Heat stress during grain filling regulates seed germination through alterations of DNA methylation in barley (Hordeum vulgare L.).

KEY MESSAGE: Alterations in DNA methylation levels of ROS, GA and ABA related gene promoters cause transcriptional changes upon imbibition to induce seed germination in barley seeds exposed to heat stress during grain filling. Environmental changes, especially changes in temperature, during seed development affect germination in several plant species. We have previously shown that heat stress during rice grain filling alters DNA methylation, an epigenetic mark important for gene silencing, regulates transcript levels of phytohormone metabolism genes, and delays seed germination. However, whether this phenomenon is present in other plant species remained to be elucidated. In this study, we compared seeds germination of barley (Hordeum vulgare L.) plants grown at 15 °C (control) or 25 °C (heat stress) during grain filling. Heat stress during grain filling significantly promoted seed germination in comparison with the control. The phytohormone gibberellic acid (GA) and reactive oxygen species produced by NADPH oxidases promote seed germination, whereas phytohormone abscisic acid (ABA) suppresses seed germination. We found that in heat-stressed seeds, genes related to ABA biosynthesis (HvNCED1 and 2) were significantly suppressed, whereas genes related to ABA catabolism (HvABA8'OH) and GA biosynthesis (HvHA20ox, HvGA3ox), and NADPH oxidase (HvRboh) genes were significantly upregulated after imbibition. Using MeDIP-qPCR, we showed that the promoters of HvNCED were hyper-methylated, and those of HvABA8'OH1, HvABA8'OH3, HvGA3ox2, and HvRbohF2 were hypo-methylated in heat treated seeds. Taken together, our data suggest that heat stress during grain filling affects DNA methylation of germination-related genes and promotes seed germination in barley.

Early Diastolic Left Ventricular Relaxation in Normal Neonates is Influenced by Ventricular Stiffness and Longitudinal Systolic Function.

Tissue Doppler velocity during early diastole (e') is one of the most feasible and reproducible echocardiographic assessments to reflect active relaxation of the left ventricle. Although several reports have described the mechanisms of temporal diastolic dysfunction in the early neonatal period, factors influencing diastolic function have not been determined. The purpose of this study was to elucidate factors significantly influencing e' in the early neonatal period.A total of 179 consecutive normal neonates underwent echocardiographic studies performed at 0 days and 5-10 days after birth. The statistical relationships between e' and age, body weight, mean blood pressure, heart rate, shortening fraction of the left ventricle, peak systolic motion velocity (s'), early diastolic transmitral flow velocity over annulus velocity, Tei index, and diastolic wall strain (DWS) were analyzed.Between the 0 days and 5-10-days-after birth groups, significant differences were shown in mean blood pressure, shortening fraction of left ventricle, e', and Tei index. Age, body weight, mean blood pressure, s', and DWS showed significant correlations with e'. In multivariate regression analysis within these parameters, s' (ß = 0.6119, P < 0.0001) and DWS (ß = 0.1216, P = 0.0321) showed positive correlations with e'.Longitudinal systolic motion velocity and ventricular wall stiffness of the left ventricle influence diastolic relaxation in normal neonates. Age, body weight, and circumferential systolic function are not significant factors.

Mitral annular plane systolic excursion/left ventricular length (MAPSE/L) as a simple index for assessing left ventricular longitudinal function in children.

BACKGROUND: Assessment of longitudinal left ventricular (LV) function is important for early detection of cardiac dysfunction. Although mitral annular plane systolic excursion (MAPSE) obtained by M-mode echocardiography offers a simple method for assessing longitudinal LV function, normal values of MAPSE for children change according to body size. METHODS: To minimize the effects of body size, MAPSE was divided by LV long-axis length (MAPSE/L). MAPSE/L was measured in 210 healthy children from birth to 15 years of age and classified into five subgroups. MAPSE/L was then compared with 10 parameters in 136 children (age, heart rate, mean blood pressure, ejection fraction of the LV (EF), peak atrial flow velocity/peak early diastolic flow velocity of mitral flow, tissue Doppler velocity during systole (s') and early diastole (e'), E/e' ratio, Tei index, and global longitudinal strain (GLS) of the LV by the speckle tracking method). RESULTS: MAPSE/L was significantly lower in the neonate group than in the remaining four groups. MAPSE/L then increased with age to peak at 1-5 years and gradually decreased thereafter. In all cases beyond the neonatal period, MAPSE/L was more than 0.17. Among various parameters, GLS, age, EF, Tei index and s' were significantly associated with MAPSE/L in that order. In univariate analysis, GLS was most significantly associated with MAPSE/L (r=.56). CONCLUSIONS: We have established normal reference values for MPSE/L in healthy children. MAPSE/L is expected to offer a simple parameter to evaluate LV longitudinal systolic function during daily routine echocardiography in children.

