Translating, adapting, and validating the medical student version of the patient care ownership scale for use in Japan.

BACKGROUND: Patient care ownership (PCO) among medical students is a growing area in the field of medical education. While PCO has received increasing attention, there are no instruments to assess PCO in the context of Japanese undergraduate medical education. This study aimed to translate, culturally adapt, and validate the PCO Scale - Medical students (PCOS-S) in the Japanese context. METHODS: We collected survey data from fifth- and sixth-grade medical students from five different universities varying in location and type. Structural validity, convergent validity, and internal consistency reliability were examined. RESULTS: Data from 122 respondents were analyzed. Factor analysis of the Japanese PCOS-S revealed three factors with Cronbach's alpha values exceeding the satisfactory criterion (0.70). A positive correlation was observed between the total Japanese PCOS-S scores and the global rating scores for the clinical department as a learning environment (Pearson's correlation coefficient = 0.61). CONCLUSIONS: We conducted the translation of the PCOS-S into Japanese and assessed its psychometric properties. The Japanese version has good reliability and validity. This instrument has potential value in assessing the development of medical students' PCO.

Translation, adaptation, and validation of the Tolerance of Ambiguity in Medical Students and Doctors (TAMSAD) scale for use in Japan.

BACKGROUND: Ambiguity is inherent to the medical field; hence, assessing and educating medical trainees regarding ambiguity tolerance is essential. The Tolerance of Ambiguity in Medical Students and Doctors (TAMSAD) scale-a novel instrument that assesses ambiguity tolerance in clinical settings-has been widely used for medical education research in Western countries. However, a version of this scale applicable to the intricate clinical contexts of Japan has not yet been developed. In this study, we developed the Japanese version of the TAMSAD (J-TAMSAD) scale and tested its psychometric properties. METHODS: In this multicenter study, we collected data through a cross-sectional survey in two universities (medical students) and ten hospitals (residents) across Japan, and evaluated the structural validity, criterion-related validity, and internal consistency reliability of the J-TAMSAD scale. RESULTS: We analyzed the data of 247 participants. The sample was randomly divided in half, with exploratory factor analysis (EFA) performed on one half and confirmatory factor analysis (CFA) on the other. EFA led to an 18-item J-TAMSAD scale comprising five factors. CFA showed acceptable fit for this five-factor model (comparative fit index = 0.900, root mean square error of approximation = 0.050, standardized root mean square residual = 0.069, goodness of fit index = 0.987). There was a positive correlation between the J-TAMSAD scale scores and total reverse scores on the Japanese version of the Short Intolerance of Uncertainty Scale (Pearson correlation coefficient 0.41). The internal consistency was found to be satisfactory (Cronbach's alpha 0.70). CONCLUSIONS: The J-TAMSAD scale was developed, and its psychometric properties were confirmed. The instrument can be useful for assessing tolerance of ambiguity among medical trainees in Japan. With further validation, it could be used to verify the educational effectiveness of curricula that foster ambiguity tolerance in medical trainees, or even in research assessing the relationship with other variables.

Translating and validating a Japanese version of the instrument for patient assessment of medical professionalism (J-IPAMP): a cross-sectional survey.

BACKGROUND: Although there are many tools to assess medical professionalism, they rarely address patients' perspectives. The instrument for patient assessment of medical professionalism (IPAMP) comprises 11 items and has been established and validated as a valuable tool for assessing trainees' professionalism from the patient's perspective. However, there is no instrument to assess professionalism from the patient's perspective in Japan. The purpose of the present study was to develop a Japanese version of the IPAMP (J-IPAMP) and test its validity and reliability. METHODS: We conducted a cross-sectional survey to examine the reliability and validity of the J-IPAMP in two hospitals (one each in an urban and rural area) in Japan. Receptionists or surveyors distributed the anonymous questionnaire to 276 inpatients; all participants were aged above 20 years and assigned to medical trainees. We evaluated its structural and criterion-related validity, as well as internal consistency reliability. RESULTS: Data of 235 (85.1%) patients were analyzed. Using the split-half validation technique, we performed an exploratory factor analysis (EFA) along with a confirmatory factor analysis (CFA). The EFA showed a one-factor solution. Then, to compare the model fitness between two models (the two-factor model from the original English version vs. unidimensional model suggested by the EFA), the CFA was performed. The CFA showed that almost all of the fit indices met their respective criteria and were approximately the same for the two models. Thus, we adopted a single-factor model. The Pearson correlation coefficients between the total J-IPAMP scores and the global ratings were 0.738, indicating adequate criterion-related validity. The Cronbach's alpha of the 11 items of the instrument was 0.96 (95% confidence interval: 0.96-0.97) and the omega value was 0.96, demonstrating acceptable internal consistency reliability. CONCLUSIONS: We developed the Japanese version of the IPAMP. Its validity and reliability were verified through analysis. This instrument can be utilized for professionalism education in the postgraduate training setting.

