Details of a Researcher - SAITO Atsushi

Papers - SAITO Atsushi

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肺サーファクタントタンパク質Dは変異型EGFRの活性化を阻害し、肺腺がんの進展を抑制する

長谷川喜弘, 梅田泰淳, 大塚満雄, 有木茂, 高宮里奈, 齋藤充史, 高橋弘毅, 黒木由夫, 高橋素子

生命科学系学会合同年次大会 ( 生命科学系学会合同年次大会運営事務局 ) 2017年度 [1P - 0948] 2017.12

タバコ煙中のアクロレインが肺サーファクタントタンパク質A(SP-A)の機能や構造に及ぼす影響

高宮里奈, 内田浩二, 柴田貴広, 前野敏孝, 有木茂, 長谷川喜弘, 齊藤充史, 黒木由夫, 高橋素子

生命科学系学会合同年次大会 ( 生命科学系学会合同年次大会運営事務局 ) 2017年度 [3P - 1166] 2017.12

Surfactant protein A down-regulates epidermal growth factor receptor by mechanisms different from those of surfactant protein D

Yoshihiro Hasegawa, Motoko Takahashi, Shigeru Ariki, Atsushi Saito, Yasuaki Uehara, Rina Takamiya, Koji Kuronuma, Hirofumi Chiba, Yuji Sakuma, Hiroki Takahashi, Yoshio Kuroki

JOURNAL OF BIOLOGICAL CHEMISTRY ( AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC ) 292 ( 45 ) 18565 - 18576 2017.11 [Refereed]

We recently reported that the lectin surfactant protein D (SP-D) suppresses epidermal growth factor receptor (EGFR) signaling by interfering with ligand binding to EGFR through an interaction between the carbohydrate-recognition domain (CRD) of SP-D and N-glycans of EGFR. Here, we report that surfactant protein A (SP-A) also suppresses EGF signaling in A549 human lung adenocarcinoma cells and in CHOK1 cells stably expressing human EGFR and that SP-A inhibits the proliferation and motility of the A549 cells. Results with I-125-EGF indicated that SP-A interferes with EGF binding to EGFR, and a ligand blot analysis suggested that SP-A binds EGFR in A549 cells. We also found that SP-A directly binds the recombinant extracellular domain of EGFR (soluble EGFR or sEGFR), and this binding, unlike that of SP-D, was not blocked by EDTA, excess mannose, or peptide:N-glycosidase F treatment. We prepared a collagenase-resistant fragment (CRF) of SP-A, consisting of CRD plus the neck domain of SP-A, and observed that CRF directly binds sEGFR but does not suppress EGF-induced phosphorylation of EGFR in or proliferation of A549 cells. These results indicated that SP-A binds EGFR and down-regulates EGF signaling by inhibiting ligand binding to EGFR as well as SP-D. However, unlike for SP-D, SP-A lectin activity and EGFR N-glycans were not involved in the interaction between SP-A and EGFR. Furthermore, our results suggested that oligomerization of SP-A is necessary to suppress the effects of SP-A on EGF signaling.

Surfactant protein D inhibits activation of non-small cell lung cancer-associated mutant EGFR and affects clinical outcomes of patients

Y. Umeda, Y. Hasegawa, M. Otsuka, S. Ariki, R. Takamiya, A. Saito, Y. Uehara, H. Saijo, K. Kuronuma, H. Chiba, H. Ohnishi, Y. Sakuma, H. Takahashi, Y. Kuroki, M. Takahashi

ONCOGENE ( NATURE PUBLISHING GROUP ) 36 ( 46 ) 6432 - 6445 2017.11 [Refereed]

