alpha Chain and gamma chain abnormal hemoglobins in newborn babies: structural and genetic aspects

Structural studies and quantitative analyses were conducted on the hemoglobin of 55 newborn babies. Seven alpha chain variants (G-Philadelphia, Montgomery, Inkster, I-Philadelphia, Matsue-Oki, Winnipeg, and O-Indonesia) were present in 26 heterozygous newborns (17 black, eight Caucasian, and one Indonesian). The relative amount of the alpha X containing abnormal Hb F of the Hb G-Philadelphia and Hb Winnipeg babies was less than observed in heterozygous adults, which may indicate a decreased rate of assembly of the alpha X-gamma dimer over that of the alpha X-beta dimer. Of the 29 newborns with gamma chain variants, 16 were Caucasian babies; of these 15 had a Hb A gamma F-Hull heterozygosity and one a Hb G gamma F-Marietta heterozygosity. Six black babies were heterozygous for Hb A gamma F-Texas-I and six for Hb G gamma F-Port Royal. One Japanese baby had a heterozygosity for A gamma F-Iwata and a second was heterozygous for A gamma TF-Yamaguchi. Quantitative analyses of the isolated normal Hb Fo as well as an evaluation of the relative amounts of the Hb Fx in the red cell lysates gave data useful for a speculation of the genetic condition in each of these babies. It was concluded that the babies with the Hbs F-Texas-I, F-Iwata, F-Hull, and F-Marietta were simple heterozygotes with either the G gamma x A gamma/G gamma x A gamma X or the G gamma x A gamma/G gamma X x A gamma genic arrangement. The babies with Hb F-Port Royal had a G gamma x G gamma X/G gamma x A gamma arrangement, which may result from a (to be determined) gene conversion. The newborn baby with Hb F-Yamaguchi has the G gamma x A gamma x/A gamma T-X.-. genic arrangement, suggesting the presence of three distinctly different gamma chain genes of which one, the A gamma T-X gene, produces an A gamma chain (with threonyl at position gamma 75 and an Asn at position gamma 80) at a level usually seen for G gamma rather than A gamma chains. These studies were greatly facilitated by the use of high pressure liquid chromatographic methods.     

Physical inactivity in patients with rheumatoid arthritis: Data from twenty‐one countries in a cross‐sectional,international study

Objective

Regular physical activity is associated with decreased morbidity and mortality. Traditionally, patients with rheumatoid arthritis (RA) have been advised to limit physical exercise. We studied the prevalence of physical activity and associations with demographic and disease‐related variables in patients with RA from 21 countries.

Methods

The Questionnaires in Standard Monitoring of Patients with Rheumatoid Arthritis (QUEST‐RA) is a cross‐sectional study that includes a self‐report questionnaire and clinical assessment of nonselected consecutive outpatients with RA who are receiving usual clinical care. Frequency of physical exercise (≥30 minutes with at least some shortness of breath, sweating) is queried with 4 response options: ≥3 times weekly, 1–2 times weekly, 1–2 times monthly, and no exercise.

Results

Between January 2005 and April 2007, a total of 5,235 patients from 58 sites in 21 countries were enrolled in QUEST‐RA: 79% were women, >90% were white, mean age was 57 years, and mean disease duration was 11.6 years. Only 13.8% of all patients reported physical exercise ≥3 times weekly. The majority of the patients were physically inactive with no regular weekly exercise: >80% in 7 countries, 60–80% in 12 countries, and 45% and 29% in 2 countries, respectively. Physical inactivity was associated with female sex, older age, lower education, obesity, comorbidity, low functional capacity, and higher levels of disease activity, pain, and fatigue.

Conclusion

In many countries, a low proportion of patients with RA exercise. These data may alert rheumatologists to motivate their patients to increase physical activity levels.     

