Metabolome-based discrimination of chrysanthemum cultivars for the efficient generation of flower color variations in mutation breeding.

INTRODUCTION: The color variations of ornamental flowers are often generated by ion-beam and gamma irradiation mutagenesis. However, mutation rates differ significantly even among cultivars of the same species, resulting in high cost and intensive labor for flower color breeding. OBJECTIVES: We aimed to establish a metabolome-based strategy to identify biomarkers and select promising parental lines with high mutation rates using Chrysanthemum as the case study. METHODS: The mutation rates associated with flower color were measured in 10 chrysanthemum cultivars with pink, yellow, or white flowers after soft X-ray irradiation at the floret-formation stage. The metabolic profiles of the petals of these cultivars were clarified by widely targeted metabolomics and targeted carotenoid analysis using liquid chromatography-tandem quadrupole mass spectrometry. Metabolome and carotenoid data were subjected to an un-supervised principal component analysis (PCA) and a supervised logistic regression with least absolute shrinkage and selection operator (LASSO). RESULTS: The PCA of the metabolic profile data separated chrysanthemum cultivars according to flower color rather than mutation rates. By contrast, logistic regression with LASSO generated a discrimination model to separate cultivars into two groups with high or low mutation rates, and selected 11 metabolites associated with mutation rates that can be biomarkers candidates for selecting parental lines for mutagenesis. CONCLUSION: This metabolome-based strategy to identify metabolite markers for mutation rates associated with flower color might be applied to other ornamental flowers to accelerate mutation breeding for generating new cultivars with a wider range of flower colors.

Detection of Extremely Low Level Ciguatoxins through Monitoring of Lithium Adduct Ions by Liquid Chromatography-Triple Quadrupole Tandem Mass Spectrometry.

Ciguatera poisoning (CP) is the most common type of marine biotoxin food poisoning worldwide, and it is caused by ciguatoxins (CTXs), thermostable polyether toxins produced by dinoflagellate Gambierdiscus and Fukuyoa spp. It is typically caused by the consumption of large fish high on the food chain that have accumulated CTXs in their flesh. CTXs in trace amounts are found in natural samples, and they mainly induce neurotoxic effects in consumers at concentrations as low as 0.2 µg/kg. The U.S. Food and Drug Administration has established CTX maximum permitted levels of 0.01 µg/kg for CTX1B and 0.1 µg/kg for C-CTX1 based on toxicological data. More than 20 variants of the CTX1B and CTX3C series have been identified, and the simultaneous detection of trace amounts of CTX analogs has recently been required. Previously published works using LC-MS/MS achieved the safety levels by monitoring the sodium adduct ions of CTXs ([M+Na]+ > [M+Na]+). In this study, we optimized a highly sensitive method for the detection of CTXs using the sodium or lithium adducts, [M+Na]+ or [M+Li]+, by adding alkali metals such as Na+ or Li+ to the mobile phase. This work demonstrates that CTXs can be successfully detected at the low concentrations recommended by the FDA with good chromatographic separation using LC-MS/MS. It also reports on the method's new analytical conditions and accuracy using [M+Li]+.

Frequency of elevated biomarkers in patients with cryptogenic hepatocellular carcinoma.

BACKGROUND: The incidence of hepatocellular carcinoma (HCC) continues to increase in Japan, but the clinical characteristics of Japanese patients with HCC have not been well described. The aim of this study was to determine the frequencies and utilities of elevated a-fetoprotein (AFP) and des-gamma-carboxy prothrombin (DCP) levels as biomarkers in cryptogenic HCC. MATERIAL/METHODS: A total of 2638 patients with HCC diagnosed between 1999 and 2010 in the Nagasaki Association Study of Liver (NASLD) were recruited for this study. The cause of HCC was categorized into 4 groups; HCC-B, HCC-C, HCC-BC, and HCC-nonBC. The significance of factors was examined for HCC-nonBC using logistic regression analysis in all patients. RESULTS: Multivariate analysis identified age, sex, BMI, alcohol consumption, platelet count, AST, ALT, AFP, DCP, and TNM stage as independent and significant risk factors for HCC-nonBC. According to TNM stage, the median AFP levels in HCC-nonBC with TNM stages I, II, and III were significantly lower than in either HCC-B or HCC-C. In TNM stage IV, the median AFP level in HCC-nonBC was significantly lower than in either HCC-B or HCC-BC. The median DCP levels in HCC-nonBC with TNM stages I and II were significantly higher than those in either HCC-B or HCC-C. In TNM stage III, the median DCP level in HCC-nonBC was significantly higher than that in HCC-C. CONCLUSIONS: DCP was more sensitive than AFP for the diagnosis of early stage cryptogenic HCC. DCP should be used as the main serum test for cryptogenic HCC detection.

