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11月
21
2019
学術講演会(光ピンセット顕微鏡システムの説明会)
11月 21 @ 10:00

演 題:学術講演会(光ピンセット顕微鏡システムの説明会)

日 時:11月21日(金) ①10:00~ ②13:30~ ③16:00~

場 所:総合研究1号館209室

申し込み:ナノハブの高橋様 (kyodai-hub@saci.kyoto-u.ac.jp) まで

開始時間(ご参加可能な時間帯を全て)と言語(日本語、英語)をご連絡願います。

2019年度#3機器説明会ポスター20191121

11月
27
2019
The Journey and the Destination: Confinement Mechanobiology
11月 27 @ 17:00 – 18:30

日時:2019年11月27日(水)17:00-17:45

講演者:Dr. Andrew Holle
      (Postdoctoral Fellow and AACR Basic Research Fellow, Max Planck Institute for Medical Research; Heidelberg, Germany)

演題: The Journey and the Destination: Confinement Mechanobiology

言語:英語

場所:京都大学 高等研究院(iCeMS)本館(建物番号 77)2階 セミナールーム (A207)
http://www.icems.kyoto-u.ac.jp/ja/access/

アブストラクト:
The ability of cells to live under physical confinement and navigate through tight spaces has relevance in a wide variety of biological regimes, including homeostatic processes like stem cell migration or immune cell targeting and disease states like cancer or chronic inflammation. One method we use to investigate this process utilizes photolithography-generated microchannel arrays to present long, confining passages to cancer cells. We have identified distinct mesenchymal and amoeboid migration modalities in MDA-MB-231 breast cancer cells that are dependent on time, ligand composition, channel dimension, cytoskeletal activity, and intracellular signaling. The transition from mesenchymal to amoeboid modalities (MAT) was found to be a dynamic process that is driven initially by Rac1-induced protrusions and subsequently by ROCK-dependent contractility. While the cytoskeletal response to confined spaces has been studied extensively, recent research into the role of the nucleus in confined migration has implicated nuclear biophysics in a number of confinement-dependent outcomes, including enhanced metastasis and cell death. We have investigated the expression of multiple nuclear reorganization proteins, including DNA damage repair (53BP1) and nuclear envelope repair (CHMP4B), and have found that in long microchannels, cells do not display the enhanced nuclear damage observed in ‘pinch-point’ constrictions. This suggests that cellular sensing of sustained confinement allows for cytoskeletal-nuclear adaptation. One such adaptation is dynamic changes in nuclear volume that accompany cell entrance into channels and are maintained in cells post-permeation, providing a form of ‘migrational memory’. Using a panel of CRISPR-mediated knockout cell lines, we have identified the Protein Kinase D isoform PKD1 as a crucial signaling protein required for nuclear adaptation and remodeling in response to a narrow, sustained confinement. Future investigations into­­­­ confined migration in diverse cellular models, including stem cells and immune cells, will likely play an important role in the development of new tissue engineering strategies.

***
日時:2019年11月27日(水)17:45-18:30

講演者:Dr. Jennifer L. Young
      (Postdoctoral Fellow, Max Planck Institute for Medical Research; Heidelberg, Germany)

演題: Assaying ECM-conferred chemoresistance on orthogonal gradient hydrogel systems

言語:英語

場所:京都大学 高等研究院(iCeMS)本館(建物番号 77)2階 セミナールーム (A207)
http://www.icems.kyoto-u.ac.jp/ja/access/

アブストラクト:
Cancer cell-ECM interactions have been shown to positively influence cancer cell survival and invasion by conferring adhesion-based resistance in response to chemotherapeutic drugs, and subsequently driving metastasis into surrounding tissues. While numerous promising integrin-targeted drugs have been developed, none have been successfully implemented into clinical practice due to their inconsistent performance in effectively targeting the tumor, which could stem from the enormous molecular complexity of the tumor matrix environment. Here, we present a highly-defined platform designed to identify protective matrix properties in a robust manner, examining the effects of both ECM ligand and mechanical properties in regulating chemoresistance in breast cancer cells. We have previously developed a two-step polymerization method for creating tunable stiffness gradient polyacrylamide (PA) hydrogels with values spanning the in vivo physiological and pathological mechanical landscape, i.e. from ~ 0.1 up to 40 kPa. In order to study the influence of a wide range of ECM environments, we created dual gradient hydrogels by first synthesizing shallow, non-durotactic stiffness gradient hydrogels and subsequently fabricating a gradient of ligands orthogonal to the stiffness gradient to which breast cancer cells can attach. Ligand gradients are produced by either a gradient photomask to which proteins can be coupled to the substrate via a UV-sensitive crosslinker or by depositing a gradient of highly-ordered gold nanoparticles onto the hydrogel to which thiolated integrin subtype-specific peptidomimetics, specific to integrins αvβ3 or α5β1, can readily attach. Linear nanoparticle gradients are created using the technique Block Copolymer Micelle Nanolithography, and can span ~ 35 to 80 nm interparticle spacing over ~ 20 mm. Using these dual gradient hydrogels, we have identified specific combinations of ligand density and substrate stiffness that affect cancer cell survival, i.e. stiffer regions with more dense ligands afford higher chemoresistance to breast cancer cells. Interestingly, we find a correlation between substrate stiffness and ligand density in promoting chemoresistance, whereby high cell survival is found in a linear regime spanning lower substrate stiffness with less dense ligands up to higher substrate stiffness with more dense ligands, indicating that cancer cells could be actively modulating ligand spacing in order to enhance their survival via the activation of pro-survival signaling cascades that originate at the cell membrane. Taken together, these data provide a better understanding of the interplay between substrate stiffness, ligand type, and ligand spacing in regulating adhesion-conferred chemoprotection in cancer cells, with the ultimate aim of establishing future targets for more effective, combinatorial cancer treatments.

