Generation of a human induced pluripotent stem cell line (YUCMi020-A) from peripheral blood mononuclear cells derived from a female with the Jr(a-) blood type.

The Jra antigen, the only antigen within the JR blood group system, is a high-prevalence red blood cell (RBC) antigen found in over 99 % of the global population. An induced pluripotent stem cell line (YUCMi020-A) was generated from peripheral blood drawn from a Jr(a-) phenotype individual, who was homozygous for a null mutation of ABCG2*01N.01 (rs72552713, c.376C>T; p.Gln126*). The generated line exhibited pluripotent characteristics and no chromosomal aberrations. This cell line will serve as a cell source, enabling us to produce RBCs with the Jr(a-) phenotype in vitro, which can be used for transfusing individuals with anti-Jra antibodies.

CEBPA double mutations associated with ABO antigen weakness in hematologic diseases.

ABSTRACT: ABO antigen weakness is rarely observed in ABO typing for transfusion. Hematologic diseases and associated gene mutations have been suggested as potential causes of this phenomenon, yet the precise etiology has not been elucidated. Through ABO typing and genetic analysis data conducted over 7 years, we have reconfirmed the association between ABO antigen weakness and hematologic diseases, especially acute myeloid leukemia (odds ratio [OR], 2.55; 95% confidence interval [CI], 1.12-5.83) and myelodysplastic syndrome (OR, 6.94; 95% CI, 2.86-16.83), and discovered previously unidentified candidate genes, CEBPA (OR, 43.70; 95% CI, 18.12-105.40), NRAS (OR, 3.37; 95% CI, 1.46-7.79), U2AF1 (OR, 8.12; 95% CI, 2.86-23.03), and PTPN11 (OR, 4.52; 95% CI, 1.51-13.50), seemingly associated with this phenomenon. Among these, CEBPA double mutations displayed a significant association, with ABO antigen weakness being observed in 20 of the 25 individuals (80.0%) possessing these mutations. From this study, new factors associated with ABO antigen weakness have been identified.

Evaluating the TaqMan Jra-Genotyping Method for Rapidly Predicting the Presence of Anti-Jra Antibodies.

Background: The Jra antigen is a high-prevalence red blood cell (RBC) antigen. Reports on cases of fatal hemolytic disease of the fetus and newborn and acute hemolytic transfusion reactions suggest that antibodies against Jra (anti-Jra) have potential clinical significance. Identifying anti-Jra is challenging owing to a lack of commercially available antisera. We developed an alternative approach to rapidly predict the presence of anti-Jra using the TaqMan single-nucleotide polymorphism (SNP)-genotyping method. Methods: Residual peripheral blood samples from 10 patients suspected of having the anti-Jra were collected. Two samples with confirmed Jr(a-) RBCs and anti-Jra were used to validate the TaqMan genotyping assay by comparing the genotyping results with direct sequencing. The accuracy of the assay in predicting the presence of anti-Jra was verified through crossmatching with in-house Jr(a-) O+ RBCs. Results: The TaqMan-genotyping method was validated with two Jr(a-) RBC- and anti-Jra-confirmed samples that showed concordant Jra genotyping and direct sequencing results. Jra genotyping for the remaining samples and crossmatching the serum samples with in-house Jr(a-) O+ RBCs showed consistent results. Conclusions: We validated a rapid, simple, accurate, and cost-effective method for predicting the presence of anti-Jra using a TaqMan-based SNP-genotyping assay. Implementing this method in routine practice in clinical laboratories will assist in solving difficult problems regarding alloantibodies to high-prevalence RBC antigens and ultimately aid in providing safe and timely transfusions and proper patient care.

OsWRKY114 Is a Player in Rice Immunity against Fusarium fujikuroi.

