Senescence-Associated Heterochromatin Foci Suppress γ-H2AX Focus Formation Induced by Radiation Exposure.

DNA damage is induced by both endogenous and exogenous factors. Repair of DNA double-strand break (DSB), a serious damage that threatens genome stability, decreases with senescence. However, the molecular mechanisms underlying the decline in DNA repair capacity during senescence remain unclear. We performed immunofluorescence staining for phosphorylated histone H2AX (γ-H2AX) in normal human fetal lung fibroblasts and human skin fibroblasts of different ages after chronic irradiation (total dose, 1 Gy; dose rate, 1 Gy/day) to investigate the effect of cellular senescence and organismal aging on DSB repair. Accumulation of DSBs was observed with cellular senescence and organismal aging, probably caused by delayed DSB repair. Importantly, the formation of γ-H2AX foci, an early event in DSB repair, is delayed with cellular senescence and organismal aging. These results suggest that the delay in γ-H2AX focus formation might delay the overall DSB repair. Interestingly, immediate γ-H2AX foci formation was suppressed in cells with senescence-associated heterochromatin foci (SAHF). To investigate the relationship between the γ-H2AX focus formation and SAHF, we used LiCl to relax the SAHFs, followed by irradiation. We demonstrated that LiCl rescued the delayed γ-H2AX foci formation associated with cellular senescence. This indicates that SAHF interferes with γ-H2AX focus formation and inhibits DSB repair in radiation-induced DSB. Our results suggest that therapeutic targeting of SAHFs have potential to resolve DSB repair dysfunction associated with cellular senescence.

Albumin Hydrogel-Coated Mesoporous Silica Nanoparticle as a Carrier of Cationic Porphyrin and Ratiometric Fluorescence pH Sensor.

Here, we report that a mesoporous silica nanoparticle (MSN) coated with a fluoresceine-labeled bovine serum albumin (F-BSA) hydrogel layer works as a temperature-responsive nanocarrier for tetrakis-N-methylpyridyl porphyrin (TMPyP) and as a fluorescence ratiometric pH probe. F-BSA hydrogel-coated MSN containing TMPyP (F-BSA/MSN/TMPyP) was synthesized by thermal gelation of denatured F-BSA on the external surface of MSN. The F-BSA hydrogel layer was composed of an inner hard corona layer and an outer soft layer and was stable under physiological conditions. F-BSA/MSN/TMPyP exhibited temperature-dependent exponential release of TMPyP. In this release profile, the MSN was found to be a suitable host for stable encapsulation of tetracationic TMPyP by electrostatic interactions, and the F-BSA hydrogel layer mediated the diffusion of TMPyP from the MSN pore interior into the solution phase. Increasing temperature promoted partitioning of TMPyP from the pore interior to the F-BSA hydrogel layer, from where it was spontaneously released into the bulk solution phase by cation exchange. F-BSA/MSN/TMPyP also gave a linear ratiometric fluorescence response (1.3 per pH unit) in the pH range from 6.1 to 8.9.

Prediction of late adverse events in pelvic cancer patients receiving definitive radiotherapy using radiation-induced gamma-H2AX foci assay.

Radiation can induce DNA double-stranded breaks, which are typically detected by the fluorescence of phosphorylated histone H2AX. In this study, we examined the usefulness of the dynamics of radiation-induced gamma-H2AX foci of peripheral blood lymphocytes (PBLs), as a marker of DNA repair ability, in predicting late adverse events from radiotherapy. A total of 46 patients with cervical, vaginal and anal canal cancers treated with radical radiotherapy between 2014 and 2019 were included in this analysis. Concurrent chemotherapy was administered in 36 cases (78.3%). Peripheral blood was obtained before treatment, and then irradiated ex vivo with 1 Gy X-ray. The ratio of radiation-induced gamma-H2AX foci in PBLs measured at 30 min and at 4 h was defined as the foci decay ratio (FDR). With a median follow-up of 54 months, 9 patients (19.6%) were observed to have late genitourinary or gastrointestinal (GU/GI) toxicity. The FDR ranged from 0.51 to 0.74 (median 0.59), with a significantly higher incidence of Grade 1 or higher late adverse events in the FDR ≥ 0.59 group. In multivariate analysis, FDR ≥ 0.59 and hypertension also emerged as significant factors associated with the development of late toxicities. Overall, our results suggest that measurement of radiation-induced gamma-H2AX foci in PBLs may predict the risk of late GU/GI toxicities from chemoradiotherapy, which can enable tailoring the radiation dose to minimize adverse effects.

