Accurate spatial and temporal transgene expression driven by a 3.8-kilobase promoter of the bovine β-casein gene in the lactating mouse mammary gland

The spatial, temporal, and hormonal pattern of expression of the β-casein gene is highly regulated and confined to the epithelial cells of the lactating mammary gland. Previous studies have shown that 1.7 kb of the bovine β-casein promoter were able to drive cell-specific and hormone-dependent expression to a mouse mammary cell line but failed to induce accurate expression to the mammary gland of transgenic mice. We investigated here the ability of 3.8 kb of the bovine β-casein gene promoter to drive the expression of the human growth hormone (hGH) gene in transgenic mice. A Northern blot analysis using total RNA obtained from different tissues of lactating and nonlactating females revealed the presence of hGH mRNA only in the mammary gland of lactating females. hGH mRNA was not detectable in the mammary gland of virgin females or males. A developmental analysis showed that hGH mRNA only peaked on parturition, resembling more closely the bovine β-casein temporal expression pattern rather than the murine. In situ hibridization studies performed on mammary gland sections showed that the cellular pattern of hGH expression was homogeneous in all lobules from heterozygous and homozygous transgenic mice. Silver grain counts on the tissue sections highly correlated with the hGH contents in the milk determined by radioimmunoassay (r = 0.996). Thus 3.8 kb of the bovine β-casein promoter direct a high-level expression of a reporter gene to the lactating mammary gland of transgenic mice in a tissue-specific and developmentally regulated manner. Mol. Reprod. Dev. 49:236–245, 1998. © 1998 Wiley-Liss, Inc.     

Defective neurite extension is caused by a mutation in amyloid β/A4 (Aβ) protein precursor found in Familial Alzheimer's Disease

Clonal central nervous system neuronal cells, B103, do not synthesize detectable endogenous APP or APLP. B103 cells transfected with both wild-type (B103/APP) and mutant APP construct (B103/APPΔNL) secreted comparable amounts of soluble forms of APP (sAPP). B103/APP cells produced sAPP and cleaved at amyloid β/A4 (Aβ) 16, the α-secretase site, and B103/APPΔNL cells produced sAPPβ cleaved at Aβ 1, the β-secretase site. B103/APPΔNL cells developed fewer neurites than B103/APP cells in a serum-free defined medium. Neurite numbers of parent B103 cells were increased by the 50% conditioned medium (CM) from B103/APP cells but reduced by the CM from B103/APPΔNL cells. Chemically synthesized Aβ at concentration levels higher than 1 nM reduced numbers of neurites from B103 or B103/APPΔNL cells. However, Aβ at 1–100 nM could not reduce the neurite number of B103/APP cells. The protective activity against Aβ's deleterious effect to reduce neurite numbers was attributed to sAPPα in the CM. Although sAPPα could block the effect of Aβ, sAPPβ could not do so under the identical condition, suggesting the importance of the C-terminal 15-amino acid sequence in sAPPα. Nevertheless, sAPPα's protective activity required the N-terminal sequence around RERMS, previously identified to be the active domain of sAPPβ. The overall effect of APP mutation which overproduced Aβ and sAPPβ and underproduced sAPPα was a marked decline in the neurotrophic effect of APP. We suggest that the disruption of balance between the detrimental effect of Aβ and the trophic effect of sAPP may be important in the pathogenesis of AD caused by this pathogenic APP mutation © 1997 John Wiley & Sons, Inc. J Neurobiol 32: 469–480, 1997     

Effects of Estrogen and Phytoestrogen Treatment on an In Vitro Model of Recurrent Stroke on HT22 Neuronal Cell Line

An increase of stroke incidence occurs in women with the decline of estrogen levels following menopause. This ischemic damage may recur, especially soon after the first insult has occurred. We evaluated the effects of estrogen and phytoestrogen treatment on an in vitro recurrent stroke model using the HT22 neuronal cell line. HT22 cells were treated with 17β-estradiol or genistein 1 h after the beginning of the first of two oxygen and glucose deprivation/reoxygenation (OGD/R) cycles. During the second OGD, there was a deterioration of some components of the electron transport chain, such as cytochrome c oxidase subunit 1 with a subsequent increase of reactive oxygen species (ROS) production. Accordingly, there was also an increase of apoptotic phenomena demonstrated by poly(ADP-ribose) polymerase 1 cleavage, Caspase-3 activity, and Annexin V levels. The recurrent ischemic injury also raised the hypoxia-inducible factor 1α and glucose transporter 1 levels, as well as the ratio between the lipidated and cytosolic forms of microtubule-associated protein 1A/1B-light chain 3 (LC3-II/LC3-I). We found a positive effect of estradiol and genistein treatment by partially preserving the impaired cell viability after the recurrent ischemic injury; however, this positive effect does not seem to be mediated neither by blocking apoptosis processes nor by decreasing ROS production. This work contribute to the better understanding of the molecular mechanisms triggered by recurrent ischemic damage in neuronal cells and, therefore, could help with the development of an effective treatment to minimize the consequences of this pathology.     

