Acidic growth conditions stabilize the ribosomal RNA gene cluster and extend lifespan through noncoding transcription repression.

Blackcurrant (Ribes nigrum L.) is a classical fruit that has long been used to make juice, jam, and liqueur. Blackcurrant extract is known to relieve cells from DNA damage caused by hydrogen peroxide (H2 O2 ), methyl methane sulfonate (MMS), and ultraviolet (UV) radiation. We found that blackcurrant extract (BCE) stabilizes the ribosomal RNA gene cluster (rDNA), one of the most unstable regions in the genome, through repression of noncoding transcription in the intergenic spacer (IGS) which extended the lifespan in budding yeast. Reduced formation of extrachromosomal circles (ERCs) after exposure to fractionated BCE suggested that acidity of the growth medium impacted rDNA stability. Indeed, alteration of the acidity of the growth medium to pH ~4.5 by adding HCl increased rDNA stability and extended the lifespan. We identified RPD3 as the gene responsible for this change, which was mediated by the RPD3L histone deacetylase complex. In mammals, as inflammation sites in a tissue are acidic, DNA maintenance may be similarly regulated to prevent genome instability from causing cancer.

Preoperative sleep-disordered breathing and craniofacial abnormalities are risk factors for postoperative sleep-disordered breathing in patients undergoing skin-flap oropharyngeal reconstruction surgery for oral cavity cancer: a prospective case-control study.

PURPOSE: After oropharyngeal reconstruction surgery, excessive flap volume within the oral cavity may increase the risk of pharyngeal obstruction during sleep. This prospective observational study aimed to test a hypothesis that the skin-flap oropharyngeal reconstructive surgery increases nocturnal apnea-hypopnea index (nAHI, primary variable) after surgery. METHODS: Adult patients undergoing oropharyngeal reconstruction surgery participated in this study. The hypothesis was tested by comparing the results of portable type 4 sleep study and craniofacial assessments with lateral head and neck computed tomography scout image before and after surgery. Multiple linear regression analyses were performed to identify predictors for nAHI increase after the surgery. RESULTS: In 15 patients, a postoperative sleep study was performed at 41 (27, 59) (median (IQR)) days after the surgery. nAHI did not increase after the surgery (mean (95% CI), 13.0 (7.2 to 18.7) to 18.4 (10.2 to 26.6) events.hour-1, p = 0.277), while apnea index significantly increased after the surgery (p = 0.026). Use of the pedicle flap for the oropharyngeal reconstruction (p = 0.051), small mandible (p = 0.008), longer lower face (0.005), and larger tongue size (p = 0.008) were independent predictors for worsening of nAHI after surgery. Hospital stay was significantly longer in patients with the pedicle flap (n = 8) than in those with the free flap (n = 7) (p = 0.014), and the period of hospital stay was directly associated with increase of nAHI after surgery (r = 0.788, p < 0.001, n = 15). CONCLUSIONS: Oropharyngeal reconstruction surgery worsens sleep-disordered breathing in some patients with craniofacial and surgical risk factors. TRIAL REGISTRATION: UMIN Clinical Trial Registry (UMIN000036260, March 22, 2019), https://rctportal.niph.go.jp/s/detail/um?trial_id=UMIN000036260.

Expression analysis and functional characterization of thioredoxin domain-containing protein 11.

