Biosensing with Surface-Charge-Modulated Graphene Field-Effect Transistors beyond Nonlinear Electrolytic Screening.

In field-effect transistor (FET) biosensors, charge screening in electrolyte solutions limits the sensitivity, thereby restricting the applicability of FET sensors. This is particularly pronounced in graphene FET (GFET) biosensors, where the bare graphene surface possesses a strongly negative charge, which impedes the high sensitivity of GFETs owing to nonlinear electrolytic screening at the interfaces between graphene and liquid. In this study, we counteracted the negative surface charge of graphene by decorating positively charged compounds and demonstrated the sensing of C-reactive protein (CRP) with surface-charge-modulated GFETs (SCM-GFETs). We integrated multiple SCM-GFETs with anti-CRP antibodies and nonfunctionalized GFETs into a chip and measured differentials to eliminate background changes to improve measurement reliability. The FET response corresponded to the fluorescence images, which visualized the specific adsorption of CRP. The estimated dissociation constant was consistent with previously reported values; this supports the conclusion that the results are attributed to specific adsorption. Conversely, the signal in GFETs without decoration was obscured by noise because of nonlinear electrolytic screening, further emphasizing the significance of surface-charge modulation. The limit of detection of the system was determined to be 2.9 nM. This value has the potential to be improved through further optimization of the surface charges to align with specific applications. Our devices effectively circumvent nonlinear electrolytic screening, opening the door for further advancements in GFET biosensor technology.

Stress-dependent cell stiffening by tardigrade tolerance proteins that reversibly form a filamentous network and gel.

Tardigrades are able to tolerate almost complete dehydration by entering a reversible ametabolic state called anhydrobiosis and resume their animation upon rehydration. Dehydrated tardigrades are exceptionally stable and withstand various physical extremes. Although trehalose and late embryogenesis abundant (LEA) proteins have been extensively studied as potent protectants against dehydration in other anhydrobiotic organisms, tardigrades produce high amounts of tardigrade-unique protective proteins. Cytoplasmic-abundant heat-soluble (CAHS) proteins are uniquely invented in the lineage of eutardigrades, a major class of the phylum Tardigrada and are essential for their anhydrobiotic survival. However, the precise mechanisms of their action in this protective role are not fully understood. In the present study, we first postulated the presence of tolerance proteins that form protective condensates via phase separation in a stress-dependent manner and searched for tardigrade proteins that reversibly form condensates upon dehydration-like stress. Through a comprehensive search using a desolvating agent, trifluoroethanol (TFE), we identified 336 proteins, collectively dubbed "TFE-Dependent ReversiblY condensing Proteins (T-DRYPs)." Unexpectedly, we rediscovered CAHS proteins as highly enriched in T-DRYPs, 3 of which were major components of T-DRYPs. We revealed that these CAHS proteins reversibly polymerize into many cytoskeleton-like filaments depending on hyperosmotic stress in cultured cells and undergo reversible gel-transition in vitro. Furthermore, CAHS proteins increased cell stiffness in a hyperosmotic stress-dependent manner and counteract the cell shrinkage caused by osmotic pressure, and even improved the survival against hyperosmotic stress. The conserved putative helical C-terminal region is necessary and sufficient for filament formation by CAHS proteins, and mutations disrupting the secondary structure of this region impaired both the filament formation and the gel transition. On the basis of these results, we propose that CAHS proteins are novel cytoskeleton-like proteins that form filamentous networks and undergo gel-transition in a stress-dependent manner to provide on-demand physical stabilization of cell integrity against deformative forces during dehydration and could contribute to the exceptional physical stability in a dehydrated state.

Retusone A, a Guaiane-Type Sesquiterpene Dimer from Wikstroemia retusa and Its Inhibitory Effects on Histone Acetyltransferase HBO1 Expression.

