[Association of heterozygote drepanocytosis and G6PD deficiency: apropos of a case].
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Child↗
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Adult-onset Still's disease associated with G6PD deficiency: a case report and literature review.
A 37-year-old man presented with symptoms consistent with adult-onset Still's disease. Fever and leukocytosis were prominent, and the patient was started on high-dose aspirin for possible acute rheumatic fever. He developed severe anemia as a result of glucose-6-phosphate dehydrogenase deficiency. His treatment was changed to naproxen, and he recovered with restoration of his hematologic parameters. Although Still's disease is frequently accompanied by mild-to-moderate anemia, the development of severe anemia should raise the possibilities of hemolysis secondary to glucose-6-phosphate dehydrogenase deficiency.
Adult↗
[Molecular analysis of glucose-6-dehydrogenase deficiency in Spain].
PURPOSE: G6PD deficiency is the most frequent enzymopathy-producing genetic polymorphism in humans. Up to now, over 400 putative variants of G6PD have been distinguished on the basis of biochemical characterization of the deficient enzyme. Analysis of the G6PD gene has made possible a precise classification of the G6PD molecular variants by identification of about 80 different point mutations causing much of the phenotypic heterogeneity. In the Spanish population, the analysis of G6PD has led to the identification of 15 different point mutations that underlay the phenotypic heterogeneity of G6PD previously reported by biochemical analysis. The purpose of the study has been to identify the genetic mutation responsible of the G6PD deficiency and to improve the knowledge of its genetic homogeneity. PATIENTS AND METHODS: From 50 Spanish males with G6PD deficiency 34 came from out consultation and 16 from the Spanish Study Group on Red Cell Pathology (GEHBTA-Eritropatología) The methods employed included screening of prevalent mutations by ER-PCR, SSCP-PCR, genetic segmentation and biochemical characterization of the deficient enzyme. RESULTS: In 31 cases the mutations were characteristic of the four most frequent polymorphic variants found in Spain (G6PD A-376G/202A, G6PD Mediterranean 563T G6PD Union 1360T and G6PD Seattle 344C). Since these mutations either create or abolish a specific site recognized by a restriction endonuclease (RE), they can be rapidly detected by RE digestion of a PCR-amplified product (PCR-RE). In patients where none of these mutations were present (17 cases), the G6PD gene was subjected to PCR single-strand conformation polymorphism (PCR-SSCP) analysis combined with direct PCR-sequencing. By using this procedure, 9 new mutations have been identified, five of them have been also found in other geographical areas and were associated with favism (G6PD A-376G/968C, G6PD Santamaria 376G/542T, G6PD Aures 143C and G6PD Chatham 1003A) or chronic haemolytic anaemia (G6PD Tomah 1153C). The other four mutations are unique and not reported so far: Three of them are associated with favism (G6PD Málaga 542T, G6PD Murcia 209G and G6PD Valladolid 406T) and one with chronic haemolytic anaemia (G6PD Madrid 1155G). The remaining eight cases are under study. CONCLUSION: The present study confirms the marked genetic heterogeneity of G6PD deficiency in Spain and demonstrate that the PCR-RE analysis is an easy tool for rapid diagnosis of the molecular defect in subjects with the common forms of G6PD deficiency. Furthermore the fact that G6PD A-376G/202A is the most common variant within Spanish population and the finding of G6PD Aures 43C and G6PD Santamaría 76G/542T, who are polymorphic in Algeria is consistent with a significant gene flow from Africa to Europe through Spain.
Africa, Northern↗
Multiple molecular forms of glucose-6-phosphate dehydrogenase in normal, preneoplastic, and neoplastic mammary tissues of mice.