BACKGROUNDS: The endoplasmic reticulum (ER) is a crucial organelle that regulates both the folding, modification and transport of many proteins and senses certain stimuli inside and outside of cells. ER-associated degradation (ERAD), including SEL1L is a crucial mechanism to maintain homeostasis. In this study, we performed comparative proteome analysis in wild-type (wt) and SEL1L-deficient cells. METHODS AND RESULTS: We found constitutively high expression of thioredoxin domain-containing protein 11 (TXNDC11) mRNA and protein in our SEL1L-deficient HEK293 cells by RT-PCR and Western blot analysis. The TXNDC11 gene possesses a well-conserved unfolded protein response element (UPRE) around its transcription start site, and ER stress increased TXNDC11 mRNA and luciferase reporter activity via this putative UPRE in HEK293 cells. The amounts of TXNDC11 protein in wild-type and SEL1L-deficient cells with or without thapsigargin (Tg) treatment were parallel to their mRNAs in these cells, which was almost proportional to spliced XBP1 (sXBP1) mRNA expression. The establishment and characterization of TXNDC11-deficient HEK293 cells revealed that the expression of three different ER resident stress sensors, ATF6α, CREB3 and CREB3L2, is regulated by TXNDC11. The rate of disappearance of the three proteins by CHX treatment in wt cells was remarkably different, and the full-length CREB3L2 protein was almost completely degraded within 15 min after CHX treatment. TXNDC11 deficiency increased the expression of each full-length form under resting conditions and delayed their disappearance by CHX treatment. Interestingly, the degree of increase in full-length CREB3/CREB3L2 by TXNDC11 deficiency was apparently higher than that in full-length ATF6α. The increase in these proteins by TXNDC11 deficiency was hardly correlated with the expression of each mRNA. Treatment with ER stress inducers influenced each full-length mature form, and the difference in each full-length form observed in wt and TXNDC11-deficient cells was smaller. CONCLUSION: This study demonstrated that TXNDC11 is an ER stress-inducible gene regulated by the IRE1-sXBP1 pathway. In addition, TXNDC11 is involved in the regulation of ATF6α, CREB3 and CREB3L2 protein expression, although the contribution to the stability of these proteins is quite variable. Therefore, its further characterization will provide new insights for understanding protein homeostasis in ER physiology and pathology.

Comparative Analysis of CREB3 and CREB3L2 Protein Expression in HEK293 Cells.

We performed a comparative analysis of two ER-resident CREB3 family proteins, CREB3 and CREB3L2, in HEK293 cells using pharmacological and genome editing approaches and identified several differences between the two. Treatment with brefeldin A (BFA) and monensin induced the cleavage of full-length CREB3 and CREB3L2; however, the level of the full-length CREB3 protein, but not CREB3L2 protein, was not noticeably reduced by the monensin treatment. On the other hand, treatment with tunicamycin (Tm) shifted the molecular weight of the full-length CREB3L2 protein downward but abolished CREB3 protein expression. Thapsigargin (Tg) significantly increased the expression of only full-length CREB3L2 protein concomitant with a slight increase in the level of its cleaved form. Treatment with cycloheximide and MG132 revealed that both endogenous CREB3 and CREB3L2 are proteasome substrates. In addition, kifunensine, an α-mannosidase inhibitor, significantly increased the levels of both full-length forms. Consistent with these findings, cells lacking SEL1L, a crucial ER-associated protein degradation (ERAD) component, showed increased expression of both full-length CREB3 and CREB3L2; however, cycloheximide treatment downregulated full-length CREB3L2 protein expression more rapidly in SEL1L-deficient cells than the full-length CREB3 protein. Finally, we investigated the induction of the expression of several CREB3 and CREB3L2 target genes by Tg and BFA treatments and SEL1L deficiency. In conclusion, this study suggests that both endogenous full-length CREB3 and CREB3L2 are substrates for ER-associated protein degradation but are partially regulated by distinct mechanisms, each of which contributes to unique cellular responses that are distinct from canonical ER signals.

Synthesis of Bis{meso-Tetrakis(4-N-alkylpyridiniumyl)porphyrinato}cerium and Its Redox Switching Behavior.

A novel double-decker porphyrin complex, bis{meso-tetrakis(4-N-alkylpyridiniumyl)porphyrinato}cerium, was prepared. Electrochemical measurements revealed that this complex exhibited reversible redox waves corresponding to a 1e- redox reaction of the cerium center. Treating the complex alternately with an oxidant and a reductant resulted in the reversible redox switching between the oxidized and reduced states in an organic solvent.

Spectroscopic Characterization of Halorhodopsin Reconstituted into Nanodisks Using Native Lipids.

