spi1  (MedChemExpress)

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: SPI1 is associated with the hypertranscriptional phenotype in HNSCC. a TF activity inference identifying SPI1 as the top candidate. Scatter plot of TF activity scores (weighted t statistic) versus significance. b Kaplan‒Meier analysis showing that patients with high SPI1 activity exhibit significantly worse overall survival. P values were calculated using the log-rank test. c Forest plot derived from an expanded multivariate Cox regression model. High SPI1 activity remained an independent predictor of poor prognosis after we adjusted for clinicopathological factors (age, sex, stage, smoking/alcohol history, tumor subsite, and HPV status) and tumor microenvironment characteristics (stromal, immune, and ESTIMATE scores). d UMAP plots of single cells illustrating graded SPI1 activity (left) and mRNA hypertranscription status (right). e Heatmap of representative differential genes across low-, medium-, and high-SPI1-activity groups, showing distinct gene expression modules associated with SPI1 activation. f Scatter plot showing a positive correlation between SPI1 activity and normalized total UMI counts ( R = 0.42, p < 0.0001). g Violin plots of the SPI1 activity across tertile-defined low, medium, and high mRNA hypertranscription groups ( n = 6266 cells per group; p < 0.0001). h Heatmap of the top 20 hallmark pathways enriched in cells with high SPI1 activity. i Rank-ordered distribution of TF activity t statistics from an independent single-cell HNSCC dataset ( GSE234933 )

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: Activity Assay, Derivative Assay, Gene Expression, Activation Assay, Single Cell

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: SPI1 regulates proliferation, survival, migration, and invasion in HNSCC cells in vitro. a Representative Western blot analysis showing the SPI1 protein levels in the normal human keratinocyte cell line HaCaT and the head and neck squamous cell carcinoma cell lines SAS, FaDu, CAL27, and CAL33. β-actin was used as a loading control. b Quantification of SPI1 protein expression normalized to that of β-actin and expressed relative to that in HaCaT cells. Data are presented as the mean ± SD from independent experiments. Statistical significance was assessed using one-way ANOVA; P < 0.05 is indicated by an asterisk. c Immunofluorescence staining showing the nuclear localization of SPI1 in CAL27 and CAL33 cells; the right panels show line-scan intensity profiles. Scale bar, 20 μm. d Western blot confirmation of the SPI1 knockdown efficiency in CAL27 and CAL33 cells using two independent shRNAs. e Cell viability was measured by a CCK-8 assay at 72 h posttransduction in control and SPI1-knockdown cells ( n = 3 independent replicates). Quantification ( f ) and representative images ( g ) of colony formation assays showing the clonogenic potential of CAL27 and CAL33 cells after SPI1 knockdown ( n = 3 independent replicates). h , i EdU incorporation assay for cell proliferation. h Representative immunofluorescence images showing EdU incorporation in CAL27 and CAL33 cells transduced with shNC or SPI1-targeting shRNAs (scale bar = 50 μm); i quantification of EdU-positive cells ( n = 3 independent replicates). j Representative images from Transwell migration and invasion assays in CAL27 and CAL33 cells with or without SPI1 knockdown (scale bar = 200 μm). Quantification of migrated and invaded CAL27 ( k ) and CAL33 ( l ) cells ( n = 3 independent replicates). m Western blot showing SPI1 overexpression in SAS and FaDu cells. n CCK-8 assay showing enhanced proliferation in SPI1-overexpressing SAS and FaDu cells ( n = 3 independent replicates). Colony formation assay results showing increased clonogenic capacity following SPI1 overexpression ( o ), with corresponding representative colony images shown in ( p ) ( n = 3 independent replicates). q Representative transwell migration and invasion assay images of SPI1-overexpressing SAS and FaDu cells (scale bar = 200 μm). Quantification of migrated and invaded SAS ( r ) and FaDu ( s ) cells following SPI1 overexpression ( n = 3 independent replicates). In ( d – l ), the shNC group indicates cells transduced with a lentiviral vector carrying nontargeting shRNA. In ( m – s ), the NC group indicates cells transduced with an empty lentiviral vector as the control for SPI1 overexpression. The data are presented as the mean ± SD. Statistical significance was assessed using one-way ANOVA followed by Tukey’s post hoc test for multiple group comparisons or two-tailed unpaired Student’s t test for two-group comparisons. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: Migration, In Vitro, Western Blot, Control, Expressing, Immunofluorescence, Staining, Knockdown, CCK-8 Assay, Transduction, Over Expression, Colony Assay, Invasion Assay, Plasmid Preparation, shRNA, Two Tailed Test

