C/EBPß: A transcription factor associated with the irreversible progression of Alzheimer's disease.

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative disorder distinguished by a swift cognitive deterioration accompanied by distinctive pathological hallmarks such as extracellular Aß (ß-amyloid) peptides, neuronal neurofibrillary tangles (NFTs), sustained neuroinflammation, and synaptic degeneration. The elevated frequency of AD cases and its proclivity to manifest at a younger age present a pressing challenge in the quest for novel therapeutic interventions. Numerous investigations have substantiated the involvement of C/EBPß in the progression of AD pathology, thus indicating its potential as a therapeutic target for AD treatment. AIMS: Several studies have demonstrated an elevation in the expression level of C/EBPß among individuals afflicted with AD. Consequently, this review predominantly delves into the association between C/EBPß expression and the pathological progression of Alzheimer's disease, elucidating its underlying molecular mechanism, and pointing out the possibility that C/EBPß can be a new therapeutic target for AD. METHODS: A systematic literature search was performed across multiple databases, including PubMed, Google Scholar, and so on, utilizing predetermined keywords and MeSH terms, without temporal constraints. The inclusion criteria encompassed diverse study designs, such as experimental, case-control, and cohort studies, restricted to publications in the English language, while conference abstracts and unpublished sources were excluded. RESULTS: Overexpression of C/EBPß exacerbates the pathological features of AD, primarily by promoting neuroinflammation and mediating the transcriptional regulation of key molecular pathways, including δ-secretase, apolipoprotein E4 (APOE4), acidic leucine-rich nuclear phosphoprotein-32A (ANP32A), transient receptor potential channel 1 (TRPC1), and Forkhead BoxO (FOXO). DISCUSSION: The correlation between overexpression of C/EBPß and the pathological development of AD, along with its molecular mechanisms, is evident. Investigating the pathways through which C/EBPß regulates the development of AD reveals numerous multiple vicious cycle pathways exacerbating the pathological progression of the disease. Furthermore, the exacerbation of pathological progression due to C/EBPß overexpression and its molecular mechanism is not limited to AD but also extends to other neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), and multiple sclerosis (MS). CONCLUSION: The overexpression of C/EBPß accelerates the irreversible progression of AD pathophysiology. Additionally, C/EBPß plays a crucial role in mediating multiple pathways linked to AD pathology, some of which engender vicious cycles, leading to the establishment of feedback mechanisms. To sum up, targeting C/EBPß could hold promise as a therapeutic strategy not only for AD but also for other degenerative diseases.

rs822336 binding to C/EBPß and NFIC modulates induction of PD-L1 expression and predicts anti-PD-1/PD-L1 therapy in advanced NSCLC.

Efficient predictive biomarkers are needed for immune checkpoint inhibitor (ICI)-based immunotherapy in non-small cell lung cancer (NSCLC). Testing the predictive value of single nucleotide polymorphisms (SNPs) in programmed cell death 1 (PD-1) or its ligand 1 (PD-L1) has shown contrasting results. Here, we aim to validate the predictive value of PD-L1 SNPs in advanced NSCLC patients treated with ICIs as well as to define the molecular mechanisms underlying the role of the identified SNP candidate. rs822336 efficiently predicted response to anti-PD-1/PD-L1 immunotherapy in advanced non-oncogene addicted NSCLC patients as compared to rs2282055 and rs4143815. rs822336 mapped to the promoter/enhancer region of PD-L1, differentially affecting the induction of PD-L1 expression in human NSCLC cell lines as well as their susceptibility to HLA class I antigen matched PBMCs incubated with anti-PD-1 monoclonal antibody nivolumab. The induction of PD-L1 expression by rs822336 was mediated by a competitive allele-specificity binding of two identified transcription factors: C/EBPß and NFIC. As a result, silencing of C/EBPß and NFIC differentially regulated the induction of PD-L1 expression in human NSCLC cell lines carrying different rs822336 genotypes. Analysis by binding microarray further validated the competitive allele-specificity binding of C/EBPß and NFIC to PD-L1 promoter/enhancer region based on rs822336 genotype in human NSCLC cell lines. These findings have high clinical relevance since identify rs822336 and induction of PD-L1 expression as novel biomarkers for predicting anti-PD-1/PD-L1-based immunotherapy in advanced NSCLC patients.

