Determining a Critical Threshold for G6PD Activity below which RBC Response to Oxidative Stress is Poor
Background Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme disorder in the world. Its main function is to generate NADPH that is required for anti-oxidative pathway in the cells especially in red blood cells (RBC). G6PD deficiency is X-linked and thus subject to random X-chromosome inactivation in women giving them mosaic expression of G6PD activities in their individual cells. This phenomenon makes it difficult for diagnosis with the currently available G6PD qualitative diagnostic tests. With the rolling out of newly marketed anti-malarial drug tafenoquine, which has a long half-life, screening for G6PD deficiency becomes a necessity where those with <70% G6PD activity cannot receive this drug. Thus, evidence for a quantitative cut-off for G6PD activity is needed to ensure safe drug administration.
Methods RBC models were developed to analyse the effect of oxidant on RBC oxidative markers namely total glutathione (GSH)and malondialdehyde (MDA). G6PD activity was measured using quantitative assay from Trinity Biotech and was correlated with cytofluorometric assay. RBC from twoG6PD heterozygous women with different G6PD activities were also analysed for comparison.
Results There was a negative correlation between G6PD activity and CuCl concentration and a strong association between G6PD activities and proportion of G6PD normal RBC in CuCl-treated models and in ex vivo RBC. However, in terms of oxidative stress markers analyses, unlike the hypothesis where the lower G6PD activity, the higher MDA and the lower GSH level, the CuCl RBC model showed that in low G6PD activities (10-30%) cells, the MDA level is lower compared to the rest of the models (p<0.05). The ex vivo models however were in line with the hypothesis, although the result was not significant (p=0.5). There was a significant difference between RBC with <60% and those with >80% G6PD activities in CuCl RBC model, but not in ex vivo RBC (p=0.5). Genotyping heterozygous subjects showed G6PDViangchan variant with 2.97U/gHb (33% activity) and 6.58 U/gHb (74% activity).
Conclusions The GSH analysis has pointed to the 60% G6PD activity cut-off and this data is supportive of the old World Health Organization threshold for intermediate upper limit of 60% G6PD activity. However, there are significant limitations in using MDA assay with CuCl RBC model because the RBC was already stressed due to the copper treatment and thus present a different result when compared to the ex-vivo model.
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Posted 04 Jun, 2020
On 17 Jun, 2020
On 28 May, 2020
On 27 May, 2020
On 26 May, 2020
On 26 May, 2020
On 11 May, 2020
Received 17 Mar, 2020
On 13 Mar, 2020
Invitations sent on 12 Mar, 2020
On 25 Feb, 2020
On 24 Feb, 2020
On 24 Feb, 2020
On 27 Jan, 2020
Received 26 Jan, 2020
Received 25 Jan, 2020
Received 25 Jan, 2020
On 22 Jan, 2020
On 19 Jan, 2020
On 10 Jan, 2020
Invitations sent on 09 Jan, 2020
On 22 Dec, 2019
On 21 Dec, 2019
On 21 Dec, 2019
On 19 Dec, 2019
Determining a Critical Threshold for G6PD Activity below which RBC Response to Oxidative Stress is Poor
Posted 04 Jun, 2020
On 17 Jun, 2020
On 28 May, 2020
On 27 May, 2020
On 26 May, 2020
On 26 May, 2020
On 11 May, 2020
Received 17 Mar, 2020
On 13 Mar, 2020
Invitations sent on 12 Mar, 2020
On 25 Feb, 2020
On 24 Feb, 2020
On 24 Feb, 2020
On 27 Jan, 2020
Received 26 Jan, 2020
Received 25 Jan, 2020
Received 25 Jan, 2020
On 22 Jan, 2020
On 19 Jan, 2020
On 10 Jan, 2020
Invitations sent on 09 Jan, 2020
On 22 Dec, 2019
On 21 Dec, 2019
On 21 Dec, 2019
On 19 Dec, 2019
Background Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme disorder in the world. Its main function is to generate NADPH that is required for anti-oxidative pathway in the cells especially in red blood cells (RBC). G6PD deficiency is X-linked and thus subject to random X-chromosome inactivation in women giving them mosaic expression of G6PD activities in their individual cells. This phenomenon makes it difficult for diagnosis with the currently available G6PD qualitative diagnostic tests. With the rolling out of newly marketed anti-malarial drug tafenoquine, which has a long half-life, screening for G6PD deficiency becomes a necessity where those with <70% G6PD activity cannot receive this drug. Thus, evidence for a quantitative cut-off for G6PD activity is needed to ensure safe drug administration.
Methods RBC models were developed to analyse the effect of oxidant on RBC oxidative markers namely total glutathione (GSH)and malondialdehyde (MDA). G6PD activity was measured using quantitative assay from Trinity Biotech and was correlated with cytofluorometric assay. RBC from twoG6PD heterozygous women with different G6PD activities were also analysed for comparison.
Results There was a negative correlation between G6PD activity and CuCl concentration and a strong association between G6PD activities and proportion of G6PD normal RBC in CuCl-treated models and in ex vivo RBC. However, in terms of oxidative stress markers analyses, unlike the hypothesis where the lower G6PD activity, the higher MDA and the lower GSH level, the CuCl RBC model showed that in low G6PD activities (10-30%) cells, the MDA level is lower compared to the rest of the models (p<0.05). The ex vivo models however were in line with the hypothesis, although the result was not significant (p=0.5). There was a significant difference between RBC with <60% and those with >80% G6PD activities in CuCl RBC model, but not in ex vivo RBC (p=0.5). Genotyping heterozygous subjects showed G6PDViangchan variant with 2.97U/gHb (33% activity) and 6.58 U/gHb (74% activity).
Conclusions The GSH analysis has pointed to the 60% G6PD activity cut-off and this data is supportive of the old World Health Organization threshold for intermediate upper limit of 60% G6PD activity. However, there are significant limitations in using MDA assay with CuCl RBC model because the RBC was already stressed due to the copper treatment and thus present a different result when compared to the ex-vivo model.
Figure 1
Figure 2
Figure 3
Figure 4