Multiple molecular forms of glucose-6-phosphate dehydrogenase (G6PD) in normal, preneoplastic, and neoplastic mammary tissues were separated by polyacrylamide gel electrophoresis and identified by specific straining for enzyme activity. Mammary tissue from lactating BALB/c mice showed considerable amounts (up to 50%) of a slower-migrating G6PD species, G6PD-III, which was essentially absent from glands of pregnant mice, preneoplastic nodules, and mammary carcinomas. All tissues possessed a faster-migrating species, G6PD-II, which accounted for up to 85% of the total G6PD in the glands of pregnant mice. A third species, G6PD-I, migrating more rapidly than G6PD-II, was found in both abnormal tissues (preneoplastic and neoplastic) and accounted for up to 35% of the total enzymatic activity. G6PD-I was present in moderate amounts (less than 15%) in glands from pregnant mice and was essentially absent from the lactating gland (approximately 5%). The addition of dithiothreitol did not alter the measurable G6PD activity but did increase the relative activity of G6PD-II or G6PD-I, as judged by the intensity of the bands on the gels. Mild oxidation (stirring overnight at 4 degrees in air) resulted in a loss of G6PD activity, but preparations had greater amounts of G6PD-III; presence of dithiothreitol during aeration partially prevented loss of G6PD activity and largely prvented the appearance of G6PD-III. Molecular-weight estimations with preparations from lactating mice yielded a value of 118,000 for G6PD-II and 260,000 for G6PD-III, suggesting a monomer and dimer, respectively. The addition of nicotinamide adenine dinucleotide phosphate stabilized G6PD activity by preventing heat inactivation at 47 degrees; nicotinamide adenine dinucleotide phosphate did not alter the pattern of species present. The data from heat inactivation studies suggest that G6PD-III (dimer) was the more stable species. The addition of nicotinamide adenine dinucleotide phosphate to samples after oxidation in the absence of dithiothreitol (about 70% loss of activity) resulted in no change in patterns and in recovery of full G6PD activity during heating at 47 degrees. A potential relationship between glutathione reductase activity and the pattern of G6PD species observed in the various tissues is noted.
Animals↗
Independent origin of single and double mutations in the human glucose 6-phosphate dehydrogenase gene.
The vast majority of both polymorphic and sporadic G6PD variants are due to single missense mutations. In the four polymorphic variants that have two point mutations, one of the mutations is always 376 A-->G (126 Asn-->Asp), which on its own gives rise to the nondeficient polymorphic variant, G6PD A. In a study of G6PD deficient patients who presented with clinical favism in Spain, we have found a new polymorphic variant that we have called G6PD Malaga, whose only abnormality is a 542 A-->T (181 Asp-->Val) mutation. This is the same mutation as previously found in association with the mutation of G6PD A in the double mutant, G6PD Santamaria. G6PD Malaga is associated with enzyme deficiency (class III), and the enzymic properties of G6PD Malaga and G6PD Santamaria are quite similar, indicating that in this case the effects of the two mutations are additive rather than synergistic. G6PD Santamaria might have been produced by recombination between G6PD A and G6PD Malaga; however haplotype analysis, including the use of a new silent polymorphism, suggests that the same 542 A-->T mutation has taken place independently in a G6PD B gene to give G6PD Malaga and in a G6PD A gene to give G6PD Santamaria. These findings help to outline the relationship and evolution of mutations in the human G6PD locus.
Amino Acid Sequence↗
Further investigations of glucose-6-phosphate dehydrogenase variants in Flores Island, eastern Indonesia.