The gamma chain heterogeneity of fetal hemoglobin in black beta- thalassemia and HPFH heterozygotes

High pressure liquid chromatography (HPLC) was applied to the HbF isolated from blood of numerous black patients with beta-thalassemia trait or homozygosity, G gamma-delta beta-thalassemia trait, G gamma A- gamma HPFH heterozygosity, or the G gamma-[delta+ beta+]-HPFH condition. The method allowed an accurate evaluation of the relative quantities of three types of gamma-chain (G gamma, A gamma I, A gamma T) in the fetal hemoglobins. The results have shown the following. (A) The incidence of the A gamma T-chain in beta-thal heterozygotes and G gamma A gamma-HPFH heterozygotes is about the same as has been observed in black newborn; about one of five blacks are heterozygous for this A gamma-chain variant. The A gamma T-chain was not detected in the nine G gamma-delta beta-thal heterozygotes nor in the eight G gamma-[delta+ beta+]-HPFH heterozygotes. (B) In most cases, the A gamma T-chain was produced by the A gamma gene in trans to the beta-thal or HPFH determinant. The contribution by the gamma-chain genes in trans to the beta-thal or HPFH determinant is about 15% of the total gamma-chain production in both conditions. (C) Three black beta-thal heterozygotes (and five additional relatives) had the A gamma T gene in cis to the beta-thal determinant. Four of these patients had a low levels of G gamma-chain (the "adult" level), and the contribution by the A gamma gene in cis to the beta-thal determinant was about three times that of the A gamma gene in trans. The four additional patients, all members of one family, had a high level of G gamma-chain (the "newborn" level), and the contribution of the A gamma gene in cis was half of that seen in the previously mentioned four patients while that of the A gamma gene in trans was essentially the same. These limited data suggest that the genetic anomaly causing high high G gamma levels in adult beta-thal heterozygotes is linked to the beta-thal determinant and that one of its primary effects is a decreased synthetic expression of the A gamma gene in cis to the beta-thal determinant.     

A Homozygote for the HbGγ Type of Foetal Haemoglobin in India: A Study of Two Indian and Four Negro Families

S ummary . The haemoglobin of a 9-yr-old boy from the area of Bombay has only haemoglobin F. The γ-chains of this haemoglobin F are solely of ^Gγ type, and the child is considered to be a homozygote for hereditary persistence of foetal haemoglobin of the ^Gγ type. The same kind of haemoglobin F is also present in a second Indian family and in four American Negro families. One Negro family has an appreciably lower percentage of haemoglobin F in heterozygotes than the other five families. In some of the families certain individuals also have haemoglobin E or haemoglobin S in trans to the other condition and haemoglobin A is absent. Although it has been concluded that the hereditary persistence of foetal haemoglobin is present the condition is not expressed uniformly in all these families. Some of them have certain characteristics of F-thalassaemia. It is possible that two conditions are, in reality, represented: perhaps some families have the hereditary persistence of foetal haemoglobin and others have F-thalassaemia.     

Sequence variations in the 5' hypersensitive site-2 of the locus control region of beta S chromosomes are associated with different levels of fetal globin in hemoglobin S homozygotes.

We have compared the sequence of the 5' hypersensitive site-2 (5'-HS-2) of the locus control region (LCR) from a sickle cell anemia (SS) patient homozygous for haplotype 19 and with low levels of fetal hemoglobin (HbF), with the same sequence from an SS patient homozygous for haplotype 3 and with high levels of HbF. Several nucleotide variations were present in the 5'HS-2 of the haplotype 19 individual. One is the A----G at position -10905 that creates an Sp1 binding site GCCCC (A----G)CCCC. A second is the T----G at position -10924 in a sequence that binds both erythroid and ubiquitous factors and exhibits high homology to the long terminal repeat of the Moloney leukemia viruses and Friend murine leukemia virus. Other differences were in the two AT-rich stretches of DNA, and an A----T substitution at position -10390. Dot-blot analyses of amplified DNA from several SS patients showed that these variations are specific for beta S chromosomes with haplotype 19. We also examined the 5'HS-2 sequence from an SS patient who is homozygous for haplotype 19, but has abnormally high levels of HbF (greater than 20%). We observed a cross-over that has placed sequences similar to the 5'HS-2 of haplotype 3 in juxtaposition to the 5' flanking regions of haplotype 19. Thus, a beta S chromosome with haplotype 19 but having a 5'HS-2 (LCR) characteristic for haplotype 3 is associated with high gamma-chain expression. We postulate that factors produced under conditions of hematopoietic stress, together with genetic determinants on the haplotype 3-like LCR sequences, allow for high level expression of gamma-globin genes.     

