Deficiency of 3-hydroxybutyrate dehydrogenase (BDH1) in mice causes low ketone body levels and fatty liver during fasting.

d-3-Hydroxy-n-butyrate dehydrogenase (BDH1; EC 1.1.1.30), encoded by BDH1, catalyzes the reversible reduction of acetoacetate (AcAc) to 3-hydroxybutyrate (3HB). BDH1 is the last enzyme of hepatic ketogenesis and the first enzyme of ketolysis. The hereditary deficiency of BDH1 has not yet been described in humans. To define the features of BDH1 deficiency in a mammalian model, we generated Bdh1-deficient mice (Bdh1 KO mice). Under normal housing conditions, with unrestricted access to food, Bdh1 KO mice showed normal growth, appearance, behavior, and fertility. In contrast, fasting produced marked differences from controls. Although Bdh1 KO mice survive fasting for at least 48 hours, blood 3HB levels remained very low in Bdh1 KO mice, and despite AcAc levels moderately higher than in controls, total ketone body levels in Bdh1 KO mice were significantly lower than in wild-type (WT) mice after 16, 24, and 48 hours fasting. Hepatic fat content at 24 hours of fasting was greater in Bdh1 KO than in WT mice. Systemic BDH1 deficiency was well tolerated under normal fed conditions but manifested during fasting with a marked increase in AcAc/3HB ratio and hepatic steatosis, indicating the importance of ketogenesis for lipid energy balance in the liver.

Development of Specific Fluorogenic Substrates for Human ß-N-Acetyl-D-hexosaminidase A for Cell-Based Assays.

Inhibitors of human ß-N-acetyl-D-hexosaminidase (hHEX) A and human O-GlcNAcase (hOGA) reportedly play roles in multiple diseases, suggesting their potential for pharmacological chaperone (PC) therapy of Sandhoff disease (SD) and Tay-Sachs disease (TSD), as lysosomal storage diseases, and Alzheimer's disease and progressive supranuclear palsy, respectively. In particular, hHEXA inhibitors as PCs have been shown to successfully enhance hHEXA levels, leading to the chronic form of SD and TSD. In the diagnosis of enzyme deficiencies in SD and TSD, artificial hHEXA substrates based on 4-methylumbelliferone as a fluorophore are available and generally used; however, they do not have sufficient performance to screen for potential inhibitors for a PC therapy from compound libraries. Further, there are currently few fluorogenic substrates for hHEXA suitable for such requirements and there are no substrates ideal for cell-based inhibitor screening. Here, we clarified the difference in enzyme active site structure between hHEXA and hOGA from their tertiary structures. To develop lysosome-localized hHEXA-specific fluorogenic substrates based on the difference in their active site structures, our developed quinone methide cleavage substrate design platform was applied for the molecular design of substrates. Thereafter, we synthesized via the shortest route and evaluated novel three-color fluorogenic substrates for hHEXA that exhibited excellent specificity and sensitivity in three human cell lines. The designed substrates represent the first-in-a class of new substrates that can be utilized to screen hHEXA inhibitors in adherent human cultured cells.

Recent advances in understanding beta-ketothiolase (mitochondrial acetoacetyl-CoA thiolase, T2) deficiency.

Beta-ketothiolase (mitochondrial acetoacetyl-CoA thiolase, T2) deficiency (OMIM #203750, *607809) is an inborn error of metabolism that affects isoleucine catabolism and ketone body metabolism. This disorder is clinically characterized by intermittent ketoacidotic crises under ketogenic stresses. In addition to a previous 26-case series, four series of T2-deficient patients were recently reported from different regions. In these series, most T2-deficient patients developed their first ketoacidotic crises between the ages of 6 months and 3 years. Most patients experienced less than three metabolic crises. Newborn screening (NBS) for T2 deficiency is performed in some countries but some T2-deficient patients have been missed by NBS. Therefore, T2 deficiency should be considered in patients with severe metabolic acidosis, even in regions where NBS for T2 deficiency is performed. Neurological manifestations, especially extrapyramidal manifestations, can occur as sequelae to severe metabolic acidosis; however, this can also occur in patients without any apparent metabolic crisis or before the onset of metabolic crisis.

