Biology subjects

Sakai, J.

Publications and source records attributed to Sakai, J..

8 recordsLinked to original sources

bioRxiv2026

The live attenuated DGAT1-knockout whole-cell Toxoplasma vaccine confers protective immunity against acute and chronic toxoplasmosis

The intravacuolar parasite Toxoplasma gondii scavenges fatty acids from host mammalian cells and stores excess in lipid droplets. To investigate the physiological relevance of neutral lipid storage in Toxoplasma, we generated a mutant lacking DGAT1, an ER-localized enzyme that synthesizes triacylglycerols, from the virulent type I RH strain of T. gondii. Compared to WT, RH {Delta}DGAT1 parasites grow poorly in mammalian cells, form few LD, suffer from lipotoxicity, and do not cause disease or lethality in immunocompetent or immunodeficient mice. Importantly, mice immunized with RH {Delta}DGAT1 parasites mount strong, long-term immune responses involving both cellular and humoral components, with higher levels of T. gondii-specific IgG antibodies, effector memory T cells, and both pro-inflammatory and anti-inflammatory cytokines, indicating a mixed Th1/Th2 response with Th1 predominance. This immunity provides complete, long-lasting protection (up to 6 months) against rechallenge from homologous type I (acute infection) and heterologous cyst-forming type II (chronic infection) T. gondii strains. Additional analyses reveal that IFN-{gamma}, CD8+ T cells, as well as B cells are crucial for defending against type I T. gondii in immunized mice. Overall, our live-attenuated RH {Delta}DGAT1 strain is a promising vaccine candidate and a model for studying immune responses that control T. gondii infections.

microbiology↗

bioRxiv2026

BAT protects against hepatic oxidative stress by remodeling the circulating metabolome

Brown adipose tissue (BAT) regulates systemic metabolism beyond thermogenesis, yet the circulating mediators through which BAT communicates with other organs remain less explored. Here, we performed comprehensive serum metabolomics and lipidomics in BAT-ablated mice and human cohorts with varying BAT activity to delineate how BAT activity shapes the circulating metabolome. By integrating datasets across serum, tissues, extracellular fluids, and conditioned media, we assembled BAT-linked circulating molecular signatures. The analyses support a critical role for BAT in the clearance of circulating branched-chain amino acids and triglycerides. We also identified a cold-inducible metabolite, 3-hydroxystearic acid (3-OHSA), produced primarily by BAT and released into circulation. 3-OHSA serves as a circulating readout of cold-activated BAT and acts on the liver to reduce mitochondrial membrane potential and reactive oxygen species production, thereby limiting oxidative stress. This work provides a framework for identifying BAT-derived mediators and uncovers a BAT-liver axis that coordinates adaptation to metabolic stress.

physiology↗

bioRxiv2024

Large-scale generation of uniform sub-100 μm adipocyte spheroids in hydrogel microcapsules using a flow-focusing microfluidic device

Adipocyte spheroids are a promising three-dimensional (3D) cell culture model for obesity research because they reproduce 3D adipose tissue structures and cell-cell interactions better than 2D cultures. However, current methods fail to produce uniformly sized, small adipocyte spheroids at large scales, significantly limiting their use in analysis such as large-scale drug screening. Here, we develop a scalable method that combines simple microfluidics with templated emulsification to generate small, uniformly sized adipocyte spheroids. By encapsulating preadipocytes in numerous hollow agarose microcapsules and incubating them for two days, we reproducibly produced more than 100,000 uniform spheroids with diameters of approximately 50 {micro}m (CV: <13%); we then differentiated preadipocyte spheroids into adipocyte spheroids after an 8-day induction period. Our platform enhances large-scale 3D analysis using adipocyte spheroids for obesity research and can be adapted to generate various spheroid and organoid models, advancing biomedical research across diverse fields.

bioengineering↗

bioRxiv2023

Executive function and underlying brain network distinctions for callous-unemotional traits and conduct problems in adolescents

The complexity of executive function (EF) impairments in youth antisocial phenotypes of callous-unemotional (CU) traits and conduct problems (CP) challenge identifying phenotypic specific EF deficits. We can redress these challenges by (1) accounting for EF measurement error and (2) testing distinct functional brain properties accounting for differences in EF. Thus, we employed a latent modeling approach for EFs (inhibition, shifting, fluency, common EF) and extracted connection density from matching contemporary EF brain models with a sample of 112 adolescents (ages 13-17, 42% female). Path analysis indicated CU traits associated with lower inhibition. Inhibition network density positively associated with inhibition, but this association was strengthened by CU and attenuated by CP. Common EF associated with three-way interactions between density*CP by CU for the inhibition and shifting networks. This suggests those higher in CU require their brain to work harder for lower inhibition, whereas those higher in CP have difficulty engaging inhibitory brain responses. Additionally, those with CP interacting with CU show distinct brain patterns for a more general EF capacity. Importantly, modeling cross-network connection density in contemporary EF models to test EF involvement in core impairments in CU and CP may accelerate our understanding of EF in these phenotypes.

