Association between vascular access failure and microparticles in hemodialysis patients.

BACKGROUND: Vascular access failure, a major cause of morbidity in hemodialysis (HD) patients, occurs mainly at stenotic endothelium following an acute thrombotic event. Microparticles (MPs) are fragments derived from injured cell membrane and are closely associated with coagulation and vascular inflammatory responses. METHODS: We investigated the relationship between levels of circulating MPs and vascular access patency in HD patients. A total of 82 HD patients and 28 healthy patients were enrolled. We used flow cytometry to measure endothelial MPs (EMPs) identified by CD31+CD42- or CD51+ and platelet-derived MPs (PMPs) identified by CD31+CD42+ in plasma samples of participants. Vascular access patency was defined as an interval from the time of access formation to the time of first access stenosis in each patient. MP counts were compared according to access patent duration. RESULTS: The levels of EMP (both CD31+CD42- and CD51+) and CD31+CD42+PMP were significantly higher in patients than in healthy participants. Levels of CD31+CD42-EMP and CD31+CD42+PMP showed a positive correlation. In non-diabetic HD patients, CD31+CD42-EMPs and CD31+CD42+PMPs were more elevated in the shorter access survival group (access survival <1 year) than in the longer survival group (access survival ≥ 4 years). CONCLUSION: Elevated circulating EMP or PMP counts are influenced by end-stage renal disease and increased levels of EMP and PMP may be associated with vascular access failure in HD patients.

Investigation of molecular factors associated with malignant transformation of oligodendroglioma by proteomic study of a single case of rapid tumor progression.

PURPOSE: Frozen tumor tissues from a patient who showed rapid progression to anaplastic oligodendroglioma after near total resection of oligodendroglioma were used to examine differential expression of proteins to gain better understanding of the pathogenesis of malignant transformation. METHODS: We have determined their protein profiles using a 2D gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry approach. RESULTS: Among 23 differentially expressed spots, overexpression of peroxiredoxin 6 and underexpression of rho GDP dissociation inhibitor alpha were confirmed to be valid after western blot and immunocytochemical analysis of oligodendroglioma tissue. CONCLUSIONS: Abnormal expression of peroxiredoxin 6 and rho GDP dissociation inhibitor alpha may be associated with malignant transformation in oligodendroglioma and these proteins might be candidates of molecular predictive factors.

Radiation-induced cerebellar glioblastoma at the site of a treated medulloblastoma: case report.

Radiation-induced glioblastoma multiforme (GBM) is a rare complication of radiotherapy. The authors report such a case occurring 10 years after treatment of cerebellar medulloblastoma. The patient was a 15-year-old boy who had undergone a gross-total removal of a medulloblastoma and received radiation therapy at the age of 5 years. He had experienced no tumor recurrences for 10 years until a new enhancing mass was found at the original site of the medulloblastoma. Following its resection the new lesion was found to be a GBM and there was no evidence of a medulloblastoma. The second tumor developed at the same site as the previous one after a sufficient latent period and fulfilled the criteria for a radiation-induced neoplasm. The original tumor cells expressed synaptophysin without p53 overexpression, a characteristic feature of medulloblastomas. In contrast, cells from the later tumor expressed glial fibrillary acidic protein and p53 but not synaptophysin. A sequence analysis of the p53 gene showed deletion at codon 233 and a C to G transition at codon 278 in the GBM but no mutation in the medulloblastoma. A GBM specimen revealed no amplification of the epidermal growth factor receptor compared with a normal control specimen. In conclusion, the clinical features of a radiation-induced GBM are similar to that of the primary GBM, whereas its genetic alterations render it a secondary GBM. These findings indicate that radiation-induced GBM should be considered a distinct clinical entity.

Overexpression of cyclooxygenase-2 in childhood ependymomas: role of COX-2 inhibitor in growth and multi-drug resistance in vitro.

The management of ependymomas remains one of the most frustrating issues in pediatric neuro-oncology. Gross total resection is not always possible, and intensive chemotherapy and craniospinal radiotherapy have made no clear advances. Moreover, the chemoresistance of ependymomas may be explained by the expression of membrane transport molecule P-glycoprotein (P-gp). In this study the expression of cyclooxygenase 2 (COX-2) and the role of the specific COX-2 inhibitor NS-398 in growth and multi-drug resistance of ependymomas were investigated. COX-2 protein expression was assessed in 19 ependymomas immunohistochemically, and the effect of NS-398 on growth and multi-drug resistance was investigated using two primary cultured ependymoma cell lines. COX-2 protein expression was observed in 15 (79%) of the 19 ependymomas. NS-398 was found to reduce the proliferation of monolayer cell cultures in a dose- and time-dependent manner and to induce apoptosis and lower bcl-2 protein levels. After NS-398 treatment, Western blotting showed reduced P-gp expression and a rhodamine 123 efflux assay demonstrated a significant decrease in P-gp activity. Our findings demonstrate that COX-2 is overexpressed in ependymomas and that NS-398 is able to induce apoptosis and suppress P-gp expression and activity.

