BMB Reports最新文献

Hypoxia-inducible factors (HIFs) act as master regulators of hypoxia-induced pathological angiogenesis, a hallmark of various ischemic ocular diseases. Recent evidence highlights the pivotal role of HIF-2α among HIF isoforms in endothelial cells (ECs) undergoing pathological angiogenesis under hypoxic conditions. However, the regulatory mechanisms underlying endothelial HIF-2α expression during hypoxia remain incompletely understood. Here, we demonstrated that stem cell factor (SCF) and its receptor, cKIT, upregulate HIF-2α expression in hypoxic ECs. Using the oxygen-induced retinopathy (OIR) mouse model, we observed that HIF-2α was highly expressed in cKIT-positive pathological neovessels in the retina, and SCF was upregulated mainly at astrocytes in the inner retinal layer. Experiments using cKIT mutant mice and anti-SCF neutralizing antibody demonstrated that inhibition of SCF/cKIT signaling significantly reduced retinal HIF-2α expression and pathological angiogenesis in mice with OIR. Moreover, HIF-2α inhibition abolished the SCF-induced increase in the angiogenic activity of human umbilical vein ECs (HUVECs) under hypoxic conditions. Mechanistic studies in HUVECs revealed that SCF enhanced HIF-2α protein levels without affecting its mRNA levels, through AKT- and ERK1/2-dependent phosphorylation of ribosomal protein S6 kinase β-1 under hypoxia. These findings provide novel insights into the regulatory mechanisms controlling HIF-2α expression in angiogenic ECs during hypoxia and suggest that the SCF/cKIT/HIF-2 axis in hypoxic ECs represents a crucial pathway in the regulation of pathological angiogenesis in ischemic ocular diseases.

Mitochondria are crucial for energy metabolism and their dysfunction is implicated in the development of various human diseases. Direct mitochondrial transplantation has shown potential in reversing mitochondrial dysfunction in recipient cells. Mesenchymal stem cells (MSCs) present a promising approach as donor cells for such transplantation. We have previously demonstrated that tomatidine, a natural steroidal alkaloid, promotes the differentiation of human embryonic stem cells (hESCs) into mature cardiomyocytes by enhancing mitochondrial quantity and function. In this study, we assessed the capacity of hESCderived cardiomyocytes (hESC-CMs) and MSCs as donor cells for mitochondrial transplantation. Mitochondria were extracted from MSCs, immature hESC-CMs, and tomatidine-treated mature hESC-CMs. Treating MSCs with mitochondria derived from mature hESC-CMs led to a marked increase in mitochondrial protein levels, such as COX IV and MIC60, in the recipient MSCs, in comparison to those receiving mitochondria from immature hESC-CMs or MSCs. Transplantation of mature hESC-CM-derived mitochondria significantly enhanced the proliferation of recipient MSCs. These findings indicate that mature hESC-CMs are highly effective as donor cells for mitochondrial transplantation in addressing mitochondrial dysfunction.

Chimeric antigen receptor-natural killer (CAR-NK) cells are emerging as a promising platform for allogeneic, cell-based immunotherapy. Among available sources, NK cells derived from pluripotent stem cells (PSCs) provide a renewable and scalable option that overcomes the limitations of primary NK cells. Human epidermal growth factor receptor 2 (HER2), a membrane protein frequently overexpressed in many solid tumors, is an attractive target for cancer immunotherapy. In this study, we designed a green fluorescent protein (GFP)-linked anti-HER2 CAR construct and introduced it into PSCs via lentiviral transduction. Three stable PSC-derived clones (CAR-A, CAR-B, and CAR-C) were established, each co-expressing anti-HER2 CAR with GFP. After differentiation under xeno-free and feeder-free culture conditions, CAR expression was maintained in NK cells. The resulting PSC-derived CAR-NK cells displayed phenotypic and functional features comparable to wild-type PSC-derived NK cells, while showing markedly enhanced cytotoxicity against HER2-positive cancer cell lines. These findings demonstrated the potential of the use of PSC-derived anti-HER2 CAR-NK cells as a robust and scalable immunotherapy. In addition, this platform could be extended to produce CAR-NK cells directed against a wide range of tumorassociated antigens. [BMB Reports 2025; 58(11): 475-483].

