Distribution of mutations in the PEX gene in families with X-linked hypophosphataemic rickets (HYP)

Mutations in the PEX gene at Xp22.1 (phosphate-regulating gene with homologies to endopeptidases, on the X-chromosome), are responsible for X-linked hypophosphataemic rickets (HYP). Homology of PEX to the M13 family of Zn2+ metallopeptidases which include neprilysin (NEP) as prototype, has raised important questions regarding PEX function at the molecular level. The aim of this study was to analyse 99 HYP families for PEX gene mutations, and to correlate predicted changes in the protein structure with Zn2+ metallopeptidase gene function. Primers flanking 22 characterised exons were used to amplify DNA by PCR, and SSCP was then used to screen for mutations. Deletions, insertions, nonsense mutations, stop codons and splice mutations occurred in 83% of families screened for in all 22 exons, and 51% of a separate set of families screened in 17 PEX gene exons. Missense mutations in four regions of the gene were informative regarding function, with one mutation in the Zn2+-binding site predicted to alter substrate enzyme interaction and catalysis. Computer analysis of the remaining mutations predicted changes in secondary structure, N-glycosylation, protein phosphorylation and catalytic site molecular structure. The wide range of mutations that align with regions required for protease activity in NEP suggests that PEX also functions as a protease, and may act by processing factor(s) involved in bone mineral metabolism.      

Alternative splicing of exon 14 determines nuclear or cytoplasmic localisation of fmr1 protein isoforms

Impaired expression of the FMR1 gene is responsible for the fragile X mental retardation syndrome. The FMR1 gene encodes a cytoplasmic protein with RNA-binding properties. Its complex alternative splicing leads to several isoforms, whose abundance and specific functions in the cell are not known. We have cloned in expression vectors, cDNAs corresponding to several isoforms. Western blot comparison of the pattern of endogenous FMR1 proteins with these transfected isoforms allowed the tentative identification of the major endogenous isoform as ISO 7 and of a minor band as an isoform lacking exon 14 sequences (ISO 6 or ISO 12), while some other isoforms (ISO 4, ISO 5) were not expressed at detectable levels. Surprisingly, in immunofluorescence studies, the transfected splice variants that exclude exon 14 sequences (and have alternate C-terminal regions) were shown to be nuclear. Such differential localisation was however not seen in subcellular fractionation studies. Analysis of various deletion mutants suggests the presence of a cytoplasmic retention domain encoded in exon 14 and of a nuclear association domain encoded within the first eight exons that appear however to lack a typical nuclear localisation signal.      

Combination drug therapy in children with exercise-induced bronchospasm

Fifty-three children with moderately severe asthma were studied to determine the capacity of drugs to block exercise-induced bronchospasm (EIB). All 53 children demonstrated EIB (as defined by a 20% fall in FEV1 or 30% fall in FEF25-75) while receiving therapeutic theophylline (serum level 10-20 micrograms/ml). EIB was completely blocked in 47 children by an inhaled metaproterenol 10 minutes prior to exercise, in addition to the theophylline. In six children, EIB was only partially blocked when either metaproterenol or cromolyn was added to the theophylline, but was completely blocked when all three drugs were used. A small group of children may benefit from combination therapy for EIB.     

Monoamine distribution in primate brain—IV. Indoleamine-containing perikarya in the brain stem of Macaca arctoides

The histochemical fluorescence technique for the demonstration of monoamines in the central nervous system was employed to assess the distribution of serotonin-containing neurons within the brain stem of the immature and adult stump-tailed macaque (Macaco arctoides). Microspectrofluorometric analysis was performed in order to verify the existence of serotonin within perikarya which contained yellow histofluorescence. Serotonin-containing perikarya were found within raphe nuclei including nucleus raphe-pallidus, -obscurus, -pontis, -magnus, -dorsalis, and -centralis superioralis. Serotoninergic perikarya did not appear confined exclusively to the raphe, but were observed in the reticular formation and other brain stem nuclei including the locus coeruleus and nucleus subcoeruleus. Serotoninergic cells were not seen within the brain stem at superior collicular levels.The localization of Serotoninergic perikarya in regions other than the raphe nuclei presents certain dissimilarities in relation to patterns reported in other mammalian species.     

