Effect of a novel siRNA delivery system, siRNA ternary complex, on melanoma lung metastasis.

In this study, we determined effects of an anionic siRNA delivery vector, siRNA ternary complex, which is constructed with biodegradable dendrigraft poly-L-lysine (DGL) and γ-polyglutamic acid (γ-PGA) on the melanoma cells and melanoma lung metastasis. The siRNA ternary complex showed high cellular uptake and silencing effect in melanoma cell line B16-F10/Luc cells. After intravenous administration of the siRNA ternary complex, high silencing effect was also observed in the lung of B16-F10/Luc melanoma metastasis model mice. Therefore, we applied vascular endothelial growth factor (VEGF)-siRNA on the siRNA ternary complex and determined the effect on the melanoma lung metastasis. The siRNA ternary complex containing VEGF-siRNA reduced VEGF protein levels significantly in in vitro and in vivo, and the complex successfully inhibited melanoma lung metastasis. This biodegradable and effective siRNA delivery vector, siRNA ternary complex, could be available for clinical trials.

Suitable Promoter for DNA Vaccination Using a pDNA Ternary Complex.

In this study, we evaluated the effect of several promoters on the transfection activity and immune-induction efficiency of a plasmid DNA (pDNA)/polyethylenimine/γ-polyglutamic acid complex (pDNA ternary complex). Model pDNAs encoding firefly luciferase (Luc) were constructed with several promoters, such as simian virus 40 (SV40), eukaryotic elongation factor 1 alpha (EF1), cytomegalovirus (CMV), and chicken beta actin hybrid (CBh) (pSV40-Luc, pEF1-Luc, pCMV-Luc, and pCBh-Luc, respectively). Four types of pDNA ternary complexes, each with approximately 145-nm particle size and -30-mV ζ-potential, were stably constructed. The pDNA ternary complex containing pSV40-Luc showed low gene expression, but the other complexes containing pEF1-Luc, pCMV-Luc, and pCBh-Luc showed high gene expression in DC2.4 cells and spleen after intravenous administration. After immunization using various pDNA encoding ovalbumin (OVA) such as pEF1-OVA, pCMV-OVA, and pCBh-OVA, only the pDNA ternary complex containing pCBh-OVA showed significant anti-OVA immunoglobulin G (IgG) induction. In conclusion, our results showed that the CBh promoter is potentially suitable for use in pDNA ternary complex-based DNA vaccination.

Interaction of γ-Polyglutamic Acid/Polyethyleneimine/Plasmid DNA Ternary Complexes with Serum Components Plays a Crucial Role in Transfection in Mice.

Typical examples of non-viral vectors are binary complexes of plasmid DNA with cationic polymers such as polyethyleneimine (PEI). However, problems such as cytotoxicity and hemagglutination, owing to their positively charged surfaces, hinder their in vivo use. Coating binary complexes with anionic polymers, such as γ-polyglutamic acid (γ-PGA), can prevent cytotoxicity and hemagglutination. However, the role of interactions between these complexes and serum components in in vivo gene transfer remains unclear. In this study, we analyzed the contribution of serum components to in vivo gene transfer using PEI/plasmid DNA binary complexes and γ-PGA/PEI/plasmid DNA ternary complexes. In binary complexes, heat-labile components in the serum greatly contribute to the hepatic and splenic gene expression of the luciferase gene. In contrast, serum albumin and salts affected the hepatic and splenic gene expression in the ternary complexes. Changes in physicochemical characteristics, such as increased particle size and decreased absolute values of ζ-potential, might be involved in the enhanced gene expression. These findings would contribute to a better understanding of in vivo non-viral gene transfer using polymers, such as PEI and γ-PGA.

Utility of Blood Culture in Patients with Community-Acquired Pneumonia: A Propensity Score-Matched Analysis Based on a Japanese National Health Insurance Database.

