Multi-omics profiling of retinal pigment epithelium reveals enhancer-driven activation of RANK-NFATc1 signaling in traumatic proliferative vitreoretinopathy.

During the progression of proliferative vitreoretinopathy (PVR) following ocular trauma, previously quiescent retinal pigment epithelial (RPE) cells transition into a state of rapid proliferation, migration, and secretion. The elusive molecular mechanisms behind these changes have hindered the development of effective pharmacological treatments, presenting a pressing clinical challenge. In this study, by monitoring the dynamic changes in chromatin accessibility and various histone modifications, we chart the comprehensive epigenetic landscape of RPE cells in male mice subjected to traumatic PVR. Coupled with transcriptomic analysis, we reveal a robust correlation between enhancer activation and the upregulation of the PVR-associated gene programs. Furthermore, by constructing transcription factor regulatory networks, we identify the aberrant activation of enhancer-driven RANK-NFATc1 pathway as PVR advanced. Importantly, we demonstrate that intraocular interventions, including nanomedicines inhibiting enhancer activity, gene therapies targeting NFATc1 and antibody therapeutics against RANK pathway, effectively mitigate PVR progression. Together, our findings elucidate the epigenetic basis underlying the activation of PVR-associated genes during RPE cell fate transitions and offer promising therapeutic avenues targeting epigenetic modulation and the RANK-NFATc1 axis for PVR management.

PTHrP participates in the bone destruction of middle ear cholesteatoma via promoting macrophage differentiation into osteoclasts induced by RANKL. / PTHrP促进RANKLè¯±å¯¼å·¨å™¬ç»†èƒžåˆ†åŒ–ä¸ºç ´éª¨ç»†èƒžå‚ä¸Žä¸­è€³èƒ†è„‚ç˜¤éª¨ç ´å.

OBJECTIVES: Progressive bone resorption and destruction is one of the most critical clinical features of middle ear cholesteatoma, potentially leading to various intracranial and extracranial complications. However, the mechanisms underlying bone destruction in middle ear cholesteatoma remain unclear. This study aims to explore the role of parathyroid hormone-related protein (PTHrP) in bone destruction associated with middle ear cholesteatoma. METHODS: A total of 25 cholesteatoma specimens and 13 normal external auditory canal skin specimens were collected from patients with acquired middle ear cholesteatoma. Immunohistochemical staining was used to detect the expressions of PTHrP, receptor activator for nuclear factor-kappa B ligand (RANKL), and osteoprotegerin (OPG) in cholesteatoma and normal tissues. Tartrate-resistant acid phosphatase (TRAP) staining was used to detect the presence of TRAP positive multi-nucleated macrophages in cholesteatoma and normal tissues. Mono-nuclear macrophage RAW264.7 cells were subjected to interventions, divided into a RANKL intervention group and a PTHrP+ RANKL co-intervention group. TRAP staining was used to detect osteoclast formation in the 2 groups. The mRNA expression levels of osteoclast-related genes, including TRAP, cathepsin K (CTSK), and nuclear factor of activated T cell cytoplasmic 1 (NFATc1), were measured using real-time polymerase chain reaction (real-time PCR) after the interventions. Bone resorption function of osteoclasts was assessed using a bone resorption pit analysis. RESULTS: Immunohistochemical staining showed significantly increased expression of PTHrP and RANKL and decreased expression of OPG in cholesteatoma tissues (all P<0.05). PTHrP expression was significantly positively correlated with RANKL, the RANKL/OPG ratio, and negatively correlated with OPG expression (r=0.385, r=0.417, r=-0.316, all P<0.05). Additionally, the expression levels of PTHrP and RANKL were significantly positively correlated with the degree of bone destruction in cholesteatoma (r=0.413, r=0.505, both P<0.05). TRAP staining revealed a large number of TRAP-positive cells, including multi-nucleated osteoclasts with three or more nuclei, in the stroma surrounding the cholesteatoma epithelium. After 5 days of RANKL or PTHrP+RANKL co-intervention, the number of osteoclasts was significantly greater in the PTHrP+RANKL co-intervention group than that in the RANKL group (P<0.05), with increased mRNA expression levels of TRAP, CTSK, and NFATc1 (all P<0.05). Scanning electron microscopy of bone resorption pits showed that the number (P<0.05) and size of bone resorption pits on bone slices were significantly greater in the PTHrP+RANKL co-intervention group compared with the RANKL group. CONCLUSIONS: PTHrP may promote the differentiation of macrophages in the surrounding stroma of cholesteatoma into osteoclasts through RANKL induction, contributing to bone destruction in middle ear cholesteatoma.