Association of demographics, HCV co-infection, HIV-1 subtypes and genetic clustering with late HIV diagnosis: a retrospective analysis from the Japanese Drug Resistance HIV-1 Surveillance Network.

INTRODUCTION: Late diagnosis of the human immunodeficiency virus (HIV) is a major concern epidemiologically, socially and for national healthcare systems. Although the association of certain demographics with late HIV diagnosis has been reported in several studies, the association of other factors, including clinical and phylogenetic factors, remains unclear. In the present study, we conducted a nationwide analysis to explore the association of demographics, clinical factors, HIV-1 subtypes/circulating recombinant form (CRFs) and genetic clustering with late HIV diagnosis in Japan, where new infections mainly occur among young men who have sex with men (MSM) in urban areas. METHODS: Anonymized data on demographics, clinical factors and HIV genetic sequences from 39.8% of people newly diagnosed with HIV in Japan were collected by the Japanese Drug Resistance HIV-1 Surveillance Network from 2003 to 2019. Factors associated with late HIV diagnosis (defined as HIV diagnosis with a CD4 count <350 cells/µl) were identified using logistic regression. Clusters were identified by HIV-TRACE with a genetic distance threshold of 1.5%. RESULTS: Of the 9422 people newly diagnosed with HIV enrolled in the surveillance network between 2003 and 2019, 7752 individuals with available CD4 count at diagnosis were included. Late HIV diagnosis was observed in 5522 (71.2%) participants. The overall median CD4 count at diagnosis was 221 (IQR: 62-373) cells/µl. Variables independently associated with late HIV diagnosis included age (adjusted odds ratio [aOR] 2.21, 95% CI 1.88-2.59, ≥45 vs. ≤29 years), heterosexual transmission (aOR 1.34, 95% CI 1.11-1.62, vs. MSM), living outside of Tokyo (aOR 1.18, 95% CI 1.05-1.32), hepatitis C virus (HCV) co-infection (aOR 1.42, 95% CI 1.01-1.98) and not belonging to a cluster (aOR 1.30, 95% CI 1.12-1.51). CRF07_BC (aOR 0.34, 95% CI 0.18-0.65, vs. subtype B) was negatively associated with late HIV diagnosis. CONCLUSIONS: In addition to demographic factors, HCV co-infection, HIV-1 subtypes/CRFs and not belonging to a cluster were independently associated with late HIV diagnosis in Japan. These results imply the need for public health programmes aimed at the general population, including but not limited to key populations, to encourage HIV testing.

Identification of Novel Senescent Markers in Small Extracellular Vesicles.

Senescent cells exhibit several typical features, including the senescence-associated secretory phenotype (SASP), promoting the secretion of various inflammatory proteins and small extracellular vesicles (EVs). SASP factors cause chronic inflammation, leading to age-related diseases. Recently, therapeutic strategies targeting senescent cells, known as senolytics, have gained attention; however, noninvasive methods to detect senescent cells in living organisms have not been established. Therefore, the goal of this study was to identify novel senescent markers using small EVs (sEVs). sEVs were isolated from young and senescent fibroblasts using three different methods, including size-exclusion chromatography, affinity column for phosphatidylserine, and immunoprecipitation using antibodies against tetraspanin proteins, followed by mass spectrometry. Principal component analysis revealed that the protein composition of sEVs released from senescent cells was significantly different from that of young cells. Importantly, we identified ATP6V0D1 and RTN4 as novel markers that are frequently upregulated in sEVs from senescent and progeria cells derived from patients with Werner syndrome. Furthermore, these two proteins were significantly enriched in sEVs from the serum of aged mice. This study supports the potential use of senescent markers from sEVs to detect the presence of senescent cells in vivo.