Community-acquired pneumonia (CAP) is an acute pulmonary parenchymal infection acquired outside the hospital. The utility of blood cultures in inpatients with CAP to reduce mortality and length of hospital stay is controversial. This study aimed to determine the utility of blood cultures on the first day of hospitalization for CAP inpatients and its influence on mortality, length of hospital stay, and antibiotics use. We conducted a fact-finding survey on the implementation of blood culture in inpatients with CAP in Japan. A propensity score (PS)-matched analysis based on the National Database of Health Insurance Claims and Specific Health Check-ups of Japan database was conducted. Overall, 163173 patients were included in the analysis, and PS matching extracted 68104 pairs. The results of the comparison between the PS-matched blood culture group and PS-matched control group were as follows: mortality and length of hospital stay were significantly lower in the PS-matched blood culture group than in the control group. The adjusted odds ratio (OR) (95% confidence interval (CI)) for in-hospital mortality with blood culture test was 0.73 (0.68-0.79). Moreover, for days of antibiotic usage, number of antibiotics used were significantly higher in the PS-matched blood culture group than that in the control group. Our findings indicated that performing a blood culture on the first day of hospitalization for inpatients with CAP was associated with reduced mortality. To our knowledge, this is the largest epidemiological study to assess the utility of blood culture in Japanese inpatients with CAP. This testing method shows potential for application in clinical practice.

Auditory noise improves balance control by cross-modal stochastic resonance.

It is known that enhanced somatosensory function leads to improved balance, and somatosensory function can be enhanced by the appropriate level of mechanical, visual, or auditory noise. In this study, we tested the potential benefit of an auditory noise on balance control. We first assessed static balance by measuring 10 times the duration of standing on the toes of one leg with closed eyes. For the 18 healthy adult participants, the median standing times ranged from 2.1 to 45.6 s, and the median of the distribution was 9.9 s. From the above, the participants were divided into two groups: lower (below 10 s, n = 9) and higher (above 10 s, n = 9) balance groups. We then investigated the effect on balance control of an auditory white noise emitted at the detection threshold. Each individual performed 20 trials. The auditory noise was applied in half the trials, while the remaining trials were conducted without noise. The order of the noise and no-noise trials was quasi-random. In the lower-balance group, the median standing time significantly increased during the noise trials (10.3 s) compared with the time in the no-noise controls (5.2 s). On the other hand, noise had no significant effect in the higher-balance group, presumably because of a ceiling effect. These findings suggest that static balance in the lower-balance participants can be improved by applying a weak noise through cross-modal stochastic resonance.

Gene-Activated Matrix with Self-Assembly Anionic Nano-Device Containing Plasmid DNAs for Rat Cranial Bone Augmentation.

We have developed nanoballs, a biocompatible self-assembly nano-vector based on electrostatic interactions that arrange anionic macromolecules to polymeric nanomaterials to create nucleic acid carriers. Nanoballs exhibit low cytotoxicity and high transfection efficiently in vivo. This study investigated whether a gene-activated matrix (GAM) composed of nanoballs containing plasmid (p) DNAs encoding bone morphogenetic protein 4 (pBMP4) could promote bone augmentation with a small amount of DNA compared to that composed of naked pDNAs. We prepared nanoballs (BMP4-nanoballs) constructed with pBMP4 and dendrigraft poly-L-lysine (DGL, a cationic polymer) coated by γ-polyglutamic acid (γ-PGA; an anionic polymer), and determined their biological functions in vitro and in vivo. Next, GAMs were manufactured by mixing nanoballs with 2% atelocollagen and ß-tricalcium phosphate (ß-TCP) granules and lyophilizing them for bone augmentation. The GAMs were then transplanted to rat cranial bone surfaces under the periosteum. From the initial stage, infiltrated macrophages and mesenchymal progenitor cells took up the nanoballs, and their anti-inflammatory and osteoblastic differentiations were promoted over time. Subsequently, bone augmentation was clearly recognized for up to 8 weeks in transplanted GAMs containing BMP4-nanoballs. Notably, only 1 µg of BMP4-nanoballs induced a sufficient volume of new bone, while 1000 µg of naked pDNAs were required to induce the same level of bone augmentation. These data suggest that applying this anionic vector to the appropriate matrices can facilitate GAM-based bone engineering.

Delivery of pDNA to the Lung by Lipopolyplexes Using N-Lauroylsarcosine and Effect on the Pulmonary Fibrosis.

