Sumiko Kobayashi

Progressive disappearance of anti-hepatitis B surface antigen antibody and reverse seroconversion after allogeneic hematopoietic stem cell transplantation in patients with previous hepatitis B virus infection

Reactivation of resolved hepatitis B virus (HBV) infection, which is known as reverse seroconversion (RS), has been reported as a rare complication of allogeneic hematopoietic stem cell transplantation. We retrospectively studied HBV serologic markers in 14 recipients with pretransplant anti-hepatitis B surface antigen antibody (anti-HBs). Progressive decreases in anti-HBs titer were observed in all cases. In 12 cases, anti-HBs titer had decreased to under the protective value. RS occurred in seven cases after disappearance of anti-HBs. Although reseroconversion occurred in five cases, two cases remained in an HBV-carrier status after resolution of hepatitis. In the other five cases, RS did not occur even after disappearance of anti-HBs. The actual risks of anti-HBs disappearance and RS were estimated to be 75.0% and 39.8% at 2 years and 100.0% and 70.0% at 5 years, respectively. In conclusion, RS is a late-onset complication with high frequency that can be predicted by careful monitoring of progressive decrease in anti-HBs titer.

The important balance between cytokines derived from type 1 and type 2 helper T cells in the control of graft-versus-host disease

We have investigated cytokine mRNA expression in the peripheral blood mononuclear cells of 20 patients who received allogeneic hematopoietic stem cell transplants to assess the cytokine network after transplantation. IL-4 mRNA expression decreased in five of five (100%) patients with ⩾grade III (severe) acute GVHD and increased in 10 of 22 (45%) patients without severe GVHD. In contrast, IL-12 mRNA expression increased in two of two (100%) patients with severe GVHD, but increased in only six of 18 (33%) patients without severe GVHD. Furthermore, IL-10 and/or IL-13 mRNA expression increased in 19 of 22 (86%) patients without severe GVHD, but increased in only one of three (33%) patients with severe GVHD. In patients with allogeneic PBSCT who had severe acute GVHD, the cytokine mRNA expression in patients with allogeneic PBSCT, who had no severe GVHD, showed a similar pattern to that in patients with allogeneic BMT. IL-4 mRNA expression increased in three of five (60%) patients and IL-10 and/or IL-13 mRNA expression increased in five of five (100%) patients. In contrast, IL-12 mRNA expression increased in only one of three (33%) patients. Serum IL-4 concentration in allogeneic PBSCT patients in the early engraftment phase was relatively high, while serum IL-12 concentration was low. These findings suggest that severe GVHD may be related to the cytokine imbalance between type 1 helper T (Th1) cells and type 2 helper T (Th2) cells.

Th2 cytokines (IL-4, IL-10 and IL-13) and IL-12 mRNA expression by concanavalin A-stimulated peripheral blood mononuclear cells during chronic graft-versus-host disease

To the Editor: We have reported previously that the response of IFN-)I mRNA expression to the stimulation of concanavalin A (ConA) in peripheral blood mononuclear cells (PBMC) in patients who had extensive chronic graft-versus-host disease (cGVHD) after allogeneic bone marrow transplantation (allo BMT) was not increased compared with those without extensive cGVHD. A similar low response of IL-2 and IL-5 mRNA expression to ConA was observed in such patients with extensive cGVHD (1). Based on these findings, we have investigated the response of IL-4, IL-10 and IL-13 (Th2 cytokines) as well as IL12 (Th2-suppressing cytokine) mRNA expression in PBMC to the stimulation of ConA in patients receiving allo-BMT, since IFN-)I and IL-2 are produced mainly by Thl cells and IL-5 is produced mainly by Th2 cells. Three patients with extensive cGVHD, 5 patients with limited cGVHD, 4 patients without cGVHD, 2 patients in acute phase (3 months after allo-BMT), 1 patient each after autoand syngeneic BMT and 2 normal individuals were analysed in the present study. PBMC were obtained from heparinized fresh blood samples and cultured at 106/ml in RPMI-1640 medium containing 10% fetal calf serum and 5 x M 2-mercaptoethanol. Replicate cultures then received either no stimulation or stimulation by addition of Con A (Pharmacia Fine Chemicals, Uppsala, Sweden) at a final concentration of 5 pg/ml for 12 h. The cells were harvested and the total RNA was extracted and then cytokine gene expression was analysed by semiquantitative RT-PCR as reported previously (Table 1) ( 1-5). Each 5 pg of total RNA was reverse-transcribed with 600 U of murine Moloney leukemia virus reverse transcriptase (BRL, Grand Island, NY, USA) and 150 pmol of random hexamer. An aliquot (1/20th) of the resulting cDNA was used for the semiquantitative polymerase chain reaction (PCR). The following primers were synthesized using a 380B DNA synthesizer (Applied Biosystems):

