Late-maturing cooking rice Sensyuraku has excellent properties,equivalent to sake rice,for high-quality sake brewing

Low protein content and sufficient grain rigidity are desired properties for the rice used in high-quality sake brewing such as Daiginjo-shu (polishing ratio of the rice, less than 50%). Two kinds of rice, sake rice (SR) and cooking rice (CR), have been used for sake brewing. Compared with those of SR, analyses of CR for high-quality sake brewing using highly polished rice have been limited. Here we described the original screening of late-maturing CR Sensyuraku (SEN) as rice with low protein content and characterization of its properties for high-quality sake brewing. The protein content of SEN was lower than those of SR Gohyakumangoku (GOM) and CR Yukinosei (YUK), and its grain rigidity was higher than that of GOM. The excellent properties of SEN with respect to both water-adsorption and enzyme digestibility were confirmed using a Rapid Visco Analyzer (RVA). Further, we confirmed a clear taste of sake produced from SEN by sensory evaluation. Thus, SEN has excellent properties, equivalent to those of SR, for high-quality sake brewing.     

Improvement of the post-thaw qualities of Okinawan native Agu pig sperm frozen in an extender supplemented with antiapoptotic PTD-FNK protein

The technical establishment of boar sperm cryopreservation is indispensable for effective breeding of the scarce Okinawan native Agu pig. The objective was to determine whether an artificial anticell death protein (PTD-FNK protein) was capable of improving the quality of cryopreserved Agu sperm. Ejaculated Agu sperm frozen in an extender supplemented with 0, 100, 200, 300, or 400 nm PTD-FNK protein was thawed, and mitochondrial integrity and other sperm characteristics were evaluated. Treatment with 300 nm PTD-FNK protein had the most beneficial effect (P < 0.05) on mitochondrial integrity (45-59%) and sperm motility (56-67%) after freezing-thawing. In particular, the proportion of post-thaw sperm with activated caspase-9 and -3 but not caspase-8 was markedly reduced among sperm frozen in the presence of PTD-FNK protein (P < 0.05), implying protection against apoptotic-cell death in response to mitochondrial damage. There were high levels of intracellular ATP (9.4-10.5 nmol/10⁸ sperm) in post-thaw sperm treated with PTD-FNK protein, and the inhibitory effect of PTD-FNK protein on activation of caspases influenced the increase in the number of sperm with intact DNA (36-53%; P < 0.05). Furthermore, the addition of PTD-FNK protein to the freezing extender strongly preserved the ability of the sperm to penetrate to mature oocytes in all individuals (60-80%; P < 0.05). In conclusion, treatment with PTD-FNK protein in the freezing extender effectively improved post-thaw qualities of fragile Agu sperm through prevention of mitochondrial dysfunction leading to apoptotic-cell death during cryopreservation.     

Direct evidence for non-enzymatic fragmentation of chloroplastic glutamine synthetase by a reactive oxygen species

Chloroplastic glutamine synthetase (GS: EC 6·3·1·2), the octamer of the 44 kDa subunit, is rapidly degraded under photo‐oxidative stress conditions in leaves, chloroplasts, and chloroplast lysates. Recent studies have suggested that chloroplastic GS might be cleaved by the hydroxyl radical under such conditions ( Thoenen & Feller 1998 ; Australian Journal of Plant Physiology 25, 279–286; Palatnik, Carrillo & Valle 1999 , Plant Physiology 121, 471–478). Herein, we present evidence which supports the above hypothesis. When the purified GS from wheat (Triticum aestivum L.) chloroplasts was exposed to the hydroxyl radical‐generating system comprising H₂O₂–FeSO₄–ascorbic acid or FeCl₃–ascorbic acid, the GS subunit was degraded into four distinct fragments having apparent molecular masses of 39, 35, 32 and 28 kDa. The apparent molecular masses and isoelectric points of these fragments were identical to those of the respective fragments found in the illuminated lysates of chloroplasts. In addition, the appearance of the GS fragments was completely suppressed in the presence of the scavenger for the hydroxyl radical, n‐propyl gallate, in the illuminated lysates of chloroplasts. These results strongly support the hypothesis that the primary cleavage of GS is directly driven by the hydroxyl radical, formed by Fenton reaction under photo‐oxidative stress conditions in chloroplasts.     