In a previous study, we constructed a lung-targeting lipopolyplex containing polyethyleneimine (PEI), 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA), and N-lauroylsarcosine (LS). The lipopolyplex exhibited an extremely high gene expression in the lung after intravenous administration. Here, we optimized the lipopolyplex and used it to deliver a TGF-ß1 shRNA to treat refractory pulmonary fibrosis. We constructed several lipopolyplexes with pDNA, various cationic polymers, cationic lipids, and LS to select the most effective formulation. Then, the pDNA encoding shRNA against mouse TGF-ß1 was encapsulated in the lipopolyplex and injected into mice with bleomycin-induced pulmonary fibrosis. After optimizing the lipopolyplex, dendrigraft poly-L-lysine (DGL) and DOTMA were selected as the appropriate cationic polymer and lipid, respectively. The lipopolyplex was constructed with a pDNA, DGL, DOTMA, and LS charge ratio of 1:2:2:4 showed the highest gene expression. After intravenous administration of the lipopolyplex, the highest gene expression was observed in the lung. In the in vitro experiment, the lipopolyplex delivered pDNA into the cells via endocytosis. As a result, the lipopolyplex containing pDNA encoding TGF-ß1 shRNA significantly decreased hydroxyproline in the pulmonary fibrosis model mice. We have successfully inhibited pulmonary fibrosis using a novel lung-targeting lipopolyplex.

Analysis of Predictive Factors for Diarrhea after the Administration of Naldemedine.

Naldemedine (NAL), a peripherally acting µ-opioid receptor antagonist, is effective for opioid-induced constipation (OIC). However, diarrhea is the most common adverse event. We investigated the incidence of NAL-induced diarrhea in patients who started NAL at Nagasaki University Hospital between June 2017 and March 2019. Predictors of NAL-induced diarrhea were analyzed using a multivariate logistic regression model. Two hundred and forty-two patients were included in the present study, and NAL-induced diarrhea was observed in 17.8% (43 patients). The results of multiple logistic regression analyses identified the administration of opioid analgesics for 8 d or longer before the initiation of NAL (odds ratio (OR): 2.20, 95% confidence interval (95% CI): 1.04-4.64, p = 0.039), the combination of a laxative (OR: 2.22, 95%CI: 1.03-4.81, p = 0.042), and the combination of CYP3A4 inhibitors (strong/moderate) (OR: 2.80, 95%CI: 1.02-7.67, p = 0.045) as risk factors. Therefore, the development of diarrhea needs to be considered in patients with these risk factors. Furthermore, diarrhea may be controlled by the initiation of NAL within 7 d of opioid analgesics and, where possible, the discontinuation of or change in the combination of moderate or strong CYP3A4 inhibitors.

Induction of mucosal immunity by pulmonary administration of a cell-targeting nanoparticle.

We previously found that a nanoparticle constructed with an antigen, benzalkonium chloride (BK) and γ-polyglutamic acid (γ-PGA) showed high Th1 and Th2-type immune induction after subcutaneous administration. For prophylaxis of respiratory infections, however, mucosal immunity should be induced. In this study, we investigated the effect of pulmonary administration of a nanoparticle comprising ovalbumin (OVA) as a model antigen, BK, and γ-PGA on induction of mucosal immunity in the lungs and serum. The complex was strongly taken up by RAW264.7 and DC2.4cells. After pulmonary administration, lung retention was longer for the OVA/BK/γ-PGA complex than for OVA alone. OVA-specific serum immunoglobulin (Ig)G was highly induced by the complex. High IgG and IgA levels were also induced in the bronchoalveolar lavage fluid, and in vivo toxicities were not observed. In conclusion, we effectively and safely induced mucosal immunity by pulmonary administration of an OVA/BK/γ-PGA complex.

Mesenchymal stem cell-derived extracellular vesicles as probable triggers of radiation-induced heart disease.

