CRISPR/Cas9-mediated disruption of CjACOS5 confers no-pollen formation on sugi trees (Cryptomeria japonica D. Don).

Sugi (Cryptomeria japonica D. Don) is an economically important coniferous tree in Japan. However, abundant sugi pollen grains are dispersed and transported by the wind each spring and cause a severe pollen allergy syndrome (Japanese cedar pollinosis). The use of pollen-free sugi that cannot produce pollen has been thought as a countermeasure to Japanese cedar pollinosis. The sugi CjACOS5 gene is an ortholog of Arabidopsis ACOS5 and rice OsACOS12, which encode an acyl-CoA synthetase that is involved in the synthesis of sporopollenin in pollen walls. To generate pollen-free sugi, we mutated CjACOS5 using the CRISPR/Cas9 system. As a result of sugi transformation mediated by Agrobacterium tumefaciens harboring the CjACOS5-targeted CRISPR/Cas9 vector, 1 bp-deleted homo biallelic mutant lines were obtained. Chimeric mutant lines harboring both mutant and wild-type CjACOS5 genes were also generated. The homo biallelic mutant lines had no-pollen in male strobili, whereas chimeric mutant lines had male strobili with or without pollen grains. Our results suggest that CjACOS5 is essential for the production of pollen in sugi and that its disruption is useful for the generation of pollen-free sugi. In addition to conventional transgenic technology, genome editing technology, including CRISPR/Cas9, can confer new traits on sugi.

Targeted deletion of grape retrotransposon associated with fruit skin color via CRISPR/Cas9 in Vitis labrascana 'Shine Muscat'.

Transposition of transposable elements affect expression levels, splicing and epigenetic status, and function of genes located in, or near, the inserted/excised locus. For example, in grape, presence of the Gret1 retrotransposon in the promoter region of the VvMYBA1a allele at the VvMYBA1 locus suppress the expression of the VvMYBA1 transcription factor gene for the anthocyanin biosynthesis and this transposon insertion is responsible for the green berry skin color of Vitis labrascana, 'Shine Muscat', a major grape cultivar in Japan. To prove that transposons in grape genome can be removed by genome editing, we focused on Gret1 in the VvMYBA1a allele as a target of CRISPR/Cas9 mediated transposon removal. PCR amplification and sequencing detected Gret1 eliminated cells in 19 of 45 transgenic plants. Although we have not yet confirmed any effects on grape berry skin color, we were successful in demonstrating that cleaving the long terminal repeat (LTR) present at both ends of Gret1 can efficiently eliminate the transposon.

A MAPKKK gene from rice, RBG1res, confers resistance to Burkholderia glumae through negative regulation of ABA.

Burkholderia glumae causes bacterial seedling rot (BSR) of rice and is a threat to a consistent food supply. When previously screening for resistance against B. glumae in the resistant cultivar Nona Bokra (NB) versus the susceptible cultivar Koshihikari (KO), we detected a gene, Resistance to Burkholderia glumae 1 (RBG1), at a quantitative trait locus (QTL). Here, we found that RBG1 encodes a MAPKKK gene whose product phosphorylates OsMKK3. We also found that the kinase encoded by the RBG1 resistant (RBG1res) allele in NB presented higher activity than did that encoded by the RBG1 susceptible (RBG1sus) allele in KO. RBG1res and RBG1sus differ by three single-nucleotide polymorphisms (SNPs), and the G390T substitution is essential for kinase activity. Abscisic acid (ABA) treatment of inoculated seedlings of RBG1res-NIL (a near-isogenic line (NIL) expressing RBG1res in the KO genetic background) decreased BSR resistance, indicating that RBG1res conferred resistance to B. glumae through negative regulation of ABA. The results of further inoculation assays showed that RBG1res-NIL was also resistant to Burkholderia plantarii. Our findings suggest that RBG1res contributes to resistance to these bacterial pathogens at the seed germination stage via a unique mechanism.