DNA sequence analysis of the apocytochrome b gene of Podospora anserina: a new family of intronic open reading frame

Summary The 5,969 by (base pair) DNA sequence of the apocytochrome b mitochondrial (mt) gene of race A Podospora anserina was located in a 8.5 Kbp region. This gene contained a 2,499 by subgroup IB and a 1,306 by subgroup ID intron as well as a 990 bp subgroup IB intron which is present in race A but not race s. The large subgroup IB intron and the race A specific IB intron both contained potential alternate splice sites which brought their open reading frames into phase with their upstream exon sequences. All three introns were compared with regard to their secondary structures and open reading frames to the other 30 group I introns in Podospora anserina, as well as to other fungal introns. We detected a new family of intronic ORFs comprising seven P. anserina introns, several N. crassa introns, as well as the T4td bacteriophage intron. Sequence similarities to intron-encoded endonucleases were noteworthy. The DNA sequences reported here and in the accompanying paper complete the analysis of race s and race A mitochondrial DNA.     

Obliterative muscularization of the small bowel submucosa in Crohn disease: a possible mechanism of small bowel obstruction

CONTEXT: The pathology of small bowel obstruction in Crohn disease has not been studied extensively. Stricture formation has been attributed mainly to fibrosis, although muscularization of the submucosa has been discussed previously. OBJECTIVE: To identify additional pathologic changes in Crohn disease that could be involved in the formation of strictures. DESIGN: We reviewed 50 ileal resections from patients with Crohn disease. The histopathologic slides were reviewed initially without knowledge of the macroscopic or clinical findings. We identified an unusual muscular proliferation that we refer to as obliterative muscularization of the submucosa, defined as a thick and continuous muscle layer from the mucosal base to the muscularis propria that is at least 1 cm in length. Subsequently, histopathologic findings were correlated with macroscopic and clinical findings. RESULTS: Obliterative muscularization of the submucosa was present in 14 specimens, and in 11 of these 14 it was topographically restricted to strictures. Submucosal fibrosis was observed in sections from adjacent regions. Obliterative muscularization of the submucosa, including thick-walled vessels and hyperplastic nerves but not prominent scarring, was more common in specimens with strictures; the difference was statistically significant (P <.001). CONCLUSIONS: Obliterative muscularization of the submucosa may be pathogenetically involved in the formation of strictures either directly by causing a sustained spasm, or indirectly by minimizing the vasoprotective role of the submucosa, impairing repair and enhancing scarring.     

Prevalence of antibody to human immunodeficiency virus and hepatitis B surface antigen in blood samples submitted to a hospital laboratory. Implications for handling specimens

The prevalence of hepatitis B surface antigen (HBsAg) and antibody to human immunodeficiency virus (HIV) was determined in serum or plasma specimens of 506 patients submitted to the clinical chemistry laboratory of an urban teaching hospital, and the results were correlated with "biohazard" warning labels on the specimens. Hepatitis B surface antigen, HIV antibody, or either of these were present in 32 (6.3%), 15 (3.0%), and 44 specimens (8.7%), respectively. Ten (67%) of 15 specimens with HIV antibody and nine (28%) of 32 with HBsAg bore biohazard labels. Among 473 unlabeled specimens, HIV antibody was present in five (1.1%), HBsAg was present in 23 (4.9%), and 27 (5.7%) contained either or both of these markers. All clinical and laboratory personnel should be vaccinated against hepatitis B and should handle all blood specimens as if they were infected, regardless of biohazard labeling. By fostering complacency in handling unlabeled specimens, the use of biohazard labels may paradoxically increase the risk that health care workers will be exposed to HIV and hepatitis B virus.     

Occurrence of circulating 7-deoxyaglycone metabolites of 4′-deoxydoxorubicin in man

Summary In five cancer patients we have determined the pharmacokinetics of 4-deoxydoxorubicin (4-DOX), its alcoholic metabolite 4-deoxydoxorubicinol and the occurrence of circulating 7-deoxyaglycone metabolites. The 7-deoxyaglycone of the alcohol metabolite, the major aglycone of Adriamycin (ADR) present in man, was not detected in any serum sample. The 7-deoxyaglycone of the parent drug, which appears in concentrations in excess of 30ng/ml after ADR administration, was detected in only 2/5 patients in trace amounts. These preliminary data indicate a difference in biotransformation between ADR and 4-DOX despite their close structural similarities.