Alexandros Lambropoulos

Revisiting OCT4 Expression in Peripheral Blood Mononuclear Cells

The transcription factor OCT4 (officially POU5F1; alternatively OCT3, OCT3/4, OTF3, and OTF4) is currently considered a main regulator of human embryonic stem cell pluripotency and self‐renewal capacities. Importantly, these stemness properties are attributed to OCT4A, which is one of the two isoforms produced by the OCT4 gene. The second OCT4 isoform, OCT4B, does not share the stemness factor characteristics of OCT4A and is currently considered of unknown function. Hence, when investigating OCT4 expression at the mRNA and protein level, it is important to specify which OCT4 isoform is detected by the applied methods, such as polymerase chain reaction assays and immunocytochemistry antibodies. Here, we discuss the need to distinguish between OCT4A and OCT4B when interpreting OCT4 expression in differentiated cells, such as peripheral blood mononuclear cells.

Methylenetetrahydrofolate reductase polymorphism C677T is not associated to the risk of cervical dysplasia

The aim of the study was to explore a possible association between methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism and cervical neoplasia. A total of 229 women were subjected to cytologic and colposcopic evaluation. Ninety-one of them were found to be normal, and served as the control group, while the other 138 of them had present or past histologically proven cervical pathology (patients group). All patients and controls were investigated for the MTHFR C677T polymorphism. Statistical analysis between the groups of cases with cervical intraepithelial neoplasia or invasive cervical cancer and the control group did not reveal any statistically significant difference in the frequency of the MTHFR C677T polymorphism.

Factor V Leiden and prothrombin G20210A mutations in pregnancies with adverse outcome

Background: Inherited thrombophilia has been associated with obstetric complications through mechanisms that are not yet fully elucidated. The aim of this study was to investigate the relationship between specific obstetric adverse outcomes and factor V Leiden and prothrombin G20210A mutations. Methods: Forty-five women with adverse pregnancy outcome defined as severe pre-eclampsia, abruptio placentae, intrauterine growth restriction and stillbirth, were tested for factor V Leiden and prothrombin G20210A mutations. The control group comprised 100 women with at least one normal pregnancy and no history of thrombosis. Results: Overall, 13 women with one or more of the above-mentioned pregnancy complications (28%) had either thrombophilic mutation, as compared with six in the control group (6%) (p < 0.001, odds ratio (OR) 6.1; 95% confidence interval (CI) 1.9-20). The factor V Leiden mutation was detected in ten of the women with complicated pregnancies (22%) and in four of the controls (4%) (p < 0.001, OR 6.6; 95% CI 1.7-27.2). The prothrombin G20210A mutation was detected in three women in the group with complications (6%) and in two of the controls (2%) (p = 0.17, OR 3.4; 95% CI 0.4-30.5). Compared to controls, the prevalence of the factor V Leiden mutation was significantly higher in the subgroups of severe pre-eclampsia, abruptio placentae and fetal growth restriction. The prevalence of the prothrombin G20210A mutation does not appear to be significantly different from that in the controls in any of the groups studied. Conclusions: Our data suggest that inherited thrombophilia, and specifically the factor V Leiden mutation, may be associated with adverse pregnancy outcome. The role of the prothrombin G20210A mutation remains to be elucidated.

OCT4B1 ISOFORM: THE NOVEL OCT4 ALTERNATIVE SPLICED VARIANT AS A PUTATIVE MARKER OF STEMNESS

A novel OCT4 alternative spliced variant (OCT 4B1) is deduced to be the isoform present in 59 hESC lines characterised by the International Stem Cell Initiative (ISCI) rather than OCT4A as previously assumed. The new variant may be a more reliable marker of stemness than OCT4A and studies are needed to test this.

