OBJECTIVETo construct a eukaryotic expression vector for alpha-1-antitrypsin (AAT) and detect its expression and localization in NIH 3T3 cells.

METHODSThe total RNA was extracted from the liver tissue of BALB/c mice, and the corresponding coding sequences for mouse AAT (GenBank accession No. NM_009243) were amplified by RT-PCR and cloned into hemagglutinin (HA)-tagged vector pcDNA3-HA. The construct was then transfected into NIH 3T3 cells, which were observed under fluorescence microscope.

RESULTSThe recombinant plasmid was verified by PCR, enzyme digestion and sequence analysis, and the fusion protein was highly expressed in NIH 3T3 cells. Under fluorescence microscope, the fusion protein was found to distribute mainly in the cytoplasm.

CONCLUSIONThe expression vector for AAT-HA fusion protein has been successfully constructed and effectively expressed in mammalian cells to allow future functional study of AAT in mammalian cells.

Animals ; Genetic Vectors ; biosynthesis ; genetics ; Hemagglutinins ; genetics ; metabolism ; Mice ; Mice, Inbred BALB C ; NIH 3T3 Cells ; Plasmids ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; alpha 1-Antitrypsin ; biosynthesis ; genetics

Malignant fibrous histiocytoma(MFH) is a rare primary neoplasm that constitutes less than 1% of the malignant tumors of bone, and involvement of the skull is very rare. We present a case of malignant fibrous histiocytoma of the skull, presenting an intraosseous lesion in a 43-yr-old woman. She had a rapidly growing, tender mass in the right parietal region. A plain radiograph showed an osteolytic lesion of the right parietal bone. Magnetic resonance imaging revealed that the lesion showed heterogeneous low signal intensity on T1-weighted images and slightly high signal intensity on T2-weighted images. No evidence of an extraosseous extension to the adjacent dura and soft tissue was found, and a wide excision of the parietal bone was performed. Histologically, the tumor was a typical MFH displaying pleomorphic spindle cells in a storiform pattern. The results of immunohistochemical stainings revealed that the tumor cells were positive for vimentin, alpha-1-antitryp-sin, and p53, and negative for smooth muscle actin, S100 protein, desmin, and MyoD1. Three months later, a mainly cystic, recurrent mass was developed at the previously operated site. Before the resection, we first performed the percutaneous aspiration cytology, revealing diagnostic multinucleated pleomorphic cells. There-after, she had to receive repetitive resections of recurrent or residual lesions, and she died of postoperative meningoencephalitis two years after the first operation.
Actins/biosynthesis ; Adult ; Brain/pathology ; Desmin/biosynthesis ; Female ; Giant Cells/metabolism ; Histiocytoma, Fibrous/*diagnosis ; Human ; Immunohistochemistry ; Magnetic Resonance Imaging ; Mitosis ; Muscle, Smooth/metabolism ; MyoD Protein/biosynthesis ; Protein p53/biosynthesis ; S100 Proteins/biosynthesis ; Skull Neoplasms/*diagnosis ; Tomography, X-Ray Computed ; Vimentin/biosynthesis ; alpha 1-Antitrypsin/biosynthesis