Objective： To assess if arachnoid cells have the capability to present antigen and activate T-lymphocytes after stimulation by bloody cerebrospinal fluid （CSF）, and to illuminate the mechanism of coagulation-initia...Objective： To assess if arachnoid cells have the capability to present antigen and activate T-lymphocytes after stimulation by bloody cerebrospinal fluid （CSF）, and to illuminate the mechanism of coagulation-initiated inflammation in the subarachnoid space after subarachnoid hemorrhage （SAH）. Methods： Arachnoid cells were cultured, characterized, and examined by immunofluorescence for the basal expression of human leukocyte antigen-DR （HLA-DR）, Expression of HLA-DR, after co-culturing arachnoid cells in vitro with bloody CSF, was investigated by immunofluorescence and flow cytometry （FCM）. The variation of arachnoid cells＇ ultrastructure was observed by transmission electron microscope （TEM）. Arachnoid cells were co-cultured with peripheral blood mononuclear cells （PBMCs）. The content of soluble interleukin-2 receptor （slL-2r） in culture medium was detected by enzyme-linked immunosorbent assay （ELISA）. Results： （1） Arachnoid cells were successfully cultured for many passages. The immunofluorescent staining was positive for HLA-DR in over 95% of the human arachnoid cells. The punctate HI_A-DR was distributed in cytoplasm and not in the karyon. （2） After co-culturing arachnoid cells in vitro with bloody CSF, numerous particles with strong fluorescence appeared in the cytoplasm on Day 6. On Day 8, the quantity of particles and fluorescent intensity were maximal. FCM showed that the percentage of HLA-DR expressing cells was （2.5＋_0.4）% at the first 5 d, increasing to （60.8＋_3.6）% on Day 7. （3） After co-culturing arachnoid cells in vitro with bloody CSF, many lysosome and secondary lysosome particles were present in the cytoplasm. Hyperplasia of rough endoplasmic re- ticulum and enlarged cysts were observed, with numerous phagocytizing vesicles also observed at the edge of the arachnoid cells. （4） Arachnoid cells stimulated by bloody CSF were co-cultured in vitro with PBMCs. The content of slL-2r in the culture medium, having been maintained at around 1.30 ng/ml during the first 3 d, had increased by Day 4. The content of slL-2r peaked 7.53 ng/ml on Day 7 and then reduced gradually. Conclusions： （1） Basic HLA-DR expression is present in arachnoid cells. （2） After stimulation by bloody CSF, arachnoid cells have the potential to serve as antigen presenting cells （APCs） and the ability to activate T-lymphocytes, indicating that arachnoid cells are involved in the mechanism of coagulation-initiated inflammation in the subarachnoid space after SAH.展开更多
基金Project supported by the National Natural Science Foundation of China (No. 30370497)the Science and Technology Research Program of Zhejiang Province,China (No. 2007-C-33039)
文摘Objective： To assess if arachnoid cells have the capability to present antigen and activate T-lymphocytes after stimulation by bloody cerebrospinal fluid （CSF）, and to illuminate the mechanism of coagulation-initiated inflammation in the subarachnoid space after subarachnoid hemorrhage （SAH）. Methods： Arachnoid cells were cultured, characterized, and examined by immunofluorescence for the basal expression of human leukocyte antigen-DR （HLA-DR）, Expression of HLA-DR, after co-culturing arachnoid cells in vitro with bloody CSF, was investigated by immunofluorescence and flow cytometry （FCM）. The variation of arachnoid cells＇ ultrastructure was observed by transmission electron microscope （TEM）. Arachnoid cells were co-cultured with peripheral blood mononuclear cells （PBMCs）. The content of soluble interleukin-2 receptor （slL-2r） in culture medium was detected by enzyme-linked immunosorbent assay （ELISA）. Results： （1） Arachnoid cells were successfully cultured for many passages. The immunofluorescent staining was positive for HLA-DR in over 95% of the human arachnoid cells. The punctate HI_A-DR was distributed in cytoplasm and not in the karyon. （2） After co-culturing arachnoid cells in vitro with bloody CSF, numerous particles with strong fluorescence appeared in the cytoplasm on Day 6. On Day 8, the quantity of particles and fluorescent intensity were maximal. FCM showed that the percentage of HLA-DR expressing cells was （2.5＋_0.4）% at the first 5 d, increasing to （60.8＋_3.6）% on Day 7. （3） After co-culturing arachnoid cells in vitro with bloody CSF, many lysosome and secondary lysosome particles were present in the cytoplasm. Hyperplasia of rough endoplasmic re- ticulum and enlarged cysts were observed, with numerous phagocytizing vesicles also observed at the edge of the arachnoid cells. （4） Arachnoid cells stimulated by bloody CSF were co-cultured in vitro with PBMCs. The content of slL-2r in the culture medium, having been maintained at around 1.30 ng/ml during the first 3 d, had increased by Day 4. The content of slL-2r peaked 7.53 ng/ml on Day 7 and then reduced gradually. Conclusions： （1） Basic HLA-DR expression is present in arachnoid cells. （2） After stimulation by bloody CSF, arachnoid cells have the potential to serve as antigen presenting cells （APCs） and the ability to activate T-lymphocytes, indicating that arachnoid cells are involved in the mechanism of coagulation-initiated inflammation in the subarachnoid space after SAH.