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: a Representative images of xenograft tumors derived from CAL33 cells transduced with nontargeting shRNA (shNC) or two independent SPI1-targeting shRNAs ( n = 6 biologically independent mice per group). b Growth curves of CAL33 xenografts with or without SPI1 knockdown ( n = 6 biologically independent mice per group). c Tumor weights at the endpoint in CAL33 xenografts ( n = 6 biologically independent mice per group). d Quantification of the percentage of SPI1⁺ cells among EpCAM⁺ tumor cells in CAL33 xenografts on the basis of immunofluorescence analysis ( n = 6 independent tumors). e Quantification of the percentage of Ki-67⁺ cells among EpCAM⁺ tumor cells in CAL33 xenografts on the basis of immunofluorescence analysis ( n = 6 independent tumors). f Quantification of apoptotic cell percentages in CAL33 xenografts by TUNEL staining ( n = 6 independent tumors). g Representative histological and immunofluorescence images of CAL33 xenograft tumors. Hematoxylin and eosin (H&E) staining is shown in the top row. Immunofluorescence images show costaining of SPI1 (magenta) and EpCAM (yellow), Ki-67 (green) and EpCAM (yellow) with DAPI (blue), and TUNEL-positive apoptotic cells (green) with DAPI (blue). SPI1/EPCAM and Ki-67/EPCAM images were derived from the same multiplex-stained section and identical field, with channels displayed separately for visualization. Scale bar, 100 μm. h Representative images of xenograft tumors derived from SAS cells with SPI1 overexpression or control cells ( n = 6 biologically independent mice per group). i Tumor weights at the endpoint in SAS xenografts ( n = 6 biologically independent mice per group). j Growth curves of SAS xenografts with or without SPI1 overexpression ( n = 6 biologically independent mice per group). k Representative images of xenograft tumors derived from FaDu cells with SPI1 overexpression or control cells ( n = 6 biologically independent mice per group). l Tumor weights at the endpoint in the FaDu xenografts ( n = 6 biologically independent mice per group). m Growth curves of FaDu xenografts with or without SPI1 overexpression ( n = 6 biologically independent mice per group). Quantification of the percentage of SPI1⁺ cells among EpCAM⁺ tumor cells in SAS ( n ) and FaDu ( o ) xenografts on the basis of immunofluorescence analysis ( n = 6 independent tumors). Representative immunofluorescence images showing the costaining of SPI1 (magenta) with EpCAM (yellow) and Ki-67 (green) with EpCAM (yellow) in SAS ( p ) and FaDu ( q ) xenografts; DAPI (blue). SPI1/EPCAM and Ki-67/EPCAM images were derived from the same multiplex-stained section and identical field, with channels displayed separately for visualization. Scale bar, 100 μm. Quantification of the percentage of Ki-67⁺ cells among EpCAM⁺ tumor cells in SAS ( r ) and FaDu ( s ) xenografts on the basis of immunofluorescence analysis ( n = 6 independent tumors). In ( a – g ), the shNC group indicates cells transduced with a lentiviral vector carrying nontargeting shRNA. In ( h – s ), the NC group indicates cells transduced with an empty lentiviral vector as the control for SPI1 overexpression. In ( b , j , m ), the data are presented as the mean ± SEM. Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test for multiple group comparisons or two-tailed unpaired Student’s t test for two-group comparisons. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: Derivative Assay, Transduction, shRNA, Knockdown, Immunofluorescence, TUNEL Assay, Staining, Multiplex Assay, Over Expression, Control, Plasmid Preparation, Two Tailed Test