問い合わせ先:京都大学物質-細胞統合システム拠点(iCeMS) 見学グループ 中澤直高
       kengaku-g*icems.kyoto-u.ac.jp(*を@に変換ください)(内線9833)

*******************************************************************************************

** Seminar announcement **

This email is to inform you of upcoming seminar, all researchers are welcomed to attend. No registration is required.

Date/Time: Wednesday, November 27, 2019, 17:00-17:45

Title: The Journey and the Destination: Confinement Mechanobiology

Speaker: Dr. Andrew Holle
      (Postdoctoral Fellow and AACR Basic Research Fellow, Max Planck Institute for Medical Research; Heidelberg, Germany)

Venue: 2nd floor Seminar Room (#A207), KUIAS, iCeMS Main Building (#77), Kyoto University
http://www.icems.kyoto-u.ac.jp/en/access/

Abstract:
The ability of cells to live under physical confinement and navigate through tight spaces has relevance in a wide variety of biological regimes, including homeostatic processes like stem cell migration or immune cell targeting and disease states like cancer or chronic inflammation. One method we use to investigate this process utilizes photolithography-generated microchannel arrays to present long, confining passages to cancer cells. We have identified distinct mesenchymal and amoeboid migration modalities in MDA-MB-231 breast cancer cells that are dependent on time, ligand composition, channel dimension, cytoskeletal activity, and intracellular signaling. The transition from mesenchymal to amoeboid modalities (MAT) was found to be a dynamic process that is driven initially by Rac1-induced protrusions and subsequently by ROCK-dependent contractility. While the cytoskeletal response to confined spaces has been studied extensively, recent research into the role of the nucleus in confined migration has implicated nuclear biophysics in a number of confinement-dependent outcomes, including enhanced metastasis and cell death. We have investigated the expression of multiple nuclear reorganization proteins, including DNA damage repair (53BP1) and nuclear envelope repair (CHMP4B), and have found that in long microchannels, cells do not display the enhanced nuclear damage observed in ‘pinch-point’ constrictions. This suggests that cellular sensing of sustained confinement allows for cytoskeletal-nuclear adaptation. One such adaptation is dynamic changes in nuclear volume that accompany cell entrance into channels and are maintained in cells post-permeation, providing a form of ‘migrational memory’. Using a panel of CRISPR-mediated knockout cell lines, we have identified the Protein Kinase D isoform PKD1 as a crucial signaling protein required for nuclear adaptation and remodeling in response to a narrow, sustained confinement. Future investigations into­­­­ confined migration in diverse cellular models, including stem cells and immune cells, will likely play an important role in the development of new tissue engineering strategies.

***
Date/Time: Wednesday, November 27, 2019, 17:45-18:30

Title: Assaying ECM-conferred chemoresistance on orthogonal gradient hydrogel systems

Speaker: Dr. Jennifer L. Young
      (Postdoctoral Fellow, Max Planck Institute for Medical Research; Heidelberg, Germany)

Venue: 2nd floor Seminar Room (#A207), KUIAS, iCeMS Main Building (#77), Kyoto University
<http://www.icems.kyoto-u.ac.jp/en/access/>

