African Swine Fever Virus Envelope Glycoprotein CD2v Interacts with Host CSF2RA to Regulate the JAK2-STAT3 Pathway and Inhibit Apoptosis to Facilitate Virus Replication.

African swine fever (ASF) is a highly infectious disease caused by the African swine fever virus (ASFV) in swine. It is characterized by the death of cells in infected tissues. However, the molecular mechanism of ASFV-induced cell death in porcine alveolar macrophages (PAMs) remains largely unknown. In this study, transcriptome sequencing of ASFV-infected PAMs found that ASFV activated the JAK2-STAT3 pathway in the early stages and apoptosis in the late stages of infection. Meanwhile, the JAK2-STAT3 pathway was confirmed to be essential for ASFV replication. AG490 and andrographolide (AND) inhibited the JAK2-STAT3 pathway, promoted ASFV-induced apoptosis, and exerted antiviral effects. Additionally, CD2v promoted STAT3 transcription and phosphorylation as well as translocation into the nucleus. CD2v is the main envelope glycoprotein of the ASFV, and further investigations showed that CD2v deletion downregulates the JAK2-STAT3 pathway and promotes apoptosis to inhibit ASFV replication. Furthermore, we discovered that CD2v interacts with CSF2RA, which is a hematopoietic receptor superfamily member in myeloid cells and a key receptor protein that activates receptor-associated JAK and STAT proteins. In this study, CSF2RA small interfering RNA (siRNA) downregulated the JAK2-STAT3 pathway and promoted apoptosis to inhibit ASFV replication. Taken together, ASFV replication requires the JAK2-STAT3 pathway, while CD2v interacts with CSF2RA to regulate the JAK2-STAT3 pathway and inhibit apoptosis to facilitate virus replication. These results provide a theoretical basis for the escape mechanism and pathogenesis of ASFV. IMPORTANCE African swine fever is a hemorrhagic disease caused by the African swine fever virus (ASFV), which infects pigs of different breeds and ages, with a fatality rate of up to 100%. It is one of the key diseases affecting the global livestock industry. Currently, no commercial vaccines or antiviral drugs are available. Here, we show that ASFV replicates via the JAK2-STAT3 pathway. More specifically, ASFV CD2v interacts with CSF2RA to activate the JAK2-STAT3 pathway and inhibit apoptosis, thereby maintaining the survival of infected cells and promoting viral replication. This study revealed an important implication of the JAK2-STAT3 pathway in ASFV infection and identified a novel mechanism by which CD2v has evolved to interact with CSF2RA and maintain JAK2-STAT3 pathway activation to inhibit apoptosis, thus elucidating new information regarding the signal reprogramming of host cells by ASFV.

M2 macrophages-derived exosomes regulate osteoclast differentiation by the CSF2/TNF-α axis.

BACKGROUND: Osteoclast-mediated bone resorption cause bone loss in several bone diseases. Exosomes have been reported to regulate osteoclast differentiation. M2-polarized macrophages exhibit anti-inflammatory activity. This study aimed to explore the effect of exosomes from M2 polarized macrophages (M2-exos) on osteoclastogenesis and molecular mechanisms. METHODS: M2-exos were isolated from IL-4-induced Raw264.7 cells (M2 macrophages) and used to treat osteoclasts (RANKL-induced Raw264.7 cells). Osteoclast differentiation was visualized using tartrate resistant acid phosphatase staining. Quantitative real-time PCR (qPCR) was conducted to measure the levels of osteoclastogenesis-related genes. The underlying mechanisms of M2-exos were evaluated using qPCR and western blotting. RESULTS: M2-exos suppressed osteoclast differentiation induced by RANKL. Additionally, CSF2 was highly expressed in M2 macrophages, and knockdown of CSF2 further enhanced the effects of M2-exos on osteoclast differentiation. Moreover, CSF2 positively regulated TNF-α signaling, which inhibition promoted differentiation of M2-exo-treated osteoclasts. CONCLUSION: M2-exos inhibited RANKL-induced osteoclast differentiation by downregulating the CSF2 expression through inactivating the TNF-α signaling, suggesting the potential application of exosomes in bone disease therapy.