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: Transcriptional reprogramming and phenotypic alterations driven by SPI1 perturbation in HNSCC cells. a Violin plot showing inferred SPI1 TF activity in CAL33 cells transduced with shNC or shSPI1 #1. b Heatmap displaying the top differentially active TFs between shNC and shSPI1 #1 inferred by decoupleR. The red arrow highlights the specific downregulation of SPI1 activity. c Violin plot confirming significantly increased SPI1 TF activity in SAS cells upon SPI1 overexpression (OE) compared with that in the negative control (NC). d Heatmap showing the expression profiles of key EMT- and invasion-related markers (e.g., MMP9, VIM, and CDH2 ) in SAS-NC versus SAS-OE cells. The side bar indicates the log2-fold change (log2FC). e Gene set enrichment analysis (GSEA) of cancer hallmark pathways in SPI1-overexpressing cells. Red circles indicate upregulated pathways, while blue circles indicate downregulated pathways. Dot size represents statistical significance (−log10 FDR). *, P < 0.05

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: Activity Assay, Transduction, Over Expression, Negative Control, Expressing

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: SPI1 promotes mRNA hypertranscription in HNSCC through a time-dependent mechanism. Representative images ( a ) and quantification ( b ) of EU incorporation in CAL27 and CAL33 cells after stable knockdown of SPI1 using two independent shRNAs ( n = 3 independent replicates). Red: EU-labeled nascent RNA; blue: DAPI-stained nuclei. Scale bar, 100 μm. c Relative per-cell mRNA yield in CAL27 and CAL33 cells, determined after mRNA purification and normalized by cell number ( n = 3 independent replicates). Representative EU-stained images ( d ) and quantification ( e ) of SAS and FaDu cells with or without SPI1 overexpression ( n = 3 independent replicates). f Relative per-cell mRNA yield in SPI1-overexpressing SAS and FaDu cells, based on poly(A)-selected RNA quantification ( n = 3 independent replicates). g Time-course RT‒qPCR analysis of SPI1 expression in Dox-inducible Tet-ON SAS and FaDu cells following doxycycline treatment ( n = 3 independent replicates). The data were normalized to that of GAPDH and expressed relative to 0 h. h Immunoblot analysis of the SPI1 and c-MYC protein levels. Representative images ( i ) and quantification of the fluorescence intensity ( j ) of cells with incorporated EU at the indicated time points after Dox induction ( n = 3 independent replicates). The data are presented as the mean ± SD. Scale bars, 50 μm. In ( a – c ), the shNC group indicates cells transduced with a lentiviral vector carrying nontargeting shRNA. In ( d – f ), the NC group indicates cells transduced with an empty lentiviral vector as the control for SPI1 overexpression. Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test or an unpaired two-tailed Student’s t test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: Knockdown, Labeling, Staining, Purification, Over Expression, Expressing, Western Blot, Fluorescence, Transduction, Plasmid Preparation, shRNA, Control, Two Tailed Test

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: Integrative transcriptomic and epigenomic analyses of SPI1 function in HNSCC. a Heatmaps and averaged ChIP-seq signal profiles showing SPI1 occupancy around transcription start sites (TSSs) in SPI1-overexpressing SAS cells, with SPI1 ChIP-seq Rep1 and its corresponding input control shown separately. b Aggregated metagene profiles illustrating normalized SPI1 ChIP-seq read density relative to the TSS across two independent SPI1-OE ChIP replicates and matched input samples. c Circos plot showing the genome-wide chromosomal distribution of SPI1 ChIP-seq binding peaks. d Volcano plot of DEGs identified by RNA-seq following SPI1 overexpression. DEGs were defined using an FDR < 0.05 and a |log2FC| > 1. e Representative gene set enrichment analysis (GSEA) plots showing hallmark pathways enriched in SPI1-overexpressing cells. f Venn diagram depicting the overlap between genes upregulated upon SPI1 overexpression and genes annotated with SPI1 ChIP-seq binding peaks. g Bubble plot showing Gene Ontology (GO) biological process enrichment of genes coregulated by SPI1 transcriptional activation and direct chromatin binding; the dot size indicates the gene count, and the color represents the adjusted P value. h De novo motif analysis of SPI1 ChIP-seq binding peaks. i Dual-luciferase reporter assay measuring SPI1-dependent transcriptional activity in a doxycycline-inducible system over time ( n = 3 independent experiments; mean ± SD; one-way ANOVA with Tukey’s post hoc test)

