OBJECTIVETo investigate the morphology alterations and proteomics changes in primary astrocytes following fluid percussion injury.

METHODSPrimary cultures of astrocytes were prepared from cerebral hemispheres of 1-3 d-old SD rats, then, astrocytes were randomly divided into control group and injury group which were subjected to (0.2 +/- 0.01) MPa fluid percussion injury. The changes of protein expression pattern in astrocytes between injury and control groups were monitored with two dimensional gel electrophoresis.

RESULTSAstrocytes' s abnonmalities of morphology after injury were apparent. The fluid percussion injury caused astrocytes edema, shrinkage, cell junction disconnection and necrosis at 2 h after injury. 24 h and 48 h after injury, most part of astrocytes's dendrites and soma became hypertrophy and showed a higher rate of cell proliferation. The dynamic proteomics changes were identified and total different 13 spots were detected in this study from the 2DE gels. The different displayed 5 spots were identified via MALDI-TOF: cofilin 1, destrin, phosphoglycerate mutase 1, NADH dehydrogenase (ubiquinone) 1 alpha subcomplex 10, annexin 1.

CONCLUSIONThe obvious alteration of morphology and protein expression pattern in primary cultured astrocytes could be induced after fluid percussion injury. The differential proteins detected were probably related to stress responses.

Animals ; Animals, Newborn ; Astrocytes ; cytology ; metabolism ; Brain Injuries ; metabolism ; physiopathology ; Cerebral Cortex ; cytology ; Female ; Male ; Primary Cell Culture ; methods ; Proteins ; metabolism ; Proteome ; analysis ; Proteomics ; methods ; Rats ; Rats, Sprague-Dawley ; Stress, Physiological ; physiology

OBJECTIVETo establish differential proteomics profiles of glioblastoma cell lines from Chinese, and to provide reference for future basic studies.

METHODSTotal protein was extracted from 3 glioblastoma cell lines, CHG-5, TJ899 and TJ905. After normalization, the total protein was presented by two-dimensional (2D) electrophoresis, scanned and analyzed. Some of the identified protein spots were verified by immunocytochemistry of cell lines and immunohistochemistry of solid tumors. The glia cells were used as the control throughout the study.

RESULTSA total of 13 differential protein spots were selected, and eventually 10 were identified as unique proteins. These 10 proteins were involved in cytoskeleton forming, cellular metabolism, tumor migration, stress and inflammatory reaction. Immunocytochemistry and immunohistochemistry further confirmed these proteins present in the solid tumors.

CONCLUSIONDistinct differential proteomics profiles exist in glioblastoma cell lines and normal glia cells, likely related to the transformation of normal glia to glioma.

Brain Neoplasms ; genetics ; metabolism ; Cathepsin D ; metabolism ; Cell Line, Tumor ; Gene Expression Profiling ; Glial Fibrillary Acidic Protein ; metabolism ; Glioblastoma ; genetics ; metabolism ; Humans ; Microfilament Proteins ; metabolism ; Neuroglia ; metabolism ; Proteomics ; methods

OBJECTIVETo study the therapeutic efficacy of siRNA fragments silencing p75 neurotrophin receptor (p75(NTR)), which may be a key regulator of glioma cell apoptosis and invasion.

METHODSThe siRNA sequence fragments targeting p75(NTR) were designed and transferred into human glioma cell line U251. RT-PCR and immunocytochemistry method were used to explore the expression of p75(NTR) mRNA and protein. Cell adhesion assay was employed to detect cellular adhesion ability, and soft agar clone formation assay was adopted to identify oncogenicity, and a U251 glioma model was established in nude mice. The intracranial tumor volume was detected by MRI. The expression of p75(NTR), NGF and cyclin D2 were identified using immunohistochemistry. Cell apoptosis was detected by apoptosis kit in situ.

RESULTSThe siRNA fragments targeting p75(NTR) were capable of decreasing mRNA and protein expression of p75(NTR) in U251 glioma cell line. Both the cellular adhesion ability and oncogenicity were weakly relevant. The p75(NTR) expression level was negatively correlated with cyclin D2 and apoptosis, and positively correlated with NGF expression. The siRNA sequence fragments targeting p75(NTR) were effective in decreasing the gross volume of tumor; prolonged the survival time of mice, and the edge of tumor was much sharper than that of the control group.

CONCLUSIONSThe gene silencing technique by siRNA targeting p75(NTR) is capable of decreasing tumor invasion and cell proliferation as well as inducing cell apoptosis. It is expected to be a new choice for glioma gene therapy.

Animals ; Apoptosis ; Brain Neoplasms ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Cyclin D2 ; metabolism ; Gene Silencing ; Glioma ; genetics ; metabolism ; pathology ; Humans ; Male ; Mice ; Mice, Nude ; Neoplasm Invasiveness ; Neoplasm Transplantation ; Nerve Growth Factor ; metabolism ; RNA, Messenger ; metabolism ; RNA, Small Interfering ; genetics ; Random Allocation ; Receptor, Nerve Growth Factor ; genetics ; metabolism