Human aquaporin adipose (AQPap) gene. Genomic structure, promoter analysis and functional mutation.

Aquaporin adipose (AQPap), which we identified from human adipose tissue, is a glycerol channel in adipocyte [Kishida et al. (2000) J. Biol. Chem. 275, 20896-20902]. In the current study, we determined the genomic structure of the human AQPap gene, and identified three AQPap-like genes that resembled (approximately 95%) AQPap, with little expression in human tissues. The AQPap promoter contained a putative peroxisome proliferator response element (PPRE) at -46 to -62, and a putative insulin response element (IRE) at -542/-536. Deletion of the PPRE abolished the pioglitazone-mediated induction of AQPap promoter activity in 3T3-L1 adipocytes. Deletion and single base pair substitution analysis of the IRE abolished the insulin-mediated suppression of the human AQPap gene. Analysis of AQPap sequence in human subjects revealed three missense mutations (R12C, V59L and G264V), and two silent mutations (A103A and G250G). The cRNA injection of the missense mutants into Xenopus oocytes revealed the absence of the activity to transport glycerol and water in the AQPap-G264V protein. In the subject homozygous for AQPap-G264V, exercise-induced increase in plasma glycerol was not observed in spite of the increased plasma noradrenaline. We suggest that AQPap is responsible for the increase of plasma glycerol during exercise in humans.     

Effect of lard and corn oil intake on serum lipids in young men

An experimental diet with lard (30 g/day for 7 days) and corn oil (30 g/day for 7 days) on high carbohydrate (basal diet) was given to four healthy Japanese young men and the effect of diets containing different fat on serum lipids was examined. Serum total cholesterol was increased significantly from a basal diet of 106 +/- 23 to 141 +/- 26 mg/dl on lard diet, and then decreased significantly (p less than 0.05) to 111 +/- 22 mg/dl on corn oil diet. Serum triglycerides increased significantly (p less than 0.01) from 66 +/- 38 to 173 +/- 32 mg/dl on basal diet. Serum HDL-cholesterol was decreased significantly (p less than 0.01) from 41.9 +/- 1.6 to 31.2 +/- 3.8 mg/dl on lard diet and increased significantly (p less than 0.05) to 41.9 +/- 4.6 mg/dl on corn oil diet. Serum HDL-cholesterol fraction was decreased significantly (p less than 0.01) from 41.6 +/- 4.9 to 28.1 +/- 3.2% on basal diets, but increased significantly (p less than 0.05) to 44.3 +/- 3.1% on lard diet, and then decreased to 36.3 +/- 2.5% on corn oil diet. Serum HDL phospholipid fraction decreased significantly (p less than 0.05) from 62.5 +/- 6.7 to 50.7 +/- 1.8% on basal diet and increased significantly (p less than 0.05) to 60.4 +/- 1.0% on lard and corn oil diet. Serum phospholipids did not change by experimental diets. It is concluded that lard and corn oil have different and specific roles in lipid metabolism.     

Development of infant common marmosets’ (<Emphasis Type=Italic>Callithrix jacchus</Emphasis>) preference for their parents over adults from another group

Parent–offspring attachment is important for animals which have offspring that require parental care for their development. Infant attachment to the mother has been examined in macaques, but it remains poorly understood in common marmosets. Here, we examined the abilities of 14 common marmoset infants to show preference for their parents over adults from another group at the ages of 4, 10, and 15 weeks. Each infant was exposed to its parent and an adult from another group in an I-shaped maze. Although 4-week-old infants did not show a significant difference between approach behaviors toward their parents and other adults, 10- and 15-week-old infants approached and stayed longer near their parents than adults from another group. These results suggest selective approach behavior develops in marmosets by the age of 10 weeks.     

Purification and characterization of the hydantoin racemase of Pseudomonas sp. strain NS671 expressed in Escherichia coli.

