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健常人尿由来N結合型糖鎖の微小不均一性と個体差の解析研究
Get PDF新潟大学Niigata University博士（理学）Urine is considered an outstanding biological fluid for biomarker discovery, reflecting both systemic and urogenital physiology. However, analyzing the N-glycome in urine in detail has been challenging due to the low abundance of glycans attached to glycoproteins compared to free oligosaccharides. Therefore, this study aims to thoroughly analyze urinary N-glycome using LC-MS/MS. The N-glycans were released using hydrazine and labeled with 2-aminopyridine (PA), followed by anion-exchange fractionation before LC-MS/MS analysis. A total of 109 N-glycans were identified and quantified, of which 58 were identified and quantified repeatedly in at least 80% of samples and accounted for approximately 85% of the total urinary glycome signal. Interestingly, a comparison between urine and serum N-glycome revealed that approximately 50% of the urinary glycome could originate from the kidney and urinary tract, where they were exclusively identified in urine, while the remaining 50% were common in both. Additionally, a correlation was found between age/sex and the relative abundances of urinary N-glycome, with more age-related changes observed in women than men. The results of this study provide a reference for human urine N-glycome profiling and structural annotations.Biomolecules. 2023, 13(5), 756.新大院博(理)第491
サステナブルなダイヤモンド電極を用いた,安価な血漿中薬物濃度測定法の創出
Get PDF新潟大学Niigata University博士（医学）On-site monitoring of plasma drug concentrations is required for effective therapies. Recently developed handy biosensors are not yet popular owing to insufficient evaluation of accuracy on clinical samples and the necessity of complicated costly fabrication processes. Here, we approached these bottlenecks via a strategy involving engineeringly unmodified boron-doped diamond (BDD), a sustainable electrochemical material. A sensing system based on a ~1 cm2 BDD chip, when analysing rat plasma spiked with a molecular-targeting anticancer drug, pazo-panib, detected clinically relevant concentrations. The response was stable in 60 sequential measurements on the same chip. In a clinical study, data obtained with a BDD chip were consistent with liquid chromatography–mass spectrometry results. Finally, the portable system with a palm-sized sensor containing the chip analysed ~40 μL of whole blood from dosed rats within ~10 min. This approach with the ‘reusable’ sensor may improve point-of-monitoring systems and personalised medicine while reducing medical costs."On-site monitoring of plasma drug concentrations is required for effective therapies. Recently developed handy biosensors are not yet popular owing to insufficient evaluation of accuracy on clinical samples and the necessity of complicated costly fabrication processes. Here, we approached these bottlenecks via a strategy involving engineeringly unmodified boron-doped diamond (BDD), a sustainable electrochemical material. A sensing system based on a ~1 cm2 BDD chip, when analysing rat plasma spiked with a molecular-targeting anticancer drug, pazo-panib,detected clinically relevant concentrations. The response was stable in 60 sequential measurements on the same chip. In a clinical study, data obtained with a BDD chip were consistent with liquid chromatography–mass spectrometry results. Finally, the portable system with a palm-sized sensor containing the chip nalysed ~40 μL of whole blood from dosed rats within ~10 min. This approach with the ‘reusable’ sensor may improve point-of-monitoring systems and personalised medicine while reducing medical costs."Heliyon. 2023, 9(5), e15963新大院博（医）第1137
Visiumシステムを用いたマウス鼻腔内ワクチン接種後の空間トランスクリプトーム解析への応用