Modulation of the sphingolipid rheostat is involved in paclitaxel resistance of the human prostate cancer cell line PC3-PR.

Taxoids are anti-cancer drugs frequently used to treat solid tumors, but they are sometimes ineffective and tumors may become resistant to their action. Here, we examined the involvement of sphingolipid metabolic enzymes in paclitaxel (PTX) resistance using a human prostate cancer cell line, PC3, and its PTX-resistant subline, PC3-PR. PTX (20 nM) suppressed cell proliferation and increased various ceramide species in PC3, but not PC3-PR, cells. PC3-PR contained higher S1P levels than did PC3, regardless of PTX treatment. Western blotting revealed that PC3-PR cells expressed higher levels of sphingosine kinase 1 (SPHK1) and glucosylceramide synthase (GCS) but lower levels of acid sphingomyelinase (ASMase) and neutral sphingomyelinase 2 than did PC3 cells. Inhibition of SPHK1 using siRNA or a pharmacological inhibitor decreased S1P levels in PC3-PR cells and inhibited proliferation in the presence or absence of PTX, suggesting that SPHK1 is at least partially responsible for PTX resistance. Similarly, GCS inhibitors (PDMP and PPMP) increased cellular ceramides and suppressed the proliferation of PC3-PR. However, inhibition of proteasome function or histone deacetylase activity increased SMase and ceramide levels and suppressed PC3-PR proliferation. These results suggest that modulation of metabolic enzyme expression and alteration of the sphingolipid rheostat protects cancer cells against PTX.

Heterozygous carriers of succinyl-CoA:3-oxoacid CoA transferase deficiency can develop severe ketoacidosis.

Succinyl-CoA:3-oxoacid CoA transferase (SCOT, gene symbol OXCT1) deficiency is an autosomal recessive disorder in ketone body utilization that results in severe recurrent ketoacidotic episodes in infancy, including neonatal periods. More than 30 patients with this disorder have been reported and to our knowledge, their heterozygous parents and siblings have had no apparent ketoacidotic episodes. Over 5 years (2008-2012), we investigated several patients that presented with severe ketoacidosis and identified a heterozygous OXCT1 mutation in four of these cases (Case1 p.R281C, Case2 p.T435N, Case3 p.W213*, Case4 c.493delG). To confirm their heterozygous state, we performed a multiplex ligation-dependent probe amplification analysis on the OXCT1 gene which excluded the presence of large deletions or insertions in another allele. A sequencing analysis of subcloned full-length SCOT cDNA showed that wild-type cDNA clones were present at reasonable rates to mutant cDNA clones. Over the following 2 years (2013-2014), we analyzed OXCT1 mutations in six more patients presenting with severe ketoacidosis (blood pH ≦7.25 and total ketone body ≧10 mmol/L) with non-specific urinary organic acid profiles. Of these, a heterozygous OXCT1 mutation was found in two cases (Case5 p.G391D, Case6 p.R281C). Moreover, transient expression analysis revealed R281C and T435N mutants to be temperature-sensitive. This characteristic may be important because most patients developed ketoacidosis during infections. Our data indicate that heterozygous carriers of OXCT1 mutations can develop severe ketoacidotic episodes in conjunction with ketogenic stresses.

Characterization and outcome of 41 patients with beta-ketothiolase deficiency: 10 years' experience of a medical center in northern Vietnam.

Beta-ketothiolase (T2) deficiency is an inherited disease of isoleucine and ketone body metabolism caused by mutations in the ACAT1 gene. Between 2005 and 2016, a total of 41 patients with T2 deficiency were identified at a medical center in northern Vietnam, with an estimated incidence of one in 190,000 newborns. Most patients manifested ketoacidotic episodes of varying severity between 6 and 18 months of age. Remarkably, 28% of patients showed high blood glucose levels (up to 23.3 mmol/L). Ketoacidotic episodes recurred in 43% of patients. The age of onset, frequency of episodes, and identified genotype did not affect patient outcomes that were generally favorable, with the exception of seven cases (five died and two had neurological sequelae). Custom-tailored acute and follow-up management was critical for a positive clinical outcome. Two null mutations, c.622C>T (p.Arg208*) and c.1006-1G>C (p.Val336fs), accounted for 66% and 19% of all identified ACAT1 mutant alleles, respectively. Most patients showed characteristic biochemical abnormalities. A newborn screening program could be expected to have a high yield in Vietnam. Investigation findings of haplotypes linked to the most common ACAT1 mutation (c.622C>T) are consistent with an ancient common founder of mutation-bearing chromosomes belonging to the Kinh ethnic population. The direct management and long-term follow-up of a large number of T2-deficient patients enabled us to study the natural history of this rare disease.