We conducted field surveys for malaria and glucose-6-phosphate dehydrogenase (G6PD) deficiency in the eastern part of Flores Island, East Nusa Tenggara Province, Indonesia. A total of 1,108 volunteers (642 males and 466 females) belonging to three ethnic groups (Sikka, Ende and Bajo) were examined, and 55 G6PD-deficient individuals (38 males and 17 females) were detected. Among them, 50 samples were analyzed molecularly, in addition to three deficient cases in a Bajo family. In the Sikka population, G6PD Kaiping (1388G>A), one of the two common variants in the Chinese population, was unexpectedly found as the most dominant variant (11/22, 50.0%), followed by G6PD Chatham (1003G>A, 36.4%), G6PD Coimbra (592C>T, 9.1%) and G6PD Vanua Lava (383T>C, 4.5%). Frequency of G6PD Kaiping in the Sikka might be the highest among non-Chinese populations reported so far. In the Ende population, G6PD Vanua Lava (9/14, 64.3%) was the highest, followed by G6PD Kaiping (14.3%), G6PD Chinese-5 (1024C>T, 14.3%) and G6PD Chatham (7.1%). In the Bajo population, a total of 18 deficient cases were analyzed, and a novel mutation (844G>T) in exon 8 with a predicted amino acid change of 282 Asp>Tyr was found in a 7-year-old boy at a Bajo village near Maumere. This new Class II (mild type) variant was also confirmed in his mother and sister, and designated as G6PD Bajo Maumere. The missense mutation at the same nucleotide 844 has been known as G6PD Seattle/Lodi/Modena/Ferrara II, but this mutation is caused by a G>C substitution (282 Asp>His). In the Bajo population, G6PD Viangchan (871G>A, IVS 11 nt93 T>C, 1311C>T), the most common variant in continental Southeast Asian populations, was found to be the dominant (11/18, 61.1%), followed by G6PD Vanua Lava and the new variant (each 16.7%), and G6PD Coimbra (5.6%). These results strongly suggest that the Bajo peoples may have different ancestors from those for Sikka and Ende, and may be much closer to continental Southeast Asian populations. It is interesting that G6PD Canton (1376G>T), another common variant in Chinese, was not seen in the Flores population.
DNA Mutational Analysis↗
Glucose-6-phosphate dehydrogenase modulates vascular endothelial growth factor-mediated angiogenesis.
Glucose-6-phosphate dehydrogenase (G6PD), the first enzyme of the pentose phosphate pathway, is the principal intracellular source of NADPH. NADPH is utilized as a cofactor by vascular endothelial cell nitric-oxide synthase (eNOS) to generate nitric oxide (NO*). To determine whether G6PD modulates NO*-mediated angiogenesis, we decreased G6PD expression in bovine aortic endothelial cells using an antisense oligodeoxynucleotide to G6PD or increased G6PD expression by adenoviral gene transfer, and we examined vascular endothelial growth factor (VEGF)-stimulated endothelial cell proliferation, migration, and capillary-like tube formation. Deficient G6PD activity was associated with a significant decrease in endothelial cell proliferation, migration, and tube formation, whereas increased G6PD activity promoted these processes. VEGF-stimulated eNOS activity and NO* production were decreased significantly in endothelial cells with deficient G6PD activity and enhanced in G6PD-overexpressing cells. In addition, G6PD-deficient cells demonstrated decreased tyrosine phosphorylation of the VEGF receptor Flk-1/KDR, Akt, and eNOS compared with cells with normal G6PD activity, whereas overexpression of G6PD enhanced phosphorylation of Flk-1/KDR, Akt, and eNOS. In the Pretsch mouse, a murine model of G6PD deficiency, vessel outgrowth from thoracic aorta segments was impaired compared with C3H wild-type mice. In an in vivo Matrigel angiogenesis assay, cell migration into the plugs was inhibited significantly in G6PD-deficient mice compared with wild-type mice, and gene transfer of G6PD restored the wild-type phenotype in G6PD-deficient mice. These findings demonstrate that G6PD modulates angiogenesis and may represent a novel angiogenic regulator.
Animals↗
Distribution of glucose-6-phosphate dehydrogenase mutations in Southeast Asia.
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a heterogeneous enzyme abnormality with high frequency in tropical areas. We performed population screening and molecular studies of G6PD variants to clarify their distribution and features in Southeast Asia. A total of 4317 participants (2019 males, 2298 females) from 16 ethnic groups in Myanmar, Lao in Laos, and Amboinese in Indonesia were screened with a single-step screening method. The prevalence of G6PD-deficient males ranged from 0% (the Akha) to 10.8% (the Shan). These G6PD-deficient individuals and 12 G6PD-deficient patients who had been diagnosed at hospitals in Indonesia and Malaysia were subjected to molecular analysis by a combination of polymerase-chain-reaction-based single-strand conformation polymorphism analysis and direct sequencing. Ten different missense mutations were identified in 63 G6PD-deficient individuals (50 hemizygotes, 11 heterozygotes, and 2 homozygotes) from 14 ethnic groups. One missense mutation (1291 G-->A) found in an Indonesian Chinese, viz., G6PD Surabaya, was previously unknown. The 487 G-->A (G6PD Mahidol) mutation was widely seen in Myanmar, 383 T-->C (G6PD Vanua Lava) was specifically found among Amboinese, 871 G-->A (G6PD Viangchan) was observed mainly in Lao, and 592 C-->T (G6PD Coimbra) was found in Malaysian aborigines (Orang Asli). The other five mutations, 95 A-->G (G6PD Gaohe), 1003 G-->A (G6PD Chatham), 1360 C-->T (G6PD Union), 1376 G-->T (G6PD Canton), and 1388 G-->A (G6PD Kaiping) were identified mostly in accordance with distributions reported previously.
