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Abstract The overexpression of Polo-like kinase 1 (PLK1) is associated with poor clinical outcomes in various malignancies, making it an attractive target for anticancer therapies. Although recent studies suggest PLK1’s involvement in homologous recombination (HR), the impact of its overexpression on HR remains unclear. In this study, we investigated the effect of PLK1 overexpression on HR using bioinformatics and experimental approaches. Analyzing The Cancer Genome Atlas (TCGA) and Cancer Cell Line Encyclopedia (CCLE) datasets with the Homologous Recombination Deficiency (HRD) score, we found a positive correlation between PLK1 expression and HRD score, indicating that increased PLK1 expression suppresses HR. To validate these findings, we performed cell line-based experiments, demonstrating that PLK1 overexpression attenuates RAD51 focus formation and HR, as measured by ASHRA in T47D cells. Since HR-deficient cells are hypersensitive to PARP inhibitors, we further confirmed that PLK1 overexpression increases sensitivity to PARP inhibitors, both in CCLE dataset analysis and experiments using T47D cells. Additionally, we found that the effects of PLK1 overexpression on HR suppression and increased PARP inhibitor sensitivity were mitigated by either a PLK1 kinase inhibitor or the kinase-dead mutant [T210A]. This suggests that PLK1’s impact on HR and PARP inhibitor sensitivity is mediated through its kinase activity. Moreover, analysis of clinical ovarian cancer samples revealed that higher PLK1 expression correlates with increased sensitivity to PARP inhibitors. Our results suggest that PLK1 overexpression suppresses homologous recombination, leading to enhanced sensitivity to PARP inhibition, presenting a potential therapeutic strategy for targeting cancers with overexpression of PLK1.

Abstract Optimal conditions for wound healing require a smooth transition from the early stage of inflammation to proliferation, and during this time alternatively activated (M2) macrophages play a central role. Recently, heat-killed lactic acid bacteria (LAB), such as Lactobacillus plantarum (L. plantarum) have been reported as possible modulators affecting the immune responses in wound healing. However, how signaling molecules regulate this process after the administration of heat-killed LAB remains unclear. In this study, we examined the effect of heat-killed L. plantarum KB131 (KB131) administration on wound healing and the contribution of CARD9, which is an essential signaling adaptor molecule for NF-kB activation upon triggering through C-type lectin receptors, in the effects of this bacterium. We analyzed wound closure, histological findings, and inflammatory responses. We found that administration of KB131 accelerated wound closure, re-epithelialization, granulation area, CD31-positive vessels, and α-SMA-positive myofibroblast accumulated area, as well as the local infiltration of leukocytes. In particular, M2 macrophages were increased, in parallel with CCL5 synthesis. The acceleration of wound healing responses by KB131 was canceled in CARD9-knockout mice. These results indicate that the topical administration of KB131 accelerates wound healing, accompanying increased M2 macrophages, which suggests that CARD9 may be involved in these responses.

Abstract The identification of acquired resistance mutations has been essential in non-small-cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) active mutations. Rebiopsy plays a pivotal role in selecting the optimal treatment for patients who develop resistance to initial EGFR-tyrosine kinase inhibitors (EGFR-TKIs). This multicenter, observational study was conducted to investigate the details of rebiopsy in Japanese clinical practice. The primary endpoints were the implementation rate of rebiopsy and the concordance rate for T790M mutation detection between histological and cytological specimens using the cobas EGFR Mutation Test, version 2. One hundred ninety-four patients with EGFR-mutant NSCLC were enrolled, and 120 patients developed acquired resistance to EGFR-TKIs. The median age was 68 years (range 20–87), and 52.5% of the patients were women. Rebiopsy was performed in 109 patients, and the implementation rate of rebiopsy was 90.8%. The success rates of rebiopsy in the total, histology, cytology and liquid biopsy populations were 67.9%, 81.3%, 66.7% and 43.8%, respectively. The positive percent agreement and the negative percent agreement in the detection of the T790M mutation between the histological and cytological specimens were both 90.9%. Obtaining histological or cytological tissue samples at rebiopsy may contribute to improving the detection rate of the T790M mutation (trial registration number: UMIN000026019).