Retusone A (1), a new sesquiterpene dimer consisting of two guaiane-type sesquiterpenoids, and oleodaphnal (2) were isolated from heartwood of Wikstroemia retusa (Thymelaeaceae). The planar structure of 1 was elucidated on the basis of HRESIMS and NMR spectroscopic data, and the relative stereochemistry was established by X-ray diffraction analysis. The absolute configuration of 1 was determined by electronic circular dichroism. Compound 1 suppressed luciferase reporter gene expression driven by the HBO1 (histone acetyltransferase binding to ORC1) gene promoter in human breast cancer MCF7 cells. Compound 1 also decreased the expression of endogenous HBO1 mRNA and protein, and inhibited proliferation of the cells. These results suggest that retusone A (1), which has a unique dimeric sesquiterpenoid structure with inhibitory activity against HBO1 expression, may contribute to the development of a novel therapeutic candidate for the treatment of breast cancer.

Relationship between pre-sarcopenia and quality of life in patients with chronic liver disease: a cross-sectional study.

AIM: This study aimed to clarify the relationship between pre-sarcopenia (PS) and quality of life (QOL) in patients with chronic liver disease (CLD). PATIENTS AND METHODS: This cross-sectional study evaluated 335 patients with CLD. PS was diagnosed on the basis of the assessment criteria by the Japan Society of Hepatology. QOL was evaluated using the short form-36. RESULTS: Patients' mean age was 69.52 ± 10.17 years, and 169 (50.4%) participants were men. The prevalence of PS was 53.7%. Patients were divided into the PS and non-pre-sarcopenia (NPS) groups. Patients in the PS group were older (71.84 ± 9.78 vs. 66.81 ± 9.97, P < 0.01) and mostly women (65.2 vs. 37.8%, P < 0.01) compared with those in the NPS group. QOL, physical function (38.30 ± 17.63 vs. 44.02 ± 14.76, P < 0.01), physical role functioning (RP) (40.63 ± 15.38 vs. 44.88 ± 13.89, P < 0.01), and bodily pain (BP) (48.42 ± 11.45 vs. 51.24 ± 10.19, P = 0.02) were significantly lower in the PS group than in the NPS group. Logistic regression analyses identified that the independent predictive factors for PS were female sex (odds ratio: 3.16, 95% confidence interval: 2.01-4.98; P < 0.01) and RP (odds ratio: 1.97, 95% confidence interval: 1.24-3.12; P < 0.01). CONCLUSION: QOL characteristics of PS patients with CLD were low physical function, RP, and BP in short form-36. In addition, social role functioning was low in the PS patients aged 65-74 years, whereas RP and BP were low in those aged at least 75 years. Female sex and RP were independent predictors of PS according to the multivariate analysis. Maintaining and increasing muscle mass in patients with CLD may contribute toward improving physical QOL.

Health-related quality of life on the clinical course of patients with chronic hepatitis C receiving daclatasvir/asunaprevir therapy: A prospective observational study comparing younger (<70) and elderly (≥70) patients.

Interferon-free direct acting antiviral agent regimens for chronic hepatitis C (CHC) have been developed. These regimens have shown a high rate of sustained virologic response (SVR), and a reduction in side effects during treatment is also anticipated. However, the impact of the regimens on health-related quality of life (HRQOL) and side effects during treatment is not fully understood. The purpose of the present study was to evaluate HRQOL in the clinical course of patients with CHC receiving daclatasvir/asunaprevir (DCV/ASV) therapy using the Short Form-36 (SF-36) method. Twenty-eight patients with CHC receiving DCV/ASV therapy were analyzed in the present study, and HRQOL was measured by SF-36. Patients were asked to fill out the SF-36 prior to therapy (baseline), following 12 weeks of therapy, at the end of treatment and at SVR week 24 (SVR24) to evaluate HRQOL. Laboratory data were also investigated during the same period, and associations between these results and SF-36 were investigated. Aspartate aminotransferase, alanine aminotransferase, serum albumin, α-fetoprotein, platelet counts and Fibrosis (Fib)-4 index were all significantly improved at each time point when compared with baseline. With regard to alterations in HRQOL during therapy, the ≥70-year-old group displayed a significantly greater improvement in physical functioning during the period between baseline and 12 weeks when compared with the <70-year-old group. In the analysis of the SF-36 differences within each group, general health improved significantly in the ≥70-year-old group, as well as albumin levels. In addition, Fib-4-index significantly improved at all time points (12 and 24 weeks, and SVR24) when compared with baseline in the ≥70-year-old group. Therefore, DCV/ASV therapy may improve HRQOL and hepatic functional reserve, particularly in elderly patients.