The hydantoin racemase gene of Pseudomonas sp. strain NS671 had been cloned and expressed in Escherichia coli. Hydantoin racemase was purified from the cell extract of the E. coli strain by phenyl-Sepharose, DEAE-Sephacel, and Sephadex G-200 chromatographies. The purified enzyme had an apparent molecular mass of 32 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. By gel filtration, a molecular mass of about 190 kDa was found, suggesting that the native enzyme is a hexamer. The optimal conditions for hydantoin racemase activity were pH 9.5 and a temperature of 45 degrees C. The enzyme activity was slightly stimulated by the addition of not only Mn2+ or Co2+ but also metal-chelating agents, indicating that the enzyme is not a metalloenzyme. On the other hand, Cu2+ and Zn2+ strongly inhibited the enzyme activity. Kinetic studies showed substrate inhibition, and the Vmax values for D- and L-5-(2-methylthioethyl)hydantoin were 35.2 and 79.0 mumol/min/mg of protein, respectively. The purified enzyme did not racemize 5-isopropylhydantoin, whereas the cells of E. coli expressing the enzyme are capable of racemizing it. After incubation of the purified enzyme with 5-isopropylhydantoin, the enzyme no longer showed 5-(2-methylthioethyl)hydantoin-racemizing activity. However, in the presence of 5-(2-methylthioethyl)hydantoin, the purified enzyme racemized 5-isopropylhydantoin completely, suggesting that 5-(2-methylthioethyl)hydantoin protects the enzyme from inactivation by 5-isopropylhydratoin. Thus, we examined the protective effect of various compounds and found that divalent-sulfur-containing compounds (R-S-R' and R-SH) have this protective effect.     

Functional morphology of the luminescence system of Siphamia versicolor (Perciformes: Apogonidae), a bacterially luminous coral reef fish

Previous studies of the luminescence system of Siphamia versicolor (Perciformes: Apogonidae) identified a ventral light organ, reflector, lens, duct, and a ventral diffuser extending from the throat to the caudal peduncle. The control and function of luminescence in this and other species of Siphamia, however, have not been defined. Morphological examination of fresh and preserved specimens identified additional components of the luminescence system involved in control and ventral emission of luminescence, including a retractable shutter over the ventral face of the light organ, contiguity of the ventral diffuser from the caudal peduncle to near the chin, and transparency of the bones and other tissues of the lower jaw. The shutter halves retract laterally, allowing the ventral release of light, and relax medially, blocking ventral light emission; topical application of norepinephrine to the exposed light organ resulted in retraction of the shutter halves, which suggests that operation of the shutter is under neuromuscular control. The extension of the diffuser to near the chin and transparency of the lower jaw allow a uniform emission of luminescence over the entire ventrum of the fish. The live aquarium‐held fish were found to readily and consistently display ventral luminescence. At twilight, the fish left the protective association with their longspine sea urchin, Diadema setosum, and began to emit ventral luminescence and to feed on zooplankton. Ventral luminescence illuminated a zone below and around the fish, which typically swam close to the substrate. Shortly after complete darkness, the fish stopped feeding and emitting luminescence. These observations suggest that S. versicolor uses ventral luminescence to attract and feed on zooplankton from the reef benthos at twilight. Ventral luminescence may allow S. versicolor to exploit for feeding the gap at twilight in the presence of potential predators as the reef transitions from diurnally active to nocturnally active organisms. J. Morphol., 2011. © 2011 Wiley‐Liss, Inc     

Production of (R)-3-Chloro-1,2-Propanediol from Prochiral 1,3-Dichloro-2-Propanol by Corynebacterium sp. Strain N-1074

The production of (R)-3-chloro-1,2-propanediol [(R)-MCP] from prochiral 1,3-dichloro-2-propanol (DCP) was examined with a bacterial strain identified as a Corynebacterium strain. The addition of glycerol as a carbon source or some chlorinated alcohols to a medium was effective for the induction of activity catalyzing the transformation of DCP into MCP. The optimum pH for (R)-MCP production by the resting cell reaction was around 8.0. The optical purity of (R)-MCP formed was improved by keeping the level of DCP in the reaction mixture at a low concentration. (R)-MCP was obtained from 77.5 mM DCP with a 97.3% molar conversion yield and an 83.8% enantiomeric excess of its optical purity by periodic feeding of the substrate.     

