Kyushu University Academic Staff Educational and Research Activities Database
List of Papers
Eiichi Hasegawa Last modified date:2020.06.08

Assistant Professor / Ophthalmology / Kyushu University Hospital

1. Eiichi Hasegawa, Atsunobu Takeda, Nobuyo Yawata, Koh Hei Sonoda, The effectiveness of adalimumab treatment for non-infectious uveitis, Immunological Medicine, 10.1080/25785826.2019.1642080, 42, 2, 79-83, 2019.04, Uveitis, which is a major cause of blindness worldwide, is defined as intraocular inflammation that affects the iris, ciliary body, vitreous, retina and choroid. Tumor necrosis factor-alpha (TNF-α) is a key cytokine involved in the pathogenesis of many inflammatory diseases including uveitis. Corticosteroids and immunosuppressive agents are the conventional therapy to treat non-infectious uveitis. In cases that are resistant to these therapies, anti-TNF agents are added. An anti-TNF-α agent, adalimumab, was recently approved for the treatment of refractory non-infectious uveitis. In this review, we provide an introduction to uveitis and summarize the effectiveness and safety of adalimumab in the treatment of non-infectious uveitis..
2. Keijiro Ishikawa, Ri Ichiro Kohno, Eiichi Hasegawa, shintaro nakao, Shigeo Yoshida, Kohei Sonoda, Preoperative estimation of distance between retinal break and limbus with wide-field fundus imaging
Potential clinical utility for conventional scleral buckling, PloS one, 10.1371/journal.pone.0212284, 14, 2, 2019.02, Objective :Accurate scleral marking of retinal breaks is essential for successful scleral buckling. This study aimed to investigate the use of wide-field fundus images obtained with an Optos for preoperative estimation of the distance from the limbus to the retinal breaks. Methods and analysis: This is a retrospective review of 29 eyes from 26 patients with rhegmatogenous retinal detachment who received scleral buckling with anatomically successful repair. They underwent wide-field fundus photography with Optos California. In the pre- and postoperative fundus images, we measured distances from the macula to the retinal tears (T
), to the center of the vortex veins (V
), to the optic disc (D
), and to the posterior edge of the scleral buckle (B
). Results: (BM-V
) / D
was significantly correlated with the distance from the limbus to the posterior edge of the scleral buckle that had been determined intraoperatively. (r = 0.705; p<0.001) We applied a regression line derived from this correlation with the value of (T
) / D
in order to calculate estimated distances between retinal breaks and the limbus. The calculated distances were all within the range of distances from the limbus to the anterior and posterior edges of the scleral buckles. Conclusion: Preoperative analysis of Optos images may be useful for estimating the distance from the limbus to retinal breaks, which might aid scleral marking during scleral buckling surgery..
3. Ryo Mukai, Dong Ho Park, Yoko Okunuki, Eiichi Hasegawa, Garrett Klokman, Clifford B. Kim, Anitha Krishnan, Meredith Gregory-Ksander, Deeba Husain, Joan W. Miller, Kip M. Connor, Mouse model of ocular hypertension with retinal ganglion cell degeneration, PloS one, 10.1371/journal.pone.0208713, 14, 1, 2019.01, Objectives Ocular hypertension is a primary risk factor for glaucoma and results in retinal ganglion cell (RGC) degeneration. Current animal models of glaucoma lack severe RGC cell death as seen in glaucoma, making assessment of physiological mediators of cell death difficult. We developed a modified mouse model of ocular hypertension whereby long-lasting elevation of intraocular pressure (IOP) is achieved, resulting in significant reproducible damage to RGCs. Results In this model, microbeads are mixed with hyaluronic acid and injected into the anterior chamber of C57BL/6J mice. The hyaluronic acid allows for a gradual release of microbeads, resulting in sustained blockage of Schlemm’s canal. IOP elevation was bimodal during the course of the model’s progression. The first peak occurred 1 hours after beads injection, with an IOP value of 44.69 ± 6.00 mmHg, and the second peak occurred 6–12 days post-induction, with an IOP value of 34.91 ± 5.21 mmHg. RGC damage was most severe in the peripheral retina, with a loss of 64.1% compared to that of untreated eyes, while the midperiphery exhibited a 32.4% loss, 4 weeks following disease induction. Conclusions These results suggest that sustained IOP elevation causes more RGC damage in the periphery than in the midperiphery of the retina. This model yields significant and reproducible RGC degeneration..