Evaluation of the health-related quality of life using the 36-item short form health survey in patients with chronic hepatitis C receiving pegylated interferon/ribavirin/telaprevir triple treatment.

The rate of sustained virologic response (SVR) has increased in patients with chronic hepatitis C (CHC; genotype 1) since triple treatment with pegylated interferon (PEG-IFN), ribavirin (RBV) and telaprevir (TVR) was included in Japanese health insurance. However, side effects such as high-grade anemia and skin disorders means it is important to investigate the extent to which quality of life (QOL) is maintained during treatment. The impact on health-related (HR) QOL, as a result of TVR-based triple treatment was investigated long-term (48 weeks) in 34 patients (18 men, 16 women) following TVR-based triple treatment, using the 36-item short form health survey (SF-36). While scores for physical health were significantly lower during treatment, an improvement was seen in patients who showed complete response to treatment from 12 weeks following treatment (P<0.05). HRQOL improved significantly following completion of TVR-based triple treatment in these complete-responders, with higher scores compared with those prior to treatment. Anemia and skin symptoms appeared frequently during treatment and scores for physical health dropped. Particular care needs to be taken in regards to the management of side effects during TVR treatment. Further evaluations using the SF-36 may help in controlling doses to achieve SVR.

[Influence of interest region and interest value settings on exposure index-analysis using chest clinical images].

With the digitization of general radiography, there are some concerns about an increase in radiation exposure doses. Therefore, the exposure index (EI) as a new dose index was proposed in 2008 by IEC. However, the settings for the interest region and the interest value in the clinical image do not show concrete prescribed values. Therefore, we inspected the distribution of EI by changing the interest region and the interest value for a standing-position chest radiograph image in students' medical examinations. EI50f, which is generally used, fluctuated between 41 and 136. In addition, as the area of the interest region became smaller, EI increased and the variation index increased. For the interest value, 50% (EI50h) was smaller than 85% (EI85h), and the variation index was also smaller. EI was not the absolute value in the clinical image, and immediate display of the deviation index (DI) with target exposure index (EIT) was effective of the adequacy of the radiography condition.

Tetanus as cause of mass die-off of captive Japanese macaques, Japan, 2008.

In 2008 in Japan, 15/60 captive Japanese macaques died. Clostridium tetani was isolated from 1 monkey, and 11 had tetanus-specific symptoms. We conclude the outbreak resulted from severe environmental C. tetani contamination. Similar outbreaks could be prevented by vaccinating all monkeys, disinfecting housing areas/play equipment, replacing highly C. tetani-contaminated soil, and conducting epidemiologic surveys.

Oxidative stress-responsive intracellular regulation specific for the angiostatic form of human tryptophanyl-tRNA synthetase.