A Case of a Malignant Lymphoma Patient Persistently Infected with SARS-CoV-2 for More than 6 Months.

Coronavirus disease 2019 (COVID-19) is an emerging infectious disease caused by severe acute respiratory syndrome 2 (SARS-CoV-2). There are many unknowns regarding the handling of long-term SARS-CoV-2 infections in immunocompromised patients. Here, we describe the lethal disease course in a SARS-CoV-2-infected patient during Bruton's tyrosine kinase inhibitor therapy. We performed whole-genome analysis using samples obtained during the course of the disease in a 63-year-old woman who was diagnosed with intraocular malignant lymphoma of the right eye in 2012. She had received treatment since the diagnosis. An autologous transplant was performed in 2020, but she experienced a worsening of the primary disease 26 days before she was diagnosed with a positive SARS-CoV-2 RT-PCR. Tirabrutinib was administered for the primary disease. A cluster of COVID-19 infections occurred in the hematological ward while the patient was hospitalized, and she became infected on day 0. During the course of the disease, she experienced repeated remission exacerbations of COVID-19 pneumonia and eventually died on day 204. SARS-CoV-2 whole-viral sequencing revealed that the patient shed the virus long-term. Viral infectivity studies confirmed infectious virus on day 189, suggesting that the patient might be still infectious. This case report describes the duration and viral genetic evaluation of a patient with malignant lymphoma who developed SARS-CoV-2 infection during Bruton's tyrosine kinase inhibitor therapy and in whom the infection persisted for over 6 months.

Detection of Respiratory Viruses in SARS-CoV-2-Negative Specimens from January to March 2020 in Fukuoka, Japan.

Coronavirus disease 2019, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was first confirmed in Japan on January 15, 2020. The Fukuoka Institute of Health and Environmental Sciences conducted testing using polymerase chain reaction (PCR) for SARS-CoV-2 from January 31 to March 4, 2020. Samples (n = 119) were collected from 81 patients suspected of having SARS-CoV-2 infection, presenting with fever, cough, fatigue, pneumonia, and other symptoms; all the samples tested during that period were negative. To identify the pathogens responsible for these symptoms, we conducted multiplex PCR. Respiratory viruses, human metapneumovirus (hMPV) was detected in 10 patients (12%), human rhinovirus (HRV) in 3 patients (4%), and influenza B virus in 1 patient (1%). In addition, the patients who had the viruses were significantly older than those who did not. Infections with hMPV and HRV have been associated with a risk of severe illness and death among older adults. Therefore, differentiating SARS-CoV-2 from other respiratory viruses, such as hMPV and HRV, is necessary to prevent and control the spread of infection, especially in older adults.

Beyond visualization of DNA double-strand breaks after radiation exposure.

PURPOSE: Radiation science and radiation biology are fields where milestones have been set by numerous woman researchers, as represented by Marie Curie. This shows that it is a research field that is like a model of research diversity in modern society. In this review, I will describe what kind of research activities I have conducted as a Japanese woman researcher in the field of radiation science research. In addition, as a Japanese woman radiobiologist, I will describe the sense of mission I felt after the Fukushima Nuclear Power Plant accident and the research issues we must challenge in the future. CONCLUSION: As a Japanese woman researcher, I have felt a bias in gender balance in the field of science in Japan. Also, after the Fukushima nuclear Power Plant accident, I sometimes felt that woman researchers would be more suitable when sharing research results and specialized knowledge with the general public. In recent years, the importance of STEAM (Science-Technology-Engineering-Art-Mathematics) education has been highlighted all over the world, and I believe that the field of radiation science falls exactly into the STEAM education category. STEAM education is for people of all gender. I hope that radiation science research will lead to various younger generations, and that the gender balance of Japanese scientific researchers will increase.

An Enriched Environment Alters DNA Repair and Inflammatory Responses After Radiation Exposure.

