Distinctive accumulation of globotriaosylceramide and globotriaosylsphingosine in a mouse model of classic Fabry disease.

Fabry disease (FD) is an inherited disease caused by deficient α-galactosidase A activity that is characterized by the accumulation of globotriaosylceramide (Gb3) and globotriaosylsphingosine (lyso-Gb3). Although plasma lyso-Gb3 is a sensitive biomarker of FD, the correlation between its concentration and clinical symptoms remains unclear. To clarify the influence of plasma Gb3 and lyso-Gb3 in a symptomatic Gla tm Tg(CAG-A4GALT) FD mouse model, the total contents of Gb3, lyso-Gb3 and their analogs in various organs and plasma were determined in mice with early- (5-week-old) and late-stage (20-week-old) renal dysfunction. A marked increase in total Gb3 content in the heart, kidneys, spleen, liver, small intestine, lungs, brain, and plasma was observed in the 20-week-old mice compared to that in 5-week-old mice. In contrast, the increase in lyso-Gb3 was relatively small, and the total content in the lungs and plasma was unchanged. Lyso-Gb3 analogs {lyso-Gb3(-2) and lyso-Gb3(+18)} and Gb3 analogs {Gb3(-2) and Gb3(+18)} were observed in all organs and plasma at both ages, and the percentages of the analogs were unique to specific organs. The pattern of 37 Gb3 analogs/isoforms of liver Gb3 corresponded well with that of plasma Gb3. Although the analog pattern of plasma lyso-Gb3 did not resemble that of any organ lyso-Gb3, the relative content {lyso-Gb3: lyso-Gb3(-2)} in the sum of all organs corresponded well to that of the plasma at both ages. These data indicate that liver Gb3 may contribute to the plasma Gb3 level, while plasma lyso-Gb3 may be released from all organs, and the capacity of the plasma lyso-Gb3 pool may reach a maximum at an early stage of renal dysfunction.

Plasma Globotriaosylsphingosine and α-Galactosidase A Activity as a Combined Screening Biomarker for Fabry Disease in a Large Japanese Cohort.

Fabry disease is an X-linked disorder of α-galactosidase A (GLA) deficiency. Our previous interim analysis (1 July 2014 to 31 December 2015) revealed plasma globotriaosylsphingosine as a promising primary screening biomarker for Fabry disease probands. Herein, we report the final results, including patients enrolled from 1 January to 31 December 2016 for evaluating the potential of plasma globotriaosylsphingosine and GLA activity as a combined screening marker. We screened 5691 patients (3439 males) referred from 237 Japanese specialty clinics based on clinical findings suggestive of Fabry disease using plasma globotriaosylsphingosine and GLA activity as primary screening markers, and GLA variant status as a secondary screening marker. Of the 14 males who tested positive in the globotriaosylsphingosine screen (≥2.0 ng/mL), 11 with low GLA activity (<4.0 nmol/h/mL) displayed GLA variants (four classic, seven late-onset) and one with normal GLA activity and no pathogenic variant displayed lamellar bodies in affected organs, indicating late-onset biopsy-proven Fabry disease. Of the 19 females who tested positive in the globotriaosylsphingosine screen, eight with low GLA activity displayed GLA variants (six classic, two late-onset) and five with normal GLA activity displayed a GLA variant (one classic) and no pathogenic variant (four late-onset biopsy-proven). The combination of plasma globotriaosylsphingosine and GLA activity can be a primary screening biomarker for classic, late-onset, and late-onset biopsy-proven Fabry disease probands.

Low bone mineral density due to secondary hyperparathyroidism in the GlatmTg(CAG-A4GALT) mouse model of Fabry disease.

Low bone mineral density (BMD)-diagnosed as osteoporosis or osteopenia-has been reported as a new characteristic feature of Fabry disease; however, the mechanism underlying the development of low BMD is unknown. We previously revealed that a mouse model of Fabry disease [GlatmTg(CAG-A4GALT)] exhibits impaired functioning of medullary thick ascending limb (mTAL), leading to insufficient Ca2+ reabsorption and hypercalciuria. Here, we investigated bone metabolism in GlatmTg(CAG-A4GALT) mice without marked glomerular or proximal tubular damage. Low BMD was detected by 20 weeks of age via micro-X-ray-computed tomography. Bone histomorphometry revealed that low BMD results by accelerated bone resorption and osteomalacia. Plasma parathyroid hormone levels increased in response to low blood Ca2+-not plasma fibroblast growth factor 23 (FGF-23) elevation-by 5 weeks of age and showed progressively increased phosphaturic action. Secondary hyperparathyroidism developed by 20 weeks of age and caused hyperphosphatemia, which increased plasma FGF-23 levels with phosphaturic action. The expression of 1α-hydroxylase [synthesis of 1α,25(OH)2D3] in the kidney did not decrease, but that of 24-hydroxylase [degradation of 1α,25(OH)2D3] decreased. Vitamin D deficiency was ruled out as the cause of osteomalacia, as plasma 1α,25(OH)2D3 and 25(OH)D3 levels were maintained. Results demonstrate that secondary hyperparathyroidism due to mTAL impairment causes accelerated bone resorption and osteomalacia due to hyperphosphaturia and hypercalciuria, leading to low BMD in Fabry model mice.