A STUB1 ubiquitin ligase/CHIC2 protein complex negatively regulates the IL-3, IL-5, and GM-CSF cytokine receptor common ß chain (CSF2RB) protein stability.

The IL-3, IL-5, and GM-CSF family of cytokines play an essential role in the growth, differentiation, and effector functions of multiple hematopoietic cell types. Receptors in this family are composed of cytokine-specific α chains and a common ß chain (CSF2RB), responsible for the majority of downstream signaling. CSF2RB abundance and stability influence the magnitude of the cellular response to cytokine stimulation, but the exact mechanisms of regulation are not well understood. Here, we use genetic screens in multiple cellular contexts and cytokine conditions to identify STUB1, an E3 ubiquitin ligase, and CHIC2 as regulators of CSF2RB ubiquitination and protein stability. We demonstrate that Stub1 and Chic2 form a complex that binds Csf2rb and that genetic inactivation of either Stub1 or Chic2 leads to reduced ubiquitination of Csf2rb. The effects of Stub1 and Chic2 on Csf2rb were greatest at reduced cytokine concentrations, suggesting that Stub1/Chic2-mediated regulation of Csf2rb is a mechanism of reducing cell surface accumulation when cytokine levels are low. Our study uncovers a mechanism of CSF2RB regulation through ubiquitination and lysosomal degradation and describes a role for CHIC2 in the regulation of a cytokine receptor.

Colony stimulating factor 2 protects the preimplantation bovine embryo from heat shock.

Heat stress can have severe deleterious effects on embryo development and survival. The present study evaluated whether CSF2 can protect the developmental competence of the bovine embryo following exposure to a heat shock of 41°C at the zygote and morula stages. In the first experiment, putative zygotes and 2-cell embryos were assigned to receive either 10 ng/ml CSF2 or vehicle, and then cultured for 15 h at either 38.5°C or 41°C and then at 38.5°C until day 7.5. Heat shock reduced blastocyst development for embryos treated with vehicle but not for embryos cultured with CSF2. In the second experiment, day 5 embryos (morula) were treated with CSF2 or vehicle and then cultured for 15 h at either 38.5°C or 41°C and then at 38.5°C until day 7.5. Temperature treatment did not affect development to the blastocyst stage and there was no effect of CSF2 treatment or the interaction. Results indicate that CSF2 can reduce the deleterious effects of heat shock at the zygote or two-cell stage when the embryo is transcriptionally inactive.

Microglial reduction of colony stimulating factor-1 receptor expression is sufficient to confer adult onset leukodystrophy.

Adult onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP) is a dementia resulting from dominantly inherited CSF1R inactivating mutations. The Csf1r+/- mouse mimics ALSP symptoms and pathology. Csf1r is mainly expressed in microglia, but also in cortical layer V neurons that are gradually lost in Csf1r+/- mice with age. We therefore examined whether microglial or neuronal Csf1r loss caused neurodegeneration in Csf1r+/- mice. The behavioral deficits, pathologies and elevation of Csf2 expression contributing to disease, previously described in the Csf1r+/- ALSP mouse, were reproduced by microglial deletion (MCsf1rhet mice), but not by neural deletion. Furthermore, increased Csf2 expression by callosal astrocytes, oligodendrocytes, and microglia was observed in Csf1r+/- mice and, in MCsf1rhet mice, the densities of these three cell types were increased in supraventricular patches displaying activated microglia, an early site of disease pathology. These data confirm that ALSP is a primary microgliopathy and inform future therapeutic and experimental approaches.

Regulation of gene expression in the bovine blastocyst by colony-stimulating factor 2 is disrupted by CRISPR/Cas9-mediated deletion of CSF2RA.