After the Fukushima Daiichi Nuclear Power Plant accident, there is growing concern about radiation-induced carcinogenesis. In addition, living in a long-term shelter or temporary housing due to disasters might cause unpleasant stress, which adversely affects physical and mental health. It's been experimentally demonstrated that "eustress", which is rich and comfortable, has beneficial effects for health using mouse models. In a previous study, mice raised in the enriched environment (EE) has shown effects such as suppression of tumor growth and enhancement of drug sensitivity during cancer treatment. However, it's not yet been evaluated whether EE affects radiation-induced carcinogenesis. Therefore, to evaluate whether EE suppresses a radiation-induced carcinogenesis after radiation exposure, in this study, we assessed the serum leptin levels, radiation-induced DNA damage response and inflammatory response using the mouse model. In brief, serum and tissues were collected and analyzed over time in irradiated mice after manipulating the raising environment during the juvenile or adult stage. To assess the radiation-induced DNA damage response, we performed immunostaining for phosphorylated H2AX which is a marker of DNA double-strand break. Focusing on the polarization of macrophages in the inflammatory reaction that has an important role in carcinogenesis, we performed analysis using tissue immunofluorescence staining and RT-qPCR. Our data confirmed that EE breeding before radiation exposure improved the responsiveness to radiation-induced DNA damage and basal immunity, further suppressing the chronic inflammatory response, and that might lead to a reduction of the risk of radiation-induced carcinogenesis.

Health effects triggered by tritium: how do we get public understanding based on scientifically supported evidence?

The Commission for 'Corresponding to Radiation Disaster of the Japanese Radiation Research Society' formulated a description of potential health effects triggered by tritium. This was in response to the issue of discharging water containing tritium filtered by the Advanced Liquid Processing System (ALPS), generated and stored in Fukushima Daiichi Nuclear Power Station after the accident. In this review article, the contents of the description, originally provided in Japanese, which gives clear and detailed explanation about potential health effects triggered by tritium based on reliable scientific evidence in an understandable way for the public, were summarized. Then, additional information about biochemical or environmental behavior of organically bound tritium (OBT) were summarized in order to help scientists who communicate with general public.

Repair Kinetics of DNA Double Strand Breaks Induced by Simulated Space Radiation.

Radiation is unavoidable in space. Energetic particles in space radiation are reported to induce cluster DNA damage that is difficult to repair. In this study, normal human fibroblasts were irradiated with components of space radiation such as proton, helium, or carbon ion beams. Immunostaining for γ-H2AX and 53BP1 was performed over time to evaluate the kinetics of DNA damage repair. Our data clearly show that the repair kinetics of DNA double strand breaks (DSBs) induced by carbon ion irradiation, which has a high linear energy transfer (LET), are significantly slower than those of proton and helium ion irradiation. Mixed irradiation with carbon ions, followed by helium ions, did not have an additive effect on the DSB repair kinetics. Interestingly, the mean γ-H2AX focus size was shown to increase with LET, suggesting that the delay in repair kinetics was due to damage that is more complex. Further, the 53BP1 focus size also increased in an LET-dependent manner. Repair of DSBs, characterized by large 53BP1 foci, was a slow process within the biphasic kinetics of DSB repair, suggesting non-homologous end joining with error-prone end resection. Our data suggest that the biological effects of space radiation may be significantly influenced by the dose as well as the type of radiation exposure.

DNA Damage Regulates Senescence-Associated Extracellular Vesicle Release via the Ceramide Pathway to Prevent Excessive Inflammatory Responses.

DNA damage, caused by various oncogenic stresses, can induce cell death or cellular senescence as an important tumor suppressor mechanism. Senescent cells display the features of a senescence-associated secretory phenotype (SASP), secreting inflammatory proteins into surrounding tissues, and contributing to various age-related pathologies. In addition to this inflammatory protein secretion, the release of extracellular vesicles (EVs) is also upregulated in senescent cells. However, the molecular mechanism underlying this phenomenon remains unclear. Here, we show that DNA damage activates the ceramide synthetic pathway, via the downregulation of sphingomyelin synthase 2 (SMS2) and the upregulation of neutral sphingomyelinase 2 (nSMase2), leading to an increase in senescence-associated EV (SA-EV) biogenesis. The EV biogenesis pathway, together with the autophagy-mediated degradation pathway, functions to block apoptosis by removing cytoplasmic DNA fragments derived from chromosomal DNA or bacterial infections. Our data suggest that this SA-EV pathway may play a prominent role in cellular homeostasis, particularly in senescent cells. In summary, DNA damage provokes SA-EV release by activating the ceramide pathway to protect cells from excessive inflammatory responses.

