Background: Exosomes are small vesicles of sizes ranging from 30 to 150 nm that originate from the endosomal luminal membrane and are constantly released by fusion with cell membranes. Several studies have revealed that exosomes play an important role in mediating local and systemic cellular communication through the transmission of nucleic acid materials and information in the form of proteins. Here, we demonstrated a unique layered 3D stromal tissue model of visualization of promoting angiogenesis by exosomes in colorectal cancer (CRC) patients.

Method: The CRC specimens of exosome were obtained from 7 patients in stage IV. Isolation of total exosome was performed according to standard procedure by ultracentrifuge. Layered 3D stromal tissue model was constructed with human umbilical vein endothelial cells (HUVEC) and normal human dermal fibroblasts. Cells were aggregated by heparin and collagen to suspension under cationic condition. The cell suspension was seeded to transwell inserts membrane at a density of 2×106. Media were changed after 24h, and serum, exosome or supernatant after ultracentrifuge of serum (SAUS) was added to the 3D stromal tissue after 6 days. The isolated exosomes were diluted with an equal volume of PBS buffer to the serum sample. After cultivation, the tube formation of HUVEC was stained with Immunofluorescence, and the degree of hypervascularity was compared with healthy volunteers.

Result: Except for one serum specimen, hypervascularization was observed in the 3D stromal tissue model in an additive-dependent manner. Especially when exosome was added, hypervascularization was observed in the layered 3D stromal tissue model more efficiently than serum in all specimens (see Table). Interestingly, the frequency of hypervascularization was drastically reduced in the SAUS.

Conclusion: Our data encourage that this layered 3D stromal tissue model may be applicable to cancer diagnosis using exosomes from CRC patients.

Table. Frequency of hypervascularization by exosomes derived from CRC

 5μl 10μl 50μl 
Exosome 5/7 (71.4%) 7/7 (100.0%) 7/7 (100.0%) 
Serum 4/7 (57.1%) 6/7 (85.7%) 6/7 (85.7%) 
SAUS 1/7 (14.3%) 0/7 (0.0%) 0/7 (0.0%) 
 5μl 10μl 50μl 
Exosome 5/7 (71.4%) 7/7 (100.0%) 7/7 (100.0%) 
Serum 4/7 (57.1%) 6/7 (85.7%) 6/7 (85.7%) 
SAUS 1/7 (14.3%) 0/7 (0.0%) 0/7 (0.0%) 

Citation Format: Shiro Kitano, Kei Tsukamoto, Rii Morimura, Shinji Irie, Eiji Shinozaki, Kensei Yamaguchi. Visualization of cancer-derived exosome-mediated angiogenesis in a layered 3D stromal tissue model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5036.