Expression Of Granulocyte Colony-stimulating Factor Research Articles

Potential diagnostic and drug target markers in glioblastoma

Glioblastoma multiforme (GBM) IDH-wildtype is the most prevalent brain malignancy in adults. However, molecular mechanisms, which leads to GBM have not been completely elucidated. Granulocyte colony-stimulating factor (GCSF), Granulocyte colony-stimulating factor receptor GCSFR, and Signal transducers and activators of transcription 3 (STAT3) have been involved in the occurrence and development of various cancers, but their role in GBM is little known. Herein, we have investigated the gene and protein expression of GCSF, GCSFR, and STAT3 in 21 tissue biopsy samples and also in tumor associated normal tissue (TANT) samples derived from glioblastoma patients, which revealed significantly differential expression of these genes. To validate our findings, we performed a comprehensive integrated analysis of transcriptomic and proteomic profiling of respective genes by retrieving GBM RNA-sequence data from Genome Atlas Databases. GO and KEGG analysis revealed enrichment in disease-related pathways, such as JAK/STAT pathway activation, which were associated with GBM progression. We further performed computational docking analysis of potential drug candidate Nisin against GCSF, and the results were validated in vitro through cytotoxic activity assay using a human glioblastoma cell line SF-767 in a dose-dependent manner. Our comprehensive analysis reveals that GCSF augments glioma progression, and its blockade with anticancer bacteriocin peptide Nisin can potentially inhibit the growth and metastasis of GBM.

Saharan dust induces the lung disease-related cytokines granulocyte–macrophage colony-stimulating factor and granulocyte colony-stimulating factor

Desert dust exposure is associated with adverse respiratory health effects. Desert dust is a complex pollutant mixtures that includes respirable crystalline and amorphous particles, metals, and microbial constituents. Given the health effects of desert dust and its heterogeneity, as yet unidentified harmful biological pathways may be triggered. Therefore, we exposed human in vitro air–liquid interface co-cultures of alveolar epithelial A549 cells and THP-1 macrophages to Saharan dust (SD). For comparison, we used the known pulmonary toxicant DQ12 quartz dust. Via RNA sequencing, we identified that SD but not DQ12 increased the gene expression of granulocyte–macrophage colony-stimulating factor (GMCSF) and granulocyte colony-stimulating factor (GCSF). These findings were confirmed by quantitative reverse transcriptase PCR. SD dose-dependently upregulated GMCSF and GCSF expression with significant 7 and 9-fold changes, respectively, at the highest tested concentration of 31 µg/cm2. Furthermore, we observed that SD significantly enhanced the secretion of GM-CSF and G-CSF by 2-fold. Both cytokines have previously been associated with lung diseases such as asthma and fibrosis. Hence, we present two molecular messengers that may contribute to the adverse health effects of desert dust and might serve as drug targets for this globally relevant non-anthropogenic air pollutant.

Scutellarin suppresses the metastasis of triple-negative breast cancer via targeting TNFα/TNFR2-RUNX1-triggered G-CSF expression in endothelial cells

Triple-negative breast cancer (TNBC) is heterogeneous and aggressive, with high vascularity and frequent metastasis. We have already found natural flavonoid scutellarin (SC) suppressed spontaneous TNBC metastasis via normalizing tumor vasculature in vivo. In this study, supernatant from tumor necrosis factorα (TNFα)-treated human mammary microvascular endothelial cell (HMMEC) promoted cell migration and pseudopod formation in TNBC cells, but these phenomena were disappeared in SC-co-treated HMMEC. TNFα enhanced the expression of granulocyte colony-stimulating factor (G-CSF) and granulocyte–macrophage colony-stimulating factor (GM-CSF) in both HMMEC and human umbilical vein endothelial cell (HUVEC). G-CSF promoted TNBC migration and invasion in vitro, while G-CSF neutralization antibody and SC both inhibited TNBC metastasis in Balb/c mice. SC had no inhibition on the G-CSF-induced TNBC cell migration, but reduced G-CSF content in TNBC tumor tissues and TNFα-stimulated endothelial cells (ECs). SC restricted the nuclear translocation of runt-related transcription factor 1 (RUNX1) in TNBC tumor vessels and TNFα-treated ECs. RUNX1 was found to directly bind to the promoter of G-CSF in TNBC tumor vessels and regulated G-CSF expression. TNF receptor 2 (TNFR2) was crucial for regulating the TNFα-induced RUNX1 activation and G-CSF expression. Notably, SC hindered the interaction between TNFα and TNFR2 via binding to TNFR2. This work demonstrated that SC reduced TNBC metastasis by targeting TNFα/TNFR2-initiated RUNX1 activation and subsequent G-CSF production in TNBC-associated ECs.