Transcriptional upregulation of the myo-inositol biosynthesis pathway is enhanced by NFAT5 in hyperosmotically stressed tilapia cells.

Euryhaline fish experience variable osmotic environments requiring physiological adjustments to tolerate elevated salinity. Mozambique tilapia (Oreochromis mossambicus) possess one of the highest salinity tolerance limits of any fish. In tilapia and other euryhaline fish species, the myo-inositol biosynthesis (MIB) pathway enzymes, myo-inositol phosphate synthase (MIPS) and inositol monophosphatase 1 (IMPA1.1), are among the most upregulated mRNAs and proteins indicating the high importance of this pathway for hyperosmotic (HO) stress tolerance. These abundance changes must be precluded by HO perception and signaling mechanism activation to regulate the expression of MIPS and IMPA1.1 genes. In previous work using a O. mossambicus cell line (OmB), a reoccurring osmosensitive enhancer element (OSRE1) in both MIPS and IMPA1.1 was shown to transcriptionally upregulate these enzymes in response to HO stress. The OSRE1 core consensus (5'-GGAAA-3') matches the core binding sequence of the predominant mammalian HO response transcription factor, nuclear factor of activated T-cells (NFAT5). HO-challenged OmB cells showed an increase in NFAT5 mRNA suggesting NFAT5 may contribute to MIB pathway regulation in euryhaline fish. Ectopic expression of wild-type NFAT5 induced an IMPA1.1 promoter-driven reporter by 5.1-fold (P < 0.01). Moreover, expression of dominant negative NFAT5 in HO media resulted in a 47% suppression of the reporter signal (P < 0.005). Furthermore, reductions of IMPA1.1 (37-49%) and MIPS (6-37%) mRNA abundance were observed in HO-challenged NFAT5 knockout cells relative to control cells. Collectively, these multiple lines of experimental evidence establish NFAT5 as a tilapia transcription factor contributing to HO-induced activation of the MIB pathway.NEW & NOTEWORTHY In our study, we use a multi-pronged synthetic biology approach to demonstrate that the fish homolog of the predominant mammalian osmotic stress transcription factor nuclear factor of activated T-cells (NFAT5) also contributes to the activation of hyperosmolality inducible genes in cells of extremely euryhaline fish. However, in addition to NFAT5 the presence of other strong osmotically inducible signaling mechanisms is required for full activation of osmoregulated tilapia genes.

ARID1A safeguards the canalization of the cell fate decision during osteoclastogenesis.

Chromatin remodeler ARID1A regulates gene transcription by modulating nucleosome positioning and chromatin accessibility. While ARID1A-mediated stage and lineage-restricted gene regulation during cell fate canalization remains unresolved. Using osteoclastogenesis as a model, we show that ARID1A transcriptionally safeguards the osteoclast (OC) fate canalization during proliferation-differentiation switching at single-cell resolution. Notably, ARID1A is indispensable for the transcriptional apparatus condensates formation with coactivator BRD4/lineage-specifying transcription factor (TF) PU.1 at Nfatc1 super-enhancer during safeguarding the OC fate canalization. Besides, the antagonist function between ARID1A-cBAF and BRD9-ncBAF complex during osteoclastogenesis has been validated with in vitro assay and compound mutant mouse model. Furthermore, the antagonistic function of ARID1A-"accelerator" and BRD9-"brake" both depend on coactivator BRD4-"clutch" during osteoclastogenesis. Overall, these results uncover sophisticated cooperation between chromatin remodeler ARID1A, coactivator, and lineage-specifying TF at super-enhancer of lineage master TF in a condensate manner, and antagonist between distinct BAF complexes in the proper and balanced cell fate canalization.

