Time-Dependent Effects of Progressive Gamma -Linolenate Feeding on Hyperphagia,Weight Gain,and Erythrocyte Fatty Acid Composition During Growth of Zucker Obese Rats

Obese Zucker rats (fa/fa) have low levels of arachidonic acid (AA) in liver phospholipids (PL). We have previously shown that a 70% gamma-linolenate concentrate (GLA; an AA intermediate) fed at a fixed dose (0.07 g/day) normalized hepatic PL AA and reduced weight gain selectively in the obese animals. In a follow-up study, 16 obese (fa/fa) and 16 lean (Fa/Fa) 4-week-old male rats were randomized into 4 groups of 8 each and gavaged daily with soybean oil (SOY) containing 55% 18:2ω6 (an AA precursor) or GLA, using a progressive dose (≤ 5% of total calories) based on body weight. A defined diet with 11% of energy as SOY was fed ad libitum for 60 days. GLA obese had lower body weight (p<0.0001) and 60-day cumulative food intake (p<0.05) compared to SOY obese, but neither parameter differed between the lean groups. For the last twenty days cumulative food intake was identical for GLA obese and SOY lean, whereas SOY obese consumed 18% more (p<0.05). Thus the progressive dose of GLA selectively suppressed hyperphagia in obese Zucker rats. Erythrocytes collected at 15-day intervals showed parallel increases in AA in both genotypes over time, suggesting normal AA availability during rapid growth. Thus, the reduced PL AA in the livers from the obese rats probably reflects impaired distribution in selected tissues rather than reduced hepatic production. Due to the potential health risks of enriching tissue lipids with AA, great caution is advised in considering GLA as therapy for human obesity.     

Increased anti-oxidative action compensates for collagen tissue degeneration in vitiligo dermis

Vitiligo is a common depigmentation disorder characterized by the selective loss of melanocytes. In our daily clinic experience, we noticed that the skin tightness of hypopigmented lesions would be more evident in comparison to that of uninvolved perilesional skin in vitiligo patients. Therefore, we hypothesized that collagen homeostasis might be maintained in vitiligo lesions, irrespective of the substantial excessive oxidative stress that occurs in association with the disease. We found that the expression levels of collagen-related genes and anti-oxidative enzymes were upregulated in vitiligo-derived fibroblasts. Abundant collagenous fibers were observed in the papillary dermis of vitiligo lesions in comparison to uninvolved perilesional skin by electron microscopy. The production of matrix metalloproteinases that degraded collagen fibers was suppressed. The deposition of acrolein adduct protein, which is a product of oxidative stress, was significantly reduced in vitiligo dermis and fibroblasts. As part of the mechanism, we found upregulation of the NRF2 signaling pathway activity, which is an important defense system against oxidative stress. Taken together, we demonstrated that the anti-oxidative action and collagen production were upregulated and that the collagen degeneration was attenuated in vitiligo dermis. These new findings may provide important clues for the maintenance of antioxidant ability in vitiligo lesions.     

The periodic loss of metabolically unstable DNA during replication of chromosomal DNA of CHO cells

The fate of ³H-thymidine incorporated into newly synthesized DNA of CHO cells was analyzed by either the estimation of the incorporated radioactivity per cell or sedimentation in alkaline sucrose gradient. Under conditions in which DNA synthesis proceeded continuously, of incorporated radioactivity was periodically lost and regained during a 90 min chase, corresponding to a cyclic change in the sedimentation profiles. When DNA synthesis was inhibited by hydroxyurea no cyclic change of the incorporated radioactivity was observed. The cyclic changes were regarded as the result of an actual metabolic change in³H-labelled DNA probaly joining to one of the newly formed sister strands of DNA and the loss of radioactivity seems to require active continued DNA synthesis.     