Gene expression profile analyses of cortical dysplasia by cDNA arrays.

Cortical dysplasia (CD) is a well-recognized cause of intractable epilepsy, especially in children and is characterized histologically by derangements in cortical development and organization. The objective of this study was to expand the current knowledge of altered gene expression in CD as a first step towards in the identification of additional genes operative in the evolution of CD. Surgical specimens were obtained from eight patients (4 males and 4 females; age range 2-38 years; mean 15 years) with a pathologic diagnosis of CD. Nondysplastic temporal neocortex was obtained from a 2-year-old boy with intractable epilepsy and medial temporal lobe ganglioglioma. After total RNA isolation from frozen brain tissues, we carried out gene expression profiling using a cDNA expression array. Differences in gene expressions between CD and the nondysplastic neocortex were confirmed by semi-quantitative conventional reverse transcription-PCR. Three genes (recombination activating gene 1 (RAG1), heat shock 60 kDa protein 1 (HSP-60), and transforming growth factor beta1 (TGF beta1)) were found to be up-regulated more than two-fold in CD, whereas four genes (phosphoinositide-3-kinase regulatory subunit polypeptide 1 [p85 alpha] (PI3K), frizzled homolog 2 [Drosophila], Bcl-2/adenovirus E1B 19 kDa interacting protein (NIP3), and glia maturation factor beta (GMF beta)) were down-regulated to less than 50% of their normal levels. Interestingly, the majority of genes showing altered expression were associated with apoptosis. Our study demonstrates diverse changes in gene expression in CD. However, it remains to be shown which of these are causally related to the evolution of CD.

Adenoviral p16/CDKN2 gene transfer to malignant glioma: role of p16 in growth, invasion, and senescence.

In gliomas, a high frequency of homozygous p16 gene deletions have been demonstrated, which are believed to be linked with malignant progression. The aim of this study was to assess the role of p16 in growth, invasion, and senescence. The human glioma cell lines U87 MG and U373 MG were transduced with Ad-p16, and cell viability was assessed by trypan blue staining. To examine the mechanism of cell growth inhibition, cell cycle analyses and annexin assays were performed. The invasive potential of Ad-p16 transduced cells was evaluated using a Matrigel invasion assay, and trimolecular complex (MMP-2/MT1-MMP/TIMP-2) synthesis was proven by zymography and Western blotting. To establish the link between p16 and cell senescence, we stained for Senescence-Associated beta-galactosidase activity. A cell proliferation assay demonstrated that Ad-p16 treatment significantly inhibits cell growth. Moreover, this cell growth inhibition was induced by cell cycle arrest, not by apoptosis. In vitro treatment of malignant glioma cells with Ad-p16 significantly decreased their invasive potential by Matrigel invasion assay. However, we were unable to demonstrate any differences in the constitutive productions and secretions of MMP-2, MT1-MMP, and TIMP-2, among the mock-treated, Ad-lacZ-transduced, and Ad-p16-transduced cells. p16 expression caused an enlargement of all cells, and these were morphologically similar to senescent cells. Staining for Senescence-Associated beta-galactosidase activity showed that the enlarged cells stained positively. Taken together these data strongly suggest that the anti-cancer effect of p16 is modulated by p16-mediated cell cycle arrest and by the induction of senescence.

Interaction between p53 and p16 expressed by adenoviral vectors in human malignant glioma cell lines.

OBJECT: Multiple gene replacements have been examined as a potential treatment modality for malignant gliomas. Nevertheless, no reports are available that detail the synergy, additivity, or antagonism of multiple genes. The aim of this study was to assess the interaction between p53 and p16 genes in the growth of glioma cell lines. METHODS: The human glioma cell lines U87MG and U373MG were transduced using an adenoviral vector with Ad-p53, Ad-p16, or both. Western blotting was performed to determine the expression of the protein products of the transduced p53 and p16 genes. To establish whether the combination of Ad-p53 and Ad-p16 would be beneficial, the effects of gene combinations at the median inhibitory concentration level were analyzed using the isobologram method. Annexin assays and cell cycle analyses were performed on the transduced cells. Western blotting demonstrated the expression of p53 and p16 in transduced cells. Simultaneous exposure to Ad-p53 and Ad-p16 produced additive effects in both glioma cell lines. Experimental data points in U373MG lay near the Mode I line, indicating that the vectors had a different mode of action. The restoration of normal p53-encoded protein in the mutant cell lines induced apoptosis, whereas in the wild-type p53 cell lines, the overexpression of wild-type p53 resulted in a moderate degree of apoptosis and G1 arrest. Furthermore, Ad-p16 induced more marked G1 arrest than Ad-p53 in cells with wild-type p53. CONCLUSIONS: The results show that interaction between Ad-p53 and Ad-p16 is additive, regardless of p53 gene status.