Monolayered mesenchymal stem cells repair scarred myocardium after myocardial infarction

Mesenchymal stem cells are multipotent cells that can differentiate into cardiomyocytes and vascular endothelial cells. Here we show, using cell sheet technology, that monolayered mesenchymal stem cells have multipotent and self-propagating properties after transplantation into infarcted rat hearts. We cultured adipose tissue-derived mesenchymal stem cells characterized by flow cytometry using temperature-responsive culture dishes. Four weeks after coronary ligation, we transplanted the monolayered mesenchymal stem cells onto the scarred myocardium. After transplantation, the engrafted sheet gradually grew to form a thick stratum that included newly formed vessels, undifferentiated cells and few cardiomyocytes. The mesenchymal stem cell sheet also acted through paracrine pathways to trigger angiogenesis. Unlike a fibroblast cell sheet, the monolayered mesenchymal stem cells reversed wall thinning in the scar area and improved cardiac function in rats with myocardial infarction. Thus, transplantation of monolayered mesenchymal stem cells may be a new therapeutic strategy for cardiac tissue regeneration.     

Hepatoprotective effects of purple potato extract against D-galactosamine-induced liver injury in rats

We investigated the hepatoprotective effect of purple potato extract (PPE) against D-galactosamine (GalN)-induced liver injury in rats. PPE (400 mg) was administered once daily for 8 d, and then GalN (250 mg/kg of body weight) was injected at 22 h before the rats were killed. Serum tumor necrosis factor alpha (TNF-alpha), lactate dehydrogenase (LDH), alanine aminotransferase (ALT), and asparate aminotranferase (AST) levels increased significantly after injection of GalN, but PPE inhibited GalN-induced alterations in serum TNF-alpha, LDH, ALT, and AST levels. Hepatic lipid peroxide and glutathione levels in the control + GalN group were higher and lower respectively than those in the control group, and those in the PPE + GalN group did not differ from that in the control group. The lipid peroxide level in hepatic microsomes treated with 2,2'-azobis (2-amidinopropane) dihydrochloride in the PPE group was significantly lower than that in the control group. This suggests that PPE has hepatoprotective effects against GalN-induced hepatotoxicity via inhibition lipid peroxidation and/or inflammation in rats.     

Dikaryotic arthroconidiation of Pleurotus subgenus Coremiopleurotus

This study was carried out to elucidate the poorly understood processes of arthroconidiation through coremium formation using Pleurotus cystidiosus subsp. abalonus. The coremia exclusively produced dikaryotic arthroconidia with the remnant of a clamp connection. Cells in the subapical zone of the hyphal bundle reduced their length by division before arthroconidiation. Approximately 400 000 arthroconidia were produced by a coremium in 1 day, with constant productivity over a 2-week period. Continuous cell extension and division in the coremium stipe supplied cells for arthroconidiation at the coremium apex, which is surrounded by a liquid droplet (coremioliquid). Maintenance of moisture with coremioliquid was necessary for arthroconidiation. The coremioliquid formation was performed by active liquid transportation in the hyphae, a process that was blocked by the microtubule depolymerization agent thiabendazole.     

Production-dependent reproductive allocation of a tall tree speciesQuercus serrata

The reproductive allocation (allocation of net production to acorns) of a tall tree speciesQuercus serrata was estimated by a method combining branch diameter distribution and the sampling of acorns per branch. Acorn production per branch of 0.5-cm diameter significantly varied among individuals and years, but not significantly across tree size. Leaf production per branch of 0.5-cm diameter had little annual fluctuation, while that of acorns fluctuated about 6 times between the maximum and minimum. The reproductive allocation was regressed against vegetative allocation, following the general model of klinkhameret al. (1992); the reproduction had a beginning threshold and reproductive allocation increased drastically at relatively young stage after the tree began reproduction. With its small threshold production, coppicing traits and large maximum size,Q. serrata seemed adaptive to infrequent but large-scale disturbances like fires.     