Mumps encephalitis with akinesia and mutism.

Measles-rubella-mumps vaccination is routine in many countries, but the mumps vaccine remains voluntary and is not covered by insurance in Japan. A 5-year-old Japanese boy who had not received the mumps vaccine was affected by mumps parotitis. Several days later, he presented with various neurological abnormalities, including akinesia, mutism, dysphagia, and uncontrolled respiratory disorder. Mumps encephalitis was diagnosed. Despite steroid pulse and immunoglobulin treatment, the disease progressed. Magnetic resonance imaging showed necrotic changes in bilateral basal ganglia, midbrain, and hypothalamus. At 1 year follow up, he was bedridden and required enteral feeding through a gastric fistula and tracheostomy. Mumps vaccination should be made routine as soon as possible in Japan, because mumps encephalitis carries the risk of severe sequelae.

[Symptoms of Cancer Patients and Kampo Formulas Effective for Them].

Patients with cancer exhibit various symptoms induced by cancer itself and its therapy leadingto fatigue; however, their vital energy can be restored by administration of Kampo, which is a traditional Japanese herbal medicine. Restoration and maintenance of mental and physical energy are important for successful cancer treatment. For this purpose, appropriate use of Kampo formulas, such as"Ho-zai", formulas to vitalize fatigued patients (eg, Hochu-ekki-to, Juzen-taiho-to, Ninjin-yoeito), "Hojin-zai", formulas to restore energy (eg, Gosha-jinki-gan), and"Kuoketsu-zai ", formulas to resolve stagnant blood flow (eg, Keishi-bukuryo-gan, Tokaku-joki-to, Toki-shakuyaku-san) are administered in combination. Consequently, basic autonomic functions, such as appetite, sleep, defecation, and urination normalize and the nutritional and mental conditions are restored. These favorable changes in the patients' condition allow completion of the standard cancer therapy course, resultingin an improved outcome of cancer therapy and successful treatment. Kampo therapy can be administered as the final treatment option for patients with last-stage cancer who do not have any other effective therapy options. If patients with cancer are administered Kampo formulas, their vital energy is restored, and they develop a will to fight the cancer. As a result, communication becomes easier.

Left atrial volume change throughout the cardiac cycle in children with congenital heart disease associated with increased pulmonary blood flow: evaluation using a novel left atrium-tracking method.

There is a paucity of data regarding the significance of left atrial (LA) volume and its changes throughout the cardiac cycle in pediatric patients with heart disease. The recently developed LA volume-tracking (LAVT) method can automatically construct the LA volume curve. The study group consisted of 48 pediatric patients with ventricular septal defect (n = 34) or patent ductus arteriosus (n = 14) and age-matched healthy controls. Maximum and minimum LA volumes (LAVmax and LAVmin, respectively) were measured. The total LA emptying volume (LAVtotal) was defined as LAVmax--LAVmin. Volume parameters were standardized by dividing by body surface area (BSA). The total LA emptying fraction (%LAVtotal) was defined as the ratio of LAVtotal to LAVmax. In the patient group, there was a positive correlation between the ratio of pulmonary to systemic blood flow (Qp/Qs) and LAVmax/BSA, LAVmin/BSA, and LAVtotal/BSA (r = 0.42, 0.44, and 0.34, respectively). LAVmin/BSA was positively correlated with the ratio of early mitral inflow velocity to early mitral annular diastolic tissue Doppler velocity (E/E') (r = 0.32). The %LAVtotal had a negative correlation with left-ventricular (LV) end-diastolic pressure (r = -0.32). There were significant correlations between serum B-type natriuretic peptide level and LAVmax/BSA, LAVmin/BSA, and %LAVtotal (r = 0.38, 0.49, and -0.35, respectively). The LAVT method is useful in the evaluation of LV diastolic function in pediatric patients with chronic LV volume overload.