Colony-stimulating factor 2 (CSF2) functions in the reproductive tract to modulate the function of the preimplantation embryo. The ß subunit of the CSF2 receptor (CSF2RB) is not expressed in the embryo, and signal transduction is therefore different than for myeloid cells where the receptor is composed of α (CSF2RA) and ß subunits. Here, we produced embryos in which exons 5 and 6 of CSF2RA were disrupted using the CRISPR/Cas 9 system to test whether CSF2RA signaling was essential for actions of CSF2 in the bovine embryo. Wild-type and CSF2RA knockout embryos were treated with 10 ng/mL CSF2 or vehicle at day 5 of development. Blastocysts were harvested at day 8 to determine transcript abundance of 90 genes by real-time polymerase chain reaction (PCR). Responses in female blastocysts were examined separately from male blastocysts because actions of CSF2 are sex-dependent. For wild-type embryos, CSF2 altered expression of 10 genes in females and 20 in males. Only three genes were affected by CSF2 in a similar manner for both sexes. Disruption of CSF2RA prevented the effect of CSF2 on expression for 9 of 10 CSF2-regulated genes in females and 19 of 20 genes in males. The results confirm the importance of CSF2RA for regulation of gene expression by CSF2 in the blastocyst.

Roles of acyl-CoA synthetase long-chain family member 5 and colony stimulating factor 2 in inhibition of palmitic or stearic acids in lung cancer cell proliferation and metabolism.

Lung cancer is a heterogeneous and complex disease with the highest incidence and mortality rate. The present study aims at defining the lung cancer phenome specificity of lipidomic profiles, screening target lipid-dependent transcriptional alternations, identifying target lipid-associated target genes, and exploring molecular mechanisms. Lung cancer-specific and lung cancer subtype-specific target lipids palmitic acid (C16:0) and stearic acid (C18:0) were found as target lipids by integrating clinical phenomics, lipidomics, and transcriptomics and exhibited antiproliferative effects in sensitive cells. The metabolism-associated gene ACSL5 or inflammation-associated gene CCL3 was identified in lung adenocarcinoma or small lung cancer cells, respectively. C16:0 or C18:0 could upregulate ACSL5 or CSF2 expression in a time- and dose-dependent pattern, and the deletion of both genes led to the insensitivity of cells. Target lipids increased the expression of PDK4 gene in different patterns and inhibited cell proliferation through alterations of intracellular energy. Thus, our data provide a new strategy to investigate the trans-points between clinical and phenomics and lipidomics and target lipid-associated molecular mechanisms to benefit from the discovery of new therapies.

Validation of novel hub genes and molecular mechanisms in acute lung injury using an integrative bioinformatics approach.

Acute lung injury (ALI) is an inflammatory pulmonary disease that can easily develop into serious acute respiratory distress syndrome, which has high morbidity and mortality. However, the molecular mechanism of ALI remains unclear, and few molecular biomarkers for diagnosis and treatment have been identified. In this study, we aimed to identify novel molecular biomarkers using a bioinformatics approach. Gene expression data were obtained from the Gene Expression Omnibus database, co-expressed differentially expressed genes (CoDEGs) were identified using R software, and further functional enrichment analyses were conducted using the online tool Database for Annotation, Visualization, and Integrated Discovery. A protein-protein interaction network was established using the STRING database and Cytoscape software. Lipopolysaccharide (LPS)-induced ALI mouse model was constructed and verified. The hub genes were screened and validated in vivo. The transcription factors (TFs) and miRNAs associated with the hub genes were predicted using the NetworkAnalyst database. In total, 71 CoDEGs were screened and found to be mainly involved in the cytokine-cytokine receptor interactions, and the tumor necrosis factor and malaria signaling pathways. Animal experiments showed that the lung injury score, bronchoalveolar lavage fluid protein concentration, and wet-to-dry weight ratio were higher in the LPS group than those in the control group. Real-time polymerase chain reaction analysis indicated that most of the hub genes such as colony-stimulating factor 2 (Csf2) were overexpressed in the LPS group. A total of 20 TFs including nuclear respiratory factor 1 (NRF1) and two miRNAs were predicted to be regulators of the hub genes. In summary, Csf2 may serve as a novel diagnostic and therapeutic target for ALI. NRF1 and mmu-mir-122-5p may be key regulators in the development of ALI.

Molecular fingerprint of female bovine embryos produced in vitro with high competence to establish and maintain pregnancy†.