[Impact of preventive administration of antiemetic drugs on risk of acute nausea and vomiting induced by transcatheter arterial chemoembolization in patients with hepatocellular carcinomas - a retrospective study].

This study aimed to evaluate the impact of the preventive administration of antiemetic drugs on the risk of acute nausea and vomiting induced by transcatheter arterial chemoembolization(TACE)in patients with hepatocellular carcinomas(HCCs). From January 2007 to June 2009, a total of 536 patients with HCCs who underwent TACE with a mixture of iodized oil, epirubicin, and porous gelatin particles were included in this retrospective study. Of those patients, 23 out of 357(6.4% ) who had received the 5-HT(3) receptor antagonist before TACE, and 18 out of 179(10.1% )without the medication, experienced vomiting. The multivariate logistic regression model with a predictive success of 92. 4% for vomiting identified significant associations between female gender(odds ratio: 3.73, p<0.001 ), the number of tumors(1.29, p<0.01 ), and administration of pentazocine(11.70, p<0.05)with the risk of vomiting. In contrast, the preventive administration of antiemetic drugs was not included in the model as a significant predictive variable. We therefore conclude from this retrospective study that the 5-HT(3) receptor antagonist did not significantly contribute to preventing the TACE-induced emesis.

Applicability of supercritical fluid chromatography for oligonucleotide analysis: A proof-of-concept study.

We evaluated the suitability of supercritical fluid chromatography (SFC) for oligonucleotide analysis using 4-mer oligonucleotides with various phosphorothioate (PS) contents as model compounds. Column screening showed that the diol-modified column was able to separate sequences with different PS contents. Optimization of the column body and additives allowed us to analyze polar oligonucleotides using SFC. Various sequences were also analyzed using the optimized method. A good peak shape was obtained when the guanine plus cytosine content of the analyte was two or less in the 4-mer oligonucleotides. Furthermore, we found that the retention times of the selected sequences were positively correlated with polar surface areas, indicating that oligonucleotides interact with polar stationary phases. In contrast, more hydrophobic full PS sequences were retained more strongly in the diol column than the full phosphodiester (PO) sequences. This suggests that the diol column has unique selectivity for PO and PS linkages. These results indicate that SFC is potentially applicable to oligonucleotide analysis with a separation mechanism that is different from that of ion-pair reversed-phase liquid chromatography.

Adherence to anti-retroviral therapy, decisional conflicts, and health-related quality of life among treatment-naïve individuals living with HIV: a DEARS-J observational study.

BACKGROUND: Supporting people living with HIV using anti-retroviral therapy (ART) is important due to the requirement for strict medication adherence. To date, no data from longitudinal studies evaluating adherence by treatment-naïve people living with HIV are currently available. We investigated the adherence of treatment-naïve people living with HIV over time and examined the relationships among decisional conflicts, adherence, and health-related quality of life (HRQL). METHODS: The survey items included adherence (visual analogue scale [VAS]), decisional conflict (decisional conflict scale [DCS]), and HRQL (Medical Outcomes Study HIV Health Survey [MOS-HIV]). The DCS and MOS-HIV scores and the VAS and MOS scores were collected electronically at the ART initiation time point and at 4-, 24-, and 48-week post-treatment time points. RESULTS: A total of 215 participants were enrolled. The mean DCS score was 27.3 (SD, 0.9); 23.3% of participants were in the high-score and 36.7% in the low-score groups. The mean adherence rates at 4, 24, and 48 weeks were 99.2% (standard error [SE], 0.2), 98.4% (SE, 0.4), and 96.0% (SE, 1.2), respectively. The least-square means of the MOS-HIV for the DCS (high vs. low scores) were 64.4 vs. 69.2 for general health perceptions and 57.7 vs. 64.0 for HRQL, respectively. CONCLUSION: Adherence among treatment-naïve people living with HIV was maintained at a higher level, and HRQL tended to improve with ART. People with high levels of decisional conflict tended to have lower HRQL scores. Support for people living with HIV during ART initiation may be related to HRQL.

