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Developmental toxicity testing is essential to identify substances that may harm embryonic development. This study aimed to establish a protocol for evaluating developmental toxicity using human induced pluripotent stem cells (iPSCs) by analyzing cellular activity and gene expression changes. Two ICH S5(R3) positive substances, valproic acid (VPA), which is a substance previously detected as positive by other test methods, and thalidomide (Thalido), were examined during early trichoderm differentiation without fetal bovine serum. RNA-seq analysis identified seven candidate genes, including TP63, associated with altered expression following exposure to VPA or Thalido. These genes were implicated in pathways related to tissue development, cell growth, and molecular interactions. While the assay effectively detected VPA and Thalido, its limitations include testing only soluble substances and focusing on early differentiation stages. Nevertheless, the protocol demonstrates potential for the classification and evaluation of emerging modality drugs based on physical properties such as solubility, polarity, and pH. Integration with AI analysis may enhance its capacity to uncover genetic variations and evaluate previously uncharacterized substances. This study provides a foundation for alternative developmental toxicity testing methods, with further refinements in the culture method expected to improve accuracy and applicability in regulatory toxicology. [ABSTRACT FROM AUTHOR]

Author(s): Tyson J. Moyer [sup.1] [sup.2] , Yu Kato [sup.2] [sup.3] , Wuhbet Abraham [sup.1] , Jason Y. H. Chang [sup.1] , Daniel W. Kulp [sup.2] [sup.4] [sup.5] , Nicki [...], Adjuvants are central to the efficacy of subunit vaccines. Aluminum hydroxide (alum) is the most commonly used vaccine adjuvant, yet its adjuvanticity is often weak and mechanisms of triggering antibody responses remain poorly understood. We demonstrate that site-specific modification of immunogens with short peptides composed of repeating phosphoserine (pSer) residues enhances binding to alum and prolongs immunogen bioavailability. The pSer-modified immunogens formulated in alum elicited greatly increased germinal center, antibody, neutralizing antibody, memory and long-lived plasma cell responses compared to conventional alum-adsorbed immunogens. Mechanistically, pSer-immunogen:alum complexes form nanoparticles that traffic to lymph nodes and trigger B cell activation through multivalent and oriented antigen display. Direct uptake of antigen-decorated alum particles by B cells upregulated antigen processing and presentation pathways, further enhancing B cell activation. These data provide insights into mechanisms of action of alum and introduce a readily translatable approach to significantly improve humoral immunity to subunit vaccines using a clinical adjuvant. Alum coupled to protein immunogens via site-specific phosphoserine-containing linkers enhances long-lived B cell responses and can selectively direct antibodies toward protective neutralizing epitopes.

Ionizing radiation (IR)-induced double-strand breaks (DSBs) are primarily repaired by non-homologous end joining or homologous recombination (HR) in human cells. DSB repair requires adenosine-5′-triphosphate (ATP) for protein kinase activities in the multiple steps of DSB repair, such as DNA ligation, chromatin remodeling, and DNA damage signaling via protein kinase and ATPase activities. To investigate whether low ATP culture conditions affect the recruitment of repair proteins at DSB sites, IR-induced foci were examined in the presence of ATP synthesis inhibitors. We found that p53 binding protein 1 foci formation was modestly reduced under low ATP conditions after IR, although phosphorylated histone H2AX and mediator of DNA damage checkpoint 1 foci formation were not impaired. Next, we examined the foci formation of breast cancer susceptibility gene I (BRCA1), replication protein A (RPA) and radiation 51 (RAD51), which are HR factors, in G2 phase cells following IR. Interestingly, BRCA1 and RPA foci in the G2 phase were significantly reduced under low ATP conditions compared to that under normal culture conditions. Notably, RAD51 foci were drastically impaired under low ATP conditions. These results suggest that HR does not effectively progress under low ATP conditions; in particular, ATP shortages impair downstream steps in HR, such as RAD51 loading. Taken together, these results suggest that the maintenance of cellular ATP levels is critical for DNA damage response and HR progression after IR. [ABSTRACT FROM AUTHOR]

