## 3 The Mesotheliomagenic Activity of CNT: A Remaining Issue
### Toxic Needled-Like (CNT-N) Versus Non-Toxic Tangled-Like (CNT-T) Carbon Nanotubes  
Several animal models have been established to study mesothelioma by using asbestos fibers. Intraperitoneal injection is the preferred administration route because long, straight, fibrous and solid fibers directly reach mesothelial cells of the peritoneal cavity. In addition, the clearance mechanisms of this cavity are similar to those of pleural cavity, which is the predilection site for malignant mesothelioma [22; 56]. Not all rodents are equally sensitive to mesothelioma fibers. Indeed, the incidence in rats is higher than in mice, which therefore appears to be more resistant to mesothelioma development [68]. The mesotheliomagenic properties of CNT represent an interesting tool for investigating mesothelioma in animals. We have shown that CNT-induced mesothelioma affects all treated rats (only 30% to 50% of animals injected with asbestos) in a limited period (6 months for CNT instead of 2 years for asbestos) [92; 101]. The other key advantage of CNT compared to natural asbestos is the existence of a very wide range of manufactured CNT. These particles can be categorized from a structural and toxicological point of view by using their morphology (tangled CNT T versus needlesled/straight/rod CNT N). CNT-T are thin enough to fold and self-assemble into short, tangled aggregates, while straight CNT-N are fibrous, resistant and long. Unlike asbestos fibers, which are all considered carcinogenic, several studies demonstrate that only CNT-N are associated with the development of chronic pathologies (including mesothelioma) unlike CNT-T, which are poorly reactive and toxic [92; 101; 102; 103]. The exact reasons for this discrepancy have not yet been fully elucidated. The greater bio-persistence of CNT-N (longer and therefore more difficult to clear) is one possible explanation. In that respect, [PERSON] and co-authors reported frustrated phagocytosis by macrophages in rodents treated with CNT-N. Accordingly, these needlesd CNT-N are not completely covered by phagocytes and remain biopersistent in the tissue. In contrast, tangled CNT-T are entirely phagocytosed and taken up by macrophages resulting in tissue translocation, particle biodegradation and accelerated rate of clearance [18; 104] Nanotube geometry is also crucial in the development of inflammation, lymphoid infiltration and granuloma formation in animals. Only CNT-N exposure resulted in neutrophilization inflammation and larger lymphoid and fibrotic granulomas contrary to CNT-T [37; 105]. Needled CNT-N were more potent than tangled CNT T to elicit inflammatory effects towards macrophages [106; 107]. For [PERSON] and colleagues, the shape- and length-related structural determinants of CNT are also crucial factors for their potential carcinogenic activity. Straight and fibrous CNT-N were the strongest inducers of chromosomal aberrations in cell cultures [65]. CNT-N elicit a more pronounced primary genotoxicity effect than CNT-T, as assessed by DNA damage and micronuclei formation [108]. These in vitro observations are in accordance with data obtained _in vivo_. CNT-N cause secondary genotoxicity and mesothelioma in rodents (absolute incidences of 100%) contrary to CNT-T (no genotoxicity and tumor) [92; 101; 102; 103; 108; 109]. A New Strategy to Identify Pathogenic Immune Pathways: Gene or Protein Expression by Purified Macrophages after Needled and Tangled CNT Exposure  
The paradigm associating physicochemical characteristics and carcinogenicity of CNT represents a major asset for risk management and predictive toxicology. However, these data do not entirely explain the reasons why some CNT induce or not mesothelioma, and do not offer specific mechanisms or targets for clinical management or drug development against mesothelioma. RNA-related profiling methods and next-generation sequencing (NGS) technologies were elaborated to cartography genes expressed by purified cell populations.