Tryptophanyl-tRNA synthetase (TrpRS) exists in two forms in human cells, i.e., a major form which represents the full-length protein and a truncated form (mini TrpRS) in which an NH(2)-terminal extension is deleted because of alternative splicing of its pre-mRNA. Mini TrpRS can act as an angiostatic factor, while full-length TrpRS is inactive. We herein show that an oxidized form of human glyceraldehyde-3-phosphate dehydrogenase (GapDH) interacts with both full-length and mini TrpRSs and specifically stimulates the aminoacylation potential of mini, but not full-length, TrpRS. In contrast, reduced GapDH did not bind to TrpRSs and did not influence their aminoacylation activity. Mutagenesis experiments clarified that the NH(2)-terminal Rossmann fold region of GapDH is crucial for its interaction with mini TrpRS as well as tRNA and for the regulation of its aminoacylation potential and suggested that monomeric GapDH can bind to mini TrpRS and stimulate its aminoacylation activity. These results suggest that the angiostatic human mini, but not the full-length, TrpRS may play an important role in the intracellular regulation of protein synthesis under conditions of oxidative stress.

Association of human neuroglobin with cystatin C, a cysteine proteinase inhibitor.

Neuroglobin (Ngb) is a newly discovered globin that is expressed in vertebrate brain. It has been reported that Ngb levels increase in neurons in response to oxygen deprivation, and that Ngb protects neurons from hypoxia. However, the mechanism of this neuroprotection remains unclear. In the present study, we identified human cystatin C, a cysteine proteinase inhibitor, as an Ngb-binding protein by using a yeast two-hybrid system. Surface plasmon resonance experiments verified that Ngb binds to cystatin C dimers, not to the monomers. Because both intracellular cystatin C and the amyloidogenic variant of cystatin C form dimers, Ngb may modulate the intracellular transport (or secretion) of cystatin C to protect against neuronal death under conditions of oxidative stress and/or it may have a role in the development of neurodegenerative diseases.

Human neuroglobin interacts with flotillin-1, a lipid raft microdomain-associated protein.

Neuroglobin (Ngb) is a newly discovered vertebrate globin that is expressed in the brain and that can reversibly bind oxygen. It has been reported that Ngb levels increase in neurons in response to oxygen deprivation, and that it protects neurons from hypoxia. However, the mechanism of this neuroprotection remains unclear. Recently, we found that oxidized human Ngb bound to the alpha-subunits of heterotrimeric G proteins (Galpha) and acted as a guanine nucleotide dissociation inhibitor for Galpha. To identify other Ngb-binding proteins, we herein screened a human brain cDNA library by using a yeast two-hybrid system. Among the plasmids isolated from positive clones, one contained an insert with 100% sequence identity to human flotillin-1. The interaction of Ngb with flotillin-1 was confirmed by glutathione S-transferase pull-down experiments. Since Galpha exists within lipid rafts critical for signal transduction and flotillin-1 recruits signaling proteins to lipid rafts, flotillin-1 might recruit Ngb to lipid rafts as a means of preventing neuronal death.

Oxidized human neuroglobin acts as a heterotrimeric Galpha protein guanine nucleotide dissociation inhibitor.

Neuroglobin (Ngb) is a newly discovered vertebrate heme protein that is expressed in the brain and can reversibly bind oxygen. It has been reported that Ngb expression levels increase in response to oxygen deprivation and that it protects neurons from hypoxia in vitro and in vivo. However, the mechanism of this neuroprotection remains unclear. In the present study, we tried to clarify the neuroprotective role of Ngb under oxidative stress in vitro. By surface plasmon resonance, we found that ferric Ngb, which is generated spontaneously as a result of the rapid autoxidation, binds exclusively to the GDP-bound form of the alpha subunit of heterotrimeric G protein (Galphai). In GDP dissociation assays or guanosine 5'-O-(3-thio)triphosphate binding assays, ferric Ngb behaved as a guanine nucleotide dissociation inhibitor (GDI), inhibiting the rate of exchange of GDP for GTP. The interaction of GDP-bound Galphai with ferric Ngb will liberate Gbetagamma, leading to protection against neuronal death. In contrast, ferrous ligand-bound Ngb under normoxia did not have GDI activities. Taken together, we propose that human Ngb may be a novel oxidative stress-responsive sensor for signal transduction in the brain.