Comparison of cytokine levels in prostatic secretion between the IIIa and IIIb subtypes of prostatitis.

Chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS), also known as National Institutes of Health (NIH) type III prostatitis, is a common disorder with an unclear etiology and no known curative treatments. Based on the presence or absence of leukocytes in expressed prostatic secretion (EPS), CP/CPPS is classified further into IIIa (inflammatory) and IIIb (noninflammatory) subtypes. However, the severity of symptoms is not entirely consistent with the white blood cell (WBC) count. Following the preliminary finding of a link between inflammatory cytokines and CP/CPPS, we performed this clinical study with the aim of identifying cytokines that are differentially expressed according to whether the prostatitis subtype is IIIa or IIIb. We found that granulocyte colony-stimulating factor (G-CSF), interleukin-18 (IL-18), and monocyte chemoattractant protein-1 (MCP-1) levels were significantly elevated and interferon-inducible protein-10 (IP-10) and platelet-derived growth factor-BB (PDGF-BB) levels were downregulated in the EPS of patients with type IIIa prostatitis. In a word, it is a meaningful study in which we investigate the levels of various cytokines in EPS according to whether prostatitis is the IIIa or IIIb subtype. The combination of G-CSF, IL-18, MCP-1, IP-10, and PDGF-BB expression levels could form a basis for classification, diagnosis, and therapeutic targets in clinical CP/CPPS.

Evaluation of the Effect of TNF-α on the Expression of G-CSF and M-CSF in the Monocytes of Multiple Sclerosis Patients

Background: Multiple sclerosis (MS) is an inflammatory autoimmune disease in which the myelin sheaths of nerve cells in the brain and spinal cord are damaged. This damage can disrupt the abilities of parts of the nervous system that are responsible for communication and cause many physical signs and symptoms. Granulocyte colony-stimulating factor (G-CSF) is a major extracellular regulator of hematopoiesis and the innate immune system. Macrophage colony-stimulating factor (M-CSF) is a glycoprotein that is effective in hematopoiesis and stimulates the production of numerous cytokines. It also plays a key role in stimulating the clearance of myelin by the brain’s innate immune cells, which is a prerequisite for proper remyelination and myelin repair. The aim of the study was to investigate the effect of overlapping and non-overlapping cytokines in MS. Methods: Overall, 20 MS patients referred to the MS clinic of Imam Reza hospital with mild, moderate, and severe stages were selected for investigation. The blood samples were taken from the patients, and magnetic-activated cell-sorting monocyte cells were separated from peripheral blood mononuclear cells (PBMCs). The purity of the monocyte cells was determined using flow cytometry and the CD14 antibody. The monocyte cells were then cultured and stimulated with tumor necrosis factor (TNF) over a time course of 0, 6, 12, 24, and 48 hours. After the end of stimulation, the cells were collected, and the expression level of G-CSF and M-CSF was evaluated by real-time polymerase chain reaction (RT-PCR). Results: The expression of G-CSF and M-CSF receptors decreased significantly over time as a time course of stimulation, with the lowest value 12 hours after stimulation. Comparing G-CSF and M-CSF, the decreasing ratio has been the same. The level of TNF-α function was evaluated as a time course. Conclusion: The results of RT-PCR showed that TNF-α significantly decreases the expression of innate immune system receptors such as G-CSF and M-CSF. This reduction was in surface receptors such as G-CSF and M-CSF.

Increased Tumor-Associated CD66b+ Myeloid-Derived Suppressor Cells in Waldenstrom Macroglobulinemia Inhibit T-Cell Immune Function

