Assessment of Nuclear Homogenicity in Chronic Lymphocytic Leukemia by Image Analysis.

Maturation of progenitor cells involves gradual chromatin condensation. We have shown that B-CLL cells represnet specific specrtral patterns by multipixel spectral imaging (J. Histocem. Cytohem. 46:113,1998). We further used image analysis techniques to quantify the alteration of chromatin homogenicity in lymphocytes from healthy subjects and B-CLL patients, as well as during maturation of myeloid and erythroid series, using light microscopic digital images of May-Grünwald Giemsa-stained (MGG) cells (B-CLL n=60, control n=22) and transmission electron microscopy (TEM). Nuclei were outlined by mouse cursor. Light microscopy images were converted to grayscale and designated as vector of the red, green and blue channels (sqrt(R*R+G*G+B*B)). Average color intensity (0–255), entropy (0–8), standard deviation, area diversity (0–100) and positive percent area (PPA) measurements were obtained from each nucleus. The entropy and area diversity of B-CLL lymphocyte nuclei was significantly lower than control lymphocytes (entropy 5.6±0.19 vs. 5.44±0.2, respectively, p<0.0001 and area diversity 33.2±4.3 vs. 30.5±2, respectively, p=0.006). In addition, we analyzed MGG-stained and TEM myeloid and erythroid cells at various stages of differentiation (n=155). Both the myeloid and erythroid cell lines showed gradual reduced color intensity, increased entropy and area diversity. Furthermore, MGG-stained megaloblastic proerythroblasts showed significantly higher levels of entropy and area diversity as compared with control proerythroblasts (n=73). Thus, assessment of nuclear entropy and area diversity may help to in the differentiation between normal lymphocytes and B-CLL cells.