In vitro and in vivo properties of distinct populations of amniotic fluid mesenchymal progenitor cells

Human mesenchymal progenitor cells (MPCs) are considered to be of great promise for use in tissue repair and regenerative medicine. MPCs represent multipotent adherent cells, able to give rise to multiple mesenchymal lineages such as osteoblasts, adipocytes or chondrocytes. Recently, we identified and characterized human second trimester amniotic fluid (AF) as a novel source of MPCs. Herein, we found that early colonies of AF-MPCs consisted of two morphologically distinct adherent cell types, termed as spindle-shaped (SS) and round-shaped (RS). A detailed analysis of these two populations showed that SS-AF-MPCs expressed CD90 antigen in a higher level and exhibited a greater proliferation and differentiation potential. To characterize better the molecular identity of these two populations, we have generated a comparative proteomic map of SS-AF-MPCs and RS-AF-MPCs, identifying 25 differentially expressed proteins and 10 proteins uniquely expressed in RS-AF-MPCs. Furthermore, SS-AF-MPCs exhibited significantly higher migration ability on extracellular matrices, such as fibronectin and laminin in vitro, compared to RS-AF-MPCs and thus we further evaluated SS-AF-MPCs for potential use as therapeutic tools in vivo. Therefore, we tested whether GFP-lentiviral transduced SS-AF-MPCs retained their stem cell identity, proliferation and differentiation potential. GFP-SS-AF-MPCs were then successfully delivered into immunosuppressed mice, distributed in different tissues and survived longterm in vivo. In summary, these results demonstrated that AF-MPCs consisted of at least two different MPC populations. In addition, SS-AF-MPCs, isolated based on their colony morphology and CD90 expression, represented the only MPC population that can be expanded easily in culture and used as an efficient tool for future in vivo therapeutic applications

Origin and spread of human mitochondrial DNA haplogroup U7

Human mitochondrial DNA haplogroup U is among the initial maternal founders in Southwest Asia and Europe and one that best indicates matrilineal genetic continuity between late Pleistocene hunter-gatherer groups and present-day populations of Europe. While most haplogroup U subclades are older than 30 thousand years, the comparatively recent coalescence time of the extant variation of haplogroup U7 (~16–19 thousand years ago) suggests that its current distribution is the consequence of more recent dispersal events, despite its wide geographical range across Europe, the Near East and South Asia. Here we report 267 new U7 mitogenomes that – analysed alongside 100 published ones – enable us to discern at least two distinct temporal phases of dispersal, both of which most likely emanated from the Near East. The earlier one began prior to the Holocene (~11.5 thousand years ago) towards South Asia, while the later dispersal took place more recently towards Mediterranean Europe during the Neolithic (~8 thousand years ago). These findings imply that the carriers of haplogroup U7 spread to South Asia and Europe before the suggested Bronze Age expansion of Indo-European languages from the Pontic-Caspian Steppe region

Gene Therapy for Malignant and Benign Gynaecological Disorders: A Systematic Review of an Emerging Success Story

Despite the major advances in screening and therapeutic approaches, gynaecological malignancies still present as a leading cause of death among women of reproductive age. Cervical cancer, although largely preventable through vaccination and regular screening, remains the fourth most common and most lethal cancer type in women, while the available treatment schemes still pose a fertility threat. Ovarian cancer is associated with high morbidity rates, primarily due to lack of symptoms and high relapse rates following treatment, whereas endometrial cancer, although usually curable by surgery, it still represents a therapeutic problem. On the other hand, benign abnormalities, such as fibroids, endometriosis, placental, and embryo implantation disorders, although not life-threatening, significantly affect women’s life and fertility and have high socio-economic impacts. In the last decade, targeted gene therapy approaches toward both malignant and benign gynaecological abnormalities have led to promising results, setting the ground for successful clinical trials. The above therapeutic strategies employ both viral and non-viral systems for mutation compensation, suicide gene therapy, oncolytic virotherapy, antiangiogenesis and immunopotentiation. This review discusses all the major advances in gene therapy of gynaecological disorders and highlights the novel and potentially therapeutic perspectives associated with such an approach

Metabolic rewiring is associated with HPV-specific profiles in cervical cancer cell lines

Both HPV-positive and HPV-negative cervical cancers are associated with aberrant metabolism, although the oncogenic drivers remain elusive. Here we show the assessment of the metabolomic profiles of four distinct cervical cell lines, a normal and three cancer cell lines, one HPV-negative (C33A) and two HPV-positive (SiHa HPV16+, HeLa HPV18+), employing an ultra performance liquid chromatography and a high resolution mass spectrometry. Out of the total 462 metabolites, 248 to 326 exhibited statistically significant differences, while Random Forests analysis identified unique molecules for each cell line. The two HPV+ cell lines exhibited features of Warburg metabolism, consistent with the role of the HPV E6 protein. SiHa and HeLa cells displayed purine salvage pathway activity, while C33A cells revealed synthesis of cytidine, via a novel mechanism. These data document a highly dynamic HPV-specific rewiring of metabolic pathways occurring in cervical cancer. Therefore, this approach can eventually provide novel mechanistic insights into cervical carcinogenesis

