenile neuronal ceroid lipofuscinosis (JNCL) is a neurodegenerative lysosomal storage disease caused by the loss of the endolysosomal transmembrane protein, CLN3. The main protein component of lysosomal storage material in JNCL is subunit c of mitochondrial ATP synthase (SCMAS), which is normally degraded within the lysosome by tripeptidyl-peptidase 1 (TPP1) during mitophagy. Previous studies have shown that TPP1 expression is elevated in JNCL, a potential compensatory response, while reduced levels of TPP1 exacerbate disease in a JNCL mouse model. These observations suggest a role for TPP1 in JNCL pathogenesis, and it is possible that lysosomal perturbations from the loss of CLN3 in JNCL could reduce the ability of TPP1 to degrade SCMAS. To test this hypothesis, we introduced a transgene that overexpresses TPP1 in a mouse model of JNCL and find that constitutively elevated TPP1 prevents SCMAS storage. This is associated with correction or significant reduction of other phenotypes of disease including neuroinflammation, an elevated plasma biomarker of neurodegeneration, and a disease-associated loss of brain mass with aging. From a clinical perspective, these results suggest that TPP1 augmentation could be a viable therapeutic strategy for JNCL and other lysosomal diseases that accumulate SCMAS where addressing the primary defect may be difficult or impossible. Show less

Nabumetone, a nonsteroidal anti-inflammatory prodrug, is converted to a pharmacologically active metabolite, 6-methoxy-2-naphthylacetic acid (6-MNA); however, it is 11-fold more efficiently converted to 4-(6-methoxy-2-naphthyl)butan-2-ol (MNBO) via a reduction reaction in human hepatocytes. The goal of this study was to identify the enzyme(s) responsible for MNBO formation from nabumetone in the human liver. MNBO formation by human liver microsomes (HLM) was 5.7-fold higher than in the liver cytosol. In a panel of 24 individual HLM samples with quantitative proteomics data, the 17β-hydroxysteroid dehydrogenase 12 (HSD17B12) protein level had the high correlation coefficient (r = 0.80, P < 0.001) among 4457 proteins quantified in microsomal fractions during MNBO formation. Recombinant HSD17B12 expressed in HEK293T cells exhibited prominent nabumetone reductase activity, and the contribution of HSD17B12 to the activity in the HLM was calculated as almost 100%. MNBO formation in HepG2 and Huh7 cells was significantly decreased by the knockdown of HSD17B12. We also examined the role of HSD17B12 in drug metabolism and found that recombinant HSD17B12 catalyzed the reduction reactions of pentoxifylline and S-warfarin, suggesting that HSD17B12 prefers compounds containing a methyl ketone group on the alkyl chain. In conclusion, our study demonstrated that HSD17B12 is responsible for the formation of MNBO from nabumetone. Together with the evidence for pentoxifylline and S-warfarin reduction, this is the first study to report that HSD17B12, which is known to metabolize endogenous compounds, such as estrone and 3-ketoacyl-CoA, plays a role as a drug-metabolizing enzyme. Show less

Although relatively uncommon, pathologists may encounter minimal inflammatory foci in the absence of typical structural heart disease; however, the clinicopathological significance of minimal inflammatory foci, including correlation with sudden unexpected death, is unexplored. From 1072 serial autopsy subjects, cases with unexplained minimal inflammatory foci, the extent of which was under 1% of the whole examined ventricle, were extracted to exclude cases with borderline/focal myocarditis resulting from local, systemic infection, or autoimmune mechanisms. Immunohistochemistry and genetic analysis targeting viral genomes and heart disease-related genes using next generation sequencing were performed. We detected 10 cases with unexplained minimal inflammatory foci (five males, five females, aged 15-68 years). The cause and/or manner of death were sudden unexpected death (6 cases, 60%), sudden unexpected death with epilepsy (1 case, 10%), drowning in a hot bath (1 case, 10%), and suicide (2 cases, 20%). In none of these cases was pathogen-derived DNA or RNA detected. In 8 of the 10 cases (80%), 17 possible pathogenic genetic variants causative for arrhythmogenic right ventricular cardiomyopathy or dilated cardiomyopathy; DSP was the most frequently involved gene (three cases with two different variants), followed by LAMA4 and MYBPC3 (two cases, two variants for each gene), LDB3 (two cases, one variant), and the remaining 10 variants occurred in seven cases (DSC2, RYR2, SOS1, SCN5A, SGCD, LPL, PKP2, MYH11, GATA6, and DSG2). All mutations were missense mutations. DSP_Lys1581Glu and DSC2_p.Thr275Met were classified according to American College of Medical Genetics and Genomics consensus statement guidelines as pathogenic or likely pathogenic for arrhythmogenic cardiomyopathy in three patients (30%). The remaining 15 variants were classified as potentially pathogenic variants. Unexplained minimal inflammatory foci may be an early sign of inherited cardiomyopathy, and such cases might already have arrhythmogenic potential that can lead to sudden unexpected death. Detection of minimal inflammatory foci by careful pathological examination may indicate the value of conducting comprehensive genetic analysis, even if significant structural abnormalities are not evident. Show less