Tiny keratotic brown lesions on the interdigital web between the toes of a healthy man caused by Curvularia species infection and a review of cutaneous Curvularia infections

A 74-year-old healthy Japanese man had noticed an itchy lesion with two tiny thick brown scales on the fourth interdigital web of his right foot 3 months earlier. The brown fungus isolated from the scales had demonstrated brown, ellipsoidal, obovoid or clavate, 3-septate conidia. Morphologically, the fungus was identified as Curvularia sp., and was differentiated from other human pathogenic species of the genus, such as C. lunata, C. pallescens, C. trifolii, C. clavata, and C. brachyspora, by molecular analysis based on the DNA sequence data. The fungus grows well on keratotic materials (hairs, nails, and callus), which indicates that it might have the ability to infect the skin surface.     

Three-dimensional reconstruction of the human capillary network and the intramyocardial micronecrosis

Three-dimensional reconstruction of the human heart was performed to define the structure of the intramyocardial microvasculature. A total of 200 consecutive serial sections of 6 μm each were prepared from the left ventricular tissue of an autopsied human heart with normal coronary arteries. The corresponding arteriole, venule, and all capillaries were reconstructed using three-dimensional software. The capillary network extended right and left along the cardiomyocyte with major and minor axes of about 130 and 120 μm, respectively. The capillary length from an arteriole to an adjacent venule was about 350 μm. Two types of sack-like structures, the precapillary sinus and the capillary sinus, were present in the capillary network, and many capillaries diverged from these sinuses. The cardiomyocytes were covered with reticular capillaries. In contrast, the precapillary and capillary sinuses were surrounded by many cardiomyocytes. The arterial and venous capillaries were positioned alternately, forming a lattice pattern. Intramyocardial microcirculatory units forming a capillary network from an arteriole to adjacent venules on both sides were present. The sizes of myocardial micronecroses corresponded to that of the intramyocardial microcirculatory unit. These results show that the capillary network is an ordered and anatomically regulated structure and that the microcirculatory unit and the precapillary and capillary sinuses may play an important role in maintaining the intramyocardial microcirculation during contraction and relaxation.     

Spatial localizations of Mam22 and Mam12 in the magnetosomes of Magnetospirillum magnetotacticum

Magnetospirillum magnetotacticum possesses intracellular magnetite particles with a chain-like structure, termed magnetosomes. The bacterium expresses 22-kDa and 12-kDa magnetosome-associated proteins, termed Mam22 (MamA) and Mam12 (MamC), respectively. In this study, we investigated the structure of the purified magnetosomes with transmission electron microscopic techniques and found that the magnetosomes consisted of four compartments, i.e., magnetite crystal, magnetosomal membrane, interparticle connection, and magnetosomal matrix. Furthermore, we determined the precise localizations of Mam22 and Mam12 using immunogold staining of the purified magnetosomes and ultrathin sections of the bacterial cells. Interestingly, most Mam22 existed in the magnetosomal matrix, whereas Mam12 was strictly localized in the magnetosomal membrane. Moreover, the recombinant Mam22 was attached to the magnetosomal matrix of the Mam22-deficient magnetosomes prepared by alkaline treatment, such as 0.1 M Caps-NaOH buffer (pH 11.0). The spatial localization of the magnetosome-associated proteins in the magnetosomal chain provides useful information to elucidate the functional roles of these proteins.     

Expression of SGLT1 in Human Hearts and Impairment of Cardiac Glucose Uptake by Phlorizin during Ischemia-Reperfusion Injury in Mice

Objective

Sodium-glucose cotransporter 1 (SGLT1) is thought to be expressed in the heart as the dominant isoform of cardiac SGLT, although more information is required to delineate the subtypes of SGLTs in human hearts. Moreover, the functional role of SGLTs in the heart remains to be fully elucidated. We herein investigated whether SGLT1 is expressed in human hearts and whether SGLTs significantly contribute to cardiac energy metabolism during ischemia-reperfusion injury (IRI) via enhanced glucose utilization in mice.

