Metabolism of D-arabinose by Escherichia coli B-r

The pathway for d-arabinose metabolism in Escherichia coli B/r has been determined. Evidence is presented to support the following metabolic scheme: d-arabinose d-ribulose right harpoon-up d-ribulose-5-phosphate d-xylulose-5-phosphate.     

Seasonal adaptation of thermal and metabolic responses in men wearing different clothing at 10° C

Thermoregulatory responses at ambient temperatures of 20 and 10° C in six male subjects wearing two different kinds of clothing were compared between summer and winter. The two different kinds of clothing were one insulating the upper half of the body lightly and the lower half of the body heavily (clothing A, the weight in the upper and lower halves of the body being, respectively, 489 g and 1278 g) and the other insulating the upper half of the body heavily and the lower half of the body lightly (clothing B: 1212 g and 559 g). The major findings are summarized as follow. (i) Rectal temperature was kept significantly higher in clothing B than in clothing A both in summer and winter. (ii) The fall of rectal temperature was significantly greater in summer than in winter in both types of clothing. (iii) Mean skin temperatures and skin temperatures in the face, chest, thigh and leg were significantly lower atT _a of 10° C in summer than in winter in clothing A, while skin temperatures in the face and thigh were also significantly lower atT _a of 10° C in summer than in winter in clothing B. (iv) Metabolic heat production was higher in summer than in winter at 20 and 10° C in both types of clothing. (v) The subjects felt cooler and colder toT _a of 10° C in summer than in winter in both types of clothing. These different responses occurring between summer and winter are discussed mainly in terms of total conductance and dry heat loss.     

Mycobacterium kansasii-induced death of murine macrophages involves endoplasmic reticulum stress responses mediated by reactive oxygen species generation or calpain activation

Although pathogenic mechanisms of tuberculosis have been extensively studied, little is known about the pathogenic mechanisms of Mycobacterium kansasii. In this work the influence of virulence and ER-stress mediated apoptosis of macrophages during two different strains of M. kansasii infection was investigated. We show that M. kansasii infection is associated with ER stress-mediated apoptosis in the murine macrophage cell line RAW 264.7. Infection of RAW 264.7 cells in vitro with apoptosis-inducing a clinical isolate of M. kansasii SM-1 (SM-1) resulted in strong induction of ER stress responses compared with M. kansasii type strain (ATCC 12478)-infected RAW 264.7 cells. Interestingly, inhibition of calpain prevented the induction of CHOP and Bip in ATCC 12478-infected RAW 264.7 cells but not in RAW 264.7 cells infected with SM-1. In contrast, reactive oxygen species (ROS) were significantly increased only in RAW 264.7 cells infected with SM-1. We propose that ROS generation is important for triggering ER stress-mediated apoptosis during SM-1 infection, whereas ATCC 12478-induced, ER stress-mediated apoptosis is associated with calpain activation. Our results demonstrate that the ER stress pathway plays important roles in the pathogenesis of M. kansasii infections, and that different strains of M. kansasii induce different patterns of ER stress-mediated apoptosis.     

Temperature-dependent post-diapause development and prediction of spring emergence of the pine needle gall midge (Dipt., Cecidomyiidae)

Abstract:  We determined the influence of temperature on post-diapause development of overwintered Thecodiplosis japonensis Uchida et Inouye (Dipt., Cecidomyiidae) under various treatments (12, 15, 18, 21, 24, 27 and 30°C) in an effort to predict its spring emergence. Survival and developmental period for the overwintered larvae and pupae were significantly influenced by temperature. Linear and nonlinear regression models quantitatively described temperature-dependent development and survival of T. japonensis . The survival models exhibited right-skewed bell shape patterns for all stages, indicating a more detrimental impact on survival at high temperatures. Theoretical optimum temperatures with highest survival were 22.3, 24.0 and 24.0°C for the overwintered larvae, pupae and total post-diapause development (the larvae to adults) respectively. Pupal mortality was higher at all temperatures than larval mortality and the suitable range of temperature for pupae was narrower than that of larvae. The nonlinear Briere model estimated that optimum temperatures with the fastest development were 29.1°C for larvae, 27.6°C for pupae and 27.0°C for larvae to adults. In a linear model, the lower threshold temperatures were 5.1, 7.1 and 5.9°C for larvae, pupae, and larvae to adults respectively. A predictive degree-day model was developed using trap catches of T. japonensis adult emergence during 1991–1995. The model accounted for 84.6% of year-to-year variation in adult emergence and predicted accurately the median emergence time in 1996.     