Abstract:
Cancer cell-ECM interactions have been shown to positively influence cancer cell survival and invasion by conferring adhesion-based resistance in response to chemotherapeutic drugs, and subsequently driving metastasis into surrounding tissues. While numerous promising integrin-targeted drugs have been developed, none have been successfully implemented into clinical practice due to their inconsistent performance in effectively targeting the tumor, which could stem from the enormous molecular complexity of the tumor matrix environment. Here, we present a highly-defined platform designed to identify protective matrix properties in a robust manner, examining the effects of both ECM ligand and mechanical properties in regulating chemoresistance in breast cancer cells. We have previously developed a two-step polymerization method for creating tunable stiffness gradient polyacrylamide (PA) hydrogels with values spanning the in vivo physiological and pathological mechanical landscape, i.e. from ~ 0.1 up to 40 kPa. In order to study the influence of a wide range of ECM environments, we created dual gradient hydrogels by first synthesizing shallow, non-durotactic stiffness gradient hydrogels and subsequently fabricating a gradient of ligands orthogonal to the stiffness gradient to which breast cancer cells can attach. Ligand gradients are produced by either a gradient photomask to which proteins can be coupled to the substrate via a UV-sensitive crosslinker or by depositing a gradient of highly-ordered gold nanoparticles onto the hydrogel to which thiolated integrin subtype-specific peptidomimetics, specific to integrins αvβ3 or α5β1, can readily attach. Linear nanoparticle gradients are created using the technique Block Copolymer Micelle Nanolithography, and can span ~ 35 to 80 nm interparticle spacing over ~ 20 mm. Using these dual gradient hydrogels, we have identified specific combinations of ligand density and substrate stiffness that affect cancer cell survival, i.e. stiffer regions with more dense ligands afford higher chemoresistance to breast cancer cells. Interestingly, we find a correlation between substrate stiffness and ligand density in promoting chemoresistance, whereby high cell survival is found in a linear regime spanning lower substrate stiffness with less dense ligands up to higher substrate stiffness with more dense ligands, indicating that cancer cells could be actively modulating ligand spacing in order to enhance their survival via the activation of pro-survival signaling cascades that originate at the cell membrane. Taken together, these data provide a better understanding of the interplay between substrate stiffness, ligand type, and ligand spacing in regulating adhesion-conferred chemoprotection in cancer cells, with the ultimate aim of establishing future targets for more effective, combinatorial cancer treatments.

Contact: Naotaka Nakazawa, Kengaku Group,  iCeMS
             kengaku-g*icems.kyoto-u.ac.jp (change*to@) (x 9833)

20191127 KengakuG seminar flyer

11月
29
2019
Optogenetic Dissection Erk-dependent Cell Fates During Embryogenesis
11月 29 @ 00:00

日時:11月29日
演者(所属)1: Heath Johnson(Princeton University)
演題1:Optogenetic Dissection Erk-dependent Cell Fates During Embryogenesis

演者(所属)2: 小崎 健次郎(慶応義塾大学)
演題2:Learning Biologic Insights from Patients with Undiagnosed Diseases

場所:基礎医学記念講堂・医学部資料館
世話人(分野):松田 道行

12月
6
2019
Mechanisms of short- and long-term mechanosensing
12月 6 @ 17:00 – 18:00

日時:2019年12月6日(金)17:00-18:00

講演者:Dr. Haguy Wolfenson
   (Assistant Professor, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel)

演題: Mechanisms of short- and long-term mechanosensing

言語:英語

場所:京都大学 高等研究院(iCeMS)本館(建物番号 77)2階 セミナールーム (A207)
http://www.icems.kyoto-u.ac.jp/ja/access/

アブストラクト:
Cells respond to mechanical signals from their environment in a variety of ways. In particular, the rigidity of the extracellular matrix (ECM) to which cells adhere is a critical determinant of the most fundamental cellular processes, including cell migration, differentiation, death, and growth. In order to test the rigidity of the ECM, cells apply cytoskeletal-based forces to it; however, there are fundamental aspects of this ‘mechanosensing’ process that are poorly understood. In my talk, I will discuss our recent findings on the kinetics of contractile force generation during mechanosensing. Based on a mathematical model of cell contractility, we predict that for a broad range of physiological and laboratory conditions the contractile force scales with the environment‘s rigidity, where the proportionality factor is an intrinsic, cell-specific time-dependent contractile displacement that is non-mechanosensitive, i.e. independent of the environment’s rigidity. Our extensive experiments on various adherent cells show that, as predicted, the time-dependent contractile displacement is independent of the rigidity, varied more than 15-fold. Furthermore, we show that the intrinsic time-dependent contractile displacement is directly related to the evolution of the actomyosin network, properly quantified by the time-dependent concentration of F-actin. The emerging picture unifies various other existing observations on cellular contractility and provides a novel framework to address the process of mechanosensing.

Key Words:Mechanosensing, ECM

問い合わせ先:京都大学物質-細胞統合システム拠点(iCeMS) 見学グループ 中澤直高
       kengaku-g*icems.kyoto-u.ac.jp(*を@に変換ください)(内線9833)

191206 Dr.Wolfenson seminar flyer

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