A selective human beta3 adrenergic receptor agonist increases metabolic rate in rhesus monkeys

Activation of beta3 adrenergic receptors on the surface of adipocytes leads to increases in intracellular cAMP and stimulation of lipolysis. In brown adipose tissue, this serves to up-regulate and activate the mitochondrial uncoupling protein 1, which mediates a proton conductance pathway that uncouples oxidative phosphorylation, leading to a net increase in energy expenditure. While chronic treatment with beta3 agonists in nonprimate species leads to uncoupling protein 1 up-regulation and weight loss, the relevance of this mechanism to energy metabolism in primates, which have much lower levels of brown adipose tissue, has been questioned. With the discovery of L-755,507, a potent and selective partial agonist for both human and rhesus beta3 receptors, we now demonstrate that acute exposure of rhesus monkeys to a beta3 agonist elicits lipolysis and metabolic rate elevation, and that chronic exposure increases uncoupling protein 1 expression in rhesus brown adipose tissue. These data suggest a role for beta3 agonists in the treatment of human obesity.     

A novel synaptobrevin/VAMP homologous protein (VAMP5) is increased during in vitro myogenesis and present in the plasma membrane

cDNA clones encoding a novel protein (VAMP5) homologous to synaptobrevins/VAMPs are detected during database searches. The predicted 102-amino acid VAMP5 harbors a 23-residue hydrophobic region near the carboxyl terminus and exhibits an overall amino acid identity of 33% with synaptobrevin/VAMP1 and 2 and cellubrevin. Northern blot analysis reveals that the mRNA for VAMP5 is preferentially expressed in the skeletal muscle and heart, whereas significantly lower levels are detected in several other tissues but not in the brain. During in vitro differentiation (myogenesis) of C2C12 myoblasts into myotubes, the mRNA level for VAMP5 is increased approximately 8- to 10-fold. Immunoblot analysis using antibodies specific for VAMP5 shows that the protein levels are also elevated approximately 6-fold during in vitro myogenesis of C2C12 cells. Indirect immunofluorescence microscopy and immunoelectron microscopy reveal that VAMP5 is associated with the plasma membrane as well as intracellular perinuclear and peripheral vesicular structures of myotubes. Epitope-tagged versions of VAMP5 are similarly targeted to the plasma membrane.     

Homocysteine and vascular disease

Teeth agenesis is an anomaly of frequent observation with a familiar heredity transmission pattern. In a frequency order the most common agenetic teeth are: third molars, upper lateral incisors and second lower bicuspids. Of exceptional rarity is the absence of second molars. Our observation confirm the heredity pattern of congenitally missing permanent teeth; in this case anomaly showed variable expressivity and different degree of severity.     

Methodological considerations in the neuropsychological study of central nervous system underarousal with a specific emphasis on coma

Several obstacles exist that impede the scientific study of pathological underarousal (stupor and coma). For instance, there is a lack of a clear, rational, standardized taxonomy with which to describe the phenomenon. Moreover, there is considerable confusion of the construct of arousal with other neurobehavioral constructs. The field also suffers from a general lack of acceptably reliable and valid instruments, especially measures of long-term outcome. Additionally, current treatments for chronic pathological underarousal are frequently presumptive and applied haphazardly, and thus do little to elucidate the process of recovery. It is suggested that biomedical and neuropsychological approaches to the study and treatment of pathological underarousal are complementary and that it is useful to view pathological underarousal as a behavioral, as well as a medical, phenomenon.     

A systematic mutation screen of 10 nuclear and 25 mitochondrial candidate genes in 21 patients with cytochrome c oxidase (COX) deficiency shows tRNA(Ser)(UCN) mutations in a subgroup with syndromal encephalopathy

COX deficiency is believed to be the most common defect in neonates and infants with mitochondrial diseases. To explore the causes of this group of disorders, we examined 25 mitochondrial genes (three COX subunit genes and 22 tRNA genes) and 10 nuclear COX subunit genes for disease associated mutations using PCR-SSCP and direct sequencing of polymorphic SSCP fragments. DNA from one patient with severe COX deficiency and with consanguineous parents was entirely sequenced. The patient population consisted of 21 unrelated index patients with mitochondrial disorders and predominant (n=7) or isolated (n=14) COX deficiency. We detected two distinct tRNA(Ser)(UCN) mutations, which have been recently described in single kindreds, in a subgroup of four patients with COX deficiency, deafness, myoclonic epilepsy, ataxia, and mental retardation. Besides a number of nucleotide variants, a single novel missense mutation, which may contribute to the disease phenotype, was found in the mitochondrial encoded COX 1 gene (G6480A). Mutations in nuclear encoded COX subunit genes were not detected in this study.     