Waldenstrom macroglobulinemia (WM) is a discrete clinicopathologic entity resulting from the accumulation of lymphoplasmacytic lymphoma (LPL) cells in the bone marrow (BM) that secrete a monoclonal immunoglobulin (Ig)M protein. MYD88 (>90%) and CXCR4 (30-35%) mutations are the most commonly identified mutations in this disease. While much is known about the genomics of WM, far less is known about the role of the BM microenvironment in this disease. Emerging evidence has attested to the role of myeloid-derived suppressor cells (MDSCs) and their potent immunosuppressive activity, however, there is limited information about these cells in WM and monoclonal gammopathy of undetermined significance (MGUS). This study was therefore designed to explore the phenotype of MDSCs and their interaction with other cells in the BM in WM. A multiparametric approach was combined with single-cell genomic and proteomic tools to better understand MDSC immunology and the antitumor immune response in WM. In the present study, using flow cytometry, we first analyzed the number of MDSCs (by gating on Lin- HLA-DR- CD11b+ CD33+) in normal BM (NBM; n=11) and WM symptomatic /MGUS (n=18) and found an increased number of MDSCs in WM specimens compared to controls (p=0.005). We also found that most MDSCs in WM specimens had a granulocytic phenotype (Lin- HLA-DR- CD11b+ CD33+ CD15+). A detailed phenotypic study using CyTOF (mass spectrometry) was performed on MDSCs from NBM (n=4) and WM (n=8). The CyTOF analysis identified 3 subtypes of MDSCs including CD66b+ MDSCs with a granulocytic phenotype (G-MDSCs) and confirmed the expansion of CD66b+ MDSCs in WM symptomatic patients when compared to smoldering WM and normal controls. Our previous studies have shown that granulocyte colony-stimulating factor (G-CSF) expression is increased in the bone marrow of WM (Jalali et al., 2018). Therefore, to determine whether G-CSF accounted for the expansion of CD66b+ G-MDSCs, we treated WM-BM MDSCs with G-CSF and found that the CD66b+ G-MDSC population substantially increased (p=0.0017). Further, to establish a better understanding of the transcriptome profile and a unique phenotype of CD66b+ G-MDSCs, we performed Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) on WM (n=3) and NBM (n=2) specimens. A significantly high expression of NLRP12, IGFBP7, CXCR4, IL2, CD55, SOX4, and FoxP1 genes was observed in CD66b+ MDSCs as compared to other MDSC populations. This gene expression signature suggested upregulation of inflammatory pathways potentially affecting immune function. Therefore, to evaluate the effect of CD66b+ G-MDSCs on the immune function of other cells in the BM, CD66b+ and CD66b- MDSCs were co-cultured with activated autologous T-cells. We found that while all populations of MDSCs suppressed T-cell activation, CD66b+ G-MDSCs had the greatest suppressive effect. In conclusion, this study has shown that CD66b+ G-MDSCs are expanded in WM patients, exhibit a unique inflammatory transcriptome, and substantially inhibit T-cell activation and proliferation. Furthermore, CD66b+ G-MDSCs are expanded due to increased G-CSF in the BM microenvironment in WM. Clinical strategies to inhibit the expansion of CD66b+ G-MDSCs may therefore enhance the efficacy of immunological therapies in patients with WM.

Mechanism of human airway epithelial cell injury induced by Candida albicans infection

Objective: To investigate the immune mechanism of human airway epithelial cell injury induced by invasion of Candida albicans with different biofilm formation abilities. Methods: Twenty-five strains of Candida albicans isolated and cultured in General Hospital of Ningxia Medical University from June to December 2019 were selected, and quality control strain SC5314 was used as the standard strain. An in vitro model of Candida albicans biofilm was established, and the biofilm formation ability of different Candida albicans was detected by crystal violet staining and enzyme plate method. The absorbance value at 570 nm (A570) was determined by enzyme plate method. A570≥0.5, 0.25<A570<0.5 and A570≤0.25 indicated strong biofilm Candida albicans form (SBF), moderate biofilm Candida albicans form (DRF) and weak biofilm Candida albicans form (WBF), respectively. The gas-liquid phase culture model of human airway epithelial cells was isolated and established in vitro and divided into five groups, including blank control group (n=20), standard strain group (n=20), strong biofilm group (n=19), weak biofilm group (n=17) and fluconazole-resistant group (n=18).The morphology of the epithelial cells was observed by scanning electron microscope (SEM), and the expression of marker protein in the model was detected by immunofluorescence in vitro. The level of lactate dehydrogenase (LDH) in cells was detected by microplate method, and the secretion of β-defensin (hBD2), granulocyte macrophage colony stimulating factor(GM-CSF) and granulocyte colony-stimulating factor (G-CSF) was detected by enzyme-linked immunosorbent assay (ELISA). Results: The strong biofilm strains grew with interlacing mycelia, and very few yeast cells could be seen wrapped in them.SEM observed that the mycelia of epithelial cells in gas-liquid phase culture could actively invade epithelial cells, and the expression of acetylated tubulin and keratin in cilia were significantly reduced, while the expression of Ki67 was down-regulated.The LDH levels in the blank control group, standard strain group, strong biofilm group, weak biofilm group, and fluconazole-resistant group were (12.21±5.68), (46.35±6.35), (18.69±4.38), (12.56±3.69), and (13.48±4.28) U/L, respectively, with statistically significant differences (P<0.001). Compared with standard strain group, LDH level in strong biofilm group, weak biofilm group and fluconazole resistant group were significantly decreased (all P<0.01). The hBD2 levels of the five groups were (26.14±0.77), (56.18±0.83), (30.66±2.59), (29.22±0.48), (28.28±1.56) ng/L, respectively, with statistically significant differences(P<0.001). Compared with the blank control group, SC5314-treated epithelial cells induced an increase of intracellular hBD2 expression (P<0.001). The differences in the expression of GM-CSF and G-CSF between different groups were not statistically significant(all P>0.05). Conclusion: Strong biofilm Candida albican can inhibit cell proliferation, disrupt the integrity of epithelial cells and induce cell damage by down-regulating the expression of cell proliferation-related protein.