Complete primary structure of a galactose-specific lectin from the venom of the rattlesnake Crotalus atrox. Homologies with Ca2(+)-dependent-type lectins

The complete primary structure of a galactose-specific lectin contained in the venom of the rattlesnake, Crotalus atrox, was determined. The lectin is composed of two covalently linked, identical subunits, each consisting of 135 amino acid residues. Under physiological conditions the lectin proved to be highly aggregated. The venom lectin contained 9 half-cystines, 8 of which formed four intrasubunit disulfide bridges (Cys3-Cys14, Cys31-Cys131, Cys38-Cys133, and Cys106-Cys123), while Cys86 was involved in an intersubunit disulfide bridge. Because of the high content of disulfide bridges, the intact lectin was extremely resistant to tryptic digestion. The determined amino acid sequence was found to be homologous with those of the so-called carbohydrate recognition domains of Ca2(+)-dependent-type lectins in animal. Among them, 8 amino acid residues (Cys31, Gly69, Trp92, Pro97, Cys106, Asp120, Cys123, and Cys131) were completely conserved. Leu40, Trp67, and Trp81 were also well conserved. The rattlesnake venom lectin showed high hemagglutinating activity. These results, together with the occurrence of similar lectins in crotalid venoms, suggest that these lectins have evolved in order to make the venom a more effective weapon to capture prey animals.     

Prostaglandin E2 Activates Ca2+ Channels in Bovine Adrenal Chromaffin Cells

We have demonstrated that prostaglandin E2 (PGE2) treatment of bovine adrenal chromaffin cells results in a sustained elevation of intracellular Ca2+ concentration ([Ca2+]i) in these cells. Because the continued elevation of [Ca2+]i was dependent on extracellular Ca2+ concentration, it can be assumed that the PGE2-induced [Ca2+]i increase is due, at least in part, to an opening of membrane Ca2+ channels. In this study, we used electrophysiological methods to examine the mechanism of the PGE2-induced [Ca2+]i increase directly. Puff application of PGE2 to the external medium resulted in a prolonged depolarization in about half of the chromaffin cells examined. In whole-cell voltage-clamp recordings, an increase in inward current was observed over a 6-7 min period following bath application of PGE2 (greater than or equal to 10 microM), even in the absence of external Na+. This inward current was abolished when the recordings were made with the cells in a Ca2(+)-free medium, but it was not inhibited by Mn2+, a blocker of voltage-dependent Ca2+ channels. In cell-attached patch-clamp configuration, PGE2 produced an increase in the opening frequency of inward currents. The reversal potential of the PGE2-induced currents was about +40 mV, which is close to the reversal potential of the Ca2+ channel. The opening frequency was not affected by membrane potential changes. In inside-out patch-clamp configuration, inositol 1,4,5-trisphosphate (2 microM) added to the cytoplasmic side activated the Ca2(+)-channel currents, but PGE2 was ineffective when applied to the cytoplasmic side. These results suggest that PGE2 activates voltage-independent Ca2+ channels in chromaffin cells through a diffusible second messenger, possibly inositol 1,4,5-trisphosphate.     

Proposal of Mycobacterium peregrinum sp. nov., nom. rev., and elevation of Mycobacterium chelonae subsp. abscessus (Kubica et al.) to species status: Mycobacterium abscessus comb. nov.

We studied the taxonomic positions of the rapidly growing organism Mycobacterium fortuitum and phenotypically related organisms. We confirmed that "Mycobacterium peregrinum" ATCC 14467T (T = type strain) is genetically independent of M. fortuitum ATCC 6841T by using various DNA hybridization conditions. Strains that were genetically identified as "M. peregrinum" were phenotypically differentiated from M. fortuitum ATCC 6841T. Thus, we propose that "M. peregrinum" should be revived as an independent species, Mycobacterium peregrinum sp. nov., nom. rev. The type strain is strain ATCC 14467. M. fortuitum subsp. acetamidolyticum ATCC 35931T exhibited a high level of DNA relatedness to M. fortuitum ATCC 6841T. The hybridized DNAs maintained stable heteroduplexity at high stringency; thus, we confirmed that M. fortuitum subsp. acetamidolyticum is identical to M. fortuitum ATCC 6841T. We found that M. chelonae subsp. abscessus ATCC 19977T is genetically different from M. chelonae subsp. chelonae NCTC 946T on the basis of the results of quantitative hybridization even under optimal conditions. There was no reason to maintain this organism as a subspecies of M. chelonae. Thus, we propose that M. chelonae subsp. abscessus should be elevated to species status as Mycobacterium abscessus (Kubica et al.) comb. nov. The type strain is strain ATCC 19977.     

Structural and functional modifications of the manganese cluster in Ca(2+)-depleted S1 and S2 states: electron paramagnetic resonance and X-ray absorption spectroscopy studies.