Abstract Cryptococcus deneoformans is an opportunistic fungal pathogen that infects the lungs via airborne transmission and frequently causes fatal meningoencephalitis. Claudins (Cldns), a family of proteins with 27 members found in mammals, form the tight junctions within epithelial cell sheets. Cldn-4 and 18 are highly expressed in airway tissues, yet the roles of these claudins in respiratory infections have not been clarified. In the present study, we analyzed the roles of Cldn-4 and lung-specific Cldn-18 (luCldn-18) in host defense against C. deneoformans infection. luCldn-18-deficient mice exhibited increased susceptibility to pulmonary infection, while Cldn-4-deficient mice had normal fungal clearance. In luCldn-18-deficient mice, production of cytokines including IFN-γ was significantly decreased compared to wild-type mice, although infiltration of inflammatory cells including CD4+ T cells into the alveolar space was significantly increased. In addition, luCldn-18 deficiency led to high K+ ion concentrations in bronchoalveolar lavage fluids and also to alveolus acidification. The fungal replication was significantly enhanced both in acidic culture conditions and in the alveolar spaces of luCldn-18-deficient mice, compared with physiological pH conditions and those of wild-type mice, respectively. These results suggest that luCldn-18 may affect the clinical course of cryptococcal infection indirectly through dysregulation of the alveolar space microenvironment.

Abstract Integration of multiple properties of an object is a fundamental function of the visual cortex in object recognition. For instance, surface patterns and contour shapes are thought to be crucial characteristics that jointly contribute to recognition. However, the mechanisms of integration and corresponding cortical representations have not been fully clarified. We investigated the integration of surfaces and shapes by examining the tilt after effects (TAEs) evoked by the symmetry of patterns and contours. As symmetry in both pattern and contour evokes TAEs, we can directly measure the interaction between the two. The measured TAEs exhibited mutual transfer between the symmetry of the pattern (SP) and that of the contour shape (SS), i.e., adaptation by SP (SS) evoked TAEs when tested by SS (SP), suggesting the existence of an integrated representation. Next, we examined the interaction between SP and SS when both were simultaneously presented in adaptation. Congruent adaptors wherein their symmetry axes aligned evoked compressive interaction, whereas incongruent adaptors wherein the axes of SP and SS tilted to the opposite directions evoked subtractive interaction. These results suggest the existence of a cortical representation that integrates the properties of the surface and shape with suppressive interactions, which can provide crucial insights into the formation of object representation as well as the integration of visual information in the cortex.

Abstract A subset of prostate cancer displays a poor clinical outcome. Therefore, identifying this poor prognostic subset within clinically aggressive groups (defined as a Gleason score (GS) ≧8) and developing effective treatments are essential if we are to improve prostate cancer survival. Here, we performed a bioinformatics analysis of a TCGA dataset (GS ≧8) to identify pathways upregulated in a prostate cancer cohort with short survival. When conducting bioinformatics analyses, the definition of factors such as “overexpression” and “shorter survival” is vital, as poor definition may lead to mis-estimations. To eliminate this possibility, we defined an expression cutoff value using an algorithm calculated by a Cox regression model, and the hazard ratio for each gene was set so as to identify genes whose expression levels were associated with shorter survival. Next, genes associated with shorter survival were entered into pathway analysis to identify pathways that were altered in a shorter survival cohort. We identified pathways involving upregulation of GRB2. Overexpression of GRB2 was linked to shorter survival in the TCGA dataset, a finding validated by histological examination of biopsy samples taken from the patients for diagnostic purposes. Thus, GRB2 is a novel biomarker that predicts shorter survival of patients with aggressive prostate cancer (GS ≧8).

Neurons in visual area V4 modulate their responses depending on the figure-ground (FG) organization in natural images containing a variety of shapes and textures. To clarify whether the responses depend on the extents of the figure and ground regions in and around the classical receptive fields (CRFs) of the neurons, we estimated the spatial extent of local figure and ground regions that evoked FG-dependent responses (RF-FGs) in natural images and their variants. Specifically, we applied the framework of spike triggered averaging (STA) to the combinations of neural responses and human-marked segmentation images (FG labels) that represent the extents of the figure and ground regions in the corresponding natural image stimuli. FG labels were weighted by the spike counts in response to the corresponding stimuli and averaged over. The bias due to the nonuniformity of FG labels was compensated by subtracting the ensemble average of FG labels from the weighted average. Approximately 50% of the neurons showed effective RF-FGs, and a large number exhibited structures that were similar to those observed in virtual neurons with ideal FG-dependent responses. The structures of the RF-FGs exhibited a subregion responsive to a preferred side (figure or ground) around the CRF center and a subregion responsive to a non-preferred side in the surroundings. The extents of the subregions responsive to figure were smaller than those responsive to ground in agreement with the Gestalt rule. We also estimated RF-FG by an adaptive filtering (AF) method, which does not require spherical symmetry (whiteness) in stimuli. RF-FGs estimated by AF and STA exhibited similar structures, supporting the veridicality of the proposed STA. To estimate the contribution of nonlinear processing in addition to linear processing, we estimated nonlinear RF-FGs based on the framework of spike triggered covariance (STC). The analyses of the models based on STA and STC did not show inconsiderable contribution of nonlinearity, suggesting spatial variance of FG regions. The results lead to an understanding of the neural responses that underlie the segregation of figures and the construction of surfaces in intermediate-level visual areas.

