Effects of magnesium and sodium ions on the outer membrane permeability of cephalosporins in Escherichia coli

Both Mg2+ and Na+ stimulated the outer membrane permeation of negatively charged cephalosporins in Escherichia coli without any significant alteration of the permeation of a zwitterionic cephalosporin. Such stimulation was not observed in an E. coli mutant lacking porins. The stimulation was caused by the direct interaction between the cations and the porin pores, which resulted in a decrease in cation selectivity of both the Omp F and Omp C porin pores.     

Stroma-dependent maintenance of cytokine responsive hematopoietic progenitor cells derived from long-term bone marrow culture

Hematopoietic cells maintained for long periods on primary cultures of bone marrow stromal cells formed cobblestone colonies (Dexter's long-term bone marrow culture, LTBC). These stably maintained hematopoietic cells (for 4 months) were transferred to a coculture on an established spleen stromal cell line (MSS62), and maintained under stromal cell layer, where they retained their invasive ability in the restricted space between the stromal cell layer and culture substratum (DFC culture). DFC contained lineage-negative (Lin-), c-Kit+, Sca-1- cells and spontaneously produced Mac-1+, Gr-1+ cells. DFC could not grow in the absence of MSS62 stromal cells, although, GM-CSF, IL-3, or IL-7 stimulated its growth. Production of granulocyte and monocytic cells was maintained by GM-CSF or IL-3 while it was decreased by IL-7. RT-PCR analysis showed that the IL-7 responsive cell population expressed early lymphoid markers (Ikaros, Pax-5, Oct-2, Rag-1, TdT, IL-7R and Imu), while lacking expression of receptors for G-CSF (G-CSFR) and for M-CSF (M-CSFR), or myeloperoxidase (MPO). These results suggested that DFC simultaneously contained lymphoid-committed progenitors and myeloid-committed progenitors, and that cytokines may expand their responding progenitor cells under the influence of signals provided by the stromal cells. Such a stromal cell-dependent culture system may be useful to analyze the switching mechanism from constitutive to inducible hematopoiesis in vitro.     

The contraction-induced increase in Achilles tendon moment arm: A three-dimensional study

The present study aimed to re-examine the influence of the isometric plantarflexors contraction on the Achilles tendon moment arm (ATMA) and the factors influencing the ATMA in three-dimensions. A series of coronal magnetic resonance images of the right ankle were recorded at foot positions of 10° of dorsiflexion, neutral position, and 10° of plantarflexion for the rest condition and the plantarflexors contraction condition at 30% maximal voluntary effort. The shortest distance between the talocrural joint axis and the line of action of the Achilles tendon force projected to the orthogonal plane of the talocrural joint axis was determined as the ATMA. The ATMA determined in the contraction condition was significantly greater by 8 mm than that determined in the rest condition. The talocrural joint axis was displaced anteriorly by 3 mm and distally by 2 mm due to the muscle contraction. As the same time, the line of action of the Achilles tendon force was displaced posteriorly by 5 mm and medially by 2 mm. These linear displacements of the talocrural joint axis and the line of action of the Achilles tendon force accounted for the difference in the ATMAs between the two conditions by 35.9 and 62.4%, respectively. These angular displacements accounted for the total of 0.4% increase in the ATMA. These results confirm the previous findings reported in two-dimensional studies and found that the linear displacement of the line of action of the Achilles tendon force is the primary source of the contraction-induced increase in the ATMA.     

Molecular cloning of endo-beta-D-1,4-glucanase genes,rce1, rce2, and rce3, from Rhizopus oryzae

Three endoglucanase genes, designated the rce1, rce2, and rce3 genes, were isolated from Rhizopus oryzae as the first cellulase genes from the subdivision ZYGOMYCOTA: All the amino acid sequences deduced from the rce1, rce2, and rce3 genes consisted of three distinct domains: cellulose binding domains, linker domains, and catalytic domains belonging to glycosyl hydrolase family 45. The rce3 gene had two tandem repeated sequences of cellulose binding domains, while rce1 and rce2 had only one. rce1, rce2, and rce3 had various lengths of linker sequences.     

Measurement of hypocretin/orexin content in the mouse brain using an enzyme immunoassay: the effect of circadian time,age and genetic background

The hypocretins (1 and 2) have emerged as key regulators of sleep and wakefulness. We developed a high-throughput enzyme immunoassay (EIA) to measure total brain hypocretin levels from large numbers of mice. Hypocretin levels were not altered by circadian time or age. However, significant differences in one or both hypocretin peptides were observed between different mouse strains. We studied hypocretin levels in knockout and transgenic mouse models with obesity, circadian gene mutations or monoaminergic defects. Compared to controls, only histamine receptor knockouts had lower hypocretin levels. This was most pronounced in H1 receptor knockouts suggesting the existence of a positive feedback loop between hypocretin and histaminergic neurons.     

