An improved method for primary culture of ovarian androgen-producing cells in serum-free medium: Effect of lipoproteins,insulin, and insulinlike growth factor-I

Summary Although luteinizing hormone (LH) alone stimulates ovarian interstitial cells cultured in serum-free medium to synthesize large amounts of androgens, there seem to be additional factors in vivo that modulate the time course and magnitude of the cellular responses to LH. In an attempt to develop a more nearly physiologic cell culture model, lipoproteins, insulin, and insulinlike growth factor-I (IGF-I) were added to the serum-free medium. The effects of these modifications on androgen biosynthesis by dispersed cells from ovaries of hypophysectomized immature rats cultured in 96-well tissue culture plates were examined. A saturating dose of LH stimulated a 25-fold increase in androsterone synthesis at 2 d, which decreased at 4 and 6 d. Addition of human high density (hHDL) or human low density lipoprotein (hLDL) caused a 2.5-fold increase in LH-stimulated androsterone synthesis. Cells were approximately twice as sensitive to hHDL (ED₅₀=5.5±0.5 μg cholesterol/ml) compared to hLDL (ED₅₀=9.1±1.1 μg cholesterol/ml). Surprisingly, rat HDL caused only a 40% increase in LH-stimulated androsterone synthesis. When insulin alone was added to cells cultured with a saturating dose of LH, there was a 2.8-fold increase in androsterone synthesis. Addition of hHDL and insulin together caused a synergistic increase in LH-stimulated androsterone synthesis. In contrast to hHDL, which did not change the time course of LH-stimulated androsterone production, insulin prolonged maximal LH-stimulated androsterone synthesis at 4 and 6 d. Inasmuch as the ED₅₀ for insulin action (1.3±0.1 μg/ml) was supraphysiologic, the effects of IGF-I on LH-stimulated androgen synthesis were examined. IGF-I mimicked all of the effects of insulin, but at a physiologic concentration (ED₅₀=2.5±0.3 ng/ml). Ovarian cells cultured in serum-free medium supplemented with hHDL and insulin or IGF-I exhibit responses that closely approximate the physiologic responses observed in vivo. These results suggest that lipoproteins and IGF-I are important physiologic stimulators of ovarian theca-interstitial cell androgen biosynthesis which, when added to the serum-free medium, make the cellular responses in this in vitro model more nearly approximate the responses in vivo. This research was supported by research center grant HD 12303 from the National Institute of Child Health and Human Development, Bethesda, MD, and USCD Academic Senate grant RM-169M     

Somatic embryogenesis and plant regeneration from zygotic embryo explants in mexican weeping bamboo,Otatea acuminata aztecorum

An efficient protocol has been developed for the in vitro propagation of Mexican Weeping Bamboo through somatic embryogenesis from zygotic embryo explants. Mature seeds and excised embryos were cultured in the light or in the dark on both Murashige and Skoog's and Gamborg's B5 basal media with various supplements. Optimal somatic embryogenesis and plant regeneration were obtained by culture in the dark on Murashige and Skoog's basal medium supplemented with 3 mg/1 2,4-dichlorophenoxyacetic acid, 0.5 mg/1 6-benzylaminopurine and 2.0% sucrose. More than 95% of the germinating somatic embryos developed shoots and roots, and were transferred to soil with 85% success.     

Nucleoside triphosphates are required to open the CFTR chloride channel.

The CFTR Cl- channel contains two predicted nucleotide-binding domains (NBD1 and NBD2); therefore, we examined the effect of ATP on channel activity. Once phosphorylated by cAMP-dependent protein kinase (PKA), channels required cytosolic ATP to open. Activation occurred by a PKA-independent mechanism. ATP gamma S substituted for ATP in PKA phosphorylation, but it did not open the channel. Several hydrolyzable nucleotides (ATP greater than GTP greater than ITP approximately UTP greater than CTP) reversibly activated phosphorylated channels, but nonhydrolyzable analogs and Mg(2+)-free ATP did not. Studies of CFTR mutants indicated that ATP controls channel activity independent of the R domain and suggested that hydrolysis of ATP by NBD1 may be sufficient for channel opening. The finding that nucleoside triphosphates regulate CFTR begins to explain why CF-associated mutations in the NBDs block Cl- channel function.     

On the interpretation of host-parasite ecology: Heligmosomoides polygyrus (Nematoda) in wild wood mouse (Apodemus sylvaticus) populations

This paper describes epidemiological and seasonal patterns in the interaction between wood mice, Apodemus sylvaticus and Heligmosomoides polygyrus. Data used in the analysis were collected by C. S. Elton and co-workers at Bagley Wood, Oxfordshire in the late 1920s. Heligmosomoides polygyrus was by far the most common helminth parasite with 70% of all wood mice infected and average intensity around 12 worms per mouse. Male and female mice were shown to harbour similar parasite burdens. Parasite numbers per host were highly overdispersed and were well described by the negative binomial distribution. There was little evidence for convexity in age (= weight)-intensity curves, either within or across sexes.
Host and parasite numbers showed predictable seasonal patterns, with mouse populations at their largest at the end of the breeding season, in August and September, and parasite populations at their largest in the late spring, around May. Results are discussed in relation to the ecology of H. polygyrus in wood and laboratory mice, and tentative comparison is made with human helminth infection. The interpretation of epidemiological patterns in these data was problematic. Of particular importance was the statistical distribution of parasites within the host population, and possible differences between mouse sexes in relation to growth, survival and trapping. Such difficulties are relevant to a range of similar field data.     

