American Journal of Hematology

Shiokawa, Satoshi; Suehiro, Youko; Uike, Naokuni; Muta, Koichiro; Nishimura, Junji

doi: 10.1002/ajh.1169pmid: 11754393

Waldenström's macroglobulinemia (WM) is a malignant lymphoplasmo‐proliferative disorder with monoclonal pentameric immunoglobulin (Ig)M production. The most consistent feature of clonal B cells in the bone marrow (BM) and/or lymph nodes of patients with WM is the presence of pleomorphic B‐lineage cells at different stages of maturation, such as small lymphocytes, lymphoplasmacytoid cells, and plasma cells. Monoclonal lymphocytes express μ chains with or without δ chains. A recent DNA analysis of WM tumor clones showed WM to be derived from B cells that have been selected by antigen at a relatively late stage of differentiation. To further clarify the origin of WM tumor cells, we analyzed the variable (V) domain sequences of tumor derived μ and δ transcripts. The expression of δ transcripts was also examined in peripheral blood (PB) and BM using the reverse transcriptase polymerase chain reaction (RT‐PCR) combined with a single‐strand conformation polymorphism (SSCP) analysis. The sequences were identical among the μ and δ transcripts in each patient and the level of somatic mutation in the VH regions expressed by tumor cells was in the same range as that of IgM‐only B cells and IgM+IgD+ memory B cells. In our previous RT‐PCR‐SSCP analysis, a single dominant band of the μ isotype was observed in BM and PB in all patients. However, common dominant bands in BM and PB were detected in only one patient in a δ transcript analysis. In the rest of the patients, monoclonal δ transcripts were only detected in BM. Our results suggest that a normal counterpart of WM cells is somatically mutated IgM+IgD+ and/or IgM‐only B cells and the expression patterns of monoclonal μ and δ transcripts differ between BM and PB in some cases of WM. Am. J. Hematol. 68:139–143, 2001. © 2001 Wiley‐Liss, Inc.

Martínez‐Jaramillo, Guadalupe; Flores‐Figueroa, Eugenia; Gómez‐Morales, Enrique; Sánchez‐Valle, Elizabeth; Mayani, Hector

doi: 10.1002/ajh.1170pmid: 11754394

We have previously shown that the levels of hematopoietic progenitors in long‐term marrow cultures (LTMC) from patients with aplastic anemia (AA) are drastically reduced, as compared to normal LTMC. We have also reported that when LTMC from AA patients are supplemented with recombinant human granulocyte‐macrophage colony‐stimulating factor (rhGM‐CSF) there is an increase in colony‐forming cell (CFC) levels. However, such a stimulation is only transient and it is followed by an inhibition in CFC growth. Based on these observations, in the present study we have tested the hypothesis that the levels of tumor necrosis factor‐α (TNF‐α), an inhibitor of hematopoiesis, are increased in AA LTMC and that such levels are further increased after rhGM‐CSF has been added to the cultures for several weeks. Accordingly, we have determined the levels of TNF‐α in the supernatant of LTMC established from normal (n = 8) and AA (n = 6) bone marrow and in AA LTMC supplemented with rhGM‐CSF (n = 6). At the time of culture initiation, TNF‐α levels were below detection in all the samples analyzed. After 5 weeks of culture, TNF‐α levels in normal LTMC were very low, with a median of 7.3 pg/mL. In contrast, AA LTMC contained higher levels of TNF‐α (median of 49.6 pg/mL). In keeping with our hypothesis, addition of rhGM‐CSF to AA LTMC resulted in a significant further increase of TNF‐α levels (median of 135.4 pg/mL). Our results demonstrate an inverse correlation between reduced hematopoiesis in AA LTMC and increased levels of TNF‐α in this culture system. Based on the results presented here, together with previous reports indicating that TNF‐α is a potent inducer of apoptosis in hematopoietic progenitor cells, it seems reasonable to suggest that TNF‐α is implicated in the pathophysiology of AA. Am. J. Hematol. 68:144–148, 2001. © 2001 Wiley‐Liss, Inc.

