Nutrient Condition in the Microenvironment Determines Essential Metabolisms of CD8(+) T Cells for Enhanced IFN gamma Production by Metformin

Metformin (Met), a first-line drug for type 2 diabetes, lowers blood glucose levels by suppressing gluconeogenesis in the liver, presumably through the liver kinase B1-dependent activation of AMP-activated protein kinase (AMPK) after inhibiting respiratory chain complex I. Met is also implicated as a drug to be repurposed for cancers; its mechanism is believed identical to that of gluconeogenesis inhibition. However, AMPK activation requires high Met concentrations at more than 1 mM, which are unachievable in vivo. The immune-mediated antitumor response might be the case in a low dose Met. Thus, we proposed activating or expanding tumor-infiltrating CD8(+) T cells (CD8TILs) in a mouse model by orally administering Met in free drinking water. Here we showed that Met, at around 10 mu M and a physiologically relevant concentration, enhanced production of IFN gamma,TNF alpha and expression of CD25 of CD8(+) T cells upon TCR stimulation. Under a glucose-rich condition, glycolysis was exclusively involved in enhancing IFN gamma production. Under a low-glucose condition, fatty acid oxidation or autophagy-dependent glutaminolysis, or both, was also involved. Moreover, phosphoenolpyruvate carboxykinase 1 (PCK1), converting oxaloacetate to phosphoenolpyruvate, became essential. Importantly, the enhanced IFN gamma production was blocked by a mitochondrial ROS scavenger and not by an inhibitor of AMPK. In addition, IFN gamma production by CD8TILs relied on pyruvate translocation to the mitochondria and PCK1. Our results revealed a direct effect of Met on IFN gamma production of CD8(+) T cells that was dependent on differential metabolic pathways and determined by nutrient conditions in the microenvironment

高齢者における視機能と認知機能障害の関連 : 藤原京アイスタディより

Both visual impairment and cognitive impairment are essential factors that determine the quality of life in the aged population. The aim of this study was to determine if a correlation existed between visual acuity and cognitive impairment in an elderly Japanese population. The Fujiwara-kyo Eye Study was a cross-sectional study of individuals aged ≥68 years who lived in Nara Prefecture of Japan. Participants underwent ophthalmological examinations and cognitive function test. A mild visual impairment was defined as having a best corrected visual acuity (BCVA) >0.2 logarithm of the minimum angle of resolution (logMAR) units in the better eye. Cognitive impairment was defined as having a Mini-Mental State Examination (MMSE) score of ≤23 points. A total to 2818 individuals completed the examinations. The mean age of the participants was 76.3 ± 4.8 years (mean ± standard deviation). The mean BCVA of the better eye was -0.02 ± 0.13 logMAR units and 6.6% subjects were classified as being mildly visually impaired. The mean MMSE score was 27.3 ± 2.3 and 5.7% subjects were classified as being cognitively impaired. The proportion of subjects with cognitive or moderate visual impairment increased with age, and there was a significant correlation between the visual acuity and MMSE score (r = -0.10, p < 0.0001). Subjects with mild visual impairments had 2.4 times higher odds of having cognitive impairment than those without visual impairment (odds ratio 2.4, 95% confidence interval, 1.5-3.8, p < 0.001) after adjusting for age, sex, and length of education. We conclude that it may be important to maintain good visual acuity to reduce the risk of having cognitive impairment.博士（医学）・乙第1396号・平成29年3月15日© Masashi Mine et al. 2016; Published by Mary Ann Liebert, Inc. This Open Access article is distributed under the terms of the Creative Commons License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited

Perinatal Management for a Pregnant Woman with an MYH9

We diagnosed a primipara woman with an MYH9 disorder during her pregnancy. A peripheral blood smear with an immunofluorescence analysis is the established method of diagnosing MYH9 disorders. We provided genetic counseling, as required, which included apprising the woman of the inheritance pattern, the importance of a genetic analysis, and the potential delivery risks for the patient and her offspring. Given that the potential delivery risks are reportedly low, special perinatal management is not necessary for patients with an MYH9 disorder whose platelet count is above 5.0 × 104/μL, except for rapid blood access