NFATc1 Is Required for Vitamin D- and Phosphate-Mediated Regulation of Osteocyte Lacuno-Canalicular Remodeling.

Osteocytes are embedded in lacunae and connected by canaliculi (lacuno-canalicular network, LCN). Bones from mice with X-linked hypophosphatemia (Hyp), which have impaired production of 1,25 dihydroxyvitamin D (1,25D) and hypophosphatemia, have abnormal LCN structure that is improved by treatment with 1,25D or an anti-FGF23 targeting antibody, supporting roles for 1,25D and phosphate in regulating LCN remodeling. Bones from mice lacking the vitamin D receptor (VDR) in osteocytes (Vdrf/f;Dmp1Cre+) and mice lacking the sodium phosphate transporter 2a (Npt2aKO), which have low serum phosphate with high serum 1,25D, have impaired LCN organization, demonstrating that osteocyte-specific actions of 1,25D and hypophosphatemia regulate LCN remodeling. In osteoclasts, nuclear factor of activated T cells cytoplasmic 1 (NFATc1) is critical for stimulating bone resorption. Since osteocytes also resorb matrix, we hypothesize that NFATc1 plays a role in 1,25D and phosphate-mediated LCN remodeling. Consistent with this, 1,25D and phosphate suppress Nfatc1 mRNA expression in IDG-SW3 osteocytes, and knockdown of Nfatc1 expression in IDG-SW3 cells blocks 1,25D- and phosphate-mediated suppression of matrix resorption gene expression and 1,25D- and phosphate-mediated suppression of RANKL-induced acidification of the osteocyte microenvironment. To determine the role of NFATc1 in 1,25D- and phosphate-mediated LCN remodeling in vivo, histomorphometric analyses of tibiae from mice lacking osteocyte-specific Nfatc1 in Vdrf/f;Dmp1Cre+ and Npt2aKO mice were performed, demonstrating that bones from these mice have decreased lacunar size and expression of matrix resorption genes, and improved canalicular structure compared to Vdrf/f;Dmp1Cre+ and Npt2aKO control. This study demonstrates that NFATc1 is necessary for 1,25D- and phosphate-mediated regulation of LCN remodeling.

Arsenic sulfide enhances radiosensitivity in rhabdomyosarcoma via activating NFATc3-RAG1 mediated DNA double strand break (DSB).

Rhabdomyosarcoma (RMS) represents one of the most lethal soft-tissue sarcomas in children. The toxic trace element arsenic has been reported to function as a radiosensitizer in sarcomas. To investigate the role of arsenic sulfide (As4S4) in enhancing radiation sensitization in RMS, this study was conducted to elucidate its underlying mechanism in radiotherapy. The combination of As4S4 and radiotherapy showed significant inhibition in RMS cells, as demonstrated by the cell counting kit-8 (CCK-8) assay and flow cytometry. Subsequently, we demonstrated for the first time that As4S4, as well as the knockdown of NFATc3 led to double-strand break (DSB) through increased expression of RAG1. In vivo experiment confirmed that co-treatment efficiently inhibited RMS growth. Furthermore, survival analysis of a clinical cohort consisting of 59 patients revealed a correlation between NFATc3 and RAG1 expression and overall survival (OS). Cox regression analysis also confirmed the independent prognostic significance of NFATc3 and RAG1.Taken together, As4S4 enhances radiosensitivity in RMS via activating NFATc3-RAG1 mediated DSB. NFATc3 and RAG1 are potential therapeutic targets. As4S4 will hopefully serve as a prospective radio-sensitizing agent for RMS.