Losco, Patricia; Nash, Gerard; Stone, Phil; Ventre, John

doi: 10.1002/ajh.1171pmid: 11754395

Iodinated radiographic contrast media have traditionally been contraindicated in patients with sickle cell disease because their high osmolality may induce osmotic shrinkage of red blood cells, impair blood flow through the microcirculation, and precipitate or exacerbate a sickle cell crisis. This study investigated that concept by comparing the hematological and rheological effects in vitro of four X‐ray contrast media of differing osmolalities: Visipaque (290 mOsm/kg), Hexabrix (600 mOsm/kg), Omnipaque (844 mOsm/kg), and RenoCal‐76 (1940 mOsm/kg). Blood was tested from 10 normal and 10 sickle cell donors at drug concentrations of 0, 1, 10, and 30% w/v in an attempt to approximate the relative concentrations of contrast medium to blood that might occur during the bolus‐injection and circulation‐diluted phases of drug administration. Parameters evaluated included hematology, red cell morphology, and red cell flow resistance through a micropore filter to approximate the microcirculatory effects. Significant hematological effects for both normal and sickle cell donors included a concentration dependent decrease in hematocrit and MCV, and increase in MCHC, all of which varied directly with the osmolality of the contrast media in the order of RenoCal‐76 > Omnipaque > Hexabrix > Visipaque. The contrast media had minor effects on red blood cell morphology except for RenoCal‐76, 10–30% in which marked echinocytosis was observed. There was no significant increase in the number of irreversibly sickled cells in donors with hemoglobin S. Filterability of red cell suspensions through capillary size pores was impaired in both normal and sickle cell samples in direct proportion to the osmolality of the contrast media, as listed above. Filterability effects were greater for sickle cells than for normal red cells. Visipaque, which was closest to isotonicity, had little effect on red cell volume and had no significant effect on filterability of normal or sickle cells. These results suggest that microcirculatory impairment following infusion of contrast media may occur in sickle patients because of the unusual rheological sensitivity of HbSS red cells, and may be avoided by choice of an isotonic medium. Am. J. Hematol. 68:149–158, 2001. © Wiley‐Liss, Inc.

Kamali, Farhad; Edwards, Clive; Wood, Peter; Wynne, Hilary A.; Kesteven, Patrick

doi: 10.1002/ajh.1172pmid: 11754396

There is no information available on temporal variability in plasma vitamin K concentrations and its relationship to coagulation processes. We investigated the possible existence of temporal changes in plasma vitamin K and lipid concentrations and activity of clotting factors II, VII, IX, and X and relationships between these variables. Plasma vitamin K and lipid concentrations and clotting factor activity were measured at four‐hour intervals for 28 hours in a group of healthy volunteers. Temporal variations existed in plasma vitamin K concentrations, with a mean maximum at 22:00 hr and a mean minimum (32% of the maximum) at 10:00 hr. Plasma triglycerol concentrations mirrored the changes in vitamin K concentrations. Mean factor VII activity was positively correlated with mean total plasma cholesterol concentrations (r = 0.714; P < 0.0001) and with mean plasma low density lipoprotein (LDL) cholesterol concentrations (r = 0.461; P < 0.0001). No distinct correlations were found between plasma vitamin K concentrations and either high density lipoprotein (HDL) or LDL cholesterol concentrations, or between triglycerol, HDL, or LDL cholesterol concentrations and functional activity of factors II, IX, and X. Plasma vitamin K concentrations did not correlate with the functional activity of any of the clotting factors. The presence of a correlation between plasma cholesterol concentrations and factor VII activity for blood samples collected at four‐hour intervals suggests that plasma cholesterol concentrations may have a more acute effect on factor VII activity. Temporal variations in plasma vitamin K concentrations indicate that a single time point measurement may be an inappropriate method of establishing vitamin K status in an individual. Am. J. Hematol. 68:159–163, 2001. © 2001 Wiley‐Liss, Inc.