Translating and validating a Japanese version of the Patient Care Ownership Scale: a multicenter cross-sectional study.

BACKGROUND: Patient care ownership (PCO) is an essential component in medical professionalism and is crucial for delivering high-quality care. The 15-item PCO Scale (PCOS) is a validated questionnaire for quantifying PCO in residents; however, no corresponding tool for assessing PCO in Japan exists. This study aimed to develop a Japanese version of the PCOS (J-PCOS) and validate it among Japanese medical trainees. METHODS: We performed a multicenter cross-sectional survey to test the validity and reliability of the J-PCOS. The study sample was trainees of postgraduate years 1-5 in Japan. The participants completed the J-PCOS questionnaire. Construct validity was assessed through exploratory and confirmatory factor analyses. Internal consistency reliability was examined by calculating Cronbach's alpha coefficients and inter-item correlations. RESULTS: During the survey period, 437 trainees at 48 hospitals completed the questionnaire. Exploratory factor analysis of the J-PCOS extracted four factors: assertiveness, sense of ownership, diligence, and being the "go-to" person. The second factor had not been identified in the original PCOS, which may be related to a unique cultural feature of Japan, namely, a historical code of personal conduct. Confirmatory factor analysis supported this four-factor model, revealing good model fit indices. The analysis results of Cronbach's alpha coefficients and inter-item correlations indicated adequate internal consistency reliability. CONCLUSIONS: We developed the J-PCOS and examined its validity and reliability. This tool can be used in studies on postgraduate medical education. Further studies should confirm its robustness and usefulness for improving PCO.

A subset of virus-specific CD161+ T cells selectively express the multidrug transporter MDR1 and are resistant to chemotherapy in AML.

The establishment of long-lived pathogen-specific T cells is a fundamental property of the adaptive immune response. However, the mechanisms underlying long-term persistence of antigen-specific CD4+ T cells are not well-defined. Here we identify a subset of memory CD4+ T cells capable of effluxing cellular toxins, including rhodamine (Rho), through the multidrug efflux protein MDR1 (also known as P-glycoprotein and ABCB1). Drug-effluxing CD4+ T cells were characterized as CD161+CD95+CD45RA-CD127hiCD28+CD25int cells with a distinct chemokine profile and a Th1-polarized pro-inflammatory phenotype. CD4+CD161+Rho-effluxing T cells proliferated vigorously in response to stimulation with anti-CD3/CD28 beads and gave rise to CD161- progeny in vitro. These cells were also capable of self-renewal and maintained their phenotypic and functional characteristics when cultured with homeostatic cytokines. Multidrug-effluxing CD4+CD161+ T cells were enriched within the viral-specific Th1 repertoire of healthy donors and patients with acute myeloid leukemia (AML) and survived exposure to daunorubicin chemotherapy in vitro. Multidrug-effluxing CD4+CD161+ T cells also resisted chemotherapy-induced cytotoxicity in vivo and underwent significant expansion in AML patients rendered lymphopenic after chemotherapy, contributing to the repopulation of anti-CMV immunity. Finally, after influenza vaccination, the proportion of influenza-specific CD4+ T cells coexpressing CD161 was significantly higher after 2 years compared with 4 weeks after immunization, suggesting CD161 is a marker for long-lived antigen-specific memory T cells. These findings suggest that CD4+CD161+ T cells with rapid efflux capacity contribute to the maintenance of viral-specific memory T cells. These data provide novel insights into mechanisms that preserve antiviral immunity in patients undergoing chemotherapy and have implications for the development of novel immunotherapeutic approaches.

IL-10+ regulatory B cells are enriched in cord blood and may protect against cGVHD after cord blood transplantation.

