Rapid Cohort Generation and Analysis of Disease Spectrum of Large Animal Model of Cone Dystrophy

Large animal models are an important resource for the understanding of human disease and for evaluating the applicability of new therapies to human patients. For many diseases, such as cone dystrophy, research effort is hampered by the lack of such models. Lentiviral transgenesis is a methodology broadly applicable to animals from many different species. When conjugated to the expression of a dominant mutant protein, this technology offers an attractive approach to generate new large animal models in a heterogeneous background. We adopted this strategy to mimic the phenotype diversity encounter in humans and generate a cohort of pigs for cone dystrophy by expressing a dominant mutant allele of the guanylate cyclase 2D (GUCY2D) gene. Sixty percent of the piglets were transgenic, with mutant GUCY2D mRNA detected in the retina of all animals tested. Functional impairment of vision was observed among the transgenic pigs at 3 months of age, with a follow-up at 1 year indicating a subsequent slower progression of phenotype. Abnormal retina morphology, notably among the cone photoreceptor cell population, was observed exclusively amongst the transgenic animals. Of particular note, these transgenic animals were characterized by a range in the severity of the phenotype, reflecting the human clinical situation. We demonstrate that a transgenic approach using lentiviral vectors offers a powerful tool for large animal model development. Not only is the efficiency of transgenesis higher than conventional transgenic methodology but this technique also produces a heterogeneous cohort of transgenic animals that mimics the genetic variation encountered in human patients.     

Invasive aquatic pets: failed policies increase risks of harmful invasions

Businesses in the pet trade collect and transport many aquatic species around the globe, and some of these individuals are released into new habitats. Some jurisdictions have introduced laws intended to regulate this trade, but these regulations have rarely had the desired effects. Laws regarding pets and the pet trade are often poorly communicated, poorly enforced, and not aligned with hobbyists’ beliefs. Consequently, some laws may increase the number of unwanted introductions instead of decreasing them. A significant change in approach is needed, involving far greater communication with scientists, administrations, politicians, the pet industry, and pet owners, promoting euthanasia of unwanted pets rather than release, and the creation and promotion “white lists” of low risk species that can be sold in the pet trade.     

Two Novel Genes Rapidly and Transiently Activated in Suspension-Cultured Rice Cells by Treatment with N-Acetylchitoheptaose, a Biotic Elicitor for Phytoalexin Production

By using subtracted probes, two cDNA clones of rice, EL2 andEL3, were isolated as genes responsive within 6 min to N-acetylchitoheptaose,a potent biotic elicitor for phytoalexin biosynthesis. Analysesof the sequence of the cDNAs showed that both of EL2 and EL3encoded basic proteins with no significant similarities to thoseof known genes. (Received April 18, 1996; Accepted May 7, 1996)     

Improved synthesis of a protected 11-oxoestrone

An improved synthesis of 11-oxoestrone-3-acetate-17-ethyleneketal is reported. Adjustments are proposed for the oxidation of estrone by 2,3-dichloro-5,6-dicyano-1,4-benzoquinone into 9(11)-dehydroestrone. A complete hydroboration-oxidation of the resulting ketal, by means of borane-methylsulfide complex, gives the corresponding 11-hydroxy derivative. This latter compound is then acetylated for successful oxidation with pyridinium chlorochromate on alumina. The overall yield is 30%.     

Distribution of enteric neural peptide YY in the dog gastrointestinal tract

Various regions of the dog gastrointestinal tract were investigated for the distribution of peptide YY (PYY) neurons using immunocytochemistry and radioimmunoassay. PYY neurons that encircled non-PYY-immunoreactive neurons were mainly observed in the myenteric plexus from the stomach to the colon. There was more PYY-like immunoreactivity in the muscle layer of the stomach and ileum than in the other intestines. The results of high performance liquid chromatography revealed that neural PYY-immunoreactive substance is identical to authentic PYY. PYY was not localized in the cholinergic neurons. These results indicate that PYY, as a neuropeptide, is involved in the regulation of gastrointestinal function.     