Tyrosine kinase inhibitor (TKI)-sensitive and TKI-resistant mutations of epidermal growth factor receptor (EGFR) are associated with lung adenocarcinoma. EGFR mutants were previously shown to exhibit ligand-independent activation. We have previously demonstrated that pulmonary surfactant protein D (SP-D, SFTPD) suppressed wild-type EGFR signaling by blocking ligand binding to EGFR. We herein demonstrate that SFTPD downregulates ligand-independent signaling in cells harboring EGFR mutations such as TKI-sensitive exon 19 deletion (Ex19del) and L858R mutation as well as TKI-resistant T790M mutation, subsequently suppressing cellular growth and motility. Lectin blotting and ligand blotting in lung cancer cell lines suggested that EGFR mutants express oligomannose-type N-glycans and interact with SFTPD directly. Cross-linking assay indicated that SFTPD inhibits ligand-independent dimerization of EGFR mutants. We also demonstrated that SFTPD reduced dimerization-independent phosphorylation of Ex19del and T790M EGFR mutants using point mutations that disrupted the asymmetric dimer interface. It was confirmed that SFTPD augmented the viability-suppressing effects of EGFR-TKIs. Furthermore, retrospective analysis of 121 patients with lung adenocarcinoma to examine associations between serum SFTPD levels and clinical outcome indicated that in TKI-treated patients with lung cancer harboring EGFR mutations, including Ex19del or L858R, high serum SFTPD levels correlated with a lower number of distant metastases and prolonged overall survival and progression-free survival. These findings suggest that SFTPD downregulates both TKI-sensitive and -resistant EGFR mutant signaling, and SFTPD level is correlated with clinical outcome. These findings illustrate the use of serum SFTPD level as a potential marker to estimate the efficacy of EGFR-TKIs.

Serum surfactant protein D predicts the outcome of patients with idiopathic pulmonary fibrosis treated with pirfenidone

Kimiyuki Ikeda, Masanori Shiratori, Hirofumi Chiba, Hirotaka Nishikiori, Keiki Yokoo, Atsushi Saito, Yoshihiro Hasegawa, Koji Kuronuma, Mitsuo Otsuka, Gen Yamada, Hiroki Takahashi

RESPIRATORY MEDICINE ( W B SAUNDERS CO LTD ) 131 184 - 191 2017.10 [Refereed]

Background: Idiopathic pulmonary fibrosis (IPF) is a fatal pulmonary disease with poor prognosis. Pirfenidone, the first antifibrotic drug, suppresses the decline in forced vital capacity (FVC) and improves prognosis in some, but not all, patients with IPF; therefore, an indicator for identifying improved outcomes in pirfenidone therapy is desirable. This study aims to clarify whether baseline parameters can be predictors of disease progression and prognosis in patients with IPF treated with pirfenidone. Methods: We retrospectively investigated patients with IPF who started treatment with pirfenidone between December 2008 and November 2014 at the Sapporo Medical University Hospital. Patients treated with pirfenidone for >6 months were enrolled in this study and were observed until November 2015. We investigated the association of clinical characteristics, pulmonary function test results, and blood examination results at the start of pirfenidone with the outcome of patients. Results: Sixty patients were included in this study. In multivariate logistic regression analysis, % predicted FVC and serum surfactant protein (SP)-D levels were predictors of a >10% decline in FVC in the initial 12 months. In the Cox proportional hazards model, these two factors predicted progression-free survival. Pack-years, % predicted diffusing capacity for carbon monoxide, and SP-D levels predicted overall survival. Conclusions: The serum SP-D level was a predictor of disease progression and prognosis in patients with IPF treated with pirfenidone. In addition, this analysis describes the relative usefulness of other clinical parameters at baseline in estimating the prognosis of patients with IPF who are candidates for pirfenidone therapy. (C) 2017 Elsevier Ltd. All rights reserved.