Tang, Delia C.; Prauner, Ron; Liu, Wenli; Kim, Kye‐Hyun; Hirsch, Robert P.; Driscoll, M. Catherine; Rodgers, Griffin P.

doi: 10.1002/ajh.1173pmid: 11754397

Stroke is one of the most devastating complications of patients with sickle cell disease (SCD). Currently, there are no known molecular or genetic markers that can be used to assess the risk of stroke in this population. We have previously shown that relative hypertension may be one risk factor for stroke in SCD. In a case‐control study, we investigated the association between GT‐repeat polymorphism within the angiotensinogen (AGT) gene and the risk of stroke in pediatric patients with SCD. After informed consent was obtained, 63 patients (21 stroke subjects and 42 nonstroke control subjects matched according to age and sex) with SCD followed at local pediatric hematology clinics were genotyped to test the association of specific GT‐repeat alleles of the AGT gene and occurrence of stroke. There were statistical differences in the distribution of the genotypes among stroke and nonstroke SCD patients (χ2 = 10.82, df = 11, P < 0.05). We also found GT‐repeat alleles A3 and/or A4 of the AGT gene conferred a four‐fold increase in the risk of stroke (odds ratio [OR] = 4, P < 0.05). The attributable odds ratio for allele A3 and A4 is 2.24 and 4.33, respectively (P < 0.005). Our results suggest that GT‐repeat within the AGT gene may be associated with risk of stroke in pediatric SCD. The relative risk of stroke in the presence of alleles A3 and/or A4 is fourfold greater than in the absence of these alleles. If these data are substantiated in a larger cohort of patients, our results indicate that the determination of GT‐repeat of AGT gene may be a useful genetic marker to assess the risk for stroke of patients with SCD. Am. J. Hematol. 68:164–169, 2001. Published 2001 Wiley‐Liss, Inc.

Lavelle, Donald; Chen, Yi‐Hsiang; Hankewych, Maria; DeSimone, Joseph

doi: 10.1002/ajh.1174pmid: 11754398

Histone deacetylase (HDAC) inhibitors cause growth arrest and apoptosis of cancer cells by both p21‐dependent and independent mechanisms. Decreased expression of growth factor receptors may be a key factor in the p21‐independent mechanism, although this has not been directly tested. We have tested the effects of sodium butyrate and trichostatin A on human myeloma cell lines and have observed G1 arrest and apoptosis associated with increased expression of p21WAF1, Bax, Rb dephosphorylation, and decreased IL‐6 receptor (IL‐6R) expression. Experiments to determine the role of disruption of IL‐6 signaling as a result of decreased IL‐6 receptor expression in mediating these effects were conducted using a stable transfectant of the OPM‐2 line which constitutively expressed the IL‐6 receptor. Our results indicated that decreased IL‐6R expression was not required for induction of p21WAF1 or apoptosis. Thus, HDAC inhibitors appear to activate multiple cellular pathways, leading to growth arrest and apoptosis, and their use in the treatment of myeloma, particularly in combination with other agents, warrants further investigation. Am. J. Hematol. 68:170–178, 2001. © 2001 Wiley‐Liss, Inc.

Schnog, J.B.; Kater, A.P.; Mac Gillavry, M.R.; Duits, A.J.; Lard, L.R.; van der Dijs, F.P.L.; Brandjes, D.P.M.; ten Cate, H.; Statius van Eps, L.W.; Rojer, R.A.

doi: 10.1002/ajh.1175pmid: 11754399

Vasoocclusion is a continuous process in sickle cell disease (SCD) and accumulates to significant end organ damage, mostly irrespective of the occurrence of manifest acute vasoocclusive events. As there are indications that reversing the hypercoagulable state may be of clinical benefit in sickle cell patients, we performed a randomized, double blind, placebo‐controlled, cross‐over pilot study to assess the efficacy and safety of low‐adjusted dose acenocoumarol therapy (International Normalized Ratio: 1.6–2.0) in SCD. Treatment consisted of either acenocoumarol or placebo for 14 weeks, after which treatment was discontinued for a period of five weeks. Then, patients initially on acenocoumarol received placebo (and vice versa) for 14 weeks. Therapy efficacy was assessed by comparing the frequency of vasoocclusive complications, the occurrence of bleeding, and clotting activation between acenocoumarol and placebo treatment of each individual patient. Twenty‐two patients (14 homozygous [HbSS] and 8 double heterozygous sickle‐C [HbSC]; aged 20–59 years) completed the entire study. Acenocoumarol treatment did not result in a significant reduction of acute vasoocclusive events (three painful crises during acenocoumarol, five painful crises during placebo). There was a marked reduction of the hypercoagulable state (depicted by a decrease in plasma levels of prothrombin F1.2 fragments [P = 0.002], thrombin‐antithrombin complexes [P = 0.003], and D‐dimer fragments [P = 0.001]) without the occurrence of major bleeding. Even though no clinical benefit (pertaining to the frequency of painful crises) was detected in this pilot study, the value of low adjusted‐dose acenocoumarol for preventing specific events (such as strokes) and as a long‐term treatment of sickle cell patients should be subject of further study. Am. J. Hematol. 68:179–183, 2001. © 2001 Wiley‐Liss, Inc.

