Enhancement of ATRA-induced differentiation of neuroblastoma cells with LOX/COX inhibitors: an expression profiling study

<p>Abstract</p> <p>Background</p> <p>We performed expression profiling of two neuroblastoma cell lines, SK-N-BE(2) and SH-SY5Y, after combined treatment with all-<it>trans </it>retinoic acid (ATRA) and inhibitors of lipoxygenases (LOX) and cyclooxygenases (COX). This study is a continuation of our previous work confirming the possibility of enhancing ATRA-induced cell differentiation in these cell lines by the application of LOX/COX inhibitors and brings more detailed information concerning the mechanisms of the enhancement of ATRA-induced differentiation of neuroblastoma cells.</p> <p>Methods</p> <p>Caffeic acid, as an inhibitor of 5-lipoxygenase, and celecoxib, as an inhibitor on cyclooxygenase-2, were used in this study. Expression profiling was performed using Human Cancer Oligo GEArray membranes that cover 440 cancer-related genes.</p> <p>Results</p> <p>Cluster analyses of the changes in gene expression showed the concentration-dependent increase in genes known to be involved in the process of retinoid-induced neuronal differentiation, especially in cytoskeleton remodeling. These changes were detected in both cell lines, and they were independent of the type of specific inhibitors, suggesting a common mechanism of ATRA-induced differentiation enhancement. Furthermore, we also found overexpression of some genes in the same cell line (SK-N-BE(2) or SH-SY5Y) after combined treatment with both ATRA and CA, or ATRA and CX. Finally, we also detected that gene expression was changed after treatment with the same inhibitor (CA or CX) in combination with ATRA in both cell lines.</p> <p>Conclusions</p> <p>Obtained results confirmed our initial hypothesis of the common mechanism of enhancement in ATRA-induced cell differentiation via inhibition of arachidonic acid metabolic pathway.</p

Nestin expression in osteosarcomas and derivation of nestin/CD133 positive osteosarcoma cell lines

<p>Abstract</p> <p>Background</p> <p>Nestin was originally identified as a class VI intermediate filament protein that is expressed in stem cells and progenitor cells in the mammalian CNS during development. This protein is replaced in the adult organism by other intermediate filament proteins; however, nestin may be re-expressed under certain pathological conditions such as ischemia, inflammation, brain injury, and neoplastic transformation. Nestin has been detected in many kinds of tumors, especially in tumors derived from the CNS. Co-expression of nestin and the CD133 surface molecule is considered to be a marker for cancer stem cells in neurogenic tumors. Our work was aimed at a detailed study of nestin expression in osteosarcomas and osteosarcoma-derived cell lines.</p> <p>Methods</p> <p>Using immunodetection methods, we examined nestin in tumor tissue samples from 18 patients with osteosarcomas. We also successfully established permanent cell lines from the tumor tissue of 4 patients and immunodetection of nestin and CD133 was performed on these cell lines.</p> <p>Results</p> <p>Nestin-positive tumor cells were immunohistochemically detected in all of the examined osteosarcomas, but the proportion of these cells that were positively stained as well as the intensity of staining varied. Nestin-positive cells were rarely observed in 2 tumor samples, and the remaining 16 tumor samples showed various nestin expression patterns ranging from very sporadic occurrence to an overwhelming proportion of cells with strong positive staining. Three of the established osteosarcoma cell lines were demonstrated to be nestin-positive, and only one cell line showed no expression of nestin; this finding corresponds with the rare occurrence of nestin-positive cells in the respective tumor sample. Moreover, three of these osteosarcoma cell lines were undoubtedly proven to be Nes+/CD133+.</p> <p>Conclusion</p> <p>Our results represent the first evidence of nestin expression in osteosarcomas and suggest the possible occurrence of cells with a stem-like phenotype in these tumors.</p

Prolonged survival in patients with local chronic infection after high-grade glioma treatment: Two case reports