Disruption of the structural and functional features of surfactant protein A by acrolein in cigarette smoke

Rina Takamiya, Koji Uchida, Takahiro Shibata, Toshitaka Maeno, Masaki Kato, Yoshiki Yamaguchi, Shigeru Ariki, Yoshihiro Hasegawa, Atsushi Saito, Soichi Miwa, Hiroki Takahashi, Takaaki Akaike, Yoshio Kuroki, Motoko Takahashi

SCIENTIFIC REPORTS ( NATURE PUBLISHING GROUP ) 7 ( 1 ) 8304 2017.08 [Refereed]

The extent to which defective innate immune responses contribute to chronic obstructive pulmonary disease (COPD) is not fully understood. Pulmonary surfactant protein A (SP-A) plays an important role in regulating innate immunity in the lungs. In this study, we hypothesised that cigarette smoke (CS) and its component acrolein might influence pulmonary innate immunity by affecting the function of SP-A. Indeed, acrolein-modified SP-A was detected in the lungs of mice exposed to CS for 1 week. To further confirm this finding, recombinant human SP-A (hSP-A) was incubated with CS extract (CSE) or acrolein and then analysed by western blotting and nanoscale liquid chromatography-matrix-assisted laser desorption/ionisation time-of-flight tandem mass spectrometry. These analyses revealed that CSE and acrolein induced hSP-A oligomerisation and that acrolein induced the modification of six residues in hSP-A: His39, His116, Cys155, Lys180, Lys221, and Cys224. These modifications had significant effects on the innate immune functions of hSP-A. CSE- or acrolein-induced modification of hSP-A significantly decreased hSP-A's ability to inhibit bacterial growth and to enhance macrophage phagocytosis. These findings suggest that CS-induced structural and functional defects in SP-A contribute to the dysfunctional innate immune responses observed in the lung during cigarette smoking.

Mitogenic stimulation accelerates influenza-induced mortality by increasing susceptibility of alveolar type II cells to infection

Nikolaos M. Nikolaidis, John G. Noel, Lori B. Pitstick, Jason C. Gardner, Yasuaki Uehara, Huixing Wu, Atsushi Saito, Kara E. Lewnard, Huan Liu, Mitchell R. White, Kevan L. Hartshorn, Francis X. McCormack

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA ( NATL ACAD SCIENCES ) 114 ( 32 ) E6613 - E6622 2017.08 [Refereed]

Development of pneumonia is the most lethal consequence of influenza, increasing mortality more than 50-fold compared with uncomplicated infection. The spread of viral infection from conducting airways to the alveolar epithelium is therefore a pivotal event in influenza pathogenesis. We found that mitogenic stimulation with keratinocyte growth factor (KGF) markedly accelerated mortality after infectious challenge with influenza A virus (IAV). Coadministration of KGF with IAV markedly accelerated the spread of viral infection from the airways to alveoli compared with challenge with IAV alone, based on spatial and temporal analyses of viral nucleoprotein staining of lung tissue sections and dissociated lung cells. To better define the temporal relationship between KGF administration and susceptibility to IAV infection in vivo, we administered KGF 120, 48, 24, and 0 h before intrapulmonary IAV challenge and assessed the percentages of proliferating and IAV-infected, alveolar type II (AECII) cells in dispersed lung cell populations. Peak AECII infectivity coincided with the timing of KGF administration that also induced peak AECII proliferation. AECII from mice that were given intrapulmonary KGF before isolation and then infected with IAV ex vivo exhibited the same temporal pattern of proliferation and infectious susceptibility. KGF-induced increases in mortality, AECII proliferation, and enhanced IAV susceptibility were all reversed by pretreatment of the animals with the mTOR inhibitor rapamycin before mitogenic stimulation. Taken together, these data suggest mTOR signalingdependent, mitogenic conditioning of AECII is a determinant of host susceptibility to infection with IAV.