MMP9 but Not EGFR, MET, ERCC1, P16, and P-53 Is Associated with Response to Concomitant Radiotherapy, Cetuximab, and Weekly Cisplatin in Patients with Locally Advanced Head and Neck Cancer

Concomitant administration of radiotherapy with cisplatin or radiotherapy with cetuximab appear to be the treatment of choice for patients with locally advanced head and neck cancer. In the present retrospective analysis, we investigated the predictive role of several biomarkers in an unselected cohort of patients treated with concomitant radiotherapy, weekly cisplatin, and cetuximab (CCRT). We identified 37 patients treated with this approach, of which 13 (35%) achieved a complete response and 10 (27%) achieved a partial response. Severe side effects were mainly leucopenia, dysphagia, rash, and anemia. Tumor EGFR, MET, ERCC1, and p-53 protein and/or gene expression were not associated with treatment response. In contrast, high MMP9 mRNA expression was found to be significantly associated with objective response. In conclusion, CCRT is feasible and active. MMP9 was the only biomarker tested that appears to be of predictive value in cetuximab treated patients. However, this is a hypothesis generating study and the results should not be viewed as definitive evidence until they are validated in a larger cohort.

Prevalence and distribution of high-risk human papillomavirus in Greece.

As knowledge of regional human papillomavirus (HPV) type distribution is essential for the optimization of prevention strategies, this study was carried out to explore the prevalence and type distribution of high-risk HPV in a screening population across Greece. Cervical samples were collected by local physicians and nurses in hospitals and health centers across the country from 4139 women attending for cervical cancer screening. High-risk HPV-DNA was detected by using Hybrid Capture-2 (HC2) and positive samples with adequate cellular content were further typed by restriction fragment length polymorphism-polymerase chain reaction. Almost six percent (5.9%) of women tested positive in HC2. The most common type was HPV16 (1.4% in the whole sample and 32.4% of the typed samples), followed by HPV53 (0.6 and 14.0%, respectively), HPV31 (0.6 and 12.9%, respectively), HPV35 (0.5 and 12.3%, respectively), HPV51 (0.4 and 7.8%, respectively), HPV18 (0.3 and 7.3%, respectively) and 22 more types. Almost 15% of the typed samples showed a coinfection with two HPV types and 2.1% with three types. There was a bimodal distribution by age, with the highest peak in women 20–29 years old and a lower peak in women 50–59 years old. Apart from the types originally included in HC2 cocktail, PCR analysis identified 15 more types (HPV6, HPV11, HPV34, HPV37, HPV38, HPV42, HPV53, HPV54, HPV55, HPV61, HPV62, HPV66, HPV73, HPV82, HPV83). Eleven percent of HC2-positive results arose from single-type infections with HPV53 (10%) and HPV66 (1%), which are potentially high-risk types. In conclusion, HPV16 is the most common type in the largest Greek screening sample used to date and, together with its related types, accounts for more than half of high-risk HPV infections. Approximately 10% of positive HC2 results arise from HPV53, which is not normally detected by the test, but may be clinically significant.

Subspecies variation in Greek strains of Chlamydophila abortus.

The Greek chlamydial strains FAS, FAG, VPG and LLG, isolated from aborted sheep or goat foetuses, had been previously characterized as divergent on the basis of mouse cross-protection experiments, with LLG and its homologous POS significantly different from the rest in inclusion morphology, polypeptide profiles and reactivity with monoclonal antibodies. To determine the genetic basis of their divergence the 16S-23S ribosomal intergenic spacer was analysed by RFLP analysis of PCR 16SF2/23R amplicons. Using the restriction enzymes BfaI, SfcI, HpaI, BclI, DdeI and AclI, the strains were classified as Chlamydophila abortus. However, digestion with RsaI made it possible to differentiate strains FAS, FAG and VPG from strains LLG and POS, generating DNA fragments of 530/55 and 585bp, respectively. By subsequent sequence analysis of the 23S domain I rRNA gene only strain FAS was identical to reference strain A22 of C. abortus. Strains FAG and VPG presented an identical nucleotide deviation at position 593 of signature sequences. Strains LLG and POS presented three identical nucleotide deviations at positions 156, 186 and 307. Variation within the domain I signature sequences for the examined abortion strains was < or =0.69%. In conclusion, substantial genetic and biological diversity among strains of C. abortus was demonstrated, suggesting that subspecies variation status for certain strains may be applicable. Our findings suggest that differentiation may be possible at a subspecies level by RFLP analysis.

Factor V Leiden in Greek thrombophilic patients: relationship with activated protein C resistance test and levels of thrombin-antithrombin complex and prothrombin fragment 1 + 2.