neuroscience↗

bioRxiv2022

IL-6 suppresses vaccine responses in neonatal mice by enhancing IL-2 activity on T follicular helper cells

The inability of neonates to develop CD4+CXCR5+PD-1+ T follicular helper (TFH) cells contributes to their weak vaccine responses. In adult mice, IL-6 promotes TFH-cell expansion by suppressing the expression of IL-2R{beta} on TFH cells. Here, we found a totally opposite role for IL-6 in neonatal mice TFH response. Whereas co-injection of neonatal mice with IL-6 and a conjugate polysaccharide vaccine suppressed TFH response by increasing the production of IL-2 and expression of IL-2R and IL-2R{beta} on TFH cells, immunization of IL-6 knock-out neonatal mice led to improved antibody responses accompanied by expanded TFH cells as well as lower levels of IL-2 and IL-2 receptors on TFH cells. Moreover, CpG containing vaccine improved TFH response in neonates while suppressing the expression of IL-2 receptors on TFH cells, suggesting that CpG protects TFH cells by inhibiting IL-2 activity. These findings unveil age specific differences in IL-6 mediated vaccine responses and highlight the need to consider age related immunobiological attributes in designing vaccines.

immunology↗

bioRxiv2022

B cell receptor induced IL-10 production from neonatal CD19+CD43- cells depends on STAT5 mediated IL-6 secretion

Newborns are unable to reach the adult-level humoral immune response partly due to the potent immunoregulatory role of IL-10. Increased IL-10 production by neonatal B cells has been attributed to the larger population of IL-10-producting CD43+ B-1 cells in neonates. Here, we show that neonatal CD43- non-B-1 cells also produce substantial amounts of IL-10 following B cell antigen receptor (BCR) activation. In neonatal CD43- non-B-1 cells, BCR engagement activated STAT5 under the control of phosphorylated forms of signaling molecules Syk, Btk, PKC, FAK and Rac1. STAT5 activation led to IL-6 production, which in turn was responsible for IL-10 production in an autocrine/paracrine fashion through the activation of STAT3. In addition to the increased IL-6 production in response to BCR stimulation, elevated expression of IL-6R expression in neonatal B cells rendered them highly susceptible to IL-6 mediated STAT3 phosphorylation and IL-10 production. Finally, IL-10 secreted from neonatal CD43- non-B-1 cells was sufficient to inhibit TNF- secretion by macrophages. Our results unveil a distinct mechanism of IL-6-dependent IL-10 production in BCR-stimulated neonatal CD19+CD43- B cells.

immunology↗

bioRxiv2021

Glutamine deficiency in solid tumors confers resistance to ribosomal RNA synthesis inhibitors

Ribosome biogenesis involves the processing of precursor ribosomal RNAs (pre-rRNAs) and sequential assembly with ribosomal proteins. Here we report that nutrient deprivation severely impairs pre-rRNA processing and leads to the accumulation of unprocessed rRNAs. Upon nutrient restoration, the accumulated pre-rRNAs are processed into mature rRNAs that are utilized for ribosome biogenesis. Failure to accumulate pre-rRNAs under nutrient deprivation leads to perturbed ribosome assembly during nutrient restoration and subsequent apoptosis via uL5/uL18-mediated activation of p53. Restoration of glutamine alone activates p53 by triggering uL5/uL18 translation. Induction of uL5/uL18 protein synthesis by glutamine was dependent on the translation factor eukaryotic elongation factor 2 (eEF2), which was in turn dependent on Raf/MEK/ERK signalling. Depriving cells of glutamine prevents the activation of p53 by rRNA synthesis inhibitors. Our data reveals a mechanism that cancer cells can exploit to suppress p53-mediated apoptosis during fluctuations in environmental nutrient availability.

cancer biology↗

bioRxiv2019

Criticality of plasma membrane lipids reflects activation state of macrophage cells

Signalling is of particular importance in immune cells, and upstream in the signalling pathway many membrane receptors are functional only as complexes, co-locating with particular lipid species. Work over the last 15 years has shown that plasma membrane lipid composition is close to a critical point of phase separation, with evidence that cells adapt their composition in ways that alter the proximity to this thermodynamical point. Macrophage cells are a key component of the innate immune system, responsive to infections, regulating the local state of inflammation. We investigate changes in the plasma membranes proximity to the critical point, as a response to stimulation by various pro- and anti-inflammatory agents. Pro-inflammatory (IFN-{gamma}, Kdo-LipidA, LPS) perturbations induce an increase in the transition temperature of the GMPVs; anti-inflammatory IL4 has the opposite effect. These changes recapitulate complex plasma membrane composition changes, and are consistent with lipid criticality playing a master regulatory role: being closer to critical conditions increases membrane protein activity.

biophysics↗