Metabolism of [13C]-labelled photosynthate in plant cytosol and bacteroids of root nodules of Glycine max

Well-nodulated soybean ( Glycine max L. Merr. cv. Akisengoku) plants were allowed to assimilate ¹³CO₂. Plant cytosol and bacteroid fractions were isolated from nodules, and the kinetics of [¹³C]-labelling of soluble carbohydrates, organic acids and amino acids were investigated.
The concentrations of all metabolites, with the exception of trehalose and 3-hydroxy-butyrate, were 10- to 1000-fold higher in plant cell cytosol than in bacteroids. The major portion of trehalose was found in bacteroids and 3-hydroxybutyrate only in bacteroids. Sucrose was most highly labelled with ¹³C in nodules, and the levels and time-course of labelling of sucrose were in good agreement with those of respired CO₂ from the nodules. The levels and time-courses of labelling of sucrose were closely similar in cytosol and bacteroids. Glucose was less labelled than sucrose and the level of labelling was consistently higher in cytosol than in bacteroids. The levels of [¹³C]-labelling of organic acids and amino acids in nodules were lower than those of sucrose and of respired CO₂. Tricarboxylic acid cycle intermediates, particularly succinate, were considerably less labelled in bacteroids than in the cytosol. All amino acids detected were also much more rapidly labelled in the cytosol. The results are discussed in relation to the utilization and possible compartmentation of carbon substrates in nodule tissues.     

Accumulation of Rare Phytosterols in Plant Cells on Treatment with Metabolic Inhibitors and Mevalonic Acid

Root application of the metabolic inhibitors D-ethionine (100µM) and L-ethionine (30 µM), which were expectedto inhibit alkylation of the sterol side chains, merely reducedthe sterol content in the roots of Medicago sativa seedlings.The major sterol was stigmasterol. However, when (3RS)-mevalonicacid (2 mM) was applied together with ethionine, cydoartenol(about 50% of the total sterols) accumulated in the roots. Thehypocotyls of the ethionine-treated seedlings accumulated cholesterol(34% of the total sterols), and mevalonic acid showed no additionaleffect in this case. In a suspension culture of Nicotiana tabacum, the most abundantsterol was campesterol. When cells were treated with buthiobate(100 µM), a potent inhibitor of lanosterol 14-demethylationin yeasts and fungi, obtusifoliol accumulated in the treatedcells (50% of the total sterols). 14-Methylfecosterol also accumulatedin the treated cells. The addition of mevalonic acid (1 mM togetherwith buthiobate increased the obtusifoliol content (63% of thetotal sterols). (Received October 16, 1986; Accepted April 2, 1987)     

Variation in the Content and Composition of Sterols in Alfalfa Seedlings Treated with Compactin (ML-236B) and Mevalonic Acid

Compactin (ML-236B), a specific inhibitor of 3-hydroxy-3-methylglutarylCoA reductase, inhibited the elongation of roots and hypocotylsof Medicago sativa seedlings when it was applied to the roots.Addition of mevalonic acid, the direct product of the enzyme,together with compactin relieved the growth inhibition of roots. The contents and compositions of sterols were studied in threeparts of M. sativa seedlings—roots, hypocotyls and cotyledons.Compactin (20 µM) decreased the sterol contents of rootsand hypocotyls by about a half but did not affect that of cotyledons.On the other hand, mevalonic acid (2 mM) increased the sterolcontent of roots more than threefold the nontreated controllevel but not the contents of hypocotyls and cotyledons. Mevalonicacid added in combination with compactin had a similar effecton the sterol content of roots as when it was added alone. The major sterol in all three parts was stigmasterol whetheror not compactin or mevalonic acid was present. However, thevariation of the sterol composition in the roots was distinct;mevalonic acid-treated roots markedly accumulated ⁷-sitosterol,24-methylenecycloartanol and squalene. (Received October 16, 1986; Accepted April 2, 1987)