4. Takako Ito, Atsunobu Takeda, Kohta Fujiwara, Eiichi Hasegawa, shintaro nakao, Yoshihiro Ohishi, Yoshinao Oda, Hiroshi Yoshikawa, Kohei Sonoda, Risk factors for failure of vitrectomy cell block technique in cytological diagnosis of vitreoretinal lymphoma, Graefe's Archive for Clinical and Experimental Ophthalmology, 10.1007/s00417-019-04266-6, 2019.01, Purpose: To determine the factors that may affect the accuracy of vitrectomy cell block technique in detecting atypical lymphoid cells in patients with vitreoretinal lymphoma (VRL). Methods: We retrospectively reviewed 43 eyes in 39 patients who underwent vitrectomy for definitive histological diagnosis of VRL with vitrectomy cell block technique and/or smear preparation at Kyushu University Hospital from January 2001 to March 2016. The association of detection of atypical lymphoid cells using vitrectomy cell block technique with the following factors was assessed using logistic regression analysis: age at diagnosis, sex, presence or absence of concurrent cataract surgery with vitrectomy, clinical grading of vitreous haze, presence or absence of subretinal tumor infiltration, interval between initial symptoms and vitrectomy, and presence or absence of systemic corticosteroid therapy before vitrectomy. Results: Atypical lymphoid cells were more significantly detected using vitrectomy cell block technique compared to that using smear preparation (p = 0.018). After adjusting for age and sex, concurrent cataract surgery (odds ratio [OR], 10.41; 95% confidence interval [CI], 1.42–76.41) and subretinal tumor infiltration (OR, 5.06; 95% CI, 1.06–24.32) were significantly associated with failure of histological analysis with vitrectomy cell blocks. In multivariable logistic regression analysis, similar results were obtained, although subretinal tumor infiltration was only marginally associated with the detective capability of the technique. Conclusion: Vitrectomy cell block technique significantly improved the definitive diagnosis of VRL. Concurrent cataract surgery with vitrectomy and subretinal tumor infiltration were risk factors for failure in vitrectomy cell blocks..
5. Takashi Ueta, Kenji Ishihara, Shoji Notomi, Jong Jer Lee, Daniel E. Maidana, Nikolaos E. Efstathiou, Yusuke Murakami, Eiichi Hasegawa, Kunihiro Azuma, Tetsuya Toyono, Eleftherios I. Paschalis, Makoto Aihara, Joan W. Miller, Demetrios G. Vavvas, RIP1 kinase mediates angiogenesis by modulating macrophages in experimental neovascularization, Proceedings of the National Academy of Sciences of the United States of America, 10.1073/pnas.1908355116, 116, 47, 23705-23713, 2019.01, Inflammation plays an important role in pathological angiogenesis. Receptor-interacting protein 1 (RIP1) is highly expressed in inflammatory cells and is known to play an important role in the regulation of apoptosis, necroptosis, and inflammation; however, a comprehensive description of its role in angiogenesis remains elusive. Here, we show that RIP1 is abundantly expressed in infiltrating macrophages during angiogenesis, and genetic or pharmacological inhibition of RIP1 kinase activity using kinase-inactive RIP1K45A/K45A mice or necrostatin-1 attenuates angiogenesis in laser-induced choroidal neovascularization, Matrigel plug angiogenesis, and alkali injury-induced corneal neovascularization in mice. The inhibitory effect on angiogenesis is mediated by caspase activation through a kinase-independent function of RIP1 and RIP3. Mechanistically, infiltrating macrophages are the key target of RIP1 kinase inhibition to attenuate pathological angiogenesis. Inhibition of RIP1 kinase activity is associated with caspase activation in infiltrating macrophages and decreased expression of proangiogenic M2-like markers but not M1-like markers. Similarly, in vitro, catalytic inhibition of RIP1 down-regulates the expression of M2-like markers in interleukin-4–activated bone marrow-derived macrophages, and this effect is blocked by simultaneous caspase inhibition. Collectively, these results demonstrate a nonnecrotic function of RIP1 kinase activity and suggest that RIP1-mediated modulation of macrophage activation may be a therapeutic target of pathological angiogenesis..
6. Atsunobu Takeda, Hisakata Yamada, Eiichi Hasegawa, Mitsuru Arima, Shoji Notomi, Sayaka Myojin, Takeru Yoshimura, Toshio Hisatomi, Hiroshi Enaida, Ryoji Yanai, Kazuhiro Kimura, Tatsuro Ishibashi, Kohei Sonoda, Crucial role of P2X7 receptor for effector T cell activation in experimental autoimmune uveitis, Japanese Journal of Ophthalmology, 10.1007/s10384-018-0587-4, 62, 3, 398-406, 2018.05, Purpose: To investigate the roles of P2X7 receptors (P2RX7) in the pathogenesis of experimental autoimmune uveoretinitis (EAU). Study design: Experimental. Methods: Either wild-type (P2rx7+/+) or P2rx7-deficient (P2rx7) mice were immunized with interphotoreceptor retinoid-binding protein (IRBP) peptide 1-20. Severity of EAU was evaluated clinically and histopathologically. The induction of IRBP-specific proliferation and cytokines in draining lymph nodes was assessed by enzyme-linked immunosorbent assays (ELISA). The frequency of activation markers was examined by flow cytometry. Furthermore, inhibitory roles of systemic administration of Brilliant Blue G (BBG), an antagonist for P2RX7, in EAU were also assessed in the wild-type mice. Results: The severity of EAU in P2rx7 mice was reduced as compared with that in P2rx7+/+ mice, both clinically and histopathologically. IRBP-specific proliferation in P2rx7 on day 16 was slightly decreased compared to that in P2rx7+/+ mice. The induction of IRBP-specific interferon (IFN)-γ and interleukin (IL)-17 in P2rx7 mice on day 16 was lower than that in P2rx7+/+ mice. The up-regulation of surface expression of activation markers such as CD25, CD44, and CD69 in response to TCR stimulation in P2rx7 mice was decreased as compared with that in P2rx7+/+ mice. Furthermore, neutralization of P2RX7 in vivo by BBG suppressed EAU clinically and histopathologically. IRBP-specific IFN-γ and IL-17 induction in BBG-treated mice was significantly lower than that in vehicle-treated mice. Conclusion: The results suggest that P2RX7 is a novel preventative therapeutic target for uveitis as it suppresses the effector functions of both Th1 and Th17 cell responses..