Establishment and maintenance of human embryonic stem cell lines on human feeder cells derived from uterine endometrium under serum-free condition

Human embryonic stem (hES) cells are usually established and maintained on mouse embryonic fibroblast (MEFs) feeder layers. However, it is desirable to develop human feeder cells because animal feeder cells are associated with risks such as viral infection and/or pathogen transmission. In this study, we attempted to establish new hES cell lines using human uterine endometrial cells (hUECs) to prevent the risks associated with animal feeder cells and for their eventual application in cell-replacement therapy. Inner cell masses (ICMs) of cultured blastocysts were isolated by immunosurgery and then cultured on mitotically inactivated hUEC feeder layers. Cultured ICMs formed colonies by continuous proliferation and were allowed to proliferate continuously for 40, 50, and 55 passages. The established hES cell lines (Miz-hES-14, -15, and -9, respectively) exhibited typical hES cells characteristics, including continuous growth, expression of specific markers, normal karyotypes, and differentiation capacity. The hUEC feeders have the advantage that they can be used for many passages, whereas MEF feeder cells can only be used as feeder cells for a limited number of passages. The hUECs are available to establish and maintain hES cells, and the high expression of embryotrophic factors and extracellular matrices by hUECs may be important to the efficient growth of hES cells. Clinical applications require the establishment and expansion of hES cells under stable xeno-free culture systems.     

A novel in vitro retinal differentiation model by co-culturing adult human bone marrow stem cells with retinal pigmented epithelium cells

Human retinal pigment epithelium (HRPE) cells are important in maintaining the normal physiology within the neurosensory retina and photoreceptors. Recently, transplantation of HRPE has become a possible therapeutic approach for retinal degeneration. By negative immunoselection (CD45 and glycophorin A), in this study, we have isolated and cultivated adult human bone marrow stem cells (BMSCs) with multilineage differentiation potential. After a 2- to 4-week culture under chondrogenic, osteogenic, adipogenic, and hepatogenic induction medium, these BMSCs were found to differentiate into cartilage, bone, adipocyte, and hepatocyte-like cells, respectively. We also showed that these BMSCs could differentiate into neural precursor cells (nestin-positive) and mature neurons (MAP-2 and Tuj1-positive) following treatment of neural selection and induction medium for 1 month. Furthermore, the plasticity of BMSCs was confirmed by initiating their differentiation into retinal cells and photoreceptor lineages by co-culturing with HRPE cells. The latter system provides an ex vivo expansion model of culturing photoreceptors for the treatment of retinal degeneration diseases.     

Flavin containing monooxygenase 3 exerts broad effects on glucose and lipid metabolism and atherosclerosis

We performed silencing and overexpression studies of flavin containing monooxygenase (FMO) 3 in hyperlipidemic mouse models to examine its effects on trimethylamine N-oxide (TMAO) levels and atherosclerosis. Knockdown of hepatic FMO3 in LDL receptor knockout mice using an antisense oligonucleotide resulted in decreased circulating TMAO levels and atherosclerosis. Surprisingly, we also observed significant decreases in hepatic lipids and in levels of plasma lipids, ketone bodies, glucose, and insulin. FMO3 overexpression in transgenic mice, on the other hand, increased hepatic and plasma lipids. Global gene expression analyses suggested that these effects of FMO3 on lipogenesis and gluconeogenesis may be mediated through the PPARα and Kruppel-like factor 15 pathways. In vivo and in vitro results were consistent with the concept that the effects were mediated directly by FMO3 rather than trimethylamine/TMAO; in particular, overexpression of FMO3 in the human hepatoma cell line, Hep3B, resulted in significantly increased glucose secretion and lipogenesis. Our results indicate a major role for FMO3 in modulating glucose and lipid homeostasis in vivo, and they suggest that pharmacologic inhibition of FMO3 to reduce TMAO levels would be confounded by metabolic interactions.     