We successfully reconstituted single Natronomonas pharaonis halorhodopsin (NpHR) trimers into a nanodisk (ND) using the native archaeal lipid (NL) and an artificial lipid having a zwitterionic headgroup, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). Incorporation of single trimeric NpHR into NDs was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis, size-exclusion chromatography, and visible circular dichroism spectroscopy. The Cl- binding affinity of NpHR in NDs using NL (NL-ND NpHR) or POPC (POPC-ND NpHR) was examined by absorption spectroscopy, showing that the Cl--releasing affinities (Kd,N↔O) of these ND-reconstituted NpHRs are more than 10 times higher than that obtained from native NpHR membrane fragments (MFs) harvested from a NpHR-overexpressing archaeal strain (MF NpHR). The photoreaction kinetics of these ND-reconstituted NpHRs revealed that the Cl- uptake was faster than that of MF NpHR. These differences in the Cl--releasing and uptake properties of ND-reconstituted NpHRs and MF NpHR may arise from suppression of protein conformational changes associated with Cl- release from the trimeric NpHR caused by ND reconstitution, conformational perturbation in the trimeric state, and loss of the trimer-trimer interactions. On the other hand, POPC-ND NpHR demonstrated accelerated Cl- uptake compared to NL-ND NpHR, suggesting that the negative charge on the archaeal membrane surface regulates the photocycle of NpHR. Although NL-ND NpHR and MF NpHR are embedded in the same lipid, the lower Cl--binding affinity at the initial state (Kd,initial) and faster recovering from the NpHR' state to the original state of the photoreaction cycle were observed for NL-ND NpHR, probably because of insufficient interactions with a chromophore in the native membrane, bacterioruberin in reconstituted NDs. Our results indicate that specific interactions of NpHR with surrounding lipids and bacterioruberin, structural flexibility of the membrane, and interactions between trimeric NpHRs may be necessary for efficient Cl- pumping.

Protective effects of raw and cooked blackcurrant extract on DNA damage induced by hydrogen peroxide in human lymphoblastoid cells.

CONTEXT: Blackcurrant (Ribes nigrum L.) is a classical fruit that has long been used to make juice, liqueur and sometimes medicines in Europe. The beneficial effects of blackcurrant, which are inhibition of lipopolysaccharide-stimulated inflammatory, anticarcinogenesis and other health effects, have been reported. OBJECTIVE: Previously, we reported the antimutagenic activities of blackcurrant using a yeast gene mutation assay. In this study, we investigated whether this antimutagenicity of blackcurrant was confirmed in human cells. MATERIALS AND METHODS: We prepared four types of aqueous blackcurrant extracts (BCE) from mature and premature with or without heat treatment by microwave. Antioxidant activities of BCE were measured by the DPPH radical scavenger assay. In the DPPH radical scavenger assay, the maximum concentration of BCE was 1.6 mg/reaction. We investigated the antigenotoxic activities of BCE by the comet assay and micronucleus test using the human lymphoblastoid cell line TK6. In the comet assay, TK6 was treated with 300 µM H2O2 without or with BCE at concentrations of 0.5, 1.0, 2.0 and 3.0 mg/mL. In the micronucleus test, TK6 was treated with 1 mg/mL BCE without or with H2O2. RESULTS: All BCEs exhibited more than 90% of inhibition rates of DPPH radicals at the maximum concentration of BCE. DNA damage and micronuclei induced by H2O2 significantly decreased in the each BCE-treated condition. CONCLUSION: The results suggest that BCE treatment can reduce the genomic instability induced by H2O2 in human cells. We consider that these antigenotoxic effects are related to polyphenols, l-ascorbic acid and other antioxidant compounds.

Lemon Myrtle (Backhousia citriodora) Extract and Its Active Compound, Casuarinin, Activate Skeletal Muscle Satellite Cells In Vitro and In Vivo.