Extracellular C1qbp inhibits myogenesis by suppressing NFATc1.

Aging and lack of exercise are the most important etiological factors for muscle loss. We hypothesized that new factors that contribute to muscle loss could be identified from ones commonly altered in expression in aged and exercise-limited skeletal muscles. Mouse gastrocnemius muscles were subjected to mass spectrometry-based proteomic analysis. The muscle proteomes of hindlimb-unloaded and aged mice were compared to those of exercised and young mice, respectively. C1qbp expression was significantly upregulated in the muscles of both hindlimb-unloaded and aged mice. In vitro myogenic differentiation was not affected by altering intracellular C1qbp expression but was significantly suppressed upon recombinant C1qbp treatment. Additionally, recombinant C1qbp repressed the protein level but not the mRNA level of NFATc1. NFATc1 recruited the transcriptional coactivator p300, leading to the upregulation of acetylated histone H3 levels. Furthermore, NFATc1 silencing inhibited p300 recruitment, downregulated acetylated histone H3 levels, and consequently suppressed myogenic differentiation. The expression of C1qbp was inversely correlated with that of NFATc1 in the gastrocnemius muscles of exercised or hindlimb-unloaded, and young or aged mice. These findings demonstrate a novel role of extracellular C1qbp in suppressing myogenesis by inhibiting the NFATc1/p300 complex. Thus, C1qbp can serve as a novel therapeutic target for muscle loss.

RAC1 inhibition ameliorates IBSP-induced bone metastasis in lung adenocarcinoma.

Macrophage-to-osteoclast differentiation (osteoclastogenesis) plays an essential role in tumor osteolytic bone metastasis (BM), while its specific mechanisms remain largely uncertain in lung adenocarcinoma BM. In this study, we demonstrate that integrin-binding sialoprotein (IBSP), which is highly expressed in the cancer cells from bone metastatic and primary lesions of patients with lung adenocarcinoma, can facilitate BM and directly promote macrophage-to-osteoclast differentiation independent of RANKL/M-CSF. In vivo results further suggest that osteolytic BM in lung cancer specifically relies on IBSP-induced macrophage-to-osteoclast differentiation. Mechanistically, IBSP regulates the Rac family small GTPase 1 (Rac1)-NFAT signaling pathway and mediates the forward shift of macrophage-to-osteoclast differentiation, thereby leading to early osteolysis. Moreover, inhibition of Rac1 by EHT-1864 or azathioprine in mice models can remarkably alleviate IBSP-induced BM of lung cancer. Overall, our study suggests that tumor-secreted IBSP promotes BM by inducing macrophage-to-osteoclast differentiation, with potential as an early diagnostic maker for BM, and Rac1 can be the therapeutic target for IBSP-promoted BM in lung cancer.

Shifting phenotype and differentiation of CD11b+Gr.1+ immature heterogeneous myeloid derived adjuster cells support inflammation and induce regulators of IL17A in imiquimod induced psoriasis.

OBJECTIVE AND DESIGN: The exact immunological mechanism of widespread chronic inflammatory skin disorder psoriasis has not been fully established. CD11b+Gr.1+ myeloid-derived cells are immature heterogeneous cells with T-cell suppressive property in neoplasia; however, influence of these cells on adaptive immunity is highly contextual; therefore, we dubbed these cells as myeloid-derived adjuster cells (MDAC). We studied imiquimod induced psoriasis in mouse model and evaluated for the first time the RORγt-NFAT1 axis in MDACs and the function, differentiation and interaction of these cells with T cells. MATERIALS AND METHODS: The status of T cells and MDACs; their functionality and differentiation properties, and the roles of RORγt and NFAT1 in MDACs were evaluated using flow cytometry, qRT-PCR and confocal imaging. RESULTS: We found gradual increase in T cells and MDACs and an increase in the number of IL17 -secreting MDACs and T cells in the skin of psoriatic animals. We also noted that MDAC differentiation is biased toward M1 macrophages and DCs which perpetuate inflammation. We found that psoriatic MDACs were unable to suppress T-cell proliferation or activation but seemingly helped these T cells produce more IL17. Inhibition of the RORγt/NFAT1 axis in MDACs increased the suppressive nature of MDACs, allowing these cells to suppress the activity of psoriatic T-cells. CONCLUSION: Our results indicate that altered MDAC properties in psoriatic condition sustains pathological inflammation and RORγt and NFAT1 as promising intervention target for psoriasis management.