GSK-3α Is a Novel Target of CREB and CREB-GSK-3α Signaling Participates in Cell Viability in Lung Cancer

Overexpression or activation of cyclic AMP-response element-binding protein (CREB) has been known to be involved in several human malignancies, including lung cancer. Genes regulated by CREB have been reported to suppress apoptosis, induce cell proliferation, inflammation, and tumor metastasis. However, the critical target genes of CREB in lung cancer have not been well understood. Here, we identified GSK-3α as one of the CREB target genes which is critical for the viability of lung cancer cells. The CREB knockdown significantly reduced the expression of GSK-3α and the direct binding of CREB on the promoter of GSK3A was identified. Kaplan-Meier analysis with a public database showed a prognostic significance of aberrant GSK-3α expression in lung cancer. Inhibition of GSK-3α suppressed cell viability, colony formation, and tumor growth. For the first time, we demonstrated that GSK-3α is regulated by CREB in lung cancer and is required for the cell viability. These findings implicate CREB-GSK-3α axis as a novel therapeutic target for lung cancer treatment.     

N-Acetylcysteine ameliorates lipopolysaccharide-induced organ damage in conscious rats

Lipopolysaccharide is strongly associated with septic shock, leading to multiple organ failure. It can activate monocytes and macrophages to release proinflammatory mediators such as tumor necrosis factor- (TNF-), interleukin-1 (IL-1), and nitric oxide (NO). The present experiments were designed to induce endotoxin shock by an intravenous injection ofKlebsiella pneumoniae lipopolysaccharide (LPS, 10 mg/kg) in conscious rats. Arterial pressure and heart rate (HR) were continuously monitored for 48 h after LPS administration. N-Acetyl-cysteine was used to study its effects on organ damage. Biochemical substances were measured to reflect organ functions. Biochemical factors included blood urea nitrogen (BUN), creatinine (Cre), lactic dehydrogenase (LDH), creatine phosphokinase (CPK), aspartate transferase (GOT), alanine transferase (GPT), TNF-, IL-1, methyl guanidine (MG), and nitrites/nitrates. LPS caused significant increases in blood BUN, Cre, LDH, CPK, GOT, GPT, TNF-, IL-1, MG levels, and HR, as well as a decrease in mean arterial pressure and an elevation of nitrites/nitrates. N-Acetylcysteine suppressed the release of TNF-, IL-1, and MG, but enhanced NO production. These actions ameliorate LPS-induced organ damage in conscious rats. The beneficial effects may suggest a potential chemopreventive effect of this compound in sepsis prevention and treatment.     

Heterology expression of the Arabidopsis C-repeat/dehydration response element binding factor 1 gene confers elevated tolerance to chilling and oxidative stresses in transgenic tomato

In an attempt to improve stress tolerance of tomato (Lycopersicon esculentum) plants, an expression vector containing an Arabidopsis C-repeat/dehydration responsive element binding factor 1 (CBF1) cDNA driven by a cauliflower mosaic virus 35S promoter was transferred into tomato plants. Transgenic expression of CBF1 was proved by northern- and western-blot analyses. The degree of chilling tolerance of transgenic T(1) and T(2) plants was found to be significantly greater than that of wild-type tomato plants as measured by survival rate, chlorophyll fluorescence value, and radical elongation. The transgenic tomato plants exhibited patterns of growth retardation; however, they resumed normal growth after GA(3) (gibberellic acid) treatment. More importantly, GA(3)-treated transgenic plants still exhibited a greater degree of chilling tolerance compared with wild-type plants. Subtractive hybridization was performed to isolate the responsive genes of heterologous Arabidopsis CBF1 in transgenic tomato plants. CATALASE1 (CAT1) was obtained and showed activation in transgenic tomato plants. The CAT1 gene and catalase activity were also highly induced in the transgenic tomato plants. The level of H(2)O(2) in the transgenic plants was lower than that in the wild-type plants under either normal or cold conditions. The transgenic plants also exhibited considerable tolerance against oxidative damage induced by methyl viologen. Results from the current study suggest that heterologous CBF1 expression in transgenic tomato plants may induce several oxidative-stress responsive genes to protect from chilling stress.