Sarcopenia is an age-related skeletal muscle atrophy. Exercise is effective in improving sarcopenia via two mechanisms: activation of skeletal muscle satellite cells (SCs) and stimulation of muscle protein synthesis. In contrast, most nutritional approaches for improving sarcopenia focus mainly on muscle protein synthesis, and little is known about SC activation. Here, we investigated the effect of lemon myrtle extract (LM) on SC activation both in vitro and in vivo. Primary SCs or myoblast cell lines were treated with LM or its derived compounds, and incorporation of 5-bromo-2'-deoxyuridine, an indicator of cell cycle progression, was detected by immunocytochemistry. We found that LM significantly activated SCs (p < 0.05), but not myoblasts. We also identified casuarinin, an ellagitannin, as the active compound in LM involved in SC activation. The structure−activity relationship analysis showed that rather than the structure of each functional group of casuarinin, its overall structure is crucial for SC activation. Furthermore, SC activation by LM and casuarinin was associated with upregulation of interleukin-6 mRNA expression, which is essential for SC activation and proliferation. Finally, oral administration of LM or casuarinin to rats showed significant activation of SCs in skeletal muscle (p < 0.05), suggesting that LM and casuarinin may serve as novel nutritional interventions for improving sarcopenia through activating SCs.

Analysis of airborne sputum droplets flow dynamic behaviors under different ambient conditions and aerosol size effects.

The coronavirus (COVID-19) is becoming more threatening with the emergence of new mutations. New virus transmission and infection processes remain challenging and re-examinations of proper protection methods are urgently needed. From fluid dynamic viewpoint, the transmission of virus-carrying droplets and aerosols is one key to understanding the virus-transmission mechanisms. This study shows virus transmission by incorporating flow-evaporation model into the Navier-Stokes equation to describe the group of airborne sputum droplets exhaled under Rosin-Rammler distribution. Solid components and humidity field evolution are incorporated in describing droplet and ambient conditions. The numerical model is solved by an inhouse code using advection-diffusion equation for the temperature field and the humidity field, discretized by applying the total-variation diminishing Runge-Kutta method. The results of this study are presented in detail to show the different trends under various ambient conditions and to reveal the major viral-transmission routes as a function of droplet size.

Assessment of Dysphonia Using the Japanese Version of the Voice Handicap Index and Determination of Cutoff Points for Screening.

INTRODUCTION: The Voice Handicap Index (VHI) is recognized as a useful subjective assessment method for dysphonia. The original VHI has been translated into numerous other languages, including Japanese (J-VHI). Although the reliability and validity of the J-VHI have already been established, the cutoff point has not been determined. The aims of this study were to investigate the relationship between the J-VHI and other voice laboratory measurements, and determine the cutoff point. METHOD: This study included 167 dysphonic patients and 55 healthy volunteers. All patients and volunteers completed the J-VHI at the initial visit, and the following outcomes were determined: VHI scores of patients with dysphonia and healthy volunteers, VHI scores according to disease, cutoff point, and correlations between VHI scores and other voice laboratory measurements. RESULTS: Both the total VHI (VHI-T) and individual domain (functional domain [VHI-F], emotional domain [VHI-E], physical domain [VHI-P]) scores were significantly higher in the dysphonia group compared to the healthy volunteer group. VHI-T, VHI-F, and VHI-E scores were significantly lower in the benign mucosal lesion subgroup, compared to the other disease subgroups. The G scale and B scale of the grade-roughness-breathiness-asthenia-strain scale showed a significant association with VHI-T, VHI-F, and VHI-P scores. Similarly, the A scale showed a significant association with VHI-T, VHI-F, and VHI-E scores. The cutoff point (12) for VHI-T was chosen from the receiver operating characteristic curve to maximize sensitivity and specificity. Similarly, the cutoff points for VHI-F (5), VHI-P (5), and VHI-E (3) were also obtained. Significant differences in maximum phonation time, pitch range, G scale, and B scale were observed between the VHI-T negative (VHI ≤ 12) and positive (VHI-T > 13) groups. CONCLUSION: These findings suggest that self-evaluation using the VHI could serve as an independent assessment and screening tool for patients with dysphonia.

Involvement of mismatch repair proteins in adaptive responses induced by N-methyl-N'-nitro-N-nitrosoguanidine against γ-induced genotoxicity in human cells.