BACKGROUND: Radiation-induced heart disease has been reported, but the underlying mechanisms remain unclear. Mesenchymal stem cells (MSCs), also residing in the heart, are highly susceptible to radiation. We examined the hypothesis that the altered secretion of extracellular vesicles (EVs) from MSCs is the trigger of radiation-induced heart disease. METHODS: By exposing human placental tissue-derived MSCs to 5 Gy γ-rays, we then isolated EVs from the culture medium 48 h later and evaluated the changes in quantity and quality of EVs from MSCs after radiation exposure. The biological effects of EVs from irradiated MSCs on HUVECs and H9c2 cells were also examined. RESULTS: Although the amount and size distribution of EVs did not differ between the nonirradiated and irradiated MSCs, miRNA sequences indicated many upregulated or downregulated miRNAs in irradiated MSCs EVs. In vitro experiments using HUVEC and H9c2 cells showed that irradiated MSC-EVs decreased cell proliferation (P < 0.01), but increased cell apoptosis and DNA damage. Moreover, irradiated MSC-EVs impaired the HUVEC tube formation and induced calcium overload in H9c2 cells. CONCLUSIONS: EVs released from irradiated MSCs show altered miRNA profiles and harmful effects on heart cells, which provides new insight into the mechanism of radiation-related heart disease risks.

A Radiolabeled Self-assembled Nanoparticle Probe for Diagnosis of Lung-Metastatic Melanoma.

Melanoma is a highly malignant skin cancer that frequently metastasizes to the lung, bone, and brain at an early phase. Therefore, noninvasive detection of metastasized melanoma could be beneficial to determine suitable therapeutic strategies. We previously reported a biocompatible ternary anionic complex composed of plasmid DNA (pDNA), polyethyleneimine (PEI), and γ-polyglutamic acid (γ-PGA) based on an electrostatic interaction, which was highly taken up by melanoma cells (B16-F10), even if it was negatively charged. Here, we developed a radiolabeled γ-PGA complex by using indium-111 (111In)-labeled polyamidoamine dendrimer (4th generation; G4) instead of pDNA and iodine-125 (125I)-labeled PEI instead of native PEI, and evaluated its effectiveness as a melanoma-targeted imaging probe. This ternary complex was synthesized at a theoretical charge ratio; carboxyl groups of 111In-diethylenetriaminepentaacetic acid (DTPA)-G4 : amino groups of 125I-PEI : carboxyl groups of γ-PGA was 1 : 8 : 16, and the size and zeta potential were approximately 29 nm and -33 mV, respectively. This complex was taken up by B16-F10 cells with time. Furthermore, a biodistribution study, using normal mice, demonstrated its accumulation in the liver, spleen, and lung, where macrophage cells are abundant. Almost the same level of radioactivity derived from both 111In and 125I was observed in these organs at an early phase after probe injection. Compared with the normal mice, significantly higher lung-to-blood ratios of radioactivity were observed in the B16-F10-lung metastatic cancer model. In conclusion, the radiolabeled γ-PGA complex would hold potentialities for nuclear medical imaging of lung metastatic melanoma.

Evaluation of transgene expression characteristics and DNA vaccination against melanoma metastasis of an intravenously injected ternary complex with biodegradable dendrigraft poly-L-lysine in mice.

We developed a biocompatible splenic vector for a DNA vaccine against melanoma. The splenic vector is a ternary complex composed of plasmid DNA (pDNA), biodegradable dendrigraft poly-L-lysine (DGL), and γ-polyglutamic acid (γ-PGA), the selective uptake of which by the spleen has already been demonstrated. The ternary complex containing pDNA encoding luciferase (pCMV-Luc) exhibited stronger luciferase activity for RAW264.7 mouse macrophage-like cells than naked pCMV-Luc. Although the ternary complex exhibited strong luciferase activity in the spleen after its tail vein injection, luciferase activity in the liver and spleen was significantly decreased by a pretreatment with clodronate liposomes, which depleted macrophages in the liver and spleen. These results indicate that the ternary complex is mainly transfected in macrophages and is a suitable formulation for DNA vaccination. We applied the ternary complex to a pUb-M melanoma DNA vaccine. The ternary complex containing pUb-M suppressed the growth of melanoma and lung metastasis by B16-F10 mouse melanoma cells. We also examined the acute and liver toxicities of the pUb-M ternary complex at an excess pDNA dose in mice. All mice survived the injection of the excess amount of the ternary complex. Liver toxicity was negligible in mice injected with the excess amount of the ternary complex. In conclusion, we herein confirmed that the ternary complex was mainly transfected into macrophages in the spleen after its tail vein injection. We also showed the prevention of melanoma metastasis by the DNA vaccine and the safety of the ternary complex.