Mechanism of paclitaxel resistance in a human prostate cancer cell line, PC3-PR, and its sensitization by cabazitaxel.

Paclitaxel (PTX) is a microtubule-targeting drug widely used for the treatment of a variety of cancers. However, drug resistance can emerge after a series of treatments, and this can seriously affect the patient's prognosis. Here, we analyzed the mechanism of PTX resistance using a human prostate cancer cell line, PC3, and its PTX-resistant subline, PC3-PR. Compared with PC3, PC3-PR exhibited some unique phenotypes that might be associated with PTX resistance, including decreased expression of acetylated α-tubulin and the cell cycle regulator p21, and increased expression of ßIII tubulin, histone deacetylase 6 (HDAC6), and the anti-apoptotic protein Bcl2. The drug exporters MDR1 and MRP1 were not involved in PTX resistance. Although cabazitaxel (CTX), a novel taxoid, has been reported to overcome PTX resistance, its mechanism of action is unknown. We found that treatment of PC3-PR cells with CTX induced expression of acetylated α-tubulin and p21, but not the related regulators p27, p15, and p16 or the Bcl2 family proteins. The pan-HDAC inhibitors trichostatin A and suberanilohydroxamic acid and the HDAC6-specific inhibitor tubacin inhibited PC3-PR proliferation and increased expression of p21 and acetylated α-tubulin in a manner similar to CTX. Our data shed light on the cellular response to PTX and CTX.

Chain Walking as a Strategy for Carbon-Carbon Bond Formation at Unreactive Sites in Organic Synthesis: Catalytic Cycloisomerization of Various 1,n-Dienes.

Carbon-carbon bond formation at unreactive sp(3)-carbons in small organic molecules via chain walking was achieved for the palladium-catalyzed cycloisomerization of 1,n-dienes. Various 1,n-dienes (n = 7-14) such as those containing cyclic alkenes, acyclic internal alkenes, and a trisubstituted alkene can be used for the chain-walking cycloisomerization/hydrogenation process, and five-membered ring compounds including simple cyclopentane and pyrrolidine derivatives can easily be prepared. Chain walking over a tertiary carbon was also found to be possible in the cycloisomerization. It is not necessary for the linker portion of the diene to contain a quaternary center, and diene substrates with two alkene moieties linked by a tertiary carbon or a nitrogen atom can also be used as substrates. Column chromatography using silica gel containing silver nitrate was found to be effective for isolating some of the cycloisomerization products without hydrogenation. Deuterium-labeling experiments provided direct evidence to show that the reaction proceeds via a chain-walking mechanism.

The first case in Asia of 2-methyl-3-hydroxybutyryl-CoA dehydrogenase deficiency (HSD10 disease) with atypical presentation.

2-Methyl-3-hydroxybutyryl-CoA dehydrogenase (2M3HBD) deficiency (HSD10 disease) is a rare inborn error of metabolism, and <30 cases have been reported worldwide. This disorder is typically characterized by progressive neurodegenerative disease from 6 to 18 months of age. Here, we report the first patient with this disorder in Asia, with atypical clinical presentation. A 6-year-old boy, who had been well, presented with severe ketoacidosis following a 5-day history of gastroenteritis. Urinary organic acid analysis showed elevated excretion of 2-methyl-3-hydroxybutyrate and tiglylglycine. He was tentatively diagnosed with ß-ketothiolase (T2) deficiency. However, repeated enzyme assays using lymphocytes showed normal T2 activity and no T2 mutation was found. Instead, a hemizygous c.460G>A (p.A154T) mutation was identified in the HSD17B10 gene. This mutation was not found in 258 alleles from Japanese subjects (controls). A normal level of the HSD17B10 protein was found by immunoblot analysis but no 2M3HBD enzyme activity was detected in enzyme assays using the patient's fibroblasts. These data confirmed that this patient was affected with HSD10 disease. He has had no neurological regression until now. His fibroblasts showed punctate and fragmented mitochondrial organization by MitoTracker staining and had relatively low respiratory chain complex IV activity to those of other complexes.