ERK1/2-CEBPB Axis-Regulated hBD1 Enhances Anti-Tuberculosis Capacity in Alveolar Type II Epithelial Cells.

Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), remains a global health crisis with substantial morbidity and mortality rates. Type II alveolar epithelial cells (AEC-II) play a critical role in the pulmonary immune response against Mtb infection by secreting effector molecules such as antimicrobial peptides (AMPs). Here, human ß-defensin 1 (hBD1), an important AMP produced by AEC-II, has been demonstrated to exert potent anti-tuberculosis activity. HBD1 overexpression effectively inhibited Mtb proliferation in AEC-II, while mice lacking hBD1 exhibited susceptibility to Mtb and increased lung tissue inflammation. Mechanistically, in A549 cells infected with Mtb, STAT1 negatively regulated hBD1 transcription, while CEBPB was the primary transcription factor upregulating hBD1 expression. Furthermore, we revealed that the ERK1/2 signaling pathway activated by Mtb infection led to CEBPB phosphorylation and nuclear translocation, which subsequently promoted hBD1 expression. Our findings suggest that the ERK1/2-CEBPB-hBD1 regulatory axis can be a potential therapeutic target for anti-tuberculosis therapy aimed at enhancing the immune response of AEC-II cells.

Transcription factor CEBPB mediates intracranial aneurysm rupture by inflammatory and immune response.

INTRODUCTION: Genetic factors play a major part in mediating intracranial aneurysm (IA) rupture. However, research on the role of transcription factors (TFs) in IA rupture is rare. AIMS: Bioinformatics analysis was performed to explore the TFs and related functional pathways involved in IA rupture. RESULTS: A total of 63 differentially expressed transcription factors (DETFs) were obtained. Significantly enriched biological processes of these DETFs were related to regulation of myeloid leukocyte differentiation. The top 10 DETFs were screened based on the MCC algorithm from the protein-protein interaction network. After screening and validation, it was finally determined that CEBPB may be the hub gene for aneurysm rupture. The GSEA results of CEBPB were mainly associated with the inflammatory response, which was also verified by the experimental model of cellular inflammation in vitro. CONCLUSION: The inflammatory and immune response may be closely associated with aneurysm rupture. CEBPB may be the hub gene for aneurysm rupture and may have diagnostic value. Therefore, CEBPB may serve as the diagnostic signature for RIAs and a potential target for intervention.

MiR-155-5p improves the insulin sensitivity of trophoblasts by targeting CEBPB in gestational diabetes mellitus.

INTRODUCTION: Gestational diabetes mellitus (GDM) is a prevalent pregnancy complication featuring impaired insulin sensitivity. MiR-155-5p is associated with various metabolic diseases. However, its specific role in GDM remains unclear. CCAAT enhancer binding protein beta (CEBPB), a critical role in regulating glucolipid metabolism, has been identified as a potential target of miR-155-5p. This study aims to investigate the impact of miR-155-5p and CEBPB on insulin sensitivity of trophoblasts in GDM. METHODS: Placental tissues were obtained from GDM and normal pregnant women; miR-155-5p expression was then evaluated by RT‒qPCR and CEBPB expression by western blot and immunohistochemical staining. To investigate the impact of miR-155-5p on insulin sensitivity and CEBPB expression, HTR-8/SVneo cells were transfected with either miR-155-5p mimic or inhibitor under basal and insulin-stimulated conditions. Cellular glucose uptake consumption was quantified using a glucose assay kit. Furthermore, the targeting relationship between miR-155-5p and CEBPB was validated using a dual luciferase reporter assay. RESULTS: Reduced miR-155-5p expression and elevated CEBPB expression were observed in GDM placentas and high glucose treated HTR8/SVneo cells. The overexpression of miR-155-5p significantly enhanced insulin signaling and glucose uptake in trophoblasts. Conversely, inhibiting miR-155-5p induced the opposite effects. Additionally, CEBPB was directly targeted and negatively regulated by miR-155-5p in HTR8/SVneo cells. Silencing CEBPB effectively restored the inhibitory effect of miR-155-5p downregulation on insulin sensitivity in trophoblasts. DISCUSSION: These findings suggest that miR-155-5p could enhance insulin sensitivity in trophoblasts by targeting CEBPB, highlighting the potential of miR-155-5p as a therapeutic target for improving the intrauterine hyperglycemic environment in GDM.

