Cytogenetically marked clones in human fibroblasts cultured from normal subjects.

Clones of cytogenetically abnormal cells have been recognized in fibroblasts cultured from normal human adult skin. No such clones have been observed in human embryo skin fibroblasts cultured in the same way. Although the culture conditions may have played some part in the emergence of these clones, it is possible that the abnormal cells from which the clones were derived were present in vivo.     

Whole body elimination routes of gold in humans after a single-dose application of the antirheumatic Auranofin

After removal of the gallbladder (cholecystectomy) 7 patients were administered the antirheumatic agent Auranofin in its commercially available tablet form (Ridaura) 3 d postoperative. The single dose consisted of 5 tablets (4.35 mg of gold). Gold was determined in samples of blood, plasma, urine, bile, and feces (1–2 mL and 2.5 g, respectively). The specimens were drawn 10 min to 8 d p.a. and on d 14 p.a. The determinations were performed by instrumental neutron activation analysis (INAA). The limit of detection was less than 1 ng of gold. The courses of the mean gold concentrations show maxima after 1.9 h in blood and plasma, and after 16 h in urine and bile. The mean half-lives (terminal phases) are 7.6 d (blood), 23 d (plasma), and 6.5 d (bile).     

Isolation, characterization, and chromosomal mapping of mouse P450 17 alpha-hydroxylase/C17-20 lyase.

Cytochrome P450 17 alpha-hydroxylase/C17-20 lyase (P45017 alpha) catalyzes the conversion of C-21 steroids to C-19 steroids in gonads. A full-length mouse cDNA encoding P450 17 alpha was isolated from a mouse Leydig cell library and characterized by restriction mapping and sequencing. The predicted amino acid sequence has 83% homology to rat, 66% homology to human, and 62% homology to bovine P45017 alpha amino acid sequences. The protein is 507 amino acids in length, which is 1 amino acid shorter than the human protein and 2 amino acids shorter than the bovine protein. The structural gene encoding P450 17 alpha (Cyp17) was localized utilizing an interspecific testcross to mouse chromosome 19, distal to Got-1. Another cytochrome P450, P4502c (Cyp2c), also is located at the distal end of chromosome 19. CYP17, CYP2c, and GOT1 have been mapped to human chromosome 10, with CYP2C and GOT1 mapped to the distal region, q24.3 and q25.3, respectively. The data in the present study indicate conserved syntenic loci on mouse chromosome 19 and human chromosome 10 and predict that the structural gene encoding P45017 alpha will be found distal to GOT1 on human chromosome 10.     

cis-6-oxo-hexahydro-2-oxa-1,4-diazapentalene and cis-6-oxo-hexahydropyrrolo[3,2-c]pyrazole based scaffolds: design rationale, synthesis and cysteinyl proteinase inhibition

The 5,5-bicycles cis-6-oxo-hexahydro-2-oxa-1,4-diazapentalene 3 and cis-6-oxo-hexahydropyrrolo[3,2-c]pyrazole 4 were designed as rotationally restricted templates towards the preparation of inhibitors of CAC1 cysteinyl proteinases. The design strategy was exemplified through the solution and solid phase preparation of potent inhibitors of human cathepsin K and may potentially be applied to inhibitors of other CAC1 proteinases.     

BCG Vaccination Protects against Experimental Viral Infection in Humans through the Induction of Cytokines Associated with Trained Immunity

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Growth and Muscle Defects in Mice Lacking Adult Myosin Heavy Chain Genes

The three adult fast myosin heavy chains (MyHCs) constitute the vast majority of the myosin in adult skeletal musculature, and are >92% identical. We describe mice carrying null mutations in each of two predominant adult fast MyHC genes, IIb and IId/x. Both null strains exhibit growth and muscle defects, but the defects are different between the two strains and do not correlate with the abundance or distribution of each gene product. For example, despite the fact that MyHC-IIb accounts for >70% of the myosin in skeletal muscle and shows the broadest distribution of expression, the phenotypes of IIb null mutants are generally milder than in the MyHC-IId/x null strain. In addition, in a muscle which expresses both IIb and IId/x MyHC in wild-type mice, the histological defects are completely different for null expression of the two genes. Most striking is that while both null strains exhibit physiological defects in isolated muscles, the defects are distinct. Muscle from IIb null mice has significantly reduced ability to generate force while IId null mouse muscle generates normal amounts of force, but has altered kinetic properties. Many of the phenotypes demonstrated by these mice are typical in human muscle disease and should provide insight into their etiology.     

A synthetic operon containing 14 bovine pancreatic trypsin inhibitor genes is expressed in E. coli.

A synthetic gene encoding the protein sequence of mature bovine pancreatic trypsin inhibitor (BPTI) has been cloned into a novel E. coli expression vector. After in vitro gene amplification by successive DNA duplications, more than 600 000 mostly inactive inhibitor molecules may be recovered from a single cell. After purification the inhibitory activity can be reconstituted almost completely. The specificity of BPTI for trypsin is abolished by a single amino acid exchange from lysine to isoleucine at position 15. The altered protein is shown to be an efficient inhibitor of human leukocyte elastase.     

The binding of 1,10-phenanthroline to specifically active-site cobalt(II)-substituted horse-liver alcohol dehydrogenase. A probe for the open-enzyme conformation

We have studied the binding of 1,10-phenanthroline to specifically active-site cobalt(II)-substituted horse-liver alcohol dehydrogenase [Co(II)-LADH]. The dissociation constant is a factor of 6500 smaller than in the native enzyme. Spectral evidence is given which shows that 1,10-phenanthroline does not remove the catalytic Co(II) ion and that binding of 1,10-phenanthroline renders the catalytic metal ion pentacoordinate. The maximum limiting rate constant for the association of 1,10-phenanthroline to Co(II)-LADH is about 60 s-1. This is about a third of the value (169 s-1) determined for native horse-liver alcohol dehydrogenase, Zn(II)LADH [Frolich et al. (1978) Arch. Biochem. Biophys. 189, 471-480]. For cadmium(II)-substituted horse-liver alcohol dehydrogenase, [Cd(II)LADH] the maximum limiting rate constant for association of 1,10-phenanthroline increased to 590 s-1. These findings demonstrate that the rate-limiting step is strongly dependent on the chemical nature of the catalytic metal ion and its immediate environment. 1,10-Phenanthroline is shown to bind to the Co(II)-LADH.NAD+ complex in the open conformation. The maximum limiting rate constant remains unchanged in the presence of NAD+. The data have been used to derive a kinetic scheme for the formation of ternary complexes including NAD+ that involves a slow intermediary step.