Nitrogen balance in patients with hemiparetic stroke during the subacute rehabilitation phase.

BACKGROUND: In highly invasive diseases, metabolism commonly changes. Hypercatabolism is frequent in acute stroke, and nitrogen balance tends to be negative. However, there has been no study describing nitrogen balance in subacute and chronic stroke patients. The present study aimed to examine nitrogen balance in the subacute and chronic phases and to identify the factors related to it. METHODS: Nitrogen balance was calculated from the collected urine of 56 patients with subacute stroke [mean (SD) 53.8 (18.4) days post-stroke] who were admitted for rehabilitation for their first-ever ischaemic or nonsurgical haemorrhagic stroke. In the first experiment, their nitrogen balance was measured during the rehabilitation phase, and factors (type, severity of hemiparesis, activities of daily living, dysphagia and malnutrition status) related to it were evaluated. The second experiment was performed to describe the time course of nitrogen balance in 31 consecutive patients, with assessments made at admission and at discharge. RESULTS: Nitrogen balance was positive in all patients in the subacute phase. A significant difference was seen in nitrogen balance between high and low fat-free mass in male patients. In the chronic phase, nitrogen balance was positive in 96% of the patients. There was no significant difference in nitrogen balance between discharge and admission. CONCLUSIONS: In the subacute and chronic phases of stroke, it was confirmed that hypercatabolism had resolved and that intensive rehabilitation is possible in the convalescent period of stroke.

Observations in the diagnosis of cervical myelopathy in patients suffering from diabetes mellitus.

OBJECTIVES: To identify any observations that could aid in the diagnosis of cervical myelopathy in patients suffering from diabetes mellitus (DM). We compared the preoperative neurological findings in patients with cervical myelopathy among non-diabetics, mild diabetics and severe diabetics. STUDY DESIGN: A retrospective comparative study. SETTING: Department of Orthopaedic Surgery, Wakayama Medical University, Japan. METHODS: We retrospectively reviewed 111 patients who had undergone laminoplasty for cervical compressive myelopathy: 56 without DM and 29 with severe diabetes more than 10 years of medication; more than 7.0% HbA1c; diabetic retinopathy; and delayed conduction velocity of peripheral nerves. For preoperative neurological assessment we compared the following among the three groups: the 10 s test whereby the myelopathy in the hand was quantified; sensory disturbance; deep tendon reflexes; Hoffmann's, Trömner's and Babinski's reflexes; and bladder dysfunction. RESULTS: There was no significant difference preoperatively in the 10 s test between the groups. Deep tendon reflexes were significantly decreased in group S. There were no significant differences in sensory disturbance and bladder dysfunction. Although Hoffmann's and Trömner's reflexes significantly disappeared in group S, there was no significant difference in positivity of Babinski's reflex between the groups. CONCLUSIONS: The 10 s test and Babinski's reflex are helpful for the diagnosis of cervical myelopathy in patients suffering from DM.

Difference in coronary artery intima and media calcification in autopsied patients with chronic kidney disease.

BACKGROUND: In patients with chronic kidney disease (CKD), coronary artery calcification occurs at two distinct sites in the vessel wall: the intima and the media. Arterial media calcification (AMC), a nonocclusive condition, affects hemodynamics differently compared to arterial intima calcification (AIC), which occurs in atherosclerotic plaques. Arterial calcification is considered a cell-regulated process resembling intramembranous bone formation. The purpose of this retrospective observational study was to clarify the morphological differences between AIC and AMC and to evaluate the role of vascular smooth muscle cells (VSMCs) and macrophages in AIC and AMC formation. METHODS: We histologically analyzed 14 tissue specimens from 14 autopsies of patients with CKD Stage 5D who underwent hemodialysis and 5 specimens from 5 patients with CKD Stage 2 - 3 (90 ml/min/1.73 m2 > estimated GFR >= 30 ml/min/1.73 m2). We performed immunohistochemical staining of osteopontin (OPN) as a marker for bone matrix protein, alpha-smooth muscle actin (alphaSMA) for VSMCs, Cbfa1/Runx2 as a marker for osteoblastic differentiation of VSMCs, and CD68 for macrophages. RESULTS: In the CKD 2/3 group, we also found AIC and AMC. OPN and CD68 expression in the CKD 2/3 group was similar to that in the CKD 5D group. Although we did not find Cbfa1/Runx2 positive cell expression in the CKD 2/3 group, we did find it in the CKD 5D group. We found CD68-positive cells predominantly in AIC and absent in AMC in both groups. CONCLUSIONS: These findings suggest that the influence of Cbfa1/Runx2 pathway in coronary artery calcification depends on the CKD Stage. Expression of CD68-positive cells depends on the location of the coronary artery calcification.