The objective was to identify the transcriptomic profile of in vitro-derived embryos with high competence to establish and maintain gestation. Embryos produced with X-sorted sperm were cultured from day 5 to day 7 in serum-free medium containing 10 ng/ml recombinant bovine colony-stimulating factor 2 (CSF2) or vehicle. The CSF2 was administered because this molecule can increase blastocyst competence for survival after embryo transfer. Blastocysts were harvested on day 7 of culture and manually bisected. One demi-embryo from a single blastocyst was transferred into a synchronized recipient and the other half was used for RNA-seq analysis. Using P < 0.01 and a fold change >2-fold or <0.5 fold as cutoffs, there were 617 differentially expressed genes (DEG) between embryos that survived to day 30 of gestation vs those that did not, 470 DEG between embryos that survived to day 60 and those that did not, 432 DEG between embryos that maintained pregnancy from day 30 to day 60 vs those where pregnancy failed after day 30, and 635 DEG regulated by CSF2. Pathways and ontologies in which DEG were overrepresented included many related to cellular responses to stress and cell survival. It was concluded that gene expression in the blastocyst is different between embryos that are competent to establish and maintain pregnancy vs those that are not. The relationship between expression of genes related to cell stress and subsequent embryonic survival probably reflects cellular perturbations caused by embryonic development taking place in the artificial environment associated with cell culture.

A Novel Endogenous Damage Signal, CSF-2, Activates Multiple Beneficial Functions of Adipose Tissue-Derived Mesenchymal Stem Cells.

The major challenges of current mesenchymal stem cell (MSC)-based therapeutics are their low differentiation potential into specialized cell types and their homing ability to sites of injury. Therefore, many researchers have directed their efforts toward finding a novel stimulatory factor that can significantly enhance the therapeutic effects of MSCs. Colony-stimulating factor 2 (CSF-2) is previously known as a hematopoietic growth factor involved in the differentiation of various myeloid cells from hematopoietic progenitor cells. In addition to this canonical hematopoietic function, we identified for the first time that CSF-2 is actively secreted by stem cells, in response to various types of injuries, as an endogenous damage signal that promotes the therapeutic effects of MSCs by enhancing their multi-lineage differentiation and migratory capacities, possibly through its receptor CD116. Our results also revealed that CSF-2 exerts its stimulatory effects on MSCs via PI3K/Akt- and/or FAK/ERK1/2-signaling pathways. More importantly, we also found that MSCs stimulated with CSF-2 show markedly enhanced differentiation and migratory capacities and subsequent in vivo therapeutic effects in an endometrial ablation animal model. Collectively, our findings provide compelling evidence for a novel non-hematopoietic function of CSF-2 in promoting multiple beneficial functions of MSCs via a non-canonical mechanism as an endogenous damage signal.

Long-Term Safety and Efficacy of Gene-Pulmonary Macrophage Transplantation Therapy of PAP in Csf2ra-/- Mice.

Hereditary pulmonary alveolar proteinosis (PAP) is a genetic lung disease characterized by surfactant accumulation and respiratory failure arising from disruption of GM-CSF signaling. While mutations in either CSF2RA or CSF2RB (encoding GM-CSF receptor α or ß chains, respectively) can cause PAP, α chain mutations are responsible in most patients. Pulmonary macrophage transplantation (PMT) is a promising new cell therapy in development; however, no studies have evaluated this approach for hereditary PAP (hPAP) caused by Csf2ra mutations. Here, we report on the preclinical safety, tolerability, and efficacy of lentiviral-vector (LV)-mediated Csf2ra expression in macrophages and PMT of gene-corrected macrophages (gene-PMT therapy) in Csf2ra gene-ablated (Csf2ra-/-) mice. Gene-PMT therapy resulted in a stable transgene integration and correction of GM-CSF signaling and functions in Csf2ra-/- macrophages in vitro and in vivo and resulted in engraftment and long-term persistence of gene-corrected macrophages in alveoli; restoration of pulmonary surfactant homeostasis; correction of PAP-specific cytologic, histologic, and biomarker abnormalities; and reduced inflammation associated with disease progression in untreated mice. No adverse consequences of gene-PMT therapy in Csf2ra-/- mice were observed. Results demonstrate that gene-PMT therapy of hPAP in Csf2ra-/- mice was highly efficacious, durable, safe, and well tolerated.

