Mitochondrial haplotype mutation alleviates respiratory defect of MELAS by restoring taurine modification in tRNA with 3243A > G mutation.

The 3243A > G in mtDNA is a representative mutation in mitochondrial diseases. Mitochondrial protein synthesis is impaired due to decoding disorder caused by severe reduction of 5-taurinomethyluridine (τm5U) modification of the mutant mt-tRNALeu(UUR) bearing 3243A > G mutation. The 3243A > G heteroplasmy in peripheral blood reportedly decreases exponentially with age. Here, we found three cases with mild respiratory symptoms despite bearing high rate of 3243A > G mutation (>90%) in blood mtDNA. These patients had the 3290T > C haplotypic mutation in addition to 3243A > G pathogenic mutation in mt-tRNALeu(UUR) gene. We generated cybrid cells of these cases to examine the effects of the 3290T > C mutation on mitochondrial function and found that 3290T > C mutation improved mitochondrial translation, formation of respiratory chain complex, and oxygen consumption rate of pathogenic cells associated with 3243A > G mutation. We measured τm5U frequency of mt-tRNALeu(UUR) with 3243A > G mutation in the cybrids by a primer extension method assisted with chemical derivatization of τm5U, showing that hypomodification of τm5U was significantly restored by the 3290T > C haplotypic mutation. We concluded that the 3290T > C is a haplotypic mutation that suppresses respiratory deficiency of mitochondrial disease by restoring hypomodified τm5U in mt-tRNALeu(UUR) with 3243A > G mutation, implying a potential therapeutic measure for mitochondrial disease associated with pathogenic mutations in mt-tRNAs.

Expression analysis of gibberellin-regulated protein in peach by reverse transcription-quantitative PCR.

Gibberellin-regulated protein (GRP) is a fruit severe allergen. The amounts of GRP expression normalized against actin in peach were determined by reverse transcription-quantitative PCR (RT-qPCR). The results were consistent with those determined by enzyme-linked immunosorbent assay (ELISA). The GRP expression was more evident in flesh than peel and increased rapidly in the maturing period. This approach is applicable to estimate the amount of GRP in other plants.

Genomic characteristics revealed by targeted exon sequencing of testicular germ cell tumors in Japanese men.

OBJECTIVE: To investigate the somatic mutation profiles of testicular germ cell tumors in Japanese men. METHODS: We analyzed the somatic missense mutation profile of testicular germ cell tumors among 21 Japanese men with seminoma (n = 14), pure embryonic carcinoma (n = 3) and mixed testicular germ cell tumor (n = 4) by targeted next-generation sequencing of 409 cancer-related genes covering 1.23 Mb of the genome. RESULTS: We identified a total of 22 missense mutations in 21 primary testicular germ cell tumor samples (0.89 mutations/Mb), of which seven mutations were confirmed to be absent from the germline. KIT:p.Asn822Tyr, KIT:p.Leu576Pro, PIK3CA:p.Glu542Lys and FBXW7:p.Arg505His were statistically and functionally potential. A total of 18 missense mutations were previously unknown in testicular germ cell tumors. PDGFRA amplification from one patient with seminoma was detected. KIT, BCR,PIK3CG, PIK3CA and PDGFRA mutations involved in aberrant signaling of the KIT-PI3K-AKT pathway was detected in 27.3% of detected mutations. CONCLUSIONS: The present investigation identified a low mutation rate in testicular germ cell tumors among Asian patients, 18 novel mutations and PDGFRA amplification. Limitations of the present study are the small sample and missing normal DNA for some testicular germ cell tumors.

Adverse effect of switching only once low-carbohydrate diet to high-carbohydrate diet on postprandial glucose concentration in healthy women.