As humans are exposed to a variety of chemical agents as well as radiation, health effects of radiation should be evaluated in combination with chemicals. To explore combined genotoxic effects of radiation and chemicals, we examined modulating effects of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), a direct-acting methylating agent, against genotoxicity of γ-radiation. Human lymphoblastoid TK6 cells and its mismatch-deficient derivative, i.e., MT1 cells, were treated with MNNG for 24h before they were exposed to γ-irradiation at a dose of 1.0 Gy, and the resulting genotoxicity was examined. In TK6 cells, the pretreatments with MNNG at low doses suppressed frequencies of the thymidine kinase (TK) gene mutation and micronucleus (MN) formation induced by γ-irradiation and thus the dose responses of TK and MN assays were U-shaped along with the pretreatment doses of MNNG. In contrast, the genotoxic effects of MNNG and γ-irradiation were additive in MT1 cells and the frequencies of TK mutations and MN induction increased along with the doses of MNNG. Apoptosis induced by γ-radiation was suppressed by the pretreatments in TK6 cells, but not in MT1 cells. The expression of p53 was induced and cell cycle was delayed at G2/M phase in TK6, but not in MT1 cells, by the treatments with MNNG. These results suggest that pretreatments of MNNG at low doses suppress genotoxicity of γ-radiation in human cells and also that mismatch repair proteins are involved in the apparent adaptive responses.

Induction of TK mutations in human lymphoblastoid TK6 cells by the rat carcinogen 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX).

3-Chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), a chlorine disinfection by-product in drinking water, is carcinogenic in rats and genotoxic in mammalian cells in vitro. In the current study, the mechanism of genotoxicity of MX in human lymphoblastoid TK6 cells was investigated by use of the Comet assay, the micronucleus test, and the thymidine kinase (TK) gene-mutation assay. MX induced a concentration-dependent increase in micronuclei and TK mutations. The lowest effective concentrations in the MN test and the TK gene-mutation assay were 37.5µM and 25µM, respectively. In the Comet assay, a slight although not statistically significant increase was observed in the level of DNA damage induced by MX in the concentration range of 25-62.5µM. Molecular analysis of the TK mutants revealed that MX induced primarily point mutations or other small intragenic mutations (61%), while most of the remaining TK mutants (32%) were large deletions at the TK locus, leading to the hemizygous-type loss-of-heterozygosity (LOH) mutations. These findings show that aside from inducing point mutations, MX also generates LOH at the TK locus in human cells and may thus cause the inactivation of tumour-suppressor genes by LOH.

Evaporation flow characteristics of airborne sputum droplets with solid fraction: Effects of humidity field evolutions.

The continuance of the COVID-19 pandemic largely depends on the spread of virus-carrying aerosols in ambient air. The mechanism of virus transmission and infection remains under intense investigation. In this study, an evaporation flow model of airborne sputum droplets is proposed which considers the evolution effects of the humidity field under different particle distributions and solid/salt fraction interactions. The incompressible Navier-Stokes equations characterize a stream of airflow jets, and the convection-diffusion-evaporation process is used to account for the inhomogeneous humidity field caused by the respiratory tract. Momentum equations for droplet dynamics which involve the effects of drag, gravity, and Brownian motion on sputum droplets are introduced to quantify the transport of droplets in a humidity field. The Lattice Boltzmann method is used to track the evolution of the aerosol in space and time under different ambient temperature and relative humidity conditions. The results of the simulation demonstrate that airborne humidity accelerates the evaporation rate of droplet, while supersaturated humid air forms a vapor mass in front of the respiratory tract. Despite the short lifespan of this phenomenon, it significantly hinders the evaporation of the droplets. Besides, the droplet vortex dynamics in a humidity field are sensitive to the droplet size.

A new Late Miocene great ape from Kenya and its implications for the origins of African great apes and humans.

Extant African great apes and humans are thought to have diverged from each other in the Late Miocene. However, few hominoid fossils are known from Africa during this period. Here we describe a new genus of great ape (Nakalipithecus nakayamai gen. et sp. nov.) recently discovered from the early Late Miocene of Nakali, Kenya. The new genus resembles Ouranopithecus macedoniensis (9.6-8.7 Ma, Greece) in size and some features but retains less specialized characters, such as less inflated cusps and better-developed cingula on cheek teeth, and it was recovered from a slightly older age (9.9-9.8 Ma). Although the affinity of Ouranopithecus to the extant African apes and humans has often been inferred, the former is known only from southeastern Europe. The discovery of N. nakayamai in East Africa, therefore, provides new evidence on the origins of African great apes and humans. N. nakayamai could be close to the last common ancestor of the extant African apes and humans. In addition, the associated primate fauna from Nakali shows that hominoids and other non-cercopithecoid catarrhines retained higher diversity into the early Late Miocene in East Africa than previously recognized.

