MicroRNA expression in pediatric intracranial ependymomas and their potential value for tumor grading.

Intracranial ependymoma represents one of the most common pediatric central nervous system malignancies, and exhibits a wide range of clinical behavior from relatively indolent lesions to highly malignant anaplastic ependymomas. Due to the heterogeneous nature of this disease there is lack of prognostic markers, which would reliably predict the outcome of patients. MicroRNAs (miRNAs) have emerged as important molecules in cancer biology during past decade; however, very little is known about their role in ependymomas. The aim of the present study was to evaluate expression of miRNAs in archived formalin-fixed paraffin-embedded (FFPE) samples of pediatric intracranial ependymomas. The expression of miRNAs were examined in 29 samples of ependymoma and we observed that miR-135a-3p, miR-137, miR-17-5p, miR-181d and let-7d-5p were upregulated. In addition, a significantly higher expression of miR-203a was detected in Grade III tumors suggesting its possible use as a prognostic or diagnostic marker. The present study also demonstrated that storage of (FFPE) ependymoma samples for >20 years did not result in a deterioration of miRNAs. The present findings broaden the presently available knowledge regarding miRNA expression in ependymomas and provide further evidence for the employment of miRNA analysis as a supplementary method for the morphological assessment of ependymoma samples.

Mechanical Chest Compressions in Prolonged Cardiac Arrest due to ST Elevation Myocardial Infarction Can Cause Myocardial Contusion.

Acute coronary syndrome is a common cause of sudden cardiac death. We present a case report of a 60-year-old man without a history of coronary artery disease who presented with ST-elevation myocardial infarction. During transportation to the hospital, he developed ventricular fibrillation (VF) and later pulseless electrical activity. Chest compressions with LUCAS 2 (Medtronic, Minneapolis, MN) automated mechanical compression-decompression device were initiated. Coronary angiography showed total occlusion of the left main coronary artery and primary percutaneous coronary intervention (PCI) was performed. After the PCI, his heart started to generate effective contractions and LUCAS could be discontinued. Return of spontaneous circulation was achieved after 90 minutes of cardiac arrest. The patient died of cardiogenic shock 11 hours later. An autopsy revealed a transmural anterolateral myocardial infarction but also massive subepicardial hemorrhage and interstitial edema and hemorrhages on histologic samples from regions of the myocardium outside the infarction itself and also from the right ventricle. These lesions were concluded to be a myocardial contusion. The true incidence of myocardial contusion as a consequence of mechanical chest compressions is not known. We speculate that severe myocardial contusion might have influenced outcome of our patient.

[Cell cultures]. / Tkánové kultury.

Cell or tissue cultures (both terms are interchangeable) represent a complex process by which eukaryotic cells are maintained in vitro outside their natural environment. They have a broad usage covering not only scientific field but also diagnostic one since they represent the most important way of monoclonal antibodies production which are used for both diagnostic and therapeutic purposes. Cell cultures are also used as a "cultivation medium" in virology and for establishing proliferating cells in cytodiagnostics. They are well-established and easy-to-handle models in the area of research, e.g. as a precious source of nucleic acids or proteins. This paper briefly summarizes their importance and methods as well as the pitfalls of the cultivation and new trends in this field.

Changes in MYCN expression in human neuroblastoma cell lines following cisplatin treatment may not be related to MYCN copy numbers.

Neuroblastoma is a tumor accounting for approximately 10% of all childhood malignancies and 50% of all childhood cancer-related deaths. MYCN gene copy number variation represents the most important prognostic factor in neuroblastoma. Prognostic significance of MYCN gene expression is more complicated and may depend on other factors such as MYCN gene copy number status. In the present study, we assessed MYCN gene expression using real-time RT-PCR following cisplatin treatment in three human neuroblastoma cell lines (UKF-NB-3, UKF-NB-4 and SK-N-AS) and their cisplatin-resistant counterparts. We also examined MYCN gene status and copy number (gain and amplification) variations using interphase and metaphase fluorescent in situ hybridization (FISH) and multiplex ligation-dependent probe amplification (MLPA). Only cisplatin-sensitive UKF-NB-4 cells exhibited decreased MYCN expression following treatment with cisplatin. Other sensitive neuroblastoma cells did not exhibit a change in MYCN expression. In contrast, cisplatin-resistant UKF-NB-4 and SK-N-AS cells exhibited increased MYCN expression irrespective of the number of MYCN copies or concentration of cisplatin in the medium. In MYCN-amplified neuroblastoma cells we did not observe any significant change in the number of MYCN copies after cisplatin treatment, whereas MYCN-non-amplified SK-N-AS cells revealed during cisplatin treatment an increased number of MYCN gene copies caused by 2p gain in the majority of cells by FISH. We postulated that cisplatin treatment does not result directly in altered transcription of MYCN. A functional change in MYCN mRNA levels and increased MYCN expression in cisplatin-resistant neuroblastoma cells do not have a clear relationship to MYCN copy numbers. These findings may further contribute to the understanding of cisplatin chemotherapy in connection with MYCN expression, and the possible copy number variations in MYCN neuroblastoma cells may be of importance since targeting of MYCN is being tested as neuroblastoma therapy.

