Maria Samara

Molecular evidence for transferrin receptor 2 expression in all FAB subtypes of acute myeloid leukemia

We examined transferrin receptor (TfR) 1 and TfR2 mRNA expression in 50 acute myeloid leukemia (AML) patients by RT-PCR, with primers specific for exons 15-17 (TfR1), 3-5 (TfR2-alpha) and 4-5 (TfR2-beta) of the corresponding gene. There were 4/50 TfR1-negative (-), 3/50 TfR2-alpha mRNA (-) and 13/50 TfR2-beta mRNA (-) cases; only three cases were TfR1/2 mRNA (-). No significant correlations were identified between TfR2-beta mRNA negativity and specific FAB subtypes, karyotype or attainment of complete remission. These findings suggest that: (i) TfR2 expression is not restricted to erythroid cells, and (ii) iron import proteins might complement each other in AML cells.

Impact of ACE and ApoE polymorphisms on myocardial perfusion: correlation with myocardial single photon emission computed tomographic imaging

Coronary artery disease is associated with multiple genetic and environmental risk factors. In this study, we evaluated the correlation of angiotensin l-converting enzyme (ACE) (I/D) and ApoE gene polymorphisms (E2, E3, E4 and g.-219G/T) with myocardial perfusion. We examined 410 patients using exercise–rest myocardial perfusion single photon emission computed tomography (SPECT), in which the summed stress score (SSS), summed rest score (SRS) and summed difference score (SDS) indexes were calculated. Homozygotes for the ACE D allele had greater mean values of SSS (P<0.001) and SDS (P<0.001). In addition, E3 homozygotes, E4 heterozygotes and E4 homozygotes had significantly higher values of SSS and SDS compared with E3 heterozygotes (P<0.001); E4 homozygotes had significantly higher values of SSS and SDS compared with E3 homozygotes. Furthermore, for the g.-219G>T polymorphic site at the promoter region of ApoE gene, the mean values of SSS and SDS were significantly higher for T heterozygotes/homozygotes than for GG homozygotes. Adjusting for all demographic and clinical data using multiple linear regression analysis it was found that ACE D and both ApoE genotypes were independent predictors with a cumulative contribution for the prediction of SSS and SDS. Furthermore, logistic regression analysis revealed that all three genotypes had an independent predictive ability for abnormal SSS (SSS>2). These data provide the first evidence of an association and significant cumulative contribution of the aforementioned genotypes in myocardial perfusion with E4 allele having the strongest association followed by ACE D and ApoE g.-219T alleles.

The 5′ Regulatory Region of the Human Fetal Globin Genes is a Gene Conversion Hotspot

The human fetal globin genes consist of the first mammalian genomic loci for which gene conversion was reported. To date, 14 gene conversions have been described in the human Gγ- and Aγ-globin genes, the vast majority of which are restricted to the coding sequences. Here, we provide evidence for three new gene conversion events in the 5′ regulatory region of the human fetal globin genes, identified during a large genetic screening effort in adult individuals with high fetal hemoglobin (Hb) levels. The sequence variations, resulting from these conversion events, are transcriptionally silent polymorphisms that do not contribute to increased fetal Hb levels. Our results suggest that the 5′ regulatory region of the human fetal globin genes is a gene conversion hotspot that prevent globin gene promoter sequence diversification, further underlining the need for two functional fetal globin genes in fetal erythropoiesis.

Aurora B kinase in Hodgkin lymphoma: immunohistochemical pattern of expression in neoplastic Hodgkin and Reed-Sternberg cells

Aurora B is a member of the chromosomal passenger complex, which is essential for proper completion of mitosis and cell division (cytokinesis). Inappropriate chromosomal segregation and cytokinesis due to deregulated expression of chromosome passenger proteins may lead to aneuploidy and cancer including lymphomas. According to our knowledge there are extremely limited studies investigating the immunohistochemical expression of Aurora B in tumor specimens of Hodgkin lymphoma. Our purpose was to characterize the expression of Aurora B in biopsies of Hodgkin lymphomas, and to evaluate the pattern of immunoreactivity in neoplastic Hodgkin and Reed-Sternberg cells (RS cells). We examined Aurora B immunoreactivity in paraffin sections of 15 samples of Hodgkin lymphomas, obtained from 15 patients, 8 men and 7 women. Ten were of nodular sclerosis type and five were of mixed cellularity. Our results showed immunoexpression of Aurora B in mononuclear lymphoid cells as well as in bi- and multinucleated RS cells. In addition, positive neoplastic cells in mitosis were observed, whereas a subpopulation without evidence of immunoreaction was also detected in each case. Taken together our results point to a possible association between Aurora B expression and mitotic deregulation in Hodgkin lymphoma, which may provide novel targets for treatment.

Correlation between cyclin B1 immunostaining in cervical biopsies and HPV detection by PCR.

In recent years, experimental data on the roles of viruses and cyclin B1 in G2/M arrest has accumulated. The aim of our study was to immunohistochemically examine a series of cervical squamous intraepithelial lesions for the presence of cyclin B1 and test for a possible correlation with morphologic findings or human papillomavirus (HPV) types detected by polymerase chain reaction. One hundred and three biopsies and/or conization specimens were examined, including high-grade lesions (25), low-grade lesions (52), biopsies showing nondiagnostic atypia (15), and biopsies without histopathologic alterations (11). Three patterns of immunopositivity were recognized, according to the level of epithelium exhibiting positively stained cells. Immunoreactivity for cyclin B1 above the basal/parabasal cells correlated with HPV presence. Cyclin B1-stained cells may represent “prekoilocytes,” whose eventual progression to koilocytes would depend on several parameters related to the intricacies of HPV infection. In cases of nondiagnostic atypia immunoreactivity, cyclin B1 could be considered as a surrogate test.