BACKGROUND AND OBJECTIVES: Our aim was to evaluate the acute effect of switching low-carbohydrate diet (LCD) to high-carbohydrate diet (HCD) on glycemic parameters in healthy women. METHODS AND STUDY DESIGN: Twen-ty-two women (age 21.7±4.0 years; HbA1c 5.3±0.3 %, mean±SD) wore flash glucose monitoring system and consumed test meals for 3 days from Day 4 to 6. Participants consumed identical HCD meals except LCD dinner on Day 5. The energy ratio of carbohydrate, fat, and protein were 64%, 21%, and 15% for HCD and 47%, 35%, and 18% for Day 5 with LCD dinner (19%, 59%, and 22%). RESULTS: The incremental glucose peak (IGP, both p<0.001) and incremental area under the curve for glucose (IAUC, both p<0.001) 3h of LCD dinner were all sig-nificantly lower than those of HCD dinner on Day 4 and 6. However, after consuming LCD dinner on Day 5, IGP breakfast (2.33±0.15 vs 1.71±0.15 mmo/L, p<0.01), IGP lunch (3.31±0.25 vs 2.54±0.18 mol/L, p<0.01), IAUC 3h of breakfast (210±18 vs 136±14 mmol/L×min, p<0.001), mean blood glucose (5.72±0.11 vs 5.40±0.11 mmol/L, p<0.01), and standard deviation (1.11±0.06 vs 0.88±0.04 mmol/L, p<0.01) on Day 6 were all signifi-cantly higher than those of corresponding meals before LCD dinner on Day 4, in spite of consuming all identical HCD meals. The glycemic parameters returned to the levels before consuming LCD on Day 7. CONCLUSIONS: Consuming LCD only once is enough to cause 24-h higher postprandial blood glucose concentration in subse-quent consumption of HCD in healthy women.

Comparison of electron microscopic findings and clinical presentation in three patients with mitochondrial cardiomyopathy caused by the mitochondrial DNA mutation m.3243A > G.

Mitochondrial cardiomyopathy can be described as a condition characterized by abnormal heart-muscle structure and/or function, secondary to mutations in nuclear or mitochondrial DNA. Its severity can range from subclinical to critical conditions. We presented three cases of mitochondrial cardiomyopathy with m.3243A > G mutation and compared the clinical manifestations with the histological findings for each of these cases. All cases showed cardiac hypertrophy, juvenile-onset diabetes mellitus, and hearing loss. Case 1 (43-year-old male) showed less cardiac involvement and shorter duration of mitochondrial disease-related symptoms than case 2 (67-year-old female) and case 3 (51-year-old male), who showed the most advanced cardiac condition and longest duration from the manifestation of heart failure. The histological findings revealed that cardiomyocytes from case 1 showed no hypertrophy and mitochondrial degeneration in electron microscopy. Alternatively, cases 2 and 3 showed hypertrophy in their cardiomyocytes, and mitochondrial degeneration (e.g. onion-like lesions, swollen cristae, and lamellar bodies) was most apparent in case 3. These results suggested that mitochondrial degeneration, as evaluated by electron microscopy, might be correlated with impaired heart function in patients with mitochondrial cardiomyopathy.

Late-night-dinner deteriorates postprandial glucose and insulin whereas consuming dinner dividedly ameliorates them in patients with type 2 diabetes: A randomized crossover clinical trial.

BACKGROUND AND OBJECTIVES: The aims of this study is to explore the acute effect of consuming dinner at different timing on postprandial glucose and hormone in patients with type 2 diabetes. METHODS AND STUDY DESIGN: Eight patients (age 70.8±1.9 years, HbA1c 7.6±0.6 %, BMI 23.3±3.2, mean±SD) were randomly assigned in this crossover study. Patients consumed the test meals of dinner at 18:00 on the first day, and dinner at 21:00 or divided dinner (vegetable and rice at 18:00 and vegetable and the main dish at 21:00) on the second or third day. Postprandial glucose, insulin, glucagon, free fatty acid (FFA), active glucagon-like peptide-1 (GLP-1), and active glucose- dependent insulinotropic polypeptide (GIP) concentration after dinner were evaluated. RESULTS: Both incremental area under the curve (IAUC) 2h for glucose and insulin were higher in dinner at 21:00 than those in dinner at 18:00 (IAUC glucose: 449±83 vs 216±43 mmol/L×min, p<0.01, IAUC insulin:772±104 vs 527±107 µU/mL×min, p<0.01, mean±SEM). However, in divided dinner both IAUC 4h for glucose and insulin tended to be lower than those of dinner at 21:00 (IAUC glucose: 269±76 mmol/L×min, p=0.070, IAUC insulin: 552±114 µU/mL×min, p=0.070). IAUC of active GLP-1 and active GIP demonstrated no difference among different dinner regimen. CONCLUSIONS: Consuming late-night-dinner (21:00) deteriorates postprandial glucose and insulin compared with those of early-evening-dinner (18:00) whereas consuming dinner dividedly ameliorates them.

