Biology subjects

Maeda, K.

Publications and source records attributed to Maeda, K..

At least 55 records · Page 3Linked to original sources

bioRxiv2022

Impact of reinfection with SARS-CoV-2 Omicron variants in previously infected hamsters

The diversity of SARS-CoV-2 mutations raises the possibility of reinfection of individuals previously infected with earlier variants, and this risk is further increased by the emergence of the B.1.1.529 Omicron variant. In this study, we used an in vivo, hamster infection model to assess the potential for individuals previously infected with SARS-CoV-2 to be reinfected with Omicron variant and we also investigated the pathology associated with such infections. Initially, Syrian hamsters were inoculated with a lineage A, B.1.1.7, B.1.351, B.1.617.2 or a subvariant of Omicron, BA.1 strain and then reinfected with the BA.1 strain 5 weeks later. Subsequently, the impact of reinfection with Omicron subvariants (BA.1 and BA.2) in individuals previously infected with the BA.1 strain was examined. Although viral infection and replication were suppressed in both the upper and lower airways, following reinfection, virus-associated RNA was detected in the airways of most hamsters. Viral replication was more strongly suppressed in the lower respiratory tract than in the upper respiratory tract. Consistent amino acid substitutions were observed in the upper respiratory tract of infected hamsters after primary infection with variant BA.1, whereas diverse mutations appeared in hamsters reinfected with the same variant. Histopathology showed no acute pneumonia or disease enhancement in any of the reinfection groups and, in addition, the expression of inflammatory cytokines and chemokines in the airways of reinfected animals was only mildly elevated. These findings are important for understanding the risk of reinfection with new variants of SARS-CoV-2. IMPORTANCEThe emergence of SARS-CoV-2 variants and the widespread use of COVID-19 vaccines has resulted in individual differences in immune status against SARS-CoV-2. A decay in immunity over time and the emergence of variants that partially evade the immune response can also lead to reinfection. In this study, we demonstrated that, in hamsters, immunity acquired following primary infection with previous SARS-CoV-2 variants was effective in preventing the onset of pneumonia after reinfection with the Omicron variant. However, viral infection and multiplication in the upper respiratory tract were still observed after reinfection. We also showed that more diverse nonsynonymous mutations appeared in the upper respiratory tract of reinfected hamsters that had acquired immunity from primary infection. This hamster model reveals the within-host evolution of SARS-CoV-2 and its pathology after reinfection, and provides important information for countermeasures against diversifying SARS-CoV-2 variants.

microbiology↗

bioRxiv2022

Characterization of SARS-CoV-2 Omicron BA.2.75 clinical isolates

The prevalence of the Omicron subvariant BA.2.75 is rapidly increasing in India and Nepal. In addition, BA.2.75 has been detected in at least 34 other countries and is spreading globally. However, the virological features of BA.2.75 are largely unknown. Here, we evaluated the replicative ability and pathogenicity of BA.2.75 clinical isolates in Syrian hamsters. Although we found no substantial differences in weight change among hamsters infected with BA.2, BA.5, or BA.2.75, the replicative ability of BA.2.75 in the lungs was higher than that of BA.2 and BA.5. Of note, BA.2.75 caused focal viral pneumonia in hamsters, characterized by patchy inflammation interspersed in alveolar regions, which was not observed in BA.5-infected hamsters. Moreover, in competition assays, BA.2.75 replicated better than BA.5 in the lungs of hamsters. These results suggest that BA.2.75 can cause more severe respiratory disease than BA.5 and BA.2 and should be closely monitored.

microbiology↗

bioRxiv2022

Circadian ribosome profiling reveals a role for the Period2 upstream opening reading frame in sleep