Contrasting Results from Two Commercial Kits Testing for the Presence of Clostridium perfringens Enterotoxin in Feces from Norovirus-Infected Human Patients.

BACKGROUND: Detection of Clostridium perfringens enterotoxin (CPE) is critical for disease surveillance; however, commercial testing kits produce contrasting results. METHODS: We examined the cause of the differing results from a reversed passive latex agglutination (RPLA) assay (PET-RPLA Toxin Detection Kit) and an enzyme-linked immunosorbent assay (C. perfringens Enterotoxin ELISA Kit) using 73 human norovirus-positive fecal samples from gastroenteritis patients across 22 episodes in Japan. RESULTS: CPE was detected in 39/73 samples using the RPLA method; however, ELISA-based examination of 10 RPLA-positive samples produced negative results. Moreover, cpe was not detected in any of the RPLA-positive (n = 32) or -negative (n = 5) samples, and C. perfringens was only isolated from one RPLA-positive sample. CONCLUSIONS: An ELISA-based testing approach may be more reliable than RPLA assays for CPE detection from human fecal samples. These findings may also be applicable to the detection of other foodborne diseases.

Space Radiation Biology for "Living in Space".

Space travel has advanced significantly over the last six decades with astronauts spending up to 6 months at the International Space Station. Nonetheless, the living environment while in outer space is extremely challenging to astronauts. In particular, exposure to space radiation represents a serious potential long-term threat to the health of astronauts because the amount of radiation exposure accumulates during their time in space. Therefore, health risks associated with exposure to space radiation are an important topic in space travel, and characterizing space radiation in detail is essential for improving the safety of space missions. In the first part of this review, we provide an overview of the space radiation environment and briefly present current and future endeavors that monitor different space radiation environments. We then present research evaluating adverse biological effects caused by exposure to various space radiation environments and how these can be reduced. We especially consider the deleterious effects on cellular DNA and how cells activate DNA repair mechanisms. The latest technologies being developed, e.g., a fluorescent ubiquitination-based cell cycle indicator, to measure real-time cell cycle progression and DNA damage caused by exposure to ultraviolet radiation are presented. Progress in examining the combined effects of microgravity and radiation to animals and plants are summarized, and our current understanding of the relationship between psychological stress and radiation is presented. Finally, we provide details about protective agents and the study of organisms that are highly resistant to radiation and how their biological mechanisms may aid developing novel technologies that alleviate biological damage caused by radiation. Future research that furthers our understanding of the effects of space radiation on human health will facilitate risk-mitigating strategies to enable long-term space and planetary exploration.

Management of Antithrombotic Agents During Surgery or Other Kinds of Medical Procedures With Bleeding: The MARK Study.

Background Optimal management of antithrombotic agents during surgery has yet to be established. We performed a prospective multicenter observational study to determine the current status of the management of antithrombotic agents during surgery or other medical procedures with bleeding (MARK [Management of Antithrombotic Agents During Surgery or Other Kinds of Medical Procedures With Bleeding] study) in Japan. Methods and Results The participants were 9700 patients who received oral antithrombotic agents and underwent scheduled medical procedures with bleeding at 59 National Hospital Organization institutions in Japan. Primary outcomes were thromboembolic events, bleeding events, and death within 2 weeks before and 4 weeks after the procedures. We investigated the relationships between each outcome and patient demographics, comorbidities, type of procedure, and management of antithrombotic therapy. With respect to the periprocedural management of antithrombotic agents, 3551 patients continued oral antithrombotic agents (36.6%, continuation group) and 6149 patients discontinued them (63.4%, discontinuation group). The incidence of any thromboembolic event (1.7% versus 0.6%, P<0.001), major bleeding (7.6% versus 0.4%, P<0.001), and death (0.8% versus 0.4%, P<0.001) was all greater in the discontinuation group than the continuation group. In multivariate analysis, even after adjusting for confounding factors, discontinuation of anticoagulant agents was significantly associated with higher risk for both thromboembolic events (odds ratio: 4.55; 95% CI, 1.67-12.4; P=0.003) and major bleeding (odds ratio: 11.1; 95% CI, 2.03-60.3; P=0.006) in procedures with low bleeding risk. In contrast, heparin bridging therapy was significantly associated with higher risk for both thromboembolic events (odds ratio: 2.03; 95% CI, 1.28-3.22; P=0.003) and major bleeding (odds ratio: 1.36; 95% CI, 1.10-1.68; P=0.005) in procedures with high bleeding risk. Conclusions Discontinuation of oral antithrombotic agents and addition of low-dose heparin bridging therapy appear to be significantly associated with adverse events in the periprocedural period.

