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1.  Sorghum Dw1, an agronomically important gene for lodging resistance, encodes a novel protein involved in cell proliferation 
Scientific Reports  2016;6:28366.
Semi-dwarfing genes have contributed to enhanced lodging resistance, resulting in increased crop productivity. In the history of grain sorghum breeding, the spontaneous mutation, dw1 found in Memphis in 1905, was the first widely used semi-dwarfing gene. Here, we report the identification and characterization of Dw1. We performed quantitative trait locus (QTL) analysis and cloning, and revealed that Dw1 encodes a novel uncharacterized protein. Knockdown or T-DNA insertion lines of orthologous genes in rice and Arabidopsis also showed semi-dwarfism similar to that of a nearly isogenic line (NIL) carrying dw1 (NIL-dw1) of sorghum. A histological analysis of the NIL-dw1 revealed that the longitudinal parenchymal cell lengths of the internode were almost the same between NIL-dw1 and wildtype, while the number of cells per internode was significantly reduced in NIL-dw1. NIL-dw1dw3, carrying both dw1 and dw3 (involved in auxin transport), showed a synergistic phenotype. These observations demonstrate that the dw1 reduced the cell proliferation activity in the internodes, and the synergistic effect of dw1 and dw3 contributes to improved lodging resistance and mechanical harvesting.
doi:10.1038/srep28366
PMCID: PMC4916599  PMID: 27329702
2.  Bone management in Japanese patients with prostate cancer: hormonal therapy leads to an increase in the FRAX score 
BMC Urology  2016;16:32.
Background
Osteoporosis is a common consequence of androgen deprivation therapy (ADT) for prostate cancer. Up to 20 % of men on ADT have suffered from fractures within 5 years. The WHO Fracture Risk Assessment Tool (FRAX) has been utilized to predict the 10-year probability of major osteoporotic and hip fracture. However, to date, no large studies assessing the utility of the FRAX score in prostate cancer patients with or without ADT have been performed. We herein evaluated the impact of ADT on the FRAX score in prostate cancer patients.
Methods
The assessment of the FRAX score was performed in a total of 1220 prostate cancer patients, including patients who underwent brachytherapy (n = 547), radical prostatectomy (n = 200), external beam radiation therapy (n = 264) and hormonal therapy alone (n = 187) at Yokohama City University Hospital (Yokohama, Japan). We evaluated the effect of ADT on the FRAX score.
Results
Using the FRAX model, the median and mean 10-year probability of a major osteoporotic fracture according to the clinical risk factors alone was 7.9 % (8.8 ± 4.3 %), while the 10-year probability of hip fracture risk was 2.7 % (3.5 ± 3.1 %). In the ADT group, the duration of ADT was correlated with both major osteoporotic risk and hip fracture risk (R2 = 0.141, p < 0.001 and R2 = 0.166, p < 0.001, respectively). A comparison between the ADT (n = 187) and non-ADT (n = 399) groups demonstrated that the major fracture risk was > 20 % higher and the hip fracture risk was > 3 % higher in the ADT group than in the non-ADT group (ADT: 10 (5.3 %) and 118 (63.1 %), non-ADT 13 (3.3 %) and 189 (47.4 %), p < 0.001 and p < 0.001, respectively).
Conclusions
These results suggested that the longer duration of ADT led to an increased FRAX score, and the FRAX score may be a predictor of bone management treatment, particularly in prostate cancer patients.
doi:10.1186/s12894-016-0151-9
PMCID: PMC4912823  PMID: 27316330
Androgen deprivation therapy; FRAX; Prostate cancer; Bone fracture
3.  Expression of androgen receptor in non-muscle-invasive bladder cancer predicts the preventive effect of androgen deprivation therapy on tumor recurrence 
Oncotarget  2016;7(12):14153-14160.