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: ChIP-sequencing, Control, Genome Wide, Binding Assay, RNA Sequencing, Over Expression, Activation Assay, Luciferase, Reporter Assay, Activity Assay

Journal: Signal Transduction and Targeted Therapy

Article Title: Spi-1 proto-oncogene regulates mRNA hypertranscription and malignant progression in head and neck cancer

doi: 10.1038/s41392-026-02669-6

Figure Lengend Snippet: Elevated SPI1 expression is associated with poor prognosis in laryngeal and hypopharyngeal cancer patients. a Validation of SPI1 expression in a local cohort of laryngeal cancer patients. b Kaplan‒Meier survival curve and c multivariate Cox forest plot for laryngeal cancer patients. d Validation of SPI1 expression in a local cohort of hypopharyngeal cancer patients. e Kaplan‒Meier survival curve and f multivariate Cox forest plot for hypopharyngeal cancer patients. For Kaplan‒Meier analyses ( b , e ), follow-up was truncated at 1000 days to ensure stable risk set sizes and avoid overinterpretation of sparse late follow-up

Article Snippet: The membranes were then blocked with 5% nonfat milk powder and incubated overnight at 4 °C with primary antibodies against SPI1 (1:500; Cell Signaling Technology, cat. 2258), RB1 (1:1000; Proteintech, 10048-2-Ig), phospho-RB1 (Ser780) (1:1,000; Proteintech, 84692-1-RR), Akt (pan) (1:1000; Cell Signaling Technology, #4691), phospho-Akt (Ser473) (1:1000; Cell Signaling Technology, #4060), and β-actin (1:10,000; Proteintech, 60008-1-Ig).

Techniques: Expressing, Biomarker Discovery

Journal: Journal of Advanced Research

Article Title: Developmentally endothelial locus-1 facilitates intestinal inflammation resolution by suppressing the Cmpk2-cGAS-STING pathway and promoting reparatory macrophage transition

doi: 10.1016/j.jare.2025.04.030

Figure Lengend Snippet: DEL-1 knockdown inhibits inflammation resolution of DSS-induced colitis in the recovery phase. (A) Diagram of the modeling and treatment strategy for the induced repair model. Briefly, mice were treated with AAV-DEL-1 for 4 weeks, followed by DSS feeding for 7 days. Subsequently, DSS was withdrawn and replaced with sterile water for 6 days (n = 5). (B) Body weight loss was calculated as the percent change relative to day 0. (C) Disease activity index (DAI) scores. (D) Representative images of colons. (E) Colonic length. (F) Representative images of hematoxylin and eosin (H&E) staining. (G) Histological score. (H) RT-qPCR of cytokines ( Il1β , Il6 , Tnfα , Il10 , Arg1 , Ifnα , and Ifnβ ), normalized to β-actin . (I, J) Western blot of SPI1, CMPK2 and cGAS-STING pathway related protein expression. The intensity ratio of the target protein to corresponding controls quantified using densitometric analysis, including SPI1/GAPDH, CMPK2/GAPDH, CGAS/GAPDH, p-STING/STING, p-TBK1/TBK1, and p-IRF3/IRF3. Statistical analysis was calculated by student’s t tests. ns (not significant), p > 0.05; *p < 0.05; **p < 0.01; ***p < 0.001.

Article Snippet: Transfection of plasmids DNA-mediated gene overexpression DEL-1, Spi1, Cmpk2 overexpression plasmids and corresponding controls were purchased from Genechem.

Techniques: Knockdown, Sterility, Activity Assay, Staining, Quantitative RT-PCR, Western Blot, Expressing