Promotion of proper use of anti-SARS-CoV-2 drugs and SARS-CoV-2 vaccines by hospital pharmacists and establishment of an adverse drug reaction reporting system.

Newly developed anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) drugs are being rapidly approved in countries worldwide. These new drugs are being approved after testing with a limited number of cases, and in real-world clinical practice, unknown and potentially serious adverse events that could not be detected in clinical trials may emerge. Accordingly, in the event of an adverse drug reaction for which a causal relationship with these new drugs cannot be ruled out, it is vital to promptly report the details of the case to the regulatory authorities. To date, through close cooperation between physicians and pharmacists, we have reported four cases of adverse drug reactions for which a causal relationship to anti-SARS-CoV-2 drugs cannot be ruled out. Herein, we introduce safety measures taken by pharmacists when using these new drugs in the hospital, and a system for reporting to the regulatory authorities when adverse events occur.

Extracorporeal membrane oxygenation may decrease the plasma concentration of remdesivir in a patient with severe coronavirus disease 2019.

Remdesivir is an antiviral drug that results in clinical improvement after five days of treatment and accelerates recovery by 31%. No studies have discussed the pharmacokinetic analysis of remdesivir in patients with severe COVID-19 requiring extracorporeal membrane oxygenation (ECMO). A 63-year-old American man who underwent mechanical ventilation and ECMO for severe COVID-19 was administered remdesivir for ten days. The loading dosage was 200 mg at 7 PM on day 12 and 100 mg daily at 0:00 PM from day 13-21, administered within 1 h. The pharmacokinetic analysis was performed. The serum creatinine concentration was within the normal range of 0.5-0.7 mg/dL during treatment. According to the pharmacokinetic analysis, the plasma concentrations of remdesivir and GS-441524 4 h after administration (C4) were 662 ng/mL and 58 ng/mL, respectively, and the concentrations 18 h after administration (C18) were 32 ng/mL and 44 ng/mL, respectively. Therefore, the half-life of remdesivir and GS-441524 was 3.2 and 35.1 h, respectively. Monitoring the plasma concentrations of remdesivir and GS-441524 in patients undergoing ECMO may be necessary.

Transplantation of human mesenchymal stem cells promotes functional improvement and increased expression of neurotrophic factors in a rat focal cerebral ischemia model.

Previous studies have suggested that intravenous transplantation of mesenchymal stem cells (MSCs) in rat ischemia models reduces ischemia-induced brain damage. Here, we analyzed the expression of neurotrophic factors in transplanted human MSCs and host brain tissue in rat middle cerebral artery occlusion (MCAO) ischemia model. At 1 day after transient MCAO, 3 x 10(6) immortalized human MSC line (B10) cells or PBS was intravenously transplanted. Behavioral tests, infarction volume, and B10 cell migration were investigated at 1, 3, 7, and 14 days after MCAO. The expression of endogenous (rat origin) and exogenous (human origin) neurotrophic factors and cytokines was evaluated by quantitative real-time RT-PCR and Western blot analysis. Compared with PBS controls, rats receiving MSC transplantation showed improved functional recovery and reduced brain infarction volume at 7 and 14 days after MCAO. In MSC-transplanted brain, among many neurotrophic factors, only human insulin-like growth factor 1 (IGF-1) was detected in the core and ischemic border zone at 3 days after MCAO, whereas host cells expressed markedly higher neurotrophic factors (rat origin) than control rats, especially vascular endothelial growth factor (VEGF) at 3 days and epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) at 7 days after MCAO. Intravenously transplanted human MSCs induced functional improvement, reduced infarct volume, and neuroprotection in ischemic rats, possibly by providing IGF-1 and inducing VEGF, EGF, and bFGF neurotrophic factors in host brain.