Surfactant Protein A Inhibits Growth and Adherence of Uropathogenic Escherichia coli To Protect the Bladder from Infection

Jiro Hashimoto, Motoko Takahashi, Atsushi Saito, Masaki Murata, Yuichiro Kurimura, Chiaki Nishitani, Rina Takamiya, Yasuaki Uehara, Yoshihiro Hasegawa, Yoshiki Hiyama, Norimasa Sawada, Satoshi Takahashi, Naoya Masumori, Yoshio Kuroki, Shigeru Ariki

JOURNAL OF IMMUNOLOGY ( AMER ASSOC IMMUNOLOGISTS ) 198 ( 7 ) 2898 - 2905 2017.04 [Refereed]

Surfactant protein A (SP-A) is a multifunctional host defense collectin that was first identified as a component of pulmonary surfactant. Although SP-A is also expressed in various tissues, including the urinary tract, its innate immune functions in nonpulmonary tissues are poorly understood. In this study, we demonstrated that adherence of uropathogenic Escherichia coli (UPEC) to the bladder was enhanced in SP-A-deficient mice, which suggests that SP-A plays an important role in innate immunity against UPEC. To understand the innate immune functions of SP-A in detail, we performed in vitro experiments. SP-A directly bound to UPEC in a Ca2+-dependent manner, but it did not agglutinate UPEC. Our results suggest that a bouquet-like arrangement seems unsuitable to agglutinate UPEC. Meanwhile, SP-A inhibited growth of UPEC in human urine. Furthermore, the binding of SP-A to UPEC decreased the adherence of bacteria to urothelial cells. These results indicate that direct action of SP-A on UPEC is important in host defense against UPEC. Additionally, adhesion of UPEC to urothelial cells was decreased when the cells were preincubated with SP-A. Adhesion of UPEC to urothelial cells is achieved via interaction between FimH, an adhesin located at bacterial pili, and uroplakin Ia, a glycoprotein expressed on the urothelium. SP-A directly bound to uroplakin Ia and competed with FimH for uroplakin Ia binding. These results lead us to conclude that SP-A plays important roles in host defense against UPEC.

Surfactant protein A (SP-A) and SP-A-derived peptide attenuate chemotaxis of mast cells induced by human beta-defensin 3

Yasuaki Uehara, Motoko Takahashi, Masaki Murata, Atsushi Saito, Rina Takamiya, Yoshihiro Hasegawa, Koji Kuronuma, Hirofumi Chiba, Jiro Hashimoto, Norimasa Sawada, Hiroki Takahashi, Yoshio Kuroki, Shigeru Ariki

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS ( ACADEMIC PRESS INC ELSEVIER SCIENCE ) 485 ( 1 ) 107 - 112 2017.03 [Refereed]

Human beta-defensin 3 (hBD3) is known to be involved in mast cell activation. However, molecular mechanisms underlying the regulation of hBD3-induced mast cell activation have been poorly understood. We previously reported that SP-A and SP-A-derived peptide 01 (SAP01) regulate the function of hBD3. In this study, we focused on the effects of SP-A and SAP01 on the activation of mast cells induced by hBD3. SAP01 directly bound to hBD3. Mast cell-mediated vascular permeability and edema in hBD3 administered rat ears were decreased when injected with SP-A or SAP01. Compatible with the results in rat ear model, both SP-A and SAP01 inhibited hBD3-induced chemotaxis of mast cells in vitro. Direct interaction between SP-A or SAP01 and hBD3 seemed to be responsible for the inhibitory effects on chemotaxis. Furthermore, SAP01 attenuated hBD3-induced accumulation of mast cells and eosinophils in tracheas of the OVA-sensitized inflammatory model. SP-A might contribute to the regulation of inflammatory responses mediated by mast cells during infection. (C) 2017 Elsevier Inc. All rights reserved.

非HIV患者におけるST合剤使用困難ニューモシスチス肺炎に対するアトバコンの使用経験

小林智史, 齊藤充史, 錦織博貴, 黒沼幸治, 高橋弘毅

感染症学雑誌 ( (一社)日本感染症学会 ) 91 ( 1 ) 93 - 93 2017.01

Pulmonary Alveolar Microlithiasis

Atsushi Saito, Francis X. McCormack

CLINICS IN CHEST MEDICINE ( W B SAUNDERS CO-ELSEVIER INC ) 37 ( 3 ) 441 - + 2016.09 [Refereed]

Pulmonary alveolar microlithiasis (PAM) is a genetic lung disorder that is characterized by the accumulation of calcium phosphate deposits in the alveolar spaces of the lung. Mutations in the type II sodium phosphate cotransporter, NPT2b, have been reported in patients with PAM. PAM progresses gradually, often producing incremental dyspnea on exertion, desaturation in young adulthood, and respiratory insufficiency by late middle age. Treatment remains supportive, including supplemental oxygen therapy. For patients with end-stage disease, lung transplantation is available as a last resort. The recent development of a laboratory animal model has revealed several promising treatment approaches for future trials.

