Acute cold exposure may disturb the physiological homeostasis of the body in ectotherms. To date, there has been no information on the effects of cold exposure on homeostasis of reactive oxygen species (ROS) or antioxidant defense response in the Chinese soft-shelled turtle, Pelodiscus sinensis. In this study, P. sinensis juveniles were acclimated at 28 °C, transferred to 8 °C as cold exposure for 12 h, then moved back to 28 °C rewarming for 24 h. We measured the ROS level and total antioxidant capacity (TAC) in the brain, liver, kidney and spleen at 2 and 12 h cold exposure, and at the end of the rewarming period. Malonaldehyde (MDA) and carbonyl protein were used as markers of oxidative damage. Turtles being maintained simultaneously at 28 °C were used as the control group. Cold exposure did not disturb the ROS balance in all 4 tissues, while rewarming raised the ROS level in the brain and kidney of P. sinensis. Cold exposure and rewarming decreased the TAC in the brain, liver and spleen but did not change the TAC in the kidney. MDA and carbonyl protein levels did not increase during the treatment, indicating no oxidative damage in all 4 tissues of P. sinensis. Our results indicated that extreme cold exposure did not impact the inner oxidative balance of P. sinensis, but more ROS was produced during rewarming. P. sinensis showed good tolerance to the harsh temperature change through effective protection of its antioxidant defense system to oxidative damage. This study provides basic data on the stress biology of P. sinensis.

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In adult newts, when a limb is amputated, a mesenchymal cell mass called the blastema is formed on the stump, where blood vessels filled with premature erythrocytes, named polychromatic normoblasts (PcNobs), elongate. We previously demonstrated that PcNobs in the blastema express an orphan gene, Newtic1, and that they secrete growth factors such as BMP2 and TGFβ1 into the surrounding tissues. However, the relationship between Newtic1 expression and growth factor secretion was not clear since Newtic1 was thought to encode a membrane protein. In this study, we addressed this issue using morphological techniques and found that the Newtic1 protein is a component of globular structures that accumulate at the marginal band in the cytoplasm along the equator of PcNobs. Newtic1-positive (Newtic1(+)) globular structures along the equator were found only in PcNobs with a well-developed marginal band in the blastema. Newtic1(+) globular structures were associated with microtubules and potentially incorporated TGFβ1. Based on these observations, we propose a hypothesis that the Newtic1 protein localizes to the membrane of secretory vesicles that primarily carry TGFβ1 and binds to microtubules, thereby tethering secretory vesicles to microtubules and transporting them to the cell periphery as the marginal band develops.