Long double-stranded RNAs (dsRNAs) are recognized by innate immune response proteins, thereby initiating the integrated stress response. As these RNAs adopt an A-form helical structure, immune sensors recognize dsRNAs primarily based on their structural features, such as the length of the doublestranded stretch and the triphosphate at the 5' end, rather than on specific sequences. This structure-dependent, sequenceindependent mode of RNA recognition is also characteristic of many dsRNA-binding proteins (dsRBPs). Consequently, multiple dsRBPs share a common pool of dsRNA substrates, leading to a complex regulatory network in which proteins modulate each other's activation status and signaling activities. With the development of advanced analytical techniques capable of studying RNA sequences and structures at single-nucleotide resolution, research into dsRNA-protein interactions has advanced significantly. This review discusses the long dsRNAinteracting dsRBPs encoded in the human genome, their RNA substrates, recognition mechanisms, and the downstream effects of protein-RNA interactions, with the aim of deepening our understanding of dsRNA recognition and signaling. [BMB Reports 2025; 58(11): 451-466].

Glucosamine (GlcN), a critical substrate in the hexosamine biosynthetic pathway, is known to modulate inflammatory responses in macrophages depending on extracellular glucose concentration. In hyperglycemic conditions (25 mM glucose), GlcN suppresses the production of nitric oxide (NO) and decreases the expression of inducible nitric oxide synthase (iNOS). Conversely, under normoglycemic conditions (5 mM glucose), GlcN paradoxically enhances lipopolysaccharide (LPS)-induced iNOS expression, NO production, and the upregulation of additional proinflammatory mediators. In this study, we examined the effect of alloxan, a known O-GlcNAc transferase (OGT) inhibitor, on GlcN- and/or LPS-mediated inflammatory responses in RAW264.7 macrophage cells. Under hyperglycemic conditions, alloxan exhibited little effect on the LPS-induced or LPS plus GlcN-induced expression of iNOS, cyclooxygenase-2 (COX-2), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). In contrast, under normoglycemic conditions, alloxan significantly inhibited the induction of these inflammatory genes in response to LPS plus GlcN. At the mechanistic level, alloxan reduced NF-κB DNA-binding activity and prevented its recruitment to the iNOS promoter. In addition, alloxan attenuated GlcN-induced increase in OGlcNAcylation of the NF-κB subunit p65. Collectively, these results indicate that OGT-mediated O-GlcNAcylation of NF-κB is critical for GlcN-induced proinflammatory signaling under normoglycemia. Our work highlights the glucose dependency of O-GlcNAc cycling in macrophage responses and provides new perspectives on the metabolic regulation of innate immune responses. [BMB Reports 2025; 58(11): 467-474].

Lipocalin-2 (LCN2) is a protein secreted by activated astrocytes, and its signal peptide (SP) is essential for secretion and recruitment to the autophagic pathway. SP is a short sequence present at the N-terminus of secreted proteins, such as LCN2, which facilitates transport to the endoplasmic reticulum (ER). Although SP is cleaved during the initial stages of translation in the ER, it influences the subsequent pathways of mature proteins produced in the ER lumen. ER-generated proteins are secreted or recruited to the autophagic pathway. To explore this further, we sought to determine the functional role of SP from a novel perspective. In this study, we fused LCN2 SP to the N-terminus of ubiquitin (Ub), an intracellular protein used for the proteasomal degradation of misfolded proteins and autophagic degradation of protein aggregates. We demonstrated that SP enabled the secretion of free Ub and facilitated the targeting of Ub conjugates to the autophagic pathway. We also found that SP affected intracellular Ub conjugate levels by regulating their degradation via the autophagic pathway. Furthermore, the ER-generated Ub (UbE) showed increased participation in polyubiquitinating protein aggregates generated under proteotoxic stress conditions, promoting the formation of perinuclear aggresome-like structures, and recruitment to the autophagosome. It is highly likely that UbE shares a common route with protein aggregates before being recruited to autophagosomes. Thus, this study suggests that UbE confers an altered trafficking pathway compared with endogenous Ub, thereby facilitating protein aggregate clearance without altering Ub's intrinsic biochemical activity.

Atopic dermatitis (AD) is a chronic dermatological disorder characterized by intense pruritus and eczematous lesions. Repeated topical application of 2,4-dinitrofluorobenzene (DNFB) in NC/Nga mice produces AD-like clinical symptoms that closely resemble human AD. N-acetyl-L-alanine (L-NAA), a derivative of L-Alanine, has unknown biological and physiological effects on cutaneous tissue. In this study, we investigated whether L-NAA modifies AD-like symptoms elicited by ongoing DNFB exposure in NC/Nga mice. Topical administration of L-NAA markedly attenuated the development of AD-like cutaneous lesions triggered by DNFB. L-NAA treatment further suppressed DNFB-induced infiltration of eosinophils and mast cells and prevented the increase of serum IgE resulting from DNFB application. L-NAA treatment decreased DNFB-stimulated expression of IL-4, a Th2-associated cytokine, but increased IFN-γ expression, indicative of Th1 activity, within the skin lesions. In addition, L-NAA prevented the DNFB-driven upregulation of GATA3, a central regulator of Th2 lineage differentiation, in CD4＋ cells, with no effect on T-bet, the principal regulator of Th1 cells. These findings indicate that L-NAA can limit Th2 differentiation in the AD mouse model. Therefore, L-NAA may serve as a promising therapeutic and immunomodulatory compound against AD. [BMB Reports 2025; 58(10): 437-443].

