A racial difference in serum vitamin B12 levels.

Measurements of the serum Vitamin B12 concentrations of 49 black and 49 white healthy adults demonstrate a significantly higher mean serum Vitamin B12 level in blacks when compared to whites (p less than 0.001). The reason for this difference appears to be genetic, although environmental factors may also be involved. It is suggested that clinical laboratories should establish their own separate reference values of serum Vitamin B12 for blacks and whites in order to prevent misinterpretation of test results.

Laboratory studies of erythrocytic pyruvate kinase deficiency. Pathogenesis of the hemolysis.

Genetic polymorphism exists in erythrocytic pyruvate kinase (PK)-deficient hemolytic anemia, as briefly described. Destruction of erythrocytes varied in extent, but its mechanisms in different PK-deficient polymorphic persons investigated were similar. A paradoxical post-splenectomy reticulocytosis regularly occurred. Qualitative enzymatic differences, the biochemistry, and measurements of erythrocytic destruction were made in several PK variants. 51Cr autologous and cross-transfusions of PK-deficient erythrocytes into volunteers showed multimodel regression lines of several erythrocytic cohorts. 59Felabeled PK-deficient reticulocytes donated by PK-deficient splenectomized subjects were transfused 20 hours before splenectomy into two PK-deficient infants with hemolysis, and into two adult volunteers with autoimmune thrombocytopenia. The highest reticulocyte concentration in any organ initially was within the spleen. Radioiron-labeled erythrocytes and cytologic data showed large splenic reticulocyte pools. Splenic macrophages ingesting reticulocytes and erythrocytes were seen by both light and electron microscopy of the splenic pulp. After cyanide additives inhibiting reticulocyte oxidative phosphorylation, a bizarre erythrocytic cytologic configuration was found by scanning electron microscopy. These studies of PK-mutant subjects with PK-deficient erythrocytic hemolysis showed age dependent destruction of erythrocytes. Bimodal Cr survival data suggested reticuloendothelial removal of short-lived erythrocytes and reticulocytes. The transfusions of radioironlabeled PK reticulocytes and the data obtained by scanning electron microscopy suggested that the spleen was the initial hostile organ destroying a cohort of susceptible erythrocytes, prinicpally reticulocytes.

Transfusion reaction with pulmonary infiltration associated with HL-A-specific leukocyte antibodies.

A case of a non-hemolytic transfusion reaction with pulmonary infiltration secondary to leukocyte antibodies is described, and previously reported cases are reviewed. This type of reaction can be diagnosed at the bedside when a patient develops fever, hypotension and dyspnea within a few hours following transfusion of whole blood or a plasma product. The roentgenogram of the chest shows pulmonary infiltrates with a normal cardiac silhouette constituting non-cardiac pulmonary edema. To provide laboratory confirmation of this reaction, it is essential to search for leukocyte antibodies by both leukoagglutinin and cytotoxic technics, as well as to determine HL-A phenotypes of both donor and recipient. As the plasma products involved usually come from multiparous women, donor parity should be a routine question in the donor interview in transfusion services. To prevent this reaction, which may prove fatal, blood donated by women who have two or more children should be used for packed cells only.

Effectiveness of prophylactic Rh immunosuppression after transfusion with D-positive blood.

In a published investigation 42 D-negative men were transfused with single units of D-positive R1r (CDe/cde) blood to mimic the D-incompatible transfusion accidents occasionally seen in young D-negative women. Twenty-two men received no therapy and 20 were treated with Rh immunoglobulin in a calculated dose of 20 mug of anti-Rh per milliliter of infused red cells. The reported incidence of serologic D alloimmunization was 81.8 per cent in control and zero per cent in treated recipients. Posttransfusion 51Cr clearance studies with a challenge with the use of 5 ml. of D-positive cells from the same donor at five months in these identical subjects vary from the results based upon the presence of circulating anti-D active antibody. Exponential immune clearance curves occurred in two control and nine treated recipients. As the first evidence of D alloimmunization is an immune Cr clearance curve followed at some later time by circulating active anti-D antibodies, these data could suggest that Rh immunization actually was 90.9 per cent in untreated control subjects and 45 per cent in treated men. The dose of passive Rh immunoglobulin effective in prophylaxis of transfusion Rh immunization of a D-negative person is still not definitely established, but apparently it is greater than 20 mug of anti-Rh per milliliter of D-positive erythrocytes received.

Carriers with excessively low factor VIII procoagulant activity (VIII AHF): a study of two unrelated families with mild hemophilia A.