Every year, invasive pathogens cause significant damage to crops. Thus, identifying genes conferring broad-spectrum resistance to invading pathogens is critical for plant breeding. We previously demonstrated that OsWRKY114 contributes to rice (Oryza sativa L.) immunity against the bacterial pathovar Xanthomonas oryzae pv. oryzae (Xoo). However, it is not known whether OsWRKY114 is involved in defense responses to other pathogens. In this study, we revealed that OsWRKY114 enhances innate immunity in rice against the fungal pathogen Fusarium fujikuroi, which is the causal agent of bakanae disease. Transcript levels of various gibberellin-related genes that are required for plant susceptibility to F. fujikuroi were reduced in rice plants overexpressing OsWRKY114. Analysis of disease symptoms revealed increased innate immunity against F. fujikuroi in OsWRKY114-overexpressing rice plants. Moreover, the expression levels of OsJAZ genes, which encode negative regulators of jasmonic acid signaling that confer immunity against F. fujikuroi, were reduced in OsWRKY114-overexpressing rice plants. These results indicate that OsWRKY114 confers broad-spectrum resistance not only to Xoo but also to F. fujikuroi. Our findings provide a basis for developing strategies to mitigate pathogen attack and improve crop resilience to biotic stress.

OsWRKY114 Negatively Regulates Drought Tolerance by Restricting Stomatal Closure in Rice.

The WRKY family of transcription factors plays a pivotal role in plant responses to biotic and abiotic stress. The WRKY Group III transcription factor OsWRKY114 is a positive regulator of innate immunity against Xanthomonas oryzae pv. oryzae; however, its role in abiotic stress responses is largely unknown. In this study, we showed that the abundant OsWRKY114 transcripts present in transgenic rice plants are reduced under drought conditions. The overexpression of OsWRKY114 significantly increased drought sensitivity in rice, which resulted in a lower survival rate after drought stress. Moreover, we showed that stomatal closure, which is a strategy to save water under drought, is restricted in OsWRKY114-overexpressing plants compared with wild-type plants. The expression levels of PYR/PYL/RCAR genes, such as OsPYL2 and OsPYL10 that confer drought tolerance through stomatal closure, were also markedly lower in the OsWRKY114-overexpressing plants. Taken together, these results suggest that OsWRKY114 negatively regulates plant tolerance to drought stress via inhibition of stomatal closure, which would otherwise prevent water loss in rice.

Human induced pluripotent stem cell line banking for the production of rare blood type erythrocytes.

BACKGROUND: The in vitro production of mature human red blood cells (RBCs) from induced pluripotent stem cells (iPSCs) has been the focus of research to meet the high demand for blood transfusions. However, limitations like high costs and technological requirements restrict the use of RBCs produced by iPSC differentiation to specific circumstances, such as for patients with rare blood types or alloimmunized patients. In this study, we developed a detailed protocol for the generation of iPSC lines derived from peripheral blood of donors with O D-positive blood and rare blood types (D-and Jr(a-)) and subsequent erythroid differentiation. METHODS: Mononuclear cells separated from the peripheral blood of O D-positive and rare blood type donors were cultured to produce and expand erythroid progenitors and reprogrammed into iPSCs. A 31-day serum-free, xeno-free erythroid differentiation protocol was used to generate reticulocytes. The stability of iPSC lines was confirmed with chromosomal analysis and RT-PCR. Morphology and cell counts were determined by microscopy observations and flow cytometry. RESULTS: Cells from all donors were successfully used to generate iPSC lines, which were differentiated into erythroid precursors without any apparent chromosomal mutations. This differentiation protocol resulted in moderate erythrocyte yield per iPSC. CONCLUSIONS: It has previously only been hypothesized that erythroid differentiation from iPSCs could be used to produce RBCs for transfusion to patients with rare blood types or who have been alloimmunized. Our results demonstrate the feasibility of producing autologous iPSC-differentiated RBCs for clinical transfusions in patients without alternative options.

EXPLORING PERSONALITY DIAGNOSIS STABILITY FOLLOWING ACUTE PSYCHOTHERAPY FOR CHRONIC POSTTRAUMATIC STRESS DISORDER.

