Soluble Interleukin-2 Receptor Index Predicts Outcomes After Cord Blood Transplantation.

BACKGROUND: Our previous study demonstrated that the soluble interleukin-2 receptor (sIL-2R) index, defined as the ratio of serum sIL-2R levels at neutrophil engraftment to that before conditioning, is a biomarker that can predict acute graft-vs-host disease (GVHD) after unrelated bone marrow transplantation. In the present study, we evaluated the significance of the sIL-2R index among patients who underwent cord blood transplantation (CBT). METHODS: We retrospectively analyzed 31 patients who underwent single-unit CBT as their first transplantation for hematologic malignancies. RESULTS: The median sIL-2R index was 4.2. The cumulative incidence of grade II to IV acute GVHD was not associated with the sIL-2R index. However, the cumulative incidence of relapse at 3 years after transplantation was significantly lower, with an sIL-2R index ≥ 3.7 than with an index < 3.7 (12.8% vs 50.0%; P = .04). As a result, the probability of overall survival at 3 years after transplantation was significantly higher in the former group than in the latter (79.8% vs 20.0%; P < .01). Only the dose of corticosteroid administered in the pre-engraftment period influenced the sIL-2 index. CONCLUSION: The sIL-2R index can predict the incidence of relapse and probability of survival after CBT, possibly reflecting a graft-vs-leukemia effect.

TERT and JAK2 polymorphisms define genetic predisposition to myeloproliferative neoplasms in Japanese patients.

Myeloproliferative neoplasms (MPNs), including polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF), are often characterized by specific somatic mutations in any of the three genes: JAK2, CALR, or MPL. A single nucleotide polymorphism (SNP), rs2736100, in the reverse transcriptase gene (TERT) and a germline JAK2 46/1 haplotype have been associated with MPNs in North American and European patients. We examined 201 Japanese MPN patients, including 52 with PV, 131 with ET, and 18 with PMF, as well as 366 control individuals for TERT rs2736100 and JAK2 rs10974944, a tagging SNP of the 46/1 haplotype. Furthermore, correlations between the JAK2 V617F allele burden at diagnosis and TERT rs2736100 or JAK2 rs10974944 were evaluated using a digital PCR assay for accurate quantitation. The JAK2 46/1 haplotype, but not the TERT rs2736100 SNP, was correlated to the JAK2 V617F mutant allele burden in JAK2 V617F-positive MPN patients. In conclusion, we demonstrated that both TERT rs2736100_C and JAK2 46/1 haplotype are predisposing factors for MPNs in Japanese patients. While TERT rs2736100_C tended to have a more general, non-specific effect on all MPNs, the JAK2 46/1 haplotype was essentially predisposed to the JAK2 V617F-positive MPNs.

[Refractory primary immune thrombocytopenia in pregnancy requiring splenectomy and repeated intravenous immunoglobulin therapy].

A 30-year-old primigravid woman without a history of thrombocytopenia was referred to our hospital because of severe thrombocytopenia (<1,000 thrombocytes/µl) at 16 weeks of gestation and diagnosed with idiopathic thrombocytopenic purpura (ITP). There was no improvement in the platelet count after treatment with 0.5-1.0 mg/kg/day prednisolone, and the 400 mg/day intravenous immunoglobulin (IVIg) administered for 5 days gradually became ineffective; therefore, a laparoscopic splenectomy was performed at 25 weeks of gestation. The increase in the platelet count after the splenectomy was temporary, but the effects of IVIg therapy improved, and the patient received IVIg therapy seven times in total during her pregnancy. Her platelet count ranged between 10,000 and 70,000/µl after the splenectomy, compared with <5,000/µl before the surgery. The patient underwent an elective cesarean section at 34 weeks of gestation without any significant bleeding. The baby was diagnosed with thrombocytopenia at birth (34,000 thrombocytes/µl) and was administered only one dose of IVIg, which increased the baby's platelet count to a normal level after 14 days. The patient's platelet count also increased after delivery. Splenectomy and repeated IVIg therapy can be considered for refractory severe ITP during pregnancy.

Successful Treatment of Behçet's Disease Associated with Acute Myeloid Leukemia with Myelodysplasia-related Changes Using Azacitidine and Tacrolimus before Allogeneic Hematopoietic Stem Cell Transplantation.

