American Journal of Hematology

doi: 10.1002/ajh.2830380301pmid: N/A

Marx, Gerard; Krugliak, Judit; Shaklai, Matityahu

doi: 10.1002/ajh.2830380302pmid: 1951315

After ingestion of 220 mg zinc sulfate, platelet aggregation was evaluated at various time intervals (i.e., T = 0, 1, and 3 hr) and the autologous plasma analyzed by atomic absorption analysis. The zinc levels increased maximally some 0.4 ± 0.2 μg/ml within 3 hr after ingestion, which tor the entire blood pool corresponds to only 5% of the ingested zinc. Aggregation responses of platelet rich plasma (PRP), instigated with suboptimal levels of thrombin (<0.2 U/ml), ADP (<2 μM), epinephrine (<2 μM), collagen (<2 μg/ml), or PAF (<50 ng/ml), show significant improvement to at least one aggregant. Mean ± SEM values for Δ% aggregation increase are as follows: thrombin, 51 ± 10%; epinephrine, 21 ± 6%; ADP, 31 ± 6%; collagen 23 ± 6%; and platelet aggregating factor (PAF), 56 ± 6%. For controls, the platelets from one individual with Glanzmann thrombasthenia as well as four undosed volunteers exhibited no significant changes in platelet responsiveness. Increased platelet responsiveness to agonists after zinc sulfate ingestion was observed in PRP from blood collected in either citrate or heparin. We demonstrate that within a relatively short time period, single bolus of nutritional zinc intake can significantly increase platelet reactivity. These findings show that nutritional zinc availability is relevant to hemostasis and may pertain to the viability of platelet concentrates in blood banks.

Brissette, Renee E.; Swislocki, Norbert I.; Cunningham, Earlene Brown

doi: 10.1002/ajh.2830380303pmid: 1659185

A p‐nitrophenylphosphatase activity has been identified as a component of the human erythrocyte membrane. This activity is distinct from that associated with the cell's Na+ + K ‐dependent ATPase, Ca2+ ‐dependent ATPase, or spectrin phosphatase. The activity described here is stimulated by Mn2+ but not by Ca2+ with or without calmodulin. A potential erythrocyte membrane substrate for this activity is a 95 kDa phosphoprotein that can be shown to undergo Mn2+ ‐stimulated but not Mg2+ ‐stimulated dephosphorylation.

Takahashi, Hoyu; Hanano, Masaharu; Wada, Ken; Tatewaki, Wataru; Niwano, Hiroe; Shibata, Akira; Tsubouchi, Jiro; Nakano, Masahiko; Nakamura, Tadao

doi: 10.1002/ajh.2830380304pmid: 1659186

Endothelial cell injury is thought to be one of the causative factors in thrombotic thrombocytopenic purpura (TTP). A novel index of endothelial injury, plasma thrombomodulin, was measured in 13 patients with acute TTP. The mean plasma concentration of thrombomodulin was elevated in patients with TTP (34.23 ± 19.08 ng/ml) as compared with healthy subjects (16.99 ± 2.63 ng/ml, P <0.001). Eight (61.5%) of 13 patients had high thrombomodulin values. Markedly elevated thrombomodulin levels were observed in TTP patients who had suffered from systemic lupus erythematosus, in whom plasma thrombomodulin was still elevated when they achieved remission. Five of these 13 patients with TTP had normal plasma levels of thrombomodulin. In addition, the plasma thrombomodulin concentrations were correlated well with von Willebrand factor antigen and tissuetype plasminogen activator antigen levels, both of which are released from stimulated or damaged endothelial cells. No difference was found in plasma thrombomodulin levels between patients who achieved remission and who did not. These findings suggest that the magnitude of the endothelial damage in TTP is variable among patients and that plasma thrombomodulin has limited clinical relevance to the severity of TTP.

Nakatsumi, Tomoko; Nakao, Shinji; Ohtaguro, Mamiko; Chujo, Tatsuya; Tsuchiya, Haruo; Niki, Takeo; Mori, Takao; Matsuda, Tamotsu; Shiobara, Shintaro; Nagai, Kozo

doi: 10.1002/ajh.2830380305pmid: 1951316

Galli, Monica; Cortelazzo, Sergio; Barbui, Tiziano

doi: 10.1002/ajh.2830380306pmid: 1719809

The authors have investigated the presence in commercially available intravenous gammaglobulins (IVlg) of anti‐idiotypic antibodies directed to Lupus Anticoagulant (LA). In vitro incubation of 4 LA plasmas with increasing concentrations of IVlg (from 0 to 39 mg/ml) resulted in a dose‐dependent inhibition of LA activity (the highest inhibitions ranged from 14.0 to 53.4%). Similar results were obtained when patients' plasma was substituted with total IgG (the highest inhibitions ranged from 43.0 to 55.0% and were obtained at IgG:IVIg molar ratios ranging from 1:15 to 1:50). Also the incubation of patients' F(ab's)2 with F(ab')2 from IVIg produced a similar dose‐dependent inhibition of LA activity. These data are suggestive of an in vitro idiotypic‐anti‐idiotypic interaction between LA and IVIg. However, when injected in patients with LA, IVIG do not seem to operate by this mechanism of action. In fact, reduction or disappearance of LA was only observed in 2 out of 4 patients; also the quick reappearance of LA activity was not consistent with the time course of anti‐idiotypic response. Finally, this effect was reached by half the IVIg concentrations necessary to produce an appreciable inhibitory effect on LA activity in vitro. Thus, it is concluded that, even if IVIg contain anti‐idiotypic antibodies reacting with LA, the clinical efficacy of IVIg treatment in patients with these autoantibodies should be attributed to other mechanisms.