The adult human neural stem cell (ahNSC)-conditioned medium (CM) contains various secreted factors that promote tissue repair and neuroprotection. This study aimed to identify the key secreted proteins in ahNSC-CM and investigate the role of tissue inhibitor of metalloproteinases-1 (TIMP-1) in wound healing, angiogenesis, and neuroprotection against oxygenglucose deprivation. Cytokine array and liquid chromatography- tandem mass spectrometry analysis of ahNSC-CM revealed that monocyte chemoattractant protein-1 (MCP-1) and TIMP-1 were highly abundant. Enzyme-linked immunosorbent assay confirmed the enrichment of both in ahNSC-CM. Recombinant human TIMP-1 promoted wound closure and angiogenesis, whereas neutralizing TIMP-1 in ahNSC-CM attenuated these effects, with wound healing involving the extracellular signalregulated kinases 1 and 2 and protein kinase B pathways. Furthermore, ahNSC-CM protected neurons from oxygen-glucose deprivation-induced apoptosis, and this neuroprotective effect was partially reversed by TIMP-1 neutralization, potentially through the modulation of B-cell lymphoma 2 and B-cell lymphoma 2-associated X protein expression. Consistent with findings in ahNSCs, where monocyte chemoattractant protein-1 is highly expressed and contributes to pro-angiogenic effects, our study highlights the importance of secreted factors such as TIMP-1 in the therapeutic potential of ahNSC-CM. These findings suggest that TIMP-1 is a significant component of ahNSC-CM and contributes to its regenerative and neuroprotective properties. [BMB Reports 2025; 58(10): 444-449].

Memory consolidation transforms newly acquired experiences into stable long-term memories essential for learning and cognition. This process involves systems consolidation, where memory traces are reorganized across brain regions, and synaptic consolidation, which fine-tunes local neural connections. Sleep plays a critical role in both, coordinating memory reactivation, synaptic remodeling, and long-range neural communication. Systems consolidation is supported by stagespecific brain oscillations: during NREM sleep, the coupling of slow-oscillations, spindles, and sharp-wave ripples facilitates hippocampal-cortical transfer of memory representations, while REM sleep theta oscillations contribute to memory integration, abstraction, and emotional tagging. Complementary neuromodulatory dynamics, particularly involving norepinephrine and dopamine, regulate the timing and prioritization of memory processing. At the synaptic level, sleep balances strengthening and weakening of connections through a coordinated interplay of NREM and REM activity. Recent findings also suggest that dreaming may reflect the subjective correlate of these processes, particularly through the integration of recent and remote memory fragments. Although the precise relationship between systems-level reorganization and local synaptic refinement remains unclear-partly due to current technical limitations-emerging approaches are beginning to bridge these scales. Together, these findings underscore the integrative role of sleep in optimizing memory consolidation and offer promising avenues for clinical and translational research. [BMB Reports 2025; 58(10): 425-436].

Flavonoids are plant-derived polyphenols that influence nematode behavior, yet their neuronal and molecular targets remain poorly understood. Here, we show that the free-living nematode Caenorhabditis elegans detects and avoids a green tea catechin epigallocatechin gallate (EGCG) via the SRXA-7 Gprotein coupled receptor (GPCR) in the ASH nociceptive neurons. EGCG and epicatechin gallate (ECG), both containing a galloyl group, trigger strong avoidance, unlike other green tea catechins lacking this moiety. EGCG avoidance behavior displays species- and strain-specific differences among Caenorhabditis species and wild C. elegans isolates. Moreover, it is dynamically modulated by prior experience and feeding state. The ASH chemosensory neurons are required for EGCG detection, with avoidance mediated through canonical GPCR signaling components including GRK-2, GPA-3, ODR-3, and TRPV channels OCR-2 and OSM-9. A targeted GPCR screen revealed that srxa-7 mutants exhibited specific defects in EGCG avoidance. EGCG-evoked calcium responses in ASH are reduced in srxa-7 mutants and restored by ASH-specific expression of srxa-7 cDNA. These findings indicate that SRXA-7 is a sensory receptor for galloylated polyphenols, uncovering the neuronal and molecular basis of adaptive aversive responses to dietary plant compounds in nematodes.