Genome editing by introduction of Cas9/sgRNA into plant cells using temperature-controlled atmospheric pressure plasma.

Previously, we developed a technique to introduce a superfolder green fluorescent protein (sGFP) fusion protein directly into plant cells using atmospheric-pressure plasma. In this study, we attempted genome editing using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9) system using this protein introduction technique. As an experimental system to evaluate genome editing, we utilized transgenic reporter plants carrying the reporter genes L-(I-SceI)-UC and sGFP-waxy-HPT. The L-(I-SceI)-UC system allowed the detection of successful genome editing by measuring the chemiluminescent signal observed upon re-functionalization of the luciferase (LUC) gene following genome editing. Similarly, the sGFP-waxy-HPT system conferred hygromycin resistance caused by hygromycin phosphotransferase (HPT) during genome editing. CRISPR/Cas9 ribonucleoproteins targeting these reporter genes were directly introduced into rice calli or tobacco leaf pieces after treatment with N2 and/or CO2 plasma. Cultivation of the treated rice calli on a suitable medium plate produced the luminescence signal, which was not observed in the negative control. Four types of genome-edited sequences were obtained upon sequencing the reporter genes of genome-edited candidate calli. sGFP-waxy-HPT-carrying tobacco cells exhibited hygromycin resistance during genome editing. After repeated cultivation of the treated tobacco leaf pieces on a regeneration medium plate, the calli were observed with leaf pieces. A green callus that was hygromycin-resistant was harvested, and a genome-edited sequence in the tobacco reporter gene was confirmed. As direct introduction of the Cas9/sgRNA (single guide RNA) complex using plasma enables genome editing in plants without any DNA introduction, this method is expected to be optimized for many plant species and may be widely applied for plant breeding in the future.

Current status and perspectives for endoscopic diagnosis of superficial nonampullary duodenal epithelial tumors.

In recent years, there have been significant advances in the endoscopic resection (ER) procedures of superficial nonampullary duodenal epithelial tumors (SNADETs). A preoperative endoscopic diagnosis is thus deemed necessary in determining the indication for subsequent ER. For the histologic and endoscopic diagnosis of SNADETs, understanding the mucin phenotype is inevitable. Recently, two diagnostic algorithms for the differential diagnosis of SNADETs from nonneoplastic lesions under magnifying endoscopy with narrow-band imaging have been proposed. In addition, various endoscopic approaches have been proposed to differentiate low- and high-grade adenomas/carcinomas, including white light endoscopy, magnifying image-enhanced endoscopy, and endocytoscopy. These methods, however, have not been standardized with respect to the classification of their findings and the validation of their diagnostic accuracy. Moreover, there are still concerns with respect to the histologic criteria required to establish a SNADETs diagnosis. Standardization in the histologic and endoscopic diagnosis of SNADETs is needed.

Oxicam-type nonsteroidal anti-inflammatory drugs enhance Agrobacterium-mediated transient transformation in plants.

Agrobacterium-mediated transformation is a key innovation for plant breeding, and routinely used in basic researches and applied biology. However, the transformation efficiency is often the limiting factor of this technique. In this study, we discovered that oxicam-type nonsteroidal anti-inflammatory drugs, including tenoxicam (TNX), increase the efficiency of Agrobacterium-mediated transient transformation. TNX treatment increased the transformation efficiency of Agrobacterium-mediated transformation of Arabidopsis thaliana mature leaves by agroinfiltration. The increase of efficiency by TNX treatment was not observed in dde2/ein2/pad4/sid2 quadruple mutant, indicating that TNX inhibits the immune system mediated by jasmonic acid, ethylene, and salicylic acid against to Agrobacterium. We also found that TNX-treatment is applicable for the transient expression and subcellular localization analysis of fluorescent-tagged proteins in Arabidopsis leaf cells. In addition, we found that TNX increases the efficiency of Agrobacterium-mediated transient transformation of Jatropha. Given that treatment with oxicam compounds is a simple and cost effective method, our findings will provide a new option to overcome limitations associated with Agrobacterium-mediated transformation of various plant species.

