Six1 is essential for early neurogenesis in the development of olfactory epithelium

The olfactory epithelium (OE) is derived from the olfactory placode (OP) during mouse development. At embryonic day (E) 10.0-E10.5, “early neurogenesis” occurs in the OE, which includes production of pioneer neurons that emigrate out of the OE and other early-differentiated neurons. Around E12.5, the OE becomes organized into mature pseudostratified epithelium and shows “established neurogenesis,” in which olfactory receptor neurons (ORNs) are differentiated from basal progenitors. Little is known about the molecular pathway of early neurogenesis. The homeodomain protein Six1 is expressed in all OP cells and neurogenic precursors in the OE. Here we show that early neurogenesis is severely disturbed despite the unaltered expression of Mash1 at E10.5 in the Six1-deficient mice (Six1^−/−). Expression levels of neurogenin1 (Ngn1) and NeuroD are reduced and those of Hes1 and Hes5 are augmented in the OE of Six1^−/− at E10.5. Pioneer neurons and cellular aggregates, which are derived from the OP/OE and situated in the mesenchyme between the OE and forebrain, are completely absent in Six1^−/−. Moreover, ORN axons and the gonadotropin-releasing hormone-positive neurons fail to extend and migrate to the forebrain, respectively. Our study indicates that Six1 plays critical roles in early neurogenesis by regulating Ngn1, NeuroD, Hes1, and Hes5.     

Personalized medicine and proteomics: lessons from non-small cell lung cancer

Personalized medicine allows the selection of treatments best suited to an individual patient and disease phenotype. To implement personalized medicine, effective tests predictive of response to treatment or susceptibility to adverse events are needed, and to develop a personalized medicine test, both high quality samples and reliable data are required. We review key features of state-of-the-art proteomic profiling and introduce further analytic developments to build a proteomic toolkit for use in personalized medicine approaches. The combination of novel analytical approaches in proteomic data generation, alignment and comparison permit translation of identified biomarkers into practical assays. We further propose an expanded statistical analysis to understand the sources of variability between individuals in terms of both protein expression and clinical variables and utilize this understanding in a predictive test.     

Four distinct trimeric forms of light-harvesting complex II isolated from the green alga Chlamydomonas reinhardtii

Photosynthesis Research - Crassulacean acid metabolism (CAM) is a specialized photosynthetic pathway present in a variety of genera including many epiphytic orchids. CAM is under circadian control...     

Norlichexanthone produced by cultured endolichenic fungus induced from Pertusaria laeviganda and its antioxidant activity

Endolichenic fungi, nonobligate microfungi that live in lichen, are promising as new bioresources of pharmacological compounds. We found that norlichexanthone isolated from the endolichenic fungus in Pertusaria laeviganda exhibited high antioxidant activity. Norlichexanthone produced by endolichenic fungus had the antioxidant activity with same level of ascorbic acid. This is the first report of high antioxidant activity of norlichexanthone.

Abbreviations: AAPH: 2,2?-azobis (2-methylpropionamidine) dihydrochloride; DPPH: 2,2-diphenyl-1-picrylhydrazyl; FL: fluorescein sodium salt; HPLC-PDA: high-performance liquid chromatography with photodiode array; LC-ESI-MS: liquid chromatography with electrospray ionization mass spectrometry; ORAC: oxygen radical absorbance capacity; PB: phosphate buffer; ROS: reactive oxygen species; TLC: thin-layer chromatography     

Developmentally regulated expression of a cell surface protein,neuropilin, in the mouse nervous system

Neuropilin (previously A5) is a cell surface glycoprotein that was originally identified in Xenopus tadpole nervous tissues. In Xenopus, neuropilin is expressed on both the presynaptic and postsynaptic elements in the visual and general somatic sensory systems, suggesting a role in neuronal cell recognition. In this study, we identified a mouse homologue of neuropilin and examined its expression in developing mouse nervous tissues. cDNA cloning and sequencing revealed that the primary structure of the mouse neuropilin was highly similar to that of Xenopus and that the extracellular segment of the molecule possessed several motifs that were expected to be involved in cell-cell interaction. Immunohistochemistry and in situ hybridization analyses in mice indicated that the expression of neuropilin was restricted to particular neuron circuits. Neuropilin protein was localized on axons but not on the somata of neurons. The expression of neuropilin persisted through the time when axons were actively growing to form neuronal connections. These observations suggest that neuropilin is involved in growth, fasciculation, and targeting for a particular groups of axons. © 1996 John Wiley & Sons, Inc.     

A cryptic melibiose transporter gene possessing a frameshift from Citrobacter freundii

Wild-type Citrobacter freundii cannot grow on melibiose as a sole source of carbon. The melibiose transporter gene melB was cloned from a C. freundii mutant M4 that could utilize melibiose as a sole carbon source. Although the cloned melB gene is closely similar to the melB genes of other bacteria, it is cryptic because of a frameshift mutation. Site-directed mutagenesis was used to construct a functional melB gene by deleting one nucleotide, resulting in the production of an active melibiose transporter. The active MelB transporter could utilize Na(+) and H(+) as coupling cations to melibiose transport. The amino acid sequence of the C. freundii MelB was found to be most similar to those of Salmonella typhimurium and Escherichia coli MelB. These facts are consistent with the phylogenetic relationship of bacteria and the cation coupling properties of the melibiose transporters.     