An Embolic Stroke in a Patient With PROC p.Lys193del.

We report a 58-year-old woman who suddenly developed brain infarction with weakness of the left lower extremity and left perioral dysesthesia during postoperative tamoxifen therapy for breast cancer and prednisolone therapy for rheumatoid arthritis. Diffusion-weighted images detected multiple areas of hyperintensity in the posterior circulation system of the brain. Despite extensive examinations, we could not identify any embolic sources except hypoplasia of the right vertebral artery. We found decreased activity of protein C against its antigen level (activity: 59% versus antigen: 122%) with enhanced activity of coagulation factor VIII (178%) and von Willebrand factor (285%). DNA sequencing identified trinucleotide deletion of the PROC gene leading to 1 amino acid deletion at Lys-193 (p.Lys193del). We speculate that the PROC gene polymorphism may have participated in tamoxifen- and prednisolone- associated hypercoagulable state, leading to development of an embolic stroke in this patient.

Ischemic Stroke with Protein S Deficiency Treated by Apixaban.

A 57-year-old man with atherosclerosis obliterans was admitted with sudden-onset sensory aphasia and right hemiparesis. Brain MRI revealed acute cerebral infarctions in the left temporal lobe and magnetic resonance angiography showed occlusion of the posterior branch of the left middle cerebral artery. Transesophageal echocardiography and ultrasonography respectively confirmed a patent foramen ovale and deep vein thrombosis in the bilateral femoral veins. Blood findings showed low protein S antigen, low protein S activity, and a missense mutation of the PROS 1 gene. The administration of apixaban 10 mg BID prevented ischemic stroke recurrence and decreased the deep vein thrombosis. These outcomes indicated that apixaban may be alternative to warfarin for the secondary prevention of ischemic stroke in a patient with a protein S deficiency.

High genetic stability in MDCK-SIAT1 passaged human influenza viruses.

MDCK-induced amino acid (AA) mutation, such as D151G/N in the neuraminidase (NA) of influenza A/H3N2 viruses, is of concern. MDCK-SIAT1 cells, modified derivatives with an increased expression of α2,6-linked sialic acid receptors are increasingly used due to their superiority in a viral recovery. However, MDCK-SIAT1 induced AA mutations have not been fully examined. In this study, we compared NA and hemagglutinin (HA) genes of recent circulating influenza viruses isolated after an MDCK-SIAT1 passage with those directly obtained from the original samples. A total of 22 samples collected during the 2016-17 seasons included 9 A/H3N2, 5 H1N1pdm, and 8 B viruses. None of the deduced AA mutations in the NA or HA segments were detected after an MDCK-SIAT1 passage, except for one AA mutation in the NA of an influenza B virus sample. NA D151G/N changes were not seen in any of the MDCK-SIAT1 passaged A/H3N2 viruses, even in the small variants analysis conducted using deep sequencing. AA mutations induced by an MDCK-SIAT1 passage are currently rare, although careful observation is needed in the future.

Fluorescence tumor imaging by i.v. administered indocyanine green in a mouse model of colitis-associated colon cancer.

Fluorescence tumor imaging using exogenous fluorescent tumor-targeting agents has potential to improve early tumor detection. The fluorescent contrast agent indocyanine green (ICG) is used in medical diagnostics. The aim of the present study is to investigate the tumor imaging capability and the imaging mechanism of i.v. administered ICG in a mouse model of colitis-associated colon cancer. To do this, an azoxymethane/dextran sodium sulfate-induced colon cancer mouse model was used. Ex vivo imaging experiments were carried out 1 hour after i.v. injection of ICG. The ICG fluorescence was observed in the colon tumor tissues, with sufficient tumor to normal tissue ratio, correlating with tumor malignancy. In the tumor tissues, ICG fluorescence was localized in the vascular interstitial tissue. Immunofluorescence microscopy revealed that tumor cells formed tight junctions normally, suggesting an inability of tumor cellular uptake of ICG. In contrast, tumor tissues increased the CD31-immunoreactive endothelial cell area, and accumulated stromal cells immunoreactive for COX-2 and tumor cell population immunoreactive for inducible nitric oxide synthase. In vivo vascular permeability assay revealed that prostaglandin E2 promoted the endothelial cell permeability of ICG. In conclusion, our data indicated that fluorescence contrast-enhanced imaging following i.v. administered ICG can be applied to the detection of colon tumors in a mouse colitis-associated colon cancer model. The tumor tissue preference of ICG in the present model can be attributed to the enhanced vascular leakage of ICG involving inflammatory mediators, such as COX-2 and inducible nitric oxide synthase, in conjunction with increased tumor vascularity.

