Fluoxetine activity and an angiogenic-related marker.

<p>(<b>A</b>) Representative images of colon sections labeled with anti-CD133 and anti-CD31 antibodies, and nuclei stained with DAPI. White arrows indicate single-stained and double-stained positive cells into PCCS areas in colon sections from (<b>A1</b> to <b>A3</b>) MNNG without FLX treatment, and (<b>A4</b> to <b>A6</b>) MNNG+FLX treatment groups. (A3 and A6) Double-stained positive cells are shown by white arrows in microvessel-like structures nearby cryptal bottoms. Pictures were taken with FITC (495–521 nm), ultraviolet (358–461 nm), and Texas Red (595–605 nm) filters. All pictures were taken at 600x magnification, scale bars represent 20 µm. (<b>B</b>) Representative images of single colon-stained sections with anti-CD31 antibody (positive cells are seen with dark-brown cytoplasm) from (<b>B1</b>) MNNG without FLX treatment, and (<b>B2</b>) MNNG+FLX treatment groups. Pictures were taken at 400x magnification, scale bars represent 50 µm. (<b>B3</b>) Relative number of CD31 positive cell clusters detected <i>per</i> PCCS area (**p<0.01; MNNG without FLX, <i>n</i> = 4; FLX+MNNG, <i>n = </i>4).</p

Chemopreventive activity of fluoxetine in colon tissue.

<p>(<b>A</b>) Representative histological image of a severely dysplastic area (MNNG-exposed mouse). The picture inset (enlarged from the boxed region) shows the characteristic severe dysplastic features; e.g., partial loss of cell polarity, none goblet cells, presence of Paneth cells (blue arrow), and mitosis (white arrows). Pictures were taken at 200x magnification, scale bars represent 20 µm. High-magnification images were taken at 1000x magnification. (<b>B</b>) Representative image of a moderate dysplasia (MNNG+FLX treated mouse). The inset shows a compressed cryptal luminal opening, elongated nuclei (red arrow), a crowded and pseudostrafied area (sectioned black line), but with a generally still preserved cell polarity, and a lower number of globet cells (yellow arrow). Magnifications are described above. (<b>C</b>) Quantification of dysplastic lesions. Aberrant crypt foci index (ACF-i) shown as number of dysplastic lesions <i>per</i> µm² (*P<0.05; MNNG without FLX, <i>n</i> = 5; FLX+MNNG, <i>n = </i>4). (<b>D</b>) Aberrant crypt index (AC-i) shown as number of dysplastic single crypts <i>per</i> µm² (*P<0.05; MNNG without FLX, <i>n</i> = 5; FLX+MNNG, <i>n = </i>4). (<b>E</b>) Representative image of a dysplastic area (sectioned black line), and its relative vascularization spreading inside. The inset (enlarged from the boxed region) shows two microvessels towards the inner region of the dysplastic area. The yellow arrow points to a microvessel wall, and the red arrow to erythrocytes inside the microvessel walls. Pictures were taken at 400x magnification, and further details are described above. (<b>F</b>) Relative dysplastic vascularization, shown as the number of microvessels <i>per</i> lesion (*P<0.05; MNNG without FLX, <i>n</i> = 5; FLX+MNNG, <i>n = </i>4).</p

Antiproliferative activity of fluoxetine in colon tissue.

<p>(<b>A</b>) Proliferating cells detected by staining with anti-Ki67 antibody (MNNG-treated mouse). (1) Red arrows show dark-brown positive cells migrating upward in the epithelial proliferative zone. (2) Stromal positive cells are shown by a red arrow near to the cryptal-bottom. Pictures were taken as described above. (<b>B</b>) Proliferating cells (anti-Ki67 antibody; red arrows) in a MNNG+FLX-treated mouse are shown at the cryptal bottom. Pictures were taken as described above. (<b>C</b>) Proliferation in epithelial areas shown by labeling with anti-Ki67 antibody (***P<0.001; MNNG without FLX, <i>n</i> = 5; FLX+MNNG, <i>n = </i>4). (<b>D</b>) Proliferation in PCCS areas by labeling with anti-Ki67 antibody (***P<0.001; MNNG without FLX, <i>n</i> = 5; FLX+MNNG, <i>n = </i>4). (<b>E</b>) Proliferation in epithelial areas shown by labeling with anti-PCNA antibody (**P<0.01; MNNG without FLX, <i>n</i> = 4; FLX+MNNG, <i>n = </i>4). (<b>F</b>) Proliferation in PCCS areas shown by labeling with anti-PCNA antibody (**P<0.01; MNNG without FLX, <i>n</i> = 4; FLX+MNNG, <i>n = </i>4). (<b>G</b>) Expression of c-Myc in epithelia of colon tissue (***P<0.001; MNNG without FLX, <i>n</i> = 3; FLX+MNNG, <i>n = </i>4). (<b>H</b>) Expression of c-Myc in PCCS areas of colon tissue (***P<0.001; MNNG without FLX, <i>n</i> = 3; FLX+MNNG, <i>n = </i>4).</p

Fluoxetine activity and angiogenic-related markers.

<p>(<b>A</b>) Representative images of colon sections labeled with anti-CD133 and anti-VEGF antibodies, and nuclei stained with DAPI. White arrows indicate single-stained and double-stained positive cells into PCCS areas in colon sections from (<b>A1</b> to <b>A3</b>) MNNG without FLX treatment, and (<b>A4</b> to <b>A6</b>) MNNG+FLX treatment groups. Pictures were taken with FITC (495–521 nm), ultraviolet (358–461 nm), and Texas Red (595–605 nm) filters. All pictures were taken at 600x magnification, scale bars represent 20 µm. (<b>B</b>) Representative images of single colon-stained sections with anti-VEGF antibody (positive cells are seen with dark-brown cytoplasm) from (<b>B1</b>) MNNG without FLX treatment, and (<b>B2</b>) MNNG+FLX treatment groups. (<b>B3</b>) Relative number of cells expressing VEGF within PCCS areas (***p<0.001; MNNG without FLX, <i>n</i> = 3; FLX+MNNG, <i>n = </i>4). All pictures were taken at 400x magnification, scale bars represent 50 µm. (<b>C</b>) Representative images of colon sections labeled with anti-CD133 and anti-CD34 antibodies, and nuclei stained with DAPI. White arrows indicate single-stained and double-stained positive cells into PCCS areas in colon sections from (<b>C1</b> to <b>C3</b>) MNNG without FLX treatment, and (<b>C4</b> to <b>C6</b>) MNNG+FLX treatment groups. Pictures were taken as described above. (<b>D</b>) Representative images of single colon-stained sections with anti-CD34 antibody (positive cells are seen with dark-brown cytoplasm) from (<b>D1</b>) MNNG without FLX treatment, and (<b>D2</b>) MNNG+FLX treatment groups. (<b>D3</b>) Relative number of CD34 positive cells in PCCS areas (***p<0.001; MNNG without FLX, <i>n</i> = 4; FLX+MNNG, <i>n = </i>4). All pictures were taken as described above.</p