Regulation of germination by targeted mutagenesis of grain dormancy genes in barley.

High humidity during harvest season often causes pre-harvest sprouting in barley (Hordeum vulgare). Prolonged grain dormancy prevents pre-harvest sprouting; however, extended dormancy can interfere with malt production and uniform germination upon sowing. In this study, we used Cas9-induced targeted mutagenesis to create single and double mutants in QTL FOR SEED DORMANCY 1 (Qsd1) and Qsd2 in the same genetic background. We performed germination assays in independent qsd1 and qsd2 single mutants, as well as in two double mutants, which revealed a strong repression of germination in the mutants. These results demonstrated that normal early grain germination requires both Qsd1 and Qsd2 function. However, germination of qsd1 was promoted by treatment with 3% hydrogen peroxide, supporting the notion that the mutants exhibit delayed germination. Likewise, exposure to cold temperatures largely alleviated the block of germination in the single and double mutants. Notably, qsd1 mutants partially suppress the long dormancy phenotype of qsd2, while qsd2 mutant grains failed to germinate in the light, but not in the dark. Consistent with the delay in germination, abscisic acid accumulated in all mutants relative to the wild type, but abscisic acid levels cannot maintain long-term dormancy and only delay germination. Elucidation of mutant allele interactions, such as those shown in this study, are important for fine-tuning traits that will lead to the design of grain dormancy through combinations of mutant alleles. Thus, these mutants will provide the necessary germplasm to study grain dormancy and germination in barley.

The mucin phenotype does not affect the endoscopic resection outcome of non-ampullary duodenal epithelial tumors.

Background and study aims Some studies have reported an association between clinicopathological features and mucin phenotypes of non-ampullary duodenal epithelial tumors (NADETs). However, the association between clinical outcomes of endoscopic resection (ER) and mucin phenotypes has not been elucidated. The aim of this retrospective study was to analyze clinical outcomes of ER of NADETs with reference to mucin phenotypes. Patients and methods We retrospectively evaluated the clinical outcomes of ER for NADETs performed from 2006 to 2019 and compared clinicopathological characteristics, ER procedures, and outcomes, including adverse events (AEs) among tumors classified by mucin phenotype. Mucin phenotypes were classified as gastric, gastrointestinal, and intestinal based on immunohistochemical examination. Grade of dysplasia was determined according to the Vienna classification (VCL). Results The proportion of VCL 4/5 was higher in the gastric type (50 %) compared with that in the gastrointestinal (39.1 %, P  = 0.009) and intestinal types (5.4 %, P  = 0.008), respectively. With no statistical difference in tumor size and ER method among the three groups, no significant difference was observed for ER outcomes, i. e., en bloc and R0 resection rates. In the gastrointestinal and intestinal types, AEs occurred in four cases treated with ESD, but none developed in the gastric type. Conclusions This study suggests that the mucin phenotype does not affect resection outcome. However, considering high malignant potential and tendency for low AE rates, the gastric type NADETs may be more appropriate for proactive ER than the others.

CRISPR/Cas9-mediated targeted mutagenesis in Japanese cedar (Cryptomeria japonica D. Don).

Cryptomeria japonica (Japanese cedar or sugi) is one of the most important coniferous tree species in Japan and breeding programs for this species have been launched since 1950s. Genome editing technology can be used to shorten the breeding period. In this study, we performed targeted mutagenesis using the CRISPR/Cas9 system in C. japonica. First, the CRISPR/Cas9 system was tested using green fluorescent protein (GFP)-expressing transgenic embryogenic tissue lines. Knock-out efficiency of GFP ranged from 3.1 to 41.4% depending on U6 promoters and target sequences. The GFP knock-out region was mottled in many lines, indicating genome editing in individual cells. However, in 101 of 102 mutated individuals (> 99%) from 6 GFP knock-out lines, embryos had a single mutation pattern. Next, we knocked out the endogenous C. japonica magnesium chelatase subunit I (CjChlI) gene using two guide RNA targets. Green, pale green, and albino phenotypes were obtained in the gene-edited cell lines. Sequence analysis revealed random deletions, insertions, and replacements in the target region. Thus, targeted mutagenesis using the CRISPR/Cas9 system can be used to modify the C. japonica genome.