Hexaphenolic Rigid Cages Prepared by Self-Organization of C3 v Tridentates.

Coordination cages were composed by self-organization of rigid C3 v-symmetric heptaarene tridentates and Pd(II) precursors. The heptaarene framework involves one mesitylene, three phenol, and three pyridine moieties, which were connected by Suzuki coupling reactions. The treatment of the tridentates with Pd(dppp)(OTf)2 or Pd(en)(NO3)2 in a 2:3 molar ratio furnished coordination cages, which was ascertained by crystallography, 1H NMR and DOSY measurements, and ESI-TOFMS and UV-vis spectra. The cages have six phenolic hydroxy groups inside and were expected to incorporate hydrogen-bonding guest molecules such as saccharides. CD and DOSY measurements showed that octyl hexoside guests could be incorporated into the cage.

Spontaneous Helix Formation of " meta"-Ethynylphenol Oligomers by Sequential Intramolecular Hydrogen Bonding inside the Cavities.

Phenol-based oligomers linked with acetylenes at their meta positions, " meta"-ethynylphenol oligomers, were developed as a synthetic helical foldamer. The architecturally simple oligomers spontaneously formed helical higher-order structures by sequential intramolecular hydrogen bonds through the multiple phenolic hydroxy groups inside the cavities. The hydrogen bonds forced C-C≡C-C bond angles to largely bend toward the inside. Addition of chiral amines caused the helices to be chiral by electrostatic interactions between the resulting chiral ammonium cations and the phenolate anions.

AldB controls persister formation in Escherichia coli depending on environmental stress.

Persisters are multidrug-tolerant cells that are present within antibiotic-sensitive populations. Persister formation is not induced by genetic mutations, but rather by changes in the degree of expression of some genes. High redundancy has been observed among the pathways that have been hypothesized to respond to specific stresses. In this study, we conducted RNA sequencing of Escherichia coli persisters under various stress conditions to identify common mechanisms. We induced stresses such as glucose or amino acid exhaustion, acid stress and anaerobic conditions, all of which are encountered during bacterial pathogenesis. We found that most genes are differentially expressed depending on the specific stress condition; however, some genes were commonly expressed in persisters in most stress conditions. Commonly expressed genes are expected to be promising therapeutic targets for combating persistent infections. We found that knockdown of aldehyde dehydrogenase (aldB), which was expressed in every condition except for acid stress, decreased persisters in the non-stressed condition. However, the same strain unexpectedly showed an increased number of persisters in the amino acid-limited condition. Because the increase in persister number is glycolytic metabolite-dependent, metabolic flow may play a crucial role in aldB-mediated persister formation. These data suggest that environmental stresses alter persister mechanisms. Identification of environmental influences on persister formation during pathogenesis is therefore necessary to enabling persister eradication.

Long-Term Health Effects of PCBs and Related Compounds: A Comparative Analysis of Patients Suffering from Yusho and the General Population.

Yusho, which refers to a mass poisoning caused by the ingestion of rice bran oil contaminated with polychlorinated biphenyls, polychlorinated dibenzo-p-dioxins, and polychlorinated dibenzofurans, was first reported in October 1968 in Japan. Yusho patients suffer from various symptoms; however, after 40 years, some emerging symptoms have been attributed to aging. The prevalence of symptoms and diseases among Yusho patients and the general population was compared in this study. The data obtained from the survey among Yusho patients (1131 patients) by the Ministry of Health, Labour, and Welfare of Japan in 2008 were compared with the data from a survey conducted among the general population. When selecting the comparison group, the age and residential area (prefecture) were taken into account to match the baseline characteristics of Yusho patients. A logistic regression analysis was performed to identify the association between Yusho and the prevalence of symptoms and was adjusted for various potential confounding factors (age, sex, body mass index, cigarette smoking, frequency of drinking, and walking time). Skin pigmentation and acneiform eruption were found to be characteristic symptoms of Yusho and were more prevalent in these patients. Other symptoms and diseases associated with Yusho included orthostatic hypotension, hypohidrosis, dysgeusia, Basedow's disease, hoarseness, cardiac insufficiency, tachycardia, eczema, and hair loss. Symptoms related to aging, such as general fatigue, arthralgia, and numbness in the extremities, were significantly higher in Yusho patients after adjusting for age and lifestyle. This study demonstrated that, 40 years after the outbreak of Yusho, the prevalence of various symptoms and diseases in Yusho patients, including age-related diseases, was higher than that in the general population.