Ratio of early diastolic tricuspid inflow to tricuspid lateral annulus velocity reflects pulmonary regurgitation severity but not right ventricular diastolic function in children with repaired Tetralogy of Fallot.

The current study assessed relationships between the ratio of early diastolic tricuspid inflow to tricuspid lateral annular velocity (tricuspid E/e') and right ventricular (RV) function in children after tetralogy of Fallot (TOF) repair. The RV function of 25 asymptomatic children with surgically repaired TOF (age 3.3 ± 2.0 years) was assessed by echocardiography and cardiac catheterization. Right ventricular end-diastolic pressure and volume (RVEDP and RVEDV), systolic pressure, and ejection fraction, as well as mean pulmonary arterial pressure, mean right atrial pressure (RAP), and the severity of both pulmonary regurgitation (PR) and tricuspid regurgitation (TR) were assessed in terms of the contribution to tricuspid E/e'. Univariate analysis discovered a relationship between tricuspid E/e' and RVEDV (R(2) = 0172), pressure half-time of PR (PR-PHT) (R(2) = 0.173), and TR grade (R(2) = 0.145) (p < 0.01 for each). After multivariate adjustment, PR-PHT was significantly associated with tricuspid E/e' (ß = 0.210; p < 0.001). Tricuspid E/e' was not significantly associated with RVEDP or RAP. In conclusion, tricuspid E/e' does not indicate RV diastolic function but reflects the severity of PR in asymptomatic children after TOF repair.

Subclavian and pulmonary artery steal phenomenon in a patient with isolated left subclavian artery and right aortic arch.

We describe a patient with an isolated left subclavian artery associated with right aortic arch, patent ductus arteriosus, and ventricular septal defect. As the isolated left subclavian artery is supplied by the left vertebral artery in which blood flows in the retrograde direction, this anomaly is usually responsible for a congenital subclavian steal phenomenon. Atrophy of the left cerebral hemisphere and inverted left vertebral arterial flow were clearly depicted by echoencephalography in this patient, whose subclavian artery was connected to the main pulmonary artery by a patent ductus arteriosus.

Time-dependent interaction between differentiated embryo chondrocyte-2 and CCAAT/enhancer-binding protein α underlies the circadian expression of CYP2D6 in serum-shocked HepG2 cells.

Differentiated embryo chondrocyte-2 (DEC2), also known as bHLHE41 or Sharp1, is a pleiotropic transcription repressor that controls the expression of genes involved in cellular differentiation, hypoxia responses, apoptosis, and circadian rhythm regulation. Although a previous study demonstrated that DEC2 participates in the circadian control of hepatic metabolism by regulating the expression of cytochrome P450, the molecular mechanism is not fully understood. We reported previously that brief exposure of HepG2 cells to 50% serum resulted in 24-h oscillation in the expression of CYP3A4 as well as circadian clock genes. In this study, we found that the expression of CYP2D6, a major drug-metabolizing enzyme in humans, also exhibited a significant oscillation in serum-shocked HepG2 cells. DEC2 interacted with CCAAT/enhancer-binding protein (C/EBPα), accompanied by formation of a complex with histone deacetylase-1, which suppressed the transcriptional activity of C/EBPα to induce the expression of CYP2D6. The oscillation in the protein levels of DEC2 in serum-shocked HepG2 cells was nearly antiphase to that in the mRNA levels of CYP2D6. Transfection of cells with small interfering RNA against DEC2 decreased the amplitude of CYP2D6 mRNA oscillation in serum-shocked cells. These results suggest that DEC2 periodically represses the promoter activity of CYP2D6, resulting in its circadian expression in serum-shocked cells. DEC2 seems to constitute a molecular link through which output components from the circadian clock are associated with the time-dependent expression of hepatic drug-metabolizing enzyme.

Tmsb10 triggers fetal Leydig differentiation by suppressing the RAS/ERK pathway.