Cortisol stress response and histopathological alteration index in Clarias gariepinus exposed to sublethal concentrations of Qua Iboe crude oil and rig wash

The histological effect on and stress response of post juvenile Clarias gariepinus exposed to Qua Iboe crude oil and rig wash were investigated. Fish weighing 60–90 g and measuring 16–18 cm were exposed for 7–28 days to 8.00 ml^?1 Qua Iboe crude oil and 0.0018 ml^–1 rig wash, both being 0.1 of the 96 hr LC₅₀. Blood samples of C. gariepinus were collected every seven days and evaluated for stress by measuring cortisol concentration. The gills and liver were studied and scored for Gill Alteration Index (GAI) and Hepatic Alteration Index (HAI), respectively. There was an increase in cortisol level up to the 7th and 14th day among the group exposed to Qua Iboe crude oil, with a decrease on the 21st and 28th day. The rig wash group increased in cortisol level up to the 7th day and decreased slightly on the 14th day, after which the trend became irregular. The toxic effects of the Qua Iboe crude oil and rig wash were time dependent, as shown by the histopathological alteration index (HAI) of gill and liver. After 28 days of exposure, the gills had irreparable damage due to high frequency of cellular necrosis and degeneration, whereas the liver had from moderate to severe damage due to the high frequency of cellular degeneration and inflammation. Qua Iboe crude oil and rig wash are both toxic to C. gariepinus, therefore their indiscriminate discharge to the environment must be discouraged.     

The Thr505 and Ser557 residues of the AGT1-encoded alpha-glucoside transporter are critical for maltotriose transport in Saccharomyces cerevisiae

Aims:  The main objective of this study was to identify amino acid residues in the AGT1‐encoded α‐glucoside transporter (Agt1p) that are critical for efficient transport of maltotriose in the yeast Saccharomyces cerevisiae. Methods and Results:  The sequences of two AGT1‐encoded α‐glucoside transporters with different efficiencies of maltotriose transport in two Saccharomyces strains (WH310 and WH314) were compared. The sequence variations and discrepancies between these two proteins (Agt1p_WH310 and Agt1p_WH314) were investigated for potential effects on the functionality and maltotriose transport efficiency of these two AGT1‐encoded α‐glucoside transporters. A 23‐amino‐acid C‐terminal truncation proved not to be critical for maltotriose affinity. The identification of three amino acid differences, which potentially could have been instrumental in the transportation of maltotriose, were further investigated. Single mutations were created to restore the point mutations I505T, V549A and T557S one by one. The single site mutant V549A showed a decrease in maltotriose transport ability, and the I505T and T557S mutants showed complete reduction in maltotriose transport. Conclusions:  The amino acids Thr⁵⁰⁵ and Ser⁵⁵⁷, which are respectively located in the transmembrane (TM) segment TM¹¹ and on the intracellular segment after TM¹² of the AGT1‐encoded α‐glucoside transporters, are critical for efficient transport of maltotriose in S. cerevisiae. Significance and Impact of the Study:  Improved fermentation of starch and its dextrin products, such as maltotriose and maltose, would benefit the brewing and whisky industries. This study could facilitate the development of engineered maltotriose transporters adapted to starch‐efficient fermentation systems, and offers prospects for the development of yeast strains with improved maltose and maltotriose uptake capabilities that, in turn, could increase the overall fermentation efficiencies in the beer and whisky industries.     

Synthesis and biological evaluation of asymmetric gramicidin S analogues containing modified d-phenylalanine residues

The synthesis of new analogues of the cationic antimicrobial peptide gramicidin S, having a modified d-phenylalanine residue, their antibacterial properties against several Gram positive and negative strains, as well as their hemolytic activity is reported.     

Winter and spring ecology of <Emphasis Type="Italic">Anaphes nitens</Emphasis>, a solitary egg-parasitoid of the <Emphasis Type="Italic">Eucalyptus</Emphasis> snout-beetle <Emphasis Type="Italic">Gonipterus scutellatus</Emphasis>

We investigated the effects of temperature, photoperiod, food and host availability, and body size on the overwintering abilities of the egg parasitoid Anaphes nitens Girault (Hymenoptera, Mymaridae) under natural conditions. Seven groups of eighty females received one of four treatments (n = 20): (i) honey and hosts, (ii) water and hosts, (iii) honey, or (iv) water. Seven groups of forty males received only honey or water (n = 20). To test if short day-length is the main cue for larval dormancy, the experiment was replicated inside a climate chamber at 20°C and under a winter photoperiod. A. nitens overwinters because of quiescence or oligopause inside the hosts and increased adult longevity. Mean pre-emergence mortality was up to 26% indoors and 15.2% outdoors, males being more affected. Development time had a significant and positive effect on body size. Honey-fed females without hosts had the highest longevity (53 days). Mother’s diet and size affected development time, body size, longevity, and fecundity of the progeny. The results confirm the good adaptation of the parasitoid to the environmental conditions of NW Spain and its ability to synchronize its life cycle with the phenology of the host. Handling editor: Drik Babendreier.     

The impact of the International Atomic Energy Agency (IAEA) program on radiation and tissue banking in Cuba

The first multi-tissue bank was founded at Havana in 1958. At that time, freeze-drying was used at the bank as a method of preserving, as well as Cobalt 60 irradiation to sterilise bone tissue, heart valves and others. The impact of the IAEA program in tissue banking activities in Cuba can be summarised as follows: (a) Increase in the production of sterilised tissues using ionising radiation (bone, pig skin and amnion) for medical treatment in the tissue bank of the Hospital Frank Pais; (b) increase of the quality of the productions of bone tissues, pig skin and amnion; (c) reduction in the import of tissues by increasing the local production of tissues; (d) sustainability in the number of donors through the implementation of a public and professional awareness campaign; (e) training of six persons in the Regional Training Centre of Buenos Aires; (f) qualification of one person in the administration of a tissue bank and in the implementation of a Quality System. The amount of tissues produced and sterilised using the ionising radiation techniques in the established banks was 25,510 units. The amount of patients treated with sterilised tissues produced by the established banks was 2,448.