7. Kota Sato, Atsunobu Takeda, Eiichi Hasegawa, Young Joon Jo, Mitsuru Arima, Yuji Oshima, Yanai Ryoji, Toru Nakazawa, Mitsuko Yuzawa, Hiroyuki Nakashizuka, Hiroyuki Shimada, Kazuhiro Kimura, Tatsuro Ishibashi, Kohei Sonoda, Interleukin-6 plays a crucial role in the development of subretinal fibrosis in a mouse model, Immunological Medicine, 10.1080/09114300.2018.1451609, 41, 1, 23-29, 2018.01, Subretinal fibrosis has been recognized as a feature of an advanced stage of exudative age-related macular degeneration (AMD) that leads to irreversible loss of vision. This study was aimed at elucidating roles of interlukin-6 (IL-6) in the development of subretinal fibrosis. Immunohistochemistry (IHC) was performed with anti-human IL-6 antibody in surgically excised choroidal neovascular tissues from patients with exudative AMD. The area of subretinal fibrosis was measured in a mouse subretinal fibrosis model with injection of control small interfering RNA(siRNA) or IL-6 siRNA, or isotype control antibody or anti-IL-6 receptor antibody after peritoneal exudative cells (PECs) injection into the vitreous cavity. PECs derived from IL-6
or IL-6
mice were placed into the subretinal space of IL-6
mice. IL-6 was expressed in the stroma and retinal pigment epithelial (RPE) layer in the choroidal neovascular tissues. IL-6 knockdown or blocking of the IL-6 receptor suppressed the formation of subretinal fibroblastic scars. The area of subretinal fibrosis induced by PECs derived from IL-6
mice was less than that induced by PECs from IL-6
mice. The results suggested that IL-6, expressed by activated macrophages, is a crucial mediator that promotes subretinal fibrosis. Targeting IL-6 and the corresponding signaling pathway would be an attractive therapeutic approach not only in choroidal neovascularization, but also in subretinal fibrosis..
8. Eiichi Hasegawa, Saori Inafuku, Lama Mulki, Yoko Okunuki, Ryoji Yanai, Kaylee E. Smith, Clifford B. Kim, Garrett Klokman, Diane R. Bielenberg, Narender Puli, John R. Falck, Deeba Husain, Joan W. Miller, Matthew L. Edin, Darryl C. Zeldin, Kin Sing Stephen Lee, Bruce D. Hammock, Wolf Hagen Schunck, Kip M. Connor, Cytochrome P450 monooxygenase lipid metabolites are significant second messengers in the resolution of choroidal neovascularization, Proceedings of the National Academy of Sciences of the United States of America, 10.1073/pnas.1620898114, 114, 36, E7545-E7553, 2017.09, Age-related macular degeneration (AMD) is the most common cause of blindness for individuals age 50 and above in the developed world. Abnormal growth of choroidal blood vessels, or choroidal neovascularization (CNV), is a hallmark of the neovascular (wet) form of advanced AMD and leads to significant vision loss. A growing body of evidence supports a strong link between neovascular disease and inflammation. Metabolites of long-chain polyunsaturated fatty acids derived from the cytochrome P450 (CYP) monooxygenase pathway serve as vital second messengers that regulate a number of hormones and growth factors involved in inflammation and vascular function. Using transgenic mice with altered CYP lipid biosynthetic pathways in a mouse model of laser-induced CNV, we characterized the role of these lipid metabolites in regulating neovascular disease. We discovered that the CYP-derived lipid metabolites epoxydocosapentaenoic acids (EDPs) and epoxyeicosatetraenoic acids (EEQs) are vital in dampening CNV severity. Specifically, overexpression of the monooxygenase CYP2C8 or genetic ablation or inhibition of the soluble epoxide hydrolase (sEH) enzyme led to increased levels of EDP and EEQ with attenuated CNV development. In contrast, when we promoted the degradation of these CYP-derived metabolites by transgenic overexpression of sEH, the protective effect against CNV was lost. We found that these molecules work in part through their ability to regulate the expression of key leukocyte adhesion molecules, on both leukocytes and endothelial cells, thereby mediating leukocyte recruitment. These results suggest that CYP lipid signaling molecules and their regulators are potential therapeutic targets in neovascular diseases..