Our recent retrospective study revealed a significantly reduced risk of bladder cancer (BC) recurrence in men who received androgen deprivation therapy (ADT) for their prostate cancer. However, whether androgen receptor (AR) signals contributed to the preventive effect of ADT remained unclear because ADT could reduce serum estrogens as well. The purpose of this study is to investigate the associations between the expression of AR/estrogen receptors (ERs) and BC recurrence in patients treated with ADT. We immunohistochemically stained 72 BCs and 42 corresponding normal urothelial tissues. AR/ERα/ERβ were positive in 44(61%)/22(31%)/39(54%) tumors and 35(83%)/24(57%)/34(81%) corresponding normal urothelial tissues, respectively. There were no statistically significant correlations between AR/ERα/ERβ expression and clinicopathological features of BC. With a median follow-up of 31.3 months, 12 (43%) of 28 patients with AR-negative tumor versus 11 (23%) of 44 patients with AR-positive tumor experienced BC recurrence. Thus, patients with AR-positive tumor had a significantly lower risk of BC recurrence (P=0.031), compared with those with AR-negative tumor. Meanwhile, the expression of ERα/ERβ in tumors and that of AR/ERα/ERβ in normal urothelial tissues were not significantly correlated with BC recurrence. A multivariate analysis revealed AR positivity in tumors as an independent prognosticator (hazard ratio: 0.27; 95% confidence interval: 0.11-0.67) for BC recurrence. These results indicate that ADT prevents BC recurrence via the AR pathway, but not via the ERα/ERβ pathways.
doi:10.18632/oncotarget.7358
PMCID: PMC4924704  PMID: 26885620
bladder cancer; recurrence; androgen receptor; androgen deprivation therapy
4.  Neutrophil-to-lymphocyte ratio is a prognostic marker in bladder cancer patients after radical cystectomy 
BMC Cancer  2016;16:185.
Background
There is no reliable biomarker for predicting the prognosis of patients who undergo radical cystectomy for bladder cancer. Recent studies have shown that the neutrophil-to-lymphocyte ratio (NLR) could function as a useful prognostic factor in several types of malignancies. This study aimed to assess the usefulness of NLR in bladder cancer.
Methods
A total of 74 patients who underwent radical cystectomy in our institutions from 1999 to 2014 were analyzed. The NLR was calculated using the patients’ neutrophil and lymphocyte counts before radical cystectomy. An immunohistochemical analysis was also performed to detect tumor infiltrating neutrophils (CD66b) and lymphocytes (CD8) in bladder cancer specimens.
Results
A univariate analysis showed that the patients with a high NLR (≥2.38; HR = 4.84; p = 0.007), high C-reactive protein level (>0.08; HR = 10.06; p = 0.030), or pathological lymph node metastasis (HR = 4.73; p = 0.030) had a significantly higher risk of cancer-specific mortality. Kaplan-Meier and log-rank tests further revealed that NLR was strongly correlated with overall survival (p = 0.018), but not progression-free survival (p = 0.137). In a multivariate analysis, all of these were found to be independent risk factors (HR = 4.62, 10.8, and 12.35, respectively). The number of CD8-positive lymphocytes was significantly increased in high-grade (p = 0.001) and muscle-invasive (p = 0.012) tumors, in comparison to low-grade and non-muscle-invasive tumors, respectively.
Conclusions
The NLR predicted the prognosis of patients who underwent radical cystectomy and might therefore function as a reliable biomarker in cases of invasive bladder cancer.
doi:10.1186/s12885-016-2219-z
PMCID: PMC4779264  PMID: 26944862
Bladder cancer; Radical cystectomy; Biomarker; Neutrophil-to-lymphocyte ratio
5.  Pretreatment neutrophil-to-lymphocyte ratio predicts the prognosis in patients with metastatic prostate cancer 
BMC Cancer  2016;16:111.
Background
The neutrophil-to-lymphocyte ratio (NLR), a simple marker of the systemic inflammatory response in critical care patients, has been suggested as an independent prognostic factor for several solid malignancies. We investigated the utility of pretreatment NLR as a prognosticator in patients who presented with metastatic prostate cancer.
Methods
We first investigated the correlation between NLR and prostate-specific antigen (PSA) levels in 1464 men who had both tests and were found to have prostate cancer on their biopsies at our institution from 1999 to 2015. We then assessed the relationship between pretreatment NLR and the prognosis in 48 patients who were diagnosed with prostate cancer metastasized to the lymph node and/or bone.
Results
The NLR value was significantly elevated in men with higher PSA than in those with lower PSA (p < 0.001). In patients with metastatic prostate cancer, NLR (cut-off point of 3.37 determined by the AUROC curve) was correlated with both cancer-specific (p = 0.018) and overall (p = 0.008) survivals.
