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Biomedical subjects

A J Reuser

Publications and source records attributed to A J Reuser.

At least 19 recordsLinked to original sources
PubMed2001

TSC2 missense mutations inhibit tuberin phosphorylation and prevent formation of the tuberin-hamartin complex.

Tuberous sclerosis (TSC) is an autosomal dominant disorder characterized by a broad phenotypic spectrum that includes seizures, mental retardation, renal dysfunction and dermatological abnormalities. Inactivating mutations to either of the TSC1 and TSC2 tumour suppressor genes are responsible for the disease. TSC1 and TSC2 encode two large novel proteins called hamartin and tuberin, respectively. Hamartin and tuberin interact directly with each other and it has been reported that tuberin may act as a chaperone, preventing hamartin self-aggregation and maintaining the tuberin-hamartin complex in a soluble form. In this study, the ability of tuberin to act as a chaperone for hamartin was used to investigate the tuberin-hamartin interaction in more detail. A domain within tuberin necessary for the chaperone function was identified, and the effects of TSC2 missense mutations on the tuberin-hamartin interaction were investigated to allow specific residues within the central domain of tuberin that are important for the interaction with hamartin to be pin-pointed. In addition, the results confirm that phosphorylation may play an important role in the formation of the tuberin-hamartin complex. Although mutations that prevent tuberin tyrosine phosphorylation also inhibit tuberin-hamartin binding and the chaperone function, our results indicate that only hamartin is phosphorylated in the tuberin-hamartin complex.

Amino Acid Substitution↗
PubMed2001

Cardiac remodeling and contractile function in acid alpha-glucosidase knockout mice.

Pompe's disease is an autosomal recessive and often fatal condition, caused by mutations in the acid alpha-glucosidase gene, leading to lysosomal glycogen storage in heart and skeletal muscle. We investigated the cardiac phenotype of an acid alpha-glucosidase knockout (KO) mouse model. Left ventricular weight-to-body weight ratios were increased 6.3 +/- 0.8 mg/g in seven KO compared with 3.2 +/- 0.2 mg/g in eight wild-type (WT) mice (P < 0.05). Echocardiography under ketamine-xylazine anesthesia revealed an increased left ventricular (LV) wall thickness (2.17 +/- 0.16 in KO vs. 1.18 +/- 0.10 mm in WT mice, P < 0.05) and a decreased LV lumen diameter (2.50 +/- 0.32 in KO vs. 3.21 +/- 0.14 mm in WT mice, P < 0.05), but LV diameter shortening was not different between KO and WT mice. The maximum rate of rise of left ventricular pressure (LV dP/dt(max)) was lower in KO than in WT mice under basal conditions (2,720 +/- 580 vs. 4,440 +/- 440 mmHg/s) and during dobutamine infusion (6,220 +/- 800 vs. 8,730 +/- 790 mmHg/s, both P < 0.05). Similarly, during isoflurane anesthesia LV dP/dt(max) was lower in KO than in WT mice under basal conditions (5,400 +/- 670 vs. 8,250 +/- 710 mmHg/s) and during norepinephrine infusion (10,010 +/- 1,320 vs. 14,710 +/- 220 mmHg/s, both P < 0.05). In conclusion, the markedly increased LV weight and wall thickness, the encroachment of the LV lumen, and LV dysfunction reflect cardiac abnormalities, although not as overt as in humans, of human infantile Pompe's disease and make these mice a suitable model for further investigation of pathophysiology and of novel therapies of Pompe's disease.

Anesthetics, Inhalation↗
PubMed2001

Enzyme therapy for pompe disease with recombinant human alpha-glucosidase from rabbit milk.

Pompe disease is a metabolic myopathy caused by deficiency of lysosomal acid alpha-glucosidase. In this report we review the first 36 weeks of a clinical study on the safety and efficacy of enzyme therapy aimed at correcting the deficiency. Four patients with infantile Pompe disease were enrolled. They received recombinant human alpha-glucosidase from transgenic rabbit milk. The product is generally well tolerated and reaches the primary target tissues. Normalization of alpha-glucosidase activity in skeletal muscle was obtained and degradation of PAS-positive material was seen in tissue sections. The clinical condition of all patients improved. The effect on heart was most significant, with an impressive reduction of the left ventricular mass index (LVMI). Motor function improved. The positive preliminary results stimulate continuation and extension of efforts towards the realization of enzyme therapy for Pompe disease.

