Shlomo Friedman

α-Thalassemia in Negro Infants

Extract: A total of 104 of 693 Negro infants (15.0%) had moderate or small amounts of hemoglobin Barts visible on starch gel electrophoresis. Moderate amounts were found in 21 infants (3.0%) and small amounts in 83 infants (12.0%). In 17 Negro infants judged to have moderate amounts of Hb Barts, the quantitation showed 2.0–9.3% with a mean of 5.4 ± 2.1 (1 SD). A significant decrease in mean cell volume and mean cell hemoglobin was found in the Negro neonate with more than 2% Hb Barts studied at 4 days of age. In 10 Negro infants with more than 2.0% Hb Barts studied at 4 days of age, the α α/(β + γ) ratio was 0.97 ± 0.06 (1 SD) (range 0.88–1.06). In nine infants aged 5—24 months who had more than 2.0% Hb Barts in the newborn period, including six infants studied in the first group, the mean α/β ratio was 0.74 ± 0.06, (range 0.65–0.83). Each of the nine infants with more than 2.0% Hb Barts at birth had marked microcytosis and hypochromia at 5—24 months despite adequate iron therapy. Two newborn infants with moderate levels of Hb Barts at birth (8.2% and 6.8%) and balanced total globin synthesis had no free radioactive α chain by gel filtration studies. Our studies indicate clearly that the presence of more than 2% Hb Barts in the newborn period denotes the presence of α-thalassemia trait.Speculation: In a group of Negro newborn infants, 3% had more than 2% Hb Barts. These infants had the genetic disorder, α-thalassemia trait. An additional 12% of the infants had elevated levels of Hb Barts between 1% and 2%. This group may also have an α-thalassemia disorder, as has been shown in other racial groups. The absence of hydrops fetalis due to α-thalassemia in Negro neonates suggests that the molecular defect of α-thalassemia detected in Negro neonates differs from that seen in Orientals in that it is not associated with a complete absence of α-chain synthesis.

Mechanical stability of hemoglobin subunits: An abnormality in βS-subunits of sickle hemoglobin

Abstract In order to study the mechanism of the ease of precipitation of oxyhemoglobin S by mechanical shaking, the rates of precipitation of α- and β-subunits of oxyhemoglobin A and oxyhemoglobin S were compared. At pH 8.0, the α A -subunits precipitated rapidly, while the β A -subunits were very stable, although a part of β A -bunits converted to the hemichrome form. At pH 6.0, the β A -subunits precipitated rapidly while the α A -subunits were stable. Similar studies with β S -subunits showed that β S -subunits precipitated rapidly both at acidic and alkaline pHs. The abnormal precipitation of tetrameric oxyhemoglobin S during mechanical shaking may be due to this instability of the β S -subunits.

Variations in Globin Chain Synthesis in Hereditary Persistence of Fetal Haemoglobin

Globin synthesis was studied in four Negro families including 10 members with Hb A‐HPFH and four with Hb S‐HPFH. The β/α specific activity ratios in 10 of these HPFH heterozygotes were similar to those of the control group. In two patients with Hb A‐HPFH, the β/α ratio was slightly decreased in one (0.84) and clearly decreased in another (0.78). In two of the patients with Hb S‐HPFH the ratios were clearly decreased (0.71 and 0.75). The extended range of β/α ratios in these 14 patients is similar to that of Negro patients with β‐thalassaemia trait. These studies indicate that a decreased β/α ratio may be found in HPFH, as well as in β‐thalassaemia.

Globin synthesis in the Jamaican Negro with beta-thalassaemia.

Summary. Globin synthesis was studied in three Jamaican Negro families with 18 heterozygotes and five homozygotes for β‐thalassaemia. Synthesis of the β‐chain of Hb A in the peripheral blood of heterozygotes was equal to that of α‐chain in 10 patients and was decreased in the remainder. In one patient with Hb C β‐thalassaemia, the β/α ratio was normal. These findings were similar to those in American Negroes, but differed from those in Caucasians with β‐thalassaemia trait, in each of whom the β/α ratio was decreased. Globin synthesis was balanced in the bone marrows of Negro and Caucasian heterozygotes. Despite the milder clinical disease in Negro homozygotes as compared to Caucasian patients, the β/α synthesis ratios were similar in both groups.