9. J. Harry Sweigard, Hidetaka Matsumoto, Kaylee E. Smith, Leo A. Kim, Eleftherios I. Paschalis, Yoko Okonuki, Alexandra Castillejos, Keiko Kataoka, Eiichi Hasegawa, Ryoji Yanai, Deeba Husain, John D. Lambris, Demetrios Vavvas, Joan W. Miller, Kip M. Connor, Inhibition of the alternative complement pathway preserves photoreceptors after retinal injury, Science Translational Medicine, 10.1126/scitranslmed.aab1482, 7, 297, 2015.07, Degeneration of photoreceptors is a primary cause of vision loss worldwide, making the underlying mechanisms surrounding photoreceptor cell death critical to developing new treatment strategies. Retinal detachment, characterized by the separation of photoreceptors from the underlying retinal pigment epithelium, is a sight-threatening event that can happen in a number of retinal diseases. The detached photoreceptors undergo apoptosis and programmed necrosis. Given that photoreceptors are nondividing cells, their loss leads to irreversible visual impairment even after successful retinal reattachment surgery. To better understand the underlying disease mechanisms, we analyzed innate immune system regulators in the vitreous of human patients with retinal detachment and correlated the results with findings in a mouse model of retinal detachment. We identified the alternative complement pathway as promoting early photoreceptor cell death during retinal detachment. Photoreceptors down-regulate membrane-bound inhibitors of complement, allowing for selective targeting by the alternative complement pathway. When photoreceptors in the detached retina were removed from the primary source of oxygen and nutrients (choroidal vascular bed), the retina became hypoxic, leading to an up-regulation of complement factor B, a key mediator of the alternative pathway. Inhibition of the alternative complement pathway in knockout mice or through pharmacological means ameliorated photoreceptor cell death during retinal detachment. Our current study begins to outline the mechanism by which the alternative complement pathway facilitates photoreceptor cell death in the damaged retina..
10. James F. Sampson, Eiichi Hasegawa, Lama Mulki, Amol Suryawanshi, Shuhong Jiang, Wei Sheng Chen, Gabriel A. Rabinovich, Kip M. Connor, Noorjahan Panjwani, Galectin-8 ameliorates murine autoimmune ocular pathology and promotes a regulatory T cell response, PLoS One, 10.1371/journal.pone.0130772, 10, 6, 2015.06, Galectins have emerged as potent immunoregulatory agents that control chronic inflammation through distinct mechanisms. Here, we report that treatment with Galectin-8 (Gal-8), a tandem-repeat member of the galectin family, reduces retinal pathology and prevents photoreceptor cell damage in a murine model of experimental autoimmune uveitis. Gal-8 treatment increased the number of regulatory T cells (Treg) in both the draining lymph node (dLN) and the inflamed retina. Moreover, a greater percentage of Treg cells in the dLN and retina of Gal-8 treated animals expressed the inhibitory coreceptor cytotoxic T lymphocyte antigen (CTLA)-4, the immunosuppressive cytokine IL-10, and the tissue-homing integrin CD103. Treg cells in the retina of Gal-8-treated mice were primarily inducible Treg cells that lack the expression of neuropilin-1. In addition, Gal-8 treatment blunted production of inflammatory cytokines by retinal T helper type (TH) 1 and TH17 cells. The effect of Gal-8 on T cell differentiation and/or function was specific for tissues undergoing an active immune response, as Gal-8 treatment had no effect on T cell populations in the spleen. Given the need for rational therapies for managing human uveitis, Gal-8 emerges as an attractive therapeutic candidate not only for treating retinal autoimmune diseases, but also for other TH1- and TH17-mediated inflammatory disorders..
11. Eiichi Hasegawa, Harry Sweigard, Deeba Husain, Ana M. Olivares, Bo Chang, Kaylee E. Smith, Amy E. Birsner, Robert J. D'Amato, Norman A. Michaud, Yinan Han, Demetrios G. Vavvas, Joan W. Miller, Neena B. Haider, Kip M. Connor, Characterization of a spontaneous retinal neovascular mouse model, PLoS One, 10.1371/journal.pone.0106507, 9, 9, 2014.09, Background: Vision loss due to vascular disease of the retina is a leading cause of blindness in the world. Retinal angiomatous proliferation (RAP) is a subgroup of neovascular age-related macular degeneration (AMD), whereby abnormal blood vessels develop in the retina leading to debilitating vision loss and eventual blindness. The novel mouse strain, neoretinal vascularization 2 (NRV2), shows spontaneous fundus changes associated with abnormal neovascularization. The purpose of this study is to characterize the induction of pathologic angiogenesis in this mouse model. Methods: The NRV2 mice were examined from postnatal day 12 (p12) to 3 months. The phenotypic changes within the retina were evaluated by fundus photography, fluorescein angiography, optical coherence tomography, and immunohistochemical and electron microscopic analysis. The pathological neovascularization was imaged by confocal microscopy and reconstructed using three-dimensional image analysis software. Results: We found that NRV2 mice develop multifocal retinal depigmentation in the posterior fundus. Depigmented lesions developed vascular leakage observed by fluorescein angiography. The spontaneous angiogenesis arose from the retinal vascular plexus at postnatal day (p)15 and extended toward retinal pigment epithelium (RPE). By three months of age, histological analysis revealed encapsulation of the neovascular lesion by the RPE in the photoreceptor cell layer and subretinal space. Conclusions: The NRV2 mouse strain develops early neovascular lesions within the retina, which grow downward towards the RPE beginning at p15. This retinal neovascularization model mimics early stages of human retinal angiomatous proliferation (RAP) and will likely be a useful in elucidating targeted therapeutics for patients with ocular neovascular disease..