Amino Acid Substitution↗
Molecular heterogeneity of the glucose-6-phosphate dehydrogenase deficiency in the Hellenic population.
We report results from a systematic study to identify the molecular basis of glucose-6-phosphate dehydrogenase (G6PD) deficiency on a sample of 299 male subjects from the Hellenic population. Our stepwise approach involved partial biochemical characterization and quantitation of the enzyme's activity, MboII restriction endonuclease digestion to identify the G6PD Mediterranean variant, which represents the most frequent G6PD variant in our population and a nonradioactive polymerase chain reaction-single-strand conformation polymorphism methodology for the detection of the underlying molecular defect(s) in the rest of the non-Mediterranean G6PD-deficient individuals. Through this approach, six different G6PD variants were identified (G6PD Mediterranean, G6PD Hermoupolis, G6PD Cassano, G6PD Seattle, G6PD Ierapetra and G6PD Acrokorinthos), two of which were new (G6PD Hermoupolis, G6PD Acrokorinthos). In essence, this study underlines the remarkable genetic heterogeneity of the G6PD deficiency in the Hellenic population, while the finding of the double mutant, G6PD Hermoupolis, may help to outline the relationship and evolution of mutations in the human G6PD locus.
Exons↗
Glucose-6 phosphate dehydrogenase deficiency decreases the vascular response to angiotensin II.
BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) regulates production of the reduced form of NADPH through the pentose phosphate pathway. G6PD may therefore affect superoxide anion production via vascular NADPH oxidase, which is key in mediating the vascular response to angiotensin II (Ang II). We determined the hypertensive and vascular hypertrophic response to Ang II in G6PD-deficient mice. METHODS AND RESULTS: Ang II (0.7 mg/kg per day) was infused via subcutaneous osmotic pumps for 6 days in male hemizygote G6PD mutant (G6PD(mut)) and wild-type (WT) C3H mice. (1) Compared with WT, G6PD(mut) mouse aorta had 10% to 20% of G6PD activity and 50% less NADPH. (2) Basal systolic blood pressure was not significantly different in G6PD(mut) mice (WT 88+/-4 mm Hg versus G6PD(mut) 95+/-4 mm Hg), but Ang II increased blood pressure to a lower level in G6PD(mut) mice (WT 139+/-4 mm Hg versus G6PD(mut) 123+/-5 mm Hg; P<0.05). (3) Ang II increased aortic medial thickness less in G6PD(mut) mice (WT 71+/-2 mum versus G6PD(mut) 62+/-1 mum; P<0.01). (4) 3-o-Nitrotyrosine staining and dihydroethidium oxidation in the aorta was increased by Ang II less in G6PD(mut) mice. (5) Smooth muscle cells isolated from G6PD(mut) mice showed less Ang II-induced phosphorylation of Akt and p42/44 ERK. CONCLUSIONS: G6PD deficiency may reduce vascular superoxide anion production by limiting production of the substrate for NADPH oxidase, thereby inhibiting oxidant-mediated Ang II-induced signaling pathways that contribute to hypertension and smooth muscle hypertrophy.
Angiotensin II↗
[Identification of molecular variants of the enzyme glucose-6-phosphate dehydrogenase by the polymerase chain reaction technique].