Conclusions
Pretreatment NLR may function as a new biomarker that precisely predicts the prognosis in patients with metastatic prostate cancer.
doi:10.1186/s12885-016-2134-3
PMCID: PMC4754823  PMID: 26883640
Prostate cancer; Biomarker; Neutrophil-to-lymphocyte ratio; Metastasis
6.  JunB promotes cell invasion, migration and distant metastasis of head and neck squamous cell carcinoma 
Background
While treatment failure in cases of head and neck squamous cell carcinoma (HNSCC) frequently takes the form of locoregional recurrences and distant metastasis, our understanding of the mechanisms of metastasis in HNSCC is limited. We initially performed the upstream and key nodes analysis together with whole gene microarray analysis characterized by distant metastatic potential in vivo with HNSCC cell lines and identified JunB, a member of the activator protein-1 (AP-1) family, as a key molecule in the regulation of the pathways related to distant metastasis in HNSCC. We have therefore tested the hypothesis that JunB plays a crucial role in distant metastasis in HNSCC.
Methods
To study the role of JunB on metastatic potential of HNSCC, small interfering RNA (siRNA)-mediated knockdown and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (cas9) system (CRISPR/Cas9)-mediated knockout of JunB in HNSCC cells were established and the abilities of cell invasion and migration in vitro were examined. The efficacy of knockout of JunB was also examined using an experimental lung metastatic mouse model of HNSCC. In addition, to study if the role of JunB in HNSCC cell migration and invasiveness is related to epithelial-to-mesenchymal transition (EMT), cell morphology and expression of mesenchymal or epithelial marker on siRNA mediated JunB knockdown in HNSCC cells were examined with or without TGF-β stimulation.
Results
siRNA knockdown and sgRNA knockout of JunB in metastatic HNSCC cells significantly suppressed both cell invasion and migration in vitro. In addition, the knockout of JunB in metastatic HNSCC cells significantly repressed the incidence of lung metastases and prolonged the survival in vivo. However, we did not observe any change in cell morphology with the down-regulation of mesenchymal markers and up-regulation of epithelial markers in response to siRNA-mediated JunB knockdown in HNSCC cells.
Conclusion
These results suggested that JunB could play an important role in promoting cell invasion, migration and distant metastasis in HNSCC via pathways other than EMT and that the down-regulation of JunB may become an effective strategy for patients with invasive HNSCC.
Electronic supplementary material
The online version of this article (doi:10.1186/s13046-016-0284-4) contains supplementary material, which is available to authorized users.
doi:10.1186/s13046-016-0284-4
PMCID: PMC4709939  PMID: 26754630
JunB; AP-1; Head and neck squamous cell carcinoma (HNSCC); Distant metastasis; CRISPR/Cas9-mediated knockout
7.  Neutrophil-to-lymphocyte ratio predicts prostatic carcinoma in men undergoing needle biopsy 
Oncotarget  2015;6(31):32169-32176.
Neutrophil-to-lymphocyte ratio (NLR), a simple marker of systemic inflammatory response, has been demonstrated as an independent prognosticator for some solid malignancies, including prostate cancer. In the present study, we evaluated the role of NLR in men who underwent prostate needle biopsy for their initial diagnosis of prostatic carcinoma. Both complete blood counts and free/total (F/T) prostate-specific antigen (PSA) ratio were examined in a total of 3,011 men in our institution. Of these, 1,207 had a PSA level between 4 and 10 ng/mL, and 357 of 810 who subsequently underwent prostate needle biopsy were found to have prostatic adenocarcinoma. NLR value was significantly higher in men with PSA of ≥ 20 ng/mL than in those with PSA of < 20 ng/mL (p < 0.001). NLR was also significantly higher in men with positive biopsy than in those with negative biopsy (p < 0.001). Using NLR cut-off point of 2.40 determined by the AUROC curve, positive/negative predictive values of NLR alone and NLR combined with F/T PSA ratio (cut-off: 0.15) were 56.6%/60.8% and 80.7%/60.1%, respectively. Multivariate analysis revealed that not only F/T PSA ratio (HR = 3.13) but also NLR (HR = 2.21) was an independent risk factor for prostate cancer. NLR is thus likely elevated in patients with prostate cancer. Accordingly, NLR, with or without combination with F/T PSA ratio, may function as a new biomarker to predict prostate cancer in men undergoing prostate needle biopsy.