Many mammalian proteins have circadian cycles of production and degradation, and many of these rhythms are altered post-transcriptionally. We used ribosome profiling to examine post-transcriptional control of circadian rhythms by quantifying RNA translation in the liver over a 24-h period from circadian-entrained mice transferred to constant darkness conditions and by comparing ribosome binding levels to protein levels for 16 circadian proteins. We observed large differences in ribosome binding levels compared to protein levels, and we observed delays between peak ribosome binding and peak protein abundance. We found extensive binding of ribosomes to upstream open reading frames (uORFs) in circadian mRNAs, including the core clock gene Period2 (Per2). An increase in the number of uORFs in the 5UTR was associated with a decrease in ribosome binding in the main coding sequence and a reduction in expression of synthetic reporter constructs. Mutation of the Per2 uORF increased luciferase and fluorescence reporter expression in 3T3 cells and increased luciferase expression in PER2:LUC MEF cells. Mutation of the Per2 uORF in mice increased Per2 mRNA expression, enhanced ribosome binding on Per2, and reduced total sleep time compared to that in wild-type mice. These results suggest that uORFs affect mRNA post-transcriptionally, which can impact physiological rhythms and sleep. Significance StatementPeriod (Per) is an iconic gene in the field of circadian rhythms since its discovery in 1971 by Seymour Benzer and Ronald Konopka in fruit flies. The inhibitory feedback loop of PER protein drives circadian rhythms. We show that Per2 is regulated by an upstream open reading frame (uORF) in the 5 untranslated region of Period2 mRNA. Mutation of the Per2 uORF altered the amplitude of luciferase reporter expression in well-characterized cell culture models. Per2 uORF mutant mice had significantly elevated Per2 mRNA levels and exhibited sleep loss, particularly during light-to-dark and dark-to-light transitions, which suggests a role for uORFs in modulating molecular and physiological circadian rhythms.

cell biology↗

bioRxiv2022

Potential anti-monkeypox virus activity of atovaquone, mefloquine, and molnupiravir, and their potential use as treatments

Monkeypox virus (MPXV) is a zoonotic orthopoxvirus that causes smallpox-like symptoms in humans and caused an outbreak in May 2022 that led the WHO to declare global health emergency. In this study, from a screening of approved-drug libraries using an MPXV infection cell system, atovaquone, mefloquine, and molnupiravir exhibited anti-MPXV activity, with 50% inhibitory concentrations of 0.51-5.2 M, which is more potent than cidofovir. Whereas mefloquine was suggested to inhibit viral entry, atovaquone and molnupiravir targeted post-entry process to impair intracellular virion accumulation. Inhibitors of dihydroorotate dehydrogenase, an atovaquones target enzyme, showed conserved anti-MPXV activities. Combining atovaquone with tecovirimat enhanced the anti-MPXV effect of tecovirimat. Quantitative mathematical simulations predicted that atovaquone can promote viral clearance in patients by seven days at clinically relevant drug concentrations. Moreover, atovaquone and molnupiravir exhibited pan-Orthopoxvirus activity against vaccinia and cowpox viruses. These data suggest that atovaquone would be potential candidates for treating monkeypox.

microbiology↗

bioRxiv2022

SARS-CoV-2-neutralizing humoral IgA response occurs earlier but modest and diminishes faster compared to IgG response.

