rabbit anti sdf4 polyclonal antibody  (Proteintech)

Journal: Cell Death Discovery

Article Title: Fibroblast CEBPD/SDF4 axis in response to chemotherapy-induced angiogenesis through CXCR4

doi: 10.1038/s41420-021-00478-0

Figure Lengend Snippet: A SDF4 expression was induced following CEBPD induction by CDDP or 5-FU treatment in HFL1 cells infected with shβ-galactosidase (shC) or shCEBPD (shD) lentiviruses and treated with or without CDDP or 5-FU for 6 h. RT-PCR assays were conducted to examine the CEBPD , SDF4, SDF1 , and GAPDH transcript levels. B SDF4 expression was examined in conditioned medium (C.M.) or cell lysates from HFL1 cells after CDDP or 5-FU treatment for 6 h. p84 was used as an internal control. C CEBPD activates SDF4 reporter activity. Representation of reporter constructs (left panel). A reporter assay was conducted to assess the activity of the SDF4 reporter with or without CEBPD expression vector (right panel). D A ChIP assay was performed with the indicated antibodies. Three independent experiments were performed in triplicate. All data are expressed as the mean ± S.D. Differences between groups were analyzed with the unpaired two-tailed t -test. *** p < 0.001.

Article Snippet: The primary antibodies used in this study were mouse anti-CD31 monoclonal antibody (1:100 dilution; Zhongshan JinQiao, ZM-0044, China), rabbit anti-SDF4 polyclonal antibody (1:80 dilution; Proteintech, 10517-1-AP, USA), and mouse anti-VEGF-D monoclonal antibody (1:50 dilution; R&D systems, MAB286, USA).

Techniques: Expressing, Infection, Reverse Transcription Polymerase Chain Reaction, Control, Activity Assay, Construct, Reporter Assay, Plasmid Preparation, Two Tailed Test

Journal: Cell Death Discovery

Article Title: Fibroblast CEBPD/SDF4 axis in response to chemotherapy-induced angiogenesis through CXCR4

doi: 10.1038/s41420-021-00478-0

Figure Lengend Snippet: A The proliferation of HUVECs cultured in conditioned medium from HFL1 cells infected with shβ-galactosidase (shC) or shSDF4 (shS) lentiviruses and treated with or without CDDP or 5-FU for 6 h, followed by 18-h recovery with fresh medium was assessed. B The migration of HUVECs was assessed by determining the number of HUVECs cultured in conditioned medium from HFL1 cells infected with shC and shS lentiviruses and treated with or without CDDP or 5-FU for 6 h, followed by 18-h recovery with fresh medium. C Angiogenesis was assessed by counting the intersection number between branches of HUVECs growing in conditioned medium from HFL1 cells infected with shC and shS lentiviruses and treated with or without CDDP or 5-FU for 6 h, followed by 18 h recovery with fresh medium. D Assays to assess migration and in vitro tube formation were conducted as described in materials and methods. HUVECs were treated with SDF4 at 0.25, 0.5, and 1 μg/ml. E Cebpd +/+ (WT) mice were subcutaneously inoculated with 0.5-μg/ml SDF4, 1-μg/ml SDF4, or 0.2-μg/ml VEGF Matrigel plugs. The experimental mice were sacrificed, and the Matrigel plugs were removed to assess the newly formed blood vessels; hemoglobin levels were measured in the plugs using a Drakin’s reagent kit ( n = 3 per group). VEGF was used as a positive control. F A549 cells mixed with HFL1 cells carrying a shβ-galactosidase knockdown vector with GFP (shC-GFP) or a SDF4 knockdown vector with GFP (shS-GFP) were inoculated subcutaneously into the dorsal rear flanks of NOD-SCID mice, and the mice were treated with or without CDDP (5 mg/kg). The mice with A549-xenografted tumors were sacrificed in the 12th week. Tumor tissues were stained for CD31 (red) and GFP (green), and nuclei were stained with DAPI (blue). G The number of metastasis nodules from A549-xenografted tumors in the lungs was determined. H Tumor volume was measured with external calipers and calculated using the standard formula: V = ( w × l 2 ) × 0.52, where l is the length and w is the width of the tumor ( n = 6 per group). Three independent experiments were performed in triplicate. All data are expressed as the mean ± S.D. Differences among groups were analyzed with one-way ANOVA followed by the Tukey’s multiple comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The primary antibodies used in this study were mouse anti-CD31 monoclonal antibody (1:100 dilution; Zhongshan JinQiao, ZM-0044, China), rabbit anti-SDF4 polyclonal antibody (1:80 dilution; Proteintech, 10517-1-AP, USA), and mouse anti-VEGF-D monoclonal antibody (1:50 dilution; R&D systems, MAB286, USA).

Techniques: Cell Culture, Infection, Migration, In Vitro, Positive Control, Knockdown, Plasmid Preparation, Staining, Comparison

Journal: Cell Death Discovery

Article Title: Fibroblast CEBPD/SDF4 axis in response to chemotherapy-induced angiogenesis through CXCR4

doi: 10.1038/s41420-021-00478-0

Figure Lengend Snippet: A Cell membrane fractions were incubated with or without GST-SDF4, and then, a co-immunoprecipitation assay was performed with antibody against SDF4, GST or CXCR4. B An immunofluorescence assay was conducted to assess GST, SDF4, and CXCR4 signals. C Angiogenesis was assessed by counting the branched intersection number of SDF4-treated HUVECs with or without AMD3100 treatment as indicated. D , E The activity of AKT1, ERK1/2, and p38 in response to SDF4 (0.5 μg/ml) and with or without AMD3100 (20 μg/ml) treatment in the indicated time courses. F Angiogenesis was assessed by counting the branched intersection number of HUVECs treated with SDF4 and/or the indicated kinase inhibitors or SDF4 combined with pretreatment with increasing concentrations of wortmannin, PD98059 or SB203580. G VEGFD expression in response to various signaling inhibitors on SDF4-treated HUVECs. Three independent experiments were performed in triplicate. H LLC1-Luc2 cells were orthotopically inoculated into the lung of C57BL/6 mice. The experimental mice were treated with CDDP or AMD3100 as indication after inoculation with tumor cells. Representative in vivo bioluminescent images and total tumor flux of LLC1-Luc2-bearing mice in each group shown at 5th week. n = 8 per group. All data are expressed as the mean ± S.D. Differences among groups were analyzed with the one-way ANOVA followed by Tukey’s multiple comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The primary antibodies used in this study were mouse anti-CD31 monoclonal antibody (1:100 dilution; Zhongshan JinQiao, ZM-0044, China), rabbit anti-SDF4 polyclonal antibody (1:80 dilution; Proteintech, 10517-1-AP, USA), and mouse anti-VEGF-D monoclonal antibody (1:50 dilution; R&D systems, MAB286, USA).

Techniques: Membrane, Incubation, Co-Immunoprecipitation Assay, Immunofluorescence, Activity Assay, Expressing, In Vivo, Comparison

Journal: Cell Death Discovery

Article Title: Fibroblast CEBPD/SDF4 axis in response to chemotherapy-induced angiogenesis through CXCR4

doi: 10.1038/s41420-021-00478-0

Figure Lengend Snippet: A Tissue specimens from 57 patients with lung cancer treated with or without cisplatin were stained for SDF4 expression. SDF4 was positively correlated with fibroblasts in cisplatin-treated lung cancer patients. B , C SDF4, VEGF-D, and CD31 staining was performed via IHC with their individual specific antibodies. SDF4 abundance was correlated with VEGF-D and CD31 signals in 57 lung cancer patients. D Correlation between SDF4 abundance and the survival rate of cisplatin-treated lung cancer patients. Differences between patient subsets in overall survival were determined via Kaplan–Meier plot analysis and log-rank tests. A two-tailed p < 0.05 was considered statistically significant. Statistical analysis was performed with SPSS v. 17.0 software (SPSS, USA).

Article Snippet: The primary antibodies used in this study were mouse anti-CD31 monoclonal antibody (1:100 dilution; Zhongshan JinQiao, ZM-0044, China), rabbit anti-SDF4 polyclonal antibody (1:80 dilution; Proteintech, 10517-1-AP, USA), and mouse anti-VEGF-D monoclonal antibody (1:50 dilution; R&D systems, MAB286, USA).

Techniques: Staining, Expressing, Two Tailed Test, Software