Journal: Journal of Advanced Research

Article Title: Developmentally endothelial locus-1 facilitates intestinal inflammation resolution by suppressing the Cmpk2-cGAS-STING pathway and promoting reparatory macrophage transition

doi: 10.1016/j.jare.2025.04.030

Figure Lengend Snippet: DEL-1 regulates transcription of Cmpk2 and reparative gene Il10 though transcription factor Spi1. (A, B) The expression of nuclear SPI1 and cytoplasm SPI1 were measured by western blot in DEL-1 overexpressed RAW264.7 macrophages pulsed with LPS and 24 h post LPS withdrawal, and the intensity ratio of the target protein to HISTONE 3 or GAPDH quantified using densitometric analysis (n = 3–4). (C, D) The expression of SPI1 were measured by western blot in BMDMs in the acute and repair model, and densitometric analysis quantified the intensity ratio of the SPI1 to GAPDH (n = 3–4). (E, F) The mRNA expression of Cmpk2 (E) and Il10 (F) were measured by RT-qPCR in RAW264.7 macrophages with Spi1 overexpression, normalized to β-actin . (G-N) Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays analyzed the regulatory role of the transcription factor Spi1 on target genes in RAW264.7 and HEK293T cells. (G) ChIP analyzed the association between Spi1 and the promoter of Cmpk2 gene without intervention. Agarose gel electrophoresis of products (top), and %input calculated from CT values of RT-qPCR (bottom). (H, I) ChIP analyzed the association between Spi1 and the promoter of Cmpk2 gene in the acute and repair model. (J) ChIP analyzed the association between Spi1 and the promoter of Il10 gene without intervention. Agarose gel electrophoresis of products (top), and %input calculated from CT values of RT-qPCR (bottom). (K, L) ChIP analyzed the association between Spi1 and the promoter of Il10 gene in the acute and repair model. (M) The luciferase activity of the Cmpk2 promoter with Spi1 binding sites in HEK293T cells. (N) The luciferase activity of the Il10 promoter with Spi1 binding sites in HEK293T cells. Statistical analysis was calculated by student’s t tests or one-way-analysis of variance (ANOVA). ns (not significant), p > 0.05; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: Transfection of plasmids DNA-mediated gene overexpression DEL-1, Spi1, Cmpk2 overexpression plasmids and corresponding controls were purchased from Genechem.

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Over Expression, Chromatin Immunoprecipitation, Luciferase, Agarose Gel Electrophoresis, Activity Assay, Binding Assay

Journal: Journal of Advanced Research

Article Title: Developmentally endothelial locus-1 facilitates intestinal inflammation resolution by suppressing the Cmpk2-cGAS-STING pathway and promoting reparatory macrophage transition

doi: 10.1016/j.jare.2025.04.030

Figure Lengend Snippet: DEL-1 induces the ubiquitin – proteasome-dependent degradation of transcription factor Spi1, and further inhibits the Cmpk2-cGAS-STING pathway in macrophages. (A-C) Cmpk2 mRNA (A) and protein (B, C) expression in RAW264.7 macrophages with transfection Cmpk2 overexpression plasmid was determined using RT-qPCR and western blot (n = 4). (D-F) RAW264.7 macrophages were transfected with DEL-1, Spi1, and Cmpk2 overexpression plasmids and corresponding controls for 24–36 h. Cells were pulsed with LPS (1 μg/ml) and the STING pathway agonist DMXAA (1 μg/ml) for 4 h in the acute phase, or withdrawn LPS stimulation and treated with DMXAA for 24 h in the recovery phase (n = 4). The expression of CMPK2 and cGAS-STING pathway related protein were measured by western blot, and densitometric analysis quantified the intensity ratio of the target protein to relevant controls: CMPK2/GAPDH, CGAS/GAPDH, p-STING/STING, p-TBK1/TBK1, and p-IRF3/IRF3. (G) The mRNA expression of Spi1 were determined by RT-qPCR in DEL-1 overexpressed macrophages pulsed with LPS and 24 h post LPS withdrawal. (H, I) DEL-1 overexpressed macrophages treated with cycloheximide (CHX, 60 μg/ml), MG132 (20 uM), and chloroquine (50 uM) in the acute and repair model, and the expression of SPI1 was measured using western blot. Statistical analysis was calculated by student’s t tests or one-way-analysis of variance (ANOVA). ns (not significant), p > 0.05; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: Transfection of plasmids DNA-mediated gene overexpression DEL-1, Spi1, Cmpk2 overexpression plasmids and corresponding controls were purchased from Genechem.

Techniques: Ubiquitin Proteomics, Expressing, Transfection, Over Expression, Plasmid Preparation, Quantitative RT-PCR, Western Blot