We studied 172 Greek patients (72 men aged 44.0 × 16.7 years and 100 women aged 46.5 × 14.1 years) with an unexplained thrombophilic tendency. One hundred and four apparently healthy persons (63 men aged 34.2 × 10.0 years and 41 women aged 37.1 × 13.3 years) were included as a control group. We performed the activated protein C resistance (APC-r) test using a clotting test (Chromogenix kit), detection of factor V Leiden using polymerase chain reaction (PCR)-restriction fragment length polymorphisms and measurement of thrombin-antithrombin complexes (TAT) and prothrombin fragment 1+2 (F1+2) levels with an immunoenzymatic assay. The normal range for the APC-r test (> 2.12) was determined from the controls. The factor V Leiden mutation was found in 31.9% of all the patients tested, in 28.1% of the unrelated patients with documented thrombophilic tendency of unknown origin and in 4.8% of the healthy controls. The APC-r test had a sensitivity of 0.42 and a specificity of 0.91 for the detection of factor V Leiden. Furthermore, we found no significant difference in levels of TAT and F1+2 between patients with and without the mutation and there was no correlation between aPC-r values and levels of TAT and F1+2.

Glutathione-S-transferase M1 and T1 and cytochrome P1A1 genetic polymorphisms and susceptibility to cervical intraepithelial neoplasia in Greek women.

The aim of the study was to determine the importance of genetic polymorphisms of glutathione-S-transferase T1 and M1 and cytochrome P1A1 genes in the development of cervical intraepithelial neoplasia in Greek women. This was a prospective, case–control study conducted by the Cervical Pathology and Colposcopy Unit of a University Ob/Gyn Department from 1999 to 2003. Cervical smears from 114 controls without any cytological and/or colposcopical evidence of cervical pathology and from 166 women with history of cervical intraepithelial neoplasia (56 CIN I, 54 CIN II and 56 CIN III) were examined with polymerase chain reaction for the above-mentioned genetic polymorphisms, taking also in mind their smoking attitudes. Statistical analysis was performed to detect any association between the null genotype of GSTM1 and GSTT1 genes and the CYP1A1 m1 polymorphism and the severity of cervical intraepithelial neoplasia. The distributions of the GSTT1 and GSTM1 wild-type genotypes were 57.48 and 39.75%, respectively. No woman with homozygous GSTT1 and GSTM1 null/null genotype was identified. CYP1A1 m1 polymorphism frequency was 24.49%. No woman with homozygous CYP1A1 m1/m1 genotype was detected as well. No significant difference in the frequencies of the GSTM1 and GSTT1 null alleles, and the CYP1A1 m1 polymorphism, was found between cases and controls. After application of Mantel–Haenszel χ2 procedure, there was no linear severity of the lesion and the frequency of these polymorphisms. According to our results, glutathione-S-transferase T1 and M1 and cytochrome P1A1 genetic polymporphisms do not appear to be a risk factor for cervical disease irrespective of smoking habits.

Genetics in rheumatoid arthritis beyond HLA genes: What meta-analyses have shown?

OBJECTIVE Rheumatoid arthritis (RA) is a complex disorder with many genetic and environmental factors to account for disease susceptibility. Individual genetic association studies usually suffer from small sample size leading to biased results of polymorphisms association with RA liability. Therefore, meta-analyses seem to resolve this limitation, up to a point, increasing the power of statistical analyses. In this review, we summarize the current knowledge of non-HLA genetic factors contributing to RA predisposition based on meta-analyses. METHODS Using the key words: rheumatoid arthritis, meta-analysis, and polymorphism, we searched the PubMed database for the associated articles. Up to the middle of November 2012, seventy-nine articles fulfilled the criteria and highlighted the current findings on the genetic factors contributing to RA susceptibility. RESULTS The association with RA was confirmed for 32 gene polymorphisms, being population specific in some cases. However, meta-analyses did not confirm an association in case of 16 gene variants, previously studied in individual studies for their association with RA. CONCLUSIONS The use of bioinformatics tools and functional studies of the summarized implicated genes in RA pathogenesis could shed light on the molecular pathways related to the disorder, helping to the development of new drug targets for a better treatment of RA.