Correlation between Serum G-CSF Level and Immune Function in Children with Aplastic Anemia

To observe the occurrence of immune dysfunction in children with aplastic anemia (AA) and the factors that may lead to immune dysfunction, analyze the relationship between the expression of granulocyte colony stimulating factor (G-CSF) and immune dysfunction. A total of 34 children with AA treated in our hospital from December 2016 to September 2018 were selected. All the children received immunosuppressive therapy (IST) for 6 months. According to whether the children had immune dysfunction after 6 months of treatment, they were divided into occurrence group and non occurrence group. General information and laboratory indices were compared between the two groups, and serum G-CSF level was tested, the relationship between serum G-CSF level and immune dysfunction in AA children after treatment with IST was observed and analyzed. After treatment with IST for 6 months, 12 cases developed immune dysfunction (35.29%). Serum interferon (IFN)-γ level of the occurrence group was higher but G-CSF level was lower than those of the non occurrence group (P<0.05), while the difference of other baseline data was not statistically significant (P>0.05). Multiple regression analysis showed that overexpression of serum IFN-γ and low expression of G-CSF were both the influencing factors of immune dysfunction in AA children after IST treatment (OR>1, P<0.05). ROC curve was drawn, and the result showed that the area under the curve (AUC) of serum G-CSF level predicted the risk of immune dysfunction after IST was 0.843>0.80, when the index cut-off value was set at 6.614 pg/ml, the predictive value was ideal. AA children have a higher risk of immune dysfunction after IST, which may be related to the low expression of serum G-CSF. The detection of serum G-CSF expression can be considered to predict the risk of immune dysfunction in AA children after IST, so as to guide early clinical intervention.

Injection of Cerium Oxide Nanoparticles to Treat Spinal Cord Injury in Rats.

This study investigated the effects of local injection of cerium oxide nanoparticles (CeONPs) in a rat spinal cord injury (SCI) model. Thirty-six adult male Wistar rats were divided into 4 groups: controls (healthy animals), sham (laminectomy), SCI (laminectomy+SCI induction), and treatment (laminectomy+SCI induction+intrathecal injection of CeONPs immediately after injury). SCI was induced using an aneurysm clip at the T12-T13 vertebral region. Motor performance and pain threshold tests were performed weekly; H&E staining and measurement of cavity sizes were performed 6 weeks after injury. The expression of granulocyte colony-stimulating factor (GCSF), P44/42 MAPK, P-P44/42 MAPK, Tau, myelin-associated glycoprotein(MAG) was evaluated after 6 weeks by Western blot. The Basso, Beattie, and Bresnahan locomotor scoring scales improved in animals receiving CeONPs compared with SCI animals. The cavity sizes were less in the treatment group. GCSF expression was similar in the animals receiving CeONPs compared with the SCI group but the expression of ERK1/ERK2 and phospho-ERK was lower than in the SCI group. Expression levels of Tau and MAG were significantly increased in treated animals compared to the SCI group. These data indicate that the use of CeONPs may improve motor functional recovery in SCI.