12. Ryoji Yanai, Lama Mulki, Eiichi Hasegawa, Kimio Takeuchi, Harry Sweigard, Jun Suzuki, Philipp Gaissert, Demetrios G. Vavvas, Kohei Sonoda, Michael Rothe, Wolf Hagen Schunck, Joan W. Miller, Kip M. Connor, Cytochrome P450-generated metabolites derived from ω-3 fatty acids attenuate neovascularization, Proceedings of the National Academy of Sciences of the United States of America, 10.1073/pnas.1401191111, 111, 26, 9603-9608, 2014.07, Ocular neovascularization, including age-related macular degeneration (AMD), is a primary cause of blindness in individuals of industrialized countries. With a projected increase in the prevalence of these blinding neovascular diseases, there is an urgent need for new pharmacological interventions for their treatment or prevention. Increasing evidence has implicated eicosanoid-like metabolites of long-chain polyunsaturated fatty acids (LCPUFAs) in the regulation of neovascular disease. In particular, metabolites generated by the cytochrome P450 (CYP)-epoxygenase pathway have been shown to be potent modulators of angiogenesis, making this pathway a reasonable previously unidentified target for intervention in neovascular ocular disease. Here we show that dietary supplementation with ω-3 LCPUFAs promotes regression of choroidal neovessels in a well-characterized mouse model of neovascular AMD. Leukocyte recruitment and adhesion molecule expression in choroidal neovascular lesions were down-regulated in mice fed ω-3 LCPUFAs. The serum of these mice showed increased levels of anti-inflammatory eicosanoids derived from eicosapentaenoic acid and docosahexaenoic acid. 17,18-epoxyeicosatetraenoic acid and 19,20-epoxydocosapentaenoic acid, the major CYP-generated metabolites of these primary ω-3 LCPUFAs, were identified as key lipid mediators of disease resolution. We conclude that CYP-derived bioactive lipid metabolites from ω-3 LCPUFAs are potent inhibitors of intraocular neovascular disease and show promising therapeutic potential for resolution of neovascular AMD..
13. Atsunobu Takeda, Eiichi Hasegawa, Takako Fukuhara, Sayaka Hirakawa, Hisakata Yamada, Yang Yang, Takeru Yoshimura, Toshio Hisatomi, Yuji Oshima, Hiroki Yoshida, Kohei Sonoda, Tatsuro Ishibashi, EBI3 is pivotal for the initiation of experimental autoimmune uveitis, Experimental Eye Research, 10.1016/j.exer.2014.06.004, 125, 107-113, 2014.01, Murine experimental autoimmune uveitis (EAU) is a model for human autoimmune uveitis, whose pathogenesis is caused by both Th1 and Th17cell responses. Epstein-Barr virus-induced gene 3 (EBI3) is a component of the heterodimeric cytokines: interleukin (IL)-27 and IL-35. Although IL-27 was shown to initiate Th1 cell development, it is also recognized as a negative regulator of fully activated CD4+ T cells, including Th17cells. Recently, IL-35 also has also been reported to play immunosuppressive roles in autoimmunity. To investigate the roles of EBI3 in EAU, EBI3-/- mice were immunized with human interphotoreceptor retinoid binding protein peptide 1-20 (IRBP) to induce EAU. We observed that the clinical score in EBI3-/- mice was diminished compared with that in EBI3+/+ mice up to day 22 after immunization, whereas the score in EBI3-/- mice reached the same levels as that of EBI3+/+ mice after day 28. Histological analysis revealed a significant reduction of cellular infiltration into the retina in EBI3-/- mice on day 16. Although Th1 cell responses and IRBP-specific IL-10 production were reduced in EBI3-/- mice, the development of Th17cell responses was unaffected on day 9. On day 21, Th1 cell responses and IRBP-specific IL-10 production was restored to the same levels as that in EBI3+/+ mice, and Th17cell responses significantly increased in EBI3-/- mice. Furthermore, Foxp3 expression in CD4+ T cells was comparable between EBI3+/+ and EBI3-/- mice on days 9 and 21. Therefore, these results indicate that EBI3 may be important in EAU initiation by Th1 cell responses and may suppress EAU by inhibition of both Th1 and Th17cell responses in the late/maintenance phase..
