Journal: Blood Advances

Article Title: Modulating endothelial cells with EGFL7 to diminish aGVHD after allogeneic bone marrow transplantation in mice

doi: 10.1182/bloodadvances.2021005498

Figure Lengend Snippet: Effect of EGFL7 on GVHD severity in B6→B6D2F1 mice. (A) Weights of animals that had undergone transplantation were measured daily and averaged for the group (green circles: syngeneic T cells [no GVHD]; red triangles: allogeneic treated with PBS; blue squares: allogeneic treated with rEGFL7). Daily EGFL7 treatment was initiated at day +21 post-BMT. Data were pooled from 3 experiments with 6 to 13 mice per group. (B) Survival curve of transplanted mice (dotted green line: syngeneic control [SYN], blue line: allogeneic treated with rEGFL7, and thin dotted red line: allogeneic treated with PBS). Data were pooled from 3 experiments with 6 to 13 mice per group. (C) Clinical scores of GVHD + PBS, GVHD + rEGFL7, and syngeneic mice at day +28 post-BMT. (D) Histopathology of the gut 28 days after allo-HSCT. Left: magnification ×200 and right: magnification ×400. (E) Gastrointestinal (GI) histopathology score of the gut of GVHD + rEGFL7 and GVHD + PBS-treated mice. Results show mean ± standard error of the mean (SEM). (F) Immunofluorescence analysis of the intestine from PBS or rEGFL7-treated mice transplanted with allogenic splenocytes that were stained for immunofluorescence. Cells were stained with CD31 (secondary antibody: donkey anti-goat alexa fluor 488), CD45 (secondary antibody: donkey anti-rabbit alexa fluor 647), and Ki-67 antibodies (secondary antibody donkey anti-rat alexa fluor 594). Nuclei were counterstained with 4′,6-diamidino-2-phenylindole (DAPI). Magnification ×400. (G) Cells were stained with CD3 (secondary antibody donkey anti-goat alexa fluor 488), CD4 (secondary antibody donkey anti-rabbit alexa fluor 647), and CD8 antibodies (secondary antibody donkey anti-rat alexa fluor 594). Nuclei were counterstained with 4′,6-diamidino-2-phenylindole. Magnification ×400. Data show 1 representative sample from each experimental group, which consists of 3 to 4 mice per group.

Article Snippet: Recombinant human EGFL7 (rEGFL7) was purchased from Peprotech (Rocky Hill, NJ).

Techniques: Transplantation Assay, Control, Histopathology, Immunofluorescence, Staining

Journal: Blood Advances

Article Title: Modulating endothelial cells with EGFL7 to diminish aGVHD after allogeneic bone marrow transplantation in mice

doi: 10.1182/bloodadvances.2021005498

Figure Lengend Snippet: Effect of EGFL7 on immune reconstitution and graft-versus-leukemia effect. (A-C) Splenocytes and thymocytes were collected from PBS-treated mice as they reached their endpoints (day +28 to 35) and from EGFL7-treated mice (day +30 to 35 post-BMT). (A) Absolute counts of TCR + , CD4 + , and CD8 + lymphocytes in the spleen of Syn, GVHD + rEGFL7, and GVHD + PBS treated (B6→B6D2F1) mice. Data were pooled from 3 experiments with 4 to 7 mice per group. Gating was performed on CD45.1 + and CD45.2 + cells. (B) Absolute counts of BM-derived thymocytes (CD45.2 + ) in Syn, GVHD + rEGFL7, and GVHD + PBS treated (B6→B6D2F1) mice. Data were pooled from 3 experiments with 4 to 8 mice per group. Results show mean ± SEM. Statistical analysis compared GVHD + rEGFL7 vs GVHD + PBS treated (B6→B6D2F1) mice, and P values were determined by a Mann-Whitney U test. (C) Dot-plot analysis of thymocytes (derived from the BM cells [CD45.2 + CD45.1 − ]), based on the expression of CD4, CD8, and CD45.2 antigens, in the thymus of SYN control, GVHD + rEGFL7, and GVHD + PBS mice. These data are representative of 3 or more experiments with 6 to 8 mice per group. Histogram showed mean ± SEM. Statistical analysis compared GVHD + rEGFL7 vs GVHD + PBS treated (B6→B6D2F1) mice, and P values were determined by a Mann-Whitney U test. Transplant for GVL was performed in BALB/c mice as described in “Methods.” (D-E) Whole-body bioluminescent signal intensity of recipient mice (n = 4-5 per cohort). Mice were imaged on indicated days. Average radiance expressed as mean ± SEM. One representative transplant experiment of 2 is shown. (F) Splenocytes were isolated at the time when mice reached their endpoints, days 28 to 33 post-BMT for PBS vehicle and days 30 to 35 post-BMT in rEGFL7-treated mice. Percentage GFP positivity representing P815 leukemic cell infiltration in the spleen. Each dot represents a single mouse. (G) Representative flow cytometric contour plots. Allo. spl., allogeneic splenocytes; GFP, green fluorescent protein; Min, minimum; Max, maximum; SSC-A, side-scatter-area; SYN, syngeneic control; TCD-BM, T cell depleted bone marrow. *** P < .001.