BACKGROUND: An assessment of the usefulness of polymerase chain reaction (PCR) in the identification of the most frequent molecular variants of the glucose-6-phosphate dehydrogenase (G6PD) in Spain: G6PD A-, G6PD Mediterranean and G6PD Seattle through the screening of the mutations: 376 A-->G; 202 G-->A; 680 G-->T; 968 T-->C; 563 C-->T and 844 G-->C. METHODS: Three groups of patients have been studied: 1) males (40 cases); 2) relatives from the preceding group (31 cases: 7 males and 24 females), and 3) samples classified according to their fast electrophoretic mobility as G6PD A-(17 cases). The method used has been the PCR followed by digestion with specific restriction endonucleases. RESULTS: Group 1: 23 out of 40 samples (57%), were identified as G6PD Med563T variant (8 cases), G6PD A-376G/202A (13 cases) and G6PD Seattle844C (2 cases). Group 2: The study of relatives from 13 of the 23 identified samples allowed the study of additional 31 samples (7 males, 24 females): hemizygous G6PD Med563T (3 cases), heterozygous GdB/Gd Med563T (5 cases), hemizygous G6PD A-376G/202A (4 cases), heterozygous GdB/Gd A-376G/202A (11 cases), heterozygous GdB/Gd Seattle844C (1 case) and normal females (7 cases). Group 3: In all electrophoretically fast samples classified as G6PD A-was detected the 376 A-->G mutation (characteristic of G6PD A+). In 15 of these cases a second mutation was found at nucleotide 202 G-->A (G6PD A-376G/202A); and in two, at nucleotide 968 T-->C (G6PD A-376G/968C). CONCLUSIONS: The PCR method is fast and simple enough to allow the identification of known G6PD deficient variant, avoiding the need of its molecular characterization, which is more cumbersome and time consuming. In addition, the PCR is a very useful tool for demonstrating the carrier condition of G6PD deficiency in females with enzyme activity within normal range.
Base Sequence↗
Genetic heterogeneity at the glucose-6-phosphate dehydrogenase locus in southern Italy: a study on a population from the Matera district.
Glucose-6-phosphate dehydrogenase (G6PD) has been analyzed by gel electrophoresis and by quantitative assay in an unselected sample of 1524 schoolboys from the province of Matera (Lucania) in southern Italy. We have identified 43 subjects with a G6PD variant. Of these, 31 had severe G6PD deficiency, nine had mild to moderate deficiency, and three had a non-deficient electrophoretic variant. The overall rate of G6PD deficiency was 2.6%. The frequency of G6PD deficiency, ranging from 7.2% on the Ionian Coast to zero on the eastern side of the Lucanian Apennines, appears to be inversely related to the distance of each town examined from the Ionian Coast, suggesting that this geographic distribution may reflect, at least in part, gene flow from Greek settlers. Biochemical characterization has shown that most of the G6PD deficiency in this population is accounted for by G6PD Mediterranean. In addition, we have found several examples of two other known polymorphic variants (G6PD Cagliari and G6PD A-); three new polymorphic variants, G6PD Metaponto (class III), G6PD Montalbano (class III), and G6PD Pisticci (class IV); and two sporadic variants, G6PD Tursi (class III) and G6PD Ferrandina (class II). These data provide further evidence for the marked genetic heterogeneity of G6PD deficiency within a relatively narrow geographic area and they prove the presence in the Italian peninsula of a gene (GdA-) regarded as characteristically African.
Child↗
Molecular analysis of eight biochemically unique glucose-6-phosphate dehydrogenase variants found in Japan.
We analyzed the molecular mutations of eight known Japanese glucose-6-phosphate dehydrogenase (G6PD) variants with unique biochemical properties. Three of them were caused by novel missense mutations: G6PD Musashino by 185 C-->T, G6PD Asahikawa by 695 G-->A, and G6PD Kamiube by 1387 C-->T. Predicted amino acid substitutions causing asymptomatic variants G6PD Musashino (62 Pro-->Phe) and G6PD Kamiube (463 Arg-->Cys) were located in regions near the amino or carboxyl end of the polypeptide chain, whereas an amino acid change 232 Cys-->Tyr causing a class 1 variant G6PD Asahikawa was located in the region where amino acid alterations in some class 1 variants were clustered. The other five variants had known missense mutations, namely, G6PD Fukushima, 1246 G-->A, G6PD Morioka, 1339 G-->A, and G6PD Iwate, G6PD Niigata and G6PD Yamaguchi, 1160 G-->A, which cause variants, G6PD Tokyo, G6PD Santiago de Cuba, and G6PD Beverly Hills, respectively.