[Criteria for differential diagnosis of epilepsy and long QT syndrome presenting with seizures and EEG abnormalities].

We report a 13-year-old girl with congenital long QT syndrome (LQTS) who developed a cluster of generalized tonic clonic seizures with post-ictal EEG abnormality. The provisional diagnosis was epilepsy. However, ECG monitoring showed torsade de pointes, and thus the final diagnosis was LQTS. Although LQTS can be potentially misdiagnosed as epilepsy when it presents with seizures, it is important to differentiate LQTS from epilepsy because patients with LQTS are at risk of sudden death. We reviewed 11 previously reported cases with LQTS and EEG abnormalities who were initially diagnosed as epilepsy. We emphasized the importance of the following five criteria in the differentiation of LQTS from epilepsy: 1) awareness that LQT2 and LQT3 can cause life-threatening arrhythmia at rest or during sleep, 2) examination of arterial pulse during seizures, 3) monitoring ECG during EEG recording, 4) careful establishment of the correct diagnosis taking into consideration the interictal EEG findings, and 5) reconsidering the possibility of cardiac origin when the attacks cannot be controlled even by therapeutic levels of antiepileptic drugs in the blood.

Temperature-sensitive photoreactivation of cyclobutane thymine dimer in soybean.

UV radiation induces the formation of two classes of photoproducts in DNA, the cyclobutane pyrimidine dimer (CPD) and the pyrimidine 6-4 pyrimidone photoproduct. CPDs in plants are repaired by class II CPD photolyase via a UV-A/blue light-dependent mechanism. The genes for the class II CPD photolyase have been cloned from higher plants such as Arabidopsis, Cucumis sativus (cucumber), Oryza sativa (rice) and Spinacia oleracea (spinach). Flavin adenine dinucleotide (FAD) has been identified as a cofactor. Here we report the isolation and characterization of the CPD photolyase cDNA from soybean (Glycin max). The sequence of amino acids predicted from the cDNA sequence was highly homologous to sequences of higher plant class II CPD photolyases. When the cDNA was expressed in a photolyase-deficient Escherichia coli, photoreactivation activity was partially restored by illumination with a fluorescent light. The purified enzyme showed CPD binding and light-dependent photoreactivation activities in vitro. When soybean CPD photolyase was heat-treated in vitro from 25 degrees C to 45 degrees C for 3 min, thymine dimer-binding activity and photoreactivation activity were decreased, and FAD was released from the enzyme. On the other hand, when the enzyme-CPD complex was heat-treated, photoreactivation activity was stable. We argue that FAD in the soybean CPD photolyase is labile for temperature, but once the enzyme-CPD complex has formed, FAD becomes tightly bound to the enzyme or complex.

Biochemical and biological properties of DNA photolyases derived from utraviolet-sensitive rice cultivars.

Class I and class II CPD photolyases are enzymes which repair pyrimidine dimers using visible light. A detailed characterization of class I CPD photolyases has been carried out, but little is known about the class II enzymes. Photolyases from rice are suitable for functional analyses because systematic breeding for long periods in Asian countries has led to the selection of naturally occurring mutations in the CPD photolyase gene. We report the biochemical characterization of rice mutant CPD photolyases purified as GST-form from Escherichia coli. We identified three amino acid changes, Gln126Arg, Gly255Ser, and Gln296His, among which Gln but not His at 296 is important for complementing phr-defective E. coli, binding UV-damage in E. coli, and binding thymine dimers in vitro. The photolyase with Gln at 296 has an apoenzyme:FAD ratio of 1 : 0.5 and that with His at 296 has an apoenzyme:FAD ratio of 1 : 0.12-0.25, showing a role for Gln at 296 in the binding of FAD not in the binding of thymine dimer. Concerning Gln or Arg at 126, the biochemical activity of the photolyases purified from E. coli and complementing activity for phr-defective E. coli are similarly proficient. However, the sensitivity to UV of cultivars differs depending on whether Gln or Arg is at 126. The role of Gln and Arg at 126 for photoreactivation in rice is discussed.