Cord blood (CB) offers a number of advantages over other sources of hematopoietic stem cells, including a lower rate of chronic graft-versus-host disease (cGVHD) in the presence of increased HLA disparity. Recent research in experimental models of autoimmunity and in patients with autoimmune or alloimmune disorders has identified a functional group of interleukin-10 (IL-10)-producing regulatory B cells (Bregs) that negatively regulate T-cell immune responses. At present, however, there is no consensus on the phenotypic signature of Bregs, and their prevalence and functional characteristics in CB remain unclear. Here, we demonstrate that CB contains an abundance of B cells with immunoregulatory function. Bregs were identified in both the naive and transitional B-cell compartments and suppressed T-cell proliferation and effector function through IL-10 production as well as cell-to-cell contact involving CTLA-4. We further show that the suppressive capacity of CB-derived Bregs can be potentiated through CD40L signaling, suggesting that inflammatory environments may induce their function. Finally, there was robust recovery of IL-10-producing Bregs in patients after CB transplantation, to higher frequencies and absolute numbers than seen in the peripheral blood of healthy donors or in patients before transplant. The reconstituting Bregs showed strong in vitro suppressive activity against allogeneic CD4(+) T cells, but were deficient in patients with cGVHD. Together, these findings identify a rich source of Bregs and suggest a protective role for CB-derived Bregs against cGVHD development in CB recipients. This advance could propel the development of Breg-based strategies to prevent or ameliorate this posttransplant complication.

Specific combinations of donor and recipient KIR-HLA genotypes predict for large differences in outcome after cord blood transplantation.

The ability of cord blood transplantation (CBT) to prevent relapse depends partly on donor natural killer (NK) cell alloreactivity. NK effector function depends on specific killer-cell immunoglobulin-like receptors (KIR) and HLA interactions. Thus, it is important to identify optimal combinations of KIR-HLA genotypes in donors and recipients that could improve CBT outcome. We studied clinical data, KIR and HLA genotypes, and NK-cell reconstitution in CBT patients (n = 110). Results were validated in an independent cohort (n = 94). HLA-KIR genotyping of recipient germline and transplanted cord blood (CB) grafts predicted for large differences in outcome. Patients homozygous for HLA-C2 group alleles had higher 1-year relapse rate and worse survival after CBT than did HLA-C1/C1 or HLA-C1/C2 (HLA-C1/x) patients: 67.8% vs 26.0% and 15.0% vs 52.9%, respectively. This inferior outcome was associated with delayed posttransplant recovery of NK cells expressing the HLA-C2-specific KIR2DL1/S1 receptors. HLA-C1/x patients receiving a CB graft with the combined HLA-C1-KIR2DL2/L3/S2 genotype had lower 1-year relapse rate (6.7% vs 40.1%) and superior survival (74.2% vs 41.3%) compared with recipients of grafts lacking KIR2DS2 or HLA-C1 HLA-C2/C2 patients had lower relapse rate (44.7% vs 93.4%) and better survival (30.1% vs 0%) if they received a graft with the combined HLA-C2-KIR2DL1/S1 genotype. Relapsed/refractory disease at CBT, recipient HLA-C2/C2 genotype, and donor HLA-KIR genotype were independent predictors of outcome. Thus, we propose the inclusion of KIR genotyping in graft selection criteria for CBT. HLA-C1/x patients should receive an HLA-C1-KIR2DL2/L3/S2 CB graft, while HLA-C2/C2 patients may benefit from an HLA-C2-KIR2DL1/S1 graft.

CD4+ virtual memory: Antigen-inexperienced T cells reside in the naïve, regulatory, and memory T cell compartments at similar frequencies, implications for autoimmunity.

