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1.  A Method for Simultaneous Determination of 25-Hydroxyvitamin D3 and Its 3-Sulfate in Newborn Plasma by LC/ESI-MS/MS after Derivatization with a Proton-Affinitive Cookson-Type Reagent 
Mass Spectrometry  2016;5(Spec Iss):S0051.
A method for the simultaneous determination of 25-hydroxyvitamin D3 [25(OH)D3] and its 3-sulfate [25(OH)D3S] in newborn plasma, which is expected to be helpful in the assessment of the vitamin D status, using stable isotope-dilution liquid chromatography/electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) has been developed and validated. The plasma was pretreated based on the deproteinization and solid-phase extraction, then subjected to derivatization with 4-(4-dimethylaminophenyl)-1,2,4-triazoline-3,5-dione (DAPTAD). The derivatization enabled the accurate quantification of 25(OH)D3 without interference from 3-epi-25(OH)D3 and also facilitated the simultaneous determination of the two metabolites by LC/positive ESI-MS/MS. Quantification was based on the selected reaction monitoring with the characteristic fragmentation of the DAPTAD-derivatives during MS/MS. This method was reproducible (intra- and inter-assay relative standard deviations of 7.8% or lower for both metabolites) and accurate (analytical recovery, 95.4–105.6%). The limits of quantification were 1.0 ng/mL and 2.5 ng/mL for 25(OH)D3 and 25(OH)D3S, respectively, when using a 20-μL sample. The developed method was applied to the simultaneous determination of plasma 25(OH)D3 and 25(OH)D3S in newborns; it was recognized that the plasma concentration of 25(OH)D3S is significantly higher than that of 25(OH)D3, and preterm newborns have lower plasma 25(OH)D3S concentrations than full-term newborns.
PMCID: PMC5023818  PMID: 27656337
25-hydroxyvitamin D3; 25-hydroxyvitamin D3 3-sulfate; LC/ESI-MS/MS; newborn plasma; derivatization
2.  AB139. Web promotion of da Vinci robotic prostatectomy exhibits varying sexual health information 
Translational Andrology and Urology  2015;4(Suppl 1):AB139.
The surgical robot has been widely adopted in Japan in spite of its high cost and controversy surroundings its benefit. Accordingly, robotic prostatectomy has become the most common treatment options for prostate cancer. The evidence isn’t strong enough to determine whether or not a robot is better than traditional minimally invasive surgery, but the evidence indicates that it is better compared to open surgery. The da Vinci surgical system continues to be the shiny new toy that hospitals boast about on websites. Clinical experience shows that patients are often given unrealistic expectations, especially about the long-term outcomes for erectile function, as well as the time to recovery of erectile function. Sexual dysfunctions other than ED, e.g., orgasmic pain, reduced orgasmic intensity, orgasmic-associated incontinence, penile volume changes with the development of Peyronie’s disease are rarely mentioned to a patient before radical prostatectomy. Hospitals advertise their da Vinci machines in part as a response to perceived consumer demand. The purpose of this analysis was to survey websites for robotic prostatectomy to evaluate the quality of the information found there as it pertains to the outcome for ED.
We identified 168 centers in Japan that were using robotic prostatectomy. Their websites were reviewed for the following factors: Has information between copied directly from the Intuitive Surgical website? Is ED mentioned as a complication of robotic prostatectomy? Is it suggested that the robotic prostatectomy approach is better than the other technique? Are there references to support the values mentioned? Did the sites give realistic expectations about time to recovery and overall recovery? Are the ED rates cited within the published rates? Are the ED rates cited specific to the individual site? Did the site mention ED treatment options? Did the site mention sexual dysfunction other than ED?
Of the 168 centers websites reviewed, 63 (38%) were academic, while 105 (62%) were community-based. On the websites, 59% of the hospitals only posted robotic prostatectomy as a treatment option for prostate cancer. Almost half (55%) of the centers websites suggested that functional outcome were better for robotic prostatectomy than for the other approach; this compared to rates being 58.7% for academic and 53.3% for community-based (P=0.525). ED was mentioned by only 15%; 28.6% of academic centers mentioned ED compared with 7.6% of community-based centers (P<0.05). Realistic expectations about time to recovery for erectile function and overall recovery were mentioned by only 4%. Only 2% offered the ED rates cited specific to the individual site.
To make informed decisions about their medical care, patients need unbiased, evidence-based information about the benefits and risks of different treatment options. The Internet is a major source of information for prostate cancer patients. However, our study of how hospitals talk about robotic surgery found that many copied directly from Intuitive’s marketing materials. Many centers claimed benefits that were unsupported by evidence and only a minority pointed to potential risks. A balanced presentation of outcomes expected after robotic prostatectomy is necessary to allow patients to make informed treatment decisions and to help them set realistic expectations, which will improve their satisfaction and minimize any regret.
