Mouse thymic stromal cell line 4 (MTSC4) is one of the stromal cell lines established in our laboratory. While losing the characteristics of epithelial cells, they express some surface markers shared with thymic dendr...Mouse thymic stromal cell line 4 (MTSC4) is one of the stromal cell lines established in our laboratory. While losing the characteristics of epithelial cells, they express some surface markers shared with thymic dendritic cells (TDCs). To further study the biological functions of these cells, we compared the capability of MTSC4 with TDCs in the induction of thymocyte apoptosis, using thymic reaggregation culture system. Apoptosis of thymocytes induced by MTSC4 and TDCs was measured by Annexin V and PI staining and analyzed by flow cytometry. We found that MTSC4 selectively augmented the apoptosis of CD4^+8^+ (DP) thymocytes. This effect was Fas/FasL independent and could not be blocked by antibodies to MHC class Ⅰ and class Ⅱ molecules. In addition, MTSC4 enhanced the apoptosis of DP thymocytes from different strains of mice, which implies that MTSC4-induced thymocyte apoptosis is not mediated by the TCR recognition of self peptide/MHC molecules. In contrast to MTSC4, thymocyte apoptosis induced by TDCs was MHC-restricted. Thus, MHC-independent fashion of stromal-DP thymocyte interaction may be one of the ways to induce thymocyte apoptosis in thymus. Our study has also shown that the interaction of MTSC4 stromal cells and thymocytes is required for the induction of thymocyte apoptosis.展开更多
Apoptosis of thymocytes is involved in the negative selection of thymus, but it remains unclear how the cell death of thymocytes is regulated by the thymic stromal cells. By using immunohistochemistry, TdT mediated dU...Apoptosis of thymocytes is involved in the negative selection of thymus, but it remains unclear how the cell death of thymocytes is regulated by the thymic stromal cells. By using immunohistochemistry, TdT mediated dUTP nick end labeling (TUNEL) and flow cytometry methods, it was found that Fas ligand was expressed in the thymic medulla and a mouse thymic medullary type dendritic cell line (MTSC4). The DNA fragmentation and TUNEL positive staining of thymocytes were detected in 6 h of cocultures with MTSC4. 97% of the thymocytes which bound to MTSC4 were Fas antigen positive cells. The DNA fragmentation of thymocytes induced by MTSC4 was inhibited by the addition of 20 mmol/L N acetyl galactosamin monosaccharide and the pretreatment of a monoclonal antibody (PF 18 3) which recognized a putative antigen on MTSC4. These results suggested that the mechanisms of induction of cell death by stromal cells may include the interactions of multiple cell surface molecules, in addition to the Fas/Fas ligand system.展开更多
Using terminal deoxynucleotide transferase mediated dUTP digoxigenin nick end labeling (TUNEL) assay and propidium iodide DNA staining flow cytometry assay, the effects of mouse thymic dendritic cells (MTSC4) on the p...Using terminal deoxynucleotide transferase mediated dUTP digoxigenin nick end labeling (TUNEL) assay and propidium iodide DNA staining flow cytometry assay, the effects of mouse thymic dendritic cells (MTSC4) on the process of programmed cell death of thymocytes in vitro were investigated. It was noticed that thymocytes bound to MTSC4 used in this study. That the percentages of apoptotic nuclei of the bound thymocytes on MTSC4 were much higher than those of medium cultured thymocytes, while the bound thymocytes on mouse thymic epithelial cell (MTEC1) showed much lower percentages of apoptosis. FACS analysis quantitatively confirmed the observation. Phenotype analysis showed that MTSC4 induced the deletion of CD4+CD8+ cells and CD4+CD8 cells in 18 h of coculture. The results suggest that the negative selection of medullary thymocytes may be achieved by thymic dendritic cells through their enhancing effects on apoptosis.展开更多
A murine CD4+ thymocyte subset with phenotype of TCRαβ + 3G11- 6C10- CD4 + CD8- CD69 + /- HSAmed/lo contains the cells in relatively functional matured status. The functional property of the cells in this subset is ...A murine CD4+ thymocyte subset with phenotype of TCRαβ + 3G11- 6C10- CD4 + CD8- CD69 + /- HSAmed/lo contains the cells in relatively functional matured status. The functional property of the cells in this subset is characterized by the unique pattern of cytokine production at transitional stage from ThO to Th2 type with the latter being the dominant type. After being co-cultured with murine thymic medullary epithelial cell line (MTEC1) cells, a murine展开更多
基金This work was funded by grants from Major State Bascic Research Development Program of China(No.G1999053904)National Natural Science Foundation of China(No.30271200)Young Scentist Cooperation Research Fund(No.39928019).
