OBJECTIVE: To study whether recombinant adeno-associated virus (rAAV) mediated foreign gene, LacZ, could pass the blood brain barrier by intra-carotid artery delivery and express in vivo in ischemic brain of the foc...OBJECTIVE: To study whether recombinant adeno-associated virus (rAAV) mediated foreign gene, LacZ, could pass the blood brain barrier by intra-carotid artery delivery and express in vivo in ischemic brain of the focal embolic stroke rats to investigate a possibility of delivering foreign gene through carotid artery to treat acute ischemic stroke. METHODS: The carotid artery territory in 41 rats was embolized with or without arterial-like fibrin rich clots to make a model of focal embolic stroke rat. rAAV containing LacZ gene (rAAV-LacZ) was constructed in 293 cells by calcium phosphate cotransfection. The rats were assigned to one of the following treatments: 1 control (without embolism) groups, including PBS treated (n = 6), pLacZ treated (n = 6 ) and rAAV-LacZ treated (n = 6): 2 embolic groups, including embolism + PBS (n =7),embolism + pLacZ (n = 8) and embolism + rAAV-LacZ (n = 8). Brains were cryosectioned and kappa-Gal stain was performed at 2, 4, and 8 weeks, respectively, after transfection, and then infarct volume was measured and the percentage of LacZ staining-positive cells was calculated. RESULTS: In all the control groups and embolism + PBS treated animal, no kappa-Gal staining-positive cells were found, but in embolism + pLacZ (n = 8) and embolism+rAAV-LacZ groups a lot of kappa-Gal staining-positive cells were found. The expression cells were in the tissues around the infarction. The gene expression persisted only nearly four weeks in embolic group with pLacZ. In the embolic group with rAAV-LacZ the expression was very stable during the experiment course (eight weeks) and the percentage of the expressed cells was significantly higher than that of its contralateral areas at the same time points, respectively. CONCLUSIONS: The plasmid vector and rAAV could enter the brain through the ischemia-damaged blood barrier and foreign gene can be expressed in brain. The positive gene expression is mainly in the peripheral areas of the infarction. rAAV as a permanent expression vector may ultimately be us展开更多
OBJECTIVE: To verify whether the extracellular domain of kinase domain region (KDR) has anti-angiogenesis activity in vivo. METHODS: cDNA was cloned into adeno-associated virus (AAV) vector pSNAV and transfected to ba...OBJECTIVE: To verify whether the extracellular domain of kinase domain region (KDR) has anti-angiogenesis activity in vivo. METHODS: cDNA was cloned into adeno-associated virus (AAV) vector pSNAV and transfected to baby hamster kidney (BHK) cells. Recombinant AAV was obtained from the cell culture supernatant after adding helper virus. Recombinant AAV-infected human bladder cancer EJ cell line (EJ cells) were injected subcutaneously into Balb-c nude mice. Tumor specimens were removed from the mice, paraffin-embedded and sliced, then stained by immunohistochemistry. Microvessel density (MVD) was determined under a microscope. RESULTS: The tumor volume developed by EJ cells transfected with the extracellular domain of KDR was significantly smaller (1.70 +/- 0.18 cm(3)) compared with that in the control (5.62 +/- 0.67 cm(3)) (P展开更多
基金ThisstudywassupportedbythegrantforKeyProjectofNationalNaturalScienceFoundationinChina (No39730170)andthegrantforGeneralProjectofNSFC (No 3 9770 810 )
文摘OBJECTIVE: To study whether recombinant adeno-associated virus (rAAV) mediated foreign gene, LacZ, could pass the blood brain barrier by intra-carotid artery delivery and express in vivo in ischemic brain of the focal embolic stroke rats to investigate a possibility of delivering foreign gene through carotid artery to treat acute ischemic stroke. METHODS: The carotid artery territory in 41 rats was embolized with or without arterial-like fibrin rich clots to make a model of focal embolic stroke rat. rAAV containing LacZ gene (rAAV-LacZ) was constructed in 293 cells by calcium phosphate cotransfection. The rats were assigned to one of the following treatments: 1 control (without embolism) groups, including PBS treated (n = 6), pLacZ treated (n = 6 ) and rAAV-LacZ treated (n = 6): 2 embolic groups, including embolism + PBS (n =7),embolism + pLacZ (n = 8) and embolism + rAAV-LacZ (n = 8). Brains were cryosectioned and kappa-Gal stain was performed at 2, 4, and 8 weeks, respectively, after transfection, and then infarct volume was measured and the percentage of LacZ staining-positive cells was calculated. RESULTS: In all the control groups and embolism + PBS treated animal, no kappa-Gal staining-positive cells were found, but in embolism + pLacZ (n = 8) and embolism+rAAV-LacZ groups a lot of kappa-Gal staining-positive cells were found. The expression cells were in the tissues around the infarction. The gene expression persisted only nearly four weeks in embolic group with pLacZ. In the embolic group with rAAV-LacZ the expression was very stable during the experiment course (eight weeks) and the percentage of the expressed cells was significantly higher than that of its contralateral areas at the same time points, respectively. CONCLUSIONS: The plasmid vector and rAAV could enter the brain through the ischemia-damaged blood barrier and foreign gene can be expressed in brain. The positive gene expression is mainly in the peripheral areas of the infarction. rAAV as a permanent expression vector may ultimately be us
文摘OBJECTIVE: To verify whether the extracellular domain of kinase domain region (KDR) has anti-angiogenesis activity in vivo. METHODS: cDNA was cloned into adeno-associated virus (AAV) vector pSNAV and transfected to baby hamster kidney (BHK) cells. Recombinant AAV was obtained from the cell culture supernatant after adding helper virus. Recombinant AAV-infected human bladder cancer EJ cell line (EJ cells) were injected subcutaneously into Balb-c nude mice. Tumor specimens were removed from the mice, paraffin-embedded and sliced, then stained by immunohistochemistry. Microvessel density (MVD) was determined under a microscope. RESULTS: The tumor volume developed by EJ cells transfected with the extracellular domain of KDR was significantly smaller (1.70 +/- 0.18 cm(3)) compared with that in the control (5.62 +/- 0.67 cm(3)) (P
