Features of Morphological Changes in Experimental CT-26 Tumors Growth
The aim of the investigation was to determine the optimal dates and morphological parameters of inoculated CT-26 tumors for evaluating the effectiveness of anti-cancer therapy. This was undertaken on the basis of morphological and morphometric investigations of the tumor growth dynamics by identifying the daily structural changes which were occurring.
Materials and Methods. The features of morphological changes in experimental tumors growth were evaluated on Balb/c mice using the mouse colon CT-26 adenocarcinoma inoculated onto the external side of mouse ear auricles (n=20). The inoculation dose in all cases was 2×105 cells. The animals were removed by cervical dislocation on days 4, 8, 11, 17–19 or 20–22 after the inoculation. Each group consisted of four animals. The excised tumor was exposed to macroscopic and histological investigation. The specimens were stained by hematoxylin and eosin and picro-fuchsin using the Van-Gieson method. There were examined the quantitative composition of the cells the areas of the stroma and tumor parenchyma, the volume of the vascular bed, the area of necrosis and the presence of hemodynamic disruptions, the mitotic activity of the tumor cells.
Results. The study of the features of morphological changes in experimental tumors growth showed that tumor growth is determined not only by the mitotic activity of the cells, but also by the degree of the vascular bed development and its capacity for performing its required functional load to the full.
It was found that tumor angiogenesis is a fluctuating process which is, to a certain degree, connected with the state of the tumor cells. The formation of the vascular bed in the form of sinusoidal capillaries begins on day 4 after inoculation and it increases continuously until day 11. Between days 17–19 after inoculation there is a decrease in the proportion of vessels in the tumor tissue, but by days 20–22 this has increased again with the formation not only of sinusoidal capillaries, but also of vessels of larger diameters (to 200 µm).
The manifestations of spontaneous tumor regration are characterized by the occurrence of cells with irreversible damage to their nuclei and cytoplasm with a parallel decrease in their mitotic activity, and by the appearance of mosaically located areas of spontaneous necrosis. Spontaneous regression is also reflected in the observation of hemodynamic and hemorheologic disorders — plethora (hyperemia), stasis, the sludge-phenomenon and diapedetic hemorrhage. Here there is a connection between the severity of the changes associated with spontaneous regression and the relationship of the tumor volume to the degree of the vascular bed development.
When the tumor is small and the vascular network is well-developed, as is seen on day 11, spontaneous regression is revealed only by irreversible cell changes in a small number of tumor cells, while, by contrast, the bigger tumor size with a small number of vessels, as is registered on days 17–19, is characterized by a considerably greater number of dead cells (85%). The tumors of larger sizes observed on days 20–22 after inoculation, despite their more developed network of blood vessels, are characterized by a high level of spontaneous regression, which leads us to the conclusion that there is a mismatch between the level of angiogenesis in relation to the needs of the proliferating parenchymal elements.
Conclusion. The development of this carcinoma is characterized by two parallel processes: the formation of the tumor as a morphological substrate and the development of spontaneous regressive changes within it. The state of the tumor cells and the degree of development of spontaneous regressive changes depend on both the size of the tumor and on the level of development and functioning of the vascular network. These findings suggest that when using CT-26 tumors inoculated into mouse ears for evaluating the effectiveness of anti-cancer therapy periods of greater than 11 days should be avoided, as from this time there are evident manifestations of spontaneous pathomorphism in the tumor.
- Rosenberg D.W., Giardina C., Tanaka T. Mouse models for the study of colon carcinogenesis. Carcinogenesis 2009; 30(2): 183–196, http://dx.doi.org/10.1093/carcin/bgn267.
- Taketo M.M., Edelmann W. Mouse models of colon cancer. Gastroenterology 2009; 136(3): 780–798, http://dx.doi.org/10.1053/j.gastro.2008.12.049.
- Tanaka Y., Eda H., Tanaka T., Udagawa T., Ishikawa T., Horii I., Ishitsuka H., Kataoka T., Taguchi T. Experimental cancer cachexia induced by transplantable colon 26 adenocarcinoma in mice. Cancer Res 1990; 50(8): 2290–2295.
- Aulino P., Berardi E., Cardillo V.M., Rizzuto E., Perniconi B., Ramina C., Padula F., Spugnini E.P., Baldi A., Faiola F., Adamo S., Coletti D. Molecular, cellular and physiological characterization of the cancer cachexia-inducing C26 colon carcinoma in mouse. BMC Cancer 2010; 10: 363, http://dx.doi.org/10.1186/1471-2407-10-363.
- Chang Y.S., di Tomaso E., McDonald D.M., Jones R., Jain R.K., Munn L.L. Mosaic blood vessels in tumors: frequency of cancer cells in contact with flowing blood. Proc Natl Acad Sci USA 2000; 97(26): 14608–14613, http://dx.doi.org/10.1073/pnas.97.26.14608.
- Kruskal J.B. Can optical imaging assist in characterization of the onset of angiogenesis in developing hepatic metastases in mice livers? Radiology 2007; 243(2): 307–308, http://dx.doi.org/10.1148/radiol.2432061988.
- Knighton D., Ausprunk D., Tapper D., Folkman J. Avascular and vascular phases of tumour growth in the chick embryo. Br J Cancer 1977; 35(3): 347–356, http://dx.doi.org/10.1038/bjc.1977.49.
- Prikaz Minzdravsotsrazvitiya RF ot 23.08.2010 №708n “Ob utverzhdenii Pravil laboratornoy praktiki v Rossiiskoy Federatsii” [Ministry of Health and Social Development of the Russian Federation Decree No.708n as of Aug 23, 2010 “On good laboratory practices of the Russian Federation”].
- Mezhdunarodnye rekomendatsii (eticheskiy kodeks) po provedeniyu mediko-biologicheskikh issledovaniy s ispol’zovaniem zhivotnykh [International guiding principles (ethical code) for biomedical research involving animals]. 1985.
- Song H.-W., Lee S.-W., Jung M.-H., Kim K.R., Yang S., Park J.W., Jeong M.-S., Jung M.Y., Kim S. Optical monitoring of tumors in BALB/c nude mice using optical coherence tomography. Journal of the Optical Society of Korea 2013; 17(1): 91–66, http://dx.doi.org/10.3807/josk.2013.17.1.091.
- Avtandilov G.G. Meditsinskaya morfometriya [Medical morphometry]. Moscow: Meditsina; 1990.