Inclusions in Red Blood Cells Containing Hb S or Hb C

Summary. To demonstrate and characterize red cell inclusions in 101 persons with Hb S or Hb C disorders three methods were used: (1) examination of unstained blood smears by dark field microscopy (DFM), (2) examination of blood smears after acid elution and staining (AE), and (3) measurement of membrane‐associated denatured haemoglobin (MADH) in ghosts. The control group had inclusions in less than 5% of red cells by DFM and AE and the mean percentage of MADH per total cellular Hb was 0.030±0.016%. The highest percentages of red cells with inclusions and of MADH were present in clinically severe haemoglobin disorders, e.g. homozygous sickle cell disease (Hb SS) with less than 10% Hb F and Hb SOArab, with successively lower percentages in moderate to severe disorders, e.g. Hb SS‐α thalassaemia, Hb S‐β0 thalassaemia, Hb SC disease, and Hb SS with more than 10% Hb F, indicating agreement in results by three methods. In asymptomatic or mild disorders, e.g. Hb S‐β+ thalassaemia, Hb CC, Hb AC and Hb AS, the results by AE and measurements of MADH were the same or similar to those in controls, while those by DFM were different. Of 56 patients with Hb SS or Hb SC, the group with functional asplenia had higher percentages of MADH and of red cells with inclusions than those with functioning spleens. Our study suggests that inclusions in sickling disorders may be due to denatured Hb S, with AE being the more accurate method for visualizing these inclusions, as results by this method correlate better with the amount of MADH than those by DFM.

THE OCCURRENCE OF 1–2% HB BART'S IN A HIGH PROPORTION OF BLACK NEONATES AS AN EXPRESSION OF α-THALASSEMIA

Levels of Hb Barts in Thai neonates of 2-10% and 1-2% indicate the presence of the α-thal2 or α-thal2 genes, respectively. We have previously shown that the 3.0% of black neonates with greater than 2.0% Hb Barts have α-thalassemia trait (Pediat.Res. 8:955, 1974). In a screening study in 1970-72, we found that 12.0% of the 693 neonates screened had 1.1-1.7% Hb Barts, using starch gel electrophoresis and CM Sephadex chromatography. Follow-up studies were done on 8 infants who had 1-2% Hb Barts at birth and on 7 without Hb Barts. The children were between 25 and 60 months of age at the time of study. There were no differences in Hb levels, RBC and reticulocyte counts between the groups. Significant differences were found in mean cell volume, 79.4±3.4 and 72.6±2.9 (p<0.01) and mean cell hemoglobin, 26.74±1.7 and 24.0±1.3 (p<0.01), with the lower values in the children who had Hb Barts at birth. The levels of Hb A2, Hb F, serum Fe, Fe-binding capacity and Pb were normal in the two groups. The results indicate a mild form of α-thalassemia in infants with 1-2% Hb Barts at birth, and some type of α-thalassemia in 15.0% of all black newborns in Philadelphia. This high incidence should be taken into account in determining normal values for red cell indices in black children, and in evaluating the diversity of expression of other hemoglobin disorders in blacks.

IMPROVEMENT OF CEREBRAL VASCULAR DISEASE IN SICKLE CELL ANEMIA FOLLOWING TRANSFUSIONS

Cerebrovascular accidents (CVA) are a major clinical problem in sickle cell disease. We have studied four affected children soon after CVA and one year later to evaluate the effect of chronic transfusion therapy on arterial abnormalities. Percutaneous femoral-cerebral angiography was performed after careful preparation of patients by partial exchange or repeated transfusions to achieve a normal hemoglobin concentration and reduction of Hb S to about 20%. There were no complications of angiography. When possible, the right and left carotid and basilar systems were visualized and cerebral blood flow was measured with 133Xe. Major cerebral artery disease, often more extensive than was clinically evident, was found in all cases. Three of the four children were transfused for one year. Two showed complete resolution of occlusive disease. In a third child in whom transfusions were not started until after a second CVA, some affected vessels had improved after a year, but others showed no change or chronic narrowing. Cerebral blood flow (133xe), initially decreased in this child, was normal on the second study. A fourth child not transfused between studies showed severe occlusive disease with progression. These data suggest cerebrovascular damage in sickle cell disease may be reversible with long-term transfusion therapy.