Liang, Raymond; Chan, Vivian; Chan, T. K.; Wong, Thomas; Chiu, Edmond; Todd, David

doi: 10.1002/ajh.2830380307pmid: 1951317

Polymerase chain reaction (PCR) was used to amplify the DNA fragments of the complementarity determining region 3 of the immunoglobulin (Ig) gene heavy chain from the leukemic cell specimens of patients with acute and chronic lymphoid leukemias of B‐cell lineage. Two different pairs of primers were tested. Fourteen of the 17 (82%) cases of acute lymphoblastic leukemia (ALL), and all 15 cases (100%) of B‐cell chronic lymphocytic leukemia, who had rearrangement of the Ig gene heavy chain by Southern analysis, were positive by PCR with either one or both pairs of primers. This technique was able to detect leukemic cells at the level of 0.1%. Applying it to study the remission marrow specimens following induction chemotherapy was more useful than morphology alone in predicting early relapse of the leukemia.

Wajcman, H.; Vasseur, C.; Poyart, C.; Blouquit, Y.; Galacteros, F.; Santo, D. Esperito; Peres, M. J.; Martins, M. C.

doi: 10.1002/ajh.2830380308pmid: 1951318

Hb Redondo (β92(F8) His→Asn) illustrates how post‐translational structural modifications may modify the phenotypic expression of an unstable hemoglobin. This variant was found in a Portuguese patient suffering from a chronic hemolytic anemia. The electrophoretic pattern demonstrated that it occurred in two forms, both being semi‐hemoglobins: the fastest one migrating like HbS and the other like HbA2; after a few days of storage at 4°C the intensity of the slowest Hb fraction decreased while that of the other increased proportionally. In both cases, the RP‐HPLC analysis of the tryptic digest of the aminoethylated β chains demonstrated the presence of an abnormal βT10 peptide carrying a His→Asx substitution. Microsequence study of these two peptides demonstrated that the slowest abnormal Hb fraction had a β92 His → Asn substitution and the fastest a His → Asp at the same site. All these results suggest that the β92 His → Asn variant loses readily its heme group and that a deamidation occurs rapidly in vitro, yielding a β92 Asp semi‐hemoglobin. The oxygen affinity of the patient's red blood cells was increased, leading to a stimulation of erythropoiesis and to a macrocytic hemolytic disease.

Kondo, Takahito; Isobe, Hiroshi; Kawakami, Yoshikazu; Sakai, Masaharu; Nishi, Shinzou; Taniguchi, Naoyuki

doi: 10.1002/ajh.2830380309pmid: 1719810

Induction of carbonic anhydrase isozyme I (CA‐I) by erythropoietin or hemin was investigated using erythroleukemia (K562) cells. Immunological estimation and purification of carbonic anhydrases showed that untreated K562 cells contained only carbonic anhydrase isozyme II(CA‐II), while incubation of the cells with 2 units of erythropoietin (EP) per ml of the incubation medium or with 50 μM hemin resulted in the induction of CA‐I. The purified CA‐I induced in K562 cells was enzymatically and immunologically identical to that from mature erythrocytes. Flow cytometric analysis showed that incubation of K562 cells with EP as well as hemin induced CA‐I at the 3rd h, while α‐globin was detected at the 8th h. Northern blot analysis of CA‐I mRNA using a cloned genomic DNA as a probe showed that mRNA of CA‐I was induced by EP. These results suggest that induction of CA‐I is regulated at the transcriptional level during developmental changes of erythroid cells, and that CA‐I may play a physiologically important role during erythroid differentiation.

Todd, A. V.; Ireland, C. M.; Radloff, T. J.; Kronenberg, H.; Iland, H. J.

doi: 10.1002/ajh.2830380310pmid: 1951319

N‐ras gene activation occurs via single base substitutions in codons 12, 13, and 61. We have developed a rapid screening method, termed allele specific restriction analysis (ASRA), for detection of N‐ras mutations at these three critical codons in acute myeloid leukemia (AML). Patient DNA samples are amplified by the polymerase chain reaction (PCR) by using primers that induce restriction sites in normal but not mutant N‐ras alleles. We have used ASRA to identify 5 point mutations in four out of 19 patients at initial presentation of de novo AML. Three patients had one mutation at codon 12, 13, or 61 respectively, while a fourth patient had concurrent mutations at codons 12 and 13. N‐ras mutations were more common in patients over 65 years of age (P < 0.04), but did not correlate with FAB classification, attainment of complete remission, disease free survival, or overall survival. ASRA can also be used as the first step in a more sensitive approach to the detection of ras mutations. When ASRA was combined with allele specific oligonucleotide (ASO) hybridization the sensitivity and specificity of these assays were increased. This allowed identification of additional low level mutations in two patients. The data presented here constitute the first complete analysis of N‐ras mutations in leukemia by ASRA and include the first identification of three concurrent N‐ras mutations in a single leukemic patient. By facilitating sensitive sequential studies, ASRA should contribute to our understanding of the role of N‐ras mutations in leukemogenesis.