The protease activated receptor 2 - CCAAT/enhancer-binding protein beta - SerpinB3 axis inhibition as a novel strategy for the treatment of non-alcoholic steatohepatitis.

OBJECTIVE: The serine protease inhibitor SerpinB3 has been described as critical mediator of liver fibrosis and it has been recently proposed as an additional hepatokine involved in NASH development and insulin resistance. Protease Activated Receptor 2 has been identified as a novel regulator of hepatic metabolism. A targeted therapeutic strategy for NASH has been investigated, using 1-Piperidine Propionic Acid (1-PPA), since this compound has been recently proposed as both Protease Activated Receptor 2 and SerpinB3 inhibitor. METHODS: The effect of SerpinB3 on inflammation and fibrosis genes was assessed in human macrophage and stellate cell lines. Transgenic mice, either overexpressing SerpinB3 or carrying Serpinb3 deletion and their relative wild type strains, were used in experimental NASH models. Subgroups of SerpinB3 transgenic mice and their controls were also injected with 1-PPA to assess the efficacy of this compound in NASH inhibition. RESULTS: 1-PPA did not present significant cell and organ toxicity and was able to inhibit SerpinB3 and PAR2 in a dose-dependent manner. This effect was associated to a parallel reduction of the synthesis of the molecules induced by endogenous SerpinB3 or by its paracrine effects both in vitro and in vivo, leading to inhibition of lipid accumulation, inflammation and fibrosis in experimental NASH. At mechanistic level, the antiprotease activity of SerpinB3 was found essential for PAR2 activation, determining upregulation of the CCAAT Enhancer Binding Protein beta (C/EBP-ß), another pivotal regulator of metabolism, inflammation and fibrosis, which in turn determined SerpinB3 synthesis. CONCLUSIONS: 1-PPA treatment was able to inhibit the PAR2 - C/EBP-ß - SerpinB3 axis and to protect from NASH development and progression, supporting the potential use of a similar approach for a targeted therapy of NASH.

CaSSiDI: novel single-cell "Cluster Similarity Scoring and Distinction Index" reveals critical functions for PirB and context-dependent Cebpb repression.

PirB is an inhibitory cell surface receptor particularly prominent on myeloid cells. PirB curtails the phenotypes of activated macrophages during inflammation or tumorigenesis, but its functions in macrophage homeostasis are obscure. To elucidate PirB-related functions in macrophages at steady-state, we generated and compared single-cell RNA-sequencing (scRNAseq) datasets obtained from myeloid cell subsets of wild type (WT) and PirB-deficient knockout (PirB KO) mice. To facilitate this analysis, we developed a novel approach to clustering parameter optimization called "Cluster Similarity Scoring and Distinction Index" (CaSSiDI). We demonstrate that CaSSiDI is an adaptable computational framework that facilitates tandem analysis of two scRNAseq datasets by optimizing clustering parameters. We further show that CaSSiDI offers more advantages than a standard Seurat analysis because it allows direct comparison of two or more independently clustered datasets, thereby alleviating the need for batch-correction while identifying the most similar and different clusters. Using CaSSiDI, we found that PirB is a novel regulator of Cebpb expression that controls the generation of Ly6Clo patrolling monocytes and the expansion properties of peritoneal macrophages. PirB's effect on Cebpb is tissue-specific since it was not observed in splenic red pulp macrophages (RPMs). However, CaSSiDI revealed a segregation of the WT RPM population into a CD68loIrf8+ "neuronal-primed" subset and an CD68hiFtl1+ "iron-loaded" subset. Our results establish the utility of CaSSiDI for single-cell assay analyses and the determination of optimal clustering parameters. Our application of CaSSiDI in this study has revealed previously unknown roles for PirB in myeloid cell populations. In particular, we have discovered homeostatic functions for PirB that are related to Cebpb expression in distinct macrophage subsets.