Pachychoroid neovasculopathy と加齢黄斑変性症における血漿 von Willebrand 因子に焦点を当てた解析

Pachychoroid neovasculopathy (PNV) is a new concept of macular disorder. Some cases diagnosed as age-related macular degeneration (AMD) have been re-diagnosed as PNV. However, the biological features of PNV are still uncertain. The purpose of this study was to compare PNV and AMD by analyses focusing on von Willebrand factor (VWF) and complement factor H (CFH). Ninety-seven patients who were previously diagnosed with treatment naïve AMD were enrolled in this study. They were re-classified as either PNV or AMD based on the clinical criteria and 33 patients were classified as PNV and 64 patients as AMD. We examined the clinical data, analyzed VWF multimer and two genetic polymorphisms (I62V and Y402H) in the CFH. PNV group was significantly younger than AMD group (P = 0.001). In both I62V and Y402H, there were no significant differences between PNV and AMD while the recessive homozygous (AA) was found only in PNV group in I62V. The presence of unusually large VWF multimers (UL-VWFMs) and subretinal hemorrhages were significantly higher in PNV than in AMD (P = 0.045, P = 0.020, respectively). Thus, the residual UL-VWFMs may result in platelet thrombosis and hemorrhages in the choriocapillaris of PNV. In conclusion, our results suggest the biological differences between PNV and AMD.博士（医学）・甲第832号・令和4年3月15日© 2021. The Author(s). Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/

Intravitreal injection of aflibercept, an anti-VEGF antagonist, down-regulates plasma von Willebrand factor in patients with age-related macular degeneration

Abstract We investigated the association between von Willebrand factor (VWF) and exudative age-related macular degeneration (AMD) in 114 Japanese patients. Intravitreal injection of vascular endothelial growth factor (VEGF) inhibitor is the most effective therapy for AMD. Therefore, we analyzed changes of VWF antigen (VWF:Ag) and VWF multimers (VWFMs) after intravitreal injection of aflibercept, an anti-VEGF antagonist. The relationship between polymorphisms in complement factor H (p.Y402H and p.I62V) and AMD was previously reported. In our patients, p.I62V, but not p.Y402H, was significantly associated with an increased risk of AMD. Pre-treatment plasma levels of VWF:Ag in patients with AMD were significantly higher than those in controls. Unusually large VWFMs (UL-VWFMs) were detected in the majority of AMD patients with concurrent vitreous or subretinal hemorrhage. After intravitreal injection of aflibercept, plasma levels of VWF:Ag and VEGF-A were significantly decreased. UL-VWFMs disappeared after aflibercept injection in three cases, but persisted even 1 month after injection in the other five cases. In conclusion, plasma VWF:Ag levels were significantly elevated in patients with AMD, and decreased after intravitreal aflibercept injection. VWF may play an important role in the pathophysiology of AMD, and aflibercept might improve AMD by reducing plasma levels of VWF in addition to VEGF-A

Effects of concentration of amyloid β (Aβ) on viability of cultured retinal pigment epithelial cells

Abstract Background Amyloid beta (Aβ) is a constituent of drusen that is a common sign of age-related macular degeneration (AMD). The purpose of this study was to investigate the effect of Aβ on human retinal pigment epithelial (RPE) cells in culture. Methods Cells from a human RPE cell line (ARPE-19) were exposed to 0 to 25 μM of Aβ 1–40 for 48 h, and the number of living cells was determined by WST-8 cleavage. Replicative DNA synthesis was measured by the incorporation of 5′-bromo-2′-deoxyuridine. The cell death pathway was investigated by the WST-8 cleavage assay after the addition of caspase-9 inhibitor, an anti-apoptotic factor. Real-time qRT-PCR was performed using Aβ-exposed cellular RNA to determine the level of vascular endothelial growth factor (VEGF)-A and pigment epithelium derived factor (PEDF). To determine the effect of receptor-for-advanced glycation end products (RAGE), the siRNA for RAGE was inserted into ARPE-19 treated with Aβ, and the levels of expression of VEGF-A and PEDF were determined. Results The number of living ARPE-19 cells was increased by exposure to 5 μM Aβ but was decreased by exposure to 25 μM of Aβ. Replicative DNA synthesis by ARPE-19 cells exposed to 25 μM of Aβ was significantly decreased indicating that 25 μM of Aβ inhibited cell proliferation. Real-time RT-PCR showed that the level of the mRNA of PEDF was increased by exposure to 5 μM Aβ, and the levels of the mRNAs of PEDF and VEGF-A were also increased by exposure to 25 μM Aβ. The addition of an inhibitor of caspase-9 blocked the decrease the number of ARPE-19 cells exposed to 25 μM Aβ. Exposure to si-RAGE attenuated the increase of VEGF-A and PEDF mRNA expression in ARPE-19 exposed to Aβ. Conclusions Exposure of ARPE-19 cells to low concentrations of Aβ increases the level of PEDF which then inhibits the apoptosis of ARPE-19 cells leading to RPE cell proliferation. Exposure to high concentrations of Aβ induces RPE cell death and enhances the expression of the mRNA of VEGF-A in RPE cells. The Aβ-RAGE pathway may lead to the expression VEGF-A and PEDF in RPE cells. These results suggest that Aβ is strongly related to the pathogenesis of choroidal neovascularization

Intravitreal injection of aflibercept, an anti-VEGF antagonist, down-regulates plasma von Willebrand factor in patients with age-related macular degeneration.

