[Show abstract][Hide abstract] ABSTRACT: Pompe disease is an autosomal recessive inherited metabolic disease caused by deficiency of acid α-glucosidase (GAA). Glycogen accumulation is seen in the affected organ such as skeletal muscle, heart, and liver. Hypertrophic cardiomyopathy is frequently seen in the infantile onset Pompe disease. On the other hand, cardiovascular complication of the late-onset Pompe disease is considered as less frequent and severe than that of infantile onset. There are few investigations which show cardiovascular complication of late onset Pompe disease due to the shortage of appropriate disease model. We have generated late-onset Pompe disease-specific induced pluripotent stem cell (iPSC) and differentiated them into cardiomyocytes. Differentiated cardiomyocyte shows glycogen accumulation and lysosomal enlargement. Lentiviral GAA rescue improves GAA enzyme activity and glycogen accumulation in iPSC. The efficacy of gene therapy is maintained following the cardiomyocyte differentiation. Lentiviral GAA transfer ameliorates the disease-specific change in cardiomyocyote. It is suggested that Pompe disease iPSC-derived cardiomyocyte is replicating disease-specific changes in the context of disease modeling, drug screening, and cell therapy.
[Show abstract][Hide abstract] ABSTRACT: We developed an immunochromatography-based assay for detecting antibodies against recombinant α-galactosidase A proteins in serum. The evaluation of 29 serum samples from Fabry patients, who had received enzyme replacement therapy with agalsidase alpha and/or agalsidase beta, was performed by means of this assay method, and the results clearly revealed that the patients exhibited the same level of antibodies against both agalsidase alpha and agalsidase beta, regardless of the species of recombinant α-galactosidase A used for enzyme replacement therapy. A conventional enzyme-linked immunosorbent assay supported the results. Considering these, enzyme replacement therapy with agalsidase alpha or agalsidase beta would generate antibodies against the common epitopes in both agalsidase alpha and agalsidase beta. Most of the patients who showed immunopositive reaction exhibited classic Fabry phenotype and harbored gene mutations affecting biosynthesis of α-galactosidase A. As immunochromatography is a handy and simple assay system which can be available at bedside, this assay method would be extremely useful for quick evaluation or first screening of serum antibodies against agalsidase alpha or agalsidase beta in Fabry disease with enzyme replacement therapy.
PLoS ONE 06/2015; 10(6):e0128351. DOI:10.1371/journal.pone.0128351 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We examined alpha-galactosidase A (GLA) gene mutations in 74 Japanese families with Fabry disease (FD) to determine the frequency of de novo mutations. In 5 of 74 families (6.8%), the probands had no positive family histories and were diagnosed as de novo because their parents had no mutations in GLA gene. The parents of Fabry patients do not necessarily have mutations in GLA gene which is an important consideration in genetic counseling for FD.
Molecular Genetics and Metabolism Reports 12/2014; 1(1):283–287. DOI:10.1016/j.ymgmr.2014.07.001
[Show abstract][Hide abstract] ABSTRACT: Mucopolysaccharidosis type II (MPS II) is a lysosomal storage disorder caused by deficient activity of the iduronate-2-sulfatase. This leads to accumulation of glycosaminoglycans (GAGs) in the lysosomes of various cells. Although it has been proposed that bone marrow transplantation (BMT) may have a beneficial effect for patients with MPS II, the requirement for donor-cell chimerism to reduce GAG levels is unknown. To address this issue, we transplanted various ratios of normal and MPS II bone marrow cells in a mouse model of MPS II and analyzed GAG accumulation in various tissues. Chimerism of whole leukocytes and each lineage of BMT recipients’ peripheral blood was similar to infusion ratios. GAGs were significantly reduced in the liver, spleen, and heart of recipients. The level of GAG reduction in these tissues depends on the percentage of normal-cell chimerism. In contrast to these tissues, a reduction in GAGs was not observed in the kidney and brain, even if 100 % donor chimerism was achieved. These observations suggest that a high degree of chimerism is necessary to achieve the maximum effect of BMT, and donor lymphocyte infusion or enzyme replacement therapy might be considered options in cases of low-level chimerism in MPS II patients.