Itraconazole inhibits tumor growth via CEBPB-mediated glycolysis in colorectal cancer.

Advanced colorectal cancer (CRC) is characterized by a high recurrence and metastasis rate, which is the primary cause of patient mortality. Unfortunately, effective anti-cancer drugs for CRC are still lacking in clinical practice. We screened FDA-approved drugs by utilizing targeted organoid sequencing data and found that the antifungal drug itraconazole had a potential therapeutic effect on CRC tumors. However, the effect and mechanism of itraconazole on CRC tumors have not been investigated. A cell line-derived xenograft model in tumor-bearing mice was established and single-cell RNA sequencing was performed on tumor samples from four mice with or without itraconazole treatment. The proportion of cell populations and gene expression profiles was significantly different between the two groups. We found that itraconazole could inhibit tumor growth and glycolysis. We revealed that CEBPB was a new target for itraconazole, and that silencing CEBPB could repress CRC glycolysis and tumor growth by inhibiting ENO1 expression. Clinical analysis showed that CEBPB expression was obviously elevated in CRC patients, and was associated with poor survival. In summary, itraconazole treatment remodeled cell composition and gene expression profiles. Itraconazole inhibited cell glycolysis and tumor growth via the CEBPB-ENO1 axis. In this study, we illustrate a new energy metabolism mechanism for itraconazole on tumor growth in CRC that will provide a theoretical basis for CRC targeting/combination therapy.

[C/EBPß mediates expressions of downstream inflammatory factors of the tumor necrosis factor-α signaling pathway in renal tubular epithelial cells with NPHP1 knockdown].

OBJECTIVE: To explore the activation of tumor necrosis factor-α (TNF-α) signaling pathway and the expressions of the associated inflammatory factors in NPHP1-defective renal tubular epithelial cells. METHODS: A human proximal renal tubular cell (HK2) model of lentivirus-mediated NPHP1 knockdown (NPHP1KD) was constructed, and the expressions of TNF-α, p38, and C/EBPß and the inflammatory factors CXCL5, CCL20, IL-1ß, IL-6 and MCP-1 were detected using RT-qPCR, Western blotting or enzyme-linked immunosorbent assay. A small interfering RNA (siRNA) was transfected in wild-type and NPHP1KDHK2 cells, and the changes in the expressions of TNF-α, p38, and C/EBPß and the inflammatory factors were examined. RESULTS: NPHP1KDHK2 cells showed significantly increased mRNA expressions of TNF-α, C/EBPß, CXCL5, IL-1ß, and IL-6 (P < 0.05), protein expressions of phospho-p38 and C/EBPß (P < 0.05), and IL-6 level in the culture supernatant (P < 0.05), and these changes were significantly blocked by transfection of cells with siRNA-C/EBPß (P < 0.05). CONCLUSION: TNF-α signaling pathway is activated and its associated inflammatory factors are upregulated in NPHP1KDHK2 cells, and C/EBPß may serve as a key transcription factor to mediate these changes.

Cebp1 and Cebpß transcriptional axis controls eosinophilopoiesis in zebrafish.

Eosinophils are a group of granulocytes well known for their capacity to protect the host from parasites and regulate immune function. Diverse biological roles for eosinophils have been increasingly identified, but the developmental pattern and regulation of the eosinophil lineage remain largely unknown. Herein, we utilize the zebrafish model to analyze eosinophilic cell differentiation, distribution, and regulation. By identifying eslec as an eosinophil lineage-specific marker, we establish a Tg(eslec:eGFP) reporter line, which specifically labeled cells of the eosinophil lineage from early life through adulthood. Spatial-temporal analysis of eslec+ cells demonstrates their organ distribution from larval stage to adulthood. By single-cell RNA-Seq analysis, we decipher the eosinophil lineage cells from lineage-committed progenitors to mature eosinophils. Through further genetic analysis, we demonstrate the role of Cebp1 in balancing neutrophil and eosinophil lineages, and a Cebp1-Cebpß transcriptional axis that regulates the commitment and differentiation of the eosinophil lineage. Cross-species functional comparisons reveals that zebrafish Cebp1 is the functional orthologue of human C/EBPεP27 in suppressing eosinophilopoiesis. Our study characterizes eosinophil development in multiple dimensions including spatial-temporal patterns, expression profiles, and genetic regulators, providing for a better understanding of eosinophilopoiesis.

