Intracellular assembly of kell and XK blood group proteins

CURRENT LITERATURE achieved with high-dose IVIG therapy. The authors conclude that high concentrations of some but not all IVIG preparations result in reduced placental transfer of maternal alloantibody by blockade of ilnmunoglobulin Fc receptor sites on the placenta. They hypothesize that different manufacturing procedures may 'affect the functional stability of the Fc portion of the IVIG molecule. Part of the variability in the efficacy of IVIG therapy may be attributable to these functional differences. It would be interesting to determine whether differences in Fc receptor function can be shown between the different IVIG preparations in other experimental model systems. (M. G.)

Pharmacological strategies to decrease excessive blood loss in cardiac surgery: A meta-analysis of clinically relevant endpoints. M. Levi, M.E. Cromheecke, E. de Jonge, et al. Lancet 354:1940-1947, 1999. Although pharmacological strategies to decrease perioperarive bleeding have been investigated in a large number of clinical trials, the primary outcome measurements have been decreased blood loss or percentage of patients requiring transfusions. Because of their small size, the studies have lacked the power to detect changes in other more significant clinical outcomes, such as rethoracotomy, perioperative myocardial infarction, and mortality rate. The authors report the results of a meta-analysis of studies using aprotinin, lysine analogs (aminocaproic acid and tranexamic acid), or desmopressin in regular or complicated cardiac surgery patients, defined as patients taking aspirin undergoing repeat surgery. Only randomized, controlled trials in adults were included in the analysis. Seventy-two trials, involving 30 to 870 patients (8,409 overall) were analyzed. Most of the trials compared treatment with one agent with placebo, and the analysis is expressed as an odds ratio, with 1 signifying equivalence between the treatment and control group, ratios below 1 signifying benefit for the treatment group, and ratios above 1 signifying benefit for the control group. The most striking results were noted for atropinin, with an almost twofold decrease in mortality rate from 2.8% to 1.5%; odds ratio, 0.55 (95% CI, 0.34 to 0.90), as well as a decrease in the rethoracotomy rate from 5.0% to 1.8%; odds ratio, 0.37 (95% CI, 0.25 to 0.55). There was no difference in the myocardial infarction rate in trials comparing aptrotinin with placebo. In trials comparing conventional-dose aprotinin with lower-dose regimens, the rate of myocardial infarction was higher in patients receiving conventional dose therapy. Analysis of mortality rates, rethoracotomy rates, and myocardial infarction rates in patients receiving lysine analogs favored the treatment group; however, the 95% confidence intervals were large and approached or exceeded 1. Treatment with desmopressin did not change the mortality rate or rethoracotomy rate but resulted in a 2.4-fold increased risk on perioperative myocardial infarction, with an odds ratio of 2.39 (95% CI, 1.02 to 5.60). The odds ratio for the decrease in the proportion of patients receiving any blood transfusion was 0.33 (95% CI, 0.26 to 0.42), 0.43 (95% CI, 0.26 to 0.72), and 0.82 (95% CI, 0.55 to 1.23) for aprotinin, lysine analogs, and desmopressin, respectively. Results were similar for patients undergoing complicated surgery. The authors hypothesize that the decrease in blood loss with aprotinin and lysine analogs results in a lower frequency of rethoracotomy, which in turn decreases the perioperative mortality rate. In addition, the nonspecific antiprotease effect of aprotinin may inhibit proinflammatory mediators. The meta-analysis does not resolve the

283 controversy over the potentially prothrombotic effect of aprotinin, because there was an increase in the myocardial infarction rate in conventional-dose compared with low-dose aptrotinin regimens, but no difference between all aprotinin regimens compared with placebo. Desmopressin had a smaller effect on reducing blood loss and did not result in improvement in clinically important end points. It is possible that the hemodynamic changes induced by desmopressin may contribute to an increased risk of myocardial infarction. The authors conclude that ,although the meta-analysis supports the use of aprotinin or tysine analogs in cardiac surgery, a large prospective controlled trial is necessary to provide definitive evidence of improvements in mortality rate. (M.G.)