Effective ex vivo expansion of hematopoietic stem cells using osteoblast-differentiated mesenchymal stem cells is CXCL12 dependent.

Effective ex vivo expansion of hematopoietic stem cells (HSCs) is a prerequisite for HSC transplantation. Growth and maintenance of HSC is dependent on cytokine and niche factors. We investigated whether mesenchymal stem cells (MSCs) or osteogenic cytokine-differentiated MSCs play a role in HSC expansion. We used the human HM3.B10 (B10) MSC cell line and the osteoblast-differentiated B10 (Ost-B10) as a feeder layer and examined ex vivo expansion of CD34(+)CD38(-) HSCs obtained from peripheral blood (PB) and cord blood (CB) with or without several growth cytokines. Both undifferentiated B10 and Ost-B10 cells exhibited similar effects on total HSC expansion; however, Ost-B10 demonstrated a higher potency in CD34(+)CD38(-) cell-specific proliferation in the presence of cytokines compared to undifferentiated B10 HSCs. Colony-forming cell assay and long-term culture initiating cell assay revealed that Ost-B10 displayed multipotent differentiation and enabled long-term ex vivo culture of HSCs. We next examined the relationship between HSC expansion and the presence of various chemokines. CXCL4 and CXCL12 expression were increased in Ost-B10 cells compared with the B10 cells. CD34(+)CD38(-) cells were significantly increased with CXCL12, but not CXCL4 treatment. siRNA inhibition of CXCL12 decreased CXCL12 secretion in both B10 and Ost-B10, whereas expansion of CD34(+)CD38(-) cells was decreased in Ost-B10 alone. These results demonstrated that ex vivo expansion of HSCs may be highly effective through osteoblast-differentiated MSCs acting as a feeder layer, and likely operates through the CXCL12 chemokines signaling pathway.

Selecting treatment for hepatocellular carcinoma based on the results of hepatic resection and local ablation therapy.

BACKGROUND: First-line treatment for

[Evaluation of perioperative management of seven patients with pulmonary artery sling in recent seven years].

BACKGROUND: Pulmonary artery sling (PA sling) is a rare vascular anomaly in which the left pulmonary artery arises from the right pulmonary artery. It can cause compression of trachea. Due to its rarity, risk factors influencing postsurgical outcome are still unclear. The purpose of our study is to investigate past cases of PA sling to clarify risk factors. METHODS: We investigated 7 patients who had undergone one-stage operation of PA sling from 2001 to 2007. Hospital stay, ICU stay and length of artificial ventilation are set as primary outcome. Patients' weight, range of tracheal stenosis, operation time, cardiopulmonary bypass time, presence of tracheal stenosis, bronchial anomaly, preoperative intubation, intracardiac anomaly and postoperative re-intubation were compared. RESULTS: In-hospital mortality was none. Two cases in patients with tracheomalacia were re-intubated. Concomitant tracheoplasty was performed in six patients. Two patients with intracardiac anomaly underwent cardiac operation. No factor except tracheomalacia affected primary outcome. CONCLUSIONS: We need more cases to clarify factors affecting outcome of patients with PA sling. However, existence of tracheomalacia is found to be a risk factor of longer artificial ventilation.

Nucleos(t)ide reverse transcriptase inhibitor-sparing regimens in the era of standard 3-drug combination therapies for HIV-1 infection.