Deficiency of circadian clock gene Bmal1 exacerbates noncanonical inflammasome-mediated pyroptosis and lethality via Rev-erbα-C/EBPß-SAA1 axis.

Circadian arrhythmia has been linked to increased susceptibility to multiple inflammatory diseases, such as sepsis. However, it remains unclear how disruption of the circadian clock modulates molecular aspects of innate immune responses, including inflammasome signaling. Here, we examined the potential role of the circadian clock in inflammasome-mediated responses through myeloid-specific deletion of BMAL1, a master circadian clock regulator. Intriguingly, Bmal1 deficiency significantly enhanced pyroptosis of macrophages and lethality of mice under noncanonical inflammasome-activating conditions but did not alter canonical inflammasome responses. Transcriptome analysis of enriched peritoneal myeloid cells revealed that Bmal1 deficiency led to a marked reduction in Rev-erbα expression at steady state and a significant increase in serum amyloid A1 (SAA1) expression upon poly(I:C) stimulation. Notably, we found that the circadian regulator Rev-erbα is critical for poly(I:C)- or interferon (IFN)-ß-induced SAA1 production, resulting in the circadian oscillation pattern of SAA1 expression in myeloid cells. Furthermore, exogenously applied SAA1 markedly increased noncanonical inflammasome-mediated pyroptosis of macrophages and lethality of mice. Intriguingly, our results revealed that type 1 IFN receptor signaling is needed for poly(I:C)- or IFN-ß-induced SAA1 production. Downstream of the type 1 IFN receptor, Rev-erbα inhibited the IFN-ß-induced association of C/EBPß with the promoter region of Saa1, leading to the reduced transcription of Saa1 in macrophages. Bmal1-deficient macrophages exhibited enhanced binding of C/EBPß to Saa1. Consistently, the blockade of Rev-erbα by SR8278 significantly increased poly(I:C)-stimulated SAA1 transcription and noncanonical inflammasome-mediated lethality in mice. Collectively, our data demonstrate a potent suppressive effect of the circadian clock BMAL1 on the noncanonical inflammasome response via the Rev-erbα-C/EBPß-SAA1 axis.

CircDYM attenuates microglial apoptosis via CEBPB/ZC3H4 axis in LPS-induced mouse model of depression.

Major depressive disorder (MDD) is a highly prevalent condition and one of the most common psychiatric disorders worldwide. Circular RNA (circRNA) has been increasingly implicated in MDD. However, a comprehensive understanding of circRNA and microglial apoptosis in depression is incomplete. Here, we show that circDYM inhibits microglial apoptosis induced by LPS via CEBPB/ZC3H4 axis. CircDYM prevents the translocation of CEBPB from cytoplasm to the nucleus by binding with CEBPB. Moreover, LPS-induced CEBPB nuclear entry downregulates the expression of ZC3H4, in which promotes autophagy and apoptosis in microglia. Taken together, our findings provide new insights into the relationship between circDYM and microglial apoptosis and shed new light on the function of this novel mechanism in depression-associated complex changes in the brain.

Melatonin alleviates the toxic effect of di(2-ethylhexyl) phthalate on oocyte quality resulting from CEBPB suppression during primordial follicle formation.