[Show abstract][Hide abstract] ABSTRACT: The authors here report a single case of a 10-year-old male patient who presented with severe vision loss associated with progressive demyelination. The patient was diagnosed with X-linked childhood cerebral adrenoleukodystrophy (ALD). Genetic analysis demonstrated a missense mutation (Gly266Arg) in exon 1 of the ABCD1 gene. His corrected visual acuity confirmed the absolute lack of light perception in both eyes. Funduscopy revealed severe pallor of the optic disc in both eyes. Spectral-domain optical coherence tomography showed thinning of the retinal ganglion cell and inner plexiform layers (GCL and IPL). Thinning of the GCL and IPL may be due to transneuronal retrograde degeneration of ganglion cells secondary to optic tract demyelination.
[Show abstract][Hide abstract] ABSTRACT: Pompe disease is an autosomal recessive myopathic disorder caused by the deficiency of lysosomal acid α-glucosidase (GAA). Recently, we showed that function of mutant GAA in fibroblasts derived from Pompe disease patient carrying c.546G>T mutation is improved by treatment with proteasome inhibitor bortezomib as well as pharmacological chaperone (PC). However, bortezomib-responsive GAA mutations are not fully characterized. In this study, we showed the effect of bortezomib on different mutants of GAA in patient fibroblasts and transiently expressed HEK293T cells. Bortezomib increased the maturation and residual activity of GAA in patient fibroblasts carrying PC-responsive M519V and PC-unresponsive C647W mutations. Enhanced colocalization of GAA with lysosomal marker LAMP2 was also observed in patient fibroblasts after treatment with bortezomib. When four distinct mutant GAAs, which show different response to PC, were overexpressed in HEK293T cells, bortezomib improved the activity of M519V, S529V, and C647W in them (1.3-5.9-fold). These results indicate that bortezomib enhances the activity of some PC-unresponsive GAA mutants as well as PC-responsive mutants.
[Show abstract][Hide abstract] ABSTRACT: Mucolipidosis type II is an autosomal recessive lysosomal storage disease caused by N-acetylglucosamine-1-phosphotransferese deficiency. We report here pathological findings of an autopsy case of mucolipidosis type II. The patient was an 8-year-old boy with mucolipidosis type II and was complicated with hypertrophic cardiomyopathy. He suddenly developed progressive respiratory failure and finally died. At autopsy, systemic accumulation of undigested lysosomal metabolites was prominent, particularly in the heart, lungs, and dorsal root ganglion. In cardiomyocyte, LC3, an autophagy marker, was positive in the cytoplasm. Ubiquitin, p62, K48 polyubiquitin, and K63 polyubiquitin were also positive in the cytoplasm. Our findings suggest that autophagic dysfunction might be associated with the cardiomyopahty of mucolipidosis type II.
[Show abstract][Hide abstract] ABSTRACT: Pompe disease (PD), which is also called glycogen storage disease type II (GSDII), is one of the lysosomal storage diseases (LSDs) caused by a deficiency in acid-α-glucosidase (GAA) in the lysosome and is characterized by the accumulation of glycogen in various cells. PD has been treated by enzyme replacement therapy (ERT). We generated induced pluripotent stem cells (iPSCs) from the cells of patients with infantile-type and late-onset-type PD using a retrovirus vector to deliver transgenes encoding four reprogramming factors, namely, OCT4, SOX2, c-MYC, and KLF4. We confirmed that the two types of PD-iPSCs exhibited an undifferentiated state, alkaline phosphatase staining, and the presence of SSEA-4, TRA-1-60, and TRA-1-81. The PD-iPSCs exhibited strong positive staining with Periodic acid-Schiff (PAS). Moreover, ultrastructural features of these iPSCs exhibited massive glycogen granules in the cytoplasm, particularly in the infantile-type but to a lesser degree in the late-onset type. Glycogen granules of the infantile-type iPSCs treated with rhGAA were markedly decreased in a dose-dependent manner. Human induced pluripotent stem cell provides an opportunity to build up glycogen storage of Pompe disease in vitro. It represents a promising resource to study disease mechanisms, screen new drug compounds and develop new therapies for Pompe disease.