A single dose of granulocyte-macrophage colony-stimulating factor induces systemic interleukin-8 release and neutrophil activation in healthy volunteers

Granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) are frequently used in the clinical management of neutropenia. These cytokines not only enhance the proliferation of myeloid precursor cells but also influence the function of mature leukocytes. In a previous study, we found that the in vivo effects of G-CSF on neutrophils differed from those in vitro. In the present study, we investigated the effects of a single dose of recombinant GM-CSF (7.5 microg/kg, subcutaneously) on neutrophils, eosinophils, and monocytes in healthy volunteers. We analyzed leukocyte kinetics, phenotypical changes, neutrophil degranulation, and systemic cytokine production. After GM-CSF injection, phenotypical changes included upregulation of CD11b on all three cell types and a decreased expression of L-selectin and Fc(gamma)RIII on neutrophils. Neutrophil degranulation was evident from the increased plasma concentrations of lactoferrin and elastase. GM-CSF induced the release of interleukin-8 (IL-8), but not of IL-6 or tumor necrosis factor alpha. In comparison to the results from our previous study with G-CSF in healthy volunteers, GM-CSF induced a stronger activation of mature neutrophils but had a much less pronounced effect on the production and maturation of neutrophil precursors. These data may help to guide the choice between the two cytokines in different clinical situations.     

Certain mutations observed in the 5' sequences of the G gamma- and A gamma-globin genes of beta S chromosomes are specific for chromosomes with major haplotypes.

In this paper we describe the distribution of some specific sequence differences in the 5' flanking regions of the A gamma- and G gamma-globin genes from 100 Black adult and 57 newborn SS patients from the southeastern United States, from 76 individuals with AS, S-beta-thal, SC, AC, or A-beta-thal, and from 31 normal individuals. Haplotypes for all adult individuals have been previously determined using various restriction endonucleases. The DNA samples were amplified, dot blotted, and hybridized with 32P-labeled specific oligonucleotide probes. All 134 chromosomes with haplotype 19 were positive for the G----T substitution at position -657 (A gamma), while 132 were also positive for the C----G mutation at -369 (G gamma). The three specific changes for the chromosome with haplotype 20 were found on all 54 chromosomes with this haplotype. The C----T mutation at -158 5' to G gamma was present on all 41 chromosomes with haplotype 3, and on two chromosomes with a related atypical haplotype. Normal and beta-thal chromosomes with each of these substitutions had the same 5' subhaplotype as beta S haplotypes 19 or 20, respectively. The close relationship between the occurrence of specific mutations and the haplotype of beta S chromosomes makes the determination of these haplotypes with specific oligonucleotide probes attractive with respect to time and expense.     

Beta zero-thalassemia in association with a gamma-globin gene quadruplication

We have studied the hematology, hemoglobin composition, and globin gene arrangements in one young Turkish boy with a beta zero-thalassemia homozygosity and in 11 of his relatives. Evidence is presented that the chromosome with the beta zero-thalassemia determinant carries a gamma- globin gene quadruplication, perhaps in a -G gamma-G gamma-G gamma-A gamma-gene arrangement. The eight gamma-globin genes in this patient produced G gamma and A gamma chains in a 95 to 5 ratio, and nearly 99% of the patient's hemoglobin was of the fetal type. The clinical condition resembled that of a thalassemia intermedia. HbF levels in eight beta-thalassemia heterozygotes varied between 0.5 and 4.2% and the percentages of G gamma in this HbF averaged at 87% or 95%; this level is to some extent related to the haplotype of the normal chromosome. All subjects carried four alpha-globin genes; a new BglII polymorphism was observed within the psi alpha-globin gene.     

Identical nucleotide sequences of the 3'A gamma globin gene enhancer elements from four different chromosomes

We have determined the nucleotide sequence of the 2,360-bp long EcoRI fragment from four chromosomes; this fragment is located 3' to the A gamma globin gene and is considered to contain the enhancer element identified by Bodine and Ley. The chromosomes were from an Arabian sickle cell anemia patient with high Hb F and a homozygosity for haplotype No 31 and from a black sickle cell anemia patient with low Hb F and a homozygosity for haplotype No 19. A third chromosome carried the determinant for a nondeletional hereditary persistence of fetal hemoglobin seen in a Chinese subject, and the fourth was a normal chromosome from a Yugoslavian subject. Twenty-one differences were observed when a comparison was made with the published sequence; no differences were seen between the sequences of the four different samples except for an additional mutation in the Chinese. These data make it unlikely that specific mutations within this sequence are associated with increases in G gamma and A gamma production.