Wearable flexible strain sensors that can sense human motion are expected to be applied in healthcare and metaverse applications. Examples of such sensors are resistive strain sensors, which are characterized by high gauge factors and simple device structures and can be fabricated using solution processes. The performance of resistive strain sensors is affected by changes in the resistance due to temperature. To improve temperature performance, researchers have studied strain sensors with a temperature coefficient of resistance (TCR) close to zero. However, solution-processed near-zero-TCR strain sensors with excellent flexibility have not yet been developed. In this study, using a sensor material consisting of a conductive polymer and Ag nanoparticles, a near-zero-TCR strain sensor is developed with a minimum bending radius of 0.14 mm at a total device thickness of 14 μm. The sensor material exhibits lower sensitivity to temperature and higher sensitivity to strain, with a TCR of 0.11% K−1 and gauge factor of 64 at a strain of 6.1%. Furthermore, it is sufficiently durable to operate even after 1000 cycles at a strain of 3.6%. When the device is attached to the inside of an arm and the arm is bent deeply in a temperature-changing environment, it operates without failure and senses arm bending without being affected by the environmental or body temperature. [ABSTRACT FROM AUTHOR]

Background The development of memory B cells after asymptomatic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is not well understood. Methods We compared spike antibody titers, pseudovirus neutralizing antibody titers, and memory B-cell responses among SARS-CoV-2 PCR-positive Marine recruits who either reported asymptomatic or symptomatic infection. Results Thirty-six asymptomatic participants exhibited similar spike IgG titers, spike IgA titers, and pseudovirus neutralization titers compared to 30 symptomatic participants. Pseudovirus neutralization and spike IgG titers showed significant positive correlations with frequency of memory B cells. Conclusions Among young adults, asymptomatic SARS-CoV-2 infection induced antibody and memory B-cell responses comparable to mild symptomatic infection. [ABSTRACT FROM AUTHOR]

Simple Summary: Human multicentric osteosarcoma (HMOS) is a variant of human osteosarcoma (HOS) in which multiple bone lesions occur simultaneously or asynchronously before lung metastasis. HOS is associated with a poor prognosis. Using the human antibody phage library, we obtained 95 types of antibody clones against HMOS cell lines and developed a fluorescence probe-based enzyme-linked immunosorbent assay (ELISA) that enables high-throughput screening of the reactivity of these antibodies. The lysate of the HMOS cell line was incubated with the selected antibody, and the antigen–antibody complex was recovered using magnetic beads. Liquid chromatography-mass spectrometry (LC/MS) analysis of the sample bands extracted from gel electrophoresis detected CAVIN1/PTRF specifically in HMOS cells. This novel high-throughput screening assay using antibody–drug conjugate targeting for these specific proteins is promising for clinical applications. Osteosarcoma is a malignant tumor that produces neoplastic bone or osteoid osteoma. In human multicentric osteosarcoma (HMOS), a unique variant of human osteosarcoma (HOS), multiple bone lesions occur simultaneously or asynchronously before lung metastasis. HMOS is associated with an extremely poor prognosis, and effective treatment options are lacking. Using the proteins in our previously generated HMOS cell lines as antigens, we generated antibodies using a human antibody phage library. We obtained antibody clones recognizing 95 independent antigens and developed a fluorescence probe-based enzyme-linked immunosorbent assay (ELISA) technique capable of evaluating the reactivity of these antibodies by fluorescence intensity, allowing simple, rapid, and high-throughput selection of antibody clones. These results were highly correlated with those using flow cytometry. Subsequently, the HMOS cell lysate was incubated with the antibody, the antigen–antibody complex was recovered with magnetic beads, and the protein bands from electrophoresis were analyzed using liquid chromatography-mass spectrometry (LC/MS). CAVIN1/polymerase I transcript release factor was specifically detected in the HMOS cells. In conclusion, we found via a novel high-throughput screening method that CAVIN1/PTRF is an HMOS-specific cell membrane biomarker and an antigen capable of producing human antibodies. In the future, antibody–drug conjugate targeting of these specific proteins may be promising for clinical applications. [ABSTRACT FROM AUTHOR]