Secretory immunoglobulin A (IgA) plays a crucial role in the mucosal immunity for preventing the invasion of the exogenous antigens, however, little has been understood about the neutralizing activity of serum IgA. Here, to examine the role of IgA antibodies against COVID-19 illnesses, we determined the neutralizing activity of serum/plasma IgG and IgA purified from previously SARS-CoV-2-infected and COVID-19 mRNA-vaccine-receiving individuals. We found that serum/plasma IgA possesses substantial but rather modest neutralizing activity against SARS-CoV-2 compared to IgG with no significant correlation with the disease severity. Neutralizing IgA and IgG antibodies achieved the greatest activity at approximately 25 and 35 days after symptom onset, respectively. However, neutralizing IgA activity quickly diminished and went down below the detection limit approximately 70 days after onset, while substantial IgG activity was observed till 200 days after onset. The total neutralizing activity in sera/plasmas of those with COVID-19 largely correlated with that in purified-IgG and purified-IgA and levels of anti-SARS-CoV-2-S1-binding IgG and anti-SARS-CoV-2-S1-binding IgA. In individuals who were previously infected with SARS-CoV-2 but had no detectable neutralizing IgA activity, a single dose of BNT162b2 or mRNA-1273 elicited potent serum/plasma neutralizing IgA activity but the second dose did not further strengthen the neutralization antibody response. The present data show that the systemic immune stimulation with natural infection and COVID-19 mRNA-vaccines elicit both SARS-CoV-2-specific neutralizing IgG and IgA response in serum, but the IgA response is modest and diminishes faster compared to IgG response. Author SummaryImmunoglobulin A (IgA) is the most abundant type of antibody in the body mostly located on mucosal surfaces as a dimeric secretory IgA. Such secretory IgA plays an important role in preventing the adherence and invasions of foreign objects by its neutralizing activity, while monomeric serum IgA is thought to relate to the phagocytic immune system activation. Here, we report that individuals with the novel coronavirus disease (COVID-19) developed both systemic neutralizing IgG and IgA active against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Although the neutralizing IgA response was quick and reached the highest activity 25 days post-symptom-onset, compared to 35 days for IgG response, neutralizing IgA activity was modest and diminished faster than neutralizing IgG response. In individuals, who recovered from COVID-19 but had no detectable neutralizing IgA activity, a single dose of COVID-19 mRNA-vaccine elicited potent neutralizing IgA activity but the second dose did not further strengthen the antibody response. Our study provides novel insights into the role and the kinetics of serum IgA against the viral pathogen both in naturally-infected and COVID-19 mRNA-vaccine-receiving COVID-19-convalescent individuals.

microbiology↗

bioRxiv2022

Pathogenic variants of sphingomyelin synthase SMS2 disrupt lipid landscapes in the secretory pathway

Sphingomyelin is a dominant sphingolipid in mammalian cells. Its production in the trans-Golgi traps cholesterol synthesized in the ER to promote formation of a sphingomyelin/sterol gradient along the secretory pathway. This gradient marks a fundamental transition in physical membrane properties that help specify organelle identify and function. We previously identified mutations in sphingomyelin synthase SMS2 that cause osteoporosis and skeletal dysplasia. Here we show that SMS2 variants linked to the most severe bone phenotypes retain full enzymatic activity but fail to leave the ER owing to a defective autonomous ER export signal. Cells harboring pathogenic SMS2 variants accumulate sphingomyelin in the ER and display a disrupted transbilayer sphingomyelin asymmetry. These aberrant sphingomyelin distributions also occur in patient-derived fibroblasts and are accompanied by imbalances in cholesterol organization, glycerophospholipid profiles and lipid order in the secretory pathway. We postulate that pathogenic SMS2 variants undermine the capacity of osteogenic cells to uphold nonrandom lipid distributions that are critical for their bone forming activity.

cell biology↗

bioRxiv2022

Primate dopamine neurons activated in rewarding environment by tonic disinhibition from amygdala

Seeking out good and avoiding bad objects is critical for survival. In practice, objects are rarely good every time or everywhere, but only at the right time or place. Whereas the basal ganglia (BG) are known to mediate goal-directed behavior, for example, saccades to rewarding objects, it remains unclear how such simple behaviors are rendered contingent on higher-order factors, including environmental context. Here we show that amygdala neurons are sensitive to environments and regulate dopamine (DA) neurons via an inhibitory projection to substantia nigra pars reticulata (SNr) neurons. We combined optogenetics and multi-channel recording to demonstrate that rewarding environments induce tonic firing changes in DA neurons. These tonic responses are mediated by disinhibition via GABAergic projection onto DA neurons from SNr, which in turn are suppressed by inhibitory projection from the amygdala. Thus, the amygdala provides an additional source of learning to BG circuits, namely contingencies imposed by the environment.

neuroscience↗

bioRxiv2022

Different efficacies of neutralizing antibodies and antiviral drugs on SARS-CoV-2 Omicron subvariants, BA.1 and BA.2

