Guaiane- and aristolane-type sesquiterpenoids of Nardostachys chinensis roots

Two guaiane-type compounds, nardoguaianone J and K (1 and 2) and two aristolane-type compounds, kanshone F and G (3 and 4), were isolated from Nardostachys chinensis roots. The structures including the absolute configurations were elucidated by spectral means and by comparison of their CD spectra.     

Identification of the sex pheromone components secreted by female moths of Peridroma saucia (Noctuidae: Noctuinae)

The variegated cutworm, Peridroma saucia Hübner, is a lepidopteran pest to a large number of crops in Canada, the United States, and Europe. It was probably naturalized in Japan in the 1970s. The pheromone glands of the female moth include two components with electroantennographic activity in a ratio of 3:1. GC-MS analyses of pheromone extracts untreated and treated with dimethyl disulfide revealed the major component to be (Z)-11-hexadecenyl acetate and the minor component to be (Z)-9-tetradecenyl acetate. The synthetic pheromone was used to attract a large number of males in a vegetable field in Tokyo, which suggests that this species has already become a harmful pest in Japan.     

Co-ordinating retinal histogenesis: early cell cycle exit enhances early cell fate determination in the Xenopus retina

The laminar arrays of distinct cell types in the vertebrate retina are built by a histogenic process in which cell fate is correlated with birth order. To explore this co-ordination mechanistically, we altered the relative timing of cell cycle exit in the developing Xenopus retina and asked whether this affected the activity of neural determinants. We found that Xath5, a bHLH proneural gene that promotes retinal ganglion cell (RGC) fate, ( Kanekar, S., Perron, M., Dorsky, R., Harris, W. A., Jan, L. Y., Jan, Y. N. and Vetter, M. L. (1997) Neuron 19, 981-994), does not cause these cells to be born prematurely. To drive cells out of the cell cycle early, therefore, we misexpressed the cyclin kinase inhibitor, p27Xic1. We found that early cell cycle exit potentiates the ability of Xath5 to promote RGC fate. Conversely, the cell cycle activator, cyclin E1, which inhibits cell cycle exit, biases Xath5-expressing cells toward later neuronal fates. We found that Notch activation in this system caused cells to exit the cell cycle prematurely, and when it is misexpressed with Xath5, it also potentiates the induction of RGCs. The potentiation is counteracted by co-expression of cyclin E1. These results suggest a model of histogenesis in which the activity of factors that promote early cell cycle exit enhances the activity of factors that promote early cellular fates.     

Gas chromatographic-mass spectrometric determination of erythrocyte 3-deoxyglucosone in diabetic patients

To determine if the erythrocyte levels of 3-deoxyglucosone (3-DG) are increased in diabetic patients, and if they correlate with glycemic status, they were measured in diabetic patients without renal disease as well as in healthy subjects. The erythrocyte levels of 3-DG were measured by a selected ion monitoring method of gas chromatography-chemical ionization mass spectrometry using [(13)C(6)]-3-DG as an internal standard. The erythrocyte levels of 3-DG were significantly higher in diabetic patients than in healthy subjects. The erythrocyte concentration of 3-DG was significantly and positively correlated with HbA1c (r=0.84, P<0.001). However, no significant correlation could be found between erythrocyte 3-DG and age, onset age of diabetes, or duration of diabetes in our group of diabetic patients. In diabetes, the production of 3-DG in the erythrocytes is increased via the polyol pathway and/or the Maillard reaction due to hyperglycemia.     

Hexameric ring structure of the ATPase domain of the membrane-integrated metalloprotease FtsH from Thermus thermophilus HB8

FtsH is a cytoplasmic membrane-integrated, ATP-dependent metalloprotease, which processively degrades both cytoplasmic and membrane proteins in concert with unfolding. The FtsH protein is divided into the N-terminal transmembrane region and the larger C-terminal cytoplasmic region, which consists of an ATPase domain and a protease domain. We have determined the crystal structures of the Thermus thermophilus FtsH ATPase domain in the nucleotide-free and AMP-PNP- and ADP-bound states, in addition to the domain with the extra preceding segment. Combined with the mapping of the putative substrate binding region, these structures suggest that FtsH internally forms a hexameric ring structure, in which ATP binding could cause a conformational change to facilitate transport of substrates into the protease domain through the central pore.     