Boosting adjuvant activity: Adjuvants can enhance immune responses against poorly immunogenic antigens, making them an important component of many vaccines despite the limited number currently approved for clinical use. Silva et al. developed a new adjuvant formed through self-assembly of saponin with the Toll-like receptor 4 agonist monophosphoryl lipid A (MPLA) into nanoparticle structures. Compared with a panel of tested adjuvants including other saponin-based formulations, saponin/MPLA nanoparticles (SMNP) induced superior antibody and germinal center responses after immunization with a poorly immunogenic HIV antigen in both mice and rhesus macaques. SMNP increased lymphatic flow in a mast cell–dependent manner and antigen delivery to draining lymph nodes, providing insight into the mechanisms by which saponin adjuvants act to enhance humoral immune responses. Saponins are potent and safe vaccine adjuvants, but their mechanisms of action remain incompletely understood. Here, we explored the properties of several saponin formulations, including immune-stimulatory complexes (ISCOMs) formed by the self-assembly of saponin and phospholipids in the absence or presence of the Toll-like receptor 4 agonist monophosphoryl lipid A (MPLA). We found that MPLA self-assembles with saponins to form particles physically resembling ISCOMs, which we termed saponin/MPLA nanoparticles (SMNP). Saponin-containing adjuvants exhibited distinctive mechanisms of action, altering lymph flow in a mast cell–dependent manner and promoting antigen entry into draining lymph nodes. SMNP was particularly effective, exhibiting even greater potency than the compositionally related adjuvant AS01B in mice, and primed robust germinal center B cell, TFH, and HIV tier 2 neutralizing antibodies in nonhuman primates. Together, these findings shed new light on mechanisms by which saponin adjuvants act to promote the immune response and suggest that SMNP may be a promising adjuvant in the setting of HIV, SARS-CoV-2, and other pathogens. [ABSTRACT FROM AUTHOR]

Adaptive immune responses to SARS-CoV-2 infection have been extensively characterized in blood; however, most functions of protective immunity must be accomplished in tissues. Here, we report from examination of SARS-CoV-2 seropositive organ donors (ages 10 to 74) that CD4+ T, CD8+ T, and B cell memory generated in response to infection is present in the bone marrow, spleen, lung, and multiple lymph nodes (LNs) for up to 6 months after infection. Lungs and lung-associated LNs were the most prevalent sites for SARS-CoV-2–specific memory T and B cells with significant correlations between circulating and tissue-resident memory T and B cells in all sites. We further identified SARS-CoV-2–specific germinal centers in the lung-associated LNs up to 6 months after infection. SARS-CoV-2–specific follicular helper T cells were also abundant in lung-associated LNs and lungs. Together, the results indicate local tissue coordination of cellular and humoral immune memory against SARS-CoV-2 for site-specific protection against future infectious challenges. [ABSTRACT FROM AUTHOR]

Myoglobin, which can cause acute kidney injury, has a relatively high molecular weight and is poorly cleared by diffusion. We compared and examined myoglobin clearance by changing the blood purification membrane and modality in patients with a myoglobin blood concentration ≥ 1000 ng/ml. We retrospectively analyzed three patient groups based on the following three types of continuous hemofiltration (CHF): AN69ST membrane, polymethylmethacrylate (PMMA) membrane, and high‐flow hemodiafiltration (HDF) with increased dialysate flow rate using the PMMA membrane. There was no significant difference in clearance in CHF between AN69ST and PMMA membranes. However, the high‐flow HDF group showed the highest myoglobin clearance (p = 0.003). In the PMMA membrane, changing the treatment modality to high‐flow HDF increased clearance above the theoretical value, possibly due to internal filtration. To remove myoglobin by kidney replacement therapy from patients with hypermyoglobinemia, a modality such as high‐flow HDF would be desirable. [ABSTRACT FROM AUTHOR]

The high mobility group box 1 protein (HMGB1) is recognized as a prototypical endogenous danger cytokine in sepsis. We previously reported that a polyacrylonitrile (AN69ST) membrane rapidly adsorbed HMGB1. Herein, an in vitro hemofiltration system was designed to assess the HMGB1 adsorption capacity, adsorption sites, and adsorption mechanism of the AN69ST membrane. HMGB1 was repeatedly added seven times during hemofiltration. A rapid decrease in circulating HMGB1 was observed after every addition with no sign of saturation. Presence of HMGB1 on the filter membrane was observed on both membrane surfaces and within the bulk layer using a high concentration of HMGB1 by immunoelectron microscopy. We hypothesized that the addition of heparin to the membrane surface or filtration rate would contribute to the adsorption mechanism. We could not measure the influence of heparin and filtration. Although the membrane was too large to saturate under the μg/mL HMGB1 conditions, our results show that the AN69ST membrane has a robust absorption capacity that could be used to treat sepsis. [ABSTRACT FROM AUTHOR]