Ketone body metabolism and its defects.

Acetoacetate (AcAc) and 3-hydroxybutyrate (3HB), the two main ketone bodies of humans, are important vectors of energy transport from the liver to extrahepatic tissues, especially during fasting, when glucose supply is low. Blood total ketone body (TKB) levels should be evaluated in the context of clinical history, such as fasting time and ketogenic stresses. Blood TKB should also be evaluated in parallel with blood glucose and free fatty acids (FFA). The FFA/TKB ratio is especially useful for evaluation of ketone body metabolism. Defects in ketogenesis include mitochondrial HMG-CoA synthase (mHS) deficiency and HMG-CoA lyase (HL) deficiency. mHS deficiency should be considered in non-ketotic hypoglycemia if a fatty acid beta-oxidation defect is suspected, but cannot be confirmed. Patients with HL deficiency can develop hypoglycemic crises and neurological symptoms even in adolescents and adults. Succinyl-CoA-3-oxoacid CoA transferase (SCOT) deficiency and beta-ketothiolase (T2) deficiency are two defects in ketolysis. Permanent ketosis is pathognomonic for SCOT deficiency. However, patients with "mild" SCOT mutations may have nonketotic periods. T2-deficient patients with "mild" mutations may have normal blood acylcarnitine profiles even in ketoacidotic crises. T2 deficient patients cannot be detected in a reliable manner by newborn screening using acylcarnitines. We review recent data on clinical presentation, metabolite profiles and the course of these diseases in adults, including in pregnancy.

Recurrent Optic Neuropathy Caused by a Mucocele of the Anterior Clinoid Process after a 5-Year Remission: A Case Report and Literature Review.

A 32-year-old male presented with acute left vision loss during a second recurrence of optic neuropathy. Steroid pulse therapy had been effective in both the first episode 9 years previously and the first recurrence 5 years previously. Magnetic resonance imaging demonstrated an anterior clinoid process mucocele compressing the optic nerve. Although surgical treatment was performed, improvement was limited. This report indicates that steroid pulse therapy could be an alternative treatment to obtain temporary remission, but surgical treatment should be considered to prevent irreversible neurological deficits. This paper also presents a review of the literature on anterior clinoid process mucoceles.

Factors Influencing Willingness to Undergo Lung Cancer Screening in the Future: A Cross-Sectional Study of Japanese University Students.

Lung cancer (LC) is currently the leading cause of cancer deaths in Japan. Early detection through lung cancer screening (LCS) is important for reducing mortality. Therefore, exploring the factors affecting willingness to undergo LCS, particularly among young people, is important. This study aimed to elucidate the inclination toward LCS and its determining factors among Japanese university students. This cross-sectional study, involving 10,969 Japanese university students, was conducted in April 2023. A Pearson's chi-square test and a binomial logistic regression analysis were used to analyze factors related to the dependent variable, willingness to undergo LCS in the future. Out of the 6779 participants (61.8%) involved in this study, 6504 (95.9%) provided valid responses, and 4609 (70.9%) expressed a willingness to undergo LCS in the future. Analysis revealed current smoking as a barrier to future willingness to undergo LCS. Other barriers included postponing the age of screening, anxiety about the screening content, and concerns about the possibility of having cancer after screening. Addressing barriers, such as current smoking and anxiety about screening, that prevent young people from undergoing LCS in the future is crucial. Therefore, universities should provide opportunities to educate students about LCS and explore various educational methods.

Association between the experience of exertional heat illness (EHI) and living conditions of collegiate student athletes.