Assessment of risk factors for low bone mineral density in Brazilian postmenopausal women.

OBJECTIVE: To assess risk factors associated with low bone mineral density (BMD) in postmenopausal women. METHODS: In this cross-sectional study, a total of 412 Brazilian postmenopausal women, aged 40-75 years, with BMD measured using central dual-energy X-ray absorptiometry, were included. The clinical risk factors assessed were: age, time since menopause, smoking, physical activity, use of hormone therapy (HT) or corticosteroids, personal fracture history, maternal history of fracture, and body mass index (BMI, weight/height(2)). Low BMD was considered when total spine and/or femoral neck T-score values were ≤ -2.0 standard deviations. Logistic regression was used to determine the odds ratio (OR) for low BMD in the presence of the influential variables analyzed. RESULTS: Low BMD, which occurred in 36.6% (151/412) of the participants, was observed in 22.4% of women aged 40-49 years, in 34.2% of those aged 50-59 years, and in 60.5% of those > 60 years (p < 0.001). Similarly, low BMD was observed in 21.9% of women with menopause duration ≤ 5 years, in 39.5% with a duration of 6-10 years, and in 57.7% with menopause duration of > 10 years (p < 0.001). Seventy percent of women with BMI < 20 kg/m(2) were osteopenic/osteoporotic (p < 0.001). The percentage of HT users was 37.4%; 27.7% took regular physical activity and 24.5% were smokers. The risk for low BMD detection increased significantly with age (OR 1.08; 95% confidence interval (CI) 1.02-1.14), time since menopause (OR 1.12; 95% CI 1.04-1.20), smoking (OR 3.43; 95% CI 1.67-6.96), fracture history (OR 2.05; 95% CI 1.11-3.78), and maternal history of fracture (OR 2.16; 95% CI 1.14-4.09). Physical activity, diet, corticotherapy and thyropathies did not influence risk. Contrarily, use of HT (OR 0.38; 95% CI 0.24-0.60) and high BMI (OR 0.89; 95% CI 0.84-0.96) reduced risk (p < 0.05). CONCLUSION: In postmenopausal women, age, time since menopause, smoking, and personal or maternal history of fracture were strong clinical indicators of risk for low BMD, whereas the use of hormone therapy and high BMI were shown to be protective factors.

Studies of endotoxin-induced decrease in lipoprotein lipase activity.

A variety of invasive stimuli have been shown to induce hyperlipidemia due to impaired removal of triglyceride from the circulation. The mechanism by which endotoxin induces a deficiency in the activity of the key enzyme of triglyceride metabolism, lipoprotein lipase (LPL), has been studied. In C3H/HeN (endotoxin-sensitive) mice, LPL activity in adipose tissue was markedly suppressed 16 h after endotoxin administration. In contrast, the endotoxin-resistant C3H/HeJ mice were less sensitive to the suppressive effect of endotoxin on LPL activity. After endotoxin administration, a transferable factor had been detected in the blood of C3H/HeN mice 2 h after the injection of endotoxin that causes a suppression of adipose tissue LPL activity in C3H/HeJ mice as well as in C3H/HeN mice. Conditioned medium from the cultures of peritoneal exudate cells of C3H/HeN mice incubated in endotoxin also suppresses adipose tissue LPL in C3H/HeJ mice. These studies demonstrate that exudate cells produce a humoral factor in response to endotoxin, which suppresses adipose tissue LPL.

Studies of insulin resistance in adipocytes induced by macrophage mediator.

An apparent insulin resistance is noted in 3T3-L1 adipocytes after the addition of an endotoxin-induced mediator from macrophages. Examination at the level of the insulin receptor has revealed that the mediator does not effect either the functional ability of the cells to bind insulin or the ability of insulin to stimulate the uptake of glucose. The resistance appears to reflect a post-receptor interference with the insulin-induced biosynthesis of the anabolic enzymes, acetyl Co-A carboxylase and fatty acid synthetase, which are required for the conversion of glucose into storage lipid. These studies offer a new in vitro model for investigating the molecular basis of insulin resistance.