A NOVEL ORALLY ACTIVE METABOLITE REVERSES CROHN’S DISEASE-ASSOCIATED INTESTINAL FIBROSIS

Abstract BACKGROUND Intestinal fibrosis is a debilitating complication of Crohn’s disease (CD) patients. Our groups and others discovered that intestinal metabolites are associated with intestinal fibrosis. We selected the most relevant metabolites and determined their anti-fibrogenic efficacies in three mouse models of intestinal fibrosis. METHODS We compared the intestinal metabolite profiles between non-IBD, stricturing CD, and non-stricturing CD patients in inflammatory bowel disease (IBD) multi-omics database to identify CD stricture-related metabolites. We compared the fecal metabolite profiles between trinitrobenzene sulfonic acid (TNBS)-treated mice with and without anti-fibrogenic drug CSA13 treatment to identify mouse intestinal fibrosis-related metabolites. RESULTS Compared to non-stricturing CD patients and non-IBD patients, stricturing CD patients had reduced fecal levels of hydrocinnamate (H), inosine (I), taurine (T), tauroursodeoxycholate (TUDCA), and sphingosine (S). Oral CSA13 increased fecal levels of inosine (I) and sphingosine (S) in the TNBS-treated mice. Overall disease activity (ODA) is a comprehensive assessment of histology and fibrosis scores and intestinal mRNA expression of fibrosis and inflammation markers. Sphingosine (10mg/kg/day for 7-14 days via oral gavages) showed the best anti-fibrogenic effects against spontaneous ileal fibrosis in 42-week-old SAMP1/YitFc mice (11% ODA), cecal fibrosis in Salmonella-infected mice (10% ODA), and colonic fibrosis in TNBS-treated mice (9% ODA). The other four metabolites were ineffective in reversing intestinal fibrosis. Sphingosine (10 microM), but not other metabolites, directly inhibited collagen expression in human colonic CCD-18Co fibroblasts and primary human intestinal fibroblasts from CD patients (CD-HIF). RNA-seq and proteomics identified that low MAPK3 and high ZEB1 mRNA expression in intestines are associated with intestinal fibrosis in CD patients. Sphingosine activated ERK1/2 phosphorylation and inhibited TGF-b1-induced ZEB1 expression in CCD-18Co fibroblasts. The sphingosine-mediated inhibition of collagen synthesis was reversed by ERK1/2 inhibitor PD98059 and ZEB1 overexpression. The anti-fibrogenic effect of sphingosine in the TNBS model was reversed by Zeb1 overexpression. Stricturing CD patients have higher circulating granulocyte-colony stimulating factor (G-CSF) levels than non-stricturing CD patients. Csf3 promoted collagen expression in CD-HIF, while Csf3 inhibition diminished cecal fibrosis in Salmonella-infected mice. Sphingosine diminished G-CSF secretion in primary human peripheral blood mononuclear cells (PBMCs). Sphingosine also inhibited cecal Csf3 mRNA expression and reduced circulating G-CSF levels in Salmonella-infected mice. CONCLUSIONS Sphingosine exerts the most potent anti-fibrogenic effects by modulating ERK1/2, ZEB1, and G-CSF expression.

Effect of Extended Window Thrombectomy on Acute Cerebral Infarction and Granulocyte Colony-Stimulating Factor Expression and Postoperative Brain Function

Effect of Extended Window Thrombectomy on Acute Cerebral Infarction and Granulocyte Colony-Stimulating Factor Expression and Postoperative Brain Function

Astaxanthin ameliorates damage to the cerebral cortex, hippocampus and cerebellar cortex caused by methotrexate

ABSTRACT We investigated the ameliorating effects of astaxanthin (AXA) on methotrexate (MTX) induced damage to the cerebral cortex, hippocampus, cerebellar cortex and blood. We used 24 female Wistar albino rats divided into three groups of eight as follows: sham/control group, single dose of saline intraperitoneally (i.p.) and 7 days orally; MTX group, single dose of 20 mg/kg MTX (i.p.); MTX + AXA group, single dose of 20 mg/kg MTX i.p.+ 100 mg/kg AXA orally for 7 days. For all groups we measured total oxidant status (TOS) and total antioxidant status (TAS) in the cerebral cortex, hippocampus and blood. Histological sections of cerebral cortex, hippocampus and cerebellar cortex were inspected microscopically. Caspase-3 (cas-3), granulocyte colony-stimulating factor (GCSF), growth related oncogene (GRO), inducible nitric oxide synthase (iNOS) and myelin basic protein (MBP) were estimated immunohistochemically in the cerebral cortex, hippocampus and cerebellar cortex. In the MTX group, TAS was decreased significantly in the cerebral cortex, hippocampus and blood, while TOS was significantly increased. AXA significantly ameliorated oxidative stress parameters in the cerebral cortex and hippocampus. Histopathological examination revealed degeneration, edema and hyperemia in the cerebral cortex, hippocampus and cerebellar cortex in the MTX group. AXA treatment ameliorated histopathological changes. MTX decreased MBP expression in cerebral cortex. Although MBP expression was decreased in the cerebral cortex, hippocampus and cerebellar cortex stimulated with MTX, the expressions of cas-3, GCSF, GRO and iNOS were significantly increased. AXA ameliorated the expression of cas-3, GCSF, GRO, iNOS and MBP. AXA exhibits anti-inflammatory, antioxidant and anti-apoptotic effects on MTX induced toxicity in the cerebral cortex, hippocampus and cerebellar cortex by increasing MBP expression, regulating inflammatory cytokine release and reducing oxidative stress.

Administration of Δ9-Tetrahydrocannabinol Following Controlled Cortical Impact Restores Hippocampal-Dependent Working Memory and Locomotor Function.