Isoforms of the Lutheran/basal cell adhesion molecule glycoprotein are differentially delivered in polarized epithelial cells: Mapping of the basolateral sorting signal to a cytoplasmic di-leucine motif. W. E~ Nemer, Y. Co~in, C. Bauvy, eta/. J Biol Chem 274:31903-31908, 1999. Lu and Lu(vl3) (synonym: BCAM) are 2 glycoprotein isoforms resulting from alternative splicing of the LU gene. Both proteins belong to the immunoglobulin supeffamily and carry Lutheran blood group antigens and the basal cell adhesion molecule epithelial cancer antigen. Lu (85 Kd) and Lu(vl3) (78 Kd) glycoproteins differ only by their cytoplasmic tail and are adhesion molecules that bind laminin. In nonerythroid tissues, the Lu/basal cell adhesion molecule antigens are predominantly expressed in the endothelium of blood vessel walls and in the basement membrane region of normal epithelial cells, whereas they exhibit a nonpolarized expression in some epithelial cancers. In this paper, the investigators studied the polarization of Lu and Lu(vl3) glycoproteins in epithelial cells by confocal microscopy and domain-selective biotinylation assays. Differentiated human colon carcinoma Caco-2 cells had a polarized expression of endogenous Lu antigens with predominant expression of the Lu isoform at the basolateral domain of the plasma membrane and a very low expression of the Lu(vl3) isoform at both the apical and basolateral domains. Similar expression was obtained in analysis of transfected Madin-Darby canine kidney cells. Studies indicated that a dileucine motif at positions 608 and 609, which was specific to the Lu isoform, was a basolateral sorting signal that prevents Lu glycoprotein from being transported to the apical surface. (M.E.R.)

Intracellular assembly of Kell and XK blood group proteins. D. Russo, S. Lee, and C. Redman. Biochim Biophys Acta 1461:10-18, 1999. Kcll is a 93-Kd type II membrane glycoprotein, and Xk is a 37-Kd membrane protein that is predicted to pass through the membrane 10 times. The 2 proteins exist as a disulfide-bonded complex in human red blood cells. Cys72 of Kell (close to the membrane-spanning domain) is linked to Cys347 of Xk (in the fifth extracelhilar domain). In this paper, the mechanism of Kell/Xk assembly was studied in transfected COS cells by co-expressing Kell and Xk proteins. Time course studies combined with endonuclease-H treatment and cell fractionation showed that the 2 proteins are assembled in the endoplasmic reticulum. The complex then travels through the Golgi to the plasma membrane. When Kell or XK were transfected in the

284 absence of the other, they were transported to the COS cell surface, indicating that linkage of Kell and Xk is not obligatory for cell surface expression. Indeed, in the recombinant system used, co-expression of Kell and Xk did not enhance the amount of Kell or Xk that is placed on the cell surface. (M.E.R.)

Emergence of FY*A nujl in a Plasmodium vivax-endemic region of Papua New Guinea. P.A. Zimmerman, L Woolley, G.L. Masinde, eta/. Proc Natl Acad Sci U S A 96:13973-13977, 1999 It has been well established that a mutation in the GATA-1 promoter region silences expression of Dully in red blood cells (RBCs). In Africans, this mutation has always been associated with the gene encoding the Fy b isofOrlTl of the Duffy protein. This paper is the first to show that the GATA mutation also occurs in the gene (FY*A n"u) encoding the Fy a isoform of the Duff'3' protein. Analysis of 1,062 people from a Plasmodium vivax-endemic region of Papau New Guinea showed 23 (2.2%) that were heterozygous FY*A/FY*A ~"ll. No homozygotes were found. By flow cytometry, using anti-Fy6, RBCs from heterozygous people expressed approximately half the amount of Duffy protein when compared with RBCs from people with 2 FY*A genes with the wild-type GATA box. Interestingly, there was a higher prevalence of P vivax infection in the FY*A/FY*A (83/508 = 0.163) when compared with FY*A/FY*A n"lt (2 of 23 = 0.087) individuals. However, the difference was not statistically significant. This same population from East Sepik has an incidence of c~+-thatassemia of 98%. This form of thalassemia is associated with increased susceptibility to uncomplicated malaria among young children. It has been proposed that e~+-thalassemia may facilitate so-called benign P vivax infection, to act later in life as a "natural vaccine" against P falciparum malaria. The authors of this paper suggest that emergence of FY*A ""ll in the Abelam-speaking population implies that P vivax is involved in selection of this polymorphism and that the mutation could ultimately compromise c~+-thalassemia/P vivax--mediated projection against severe P falciparum malaria. (M.E.R. )