Kv1.3-induced hyperpolarization is required for efficient Kaposi's sarcoma-associated herpesvirus lytic replication.

Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic herpesvirus that is linked directly to the development of Kaposi's sarcoma. KSHV establishes a latent infection in B cells, which can be reactivated to initiate lytic replication, producing infectious virions. Using pharmacological and genetic silencing approaches, we showed that the voltage-gated K+ channel Kv1.3 in B cells enhanced KSHV lytic replication. The KSHV replication and transcription activator (RTA) protein increased the abundance of Kv1.3 and led to enhanced K+ channel activity and hyperpolarization of the B cell membrane. Enhanced Kv1.3 activity promoted intracellular Ca2+ influx, leading to the Ca2+-driven nuclear localization of KSHV RTA and host nuclear factor of activated T cells (NFAT) proteins and subsequently increased the expression of NFAT1 target genes. KSHV lytic replication and infectious virion production were inhibited by Kv1.3 blockers or silencing. These findings highlight Kv1.3 as a druggable host factor that is key to the successful completion of KSHV lytic replication.

Active fraction of Polyrhachis vicina (Rogers) inhibits osteoclastogenesis by targeting Trim38 mediated proteasomal degradation of TRAF6.

BACKGROUND: Reactive Oxygen Species (ROS) is a key factor in the pathogenesis of osteoporosis (OP) primarily characterized by excessive osteoclast activity. Active fraction of Polyrhachis vicina Rogers (AFPR) exerts antioxidant effects and possesses extensive promising therapeutic effects in various conditions, however, its function in osteoclastogenesis and OP is unknown. PURPOSE: The aim of this study is to elucidate the cellular and molecular mechanisms of AFPR in OP. STUDY DESIGN AND METHODS: CCK8 assay was used to evaluate the cell viability under AFPR treatment. TRAcP staining, podosome belts staining and bone resorption were used to test the effect of AFPR on osteoclastogenesis. Immunofluorescence staining was used to observe the effect of AFPR on ROS production. si-RNA transfection, coimmunoprecipitation and Western-blot were used to clarify the underlying mechanisms. Further, an ovariectomy (OVX) -induced OP mice model was used to identify the effect of AFPR on bone loss using Micro-CT scanning and histological examination. RESULTS: In the present study, AFPR inhibited osteoclast differentiation and bone resorption induced by nuclear factor-κB receptor activator (NF-κB) ligand (RANKL) in dose-/ time-dependent with no cytotoxicity. Meanwhile, AFPR decreased RANKL-mediated ROS levels and enhanced ROS scavenging enzymes. Mechanistically, AFPR promoted proteasomal degradation of TRAF6 by significantly upregulating its K48-linked ubiquitination, subsequently inhibiting NFATc1 activity. We further observed that tripartite motif protein 38 (TRIM38) could mediate the ubiquitination of TRAF6 in response to RANKL. Moreover, TRIM38 could negatively regulate the RANKL pathway by binding to TRAF6 and promoting K48-linked polyubiquitination. In addition, TRIM38 deficiency rescued the inhibition of AFPR on ROS and NFATc1 activity and osteoclastogenesis. In line with these results, AFPR reduced OP caused by OVX through ameliorating osteoclastogenesis. CONCLUSION: AFPR alleviates ovariectomized-induced bone loss via suppressing ROS and NFATc1 by targeting Trim38 mediated proteasomal degradation of TRAF6. The research offers innovative perspectives on AFPR's suppressive impact in vivo OVX mouse model and in vitro, and clarifies the fundamental mechanism.