Exertional heatstroke (EHS), a severe form of exertional heat illness (EHI), is the third leading cause of death in athletes; thus, early detection and prevention of EHI can help prevent EHS, which is a life-threatening condition. This study aimed to clarify the association between the cognizance of experiencing EHI and living conditions and specific EHI symptoms among collegiate athletes. This study was conducted in October 2022 by administering a questionnaire to 237 male collegiate athletes. Of the 215 (90.7%) respondents, 197 (91.6%) provided valid responses; among them, 88 (44.7%) responded they had experienced EHI, while 109 (55.3%) had not. A history of medical examinations due to EHI, having experienced headaches during summer activities, and having read the EHI manual were factors indicating cognizance of EHI. The number of times meals containing a staple food, main dish, and side dish were eaten in a day was a factor in preventing EHI. Early detection of EHI is important for its prevention, and it is important that athletes themselves have knowledge of symptoms and can correctly self-diagnose EHI. Emphasizing the potential of a well-balanced dietary intake has the potential to prevent EHI is crucial.

Development of MLPA for human ACAT1 gene and identification of a heterozygous Alu-mediated deletion of exons 3 and 4 in a patient with mitochondrial acetoacetyl-CoA thiolase (T2) deficiency.

Mitochondrial acetoacetyl-CoA thiolase deficiency is an autosomal recessive disorder, characterized by intermittent ketoacidosis. We developed a multiplex ligation-dependent probe amplification method for mutation detection in the ACAT1 gene, which encodes this enzyme, and validated it using DNAs from two previously reported patients having partial deletion and duplication in this gene. Using this method, we identified a heterozygous deletion including exons 3-4 in a third patient, likely due to Alu-mediated non-equal homologous recombination between Alu sequences.

Factors affecting health-related quality of life among firefighters during the COVID-19 pandemic: A single-center study.

During the coronavirus disease 2019 (COVID-19) outbreak, firefighters have been working in an environment that is both physically and mentally taxing. This study aimed to investigate factors affecting health-related quality of life (HRQOL) among firefighters in Japan during the COVID-19 pandemic. A total of 227 firefighters from a single firefighting organization were surveyed in June 2021, during the fourth infection spread period of COVID-19 in Japan. Regression analysis was performed to examine factors affecting HRQOL of firefighters measured with the SF-8. In the present study, factors affecting HRQOL among firefighters during the COVID-19 pandemic were lack of sleep, physical abnormalities due to infection control measures, exercise habits, living with family members, and history of suspected COVID-19 infection. The present findings may help develop support services for first responders, including firefighters during the COVID-19 pandemic.

Nicotine Dependence among College Students Uninterested in Smoking Cessation during the COVID-19 Pandemic: A Cross-Sectional Survey.

This study investigated nicotine dependence among Japanese university students who had reached the smoking age (20 years or older) by the time of the coronavirus disease 2019 (COVID-19) pandemic and examined factors that encourage early smoking cessation. Social dependence on nicotine was evaluated using the Kano Total Social Nicotine Dependence Level (KTSND), and physiological dependence was evaluated using the Fagerström Nicotine Dependence Index (FTND). Of the 356 college students who smoked (4.4% of the total), 182 (51.1%) stated that they were not interested in quitting. Furthermore, 124 (68.1%) of those with no interest in quitting smoking were aware that smoking is a high-risk factor for COVID-19, and 58 (31.9%) were unaware. The group not aware of this risk had significantly higher KTSND scores than the group aware of it. The examination of cigarette type that indicated the users of non-conventional cigarette products and dual-user groups scored significantly higher than the cigarette group on FTND items. Overall, the smokers scored above the normal range for social nicotine dependence, suggesting the need to reduce nicotine dependence to encourage college students who continue to smoke to quit smoking.

University students' living conditions during the COVID-19 pandemic and predictors of their subjective health views: A cross-sectional survey.