Structure and expression of the mouse homologue of the X K gene. E. Collec, Y. Colin, F. Carbonnet, et al. Immunogenetics 50:16-21, 1999. The human Kx blood group antigen is carried on a nonglycosylated protein with an apparent molecular mass of Mr 37,000. The Xk protein consists of 444 amino acids with 10 putative transmembrane passes. This protein is absent from rare males with the McLeod syndrome. Although Xk protein has structural similarities to membrane transporters, its function has not yet been determined. This paper describes the cloning of the orthologous murine X K gene and thereby provides information necessary to develop a knockout mouse model for the X K gene to study the function of the Xk protein. The mouse X K gene is organized into 2 exons (human X K gene has 3 exons) and is predicted to encode 446 amino acid residues. The cysteine residue at position 347 (that is involved in formation of a disulfide bridge with Cys72 of the human Kell glycoprotein) is conserved. The mouse and human sequences share 82.6% identity at the nucleotide level and 82% at the amino acid level. Hydrophobicity plots showed almost identical profiles for the human and mouse protein, with 10 putative transmembrane

CURRENT LITERATURE e~-helices. Mouse Xk protein in the red blood cell (RBC) membrane has an apparent Mr of 43,000 and was shown by immunoblotting with a rabbit antibody directed against the human protein. In nonreduced RBC membrane preparations, an Mr 140,000 species was detected, suggesting that the Xk protein may be complexed with another protein in murine RBCs, which presumably is the homolog to human Kell protein. The similarity of murine Xk to human Xk is remarkable. The nucleotide sequence for the murine XK gene has been deposited in GenBank/EMBL under accession number AF064772. (M.E.R.)

A randomized, double-blind, placebo-controlled study with pegylated recombinant human mekaryocyte growth and development factor (PEG-rHuMGDF) as an adjunct to chemotherapy for adults with de novo acute myeloid leukemia. E. Archimbaud, O.G. Ottmann, J.A. Liu Yin, et al, Blood 94:3694-3701, 1999. Years ago, when the discoveries of molecular biology held promise for the development of not only erythropoietin but also neutrophil stimulatory factor and platelet stimulatory factor, there was an opinion that transfusion support of patients with leukemia might become unnecessary and obsolete. However, the use of erythropoietin and granulocyte-macrophage colony stimulating factor (GM-CSF) has had only limited impact on the need for transfusion support for most patients with leukemia, probably because marrow suppressed by lettkemia and chemotherapy cannot respond to these growth factors. This paper extends these concepts to thrombopoietin. This randomized trial studied the impact of pegylated recombinant human mekaryocyte growth and development factor (PEG-rHuMGDF). One hundred eight adult patients with leukemia were randomized to 1 of 3 groups: group A received 2.5 pg/kg/day or 5 ~g/kg/d, for up to 21 doses. Group B received 7 daily doses of 2.5 ~g/kg. Group C received placebo. The groups included patients of comparable age, gender mix, performance status, and tumor cytogenetic status. The results documented that the patients who received the greatest doses of PEG-rHuMGDF generated the highest platelet count after recovery from chemotherapy. For example, patients in group A showed a median peak platelet count of greater than 1,000 • 109/L, compared with 517 • ]09]g for group B, and 390 • 109/L for group C. More than half of the patients in group A showed platelet counts greater than 1,000 • 109/L during therapy. Most importantly, however, the response to PEGrHuMGDF had littIe impact on transfusion support. There was no effect of treatment on the median time to independence from platelet transfusions. The RBC transfusion support also was not different between the 3 groups. This large clinical trial of the use of PEG-rHuMGDF showed no evidence of serious toxicity or stimulation of the patient's underlying leukemia. No doubt the drug stimulated platelet recovery in treated patients. However, the data also suggest strongly that PEG-rHuMGDF did not lessen the need for platelet support duling induction and recovery. As we learned with erythropoietin and GM-CSF, patients with leukemia undergoing induction chemotherapy depend on transfusion support. Future clinical studies of thrombopoietins may focus on improved dosage regimens or the identification of subsets of patients who may benefit from thrombopoietin stimulation. (S.D.)