Anemia, Hemolytic, Congenital Nonspherocytic↗
Molecular heterogeneity of glucose-6-phosphate dehydrogenase deficiency in Malays in Malaysia.
Multiplex polymerase chain reaction (PCR) using multiple tandem forward primers and a common reverse primer (MPTP) was recently established as a comprehensive screening method for mutations in X-linked recessive diseases. In the work reported here, MPTP was used to scan for mutations of the glucose-6-phosphate dehydrogenase (G6PD) gene. Mutations in exons 3,4,5,6,7,9, 11, and 12 of the G6PD gene were screened by MPTP in 93 unrelated Malaysian patients with G6PD deficiency. Of the 93 patients, 80 (86%) had identified mutations. Although all of these were missense mutations, identified nucleotide changes were heterogeneous, with 9 mutations involving various parts of the exons. These 9 mutations were G-to-A nucleotide changes at nucleotide 871 of the G6PD gene (G871A), corresponding to G6PD Viangchan, G6PD Mediterranean (C563T), G6PD Vanua Lava (T383C), G6PD Coimbra (C592T), G6PD Kaiping (G1388A), G6PD Orissa (C131G), G6PD Mahidol (G487A), G6PD Canton (G1376T), and G6PD Chatham (G1003A). Our results document heterogeneous mutations of the G6PD gene in the Malaysian population.
DNA Mutational Analysis↗
Molecular characterization of glucose-6-phosphate dehydrogenase deficiency in the Kurdish population of Western Iran.
A total of 1,000 school boys ages 14-18 years, were randomly selected from six schools of Kermanshah and screened for G6PD deficiency. Fifty-three out of 1,000 were found to be severely G6PD deficient giving a frequency of 5.3% for G6PD deficiency in males in Kermanshah. We performed DNA analysis on 68 G6PD deficient children, 52 school boys and 16 children with the history of favism and hemolytic anemia using a combination of PCR-RFLP technique, single strand conformation polymorphism (SSCP) analysis and DNA sequencing. Three polymorphic G6PD mutations were identified: G6PD Mediterranean, Chatham and Cosenza. The commonest allele was found to be the G6PD Mediterranean (91.2%) followed by the G6PD Chatham (7.3%) and the G6PD Cosenza (1.5%). Haplotype analysis revealed that 56 out of 62 (90%) G6PD Mediterranean was linked to haplotype BclI(+) (1311T). In contrast, all G6PD Chatham and Cosenza had haplotype BclI(-) (1311C). The polymorphism IVS11-93 (T-->C) was present in 88.5% of the subjects studied. Only 4/55 (7.3%) of the Mediterranean alleles were associated with the T form and were always related to the nt 1311C. Our findings indicate that the allele frequency of G6PD Mediterranean mutation in Kermanshah is higher than those from two Fars ethnic groups living in Northern and Southern Iran. Nevertheless, they are in strict accordance with a previous report of the prevalence of the G6PD Mediterranean in Kurdish and Middle East population. Also, of strong association of the G6PD Mediterranean mutation and the presence of the polymorphism nt 1311C-->T in the Kermanshah population demonstrate that the presence of this mutation may be the result of migrations that have taken place through the history of Iran.
Adolescent↗
Molecular study of eight Japanese cases of glucose-6-phosphate dehydrogenase deficiency by nonradioisotopic single-strand conformation polymorphism analysis.
