In this thesis, the goal is to describe a study performed to set up a protocol to maintain and characterize in vitro cells of two immortalized cell lines previously obtained. One cell line was established from the skin samples of the cetacean Bottlenose dolphin (Tursiops truncatus) and one from the brain of the Bottlenose dolphin. Bottlenose dolphin is a protected species and the opportunities for tissues sampling from freshly dead animals suitable for cell culture are rare. For this reason, in vitro cell lines represent new opportunities for acquiring knowledge on physiological aspects of this marine mammals. The skin and brain-derived cell lines were cultured and maintained in standard conditions (at 37°C and 5% CO2) in a cell culture medium consisting of DMEM-F12 supplemented with 10% of fetal bovine serum (FBS). The cells in culture were morphologically analyzed by haematoxylin and eosin staining and the ultrastructure analysis was performed by Transmission electron microscopy observation (TEM). Moreover, cell have been characterized through immunocytochemical analysis. The cells of brain derived cell line were immunoreactive to the vimentin and ki-67 markers. The second part of this thesis focuses on investigating the effects of increasing exposure time to CAP in Bottlenose dolphin skin-derived cell line. The effects on cell viability were evaluated through the MTT assay, after different exposure to cold plasma treatment, respectively 1, 2, 5, 6, 7, 8 9 and 10 minutes, and after two incubation time: 0 hours and after 8 hours, from the treatments. Through a high-throughput screening analysis, we screened and quantified cell nuclei in the diverse phases of the mitotic cell cycle or involved in process of cell death. Our preliminary results showed an inhibition of cell proliferation and an increasing of cell death after long-time cold plasma treatment in comparison to short-time treatment in Bottlenose dolphin skin-derived cells. In this study, we showed the potential of in vitro model obtained from marine mammals, focusing on its versatility and possible application for testing the effects of novel biomedical technique as the cold atmospheric plasma (CAP) treatment in alive cells.
In this thesis, the goal is to describe a study performed to set up a protocol to maintain and characterize in vitro cells of two immortalized cell lines previously obtained. One cell line was established from the skin samples of the cetacean Bottlenose dolphin (Tursiops truncatus) and one from the brain of the Bottlenose dolphin. Bottlenose dolphin is a protected species and the opportunities for tissues sampling from freshly dead animals suitable for cell culture are rare. For this reason, in vitro cell lines represent new opportunities for acquiring knowledge on physiological aspects of this marine mammals. The skin and brain-derived cell lines were cultured and maintained in standard conditions (at 37°C and 5% CO2) in a cell culture medium consisting of DMEM-F12 supplemented with 10% of fetal bovine serum (FBS). The cells in culture were morphologically analyzed by haematoxylin and eosin staining and the ultrastructure analysis was performed by Transmission electron microscopy observation (TEM). Moreover, cell have been characterized through immunocytochemical analysis. The cells of brain derived cell line were immunoreactive to the vimentin and ki-67 markers. The second part of this thesis focuses on investigating the effects of increasing exposure time to CAP in Bottlenose dolphin skin-derived cell line. The effects on cell viability were evaluated through the MTT assay, after different exposure to cold plasma treatment, respectively 1, 2, 5, 6, 7, 8 9 and 10 minutes, and after two incubation time: 0 hours and after 8 hours, from the treatments. Through a high-throughput screening analysis, we screened and quantified cell nuclei in the diverse phases of the mitotic cell cycle or involved in process of cell death. Our preliminary results showed an inhibition of cell proliferation and an increasing of cell death after long-time cold plasma treatment in comparison to short-time treatment in Bottlenose dolphin skin-derived cells. In this study, we showed the potential of in vitro model obtained from marine mammals, focusing on its versatility and possible application for testing the effects of novel biomedical technique as the cold atmospheric plasma (CAP) treatment in alive cells.
Characterization, maintenance, and application of Bottlenose dolphin (Tursiops truncatus) cell lines: an in vitro model for cold plasma treatment
GRANDI, ELENA
2023/2024
Abstract
In this thesis, the goal is to describe a study performed to set up a protocol to maintain and characterize in vitro cells of two immortalized cell lines previously obtained. One cell line was established from the skin samples of the cetacean Bottlenose dolphin (Tursiops truncatus) and one from the brain of the Bottlenose dolphin. Bottlenose dolphin is a protected species and the opportunities for tissues sampling from freshly dead animals suitable for cell culture are rare. For this reason, in vitro cell lines represent new opportunities for acquiring knowledge on physiological aspects of this marine mammals. The skin and brain-derived cell lines were cultured and maintained in standard conditions (at 37°C and 5% CO2) in a cell culture medium consisting of DMEM-F12 supplemented with 10% of fetal bovine serum (FBS). The cells in culture were morphologically analyzed by haematoxylin and eosin staining and the ultrastructure analysis was performed by Transmission electron microscopy observation (TEM). Moreover, cell have been characterized through immunocytochemical analysis. The cells of brain derived cell line were immunoreactive to the vimentin and ki-67 markers. The second part of this thesis focuses on investigating the effects of increasing exposure time to CAP in Bottlenose dolphin skin-derived cell line. The effects on cell viability were evaluated through the MTT assay, after different exposure to cold plasma treatment, respectively 1, 2, 5, 6, 7, 8 9 and 10 minutes, and after two incubation time: 0 hours and after 8 hours, from the treatments. Through a high-throughput screening analysis, we screened and quantified cell nuclei in the diverse phases of the mitotic cell cycle or involved in process of cell death. Our preliminary results showed an inhibition of cell proliferation and an increasing of cell death after long-time cold plasma treatment in comparison to short-time treatment in Bottlenose dolphin skin-derived cells. In this study, we showed the potential of in vitro model obtained from marine mammals, focusing on its versatility and possible application for testing the effects of novel biomedical technique as the cold atmospheric plasma (CAP) treatment in alive cells.File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.12608/70637