Leydig cells in fetal testes play crucial roles in masculinizing fetuses through androgen production. Gene knockout studies have revealed that growth factors are implicated in fetal Leydig cell (FLC) differentiation, but little is known about the mechanisms regulating this process. We investigate this issue by characterizing FLC progenitor cells using single-cell RNA sequencing. The sequence datasets suggest that thymosin ß10 (Tmsb10) is transiently upregulated in the progenitors. While studying the function of Tmsb10, we reveal that platelet-derived growth factor (PDGF) regulates ciliogenesis through the RAS/ERK and PI3K/AKT pathways, and thereby promotes desert hedgehog (DHH)-dependent FLC differentiation. Tmsb10 expressed in the progenitor cells induces their differentiation into FLCs by suppressing the RAS/ERK pathway. Through characterizing the transiently expressed Tmsb10 in the FLC progenitors, this study unveils the molecular process of FLC differentiation and shows that it is cooperatively induced by DHH and PDGF.

Serum concentration of heart-type fatty acid-binding protein in children and adolescents with congenital heart disease.

BACKGROUND: Serum heart-type fatty acid-binding protein (H-FABP) is widely applied as a marker of cardiac myocyte injury. Recently, it has been reported that levels of H-FABP are elevated in adult patients with chronic heart failure and thus provide useful prognostic information. The aim of the present study was to examine the relationships between serum H-FABP levels and pathophysiological characteristics in children and adolescents with congenital heart disease (CHD). METHODS AND RESULTS: Serum H-FABP levels were preoperatively and postoperatively measured in 238 consecutive patients with CHD aged 1-31 years. The relationships between H-FABP levels and severity of heart failure, circulatory status and laboratory data were cross-sectionally analyzed. Multivariate regression analysis indicated that serum H-FABP levels are independently affected by age, New York Heart Association functional class, creatine kinase MB, creatinine and arterial oxygen saturation (standard regression coefficients, -0.378, 0.237, 0.422, 0.615, and -0.210, respectively). Neither left ventricular ejection fraction nor B-type natriuretic peptide correlated with H-FABP levels. CONCLUSIONS: H-FABP could serve as a new monitoring tool to provide information that will guide the optimal therapy and management of CHD patients.

Cell membrane stretch activates intermediate-conductance Ca2+-activated K+ channels in arterial smooth muscle cells.

The aim of this study is to determine the signal transduction of membrane stretch on intermediate-conductance Ca(2+)-activated K(+) (IKca) channels in rat aorta smooth muscle cells using the patch-clamp technique. To stretch the cell membrane, both suction to the rear end of patch pipette and hypotonic shock were used. In cell-attached and inside-out patch configurations, the open probability of IKca channels increased when 20- to 45-mmHg suction was applied. Hyposmotic swelling efficiently increased IKca channel current. When the Ca(2+)-free solution was superfused, the activation of IKca current by the hyposmotic swelling was reduced. Furthermore, gadolinium (Gd(3+)) attenuated the activation of IKca channels induced by hyposmotic swelling, whereas nicardipine did not. In the experiments with Ca(2+)-free bath solution, pretreatment with GF109203X, a protein kinase C (PKC) inhibitor, completely abolished the stretch-induced activation of IKca currents. The stretch-induced activation of IKca channels was strongly inhibited by cytochalasin D, indicating a role for the F-actin in modulation of IKca channels by changes in cell stretching. These data suggest that cell membrane stretch activates IKca channels. In addition, the activation is associated with extracellular Ca(2+) influx through stretch-activated nonselective cation channels, and is also modulated by the F-actin cytoskeleton and the activation of PKC.

Sex differences in metabolic pathways are regulated by Pfkfb3 and Pdk4 expression in rodent muscle.

Skeletal muscles display sexually dimorphic features. Biochemically, glycolysis and fatty acid ß-oxidation occur preferentially in the muscles of males and females, respectively. However, the mechanisms of the selective utilization of these fuels remains elusive. Here, we obtain transcriptomes from quadriceps type IIB fibers of untreated, gonadectomized, and sex steroid-treated mice of both sexes. Analyses of the transcriptomes unveil two genes, Pfkfb3 (phosphofructokinase-2) and Pdk4 (pyruvate dehydrogenase kinase 4), that may function as switches between the two sexually dimorphic metabolic pathways. Interestingly, Pfkfb3 and Pdk4 show male-enriched and estradiol-enhanced expression, respectively. Moreover, the contribution of these genes to sexually dimorphic metabolism is demonstrated by knockdown studies with cultured type IIB muscle fibers. Considering that skeletal muscles as a whole are the largest energy-consuming organs, our results provide insights into energy metabolism in the two sexes, during the estrus cycle in women, and under pathological conditions involving skeletal muscles.