文摘Mouse thymic stromal cell line 4 (MTSC4) is one of the stromal cell lines established in our laboratory. While losing the characteristics of epithelial cells, they express some surface markers shared with thymic dendritic cells (TDCs). To further study the biological functions of these cells, we compared the capability of MTSC4 with TDCs in the induction of thymocyte apoptosis, using thymic reaggregation culture system. Apoptosis of thymocytes induced by MTSC4 and TDCs was measured by Annexin V and PI staining and analyzed by flow cytometry. We found that MTSC4 selectively augmented the apoptosis of CD4^+8^+ (DP) thymocytes. This effect was Fas/FasL independent and could not be blocked by antibodies to MHC class Ⅰ and class Ⅱ molecules. In addition, MTSC4 enhanced the apoptosis of DP thymocytes from different strains of mice, which implies that MTSC4-induced thymocyte apoptosis is not mediated by the TCR recognition of self peptide/MHC molecules. In contrast to MTSC4, thymocyte apoptosis induced by TDCs was MHC-restricted. Thus, MHC-independent fashion of stromal-DP thymocyte interaction may be one of the ways to induce thymocyte apoptosis in thymus. Our study has also shown that the interaction of MTSC4 stromal cells and thymocytes is required for the induction of thymocyte apoptosis.
文摘Apoptosis of thymocytes is involved in the negative selection of thymus, but it remains unclear how the cell death of thymocytes is regulated by the thymic stromal cells. By using immunohistochemistry, TdT mediated dUTP nick end labeling (TUNEL) and flow cytometry methods, it was found that Fas ligand was expressed in the thymic medulla and a mouse thymic medullary type dendritic cell line (MTSC4). The DNA fragmentation and TUNEL positive staining of thymocytes were detected in 6 h of cocultures with MTSC4. 97% of the thymocytes which bound to MTSC4 were Fas antigen positive cells. The DNA fragmentation of thymocytes induced by MTSC4 was inhibited by the addition of 20 mmol/L N acetyl galactosamin monosaccharide and the pretreatment of a monoclonal antibody (PF 18 3) which recognized a putative antigen on MTSC4. These results suggested that the mechanisms of induction of cell death by stromal cells may include the interactions of multiple cell surface molecules, in addition to the Fas/Fas ligand system.
文摘Using terminal deoxynucleotide transferase mediated dUTP digoxigenin nick end labeling (TUNEL) assay and propidium iodide DNA staining flow cytometry assay, the effects of mouse thymic dendritic cells (MTSC4) on the process of programmed cell death of thymocytes in vitro were investigated. It was noticed that thymocytes bound to MTSC4 used in this study. That the percentages of apoptotic nuclei of the bound thymocytes on MTSC4 were much higher than those of medium cultured thymocytes, while the bound thymocytes on mouse thymic epithelial cell (MTEC1) showed much lower percentages of apoptosis. FACS analysis quantitatively confirmed the observation. Phenotype analysis showed that MTSC4 induced the deletion of CD4+CD8+ cells and CD4+CD8 cells in 18 h of coculture. The results suggest that the negative selection of medullary thymocytes may be achieved by thymic dendritic cells through their enhancing effects on apoptosis.
文摘A murine CD4+ thymocyte subset with phenotype of TCRαβ + 3G11- 6C10- CD4 + CD8- CD69 + /- HSAmed/lo contains the cells in relatively functional matured status. The functional property of the cells in this subset is characterized by the unique pattern of cytokine production at transitional stage from ThO to Th2 type with the latter being the dominant type. After being co-cultured with murine thymic medullary epithelial cell line (MTEC1) cells, a murine