The G9a histone methyltransferase represses osteoclastogenesis and bone resorption by regulating NFATc1 function.

Epigenetic modifications affect cell differentiation via transcriptional regulation. G9a/EHMT2 is an important epigenetic modifier that catalyzes the methylation of histone 3 lysine 9 (H3K9) and interacts with various nuclear proteins. In this study, we investigated the role of G9a in osteoclast differentiation. When we deleted G9a by infection of Cre-expressing adenovirus into bone marrow macrophages (BMMs) from G9afl/fl (Ehmt2fl/fl) and induced osteoclastic differentiation by the addition of macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL), the number of TRAP-positive multinucleated osteoclasts significantly increased compared with control. Furthermore, the mRNA expression of osteoclast markers, TRAP, and cathepsin K, and to a lesser extent, NFATc1, a critical transcription factor, increased in G9a KO cells. Infection of wild-type (WT) G9a-expressing adenovirus in G9a KO cells restored the number of TRAP-positive multinucleated cells. In G9a KO cells, increased nuclear accumulation of NFATc1 protein and decreased H3K9me2 accumulation were observed. Furthermore, ChIP experiments revealed that NFATc1 binding to its target, Ctsk promoter, was enhanced by G9a deletion. For in vivo experiments, we created G9a conditional knock-out (cKO) mice by crossing G9afl/fl mice with Rank Cre/+ (Tnfrsf11aCre/+) mice, in which G9a is deleted in osteoclast lineage cells. The trabecular bone volume was significantly reduced in female G9a cKO mice. The serum concentration of the C-terminal telopeptide of type I collagen (CTX), a bone-resorbing indicator, was higher in G9a cKO mice. In addition, osteoclasts differentiated from G9a cKO BMMs exhibited greater bone-resorbing activity. Our findings suggest that G9a plays a repressive role in osteoclastogenesis by modulating NFATc1 function.

Dynamic Foxp3-chromatin interaction controls tunable Treg cell function.

Nuclear factor Foxp3 determines regulatory T (Treg) cell fate and function via mechanisms that remain unclear. Here, we investigate the nature of Foxp3-mediated gene regulation in suppressing autoimmunity and antitumor immune response. Contrasting with previous models, we find that Foxp3-chromatin binding is regulated by Treg activation states, tumor microenvironment, and antigen and cytokine stimulations. Proteomics studies uncover dynamic proteins within Foxp3 proximity upon TCR or IL-2 receptor signaling in vitro, reflecting intricate interactions among Foxp3, signal transducers, and chromatin. Pharmacological inhibition and genetic knockdown experiments indicate that NFAT and AP-1 protein Batf are required for enhanced Foxp3-chromatin binding in activated Treg cells and tumor-infiltrating Treg cells to modulate target gene expression. Furthermore, mutations at the Foxp3 DNA-binding domain destabilize the Foxp3-chromatin association. These representative settings delineate context-dependent Foxp3-chromatin interaction, suggesting that Foxp3 associates with chromatin by hijacking DNA-binding proteins resulting from Treg activation or differentiation, which is stabilized by direct Foxp3-DNA binding, to dynamically regulate Treg cell function according to immunological contexts.

CCL22 Induces the Polarization of Immature Dendritic Cells into Tolerogenic Dendritic Cells in Radiation-Induced Lung Injury through the CCR4-Dectin2-PLC-γ2-NFATC2-Nr4a2-PD-L1 Signaling Pathway.