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariant BA.2 has spread in many countries, replacing the earlier Omicron subvariant BA.1 and other variants. Here, using a cell culture infection assay, we quantified the intrinsic sensitivity of BA.2 and BA.1 compared with other variants of concern, Alpha, Gamma, and Delta, to five approved-neutralizing antibodies and antiviral drugs. Our assay revealed the diverse sensitivities of these variants to antibodies, including the loss of response of both BA.1 and BA.2 to casirivimab and of BA.1 to imdevimab. In contrast, EIDD-1931 and nirmatrelvir showed a more conserved activities to these variants. The viral response profile combined with mathematical analysis estimated differences in antiviral effects among variants in the clinical concentrations. These analyses provide essential evidence that gives insight into variant emergences impact on choosing optimal drug treatment.

microbiology↗

bioRxiv2022

Unraveling membrane properties at the organelle-level with LipidDyn

Cellular membranes are formed from many different lipids in various amounts and proportions depending on the subcellular localization. The lipid composition of membranes is sensitive to changes in the cellular environment, and their alterations are linked to several diseases, including cancer. Lipids not only form lipid-lipid interactions but also interact with other biomolecules, including proteins, profoundly impacting each other. Molecular dynamics (MD) simulations are a powerful tool to study the properties of cellular membranes and membrane-protein interactions on different timescales and at varying levels of resolution. Over the last few years, software and hardware for biomolecular simulations have been optimized to routinely run long simulations of large and complex biological systems. On the other hand, high-throughput techniques based on lipidomics provide accurate estimates of the composition of cellular membranes at the level of subcellular compartments. The community needs computational tools for lipidomics and simulation data effectively interacting to better understand how changes in lipid compositions impact membrane function and structure. Lipidomic data can be analyzed to design biologically relevant models of membranes for MD simulations. Similar applications easily result in a massive amount of simulation data where the bottleneck becomes the analysis of the data to understand how membrane properties and membrane-protein interactions are changing in the different conditions. In this context, we developed LipidDyn, an in silico pipeline to streamline the analyses of MD simulations of membranes of different compositions. Once the simulations are collected, LipidDyn provides average properties and time series for several membrane properties such as area per lipid, thickness, diffusion motions, the density of lipid bilayers, and lipid enrichment/depletion. The calculations exploit parallelization and the pipelines include graphical outputs in a publication-ready form. We applied LipidDyn to different case studies to illustrate its potential, including membranes from cellular compartments and transmembrane protein domains. LipidDyn is implemented in Python and relies on open-source libraries. LipidDyn is available free of charge under the GNU General Public License from https://github.com/ELELAB/LipidDyn.

bioinformatics↗

bioRxiv2021

Cellular toxicity of iHAP1 and DT-061 does not occur through PP2A-B56 targeting

PP2A is an abundant phosphoprotein phosphatase that acts as a tumor suppressor. For this reason, compounds able to activate PP2A are attractive anticancer agents. The small molecule compounds iHAP1 and DT-061 have recently been reported by Leonard et al. (2020) and Morita et al. (2020) in Cell to selectively stabilize specific PP2A-B56 complexes which mediate cell killing. Here, we show that this is not the case and question key findings in these papers. Through genome wide CRISPR-Cas9 screens, we uncover the biological pathways targeted by these compounds. We find that iHAP1 directly blocks microtubule assembly both in vitro and in vivo and thus acts as a microtubule poison. In contrast, DT-061 disrupts both the Golgi apparatus and the endoplasmic reticulum and we directly visualize DT-061 in cytoplasmic granules that co-localize with Golgi markers. Our work demonstrates that iHAP1 and DT-061 cannot be used for dissecting PP2A-B56 biology.

cell biology↗

bioRxiv2021

Highly-Neutralizing COVID-19-Convalescent-Plasmas Potently Block SARS-CoV-2 Replication and Pneumonia in Syrian Hamsters.