[Show abstract][Hide abstract] ABSTRACT: We previously showed that mesenchymal stem cells (MSCs) can differentiate into a functional miniature kidney, suggesting that MSCs may be a cell source for kidney regeneration. However, MSCs from long-term dialysis patients, which have been exposed to uremic toxin, can exhibit reduced viability. Therefore, the aim of this study was to examine the gene expression profiles and differentiation capabilities of bone marrow- and adipose-derived MSCs from chronic kidney disease (CKD) model rats. CKD was induced in rats by adenine feeding, and then MSCs were isolated from bone marrow (BMSCs) and adipose tissue (ASCs). After confirming MSC surface marker expression, comprehensive gene expression profiles were obtained by RT-PCR array. MSCs were differentiated into adipocytes, osteoblasts, and chondrocytes, and histological and/or functional assays were performed. Tgfb3 expression was up-regulated, while Bmp6, Gdf15, Mmp2, and Vegfa were down-regulated in CKD-ASCs compared with Control-ASCs. There were no significant differences in the gene expression of stemness markers, and the morphology of cells that underwent adipogenesis, osteogenesis, and chondrogenesis, or GPDH activity between CKD and control groups. Comparing BMSCs with ASCs, gene expression of Bglap, Bmp4, Igf1, Itgax, Pparg, Ptprc, and Tnf were up-regulated, while Col1a1, Mmp2, Sox9, and Vegfa were down-regulated in both CKD and control groups. Uremic toxin in CKD rats had a small effect on the gene expression and differentiation of MSCs. However, long-term exposure to uremic toxin and the differences in gene expression of MSCs derived from bone marrow or adipose tissue may affect renal regeneration.
Human Cell 02/2014; 27(2). DOI:10.1007/s13577-013-0082-7 · 1.41 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Gyrate atrophy (GA) of the choroid and retina is an extremely rare inherited chorioretinal dystrophy. Ornithine aminotransferase (OAT) gene mutations are identified in patients with GA. The purpose of this study was to report a novel deletion mutation of the OAT gene and describe clinical features of two brothers with GA in a Japanese family.
We performed ophthalmic examinations, including best-corrected visual acuity, slit-lamp biomicroscopy, dilated funduscopy, fundus autofluorescence imaging, optical coherence tomography, visual field testing, and full-field electroretinography (ERG). Serum ornithine concentrations and OAT activities were analyzed. Mutation screening of the OAT gene was performed using Sanger sequencing.
Both brothers had compound heterozygous mutations (p.K169DfsX10 and p.R426X), one of which was novel. Their unaffected parents carried one of the mutations heterozygously. An arginine-restricted diet was started in the younger brother at the age of 2 years, while the diet was not initiated in the older brother until the age of 6 years. After more than 15 years of follow-up, the dietary treatment seemed to slow the progression of the chorioretinal lesions in the younger brother. However, when compared at the same age, the younger brother had more reduced ERG amplitudes and constricted visual fields than his older brother.
We identified a novel frameshift mutation (p.K169DfsX10) in the OAT gene. While an early arginine-restricted dietary treatment suppressed the fundus changes of GA to some degree in the younger brother, the efficacy of suppressing the progression of visual function loss could not be clearly determined.
[Show abstract][Hide abstract] ABSTRACT: Mucolipidosis type II is an autosomal recessive lysosomal storage disease caused by N-acetylglucosamine-1-phosphotransferese deficiency. We report here pathological findings of an autopsy case of mucolipidosis type II. The patient was an 8-year-old boy with mucolipidosis type II and was complicated with hypertrophic cardiomyopathy. He suddenly developed progressive respiratory failure and finally died. At autopsy, systemic accumulation of undigested lysosomal metabolites was prominent, particularly in the heart, lungs, and dorsal root ganglion. In cardiomyocyte, LC3, an autophagy marker, was positive in the cytoplasm. Ubiquitin, p62, K48, K63 polyubiquitin were also positive in the cytoplasm. Our findings suggest that autophagic dysfunction might be associated with the cardiomyopahty of mucolipidosis type II.