Get PDF新潟大学Niigata University博士（医学）Intranasal vaccines that elicit mucosal immunity are deemed effective against respiratory tract infections such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but their ability to induce humoral immunity characterized by immunoglobulin A (IgA) and IgG production is low. It has been reported that vaccination with a mixture of a viscous base carboxyvinyl polymer (CVP) and viral antigens induced robust systemic and mucosal immune responses. In this study, we analyzed the behavior of immunocompetent cells in the nasal cavity over time by spatial transcriptome profiling induced immediately after antigen vaccination using CVP. We established a method for performing spatial transcriptomics using the Visium system in the mouse nasal cavity and analyzed gene expression profiles within the nasal cavity after intranasal vaccination. Glycoprotein 2 (Gp2)-, SRY-box transcription factor 8 (Sox8)-, or Spi-B transcription factor (Spib)-expressing cells were increased in the nasal passage (NP) region at 3–6 hr after SARS-CoV-2 spike protein and CVP (S-CVP) vaccination. The results suggested that microfold (M) cells are activated within a short period of time (3–6 hr). Subsequent cluster analysis of cells in the nasal cavity showed an increase in Cluster 9 at 3–6 hr after intranasal vaccination with the S-CVP. We found that Il6 in Cluster 9 had the highest log2 fold values within the NP at 3–6 hr. A search for gene expression patterns similar to that 2 of Il6 revealed that the log2 fold values of Edn2, Ccl20, and Hk2 also increased in the nasal cavity after 3–6 hr. The results showed that the early response of immune cells occurred immediately after intranasal vaccination. In this study, we identified changes in gene expression that contribute to the activation of M cells and immunocompetent cells after intranasal vaccination of mice with antigen-CVP using a time-series analysis of spatial transcriptomics data. The results facilitated the identification of the cell types that are activated during the initial induction of nasal mucosal immunity.Frontiers in Immunology. 2023, 14, 1209945.新大院博（医）第1144
日本人集団における自閉症スペクトラム障害および統合失調症とGAP43の再配列決定および関連分析
Get PDF新潟大学Niigata University博士（医学）Background: Growth-associated protein 43 (GAP43), a synaptic protein involved in axonal growth and synaptic plasticity, is implicated in the pathophysiology of autism spectrum disorder (ASD) and schizophrenia. To examine the role of rare GAP43 variants in the genetic etiology of ASD and schizophrenia in a Japanese population, we performed resequencing and association analysis. Methods: First, we resequenced the GAP43 coding region in 295 ASD patients, 323 schizophrenia patients and 304 controls. Second, we genotyped rs561268447 in 273 ASD patients, 1,150 schizophrenia patients and 1,022 controls. Third, we performed an association analysis of rs561268447 in 568 ASD patients, 1,473 schizophrenia patients and 10,127 controls. Results: We identified a rare putatively damaging missense variant (rs561268447) in an ASD patient via resequencing. However, we did not detect the variant in 2,445 individuals via gen-otyping. The variant was not significantly associated with ASD or schizophrenia in the association analysis. Conclusion: This study does not provide evidence for the contribution of rare GAP43 variants to ASD or schizophrenia susceptibility in the Japanese population.Research in Autism Spectrum Disorders. 2021, 82, 101729.新大院博（医）第1154
Evaluation of teaching methods to improve parasite identification skills
Get PDF新潟大学Niigata University博士（医学）公衆衛生の向上に伴い、 日本における土壌媒介寄生虫の感染者数は減少した 。 一方、食品媒介 、性行為や海外旅行に起因する寄生虫症は増加傾向にあり、グローバリゼーションの増加に伴い寄生虫検査の需要増加が予想されている。しかし、国内の医学部医学科では 寄生虫学、教育に費やす時間は減少し 、寄生虫症診断力の低下が懸念されている 。 寄生虫検査を担う臨床検査技師の養成機関においても寄生虫学実習にかける教育時間には差異がある。寄生虫検査は形態鑑別による同定がgold standardであるため 、臨床検査技師の卒前教育の中で鏡検力を習得させておく必要がある 。そのため 、限られた時間 、限られた指導教員で実施可能である新たな教授法を考案し比較評価した 。2017年には相互依存関係の下で協同学習を行う jigsaw 法と学習支援システム （Moodle）を用いた e-learning 法を実施し、2018 年にはこれらを混合した Blended learning 法を実施した 。 Blended learning法は、他の教授法とは異なる年度に実施したため、ランダム化比較試験による評価ができなかった。そのため、本研究では傾向スコアにより学生をマッチングした集団の調整解析によって、Blended learning 法を評価し、その有用性、問題点と改善点を検討した。Blended learning 法は 、jigsaw 法や e-learning 法と比較し寄生虫検査における鏡検力を高めると研究仮説を立てたが 、鏡検試験得点は jigsaw 法 と e-learning 法の中間的な成績となり仮説は支持されなかった。 しかし、鏡検試験において jigsaw 法よりも不合格者を低減させる傾向が示唆された。 また、画像試験においても同様の傾向であったが 、経時的に実施した画像試験結果から Blended learning 法は虫卵鑑別力の維持効果を認め 、短期間であるが現職の臨床検査技師と同定程度の鑑別力を維持した。 Blended learning 法における jigsaw 活動では、事前の情報共有に問題があり、実質的な鏡検時間が不足した。また、Moodle 利用率も約 81％に留まり、全員が反転学習を実施していなかった 。今後、 jigsaw 活動の情報共有における量と内容の統一化、e-learning 教材にゲーミフィケーションを導入するなどの改善策をとることで、 Blended learning 法のさらなる教授法効果が期待される 。新潟医学会雑誌. 2023, 137(9), 319-330.新大院博（医）第1159