High-grade gliomas are primary brain tumors with poor prognosis, despite surgical treatment followed by radiotherapy and concomitant chemotherapy. We present two cases of long-term survival in patients treated for high-grade glioma and concomitant prolonged bacterial wound infection. The first patient treated for glioblastoma IDH-wildtype had been without disease progression for 61 months from the first resected recurrence. Despite incomplete chemotherapy-induced myelosuppression in the second patient with anaplastic astrocytoma IDH-mutant, she died without disease relapse after 14 years from the diagnosis due to other comorbidities. We assume that the documented prolonged survival could be related to the bacterial infection

Prevalence of Propionibacterium acnes in Intervertebral Discs of Patients Undergoing Lumbar Microdiscectomy: A Prospective Cross-Sectional Study

Background The relationship between intervertebral disc degeneration and chronic infection by Propionibacterium acnes is controversial with contradictory evidence available in the literature. Previous studies investigating these relationships were under-powered and fraught with methodical differences;moreover, they have not taken into consideration P. acnes' ability to form biofilms or attempted to quantitate the bioburden with regard to determining bacterial counts/genome equivalents as criteria to differentiate true infection from contamination. The aim of this prospective cross-sectional study was to determine the prevalence of P. acnes in patients undergoing lumbar disc microdiscectomy. Methods and Findings The sample consisted of 290 adult patients undergoing lumbar microdiscectomy for symptomatic lumbar disc herniation. An intraoperative biopsy and pre-operative clinical data were taken in all cases. One biopsy fragment was homogenized and used for quantitative anaerobic culture and a second was frozen and used for real-time PCR-based quantification of P. acnes genomes. P. acnes was identified in 115 cases (40%), coagulase-negative staphylococci in 31 cases (11%) and alpha-hemolytic streptococci in 8 cases (3%). P. acnes counts ranged from 100 to 9000 CFU/ml with a median of 400 CFU/ml. The prevalence of intervertebral discs with abundant P. acnes (>= 1x10(3) CFU/ml) was 11% (39 cases). There was significant correlation between the bacterial counts obtained by culture and the number of P. acnes genomes detected by real-time PCR (r = 0.4363, p<0.0001). Conclusions In a large series of patients, the prevalence of discs with abundant P. acnes was 11%. We believe, disc tissue homogenization releases P. acnes from the biofilm so that they can then potentially be cultured, reducing the rate of false-negative cultures. Further, quantification study revealing significant bioburden based on both culture and real-time PCR minimize the likelihood that observed findings are due to contamination and supports the hypothesis P. acnes acts as a pathogen in these cases of degenerative disc disease

Frequent and sex-biased deletion of SLX4IP by illegitimate V(D)J-mediated recombination in childhood acute lymphoblastic leukemia

Acute lymphoblastic leukemia (ALL) accounts for ∼25% of pediatric malignancies. Of interest, the incidence of ALL is observed ∼20% higher in males relative to females. The mechanism behind the phenomenon of sex-specific differences is presently not understood. Employing genome-wide genetic aberration screening in 19 ALL samples, one of the most recurrent lesions identified was monoallelic deletion of the 5′ region of SLX4IP. We characterized this deletion by conventional molecular genetic techniques and analyzed its interrelationships with biological and clinical characteristics using specimens and data from 993 pediatric patients enrolled into trial AIEOP-BFM ALL 2000. Deletion of SLX4IP was detected in ∼30% of patients. Breakpoints within SLX4IP were defined to recurrent positions and revealed junctions with typical characteristics of illegitimate V(D)J-mediated recombination. In initial and validation analyses, SLX4IP deletions were significantly associated with male gender and ETV6/RUNX1-rearranged ALL (both overall P < 0.0001). For mechanistic validation, a second recurrent deletion affecting TAL1 and caused by the same molecular mechanism was analyzed in 1149 T-cell ALL patients. Validating a differential role by sex of illegitimate V(D)J-mediated recombination at the TAL1 locus, 128 out of 1149 T-cell ALL samples bore a deletion and males were significantly more often affected (P = 0.002). The repeatedly detected association of SLX4IP deletion with male sex and the extension of the sex bias to deletion of the TAL1 locus suggest that differential illegitimate V(D)J-mediated recombination events at specific loci may contribute to the consistent observation of higher incidence rates of childhood ALL in boys compared with girl

Molekularne patologicka diagnostika dystrofinopatii se zamerenim na identifikaci prenasecek.