Neurons in visual area V4 modulate their responses depending on the figure-ground (FG) organization in natural images containing a variety of shapes and textures. To clarify whether the responses depend on the extents of the figure and ground regions in and around the classical receptive fields (CRFs) of the neurons, we estimated the spatial extent of local figure and ground regions that evoked FG-dependent responses (RF-FGs) in natural images and their variants. Specifically, we applied the framework of spike triggered averaging (STA) to the combinations of neural responses and human-marked segmentation images (FG labels) that represent the extents of the figure and ground regions in the corresponding natural image stimuli. FG labels were weighted by the spike counts in response to the corresponding stimuli and averaged over. The bias due to the nonuniformity of FG labels was compensated by subtracting the ensemble average of FG labels from the weighted average. Approximately 50% of the neurons showed effective RF-FGs, and a large number exhibited structures that were similar to those observed in virtual neurons with ideal FG-dependent responses. The structures of the RF-FGs exhibited a subregion responsive to a preferred side (figure or ground) around the CRF center and a subregion responsive to a non-preferred side in the surroundings. The extents of the subregions responsive to figure were smaller than those responsive to ground in agreement with the Gestalt rule. We also estimated RF-FG by an adaptive filtering (AF) method, which does not require spherical symmetry (whiteness) in stimuli. RF-FGs estimated by AF and STA exhibited similar structures, supporting the veridicality of the proposed STA. To estimate the contribution of nonlinear processing in addition to linear processing, we estimated nonlinear RF-FGs based on the framework of spike triggered covariance (STC). The analyses of the models based on STA and STC did not show inconsiderable contribution of nonlinearity, suggesting spatial variance of FG regions. The results lead to an understanding of the neural responses that underlie the segregation of figures and the construction of surfaces in intermediate-level visual areas.

The World Health Organization initiated a global surveillance system for respiratory syncytial virus (RSV) in 2015, and the pilot surveillance is ongoing. The real-time RT-PCR RSV assays (Pan-RSV and duplex assays) developed by the United States Centers for Disease Control and Prevention are applied as the standard assays. To introduce these as standard assays in Japan, their practicality was evaluated using 2261 specimens obtained from pediatric inpatients in Japan, which were collected from 2018 to 2021. Although the Pan-RSV and duplex assays had similar analytical sensitivities, they yielded 630 (27.9%) and 786 (34.8%) RSV-positive specimens, respectively (p < 0.001). Although sequencing analysis showed mismatches in the reverse primer used in the Pan-RSV assay, these mismatches did not affect its analytical sensitivity. The analysis of read numbers of RSV isolates from air–liquid interface culture of human bronchial/tracheal epithelial cells showed that the duplex assay had a greater number of reads than did the Pan-RSV assay. Therefore, the duplex assay has superior detection performance compared with the Pan-RSV assay, but the two assays have similar analytical sensitivities.

Segmentation of a natural scene into objects and background is a fundamental but challenging task for recognizing objects. Investigating intermediate-level visual cortical areas with a focus on local information is a crucial step towards understanding the formation of the cortical representations of figure and ground. We examined the activity of a population of macaque V4 neurons during the presentation of natural image patches and their respective variations. The natural image patches were optimized to exclude the influence of global context but included various characteristics of local stimulus. Around one fourth of the patch-responsive V4 neurons exhibited significant modulation of firing activity that was dependent on the positional relation between the figural region of the stimulus and the classical receptive field of the neuron. However, the individual neurons showed low consistency in figure-ground modulation across a variety of image patches (55-62%), indicating that individual neurons were capable of correctly signaling figure and ground only for a limited number of stimuli. We examined whether integration of the activity of multiple neurons enabled higher consistency across a variety of natural patches by training a support vector machine to classify figure and ground of the stimuli from the population firing activity. The integration of the activity of a few tens of neurons yielded discrimination accuracy much greater than that of single neurons (up to 85%), suggesting a crucial role of population coding for figure-ground discrimination in natural images.