Anionic Complex with Efficient Expression and Good Safety Profile for mRNA Delivery.

We previously found that a complex comprising plasmid DNA (pDNA), polyethylenimine (PEI), and γ-polyglutamic acid (γ-PGA) had high transgene efficiency without cytotoxicity in vitro and in vivo. However, messenger RNA (mRNA) remains an attractive alternative to pDNA. In this study, we developed a safe and effective delivery system for mRNA to prevent its degradation and efficiently deliver it into target cells. Various cationic and anionic complexes were produced containing PEI, γ-PGA, and an mRNA encoding firefly luciferase. Their physicochemical properties and cytotoxicities were analyzed and the in vitro and in vivo protein expression were determined. The cationic mRNA/PEI complex showed high in vitro protein expression with strong cytotoxicity. The anionic complex was constructed as mRNA/PEI8/γ-PGA12 complex with a theoretical charge ratio of 1:8:12 based on the phosphate groups of the mRNA, nitrogen groups of PEI, and carboxylate groups of γ-PGA. It was stable and showed high in vitro protein expression without cytotoxicity. After intravenous administration of mRNA/PEI8/γ-PGA12 complex to mice, high protein expression was observed in the spleen and liver and slight expression was observed in the lung over 24 h. Thus, the newly constructed mRNA/PEI8/γ-PGA12 complex provides a safe and effective strategy for the delivery of mRNA.

Linked color imaging improves visibility of reflux esophagitis.

BACKGROUND: With more prevalent gastroesophageal reflux disease comes increased cases of Barrett's esophagus and esophageal adenocarcinoma. Image-enhanced endoscopy using linked-color imaging (LCI) differentiates between mucosal colors. We compared LCI, white light imaging (WLI), and blue LASER imaging (BLI) in diagnosing reflux esophagitis (RE). METHODS: Consecutive RE patients (modified Los Angeles [LA] classification system) who underwent esophagogastroduodenoscopy using WLI, LCI, and BLI between April 2017 and March 2019 were selected retrospectively. Ten endoscopists compared WLI with LCI or BLI using 142 images from 142 patients. Visibility changes were scored by endoscopists as follows: 5, improved; 4, somewhat improved; 3, equivalent; 2, somewhat decreased; and 1, decreased. For total scores, 40 points was considered improved visibility, 21-39 points was comparable to white light, and < 20 points equaled decreased visibility. Inter- and intra-rater reliabilities (Intra-class Correlation Coefficient [ICC]) were also evaluated. Images showing color differences (ΔE*) and L* a* b* color values in RE and adjacent esophageal mucosae were assessed using CIELAB, a color space system. RESULTS: The mean age of patients was 67.1 years (range: 27-89; 63 males, 79 females). RE LA grades observed included 52 M, 52 A, 24 B, 11 C, and 3 D. Compared with WLI, all RE cases showed improved visibility: 28.2% (40/142), LA grade M: 19.2% (10/52), LA grade A: 34.6% (18/52), LA grade B: 37.5% (9/24), LA grade C: 27.3% (3/11), and LA grade D: 0% (0/3) in LCI, and for all RE cases: 0% in BLI. LCI was not associated with decreased visibility. The LCI inter-rater reliability was "moderate" for LA grade M and "substantial" for erosive RE. The LCI intra-rater reliability was "moderate-substantial" for trainees and experts. Color differences were WLI: 12.3, LCI: 22.7 in LA grade M; and WLI: 18.2, LCI: 31.9 in erosive RE (P < 0.001 for WLI vs. LCI). CONCLUSION: LCI versus WLI and BLI led to improved visibility for RE after subjective and objective evaluations. Visibility and the ICC for minimal change esophagitis were lower than for erosive RE for LCI. With LCI, RE images contrasting better with the surrounding esophageal mucosa were more clearly viewed.

Development of Multi-functional Nanoparticles for Clinical Application to Gene and Nucleic Acid Medicines.