Highly Oriented J-Aggregates of Nitroazo Dye and Its Surface-Induced Chromism.

Highly oriented J-aggregates of a nitroazo dye were obtained in solid thin films on aligned poly(tetrafluoroethylene) surfaces. During film deposition on a friction-transferred poly(tetrafluoroethylene) layer, a sharp peak grew in the polarized absorption spectra around 613 nm, which was red-shifted 117 nm from the peak in dilute dichloromethane solution. The peak showed remarkable optical anisotropy: dichroic ratios D of up to 22 were observed, and the intrinsic D value should substantially exceed this value. These results indicate that the peak is attributable to highly oriented J-aggregates. On glass, however, H-like aggregates grew, exhibiting an absorption peak at 410 nm. Hence, the substrate surface induced the remarkable chromism observed as a 203 nm red shift.

Change in decay rates of dioxin-like compounds in Yusho patients.

BACKGROUND: Once ingested, dioxins and dioxin-like compounds are excreted extremely slowly. Excretion can be evaluated by its half-life. Half-lives estimated from observed concentrations are affected by excretion and ongoing exposure. We investigated the change in apparent half-life using a theoretical model based on exposure to dioxin and dioxin-like compounds. METHODS: We carried out longitudinal measurements of the blood concentration of dioxins and dioxin-like compounds in a Yusho cohort during 2002 to 2010. We estimated the change in decay rates of 2,3,4,7,8-PeCDF and octachlorodibenzodioxin (OCDD) using a second-order equation. RESULTS: We found that the decay rate of OCDD increased, whereas the decay rate of 2,3,4,7,8-PeCDF of patients with a relatively high concentration of 2,3,4,7,8-PeCDF decreased. OCDD results were in accordance with decreasing levels of dioxin and dioxin-like compounds in the environment. The decay rate of OCDD in the body was affected by the decay rate of OCDD in the environment by ingestion because it was near the steady-state. In contrast, the decay rate of 2,3,4,7,8-PeCDF in the body was affected less by ingestion from the environment because it was far higher than in the steady-state. CONCLUSION: We demonstrated that the level of 2,3,4,7,8-PeCDF in the environment is decreasing. The excretion half-life is longer than the environmental half-life, thus the excretion half-life in a Yusho patient is increased.

Neuraminidase Amino Acid Sequences of Influenza A/H3N2 and B Viruses Isolated from Influenza Patients in the 2014/15 Japanese Influenza Season.

Background: Neuraminidase (NA) is a surface protein essential for influenza virus replication. NA inhibitors are commonly used for the treatment of influenza patients in Japan. Several mutations that reduce the effect of NA inhibitors have been reported. We sequenced the whole NA segment of isolated virus from influenza patients and investigated the relation between the NA amino acid sequence and the 50％ inhibitory concentration (IC_50) of four NA inhibitors. Materials and Methods: Forty A/H3N2 and 19 B influenza virus isolated from patients in the 2014/15 influenza season were analyzed. The IC_50 was determined by a neuraminidase inhibition assay using a fluorescent substrate. Viral RNA was amplified by RT-PCR and the genome was sequenced using a next generation sequencer. The deduced amino acid sequences were analyzed. Results: There was no AA change in the NA catalytic site of the A/H3N2 and B viruses isolated in the 2014-15 influenza season. There was no significant relation between the NA amino acids and the IC_50 of the four NA inhibitors for A/H3N2 or B viruses. Conclusion: The catalytic site of NA was highly conserved for these A/H3N2 and B viruses. No emergence of NA amino acid mutations related to the sensitivity of the four currently used NA inhibitors was observed.