Antibodies against influenza virus neuraminidase (NA) protein prevent releasing of the virus from host cells and spreading of infection foci and are considered the 'second line of defence' against influenza. Haemagglutinin inhibition antibody-low responders (HI-LRs) are present among influenza split vaccine recipients. The NA inhibition (NAI) antibody response in vaccinees is worth exploring, especially those in the HI-LRs population. We collected pre- and post-vaccination sera from 61 recipients of an inactivated, monovalent, split vaccine against A/H1N1pdm09 and acute and convalescent sera from 49 unvaccinated patients naturally infected with the A/H1N1pdm09 virus during the 2009 influenza pandemic. All samples were subjected to haemagglutinin inhibition (HI), NAI and neutralisation assays. Most paired sera from naturally infected patients exhibited marked elevation in the NAI activity, and seroconversion rates (SCR) among HI-LRs and HI-responders (HI-Rs) were 60% and 87%, respectively; however, those from vaccinees displayed low increase in the NAI activity, and the SCR among HI-LRs and HI-Rs were 0% and 12%, respectively. In both HI-LRs and HI-Rs, vaccination with the inactivated, monovalent, split vaccine failed to elicit the NAI activity efficiently in the sera of the naive population, compared with the natural infection. Hence, the improvement of influenza vaccines is warranted to elicit not only HI but also NAI antibodies.

Human metapneumovirus (HMPV) has been a notable etiological agent of acute respiratory infection in humans, but it was not discovered until 2001, because HMPV replicates only in a limited number of cell lines and the cytopathic effect (CPE) is often mild. To promote the study of HMPV, several groups have generated green fluorescent protein (GFP)-expressing recombinant HMPV strains (HMPVGFP). However, the growing evidence has complicated the understanding of cell line specificity of HMPV, because it seems to vary notably among HMPV strains. In addition, unique A2b clade HMPV strains with a 180-nucleotide duplication in the G gene (HMPV A2b180nt-dup strains) have recently been detected. In this study, we re-evaluated and compared the cell line specificity of clinical isolates of HMPV strains, including the novel HMPV A2b180nt-dup strains, and six recombinant HMPVGFP strains, including the newly generated recombinant HMPV A2b180nt-dup strain, MG0256-EGFP. Our data demonstrate that VeroE6 and LLC-MK2 cells generally showed the highest infectivity with any clinical isolates and recombinant HMPVGFP strains. Other human-derived cell lines (BEAS-2B, A549, HEK293, MNT-1, and HeLa cells) showed certain levels of infectivity with HMPV, but these were significantly lower than those of VeroE6 and LLC-MK2 cells. Also, the infectivity in these suboptimal cell lines varied greatly among HMPV strains. The variations were not directly related to HMPV genotypes, cell lines used for isolation and propagation, specific genome mutations, or nucleotide duplications in the G gene. Thus, these variations in suboptimal cell lines are likely intrinsic to particular HMPV strains.

The adoptive transfer of effector T cells combined with lymphodepletion has demonstrated promising antitumor effects in mice and humans, although the availability of tumor-specific T cells is limited. We and others have also demonstrated that the transfer of polyclonal naïve T cells induces tumor-specific effector T cells and enhances antitumor immunity after lymphodepletion. Because tumors have been demonstrated to induce immunosuppressive networks and regulate the function of T cells, obtaining a sufficient number of fully functional naïve T cells that are able to differentiate into tumor-specific effector T cells remains difficult. To establish culture methods to obtain a large number of polyclonal T cells that are capable of differentiating into tumor-specific effector T cells, naïve T cells were activated with anti-CD3 mAbs in vitro. These cells were stimulated with IL-2 and IL-7 for the CD8 subset or with IL-7 and IL-23 for the CD4 subset. Transfer of these hyperexpanded T cells after lymphodepletion showed significant antitumor efficacy, and tumor-specific effector T cells were primed from these expanded T cells in tumor-bearing hosts. Moreover, these ex vivo-expanded T cells maintained T cell receptor diversity and showed long-term persistence of memory against specific tumors. Further analyses revealed that combination therapy consisting of vaccination with dendritic cells that were co-cultured with irradiated whole tumor cells and the transfer of ex vivo-expanded T cells significantly enhanced antitumor immunity. These results indicate that the transfer of ex vivo-expanded polyclonal T cells can be combined with other immunotherapies and augment antitumor effects.

LSD1, a lysine-specific histone demethylase, is overexpressed in several types of cancers and linked to poor outcomes. In breast cancer, the significance of LSD1 overexpression is not clear. We have performed an in silico analysis to assess the relationship of LSD1 expression to clinical outcome. We demonstrate that LSD1 overexpression is a poor prognostic factor in breast cancer, especially in basal-like breast cancer, a subtype of breast cancer with aggressive clinical features. This link is also observed in samples of triple negative breast cancer. Interestingly, we note that overexpression of LSD1 correlates with down-regulation of BRCA1 in triple negative breast cancer. This phenomenon is also observed in in vitro models of basal-like breast cancer, and is associated with an increased sensitivity to PARP inhibitors. We propose therefore that high expression levels of the demethylase LSD1 is a potential prognostic factor of poor outcome in basal-like breast cancer, and that PARP inhibition may be a therapeutic strategy of interest in this poor prognostic subtype with overexpression of LSD1.