Keratinocyte growth factor supports pulmonary innate immune defense through maintenance of alveolar antimicrobial protein levels and macrophage function

Jason C. Gardner, Huixing Wu, John G. Noel, Benjamin J. Ramser, Lori Pitstick, Atsushi Saito, Nikolaos M. Nikolaidis, Francis X. McCormack

AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY ( AMER PHYSIOLOGICAL SOC ) 310 ( 9 ) L868 - L879 2016.05 [Refereed]

Keratinocyte growth factor (KGF) is an epithelial mitogen that has been reported to protect the lungs from a variety of toxic and infectious insults. In prior studies we found that recombinant human KGF accelerates clearance of bacteria from the murine lung by augmenting the function of alveolar macrophages (AM). In this study we tested the hypothesis that endogenous KGF plays a role in the maintenance of innate pulmonary defense against gram-negative bacterial infections. KGF-deficient mice exhibited delayed clearance of Escherichia coli from the lungs, attenuated phagocytosis by AM, and decreased antimicrobial activity in bronchoalveolar lavage (BAL) fluid, due in part to reductions in levels of surfactant protein A, surfactant protein D, and lysozyme. These immune deficits were accompanied by lower alveolar type II epithelial cell counts and reduced alveolar type II epithelial cell expression of collectin and lysozyme genes on a per cell basis. No significant between-group differences were detected in selected inflammatory cytokines or BAL inflammatory cell populations at baseline or after bacterial challenge in the wild-type and KGF-deficient mice. A single intranasal dose of recombinant human KGF reversed defects in bacterial clearance, AM function, and BAL fluid antimicrobial activity. We conclude that KGF supports alveolar innate immune defense through maintenance of alveolar antimicrobial protein levels and functions of AM. Together these data demonstrate a role for endogenous KGF in maintenance of normal pulmonary innate immune function.

Modeling pulmonary alveolar microlithiasis by epithelial deletion of the Npt2b sodium phosphate cotransporter reveals putative biomarkers and strategies for treatment

Atsushi Saito, Nikolaos M. Nikolaidis, Hassane Amlal, Yasuaki Uehara, Jason C. Gardner, Kathleen LaSance, Lori B. Pitstick, James P. Bridges, Kathryn A. Wikenheiser-Brokamp, Dennis W. McGraw, Jason C. Woods, Yves Sabbagh, Susan C. Schiavi, Goksel Altinisik, Marko Jakopovic, Yoshikazu Inoue, Francis X. McCormack

SCIENCE TRANSLATIONAL MEDICINE ( AMER ASSOC ADVANCEMENT SCIENCE ) 7 ( 313 ) 313ra181 2015.11 [Refereed]

Pulmonary alveolar microlithiasis (PAM) is a rare, autosomal recessive lung disorder associated with progressive accumulation of calcium phosphate microliths. Inactivating mutations in SLC34A2, which encodes the NPT2b sodium-dependent phosphate cotransporter, has been proposed as a cause of PAM. We show that epithelial deletion of Npt2b in mice results in a progressive pulmonary process characterized by diffuse alveolar microlith accumulation, radiographic opacification, restrictive physiology, inflammation, fibrosis, and an unexpected alveolar phospholipidosis. Cytokine and surfactant protein elevations in the alveolar lavage and serum of PAM mice and confirmed in serum from PAM patients identify serum MCP-1 (monocyte chemotactic protein 1) and SP-D (surfactant protein D) as potential biomarkers. Microliths introduced by adoptive transfer into the lungs of wild-typemice produce marked macrophage-rich inflammation and elevation of serum MCP-1 that peaks at 1 week and resolves at 1 month, concomitant with clearance of stones. Microliths isolated by bronchoalveolar lavage readily dissolve in EDTA, and therapeutic whole-lung EDTA lavage reduces the burden of stones in the lungs. A low-phosphate diet prevents microlith formation in young animals and reduces lung injury on the basis of reduction in serum SP-D. The burden of pulmonary calcium deposits in established PAM is also diminished within 4 weeks by a low-phosphate diet challenge. These data support a causative role for Npt2b in the pathogenesis of PAM and the use of the PAM mouse model as a preclinical platform for the development of biomarkers and therapeutic strategies.