Prognostic nutritional index is an independent prognostic factor for older patients aged ≥ 85 years treated by gastric endoscopic submucosal dissection.

BACKGROUND: Clinical outcomes and prognostic factors for survival after endoscopic submucosal dissection (ESD) in older patients aged ≥ 85 years with early gastric cancer (EGC) are not well defined. The aim of this study was to investigate the clinical outcomes and prognostic factors for survival after ESD in older patients aged ≥ 85 years with EGC. METHODS: Clinical outcomes of 70 patients aged ≥ 85 years with EGC treated with ESD were evaluated retrospectively. Prognostic factors for overall survival (OS) were analyzed with the Kaplan-Meier method and a Cox proportional hazards model. RESULTS: During the follow-up period, 33 patients died from any cause, none of whom died from gastric cancer. OS probability after 3 years was 90.0%. Univariate analyses revealed that a neutrophil/lymphocyte ratio ≥ 2.6, a prognostic nutritional index (PNI) < 42.5 and low serum albumin value (< 3.5 g/dl) were associated with poor OS. Cox multivariate analysis revealed low PNI (< 42.5) to be an independent prognostic factor associated with OS (hazard ratio; 3.40, 95% confidence interval; 1.47-7.86, P = 0.004). CONCLUSIONS: PNI may be a useful parameter for making the decision to perform ESD for older patients aged ≥ 85 years with EGC.

Resectability of underwater endoscopic mucosal resection for duodenal tumor: A single-center, retrospective pilot study.

BACKGROUND AND AIM: Underwater endoscopic mucosal resection (U-EMR) has been attracting much attention as treatment for patients with nonampullary duodenal epithelial tumors (NADETs). We aim to compare treatment outcomes, including submucosal resectability, between patients undergoing U-EMR and conventional endoscopic mucosal resection (C-EMR) for NADET. METHODS: We conducted a retrospective review of 38 patients with NADET treated by U-EMR or C-EMR. In the resected specimens, we measured the horizontal length, the vertical distance from the muscularis mucosa to the margin at the deepest site, and the overall submucosal area. The submucosal index (SMI) was defined as the overall submucosal area divided by the largest horizontal length. These values and other treatment outcomes were compared between NADETs resected by U-EMR and C-EMR. RESULTS: The median size of lesions was 7 mm with a range of 3-13 mm. Although the incidence of adverse events and the rates of en bloc and R0 resection were not different in the two groups, the median procedure time was significantly shorter in the U-EMR group (11 min vs 13 min; P = 0.045). The median submucosal depth at the deepest site (1.22 mm vs 1.08 mm; P = 0.38) and the median SMI (0.44 vs 0.41; P = 0.42) were not different between groups. CONCLUSIONS: The resectability between NADETs treated by U-EMR and C-EMR was comparable. These results, together with the shorter procedure time required for U-EMR, suggest that U-EMR may have the potential to be the first choice for small to medium-sized NADET.

In planta Genome Editing in Commercial Wheat Varieties.