Methods and Results

We determined that SGLT1 was highly expressed in both human autopsied hearts and murine perfused hearts, as assessed by immunostaining and immunoblotting with membrane fractionation. To test the functional significance of the substantial expression of SGLTs in the heart, we studied the effects of a non-selective SGLT inhibitor, phlorizin, on the baseline cardiac function and its response to ischemia-reperfusion using the murine Langendorff model. Although phlorizin perfusion did not affect baseline cardiac function, its administration during IRI significantly impaired the recovery in left ventricular contractions and rate pressure product, associated with an increased infarct size, as demonstrated by triphenyltetrazolium chloride staining and creatine phosphokinase activity released into the perfusate. The onset of ischemic contracture, which indicates the initiation of ATP depletion in myocardium, was earlier with phlorizin. Consistent with this finding, there was a significant decrease in the tissue ATP content associated with reductions in glucose uptake, as well as lactate output (indicating glycolytic flux), during ischemia-reperfusion in the phlorizin-perfused hearts.

Conclusions

Cardiac SGLTs, possibly SGLT1 in particular, appear to provide an important protective mechanism against IRI by replenishing ATP stores in ischemic cardiac tissues via enhancing availability of glucose. The present findings provide new insight into the significant role of SGLTs in optimizing cardiac energy metabolism, at least during the acute phase of IRI.     

Genome-wide screening for genes associated with FK506 sensitivity in fission yeast

We have been studying calcineurin signal transduction pathway in fission yeast Schizosaccharomyces pombe (S. pombe) by developing a genetic screen for mutants that show hypersensitivity to the immunosuppressive calcineurin inhibitor FK506 (tacrolimus). In the present study, to identify nonessential genes that are functionally related to the calcineurin signaling pathway, we performed a genome-wide screen of 3004 haploid deletion strains and confirmed 72 deletion strains to be FK506 sensitive. These 72 genes are classified into nine functional groups to include membrane trafficking (16 genes), signal transduction (10 genes), ubiquitination (8 genes), chromatin remodeling (6 genes), cytokinesis (4 genes), ribosomal protein (3 genes), RNA binding protein (3 genes), and a variety of other known functions (17 genes) or still unknown functions (5 genes) in the biological system. In our previous screening of FK506-sensitive mutants we isolated several membrane-trafficking mutants showing defective cell wall integrity. Here, we further examined the vacuolar fusion, the v-SNARE synaptobrevin Syb1 localization, and the sensitivity to the β-glucan synthase inhibitor micafungin in these 72 FK506-sensitive strains. Results showed that 25 deletion strains exhibited abnormal vacuole fusion, 19 deletion strains exhibited Syb1 mislocalization, and 14 deletion strains exhibited both abnormal vacuole fusion and Syb1 mislocalization, while 42 deletion strains showed both normal vacuole fusion and Syb1 localization. Likewise, 16 deletion strains showed sensitivity to micafungin. Altogether, our present study indicates that calcineurin mediates a plethora of physiological processes in fission yeast, and that calcineurin is extensively involved in cross-talk between signaling pathways.     

Quantification of Dermatophyte Viability for Evaluation of Antifungal Effect by Quantitative PCR

Dermatophytosis is a common disease caused by dermatophyte fungi such as Trichophyton rubrum and Trichophyton mentagrophytes. A method of quantifying fungal viability in the lesions of dermatophytosis is indispensable for understanding the therapeutic process and outcome; however, no such method has yet been developed. The aim of this study was to develop a method for quantifying dermatophyte viability by quantitative polymerase chain reaction (qPCR). The internal transcribed spacer (ITS) and D1/D2 regions, including each of rRNA and rDNA, were chosen as the targets, and dermatophyte-specific primer pairs were designed corresponding to ITS and D1/D2 regions. The amounts of target RNA and DNA after heat or antifungal treatment were measured by qPCR and compared with colony-forming unit (CFU) counts. RNA and DNA could extract from dermatophytes by mechanical pulverization of conidia using a Multi-Beads Shocker cell disruptor. Our method was sufficiently sensitive to detect 10 copies by qPCR using both ITS and D1/D2 primer pairs. The most sensitive target was ITS-cDNA after heat or antifungal treatment, and essentially consistent with CFU counts. On the other hands, ITS-DNA and D1/D2-DNA were not decreased soon after heat or antifungal treatment, but those were decreased significantly and reflected the CFU counts after 48 h of antifungal treatment. We conclude that ITS-cDNA is useful mainly for quantifying dermatophyte viability at early responses, but ITS-DNA and D1/D2-DNA are also available for evaluation, which does not need an early response.