PMCID: PMC4741667  PMID: 26359354
Clinical Section; prostate cancer; biomarker; neutrophil-to-lymphocyte ratio; prostate needle biopsy
8.  At what level of unconsciousness is mild therapeutic hypothermia indicated for out-of-hospital cardiac arrest: a retrospective, historical cohort study 
Background
Appropriate patient selection is very important when initiating mild therapeutic hypothermia (MTH) for patients following out-of-hospital cardiac arrest, and the extent of unconsciousness at implementation must be defined in such cases. However, there are no clear standards regarding the level of unconsciousness at which MTH would be beneficial. The effects of MTH in patients with different degrees of unconsciousness according to the motor response score of the Glasgow Coma Scale (GCS) were investigated.
Methods
The subjects consisted of witnessed non-traumatic adult out-of-hospital cardiac arrest patients admitted to our institute from April 2002 to August 2011. The patients were divided into six groups according to the GCS motor response score: 1 (GCS M1), 2 (GCS M2), 3 (GCS M3), 4 (GCS M4), 5 (GCS M5), and 6 (GCS M6). The neurological outcome was evaluated at 30 days after hospital admission using the Cerebral Performance Category. Chi-squared Automatic Interaction Detection (CHAID) analysis was performed to estimate the threshold GCS M level where therapeutic hypothermia is indicated. Odds ratios were then calculated by multiple logistic-regression analysis using factors including GCS M5–6 and MTH.
Results
A total of 289 patients were enrolled in this study. CHAID analysis demonstrated two points of significant increase in percentage of good recovery at 30 days after admission, dividing the GCS M categories into three groups. Patients classified with a GCS motor response score of 5 or higher had the highest percentage of good recovery. The odds ratio for good recovery (CPC1–2) was 2.901 (95 % CI 1.460–5.763, P = 0.002) for MTH, and that for GCS M5–6 was 159.835 (95 % CI 33.592–760.513, P < 0.001).
Conclusions
MTH may be unnecessary in patients with a GCS motor response score of 5 or higher. Consequently, because there are post cardiac arrest patients with a GCS motor response score of 4 or lower who benefit from MTH, MTH may be limited to patients with a GCS motor response score of 4 or lower.
doi:10.1186/s40560-015-0104-5
PMCID: PMC4567798  PMID: 26366291
Mild therapeutic hypothermia; Post cardiac arrest; Coma; Glasgow Coma Scale; Motor response score
9.  Gibberellin deficiency pleiotropically induces culm bending in sorghum: an insight into sorghum semi-dwarf breeding 
Scientific Reports  2014;4:5287.
Regulation of symmetrical cell growth in the culm is important for proper culm development. So far, the involvement of gibberellin (GA) in this process has not yet been demonstrated in sorghum. Here, we show that GA deficiency resulting from any loss-of-function mutation in four genes (SbCPS1, SbKS1, SbKO1, SbKAO1) involved in the early steps of GA biosynthesis, not only results in severe dwarfism but also in abnormal culm bending. Histological analysis of the bent culm revealed that the intrinsic bending was due to an uneven cell proliferation between the lower and upper sides of culm internodes. GA treatment alleviated the bending and dwarfism in mutants, whereas the GA biosynthesis inhibitor, uniconazole, induced such phenotypes in wild-type plants— both in a concentration-dependent manner, indicating an important role of GA in controlling erectness of the sorghum culm. Finally, we propose that because of the tight relationship between GA deficiency-induced dwarfism and culm bending in sorghum, GA-related mutations have unlikely been selected in the history of sorghum breeding, as could be inferred from previous QTL and association studies on sorghum plant height that did not pinpoint GA-related genes.