Phantoms with 10BF3 detectors for boron neutron capture therapy applications

Two acrylic cube phantoms have been constructed for BNCT applications that allow the depth distribution of neutrons to be measured with miniature 10BF3 detectors in 0.5-cm steps beginning at 1-cm depth. Sizes and weights of the cubes are 14 cm, 3.230 kg, and 11 cm, 1.567 kg. Tests were made with the epithermal neutron beam from the patient treatment port of the Brookhaven Medical Research Reactor. Thermal neutron depth profiles were measured with a bare 10BF3 detector at a reactor power of 50 W, and Cd-covered detector profiles were measured at a reactor power of 1 kW. The resulting plots of counting rate versus depth illustrate the dependence of neutron moderation on the size of the phantom. But more importantly the data can serve as benchmarks for testing the thermal and epithermal neutron profiles obtained with accelerator-based BNCT facilities. Such tests could be made with these phantoms at power levels about five orders of magnitude lower than that required for the treatment of patients with brain tumors.     

The nematode resistance gene Mi of tomato confers resistance against the potato aphid

Resistance against the aphid Macrosiphum euphorbiae previously was observed in tomato and attributed to a novel gene, designated Meu-1, tightly linked to the nematode resistance gene, Mi. Recent cloning of Mi allowed us to determine whether Meu-1 and Mi are the same gene. We show that Mi is expressed in leaves, that aphid resistance is isolate-specific, and that susceptible tomato transformed with Mi is resistant to the same aphid isolates as the original resistant lines. We conclude that Mi and Meu-1 are the same gene and that Mi mediates resistance against both aphids and nematodes, organisms belonging to different phyla. Mi is the first example of a plant resistance gene active against two such distantly related organisms. Furthermore, it is the first isolate-specific insect resistance gene to be cloned and belongs to the nucleotide-binding, leucine-rich repeat family of resistance genes.     

Folding of circular and permuted chymotrypsin inhibitor 2: retention of the folding nucleus

The 64-residue chymotrypsin inhibitor 2 (CI2) folds by a two-state nucleation-condensation mechanism, whereby secondary and tertiary structure coalesce concomitantly in the transition state around Ala 16 in the helical N-cap. Permutation of the SH3-domain of alpha-spectrin apparently shifts its folding nucleus to another region of the protein, suggesting that a protein's transition state may be altered by altering the protein's connectivity. We have characterized the structure of the transition state of a circular and a permuted version of CI2 by a protein engineering study encompassing 11 mutations. Circular CI2 was obtained by the introduction of cysteines at residues 3 and 63 and linking them by disulfide bond formation. Subsequent cyanogen-bromide cleavage of the scissile bond, Met 40-Glu 41, yielded permuted CI2. Circular and permuted CI2 also fold according to a two-state mechanism. Permutation does not affect the folding rate constant, but circularization increases it 7-fold. The transition states of circular and permuted CI2 are essentially unchanged from that of wild-type CI2. Importantly, the folding nucleus around Ala16 is retained. These results complement a previous observation that the transition state for association of two CI2 fragments (residues 1-40 and 41-64, generated by CNBr cleavage) is very similar to the folding transition state of intact CI2. The similarity of rate constants for folding of wild-type and permuted CI2, and their value relative to that for the association of fragments, allows us to estimate the gain in entropy of activation on having the separate fragments linked: 18.3 cal M-1 K-1; i.e. an effective molarity of 10(4) M. The contrast between the retention of the folding nucleus on permutation of CI2 and its change for the SH3-domain of alpha-spectrin probably arises because the latter was cleaved in its folding nucleus whereas cleavage at sites other than 40-41 in CI2 is very destabilizing. Whether or not a folding nucleus can be changed probably depends on the specific protein and its permissivity to permutation.