BACKGROUND: Axis I comorbidity complicates diagnosing axis II personality disorders (PDs). PDs might influence Axis I outcome. No research has examined psychotherapy effects on PDs of treating Axis I comorbidity. Secondary analysis of a randomized controlled trial examined PD diagnostic stability after brief psychotherapy of chronic posttraumatic stress disorder (PTSD). METHODS: Patients with chronic PTSD were randomly assigned to 14 weeks of prolonged exposure, interpersonal psychotherapy, or relaxation therapy. Assessments included the Structured Clinical Interview for DSM-IV, Patient Version (SCID-P) and Structured Clinical Interview for DSM-IV Axis II Disorders (SCID-II) at baseline, week 14, and for treatment responders (≥30% clinician-administered PTSD scale improvement, defined a priori) at week 26 follow-up. We hypothesized patients whose PTSD improved would retain fewer baseline PD diagnoses posttreatment, particularly with personality traits PTSD mimics, e.g. paranoid and avoidant. RESULTS: Forty-seven (47%) of 99 SCID-II patients evaluated at baseline received a SCID-II diagnosis: paranoid (28%), obsessive-compulsive (27%), and avoidant (23%) PDs were most prevalent. Among 78 patients who repeated SCID-II evaluations posttreatment, 45% (N = 35) had baseline PD diagnoses, of which 43% (N = 15/35) lost at week 14. Three (7%) patients without baseline PDs acquired diagnoses at week 14; 10 others shifted diagnoses. Treatment modality and PTSD response were unrelated to PD improvement. Of treatment responders reevaluated at follow-up (N = 44), 56% with any baseline Axis II diagnosis had none at week 26. CONCLUSION: This first evaluation of Axis I psychotherapy effects on personality disorder stability found that acutely treating a chronic state decreased apparent trait-across most PDs observed. These exploratory findings suggest personality diagnoses may have limited prognostic meaning in treating chronic PTSD.

Treating patients who strain the research psychotherapy paradigm.

Clinical trials of psychotherapy require diagnostic homogeneity, which implies a convergence of clinical presentations. Yet research study patients present diversely, and patients who do not fit a treatment paradigm may greatly complicate delivery of the study psychotherapy. The research literature has not addressed this issue. The authors use case illustrations of three psychotherapies--Prolonged Exposure, Relaxation Therapy, and Interpersonal Psychotherapy--from an ongoing psychotherapy outcome trial of posttraumatic stress disorder to describe psychotherapeutic responses to complex, "atypical" patients who strain standard treatment paradigms. Therapists required flexibility, and occasionally deviations from strict protocol, in treating heterodox patients. Such heterogeneity of presentation may have implications for psychotherapy outcome in research trials. Despite lack of discussion in the literature, many trials may face such issues.

Combined prolonged exposure therapy and paroxetine for PTSD related to the World Trade Center attack: a randomized controlled trial.

OBJECTIVE: Selective serotonin reuptake inhibitors (SSRIs) are often recommended in combination with established cognitive-behavioral therapies (CBTs) for posttraumatic stress disorder (PTSD), but combined initial treatment of PTSD has not been studied under controlled conditions. There are also few studies of either SSRIs or CBT in treating PTSD related to terrorism. The authors compared prolonged exposure therapy (a CBT) plus paroxetine (an SSRI) with prolonged exposure plus placebo in the treatment of terrorism-related PTSD. METHOD: Adult survivors of the World Trade Center attack of September 11, 2001, with PTSD were randomly assigned to 10 weeks of treatment with prolonged exposure (10 sessions) plus paroxetine (N=19) or prolonged exposure plus placebo (N=18). After week 10, patients discontinued prolonged exposure and were offered 12 additional weeks of continued randomized treatment. RESULTS: Patients treated with prolonged exposure plus paroxetine experienced significantly greater improvement in PTSD symptoms (incidence rate ratio=0.50, 95% CI=0.30-0.85) and remission status (odds ratio=12.6, 95% CI=1.23-129) during 10 weeks of combined treatment than patients treated with prolonged exposure plus placebo. Response rate and quality of life were also significantly more improved with combined treatment. The subset of patients who continued randomized treatment for 12 additional weeks showed no group differences. CONCLUSIONS: Initial treatment with paroxetine plus prolonged exposure was more efficacious than prolonged exposure plus placebo for PTSD related to the World Trade Center attack. Combined treatment medication and prolonged exposure therapy deserves further study in larger samples with diverse forms of PTSD and over longer follow-up periods.

Longitudinal and Cross-Sectional Genetic Diversity in the Korean Peninsula Based on the P vivax Merozoite Surface Protein Gene.