In vivo overexpression of IL-13 receptor alpha2 chain inhibits tumorigenicity of human breast and pancreatic tumors in immunodeficient mice.

Interleukin 13 receptor alpha2 (IL-13R(alpha)2) chain is highly expressed on some tumor cell lines and primary cell cultures. This receptor chain plays an important role in ligand binding and internalization. To determine the functional significance of overexpression of this chain, we stably transfected IL-13R(alpha)2 chain in human breast (MDA-MB-231) and pancreatic (PANC-1) cancer cell lines that naturally do not express this chain. There was no difference in growth between vector only transfected and IL-13R(alpha)2 chain transfected cells in vitro. However, surprisingly, in immunodeficient mice, tumorigenicity was profoundly inhibited in IL-13R(alpha)2 chain overexpressing tumors. Because breast tumors that grew later showed loss of IL-13R(alpha)2 gene expression, lack of tumorigenicity correlated positively with IL-13R(alpha)2 chain expression. Inflammatory cells including neutrophils and macrophages were identified in IL-13R(alpha)2 overexpressing regressing tumors and neutrophils were found to produce IL-13. IL-13 showed a modest antitumor activity to IL-13R(alpha)2 chain overexpressing tumors in vitro and in vivo. Furthermore, IL-13R(alpha)2 chain overexpressing tumors constitutively produced IL-8 that has been shown to have antitumor effect. These results establish a novel function of a cytokine receptor chain and further suggest that the presence of this chain on tumor cells by itself may play a key role in tumorigenicity.

Cytotoxicity of a perfluorocarbon blood substitute to macrophages in vitro.

Murine macrophage and macrophage-depleted splenocyte cultures were incubated under ambient oxygen with a commercially available perfluorocarbon blood substitute. The perfluorocarbon preparation was found to be selectively cytotoxic to macrophages. This finding may be significant in view of the preliminary therapeutic usage of these preparations. In addition, perfluorocarbons may be useful as a means of selectively removing macrophages from tissue and organ cultures.

Molecular cloning of the Bombyx mori prothoracicotropic hormone.

Prothoracicotropic hormone (PTTH), a brain secretory polypeptide of insects, stimulates the prothoracic glands to produce and release ecdysone, the steroid essential to insect development. The complementary DNAs encoding PTTH of the silkmoth Bombyx mori were cloned and characterized, and the complete amino acid sequence was deduced. The data indicated that PTTH is first synthesized as a 224-amino acid polypeptide precursor containing three proteolytic cleavage signals. The carboxyl-terminal component (109 amino acids) that follows the last cleavage signal represents one PTTH subunit. Two PTTH subunits are linked together by disulfide bonds, before or after cleavage from prepro-PTTH, to form a homodimeric PTTH. When introduced into Escherichia coli cells, the complementary DNA directed the expression of an active substance that was functionally indistinguishable from natural PTTH. In situ hybridization showed the localization of the prepro-PTTH mRNA to two dorsolateral neurosecretory cells of the Bombyx brain.

The critical velocity and 1500-m surface performances in Finswimming.

The purpose of this investigation was to determine whether the concepts of critical velocity (CV) and anaerobic swimming capacity (ASC) could be used by coaches as a reliable index in order to monitor 1500-m Surface (SF) performances in Finswimming. Thirteen Finswimmers (6 males and 7 females, 24+/-6 years), members of the Japanese national team, were instructed to swim three different swimming distances (400-, 800-, and 1500-m) at maximal effort in a 50m long course swimming pool. CV and the ASC were calculated using 400-m and 800-m swim times. Mean height and body mass were 170.2 cm and 69.7 kg in male and 160.5 and 61.0 kg in female. A highly positive correlation was found between the CV and the mean velocity of 1500-m SF (V1500) (r=0.91, P<0.01), but no correlation was found between the ASC and V1500. (r=0.46, P=0.11). However, a high correlation was found between the ASC and the residual error of V1500, calculated from the relationship between V1500 and the CV (r=0.89, P<0.01). These results suggest that the CV is a useful method for evaluating 1500-m SF performance and an aerobic performance expressed as the CV contributes to 1500-m SF performance.

Functional analysis of cytomegalovirus-specific T lymphocytes compared to tetramer assay in patients undergoing hematopoietic stem cell transplantation.