Somatic recombination rather than uniparental disomy suggested as another mechanism by which genetic imprinting may play a role in the etiology of Prader-Willi syndrome

Summary Six Prader-Willi syndrome (PWS) patients with normal karyotypes and their parents were analyzed to determine the nature of the molecular aberrations present in the proximal region of 15q and to determine the parental origin of the aberrant chromosome 15. In addition, the likehood that uniparental disomy plays a significant role in the etiology of PWS patients with normal karyotypes was studied. Restriction fragment length polymorphisms (RFLPs) recognized by seven probes [pML34 (D15S9), pTD3-21, pCGS0.9, pCGS1.1 (D15S10), IR4.3 (D15S11), IR10.1 (DS15S12), p189-1 (D15S13), IR39 (D15S18), and CMW-1 (D15S24)] mapping to the Prader-Willi chromosome region (PWCR) and an additional two probes [pMS1-14 (D15S1); the cDNA of neuromedin B] mapping elsewhere on chromosome 15 were analyzed in the six PWS patients and their parents. Copy number of each locus within the PWCR was determined by densitometry. Molecular rearrangements of the proximal region of 15q were observed in all of the six probands and the origin of the aberrant chromosome 15 when determined was consistently paternal in origin. While data obtained from our six patients does not support the mechanism of disomy, results obtained from three of the six patients show more complex rearrangements hypothesized to have resulted from somatic recombination. These rearrangements have resulted in acquired homozygosity and the lack of a paternal allele at various loci within the PWCR. The presence of only a maternal contribution at certain loci as the result of somatic recombination may be another mechanism by which genetic imprinting plays a role in the presentation of the PWS phenotype.     

Population structure,spite, and the iterated Prisoner's Dilemma

Evolutionary stability (sensu Maynard Smith: Evolution and the Theory of Games, Cambridge: Cambridge University Press, 1982) of TIT FOR TAT (TFT) under the social ecology of the iterated Prisoner's Dilemma is a function of the number of pure TFT groups (dyads) in the population, relative to the social position of a focal invading defector. Defecting against TFT always raises the defector's relative intragroup fitness; when Axelrod's (Am. Polit. Sci. Rev. 75:306–318, 1981; The Evolution of Cooperation. New York: Basic Books, 1984) Evolutionary stable strategy (ESS) conditions are met, defection also lowers the absolute fitness of the defector. Here the retaliatory (punishing) character of TFT converts defection into spite, permitting pure TFT groups to sufficiently outproduce the defector for the latter's evolutionary suppression. Increasing the relative impact of spiteful defection on a population lowers the range of evolutionary stability for TFT. When individuals participate in multiple dyads, those participating in the greatest number of dyads are most likely to provide a vehicle for the successful invasion of defection. Within social networks, ESS conditions for TFT are thus individual specific. This logic is generalized to the context of an interated n-person Prisoner's Dilemma, providing a cooperative solution conceptually identical with TFT in the two-person game.     

Negative interactions between Willamette mites and Pacific mites: possible management strategies for grapes

Willamette mites and Pacific mites are often negatively associated on grape vines. In particular, vineyards with early season infestations of the less damaging Willamette mite rarely develop high populations of the economically important Pacific mite. We report that Willamette mites had a negative effect on Pacific mite populations in both the greenhouse and field. In some experiments, the negative effect was more pronounced when vines had been damaged by previous feeding of Willamette mites and in other experiments concurrent feeding by both mite species was necessary to demonstrate a negative effect. Therefore, we cannot conclude if the mechanism of the response involves induced resistance against Pacific mites, more conventional interspecific competition, or both. Since predators were uncommon in our experiments, predator build up on Willamette mites did not cause the low Pacific mite population that we observed, although this mechanism may be important in many vineyards. Further, larger scale experiments are necesary to determine if growers can introduce Willamette mites to help control Pacific mites.

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Résumé E. willametti et T. pacificus sont souvent antagonistes dans les vignobles. En particulier, dans les vignobles contaminés tôt dans la saison par E. willametti,-dont les dégâts sont moins conséquents-, on observe rarement des populations élevées de T. pacificus, dont les dégâts sont économiquement importants. Nous avons constaté que E. Willametti a réduit les populations de T. pacificus, tant en serres qu'à l'extérieur. Dans quelques cas, l'effet antagoniste était plus marqué quand les vignes avaient été préalablement endommagées par l'alimentation de E. willametti. Dans d'autres expériences, la compétition alimentaire entre les deux espèces d'acariens était nécessaire pour produire un effet antagoniste. Cependant, nous ne pouvons pas en conclure que la réponse implique une résistance induite contre T. pacificus, ou une compétition interspécifique plus classique, ou les deux à la fois. Puisque les prédateurs étaient rares dans nos expériences, une explosion des prédateurs sur E. willametti n'a pu provoquer le faible niveau de population observé avec T. pacificus, bien qu'un tel méchanisme puisse être important dans certains vignobles. Quoi qu'il en soit, des expériences à plus grande échelle sont nécessaires pour déterminer si les vignerons peuvent introduire Eotetranychus willametti afin de limiter les populations de Tetranychus pacificus.