It is widely accepted that central and effector memory CD4+ T cells originate from naïve T cells after they have encountered their cognate antigen in the setting of appropriate co-stimulation. However, if this were true the diversity of T cell receptor (TCR) sequences within the naïve T cell compartment should be far greater than that of the memory T cell compartment, which is not supported by TCR sequencing data. Here we demonstrate that aged mice with far fewer naïve T cells, respond to the model antigen, hen eggwhite lysozyme (HEL), by utilizing the same TCR sequence as their younger counterparts. CD4+ T cell repertoire analysis of highly purified T cell populations from naive animals revealed that the HEL-specific clones displayed effector and central "memory" cell surface phenotypes even prior to having encountered their cognate antigen. Furthermore, HEL-inexperienced CD4+ T cells were found to reside within the naïve, regulatory, central memory, and effector memory T cell populations at similar frequencies and the majority of the CD4+ T cells within the regulatory and memory populations were unexpanded. These findings support a new paradigm for CD4+ T cell maturation in which a specific clone can undergo a differentiation process to exhibit a "memory" or regulatory phenotype without having undergone a clonal expansion event. It also demonstrates that a foreign-specific T cell is just as likely to reside within the regulatory T cell compartment as it would the naïve compartment, arguing against the specificity of the regulatory T cell compartment being skewed towards self-reactive T cell clones. Finally, we demonstrate that the same set of foreign and autoreactive CD4+ T cell clones are repetitively generated throughout adulthood. The latter observation argues against T cell-depleting strategies or autologous stem cell transplantation as therapies for autoimmunity-as the immune system has the ability to regenerate pathogenic clones.

Enforced fucosylation of cord blood hematopoietic cells accelerates neutrophil and platelet engraftment after transplantation.

Delayed engraftment is a major limitation of cord blood transplantation (CBT), due in part to a defect in the cord blood (CB) cells' ability to home to the bone marrow. Because this defect appears related to low levels of fucosylation of cell surface molecules that are responsible for binding to P- and E-selectins constitutively expressed by the marrow microvasculature, and thus for marrow homing, we conducted a first-in-humans clinical trial to correct this deficiency. Patients with high-risk hematologic malignancies received myeloablative therapy followed by transplantation with 2 CB units, one of which was treated ex vivo for 30 minutes with the enzyme fucosyltransferase-VI and guanosine diphosphate fucose to enhance the interaction of CD34(+) stem and early progenitor cells with microvessels. The results of enforced fucosylation for 22 patients enrolled in the trial were then compared with those for 31 historical controls who had undergone double unmanipulated CBT. The median time to neutrophil engraftment was 17 days (range, 12-34 days) compared with 26 days (range, 11-48 days) for controls (P = .0023). Platelet engraftment was also improved: median was 35 days (range, 18-100 days) compared with 45 days (range, 27-120 days) for controls (P = .0520). These findings support ex vivo fucosylation of multipotent CD34(+) CB cells as a clinically feasible means to improve engraftment efficiency in the double CBT setting. The trial is registered to www.clinicaltrials.gov as #NCT01471067.

A robust, good manufacturing practice-compliant, clinical-scale procedure to generate regulatory T cells from patients with amyotrophic lateral sclerosis for adoptive cell therapy.

Regulatory T cells (Tregs) play a fundamental role in the maintenance of self-tolerance and immune homeostasis. Defects in Treg function and/or frequencies have been reported in multiple disease models. Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder affecting upper and lower motor neurons. Compelling evidence supports a neuroprotective role for Tregs in this disease. Indeed, rapid progression in ALS patients is associated with decreased FoxP3 expression and Treg frequencies. Thus, we propose that strategies to restore Treg number and function may slow disease progression in ALS. In this study, we developed a robust, Good Manufacturing Practice (GMP)-compliant procedure to enrich and expand Tregs from ALS patients. Tregs isolated from these patients were phenotypically similar to those from healthy individuals but were impaired in their ability to suppress T-cell effector function. In vitro expansion of Tregs for 4 weeks in the presence of GMP-grade anti-CD3/CD28 beads, interleukin (IL)-2 and rapamcyin resulted in a 25- to 200-fold increase in their number and restored their immunoregulatory activity. Collectively, our data facilitate and support the implementation of clinical trials of adoptive therapy with ex vivo expanded and highly suppressive Tregs in patients with ALS.

Randomized phase II trial of extracorporeal phototherapy and steroids vs. steroids alone for newly diagnosed acute GVHD.