Two unrelated families are described with mild hemophilia A in whom six obligate carriers had unusually low VIII AHF levels. In each family, successive generations of males were affected with hemophilia A as determined by low VIII AHF in the presence of normal VIII AGN and VIII VWF levels. In the first family, two of five obligate carriers had low VIII AHF levels associated with clinical bleeding and one other had a history of bleeding. While receiving oral contraceptives, one of these two carriers was found to have a normal VIII AHF level. In the second family, four cousins below age 10 who were obligate carriers had significantly low VIII AHF levels, while a paternal aunt and paternal grandmother who were also obligate carriers had VIII AHF levels within the normal range. Hemorrhagic diathesis in multiple obligate carriers in these families is not readily explained by the Lyon hypothesis, and suggests that these families may be exmaples of an unusual allelic form of hemophilia A or that they may be transmitting several independent genes affecting VIII AHF levels. Our experience suggests that VIII AHF levels should be determined on all obligate or possible carriers prior to surgery to identify those individuals at risk for postoperative bleeding. Furthermore, it is suggested that hormonal therapy might be effective in the management of carriers with low levels of VIII AHF and clinical bleeding.

Molecular abnormality of erythrocyte pyruvate kinase deficiency in the Amish.

We describe the cellular and molecular biologic studies of the erythrocyte pyruvate kinase (PK) deficiency of the Amish deme in Pennsylvania. Nucleotide sequencing of the patient's PK gene showed a point mutation, CGC to CAC, corresponding to no. 1436 from the translational initiation site of the R-type PK (R-PK) mRNA, and it caused a single amino acid substitution from Arg to His at the 479th amino acid residue of the R-PK. The substituted Arg residue is located in the C domain of PK subunit, that is essential for both the intersubunit contact and the allosteric regulation. Because this enzyme shows the catalytic activity only as a dimer or tetramer, it is rational that the structural alteration would result in severe PK deficiency. To elucidate the effect of the PK deficiency on red blood cell (RBC) membrane, we performed the cellular studies of the patients' RBCs. Ouabain-insensitive K+ efflux was increased to 142% to 145% of normal controls and not inhibited by furosemide, as previously observed in HbSC disease RBCs.

Studies of the recovery and the cost of low-glycerol cryopreserved human red blood cells.

Red blood cells were equilibrated with 28 per cent (v/v) glycerol and 3 per cent mannitol in 0.65 g/100 ml sodium chloride. The units were frozen by immersion into liquid nitrogen and stored at -160 C. After thawing, they were reconstituted and washed using the IBM 2991 Blood Cell Processor. Freeze-thaw rate curves, the effect of thawing techniques, the effect of varying postthaw washing and processing techniques, estimates of red blood cell losses because of hemolysis, and in vitro recovery were determined. In vivo recovery was determined by 51Cr techniques 24 hours after infusion and Ashby survivals and subsequent life span were measured. Metabolic, scanning electronmicroscopy, cost estimates and quality control studies were done on the reconsituted red blood cells. Recipients were evaluated before and after transfusion for metabolic erythrocyte characteristics and for evidence of hemolysis. The modified method requires less wash solution and less technician time than does the standard low-glycerol method. Two units for the same recipient could be passed through the IBM software with no alteration of cell survival or loss. Revision of the IBM 2991 processing procedure provided excellent recovery of viable previously frozen red blood cells at probably a lower cost.

Hepatitis B virus, hepatitis A virus and persistently elevated aminotransferases in hemophiliacs.

To determine the exposure to hepatitis A and hepatitis B viruses (HAV, HBV) following intravenous replacement therapy in patients with classic hemophilia and to assess the role of these viruses in persistently elevated aminotransferases, sera were studied from 136 patients from 9 months to 67 years of age were transfused with either single-donor cryoprecipitate (CRYO) or Antihemophilic Factor Concentrate (AHF) for periods ranging from a few months to 15 years. Serologic evidence of past or present infection with HBV was detected in 90% of all 136 patients and in 85% of those 34 patients 10 years of age or younger. Sixty-four percent of those with serologic markers of hepatitis B had high titers of antibody to the hepatitis B surface antigen and low titers of antibody to the hepatitis B core antigen. These findings are consistent with the known high frequency of early exposure to HBV in hemophiliacs receiving replacement therapy and with recovery from these hepatitis B infections. Sixteen percent of these patients had persistently elevated aminotransferase levels; HBV could not be implicated as the cause of the enzyme elevations in most of these cases.

Immunosuppressive therapy of Factor VIII inhibitors.

Immunosuppressive therapy was used in seven hemophiliac and three nonhemophiliac patients with factor VIII inhibiors. Permanent disappearance of the inhibitor occurred in three hemophiliac and two nonhemophiliac patients following treatment with cyclophosphamide and factor VIII. Critical factors influencing the response to therapy may include both the titer and duration of the inhibitor and the degree of intervening factor VIII exposure prior to immunosuppressive therapy. Two severe hemophiliacs with low titer inhibitors that disappeared without specific therapy are also reported.