The effect of extraction of weakly bound Ca2+ by low-pH treatment on the O2-evolving apparatus was studied by use of low-temperature electron paramagnetic resonance (EPR) and X-ray absorption spectroscopy. In low-pH-treated PSII membranes, an S2 EPR multiline signal with modified line shape was induced by illumination at 0 degrees C, but its signal amplitude decreased upon lowering the excitation temperature with concomitant oxidation of cytochrome (cyt) b-559 in place of Mn. The half-inhibition temperature for formation of the modified multiline signal was found at -33 degrees C, which was much higher than that for formation of the normal S2 state in untreated control membranes. Signal IIf was normally induced down to -30 degrees C, but its dependence on excitation temperature was different from that for modified S2. This was interpreted as indicating that the low-temperature blockage of modified S2 formation is due to the incapability of electron abstraction from the Mn cluster. The Mn K-edge of X-ray absorption near-edge structure (XANES) spectrum shifted to lower energy by 0.8 eV after low-pH treatment, but the shift was reversed by addition of Ca2+. Upon illumination at 0 degrees C of treated membranes, the K-edge energy was up-shifted by 0.8 eV, but was not upon illumination at 210 K. These results were interpreted as indicating that extraction of weakly bound Ca2+ by low-pH treatment gives rise to structural and functional modulations of the Mn cluster.     

Structure of genomic DNA for rat platelet phospholipase A2

A genomic DNA for rat platelet phospholipase A2 was isolated by screening a rat genomic library with oligonucleotide probes based on its published amino acid sequence. The rat platelet phospholipase A2 gene had a total length of about 2.5 kb and contained five exons and four introns. The intron-exon structure of the rate gene was similar to that of human non-pancreatic phospholipase A2.     

Chondroitinase-Producing Bacteria in Natural Habitats

A search was undertaken for bacteria which degrade chondroitin sulfate in nature and to find bacteria with a usefully high rate of chondroitinase (ChSase) productivity. First, 253 ChSase-producing bacteria were obtained from aquatic and land environments in Japan by aerobic and anaerobic screening methods. Identification according to Bergey's Manual of Determinative Bacteriology or Bain and Shewan (1968) permitted assignment of the majority of the isolates to seven genera, Aeromonas, Vibrio, Flavobacterium, Beneckea, Proteus, Micrococcus, and Arthrobacter. Next, ChSase productivities of all the isolates were compared with those of two established ChSase-producing stock strains, Proteus vulgaris NCTC 4636 and Flavobacterium heparinum ATCC 13125. As a result, special attention was given to production by a strain of Aeromonas sp. of large quantities of extracellular ChSase-AC. None of the isolates from the current study displayed significant ChSase-ABC productivity. Finally, ChSase-AC was prepared from the culture fluid of the Aeromonas strain by fractional precipitation with ammonium sulfate, chromatography on phospho-cellulose and diethylaminoethyl-cellulose, and gel filtration on Sephadex G-200. It was concluded that the Aeromonas strain may represent a profitable source of the enzyme ChSase-AC.     

The skull evolution of oviraptorosaurian dinosaurs: the role of niche partitioning in diversification

Oviraptorosaurs are bird‐like theropod dinosaurs that thrived in the final pre‐extinction ecosystems during the latest Cretaceous, and the beaked, toothless skulls of derived species are regarded as some of the most peculiar among dinosaurs. Their aberrant morphologies are hypothesized to have been caused by rapid evolution triggered by an ecological/biological driver, but little is known about how their skull shapes and functional abilities diversified. Here, we use quantitative techniques to study oviraptorosaur skull form and mandibular function. We demonstrate that the snout is particularly variable, that mandibular form and upper/lower beak form are significantly correlated with phylogeny, and that there is a strong and significant correlation between mandibular function and mandible/lower beak shape, suggesting a form–function association. The form–function relationship and phylogenetic signals, along with a moderate allometric signal in lower beak form, indicate that similar mechanisms governed beak shape in oviraptorosaurs and extant birds. The two derived oviraptorosaur clades, oviraptorids and caenagnathids, are significantly separated in morphospace and functional space, indicating that they partitioned niches. Oviraptorids coexisting in the same ecosystem are also widely spread in morphological and functional space, suggesting that they finely partitioned feeding niches, whereas caenagnathids exhibit extreme disparity in beak size. The diversity of skull form and function was likely key to the diversification and evolutionary success of oviraptorosaurs in the latest Cretaceous.