Article Snippet: Recombinant human EGFL7 (rEGFL7) was purchased from Peprotech (Rocky Hill, NJ).

Techniques: Derivative Assay, MANN-WHITNEY, Expressing, Control, Isolation

Journal: Journal of Translational Medicine

Article Title: Intra-tumoural vessel area estimated by expression of epidermal growth factor-like domain 7 and microRNA-126 in primary tumours and metastases of patients with colorectal cancer: a descriptive study

doi: 10.1186/s12967-014-0359-y

Figure Lengend Snippet: Sample distributions according to location and analysed parameters. (EGFL7: Epidermal growth factor-like domain 7; miRNA-126: microRNA-126).

Article Snippet: 4 μl recombinant EGFL7 (Novus Biologicals H00051162-P01) and 1 μl of the anti-EGFL7 antibody were mixed in 200 μl dilution buffer (Dako EnVision FLEX WASH BUFFER K8007) to give approximately 2.4 μM antigen versus 0.6 μM antibody.

Techniques:

Journal: Journal of Translational Medicine

Article Title: Intra-tumoural vessel area estimated by expression of epidermal growth factor-like domain 7 and microRNA-126 in primary tumours and metastases of patients with colorectal cancer: a descriptive study

doi: 10.1186/s12967-014-0359-y

Figure Lengend Snippet: Visualisation of EGFL7 and miRNA-126. Representative images of (A) : the intra-tumoural expression of Epidermal Growth Factor-like Domain 7 (EGFL7), by immunohistochemical analysis, showing staining of endothelial cells (ECs, brown) and the corresponding classified image (Ac) demonstrating intense staining of ECs (green) and faint staining (dark green) of myofibroblasts. Only the intense signal was included in the quantitative estimate. (B) The microRNA-126 (miRNA-126) in situ hybridization (ISH) analysis, showing merely exclusive intra-tumoural expression of ECs (blue) and the corresponding classified image (Bc) demonstrating the ECs included in the quantitative estimate (green).

Article Snippet: 4 μl recombinant EGFL7 (Novus Biologicals H00051162-P01) and 1 μl of the anti-EGFL7 antibody were mixed in 200 μl dilution buffer (Dako EnVision FLEX WASH BUFFER K8007) to give approximately 2.4 μM antigen versus 0.6 μM antibody.

Techniques: Expressing, Immunohistochemical staining, Staining, In Situ Hybridization

Journal: Journal of Translational Medicine

Article Title: Intra-tumoural vessel area estimated by expression of epidermal growth factor-like domain 7 and microRNA-126 in primary tumours and metastases of patients with colorectal cancer: a descriptive study

doi: 10.1186/s12967-014-0359-y

Figure Lengend Snippet: Clinico-pathologic characteristics according to disease stage and relapse (N = 126)

Article Snippet: 4 μl recombinant EGFL7 (Novus Biologicals H00051162-P01) and 1 μl of the anti-EGFL7 antibody were mixed in 200 μl dilution buffer (Dako EnVision FLEX WASH BUFFER K8007) to give approximately 2.4 μM antigen versus 0.6 μM antibody.

Techniques:

Journal: Journal of Translational Medicine

Article Title: Intra-tumoural vessel area estimated by expression of epidermal growth factor-like domain 7 and microRNA-126 in primary tumours and metastases of patients with colorectal cancer: a descriptive study

doi: 10.1186/s12967-014-0359-y

Figure Lengend Snippet: Dot plot illustrating the intra-tumoural expressions of Epidermal Growth Factor-like Domain 7 (EGFL7) in primary tumours according to stage (St.) and relapse free (−R) and relapsed (+R) patients, respectively (N = 123). See Table for statistical differences between the medians (represented by black bars).

Article Snippet: 4 μl recombinant EGFL7 (Novus Biologicals H00051162-P01) and 1 μl of the anti-EGFL7 antibody were mixed in 200 μl dilution buffer (Dako EnVision FLEX WASH BUFFER K8007) to give approximately 2.4 μM antigen versus 0.6 μM antibody.

Techniques:

Journal: Journal of Translational Medicine

Article Title: Intra-tumoural vessel area estimated by expression of epidermal growth factor-like domain 7 and microRNA-126 in primary tumours and metastases of patients with colorectal cancer: a descriptive study

doi: 10.1186/s12967-014-0359-y

Figure Lengend Snippet: Epidermal growth factor-like domain 7 (EGFL7) and microRNA-126 (miRNA-126) expressions in primary tumours and associated lymph node/distant metastases

Article Snippet: 4 μl recombinant EGFL7 (Novus Biologicals H00051162-P01) and 1 μl of the anti-EGFL7 antibody were mixed in 200 μl dilution buffer (Dako EnVision FLEX WASH BUFFER K8007) to give approximately 2.4 μM antigen versus 0.6 μM antibody.

Techniques: