FT-IR Study Of The Cold Induction Process Of Fishes' Cells

It is an interesting biological topic to understand the impact of temperature, an important environmental factor for biology, on controlling effect of biochemical activity at the stage of cells. Chilling injury is a special biological concept and the nomenclature especially refers to the process that the warm-adaptable species can not get used to a relative low temperature and are caused to death. The mechanism research of chilling injury is beneficial not only to the theoretical foundation of this basic biology phenomena, but also to the practice of localization development of warm-adaptable species. In this degree thesis, CBT cell from Colossoma Brachypomum was selected for representing warm-adaptable species and CP cell from grass carp was selected as a kind of eurythermic species. The two types of cells were cold induced for 0-8 days (CBT-10℃, CP-4℃) and treated with different temperatures (-80℃, -20℃, and 60℃). By using FT-IR technique, the relative contents of the treated cells were observed, including DNA, RNA, protein, phosphates and glucose. In this way, the relationship between molecular structure and temerature can be well elucidated.In the 10 ℃ cold induction experiments of CBT cells, we found that: 1) The variation of the contents of the cells can be divided into different stages. In 0-1 day, the contents varied a lot and after the first day, the normal synthesis of RNA and DNA is obviously restrained; 2) inducing for 1-3 days, FT-IR spectra still demonstrate the existence of RNA transcription and protein translation, combining with the phosphatization of glucose. 3) after 4 days, FT-IR spectra show that most of DNA are decomposed, the transcription of RNA are stopped and the proteins are hydrolyzed, indicating the apoptosis of the cells. 4) inducing till 7-8 days, the contents of protein and RNA was decreased strongly, implying the decomposition of the cells.In the treatment of CBT cells by low (below freezing point) and high temperature, we found that:1) the damage of the intimidated cold (10 ℃) above freezing point to the cells happens mainly due to apoptosis and that of ultra high/low temperature is thephysical death; 2) As shown in FT-IR spectra, the influences of ultra low temperature -80 °C and high temperature 60 °C are quite similar, which exhibits as the denaturalization of proteins, the capability lose of utilizing the energy and the breaking of the cell membrane.In the 4 °C cold induction experiments of CP cells, we found that: 1) in 0-3 days, the variation of the cell contents is stable calculated by FT-IR results; 2) after 4 days, FT-IR spectra show that most of DNA are decomposed, the transcription of RNA are stopped and the proteins are hydrolyzed, indicating the apoptosis of the cells.In the treatment of CP cells by low (below freezing point) and high temperature, we found that:l) at -20 °C below the freezing point, an obvious accumulation of RNA and proteins are observed, suggesting that the cold stimulation reaction of the cells is induced at the first stage under such temperature; 2) the other results are similar to CBT cells.The above results show that, apoptosis occurs to warm-adaptable fish CBT cells under intimidated temperature 10 °C and to eurythermic fish CP cells under a relatively lower temperature 4 °C. The cell contents variation of the two types of cells are different mainly in the period of 1-3 days cold induction. The restrained impact of CBT cells to RNA synthesis, DNA transcription and protein synthesis at low temperature is much stronger than CP cells. Chilling injury is different with cold injury (temperature below freezing point). As shown in FT-IR spectra, the influences of ultra low temperature -80 °C and high temperature 60 °C are quite similar, which exhibits as the denaturalization of proteins, the capability lose of utilizing the energy and the breaking of the cell membrane.In this degree thesis, FT-IR technique was firstly employed in the research of cell chilling injury mechanism and proved to be very successful, which extends the application of FT-IR spectra in biological research and optimizes the methodology for continuously monitoring the contents of cells. All the results here are not reported previously, and valuable in discover the unique reaction of molecular content of cells through the process of chilling injury.