doi:10.1038/srep05287
PMCID: PMC4055941  PMID: 24924234
11.  Close Association of Carbonic Anhydrase (CA2a and CA15a), Na+/H+ Exchanger (Nhe3b), and Ammonia Transporter Rhcg1 in Zebrafish Ionocytes Responsible for Na+ Uptake 
Freshwater (FW) fishes actively absorb salt from their environment to tolerate low salinities. We previously reported that vacuolar-type H+-ATPase/mitochondrion-rich cells (H-MRCs) on the skin epithelium of zebrafish larvae (Danio rerio) are primary sites for Na+ uptake. In this study, in an attempt to clarify the mechanism for the Na+ uptake, we performed a systematic analysis of gene expression patterns of zebrafish carbonic anhydrase (CA) isoforms and found that, of 12 CA isoforms, CA2a and CA15a are highly expressed in H-MRCs at larval stages. The ca2a and ca15a mRNA expression were salinity-dependent; they were upregulated in 0.03 mM Na+ water whereas ca15a but not ca2a was down-regulated in 70 mM Na+ water. Immunohistochemistry demonstrated cytoplasmic distribution of CA2a and apical membrane localization of CA15a. Furthermore, cell surface immunofluorescence staining revealed external surface localization of CA15a. Depletion of either CA2a or CA15a expression by Morpholino antisense oligonucleotides resulted in a significant decrease in Na+ accumulation in H-MRCs. An in situ proximity ligation assay demonstrated a very close association of CA2a, CA15a, Na+/H+ exchanger 3b (Nhe3b), and Rhcg1 ammonia transporter in H-MRC. Our findings suggest that CA2a, CA15a, and Rhcg1 play a key role in Na+uptake under FW conditions by forming a transport metabolon with Nhe3b.
doi:10.3389/fphys.2013.00059
PMCID: PMC3615223  PMID: 23565095
freshwater fish; osmoregulation; mitochondria-rich cell; sodium uptake; V-ATPase; GPI anchor; proximity ligation assay; duolink
12.  Class I/II hybrid inhibitory oligodeoxynucleotide exerts Th1 and Th2 double immunosuppression 
FEBS Open Bio  2012;3:41-45.
We designed class I/II hybrid inhibitory oligodeoxynucleotides (iODNs), called iSG, and found that the sequence 5′-TTAGGG-3′, which has a six-base loop head structure, and a 3′-oligo (dG)3–5 tail sequence are important for potent immunosuppressive activity. Interestingly, splenocytes isolated from ovalbumin (OVA)-immunized mice and treated with iSG3 showed suppression of not only interleukin (IL)-6, IL-12p35, IL-12p40, and interferon (IFN) γ mRNA expression, but also IL-4 and IL-13 mRNA expression. Thus, both Th2 and Th1 immune responses can be strongly suppressed by iODNs in splenocytes from allergen-immunized mice, suggesting usefulness in the treatment of diseases induced by over-active immune activation.
Highlights
▸ Inhibitory oligodeoxynucleotides (iODNs) suppress some innate immune responses. ▸ We designed a class I/II hybrid iODN that includes the telomeric motif 5′-TTAGGG-3′. ▸ The telomere motif and a 3′-(G)3–5 polytail sequence are required for potent immunosuppression. ▸ Class I/II hybrid iODNs exert a novel effect of Th1 and Th2 double immunosuppression.
doi:10.1016/j.fob.2012.11.002
PMCID: PMC3668506  PMID: 23847756
iODN; Class I iODN; Class II iODN; Class I/II hybrid iODN; immunosuppression; ODN, oligodeoxynucleotide; IL, interleukin; TLR, Toll-like receptor; PS, phosphorothioate; PO, phosphodiester; OVA, ovalbumin; ELISA, enzyme-linked immunosorbent assay; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; IFN, interferon; Th1 cell, type 1 helper T cell; Th2 cell, type 2 helper T cell; STAT, signal transducer and activator of transcription
13.  Identification of miR-30d as a novel prognostic maker of prostate cancer 
Oncotarget  2012;3(11):1455-1471.
