A modification of determination for glucosamine and galactosamine in glycoprotein with the amino acid analyzer: Application to total acid hydrolyzate of rat renal glomerular basement membrane

A procedure was described for the rapid determination of glucosamine and galactosamine in total acid hydrolyzate of rat renal glomerular basement membrane (GBM) by the use of amino acid analyzer. Glucosamine and galactosamine, as well as other amino acids in glycoprotein hydrolyzate, were well identified and estimated simultaneously by using a short column of HITACHI I-PF-B spherical resin, eluted with a pH 6.09 buffer containing 8% methanol at 38°C.Total time consumed for elution of galactosamine was 60 min. This method is ideal for the separation of small amount of galactosamine from hydroxylysine-rich materials.     

Preparation and biological evaluation of 5-substituted retinoic acids

Various 5-substituted retinoic acids were prepared by a palladium-catalyzed cross coupling reactions of vinyl nonaflates and E- or Z-3-tributylstannyl-2-beten-1-ol as a key reaction. These coupling products were then converted to the corresponding all-E- and 9Z-retinoic acid analogs via Horner-Emmons reaction and subsequent basic hydrolysis, and their biological activities were evaluated. The all-E-derivatives, 5-butyl and isobutyl analogs exhibited stronger effects for anti-proliferative and differentiation-inducing activities in HL-60 cells. In contrast, in 9Z-derivatives, none of the analogs showed any activity.     

Design and synthesis of biologically active analogues of vitamin K2: evaluation of their biological activities with cultured human cell lines

Novel omega-oxygenated vitamin K(2) analogues were efficiently synthesized and their biological activities were evaluated. Some were biologically active and the side-chain played an important role in gamma-carboxylation and apoptosis-inducing activity. The results provide useful information on the structure-activity relationship of vitamin K(2) analogues for the development of new drugs.     

Metastatic growth of lung cancer cells is extremely reduced in Vitamin D receptor knockout mice

Lung metastatic neoplasms are the major cause of cancer mortality. Despite the progress of diagnostic techniques and improvements in surgical procedures, the prognosis of patients with lung cancer is generally poor, even in the early stages of cancer [Cancer: Principles and Practice of Oncology, vol. 1, fifth ed., Lippincott-Raven, New York, 1997, p. 849]. Epidemiological studies indicate a positive correlation with the prevalence of cancers and low serum levels of Vitamin D metabolites [Am. J. Clin. Nutr. 54 (1991) 193s; Cancer Epidemiol. Biomark. Prev. 9 (2000) 1059]. 1alpha,25-Dihydroxyvitamin D(3) [1alpha,25(OH)(2)D(3)] is a potent inhibitor of cancer cell proliferation in vitro [Proc. Natl. Acad. Sci. U.S.A. 78 (1981) 4990; Endocrinol. 139 (1998) 1046; Mol. Endocr. 15 (2001) 1127]. There is, however, no report demonstrating that 1alpha,25(OH)(2)D(3) is operative in vivo to inhibit metastatic growth of cancer cells. To verify this possibility, we generated a stable transfectant of the Lewis lung carcinoma (LLC) cell expressing green fluorescent protein (GFP) and examined its metastatic activity in wild-type mice and Vitamin D receptor (VDR) knockout mice that exhibit no Vitamin D-dependent calcemic activity and extremely high serum levels of 1alpha,25(OH)(2)D(3) due to the overexpression of the 1alpha-hydroxylase gene [Nat. Genet. 16 (1997) 391; Proc. Natl. Acad. Sci. U.S.A. 94 (1997) 9831]. Here, we show that 1alpha,25(OH)(2)D(3) inhibits metastatic growth of lung cancer cells in the defined animal model and may work as an intrinsic factor for prevention of metastasis in intact animals. These findings establish a critical role for 1alpha,25(OH)(2)D(3) in lung metastatic neoplasms and provide a new model for metastasis of malignant cells.     

Phosphorylation of neuronal nitric oxide synthase at Ser in the dentate gyrus of rat brain after transient forebrain ischemia

We previously demonstrated that calmodulin-dependent protein kinase IIα (CaM-KIIα) phosphorylates nNOS at Ser⁸⁴⁷ in the hippocampus after forebrain ischemia; this phosphorylation attenuates NOS activity and might contribute to resistance to post-ischemic damage. We also revealed that cyclic AMP-dependent protein kinase (PKA) could phosphorylate nNOS at Ser¹⁴¹²in vitro. In this study, we focused on chronological and topographical changes in the phosphorylation of nNOS at Ser¹⁴¹² after rat forebrain ischemia. The hippocampus and adjacent cortex were collected at different times, up to 24 h, after 15 min of forebrain ischemia. NOS was partially purified from crude samples using ADP agarose gel. Neuronal NOS, phosphorylated (p)-nNOS at Ser¹⁴¹², PKA, and p-PKA at Thr¹⁹⁷ were studied in the rat hippocampus and cortex using Western blot analysis and immunohistochemistry. Western blot analysis revealed that p-nNOS at Ser¹⁴¹² significantly increased between 1 and 6 h after reperfusion in the hippocampus, but not in the cortex. PKA was cosedimented with nNOS by ADP agarose gel. Immunohistochemistry revealed that phosphorylation of nNOS at Ser¹⁴¹² and PKA at Thr¹⁹⁷ occurred in the subgranular layer of the dentate gyrus. Forebrain ischemia might thereby induce temporary activation of PKA at Thr¹⁹⁷, which then phosphorylates nNOS at Ser¹⁴¹² in the subgranular layer of the dentate gyrus.     

Sociogenetic structure of Polistes (Aphanilopterus) versicolor Olivier, 1791 colonies (Hymenoptera, Vespidae, Polistini)

The observation of two distinct, well-defined oviposition areas in nests of the primitively eusocial wasp Polistes versicolor suggests the presence of multiple egg-layers and territorial behaviors. Electrophoretic analysis of enzyme loci in pupae from 35 colonies revealed an average observed heterozygosity of 0.10 and the existence of private polymorphisms, thereby indicating a low dispersion in this species. No evidence of diploid males was found. Phenotypic segregation analysis revealed the presence of more than one egg-laying female in 15 out of 35 colonies, as well as spatially preferential oviposition in 2 out of 13 nests, with distinct oviposition areas. Genetic relatedness estimates for brood were lower than expected for haplodiploid species under monogynous conditions (r = 0.75 for female broods and r = 0.5 for male) in 4 of those 13 nests, thereby inferring complex sociogenetic structuring in Polistes versicolor colonies.     

Immunological Detection of 22K Protein in Sporulating Cells of Bacillus megaterium ATCC 12872

The synthesis and deposition of 22,000-dalton (22K) spore coat protein were examined immunochemically on the sporulating cells of Bacillus megaterium ATCC 12872 using the antibody to purified 22K spore coat protein. This antibody cross-reacted with 44K and 25K proteins in immunoblot analysis of dormant spore coat proteins. Immunoblot analysis on the sporulating cells showed that 22K protein was detected from t₈ in forespore coat protein fractions. Sandwich enzyme immunoassay revealed that 22K protein in the spore coat protein fraction appeared at t₆ and reached a plateau at t₉, and 22K protein in the mother cell cytoplasmic fraction was detected at only t₇ and t₈ at a very low level.