Membrane deformability of erythrocytes with glucose-6-phosphate dehydrogenase Hamburg.

Deformability of erythrocytes of a patient with chronic nonspherocytic hemolytic anemia caused by a G-6-PD variant (G-6-PD Hamburg) in red cells was studied. The flow rate of erythrocytes from this patient through 5 mum polycarbonate sieves was higher than that of red cells from healthy adults. Even under oxidative stress in vitro, the deformability of erythrocytes of the patient was only slightly decreased. The residual hemoglobin content of red cell membranes (ghosts) from this patient was lower than that of ghosts from healthy adults and of ghosts prepared from comparable reticulocyte-rich blood withoug G-6-PD deficiency. In contrast to the low enzymatic activity of G-6-PD Hamburg in the hemolysate, a high activity of this enzyme could be demonstrated in the ghosts from the patient. In view of the flexibility of this patient's erythrocytes splenectomy is not to be recommended.     

Glucose-6-phosphate dehydrogenase mutations and haplotypes in Mexican Mestizos

In a screening for glucose-6-phosphate dehydrogenase (G-6-PD) deficiency in 1985 unrelated male subjects from the general population (Groups A and B) belonging to four states of the Pacific coast, 21 G-6-PD-deficient subjects were detected. Screening for mutations at the G-6-PD gene by PCR-restriction enzyme in these 21 G-6-PD-deficient subjects as well as in 14 G-6-PD-deficient patients with hemolytic anemia belonging to several states of Mexico showed two common G-6-PD variants: G-6-PD A-(202A/376G) (19 cases) and G-6-PD A-(376G/968C) (9 cases). In 7 individuals the mutations responsible for the enzyme deficiency remain to be determined. Furthermore, four silent polymorphic sites at the G-6-PD gene (PvuII, PstI, 1311, and NlaIII) were investigated in the 28 individuals with G-6-PD A- variants and in 137 G-6-PD normal subjects. As expected, only 10 different haplotypes were observed. To date, in our project aiming to determine the molecular basis of G-6-PD deficiency in Mexico, 60 unrelated G-6-PD-deficient Mexican males-25 in previous studies and 35 in the present work-have been studied. More than 75% of these individuals are from states of the Pacific coast (Sinaloa, Nayarit, Jalisco, Michoacán, Guerrero, Oaxaca, and Chiapas). The results show that although G-6-PD deficiency is heterogeneous at the DNA level in Mexico, only three polymorphic variants have been observed: G-6-PD A-(202A/376G) (36 cases), G-6-PD A-(376G/968C) (13 cases), and G-6-PD Seattle(844C) (2 cases). G-6-PD A- variants are relatively distributed homogeneously and both variants explain 82% of the overall prevalence of G-6-PD deficiency. The variant G-6-PD A-(202A/376G) represents 73% of the G-6-PD A- alleles. Our data also show that the variant G-6-PD A-(376G/968C)-which has been observed in Mexico in the context of two different haplotypes-is more common than previously supposed. The three polymorphic variants that we observed in Mexico are on the same haplotypes as found in subjects from Africa, the Canary Islands, and Spain.     

Red Cell Glucose-6-Phosphate Dehydrogenase Deficiency in Finland

Severe red cell glucose-6-phosphate dehydrogenase (G-6-PD) deficiency has been found in an ‘aboriginal’ Finnish family. 2 male and 9 female carriers of the variant G-6-PD were studied. The genetic pattern is consistent with x-linked recessive inheritance and the defect is associated with drug (primaquine) induced haemolysis. This was demonstrated by enzyme deficient red cell (⁵¹Cr-labelled) survival studies on a normal volunteer recipient. In addition, one of the hemizygotes studied had a slight chronic nonspherocytic haemolytic disorder. The partially purified enzyme had many of the characteristics of G-6-PD Mediterranean. The occurrence of this G-6-PD Mediterranean type variant in the Finnish population, which differs greatly from Mediterranean ethnic groups, as well as the association of slight chronic haemolysis with severe G-6-PD deficiency is discussed.     

Hematological parameters and red blood cell morphological abnormality of Glucose-6-Phosphate dehydrogenase deficiency co-inherited with thalassemia

Objective/Background

Glucose-6-phosphate dehydrogenase (G-6-PD) deficiency and thalassemia are genetically independent hemolytic disorders. Co-inheritance of both disorders may affect red blood cell pathology to a greater extent than normally seen in either disorder alone. This study determines the prevalence and evaluates hematological changes of G-6-PD deficiency and thalassemia co-inheritance.