The potential markers of NK-92 associated to cytotoxicity against K562 cells.

Markers associated to NK cytolytic activity are in a great need to regulate NK cell immunotherapy products. We assume that biomarkers which response to cytolysis will change their transcription, expression or secretion. To find NK-92 indicator to cytolytic activity, we have evaluated the potential markers by quantifying the expression of well-known cytotoxicity functional molecules (cytokine IFN-γ, Granzyme B, perforin, CD69 and CD107a), and explored candidate markers by a sweeping transcription picture of NK-92 using a direct cytolysis model (incubation with K562). We found that IFN-γ secretion was highly correlated to cytotoxicity of NK-92, neither Granzyme B, perforin secretion, nor CD69, CD107a positive population were upregulated by K562 stimulation. RNAseq revealed 432 genes expression changed during cytolysis, several genes (BIRC3, CSF2, VCAM1 and TNFRSF9) mRNA expression were validated by real time RT-PCR under K562 being killed or protected from being killed conditions. Results suggested IFN-γ secretion, BIRC3 and TNFRSF9 transcription in NK-92 were responsive to K562 cytolysis. In a word, our results confirmed one marker and reveal an array of novel candidate markers associated with NK-92 cytotoxicity. Further studies are greatly needed to determine the roles these new makers play in NK-92 cytolysis process.

Conditions of embryo culture from days 5 to 7 of development alter the DNA methylome of the bovine fetus at day 86 of gestation.

PURPOSE: We tested whether in vitro production (IVP) causes changes in DNA methylation in fetal liver and skeletal muscle and if exposure of cultured embryos to colony-stimulating factor 2 (CSF2) alters DNA methylation. METHODS: Female fetuses were produced by artificial insemination or transfer of an IVP embryo. Embryos were treated from days 5 to 7 after fertilization with CSF2 or vehicle. DNA methylation in fetal liver and skeletal muscle was determined by post-bisulfite adaptor tagging-based sequencing. The degree of DNA methylation for CpG sites in 50-bp windows of the promoter region 500 bp upstream of the transcriptional start site was compared between treatments. RESULTS: For liver, there were 12 genes (6% of those analyzed) in which DNA methylation was affected by treatment, with one 50-bp window per gene affected by treatment. For muscle, the degree of DNA methylation was affected by treatment for 32 windows (19% of the total windows analyzed) representing 28 distinct genes (23% of analyzed genes). For 19 of the 28 genes in muscle, the greatest deviation in DNA methylation was for the CSF2 group. CONCLUSION: Results are consistent with alterations in the methylome being one of the mechanisms by which IVP can result in altered fetal development and postnatal function in the resultant offspring. In addition, results indicate that maternally derived cell-signaling molecules can regulate the pattern of DNA methylation.

Regulation of present and future development by maternal regulatory signals acting on the embryo during the morula to blastocyst transition - insights from the cow.

The preimplantation embryo has a remarkable ability to execute its developmental program using regulatory information inherent within itself. Nonetheless, the uterine environment is rich in cell signaling molecules termed embryokines that act on the embryo during the morula-to-blastocyst transition, promoting blastocyst formation and programming the embryo for subsequent developmental events. Programming can not only affect developmental processes important for continuance of development in utero but also affect characteristics of the offspring during postnatal life. Given the importance of embryokines for regulation of embryonic development, it is likely that some causes of infertility involve aberrant secretion of embryokines by the uterus. Embryokines found to regulate development of the bovine embryo include insulin-like growth factor 1, colony stimulating factor 2 (CSF2), and dickkopf WNT signaling pathway inhibitor 1. Embryo responses to CSF2 exhibit sexual dimorphism, suggesting that sex-specific programming of postnatal function is caused by maternal signals acting on the embryo during the preimplantation period that regulate male embryos differently than female embryos.

Impaired RASGRF1/ERK-mediated GM-CSF response characterizes CARD9 deficiency in French-Canadians.