Despite various attempts to treat SARS-CoV-2-infected patients with COVID-19-convalescent plasmas, neither appropriate approach nor clinical utility has been established. We examined the efficacy of administration of highly-neutralizing COVID-19-convalescent plasma (hn-plasmas) and such plasma-derived IgG administration using the Syrian hamster COVID-19 model. Two hn-plasmas, which were in the best 1% of 340 neutralizing-activity-determined convalescent plasma samples, were intraperitoneally administered to SARS-CoV-2-infected hamsters, resulting in significant reduction of viral titers in lungs by up to 32-fold as compared to the viral titers in hamsters receiving control non-neutralizing plasma, while with two moderately neutralizing plasmas (mn-plasmas) administered, viral titer reduction was by up to 6-fold. IgG fractions purified from the two hn-plasmas also reduced viral titers in lungs than those from the two mn-plasmas. The severity of lung lesions seen in hamsters receiving hn-plasmas was minimal to moderate as assessed using micro-computerized tomography, which histological examination confirmed. Western blotting revealed that all four COVID-19-convalescent-plasmas variably contained antibodies against SARS-CoV-2 components including the receptor-binding domain and S1 domain. The present data strongly suggest that administering potent-neutralizing-activity-confirmed COVID-19-convalescent plasmas would be efficacious in treating patients with COVID-19. ImportanceConvalescent plasmas obtained from patients, who recovered from a specific infection, have been used as agents to treat other patients infected with the very pathogen. To treat using convalescent plasmas, despite that more than 10 randomized-controlled-clinical-trials have been conducted and more than 100 studies are currently ongoing, the effects of convalescent plasma against COVID-19 remained uncertain. On the other hand, certain COVID-19 vaccines have been shown to reduce the clinical COVID-19 onset by 94-95%, for which the elicited SARS-CoV-2-neutralizing antibodies are apparently directly responsible. Here, we demonstrate that highly-neutralizing-effect-confirmed convalescent plasmas significantly reduce the viral titers in the lung of SARS-CoV-2-infected Syrian hamsters and block the development of virally-induced lung lesions. The present data provide a proof-of-concept that the presence of highly-neutralizing antibody in COVID-19-convalescent plasmas is directly responsible for the reduction of viral replication and support the use of highly-neutralizing antibody-containing plasmas in COVID-19 therapy with convalescent plasmas.

microbiology↗

bioRxiv2021

Subacute SARS-CoV-2 replication can be controlled in the absence of CD8+ T cells in cynomolgus macaques

SARS-CoV-2 infection presents clinical manifestations ranging from asymptomatic to fatal respiratory failure. Despite the induction of functional SARS-CoV-2-specific CD8+ T-cell responses in convalescent individuals, the role of virus-specific CD8+ T-cell responses in the control of SARS-CoV-2 replication remains unknown. In the present study, we show that subacute SARS-CoV-2 replication can be controlled in the absence of CD8+ T cells in cynomolgus macaques. Eight macaques were intranasally inoculated with 105 or 106 TCID50 of SARS-CoV-2, and three of the eight macaques were treated with a monoclonal anti-CD8 antibody on days 5 and 7 post-infection. In these three macaques, CD8+ T cells were undetectable on day 7 and thereafter, while virus-specific CD8+ T-cell responses were induced in the remaining five untreated animals. Viral RNA was detected in nasopharyngeal swabs for 10-17 days post-infection in all macaques, and the kinetics of viral RNA levels in pharyngeal swabs and plasma neutralizing antibody titers were comparable between the anti-CD8 antibody treated and untreated animals. SARS-CoV-2 RNA was detected in the pharyngeal mucosa and/or retropharyngeal lymph node obtained at necropsy on day 21 in two of the untreated group but undetectable in all macaques treated with anti-CD8 antibody. CD8+ T-cell responses may contribute to viral control in SARS-CoV-2 infection, but our results indicate possible containment of subacute viral replication in the absence of CD8+ T cells, implying that CD8+ T-cell dysfunction may not solely lead to viral control failure. Author SummarySARS-CoV-2 infection presents a wide spectrum of clinical manifestations ranging from asymptomatic to fatal respiratory failure. The determinants for failure in viral control and/or fatal disease progression have not been elucidated fully. Both acquired immune effectors, antibodies and CD8+ T cells, are considered to contribute to viral control. However, it remains unknown whether a deficiency in either of these two arms is directly linked to failure in the control of SARS-CoV-2 replication. In the present study, to know the requirement of CD8+ T cells for viral control after the establishment of infection, we examined the effect of CD8+ cell depletion by monoclonal anti-CD8 antibody administration in the subacute phase on SARS-CoV-2 replication in cynomolgus macaques. Unexpectedly, our analysis revealed no significant impact of CD8+ cell depletion on viral replication, indicating that subacute SARS-CoV-2 replication can be controlled in the absence of CD8+ T cells. CD8+ T-cell responses may contribute to viral control in SARS-CoV-2 infection, but this study suggests that CD8+ T-cell dysfunction may not solely lead to viral control failure or fatal disease progression.