Myocyte disarray of >10% in the heart is broadly accepted as a diagnostic pitfall for hypertrophic cardiomyopathy (HCM) at postmortem. The present study aims to propose an additional diagnostic criterion of HCM. Heart specimens from 1387 serial forensic autopsy cases were examined. Cases with myocyte disarray were extracted and applied to morphometric analysis to determine the amount of myocyte disarray. Comprehensive genetic analysis by using next-generation sequencing was subsequently applied for cases with myocyte disarray. Fifteen cases with myocyte disarray were extracted as candidate cases (1.1%, 11 men and 4 women, aged 48⁻94 years). In terms of the cause of death, only 2 cases were cardiac or possible cardiac death, and the other was non-cardiac death. Six cases showed myocyte disarray of >10% and 3 cases showed myocyte disarray of 5% to 10%. The other 6 cases showed myocyte disarray of <5%. Nine rare variants in 5 HCM-related genes (MYBPC3, MYH7, MYH6, PRKAG2, and CAV3) were found in 8 of 9 cases with myocyte disarray of >5%. The remaining 1 and 6 cases with myocyte disarray of <5% did not have any such variant. Myocyte disarray of >5% with rare variants in related genes might be an appropriate postmortem diagnostic criterion for HCM, in addition to myocyte disarray of 10%. Show less

Although some studies have attempted to find useful prognostic factors in hypertrophic cardiomyopathy (HCM), those results are not fully helpful for use in actual clinical practice. Furthermore, several genetic abnormalities associated with HCM have been identified. However, the genotype-phenotype correlation in HCM remains to be elucidated. Here, we attempted to assess patients with different types of gene mutations causing HCM and investigate the prognosis. A total of 140 patients with HCM underwent a screening test for myofilament gene mutations by direct sequencing of eight sarcomeric genes. Patients with a single mutation in cardiac troponin T, cardiac troponin I, α-tropomyosin, and regulatory and essential light chains were excluded from the study because the number of cases was too small. The clinical presentations and outcomes of the remaining 127 patients with HCM, 31 β-myosin heavy chain (MYH7) mutation carriers, 19 cardiac myosin-binding protein C (MYBPC3) mutation carriers, and 77 mutation non-carriers were analyzed retrospectively. MYBPC3 mutation carriers had a high frequency of ventricular arrhythmia and syncope. Kaplan-Meier curves revealed no significant difference in prognosis among the three groups, but a lack of family history of sudden death (SD) and a past history of syncope were significantly related to poor prognosis. An absence of family history of SD and past history of syncope are useful prognostic factors in patients with HCM. MYH7 and MYBPC3 mutations did not significantly influence prognosis compared to non-carriers. However, patients with the MYBPC3 mutation should be closely followed for the possibility of SD. Show less

Hypertrophic cardiomyopathy (HCM), which is inherited as an autosomal dominant trait, is the most prevalent hereditary cardiac disease. Although there are several reports on the systematic screening of mutations in the disease-causing genes in European and American populations, only limited information is available for Asian populations, including Japanese. Genetic screening of disease-associated mutations in 8 genes for sarcomeric proteins, MYH7, MYBPC3, MYL2, MYL3, TNNT2, TNNI3, TPM1, and ACTC, was performed by direct sequencing in 112 unrelated Japanese proband patients with familial HCM; 37 different mutations, including 13 novel ones in 5 genes, MYH7, MYBPC3, TNNT2, TNNI3, and TPM1, were identified in 49 (43.8%) patients. Among them, 3 carried compound heterozygous mutations in MYBPC3 or TNNT2. The frequency of patients carrying the MYBPC3, MYH7, and TNNT2 mutations were 19.6%, 10.7%, and 8.9%, respectively, and the most frequently affected genes in the northeastern and southwestern parts of Japan were MYBPC3 and MYH7, respectively. Several mutations were found in multiple unrelated proband patients, for which the geographic distribution suggested founder effects of the mutations. This study demonstrated the frequency and distribution of mutations in a large cohort of familial HCM in Japan. Show less

Left ventricular noncompaction (LVNC) is a cardiomyopathy morphologically characterized by 2-layered myocardium, numerous prominent trabeculations, and deep intertrabecular recesses communicating with the left ventricular cavity. The purpose of this study was to investigate patients with LVNC for possible disease causing mutations. We screened 4 genes (TAZ, LDB3, DTNA and TPM1) in 51 patients with LVNC for mutations by polymerase chain reaction and direct DNA sequencing. A novel missense substitution in exon 1 of TPM1 (c.109A>G: p.Lys37Glu) was identified in three affected members of a family with isolated LVNC. The substitution brings about a change in amino acid charge at a highly conserved residue and could result in aberrant mRNA splicing. This variant was not identified in 200 normal control samples. Pathologic analysis of a right ventricular myocardial specimen from the proband's maternal aunt revealed endocardial and subendocardial fibrosis with prominent elastin deposition, as well as the presence of adipose tissue between muscle layers, pathologic changes that are distinct from those seen in patients with HCM or DCM. Screening of the proband and her mother for variants in other sarcomeric protein-encoding candidate genes, MYH7, MYBPC3, TNNT2, TNNI3, ACTC, MYL2, and MYL3, did not identify any other non-synonymous variants or variants in splice donor-acceptor sequences that were potentially disease causing. We conclude TPM1 is a potential candidate disease-causing gene for isolated LVNC, especially in patients experiencing sudden death. Show less