## 3 Results and Discussion
### Histopathology
This is probably due to the fact that Zn is accumulated mainly in sediments [28], and probably by extraction and disturbance of the redox potential of sediments and the chemical form of heavy metal, the efflux of metals from sediment to water and aquatic organism could be accelerated [29]. Furthermore, the diet of predators mainly consists of invertebrates and other benthic organisms, increasing metal accumulation in the organ and tissues of predatory fish and reflecting the pollution level of the sediment and its biota [30]. Moreover, the correlations between Cd, Pb, and Zn accumulation at renal level and the degree of tissue damage remain statistically significant for the omnivorous species and less significant or insignificant for the herbivorous species. Cr is mainly bioaccumulated in the gills, and this observation is significantly correlated with the histopathological changes in BB (2.6 \(\pm\) 0.24), SC (2.4 \(\pm\) 0.24) and CN (2.0 \(\pm\) 0.0) (\(p\) < 0.001) because the gills are the first pathway for metals to enter into body [31; 32]. Furthermore, the Cr concentrations in fish organs increase through biom magnification at each trophic level, taking into account that carnivorous feeders show particularly higher concentrations of this metal [33]. The liver was the target organ for Cu accumulations in all species and correlated significantly with the hepatic lesion grade for BB (2.2 \(\pm\) 0.20), SC (2.0 \(\pm\) 0.0), and CN (1.8 \(\pm\) 0.2) (\(p\) < 0.001). High levels of Cu in the hepatic tissues could be correlated with the binding proteins levels, such as metallothione (MT) [34], which stores this essential metal to fulfill enzymatic and other metabolic demands [35]. The Cu bioconcentration in the hepatic tissue of SC was in agreement with other studies in which the highest concentration of it was recorded in the pelagic species (omnivorous/herbivorous) compared with the benthic (carnivorous) fish [36; 37]. Heavy Metal Bioaccumulation Activates Inflammatory Pathways in the Gills, Liver, and Kidney of BB, CN, and SC  
Recent studies have revealed that heavy metals accumulation has generated a significant increase in TNF-\(\alpha\) expression in the gills, liver, and kidney of common carp [38] or in the monocytes/macrophages of _Chana punctatus Bloch_[39]. Figure 5 illustrates the expression of TNF-\(\alpha\) in the gills, liver, and kidney of all three species exposed to metal contaminants. A large number of DAB-positive stain cells were observed in the gills, followed by the kidneys in comparison with the livers, where immunpositivity was insignificantly detected. Inflammation was especially present in the gills of the BB samples, compared to SC and CN (\(p\) < 0.001). The correlation between the TNF-\(\alpha\) concentration in the gills and kidney of all tested species and the degree of histopathological alterations revealed a significant relationship between the two analyzed parameters, respectively: the carnivorous species with the highest expression of TNF-\(\alpha\) having extended tissue damage (Figure 5C). Our results suggested that the gills are the most sensitive organ for inflammatory processes. Taking into account the higher bioaccumulation of Cd, Cu, and Zn, this might lead to the induction of inflammation and upregulation of pro-inflammatory TNF-\(\alpha\) cytokine [40]. Western blot analysis indicated that the highest TNF-\(\alpha\) level was recorded for the carnivorous species (BB), being in agreement with the histopathological damages. Heavy Metal Bioaccumulation Activates Reparatory Mechanisms in Gills, Liver, and Kidney of Barbus barbus, Chondrostoma nasus, and Squalius cephalus  
The structural integrity of fish's gills, kidneys, and liver also depends on cell integrity [19] and can be correlated with the expression of PCNA (proliferating cell nuclear antigen) in fish organs exposed to metal contamination [24; 38], taking into account that PCNA can act as a mediator between cell cycle control and DNA repair [39].