Structural characterization andin vitroimmunogenicity evaluation of amphibian-derived collagen type II from the cartilage of Chinese Giant Salamander (Andrias davidianus)

Collagen type II (CT-II) has unique biological activities and functions, yet the knowledge on amphibian-derived CT-II is rare. Herein, acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) were successfully isolated and characterized from the cartilage of Chinese Giant Salamander (CGS). The in vitro immunogenicity of collagen was then evaluated and compared with that of the standard bovine CT-II (SCT-II) by T-lymphocyte cell proliferation activity. Results demonstrated that ASC and PSC were predominantly CT-II along with minor collagen type I and maintained intact triple-helical structure of nature collagen. Compared with SCT-II, higher glycine content (337.80 and 339.93 residues/1000 residues) and lower degree of proline hydroxylation (51.81% and 52.52%) were observed in ASC and PSC. Additionally, PSC showed comparable Td (63 °C) and higher Tm (109 °C) than SCT-II (64 °C and 103 °C, respectively), indicating its high thermal and structural stability. SEM revealed that the lyophilized ASC and PSC had interconnected porous network structures of collagen-based materials. Moreover, different from SCT-II, both ASC and PSC presented no immunogenicity because they did not cause obvious proliferation of murine T-lymphocyte regardless of the induced concentration of collagen increased from 8 to 417 μg/mL. These data suggested that the amphibian-derived CGS cartilage collagens avoid the immunogenic risk of terrestrial animal collagen, and show high thermal stability and potential advantage in biomedical application.