This study aimed to explore the factors influencing subjective health views based on the living conditions and concerns of university students during the coronavirus infection 2019 (COVID-19) pandemic. From March to April 2021, a questionnaire survey was administered to 8,547 Japanese university students, and logistic regression analysis was used to explore factors related to subjective health views. The results showed that satisfaction with quality of sleep (OR = 2.651, 95% Cl 2.370-2.966,p < 0.001), satisfaction with university life (OR = 2.486, 95%Cl 2.215-2.789, p < 0.001), satisfaction with diet (OR = 1.849, 95% CI: 1.496-2.285, p < 0.001), regular exercise (OR = 1.759, 95% CI: 1.594-1.941, p < 0.001), consciousness of nutritional balance (OR = 1.276, 95% CI: 1.147-1.420,p < 0.001), eating breakfast every day (OR = 1.247, 95% CI: 1.121-1.387, p < 0.001), and consuming soft drinks at least once a week (OR = 0.865, 95% CI: 0.755-0.966, p = 0.010) were positive factors for subjective views of health. On the other hand, anxiety about whether the necessary credits can be obtained (OR = 0.885, 95% CI: 0.799-0.980, p = 0.019), infection from minimal outings (OR = 0.881, 95% CI: 0.794-0.976, p = 0.016) building and maintaining friendships on campus (OR = 0.867, 95% CI: 0.767-0.980, p = 0.023), and being able to continue working (OR = 0.713, 95% CI: 0.640-0.795, p < 0.001) were identified as negative factors. To ensure a healthy university life during the COVID-19 pandemic or future pandemic, supports tailored to students' living conditions and measures to address their anxieties are required.

Chain-walking strategy for organic synthesis: catalytic cycloisomerization of 1,n-dienes.

The catalytic construction of carbon-carbon bonds in small organic molecules via chain walking is described. Catalytic cycloisomerization of 1,n-dienes via chain walking was achieved using a palladium-1,10-phenanthroline catalyst to form five-membered-ring products. By means of a cycloisomerization/hydrogenation protocol, 1,7- to 1,14-dienes were selectively converted to bicyclo[4.3.0]nonane derivatives. The use of chain walking provides a new method in organic synthesis to functionalize unreactive carbon-hydrogen bonds by letting the catalyst look for preferable bond-forming sites by moving around on the substrate.

Evaluation of Japanese university students' perception of smoking, interest in quitting, and smoking behavior: An examination and public health challenges during the COVID-19 pandemic.

This study examined college students' perceptions of the association between smoking and novel coronavirus disease 2019 (COVID-19), changes in smoking behavior, and interest in quitting categorized by smoking device, to identify public health challenges. A questionnaire survey was conducted among 8,547 students in a Japanese university in March and April 2021. In response to "Awareness of the increased risk of COVID-19 infection due to smoking and the tendency to develop severe disease", current smokers (70.2%) were more aware of the risk than non-smokers (49.8%) (p < 0.001), with no significant difference according to smoking device (p = 0.213). "Interest in quitting smoking" (p = 0.323), and "Changes in smoking behavior during the COVID-19 pandemic" (p = 0.146) did not differ by smoking device. However, approximately 50% of the respondents answered that they were not interested in quitting smoking, while two-thirds reported that the number of cigarettes they smoked did not change during the pandemic. During the COVID-19 pandemic, college students were found to be less interested in quitting and not likely to change their smoking behavior, despite the knowledge of the increased risk of COVID-19 transmission and severity of disease from smoking, regardless of smoking device.

Antibody response of smokers to the COVID-19 vaccination: Evaluation based on cigarette dependence.

Smokers may have lower antibody titers after vaccination with a coronavirus disease 2019 (COVID-19) mRNA vaccine. However, to the best of our knowledge, no study has evaluated antibody titers after COVID-19 vaccination based on the level of smokers' cigarette dependence. In this study, we measured the level of serum anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) spike protein receptor-binding domain (S-RBD) immunoglobulin-G (IgG) by enzyme linked immunosorbent assay of 55 actively smoking Japanese social workers (firefighters, paramedics, and rescue workers) who had received two doses of the BNT162b2 vaccine. Further, we assessed their cigarette dependence using the Fagerstrom Test for Nicotine Dependence (FTND), measured their serum cotinine levels, and tested for their correlation with anti-RBD IgG levels. Serum anti-SARS-CoV-2 S-RBD protein IgG levels after BNT162b2 vaccination showed a significant negative correlation with FTND (ρ = -0.426, p = 0.001). In addition, serum cotinine level showed a significant positive correlation with FTND (ρ = 0.470, p = 0.000). However, no significant negative correlation was noted between serum cotinine and serum anti-SARS-CoV-2 S-RBD protein IgG levels (ρ = -0.156, p = 0.256). Our results suggest that smokers with strong cigarette dependence have inadequate anti-SARS-CoV-2 S-RBD protein IgG levels after COVID-19 mRNA vaccination.