Prostate cancer (PCa) is the most common malignant carcinoma that develops in men in Western countries. MicroRNA (miRNA) have the potential to be used as biomarkers and therapeutic targets for the treatment of various cancers. We found significantly higher expression of miR-30d in 3 PCa cell lines (PC3, DU145 and LNCaP) compared with 2 normal prostate cell lines (RWPE-1 and PrSc) using miRNA microarrays and qPCR. Clinicopathological study revealed that miR-30d expression levels were significantly higher in cancer tissue samples than in the paired normal controls (P = 0.03). Furthermore, the miR-30d−high group had shorter time to biochemical recurrence (P = 0.026). MiR-30d overexpressed PCa cells promoted proliferation and invasion in vitro. Inoculation of miR-30d depleted PCa cells dramatically reduced tumor volumes in vivo. Using reporter gene assay, we identified miR-30d as a downregulator of SOCS1 expression by directly binding to 3'-UTR of SOCS1. MiR-30d regulated the expression of phospho-STAT3, MMP-2 and MMP-9 through the downregulation of SOCS1. The levels of SOCS1 mRNA and protein were significantly down-regulated in prostate cancer tissues. Consistently, miR-30d expression was inversely correlated with SOCS1 expression (P = 0.03). The miR-30d−high/SOCS1−low group was associated with an increased risk of early biochemical recurrence (P = 0.0057). Taken together, miR-30d appears to be a novel independent prognostic marker of PCa progression that allows clinicians to identify patients who need more intensive treatments.
PMCID: PMC3717805  PMID: 23231923
miRNA-30d; prostate cancer; biochemical recurrence; SOCS1
14.  A necrotic lung ball caused by co-infection with Candida and Streptococcus pneumoniae 
Introduction
A necrotic lung ball is a rare radiological feature that is sometimes seen in cases of pulmonary aspergillosis. This paper reports a rare occurrence of a necrotic lung ball in a young male caused by Candida and Streptococcus pneumoniae.
Case report
A 28-year-old male with pulmonary candidiasis was found to have a lung ball on computed tomography (CT) of the chest. The patient was treated with β-lactams and itraconazole and then fluconazole, which improved his condition (as found on a following chest CT scan) and serum β-D-glucan level. The necrotic lung ball was suspected to have been caused by coinfection with Candida and S. pneumoniae.
Conclusion
A necrotic lung ball can result from infection by Candida and/or S. pneumoniae, indicating that physicians should be aware that patients may still have a fungal infection of the lungs that could result in a lung ball, even when they do not have either Aspergillus antibodies or antigens.
doi:10.2147/IDR.S24269
PMCID: PMC3259690  PMID: 22259251
lung ball; necrotic lung ball; Candida; Streptococcus pneumoniae
15.  Identification of Zebrafish Fxyd11a Protein that is Highly Expressed in Ion-Transporting Epithelium of the Gill and Skin and its Possible Role in Ion Homeostasis 
FXYD proteins, small single-transmembrane proteins, have been proposed to be auxiliary regulatory subunits of Na+–K+-ATPase and have recently been implied in ion osmoregulation of teleost fish. In freshwater (FW) fish, numerous ions are actively taken up through mitochondrion-rich cells (MRCs) of the gill and skin epithelia, using the Na+ electrochemical gradient generated by Na+–K+-ATPase. In the present study, to understand the molecular mechanism for the regulation of Na+–K+-ATPase in MRCs of FW fish, we sought to identify FXYD proteins expressed in MRCs of zebrafish. Reverse-transcriptase PCR studies of adult zebrafish tissues revealed that, out of eight fxyd genes found in zebrafish database, only zebrafish fxyd11 (zfxyd11) mRNA exhibited a gill-specific expression. Double immunofluorescence staining showed that zFxyd11 is abundantly expressed in MRCs rich in Na+–K+-ATPase (NaK-MRCs) but not in those rich in vacuolar-type H+-transporting ATPase. An in situ proximity ligation assay demonstrated its close association with Na+–K+-ATPase in NaK-MRCs. The zfxyd11 mRNA expression was detectable at 1 day postfertilization, and its expression levels in the whole larvae and adult gills were regulated in response to changes in environmental ionic concentrations. Furthermore, knockdown of zFxyd11 resulted in a significant increase in the number of Na+–K+-ATPase–positive cells in the larval skin. These results suggest that zFxyd11 may regulate the transport ability of NaK-MRCs by modulating Na+–K+-ATPase activity, and may be involved in the regulation of body fluid and electrolyte homeostasis.
doi:10.3389/fphys.2010.00129
PMCID: PMC3059942  PMID: 21423371
calcium; Danio renio; FXYD-domain ion transport regulator; mitochondria-rich cell; Na+–K+-ATPase; osmoregulation; salinity; teleost

Results 1-15 (15)