Duchenne and Becker muscular dystrophies (DMD/BMD) are X-linked recessive neuromuscular disorders caused by dystrophin gene (DG) mutanions. The incidence of these disorders is very high (of about one DMD in 3500 newborn males, of about one BMD in 17000 newborn males) due to the extreme size of the DG. In approximately two-thirds of affected males the DG is disrupted by deletions not evenly distributed but concentrated in major and minor hot spots arround exons 45-47 and 7-9 respectively. Duplications are revealed in about 5-8% of cases and point mutation are reported to cause about one-third of total DMD/BMD cases. Discovery of the DG and its gene product led to the development of many diagnostical methods working on the protein or gene levels. Complex diagnosis of muscular distrophies has been provided in a limited extent in our country and some diagnostical methods useful in a diagnosis of preclinical stages of these diseases as well as in an identification of carriers of DMD/BMD have been put into practice in our institution. The combination of a clinical examination, muscle biopsy and the mutation analysis of mRNA using RT-PCT (reverse transcription-polymerase chain reaction) with a protein truncation test (PTT) brings the most effective results in a diagnosis of DMD/BMD. Histological examination of a muscle biopsy completed by an immunohistochemically detected deficient sarcolemmal expression of the dystrophin proved to be very reliable in a diagnosis of DMD. Normal or partially deficient sarcolemmal expression of the dystrophin in patients with a suspected BMD indicates the Western blot examination and the DG mutation analysis. RT-PCR and PTT enable the detection of deletions, duplications as well as point mutations in the DG and encompasse a large diagnostical scope in comparison with examinations on DNA level using the multiplex PCR (mPCR) which enables an identification of deletions and duplications in DG only. mPCR plays an important role especially in families with multiple incidence of DMD/BMD and with an identified and described deletion or duplication in the DG. Skipping of invasive muscular biopsies is reasonable in these cases. One of the major problems in DMD/BMD genetic counceling is still the definition of the carrier status in &quot;at risk&quot; females. The dissertation presents a diagnostical strategy using the method of fluorescence in situ hybridisation for an identification of DMD/BMD carriers. In females the detection of deletions is complicated by the presence of a second X-chromosome, carrying a normal DG. FISH allows the two X-chromosomes to be examined individually and constitutes an excellent alternative for diagnosis deletions in DMD/BMD heterozygotes. We have used a set of six cosmid probes for the detection of the most frequently deleted areas of the DG from the Department of Human Genetics, Leidel Medical Center, Nederlands. This optimized six cosmid sets shoud detect a deletion in 78% of all deletion carriers and i. e. 52% of all Duchenne/Becker muscular dystrophy cases. Data obtained in our study confirmed results of other authors in that FISH can be used to detect the deletions in the DG. FISH analysis of peripheral blood lymphocytes appears to be an effective and direct method for establishing the DMD/BMD carrier status of females. We suggest FISH to metaphase or interphase chromosomes for a rapid screening of DMD/BMD carriers.Available from STL Prague, CZ / NTK - National Technical LibrarySIGLECZCzech Republi

Comparative analysis of clinicopathological correlations of cyclooxygenase-2 expression in resectable pancreatic cancer

AIM: To perform a comparative analysis of clinicopathological correlations of cyclooxygenase-2 (COX-2) expression in pancreatic cancer, examined by monoclonal and polyclonal antibodies