Hypothesis: Administration of the phytocannabinoid Δ9-tetrahydrocannabinol (Δ9-THC) will enhance brain repair and improve short-term spatial working memory in mice following controlled cortical impact (CCI) by upregulating granulocyte colony-stimulating factor (G-CSF) and other neurotrophic factors (brain-derived neurotrophic factor [BDNF], glial-derived neurotrophic factor [GDNF]) in hippocampus (HP), cerebral cortex, and striatum. Materials and Methods: C57BL/6J mice underwent CCI and were treated for 3 days with Δ9-THC 3 mg/kg intraperitoneally (i.p.). Short-term working memory was determined using the spontaneous alternations test during exploratory behavior in a Y-maze. Locomotor function was measured as latency to fall from a rotating drum (rotometry). These behaviors were recorded at baseline and 3, 7, and 14 days after CCI. Groups of mice were euthanized at 7 and 14 days. Extent of microgliosis, astrocytosis, and G-CSF, BDNF, and GDNF expression were measured at 7 and 14 days in cerebral cortex, striatum, and HP on the side of the trauma. Levels of the most abundant endocannabinoid (2-arachidonoyl-glycerol [2-AG]) was also measured at these times. Results: Δ9-THC-treated mice exhibited marked improvement in performance on the Y-maze indicating that treatment with the phytocannabinoid could reverse the deficit in working memory caused by the CCI. Δ9-THC-treated mice ran on the rotarod longer than vehicle-treated mice and recovered to normal rotarod performance levels at 2 weeks. Δ9-THC-treated mice, compared with vehicle-treated animals, exhibited significant upregulation of G-CSF as well as BDNF and GDNF in the cerebral cortex, striatum, and HP. Levels of 2-AG were also increased in the Δ9-THC-treated mice. Conclusion: Administration of the phytocannabinoid Δ9-THC promotes significant functional recovery from traumatic brain injury (TBI) in the realms of working memory and locomotor function. This beneficial effect is associated with upregulation of brain 2-AG, G-CSF, BDNF, and GDNF. The latter three neurotrophic factors have been previously shown to mediate brain self-repair following TBI and stroke.

Production of PEGylated GCSF from Non-classical Inclusion Bodies Expressed in Escherichia coli

The recombinant human granulocyte colony stimulating factor conjugated with polyethylene glycol (PEGylated GCSF) has currently been used as an efficient drug for the treatment of neutropenia caused by chemotherapy due to its long circulating half-life. Previous studies showed that Granulocyte Colony Stimulating Factor (GCSF) could be expressed as non-classical Inclusion Bodies (ncIBs), which contained likely correctly folded GCSF inside at low temperature. Therefore, in this study, a simple process was developed to produce PEGylated GCSF from ncIBs. BL21 (DE3)/pET-GCSF cells were cultured in the LiFlus GX 1.5 L bioreactor and the expression of GCSF was induced by adding 0.5 mM IPTG. After 24 hr of fermentation, cells were collected, resuspended, and disrupted. The insoluble fraction was obtained from cell lysates and dissolved in 0.1% N-lauroylsarcosine solution. The presence and structure of dissolved GCSF were verified using SDS-PAGE, Native-PAGE, and RP-HPLC analyses. The dissolved GCSF was directly used for the conjugation with 5 kDa PEG. The PEGylated GCSF was purified using two purification steps, including anion exchange chromatography and gel filtration chromatography. PEGylated GCSF was obtained with high purity (∼97%) and was finally demonstrated as a form containing one GCSF molecule and one 5 kDa PEG molecule (monoPEG-GCSF). These results clearly indicate that the process developed in this study might be a potential and practical approach to produce PEGylated GCSF from ncIBs expressed in Escherichia coli (E. coli).

Effect of intrauterine granulocyte-colony stimulating factor administration on in vitro fertilization outcome in women with moderate-to-severe endometriosis: An RCT.

BackgroundNearly 25-50% of infertile women have endometriosis. There are reports of disorders in the expression of granulocyte colony-stimulating factor (G-CSF) receptors in women with endometriosis.ObjectiveTo examine the effect of intrauterine administration of G-CSF in in vitro fertilization (IVF) cycles on the fertility rate of infertile women with moderate-to-severe endometriosis.Materials and Methods This clinical trial was conducted on 66 infertile women with moderate-to-severe endometriosis, undergoing IVF and intracytoplasmic sperm injection (ICSI). The participants were allocated into two groups via simple randomization: the G-CSF (n = 27) and control (n = 39) groups. In the G-CSF intervention group, on the oocyte pick-up day, immediately after an ovarian puncture, 300 μg of G-CSF was injected through a transcervical catheter under abdominal ultrasound guide to visualize flushing into the uterine cavity. Women in the control group received no intervention. The two groups were evaluated for clinical pregnancy.ResultsNo significant difference was noted in the demographic characteristics of the two groups. The rate of clinical pregnancy was 28.2% in the control group and 25.9% in the G-CSF group, indicating no significant difference (p = 0.83).ConclusionThe results showed that the intrauterine injection of G-CSF had no effects on pregnancy in women with stage-3/4 endometriosis undergoing IVF.