Expression of HLA‐C‐specific natural killer cell receptors (CD158a and CD158b) on peripheral blood mononuclear cells after allogeneic bone marrow transplantation

We investigated the expression of natural killer cell receptors (NKRs) for HLA‐C on peripheral blood mononuclear cells (PBMCs) in 23 allogeneic bone marrow transplantation (allo‐BMT) patients to analyse the role of NKRs in alloresponse concerning graft‐versus‐host disease (GVHD). CD158a expression was low and there was little change in the expression after allo‐BMT. Also, there was no difference in the proportion of CD158a+/CD3− after allo‐BMT. In contrast, the proportion of CD158b+/CD3− cells, mainly NK cells, increased in the early stage (< 2 months) after allo‐BMT and then gradually decreased (3.3 ± 2.6% before BMT vs. 15.4 ± 8.6% in the early stage after BMT, 8.5 ± 4.9% during the period 3–6 months after BMT and 7.0 ± 3.0% > 6 months after BMT; P < 0.05). However, CD158b expression on CD3+ T cells increased 3 months after allo‐BMT (1.1 ± 1.1% before BMT vs. 5.1 ± 7.7% during the period 3–6 months after BMT and 3.0 ± 2.4% > 6 months after BMT, P < 0.05). The highest percentages of CD158 expression in patients without chronic GVHD (cGVHD) and those with cGVHD were compared. The percentage of CD158b+/CD3+ cells and also that of CD158b+/CD8+ cells were significantly increased in patients with cGVHD compared with those in patients without cGVHD (2.6 ± 2.0% vs. 8.0 ± 11.2% and 2.3 ± 1.5% vs. 8.3 ± 11.7% respectively; P < 0.05). The exact clinical relevance of these CD158b‐expressing cells is not clear. However, there is an interesting possibility that CD158b‐expressing cells play some role in the regulation of GVHD after allo‐BMT.

Early detection of bone marrow involvement in extramedullary plasmacytoma by whole-body F-18 FDG positron emission tomography.

The utility of positron emission tomography (PET) with the glucose analog 2-deoxy-2-[F-18] fluoro-D-glucose (F-18 FDG) in plasmacytoma has not been well evaluated. The authors report a case of extramedullary plasmacytoma (EMP) in which whole-body F-18 FDG PET was useful in the early detection of bone marrow involvement. A 34-year-old woman had EMP without histologic or radiologic evidence of bone lesions. However, the early phase of bone marrow involvement was considered because of spotty accumulation shown by F-18 FDG PET. A follow-up F-18 FDG PET study after radiotherapy showed intense multifocal accumulation in the bone marrow, which was confirmed on bone marrow aspiration as abnormal proliferation of plasma cells. Whole-body F-18 FDG PET has the potential to detect the early phase of bone marrow involvement in patients with EMP.

All-Trans Retionic ACid in the Treatment of Myelodysplastic Syndromes

Myelodysplastic syndromes (MDS) are a group of hematopoietic disorders characterized by uni- or multilineage maturation defects of the bone marrow. Controversial therapeutic results have been obtained using growth factors or differentiating agents such as 13-cis retinoic acid. In this pilot study we evaluated the effects of all-trans retinoic acid (ATRA) in 10 MDS patients (5 male, 5 female). Six patients had refractory anemia (RA), 1 had refractory anemia with excess of blasts (RAEB), and 3 had refractory anemia with excess of blasts in transformation (RAEB-t). All patients received the same dose of ATRA (45 mg/sqm/day) orally for 6 weeks. A rise in hemoglobin concentration > 1g/dl was observed in 3/10 patients, while 5/10 patients showed an increase in granulocyte count > 0.5 x 10(9)/l without concomitant increase in the percentage of blast cells in the bone marrow. A rise in the platelet count > 50 x 10(9)/l was observed in 1/10 patients. All the effects were transient and maximal responses were obtained by the fourth week of treatment. Thereafter, the peripheral blood counts started to drop again, reaching pre-therapy values by the end of the treatment. This phenomenon could be attributed either to the exhaustion of an ATRA-responding cell pool, the development of cellular resistance to ATRA or to a reduction of plasma ATRA levels after prolonged treatment. According to our results, it seems that ATRA might have therapeutic efficacy in MDS, particularly if its effect could be improved by combinations with other differentiating agents or growth factors.