14. Eiichi Hasegawa, Kohei Sonoda, Takashi Shichita, Rimpei Morita, Takashi Sekiya, Akihiro Kimura, Yuji Oshima, Atsunobu Takeda, Takeru Yoshimura, Shigeo Yoshida, Tatsuro Ishibashi, Akihiko Yoshimura, IL-23-independent induction of IL-17 from γδT cells and innate lymphoid cells promotes experimental intraocular neovascularization, Journal of Immunology, 10.4049/jimmunol.1202495, 190, 4, 1778-1787, 2013.02, Choroidal neovascularization (CNV) is a characteristic of age-related macular degeneration. Genome-wide association studies have provided evidence that the immune system is involved in the pathogenesis of age-related macular degeneration; however, the role of inflammatory cytokines in CNV has not been established. In this study, we demonstrated that IL-17 had a strong potential for promoting neovascularization in a vascular endothelial growth factor-independent manner in laser-induced experimental CNV in mice. Infiltrated γδT cells and Thy-1+ innate lymphoid cells, but not Th17 cells, were the main sources of IL-17 in injured eyes. IL-23 was dispensable for IL-17 induction in the eye. Instead, we found that IL-1β and high-mobility group box 1 strongly promoted IL-17 expression by γδT cells. Suppression of IL-1β and high-mobility group box 1, as well as depletion of γδT cells, reduced IL-17 levels and ameliorated experimental CNV. Our findings suggest the existence of a novel inflammatory cytokine network that promotes neovascularization in the eye..
15. Ryoko Yoshida, Mayu Suzuki, Ryota Sakaguchi, Eiichi Hasegawa, Akihiro Kimura, Takashi Shichita, Takashi Sekiya, Hiroshi Shiraishi, Kouji Shimoda, Akihiko Yoshimura, Forced expression of stabilized c-Fos in dendritic cells reduces cytokine production and immune responses in vivo, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2012.05.097, 423, 2, 247-252, 2012.06, Intracellular cyclic adenosine monophosphate (cAMP) suppresses innate immunity by inhibiting proinflammatory cytokine production by monocytic cells. We have shown that the transcription factor c-Fos is responsible for cAMP-mediated suppression of inflammatory cytokine production, and that c-Fos protein is stabilized by IKKβ-mediated phosphorylation. We found that S308 is one of the major phosphorylation sites, and that the S308D mutation prolongs c-Fos halflife. To investigate the role of stabilized c-Fos protein in dendritic cells (DCs) in vivo, we generated CD11c-promoter-deriven c-FosS308D transgenic mice. As expected, bone marrow-derived DCs (BMDCs) from these Tg mice produced smaller amounts of inflammatory cytokines, including TNF-α, IL-12, and IL-23, but higher levels of IL-10, in response to LPS, than those from wild-type (Wt) mice. When T cells were co-cultured with BMDCs from Tg mice, production of Th1 and Th17 cytokines was reduced, although T cell proliferation was not affected. Tg mice demonstrated more resistance to experimental autoimmune encephalomyelitis (EAE) than did Wt mice. These data suggest that c-Fos in DCs plays a suppressive role in certain innate and adaptive immune responses..
16. Takashi Shichita, Eiichi Hasegawa, Akihiro Kimura, Rimpei Morita, Ryota Sakaguchi, Ichiro Takada, Takashi Sekiya, Hiroaki Ooboshi, Takanari Kitazono, Toru Yanagawa, Tetsuro Ishii, Hideo Takahashi, Shuji Mori, Masahiro Nishibori, Kazumichi Kuroda, Shizuo Akira, Kensuke Miyake, Akihiko Yoshimura, Peroxiredoxin family proteins are key initiators of post-ischemic inflammation in the brain, Nature Medicine, 10.1038/nm.2749, 18, 6, 911-917, 2012.06, Post-ischemic inflammation is an essential step in the progression of brain ischemia-reperfusion injury. However, the mechanism that activates infiltrating macrophages in the ischemic brain remains to be clarified. Here we demonstrate that peroxiredoxin (Prx) family proteins released extracellularly from necrotic brain cells induce expression of inflammatory cytokines including interleukin-23 in macrophages through activation of Toll-like receptor 2 (TLR2) and TLR4, thereby promoting neural cell death, even though intracellular Prxs have been shown to be neuroprotective. The extracellular release of Prxs in the ischemic core occurred 12 h after stroke onset, and neutralization of extracellular Prxs with antibodies suppressed inflammatory cytokine expression and infarct volume growth. In contrast, high mobility group box 1 (HMGB1), a well-known damage-associated molecular pattern molecule, was released before Prx and had a limited role in post-ischemic macrophage activation. We thus propose that extracellular Prxs are previously unknown danger signals in the ischemic brain and that its blocking agents are potent neuroprotective tools..