microbiology↗

bioRxiv2021

RCGAToolbox: A real-coded genetic algorithm software for parameter estimation of kinetic models

SummaryKinetic modeling is essential in understanding the dynamic behavior of biochemical networks, such as metabolic and signal transduction pathways. However, parameter estimation remains a major bottleneck in the development of kinetic models. We present RCGAToolbox, software for real-coded genetic algorithms (RCGAs), which accelerates the parameter estimation of kinetic models. RCGAToolbox provides two RCGAs: the unimodal normal distribution crossover with minimal generation gap (UNDX/MGG) and real-coded ensemble crossover star with just generation gap (REXstar/JGG), using the stochastic ranking method. The RCGAToolbox also provides user-friendly graphical user interfaces. Availability and implementationRCGAToolbox is available from https://github.com/kmaeda16/RCGAToolbox under GNU GPLv3, with application examples. The user guide is provided in the Supplementary Material. RCGAToolbox runs on MATLAB in Windows, Linux, and macOS. Contactkmaeda@bio.kyutech.ac.jp Supplementary informationSupplementary Material is available at Bioinformatics online.

systems biology↗

bioRxiv2021

Biosynthesis system of Synechan, a sulfated exopolysaccharide, in the model cyanobacterium Synechocystis sp. PCC 6803

Extracellular polysaccharides of bacteria contribute to biofilm formation, stress tolerance, and infectivity. Cyanobacteria, the oxygenic photoautotrophic bacteria, uniquely and widely have sulfated extracellular polysaccharides and they may utilize the polysaccharides for survival in nature. In addition, sulfated polysaccharides of cyanobacteria and other organisms have been focused as beneficial biomaterial. However, very little is known about their biosynthesis machinery and function in cyanobacteria. Here we found that the model cyanobacterium, Synechocystis sp. PCC 6803, formed bloom-like cell aggregates using sulfated extracellular polysaccharides (designated as synechan) and identified whole set of genes responsible for synechan biosynthesis and its transcriptional regulation, thereby suggesting a model for the synechan biosynthesis apparatus. Because similar genes are found in many cyanobacterial genomes with wide variation, our findings may lead elucidation of various sulfated polysaccharides, their functions, and their potential application in biotechnology.

microbiology↗

bioRxiv2020

Development of a reverse transcription polymerase chain reaction for the detection of severe fever with thrombocytopenia syndrome virus from suspected infected animals

BackgroundSevere fever with thrombocytopenia syndrome virus (SFTSV) causes severe hemorrhagic fever in humans and cats. Clinical symptoms of SFTS-infected cats resemble to those of SFTS patients and SFTS-contracted cats shows high levels of viral RNA loads in the serum and body fluids. Due to the risk of direct infection from SFTS-infected cats to human, it is important to diagnose SFTS-suspected animals. Methodology/Principle findingsFour primer sets were newly designed from consensus sequences constructed by 108 strains of SFTSV. A reverse transcription polymerase chain reaction (RT-PCR) with these four primer sets were successfully and specifically detected several clades of SFTSV. Their limits of detection are 1-10 copies/reaction. By this RT-PCR, 5 cat cases among 56 SFTS-suspected animal cases were diagnosed as SFTS. From these cats, IgM or IgG against SFTSV were detected by enzyme-linked immunosorbent assay (ELISA), but not neutralizing antibodies by plaque reduction neutralization titer (PRNT) test. This phenomenon is similar to those of fatal SFTS patients. Conclusion/SignificanceThis newly developed RT-PCR could detect SFTSV RNA of several clades from SFTS-suspected animals. In addition to ELISA and PRNT test, the useful laboratory diagnosis systems of SFTS-suspected animals has been made in this study. Author summaryThis study developed RT-PCR to detect SFTS animal cases. This assay could detect SFTSV RNA belonging to different clades. Cats diagnosed as SFTS had IgM or IgG, but not neutralizing antibodies. SFTS cat cases were distributed in the area where SFTS patients have been reported highly, indicating the establishment of the circulation of SFTSV in the environment. These diagnostic assays could be helpful tools to detect and not to miss SFTS animal cases.