1H NMR studies of deuterated ribonuclease HI selectively labeled with protonated amino acids

Summary Two-dimensional (2D)¹H NMR experiments using deuterium labeling have been carried out to investigate the solution structure of ribonuclease HI (RNase HI) fromEscherichia coli (E. coli), which consists of 155 amino acids. To simplify the¹H NMR spectra, two fully deuterated enzymes bearing several prototed amino acids were prepared from an RNase HI overproducing strain ofE. coli grown in an almost fully deuterated medium. One enzyme was selectively labeled by protonated His, He. Val. and Leu. The other was labeled by only protonated His and Ile. The 2D¹H NMR spectra of these deuterated R Nase H1 proteins, selectively labeled with protonated amino acids, were much more simple than those of the normally protonated enzyme. The simplified spectra allowed unambiguous assignments of the resonance peaks and connectivities in COSY and NOESY for the side-chain protons. The spin-lattice relaxation times of the side-chain protons of the buried His residue of the deuterated enzyme became remarkably longer than that of the protonated enzyme. In contrast, the relaxation times of the side-chain protons of exposed His residues remained essentially unchanged.     

Peripheral tolerance and the qualitative characteristics of autoreactive T cell clones in primary biliary cirrhosis

Primary biliary cirrhosis is characterized by autoreactive T cells specific for the mitochondrial Ag PDC-E2(163-176). We studied the ability of eight T cell clones (TCC) specific for PDC-E2(163-176) to proliferate or become anergic in the presence of costimulation signals. TCC were stimulated with either human PDC-E2(163-176), an Escherichia coli 2-oxoglutarate dehydrogenase mimic (OGDC-E2(34-47)), or analogs with amino acid substitutions using HLA-matched allogeneic PBMC or mouse L-DR53 fibroblasts as APC. Based on their differential responses to these peptides (human PDC-E2(163-176), E. coli OGDC-E2(34-47)) in the different APC systems, TCC were classified as costimulation dependent or independent. Only costimulation-dependent TCC could become anergic. TCC with costimulation-dependent responses to OGDC-E2 become anergic to PDC-E2 when preincubated with mimic, even if costimulation is independent for PDC-E2(163-176). Anergic TCC produced IL-10. One selected TCC could not become anergic after preincubation with PDC-E2(163-176)-pulsed L-DR53 but became anergic using L-DR53 pulsed with PDC-E2 peptide analogs with a substitution at a critical TCR binding site. TCC that only respond to peptide-pulsed PBMC, but not L-DR53, proliferate with peptide-pulsed CD80/CD86-transfected L-DR53; however, anergy was not induced with peptide-pulsed L-DR53 transfected with only CD80 or CD86. These data highlight that costimulation plays a dominant role in maintaining peripheral tolerance to PBC-specific Ags. They further suggest that, under specific circumstances, molecular mimicry of an autoantigen may restore rather than break peripheral tolerance.     

Plasma brain natriuretic peptide as a surrogate marker for cardioembolic stroke

Background

Cardioembolic stroke generally results in more severe disability, since it typically has a larger ischemic area than the other types of ischemic stroke. However, it is difficult to differentiate cardioembolic stroke from non-cardioembolic stroke (atherothrombotic stroke and lacunar stroke). In this study, we evaluated the levels of plasma brain natriuretic peptide in acute ischemic stroke patients with cardioembolic stroke or non-cardioembolic stroke, and assessed the prediction factors of plasma brain natriuretic peptide and whether we could differentiate between stroke subtypes on the basis of plasma brain natriuretic peptide concentrations in addition to patient's clinical variables.

Methods

Our patient cohort consisted of 131 consecutive patients with acute cerebral infarction who were admitted to Kagawa University School of Medicine Hospital from January 1, 2005 to December 31, 2007. The mean age of patients (43 females, 88 males) was 69.6 ± 10.1 years. Sixty-two patients had cardioembolic stroke; the remaining 69 patients had non-cardioembolic stroke (including atherothrombotic stroke, lacunar stroke, or the other). Clinical variables and the plasma brain natriuretic peptide were evaluated in all patients.

Results

Plasma brain natriuretic peptide was linearly associated with atrial fibrillation, heart failure, chronic renal failure, and left atrial diameter, independently (F_4,126 = 27.6, p < 0.0001; adjusted R² = 0.45). Furthermore, atrial fibrillation, mitral regurgitation, plasma brain natriuretic peptide (> 77 pg/ml), and left atrial diameter (> 36 mm) were statistically significant independent predictors of cardioembolic stroke in the multivariable setting (Χ² = 127.5, p < 0.001).

Conclusion

It was suggested that cardioembolic stroke was strongly predicted with atrial fibrillation and plasma brain natriuretic peptide. Plasma brain natriuretic peptide can be a surrogate marker for cardioembolic stroke.