BACKGROUND: Caspase recruitment domain-containing protein 9 (CARD9) deficiency is an autosomal recessive primary immunodeficiency conferring human susceptibility to invasive fungal disease, including spontaneous central nervous system candidiasis (sCNSc). However, clinical characterization of sCNSc is variable, hindering its recognition. Furthermore, an in-depth understanding of the bases for this susceptibility has remained elusive. OBJECTIVES: We sought to comprehensively characterize sCNSc and to dissect the mechanisms by which a hypomorphic CARD9 mutation causes susceptibility to Candida species. METHODS: We describe the clinical and radiologic findings of sCNSc caused by CARD9 deficiency in a French-Canadian cohort. We performed genetic, cellular, and molecular analyses to further decipher its pathophysiology. RESULTS: In our French-Canadian series (n = 4) sCNSc had onset in adulthood (median, 38 years) and was often misinterpreted radiologically as brain malignancies; 1 patient had additional novel features (eg, endophthalmitis and osteomyelitis). CARD9 deficiency resulted from a hypomorphic p.Y91H mutation and allelic imbalance established in this population through founder effects. We demonstrate a consistent cellular phenotype of impaired GM-CSF responses. The ability of CARD9 to complex with B-cell CLL/lymphoma 10 (BCL10) and mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1) is intact in our series, arguing against its involvement in susceptibility to fungi. Instead, we show that the p.Y91H mutation impairs the ability of CARD9 to complex with Ras protein-specific guanine nucleotide-releasing factor 1 (RASGRF1), leading to impaired activation of nuclear factor κB and extracellular signal-regulated kinase (ERK) in monocytes and subsequent GM-CSF responses. Successful treatment of a second patient with adjunctive GM-CSF bolsters the clinical relevance of these findings. CONCLUSIONS: Hypomorphic CARD9 deficiency caused by p.Y91H results in adult-onset disease with variable penetrance and expressivity. Our findings establish the CARD9/RASGRF1/ERK/GM-CSF axis as critical to the pathophysiology of sCNSc.

rs657075 (CSF2) Is Associated with the Disease Phenotype (BAS-G) of Ankylosing Spondylitis.

Ankylosing spondylitis (AS) is a systemic autoimmune disease mainly affecting the lumbar spine and sacroiliac joints, and exhibits peripheral inflammatory arthropathy. More than 25 loci have been identified as associated with AS. Because both AS and rheumatoid arthritis (RA) are autoimmune diseases that may share some common genetic factors, we therefore examined if the newly identified RA genetic polymorphisms were associated with AS in a Taiwanese population. In this study, we enrolled 475 AS patients and 11,301 healthy subjects from a Taiwanese biobank as controls. Although none of single-nucleotide polymorphisms (SNPs) were associated with the susceptibility to AS, the AS disease index Bath AS Global (BAS-G) clinical phenotype was observed as significantly correlated to the AA genotype of rs657075 (CSF2). The significance remains after gender/age/disease duration adjustment and after group categorization by human leukocyte antigen-B 27 (HLA-B27) genotype. We further investigated the possible functions of rs657075 through bioinformatics approaches. Results revealed that polymorphism of rs657075 is able to influence the expression of acyl-CoA synthetase long-chain family member 6 (ACSL6). In conclusion, our study indicated that rs657075 (CSF2) is strongly associated with the AS disease index Bath AS Global (BAS-G) clinical phenotype.

Identification of potential biomarkers for post-traumatic complications released after trauma-hemorrhage from murine Kupffer cells and its investigation in lung and liver.

CONTEXT: Early diagnosis of complications after severe trauma by specific biomarkers remains difficult. OBJECTIVE: Identify potential new biomarkers for early diagnosis of post-traumatic complications. MATERIAL AND METHODS: Mice underwent pressure-controlled hemorrhage or sham procedure. Four hours later, genome-wide expression of isolated Kupffer cells was compared with controls using Affymetrix-Genechip-Expression-Analysis and real-time-PCR. RESULTS: Expression analysis and real-time-PCR revealed a significant increase of gene expression of Cxcl10, Il4ra, Csf2rb2, Lcn2, and Gbp5. CONCLUSION: Cxcl10, Il4ra, Csf2rb2, Lcn2, and Gbp5 might represent new biomarkers for early diagnosis of post-traumatic complications, if they are linked to the development of post-traumatic complications.