Valproic acid overcomes hypoxia-induced resistance to apoptosis.

Valproic acid (VPA), a histone deacetylase inhibitor (HDACi), has been shown to be an effective tool in cancer treatment. Although its ability to induce apoptosis has been described in many cancer types, the data come from experiments performed in normoxic (21% O2) conditions only. Therefore, we questioned whether VPA would be equally effective under hypoxic conditions (1% O2), which is known to induce resistance to apoptosis. Four neuroblastoma cell lines were used: UKF-NB-3, SK-N-AS, plus one cisplatin-resistant subline derived from each of the two original sensitive lines. All were treated with VPA and incubated under hypoxic conditions. Measurement of apoptosis and viability using TUNEL assay and Annexin V/propidium iodide labeling revealed that VPA was even more effective under hypoxic conditions. We show here that hypoxia-induced resistance to chemotherapeutic agents such as cisplatin could be overcome using VPA. We also demonstrated that apoptosis pathways induced by VPA do not differ between normoxic and hypoxic conditions. VPA-induced apoptosis proceeds through the mitochondrial pathway, not the extrinsic pathway (under both normoxia and hypoxia), since inhibition of caspase-8 failed to decrease apoptosis or influence bid cleavage. Our data demonstrated that VPA is more efficient in triggering apoptosis under hypoxic conditions and overcomes hypoxia-induced resistance to cisplatin. The results provide additional evidence for the use of VPA in neuroblastoma (NBL) treatment.

Expression of class III beta-tubulin in colorectal carcinomas: an immunohistochemical study using TU-20 & TuJ-1 antibody.

BACKGROUND & OBJECTIVE: Expression of class III beta-tubulin represents newly discovered marker of resistance to taxol-based chemotherapy in a wide spectra of carcinomas. However, very little is known about its expression in colorectal carcinomas. This study was done to determine class III beta-tubulin expression in a large series of colonic carcinomas, covering tumours with different degree of differentiation in order to evaluate its prospective significance in resistance to taxol-based chemotherapeutics and to compare the immunostaining profile of two widely used monoclonal antibodies, TU-20 and TuJ-1 METHODS: Sixty patients with colorectal carcinoma were enrolled; all of them were treated surgically by the resection. Twenty tumours were histologically assessed as G1, 20 as G2 and 20 as G3. Routine immunohistochemical procedure using TU-20 and TuJ-1 mouse monoclonal antibodies was applied to all 60 specimen and slides were evaluated using an optical microscope. RESULTS: Expression of class III beta-tubulin was detected in 14 tumours (23.3%), while remaining tumours were negative. Relatively higher frequency of class III beta-tubulin expression was observed in G3 tumours (10 cases) in comparison with G1 (3 cases) and G2 (1 case), respectively. Seven tumours displayed positive immunostaining with both tested antibodies TU-20 and TuJ-1. Six tumours showed expression of class III beta- tubulin in more than 1 per cent of neoplastic cell population. In remaining 8 tumours only individual scattered neoplastic cells exhibited class III beta-tubulin expression either with TU-20, or with TuJ-1 antibody. INTERPRETATION & CONCLUSION: Higher frequency of immunoreactivity was observed in poorly differentiated tumours. However, more than 90 per cent of neoplastic cell population did not express class III beta-tubulin in almost all tumours. These negative cells of colonic cancer could represent the potential target for taxane-based chemotherapy in the future. Our results indicate that TU-20 and TuJ-1 antibodies exhibit very similar immunoreactivity in neoplastic tissue.