17. Eiichi Hasegawa, Yuji Oshima, Atsunobu Takeda, Kazuko Saeki, Hiroki Yoshida, Kohei Sonoda, Tatsuro Ishibashi, IL-27 inhibits pathophysiological intraocular neovascularization due to laser burn, Journal of Leukocyte Biology, 10.1189/jlb.1110603, 91, 2, 267-273, 2012.02, AMD is the most common disease leading to acquired blindness in developed countries. CNV is the foremost cause of AMD and is thought to be induced by regional inflammation as a result of age-related conformational changes of the chorioretinal interface. Here, we show that IL-27, a member of the IL-6/IL-12 cytokine family, has an angiostatic effect and regulates the development of laser-induced experimental CNV in mice. In this model, IL-27 expression increased in the damaged choroid and peaked at the 24 h-time-point. IL-27 neutralization, induced by inoculating an antagonistic antibody into the vitreous cavity, enhanced VEGF production and the extent of CNV. By contrast, the administration of rIL-27 reduced VEGF production and the extent of CNV. Mice deficient in the EBI3, which lack IL-27, also showed more CNV than C57BL/6 mice, and this was reduced by IL-27 supplementation. We additionally investigated the effect of IL-27 on the function of mac-rophages, which play a critical role in CNV. IL-27 did not affect macrophage migration but inhibited its VEGF production. IL-27 therefore appears to regulate CNV and is a promising candidate target for treating sight-threatening diseases caused by ocular neovascularization..
18. Kiyokazu Hiwatashi, Taiga Tamiya, Eiichi Hasegawa, Tomohiro Fukaya, Masayuki Hashimoto, Kyosuke Kakoi, Ikko Kashiwagi, Akihiro Kimura, Naoko Inoue, Rimpei Morita, Hideo Yasukawa, Akihiko Yoshimura, Suppression of SOCS3 in macrophages prevents cancer metastasis by modifying macrophage phase and MCP2/CCL8 induction, Cancer Letters, 10.1016/j.canlet.2011.04.024, 308, 2, 172-180, 2011.09, Inflammation has been demonstrated to play important roles in tumorigenesis, tumor progression, and metastasis. STAT3 has been shown to be frequently activated in a variety of human cancer cells and STAT3 signaling promotes the growth and survival of tumor cells. However, the role of STAT3 of myeloid cells associated with tumors is currently unknown. Suppressor of cytokine signaling-3 (SOCS3) has been shown to be a negative regulator of STAT3. In this study, we used macrophage specific SOCS3 conditional knockout (cKO) mice to investigate the effect of the hyperactivation of STAT3 in macrophages on tumor development and metastasis. In a subcutaneous transplantation model of B16F10 melanoma cells, although tumor sizes were not significantly different, SOCS3-cKO mice survived longer than wild-type (WT) mice did. SOCS3-cKO mice exhibited fewer lung and liver metastatic tumor nodules than WT mice when B16F10 was challenged intravenously. SOCS3-/- macrophages stimulated with tumor lysates in vitro exhibited prolonged STAT3 phosphorylation and produced less amount of TNFα and IL-6, and higher amount of MCP2/CCL8 than WT macrophages. MCP/CCL8 was induced via STAT3 and exhibited anti-tumor metastatic effect in WT mice. These data suggest that hyperactivation of STAT3 in myeloid cells simultaneously exerted an anti-inflammatory as well as anti-tumor effects. Thus, the targeted inhibition of SOCS3 activity in macrophages may be therapeutic for the suppression of tumor metastasis..
19. T. Takimoto, Y. Wakabayashi, T. Sekiya, N. Inoue, R. Morita, K. Ichiyama, R. Takahashi, M. Asakawa, G. Muto, T. Mori, Eiichi Hasegawa, S. Shizuya, T. Hara, M. Nomura, A. Yoshimura, Smad2 and Smad3 are redundantly essential for the TGF-β-mediated regulation of regulatory T plasticity and Th1 development (Journal of Immunology (2010) 185, (842-855)), Journal of Immunology, 10.4049/jimmunol.1090121, 186, 1, 2011.01.