Molecular heterogeneity of glucose-6-phosphate dehydrogenase deficiency in Mexico: overall results of a 7-year project

Several years ago, a project aiming to determine (i) the molecular basis of G-6-PD deficiency, (ii) the distribution of four different mutant alleles previously detected, and (iii) the whole of polymorphic alleles that account for the overall prevalence of G-6-PD deficiency in Mexico was implemented. Nearly 5000 individuals-from the general population and patients with hemolytic anemia-belonging to at least 14 States were screened for G-6-PD deficiency. Seventy-six G-6-PD-deficient subjects were detected and the prevalence of G-6-PD deficiency in 4777 individuals from the general population was 0.71%. Screening for both mutations associated with enzyme deficiency and silent polymorphisms at the G-6-PD gene was performed in the enzyme-deficient individuals by PCR-SSCP combined with restriction enzyme analysis; the silent polymorphisms as well as the nondeficient variant G-6-PD A(376G) were also investigated in 366 G-6-PD normal individuals from the general population. In 88% of the enzyme-deficient individuals it was possible to define the mutation responsible and the type G-6-PD A- variants were the more common in both individuals from the general population and patients with hemolytic anemia. G-6-PD deficiency is heterogeneous at the DNA level in Mexico and up to date 10 different variants-8 in the present project and 2 previously-have been observed: G-6-PD A(-202A/376G), G-6-PD A(-376G/968C), G-6-PD Santamaria(376G/542T), G-6-PD Vanua Lava(383C), G-6-PD Tsukui(del561-563), G-6-PD "Mexico City"(680A), G-6-PD Seattle(844C), G-6PD Guadalajara(1159T),G-6-PD Nashville(1178A), and G-6-PD Union(1360T). The G-6-PD A(-) variants have a relatively homogeneous distribution and along with G-6-PD Santamaria(376G/542T), they account for 82% of the overall prevalence of G-6-PD deficiency in Mexico; all other seven variants represent 9% of the mutant alleles examined, and in the rest of the chromosomes the mutation responsible for the enzyme deficiency remains to be defined. Several of the variants observed in Mexico are common in Africa, South Europe, and Southeast Asia. The prevalence for the variant G-6-PD A(376G) was 1.64%. From 256 possible haplotypes only 14 were observed and haplotype analysis suggests that some of the G-6-PD variants probably were imported to Mexico by population flow from South Europe, Africa, and Southeast Asia. This work (i) identified the G-6-PD variants prevalent in Mexico, (ii) defines their geographical distribution, (iii) contributes to the knowledge of the genetic structure of the Mexican population, and (iv) will facilitate the molecular analysis of the G-6-PD gene in enzyme-deficient Mexican individuals.     

Red cell glucose-6-phosphate dehydrogenase deficiency in Finland. Characterization of a new variant with severe enzyme deficiency.

Severe red cell glucose-6-phosphate dehydrogenase (G-6-PD) deficiency has been found in an 'aboriginal' Finnish family. 2 male and 9 female carriers of the variant G-6-PD were studied. The genetic pattern is consistent with x-linked recessive inheritance and the defect is associated with drug (primaquine) induced haemolysis. This was demonstrated by enzyme deficient red cell (51Cr-labelled) survival studies on a normal volunteer recipient. In addition, one of the hemizygotes studied had a slight chronic nonspherocytic haemolytic disorder. The partially purified enzyme had many of the characteristics of G-6-PD Mediterranean. The occurrence of this G-6-PD Mediterranean type variant in the Finnish population, which differs greatly from Mediterranean ethnic groups, as well as the association of slight chronic haemolysis with severe G-6-PD deficiency is discussed.     

Anemia in Patients with Coinherited Thalassemia and Glucose-6-Phosphate Dehydrogenase Deficiency

Thalassemia and glucose-6-phosphate dehydrogenase (G-6-PD) deficiency are genetic disorders that cause hemolytic anemia. In areas with high frequencies of both hematological disorders, coinheritance of G-6-PD deficiency with thalassemia can be found. Whether G-6-PD deficiency, coinherited with thalassemia, enhances severe anemia is still unclear. Hematological parameters between thalassemia carriers with G-6-PD deficiency and those without G-6-PD deficiency were compared. The G-6-PD deficiency was diagnosed in 410 blood samples from thalassemia patients using a fluorescent spot test. The levels of hemoglobin (Hb), packed cell volume (PCV), mean corpuscular volume (MCV) and Hb A₂/Hb E [β26(B8)Glu→Lys; HBB: c.79G>A] were measured using an automated blood counter and high performance liquid chromatography (HPLC), respectively. The G-6-PD deficiency was found in 37 samples (9.02%). Mean levels of Hb, PCV, MCV and Hb A₂/E were similar between the two groups. Thus, G-6-PD deficiency did not enhance red blood cell pathology or induce more anemic severity in thalassemia patients.     

