A STRIPAK component Strip regulates neuronal morphogenesis by affecting microtubule stability

During neural development, regulation of microtubule stability is essential for proper morphogenesis of neurons. Recently, the striatin-interacting phosphatase and kinase (STRIPAK) complex was revealed to be involved in diverse cellular processes. However, there is little evidence that STRIPAK components regulate microtubule dynamics, especially in vivo. Here, we show that one of the core STRIPAK components, Strip, is required for microtubule organization during neuronal morphogenesis. Knockdown of Strip causes a decrease in the level of acetylated α-tubulin in Drosophila S2 cells, suggesting that Strip influences the stability of microtubules. We also found that Strip physically and genetically interacts with tubulin folding cofactor D (TBCD), an essential regulator of α- and β-tubulin heterodimers. Furthermore, we demonstrate the genetic interaction between strip and Down syndrome cell adhesion molecule (Dscam), a cell surface molecule that is known to work with TBCD. Thus, we propose that Strip regulates neuronal morphogenesis by affecting microtubule stability.This work was supported by grants from the Japanese Ministry of Education, Science, Sports, Culture and Technology (MEXT), the Japan Society for the Promotion of Science and the Japan Science and Technology Agency (to C.S., M.O., M.M. and T.C.)

Effects of anti-inflammatory steroid on the osteogenic process of tissue-engineered bone

【目的】歯科領域においては， 自家骨移植や人工骨による骨再生が行われてきたが， 近年細胞を用いた骨再生治療の実用化が期待されている． これまでの臨床研究から， 骨再生治療の安全性や有効性が示されてきたが， その一方で骨再生のメカニズムには不明な点も多い． 近年， 移植部位における炎症が骨再生を阻害する可能性が報告されている． 実際の細胞による骨再生治療の効果を最大化するためには， 局所の炎症が骨再生に与える影響の詳細を理解する必要がある． そこで， 本研究では免疫正常マウスを用いた骨形成モデルを確立した． さらに， 同マウスモデルに対して抗炎症作用を持つステロイド系抗炎症薬を投与し， その影響についても検討を行った．【材料と方法】マウスの骨髄由来間葉系幹細胞は安定した培養が困難であることから， われわれは皮質骨由来の細胞に着目した． BALB/cAJc1 の大腿骨および脛骨の皮質骨より細胞を採取し， 接着性細胞の培養を行った． この細胞をデキサメタゾン， β グリセロリン酸，アスコルビン酸およびBMP-2 を含む分化誘導培地で培養後， アルカリフォスファターゼ(ALP)活性を測定し， 骨分化を確認した． 次に， 培養細胞を担体であるβ-TCP 顆粒上に播種し， 分化誘導を行った． 得られた細胞-担体複合体(培養骨)を同系マウスの背部皮下へ移植した． 骨形成過程に対するステロイド系抗炎症薬の影響について検討するために， 実験群へは培養骨移植直後に1 回と翌日に2 回ベタメタゾンリン酸エステルナトリウムの腹腔内投与を行った． 対照群へは， 同量の生理食塩水を腹腔内に投与した． 移植3， 7 および28 日目に培養骨を摘出し， HE 染色， TRAP 染色および免疫染色（抗F4/80 抗体， 抗TNF-α 抗体， 抗Sp-7 抗体）により比較検討を行った． また， サンプルの一部は液体窒素中で直ちに凍結し， RNA を抽出後定量的PCR にてTNF-α の遺伝子発現を解析した．【結果】培養された接着性の細胞は， 間葉系幹細胞のマーカーであるCD29， CD105 およびSca-1 を発現していた． 得られた細胞を骨分化誘導培地で培養を行ったところ，ALP 活性が上昇した． さらに，免疫不全マウス（BALB/cAJc1-nu/nu）背部皮下へ移植後に骨形成が認められたことから， 骨芽細胞様の細胞が得られていることを確認した． 経時的に摘出されたサンプルの所見では， 培養骨移植後に炎症性細胞浸潤が認められ， 特に3 日目では対照群において炎症性細胞の浸潤が顕著であった．F4/80 陽性細胞は細胞の有無にかかわらず担体周囲に認められたため， 移植された担体に対するマクロファージの浸潤と考えられた． F4/80 陽性細胞数を面積比で検討2したところ， 培養骨移植7 日目において， 実験群では少ない傾向であったが， それ以降ではステロイド系抗炎症薬投与の有無による差は認められなかった． TNF-α 陽性細胞は， 移植7 日目から担体周囲に見られ， 28 日目にも局在していた． しかしながら， ステロイド系抗炎症薬投与の有無による差は認められなかった． TRAP 染色の結果では， 実験群では7 日目より陽性細胞がわずかに認められ， 28 日にかけて増加した． 一方，対照群では7日目には認められず， 28 日には実験群と同程度の陽性細胞数となっていた． Sp7 陽性細胞は， 実験群， 対照群ともに培養骨移植3 日目から認められ， その後形成された骨周囲で強陽性の細胞が出現した． しかしながら， ステロイド系抗炎症薬投与の有無による分布や発現強度の差は認められなかった．【考察】マウス下腿の皮質骨より得られた細胞は， 骨髄間質細胞と比較して安定した増殖が得られた． また， 間葉系幹細胞マーカーを発現するとともに， 分化誘導によってALP 活性が上昇することから， マウスにおける骨芽細胞の細胞源として有用と考えられた． この細胞を用いることで， 免疫正常マウスを用いた骨形成モデルの作製が可能であった． これまでの研究から， 免疫正常マウスでは， 骨形成過程の初期に起こる炎症性サイトカインの上昇によって骨形成が抑制されることが示唆されている． 培養骨移植の初期には， 移植操作による炎症や移植された担体に対する生体反応として，マクロファージなど炎症性細胞の浸潤が認められた． ステロイド系抗炎症薬の投与によって， 初期の炎症性細胞の浸潤が軽減される可能性が示唆された． しかしながら，炎症性サイトカインであるTNF-α の発現には差を認めず， また28 日以降では実験群と対照群の炎症性細胞浸潤， 破骨細胞や骨芽細胞の分布に差を認めなかった． 以上から， 培養骨移植後短期間のステロイド系抗炎症薬投与は， 培養骨移植に伴う初期の炎症反応の一部を軽減する可能性があることが示唆された． その一方で骨形成への影響は明らかでなく， 今後投与量や投与方法の違いによる影響についても， さらに検討する必要があるものと考えられた．2014博士（歯学）松本歯科大