Molecular Genetics and Metabolism 01/2014; · 2.63 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Fluorometric measurements of 4-methylumbelliferone (4-MU) are generally used to screen lysosomal storage diseases (LSDs) using dried blood spots (DBSs). However, in DBS, it is difficult to measure lysosomal acid lipase (LAL) activity due to the influence of other lipases in whole blood. Recently, Hamilton used a fluorometric enzyme assay with 4-MU derivatives to measure the LAL activity in DBS. This method requires mercury chloride as stopping reagent, and the fluorescence intensity of 4-MU was measured at an acidic pH. We report a revised method to measure the LAL activity without using toxic mercury chloride and to measure the fluorescence intensity of 4-MU at a basic pH. For this measurement, we established a more practical method that does not require mercury chloride. The LAL activity in DBS was measured in 51 normal controls, seven obligate carriers and seven patients with CESD. The average LAL activities±SD in the DBS from the normal, obligate carriers and CESD patients were 0.68±0.2 (range: 0.3-1.08), 0.21±0.1 (range: 0.11-0.41) and 0.02±0.02 (range: 0-0.06) nmol/punch/h, respectively. There was a significant difference between the normal and the CESD. Our method does not require toxic mercury chloride and is an appropriate revised enzyme assay using DBS for screening patients with CESD.
[Show abstract][Hide abstract] ABSTRACT: We report an autopsy case of advanced late-onset Pompe disease (juvenile type). At age 15, the patient experienced proximal weakness and subsequently required non-invasive positive pressure ventilation. Enzyme replacement therapy was initiated at age 37, which improved pinch power and alleviated respiratory distress. At age 41, repetitive pneumothorax led to respiratory failure and death. Autopsy excluding the brain showed few vacuoles and PAS-positive fibers in the left rectus femoris, compared with the contralateral muscle biopsy performed at age 21. Vacuolar changes were also less evident than previous autopsy cases without enzyme replacement therapy. The findings suggest that enzyme replacement therapy may have suppressed myovacuolopathy in our patient with advanced late-onset Pompe disease. Apart from vacuolopathy, the autopsied muscle showed marked myofiber atrophy and fibrosis, suggesting that disuse atrophy may have contributed to the muscle weakness. This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Objective. To determine the effects of enzyme replacement therapy (ERT) on the hearing acuity in patients with Fabry disease. Materials. The study sample comprised 34 ears of 17 affected patients who underwent pure-tone audiometry before and after ERT. Methods. The patients were studied in relation to factors such as changes in hearing, presence of accompanying symptoms, status of renal and cardiac function, age, and gender. Data of pure-tone audiometry obtained before ERT and at the final examination were compared. Results. At the end of the follow-up period, no significant worsening of hearing acuity was noted at the end of the follow-up period. SSNHL was detected in 10 ears of 6 patients. Steroid therapy successfully cured the disease in 9 of the 10 ears. Conclusions. No significant worsening of hearing acuity was noted from the beginning to the end of ERT. The rate of improvement in SSNHL of Fabry disease was excellent in the treated patients. Hearing loss is a factor that causes marked deterioration of the patients' quality of life, and it is desirable that the hearing acuity of patients be periodically evaluated and prompt treatment of SSNHL be administered, if available.
International Journal of Otolaryngology 10/2013; 2013:282487. DOI:10.1155/2013/282487
[Show abstract][Hide abstract] ABSTRACT: Before the availability of an enzyme replacement therapy (ERT) for mucopolysaccharidosis type II (MPS II), patients were treated by bone marrow transplantation (BMT). However, the effectiveness of BMT for MPS II was equivocal, particularly at addressing the CNS manifestations. To study this further, we subjected a murine model of MPS II to BMT and evaluated the effect at correcting the biochemical and pathological aberrations in the viscera and CNS. Our results indicated that BMT reduced the accumulation of glycosaminoglycans (GAGs) in a variety of visceral organs, but not in the CNS. With the availability of an approved ERT for MPS II, we investigated and compared the relative merits of the two strategies either as a mono or combination therapy. We showed that the combination of BMT and ERT was additive at reducing tissue levels of GAGs in the heart, kidney and lung. Moreover, ERT conferred greater efficacy if the immunological response against the infused recombinant enzyme was low. Finally, we showed that pathologic GAGs might potentially represent a sensitive biomarker to monitor the therapeutic efficacy of therapies for MPS II.