The promoter of Milk vetch dwarf virus component 8 confers effective gene expression in both dicot and monocot plants

The activity of a predicted promoter, PMC8, from Milk vetch dwarf virus was evaluated by comparing it with the cauliflower mosaic virus 35S RNA promoter (P35S) and PNCR, a promoter from Soybean chlorotic mottle virus. When the GUS fusion gene was introduced into tobacco, PMC8 showed a similar expression profile to P35S but with a more intense expression in proliferating tissues. The usefulness of PMC8 was confirmed by driving NPTII for selection of kanamycin-resistant tobacco plants with improved transformation efficiency. PMC8 was also effective in transgenic rice plants. Thus, PMC8 is useful as an alternative to P35S in both dicotyledonous and monocotyledonous plants, especially for gene expression in proliferating tissues.     

Acrylonitrile-induced sister-chromatid exchanges and DNA single-strand breaks in adult human bronchial epithelial cells

The ability of acrylonitrile to induce cytotoxicity, sister-chromatid exchanges and DNA single-strand breaks was studied in cultured human bronchial epithelial cells. The toxic effect as determined by cloning efficiency was observed at a dose of 600 micrograms/ml but not at doses of both 150 and 300 micrograms/ml. The frequency of sister-chromatid exchange in untreated cells was 3.7 +/- 1.3 per cell. In contrast, cells treated with acrylonitrile at 150 and 300 micrograms/ml exhibited 6.6 +/- 1.3 and 10.7 +/- 1.7 sister-chromatid exchanges per metaphase, respectively. DNA single-strand breaks were induced by acrylonitrile at dose levels of 200 and 500 micrograms/ml. The genotoxic effects on human bronchial epithelial cells that were directly exposed to acrylonitrile are of interest in relation to evidence for the higher lung cancer incidence of acrylonitrile workers in epidemiological studies.     

A smaller particle size improved the oral bioavailability of monkey head mushroom, Hericium erinaceum, powder resulting in enhancement of the immune response and disease resistance of white shrimp, Litopenaeus vannamei

The effects of different particle sizes (100–150, 74–100, and <74 μm) of powder of the dried and ground stipe from the monkey head mushroom, Hericium erinaceum, on the immune response and disease resistance of white shrimp, Litopenaeus vannamei, against the pathogen, Vibrio alginolyticus, were examined. Mushroom powder with a particle size of <74 μm had a significantly higher effect on the disease resistance of shrimp compared to particle sizes of >74 μm. Mortality of shrimp after being injected with V. alginolyticus was particle size-dependent, increasing from 66.7% ± 3.3%–93.3% ± 3.3% with diets containing stipe particle sizes of <74 and 100–150 μm, respectively. The mortality of shrimp fed the diet containing <74-μm stipe powder for 28 days was significant lower than that of shrimp fed with the control diet and the diet containing 74–100-μm stipe powder after being challenged by V. alginolyticus. The optimal concentration of the <74-μm mushroom powder for enhancing the immune response and disease resistance of shrimp was 0.2 μg (g shrimp)^?1 day^?1. No significant change in the total hemocyte count, differential hemocyte count, glutathione reductase, or phagocytic activity was found in shrimp fed the control diet and mushroom powder-containing diet at a level of up to 0.2 μg (g shrimp)^?1 day^?1. Shrimp fed 0.2 μg (g shrimp)^?1 day^?1 of a mushroom-containing diet had a significantly higher disease resistance to V. alginolyticus via an increase in phenoloxidase activity, respiratory bursts, superoxide dismutase activity, and glutathione peroxidase activity. Therefore, a diet containing the stipe powder of monkey head mushroom with a particle size <74 μm at a level of 0.2 μg (g shrimp)^?1 day^?1 was found to enhance the immunity and disease resistance of shrimp.     

Use of a low-speed,iso-density percoll centrifugation method to increase the viability of isolated rat hepatocyte preparations

Summary A simple yet effective method (iso-density percoll centrifugation) has been developed for consistently preparing isolated rat liver parenchymal cells with over 98% initial viability. The method has been applied to cells isolated by a variety of collagenase digestion techniques. This procedure involves the low-speed centrifugation (50 ×g) of the initial cell suspension through a percoll medium having a density of 1.06 g/ml and results in the separation of single and viable parenchymal cells from cell aggregates, debris, and nonparenchymal cells. The enriched parenchymal cells have been shown to be superior to untreated cells by a number of criteria including: preparation homogeneity, cell morphology, maintenance of cytochrome P-450, hormonal responsiveness (measured by the induction of tyrosine aminotransferase after treatment with glucagon or dexamethasone, or both), plasma membrane integrity (determined by both trypan blue exclusion and leakage of glutamic-oxaloacetic transaminase), and the DNA repair capability after treatment with benzo[a]pyrene or 2-acetylaminofluorene. This work was supported in part by the National Institutes of Health Biomedical Research Support Program, and National Institute of Environmental Health Services grant (ES-01737) awarded to M.T.S.H., and by National Cancer Institute grants CA-017175, CA-09135, CA-22484 awarded to H.C.P.N.S. was supported by a Cancer Research Campaign Grant (U. K.) through the International Union Against Cancer. This work was presented in part at the 24th Annual Meeting of the Society of Toxicology, 18–22 March 1985, San Diego, CA.     