OBJECTIVES: Vivax malaria has reemerged and become endemic in Korea. Our study aimed to analyze by both longitudinal and cross-sectional genetic diversity of this malaria based on the P vivax Merozoite Surface Protein (PvMSP) gene parasites recently found in the Korean peninsula. METHODS: PvMSP-1 gene sequence analysis from P vivax isolates (n = 835) during the 1996-2010 period were longitudinally analyzed and the isolates from the Korean peninsula through South Korea, the demilitarized zone and North Korea collected in 2008-2010 were enrolled in an overall analysis of MSP-1 gene diversity. RESULTS: New recombinant subtypes and severe multiple-cloneinfection rates were observed in recent vivax parasites. Regional variation was also observed in the study sites. CONCLUSION: This study revealed the great complexity of genetic variation and rapid dissemination of genes in P vivax. It also showed interesting patterns of diversity depending, on the region in the Korean Peninsula. Understanding the parasiteninsula. Under genetic variation may help to analyze trends and assess the extent of endemic malaria in Korea.

Molecular cloning of Plasmodium vivax calcium-dependent protein kinase 4.

A family of calcium-dependent protein kinases (CDPKs) is a unique enzyme which plays crucial roles in intracellular calcium signaling in plants, algae, and protozoa. CDPKs of malaria parasites are known to be key regulators for stage-specific cellular responses to calcium, a widespread secondary messenger that controls the progression of the parasite. In our study, we identified a gene encoding Plasmodium vivax CDPK4 (PvCDPK4) and characterized its molecular property and cellular localization. PvCDPK4 was a typical CDPK which had well-conserved N-terminal kinase domain and C-terminal calmodulin-like structure with 4 EF hand motifs for calcium-binding. The recombinant protein of EF hand domain of PvCDPK4 was expressed in E. coli and a 34 kDa product was obtained. Immunofluorescence assay by confocal laser microscopy revealed that the protein was expressed at the mature schizont of P. vivax. The expression of PvCDPK4-EF in schizont suggests that it may participate in the proliferation or egress process in the life cycle of this parasite.

Protein phosphatase 2Cgamma regulates the level of p21Cip1/WAF1 by Akt signaling.

PP2Cgamma is a splicing factor that dephosphorylates specific substrates required for the formation of the spliceosome. In a previous study, we reported that the degradation of p21(Cip1/WAF1)was affected by PP2Cgamma, causing an accumulation of cells in S phase. Here, we demonstrate that the PP2Cgamma-induced degradation of p21(Cip1/WAF1) is mediated by Akt signaling. In cells expressing PP2Cgamma, Akt1 protein was phosphorylated. When PP2Cgamma expression was knocked down, the phosphorylation of Akt1 was reduced and the level of p21(Cip1/WAF1) protein was increased. Interestingly, the stability of p21(Cip1/WAF1) was highly maintained in Akt1-depleted cells despite the ectopic expression of PP2Cgamma. Taken together, these results suggest that PP2Cgamma is a novel regulator of p21(Cip1/WAF1) protein stability via the Akt signaling pathway.

Prevalence and psychological correlates of complicated grief among bereaved adults 2.5-3.5 years after September 11th attacks.

A Web-based survey of adults who experienced loss during the September 11, 2001, terrorist attacks was conducted to examine the prevalence and correlates of complicated grief (CG) 2.5-3.5 years after the attacks. Forty-three percent of a study group of 704 bereaved adults across the United States screened positive for CG. In multivariate analyses, CG was associated with female gender, loss of a child, death of deceased at the World Trade Center, and live exposure to coverage of the attacks on television. Posttraumatic stress disorder, major depression, anxiety, suicidal ideation, and increase in post-9/11 smoking were common among participants with CG. A majority of the participants with CG reported receiving grief counseling and psychiatric medication after 9/11. Clinical and policy implications are discussed.

The psychology of ongoing threat: relative risk appraisal, the September 11 attacks, and terrorism-related fears.

There are now replicated findings that posttraumatic stress disorder (PTSD) symptoms related to the September 11, 2001, attacks occurred in large numbers of persons who did not fit the traditional definition of exposure to a traumatic event. These data are not explained by traditional epidemiologic "bull's eye" disaster models, which assume the psychological effects are narrowly, geographically circumscribed, or by existing models of PTSD onset. In this article, the authors develop a researchable model to explain these and other terrorism-related phenomena by synthesizing research and concepts from the cognitive science, risk appraisal, traumatic stress, and anxiety disorders literatures. They propose the new term relative risk appraisal to capture the psychological function that is the missing link between the event and subjective response in these and other terrorism-related studies to date. Relative risk appraisal highlights the core notion from cognitive science that human perception is an active, multidimensional process, such that for unpredictable societal threats, proximity to the event is only one of several factors that influence behavioral responses. Addressing distortions in relative risk appraisal effectively could reduce individual and societal vulnerability to a wide range of adverse economic and ethnopolitical consequences to terrorist attacks. The authors present ways in which these concepts and related techniques can be helpful in treating persons with September 11- or terrorism-related distress or psychopathology.

