COS cell expression studies of P86L, P86R, P480L and P480Q Hunter's disease-causing mutations

AbstractThree missense mutations identified in the IDS gene of our Hunter's disease patients (P86L, P480L and P480Q) and the previously described P86R mutation were expressed in COS cells to evaluate their functional consequence on iduronate-2-sulfatase (IDS) activity and processing. The 86-proline residue belongs to the highly conserved pentapeptide C-X-P-S-R in which cysteine modification to a formylglycine is required for sulfatase activity. The substitution of the 86-proline residue led to a severe mutation as no mature form was targeted to the lysosome in agreement with the severe phenotype observed in patients carrying P86L and P86R mutations. Expression studies with P480L and P480Q mutant cDNAs showed the presence of a small amount of 55 kDa mature form in the lysosomes of transfected COS cells. IDS activity of the P480L and P480Q mutants in cell extracts represents 16.6% and 5.4% of the wild-type, respectively

Early Neurodegeneration Progresses Independently of Microglial Activation by Heparan Sulfate in the Brain of Mucopolysaccharidosis IIIB Mice

BACKGROUND: In mucopolysaccharidosis type IIIB, a lysosomal storage disease causing early onset mental retardation in children, the production of abnormal oligosaccharidic fragments of heparan sulfate is associated with severe neuropathology and chronic brain inflammation. We addressed causative links between the biochemical, pathological and inflammatory disorders in a mouse model of this disease. METHODOLOGY/PRINCIPAL FINDINGS: In cell culture, heparan sulfate oligosaccharides activated microglial cells by signaling through the Toll-like receptor 4 and the adaptor protein MyD88. CD11b positive microglial cells and three-fold increased expression of mRNAs coding for the chemokine MIP1alpha were observed at 10 days in the brain cortex of MPSIIIB mice, but not in MPSIIIB mice deleted for the expression of Toll-like receptor 4 or the adaptor protein MyD88, indicating early priming of microglial cells by heparan sulfate oligosaccharides in the MPSIIIB mouse brain. Whereas the onset of brain inflammation was delayed for several months in doubly mutant versus MPSIIIB mice, the onset of disease markers expression was unchanged, indicating similar progression of the neurodegenerative process in the absence of microglial cell priming by heparan sulfate oligosaccharides. In contrast to younger mice, inflammation in aged MPSIIIB mice was not affected by TLR4/MyD88 deficiency. CONCLUSIONS/SIGNIFICANCE: These results indicate priming of microglia by HS oligosaccharides through the TLR4/MyD88 pathway. Although intrinsic to the disease, this phenomenon is not a major determinant of the neurodegenerative process. Inflammation may still contribute to neurodegeneration in late stages of the disease, albeit independent of TLR4/MyD88. The results support the view that neurodegeneration is primarily cell autonomous in this pediatric disease

Iduronate-sulfatase (Caractérisation des transcrits majoritaires, Mécanismes moléculaires à l'origine de la maladie de Hunter chez deux sujets féminins)

La maladie de Hunter, ou Mucopolysaccharidose de type II (MPSII) est une maladie lysosomale récessive liée à l'X due au déficit en iduronate sulfatase (IDS). Les différents phénotypes cliniques reflètent l'hétérogénéité des altérations affectant le gène IDS. Trois transcrits majoritaires de 5,7,5,4 et 2,1 kb sont générés par l'utilisation de différents signaux de plolyadénylation localisés dans l'exon IX et codent donc pour la même protéine. L'absence de corrélation entre la quantité des différents transcrits, l'activité enzymatique et la quantité de protéine IDS suggère que c'est probablement la maturation de l'IDS qui est régulée plutôt que la transcription. L'exploration moléculaire de deux filles atteintes de la maladie de Hunter a été réalisée. S.D. présente un réarrangement génique survenu de novo et une inactivation préférentielle de l'X maternel normal. C.S. est le premier cas féminin de MPS II avec deux gènes mutés. La mutation L41P a été identifiée à l'état homozygote.DIJON-BU Sciences Economie (212312102) / SudocSudocFranceF

Le psychodrame, les langues de chat et l’Amaryllis- Interview de Lucien Kroll

info:eu-repo/semantics/publishedDe la participation urbaine. La place Flage

Dosage du globotriaosylcéramide dans l’urine

La maladie de Fabry est une maladie de surchargedue à un déficit de l’α-galactosidase A.Un traitement par enzyme de substitution estdésormais possible. Le dosage du globotriaosylcéramidedans l’urine peut être effectué parspectrométrie de masse en tandem (MS/MS).Cette technique, particulièrement sensible etspécifique, est décrite, ainsi que les résultatschez des patients atteints, des femmes hétérozygoteset des malades traités par enzyme desubstitution

Liver circadian clock, a pharmacologic target of cyclin-dependent kinase inhibitor seliciclib.

Circadian disruption accelerates malignant growth and shortens survival, both in experimental tumor models and cancer patients. In previous experiments, tumor circadian disruption was rescued with seliciclib, an inhibitor of cyclin-dependent kinases (CDKs). This effect occurred at a selective dosing time and was associated with improved antitumor activity. In the current study, seliciclib altered robust circadian mRNA expression of the clock genes Rev-erb alpha, Per2, and Bmal1 in mouse liver following dosing at zeitgeber time (ZT) 3 (i.e., 3 h after the onset of the 12 h light span), when mice start to rest, but not at ZT19, near the middle of the 12 h dark span, when mice are most active. However, liver exposure to seliciclib, as estimated by the liver area under the concentration x time curve (AUC), was approximately 80% higher at ZT19 than at ZT3 (p = 0.049). Circadian clock disruption was associated with increased serum liver enzymes and modified glycogen distribution in hepatocytes, as revealed by biochemical determinations and optic and electronic microscopy. The extent of increase in liver enzymes was most pronounced following dosing at ZT3, as compared to ZT19 (p < 0.04). Seliciclib further up-regulated the transcriptional activity of c-Myc, a cell cycle gene that promotes cell cycle entry and G1-S transition (p < 0.001), and down-regulated that of Wee1, which gates cell cycle transition from G2 to M (p < 0.001). These effects did not depend upon drug dosing time. Overall, the results suggest the circadian time of seliciclib delivery is more critical than the amount of drug exposure in determining its effects on the circadian clock. Seliciclib-induced disruption of the liver molecular clock could account for liver toxicity through the resulting disruption of clock-controlled detoxification pathways. Modifications of cell cycle gene expression in the liver likely involve other mechanisms. Circadian clocks represent relevant targets to consider for optimization of therapeutic schedules of CDK inhibitors