PMCID: PMC4708819
Sexual health information; prostatectomy; web promotion
3.  The radish genome and comprehensive gene expression profile of tuberous root formation and development 
Scientific Reports  2015;5:10835.
Understanding the processes that regulate plant sink formation and development at the molecular level will contribute to the areas of crop breeding, food production and plant evolutionary studies. We report the annotation and analysis of the draft genome sequence of the radish Raphanus sativus var. hortensis (long and thick root radish) and transcriptome analysis during root development. Based on the hybrid assembly approach of next-generation sequencing, a total of 383 Mb (N50 scaffold: 138.17 kb) of sequences of the radish genome was constructed containing 54,357 genes. Syntenic and phylogenetic analyses indicated that divergence between Raphanus and Brassica coincide with the time of whole genome triplication (WGT), suggesting that WGT triggered diversification of Brassiceae crop plants. Further transcriptome analysis showed that the gene functions and pathways related to carbohydrate metabolism were prominently activated in thickening roots, particularly in cell proliferating tissues. Notably, the expression levels of sucrose synthase 1 (SUS1) were correlated with root thickening rates. We also identified the genes involved in pungency synthesis and their transcription factors.
PMCID: PMC4650646  PMID: 26056784
4.  External validation of risk classification in patients with docetaxel-treated castration-resistant prostate cancer 
BMC Urology  2014;14:31.
Castration-resistant prostate cancer (CRPC) patients have poor prognoses, and docetaxel (DTX) is among the few treatment options. An accurate risk classification to identify CRPC patient groups for which DTX would be effective is urgently warranted. The Armstrong risk classification (ARC), which classifies CRPC patients into 3 groups, is superior; however, its usefulness remains unclear, and further external validation is required before clinical use. This study aimed to examine the clinical significance of the ARC through external validation in DTX-treated Japanese CRPC patients.
CRPC patients who received 2 or more DTX cycles were selected for this study. Patients were classified into good-, intermediate-, and poor-risk groups according to the ARC. Prostate-specific antigen (PSA) responses and overall survival (OS) were calculated and compared between the risk groups. A multivariate analysis was performed to clarify the relationship between the ARC and major patient characteristics.
Seventy-eight CRPC patients met the inclusion criteria. Median PSA levels at DTX initiation was 20 ng/mL. Good-, intermediate-, and poor-risk groups comprised 51 (65%), 17 (22%), and 10 (13%) patients, respectively. PSA response rates ≥30% and ≥50% were 33%, 41%, and 30%, and 18%, 41%, and 20% in the good-, intermediate-, and poor-risk groups, respectivcixely, with no significant differences (p = 0.133 and 0.797, respectively). The median OS in the good-, intermediate-, and poor-risk groups were statistically significant (p < 0.001) at 30.1, 14.2, and 5.7 months, respectively. A multivariate analysis revealed that the ARC and PSA doubling time were independent prognostic factors.
Most of CRPC patients were classified into good-risk group according to the ARC and the ARC could predict prognosis in DTX-treated CRPC patients.
Trial registration
University Hospital Medical Information Network Clinical Trials Registry (UMIN-CTR) number, UMIN000011969.
PMCID: PMC3997751  PMID: 24742323
Castration-resistant prostate cancer; Docetaxel; Risk classification; Validation study
5.  Docetaxel with or without estramustine for estramustine refractory castration-resistant prostate cancer: a single institution experience 
BMC Urology  2012;12:3.
The significance of combination of docetaxel (DTX) with estramustine phosphate (EMP) in castration-resistant prostate cancer (CRPC) patients remains unclear. In this study, we aimed to retrospectively evaluate the efficacy and toxicity of DTX with or without EMP and to elucidate the significance of DTX and EMP combination therapy in Japanese EMP-refractory CRPC patients.
To compare the efficacy and toxicity of DTX and EMP, we divided CRPC patients, who were confirmed to be resistant to EMP, into the following two groups: group D (n = 28), which included patients treated with DTX (60 mg/m2, once in every four weeks) alone, and group DE (n = 33), which included patients treated with a combination of DTX (60 mg/m2, once in every four weeks) and EMP (twice daily oral administration at 280 mg).
Prostate specific antigen (PSA) response (> 50% decline in PSA) was observed in six patients (21%) in group D and eight patients (24%) in group DE. The median time to progression (TTP) was 12.0 months and 6.2 months and the median overall survival (OS) was 26.4 months and 24.3 months in group D and DE, respectively. There was no statistical difference between the two groups in terms of PSA response, TTP, and OS. The incidence of adverse events of grade 3/4 was low in both the groups, and there was no statistical difference between the two groups.
Although treatment with DTX at 60 mg/m2 was effective and highly tolerated in EMP-refractory Japanese CRPC patients, the DTX and EMP combination therapy might not exhibit any survival benefit for CRPC patients.