Using a newly developed nonradioisotopic method of polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis combined with the direct sequencing using the fluorescence-labeled terminator, we identified seven missense mutations, 527 A-->G, 1003 G-->A, 1159 C-->T, 1160 G-->A, 1229 G-->A, 1246 G-->A, and 1361 G-->A, in eight Japanese patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Except for the 527 A-->G, each mutation has been reported to cause variants G6PD Chatham, G6PD Guadalajara, G6PD Beverly Hills, G6PD "Japan", G6PD Tokyo, and G6PD Andalus, respectively. In addition, a single base deletion in intron 5 was found in the patients with G6PD Guadalajara or G6PD Andalus. The variant with unique 527 A-->G was characterized and designated as G6PD Shinshu. We also characterized G6PD "Japan" and found that the variant had the striking resemblance with G6PD Riverside, bearing a missense mutation in the same codon, but causing a different amino acid substitution. Our modified PCR-SSCP analysis using minigel and ethidium bromide staining could detect six of the eight diverse mutations in the G6PD gene. Because it is easy and requires no special apparatus, this modified method will be useful for screening mutations in the G6PD gene.
Base Sequence↗
Genetic heterogeneity of glucose-6-phosphate dehydrogenase deficiency revealed by single-strand conformation and sequence analysis.
We have carried out a systematic study of the molecular basis of glucose-6-phosphate dehydrogenase (G6PD) deficiency on a sample of 53 male subjects from Calabria, in southern Italy. Our sequential approach consisted of the following steps: (1) Partial biochemical characterization was used to pinpoint candidate known variants. The identity of these was then verified by restriction-enzyme or allele-specific oligonucleotide hybridization analysis of the appropriate PCR-amplified fragment. (2) On samples for which there was no obvious candidate mutation, we proceeded to amplify the entire coding region in eight fragments, followed by single-strand conformation polymorphism (SSCP) analysis of each fragment. (3) The next step was M13 phage cloning and sequencing of those individual fragments that were found to be abnormal by SSCP. Through this approach we have identified the molecular lesion in 51 of the 53 samples. In these we found a total of nine different G6PD-deficient variants, five of which (G6PD Mediterranean, G6PD A-, G6PD Coimbra, G6PD Seattle, and G6PD Montalbano) were already known, whereas four are new (G6PD Cassano, G6PD Cosenza, G6PD Sibari, and G6PD Maewo). G6PD Mediterranean is the commonest variant, followed by G6PD Seattle. At least seven of the variants are present, at polymorphic frequencies, in the Calabria region, and some have a nonrandom distribution within the region. This study shows that the genetic heterogeneity of G6PD deficiency in Calabria, when analyzed at the DNA level, is even greater than had been anticipated from biochemical characterization. The sequential approach that we have followed is fast and efficient and could be applied to other populations.
Base Sequence↗
The relationship between glucose-6-phosphate dehydrogenase deficiency and cataracts in Sardinia. An epidemiological and biochemical study.
The Italian island of Sardina occupies an important position on the map of glucose-6-phosphate dehydrogenase (G6PD)-deficiency distribution throughout the world, since in this region the condition is particularly frequent and severe (erythrocytes show only 0-7% of G6PD normal activity, while people result affect up to 35% depending on the district). In order to investigate the relationship between the deficiency of G6PD in erythrocytes and in lens, and cataractogenesis, we studied 2125 idiopathic cataractous and non-cataractous subjects, both G6PD-deficient and normal, males and females. Parameters investigated included incidence, distribution and type of cataracts, age at the moment of the first observation, geographical provenance, and G6PD activity in erythrocytes. Moreover, G6PD activity and glutathione (GSSG)-reducing activity was assessed in cataractous lenses obtained from deficient and normal individuals. G6PD deficiency was found to be significantly more frequent in males of the age-group 40-49 years (P = 0.025), while the frequency of G6PD deficiency was decisively lower in the older age-groups. In females, mainly heterozygotes, no evidence of such a relation was found. Cataractous lenses obtained from male patients with no G6PD activity in erythrocytes showed undetectable levels of G6PD activity, and lowered, but not extinguished, levels of GSSG-reducing activity. Cataractous lenses from heterozygous females showed intermediate levels of G6PD activity and GSSG-reducing activity. A preliminary study of 182 diabetic, G6PD-deficient and non-deficient subjects, failed to demonstrate that Sardinian variants of G6PD deficiency provide protection against cataract formation in diabetic patients.
Adult↗