Gene and nucleic medicines have recently gained attention as novel drugs with the advancement of molecular biology and genetics; however, they have low bioavailability and low target delivery due to their low stability and poor membrane permeability. Therefore, the development of an effective drug delivery system (DDS) is necessary for the practical use of gene and nucleic acid medicines; however, despite considerable research, both safety and efficiency remain poor. Furthermore, the healthcare needs are not met by traditional DDS. Therefore, we developed an effective multi-functional DDS, which is constructed using materials that are safe for human consumption. This DDS involves several ternary complexes as novel gene delivery carriers constructed by coating the cationic complex of the gene and nucleic acid medicines as well as the biodegradable cationic polymer with a biocompatible anionic polymer. Early implementation of the ternary complex in clinical studies is expected due to their efficacy and safety. Furthermore, these complexes may be prepared using large-scale manufacturing. In addition, personalized DDS may be prepared according to the patient's disease stage, which is useful for advanced therapy.

Development of a DNA Vaccine for Melanoma Metastasis by Inhalation Based on an Analysis of Transgene Expression Characteristics of Naked pDNA and a Ternary Complex in Mouse Lung Tissues.

The present study investigated a pulmonary delivery system of plasmid DNA (pDNA) and its application to melanoma DNA vaccines. pCMV-Luc, pEGFP-C1, and pZsGreen were used as a model pDNA to evaluate transfection efficacy after inhalation in mice. Naked pDNA and a ternary complex, consisting of pDNA, dendrigraft poly-l-lysine (DGL), and γ-polyglutamic acid (γ-PGA), both showed strong gene expression in the lungs after inhalation. The transgene expression was detected in alveolar macrophage-rich sites by observation using multi-color deep imaging. On the basis of these results, we used pUb-M, which expresses melanoma-related antigens (ubiquitinated murine melanoma gp100 and tyrosinase-related protein 2 (TRP2) peptide epitopes), as DNA vaccine for melanoma. The inhalation of naked pUb-M and its ternary complex significantly inhibited the metastasis of B16-F10 cells, a melanoma cell line, in mice. The levels of the inflammatory cytokines, such as TNF-α, IFN-γ, and IL-6, which enhance Th1 responses, were higher with the pUb-M ternary complex than with naked pUb-M and pEGFP-C1 ternary complex as control. In conclusion, we clarified that the inhalation of naked pDNA as well as its ternary complex are a useful technique for cancer vaccination.

Proliferation-associated long noncoding RNA, TMPO-AS1, is a potential therapeutic target for triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer compared with luminal or epidermal growth factor receptor 2 subtypes, thus effective therapeutic options for TNBC are yet to be developed. Nowadays, oncogenic long noncoding RNAs (lncRNAs) are applied to cancer management as a new class of therapeutic targets. We previously showed that thymopoietin antisense transcript 1 (TMPO-AS1) is a proliferation-associated lncRNA that contributes to hormone-dependent breast cancer progression by stabilizing estrogen receptor-α mRNA. We here showed that TMPO-AS1 is abundantly expressed in basal-like breast cancer subtype based on the transcriptomic data in The Cancer Genome Atlas as well as in TNBC cell lines and patient-derived cells. Small interfering RNA-based loss-of-function analyses showed that TMPO-AS1 knockdown substantially represses the proliferation and migration of TNBC cells. Expression microarray analysis showed that TMPO-AS1 alters gene signatures related to transforming growth factor-ß signaling in addition to proliferative E2F signaling pathways. TMPO-AS1-targeted siRNA treatment through engineered drug delivery systems using cancer-targeted polyion complex micelle or nanoball technology significantly impaired the in vivo growth of primary and metastatic TNBC xenograft tumors. Our findings suggest that TMPO-AS1 plays a key role in TNBC pathophysiology and could be a potential therapeutic target for TNBC.

Linked Color Imaging and the Kyoto Classification of Gastritis: Evaluation of Visibility and Inter-Rater Reliability.