The coexistence of acute myeloid leukemia (AML) with Behçet's disease (BD) is rare. The optimum treatment for AML-associated BD has not been established. We herein report a patient with BD who developed AML with myelodysplasia-related changes. Induction chemotherapy caused complete remission of the AML but worsened the BD. Thereafter, AML was treated with azacitidine. The BD was steroid-dependent. Tacrolimus was added, which improved the BD. The patient underwent allogeneic hematopoietic stem cell transplantation (HSCT) and remains in complete remission for both diseases. Allogeneic HSCT was found to be a potent therapeutic option for AML-associated BD. In addition, azacitidine and tacrolimus were shown to be a suitable bridging regimen before HSCT.

Soluble interleukin-2 receptor index predicts the development of acute graft-versus-host disease after allogeneic hematopoietic stem cell transplantation from unrelated donors.

Acute graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (HSCT) is associated with significant morbidity and mortality. In the present study, we retrospectively evaluated whether soluble interleukin-2 receptor (sIL-2R) index, defined as the ratio of serum sIL-2R levels at neutrophil engraftment to those at the pre-conditioning regimen, was predictive of acute GVHD among 51 patients who underwent allogeneic HSCT as their first transplantation and achieved engraftment. The median sIL-2R index was 3.6, and the sIL-2R values were positively associated with acute GVHD severity (grade 0-I: 3.8 ± 2.0 vs. grade II-IV: 7.1 ± 5.7; P = 0.05). Grade II-IV acute GVHD had a cumulative incidence of 31.4 %, and was significantly more frequent among patients with an sIL-2R index of ≥4.5 (≥4.5: 50.0 % vs. <4.5: 21.2 %; P = 0.03). Multivariate analysis revealed that an sIL-2R index of ≥4.5 [hazard ratio (HR) 3.5, P < 0.01] and donor age of >35 years (HR 3.8, P = 0.02) were significant risk factors for grade II-IV acute GVHD. Therefore, increased sIL-2R levels from baseline to engraftment might predict the risk of moderate-to-severe acute GVHD after allogeneic HSCT from an unrelated donor.

Altered expression of circadian clock genes during peripheral blood stem cell mobilization induced by granulocyte colony-stimulating factor.

Circulating hematopoietic stem cells exhibit robust circadian fluctuations, which influence the mobilized cell yield, even during enforced stem cell mobilization. However, alterations in the expression of circadian clock genes during granulocyte colony-stimulating factor (G-CSF)-induced peripheral blood stem cell (PBSC) mobilization are not fully elucidated. Therefore, we measured the expression of these genes in human peripheral blood leukocytes from 21 healthy donors. While CRY1 mRNA expression significantly increased by 3.9-fold (p < 0.01), the expression of PER3, CRY2 and BMAL1 mRNAs significantly decreased (by 0.2-fold, 0.2-fold, and 0.6-fold, respectively; p < 0.001) after G-CSF administration. Moreover, CRY1 mRNA expression was inversely correlated with the plasma level of noradrenaline (r = -0.36, p < 0.05), while PER3, CRY2, and BMAL1 mRNA expression directly correlated with the plasma level of noradrenaline (r = 0.55, r = 0.66, and r = 0.57, respectively; p < 0.001). Thus, significant correlations between the levels of circadian clock gene mRNAs and the plasma level of noradrenaline, a sympathetic nervous system neurotransmitter, were established. The modulation of sympathetic activation and of the circadian clock may be novel therapeutic targets for increasing stem cell yields in PBSC donors.

JAK2 46/1 haplotype is associated with JAK2 V617F-positive myeloproliferative neoplasms in Japanese patients.

Myeloproliferative neoplasms (MPNs) constitute a group of phenotypically diverse chronic myeloid malignancies, characterized by clonal hematopoiesis and excessive production of terminally differentiated myeloid blood cells. The MPNs include polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF), most of which are characterized by a somatic point mutation, V617F, in the janus kinase 2 (JAK2) gene. This mutation was recently shown to occur more frequently in a specific JAK2 haplotype, JAK2 46/1, in North American and European MPN patients. Little is known, however, about JAK2 haplotypes in Japanese MPN patients. Therefore, we examined 108 Japanese patients with MPN, including 19 with PV, 61 with ET, 10 with PMF, and 17 with unclassifiable MPN, as well as 104 control individuals for the JAK2 rs10974944(C/G) single nucleotide polymorphism, in which the G allele indicates the 46/1 haplotype. We found that the JAK2 46/1 haplotype was significantly more frequent in patients with V617F-positive MPN than in controls (odds ratio [OR], 3.6; 95 % confidence interval [CI], 2.2-5.8, p < 0.001), and in PV patients than in controls (OR, 6.3; 95 % CI, 3.0-29.4, p < 0.001). In conclusion, we demonstrated that the JAK2 46/1 haplotype is associated with JAK2 V617F-positive MPNs in Japanese patients.