Recruitment of immune cells to the injury site plays a pivotal role in the pathology of radiation-associated diseases. In this study, we investigated the impact of the chemokine CCL22 released from alveolar type II epithelial (AT2) cells after irradiation on the recruitment and functional changes of dendritic cells (DCs) in the development of radiation-induced lung injury (RILI). By examining changes in CCL22 protein levels in lung tissue of C57BL/6N mice with RILI, we discovered that ionizing radiation increased CCL22 expression in irradiated alveolar AT2 cells, as did MLE-12 cells after irradiation. A transwell migration assay revealed that CCL22 promoted the migration of CCR4-positive DCs to the injury site, which explained the migration of pulmonary CCR4-positive DCs in RILI mice in vivo. Coculture experiments demonstrated that, consistent with the response of regulatory T cells in the lung tissue of RILI mice, exogenous CCL22-induced DCs promoted regulatory T cell proliferation. Mechanistically, we demonstrated that Dectin2 and Nr4a2 are key targets in the CCL22 signaling pathway, which was confirmed in pulmonary DCs of RILI mice. As a result, CCL22 upregulated the expression of PD-L1, IL-6, and IL-10 in DCs. Consequently, we identified a mechanism in which CCL22 induced DC tolerance through the CCR4-Dectin2-PLC-γ2-NFATC2-Nr4a2-PD-L1 pathway. Collectively, these findings demonstrated that ionizing radiation stimulates the expression of CCL22 in AT2 cells to recruit DCs to the injury site and further polarizes them into a tolerant subgroup of CCL22 DCs to regulate lung immunity, ultimately providing potential therapeutic targets for DC-mediated RILI.

PDIA3 orchestrates effector T cell program by serving as a chaperone to facilitate the non-canonical nuclear import of STAT1 and PKM2.

Dysregulated T cell activation underpins the immunopathology of rheumatoid arthritis (RA), yet the machineries that orchestrate T cell effector program remain incompletely understood. Herein, we leveraged bulk and single-cell RNA sequencing data from RA patients and validated protein disulfide isomerase family A member 3 (PDIA3) as a potential therapeutic target. PDIA3 is remarkably upregulated in pathogenic CD4 T cells derived from RA patients and positively correlates with C-reactive protein level and disease activity score 28. Pharmacological inhibition or genetic ablation of PDIA3 alleviates RA-associated articular pathology and autoimmune responses. Mechanistically, T cell receptor signaling triggers intracellular calcium flux to activate NFAT1, a process that is further potentiated by Wnt5a under RA settings. Activated NFAT1 then directly binds to the Pdia3 promoter to enhance the expression of PDIA3, which complexes with STAT1 or PKM2 to facilitate their nuclear import for transcribing T helper 1 (Th1) and Th17 lineage-related genes, respectively. This non-canonical regulatory mechanism likely occurs under pathological conditions, as PDIA3 could only be highly induced following aberrant external stimuli. Together, our data support that targeting PDIA3 is a vital strategy to mitigate autoimmune diseases, such as RA, in clinical settings.

Calcineurin/NFATc1 pathway represses cellular cytotoxicity by modulating histone H3 expression.

Excess amounts of histones in the cell induce mitotic chromosome loss and genomic instability, and are therefore detrimental to cell survival. In yeast, excess histones are degraded by the proteasome mediated via the DNA damage response factor Rad53. Histone expression, therefore, is tightly regulated at the protein level. Our understanding of the transcriptional regulation of histone genes is far from complete. In this study, we found that calcineurin inhibitor treatment increased histone protein levels, and that the transcription factor NFATc1 (nuclear factor of activated T cells 1) repressed histone transcription and acts downstream of the calcineurin. We further revealed that NFATc1 binds to the promoter regions of many histone genes and that histone transcription is downregulated in a manner dependent on intracellular calcium levels. Indeed, overexpression of histone H3 markedly inhibited cell proliferation. Taken together, these findings suggest that NFATc1 prevents the detrimental effects of histone H3 accumulation by inhibiting expression of histone at the transcriptional level.

