Decreased levels of free d-aspartic acid in the forebrain of serine racemase (Srr) knock-out mice

d-Serine, an endogenous co-agonist of the N-methyl-d-aspartate (NMDA) receptor is synthesized from l-serine by serine racemase (SRR). A previous study of Srr knockout (Srr-KO) mice showed that levels of d-serine in forebrain regions, such as frontal cortex, hippocampus, and striatum, but not cerebellum, of mutant mice are significantly lower than those of wild-type (WT) mice, suggesting that SRR is responsible for d-serine production in the forebrain. In this study, we attempted to determine whether SRR affects the level of other amino acids in brain tissue. We found that tissue levels of d-aspartic acid in the forebrains (frontal cortex, hippocampus and striatum) of Srr-KO mice were significantly lower than in WT mice, whereas levels of d-aspartic acid in the cerebellum were not altered. Levels of d-alanine, l-alanine, l-aspartic acid, taurine, asparagine, arginine, threonine, γ-amino butyric acid (GABA) and methionine, remained the same in frontal cortex, hippocampus, striatum and cerebellum of WT and mutant mice. Furthermore, no differences in d-aspartate oxidase (DDO) activity were detected in the forebrains of WT and Srr-KO mice. These results suggest that SRR and/or d-serine may be involved in the production of d-aspartic acid in mouse forebrains, although further detailed studies will be necessary to confirm this finding.     

Novel scaffold for cathepsin K inhibitors

Pyrrolopyrimidine, a novel scaffold, allows to adjust interactions within the S3 subsite of cathepsin K. The core intermediate 10 facilitated the P3 optimization and identified highly potent and selective cathepsin K inhibitors 11-20.     

Analysis of exposed cellulose surfaces in pretreated wood biomass using carbohydrate‐binding module (CBM)–cyan fluorescent protein (CFP)

In enzymatic saccharification of lignocellulosics, the access of the enzymes to exposed cellulose surfaces is a key initial step in triggering hydrolysis. However, knowledge of the structure–hydrolyzability relationship of the pretreated biomass is still limited. Here we used fluorescent‐labeled recombinant carbohydrate‐binding modules (CBMs) from Clostridium josui as specific markers for crystalline cellulose (CjCBM3) and non‐crystalline cellulose (CjCBM28) to analyze the complex surfaces of wood tissues pretreated with NaOH, NaOH–Na₂S (kraft pulping), hydrothermolysis, ball‐milling, and organosolvolysis. Japanese cedar wood, one of the most recalcitrant softwood species was selected for the analysis. The binding analysis clarified the linear dependency of the exposure of crystalline and non‐crystalline cellulose surfaces for enzymatic saccharification yield by the organosolv and kraft delignification processes. Ball‐milling for 5–30 min increased saccharification yield up to 77%, but adsorption by the CjCBM–cyan fluorescent proteins (CFPs) was below 5%. Adsorption of CjCBM–CFPs on the hydrothermolysis pulp were less than half of those for organosolvolysis pulp, in coincidence with low saccharification yields. For all the pretreated wood, crystallinity index was not directly correlated with the overall saccharification yield. Fluorescent microscopy revealed that CjCBM3–CFP and CjCBM28–CFP were site‐specifically adsorbed on external fibrous structures and ruptured or distorted fiber surfaces. The assay system with CBM–CFPs is a powerful measure to estimate the initiation sites of hydrolysis and saccharification yields from chemically delignified wood pulps. Biotechnol. Bioeng. 2010; 105: 499–508. © 2009 Wiley Periodicals, Inc.     

Six family genes control the proliferation and differentiation of muscle satellite cells

Muscle satellite cells are essential for muscle growth and regeneration and their morphology, behavior and gene expression have been extensively studied. However, the mechanisms involved in their proliferation and differentiation remain elusive. Six1 and Six4 proteins were expressed in the nuclei of myofibers of adult mice and the numbers of myoblasts positive for Six1 and Six4 increased during regeneration of skeletal muscles. Six1 and Six4 were expressed in quiescent, activated and differentiated muscle satellite cells isolated from adult skeletal muscle. Overexpression of Six4 and Six5 repressed the proliferation and differentiation of satellite cells. Conversely, knockdown of Six5 resulted in augmented proliferation, and that of Six4 inhibited differentiation. Muscle satellite cells isolated from Six4^+/−Six5^−/− mice proliferated to higher cell density though their differentiation was not altered. Meanwhile, overproduction of Six1 repressed proliferation and promoted differentiation of satellite cells. In addition, Six4 and Six5 repressed, while Six1 activated myogenin expression, suggesting that the differential regulation of myogenin expression is responsible for the differential effects of Six genes. The results indicated the involvement of Six genes in the behavior of satellite cells and identified Six genes as potential target for manipulation of proliferation and differentiation of muscle satellite cells for therapeutic applications.     