Nucleos(t)ide reverse transcriptase inhibitor (NRTI)-sparing regimens have often been selected as antiretroviral therapy (ART) for HIV-1 infection recently, but data for characteristics have been lacking. This study aimed to document the current status of NRTI-sparing regimens in the era of standard 3-drug combination therapies. We cross-sectionally compared characteristics of patients treated with NRTI-sparing regimens (NRTI-sparing group) with dolutegravir plus tenofovir alafenamide fumarate/emtricitabine as a standard ART group in 2018. The NRTI-sparing and the standard ART groups included 61 and 469 patients, respectively. The mean (± standard deviation) age and serum creatinine of the NRTI-sparing group were significantly higher than those of the standard ART group (57.6 ± 12.8 years vs 42.8 ± 10.4 years (p < 0.05) and 2.09 ± 3.10 mg/dL vs. 0.93 ± 0.19 mg/dL (p < 0.05), respectively. The percentage of patients with NRTI-sparing regimens increased with age; with less than 5% in their 50s or younger, 8.4% in their 60s, and 14.1% aged ≥ 70 years. The primary reason for switching to the NRTI-sparing regimen was due to reduced renal function. According to the limited data, viral suppression was achieved at week 48 in all patients in the NRTI-sparing group. No patient had treatment failure nor developed drug resistance. The use of NRTI-sparing regimens increased with age. They were more frequently used in patients aged ≥ 60 years and those with decreased renal function.

Lysophosphatidylcholine induces glial cell activation: role of rho kinase.

Lysophosphatidylcholine (LPC), a major phospholipid component of atherogenic oxidized LDL, is implicated in atherosclerosis and, recently, in neurodegenerative diseases. We investigated the immunomodulatory functions of LPC in the central nervous system (CNS) using both an in vivo rat model, and in vitro culture systems of human primary astrocytes and a microglia cell line, HMO6. Compared with PBS injection, 20 nmol LPC-injection into the rat striatum increased astrocyte and microglial accumulation and elevated iNOS expression; concomitantly a time-dependent decrease in number of neurons was exhibited. In vitro studies on astrocytes and HMO6 cells showed that LPC increased the gene expression of proinflammatory factors IL-1beta, COX-2, and GM-CSF. LPC also induced chemotactic responses in HMO6 cells. Inhibition of rho kinase by fasudil, Y27632, or expressing a dominant negative form of rho kinase inhibited the LPC-induced IL-1beta mRNA expression in both astrocytes and HMO6. Moreover, intraperitoneal fasudil injection inhibited the LPC-induced microglial accumulation and iNOS expression and also was effective in protecting against neuronal loss. Silencing G2A, a specific receptor for LPC, inhibited proinflammatory gene expression and HMO6 migration. Overall, our results indicate that LPC induced considerable neuroinflammatory reactivity in glia mediated by rho kinase-dependent pathways with inhibition of these pathways conferring significant extents of neuroprotection.

Septic shock induced by Lecythophora mutabilis in a patient with mitochondrial encephalomyopathy.

Invasive fungal infection (IFI) caused by Lecythophora mutabilis occasionally occurs in patients with impaired host immunity; such patients had eosinophilia at onset, and surviving patients were treated with fungal cell-membrane-targeted drugs. An 18-year-old man with mitochondrial encephalomyopathy accompanied with refractory anaemia and chronic renal failure developed septic shock caused by L. mutabilis, which was detected from a blood culture, and was identified morphologically and genetically. During the course of the infection, he had eosinophilia, although beta-d-glucan levels were within the normal range. He was treated with micafungin, but deteriorated and died, despite his treatment being changed to liposomal amphotericin B. On the basis of this we suggest that IFI caused by L. mutabilis should be suspected when a compromised host develops infection and eosinophilia, and that antifungal drugs that target beta-d-glucan are not advisable.

Pure red cell precursor toxicity by linezolid in a pediatric case.

Linezolid (LZD)-induced myelosuppression has been reported in adults; however, LZD-induced pure red cell precursor toxicity rarely occurs. A 2-year-old boy diagnosed with infective endocarditis by Streptococcus mitis received LZD after developing resistance to multiple antibiotics. Although his infective symptoms were improved by LZD, progressive anemia was noticed 2 weeks after LZD therapy. Four weeks after LZD administration, his hemoglobin level was 6.5 g/dL and reticulocytes less than 0.1%. Bone marrow examination revealed markedly decreased erythropoiesis with cytoplasmic vacuolation of erythroblasts. Anemia recovered 19 days after cessation of LZD. Elevated protoporphyrin and a high LZD level in the blood suggested that mitochondrial disturbance by high-dose and long-term treatment with LZD may have been responsible for LZD-induced pure red cell precursor toxicity.