BACKGROUND/AIMS: To compare white light imaging (WLI) with linked color imaging (LCI) and blue LASER imaging (BLI) in endoscopic findings of Helicobacter pylori presently infected, previously infected, and uninfected gastric mucosae for visibility and inter-rater reliability. METHODS: WLI, LCI and BLI bright mode (BLI-bright) were used to obtain 1,092 endoscopic images from 261 patients according to the Kyoto Classification of Gastritis. Images were evaluated retrospectively by 10 experts and 10 trainee endoscopists and included diffuse redness, spotty redness, map-like redness, patchy redness, red streaks, intestinal metaplasia, and an atrophic border (52 cases for each finding, respectively). Physicians assessed visibility as follows: 5 (improved), 4 (somewhat improved), 3 (equivalent), 2 (somewhat decreased), and 1 (decreased). Visibility was assessed from totaled scores. The inter-rater reliability (intraclass correlation coefficient) was also evaluated. RESULTS: Compared with WLI, all endoscopists reported improved visibility with LCI: 55.8% for diffuse redness; LCI: 38.5% for spotty redness; LCI: 57.7% for map-like redness; LCI: 40.4% for patchy redness; LCI: 53.8% for red streaks; LCI: 42.3% and BLI-bright: 80.8% for intestinal metaplasia; LCI: 46.2% for an atrophic border. For all endoscopists, the inter-rater reliabilities of LCI compared to WLI were 0.73-0.87. CONCLUSION: The visibility of each endoscopic finding was improved by LCI while that of intestinal metaplasia was improved by BLI-bright.

Monitoring method for transgene expression in target tissue by blood sampling.

In this study, we have developed a novel method to monitor transgene expression in tissues by blood sampling. We administered plasmid DNA (pDNA) encoding non-secretory form of firefly luciferase as a reporter gene and pDNA encoding secretable Gaussia princeps luciferase as a monitor gene simultaneously into mice. Good positive correlations were found between log-transgene expression of the reporter gene and the monitor gene in the treated muscle, between the monitor gene in the treated muscle and plasma, and consequently between the reporter gene in the treated muscle and the monitor gene in plasma after naked pDNA transfer into the muscle of mice. Such positive correlations were also found with gastric serosal surface instillation of naked pDNA, intravenous injection of lipoplex, and hydrodynamics-based injection of naked pDNA. We developed monitoring method of transgene expression in tissues by blood sampling, which was named 'Therapeutic transgene monitoring (TTM)', after 'Therapeutic drug monitoring (TDM)'.

Current status of first- and second-line Helicobacter pylori eradication therapy in the metropolitan area: a multicenter study with a large number of patients.

BACKGROUND: The environment surrounding Helicobacter pylori eradication treatment is dramatically changing. Recently, vonoprazan, a first-in-class potassium-competitive acid blocker (P-CAB), was introduced onto the market in 2015. The aging of Japan's demographic structure is becoming pronounced. In this study, we examined the trend of the eradication rate of H. pylori in the metropolitan area and examined factors concerning successful eradication. METHODS: We collected data from 20 hospitals in the Tokyo metropolitan area on patients who received first-line eradication therapy with a proton-pump inhibitor (PPI)/P-CAB, amoxicillin, and clarithromycin for 1 week and second-line eradication therapy with a PPI/P-CAB, amoxicillin, and metronidazole for 1 week from 2013 to 2018. The annual eradication rate and associated factors for successful eradication were analyzed. RESULTS: We collected data of 4097 and 3572 patients in the first- and second-line eradication therapies, respectively. The eradication rate decreased from 2013 to 2014 and increased again from 2015 to 2018 with the first-line therapy [the eradication rates in 2013, 2014, 2015, 2016, 2017 and 2018 were 71.8%, 63.7%, 78.5%, 84.6%, 89.7 and 90.1%, respectively, in the per protocol (PP)]. The second-line eradication rates were 90.0%, 82.6%, 88.8%, 87.5%, 91.8% and 90.1% in 2013, 2014, 2015, 2016, 2017 and 2018, respectively, in PP. Vonoprazan was an independent factor for successful eradication in not only first-line, but also second-line eradication. Age over 75 years was an independent factor for eradication failure in both first- and second-line eradication therapies. CONCLUSION: The eradication rate improved from 2015 to 2018 with the first-line therapy because of the introduction of vonoprazan in the market. The eradication rates with first- and second-line regimens in elderly patients were lower than those in younger patients.