Gene expression and functional abnormalities in XX/Sry Leydig cells.

The SRY gene induces testis development even in XX individuals. However, XX/Sry testes fail to produce mature sperm, due to the absence of Y chromosome carrying genes essential for spermatogenesis. XX/Sry Sertoli cells show abnormalities in the production of lactate and cholesterol required for germ cell development. Leydig cells are essential for male functions through testosterone production. However, whether XX/Sry adult Leydig cells (XX/Sry ALCs) function normally remains unclear. In this study, the transcriptomes from XY and XX/Sry ALCs demonstrated that immediate early and cholesterogenic gene expressions differed between these cells. Interestingly, cholesterogenic genes were upregulated in XX/Sry ALCs, although downregulated in XX/Sry Sertoli cells. Among the steroidogenic enzymes, CYP17A1 mediates steroid 17α-hydroxylation and 17,20-lyase reaction, necessary for testosterone production. In XX/Sry ALCs, the latter reaction was selectively decreased. The defects in XX/Sry ALCs, together with those in the germ and Sertoli cells, might explain the infertility of XX/Sry testes.

Generation of ovarian follicles from mouse pluripotent stem cells.

Oocytes mature in a specialized fluid-filled sac, the ovarian follicle, which provides signals needed for meiosis and germ cell growth. Methods have been developed to generate functional oocytes from pluripotent stem cell-derived primordial germ cell-like cells (PGCLCs) when placed in culture with embryonic ovarian somatic cells. In this study, we developed culture conditions to recreate the stepwise differentiation process from pluripotent cells to fetal ovarian somatic cell-like cells (FOSLCs). When FOSLCs were aggregated with PGCLCs derived from mouse embryonic stem cells, the PGCLCs entered meiosis to generate functional oocytes capable of fertilization and development to live offspring. Generating functional mouse oocytes in a reconstituted ovarian environment provides a method for in vitro oocyte production and follicle generation for a better understanding of mammalian reproduction.

A mouse line expressing Sall1-driven inducible Cre recombinase in the kidney mesenchyme.

Sall1 is expressed in the metanephric mesenchyme in the developing kidney, and mice deficient in Sall1 show kidney agenesis or dysgenesis. Sall1 is also expressed elsewhere, including in the limb buds, anus, heart, and central nervous system. Dominant-negative mutations of Sall1 in mice and humans lead to developmental defects in these organs. Here, we generated a mouse line expressing tamoxifen-inducible Cre recombinase (CreER(T2)) under the control of the endogenous Sall1 promoter. Upon tamoxifen treatment, these mice showed genomic recombination in the tissues where endogenous Sall1 is expressed. When CreER(T2) mice were crossed with the floxed Sall1 allele, tamoxifen administration during gestation led to a significant decrease in Sall1 expression and small kidneys at birth, suggesting that Sall1 functions were disrupted. Furthermore, Sall1 expression in the kidney was significantly reduced by neonatal tamoxifen treatment. The Sall1CreER(T2) mouse is a valuable tool for in vivo time-dependent and region-specific knockout and overexpression studies.

Assessment of modified Blalock-Taussig shunt in children with congenital heart disease using multidetector-row computed tomography.