PMCID: PMC3305626  PMID: 22353627
6.  Comparative genomic analysis of 1047 completely sequenced cDNAs from an Arabidopsis-related model halophyte, Thellungiella halophila 
BMC Plant Biology  2010;10:261.
Thellungiella halophila (also known as T. salsuginea) is a model halophyte with a small size, short life cycle, and small genome. Thellungiella genes exhibit a high degree of sequence identity with Arabidopsis genes (90% at the cDNA level). We previously generated a full-length enriched cDNA library of T. halophila from various tissues and from whole plants treated with salinity, chilling, freezing stress, or ABA. We determined the DNA sequences of 20 000 cDNAs at both the 5'- and 3' ends, and identified 9569 distinct genes.
Here, we completely sequenced 1047 Thellungiella full-length cDNAs representing abiotic-stress-related genes, transcription factor genes, and protein phosphatase 2C genes. The predicted coding sequences, 5'-UTRs, and 3'-UTRs were compared with those of orthologous genes from Arabidopsis for length, sequence similarity, and structure. The 5'-UTR sequences of Thellungiella and Arabidopsis orthologs shared a significant level of similarity, although the motifs were rearranged. While examining the stress-related Thellungiella coding sequences, we found a short splicing variant of T. halophila salt overly sensitive 1 (ThSOS1), designated ThSOS1S. ThSOS1S contains the transmembrane domain of ThSOS1 but lacks the C-terminal hydrophilic region. The expression level of ThSOS1S under normal growth conditions was higher than that of ThSOS1. We also compared the expression levels of Na+-transport-system genes between Thellungiella and Arabidopsis by using full-length cDNAs from each species as probes. Several genes that play essential roles in Na+ excretion, compartmentation, and diffusion (SOS1, SOS2, NHX1, and HKT1) were expressed at higher levels in Thellungiella than in Arabidopsis.
The full-length cDNA sequences obtained in this study will be essential for the ongoing annotation of the Thellungiella genome, especially for further improvement of gene prediction. Moreover, they will enable us to find splicing variants such as ThSOS1S (AB562331).
PMCID: PMC3017837  PMID: 21106055
7.  Role of PP2C-mediated ABA signaling in the moss Physcomitrella patens 
Plant Signaling & Behavior  2009;4(9):887-889.
Plant hormone abscisic acid (ABA) is found in a wide range of land plants, from mosses to angiosperms. However, our knowledge concerning the function of ABA is limited to some angiosperm plant species. We have shown that the basal land plant Physcomitrella patens and the model plant Arabidopsis thaliana share a conserved abscisic acid (ABA) signaling pathway mediated through ABI1-related type 2C protein phosphatases (PP2Cs). Ectopic expression of Arabidopsis abi1-1, a dominant allele of ABI1 that functions as a negative regulator of ABA signaling, or targeted disruption of Physcomitrella ABI1-related gene (PpABI1A) resulted in altered ABA sensitivity and abiotic stress tolerance of Physcomitrella, as demonstrated by osmostress and freezing stress. Moreover, transgenic Physcomitrella overexpressing abi1-1 showed altered morphogenesis. These trangenic plants had longer stem lengths compared to the wild type, and continuous growth of archegonia (female organ) with few sporophytes under non-stress conditions. Our results suggest that PP2C-mediated ABA signaling is involved in both the abiotic stress responses and developmental regulation of Physcomitrella.
PMCID: PMC2802809  PMID: 19847119
ABA; ABI1; Physcomitrella patens; PP2C; signaling
8.  Group A PP2Cs evolved in land plants as key regulators of intrinsic desiccation tolerance 
Nature Communications  2013;4:2219.
Vegetative desiccation tolerance is common in bryophytes, although this character has been lost in most vascular plants. The moss Physcomitrella patens survives complete desiccation if treated with abscisic acid (ABA). Group A protein phosphatases type 2C (PP2C) are negative regulators of abscisic acid signalling. Here we show that the elimination of Group A PP2C is sufficient to ensure P. patens survival to full desiccation, without ABA treatment, although its growth is severely hindered. Microarray analysis shows that the Group A PP2C-regulated genes exclusively overlap with genes exhibiting a high level of ABA induction. Group A PP2C disruption weakly affects ABA-activated kinase activity, indicating Group A PP2C action downstream of these kinases in the moss. We propose that Group A PP2C emerged in land plants to repress desiccation tolerance mechanisms, possibly facilitating plants propagation on land, whereas ABA releases the intrinsic desiccation tolerance from Group A PP2C regulation.
Abscisic acid plays an essential role in the induction of vegetative desiccation tolerance in bryophytes. Here the authors show that elimination of protein phosphatases 2C is sufficient for the moss Physcomitrella patens to survive desiccation without the assistance of abscisic acid.
PMCID: PMC3731658  PMID: 23900426

Results 1-8 (8)