Presently, the exposure of plasticizers to humans and animals occurs daily, which pose a potential threat to reproductive health. In the present study, a pregnant mouse model exposed to di(2-ethylhexyl) phthalate (DEHP, one of the most common plasticizers) and melatonin was established, and the single-cell transcriptome technology was applied to investigate the effects of melatonin in ovarian cells against DEHP. Results showed that DEHP markedly altered the gene expression pattern of ovarian cells, and severely weakened the histone methylation modification of oocytes. The administration of melatonin recovered the expression of LHX8 and SOHLH1 proteins that essential for primordial follicle formation, and increased the expression of CEBPB, as well as key genes of histone methylation modification (such as Smyd3 and Kdm5a). In addition, the ovarian damage caused by DEHP was also relieved after the overexpression of CEBPB, which suggested melatonin could improve primordial follicle formation progress via enhancing CEBPB expression in mice. Besides, the apoptosis of ovarian cells induced by DEHP also was diminished by melatonin. The study provides evidence of melatonin preventing the damage mediated by plasticizers on the reproductive system in females and CEBPB may serve as a downstream target factor of melatonin in the process.

Transcriptional regulation of the Japanese flounder Cu,Zn-SOD (Jfsod1) gene in RAW264.7 cells during oxidative stress caused by causative bacteria of edwardsiellosis.

Edwardsiellosis is one of the most important bacterial diseases in fish, sometimes causing extensive economic losses in the aquaculture industry. Our previous studies demonstrated that the Cu,Zn-SOD (sod1) activity has significantly increased in Japanese flounder, Paralichthys olivaceus, hepatopancreas infected by causative bacteria of edwardsiellosis Edwardsiella tarda NUF251. In this study, NUF251 stimulated intracellular superoxide radical production in mouse macrophage RAW264.7 cells, which was reduced by N-acetylcysteine. This result suggests that NUF251 infection causes oxidative stress. To evaluate the regulatory mechanism of Jfsod1 at transcriptional levels under oxidative stress induced by NUF251 infection, we cloned and determined the nucleotide sequence (1124 bp) of the 5'-flanking region of the Jfsod1 gene. The sequence analysis demonstrated that the binding sites for the transcription factors C/EBPα and NF-IL6 involved in the transcriptional regulation of the mammalian sod1 gene existed. We constructed a luciferase reporter system with the 5'-flanking region (-1124/-1) of the Jfsod1 gene, and a highly increased transcriptional activity of the region was observed in NUF251-infected RAW264.7 cells. Further studies using several mutants indicated that deletion of the recognition region of NF-IL6 (-272/-132) resulted in a significant decrease in the transcriptional activity of the Jfsod1 gene in NUF251-infected RAW264.7 cells. In particular, the binding site (-202/-194) for NF-IL6 might play a major role in upregulating the transcriptional activity of the 5'-flanking region of the Jfsod1 gene in response to oxidative stress induced by NUF251 infection. These results could be provided a new insight to understand the pathogenic mechanism of causative bacteria of edwardsiellosis.

The paracrine isthmin1 transcriptionally regulated by C/EBPß exacerbates pulmonary vascular leakage in murine sepsis.

It is known that pulmonary vascular leakage, a key pathological feature of sepsis-induced lung injury, is largely regulated by perivascular cells. However, the underlying mechanisms have not been fully uncovered. In the present study, we aimed to evaluate the role of isthmin1, a secretory protein originating from alveolar epithelium, in the pulmonary vascular leakage during sepsis and to investigate the regulatory mechanisms of isthmin1 gene transcription. We observed an elevated isthmin1 gene expression in the pulmonary tissue of septic mice induced by cecal ligation and puncture (CLP), as well as in primary murine alveolar type II epithelial cells (ATII) exposed to lipopolysaccharide (LPS). Furthermore, we confirmed that isthmin1 derived from ATII contributes to pulmonary vascular leakage during sepsis. Specifically, adenovirus-mediated isthmin1 disruption in ATII led to a significant attenuation of the increased pulmonary microvascular endothelial cell (PMVEC) hyperpermeability in a PMVEC/ATII coculture system when exposed to LPS. In addition, adeno-associated virus 9 (AAV9)-mediated knockdown of isthmin1 in the alveolar epithelium of septic mice significantly attenuated pulmonary vascular leakage. Finally, mechanistic studies unveiled that nuclear transcription factor CCAAT/enhancer binding protein (C/EBP)ß participates in isthmin1 gene activation by binding directly to the cis-regulatory element of isthmin1 locus and may contribute to isthmin1 upregulation during sepsis. Collectively, the present study highlighted the impact of the paracrine protein isthmin1, derived from ATII, on the exacerbation of pulmonary vascular permeability in sepsis and revealed a new regulatory mechanism for isthmin1 gene transcription.NEW & NOTEWORTHY This article addresses the role of the alveolar epithelial-secreted protein isthmin1 on the exacerbation of pulmonary vascular permeability in sepsis and identified nuclear factor CCAAT/enhancer binding protein (C/EBP)ß as a new regulator of isthmin1 gene transcription. Targeting the C/EBPß-isthmin1 regulatory axis on the alveolar side would be of great value in the treatment of pulmonary vascular leakage and lung injury induced by sepsis.