Correction: Effectiveness of plasma lyso-Gb3 as a biomarker for selecting high-risk patients with Fabry disease from multispecialty clinics for genetic analysis.

The HTML version of this Article contained errors in Supplementary Figure S2 "Flowchart of the lyso-Gb3 screening and gene analysis in female patients", which have been detailed in the associated Correction.

Effectiveness of plasma lyso-Gb3 as a biomarker for selecting high-risk patients with Fabry disease from multispecialty clinics for genetic analysis.

PURPOSE: Plasma globotriaosylsphingosine (lyso-Gb3) is a promising secondary screening biomarker for Fabry disease. Here, we examined its applicability as a primary screening biomarker for classic and late-onset Fabry disease in males and females. METHODS: Between 1 July 2014 and 31 December 2015, we screened 2,359 patients (1,324 males) referred from 168 Japanese specialty clinics (cardiology, nephrology, neurology, and pediatrics), based on clinical symptoms suggestive of Fabry disease. We used the plasma lyso-Gb3 concentration, α-galactosidase A (α-Gal A) activity, and analysis of the α-Gal A gene (GLA) for primary and secondary screens, respectively. RESULTS: Of 8 males with elevated lyso-Gb3 levels (≥2.0 ng ml-1) and low α-Gal A activity (≤4.0 nmol h-1 ml-1), 7 presented a GLA mutation (2 classic and 5 late-onset). Of 14 females with elevated lyso-Gb3, 7 displayed low α-Gal A activity (5 with GLA mutations; 4 classic and 1 late-onset) and 7 exhibited normal α-Gal A activity (1 with a classic GLA mutation and 3 with genetic variants of uncertain significance). CONCLUSION: Plasma lyso-Gb3 is a potential primary screening biomarker for classic and late-onset Fabry disease probands.

Correction: Effectiveness of plasma lyso-Gb3 as a biomarker for selecting high-risk patients with Fabry disease from multispecialty clinics for genetic analysis.

The PDF and HTML versions of the article have been updated to include the Creative Commons Attribution 4.0 International License information.

Correction: Effectiveness of plasma lyso-Gb3 as a biomarker for selecting high-risk patients with Fabry disease from multispecialty clinics for genetic analysis.

In the above article, we noticed that one female patient in the positive group (plasma lyso-Gb3 7.6 ng/ml, α-galactosidase A activity 4.9 nmol/h/ml) who presented at the neurology clinic was already diagnosed with Fabry disease before the current study. We excluded patients with a confirmed diagnosis of Fabry disease and those with relatives known to have Fabry disease. To accurately describe the information in the current study, we must exclude this patient from the analysis. We have accurately revised this information as follows.

Medullary thick ascending limb impairment in the GlatmTg(CAG-A4GALT) Fabry model mice.

A main feature of Fabry disease is nephropathy, with polyuria an early manifestation; however, the mechanism that underlies polyuria and affected tubules is unknown. To increase globotriaosylceramide (Gb3) levels, we previously crossbred asymptomatic Glatm mice with transgenic mice that expressed human Gb3 synthase (A4GALT) and generated the GlatmTg(CAG-A4GALT) symptomatic Fabry model mice. Additional analyses revealed that these mice exhibit polyuria and renal dysfunction without remarkable glomerular damage. In the present study, we investigated the mechanism of polyuria and renal dysfunction in these mice. Gb3 accumulation was mostly detected in the medulla; medullary thick ascending limbs (mTALs) were the most vacuolated tubules. mTAL cells contained lamellar bodies and had lost their characteristic structure ( i.e., extensive infolding and numerous elongated mitochondria). Decreased expression of the major molecules-Na+-K+-ATPase, uromodulin, and Na+-K+-2Cl- cotransporter-that are involved in Na+ reabsorption in mTALs and the associated loss of urine-concentrating ability resulted in progressive water- and salt-loss phenotypes. GlatmTg(CAG-A4GALT) mice exhibited fibrosis around mTALs and renal dysfunction. These and other features were consistent with pathologic findings in patients with Fabry disease. Results demonstrate that mTAL dysfunction causes polyuria and renal impairment and contributes to the pathophysiology of Fabry nephropathy.-Maruyama, H., Taguchi, A., Nishikawa, Y., Guili, C., Mikame, M., Nameta, M., Yamaguchi, Y., Ueno, M., Imai, N., Ito, Y., Nakagawa, T., Narita, I., Ishii, S. Medullary thick ascending limb impairment in the GlatmTg(CAG-A4GALT) Fabry model mice.