Surfactant protein D suppresses lung cancer progression by downregulation of epidermal growth factor signaling

Y. Hasegawa, M. Takahashi, S. Ariki, D. Asakawa, M. Tajiri, Y. Wada, Y. Yamaguchi, C. Nishitani, R. Takamiya, A. Saito, Y. Uehara, J. Hashimoto, Y. Kurimura, H. Takahashi, Y. Kuroki

ONCOGENE ( NATURE PUBLISHING GROUP ) 34 ( 7 ) 838 - 845 2015.02 [Refereed]

Surfactant protein D (SP-D) is a member of the collectin family that has an important role in maintaining pulmonary homeostasis. In this study, we demonstrated that SP-D inhibited the proliferation, migration and invasion of A549 human lung adenocarcinoma cells. We found that SP-D suppressed epidermal growth factor (EGF) signaling in A549 cells, H441 human lung adenocarcinoma cells and human EGF receptor (EGFR) stable expression CHO-K1 cells. A binding study using I-125-EGF demonstrated that SP-D downregulated the binding of EGF to EGFR. A ligand blot indicated that SP-D bound to EGFR, and a lectin blot suggested that EGFR in A549 cells had both high-mannose type and complex type N-glycans. We purified the recombinant extracellular domain of EGFR (soluble EGFR=soluble EGFR (sEGFR)), and demonstrated that SP-D directly bound to sEGFR in a Ca2+-dependent manner. The binding of SP-D to sEGFR was suppressed by EDTA, mannose or N-glycopeptidase F treatment. Mass spectrometric analysis indicated that N-glycans in domain III of EGFR were of a high-type. These data suggest that SP-D reduces EGF binding to EGFR through the interaction between the carbohydrate recognition domain of SP-D and N-glycans of EGFR, and downregulates EGF signaling. Our finding suggests the novel type of regulation system of EGF signaling involving lectin-to-carbohydrate interaction and downregulation of ligand binding.

Surfactant protein D suppresses lung cancer progression by downregulation of epidermal growth factor signalling (vol 34, pg 838, 2014)

Hasegawa Y, Takahashi M, Ariki S, Asakawa D, Tajiri M, Wada Y, Yamaguchi Y, Nishitani C, Takamiya R, Saito A, Uehara Y, Hashimoto J, Kurimura Y, Takahashi H, Kuroki Y

Oncogene 34 ( 32 ) 4285 - 4286 2015 [Refereed]

Surfactant Protein D Inhibits Adherence of Uropathogenic Escherichia coli to the Bladder Epithelial Cells and the Bacterium-induced Cytotoxicity A POSSIBLE FUNCTION IN URINARY TRACT

Yuichiro Kurimura, Chiaki Nishitani, Shigeru Ariki, Atsushi Saito, Yoshihiro Hasegawa, Motoko Takahashi, Jiro Hashimoto, Satoshi Takahashi, Taiji Tsukamoto, Yoshio Kuroki

JOURNAL OF BIOLOGICAL CHEMISTRY ( AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC ) 287 ( 47 ) 39578 - 39588 2012.11 [Refereed]