In order to evaluate whether we could predict reactivation of CMV by monitoring the number of CMV-specific cytotoxic T-lymphocytes (CTL), tetramer analysis was performed in 37 patients who underwent hematopoietic stem cell transplantation (HSCT). The results disclosed that the mean number of CMV-specific CTL at day 30 did not differ among patients who developed CMV antigenemia (22/microl) and those who did not (12/microl). Serial tetramer analysis showed that 21% of the patients had >10/microl CMV-specific CTL at the first detection of CMV antigenemia and 67% of the patients had more than 10/microl CMV-specific CTL at the onset of CMV disease. Intracellular staining upon stimulation by CMV lysates and peptide in patients with CMV colitis revealed that both IFN-gamma producing CD4+ and CD8+ lymphocytes were suppressed at the onset of CMV colitis (1.6 and 8/microl), which increased with recovery of the disease (19 and 47/microl). These data suggest that it is difficult to predict CMV reactivation solely by the number of CMV-specific CTL. We suggest that additional functional analysis by intracellular cytokine assay may be useful for immunomonitoring against CMV.

The Wilms' tumor gene WT1-GFP knock-in mouse reveals the dynamic regulation of WT1 expression in normal and leukemic hematopoiesis.

The Wilms' tumor gene WT1 is overexpressed in most of human leukemias regardless of disease subtypes. To characterize the expression pattern of WT1 during normal and neoplastic hematopoiesis, we generated a knock-in reporter green fluorescent protein (GFP) mouse (WT1(GFP/+)) and assayed for WT1 expression in normal and leukemic hematopoietic cells. In normal hematopoietic cells, WT1 was expressed in none of the long-term (LT) hematopoietic stem cells (HSC) and very few (<1%) of the multipotent progenitor cells. In contrast, in murine leukemias induced by acute myeloid leukemia 1 (AML1)/ETO+TEL/PDGFbetaR or BCR/ABL, WT1 was expressed in 40.5 or 38.9% of immature c-kit(+)lin(-)Sca-1(+) (KLS) cells, which contained a subset, but not all, of transplantable leukemic stem cells (LSCs). WT1 expression was minimal in normal fetal liver HSCs and mobilized HSCs, both of which are stimulated for proliferation. In addition, overexpression of WT1 in HSCs did not result in proliferation or expansion of HSCs and their progeny in vivo. Thus, the mechanism by which expansion of WT1-expressing cells occurs in leukemia remains unclear. Nevertheless, our results demonstrate that the WT1(GFP/+) mouse is a powerful tool for analyzing WT1-expressing cells, and they highlight the potential of WT1, as a specific therapeutic target that is expressed in LSCs but not in normal HSCs.

The effects of time of first cleavage, developmental stage, and delipidation of nuclear-transferred porcine blastocysts on survival following vitrification.

The effect of removing cytoplasmic lipid droplets (delipidation) at the 2-cell and developmental stages on the survival of porcine somatic cell nuclear-transferred blastocysts developed from the enucleated oocytes receiving somatic cells from kidney of an adult female after cryopreservation was examined. Vitrification was performed using the Cryoloop method with a small volume of medium (0.5 microl). To select 2-cell embryos with a high potential to develop into blastocysts, the relationship between the timing of the first cleavage and the developmental potential was examined. The potential of nuclear-transferred oocytes to develop into blastocysts in the intermediate-cleavage group (20-24h after activation, 25%) was slightly or significantly (P<0.05) higher than that in fast-cleavage (<20 h after activation, 13%) and slow-cleavage groups (>24h after activation, 5%). Most non-delipidated blastocysts did not survive after thawing (0% for early-stage and 9% for advanced-stage blastocysts), but the survival rate of delipidated blastocysts 48 h after culture (54% and 72%, respectively) was not significantly different from that of non-vitrified blastocysts (80% and 92%, respectively). The survival rate of advanced-stage blastocysts after vitrification was slightly higher than that of early-stage blastocysts. The present study demonstrates that somatic cell nuclear-transferred porcine blastocysts developed from embryos selected at the 2-cell stage can be preserved by vitrification with a small volume of medium if the lipid droplets of the embryos are first removed.

The origin of the premotor potential recorded from the second lumbrical.