Glucose-6-Phosphate Dehydrogenase Deficiency in Saudi Arabia: A Survey

A survey of red cell G-6-PD deficiency has been conducted among 1296Saudi subjects. Thirteen per cent of randomly selected male subjects and 2.4per cent of the females tested were found to be G-6-PD deficient. In this study,the screening test employed did not detect female (partially G-6-PD deficient)heterozygotes. The deficiency appeared to be localized to the oasis of theEastern province, and in village studies a very high incidence was found inmale children from the Qatif and Al-Hasa oases. The geographic limits ofG-6-PD deficiency correspond precisely to the areas known to be hyperendemic for malaria in previous years. However, the population group affected represents a distinct minority in terms of religious and cultural tradition, and anthropometric type. In Saudi Arabia, falciparum malaria does notappear to be the only significant factor determining marked regional differences in the incidence of G-6-PD deficiency: this genetic marker is essentially confined to the Shiite muslim population. The Sunni population, regardless of its proximity to areas of endemic falciparum malaria, has a lowincidence of G-6-PD deficiency.

Submitted on October 17, 1963 Accepted on July 2, 1964     

Three new variants of glucose-6-phosphate dehydrogenase associated with chronic nonspherocytic hemolytic anemia: G-6-PD Lincoln Park, G-6-PD Arlington Heights, and G-6-PD West Town

Glucose-6-phosphate dehydrogenase (G-6-PD) deficiency was identified in three children who were evaluated because of chronic nonspherocytic hemolytic anemia. One child is of German extraction, another Puerto Rican, and the third Mexican. In each of the patients the hemolytic process was well compensated, but each had one or more episodes of anemia following exposure to an oxidant drug or with infections. The electrophoretic, functional, and kinetic properties of the mutant enzymes, derived both from the patients' erythrocytes and from cultured fibroblasts, allowed each to be distinguished from G-6-PD variants previously described.     

Hereditary Hemolytic Disease Secondary to Glucose-6-Phosphate Dehydrogenase Deficiency: Report of Three Cases with Special Emphasis on ATP Metabolism

1. Three cases of hereditary hemolytic disease secondary to G-6-PD deficiency are described. Two of the cases were first cousins of Scotch-Irish-English descent and the mode of inheritance was believed to be sex-linked.The third case was of Turkish origin; no family studies were availale.

2. The mothers, who were heterozygous for G-6-PD deficiency, showed onlyminimal expression of the defect, which was manifested by a slightly decreasedred cell survival in both mothers and an abnormal methemoglobin reductiontest in one of them.

3. All three cases showed a more pronounced fall in erythrocyte ATP afterincubation with phenylhydrazine than that observed in primaquine-sensitiveNegroes whose red cells were less deficient in G-6-PD.

4. It is suggested that the inability of the G-6-PD-deficient erythrocyte tomaintain adequate levels of ATP may be an important factor in the pathogenesis of the hemolytic process.

Submitted on August 26, 1963 Accepted on October 24, 1963     

Inhibitory effect of a fava bean component on the in vitro development of Plasmodium falciparum in normal and glucose-6-phosphate dehydrogenase deficient erythrocytes

We examined the hypothesis that G-6-PD deficiency associated with fava bean ingestion confers resistance to malaria by studying the in vitro interactions between malaria parasites (Plasmodium falciparum), human erythrocytes with varying degrees of G-6-PD deficiency, and isouramil (IU), a fava bean extract that is known to cause oxidant stress and hemolysis of G-6-PD-deficient erythrocytes. Untreated G-6-PD-deficient and normal erythrocytes supported the in vitro growth of P. falciparum equally well. However, after pretreatment with IU, G-6-PD-deficient erythrocytes did not support parasite growth in vitro, whereas growth remained high in normal erythrocytes. Parasite growth was proportional to the G-6-PD activity of the IU-treated erythrocytes. In contrast, when parasitized erythrocytes were exposed to IU, parasites even in normal erythrocytes were destroyed. Ring forms were much less sensitive than late trophozoites and schizonts. The results suggest that there are two modes by which IU affects the development of P. falciparum and demonstrate in vitro that G-6-PD deficiency confers resistance against malaria under conditions of fava-bean-associated oxidant stress.     