The adherence of uropathogenic Escherichia coli (UPEC) to the host urothelial surface is the first step for establishing UPEC infection. Uroplakin Ia (UPIa), a glycoprotein expressed on bladder urothelium, serves as a receptor for FimH, a lectin located at bacterial pili, and their interaction initiates UPEC infection. Surfactant protein D (SP-D) is known to be expressed on mucosal surfaces in various tissues besides the lung. However, the functions of SP-D in the non-pulmonary tissues are poorly understood. The purposes of this study were to investigate the possible function of SP-D expressed in the bladder urothelium and the mechanisms by which SP-D functions. SP-D was expressed in human bladder mucosa, and its mRNA was increased in the bladder of the UPEC infection model in mice. SP-D directly bound to UPEC and strongly agglutinated them in a Ca2+-dependent manner. Co-incubation of SP-D with UPEC decreased the bacterial adherence to 5637 cells, the human bladder cell line, and the UPEC-induced cytotoxicity. In addition, preincubation of SP-D with 5637 cells resulted in the decreased adherence of UPEC to the cells and in a reduced number of cells injured by UPEC. SP-D directly bound to UPIa and competed with FimH for UPIa binding. Consistent with the in vitro data, the exogenous administration of SP-D inhibited UPEC adherence to the bladder and dampened UPEC-induced inflammation in mice. These results support the conclusion that SP-D can protect the bladder urothelium against UPEC infection and suggest a possible function of SP-D in urinary tract.

Pulmonary Surfactant Protein A Protects Lung Epithelium from Cytotoxicity of Human beta-Defensin 3

Atsushi Saito, Shigeru Ariki, Hitoshi Sohma, Chiaki Nishitani, Kanako Inoue, Nobutaka Ebata, Motoko Takahashi, Yoshihiro Hasegawa, Koji Kuronuma, Hiroki Takahashi, Yoshio Kuroki

JOURNAL OF BIOLOGICAL CHEMISTRY ( AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC ) 287 ( 18 ) 15034 - 15043 2012.04 [Refereed]

Defensins are important molecules in the innate immune system that eliminate infectious microbes. They also exhibit cytotoxicity against host cells in higher concentrations. The mechanisms by which hosts protect their own cells from cytotoxicity of defensins have been poorly understood. We found that the cytotoxicity of human beta-defensin 3 (hBD3) against lung epithelial cells was dose-dependently attenuated by pulmonary surfactant protein A (SP-A), a collectin implicated in host defense and regulation of inflammatory responses in the lung. The direct interaction between SP-A and hBD3 may be an important factor in decreasing this cytotoxicity because preincubation of epithelial cells with SP-A did not affect the cytotoxicity. Consistent with in vitro analysis, intratracheal administration of hBD3 to SP-A(-/-) mice resulted in more severe tissue damage compared with that in WT mice. These data indicate that SP-A protects lung epithelium from tissue injury caused by hBD3. Furthermore, we found that the functional region of SP-A lies within Tyr(161)-Lys(201). Synthetic peptide corresponding to this region, tentatively called SP-A Y161-G200, also inhibited cytotoxicity of hBD3 in a dose-dependent manner. The SP-A Y161-G200 is a candidate as a therapeutic reagent that prevents tissue injury during inflammation.

肺コレクチンはレジオネラ感染により誘導されるオートファジーを抑制する

齋藤充史, 有木茂, 長谷川喜弘, 栗村雄一郎, 西谷千明, 高橋素子, 高橋弘毅, 黒木由夫

日本肺サーファクタント・界面医学会雑誌 ( 日本肺サーファクタント・界面医学会 ) 42 32 - 32 2011.10

Pulmonary Collectins Play Distinct Roles in Host Defense against Mycobacterium avium

Shigeru Ariki, Takashi Kojima, Shinsei Gasa, Atsushi Saito, Chiaki Nishitani, Motoko Takahashi, Takeyuki Shimizu, Yuichiro Kurimura, Norimasa Sawada, Nobuhiro Fujii, Yoshio Kuroki