Radial Bargmann representation for the Fock space of type B

Let $\nu_{\alpha,q}$ be the probability and orthogonality measure for the $q$-Meixner-Pollaczek orthogonal polynomials, which has appeared in \cite{BEH15} as the distribution of the $(\alpha,q)$-Gaussian process (the Gaussian process of type B) over the $(\alpha,q)$-Fock space (the Fock space of type B). The main purpose of this paper is to find the radial Bargmann representation of $\nu_{\alpha,q}$. Our main results cover not only the representation of $q$-Gaussian distribution by \cite{LM95}, but also of $q^2$-Gaussian and symmetric free Meixner distributions on $\mathbb R$. In addition, non-trivial commutation relations satisfied by $(\alpha,q)$-operators are presented.Comment: 13 pages, minor changes have been mad

Drosophila Strip serves as a platform for early endosome organization during axon elongation

Early endosomes are essential for regulating cell signalling and controlling the amount of cell surface molecules during neuronal morphogenesis. Early endosomes undergo retrograde transport (clustering) before their homotypic fusion. Small GTPase Rab5 is known to promote early endosomal fusion, but the mechanism linking the transport/clustering with Rab5 activity is unclear. Here we show that Drosophila Strip is a key regulator for neuronal morphogenesis. Strip knockdown disturbs the early endosome clustering, and Rab5-positive early endosomes become smaller and scattered. Strip genetically and biochemically interacts with both Glued (the regulator of dynein-dependent transport) and Sprint (the guanine nucleotide exchange factor for Rab5), suggesting that Strip is a molecular linker between retrograde transport and Rab5 activation. Overexpression of an active form of Rab5 in strip-mutant neurons suppresses the axon elongation defects. Thus, Strip acts as a molecular platform for the early endosome organization that has important roles in neuronal morphogenesis.This work was supported by grants from the National Institute of General Medical Science of the National Institutes of Health (R01-GM085232 to V.I.G.), the National Institutes of Health (R01-DC005982 to L.L.), the Japanese Ministry of Education, Science, Sports, Culture, and Technology (MEXT), the Japan Society for the Promotion of Science, and the Japan Science and Technology Agency (to C.S., K.T., Y.Y., M.M., and T.C.)

