A novel colorimetric method for diagnosing glycerol-3-phosphate dehydrogenase 1 deficiency K. Vanhouteghem1, P. Verloo2, R. Debruyne2, M. Speeckaert3, J. Delanghe1 1Department
of Clinical Chemistry, Ghent University Hospital, Ghent, Belgium 2Department of Pediatrics, Ghent University and Ghent University Hospital, Ghent, Belgium 3Department of Nephrology, Ghent University Hospital, Ghent, Belgium
GK
INTRODUCTION Glycerol-3-phosphate dehydrogenase 1 (G3PDH1) deficiency is a disorder in which the NADH/NAD+ dependent cycling between
G3PDH1
dihydroxyacetone phosphate (DHAP) and glycerol-3-phosphate, a key metabolite in the triglyceride synthesis, is dysregulated. This poorly described, inborn error of glycerol metabolism can be detected by the presence of a pseudohypertriglyceridemia. In the present study, we explored a novel colorimetric method for assaying G3PDH1 activity. Figure 1. Illustration of glycerol metabolism
METHOD We developed a new colorimetric method, in which G3PDH1 activity was measured, based on the conversion of L-glycerol-3-phosphate into DHAP in the presence of NAD+. 1000
normolipemic hyperlipoproteinemia type IV
G3PDH1
+ NAD+
hyperlipoproteinemia type V
+ NADH
glycerol kinase deficiency postprandial sample
DHAP L-index (U)
glycerol-3-phosphate
100
Photometric reading of the formed NADH occurred at 340 nm. Mean G3PDH1 activity was measured in a healthy control group (n=20).
10
RESULTS - Two unrelated patients - a 63 year-old man
and a 17 year-old girl
Figure 2. Log-transformed scattergram of serum triglyceride and L-index for various patient groups 1
- Both presenting with hepatosplenomegaly and an image compatible with hepatic fibrosis on ultrasound elastography. 1
10
100
1000
10000
Triglyceride (mg/dL)
- Laboratory analysis revealed: Routine fasting blood analysis
high ratio TG/L-index
Triglyceride concentration (reference interval 37 – 131 mg/dL) Lipemic index (L-index)
suggesting interference of
996 mg/dL 10 U
687 mg/dL 4U
- The analysis was repeated with a glycerol-blanked triglyceride method in each case: Triglyceride concentration (reference interval 37 – 131 mg/dL)
glycerol (hyperglycerolemia)
normalisation of triglyceride levels 98,8 mg/dL
101,2 mg/dL
suggesting a diagnosis of pseudohypertriglyceridemia
- Pseudohypertriglyceridemia based on a pronounced hyperglycerolemia may be: - due to the presence of exogenous glycerol. Dietary or pharmacological causes were excluded. - due to the accumulation of endogenous glycerol. Glycerol kinase (GK) and G3PDH1 are crucial enzymes in the
triglyceride metabolism (figure 1). Figure 2 illustrates the possibility of GK deficiency in case of high triglyceride
A very low G3PDH1 activity
levels and low L-index. However, as GK deficiency is an X-linked disorder, we assayed the serum G3PDH1 activity with
was detected
our novel colorimetric method:
suggesting a G3PDH1 deficiency.
G3PDH1 activity (% of the G3DPH1 activity measured in the healthy control group)
19,5 %
7,8 %
CONCLUSION • These cases nicely illustrate that clinicians need to be aware of pseudohypertriglyceridemia, explained by glycerolemia secondary to G3PDH1 deficiency. • Our novel colorimetric method allows determining G3PDH1 activity in human serum. • The assay may be useful for studying G3PDH1 deficiency in case of pseudohypertriglyceridemia.
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