Proliferation of human lung cancer in an orthotopic transplantation mouse model

Abstract

The objective of this study was to clarify the growth and proliferation style of human lung cancer grown in an orthotopic transplantation model. The human lung squamous cell carcinoma SQ5 and adenocarcinoma A549 cell lines were used. Tumor cells suspended in serum-free medium were directly injected into the main bronchi of anesthetized female Balb/c athymic nude mice with simultaneous administration of 0.01 M ethylenediaminetetraacetic acid. Bromodeoxyuridine was injected into mice 20 min before sacrifice. Lung tissue with tumor nodules and subcutaneous tumors were fixed and confirmed by histological examinations. Bromodeoxyuridine-labeled cells in the tumor area were counted, and the proliferation index was calculated. Lung tumor colonies of various sizes were obtained in the SQ5- and A549-cell orthotopically transplanted mice. Orthotopic SQ5 tumors whose minor diameter was 40-700 μm and major diameter was 80-830 μm showed no definite necrosis. Orthotopic SQ5 tumors whose minor diameter was 540-5,200 μm and major diameter was 600-6,100 μm showed definite necrosis in the tumor center. Similar results were also found in the orthotopic A549 tumors. The proliferation index was 7.38 (3.03)/10.63 (3.10) in the orthotopic SQ5 tumors with/without necrosis and 6.99 (2.10) in the subcutaneous SQ5 tumors with necrosis, respectively. The proliferation index was 2.70 (0.88)/3.53 (1.70) in the orthotopic A549 tumors with/without necrosis and 3.91 (0.63) in the subcutaneous A549 tumors with necrosis, respectively. The data suggest that this orthotopic transplantation model may provide the proper organ microenvironment for lung cancer growth and may be suitable for the target therapy research of human lung cancer.

Figure 1.

SQ5 tumor nodules in an orthotopic transplantation model (n=20). A total of 86 tumor nodules without necrosis was analyzed. Minor and major diameters were 40–700 and 80–830 μm, respectively (A and B). A total of 50 tumor nodules with necrosis was analyzed. Minor and major diameters were 540–5,200 and 600–6,100 μm, respectively (C and D).

Figure 2.

A549 tumor nodules in an orthotopic implantation model (n=24). A total of 51 tumor nodules without necrosis was analyzed. Minor and major diameters were 20–570 and 70–750 μm, respectively (A and B). A total of 9 tumor nodules with necrosis was analyzed. Minor and major diameters were 680–5,500 and 800–15,000 μm, respectively (C and D).

Figure 3.

Immunofluorescence examination of SQ5 tumors in the orthotopic and s.c. transplantation models. PI-labeled nuclei indicate the nuclei of the total cells (top row). BrdUrd-labeled nuclei indicate the nuclei of proliferating cells (middle row). Both BrdUrd- and PI-labeled cells were counted as proliferating cells (bottom row). Left column: proliferation of the orthotopic SQ5 tumor colony without necrosis. Middle column: proliferation of the orthotopic SQ5 tumor colony with necrosis. Right column: proliferation of the s.c. SQ5 tumor colony.

Figure 4.

Immunofluorescence examination of A549 tumors in the orthotopic and s.c. transplantation models. PI-labeled nuclei indicate the nuclei of the total cells (top row). BrdUrd-labeled nuclei indicate the nuclei of proliferating cells (middle row). Both BrdUrd- and PI-labeled cells were counted as proliferating cells (bottom row). Left column: proliferation of the orthotopic A549 tumor colony without necrosis. Middle column: proliferation of the orthotopic A549 tumor colony with necrosis. Right column: proliferation of the s.c. A549 tumor.