Caspase inhibition in select olfactory neurons restores innate attraction behavior in aged Drosophila

Sensory and cognitive performance decline with age. Neural dysfunction caused by nerve death in senile dementia and neurodegenerative disease has been intensively studied; however, functional changes in neural circuits during the normal aging process are not well understood. Caspases are key regulators of cell death, a hallmark of age-related neurodegeneration. Using a genetic probe for caspase-3-like activity (DEVDase activity), we have mapped age-dependent neuronal changes in the adult brain throughout the lifespan of Drosophila. Spatio-temporally restricted caspase activation was observed in the antennal lobe and ellipsoid body, brain structures required for olfaction and visual place memory, respectively. We also found that caspase was activated in an age-dependent manner in specific subsets of Drosophila olfactory receptor neurons (ORNs), Or42b and Or92a neurons. These neurons are essential for mediating innate attraction to food-related odors. Furthermore, age-induced impairments of neural transmission and attraction behavior could be reversed by specific inhibition of caspase in these ORNs, indicating that caspase activation in Or42b and Or92a neurons is responsible for altering animal behavior during normal aging.This work was supported by grants from the Japan Society for the Promotion of Science and the Japan Ministry of Education, Culture, Sports, Science and Technology to TC and MMi, from the Uehara memorial foundation to TC, from NIH to JWW (DK092640), and from NIH to RLD (2R37 NS19904-30). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Analysis of DOC and Ram for NSCLC

Background: Current clinical trials demonstrated that combination regimens comprising chemotherapy and immunotherapy lead to better patient outcomes compared to chemotherapy alone as the first line of treatment for non-small cell lung cancer (NSCLC). In addition, the combination therapy of docetaxel (Doc) and ramucirumab (Ram) was considered one of the standard treatments for advanced or relapsed NSCLC patients. However, little is known about the therapeutic responders of this combination therapy among previously treated NSCLC patients. In the present study, we aimed to identify predictive factors for therapeutic response, including programmed death-ligand 1 (PD-L1) expression in tumors, for Doc treatment in combination with Ram. Methods: We retrospectively analyzed a total of 135 advanced or relapsed NSCLC patients who were refractory to platinum-based chemotherapy at eleven institutions in Japan between July 2016 and November 2018. Results: Our observations showed that PD-L1 expression in tumors is not associated with the efficacy of combined therapy of Doc and Ram in previously treated NSCLC patients. Analysis of the patient clinical profiles indicated that prior treatment with immune checkpoint inhibitors (ICIs) is a reliable predictor for the good progression-free survival (PFS) to this combination therapy (P=0.041). Conclusions: Our retrospective study indicated that combination regimens comprising chemotherapy and ICIs followed by Doc and Ram could be an optimal therapeutic option for NSCLC patients regardless of the PD-L1 status of tumors. Further investigations are required to strengthen clinical evidence demonstrating the effectiveness of the combination therapy of Doc plus Ram in previously treated NSCLC patients

Prevention of Apoptosis by Mitochondrial Phosphatase PGAM5 in the Mushroom Body Is Crucial for Heat Shock Resistance in Drosophila melanogaster

The heat shock (HS) response is essential for survival of all organisms. Although the machinery of the HS response has been extensively investigated at the cellular level, it is poorly understood at the level of the organism. Here, we show the crucial role of the mushroom body (MB) in the HS response in Drosophila. Null mutants of the mitochondrial phosphatase Drosophila PGAM5 (dPGAM5) exhibited increased vulnerability to HS, which was reversed by MB-specific expression of the caspase inhibitor p35, and similar vulnerability was induced in wild-type flies by knockdown of MB dPGAM5. Elimination of the MB did not affect the HS response of wild-type flies, but did increase the resistance of dPGAM5-deficient flies to HS. Thus, the MB may possess an apoptosis-dependent toxic function, the suppression of which by dPGAM5 appears to be crucial for HS resistance

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target