Steroids remain the initial therapy for acute graft-vs.-host disease (AGVHD). Strategies to improve response and minimize steroid exposure are needed. We report results of a randomized, adaptive, Bayesian-designed, phase II trial of prednisone with or without extracorporeal photopheresis (ECP) as an initial therapy for patients with newly diagnosed AGVHD. The primary endpoint was success at day 56 defined as: alive, in remission, achieving AGVHD response without additional therapy, and on <1 mg/kg at day 28 and <0.5 mg/kg on day 56 of steroids. Eighty-one patients were randomized to the ECP arm (n = 51) or steroids alone (n = 30). Median age was 54 years (range: 17-75); 90% had grade II AGVHD and 10% had grades III and IV AGVHD, with skin (85%), upper (22%)/lower (22%) gastrointestinal, and liver (10%) involvement. The ECP arm had a higher probability of success (0.815) and exceeded the predefined threshold for determining the investigational arm promising. ECP was potentially more beneficial than steroids-alone in skin-only AGVHD (response rate: 72% vs. 57%, respectively) than for visceral-organ AGVHD (47% vs. 43%, respectively). The addition of ECP to steroids may result in higher GVHD response as initial therapy for AGVHD, especially for patients with skin-only involvement.

Rapid induction of OX40 ligand on primary T cells activated under DNA-damaging conditions.

We have previously demonstrated that normal human T cells either long-term repeatedly stimulated or freshly activated in vitro in the presence of TGF-beta express the cell surface T-cell costimulating molecule OX40 ligand (OX40L). To further elucidate the kinetics of OX40L expression by human T cells, we have examined whether cell proliferation was required for the expression of OX40L. Thus, normal fresh peripheral blood mononuclear cells were stimulated with immobilized anti-CD3 antibody in the presence of the DNA synthesis-blocking agents such as mitomycin C, 5-fluorouracil, or X-ray irradiation. Flow cytometric analyses demonstrated that a significant frequency of these DNA-damaged activated primary CD4+ and CD8+ T cells became OX40L+ as early as 1 hour after treatment. The OX40L induction on the DNA-damaged activated T cells was inhibited by treatment with either RNA or protein synthesis inhibitors, actinomycin D, or cycloheximide, respectively. Induced OX40L on T cells was functional because it bound recombinant OX40. These data indicate that human primary T cells are programmed to rapidly express functional OX40L molecules after stimulation under DNA-damaging conditions, demonstrating that the induction of OX40L by T cells is independent of cell proliferation. The clinical implications of these new findings are discussed.

Generation of mature dendritic cells with unique phenotype and function by in vitro short-term culture of human monocytes in the presence of interleukin-4 and interferon-beta.

Dendritic cell (DC)-based immunotherapy has been utilized for the treatment of not only a number of human malignancies but also a select group of infectious diseases. Conventional techniques for the generation and maturation of DCs require 7 days of in vitro culture, which prompted us to seek alternative methods that would hasten the generation of functional human myeloid DCs in vitro. Following the use of a number of cytokines/growth factors, we found that in vitro culture of purified human monocytes, in media containing interleukin (IL)-4, together with interferon (IFN)-beta for 24 hrs, followed by the addition of non-specific antigenic stimuli, such as keyhole limpet hemocyanin (KLH), lipopolysaccharide (LPS), or inactivated human immunodeficiency virus (HIV)-1 induced the monocytes to differentiated by 3 days into mature DCs (4B-DCs). These 4B-DCs expressed high levels of CD83 and CD11c, as well as markers of immune activation, including CD80 and CD86, human leukocyte antigen (HLA) class I and II, and CD14, but not CD1a. Anti-CD14 blocking antibody interfered with generation of 4B-DCs by LPS, but not by KLH or HIV-1. Interestingly, 4B-DCs, but not conventional DCs generated using macrophage-colony stimulating factor and IL-4 (G4-DCs), expressed OX40 and OX40L. 4B-DCs showed phagocytic activity, and spontaneously produced IL-12 and tumor necrosis factor (TNF)-alpha, but not IL-10. 4B-DCs promoted proliferation of allogeneic naïve CD4(+) T cells, producing IFN-(lambda) at lower levels than those stimulated with G4-DCs. 4B-DCs were more potent stimulators of allogeneic bulk CD8(+) T cells producing IFN-(lambda) than G4-DCs. These data indicate that 4B-DCs are unique and may provide a relatively more rapid alternative tool for potential clinical use, as compared with conventional G4-DCs.

Requirements for the functional expression of OX40 ligand on human activated CD4+ and CD8+ T cells.