Animals↗
PubMed2001

Analysis of TSC2 stop codon variants found in tuberous sclerosis patients.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder caused by mutations to the TSC1 and TSC2 tumour suppressor genes. We detected two sequence changes involving the TSC2 stop codon and investigated the effects of these changes on the expression of tuberin, the TSC2 gene product, and on the binding between tuberin and the TSC1 gene product, hamartin. While elongation of the tuberin open reading frame by 17 amino acids did not interfere with tuberin-hamartin binding, a longer extension prevented this interaction. Our data illustrate how functional protein assays can assist in the verification and characterisation of disease-causing mutations.

Animals↗
PubMed2000

Recombinant human alpha-glucosidase from rabbit milk in Pompe patients.

Pompe's disease is a fatal muscular disorder caused by lysosomal alpha-glucosidase deficiency. In an open-label study, four babies with characteristic cardiomyopathy were treated with recombinant human alpha-glucosidase (rhGAA) from rabbit milk at starting doses of 15 mg/kg or 20 mg/kg, and later 40 mg/kg. The enzyme was generally well tolerated. Activity of alpha-glucosidase normalised in muscle. Tissue morphology and motor and cardiac function improved. The left-ventricular-mass index decreased significantly. We recommend early treatment. Long-term effects are being studied.

Animals↗
PubMed2000

Allosteric activation of acid alpha-glucosidase by the human papillomavirus E7 protein.

Changes in the cellular carbohydrate metabolism are a hallmark of malignant transformation and represent one of the earliest discernible events in tumorigenesis. In the early stages of certain epithelial cancers, a metabolic switch is regularly observed, in which slowly growing glycogenotic cells are converted to highly proliferating basophilic cells. This step is accompanied by a rapid depletion of the intracellular glycogen stores, which in liver carcinogenesis results from the activation of the enzyme acid alpha-glucosidase by an as yet unknown mechanism. We show here that acid alpha-glucosidase is a target for the E7 protein encoded by human papillomavirus type 16, a human tumor virus that plays a key role in the genesis of cervical carcinoma. We show that expression of E7 induces the catalytic activity of acid alpha-glucosidase in vivo and wild type E7, but not transformation-deficient mutants bind directly to acid alpha-glucosidase and increase the catalytic activity of the enzyme in vitro. The data suggest that the E7 protein encoded by human papillomavirus type 16 can act as an allosteric activator of acid alpha-glucosidase.

Allosteric Regulation↗
PubMed1999

A diagnostic protocol for adult-onset glycogen storage disease type II.

To analyze the diagnostic value of various laboratory tests for the confirmation of adult-onset glycogen storage disease type II (GSD II), we performed a clinical, biochemical, and genetic study of 18 patients with this disease. Measurement of acid alpha-glucosidase (GAA) activity in muscle and histopathologic analysis of muscle tissue appeared to have no additional value when GAA activity in leukocytes was clearly deficient. Our study showed that creatine kinase elevation is a sensitive marker of GSD II. A diagnostic protocol is formulated.

Age of Onset↗
PubMed1999

Pathological features of glycogen storage disease type II highlighted in the knockout mouse model.

Glycogen storage disease type II (GSDII; Pompe's disease) is an autosomal recessive disease caused by lysosomal alpha-glucosidase deficiency. Skeletal muscle weakness is the most conspicuous clinical symptom of patients suffering from GSDII and skeletal muscle also is prominently involved in the knockout mouse model of this disease. Thus far, however, little detailed information has been published on the pathological changes in other mouse tissues. This paper aims to provide these data and gives a record of the clinical course of the mouse model over a 2-year period. Four-month-old affected mice perform worse in a running wheel than their unaffected littermates, but do not yet display other clear signs of disease. The lysosomal glycogen storage, already evident at birth, becomes more severe in time, leading to muscle wasting by 9-10 months of age and then limb girdle weakness and kyphosis. The disease does not markedly shorten the animal's life span despite the serious tissue pathology, which is not limited to heart and skeletal muscle, but is also seen in the smooth muscle of blood vessels and of the respiratory, digestive, and urogenital tracts. In addition, the mice have lysosomal glycogen storage in the liver, kidney, spleen, and salivary gland; in Schwann cells of the peripheral nerves, and in a subset of neurons in the central nervous system. By pathological criteria, the knockout mouse model parallels the human infantile form of GSDII and is attractive for studying the possible reversal of tissue pathology and symptomatology under different therapeutic regimes.

Animals↗
PubMed1999

Frequency of glycogen storage disease type II in The Netherlands: implications for diagnosis and genetic counselling.