JOURNAL OF IMMUNOLOGY ( AMER ASSOC IMMUNOLOGISTS ) 187 ( 5 ) 2586 - 2594 2011.09 [Refereed]

Pulmonary collectins, surfactant protein A (SP-A) and surfactant protein D (SP-D), play important roles in the innate immunity of the lung. Mycobacterium avium is one of the well-known opportunistic pathogens that can replicate within macrophages. We examined the effects of pulmonary collectins in host defense against M. avium infection achieved via direct interaction between bacteria and collectins. Although both pulmonary collectins bound to M. avium in a Ca(2+)-dependent manner, these collectins revealed distinct ligand-binding specificity and biological activities. SP-A and SP-D bound to a methoxy group containing lipid and lipoarabinomannan, respectively. Binding of SP-D but not SP-A resulted in agglutination of M. avium. A chimeric protein with the carbohydrate recognition domain of SP-D, which chimera revealed a bouquet-like arrangement similar to SP-A, also agglutinated M. avium. The ligand specificity of the carbohydrate recognition domain of SP-D seems to be necessary for agglutination activity. The binding of SP-A strongly inhibited the growth of M. avium in culture media. Although pulmonary collectins did not increase membrane permeability of M. avium, they attenuated the metabolic rate of the bacteria. Observations under a scanning electron microscope revealed that SP-A almost completely covers bacterial surfaces, whereas SP-D binds to certain areas like scattered dots. These observations suggest that a distinct binding pattern of collectins correlates with the difference of their biological activities. Furthermore, the number of bacteria phagocytosed by macrophages was significantly increased in the presence of SP-D. These data indicate that pulmonary collectins play critical roles in host defense against M. avium. The Journal of Immunology, 2011, 187: 2586-2594.

Pulmonary Collectins Protect Macrophages against Pore-forming Activity of Legionella pneumophila and Suppress Its Intracellular Growth

Kaku Sawada, Shigeru Ariki, Takashi Kojima, Atsushi Saito, Masami Yamazoe, Chiaki Nishitani, Takeyuki Shimizu, Motoko Takahashi, Hiroaki Mitsuzawa, Shin-ichi Yokota, Norimasa Sawada, Nobuhiro Fujii, Hiroki Takahashi, Yoshio Kuroki

JOURNAL OF BIOLOGICAL CHEMISTRY ( AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC ) 285 ( 11 ) 8434 - 8443 2010.03 [Refereed]

Pulmonary collectins, surfactant proteins A (SP-A) and D (SP-D), play important roles in innate immunity of the lung. Legionella pneumophila is a bacterial respiratory pathogen that can replicate within macrophages and causes opportunistic infections. L. pneumophila possesses cytolytic activity, resulting from insertion of pores in the macrophage membrane upon contact. We examined whether pulmonary collectins play protective roles against L. pneumophila infection. SP-A and SP-D bound to L. pneumophila and its lipopolysaccharide (LPS) and inhibited the bacterial growth in a Ca(2+)-dependent manner. The addition of LPS in the culture blocked the inhibitory effects on L. pneumophila growth by the collectins, indicating the importance of LPS-collectin interaction. When differentiated THP-1 cells were infected with L. pneumophila in the presence of SP-A and SP-D, the number of permeable cells was significantly decreased, indicating that pulmonary collectins inhibit pore-forming activity of L. pneumophila. The number of live bacteria within the macrophages on days 1-4 after infection was significantly decreased when infection was performed in the presence of pulmonary collectins. The phagocytosis experiments with the pH-sensitive dye-labeled bacteria revealed that pulmonary collectins promoted bacterial localization to an acidic compartment. In addition, SP-A and SP-D significantly increased the number of L. pneumophila co-localized with LAMP-1. These results indicate that pulmonary collectins protect macrophages against contact-dependent cytolytic activity of L. pneumophila and suppress intracellular growth of the phagocytosed bacteria. The promotion of lysosomal fusion with Legionella-containing phagosomes constitutes a likely mechanism of L. pneumophila growth suppression by the collectins.

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