PURPOSE: When recording with a palm electrode, a premotor potential (PMP) precedes the compound muscle action potential evoked from the second lumbrical muscle following median nerve stimulation. The origin of the premotor potential has remained uncertain. The aim of this study was to determine whether the PMP-2L is a SNAP derived from antidromically activated digital sensory branches of the median nerve. METHODS: We recorded three active electrodes were placed over the second lumbrical muscle, the third lumbrical muscle, the fourth lumbrical muscle by multi-channel recordings. RESULTS: PMPs are recorded only over the median digital sensory branches after stimulating the median nerve, while they are recorded only over the ulnar branch after stimulating the ulnar nerve. CONCLUSIONS: We conclude that the origin of the PMP is a SNAP arising from antidromically activated digital sensory branches.

Laparoendoscopic Single-site Plus 1-port Donor Nephrectomy: Division of Roles to Shorten Warm Ischemic Time.

BACKGROUND: In this study we present our new surgical procedure, laparoendoscopic single-site surgery plus 1 for donor nephrectomy (LESS+1-DN), which shortens warm ischemic time (WIT) and improves surgical outcomes. METHODS: From January 2013 to February 2017, 15 patients who underwent LESS-DN and 41 patients who underwent LESS+1-DN at our institution were evaluated retrospectively. Patients were divided into 3 groups: group A, 15 cases of LESS-DN; group B, the first 15 patients who underwent LESS+1-DN; and group C, 26 patients who underwent subsequent LESS+1-DN. To reduce WIT, we clearly defined the roles of the surgeon and first assistant in the 26 subsequent LESS+1-DN cases. The surgeon dissected the renal pedicle and harvested the kidney graft using a recovery bag and the first assistant held the recovery bag. RESULTS: The mean operative time in group C (213.7 minutes) was significantly shorter than that in groups A (253.3 minutes) and B (253.8 minutes). The WIT in group C (195.2 seconds) was significantly shorter than that in groups A (389.8 seconds) and B (313.2 seconds). Open conversion was required in 1 case in group A. None of the donors required conversion to open surgery and no perioperative complications occurred in groups B and C. Linear regression analysis of the LESS+1-DN operative times and consecutive case numbers demonstrated a shallow learning curve (R2 = 0.392, P < .05). CONCLUSION: Our new procedure that divides the roles of the operator and the first assistant contributed significantly to a shortening of WIT. Dividing roles can facilitate a safer laparoscopic donor nephrectomy.

Optimisation of antithrombin resistance assay as a practical clinical laboratory test: Development of prothrombin activator using factors Xa/Va and automation of assay.

INTRODUCTION: Antithrombin resistance (ATR) is a novel thrombotic risk in abnormal prothrombins. A manual ATR assay using Oxyuranus scutellatus (Ox) venom as a prothrombin activator was established for detecting antithrombin-resistant prothrombin. However, this assay was limited because of Ox snake venom availability and its throughput capacity. Here, we have improved the ATR assay using bovine factors Xa and Va (FXa/Va) as prothrombin activators and have optimised assay conditions for an automated instrument (ACL TOP 500). METHODS: Diluted plasma was incubated with a prothrombin activator mix (phospholipids, CaCl2 , and bovine FXa/Va), followed by inactivation with antithrombin for 10, 20 and 30 minutes. We added a chromogenic substrate S-2238, and assessed changes in absorbance/min at 405 nm. We also adapted assay conditions for ACL TOP 500. RESULTS: Optimum conditions for FXa/Va treatment were 6.25% phospholipids, 5 mM CaCL2 , 0.01 µg/mL FXa and 0.1 µg/mL FVa. ATR assay kinetics with the FXa/Va activator was comparable with that with the Ox activator in heterozygous reconstituted plasma with the recombinant wild-type or antithrombin-resistant prothrombin. Using ACL TOP 500, optimum conditions for the FXa/Va treatment were 10.0% phospholipids, 5 mM CaCl2 , 0.02 µg/mL FXa and 0.2 µg/mL FVa. The automated ATR assay with the FXa/Va activator demonstrated good detectability for antithrombin-resistant prothrombin in plasma from a heterozygous carrier with prothrombin Yukuhashi or Belgrade. CONCLUSION: We optimised the ATR assay with the FXa/Va activator and adapted the assay for ACL TOP 500; the assay showed the ability to clearly detect antithrombin-resistant prothrombin in manual and automated procedures.

Antiapoptotic function of 17AA(+)WT1 (Wilms' tumor gene) isoforms on the intrinsic apoptosis pathway.

