Establishment of a model of sentinel lymph node metastasis using immunodeficient swine

Toshiaki Kurihara,Reina Fujiwara,Shunichi Suzuki,Moriaki Kusakabe,Junko Kuramoto,Yuki Nakamura,Kaori Kameyama,Yuko Kitagawa,Sachiko Matsuda,Masaki Sekino,Daiichiro Fuchimoto,Masatoshi Kamata,Kohei Saeki,Akira Onishi,Hiromitsu Jinno,Tetsu Hayashida,Takayuki Nakagawa

doi:10.1038/s41598-019-44171-w

Abstract

Lymph node metastasis occurs via the migration of cancer cells through the lymphatic system. Sentinel lymph node (SLN) biopsy is a common diagnostic strategy. SLNs have been studied using healthy rodents and large animals without metastasis. Here we used immunodeficient swine to establish a model of lymph node metastasis. We used RAG2-knockout immunodeficient swine. A431 human epithelial carcinoma cells expressing green fluorescent protein were injected subcutaneously into the posterior sides of the auricle, forelimb and hindlimb of knockout swine. Indigo carmine dye was injected subcutaneously 8 weeks after tumour cell transplantation. SLNs were extracted, observed using a stereoscopic fluorescence microscope and analysed histologically using haematoxylin and eosin staining, and immunohistochemistry. Lymphoid follicles were found in wild-type swine, and a few aggregated lymphocytes and immature lymphoid follicles were observed in knockout swine. Fluorescence in the lymph nodes indicated metastasis of tumour cells to the lymph nodes. Tumour cells replaced lymph node architectures, showed high-grade nuclear atypia and formed irregular tumour nests. Our model may be useful for the preclinical validation of diagnostic methods and minimally invasive treatment of metastatic cancer.

Highlights

Lymph node metastasis occurs via the migration of cancer cells through the lymphatic system
We developed a rodent model of lymph node metastasis that enabled us to show that magnetic particles are useful for identifying SLNs23 and that photodynamic therapy is effective for treating lymph node metastasis[24,25]
A published procedure is available for clinical identification of sentinel lymph nodes strained with isosulfan blue[9,10], and the modified method was used in our study

Summary

Introduction

Lymph node metastasis occurs via the migration of cancer cells through the lymphatic system. A431 human epithelial carcinoma cells expressing green fluorescent protein were injected subcutaneously into the posterior sides of the auricle, forelimb and hindlimb of knockout swine. Fluorescence in the lymph nodes indicated metastasis of tumour cells to the lymph nodes. Cancer metastasis frequently occurs via migration of cancer cells through the lymphatic system[1]. Sentinel lymph node biopsy (SLNB) is a common diagnostic method. In breast cancer, the results of SLNB reflect axillary lymph node metastasis[2,3,4]. Research on SLNB applied to gastrointestinal cancer is progressing, and the results show promise for reducing the requirement for surgery and for improving the quality of life of patients[8]. SLNB was performed on patients with early-stage gastric cancer with an identification and correct diagnosis rates of 97.5% and 99%, respectively[8]

Methods

Results

Conclusion