Fatal cerebral hemorrhage associated with cyclosporin-A/FK506-related encephalopathy after allogeneic bone marrow transplantation

Abstract We report a case of cerebral hemorrhage associated with cyclosporin A (CsA)/FK506-related encephalopathy that developed in a 16-year-old woman after allogeneic bone marrow transplantation. Hematopoietic engraftment occurred on day 15, and the patient developed systemic convulsions after CsA was replaced by FK506 for the treatment of acute graft-versus-host disease (GVHD). Based on magnetic resonance imaging, laboratory findings and cerebrospinal fluid studies, she was diagnosed as having CsA/FK506-related encephalopathy with cerebral hemorrhagic infarction. Although she recovered completely after discontinuation of FK506, she developed convulsions again 15 days after re-administration of FK506. A computed tomography scan showed cerebral hemorrhage. She died of respiratory failure. Vascular damage induced by immunosuppressive drugs and enhanced by acute GVHD seemed to be the cause of the cerebral hemorrhage. Since hypertension, which was present during both of the central nervous system events, seemed to have contributed to the development of the cerebral hemorrhage, it is proposed that CsA and FK506 should be reduced or discontinued when patients who have risk factors of hypertension become hypertensive even if they have no symptoms of neurotoxicity.

Transformation of severe aplastic anemia into acute myeloblastic leukemia with monosomy 7

A cytogenetically normal man with severe aplastic anemia was treated with granulocyte colonystimulating factor (G-CSF), erythropoietin (EPO), cyclosporin A, anti-thymocyte globulin, and interleukin-6 (IL-6), which resulted in a gradual improvement in his neutrophil count and hemoglobin level. After 2 years of the therapy, monosomy 7 was detected during cytogenetic analysis of his bone marrow, which evolved during a period of 5 months into acute myeloblastic leukemia. An in vitro proliferation assay of cytokine responses showed that leukemic blasts were sensitive only to G-CSF, and not to EPO or IL-6. Although allogeneic bone marrow transplantation from an HLA-matched unrelated donor was carried out in the non-remission stage, the patient died of systemic fungal infection on day 25, without any evidence of hematological engraftment. As long-term use of cytokines and immunomo-suppressants in patients with severe aplastic anemia may induce or hasten the onset of a malignant transformation, careful attention must be paid to clonal evolution. Due to the poor prognosis of secondary myelodysplasia and leukemia, allogeneic bone marrow transplantation for such patients must be carried out early in the course of the disease.

Cytokine gene expression in the mixed lymphocyte culture in allogeneic bone marrow transplants as a predictive method for transplantation-related complications

SUMMARY. We have investigated cytokine gene expression in the two‐way mixed lymphocyte cultures (MLC) enhanced by concanavalin A to assess whether this is a useful predictive method for severe graft‐versus‐host disease (GVHD) and graft failure in nine allogeneic bone marrow transplantation (allo‐BMT) patients. Our present study revealed that increased IL‐2, IL‐5 and IFN‐γ mRNA expression and IL‐2 and IFN‐γ production in the MLC in two cases with severe lethal transplantation‐related complications (graft failure and grade III acute GVHD). These findings suggest that increased cytokine mRNA expression and cytokine products in this assay may be predictive of the development of transplantation‐related complications.

Clinical Relevance of Serum Soluble Interleukin-2 Receptor Levels in Acute and Chronic Graft-versus-Host Disease

Soluble interleukin-2 receptor (sIL-2R) levels were analyzed in the sera from 27 patients who underwent allogeneic bone marrow transplantation (BMT) in order to to examine whether there was any correlation between sIL-2R levels and graft-versus-host disease (GVHD). The sIL-2R levels markedly increased at the engraftment period, mainly due to cytokine administration shortly after BMT. Although the sIL-2R levels increased at the onset of acute GVHD, the subsequent development of GVHD could not be predicted by the sIL-2R levels documented before acute GVHD. As acute GVHD improved, the sIL-2R levels decreased, thus showing that the sIL-2R levels correlated with the disease status. In patients without acute GVHD, the sIL-2R levels gradually decreased with time and returned to the pretransplant levels after about 12 weeks post BMT. The sIL-2R levels were higher in unrelated allogeneic BMT patients with acute GVHD when compared with related allogeneic BMT patients. There was a significant increase in the sIL-2R levels at the engraftment period and at the onset of acute GVHD. At the onset of chronic GVHD, the sIL-2R levels once again increased and then decreased as chronic GVHD improved. Prolonged increase in sIL-2R levels was followed by subsequent development of chronic GVHD. Patients with a poor prognosis had higher sIL-2R levels than those with a good prognosis. Therefore, it seems that sIL-2R is a useful marker for monitoring the disease status of acute and chronic GVHD.