20. Fumie Konoeda, Takashi Shichita, Hideyuki Yoshida, Yuki Sugiyama, Go Muto, Eiichi Hasegawa, Rinpei Morita, Norihiro Suzuki, Akihiko Yoshimura, Therapeutic effect of IL-12/23 and their signaling pathway blockade on brain ischemia model, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2010.10.058, 402, 3, 500-506, 2010.11, Recently, T cell cytokines such as IL-17 and IFN-γ have been shown to play important roles in the progression of brain injury induced by ischemia. We have shown that IL-23 from infiltrated macrophages activates γδT cells, thereby inducing IL-17 from these cells. However, deletion of the IL-23 gene in mice showed a more dramatic protective effect against brain ischemia reperfusion (I/R) model than γδT cell depletion did, suggesting that IL-23 plays some other pivotal role in brain injury in addition to its role in IL-17 induction. To develop therapeutic methods based on these findings, we examined the effect of the JAK kinase inhibitor CP-690550 and an anti-IL12/23 monoclonal antibody on an I/R model. CP-690550 efficiently inhibited IL-17 production from memory T cells in vitro and partly suppressed infarct volume increase after I/R. Anti-p40 antibody, which blocks both IL-12 and IL-23, efficiently suppressed I/R injury and improved recovery of neurological deficits. The number of IL-17-producing cells was decreased by anti-p40 antibody treatment. Thus the JAK inhibitor and anti-p40 antibody, both of which have already been under trial for the treatment of several human inflammatory diseases, appear to be promising therapeutic agents for the amelioration of stroke..
21. Takeshi Fujimoto, Kohei Sonoda, Kuniaki Hijioka, Kohta Sato, Atsunobu Takeda, Eiichi Hasegawa, Yuji Oshima, Tatsuro Ishibashi, Choroidal neovascularization enhanced by chlamydia pneumoniae via toll-like receptor 2 in the retinal pigment epithelium, Investigative Ophthalmology and Visual Science, 10.1167/iovs.09-4464, 51, 9, 4694-4702, 2010.09, PURPOSE. Choroidal neovascularization (CNV) is directly related to visual loss in persons with age-related macular degeneration (AMD) and other macular disorders. Chlamydia pneumoniae, a prokaryotic pathogen that causes chronic inflammation, is recognized as a risk factor for cardiovascular diseases. In this study, the authors investigated the association between C. pneumoniae infection and AMD using a laser-induced CNV model in mice. METHODS. C57BL/6 mice, myeloid differentiation factor (MyD) 88 knockout (KO) mice, Toll-like receptor (TLR) 2 KO mice, and TLR4 KO mice were used. Experimental CNV was induced by rupturing the Bruch's membrane by laser photocoagulation (PC). Seven days after PC, the eyes were enucleated and the areas of CNV were measured in choroidal flat mounts. Cytokine gene expression by quantitative real-time PCR in the primary cultured retinal pigment epithelium (RPE) cells was also examined. RESULTS. Vitreous injection of the C. pneumoniae antigen increased the size of CNV. Although lipopolysaccharide stimulation can induce multiple cytokines, cultured mouse RPE cells from C57BL/6 mice expressed IL-6 and VEGF, but not TNF-α mRNA, in response to C. pneumoniae antigen. RPE cells from either MyD88 KO mice or TLR2 KO mice did not respond to the C. pneumoniae antigen. TLR2 KO mice did not augment the size increase of experimental CNV by C. pneumoniae antigen in vivo. CONCLUSIONS. C. pneumoniae can trigger inflammatory responses in the eye and promote experimental CNV in a TLR2- dependent manner. These data provide experimental evidence to imply persistent C. pneumoniae infection is a risk factor for AMD..
22. Tomohito Takimoto, Yu Wakabayashi, Takashi Sekiya, Naoko Inoue, Rimpei Morita, Kenji Ichiyama, Reiko Takahashi, Mayako Asakawa, Go Muto, Tomoaki Mori, Eiichi Hasegawa, Saika Shizuya, Toshiro Hara, Masatoshi Nomura, Akihiko Yoshimura, Smad2 and Smad3 are redundantly essential for the TGF-β-mediated regulation of regulatory T plasticity and Th1 development, Journal of Immunology, 10.4049/jimmunol.0904100, 185, 2, 842-855, 2010.07, Although it has been well established that TGF-β plays a pivotal role in immune regulation, the roles of its downstream transcription factors, Smad2 and Smad3, have not been fully clarified. Specifically, the function of Smad2 in the immune system has not been investigated because of the embryonic lethality of Smad2-deficient mice. In this study, we generated T cell-specific Smad2 conditional knockout (KO) mice and unexpectedly found that Smad2 and Smad3 were redundantly essential for TGF-β-mediated induction of Foxp3-expressing regulatory T cells and suppression of IFN-γ production in CD4+ T cells. Consistent with these observations, Smad2/Smad3-double KO mice, but not single KO mice, developed fatal inflammatory diseases with higher IFN-γ production and reduced Foxp3 expression in CD4+ T cells at the periphery. Although it has been suggested that Foxp3 induction might underlie TGF-β-mediated immunosuppression, TGF-β still can suppress Th1 cell development in Foxp3-deficient T cells, suggesting that the Smad2/3 pathway inhibits Th1 cell development with Foxp3-independent mechanisms. We also found that Th17 cell development was reduced in Smad-deficient CD4+ T cells because of higher production of Th17-inhibotory cytokines from these T cells. However, TGF-β-mediated induction of RORγt, a master regulator of Th17 cell, was independent of both Smad2 and Smad3, suggesting that TGF-β regulates Th17 development through Smad2/3-dependent and -independent mechanisms..