Granulocyte colony-stimulating factor (G-CSF) enhances cocaine effects in the nucleus accumbens via a dopamine release-based mechanism.

Cocaine use disorder is associated with alterations in immune function including altered expression of multiple peripheral cytokines in humans-several of which correlate with drug use. Individuals suffering from cocaine use disorder show altered immune system responses to drug-associated cues, highlighting the interaction between the brain and immune system as a critical factor in the development and expression of cocaine use disorder. We have previously demonstrated in animal models that cocaine use upregulates the expression of granulocyte colony-stimulating factor (G-CSF)-a pleiotropic cytokine-in the serum and the nucleus accumbens (NAc). G-CSF signaling has been causally linked to behavioral responses to cocaine across multiple behavioral domains. The goal of this study was to define whether increases in G-CSF alter the pharmacodynamic effects of cocaine on the dopamine system and whether this occurs via direct mechanisms within local NAc microcircuits. We find that systemic G-CSF injection increases cocaine effects on dopamine terminals. The enhanced dopamine levels in the presence of cocaine occur through a release-based mechanism, rather than through effects on the dopamine transporter-as uptake rates were unchanged following G-CSF treatment. Critically, this effect could be recapitulated by acute bath application of G-CSF to dopamine terminals, an effect that was occluded by prior G-CSF treatment, suggesting a similar mechanistic basis for direct and systemic exposures. This work highlights the critical interaction between the immune system and psychostimulant effects that can alter drug responses and may play a role in vulnerability to cocaine use disorder.

Endogenous Expression of G-CSF in Rat Dorsal Root Ganglion Neurons after Nerve Injury.

Granulocyte colony-stimulating factor (G-CSF) has been reported to modulate pain function following nerve injury. However, the expression of endogenous G-CSF in the dorsal root ganglion (DRG) and the response to nerve injury remain unclear. In the present study, we demonstrated that G-CSF and G-CSFR are mainly expressed in both small- and medium-diameter DRG neurons in rats and are responsible for transmitting pain responses. G-CSF and G-CSFR were co-expressed in certain nociceptive DRG neurons. In addition, G-CSF was expressed in satellite glial cells around large-diameter DRG neurons. After sciatic nerve injury, the number of G-CSF-positive DRG neurons was increased in both the ipsilateral and contralateral lesion sites in rats. However, G-CSF expression in satellite glial cells was not affected by nerve injury. To clarify the role of G-CSF in pain, exogenous G-CSF was administered to a rat model of neuropathic pain induced by partial sciatic nerve transaction (PST). Our results indicate that treatment with G-CSF did not attenuate but exacerbated neuropathic pain. In summary, G-CSF may directly activate sensory neurons and contribute to nociceptive signaling.

Plasma concentrations of granulocyte colony-stimulating factor (G-CSF) in patients with substance use disorders and comorbid major depressive disorder

Granulocyte colony–stimulating factor (G-CSF) has raised much interest because of its role in cocaine addiction in preclinical models. We explored the plasma concentrations of G-CSF in patients diagnosed with substance use disorder (SUD) and highly comorbid psychiatric disorders. In particular, we investigated the association between G-CSF concentrations and comorbid major depressive disorder (MDD) in patients with cocaine and alcohol use disorders (CUD and AUD, respectively). Additionally, patients with MDD but not SUD were included in the study. Three hundred and eleven participants were enrolled in this exploratory study: 136 control subjects, 125 patients with SUD (SUD group) from outpatient treatment programs for cocaine (N = 60, cocaine subgroup) and alcohol (N = 65, alcohol subgroup), and 50 patients with MDD but not SUD (MDD group) from primary-care settings. Participants were assessed based on DSM-IV-TR criteria, and a blood sample was collected to examine the plasma concentrations of G-CSF. G-CSF concentrations were negatively correlated with age in the entire sample (r = − 0.233, p < 0.001) but not in the patients with MDD. G-CSF concentrations were lower in patients with SUD than in controls (p < 0.05), specifically in the cocaine subgroup (p < 0.05). Patients with SUD and comorbid MDD had lower G-CSF concentrations than patients with SUD but not comorbid MDD or controls (p < 0.05). In contrast, patients with MDD but not SUD showed no differences compared with their controls. The negative association between G-CSF concentrations and age in the sample was not observed in patients with MDD. G-CSF concentrations were decreased in patients with SUD and comorbid MDD but not in patients with MDD. Therefore, G-CSF may be useful to improve the stratification of patients with dual diagnosis seeking treatment. Further investigation is needed to explore the impact of sex and type of drug on the expression of G-CSF.