Type I interferons (IFNs) are secreted by many cell types upon stimulation via pattern recognition receptors and bind to IFN-α/β receptor (IFNAR), which is composed of IFNAR1 and IFNAR2. Although type I IFNs are well known as anti-viral cytokines, limited information is available on their role during fungal infection. In the present study, we addressed this issue by examining the effect of IFNAR1 defects on the host defense response to Cryptococcus neoformans. In IFNAR1KO mice, the number of live colonies was lower and the host immune response mediated not only by Th1 but also by Th2 and Th17-related cytokines was more accelerated in the infected lungs than in WT mice. In addition, mucin production by bronchoepithelial cells and expression of MUC5AC, a major core protein of mucin in the lungs, were significantly higher in IFNAR1KO mice than in WT mice. This increase in mucin and MUC5AC production was significantly inhibited by treatment with neutralizing anti-IL-4 mAb. In contrast, administration of recombinant IFN-αA/D significantly suppressed the production of IL-4, but not of IFN-γ and IL-17A, in the lungs of WT mice after cryptococcal infection. These results indicate that defects of IFNAR1 led to improved clearance of infection with C. neoformans and enhanced synthesis of IFN-γ and the IL-4-dependent production of mucin. They also suggest that type I IFNs may be involved in the negative regulation of early host defense to this infection.

Directing attention to the spatial location or the distinguishing feature of a visual object modulates neuronal responses in the visual cortex and the stimulus discriminability of subjects. However, the spatial and feature-based modes of attention differently influence visual processing by changing the tuning properties of neurons. Intriguingly, neurons' tuning curves are modulated similarly across different visual areas under both these modes of attention. Here, we explored the mechanism underlying the effects of these two modes of visual attention on the orientation selectivity of visual cortical neurons. To do this, we developed a layered microcircuit model. This model describes multiple orientation-specific microcircuits sharing their receptive fields and consisting of layers 2/3, 4, 5, and 6. These microcircuits represent a functional grouping of cortical neurons and mutually interact via lateral inhibition and excitatory connections between groups with similar selectivity. The individual microcircuits receive bottom-up visual stimuli and top-down attention in different layers. A crucial assumption of the model is that feature-based attention activates orientation-specific microcircuits for the relevant feature selectively, whereas spatial attention activates all microcircuits homogeneously, irrespective of their orientation selectivity. Consequently, our model simultaneously accounts for the multiplicative scaling of neuronal responses in spatial attention and the additive modulations of orientation tuning curves in feature-based attention, which have been observed widely in various visual cortical areas. Simulations of the model predict contrasting differences between excitatory and inhibitory neurons in the two modes of attentional modulations. Furthermore, the model replicates the modulation of the psychophysical discriminability of visual stimuli in the presence of external noise. Our layered model with a biologically suggested laminar structure describes the basic circuit mechanism underlying the attention-mode specific modulations of neuronal responses and visual perception.

Abstract Introduction Budd-Chiari syndrome is a very rare pathological entity that ultimately leads to liver failure. Several therapeutic modalities, including percutaneous transluminal angioplasty, have been attempted to save the life of patients with Budd-Chiari syndrome. Few reports have described a salvage living donor liver transplantation performed after percutaneous transluminal angioplasty in a patient with acute Budd-Chiari syndrome. Case presentation A 26-year-old Japanese man developed severe progressive manifestations, such as massive ascites and hematemesis due to rupture of esophageal varices. After making several investigations, we diagnosed the case as Budd-Chiari syndrome. We first performed percutaneous transluminal angioplasty to dilate a short-segment stenosis of his inferior vena cava. The first percutaneous transluminal angioplasty greatly improved the clinical manifestations. However, after a year, re-stenosis was detected, and a second percutaneous transluminal angioplasty failed to open the severe stricture of his inferior vena cava. Since our patient had manifestations of acute liver failure, we decided to perform salvage living donor liver transplantation from his brother. The transplantation was successfully performed and all clinical manifestations were remarkably alleviated. Conclusion In cases of recurrent Budd-Chiari syndrome, the blocked hepatic venous outflow is not always relieved, even with invasive therapies. We have to take into account the possibility of adopting alternative salvage therapies if the first therapeutic modalities fail. When invasive therapy such as percutaneous transluminal angioplasty fails, liver transplantation should be considered as an alternative option.