We investigated the association between von Willebrand factor (VWF) and exudative age-related macular degeneration (AMD) in 114 Japanese patients. Intravitreal injection of vascular endothelial growth factor (VEGF) inhibitor is the most effective therapy for AMD. Therefore, we analyzed changes of VWF antigen (VWF:Ag) and VWF multimers (VWFMs) after intravitreal injection of aflibercept, an anti-VEGF antagonist. The relationship between polymorphisms in complement factor H (p.Y402H and p.I62V) and AMD was previously reported. In our patients, p.I62V, but not p.Y402H, was significantly associated with an increased risk of AMD. Pre-treatment plasma levels of VWF:Ag in patients with AMD were significantly higher than those in controls. Unusually large VWFMs (UL-VWFMs) were detected in the majority of AMD patients with concurrent vitreous or subretinal hemorrhage. After intravitreal injection of aflibercept, plasma levels of VWF:Ag and VEGF-A were significantly decreased. UL-VWFMs disappeared after aflibercept injection in three cases, but persisted even 1 month after injection in the other five cases. In conclusion, plasma VWF:Ag levels were significantly elevated in patients with AMD, and decreased after intravitreal aflibercept injection. VWF may play an important role in the pathophysiology of AMD, and aflibercept might improve AMD by reducing plasma levels of VWF in addition to VEGF-A.博士（医学）・甲第684号・平成30年6月27日© The Author(s) 2018 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.identifier:Scientific reports Vol.8 No.1 Article No.1491 (2018 Jan)identifier:20452322identifier:http://ginmu.naramed-u.ac.jp/dspace/handle/10564/3453identifier:Scientific reports, 8(1): Article No.149

Effects of concentration of amyloid β (Aβ) on viability of cultured retinal pigment epithelial cells.

BACKGROUND: Amyloid beta (Aβ) is a constituent of drusen that is a common sign of age-related macular degeneration (AMD). The purpose of this study was to investigate the effect of Aβ on human retinal pigment epithelial (RPE) cells in culture. METHODS: Cells from a human RPE cell line (ARPE-19) were exposed to 0 to 25 μM of Aβ 1-40 for 48 h, and the number of living cells was determined by WST-8 cleavage. Replicative DNA synthesis was measured by the incorporation of 5'-bromo-2'-deoxyuridine. The cell death pathway was investigated by the WST-8 cleavage assay after the addition of caspase-9 inhibitor, an anti-apoptotic factor. Real-time qRT-PCR was performed using Aβ-exposed cellular RNA to determine the level of vascular endothelial growth factor (VEGF)-A and pigment epithelium derived factor (PEDF). To determine the effect of receptor-for-advanced glycation end products (RAGE), the siRNA for RAGE was inserted into ARPE-19 treated with Aβ, and the levels of expression of VEGF-A and PEDF were determined. RESULTS: The number of living ARPE-19 cells was increased by exposure to 5 μM Aβ but was decreased by exposure to 25 μM of Aβ. Replicative DNA synthesis by ARPE-19 cells exposed to 25 μM of Aβ was significantly decreased indicating that 25 μM of Aβ inhibited cell proliferation. Real-time RT-PCR showed that the level of the mRNA of PEDF was increased by exposure to 5 μM Aβ, and the levels of the mRNAs of PEDF and VEGF-A were also increased by exposure to 25 μM Aβ. The addition of an inhibitor of caspase-9 blocked the decrease the number of ARPE-19 cells exposed to 25 μM Aβ. Exposure to si-RAGE attenuated the increase of VEGF-A and PEDF mRNA expression in ARPE-19 exposed to Aβ. CONCLUSIONS: Exposure of ARPE-19 cells to low concentrations of Aβ increases the level of PEDF which then inhibits the apoptosis of ARPE-19 cells leading to RPE cell proliferation. Exposure to high concentrations of Aβ induces RPE cell death and enhances the expression of the mRNA of VEGF-A in RPE cells. The Aβ-RAGE pathway may lead to the expression VEGF-A and PEDF in RPE cells. These results suggest that Aβ is strongly related to the pathogenesis of choroidal neovascularization.博士（医学）・甲第712号・令和元年6月26日© The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.© 2019 BioMed Central Ltd unless otherwise stated. Part of Springer Nature.identifier:BMC ophthalmology Vol.19 No.1 Article No.70 (2019 Mar)identifier:14712415identifier:http://ginmu.naramed-u.ac.jp/dspace/handle/10564/3584identifier:BMC ophthalmology, 19(1): Article No.7