Glycogen storage disease type II (GSD H) is an autosomal recessive myopathy. Early and late-onset phenotypes are distinguished - infantile, juvenile and adult. Three mutations in the acid alpha-glucosidase gene are common in the Dutch patient population: IVS1(-13T-->G), 525delT and delexon18. 63% of Dutch GSD II patients carry one or two of these mutations, and the genotype-phenotype correlation is known. To determine the frequency of GSD II, we have screened an unselected sample of neonates for the occurrence of these three mutations. Based on the calculated carrier frequencies, the predicted frequency of the disease is 1 in 40000 divided by 1 in 138 000 for infantile GSD II and 1 in 57 000 for adult GSD II. This is about two to four times higher than previously suggested, which is a reason to become more familiar with the presentation of GSD II in its different clinical forms and to adjust the risk assessment for genetic counselling.

Alleles↗
PubMed1999

Human acid alpha-glucosidase from rabbit milk has therapeutic effect in mice with glycogen storage disease type II.

Pompe's disease or glycogen storage disease type II (GSDII) belongs to the family of inherited lysosomal storage diseases. The underlying deficiency of acid alpha-glucosidase leads in different degrees of severity to glycogen storage in heart, skeletal and smooth muscle. There is currently no treatment for this fatal disease, but the applicability of enzyme replacement therapy is under investigation. For this purpose, recombinant human acid alpha-glucosidase has been produced on an industrial scale in the milk of transgenic rabbits. In this paper we demonstrate the therapeutic effect of this enzyme in our knockout mouse model of GSDII. Full correction of acid alpha-glucosidase deficiency was obtained in all tissues except brain after a single dose of i.v. enzyme administration. Weekly enzyme infusions over a period of 6 months resulted in degradation of lysosomal glycogen in heart, skeletal and smooth muscle. The tissue morphology improved substantially despite the advanced state of disease at the start of treatment. The results have led to the start of a Phase II clinical trial of enzyme replacement therapy in patients.

Animals↗
PubMed1999

Glycogen storage disease type II: birth prevalence agrees with predicted genotype frequency.

OBJECTIVES: To compare the overall birth prevalence of diagnosed glycogen storage disease type II (GSD II) with the predicted frequency based on mutation screening, in order to determine whether GSD II is an underdiagnosed condition, and to analyze which medical disciplines recognize GSD II. METHODS: Retrospective data on all enzymatic diagnoses of GSD II were collected from diagnostic labs throughout the Netherlands, covering the period from January 1, 1972 to December 31, 1996. Age-specific diagnostic incidence rates were calculated for the entire study period. By adding together the diagnostic incidences for all age groups, we calculated the birth prevalence of diagnosed GSD II and compared these figures with the predicted frequency based on mutation screening in a random sample from the general population. The medical specialization of the referring clinicians was also recorded. RESULTS: GSD II was diagnosed in 154 individuals, including 11 prenatal diagnoses. The birth prevalences of the various phenotypes were 1/101,000 (infantile form), 1/720,000 (juvenile form) and 1/53,000 (adult form). The birth prevalence of the adult and infantile phenotype together was 1/35,000. Eighty-two percent of the patients were diagnosed in university hospitals. Of the patients with infantile GSD II, 71% were diagnosed by a pediatrician, whereas most patients with adult GSD II were diagnosed by a neurologist (80%). CONCLUSIONS: There is no evidence for the underdiagnosis of GSD II in the Netherlands, as the calculated birth prevalences of the disease are consistent with previous predictions based on mutation screening in a random sample of newborns. The worldwide birth prevalence of the disease may well be higher than 1 in 100,000. GSD II is mainly diagnosed in university hospitals.

Journal Article↗
PubMed1998

Glycogen Storage Disease type II: genetic and biochemical analysis of novel mutations in infantile patients from Turkish ancestry.

Glycogen Storage Disease type II (GSDII) is caused by the deficiency of lysosomal alpha-glucosidase (acid maltase). This paper reports on the characterization of the molecular defects in 6 infantile patients from Turkish ancestry. Five of the 6 patients had reduced levels of the lysosomal alpha-glucosidase precursor. Conversion to mature enzyme was impaired in all cases, and the lysosomal alpha-glucosidase activity in all patients fibroblasts was less than 0.5% of control. DNA sequence analysis revealed 3 new mutations. One mutation, found in 3 patients in homozygous form, was a double insertion in exon 19 (2471AG-->CAGG) leading to a frameshift after Pro 913. It is the first insertion mutation described in the lysosomal alpha-glucosidase gene. Two patients were homozygous for missense mutations leading to the substitution of Ser to Pro at amino acid 566 (S566P) in one case and of Pro to Arg at amino acid 768 (P768R) in the other. One patient was found to have a Gly to Arg missense mutation at amino acid 643 (G643R), previously identified in an adult patient (Hermans et al., 1993), combined with a silent second allele. The latter 3 mutations were introduced in the wild type lysosomal alpha-glucosidase cDNA and expressed in COS cells to analyze their effect. Precursor species of 110 kD were formed but the maturation was impaired. As a result there was an overall deficiency of catalytic activity, which is in accordance with the findings in the patients fibroblasts and with the clinical phenotype.