PP2Cgamma-mediated S-phase accumulation induced by the proteasome-dependent degradation of p21(WAF1/CIP1).

Serine/threonine phosphatases such as PP1, PP2A, and PP2B are well known to regulate the transition phase of the cell cycle. However, the function of PP2Cgamma in cell cycle progression is still unclear. In the present study, we report the characterization of PP2Cgamma in mammalian cells during the cell cycle. After release of synchronized cells from thymidine block, over-expression of PP2Cgamma led to accumulation in the S phase. The amount of endogenous p21(WAF1/CIP1) protein was markedly reduced by the expression of PP2Cgamma. The degradation of p21(WAF1/CIP1) induced by PP2Cgamma was mediated in a proteasome-dependent manner. In addition, the phosphatase activity of PP2Cgamma was capable of repressing the level of p21(WAF1/CIP1) protein. Phosphorylation of Rb was also reduced in cells expressing PP2Cgamma. Taken together, these results indicate that PP2Cgamma-induced S phase accumulation may be associated with proteasome-directed p21(WAF1/CIP1) degradation.

Sec13 induces genomic instability in U2OS cells.

Sec13p has been known as an endoplasmic reticulum-Golgi transport protein. Recently, it has also been shown to be required for the formation of septation in the fission yeast Schizosaccharomyces pombe. In the present study, we focused on the role of a human homolog of Saccharomyces cerevisiae SEC13, Sec13 protein during mitosis in U2OS cells. We found that the expression of Sec13 was constant throughout the cell cycle, and localized to the kinetochores at metaphase during mitosis. By using green fluorescent protein technology, we observed that Sec13 is required for evasion of mitotic arrest in response to spindle damage, leading to G1-like phase and apoptotic cell death. In addition, cells expressing exogenous Sec13 showed giant nuclei compared to endogenous ones in the absence of nocodazole. These results demonstrate that Sec13 is involved in the regulation of the metaphase/anaphase transition and may be functionally associated with mitotic machinery to maintain genomic stability during mitosis.

Determination of water content in skin by using a FT near infrared spectrometer.

The water content of skin was determined using a FT near infrared (NIR) spectrometer. NIR diffuse reflectance spectra were collected from hairless mouse, in vitro, and from human inner arm, in vivo. It was found that the variation of NIR absorbance band 1450 nm from OH vibration of water and 1940 nm from the combination involving OH stretching and OH deformation, depending on the absolute water content of separated hairless mouse skin, in vitro, using the FT NIR spectrometer. Partial least squares regression (PLSR) was applied to develop a calibration model. The PLS model showed good correlation. For practical use of the evaluation of human skin moisture, the PLS model for human skin moisture was developed in vivo on the basis of the relative water content of stratum corneum from the conventional capacitance method. The PLS model predicted human skin moisture with a standard errors of prediction (SEP) of 3.98 at 1130-1830 nm range. These studies showed the possibility of a rapid and nondestructive skin moisture measurement using FT NIR spectrometer.

Induction of apoptosis in chicken oviduct cells by C2-ceramide.

The chicken oviduct is a dynamic organ that produces secretory proteins such as ovalbumin and its cells undergo cell proliferation and differentiation. There has been no study of the cellular mechanism involved in cell death in the chicken oviduct. Therefore, this study has focused on the study of apoptosis in primary oviduct cells. Because ceramide is known to activate apoptosis in tumor cells and is produced in the oviduct, we used an exogenous ceramide analog to induce cell death. The viability of ceramide-treated chicken oviduct cells decreased in a dose-dependent manner and apoptotic cells were detected by staining with annexin V. The expression of apoptosis-related genes was assessed by RT-PCR and bcl-2 mRNA was found to decrease after exposure to ceramide while Bcl-x mRNA increased 12 h post-treatment. In addition, caspase-3 was expressed strongly in the early stages of apoptosis, while caspase-1 and -9 transcripts increased at later times. We conclude that ceramide induces apoptosis in oviduct-derived primary cells via a caspase- and bcl-2- dependent pathway.