In vitro micropropagation and ex vitro acclimation of Bupleurum kaoi-an endangered medicinal plant native to Taiwan

Summary This study reports an improved protocol for in vitro-shoot multiplication and ex vitro acclimation of Bupleurum kaoi, an endangered medicinal herb. Nodal segments were cultured in half-strength Murashige and Skoog (MS) basal medium supplemented with different concentrations of benzyladenine (BA) and kinetin. The presence of 0.25 mg l⁻¹ BA induced the highest number of shoots per explant after 8 wk of culture. Although BA was more effective than kinetin on shool multiplication, it induced hyperhydric shoots at all concentrations tested. The use of dispense paper (DP) instead of aluminum foil (AF) for container closure was found to reduce hyperhydricity and improve ex vitro acclimation. The best survival rate (61%) was obtained when plantlets were grown in MS basal medium containing 0.5 mg l⁻¹ indole-3-butyric acid and 0.1–0.2 mg l⁻¹ α-naphthaleneacetic acid using DP as container closure. Leaves of the plant treated with AF6 (two layers of AF as container closure and 6 wk of incubation) lacked epicuticular wax and possessed larger stomata, higher stomata density, and fewer functional stomata compared to those of plants treated with AF2+DP4 (two layers of AF for 2 wk, then replaced AF by three layers of DP for 4wk) and ex vitro-acclimated plantlets.     

Reduction of Splenic Immunosuppressive Cells and Enhancement of Anti-Tumor Immunity by Synergy of Fish Oil and Selenium Yeast

Growing evidence has shown that regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs) abnormally increase in cancer cachectic patients. Suppressions of Tregs and MDSCs may enhance anti-tumor immunity for cancer patients. Fish oil and selenium have been known to have many biological activities such as anti-inflammation and anti-oxidation. Whether fish oil and/or selenium have an additional effect on population of immunosuppressive cells in tumor-bearing hosts remained elusive and controversial. To gain insights into their roles on anti-tumor immunity, we studied the fish oil- and/or selenium-mediated tumor suppression and immunity on lung carcinoma, whereof cachexia develops. Advancement of cachexia in a murine lung cancer model manifested with such indicative symptoms as weight loss, chronic inflammation and disturbed immune functionality. The elevation of Tregs and MDSCs in spleens of tumor-bearing mice was positively correlated with tumor burdens. Consumption of either fish oil or selenium had little or no effect on the levels of Tregs and MDSCs. However, consumption of both fish oil and selenium together presented a synergistic effect-The population of Tregs and MDSCs decreased as opposed to increase of anti-tumor immunity when both fish oil and selenium were supplemented simultaneously, whereby losses of body weight and muscle/fat mass were alleviated significantly.     

Inhibition of sickle red cell adhesion and vasoocclusion in the microcirculation by antioxidants

In sickle cell anemia (SCA), inflammatory (i.e., intravascular sickling and transient vasoocclusive) events result in chronic endothelial activation. In addition to sickling behavior, sickle (SS) red blood cells exhibit abnormal interaction with the vascular endothelium, which is considered to have an important role in initiation of vasoocclusion. Upregulation of endothelial adhesion molecules caused by oxidants (and cytokines) may lead to increased SS red cell adhesion. We hypothesize that endothelial activation is indispensable in SS red cell adhesion to the endothelium and that antioxidants will have an inhibitory effect on this interaction. We examined the effect of selected antioxidants in ex vivo mesocecum vasculature, a well-established model that allows measurement of hemodynamic parameters and, by intravital microscopy, can allow quantification of adhesion. We tested antioxidant enzymes (SOD and catalase) and an intravascular SOD mimetic, polynitroxyl albumin (PNA), in the presence of platelet-activating factor (PAF); the latter causes endothelial oxidant generation and endothelial activation, which characterize SCA. In ex vivo preparations, PAF not only induced marked endothelial oxidant generation, it also enhanced SS red cell adhesion, resulting in frequent blockage of small-diameter venules. The adhesion, inversely related to venular diameter, and vasoocclusion were markedly inhibited by antioxidants, resulting in improved hemodynamics. PNA, the most effective antioxidant, also abolished SS red cell adhesion in non-PAF-activated preparations. Thus SS red cell adhesion and related vasoocclusion may be ameliorated by antioxidant therapy with a stable and long-acting molecule (e.g., PNA).