Hematopoietic progenitor cell count, but not immature platelet fraction value, predicts successful harvest of autologous peripheral blood stem cells.

Mobilized stem cells in the peripheral blood (PB) must be efficiently harvested at the appropriate time before autologous PB stem cell (PBSC) transplantation. Enumeration of CD34+ cells in the PB before apheresis predicts the number of PBSCs that can be collected, but the cytometric techniques used are complex and expensive. Therefore, it is necessary to identify an alternative to the CD34+ cell count in PBSC harvest-time monitoring. Fully automated flow cytometry using blood cell counters now allows reliable quantification of immature myeloid cells in the PB, referred to as hematopoietic progenitor cells (HPC), and reticulated platelets, expressed as the immature platelet fraction (IPF). Immature or reticulated platelets are thought to correlate with thrombopoietic activity of the marrow. Following a chemotherapy nadir, the recovery of white blood cell and platelet counts has been used to determine the right time for apheresis. Therefore, we examined whether the HPC count and IPF value could be used to predict PBSC mobilization in 20 patients with hematological malignancies. The HPC count was found to be correlated with the CD34+ cell count (r = 0.84, P < 0.01), whereas the IPF value was not (r = 0.37, P = 0.44). Therefore, the HPC count, but not the IPF value, is a possible predictor of the timing of autologous stem cell transplantation.

Alteration of adipokines during peripheral blood stem cell mobilization induced by granulocyte colony-stimulating factor.

Adipokines, soluble mediators produced by adipocytes, have been shown to play a role in various physiological and pathological conditions. We investigated the involvement of adipokines in granulocyte colony-stimulating factor (G-CSF)-induced mobilization of hematopoietic stem cells in 21 healthy donors. We found that serum visfatin and resistin levels, but not leptin and adiponectin levels, were significantly elevated by G-CSF treatment. G-CSF treatment activated signaling proteins like extracellular signal-regulated kinase and stimulated secretion of visfatin from 3T3-L1 adipocytes. These findings suggest that some adipokines may play a role in G-CSF-induced mobilization of stem cells from the bone marrow into systemic circulation.

14-3-3zeta is indispensable for aggregate formation of polyglutamine-expanded huntingtin protein.

Huntington's disease (HD) is an autosomal dominant progressive neurodegenerative disorder caused by polyglutamine (polyQ) expansions in the huntingtin (Htt) protein. A hallmark of HD is the presence of aggregates-predominantly composed of NH(2)-terminal fragments of polyQ-expanded Htt-in the nucleus and cytoplasm of affected neurons. We previously proposed that 14-3-3zeta might act as a sweeper of misfolded proteins by facilitating the formation of aggregates possibly for neuroprotection; these aggregates are referred to as inclusion bodies. However, evidence available in this regard is indirect and circumstantial. In this study, analysis of the aggregation-prone protein Htt encoded by HD gene exon 1 containing polyglutamine expansions (Htt86Q) revealed that 17 residues in the NH(2)-terminal of this protein are indispensable for its aggregate formation. Immunoprecipitation assays revealed that 14-3-3beta, gamma, eta, and zeta interact with Htt86Q transfected in N2a cells. Interestingly, the small interfering ribonucleic acid (siRNA) suppression of 14-3-3zeta exclusively abolished Htt86Q aggregate formation, whereas 14-3-3beta or eta siRNA suppression did not. This indicates that 14-3-3zeta participates in aggregate formation under nonnative conditions. Our data support a novel role for 14-3-3zeta in the aggregate formation of nonnative, aggregation-prone proteins.

Region-specific expression and sex-steroidal regulation on aromatase and its mRNA in the male rat brain: immunohistochemical and in situ hybridization analyses.

