The aim of our study was to assess the possible relationships among heme oxygenase (HMOX), bilirubin UDP-glucuronosyl transferase (UGT1A1) promoter gene variations, serum bilirubin levels, and Fabry disease (FD). The study included 56 patients with FD (M : F ratio = 0.65) and 185 healthy individuals. Complete standard laboratory and clinical work-up was performed on all subjects, together with the determination of total peroxyl radical-scavenging capacity. The (GT)n and (TA)n dinucleotide variations in the
Fabry disease (FD, OMIM 301500), an X-linked metabolic disorder, is caused by a deficiency of the lysosomal enzyme
The pathological processes in FD are associated with increased oxidative, nitrosative, and carbonyl stress as evidenced by excessive production of reactive oxygen species (ROS) [
The oxidative stress defense system consists of antioxidative enzymes and antioxidative substrates. Among the latter, bilirubin, the heme catabolic product, belongs among the most potent endogenous antioxidants [
By regulating bilirubin production and its biotransformation in the liver, heme oxygenase-1 (encoded by
The objective of this study was to determine whether genetic variations in
The study was performed on 56 FD patients (M : F ratio = 0.70) diagnosed in our center from 2000 to 2012. The diagnosis of FD was based on a demonstrated reduction of
The control group was based upon age- and sex-matched 185 clinically healthy subjects without any chronic medication, representing a general population sample from the same geographical region. These subjects were recruited from healthy blood donors and employees of the General University Hospital. All subjects in both cohorts were of Caucasian ancestry.
The study protocol and all procedures conformed to the ethical guidelines of the 1975 Declaration of Helsinki, as reflected in a priori approval by the institution’s Ethics Committee, and all subjects signed informed-consent forms.
Peripheral venous blood was obtained from all participants after overnight fasting. The serum was immediately processed, and bilirubin was determined on an automatic analyzer (modular analyzer, Roche Diagnostics GmbH, Germany), using routine laboratory assay.
Total antioxidant status (TAS) was determined fluorometrically as the serum peroxyl radical-scavenging capacity, based on the relative proportion of chain-breaking antioxidant consumption present in the serum compared to that of Trolox (a reference and calibration antioxidant compound) [
The (GT)n (dbSNP rs1805173) and (TA)n (dbSNP rs81753472) dinucleotide variations in
The data are presented as either the mean ± SD or the median and 25–75% interquartile range. Data were evaluated by a
Compared to age-matched control subjects, patients with FD displayed significantly lower systemic bilirubin concentrations (Table
Serum bilirubin concentrations in patients with FD.
FD | Controls |
| |
---|---|---|---|
Males + females | |||
Age [years] |
|
|
0.303 |
Bilirubin [ |
|
|
|
Males | |||
Age [years] |
|
|
0.77 |
Bilirubin [ |
|
|
0.719 |
Females | |||
Age [years] |
|
|
0.313 |
Bilirubin [ |
|
|
|
Data expressed as mean ± SD, or median and IQ range, depending on their normality. FD: Fabry disease.
The effect of ERT on bilirubin and total antioxidant status in patients with FD.
ERT+ ( |
ERT− ( |
Controls ( |
| |
---|---|---|---|---|
Males + females | ||||
Bilirubin [ |
|
|
|
|
|
|
|
|
|
|
|
|
||
|
|
0.503 | ||
Males |
||||
|
|
( |
|
0.794 |
Females | ||||
Bilirubin [ |
|
|
0.186 | |
|
|
|
0.201 | |
|
|
0.235 |
Out of the entire group, 55% of patients were treated with ERT (enzyme replacement therapy) (95% men and 30% women). TAS, total antioxidant status, was determined according to [
The relationship between serum bilirubin and total antioxidant status in patients with FD. Each dot represents a single subject. FD, Fabry disease.
As expected, significantly lower serum bilirubin concentrations between FD patients and control subjects were observed within all
Serum bilirubin concentrations in patients with FD and controls according to
|
FD | Controls |
|
---|---|---|---|
6/6 |
|
|
|
6/7 |
|
|
|
7/7 |
|
|
|
|
0.538 |
|
Bilirubin expressed in
No significant differences in frequencies of the L-allele in the
The impact of variability of genes regulating heme catabolism on risk of FD.
OR for risk of FD manifestation |
| ||
|
|
0.46 | |
|
|
|
|
|
|||
FD | Controls |
| |
|
|
|
|
FD: Fabry disease.
Bilirubin has been reported to be a strong negative predictor/biomarker of oxidative stress-mediated diseases. This association has been reported for atherosclerosis and cancer, as well as metabolic, autoimmune, and neurodegenerative diseases [
With respect to oxidative stress-mediated damage, FD is not the exception, and the role of increased oxidative, nitrosative, and carbonyl stress in pathogenesis of cardiovascular complications of FD is indisputable [
Indeed, increased protein nitration and oxidative DNA damage leading to accelerated apoptosis were reported in cardiomyocytes of FD patients [
In fact, low serum TAS was also observed in our FD patient cohort, together with significantly lower serum bilirubin concentrations. Since bilirubin is one of the most potent endogenous antioxidants [
Increased consumption of antioxidant substrates seems to account for this observation, and this explanation is also supported by the decreased systemic ascorbate levels in FD patients reported by Moore et al. [
Although this observation may seem surprising, even monogenic diseases can be substantially influenced by genes capable of modifying disease phenotype [
The beneficial impact of ERT on the amelioration of increased oxidative/nitrosative stress is supported by the observation of a notable decrease of nitrotyrosine staining in the dermal blood vessels of FD patients on
Bilirubin not only is a potent antioxidant but also significantly affects nitric oxide production [
Apart from the possible effects on redox status, bilirubin can also exert its protective effects through other mechanisms, such as improvement of endothelial dysfunction, known to be present in FD patients [
We believe that marked disturbances in the oxidative stress defense system observed in our study might be of clinical importance and that they may have a prognostic value. Recently, we have shown that serum uric acid, an indirect indicator of oxidative stress, is associated with the long-term prognosis of FD patients [
FD is associated with markedly lower serum bilirubin levels, most likely due to bilirubin consumption during the increased oxidative stress associated with this disease. However, we propose that genetic factors also affecting the defense against increased oxidative stress (
Enzyme replacement therapy
Fabry disease
Globotriaosylceramide
Heme oxygenase
Reactive oxygen species
Total antioxidant status
Bilirubin UDP-glucuronosyl transferase.
The authors declare that they have no competing interests.
Alena Jirásková and Giulia Bortolussi performed all the genetic analyses. Gabriela Dostálová, Lenka Eremiášová, Lubor Golaň, and Vilém Danzig performed all the clinical work-up. Alena Jirásková and Libor Vítek planned the whole study. All authors contributed to the data analysis and interpretation, and they read and approved the final manuscript.
This work was supported by Grants PROGRES Q25/LF1 and RVO-VFN64165/2017 granted by Charles University in Prague and the Czech Ministry of Health, respectively. The funding bodies had no role in the design of the study and collection, analysis, and interpretation of data and in writing the manuscript.