Involvement of serum mannan binding proteins and mannose receptors in uptake of mannosylated liposomes by macrophages

The roles of serum mannan binding protein (MBP) and the mannose receptor in the cellular uptake of mannosylated liposomes (Man-liposomes) by macrophages were studied. Man-liposomes were prepared by incorporating cholesten-5-yloxy-N-(4-((1-imino-2-beta-D-thiomannosylethyl)amino)butyl)formamide (Man-C4-Chol) into small unilamellar long circulating liposomes consisting of cholesterol (Chol) and distearoyl phosphatidylcholine (DSPC). In the in vitro cellular uptake study with cultured mouse peritoneal macrophages, [(3)H]Man-liposomes were taken up to a great extent, whereas no significant uptake was observed for [(3)H]cholesterol and DSPC liposomes without Man-C4-Chol (Bare-liposomes). The uptake of [(3)H]Man-liposomes was dose- and temperature-dependent and inhibited by an excess of mannosylated bovine serum albumin, suggesting their specific uptake via membrane mannose receptor-mediated endocytosis. Furthermore, it was demonstrated that (111)In-MBP binds strongly to Man-liposomes based on the recognition of Man-C4-Chol and markedly enhanced their uptake by macrophages. These results are supported by confocal laser microscopic images. In addition, in vivo hepatic uptake of (111)In-MBP was enhanced by Man-liposomes. On the other hand, the uptake of Man-liposomes was significantly reduced by preincubation with serum and further with MBP-depleted serum suggesting inhibitory effects of serum proteins such as albumin on mannose receptor-mediated endocytosis. The involvement of serum-type MBP and membrane mannose receptors in the uptake of Man-liposomes is thus suggested.     

Isolation of a new melanoma antigen, MART-2, containing a mutated epitope recognized by autologous tumor-infiltrating T lymphocytes

Using cDNA expression cloning, a cDNA encoding a novel human melanoma Ag, MART-2 (melanoma Ag recognized by T cells-2), recognized by HLA-A1-restricted CD8(+) T cells from tumor-infiltrating lymphocytes (TIL1362) was isolated from an autologous melanoma cell line, 1362 mel. Homologous sequences to the cDNA had been registered in the EST database. This gene encoded an uncharacterized protein expressed ubiquitously in most normal and cancer cells. A mutation (A to G transition) was found in the cDNA obtained from the1362 mel melanoma cell line in the sequences encoding the phosphate binding loop (P-loop) that resulted in loss of the ability to bind GTP. Transfection of NIH-3T3 with the mutated MART-2 did not result in the development of significant foci. By screening 36 various cancer cell lines using single-strand conformation polymorphism, a possible mutation in the P-loop of MART-2 was found in one squamous cell lung cancer cell line, EBC1. The T cell epitope for TIL1362, FLEGNEVGKTY, was identified to be encoded by the mutated sequence of the MART-2 Ag. The mutation substituted glycine in the normal peptide with glutamic acid at the third amino acid of the epitope, which is an important primary anchor amino acid for HLA-A1 peptide binding. The normal peptide, FLGGNEVGKTY, was not recognized by TIL1362, suggesting that this T cell response was specific for the autologous tumor. Although transforming activity was not detected in the NIH-3T3 assay, MART-2 with the mutation in the P-loop may be involved in the generation of melanoma through a loss of GTP binding activity.     

Effects of nitric oxide on matrix metalloproteinase-2 production by rheumatoid synovial cells

Nitric oxide (NO) is a multifunctional messenger molecule generated from L-arginine by a family of enzymes, including nitric oxide synthase (NOS). This study was performed to examine whether NO modulates the production of matrix metalloproteinases (MMPs), which degrade all components of extracellular matrix (ECM), in rheumatoid synovial cells. We investigated the effects of exogenously generated NO by a NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), on the MMPs production by rheumatoid synovial cells. Culture media conditioned by SNAP-treated synovial cells were examined by gelatin zymography and immunoblot analysis. Incubation of synovial cells with SNAP resulted in gelatinase A production in a dose-dependent fashion. Furthermore, RT-PCR analysis demonstrated that MMP-2 mRNA expression was induced in SNAP-treated synovial cells. In contrast, SNAP did not influence the production of TIMP-1 and TIMP-2, which preferentially inhibit MMP-2, by rheumatoid synovial cells. Our data indicate that NO could modulate MMP production by rheumatoid synovial cells and therefore contribute to ECM degradation of articular components in RA.     

Three-dimensional image analysis of the hepatic restructuring process in chronic active hepatitis

OBJECTIVE: To elucidate the process of hepatic restructuring in the course of chronic hepatitis from a morphologic viewpoint. STUDY DESIGN: The three-dimensional (3-D) liver structure was investigated by computer-aided reconstruction in five cases (one autopsy and four surgical cases) of chronic active hepatitis (type C), including early to late stages of restructuring. RESULTS: Our 3-D reconstruction revealed the following. At an early stage, portal and periportal inflammation and fibrosis widened the portal tracts, giving rise to the formation of portal-to-portal and portal-to-hepatic venous connections, although most central veins were still located at an almost normal site in the hepatic lobules. In a middle stage, bridging fibrosis developed to create a network of interstitium where the central veins were rather decreased in number, with regenerative nodules multiplying in the parenchyma. At the late stage, the lobular structure was destroyed, and the parenchyma consisted uniformly of regenerative nodules, with remaining but rearranged lobules among them. CONCLUSION: The above changes of liver structure suggest that in cirrhogenesis from chronic hepatitis, a combination of nodular regeneration and formation of an interstitial network come to replace the normal lobular structure, hastening the development of liver cirrhosis.