RESUMO
Surface enhanced laser desorption/ionisation time of flight (SELDI-TOF) mass spectrometry has been used to search for new protein biomarkers in the plasma of patients with mucopolysacharidoses (MPS). Differences in the levels of some plasma proteins, particularly the apolipoprotein ApoCI, were observed between MPS patients and normal controls, using the different chromatographic surfaces (ProteinChips). ApoCI was identified by both its mass and by immunological techniques. In plasma, it exists in two forms, ApoCI and a truncated form which lacks two N-terminal amino acids, ApoCI'. In controls, the ratio of ApoCI':ApoCI observed using the cation-exchange surface (CM10) was approximately 1:2 whereas in most MPS patients it varied from 1:1 to 1:0.8. The ratio of ApoCI':ApoCI in plasma is determined by the activity of dipeptidyl peptidase IV, DPP-IV (also known as the leucocyte antigen CD26), which was found to be elevated up to 3-fold in MPS patients. The DPP-IV activity decreased in MPS I patients undergoing enzyme replacement therapy, indicating that it could be a useful biomarker for monitoring the efficacy of treatment in MPS disease. As DPP-IV has an important regulatory role in metabolism, it is possible that its elevation could cause some of the secondary pathology in MPS, and inhibition of DPP-IV might have a role in MPS therapy.
Assuntos
Dipeptidil Peptidase 4/sangue , Mucopolissacaridoses/sangue , Mucopolissacaridoses/enzimologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Adolescente , Apolipoproteína C-I/sangue , Biomarcadores/sangue , Proteínas Sanguíneas/metabolismo , Estudos de Casos e Controles , Humanos , Mucopolissacaridoses/terapiaRESUMO
Based on our preliminary observation of abnormal glycosylation in a cutis laxa patient, nine cutis laxa patients were analyzed for congenital defects of glycosylation (CDG). Isoelectric focusing of plasma transferrin and apolipoproteinC-III showed that three out of nine patients had a defect in the biosynthesis of N-glycans and core 1 mucin type O-glycans, respectively. Mass spectrometric N-glycan analyses revealed a relative increase of glycans lacking sialic acid and glycans lacking sialic acid and galactose residues. Mutation analysis of the fibulin-5 gene (FBLN5), which has been reported in cases of autosomal recessive cutis laxa, revealed no mutations in the patients' DNA. Evidence is presented that extracellular matrix (ECM) proteins of skin are likely to be highly glycosylated with N- and/or mucin type O-glycans by using algorithms for predicting glycosylation. The conclusions in this study were that the clinical phenotype of autosomal recessive cutis laxa seen in three patients is not caused by mutations in the FBLN5 gene. Our findings define a novel form of CDG with cutis laxa and neurological involvement due to a defect in the sialylation and/or galactosylation of N- and O-glycans. Improper glycosylation of ECM proteins of skin may form the pathophysiological basis for the cutis laxa phenotype.
Assuntos
Erros Inatos do Metabolismo dos Carboidratos/genética , Cútis Laxa/genética , Doenças do Sistema Nervoso/genética , Doenças do Sistema Nervoso/patologia , Polissacarídeos/biossíntese , Apolipoproteínas C/metabolismo , Pré-Escolar , Consanguinidade , Cútis Laxa/metabolismo , Cútis Laxa/patologia , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Genes Recessivos , Glicosilação , Humanos , Lactente , Focalização Isoelétrica , Espectrometria de Massas , Linhagem , Polissacarídeos/química , Transferrina/metabolismoRESUMO
We report a 12-year-old boy with a vacuolar myopathy with clinical and histologic features of X-linked myopathy with excessive autophagy. This is a rare and slowly progressive disease of skeletal muscle without cardiac, nervous system, or other organ involvement. The differential diagnosis of vacuolar myopathy includes acid maltase deficiency, Danon disease, and X-linked myopathy with excessive autophagy.
Assuntos
Autofagia , Doenças Genéticas Ligadas ao Cromossomo X/patologia , Doenças Musculares/patologia , Vacúolos , Criança , Humanos , Proteína 2 de Membrana Associada ao Lisossomo , Proteínas de Membrana Lisossomal/genética , Masculino , Doenças Musculares/genéticaRESUMO
Sanfilippo syndrome type D is an autosomal recessive lysosomal storage disease that is caused by a deficiency of N-acetylglucosamine-6-sulphatase, one of the enzymes involved in the catabolism of heparan sulphate. Only 15 patients have been described in the literature and just two mutations have been reported to date. We present the clinical, biochemical and molecular analysis of two Italian Sanfilippo D families. Novel homozygous mutations were identified in the affected patients from each family: a large intragenic deletion of 8723 bp encompassing exons 2 and 3 in family 1 and a nonsense mutation, Q272X, in family 2. The deletion is the first large intragenic deletion to be reported in any of the four Sanfilippo subtypes, including Sanfilippo type C in which the gene has recently been identified.
Assuntos
Códon sem Sentido , Mucopolissacaridose III/enzimologia , Mucopolissacaridose III/genética , Deleção de Sequência , Sequência de Bases , Criança , Glicosaminoglicanos/urina , Humanos , Itália , Masculino , Mucopolissacaridose III/urina , Sulfatases/deficiência , Sulfatases/genética , Sulfatases/metabolismoRESUMO
Conditions under which the glycosylation capacity of cells is limited provide an opportunity for studying the efficiency of site-specific glycosylation and the role of glycosylation in the maturation of glycoproteins. Congenital disorders of glycosylation type 1 (CDG-I) provide such a system. CDG-I is characterized by underglycosylation of glycoproteins due to defects in the assembly or transfer of the common dolichol-pyrophosphate-linked oligosaccharide precursor of asparagine-linked glycans. Human plasma alpha1-antitrypsin is normally fully glycosylated at three asparagine residues (46, 83, and 247), but un-, mono-, di-, and fully glycosylated forms of alpha1-antitrypsin were detected by 2D PAGE in the plasma from patients with CDG-I. The state of glycosylation of the three asparagine residues was analyzed in all the underglycosylated forms of alpha1-antitrypsin by peptide mass fingerprinting using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. It was found that asparagine 46 was always glycosylated and that asparagine 83 was never glycosylated in the underglycosylated glycoforms of alpha1-antitrypsin. This showed that the asparagine residues are preferentially glycosylated in the order 46>247>83 in the mature underglycosylated forms of alpha1-antitrypsin found in plasma. It is concluded that the nonoccupancy of glycosylation sites is not random under conditions of decreased glycosylation capacity and that the efficiency of glycosylation site occupancy depends on structural features at each site. The implications of this observation for the intracellular transport and sorting of glycoproteins are discussed.