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Notes: Summary The rationale of the present investigation is the observations made by many authors of changes in the molecular structure of the cell surface during the multistep process of malignant transformation. These changes may influence cell-matrix and cell-cell interactions and thereby cause changes in cell adhesiveness and cell shape. The aim of the present work was to investigate whether the development of various grades of transformation in vivo and in vitro of human urothelial cells is accompanied by significant changes in cell shape as measured by Fourier analysis. The following transformation grades (TGr) have been defined (Christensen et al. 1984; Kieler 1984): TGr I=nonmalignant, mortal cell lines that grow independently of fibroblasts and have a prolonged life span. TGr II=nonmalignant cell lines with an infinite life span. TGr III=malignant and immortal cell lines that grow invasively in co-cultures with embryonic chick heart fragments and possess tumorigenic properties after s.c. injection into nude mice. Comparisons of 4 pairs of cell lines were performed; each pair was of the same origin. Two pairs-each including a TGr I cell line (Hu 961b and Hu 1703S) compared to a TGr III cell line (Hu 961a or Hu 1703He)-were derived from two transitional cell carcinomas (TCC) containing a heterogeneous cell population. Two additional cell lines classified as TGr II (HCV-29 and Hu 609) were compared to two TGr III sublines (HCV-29T and Hu 609T, respectively) which arose by “spontaneous” transformation during propagation in vitro of the respective maternal TGr II-cell lines. One of these TGr II cell lines (HCV-29) originated from the histologically normal bladder mucosa obtained from a patient with a previous history of bladder papillomata treated with irradiation (Fogh, personal communication). The other TGr II cell line (Hu 609) was derived from the normal ureter of a patient with renal carcinoma. In each of these 8 cell lines, the shape of 100 cells chosen at random were subjected to Fourier analysis of shape. Each of the particular harmonic amplitude values studied was used as an individual parameter for the evaluation of differences between compared cell lines, using Chi-Square test and discriminant analysis. It was found that in two of four analysed pairs of cell lines, i.e. Hu 1703S (TGr I) vs Hu 1703He (TGr III) and HCV-29 (TGr II) vs HCV-29T (TGr III), the differences in cell shape between the two populations were very well pronounced, as was shown by several statistical parameters. In the two other pairs of cell lines, i.e. Hu 961b (TGr I) vs Hu 961a (TGr III) and Hu 609 (TGr II) vs Hu 609T (TGr III) significant differences in cell shape were also found, but they were less pronounced. The conclusion is, that differences in cell shape in vitro may reveal the cellular heterogeneity of the original transitional cell carcinoma and/or the progression of in vitro propagated urothelial cells from one grade of transformation into another.

Notes: Summary In order to evaluate in mathematical terms the morphological changes occurring in the course of cell spreading, Fourier analysis of shape was applied. Human urothelial Hu 961 b cells plated on type IV collagen, fibronectin, laminin, glass and bovine serum albumine (BSA) were studied. Fourier parameters describing cell shape as well as surface areas covered by the cells on the substrate were subjected to statistical analysis. Using analysis of variance and discriminant analysis it was found that parameters describing cell shape (both gross shape of cells and their fine scale contour foldings) possessed a higher power of discrinunation between the cells spread on various substrates than the differences in cell surface areas. In the course of observation (75 and 150 min) the highest number of attached cells and highest degree of spreading were found when cells were plated on type IV collagen. Moderate alterations in cell shape and moderate increase of surface area were seen in the group of cells seeded on fibronectin, whereas the cells plated on laminin, glass and BSA revealed a moderate increase of surface area, but no changes in their shape were observed. The differences in attachment of cells and in the degree of their spreading might be due to the variation in expression of plasma membrane receptors for various substrates. The Fourier analysis of cell shape coupled with measurement of surface area is a good tool for quantitative evaluation of cell spreading and can be used for discrimination between cells spread on different substrates.