Article Figures & Data
Figures
- Figure 1.
Characteristics of two types of orthotopic transplantation CTC mouse models to the pancreas. A. The first type is the surgical transplantation model. SUIT-2 cells in HBSS were directly injected into the pancreas pulled out through surgical incision of the abdominal wall. Primary tumours are located along the pancreas-omentum-spleen hilum. B. The second type is the intraperitoneal (ip) injection model. SUIT2-GFP cells in HBSS were ip injected into the near-pancreatic region by a 27G needle. Primary tumour was similarly located at the pancreasomental-spleen hilum. Subsequent progression such as peritoneal metastasis with ascites formation and liver/lung metastasis are clearly observed using GFP fluorescence. C. Histological examination shows primary tumours located at the pancreas-omental region and liver/lung metastasis. Inset: atypical glandular formation. Arrows indicate the primary and metastatic tumour. Bars indicate 20 μm. D: Comparison of the primary tumour and metastasis pattern between the two types of orthotopic CTC models. +; always present. +/–; sometimes present.
- Figure 2.
Characteristics of the heterotopic transplantation CTC mouse model. SUIT2-GFP cells in HBSS were injected subcutaneously. A. primary subcutaneous tumour on the back and lung metastases visualized using GFP fluorescence. The figure shows significantly higher number of lung metastases in heterotopic (sc) model than in orthotopic (pancreas) transplantation model. B. Histological examination confirms that the primary sc tumour and lung metastases are adenocarcinoma with glandular formation (arrows). Bars indicate 20 μm.
- Figure 3.
Detection of CTCs in the blood from the orthotopic and heterotopic mouse CTC models using a cytology-based CTC detection platform. A. Overview of the CTC detection procedure including filtration of CTCs using a 3D metal filter, transfer of CTCs to a glass slide, subsequent staining of CTCs, and the detection of CTCs using light microscopy. B. CTCs detected using cytokeratin (CK) or GFP immunocytochemistry and Papanicolaou (Pap) staining. Bars=20 μm. C: Representative CTCs from the orthotopic and heterotopic mouse models stained for CK and Pap. Arrows indicate a CTC cluster. Inset: single CTC. Bars=20 μm. D: Comparison of the CTC number between the two types of models. The CTC number in the heterotopic (sc) model tends to be higher than that of the orthotopic (pancreas) model, but not statistically significant (NS).
- Figure 4.
Analysis of the KRAS mutation in SUIT-2 cells and CTCs from the blood of the heterotopic (sc) xenograft mouse model. A. The procedure of CTC analysis includes pinpoint picking of the CTCs from a glass slide stained by Pap, extraction of DNA, and PCR analysis. B. Analysis of KRAS mutation in SUIT-2 cells using the cycleave PCR and direct sequencing. The result showed that SUIT-2 carry the KRAS G12D mutation at codon 12. C. Analysis of the KRAS G12D mutation of CTCs from the heterotopic mouse xenograft model by droplet-based digital PCR method. The blue and green dots (arrows) indicate droplets containing the amplified KRAS G12D mutant and wild-type alleles, respectively. The grey dots correspond to droplets without any mutant and wild-type alleles.