Benzo[a]pyrene exposed to solar-simulated light inhibits apoptosis and augments carcinogenicity

Polycyclic aromatic hydrocarbons (PAHs) such as benzo[a]pyrene (BaP) are widespread environmental pollutants and several lines of experimental evidence have suggested a role in carcinogenesis. PAHs in the environment are exposed to sunlight and photomodified PAHs have been detected in contaminated sediment and air particulate matter; however, the carcinogenicity of photomodified PAHs is not well understood. In this study, we found that solar-simulated light-irradiated BaP (LBaP) inhibited apoptosis, leading to cancer. LBaP suppressed apoptosis induced by cell detachment and serum depletion in a dose and light-irradiated time-dependent manner. The antiapoptotic effect was related to the production of reactive oxygen species from degraded BaP. The cells that survived apoptosis by LBaP treatment were transformed having the ability to form colonies in soft agar and tumors in nude mice. These capabilities were specific to LBaP, not BaP itself. The results suggested that the carcinogenicity of PAHs may be attributable not only to the genetic damage induced by their metabolites, but also to the antiapoptotic effects of oxidative products on exposure to sunlight.     

Age-independent constancy of mineral contents in human ribs

On age relationships of mineral contents in human bones, the contents of the sixth rib and a piece of its compact bone were determined by inductively coupled plasma atomic emission spectrometry (ICPS). The ribs were resected from 21 subjects (14 men and 7 women) who died in age ranging from 65 to 93 yr. There were no age-dependent decreases in Ca and P contents of the ribs in the age range on ICPS. It was found that there were no age-dependent decreases in Ca and P in compact bones of ribs.     

Changes of protein profiles during pollen development in<Emphasis Type="Italic"> Lilium longiflorum</Emphasis>

Pollen proteins of Lilium longiflorum were examined at different developmental stages (young, mature and cultured) using two-dimensional differential gel electrophoresis. Quantitative changes of six proteins (MP1–MP6) during pollen development were observed in the acidic and low molecular weight region. After water absorption on the culture medium, the quantities of all six proteins were drastically changed. Mass spectrometric analysis revealed that MP2, MP3, MP4 and MP6 are late embryogenesis abundant (LEA) (D-7) protein, profilin 3, profilin 1 and enolase, respectively. The remaining two proteins (MP1 and MP5) could not be identified by mass spectrometric analysis. Immunogold electron microscopic examination showed the presence of these proteins in different regions: MP1 around lipid bodies, suggesting possible involvement in lipid metabolism, MP4 near actin in the cytoplasm, indicating the possibility of its interaction with actin in the regulatory pathways of pollen, and MP2 and MP6 in the cytoplasm.     

Stimulating the production of homoisoflavonoids in cell suspension cultures of Caesalpinia pulcherrima using cork tissue

It has previously been demonstrated that cork tissue increases the efficiency of the production of lipophilic secondary metabolites in diverse plant cell suspension cultures. In the present study, three new homoisoflavonoids--named dihydrobonducellin, 2'-methoxydihydrobonducellin, and 2'-methoxybonducellin--and bonducellin and isobonducellin were isolated from Caesalpinia pulcherrima cultured cells coincubated with cork tissue. Cork tissue increased the production of 2'-methoxybonducellin by about 7-fold relative to control cells, and more than 80% of the product was recoverable from the cork tissue. When cork tissue and methyl jasmonate or yeast extract were added simultaneously to the medium, the amount of 2'-methoxybonducellin produced increased further. The production of the other four homoisoflavonoids was enhanced by variable amounts. Our results indicate that the addition of cork tissue would be an effective technique for investigating formation of secondary metabolites that usually accumulate only in trace amounts.