Effects of diacylglycerol on glucose, lipid metabolism, and plasma serotonin levels in lean Japanese

Objective: Diacylglycerol (DAG)‐rich oil has been suggested to suppress postprandial hyperlipidemia and promote negative caloric balance by increasing energy expenditure (EE), due to small intestine physiochemical dynamics that differ from triacylglycerol (TAG). We studied the effect of DAG on postprandial glucose/insulin metabolism by loading of carbohydrate with oil. Further, to reveal the mechanism for increased EE by DAG, we measured plasma serotonin, which is mostly present in the small intestine and mediates peripheral sympathetic thermogenesis. Methods and Procedures: Randomized crossover study with 2‐week wash‐out interval between differing fat ingestion. Seven male, lean, Japanese students ingested DAG or TAG oil with 40 g of carbohydrate. Measurements of metabolic parameters were performed before and at 2, 4, and 6 h after fat ingestion. Plasma serotonin levels and cholesterol concentration in each lipoprotein were measured using high‐performance liquid chromatography (HPLC). Results: The substitution of DAG for TAG decreased very‐low‐density lipoprotein‐cholesterol (VLDL‐C) by 45.6% at 2 h, and decreased serum insulin by 41.3% at 4 h after ingestion. The incremental area under the curve (IAUC) for VLDL‐C was positively correlated with the IAUC for insulin. Concurrently, DAG elevated plasma serotonin levels by 47.3% at 2 h, while TAG did not influence. Discussion: This study indicates that the substitution of DAG for TAG suppresses the postprandial increase in serum VLDL‐C and insulin. This study also demonstrates that DAG ingestion increases plasma serotonin, proposing a possible mechanism for a postprandial increase in EE by DAG.     

Ghrelin stimulates interleukin-8 gene expression through protein kinase C-mediated NF-kappaB pathway in human colonic epithelial cells

Ghrelin, a newly identified gastric peptide, is known for its potent activity in growth hormone (GH) release and appetite. Although ghrelin is involved in several other responses such as stress and intestinal motility, its potential role in intestinal inflammation is not clear. Here, we show that expression of ghrelin and its receptor mRNA is significantly increased during acute experimental colitis in mice injected intracolonically with trinitrobenzene sulfate (TNBS). We found by PCR that ghrelin receptor mRNA is expressed in non-transformed human colonic epithelial NCM460 cells. Exposure of NCM460 cells stably transfected with ghrelin receptor mRNA to ghrelin, increased IkappaBalpha phosphorylation and its subsequent degradation. In addition, ghrelin stimulated NF-kappaB-binding activity and NF-kappaB p65 subunit phosphorylation, and induced IL-8 promoter activity and IL-8 protein secretion. Furthermore, our data show that ghrelin-induced IkappaBalpha and p65 phosphorylation was markedly reduced by pharmacological inhibitors of intracellular calcium mobilization (BAPTA/AM) and protein kinase C (GF 109203X). Pretreatment with BAPTA/AM or GF109203X also significantly attenuated ghrelin-induced IL-8 production. Together, our results strongly suggest that ghrelin may be a proinflammatory peptide in the colon. Ghrelin may participate in the pathophysiology of colonic inflammation by inducing PKC-dependent NF-kappaB activation and IL-8 production at the colonocyte level.     

Bacteriophage-induced lytic enzyme which hydrolyzes L-alanine-D-glutamic acid peptide bond in peptidoglycan

The mode of action of bacteriophage-induced lytic enzyme “LE95” was investigated. The LE95 hydrolyzed peptide portion in peptidoglycan of $\text{Ps. aeruginosa}$ and $\text{E. coli}$. The exposed amino terminal amino acid was identified as glutamic acid by analysis of terminal amino acid by dinitrophenylation. This result suggested the LE95 hydrolyzed the peptide bond between L-alanine and D-glutamic acid in the peptidoglycan of $\text{Ps. aeruginosa}$ and $\text{E. coli}$. The enzyme did not hydrolyze various peptides prepared from bacterial cell wall. This experimental result suggested that the glycan chain of peptidoglycan would be essential for the enzymic activity.     

Fine Root Growth Increases in Response to Nitrogen Addition in Phosphorus-limited Northern Hardwood Forests

Ecosystems - Resource allocation theory posits that increased soil nutrient availability results in decreased plant investment in nutrient acquisition. We evaluated this theory by quantifying fine...