Detection of underlying characteristics of nuclear chromatin patterns of thyroid tumor cells using texture and factor analyses

BACKGROUND: Aspiration biopsy cytology of thyroid tumors has been used more frequently in recent times to differentiate between malignant and benign lesions. Chromatin patterns of the tumor cell nuclei are one of most important factors for cytologic diagnosis. The interpretation of nuclear chromatin patterns is subjective and more difficult than that of nuclear size or shape. In the present report, we investigated how to detect underlying chromatin characteristics of benign and malignant thyroid tumor cells by means of texture and factor analyses. METHODS: We employed a computer-aided system in which light microscopy was combined with an image processor and monochrome camera. Using this system, 100 randomly selected cells in a Papanicolaou stained, aspiration biopsy cytologic smear in each case of 39 benign and malignant thyroid tumor cases were digitized. We applied two-dimensional and higher texture analyses with the use of co-occurrence and run-length matrices to analyze the chromatin patterns. Factor analysis was used to determine whether a large number of independent variables actually measured one or more underlying common variables. RESULTS: According to parameters with high factor-loading values, the morphologic chromatin characters were classified into three categories according to heterogeneity, contrast, and homogeneity of chromatin patterns. On the basis of analyses with these morphologic categories, nuclei of papillary carcinoma showed higher contrast of chromatin patterns than did those of the benign group. Moreover, there was a variety of contrasting chromatin patterns among cells in each papillary carcinoma case in comparison with the benign group. In contrast, follicular carcinomas showed a significant difference in the standard deviation of factor 3, which indicated more monotonous chromatin patterns among cells in each follicular carcinoma case than in each benign case. CONCLUSION: We believe that this technique, using texture and factor analyses, is useful in the detection of underlying characteristics of nuclear chromatin patterns in aspiration biopsy cytology.     

Endothelin-1 binding to endothelin receptors in the rat anterior pituitary gland: possible formation of an ETA-ETB receptor heterodimer

1. Interaction in the recognition of endothelin-1 (ET-1), a typical bivalent ET receptor-ligand, between ET_A and ET_B receptors was investigated in the rat anterior pituitary gland, using our quantitative receptor autoradiographic method with tissue sections preserving the cell-membrane structure and ET receptor-related compounds.2. In saturation binding studies with increasing concentrations (0.77–200 pM) of ¹²⁵I-ET-1 (nonselective bivalent radioligand), ¹²⁵I-ET-1 binding to the rat anterior pituitary gland was saturable and single with a K _D of 71 pM and a B _max of 120 fmol mg^–1. When 1.0 M BQ-123 (ET_A antagonist) was added to the incubation buffer, binding parameters were 8.3 pM of K _D and 8.0 fmol mg^–1 of B _max, whereas 10 nM sarafotoxin S6c (ET_B agonist) exerted little change in these binding parameters (K _D, 72 pM; B _max, 110 fmol mg^–1).3. Competition binding studies with a fixed amount (3.8 pM) of ¹²⁵I-ET-1 revealed that when 1.0 M BQ-123 was present in the incubation buffer, ET_B receptor-related compounds such as sarafotoxin S6c, ET-3, IRL1620 (ET_B agonist), and BQ-788 (ET_B antagonist) competitively inhibited ¹²⁵I-ET-1 binding with K _is of 140, 18, 350 pM, and 14 nM, respectively, however, these compounds were not significant competitors for ¹²⁵I-ET-1 binding in the case of absence of BQ-123.4. In cold-ligand saturation studies with a fixed amount (390 pM) of ¹²⁵I-IRL 1620 (ET_B radioligand), IRL1620 bound to a single population of the ET_B receptor, and no change was observed in binding characteristics in the presence of 1.0 M BQ-123. ¹²⁵I-IRL1620 binding was competitively inhibited by ET-1 and ET-3 in the absence of BQ-123, with K _is of 20 and 29 pM, respectively, the affinities being much the same as those of 29 nM, in the presence of 1.0 M BQ-123.5. Two nonbivalent ET_A antagonists, BQ-123 and PD151242, were highly sensitive and full competitors for ¹²⁵I-ET-1 binding (5.0 pM), in the presence of 10 nM sarafotoxin S6c.6. Taken together with the present finding that mRNAs encoding the rat ET_A and the ET_B receptors are expressed in the anterior pituitary gland, we tentatively conclude that although there are ET_A and ET_B receptors with a functional binding capability for ET receptor-ligands, the ET_B receptor does not independently recognize ET-1 without the aid of the ET_A receptor. If this thesis is tenable, then ET-1 can bridge between the two receptors to form an ET_A–ET_B receptor heterodimer.     

Stabilization of FtsH-unfolded protein complex by binding of ATP and blocking of protease

Mutations in small heterodimer partner (SHP) and hepatocyte nuclear factor 4alpha (HNF4alpha) are associated with mild obesity and diabetes mellitus, respectively. Both receptors work together to determine the normal pancreatic beta-cell function. We examined their subcellular localization and interaction in living cells by tagging them with yellow and cyan variants of green fluorescent protein (GFP) variants. Expressed SHP resided only in the cytoplasm in COS-7 cells which lacks HNF4alpha, but predominantly in the nucleus in insulinoma cells (MIN6). HNF4alpha was localized exclusively in the nuclei of both cells, coexpressed with HNF4alpha in COS-7 cells, redistributed in the nucleus, depending on the amount of HNF4alpha. We found fluorescence resonance energy transfer between GFP-tagged SHP and HNF4alpha, indicating a specific close association between them in the nucleus. The results strongly suggest that SHP exists primarily in the cytoplasm and is translocated into the nucleus on interacting with its nuclear receptor partner HNF4alpha.