Reactive nitrogen species-mediated cell proliferation during tail regeneration and retinoic acid as a putative modulator of tissue regeneration in the geckos.

Reactive nitrogen species (RNS), a mediator of nitrosative stress, plays a vital role during wound healing but its role during tissue regeneration is poorly understood. In the present study, the role of RNS was investigated post-tail autotomy and limb amputation in a gecko species, Hemidactylus murrayi Gleadow, 1887. Tail autotomy led to an increased expression of iNOS and nitrosative stress leading to protein S-nitrosylation that probably restricted the acute inflammatory response caused by wounding. Increased nitrosative stress was also associated with proliferation of the wound epithelium and the tail blastema. Nitric oxide synthase inhibitor (L-NAME) caused retarded growth and structural abnormalities in the regenerating tail while peroxynitrite inhibitor (FeTmPyp) arrested tail regeneration. Spermine NONOate and retinoic acid, used as NO donors generated small outgrowths post-amputation of limbs with an increased number of proliferating cells and s-nitrosylation indicating the role of nitric oxide signalling in cell proliferation during regeneration. Additionally, retinoic acid treatment caused regeneration of nerve, muscle and adipose tissue in the regenerated limb structure 105 days post-amputation suggesting it to be a putative modulator of tissue regeneration in the non-regenerating limbs.

Protein S-nitrosylation: Nitric oxide signalling during anuran tail regression.

Tail regression is a remarkable process where a complex organ like the tail is completely resorbed by cell death during anuran metamorphosis. Nitric oxide is a signalling molecule involved in various physiological processes and along with reactive nitrogen species induces apoptosis. The present study describes the contribution of nitric oxide and reactive nitrogen species (nitrosative stress) during tail regression in the tadpoles of Indian tree frog, Polypedates maculatus. Spectrophotometric estimation revealed significantly higher levels of nitrite, nitrate and peroxynitrite in the regressing tails of the late climactic stages as compared to the early climactic stages and pre-regressing tails. S-nitrosylated proteins were detected in the apoptotic cells of epidermis and muscle, denervated and fragmented myofibres, outer notochordal sheath of the degenerating notochord, endothelium of blood vessels, blood cells and spinal cord of the regressing tail of the late climactic stages using fluorescent detection methods. Thus, a higher level of nitrosative stress in the late climactic stages is suggested to cause S-nitrosylation of proteins and subsequent apoptosis in the tail tissues. Macrophages were found engulfing the apoptotic cells and cell debris at the distal end of the regressing tail. Interestingly, macrophages were always found to be associated with melanocytes suggesting a close association for clearing cell debris by phagocytosis.