Radioprotective activity of blackcurrant extract evaluated by in vitro micronucleus and gene mutation assays in TK6 human lymphoblastoid cells.

INTRODUCTION: Blackcurrant (Ribs nigrum L.) is a classical fruit that has long been used to prepare juice, jam, liqueur, and sometimes medicines in Europe. Previously, we reported a genome defense effect by the antioxidative activity of several types of blackcurrant extracts (BCEs) in yeast and human cell gene mutation assays. In this study, we determined if BCE exerted radioprotective activity against DNA damage, chromosomal aberration, and gene mutations in the TK6 human lymphoblastoid cell line. We prepared aqueous BCE extracted from mature fruits cultivated in the Aomori Prefecture, Japan. FINDINGS: In the micronucleus test and TK gene mutation assay, TK6 cells were irradiated with 0, 0.125, 0.250, 0.500, and 1.000 Gy with or without 1.0 mg/mL BCE. Intracellular hydrogen peroxide (H2O2) was measured using the fluorescent probe BES-H2O2-Ac. Induction of micronuclei and gene mutations by γ-irradiation exposure was suppressed in combination with BCE. In addition, BCE reduced intracellular H2O2 levels caused by γ-irradiation. CONCLUSIONS: Our findings clearly support the genome defense potential of blackcurrant against γ-induced DNA damage. We postulate that these genome defense activities are related to the antioxidant compounds in blackcurrant.

Mefenamic acid enhances anticancer drug sensitivity via inhibition of aldo-keto reductase 1C enzyme activity.

Resistance to anticancer medications often leads to poor outcomes. The present study explored an effective approach for enhancing chemotherapy targeted against human cancer cells. Real-time quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed overexpression of members of aldo-keto reductase (AKR) 1C family, AKR1C1, AKR1C2, AKR1C3, and AKR1C4, in cisplatin, cis-diamminedichloroplatinum (II) (CDDP)-resistant human cancer cell lines, HeLa (cervical cancer cells) and Sa3 (oral squamous cell carcinoma cells). The genes were downregulated using small-interfering RNA (siRNA) transfection, and the sensitivity to CDDP or 5-fluorouracil (5-FU) was investigated. When the genes were knocked down, sensitivity to CDDP and 5-FU was restored. Furthermore, we found that administration of mefenamic acid, a widely used non-steroidal anti-inflammatory drug (NSAID) and a known inhibitor of AKR1Cs, enhanced sensitivity to CDDP and 5-FU. The present study suggests that AKR1C family is closely associated with drug resistance to CDDP and 5-FU, and mefenamic acid enhances their sensitivity through its inhibitory activity in drug-resistant human cancer cells. Thus, the use of mefenamic acid to control biological function of AKR1C may lead to effective clinical outcomes by overcoming anticancer drug resistance.

Haem-dependent dimerization of PGRMC1/Sigma-2 receptor facilitates cancer proliferation and chemoresistance.

Progesterone-receptor membrane component 1 (PGRMC1/Sigma-2 receptor) is a haem-containing protein that interacts with epidermal growth factor receptor (EGFR) and cytochromes P450 to regulate cancer proliferation and chemoresistance; its structural basis remains unknown. Here crystallographic analyses of the PGRMC1 cytosolic domain at 1.95 Å resolution reveal that it forms a stable dimer through stacking interactions of two protruding haem molecules. The haem iron is five-coordinated by Tyr113, and the open surface of the haem mediates dimerization. Carbon monoxide (CO) interferes with PGRMC1 dimerization by binding to the sixth coordination site of the haem. Haem-mediated PGRMC1 dimerization is required for interactions with EGFR and cytochromes P450, cancer proliferation and chemoresistance against anti-cancer drugs; these events are attenuated by either CO or haem deprivation in cancer cells. This study demonstrates protein dimerization via haem-haem stacking, which has not been seen in eukaryotes, and provides insights into its functional significance in cancer.