Interaction between OX40 expressed on activated T cells and its ligand (OX40L) on antigen presenting cells (APC) provides a co-stimulatory signal for T cells to promote acquired immunity. In the present study, we have examined various culture conditions for optimum OX40L expression on T cells stimulated with immobilized anti-CD3/CD28 monoclonal antibodies (mAbs). Although the day 3 primed T cells expressed minimal OX40L, after repeated stimulations both the CD4+ and CD8+ T cells became OX40L positive as determined by flow cytometry. Interleukin (IL)-12 interfered with the OX40L expression. Among activated T cells, a higher frequency of CD8+ T cells expressed OX40L than CD4+ T cells. By blocking OX40L-OX40 interaction by an anti-OX40 mAb, the number of OX40L+ T cells significantly increased. Screening of various cytokines showed that transforming growth factor (TGF)-beta1 was capable of induction of OX40L on the activated T cells within 3 days. The OX40L expressed on T cells was functional, as they bound soluble OX40 and stimulated human immunodeficiency virus-1 (HIV-1) production from cell lines chronically infected with HIV-1 and expressing OX40. Altogether the present study findings indicate that functional OX40L is inducible on human activated CD4+ and CD8+ T cells, and that the expression is enhanced by TGF-beta1.

Evidence for B Cell Exhaustion in Chronic Graft-versus-Host Disease.

Chronic graft-versus-host disease (cGvHD) remains a major complication of allogeneic hematopoietic stem cell transplantation (HSCT). A number of studies support a role for B cells in the pathogenesis of cGvHD. In this study, we report the presence of an expanded population of CD19+CD21- B cells with features of exhaustion in the peripheral blood of patients with cGvHD. CD21- B cells were significantly increased in patients with active cGvHD compared to patients without cGvHD and healthy controls (median 12.2 versus 2.12 versus 3%, respectively; p < 0.01). Compared with naïve (CD27-CD21+) and classical memory (CD27+CD21+) B cells, CD19+CD21- B cells in cGvHD were CD10 negative, CD27 negative and CD20hi, and exhibited features of exhaustion, including increased expression of multiple inhibitory receptors such as FCRL4, CD22, CD85J, and altered expression of chemokine and adhesion molecules such as CD11c, CXCR3, CCR7, and CD62L. Moreover, CD21- B cells in cGvHD patients were functionally exhausted and displayed poor proliferative response and calcium mobilization in response to B-cell receptor triggering and CD40 ligation. Finally, the frequencies of circulating CD21- B cells correlated with cGvHD severity in patients after HSCT. Our study further characterizes B cells in chronic cGVHD and supports the use of CD21-CD27-CD10- B cell frequencies as a biomarker of disease severity.

The CXCR4-STAT3-IL-10 Pathway Controls the Immunoregulatory Function of Chronic Lymphocytic Leukemia and Is Modulated by Lenalidomide.

Chronic lymphocytic leukemia (CLL) cells possess regulatory functions comparable to those of normal B10 cells, a regulatory B cell subset that suppresses effector T-cell function through STAT3-mediated IL-10 production. However, the mechanisms governing IL-10 production by CLL cells are not fully understood. Here, we show that the CXC chemokine ligand 12 (CXCL12)-CXCR4-STAT3 axis regulates IL-10 production by CLL cells and their ability to suppress T-cell effector function through an IL-10 mediated mechanism. Knockdown of STAT3 significantly impaired the ability of CLL cells to produce IL-10. Furthermore, experiments to assess the role of lenalidomide, an immunomodulatory agent with direct antitumor effect as well as pleiotropic activity on the immune system, showed that this agent prevents a CXCL12-induced increase in p-S727-STAT3 and the IL-10 response by CLL cells. Lenalidomide also suppressed IL-10-induced Y705-STAT3 phosphorylation in healthy T cells, thus reversing CLL-induced T-cell dysfunction. We conclude that the capacity of CLL cells to produce IL-10 is mediated by the CXCL12-CXCR4-STAT3 pathway and likely contributes to immunodeficiency in patients. Lenalidomide appears to be able to reverse CLL-induced immunosuppression through including abrogation of the CXCL12-CXCR4-S727-STAT3-mediated IL-10 response by CLL cells and prevention of IL-10-induced phosphorylation of Y705-STAT3 in T cells.