Suppression of lipopolysaccharide-induced COX-2 expression via p38MAPK, JNK, and C/EBPß phosphorylation inhibition by furomagydarin A, a benzofuran glycoside from Magydaris pastinacea.

The phytochemical investigation of the methanol extract of the seeds of Magydaris pastinacea afforded two undescribed benzofuran glycosides, furomagydarins A-B (1, 2), together with three known coumarins. The structures of the new isolates were elucidated after extensive 1D and 2D NMR experiments as well as HR MS. Compound 1 was able to inhibit the COX-2 expression in RAW264.7 macrophages exposed to lipopolysaccharide, a pro-inflammatory stimulus. RT-qPCR and luciferase reporter assays suggested that compound 1 reduces COX-2 expression at the transcriptional level. Further studies highlighted the capability of compound 1 to suppress the LPS-induced p38MAPK, JNK, and C/EBPß phosphorylation, leading to COX-2 down-regulation in RAW264.7 macrophages.

Transcriptional factor CCAAT enhancer binding protein beta inhibits epithelial-mesenchymal transition in cervical cancer via regulating attractin-like 1.

Cervical cancer (CC) is a malignancy seriously endangering women's life and health worldwide. GEPIA demonstrated that attractin-like 1 (ATRNL1) presents downregulation in CC tissue. Transcription factor CCAAT enhancer binding protein beta (CEBPB) was previously revealed to present depletion in CC tissue. We attempted to clarify molecular mechanism between ATRNL1 and CEBPB underlying CC progression. Bioinformatics, RT-qPCR and western blotting revealed expression characteristics of ATRNL1 in CC. RT-qPCR measured ATRNL1 and CEBPB levels in CC cell lines. Gain-of-function assays clarified role of ATRNL1 in CC cell behaviors. Bioinformatics, Pearson correlation, ChIP and luciferase reporter experiments assessed association of ATRNL1 and CEBPB in CC cells. Rescue assays assessed regulatory function of CEBPB-ATRNL1 in CC cellular processes. ATRNL1 showed depletion in CC tissue and cells at mRNA and protein levels. ATRNL1 upregulation repressed CC cell viability, migration and EMT. CEBPB bound to ATRNL1 promoter to transcriptionally upregulate ATRNL1 in CC cells. The impact of CEBPB elevation on CC cell viability, migration and EMT were countervailed by ATRNL1 depletion. ATRNL1 and CEBPB present depletion and serve as tumor suppressors in CC cells. ATRNL1 suppresses CC cell malignancy through CEBPB activation, which may provide a potential new direction for seeking therapeutic plans for CC.

Deubiquitinase USP1 enhances CCAAT/enhancer-binding protein beta (C/EBPß) stability and accelerates adipogenesis and lipid accumulation.

