The Combined Effect of Gamma Knife Irradiation and p53 Gene Transfection in Human Malignant Glioma Cell Lines

OBJECTIVE: The purpose of this study is to elucidate in vitro responses to combined gamma knife irradiation and p53 gene transfection on human malignant glioma cell lines. METHODS: Two malignant human glioma cell lines, U87MG (p53-wild type) and U373MG (p53-mutant) were transfected with an adenoviral vector containing p53 (MOI of 50) before and after applying 20Gy of gamma irradiation. Various assessments were performed, including, cell viability by MTT assay; apoptosis by annexin assay; and cell cycle by flow cytometry, for the seven groups: mock, p53 only, gamma knife (GK) only, GK after LacZ, LacZ after GK, GK after p53, p53 after GK. RESULTS: Cell survival decreased especially, in the subgroup transfected with p53 after gamma irradiation. Apoptosis tended to increase in p53 transfected U373 MG after gamma irradiation (apoptotic rate, 38.9%). The G2-M phase cell cycle arrest markedly increased by transfecting with p53, 48 hours after gamma knife irradiation in U373 MG (G2-M phase, 90.8%). CONCLUSION: These results suggest that the in vitro effects of combined gamma knife irradiation and p53 gene transfection is an augmentation of apoptosis and G2-M phase cell cycle arrest, which are more exaggerated in U373 MG with p53 transfection after gamma knife irradiation.

Loss of Heterozygosity Analysis of PTEN and DMBT1 Loci in Pediatric Brain Tumors / 대한소아혈액종양학회지

PURPOSE: PTEN and DMBT1, candidate tumor suppressor genes on chromosome 10q, were identified based on deletions in glioblastoma and medulloblastoma cell lines. We examined the occurrences and frequencies of allelic deletions on chromosome 10q23 and 10q25 by loss of heterozygosity (LOH) analysis in 24 pediatric brain tumors to investigate the possible involvement of PTEN and DMBT1 gene deletions in the development of pediatric brain tumors. METHOD: LOH was analyzed by polymerase chain reaction (PCR) of PTEN locus on 10q23 using 2 microsatellite markers, D10S608 and D10S579, and of DMBT1 locus on 10q25-q26.1 using a microsatellite marker, D10S587, in 24 pediatric brain tumor (18 medulloblastomas, 3 ependymomas, 2 glioblastomas and 1 supratentorial primitive neuroectodermal tumor) DNAs extracted from archival tissue specimens (case 1-15, 19) or fresh frozen tissue specimens (case 16-18, 20-24). RESULTS: Allelic deletions were detected in 4 of 23 informative cases (17%) on D10S608, 6 of 24 informative cases (25%) on D10S579, and 8 of 24 informative cases (33%) on D10S587. Overall 11 of 24 cases (46%) showed LOH on chromosome 10q at PTEN or DMBT1 loci, and they were 10 medulloblastomas and 1 ependymoma pathologically. Of 18 medulloblastomas, 7 (39%) exhibited LOH at PTEN locus, 8 (44%) exhibited LOH at DMBT1 locus, and 10 (56%) exhibited LOH at one or both of loci. CONCLUSION: Our results support the notion that PTEN and DMBT1 tumor suppressor gene deletions may be involved in the pathogenesis of pediatric brain tumors. Our results also suggested that PTEN and DMBT1 tumor suppressor gene deletions may not be important in molecular mechanism of glioblastoma development in children as in adults.

Correlation of Clinical and Biological Parameters with Peritumoral Edema in Meningioma

OBJECTIVE: Peritumoral edema(PTE) in meningioma occurs variably and can adversely affect the clinical course. Moreover, the etiology of PTE in meningioma is not well documented. To examine possible correlations with PTE, the authors report an investigation of the clinical parameters and the expressions of vascular endothelial growth factor(VEGF), matrix metalloproteinases(MMPs), and their inhibitors(TIMPs) in 20 meningiomas. METHODS: Tumor volume and edema volume estimation was done using Osiris software with magnetic resonance images and the edema index was calculated. The expression of VEGF, MMP, and TIMP were estimated in all 20 meningiomas by Western blotting, zymography, and laser densitometry. RESULTS: Tumor location was closely related with PTE. Meningiomas of the frontal lobe or the frontotemporal base had large PTEs, whereas those of the occipitoparietal lobe, posterior fossa or petroclivus were small. The level of VEGF expression bore no correlation with histologic malignancy and PTE extent. MMP-2 and -9 were detected in 100% of meningiomas and these levels were significantly related with PTE. TIMP-1 and -2 were detected in 19(95%) and 12(60%) of meningiomas respectively and their presence had no significant correlations statistically with PTE. CONCLUSION: Meningiomas with severe PTE expressed high levels of MMP-9 and low levels of MMP-2. The expressions of VEGF, MMP-2, MMP-9, and TIMPs in meningioma seems to be strongly related with PTE, which might be important factors of the etiology of PTE.

Detection of Hantaan and Seoul Viruses by Reverse Transcriptase-Polymerase Chain Reaction(RT-PCR) and Restriction Fragment Length Polymorphism(RFLP) in Korean Hemorrhagic Fever Patients / 대한신장학회잡지

BACKGROUND: Korean hemorrhagic fever(KHF), a severe from of hemorrhagic fever with renal syndrome(HFRS), is the most common cause of acute renal failure in far east. Two serotypes of hantavirus, Hantaan and Seoul viruses, were identified as pathogens for KHF in Korea. To elucidate the diagnostic applicability for the serotype diagnosis in KHF patients, using a nested reverse transcriptase-PCR and restriction fragment length polymorphism(nRT-PCR /RFLP), we screened 4 prototype viruses, 11 virus isolates from KHF patients, and 69 specimens obtained from 30 KHF patients. METHODS: The nRT-PCR was performed using serotype specific primers for G1 segments for Hantaan(HF3 1140-1163, HB14 1363-1342) and Seoul(SF2 809-832, SB3 1200-1177) viruses. The PCR product was further amplified using nested primers for Hantaan(HF4, 1141-1164, HB13, 1360-1339) and Seoul(SF7 863-884, SB1 1165-1142) viruses. Amplified segments were digested with restriction enzymes specific for either Hantaan(Cla I) or Seoul(Sac I) virus sequences. In all cultured viruses, serotypes identified by nRT-PCR/RFLP were consistent with those of PRNT. RESULTS: In KHF patients, nRT-PCR/RFLP results were compatible with Hantaan virus in 10 patients and with Seoul virus in 13 patients. In 3 patients both Hantaan and Seoul specific amplified bands were visualized in serially collected samples, and in 4 patients no detectable amplicons were detected. Among 69 specimens, 55 specimens obtained from 3 to 33 day of illness were positive. The positive rate was affected by the hospital where specimens were collected, but not by clinical phases, the day of illness, or severity of HFRS. CONCLUSIONS: In general, nRT-PCR/RFLP was a rapid and convenient method for serotype diagnosis in most of the KHF patients. The presented method also make it possible to detect genetic variation of hantavirus within the same serotype. But unlike the viruses in culture, in testing patients' sera, the sensitivity of this methods needed to be improved especially by adequate sample handling.