Predictive factors for the development or regression of Fatty liver in Japanese adults.

Fatty liver is commonly associated with alcohol or metabolic syndrome. We aimed to examine the longitudinal aspects of fatty liver, and clarify the independent predictors for the development or regression of fatty liver. In the present study, the clinical features of 1578 Japanese adults (1208 men and 370 women; 35 to 69 years of age) who visited our center both in 2000 and 2007-2008 were recorded and compared, including liver status diagnosed by ultrasonography. Of the 1578 participants, 217 (13.8%) showed fatty liver development, and 74 (4.7%) showed fatty liver regression. Logistic regression analysis revealed that body mass index and percentage body fat were strongly associated with the development or regression of fatty liver. Metabolic syndrome-related disorders such as serum levels of total cholesterol, triglyceride, uric acid, and fasting blood glucose were also associated with clinical course to some degree. However, the history of alcohol intake, the presence of metabolic syndrome, blood pressure, and habitual physical exercise were not independent predictors for the development or regression of fatty liver. Our present data suggest that control of body weight in men and the percentage body fat in women are particularly important for the prevention or treatment of fatty liver.

[Clinical utility of epidural anesthesia during and after major spine surgery].

BACKGROUND: Spine surgery causes severe postoperative pain, but epidural anesthesia is not commonly used. The main reason is that it might cause some complications, such as motor block and infection. We compared the efficacy of epidural infusion with hypodermic infusion. METHODS: Thirty-two patients scheduled for spine surgery were included in the study. Patients were randomly assigned to receive either continuous epidural infusion (ropivacaine 0.18% and fentanyl 45.5 microg x ml(-1) at 2 ml x hr(-1)) or continuous hypodermic infusion via a patient-controlled analgesia system (morphine 0.04 mg x kg(-1) x ml(-1) at 0.5 ml x hr(-1); bolus: 0.5 ml x hr(-1)). We investigated pain and nausea by using visual analogue scale after surgery. RESULTS: Patient data did not differ between groups. Pain and nausea scores were significantly lower in the epidural infusion group than in the hypodermic infusion group. Epidural infusion did not cause any complications. The incidence of nausea in the hypodermic infusion group was 13 times as high as that in the epidural infusion group. Similarly, the incidence of nausea in female patients was 13 times as high as that in male patients. CONCLUSIONS: Continuous epidural infusion provides superior analgesia even if high-dose local anesthetics are not used. Besides, it decreases the incidence of nausea because systemic administration of opioids can be avoided.

An integrase of endogenous retrovirus is involved in maternal mitochondrial DNA inheritance of the mouse.

The mechanism of maternal mitochondrial DNA (mtDNA) inheritance in animals can be said to be the selective elimination of sperm mtDNA via the elimination factor of the egg and a sperm mitochondria-specific factor. In 2005, we clarified that t-tpis (Spag1 isoform 1) is a mitochondria-specific translocator and the sperm factor, and furthermore estimated that the elimination factors of the egg are the divalent cation-dependent endonuclease and s-tpis (Spag1 isoform 2 and isoform 3) as the elimination system-specific chaperone [K. Hayashida, K. Omagari, J. Masuda, H. Hazama, Y. Kadokawa, K. Ohba, S. Kohno, The sperm mitochondria-specific translocator has a key role in maternal mitochondrial inheritance, Cell Biol. Int. 29 (2005) 472-481]. This time, using a recombinant Spag1 isoform 1 protein, a pull-down assay of ovary cytosol was performed and the elimination factors searched for. Surprisingly, an endogenous retroviral integrase fragment (Eri15) was identified using mass spectrometry of the electrophoresis band of the pull-down protein. Eri15 was detected as a complex of approximately 500kDa with Spag1 isoform 2 or isoform 3 in native PAGE of the ovary cytosol. This strongly suggested that Eri15 is selectively transported into the sperm mitochondria matrix by Spag1 isoform 2 and 3 via Spag1 isoform 1 and that sperm mtDNA is destroyed, thus causing the establishment of maternal mtDNA inheritance.