Robbins, Daniel; Kulkarni, Roshni; Gera, Renuka; Scott‐Emuakpor, Ajovi B.; Bosma, Kathy; Penner, John A.

doi: 10.1002/ajh.1176pmid: 11754400

We describe two patients with mild hemophilia A (MHA) who developed high titer inhibitor (HTI) following intensive recombinant factor VIII (rFVIII) concentrate replacement for surgery and trauma. Intranasal desmopressin was instituted shortly following immunosuppressive therapy (IST) and activated prothrombin complex concentrate (APCC) in one case, and following APCC alone in the second case. Avoidance of factor VIII (FVIII) coupled with intranasal desmopressin prophylaxis three times a week resulted in undetectable inhibitor levels. Both patients have had no further bleeding episodes and have been maintained on desmopressin prophylaxis prior to activity for the past 2 to 3 years. Recombinant factor VIIa (rFVIIa) was used successfully prior to a second surgery in one patient without complication. Am. J. Hematol. 68:184–188, 2001.© 2001 Wiley‐Liss, Inc.

Goldenberg, Alec S.; Tiesinga, James J.

doi: 10.1002/ajh.1177pmid: 11754401

The SNARECOIL™ needle is a specimen capturing bone marrow biopsy needle that incorporates a tiny internal capturing coil. It was developed to minimize postinsertion needle manipulation and to facilitate specimen recovery. Forty‐four patients underwent 50 bone marrow biopsy procedures with the SNARECOIL needle for a variety of hematologic indications. Second and third procedures were done for follow‐up or staging. Each procedure retrieved a specimen with an average length of 2.1 cm. Fifty‐two percent of the specimens were ⩾2.0 cm in length. The majority of specimens demonstrated intact marrow architecture enabling a pathologic diagnosis in every case. Delicate cores of nontrabeculated marrow were recovered in three cases. The SNARECOIL bone marrow biopsy needle reliably retrieves intact bone marrow core specimens for pathologic interpretation. Am. J. Hematol. 68:189–193, 2001 © 2001 Wiley‐Liss, Inc.

Kawasaki, Hirohide; Nakano, Takahide; Kohdera, Urara; Kobayashi, Yohnosuke

doi: 10.1002/ajh.1178pmid: 11754402

Essential thrombocythemia (ET) is a clonal myeloproliferative disorder characterized by an elevation of platelets in peripheral blood and excessive proliferation of megakaryocytes in bone marrow. The pathological mechanisms for the elevation of megakaryocytes and platelets in ET remain unclear. To study the hypersensitivity of megakaryocyte progenitors to thrombopoietin (TPO), a 9‐year‐old girl, diagnosed with ET, underwent dose‐response experiments with recombinant human TPO, using her bone marrow non‐adherent mononuclear cells and CD34 positive cells. Spontaneous colony‐forming unit‐megakaryocytes (CFU‐Meg) were observed in serum‐deprived cultures of non‐adherent mononuclear cells, whereas they disappeared in cultures of CD34 positive cells. The patient's CFU‐Meg showed maximal growth at concentrations of TPO lower than those for normal children. Dose‐response curves demonstrated a 50–80 fold increase in sensitivity of the patient's CFU‐Meg to TPO. We observed hypersensitivity of megakaryocyte progenitors to TPO in a child with ET. Our results suggest that spontaneous CFU‐Meg formation in patients with ET may be due to hypersensitivity to TPO. Am. J. Hematol. 68:194–197, 2001. © 2001 Wiley‐Liss, Inc.