The brain has an estrogen-biosynthetic potential resulting from the presence of neuronal aromatase, which controls the intraneural sex-steroidal milieu and is involved in brain sexual differentiation, psychobehavioral regulation, and neuroprotection. In the rat brain, three distinct aromatase-P450-immunoreactive (AromP450-I) neural groups have been categorized in terms of their peak expression time (fetal, fetoneonatal, and young-to-adult groups), suggesting the presence of region-specific regulation on brain AromP450. In the present study, we compared the expressions between AromP450 protein and mRNA by using immunohistochemistry and in situ hybridization with an ovary-derived cRNA probe in serial sections of fetal, fetoneonatal, and adult male rat brains and then performed steroidal manipulations to evaluate the sex-steroidal effects on AromP450 in adult orchiectomized and adrenalectomized (OCX + ADX) male rats. As a result, prominent mRNA signals were detected in the fetal (i.e., the anterior medial preoptic nucleus) and fetoneonatal (i.e., the medial preopticoamygdaloid neuronal arc) groups, although no detectable signal was found in the "young-to-adult" group (i.e., the central amygdaloid nucleus). In addition, the "fetoneonatal" AromP450-I neurons were prominently reduced in number and intensity after OCX + ADX and then were reinstated by the administration of dihydrotestosterone, testosterone, or 17beta-estradiol. In contrast, none of the sex steroids had any significant effects on the young-to-adult group. Several possible explanations were explored for why the young-to-adult group may differ in aromatase expression and regulation, including the possibility that distinct splicing variants or isozymes for aromatase exist in the rat brain.

Expression of estrogen receptors (alpha, beta) and androgen receptor in serotonin neurons of the rat and mouse dorsal raphe nuclei; sex and species differences.

Sex steroids have been inferred to be involved in the regulation of affective status at least partly through the serotonergic (5-HT) system, particularly in the dorsal raphe nucleus (DRN), which innervates enormous projections to the cerebral cortex and limbic system. In the present study, the expression of estrogen receptors-alpha and -beta (ERalpha, ERbeta), androgen receptor (AR) and 5-HT was examined immunohistochemically in the rat and mouse DRN in both sexes. The results showed that large numbers of ERalpha- and/or ERbeta-immunoreactive (ERalpha-I, ERbeta-I) cells were found in the DRN of both male and female mice, whereas only small numbers of ERalpha-I cells and no ERbeta-I cells were seen in the rat DRN of each sex. With respect to AR-immunoreactive (AR-I) cells, moderate numbers of such cells were present only in male rats and mice, and no or very few could be observed in female ones. The ERalpha-I, ERbeta-I, and AR-I cells were mainly distributed in the rostral DRN. In double-immunostaining, many 5-HT-I neurons were found to show ERalpha and/or ERbeta expression specifically in the rostral DRN (particularly dorsal, ventral and interfascicular parts) of mice of both sexes, but not in that of rats. In contrast, only a few 5-HT neurons were observed to show AR expression in the DRN of both rodents. The current results strongly suggest that sex steroids can modulate the affective regulation of the serotonergic system through ERalpha and/or ERbeta in 5-HT neurons of the mouse rostral DRN (but not so much through AR), and that such effects might be different depending on the sex and species, as shown by the prominent sex differences in AR expression and prominent species differences in ERalpha and ERbeta expression.

Gonadal and adrenal effects on the glucocorticoid receptor in the rat hippocampus, with special reference to regulation by estrogen from an immunohistochemical view-point.

Focusing on the hippocampal CA1 region, effects of peripheral gonadal and adrenal steroids on the glucocorticoid receptor (GR) were immunohistochemically evaluated in male and female adult rat brains after adrenalectomy (ADX), gonadectomy (GDX), and administration of estradiol (E2) and/or corticosterone (CS). In ADXed male rats, the hippocampal nuclear GR decreased and turned back to the cytoplasm, whereas in females, nuclear localization persisted even after ADX. In GDX+ADXed female rats, the GR was dispersedly translocated from the nucleus to the cytoplasm as well as in GDX+ADXed males. The dispersed cytoplasmic GR was again translocated into the nucleus by administration of CS. In addition, administration of a small dose of E2 for 4-13 days was found to sufficiently recover the nuclear location of GR in GDX+ADXed rat brains, whereas medium-to-large doses could not do this. Also, a longer administration more strongly enhances the nuclear GR location and expression. The present study provided strong immunohistochemical evidence that the sexually dimorphic effects of ADX on hippocampal GR are attributable to gonadal hormones, and that E2 is implicated in the effects in inversely-dose- and directly-duration-dependent manner. Taken together, intriguing gonadal and adrenal crosstalk is considered to play some important role in regulating hippocampal GR morphology and to have a possibly crucial influence on stress-related disorders such as depression.