Guanabenz Downregulates Inflammatory Responses via eIF2α Dependent and Independent Signaling.

Integrated stress responses (ISR) may lead to cell death and tissue degeneration via eukaryotic translation initiation factor 2 α (eIF2α)-mediated signaling. Alleviating ISR by modulating eIF2α phosphorylation can reduce the symptoms associated with various diseases. Guanabenz is known to elevate the phosphorylation level of eIF2α and reduce pro-inflammatory responses. However, the mechanism of its action is not well understood. In this study, we investigated the signaling pathway through which guanabenz induces anti-inflammatory effects in immune cells, in particular macrophages. Genome-wide mRNA profiling followed by principal component analysis predicted that colony stimulating factor 2 (Csf2, or GM-CSF as granulocyte macrophage colony stimulating factor) is involved in the responses to guanabenz. A partial silencing of Csf2 or eIF2α by RNA interference revealed that Interleukin-6 (IL6), Csf2, and Cyclooxygenase-2 (Cox2) are downregulated by guanabenz-driven phosphorylation of eIF2α. Although expression of IL1ß and Tumor Necrosis Factor-α (TNFα) was suppressed by guanabenz, their downregulation was not directly mediated by eIF2α signaling. Collectively, the result herein indicates that anti-inflammatory effects by guanabenz are mediated by not only eIF2α-dependent but also eIF2α-independent signaling.

CSF2 Impairs Nrf2 Signaling through the Akt/Mtor Pathway in the Development of Bladder Cancer.

Bladder Cancer (BCa) is one of the most common cancers of the urinary system. Colony-stimulating factor 2 (CSF2) is involved in many cancers, but not BCa. We investigated the effect of CSF2 on BCa in this study and the underlying molecular mechanisms. CSF2 mRNA levels in BCa were analyzed using the Cancer Genome Atlas (TCGA) database. Western blot was conducted to verify CSF2 expression in BCa tissue samples and cell lines. The effect of CSF2 on the growth of BCa cells was assessed by CCK8 and colony formation. To determine the migration and invasion capabilities of BCa cells, transwell analysis and wound healing assays were conducted. Next, western blot was used to explore the underlying mechanism. In the end, a xenografted BCa mouse model was established to examine the effects of CSF2 on tumorigenesis in vivo. Results showed that CSF2 mRNA was upregulated in BCa samples. Knocking down CSF2 significantly inhibited the proliferation and tumorigenesis of BCa cells in vitro and in vivo. Mechanism analysis revealed that CSF2 knockdown inhibited the proliferation and invasion of BCa cells via AKT/mTOR signaling. Based on these results, CSF2 promotes the proliferation and tumorigenesis of BCa.

Whole Blood Transcriptome Analysis in Patients with Trigeminal Neuralgia: a Prospective Clinical Study.

Trigeminal neuralgia (TN) brings a huge burden to patients, without long-term effective treatment. This study aimed to explore the differentially expressed genes (DEGs) and related enrichment pathways in patients with TN. This was a study of transcriptome sequencing and bioinformatics analysis of human samples. Whole blood samples were collected from the TN patients and pain-free controls. RNA was extracted to conduct the RNA-sequencing and the subsequent bioinformatics analysis. DEGs between the two groups were derived. Kyoto encyclopedia of genes and genomes (KEGG) and Gene ontology (GO) was used to find the enrichment pathways of DEGs. Protein protein interaction (PPI) network was used to depict the interaction between DEGs and find the most important gene, hub gene. Compared with the control group, there were 117 up-regulated DEGs and 103 down-regulated DEGs in the whole blood of patients in the TN group. Pathway enrichment analysis showed that DEGs were mainly enriched in the neuroimmune and metabolic pathways. The PPI network demonstrated that colony stimulating factor 2 (CSF2) was the most important hub gene in the whole blood of TN patients. This study shows the expression of the transcriptome in the whole blood samples of TN patients. The neuroimmune responses and key hub gene CSF2 in the whole blood cells play a vital role in the occurrence of TN. Our research provides a theoretical basis for the diagnosis and treatments of TN. This study was registered at clinicaltrials.gov in June 2021 (No. NCT04923399).