Detection of Borrelia DNA in Tick Species Collected from Vegetation and Wild Animals in Fukuoka, Japan.

We screened for the presence of Borrelia spp. in ticks collected from vegetation by flagging and from wild animals between May 2017 and November 2018 in Fukuoka, located in the northern Kyushu region of Japan. A total of 1,601 ticks were collected and separated based on morphology into nine species, namely Ixodes turdus, I. ovatus, Amblyomma testudinarium, Haemaphysalis flava, H. formosensis, H. kitaokai, H. longicornis, H. hystricis, and H. megaspinosa. The ticks were segregated into 561 pools and nested PCR was used to detect borrelial DNA. Borrelia turdi and Borrelia sp. HM were identified in two of the 561 pools. This is the first report of the presence of the Lyme disease group of Borrelia and of the relapsing fever group of Borrelia in Fukuoka, Japan.

Estimating HIV-1 incidence in Japan from the proportion of recent infections.

The first step of the UNAIDS/WHO 90-90-90 targets to encourage early diagnosis with treatment for the control of HIV-1 epidemic is to achieve 90% HIV-1 diagnosis in infected individuals. In Japan, approximately 30% of newly reported cases have been annually identified by AIDS onset, implying that substantial numbers of HIV-1-infected individuals remain undiagnosed. However, the proportion of undiagnosed cases has not yet been determined. In the present study, the proportion of recent HIV-1 infections to newly-diagnosed cases was determined from 2006 to 2015 using a recent infection assay for three metropolitan areas in Japan: Tokyo, Osaka, and Fukuoka. Estimated median periods between infection and diagnosis were 1.0, 1.8, and 2.9 years for Tokyo, Osaka, and Fukuoka, respectively. Estimation of annual HIV-1 incidence by a back-calculation method using these data as well as HIV/AIDS national surveillance data indicated proportions of undiagnosed to new HIV-1 infections from 2006 to 2015 to be 18% in Tokyo, 22% in Osaka, 38% in Fukuoka, and 28% in Japan. This is the first report estimating HIV-1 incidence in Japan using a serological biomarker in combination with national HIV/AIDS surveillance data.

Pilot clinical study of ascorbic acid treatment in cardiac catheterization.

Clinical radiodiagnosis and radiotherapy sometimes induce tissue damage and/or increase the risk of cancer in patients. However, in radiodiagnosis, a reduction in the exposure dose causes a blockier image that is not acceptable for diagnosis. Approximately 70% of DNA damage is induced via reactive oxygen species and/or radicals created during X-ray irradiation. Therefore, treatment with anti-oxidants and/or radical scavengers is considered to be effective in achieving a good balance between image quality and damage. However, few studies have examined the effect of using radical scavengers to reduce radiation damage in the clinical setting. In this study, we administrated 20 mg/kg ascorbic acid (AA) to patients before cardiac catheterization (CC) for diagnostic purposes. We analyzed changes in the number of phosphorylated H2AX (γH2AX) foci (a marker of DNA double-strand breaks) in lymphocytes, red blood cell glutathione levels, blood cell counts, and biochemical parameters. Unfortunately, we did not find satisfactory evidence to show that AA treatment reduces γH2AX foci formation immediately after CC. AA treatment did, however, cause a higher reduced/oxidized glutathione ratio than in the control arm immediately after CC. This is a preliminary study, but this result suggests that reducing radiation damage in clinical practice can be achieved using a biological approach.