MR Virtual Endoscopy for Biliary Tract and Pancreatic Duct

取得学位 : 博士（保健学）, 学位授与番号 : 医博甲第1946号 , 学位授与年月日 : 平成20年3月22日, 学位授与大学 : 金沢大学, 審査結果の報告日 : 平成20年2月13日, 主査 :鈴木 正行 , 副査 :真田 茂, 宮地 利

CRISPR/Cas9-mediated targeted mutagenesis in grape

<div><p>RNA-guided genome editing using the CRISPR/Cas9 CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9) system has been applied successfully in several plant species. However, to date, there are few reports on the use of any of the current genome editing approaches in grape—an important fruit crop with a large market not only for table grapes but also for wine. Here, we report successful targeted mutagenesis in grape (<i>Vitis vinifera</i> L., cv. Neo Muscat) using the CRISPR/Cas9 system. When a Cas9 expression construct was transformed to embryonic calli along with a synthetic sgRNA expression construct targeting the <i>Vitis vinifera</i> phytoene desaturase (VvPDS) gene, regenerated plants with albino leaves were obtained. DNA sequencing confirmed that the VvPDS gene was mutated at the target site in regenerated grape plants. Interestingly, the ratio of mutated cells was higher in lower, older, leaves compared to that in newly appearing upper leaves. This result might suggest either that the proportion of targeted mutagenized cells is higher in older leaves due to the repeated induction of DNA double strand breaks (DSBs), or that the efficiency of precise DSBs repair in cells of old grape leaves is decreased.</p></div

Dibothriocephalus nihonkaiensis infection identified by pathological and genetic analyses -a case report and a recent literature review of human diphyllobothriasis

We report a 20-year-old male with a past history of diarrhea and discharge of a strobila of a broad tapeworm from his anus 3 months ago. He again noticed passing a tapeworm strobila and he visited our hospital with the tapeworm sample. The macroscopic appearance and the pathological features strongly suggested Diphyllobothriasis. Soon after the first visit to our hospital, a capsule endoscopy was performed with the result of no evidence of strobilae or scoleces. Praziquantel at 20 mg/kg was orally administered once. Before drug administration, egg-like structures were present in the stool. They disappeared soon after drug administration. In human Diphyllobothriasis, Dibothriocephalus nihonkaiensis (D. nihonkaiensis) infection and Dibothriocephalus latus (D. latus) infection are common. These two species are morphologically similar but genetically distinct. Accordingly, polymerase chain reaction (PCR) using genomic DNA of the tapeworm infecting our case was performed for amplifying mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. The amplified DNA fragment by PCR of our case was phylogenetically compared with other mitochondrial cox1 gene sequences of D. nihonkaiensis, D. latus, D. dendriticus, Taenia saginata, Taenia solium and Spirometra erinaceieuropaei. As a result, the tapeworm of our case was most likely within the spectrum of D. nihonkaiensis. Although human diphyllobothriasis is not a life-threatening disease for human beings, it has become a re-emerging problem even in the most developed countries with the increasing popularity of eating raw fish. The knowledge of how to cook or freeze raw fish before eating can reduce the risk of infection. However, without general awareness of human parasite infection, food-borne infections will continue to be a public health problem. Keywords: Tapeworm, Dibothriocephalus nihonkaiensis, Dibothriocephalus latus, Mitochondrial cytochrome c oxidase subunit 1 gene, Praziquante

Emergence rate of mutated plants via embryo induction method.

<p>Emergence rate of mutated plants via embryo induction method.</p

Primer sequences.

<p>Primer sequences.</p

Molecular characterization of Helicobacter pylori VacA induction of IL-8 in U937 cells reveals a prominent role for p38MAPK in activating transcription factor-2, cAMP response element binding protein, and NF-kappaB activation.

Helicobacter pylori VacA induces multiple effects on susceptible cells, including vacuolation, mitochondrial damage, inhibition of cell growth, and enhanced cyclooxygenase-2 expression. To assess the ability of H. pylori to modulate the production of inflammatory mediators, we examined the mechanisms by which VacA enhanced IL-8 production by promonocytic U937 cells, which demonstrated the greatest VacA-induced IL-8 release of the cells tested. Inhibitors of p38 MAPK (SB203580), ERK1/2 (PD98059), IκBα ((E)-3-(4-methylphenylsulfonyl)-2-propenenitrile), Ca(2+) entry (SKF96365), and intracellular Ca(2+) channels (dantrolene) blocked VacA-induced IL-8 production. Furthermore, an intracellular Ca(2+) chelator (BAPTA-AM), which inhibited VacA-activated p38 MAPK, caused a dose-dependent reduction in VacA-induced IL-8 secretion by U937 cells, implying a role for intracellular Ca(2+) in mediating activation of MAPK and the canonical NF-κB pathway. VacA stimulated translocation of NF-κBp65 to the nucleus, consistent with enhancement of IL-8 expression by activation of the NF-κB pathway. In addition, small interfering RNA of activating transcription factor (ATF)-2 or CREB, which is a p38MAPK substrate and binds to the AP-1 site of the IL-8 promoter, inhibited VacA-induced IL-8 production. VacA activated an IL-8 promoter containing an NF-IL-6 site, but not a mutated AP-1 or NF-κB site, suggesting direct involvement of the ATF-2/CREB binding region or NF-κB-binding regions in VacA-induced IL-8 promoter activation. Thus, in U937 cells, VacA directly increases IL-8 production by activation of the p38 MAPK via intracellular Ca(2+) release, leading to activation of the transcription factors, ATF-2, CREB, and NF-κB

Detection of Cas9 expression in regenerated plants.

<p>Immunological detection of Cas9. PDS-t2-1^st-#1–2 and PDS-t2-1^st-#1–3 are regenerated plants obtained from same transgenic line.</p

Detection of mutations in PDS-t2 target locus in regenerated plant.

<p>(a) Chlorophyll-deficient variegated plant due to mutation in the PDS-t2 target locus. (b) Representative sequences of PDS-t2 target locus in leaves. The wild type sequence is shown at the top, with the PAM sequence highlighted in cyan, and the target sequence in red. Dashes, deleted bases. The net change in length is noted to the right of each sequence (+, insertion;—deletion). The number of clones representing each mutant allele is shown in the column on the right.</p