The WT1 gene is overexpressed in human primary leukemia and a wide variety of solid cancers. The WT1 gene is alternatively spliced at two sites, yielding four isoforms: 17AA(+)KTS(+), 17AA(+)KTS(-), 17AA(-)KTS(+), and 17AA(-)KTS(-). Here, we showed that 17AA(+)WT1-specific siRNA induced apoptosis in three WT1-expressing leukemia cell lines (K562, HL-60, and Kasumi-1), but not in WT1-non-expressing lymphoma cell line (Daudi). 17AA(+)WT1-specific siRNA activated caspase-3 and -9 in the intrinsic apoptosis pathway but not caspase-8 in the extrinsic one. On the other hand, 17AA(-)WT1-specific siRNA did not induce apoptosis in the three WT1-expressing cell lines. The apoptosis was associated with activation of proapoptotic Bax, which was activated upstream of the mitochondria. Constitutive expression of 17AA(+)WT1 isoforms inhibited apoptosis of K562 leukemia cells induced by apoptosis-inducing agents, etoposide and doxorubicin, through the protection of mitochondrial membrane damages, and DNA-binding zinc-finger region of 17AA(+)WT1 isoform was essential for the antiapoptotic functions. We further studied the gene(s) whose expression was altered by the expression of 17AA(+)WT1 isoforms and showed that the expression of proapoptotic Bak was decreased by the expression of 17AA(+)KTS(-)WT1 isoform. Taken together, these results indicated that 17AA(+)WT1 isoforms played antiapoptotic roles at some points upstream of the mitochondria in the intrinsic apoptosis pathway.

Development of WT1 peptide cancer vaccine against hematopoietic malignancies and solid cancers.

Wild-type Wilms' tumor gene WT1 is highly expressed not only in hematopoietic malignancies, including leukemia and myelodysplastic syndromes (MDS), but also in various kinds of solid tumors. Human cytotoxic T lymphocytes (CTLs) which could specifically lyse WT1-expressing tumor cells with HLA class I restriction were generated in vitro. We have also demonstrated that mice immunized with the WT1 peptide or WT1 cDNA rejected challenges by WT1-expressing tumor cells and survived with no signs of auto-aggression to normal organs which physiologically expressed WT1 in prophylactic and therapeutic models. Furthermore, we and others detected IgM and IgG WT1 antibodies in the patients with hematopoietic malignancies, indicating that WT1 protein was highly immunogenic, and that immunoglobulin class-switch-inducing WT1-specific cellular immune responses were elicited in the patients. CD8+ WT1-specific CTLs were also detected in peripheral blood or tumor-draining lymph nodes of cancer patients. These results provided us with the rationale for elicitation of CTL responses targeting the WT1 product for cancer immunotherapy. On the basis of the findings mentioned above, we performed a phase I clinical trial of WT1 peptide cancer vaccine for the patients with malignant neoplasms. These results strongly suggested that WT1 peptide cancer vaccine had efficacy in the clinical setting, because clinical responses, including reduction of leukemic blast cells or regression of tumor masses, were observed after the WT1 vaccination in patients with hematopoietic malignancies or solid cancers. The power of TAA-derived cancer vaccine may be enhanced by combination with stronger adjuvants, helper peptide, or conventional treatments such as molecular-target-based drugs.

Th1-biased humoral immune responses against Wilms tumor gene WT1 product in the patients with hematopoietic malignancies.

The Wilms' tumor gene WT1 is highly expressed in leukemias and myelodysplastic syndrome (MDS), and WT1 expression levels increase along with the disease progression in chronic myeloid leukemia and MDS. We previously reported that IgM and IgG WT1 antibodies were detected with significantly higher detection rate and antibody titers in leukemias and MDS compared to those in healthy volunteers. In this study, whether IgG humoral immune responses against WT1 protein were Th1- or Th2-type were determined by measurement of four subclasses of IgG WT1 antibody, IgG1, IgG2, IgG3, and IgG4. In leukemias and MDS, Th1-type WT1 antibodies such as IgG1, IgG2, and IgG3 were significantly increased in both detection rate and antibody titers compared to those in healthy volunteers, whereas Th2-type WT1 antibody such as IgG4 did not increase. These results showed that Th1-biased humoral immune responses against WT1 protein were generated in leukemias and MDS. These results should allow us to consider that Th1-biased cellular immune responses against WT1 protein, which was essentially needed for cancer immunotherapy targeting WT1, should be elicited in patients with hematopoietic malignancies.