Isolation of B subunit‐specific monoclonal antibody clones that strongly neutralize the toxicity of Shiga toxin 2

Shiga toxin 2 (Stx2)‐specific mAb‐producing hybridoma clones were generated from mice. Because mice tend to produce small amounts of B subunit (Stx2B)‐specific antibodies at the polyclonal antibody level after immunization via the parenteral route, mice were immunized intranasally with Stx2 toxoids with a mutant heat‐labile enterotoxin as a mucosal adjuvant; 11 different hybridoma clones were obtained in two trials. Six of them were A subunit (Stx2A)‐specific whereas five were Stx2B‐specific antibody‐producing clones. The in vitro neutralization activity of Stx2B‐specific mAbs against Stx2 was greater than that of Stx2A‐specific mAbs on HeLa229 cells. Furthermore, even at low concentrations two of the Stx2B‐specific mAbs (45 and 75D9) completely inhibited receptor binding and showed in vivo neutralization activity against a fivefold median lethal dose of Stx2 in mice. In western blot analysis, these Stx2B‐specific neutralization antibodies did not react to three different mutant forms of Stx2, each amino acid residue of which was associated with receptor binding. Additionally, the nucleotide sequences of the V_H and V_L regions of clones 45 and 75D9 were determined. Our Stx2B‐specific mAbs may be new candidates for the development of mouse‐human chimeric Stx2‐neutralizing antibodies which have fewer adverse effects than animal antibodies for enterohemorrhagic Escherichia coli infection.     

4-Acylamino-6-arylfuro[2,3-d]pyrimidines: potent and selective glycogen synthase kinase-3 inhibitors

Modeling studies of a furo[2,3-d]pyrimidine GSK-3 hit compound 1 superimposed onto the X-ray crystal structure of a legacy pyrazolo[3,4-c]pyridazine GSK-3 inhibitor 2 led to the identification of 4-acylamino-6-arylfuro[2,3-d]pyrimidine template 3. Synthesis of analogues based on template 3 has resulted in a number of potent and selective GSK-3beta inhibitors. The most potent and selective compound was the m-pyridyl analogue 24.     

Activation of protein phosphatase 2A by cAMP-dependent protein kinase-catalyzed phosphorylation of the 74-kDa B″ (δ) regulatory subunit in vitro and identification of the phosphorylation sites

Human erythrocyte protein phosphatase 2A, which comprises a 34-kDa catalytic C subunit, a 63-kDa regulatory A subunit and a 74-kDa regulatory B″ (δ) subunit, was phosphorylated at serine residues of B″ in vitro by cAMP-dependent protein kinase (A-kinase). In the presence and absence of 0.5 μM okadaic acid (OA), A-kinase gave maximal incorporation of 1.7 and 1.0 mol of phosphate per mol of B″, respectively. The K_m value of A-kinase for CAB″ was 0.17±0.01 μM in the presence of OA. The major in vitro phosphorylation sites of B″ were identified as Ser-60, -75 and -573 in the presence of OA, and Ser-75 and -573 in the absence of OA. Phosphorylation of B″ did not dissociate B″ from CA, and stimulated the molecular activity of CAB″ toward phosphorylated H1 and H2B histones, 3.8- and 1.4-fold, respectively, but not toward phosphorylase a.     

Origin of higher affinity to RNA of the N-terminal RNA-binding domain than that of the C-terminal one of a mouse neural protein,musashi1, as revealed by comparison of their structures,modes of interaction,surface electrostatic potentials,and backbone dynamics

Musashi1 is an RNA-binding protein abundantly expressed in the developing mouse central nervous system. Its restricted expression in neural precursor cells suggests that it is involved in maintenance of the character of progenitor cells. Musashi1 contains two ribonucleoprotein-type RNA-binding domains (RBDs), RBD1 and RBD2, the affinity to RNA of RBD1 being much higher than that of RBD2. We previously reported the structure and mode of interaction with RNA of RBD2. Here, we have determined the structure and mode of interaction with RNA of RBD1. We have also analyzed the surface electrostatic potential and backbone dynamics of both RBDs. The two RBDs exhibit the same ribo-nucleoprotein-type fold and commonly make contact with RNA on the beta-sheet side. On the other hand, there is a remarkable difference in surface electrostatic potential, the beta-sheet of RBD1 being positively charged, which is favorable for binding negatively charged RNA, but that of RBD2 being almost neutral. There is also a difference in backbone dynamics, the central portion of the beta-sheet of RBD1 being flexible, but that of RBD2 not being flexible. The flexibility of RBD1 may be utilized in the recognition process to facilitate an induced fit. Thus, comparative studies have revealed the origin of the higher affinity of RBD1 than that of RBD2 and indicated that the affinity of an RBD to RNA is not governed by its fold alone but is also determined by its surface electrostatic potential and/or backbone dynamics. The biological role of RBD2 with lower affinity is also discussed.