Tumor Immunology Visualized in Realtime in In-vivo-like 3-D Gelfoam® Histoculture by Color-coded Imaging

Abstract

Background/aim: Tumor immunology and immunotherapy have been intensely researched recently, especially with the development of immune checkpoint inhibitors (ICIs). However, only a small percentage of patients respond to ICIs, and this response is limited to a minority of cancer types, such as colon cancer, lung cancer, and melanoma. The aim of the present study was to develop an imageable in-vivo-like in vitro system to directly visualize tumor immunology and immunotherapy in real-time, to further understand tumor immunology and develop improved tumor immunotherapy.

Materials and methods: Lewis lung carcinoma cells labeled with red fluorescent protein (LLC-RFP) and peripheral blood mononuclear cells labeled with green fluorescent protein (PBMCs-GFP), derived from transgenic GFP mice, were cultured in two dimensions (2-D) on plastic or in three dimensions (3-D) in Gelfoam^® histoculture, using RPMI-1640 culture medium with 10% fetal bovine serum, and 1% penicillin/streptomycin, and with concanavalin-A.

Results: LLC-RFP cells seeded on Gelfoam^® formed glandular-like structures after 2 days of histoculture. PBMCs-GFP were seeded on Gelfoam^® with LLC-RFP after one day to establish a co-culture. The PBMCs-GFP co-localized with many of the structures formed by the LLC-RFP cells after one day of seeding PBMCs-GFP, suggesting a very specific interaction necessary for an immunological interaction. In contrast, only a few randomized co-localizations of PBMCs-GFP and the LLC-RFP cells were seen on plastic in 2-D culture.

Conclusion: The present results demonstrate a new 3-D co-culture color-coded system to visualize in real-time the interaction of PBMCs and cancer cells using fluorescence color-coded imaging. Future experiments will test drugs that can stimulate immune reactions between the PBMCs and cancer cells.

Keywords: 3-D histoculture; GFP transgenic mice; Gelfoam®; LLC; Lewis lung carcinoma; PBMC; RFP; Tumor immunology; color-coded imaging; green fluorescent protein GFP; peripheral blood mononuclear cell; red fluorescent protein.