Journal: bioRxiv

Article Title: Endothelial ACKR1 expression regulates neutrophil infiltration and breast cancer metastatic engraftment in the lung metastatic niche

doi: 10.64898/2026.03.15.711832

Figure Lengend Snippet: (A) Representative image of ACKR1-positive lung vessel from LMB-bearing mouse 28 days after implantation, showing colocalization of ACKR1 with venous endothelial marker EphB4. CD31 = magenta, EphB4 = yellow, ACKR1 = green. All ACKR1 pos vessels also expressed EphB4. 22.7±1.4% of EphB4 pos venules expressed ACKR1. Quantitation shown in . (B) Representative annotated IF image for QuPath-based proximity analysis of tumor cell distance to nearest venule (EphB4 = yellow, Pan-CK = tumor cell marker pancytokeratin in red, ACKR1 = green, and DAPI = blue). For QuPath annotations, yellow line = vessel border, red circle = disseminated tumor cell (DTC), red line = shortest distance between DTC and nearest venule border. (C) Quantification of the number of DTCs within indicated distance from the border of ACKR1-positive or ACKR1-negative venules in Day 28 LMB-bearing mouse lungs (n = 116 ACKR1 pos venules, n = 100 ACKR1 neg venules, across 4 individual mice). Unpaired t-tests. Summed data shown here for clarity, individual data points shown in . (D) Percentage of ACKR1-positive and ACKR1-negative venules with at least one DTC within 50 µm of the venule border. Chi-squared test. (E) Representative IF images for QuPath proximity analysis of neutrophil distance to nearest venule, as described in panel B except Ly6G = neutrophil marker (cyan) and red lines indicate shortest distance between neutrophil and nearest venule border. (F) Quantification of the number of neutrophils within indicated distance from the border of ACKR1-positive or ACKR1-negative venules in Day 28 LMB-bearing mouse lungs (n = 16 ACKR1 pos venules, n = 16 ACKR1 neg venules, across 3 individual mice). Unpaired t-tests. Summed data shown here for clarity, individual data points shown in .

Article Snippet: Primary antibodies used included anti-mouse ACKR1 (1 μg/mL, a kind gift of Dr. Ulrich von Andrian, Harvard Medical School), anti-mouse CD31 (1:100, Cell Signaling, 77699), anti-mouse EphB4 (2.5 μg/mL, R&D Systems, AF446), anti-mouse Pan Cytokeratin (1:200, ThermoFisher/Bioss, BS-10403R), and anti-mouse Ly-6G (E6Z1T) (1:200, Cell Signaling, 87048).

Techniques: Marker, Quantitation Assay

Journal: bioRxiv

Article Title: Endothelial ACKR1 expression regulates neutrophil infiltration and breast cancer metastatic engraftment in the lung metastatic niche

doi: 10.64898/2026.03.15.711832

Figure Lengend Snippet: (A) Representative low-magnification images of lung from mice implanted with LMB tumors at Day 28, showing overall distribution of ACKR1 expression relative to EphB4 expression. (B) Quantification of vessels positive for ACKR1, EphB4, and both ACKR1/EphB4, across 4 individual mice. (C) Quantification of the number of DTCs within indicated distance from the border of ACKR1-positive or ACKR1-negative venules in Day 28 LMB-bearing mouse lungs (n = 116 ACKR1 pos venules, n = 100 ACKR1 neg venules, across 4 individual mice). Unpaired t-tests. Individual data points from graphs in . (D) Quantification of the number of neutrophils within indicated distance from the border of ACKR1-positive or ACKR1-negative venules in Day 28 LMB-bearing mouse lungs (n = 16 ACKR1 pos venules, n = 16 ACKR1 neg venules, across 3 individual mice). Unpaired t-tests. Individual data points from graphs shown in .

Article Snippet: Primary antibodies used included anti-mouse ACKR1 (1 μg/mL, a kind gift of Dr. Ulrich von Andrian, Harvard Medical School), anti-mouse CD31 (1:100, Cell Signaling, 77699), anti-mouse EphB4 (2.5 μg/mL, R&D Systems, AF446), anti-mouse Pan Cytokeratin (1:200, ThermoFisher/Bioss, BS-10403R), and anti-mouse Ly-6G (E6Z1T) (1:200, Cell Signaling, 87048).

Techniques: Expressing