Limitations for the application of genome editing technologies on elite wheat (Triticum aestivum L.) varieties are mainly due to the dependency on in vitro culture and regeneration capabilities. Recently, we developed an in planta particle bombardment (iPB) method which has increased process efficiency since no culture steps are required to create stably genome-edited wheat plants. Here, we report the application of the iPB method to commercially relevant Japanese elite wheat varieties. The biolistic delivery of gold particles coated with plasmids expressing CRISPR/Cas9 components designed to target TaQsd1 were bombarded into the embryos of imbibed seeds with their shoot apical meristem (SAM) exposed. Mutations in the target gene were subsequently analyzed within flag leaf tissue by using cleaved amplified polymorphic sequence (CAPS) analysis. A total of 9/358 (2.51%) of the bombarded plants (cv. "Haruyokoi," spring type) carried mutant alleles in the tissue. Due to the chimeric nature of the T0 plants, only six of them were inherited to the next (T1) generation. Genotypic analysis of the T2 plants revealed a single triple-recessive homozygous mutant of the TaQsd1 gene. Compared to wild type, the homozygous mutant exhibited a 7 days delay in the time required for 50% seed germination. The iPB method was also applied to two elite winter cultivars, "Yumechikara" and "Kitanokaori," which resulted in successful genome editing at slightly lower efficiencies as compared to "Haruyokoi." Taken together, this report demonstrates that the in planta genome editing method through SAM bombardment can be applicable to elite wheat varieties that are otherwise reluctant to callus culture.

Mutation of the imprinted gene OsEMF2a induces autonomous endosperm development and delayed cellularization in rice.

In angiosperms, endosperm development comprises a series of developmental transitions controlled by genetic and epigenetic mechanisms that are initiated after double fertilization. Polycomb repressive complex 2 (PRC2) is a key component of these mechanisms that mediate histone H3 lysine 27 trimethylation (H3K27me3); the action of PRC2 is well described in Arabidopsis thaliana but remains uncertain in cereals. In this study, we demonstrate that mutation of the rice (Oryza sativa) gene EMBRYONIC FLOWER2a (OsEMF2a), encoding a zinc-finger containing component of PRC2, causes an autonomous endosperm phenotype involving proliferation of the central cell nuclei with separate cytoplasmic domains, even in the absence of fertilization. Detailed cytological and transcriptomic analyses revealed that the autonomous endosperm can produce storage compounds, starch granules, and protein bodies specific to the endosperm. These events have not been reported in Arabidopsis. After fertilization, we observed an abnormally delayed developmental transition in the endosperm. Transcriptome and H3K27me3 ChIP-seq analyses using endosperm from the emf2a mutant identified downstream targets of PRC2. These included >100 transcription factor genes such as type-I MADS-box genes, which are likely required for endosperm development. Our results demonstrate that OsEMF2a-containing PRC2 controls endosperm developmental programs before and after fertilization.

Site-directed mutagenesis by biolistic transformation efficiently generates inheritable mutations in a targeted locus in soybean somatic embryos and transgene-free descendants in the T1 generation.

The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated endonuclease 9 (Cas9) system is being rapidly developed for mutagenesis in higher plants. Ideally, foreign DNA introduced by this system is removed in the breeding of edible crops and vegetables. Here, we report an efficient generation of Cas9-free mutants lacking an allergenic gene, Gly m Bd 30K, using biolistic transformation and the CRISPR/Cas9 system. Five transgenic embryo lines were selected on the basis of hygromycin resistance. Cleaved amplified polymorphic sequence analysis detected only two different mutations in e all of the lines. These results indicate that mutations were induced in the target gene immediately after the delivery of the exogenous gene into the embryo cells. Soybean plantlets (T0 plants) were regenerated from two of the transgenic embryo lines. The segregation pattern of the Cas9 gene in the T1 generation, which included Cas9-free plants, revealed that a single copy number of transgene was integrated in both lines. Immunoblot analysis demonstrated that no Gly m Bd 30K protein accumulated in the Cas9-free plants. Gene expression analysis indicated that nonsense mRNA decay might have occurred in mature mutant seeds. Due to the efficient induction of inheritable mutations and the low integrated transgene copy number in the T0 plants, we could remove foreign DNA easily by genetic segregation in the T1 generation. Our results demonstrate that biolistic transformation of soybean embryos is useful for CRISPR/Cas9-mediated site-directed mutagenesis of soybean for human consumption.

Investigation of clomazone-tolerance mechanism in a long-grain cultivar of rice.

BACKGROUND: Clomazone is a potent herbicide for controlling weeds that have evolved resistance to other herbicides due to its unique mode of action. Clomazone is used in rice cultivation, but is limited to long-grain cultivars because other cultivars are highly sensitive to it. In this study, we investigated the mechanism of clomazone tolerance in a long-grain cultivar. RESULTS: The long-grain cultivar Kasalath tolerated approximately five-fold higher doses of clomazone compared to two short-grain cultivars, Nipponbare and Koshihikari. While Arabidopsis thaliana transformed with a rice cytochrome P450, CYP81A6, showed resistance to clomazone, the cyp81a6 knockout Kasalath was unchanged in its clomazone sensitivity. The inheritance of clomazone sensitivity in the F1 and F2 of Kasalath and Nipponbare indicated the involvement of multiple loci for clomazone tolerance. Four chromosome segment substitution lines of Nipponbare/Kasalath and Koshihikari/Kasalath exhibited partial tolerance to clomazone. The overlapping DNA region among the four lines is on chromosome 5 within 11.5 Mb. CONCLUSION: Multiple loci are involved in clomazone tolerance in Kasalath, one of which is located on chromosome 5. This information will help develop short-grain cultivars tolerant to clomazone. © 2021 Society of Chemical Industry.

Immunohistochemical Analysis of Mismatch Repair Gene Proteins in Early Gastric Cancer Based on Microsatellite Status.

BACKGROUND: Microsatellite instability (MSI) is a major pathway involved in gastric carcinogenesis and is observed in 10-20% of early gastric cancers (EGCs). Early detection of EGCs with an MSI-high phenotype would be useful for elucidating the mechanisms of gastric carcinogenesis and improving outcomes in patients with GC. OBJECTIVE: We explored the usefulness of immunohistochemical expression of mismatch repair (MMR) proteins, including MLH1, PMS2, MSH2, and MSH6 in EGC. METHODS: We examined the expression of 4 MMR proteins using immunohistochemistry in 119 patients with EGC based on MS status, as determined by polymerase chain reaction-microsatellite analysis. In addition, methylation of the MLH1 gene was quantified by pyrosequencing. RESULTS: EGCs were classified into 46 MSI-high phenotypes and 73 microsatellite stable (MSS) phenotypes. Although loss of MLH1 expression was associated with loss of PMS2 expression in the MSI-high phenotype, discordant cases of loss of expression between MLH1 and PMS2 were found (MLH1 [-]/PMS2 [+], 3 cases). Loss of MLH1/PMS2 expression was observed in 2 of 73 MSS phenotypes. Loss of MSH2/MSH6 expression was found in 4 of 46 MSI-high phenotypes, whereas loss of MSH2/MSH6 expression was not detected in the MSS phenotype. In addition, loss of MLH1 expression was correlated with methylation of MLH1. However, there were discordant cases in which loss of MLH1 expression was not accompanied by methylation of MLH1. CONCLUSION: Although immunostaining of MMR proteins could help predict MSI in EGCs, immunostaining did not have the same value as genetic testing for determination of MSI.

Risk for lymph node metastasis in Epstein-Barr virus-associated gastric carcinoma with submucosal invasion.

OBJECTIVES: Epstein-Barr virus-associated gastric cancer (EBVGC) has been reported to be associated with a low risk for lymph node metastasis (LNM). However, the curative criteria for endoscopic submucosal dissection (ESD) for submucosal EBVGC (pT1b-EBVGC) remain unclear. Our study aimed to investigate the risk factors for LNM in pT1b-EBVGC. METHODS: This was a retrospective multicenter study at five institutes in Japan. We reviewed medical records and extracted all pT1b-EBVGC cases that met the following criteria: (i) histologically proven submucosal gastric cancer; (ii) surgical or endoscopic resection between January 2000 and December 2016; and (iii) presence of Epstein-Barr virus (EBV) in tumor cells verified by EBV-encoded small RNA in situ hybridization (EBER-ISH). The association between clinicopathological factors and LNM were assessed using multivariable logistic regression analysis. RESULTS: A total of 185 pT1b-EBVGC cases were included in the analysis. LNM was found in nine cases (4.9%). Multivariable logistic regression analysis demonstrated that lymphatic invasion (OR 9.1; 95% CI 2.1-46.1) and submucosal invasion ≥4000 µm (OR 9.2; 95% CI 1.3-110.3) were significant risk factors for LNM. When we focused on pT1b-EBVGC without lymphatic invasion and with submucosal invasion <2000 µm, the rate of LNM was 0% (0/96, 95% CI 0-3.8%). CONCLUSIONS: Our findings indicated that lymphatic invasion and submucosal invasion ≥4000 µm were significant risk factors for LNM. ESD could be an appropriate option for pT1b-EBVGC without lymphatic invasion and with submucosal invasion <2000 µm.

Precision genome editing in plants via gene targeting and subsequent break-induced single-strand annealing.

Genome editing via artificial nucleases such as CRISPR/Cas9 has become popular in plants now. However, small insertions or deletions are major mutations and nucleotide substitutions rarely occur when DNA cleavage is induced. To induce nucleotide substitutions, a base editor utilizing dead or nickase-type Cas9 fused with deaminase have been developed. However, the direction and position of practical substitution are still limited. In this context, homologous recombination (HR)-mediated gene targeting (GT) has advantages because any mutations existing on the donor DNA are copied and passed onto the endogenous DNA. As HR-mediated GT is extremely rare in higher plants, positive-negative selection has been used to isolate cells in which GT has occurred. After successful selection, positive selection marker is no longer needed and should ideally be eliminated. In a previous study, we reported a seamless piggyBac-transposon-mediated marker elimination system. Precision marker elimination efficiency in this system is very high. The piggyBac transposon integrates into the host genome at TTAA elements and excises without leaving a footprint at the excised site, so a TTAA sequence is necessary at the location of a positive selection marker. To compensate for this limitation, we have developed a novel marker elimination system using an I-SceI break and subsequent single-strand annealing (SSA)-mediated DNA repair system.

Protective effect of proton pump inhibitors and potassium competitive acid blockers against post-gastric endoscopic submucosal dissection bleeding: a single-center, propensity score-matched analysis.

OBJECTIVES: Both potassium-competitive acid blockers (P-CABs) and proton pump inhibitors (PPIs) are known to be protective against bleeding after gastric endoscopic dissection (ESD) for early gastric cancers. The aim was to compare the effect of PPI and P-CAB treatment against bleeding after gastric ESD. MATERIALS AND METHODS: This was a single-center, retrospective analysis. Among 541 patients who underwent gastric ESD during the period from 2014 to 2019, we recruited subjects who were treated with PPIs (intravenous lansoprazole followed by oral esomeprazole) or a P-CAB before and after ESD. The incidence of post-ESD bleeding was compared between treatment groups. The risks associated with post-ESD bleeding were examined by univariate and multivariate analyses after propensity score-matching. RESULTS: The overall incidence of post-ESD bleeding was not significantly different between patients treated with PPIs (n = 362) and those treated with a P-CAB (n = 156) (3.0% vs 2.6%, respectively; p = .77). Even after propensity score matching (n = 153 in each group), the incidence was not significantly different between groups (2.6% vs 2.6%, respectively; p = 1.00). A multivariate analysis revealed that antithrombotic therapy (OR 4.85, 95% CI 1.14-20.57) was an independent factor associated with post-ESD bleeding. CONCLUSIONS: The incidence of post gastric ESD bleeding is not different between patients treated with PPI and patients treated with P-CAB. Antithrombotic therapy is an independent risk factor associated with post-ESD bleeding.