A new glucose 6-phosphate dehydrogenase variant (G-6-PD Verona) in a patient with myelodysplastic syndrome

A 67-year-old woman investigated because of 'myelodysplastic syndrome' was found to have a 4-fold increase in G-6-PD activity in her erythrocytes. The enzyme was partially purified and characterized. On grounds of: (a) reduced electrophoretic mobility, (b) abnormal cathodic band(s) in isoelectrofocusing, (c) increased Michaelis constant for glucose 6-phosphate, (d) abnormal thermostability, and (e) abnormal interaction with the ligand NADPH, we conclude that this is a new structural variant which we designate G-6-PD Verona. G-6-PD Verona was the sole apparent source of G-6-PD activity in the patient's erythrocytes; by contrast, the patient's fibroblasts had only normal G-6-PH (type B). The patient's haematological course terminated into acute myeloid leukaemia. We believe G-6-PD Verona was the result of a somatic mutation in an X-chromosome which took place in a haemopoietic cell clone which subsequently underwent neoplastic transformation.     

Hemoglobin E and glucose-6-phosphate deficiency in the Khmer Air Force (Cambodia).

This study was designed to investigate the use of primaquine in malaria control in the Khmer Republic (Cambodia). Blood was drawn from 106 male Khmer Air Force troops for analysis of hemoglobin E and glucose-6-phosphate (G-6-PD) deficiency. A test group of 15 men with G-6-PD deficiency and a group of 31 normal controls were given 15 mg of primaquine each morning for 14 days. The patients were followed for hemoglobinuria and changes in hematocrit. Primaquine induced a significant, but not a dangerous, hemolysis in G-6pD-deficient Khmer troops. The G-6-PD deficiency seen in Khmer Air Force subjects was G-6-PD Mahidol. Statistically, G-6-PD Mahidol was linked to hemoglobin E.     

Glycosylated hemoglobin in Saudi sickle cell patients with glucose-6-phosphate dehydrogenase deficiency

Glycosylated hemoglobin (Hb) was measured in 42 Saudi sickle cell patients and in 38 sickle cell patients with glucose-6-phosphate dehydrogenase (G-6-PD) deficiency. A statistically significant decrease in the percentages of glycosylated Hb was found in patients with sickle cell anemia when compared with those SS subjects with G-6-PD deficiency. Since glycosylated Hb is considered as an index of the red blood cell's life span, it is suggested that the enzyme defect in these patients is possibly ameliorating the severity of sickle cell anemia.     

G-6-PD Tripler: A Unique Variant Associated with Chronic Hemolytic Disease

A 52-year-old Caucasian was hospitalized for recurring jaundice. Laboratorytests established a compensated hemolytic anemia due to partial erythrocyteG-6-PD deficiency. Biochemical characterization of the affected enzyme uncovered a unique variant: G-6-PD Tripler. Its significant properties includemarked thermal instability, an electrophoretic mobility slower than the normalB variant in all systems studied, and slightly increased affinity for the substrate G-6-P.

Submitted on February 9, 1970 Revised on February 9, 1970 Accepted on February 17, 1970     

Detection of glucose-6-phosphate dehydrogenase deficient heterozygotes

Heterozygotes for the Mediterranean type of severe G-6-PD deficiencywere investigated by a variety of tests.

The methemoglobin reduction test was most successful in detecting heterozygotes (about 80 per cent). Enumeration of methemoglobin containingcells on blood films (Kleihauer-Betke technic) did not improve these results.Quantitation of enzyme level was less successful (65 per cent), and determination of decolorization time by the BCB technic was least sensitivein heterozygote detection.

Methemoglobin reduction technics reflect a more indirect effect of themutant gene than enzyme assay. The superiority of these technics in heterozygote detection is probably caused by the genetically determined presenceof both normal and enzyme-deficient cells in G-6-PD deficient heterozygotes.Since methemoglobin reduction is carried out by individual cells, the population of enzyme deficient cells does not reduce methemoglobin, and thereforeeven a minority of deficient cells leads to abnormal test results. In contrast,enzyme assay is less successful for heterozygote detection, since measurementof enzyme level is carried out on hemolyzed red cells, where cellular mosaicismno longer exists.

An additional source of variation of enzyme levels in heterozygotes iscaused by the existence of genetically determined control of normal enzymelevel. Possession of a high capacity allele for G-6-PD activity may placea heterozygote in the normal range of enzyme activity.