Dysregulation of the ubiquitin-proteasome system has been implicated in the pathogenesis of several metabolic disorders, including obesity, diabetes, and non-alcoholic fatty liver disease; however, the mechanisms controlling pathogenic metabolic disorders remain unclear. Transcription factor CCAAT/enhancer binding protein beta (C/EBPß) regulates adipogenic genes. The study showed that the expression level of C/EBPß is post-translationally regulated by the deubiquitinase ubiquitin-specific protease 1 (USP1) and that USP1 expression is remarkably upregulated during adipocyte differentiation and in the adipose tissue of mice fed a high-fat diet (HFD). We found that USP1 directly interacts with C/EBPß. Knock-down of USP1 decreased C/EBPß protein stability and increased its ubiquitination. Overexpression of USP1 regulates its protein stability and ubiquitination, whereas catalytic mutant of USP1 had no effect on them. It suggests that USP1 directly deubiquitinases C/EBPß and increases the protein expression, leading to adipogenesis and lipid accumulation. Notably, the USP1-specific inhibitor ML323-originally developed to sensitize cancer cells to DNA-damaging agents-decreased adipocyte differentiation and lipid accumulation in 3T3-L1 cells without cytotoxicity. Oral gavage of ML323 was administered to HFD-fed mice, which showed weight loss and improvement in insulin and glucose sensitivity. Both fat mass and adipocyte size in white adipose tissues were significantly reduced by ML323 treatment, which also reduced the expression of genes involved in adipogenesis and inflammatory responses. ML323 also reduced lipid accumulation, hepatic triglycerides, free fatty acids, and macrophage infiltration in the livers of HFD-fed mice. Taken together, we suggest that USP1 plays an important role in adipogenesis by regulating C/EBPß ubiquitination, and USP1-specific inhibitor ML323 is a potential treatment option and further study by ML323 is needed for clinical application for metabolic disorders.

Surface-exposed chaperonin 60 derived from Propionibacterium freudenreichii MJ2 inhibits adipogenesis by decreasing the expression of C/EBPα/PPARγ.

Recent studies have shown that the health benefits of probiotics are not limited to those offered by living bacteria. It was reported that both live and killed cells of Propionibacterium freudenreichii MJ2 (MJ2) isolated from raw milk showed antiobesity activity in 3T3-L1 cells and high-fat diet-induced obese mice. This study was aimed at identifying the active component(s) responsible for the antiadipogenic activity of MJ2. Cell wall, surface protein, and cytoplasmic fractions of MJ2 were investigated for their inhibitory effects on adipogenesis in 3T3-L1 cells. Adipocytes treated with the surface protein fraction showed significantly lower lipid accumulation. Using the MASCOT algorithm following LC-MS/MS analysis, 131 surface proteins were identified and they were principally classified into three categories (network clusters related to ribosomes, carbon metabolism, and chaperones). Among them, chaperonin 60 (Cpn60) was selected as a potential candidate protein. Cpn60 inhibited lipid accumulation and adipogenesis during the early period of differentiation (days 0-2) and decreased expression of genes related to adipogenesis (Pparg and Cebpa) and lipogenesis (Fas and Scd1). The expression of Gata2/3, which suppresses adipogenesis, significantly increased in Cpn60-treated cells. Moreover, the nuclear translocation of C/EBPß was inhibited by Cpn60 treatment. In conclusion, Cpn60, a surface protein in MJ2, shows antiadipogenic activity by reducing the expression of C/EBPß through the upregulation of Gata2/3 expression followed by downregulation of Pparg and Cebpa expression.

Immune pathway activation in neurons triggers neural damage after stroke.

Ischemic brain injury is a severe medical condition with high incidences in elderly people without effective treatment for the resulting neural damages. Using a unilateral mouse stroke model, we analyze single-cell transcriptomes of ipsilateral and contralateral cortical penumbra regions to objectively reveal molecular events with single-cell resolution at 4 h and 1, 3, and 7 days post-injury. Here, we report that neurons are among the first cells that sense the lack of blood supplies by elevated expression of CCAAT/enhancer-binding protein ß (C/EBPß). To our surprise, the canonical inflammatory cytokine gene targets for C/EBPß, including interleukin-1ß (IL-1ß) and tumor necrosis factor α (TNF-α), are subsequently induced also in neuronal cells. Neuronal-specific silencing of C/EBPß or IL-1ß and TNF-α substantially alleviates downstream inflammatory injury responses and is profoundly neural protective. Taken together, our findings reveal a neuronal inflammatory mechanism underlying early pathological triggers of ischemic brain injury.