Attenuating bone loss in osteoporosis: the potential of corylin (CL) as a therapeutic agent.

The global prevalence of osteoporosis is being exacerbated by the increasing number of aging societies and longer life expectancies. In response, numerous drugs have been developed in recent years to mitigate bone resorption and enhance bone density. Nonetheless, the efficacy and safety of these pharmaceutical interventions remain constrained. Corylin (CL), a naturally occurring compound derived from the anti-osteoporosis plant Psoralea corylifolia L., has exhibited promising potential in impeding osteoclast differentiation. This study aims to evaluate the effect and molecular mechanisms of CL regulating osteoclast differentiation in vitro and its potential as a therapeutic agent for osteoporosis treatment in vivo. Our investigation revealed that CL effectively inhibits osteoclast formation and their bone resorption capacity by downregulating the transcription factors NFATc1 and c-fos, consequently resulting in the downregulation of genes associated with bone resorption. Furthermore, it has been observed that CL can effectively mitigate the migration and fusion of pre-osteoclast, while also attenuating the activation of mitochondrial mass and function. The results obtained from an in vivo study have demonstrated that CL is capable of attenuating the bone loss induced by ovariectomy (OVX). Based on these significant findings, it is proposed that CL exhibits considerable potential as a novel drug strategy for inhibiting osteoclast differentiation, thereby offering a promising approach for the treatment of osteoporosis.

Mef2d potentiates type-2 immune responses and allergic lung inflammation.

Innate lymphoid cells (ILCs) and adaptive T lymphocytes promote tissue homeostasis and protective immune responses. Their production depends on the transcription factor GATA3, which is further elevated specifically in ILC2s and T helper 2 cells to drive type-2 immunity during tissue repair, allergic disorders, and anti-helminth immunity. The control of this crucial up-regulation is poorly understood. Using CRISPR screens in ILCs we identified previously unappreciated myocyte-specific enhancer factor 2d (Mef2d)-mediated regulation of GATA3-dependent type-2 lymphocyte differentiation. Mef2d-deletion from ILC2s and/or T cells specifically protected against an allergen lung challenge. Mef2d repressed Regnase-1 endonuclease expression to enhance IL-33 receptor production and IL-33 signaling and acted downstream of calcium-mediated signaling to translocate NFAT1 to the nucleus to promote type-2 cytokine-mediated immunity.

Effects of hypernatremia on the microglia.

Signs and symptoms of hypernatremia largely indicate central nervous system dysfunction. Acute hypernatremia can cause demyelinating lesions similar to that observed in osmotic demyelination syndrome (ODS). We have previously demonstrated that microglia accumulate in ODS lesions and minocycline protects against ODS by inhibiting microglial activation. However, the direct effect of rapid rise in the sodium concentrations on microglia is largely unknown. In addition, the effect of chronic hypernatremia on microglia also remains elusive. Here, we investigated the effects of acute (6 or 24 h) and chronic (the extracellular sodium concentration was increased gradually for at least 7 days) high sodium concentrations on microglia using the microglial cell line, BV-2. We found that both acute and chronic high sodium concentrations increase NOS2 expression and nitric oxide (NO) production. We also demonstrated that the expression of nuclear factor of activated T-cells-5 (NFAT5) is increased by high sodium concentrations. Furthermore, NFAT5 knockdown suppressed NOS2 expression and NO production. We also demonstrated that high sodium concentrations decreased intracellular Ca2+ concentration and an inhibitor of Na+/Ca2+ exchanger, NCX, suppressed a decrease in intracellular Ca2+ concentrations and NOS2 expression and NO production induced by high sodium concentrations. Furthermore, minocycline inhibited NOS2 expression and NO production induced by high sodium concentrations. These in vitro data suggest that microglial activity in response to high sodium concentrations is regulated by NFAT5 and Ca2+ efflux through NCX and is suppressed by minocycline.