microbiology↗

bioRxiv2020

Reconstruction of the urinary tract at the appropriate time reduces fibrosis of the metanephros in rats as judged by imaging

Chronic kidney disease leads to high morbidity rates among humans. It is a serious disease that requires curative treatments other than kidney transplantation. Recently, we successfully established the iPS-derived generated kidney, which might produce urine. The urine can be directed to the native bladder with a stepwise peristaltic ureter system, followed by anastomosis with the recipient ureter for reconstruction of the urinary tract. However, the growth of the regenerated kidney varies significantly, whereas the time window of the anastomosis is quite narrow. Therefore, this study was conducted to evaluate the growth of transplanted metanephros with bladder periodically and noninvasively using computed tomography and ultrasonography. Ultrasonographic findings showed high correlations with computed tomographic findings and clearly evaluated metanephros with bladder. We found that the degree of growth of the metanephros with bladder after the transplantation differed in each individual. However, most of them reached the appropriate period for urinary tract reconstruction within 3 weeks after transplantation. Optimizing the stepwise peristaltic ureter system anastomosis by ultrasonography reduced long-term tubular dilation of the metanephros, thereby decreasing fibrosis caused by transforming growth factor-{beta}. This may be significantly related to long-term maturation of fetal grafts. These results provide new insights into transplanting regenerated kidneys in higher animals. We are one step closer to the first human trial of kidney generation.

developmental biology↗

bioRxiv2020

Heritable endogenization of an RNA virus in a mammalian species

Viruses are considered one of the driving forces for genome rearrangements via infection and endogenization into the host genome (Kazazian, 2004; Maksakova et al., 2006). In 2010, Horie and colleagues demonstrated that a non-retroviral RNA virus, Borna disease virus (BDV), could integrate into the genome in cultured somatic cells (Horie et al., 2010). However, germline transmission of viral-derived sequences using animal models is yet to be experimentally demonstrated. In this study, we reported a case of heritable endogenization using the encephalomyocarditis virus (EMCV) and laboratory mice. The EMCV is a small non-enveloped single-strand RNA virus without its own reverse transcriptase activity (Carocci et al., 2012). Here, we demonstrated that the EMCV genomic RNA was reverse transcribed into DNA fragments in the murine testes. The DNA sequence originated from the RNA genome of EMCV was also detected in the liver and earlobes of the offspring generated from the EMCV-infected father. This suggests that the exogenous sequence derived from the EMCV is transmitted into the host germline and inherited across subsequent generations. This first experimental demonstration of viral endogenization proposes reconsideration about the impact of viruses as a driving force for genome modification.

genetics↗

bioRxiv2019

The WIPE assay for selection and elimination of HIV-1 provirus in vitro using latency-reversing agents

Persistence of HIV-1 latent reservoir cells during antiretroviral therapy is a major obstacle for curing HIV-1. Latency-reversing agents (LRAs) are under development to reactivate and eradicate latently infected cells; however, there are few useful models for evaluating LRA activity in vitro. Here, we established a long-term cell culture system harboring thousands of different HIV-1-infected cell clones with a wide distribution of HIV-1 provirus similar to that observed in vivo. A combination of an LRA and antiretroviral therapy (ART) significantly reduced viral rebound upon treatment interruption. Experimental investigation and mathematical modeling demonstrated that addition of LRA to ART induced latency-reversing effect and contributed to the eradication of replication competent HIV-1. The widely distributed intact provirus elimination (WIPE) assay will be useful for optimizing therapeutics against HIV-1 latency and investigating mechanistic insights into the clonal selection of heterogeneous HIV-1-infected cells.

microbiology↗