DANCE In Mouse Tissue Cells And Its Biochemical Mechanism

Objective: Recently we discovered and reported a new cellular response, which wasnamed as “Density Alteration in Non-physiological Cells”(DANCE), and could bedetermined by discontinuous sucrose gradient centrifugation (DSGC). Based on ourprevious studies, the objective of the present study was to investigate the DANCEresponse in mouse tissue cells of important organs, and to study the biochemicalmechanisms of DANCE.Methods:1. At room temperature (25°C) and low temperature (4°C), mouse tissue cells wereseparated by DSGC at different times after sacrifice of the animals, and DANCEresponse of the important organs was determined.2. The kidney, liver, skin and brain tissue cells of different age of mice (juvenile,adult and aged mice) were separated by DSGC, and DANCE was determined andanalyzed for its relation to the age of the animals.3. All-trans retinoic acid (RA) was used to determine the impact of drug on DANCEresponse of tissue cells in vivo. The time course and dose dependence of RA forDANCE were determined.4. Real-time PCR and Western Blot were used to determine the mRNA and proteinlevels of some important genes in the cells of different bands, and moleculardifference between the bands was analyzed.5. The effects of Ethanol, RA, and5-fluorouracil (5-FU) on K562cells, LO2cellsand SW1990cells were anlysised, respectively. The minimum DANCEconcentration in K562cells, LO2cells and SW1990cells also were determined,respectively.6. Atomic absorption spectrophotometer was used to determine the contents ofpotassium, calcium, phosphorus and magnesium ion in K562cells. The differenceof the ion contents between bands S50and S60was compared.7. The water content analysis of S50and S60bands in K562cells found that the water content of S50band was higher than S60band, which miply that S50bandintracellular density less than S60band.Results:1．At room temperature (25°C) and low temperature (4°C), mouse tissue cells wereseparated into3-5bands by DSGC at different times after sacrifice, and DANCEevidently occurred in the important organs specially the liver, stomach and brain.2．The kidney, liver, skin and brain tissue cells of different age of mice (juvenile,adult and aged mice) exhibited DANCE significantly, especially in the liver, skinand brain, suggesting that age of the animals might play important roles inDANCE of the tissue cells.3．RA garaged to mice was found to have significant impacts on DANCE in thetissue cells of mice, showing evident changes in the DSGC bands. It was foundthat RA exhibited good time course and dose dependence.4．The results of real-time PCR and Western Blot showed that the mRNA andprotein levels of some important genes in the cells of different bands weredifferent from each other, suggesting the cells of different bands were molecularlydifferent from each other.5．On drug condition, those cell lines all appeared DANCE up phenomenon. theminimum inhibitory concentration of RA on K562cells, LO2cells and SW1990cells were4、8、12ug/ml,5-FU were40、12.5、25ug/ml, and ethanol were8000、8000、16000ug/ml, respectively.6．The results of atomic absorption spectrophotometric analysis showed that thecontents of potassium, calcium, phosphorus and magnesium ion varied withdifferent bands, higher in S50than in S60of K562cells, suggesting that ion andwater contents increased in the cells of S50, which might be a cause for thedecrease of intracellular density in S50.7．Water content was found to be lower in S60than in S50of K562cells, andtherefor, the mass density in S60was higher than S50, suggesting that DANCEmight be caused mainly by the change of mass density in the cells. Conclusion： DANCE occurred in mouse tissue cells, and the DANCE response intissue cells exhibited good time course and dose dependence. Change of water contentin cells might be the biochemical mechanism of DANCE.