The various tests were also applied to subjects with the mild Greek typeof G-6-PD deficiency. Males with this mutation had enzyme levels varyingbetween 12-45 per cent of the mean of normal males. Methemoglobin reduction test results were considerably less abnormal in hemizygotes with themild type of Mediterranean deficiency than in heterozygotes with the severedeficiency. Fewer heterozygotes with the mild deficiency were detected.

Submitted on February 4, 1966 Accepted on May 31, 1966     

The Carbohydrate Intermediates of Erythrocytes Deficient in Glucose-6-Phosphate Dehydrogenase

The carbohydrate intermediates of erythrocytes were studied before andafter incubation for 6 hours at 37 C. A comparison was made between acase of NSHA with G-6-PD deficiency, his mother, a Caucasian and a Negrowith G-6-PD deficiency (so-called primaquine sensitivity), and a normal case.

An abnormality of ATP metabolism manifested by a distinct fall in concentration of ATP after incubation was found in the case of NSHA but not inthe other cases. This is another demonstration of the heterogeneity of thecases of deficiency of G-6-PD.

Submitted on September 29, 1964 Accepted on November 11, 1964     

Glucose-6-Phosphate Dehydrogenase Deficiency and Blood Transfusion

Abstract. Seven White American male blood donors with Italian surnames were found to have red cell glucose-6-phosphate dehydrogenase (G-6-PD) deficiency among 1,285 with Greek or Italian surnames screened. Five different genetic variants were found: G-6-PDs Mediterranean (2), 'Athens-like' (2), San Juan, Columbus and 'Canton-like'. Clinical evaluation of 23 patients who received 24 units of G-6-PD-deficient blood (G-6-PD A-) failed to reveal any deleterious effects. Screening of Black donors for G-6-PD deficiency is believed unnecessary; further data are needed before a recommendation can be made concerning screening for non-Black donors.     

Comparison for functional aberration of G-6-PD deficiency variants with exon 10 mutations

Glucose-6-phosphate dehydrogenase (G-6-PD) deficiency is a common inherited enzyme deficiency in many parts of the world and there are many different variants described. Every G-6-PD deficiency variant has a unique underlying genetic defect, therefore it manifests specific properties. The single amino acid substitution in the globin chain is the commonest form of G-6-PD deficiency variant. Usually, the G-6-PD deficiency variant with the pathogenesis of a single amino acid substitution presents with only one aberration in secondary structure. Although many G-6-PD deficiency variants present similar structural abnormal points their functions sometimes are discordant. Here, the author performed a functional analysis on some alpha haemoglobinopathies using a novel bioinformatic tool, Polyphen. The mutations of five G-6-PD deficiency variants with exon 10 mutations, Guadalajara (386 Arg-->Cys), Beverly Hills (387 Arg-->His), Serres (361 Ala-->Val), Iowa (385 Lys-->Glu), and Clinic (405 Met-->Ile) were selected for further study in this investigation. According to the in silico mutation study, the functional change in the G-6-PD deficiency variants with exon 10 mutations studied is variable. Here, it indicates that the functional aberration in the G-6-PD deficiency variant is based on complex pathogenesis. The identification of the structural aberration only in a G-6-PD deficiency variant is not sufficient and should be supplemented with a further functional analysis for a better insight in this topic.     

Comparison for functional aberration of G-6-PD deficiency variants with exon 10 mutations

Abstract

Glucose-6-phosphate dehydrogenase (G-6-PD) deficiency is a common inherited enzyme deficiency in many parts of the world and there are many different variants described. Every G-6-PD deficiency variant has a unique underlying genetic defect, therefore it manifests specific properties. The single amino acid substitution in the globin chain is the commonest form of G-6-PD deficiency variant. Usually, the G-6-PD deficiency variant with the pathogenesis of a single amino acid substitution presents with only one aberration in secondary structure. Although many G-6-PD deficiency variants present similar structural abnormal points their functions sometimes are discordant. Here, the author performed a functional analysis on some alpha haemoglobinopathies using a novel bioinformatic tool, Polyphen. The mutations of five G-6-PD deficiency variants with exon 10 mutations, Guadalajara (386 Arg?Cys), Beverly Hills (387 Arg?His), Serres (361 Ala?Val), Iowa (385 Lys?Glu), and Clinic (405 Met?Ile) were selected for further study in this investigation. According to the in silico mutation study, the functional change in the G-6-PD deficiency variants with exon 10 mutations studied is variable. Here, it indicates that the functional aberration in the G-6-PD deficiency variant is based on complex pathogenesis. The identification of the structural aberration only in a G-6-PD deficiency variant is not sufficient and should be supplemented with a further functional analysis for a better insight in this topic.