748 COMPARISON OF LASER NEPHELOMETRY (Ne) & RADIAL IMMUNODIFFUSION (RID) METHODS FOR MEASURING IMMUNOGLOBULINS (ID) IN IMMUNE DEFICIENT (ID) PATIENTS (PTS)

Previous studies have shown that in adult controls with normal Ig levels, a good correlation exists between RID & Ne. IgG, IgA & IgM levels in sera from 42, 17 & 36 respective normal children were: 100-2172 & 117-2400(r=0.81), 44-856 & 10-880(r=0.99), 41-258 & 49-350(r=0.92) by RID & Ne respectively. To measure low level Ig in ID pts by Ne linear reference curves were established having limits of detection of IgG-0.6 mg%, IgA-0.1 & IgM-0.07 mg%. IgG & IgA levels in 37 sera from 11 ID pts, 9 of whom were treated, were 42-1830 & 33-2295(r=0.88), 0-44 & 0-44(r=0.89) by RID & Ne respectively. In 13 sera from 4 pts, IgA levels were 0 & 0.25-15.6 & polymeric IgA was detected by immunoelectrophoresis(IEP), in 6 sera 0-trace & 0, in 18 sera 1.5-44 & 0.65-44 (r=0.93) by RID & Ne respectively. IgM levels in 28 sera from 5 ID pts were 0.51 & 0-36.5(r=0.87) by RID & Ne respectively. In 4 sera from 2 pts, IgM levels were 0 & 4.82-7.0 & polymeric IgM was detected by IEP, in 9 sera from 1 pt, IgM levels were 0 & 2.50-10:71 & monomeric IgM was detected by IEP, in one serum 0 & 0, in 14 sera 2.85-51.0 & 2.53-36.5(r=0.94) by RID & Ne respectively. Low levels of IgA & IgM correlate poorly in pts with selective IgA & IgM deficiencies. Laser Ne appears to be more sensitive than RID in detecting low levels of IgA & monomeric or polymeric IgM in ID pts.

845 PRELIMINARY EVALUATION OF A FOUR CHANNEL ELECTROLYTE ANALYZER (PVA-4) FOR PEDIATRIC INSTITUTIONS

A PVA-4 instrument was modified to accommodate serum or plasma vols as low as 80μl & compared to a Na&K Flame Photometer, a Cl Coulometric Titrator & a total CO2(tCO2) Analyzer. Controls, specimens(spm) & standards were diluted externally 1:5. The re-producibility was tested by analyses of macrodiluted samples of an aq. standard for Na, K & Cl & of individually microdiluted samples. A mean difference (±ISD)(mEq/L) between [Na]+[K;[amp;[Cl] was 0.14(1.63) for 15 readings with a p<.83 & 1.53(1.27) for 22 readings with a p<.0001 respectively. The means (±lSD) of 33 readings of our quality control(QC) material for Na, K & Cl by the PVA-4 were 128.3(1.29), 3.80(.068) & 99.4(1.14) compared to the QC Program means by equivalent methodologies of 129.01(1.57), 3.69(.09) & 99.84(1.34) for Level I respectively. Level II means for 39 readings were 145.8(1.07), 6.41(.052) & 114.7(.86) compared to 146.65(1.57), 6.39(.13) & 114.4(1.29) respectively. Linear regression analysis of 23 patients(pts) spm for Na, K&C1 resulted in correlation coefficients(r) of 0.883, 0.972 & 0.902 respectively. The PVA-4 HCO3 data for 22 pt spm showed good correlation to the calculated HCO3(r=0.981) & to the tCO2(r=.932). However, alkalinity/acidity in pt sera resulted in an over/under estimation of HCO3 e.g. [HCO3] of 39 & 5.6 compared to [CO2] of 33 & 9 respectively. The major drawbacks of the PVA-4 are the misleading HCO3 values in pt sera with pH imbalance & the additional external dilution of spm.