Granulocyte Colony Stimulating Factor Expression in Breast Cancer and Its Association with Carbonic Anhydrase IX and Immune Checkpoints.

Simple SummaryPreclinical studies suggest that interactions between granulocyte colony-stimulating factor (G-CSF) and hypoxia-induced carbonic anhydrase IX regulate the trafficking and function of immune cells in the tumour microenvironment. We investigated the clinical significance of this crosstalk by analyzing the protein expression of G-CSF and macrophage markers by immunohistochemistry on a well-characterized tissue microarray series of invasive breast cancers. We report that high expression of G-CSF on breast carcinoma cells is linked with significantly improved survival in an important group of breast cancers that do not respond to hormonal therapy. These tumours were infiltrated by immune cells expressing biomarkers that can be targeted with immune checkpoint inhibitor drugs. In contrast, carbonic anhydrase IX expression was associated with unfavourable outcomes.Purpose: Granulocyte colony-stimulating factor (G-CSF) and hypoxia modulate the tumour immune microenvironment. In model systems, hypoxia-induced carbonic anhydrase IX (CAIX) has been associated with G-CSF and immune responses, including M2 polarization of macrophages. We investigated whether these associations exist in human breast cancer specimens, their relation to breast cancer subtypes, and clinical outcome. Methods: Using validated protocols and prespecified scoring methodology, G-CSF expression on carcinoma cells and CD163 expression on tumour-associated macrophages were assayed by immunohistochemistry and applied to a tissue microarray series of 2960 primary excision specimens linked to clinicopathologic, biomarker, and outcome data. Results: G-CSFhigh expression showed a significant positive association with ER negativity, HER2 positivity, presence of CD163+ M2 macrophages, and CAIX expression. In univariate analysis, G-CSFhigh phenotype was associated with improved survival in non-luminal cases, although the CAIX+ subset had a significantly adverse prognosis. A significant positive association was observed between immune checkpoint biomarkers on tumour-infiltrating lymphocytes and both G-CSF- and CAIX-expressing carcinoma cells. Immune checkpoint biomarkers correlated significantly with favourable prognosis in G-CSFhigh/non-luminal cases independent of standard clinicopathological features. Conclusions: The prognostic associations linking G-CSF to immune biomarkers and CAIX strongly support their immunomodulatory roles in the tumour microenvironment.

Recovery from Traumatic Brain Injury Following Treatment with Δ9-Tetrahydrocannabinol Is Associated with Increased Expression of Granulocyte-Colony Stimulating Factor and Other Neurotrophic Factors.

Introduction: The hematopoietic cytokine granulocyte-colony stimulating factor (G-CSF) is well known to stimulate proliferation of blood stem/progenitor cells of the leukocyte lineage, but is also recognized as a neurotrophic factor involved in brain self-repair processes. G-CSF administration has been shown to promote recovery from experimental models of traumatic brain injury (TBI) and to modulate components of the endocannabinoid system (eCS). Conversely, Δ9-tetrahydrocannabinol (Δ9THC) treatment of normal mice has been shown to increase blood levels of G-CSF in the periphery. Hypothesis: Administration of the phytocannabinoid Δ9THC will enhance brain repair following controlled cortical impact (CCI) by upregulating G-CSF and other neurotrophic factors (brain-derived neurotrophic factor [BDNF] and glial-derived neurotrophic factor [GDNF]) in brain regions. Materials and Methods: C57BL/6J mice underwent CCI and were treated for 3 days with THC 3 mg/kg intraperitoneally. Motor function on a rotarod was recorded at baseline and 3, 7, and 14 days after CCI. Groups of mice were euthanized at 7 and 14 days. G-CSF, BDNF, and GDNF expression were measured at 7 and 14 days in cerebral cortex, striatum, and hippocampus on the side of the trauma. Results: Δ9THC-treated mice ran on the rotarod longer than vehicle-treated mice and recovered to normal rotarod performance levels at 2 weeks. These mice, compared to vehicle-treated animals, exhibited significant upregulation of G-CSF as well as BDNF and GDNF in cerebral cortex, striatum, and hippocampus. Conclusion: Administration of the phytocannabinoid Δ9THC promotes significant recovery from TBI and is associated with upregulation of brain G-CSF, BDNF, and GDNF, neurotrophic factors previously shown to mediate brain self-repair following TBI and stroke.