Animals↗
PubMed1998

Generalized glycogen storage and cardiomegaly in a knockout mouse model of Pompe disease.

Glycogen storage disease type II (GSDII; Pompe disease), caused by inherited deficiency of acid alpha-glucosidase, is a lysosomal disorder affecting heart and skeletal muscles. A mouse model of this disease was obtained by targeted disruption of the murine acid alpha-glucosidase gene (Gaa) in embryonic stem cells. Homozygous knockout mice (Gaa -/-) lack Gaa mRNA and have a virtually complete acid alpha-glucosidase deficiency. Glycogen-containing lysosomes are detected soon after birth in liver, heart and skeletal muscle cells. By 13 weeks of age, large focal deposits of glycogen have formed. Vacuolar spaces stain positive for acid phosphatase as a sign of lysosomal pathology. Both male and female knockout mice are fertile and can be intercrossed to produce progeny. The first born knockout mice are at present 9 months old. Overt clinical symptoms are still absent, but the heart is typically enlarged and the electrocardiogram is abnormal. The mouse model will help greatly to understand the pathogenic mechanism of GSDII and is a valuable instrument to explore the efficacy of different therapeutic interventions.

Animals↗
PubMed1998

Adenovirus-mediated transfer of the acid alpha-glucosidase gene into fibroblasts, myoblasts and myotubes from patients with glycogen storage disease type II leads to high level expression of enzyme and corrects glycogen accumulation.

Glycogen storage disease type II (GSD II) is an autosomal recessive disorder caused by defects in the lysosomal acid alpha-glucosidase (GAA) gene. We investigated the feasibility of using a recombinant adenovirus containing the human GAA gene under the control of the cytomegalovirus promoter (AdCMV-GAA) to correct the enzyme deficiency in different cultured cells from patients with the infantile form of GSD II. In GAA-deficient fibroblasts infected with AdCMV-GAA, transduction and transcription of the human transgene resulted in de novo synthesis of GAA protein. The GAA enzyme activity was corrected from the deficient level to 12 times the activity of normal cells. The transduced cells overexpressed the 110 kDa precursor form of GAA, which was secreted into the culture medium and was taken up by recipient cells. The recombinant GAA protein was correctly processed and was active on both an artificial substrate 4-methylumbelliferyl-alpha-D-glucopyranoside (4MUG) and glycogen. In GAA-deficient muscle cells, a significant increase in cellular enzyme level, approximately 20-fold higher than in normal cells, was also observed after viral treatment. The transduced muscle cells were also able to efficiently secrete the recombinant GAA. Moreover, transfer of the human transgene resulted in normalization of cellular glycogen content with clearance of glycogen from lysosomes, as assessed by electron microscopy, in differentiated myotubes. These results demonstrate phenotypic correction of cultured skeletal muscle from a patient with infantile-onset GSD II using a recombinant adenovirus. We conclude that adenovirus-mediated gene transfer might be a suitable model system for further in vivo studies on delivering GAA to GSD II muscle, not only by direct cell targeting but also by a combination of secretion and uptake mechanisms.

Adenoviridae↗
PubMed1998

Recombinant human acid alpha-glucosidase: high level production in mouse milk, biochemical characteristics, correction of enzyme deficiency in GSDII KO mice.

Glycogen storage disease type II (GSDII) is caused by lysosomal acid alpha-glucosidase deficiency. Patients have a rapidly fatal or slowly progressive impairment of muscle function. Enzyme replacement therapy is under investigation. For large-scale, cost-effective production of recombinant human acid alpha-glucosidase in the milk of transgenic animals, we have fused the human acid alpha-glucosidase gene to 6.3 kb of the bovine alphaS1-casein gene promoter and have tested the performance of this transgene in mice. The highest production level reached was 2 mg/ml. The major fraction of the purified recombinant enzyme has a molecular mass of 110 kDa and resembles the natural acid alpha-glucosidase precursor from human urine and the recombinant precursor secreted by CHO cells, with respect to pH optimum, Km, Vmax, N-terminal amino acid sequence and glycosylation pattern. The therapeutic potential of the recombinant enzyme produced in milk is demonstrated in vitro and in vivo. The precursor is taken up in a mannose 6-phosphate receptor-dependent manner by cultured fibroblasts, is converted to mature enzyme of 76 kDa and depletes the glycogen deposit in fibroblasts of patients. When injected intravenously, the milk enzyme corrects the acid alpha-glucosidase deficiency in heart and skeletal muscle of GSDII knockout mice.

Animals↗