免疫制御細胞の原虫破壊機構の解明と抗原虫ペプチドの開発

2015 Fiscal Year Research-status Report

• Project/Area Number: 15K20840
• Research Institution: Hokkaido University
• Principal Investigator: テルカウィ アラー 北海道大学, 医学(系)研究科(研究院), 助教 (00723074)
• Project Period (FY): 2015-04-01 – 2017-03-31
• Keywords: Phagocytes / Macrophages / Neutrophils

Outline of Annual Research Achievements

Understanding the molecular defense mechanism of phagocytes and identifying their effector molecules against malarial parasites may provide important clues for the discovery of new therapies. In the current proposed study, gene profile of macrophages and neutrophils phagocytizing Plasmodium falciparum-parasitized erythrocytes (iRBC) was studied using DNA microarray in an attempt to define the molecular defense mechanism against infection. The transcriptional gene profile of macrophages in response to iRBCs represented 168 down-regulated genes, which were mainly involved in the cellular immune response, and 216 upregulated genes, which were involved in cellular proteolysis, growth, and adhesion. Importantly, β-defensin 130 (DEFB130) was characterized as anti-malarial agent in vitro and in vivo (The manuscript has been submitted to the Scientific Report).
The gene profiling of the neutrophils in response to iRBCs revealed a broad and vigorous set of gene expression represented in 384 downregulated and 148 genes upregulated genes. Importantly, the upregulated genes were involved in multiple cellular function including cellular signaling, development, proteolysis, immune suppression and adhesion. Next, the results generated by DNA microarray analysis was confirmed by quantitative real-time PCR (qPCR) for the randomly upregulated genes.

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

This is the first study showing the involvement of macrophage defensin in killing of malarial parasites. DEFB130 was one of the top upregulated genes in macrophages and differentiated macrophages phagocytizing iRBCs exhibited an increase in intracellular DEFB130 levels accumulate at the site of iRBC engulfment. Both in vitro and in vivo experiments using DEFB130 synthetic peptide revealed a modest toxic effect of this molecule on the parasites.
Genes including CTSL, SOC3, CISH2, CXCL10 and CD64 were the highly upregulated in neutrophils phagocytizing iRBCs and they are involved in multiple functions such as cellular proteolysis, immune response and adhesion.

Strategy for Future Research Activity

Based on DNA microarray data, five genes including CTSL, SOC3, CISH2, CXCL10 and CD64 are selected for further characterization. Manipulation of gene expression using gene overexpression and knockdown techniques using human HL60 cell line and primary neutrophils is to be perform to understand the role of the targeted genes in malarial infection.

Causes of Carryover

所属先変更に伴い、その施設に合った設備等を選定するのにあたり、少々時間を要した。

Expenditure Plan for Carryover Budget

設備等はすでに発注等しており、購入費用に使用する計画である。

Research Products

• [Journal Article] Isolation and co-cultivation of human macrophages and neutrophils with Plasmodium falciparum-parasitized erythrocytes: An optimized system to study the phagocytic activity to malarial parasites (2016)
  • Author(s): Terkawi MA, Takano, R, Kato K.
  • Journal Title: Parasitology International Volume: In press Pages: In press
  • DOI: 10.1016/j.parint.2016.03.005.
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Dextran sulfate inhibits acute Toxoplama gondii infection in pigs (2016)
  • Author(s): Kato K, Murata Y, Horiuchi N, Inomata A, Terkawi MA, Ishiwa A, Ogawa Y, Fukumoto S, Matsuhisa F, Koyama K.
  • Journal Title: Parasites & Vectors Volume: 9 Pages: 134
  • DOI: 10.1186/s13071-016-1421-9.
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Depletion of phagocytic cells during nonlethal Plasmodium yoelii infection causes severe malaria characterized by acute renal failure in Mice (2016)
  • Author(s): Terkawi MA, Nishimura M, Furuoka H, Nishikawa Y.
  • Journal Title: Infection & Immunty Volume: 84 Pages: 845-855
  • DOI: 10.1128/IAI.01005-15.
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Characterization of Toxoplasma gondii glyoxalase 1 and evaluation of inhibitory effects of curcumin on the enzyme and parasite cultures. (2015)
  • Author(s): Goo YK, Yamagishi J, Ueno A, Terkawi MA, Aboge GO, Kwak D, Hong Y, Chung DI, Igarashi M, Nishikawa Y, Xuan X.
  • Journal Title: Parasites & Vectors Volume: 8 Pages: 654
  • DOI: 10.1186/s13071-015-1268-5.
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Molecular detection and characterization of Babesia bovis, Babesia bigemina, Theileria species and Anaplasma marginale isolated from cattle in Kenya. (2015)
  • Author(s): Adjou Moumouni PF, Aboge GO, Terkawi MA, Masatani T, Cao S, Kamyingkird K, Jirapattharasate C, Zhou M, Wang G, Liu M, Iguchi A, Vudriko P, Ybanez AP, Inokuma H, Shirafuji-Umemiya R, Suzuki H, Xuan X.
  • Journal Title: Parasites & Vectors Volume: 8 Pages: 496
  • DOI: 10.1186/s13071-015-1106-9.
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Optimization of a Fluorescence-Based Assay for Large-Scale Drug Screening against Babesia and Theileria Parasites. (2015)
  • Author(s): Rizk MA, El-Sayed SA, Terkawi MA, Youssef MA, El Said el Sel S, Elsayed G, El-Khodery S, El-Ashker M, Elsify A, Omar M, Salama A, Yokoyama N, Igarashi I.
  • Journal Title: PloS One Volume: 10 Pages: e0125276
  • DOI: 10.1371/journal.pone.0125276. eCollection 2015.
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Presentation] Differentiated gene profile of neutrophils induced by phagocytosis of Plasmodium falciparum-parasitized erythrocytes. (2016)
  • Author(s): Terkawi MA
  • Organizer: The 85th Annual Meeting of the Japanese Society of Parasitology.
  • Place of Presentation: 宮崎市民プラザ（宮崎市）
  • Year and Date: 2016-03-19 – 2016-03-20
• [Presentation] Involvement of a novel cysteine-rich cationic low molecular weight protein in the macrophage microbicidal mechanisms for killing Plasmodium falciparum (2015)
  • Author(s): Terkawi MA
  • Organizer: The 14th Awaji international forum on infection and immunity
  • Place of Presentation: 淡路夢舞台国際会議場（淡路市）
  • Year and Date: 2015-09-08 – 2015-09-11