The purpose of this study was to assess the feasibility of multidetector-row computed tomography (MDCT) for the evaluation of modified Blalock-Taussig (B-T) shunt in children with congenital heart disease associated with reduced pulmonary blood flow. A total of 25 consecutive patients (mean age, 2.6 ± 3.6 years; range, 2 months-16 years) underwent MDCT angiography of the thorax with a 16-detector row scanner prior to cardiac catheterization. A total of 39 shunts (right, 22; left, 17) were included in the study. Conventional angiographic findings were used as the gold standard for the detection of B-T shunts. Shunt diameter was measured quantitatively and independently at four sites (the subclavian artery site, the pulmonary artery site, the widest site, and the stenotic site) on MDCT and on conventional invasive angiography. All B-T shunts were depicted on multiplanar reconstruction (MPR), maximum intensity projection (MIP), curved planar reconstruction (CPR), and three-dimensional volume-rendered (VR) images, enabling evaluation in all patients except for one with occluded shunt. There were excellent correlations between MDCT- and conventional angiography-based measurements of shunt diameter at the subclavian artery site, pulmonary artery site, and the widest site (R² = 0.46, 0.74 and 0.64, respectively; p < 0.0001 for each), although systematic overestimation was observed for MDCT (mean percentage of overestimation, 23.1 ± 32.4%). Stenotic site diameter and degree of stenosis showed a mild correlation (R² = 010 and 0.25, respectively; p < 0.01 for each). This study demonstrates that MDCT is a promising tool for the detection of lesions in B-T shunts.

Development of systemic-to-pulmonary collateral arteries in a patient with hypoplastic left cardiac syndrome after bilateral pulmonary artery banding.

We describe systemic-to-pulmonary collateral arteries that developed after bilateral pulmonary artery banding in a patient with hypoplastic left cardiac syndrome. The growth of collateral arteries should be evaluated carefully because bilateral pulmonary artery banding under prostaglandin E1 administration is considered an initial palliative option.

Coordination of Multiple Cellular Processes by NR5A1/Nr5a1.

The agenesis of the gonads and adrenal gland in revealed by knockout mouse studies strongly suggested a crucial role for Nr5a1 (SF-1 or Ad4BP) in organ development. In relation to these striking phenotypes, NR5A1/Nr5a1 has the potential to reprogram cells to steroidogenic cells, endow pluripotency, and regulate cell proliferation. However, due to limited knowledge regarding NR5A1 target genes, the mechanism by which NR5A1/Nr5a1 regulates these fundamental processes has remained unknown. Recently, newlyestablished technologies have enabled the identification of NR5A1 target genes related to multiple metabolic processes, as well as the aforementioned biological processes. Considering that active cellular processes are expected to be accompanied by active metabolism, NR5A1 may act as a key factor for processes such as cell differentiation, proliferation, and survival by coordinating these processes with cellular metabolism. A complete and definite picture of the cellular processes coordinated by NR5A1/Nr5a1 could be depicted by accumulating evidence of the potential target genes through whole genome studies.

Drug-induced lung disease adverse effect with Ledipasvir Acetonate/Sofosbuvir.

BACKGROUND: Interferon and ribavirin have been used as therapeutic agents for chronic hepatitis C infection or C-compensated cirrhosis in the conventional treatment. Hepatitis C virus (HCV) -specific direct-acting antiviral agents that directly inhibit the growth process of HCV have been approved since 2011. However, in the early post-marketing vigilance phase of ledipasvir acetonate/sofosbuvir (LDV/SOF), there were reports of interstitial lung disease in 4 out of 32,700 cases with death in 1 case; the onset mechanism is unknown. CASE PRESENTATION: Treatment for hepatitis C was deemed to be necessary, and the patient was referred to our hospital. Oral administration of LDV/SOF was started. On day 8 of administration, a fever of 38-39 °C and coughing were observed followed by the gradual appearance of shortness of breath. As there was no improvement, the patient visited her primary care physician on day 16 of administration and the patient was brought urgently to our hospital on the same day. Blood tests and imaging tests were conducted at our hospital on the day of emergency transport; inflammatory response markers showed abnormal values, and sialylated carbohydrate antigen Krebs von den Lungen-6 was within the normal value range at 303 U/mL. Because the possibility of infection was low based on results of imaging and bronchoalveolar lavage, drug-induced lung disease was suspected, LDV/SOF administration was discontinued, and steroid administration was started. Following steroid pulse therapy, treatment with oral prednisolone tablets was gradually tapered. The patient's symptoms were relieved and she was discharged. CONCLUSIONS: The patient's medication history in this case indicated that there were no drugs taken before or after administration of LDV/SOF until the adverse reaction occurred, and there were no supplements or dietary supplements taken. Therefore, LDV/SOF has been proposed as the cause of the suspected adverse effect. Pharmacists should try to collect adverse effect reports to identify adverse effects early.