Mutation profile of KRAS and BRAF genes in patients with colorectal cancer: association with morphological and prognostic criteria

KRAS and BRAF mutations are well-recognized molecular alterations during colorectal carcinogenesis, but there is little agreement on their effect on tumor characteristics. Therefore, we aimed to evaluate the distribution of the most common KRAS and BRAF mutations in Greek patients with colorectal cancer and their possible associations with clinical histopathological parameters. In this study, 322 and 188 colorectal carcinomas were used for the mutation analysis of KRAS (exon 2) and BRAF (exon 15) genes, respectively. The mutational status of both genes was evaluated by polymerase chain reaction and sequencing analysis. Although the overall frequency of KRAS mutations (36.6%) seemed to be similar to those reported for other populations, the rate of point mutations at codon 13 was significantly lower (12%) in Greek patients with colorectal cancer and associated with male gender (P < 0.05). Tumors with G>T codon 12 transversions and G>C transitions showed more frequent lymph node metastasis (P < 0.05, P < 0.005, respectively). The rate of KRAS mutations gradually decreased with increasing histological grade (P < 0.05), as opposed to BRAF mutations, which were strongly associated with poorly differentiated tumors (P < 0.005). Additionally, we found that the histological features of preexisting adenoma were associated with the absence of BRAF mutations, in contrast to KRAS (P < 0.05). Our data suggested that there seems to be a correlation between morphological criteria and discrete genetic pathways in colorectal carcinogenesis. Moreover, ethnic or geographic factors may have an impact on genetic background of colorectal carcinomas, and specific types of KRAS mutations may influence the metastatic potential of colorectal tumors.

Darwinian molecular imaging in nuclear cardiology

Dear Sir, We read with great interest the recent editorial by Bertrand Tavitian and Uwe Haberkorn published in the September 2009 issue concerning the role of Darwin’s theory of evolution through natural selection in the fields of molecular imaging and nuclear medicine [1]. As the authors suggest, at first sight molecular imaging appears to have little if any connection with Darwin’s theory apart from the “minor” point that the environment exerts a selective pressure on the “survival” of the tracer, i.e. its escape from the body’s excretory and metabolic processes. However, new probes can now be synthesized by artificial adaptation of natural molecular synthesis mechanisms for the production of macromolecules with excellent recognition capacity, hence making their use in molecular imaging tempting. In these cases the scientist is responsible for designing the variations in the pool of the species (molecules) which have to encompass and survive the selection pressure imposed on them. Apart from the artificial character of the procedure, the mechanisms at work are basically Darwinian processes [1]. Of significant importance is the authors’ report on the implementation of Darwinian approaches to molecular evolution of oligonucleotide binders and in vivo nuclear imaging with aptamers, to cell-based display systems (i.e. phage display or bacterial peptide display) and to imaging and therapy using molecules identified by phage display, and also to cell-free (in vitro selection) systems such as ribosome display and mRNA display [1–8]. Moreover, the authors comment on the DNA shufflingmethodwhichmimics natural recombination by allowing in vitro homologous recombination of DNA, and it seems of potential interest for the generation (although not applied yet) of highaffinity radioisotopic binders for use in nuclear medicine in the future [1]. Finally, the authors conclude that the considerable “creationist” component that may be considered to exist in the presented techniques only applies to the design of the starting point of these experiments. The mechanisms at work during the generation of a variety of species and the process of selection are purely evolutionary. In addition, high throughput methods for ligand selection enter the field of nuclear molecular imaging, and these add new classes of molecules with improved properties with respect to stability, clearance, transport and affinity [1]. Nuclear medicine, even from its first applications (i.e. bone and thyroid imaging), has the unique ability to provide functional and more recently molecular imaging, allowing early-stage disease diagnosis, as functional alterations usually precede anatomical lesions. Moreover, the functional character of radioisotopic imaging has led to an important correlation between genetics and nuclear medicine (i.e. identifying P-glycoproteins using Tc-sestamibi P. Georgoulias (*) :V. Valotassiou Nuclear Medicine Department, University Hospital of Larissa, Mezourlo, Larissa 41110, Hellas e-mail: pgeorgoul@med.uth.gr

Smad4 and epithelial–mesenchymal transition proteins in colorectal carcinoma: an immunohistochemical study

Epithelial–mesenchymal transition (EMT) plays an important role in cancer metastasis. During EMT, tumor cells acquire the capacity to migrate and invade the stroma. Activation of the transforming growth factor-b (TGF-b) signaling pathway is of major importance for the initiation of EMT. Smad4, an essential protein of this pathway, is known to complex with multiple transcription factors (e.g. Snail-1, Slug, Twist-1), in various types of cancer, promoting the repression or activation of target genes. The role of Smad4 in colorectal cancer (CRC) is not straightforward so far. In the present study forty eight resected CRC tumor specimens were immunohistochemically examined in order to assess the expression of Smad4 and its association with E-cadherin, Snail-1, Slug, Twist-1 protein expression and with various pathological parameters. Smad4 was found to be positively correlated with Snail-1, Slug and Twist-1 expression (p < 0.001). On the other hand it was negatively correlated with the expression of E-cadherin (p < 0.001). Furthermore, lymphatic invasion could be clearly associated with Smad4 expression, a finding complying with the metastatic ability of EMT cells. In conclusion, Smad4 could be considered as a central component of EMT transition in human colorectal cancer that combines with transcriptional factors to reduce E-cadherin and alter the expression of the epithelial phenotype.