Isolation, study of activity, and characteristics of an immunosuppressive liver factor causing apoptosis of EL-4 thymoma cells in vitro

Immunosuppressive factor (ISFnp) which inhibits proliferation and viability of thymoma EL-4 cells was isolated from mouse liver. The testing procedure based on the biotransformation of MTT-tetrazolium by mitochondrial enzymes of viable cells allowed us to purify this factor as individual peak of protein, that allowed to obtain polyclonal rabbit antibodies to this factor. By the methods of double immunodiffusion, gel-filtration and SDS-PAGE with subsequent immunoblotting we shown that this factor specifically localized in liver and consists two subunits of 40 and 42 kDa which form dimers with apparent M(r) about 70-80 kDa. This factor induced olygonucleosomal DNA cleavage in EL-4 cells in vitro similar to dexamethasone-induced DNA-degradation in thymocytes. This cleavage preceded to lysis of EL-4 cells assessed by 51Cr-release, that strongly suggested an involvement of apoptosis in cell death mechanism. ISFnp strongly inhibited blast-transformation and proliferation in MLC-responses to mutant MHC class 2 molecule. This effect was not due to deletion of allo-reactive clones, because removing of this factor from MLC cultures treated with one for 4 days resulted in blast-transformation without any reduction of the number of viable cells as well as their capacity for secondary responses to the same antigen as compared with control cultures.     

Electron-microscopic radioautography of DNA-synthesizing cells of growing rat smooth muscle tissue

DNA-synthesizing cells of rat stomach muscle tissue following 50% resection of the fundal part, identical cells of the vena cava after disturbance of the blood out flow, and of the appendix on contraction of the ascending part of the large intestine were studied by electron microscopic autoradiography. DNA synthesis in the differentiated myocyte nuclei of the stomach muscle tissue, and of the "activated" myocytes of the stomach and the vein was observed. There were signs of asynchronous DNA synthesis in the nuclei of some smooth muscle cells and fibroblasts.