# Atrazine-Induced Hepato-Renal Toxicity in Adult Male Xenopus laevis Frogs
## 3 Results
### Histopathology
#### 3.2.3 Liver and Kidney Caspase-3 Immunohistochemistry  
The expression of caspase-3 was predominantly in the epithelial and immune cells of the liver and not in the cytoplasm of hepatocytes. Controls showed no positive immuno-reaction (Figures 5A and 6A). In the liver, caspase-3 expression was observed in Kupffer cells within the peri-sinusoidal space surrounding the hepatocytes (Figure 5B-D; red arrow) and in monocytes within the sinusoids (Figure 5B,D; black arrow) and sinusoidal endothelial cells (Figure 5B-D; blue arrowhead). Caspase expression was also observed in the bile duct epithelial cells of the 0.01 ug/L and 500 ug/L groups (Figure 5E,F, blue arrowhead). The number of melanocrophases (MMCs) significantly decreased in the 0.01 ug/L and 500 ug/L groups (_p_ < 0.0001) and in the 200 ug/L group (_p_ < 0.03) compared with the control group (Figure 5G). In the kidney, caspase-3 expression was not observed in the control group (Figure 6A) but in the epithelial cells of cortical proximal tubules (Figure 6B,C,E; black arrow), distal tubule (Figure 6B,C,E; red arrow) and collecting tubules (Figure 6B,C,E white arrow) in all the atrazine-exposed groups instead. However, weak expression in the glomeruli was observed in the 0.01 ug/L group (Figure 6B,G). In the 200 ug/L (Figure 6D) and 500 \(\upmu\)g/L (Figure 6E) groups, intensely expressed caspase-3 was observed in the podocytes (black arrowhead), simple squamous epithelial cells (white arrowhead) of the parietal layer and within the macula densa cells (circle) of the glomeruli.  
Figure 5: Expression of caspase-3 photomicrographs in _Xenopus laevis_ liver cells with sub-chronic exposure to atrazine. (**A**) Liver tissue section showing melanomacrophages (white arrowhead) and lack of positive caspase-3 expression. (**B**) 0.01 ug/L, (**C**) 200 ug/L and (**D**) 500 ug/L) caspase-3 expression in monocytes in sinusoids (black arrow), Kupffer cells in the peri-sinusoidal space (red arrow) and flattened squamous endothelial cells lining sinusoids (blue arrowhead). (**E**,**F**) Caspase-3 expression in epithelial cells of bile ducts of the 0.01 ug/L and 500 ug/L groups, respectively. (**G**) The population percentage of melanomacrophages (MMC) in liver sections, * significantly decreased (\(p\) < 0.05) in comparison to control group; * significantly decreased (\(p\) < 0.001) in comparison to the control group. Scale bar in A to F = 38 μm (40\(\times\) magnification). Data presented as mean \(\pm\) SD (\(n\) = 10).  
Figure 6: Photomicrographs of the expression of caspase-3 in frog kidneys following chronic exposure to atrazine. (**A**) Control with absence of caspase-3 expression. (**B**) 0.01 \(\mathrm{\SIUnitSymbolMicro g}\)/L, (**C**) 200 \(\mathrm{\SIUnitSymbolMicro g}\)/L and (**E**) 500 \(\mathrm{\SIUnitSymbolMicro g}\)/L caspase-3 expression in epithelial cells of proximal tubules (black arrow), distal tubules (red arrow) and collecting tubules (white arrow). (**D**) 200 \(\mathrm{\SIUnitSymbolMicro g}\)/L and (**E**) 500 \(\mathrm{\SIUnitSymbolMicro g}\)/L caspase-3 expression in the glomeruli, macdens cells (circle), podocytes (black arrowhead) and epithelial cells within the parietal layer (white arrowhead). (**F**) Control with low power absence of immuno-positive caspase-3 expression. (**G**) 0.01 \(\mathrm{\SIUnitSymbolMicro g}\)/L, (**H**) 200 \(\mathrm{\SIUnitSymbolMicro g}\)/L and (**I**) 500 \(\mathrm{\SIUnitSymbolMicro g}\)/L caspase-3 expression in the cortex and medulla. Scale bar in A to E = 38 \(\mathrm{\SIUnitSymbolMicro m}\) (40\(\times\) magnification). Scale bar in F to I = 240 \(\mathrm{\SIUnitSymbolMicro m}\) (10\(\times\) magnification). C: cortex; M: medulla; G: glomerulus.