Transcription factor ATF-3 regulates allele variation phenotypes of the human SLC11A1 gene

Genetic polymorphisms in the human solute carrier family 11 member 1 (SLC11A1) gene predispose to susceptibility to infectious/inflammatory diseases and cancer. Human susceptibility to these diseases exhibits allelic association with a polymorphic regulatory Z-DNA-forming microsatellite of a (GT/AC)n repeat. The carriage of different alleles may influence chromatin remodeling and accessibility by transcription factors. Of particular importance is the binding site for the Activating Protein 1 (AP-1) elements, (ATF-3 and c-Jun), adjacent to the 5’ sequence of the Z-DNA-forming polymorphism. The aim of the study was to characterize the transcriptional mechanisms controlling different alleles of SLC11A1 expression by ATF-3 and c-Jun. Allele 2, [T(GT)(5)AC(GT)(5)AC(GT)(10)GGCAGA(G)(6)], and Allele 3, [T(GT)(5)AC(GT)(5)AC(GT)(9)GGCAGA(G)(6)], were subcloned into the PGL2Basic vector. Transient transfections of THP-1 cells with the constructs, in the presence or absence of pATF-3 were preformed. Luciferase expression was determined. To document the recruitment of ATF-3 and c-Jun, to the polymorphic promoter alleles in vivo, we performed ChIP assays with transient transfected THP-1 cells treated with or without lipopolyssacharides. Our data documented that ATF-3 suppresses the transcriptional activation of Allele-3, and this suppression is enhanced in the presence of lipopolyssacharides. Our findings suggest that ATF-3 and c-Jun may influence heritable variation in SLC11A1-dependent innate resistance to infection and inflammation both within and between populations

The Q192R polymorphism of the paraoxonase-1 (PON1) gene is associated with susceptibility to gestational diabetes mellitus in the Greek population

A key factor protecting from oxidative stress in gestational diabetes mellitus (GDM) and in type 2 diabetes (T2D) is paraoxonase-1 (PON1). Inconclusive and limited data exist regarding the effect of a coding polymorphism (Q192R) of the PON1 gene in conferring susceptibility to both states. In the present study, we investigated the association between the PON1 gene and the risk for GDM in the Greek population and assessed for the first time its transcriptional efficiency. We studied 185 women with GDM and 104 non-diabetic controls for the PON1 polymorphism. For PON1 mRNA expression, peripheral leucocytes were harvested from 20 GDM and 20 control women, harboring different genotypes for the polymorphism, using real-time quantitative PCR. The RR genotype and the R allele of the PON1 Q192R polymorphism were significantly associated with an increased risk for GDM (p=0.012 and p<0.0001, respectively). Furthermore, there was no statistical correlation between the individual metabolic parameters tested and the three genotypes. Finally, the expression levels of PON1 mRNA in GDM patients did not exhibit any statistical difference compared with normal controls (p=0.138). These data independently document that the Q192R polymorphism is closely associated with GDM susceptibility, while the PON1 gene expression is not impaired in GDM

The major circadian pacemaker ARNT-like protein-1 (BMAL1) is associated with susceptibility to gestational diabetes mellitus

Aims: Recently a relationship between circadian clock function and the risk for type 2 diabetes (T2D) has been shown. BMAL1 is a key component of the mammalian molecular clock. Two SNPs in the BMAL1 gene have been identified to confer T2D susceptibility. In the present study we investigated for the first time the association between the BMAL1 gene and the risk for GDM, in a Greek population. Methods: We studied 185 women with GDM and 161 non-diabetic controls for BMAL1 polymorphisms. For BMAL1 mRNA expression, peripheral leukocytes were harvested from 20 GDM and 20 control women, harboring different genotypes for the tested polymorphisms, using real-time quantitative PCR. Results: The minor allele (A) of the BMAL1 rs7950226 (G > A) polymorphism was found to be significantly associated with an increased risk of GDM (P = 0.025). Analysis of the second BMAL1 rs11022775 (T > C) polymorphism, showed that the C-allele frequency was strongly increased in women with GDM (P = 4.455e-06). The CC genotype was also significantly overrepresented in GDM vs. controls (P = 0.00001). Additionally, the rs7950226G/rs11022775C and rs7950226A/rs11022775C haplotypes were also found to be associated with increased susceptibility to GDM. Furthermore, the expression levels of BMAL1 mRNA were significantly lower in GDM patients than in controls. Conclusion: These data suggest that the impairment of the BMAL1 clock gene expression is closely associated with GDM susceptibility. (C) 2012 Elsevier Ireland Ltd. All rights reserved

Circadian clock gene expression is impaired in gestational diabetes mellitus

Dysfunction of the circadian clock genes is involved in the development of obesity and type 2 diabetes (T2D). Since gestational diabetes mellitus (GDM) and T2D share common genetic and phenotypic features, in the present study, we investigated the status of the circadian clock in a cohort of 40 Greek pregnant women with GDM, four with T2D and 20 normal controls. Peripheral blood mRNA transcript levels of 10 clock genes (CLOCK1, BMAL1, PERI, PER2, PER3, PPARA, PPARD, PPARG, CRY1 and CRY2) were determined by real-time quantitative PCR. GDM patients expressed significantly lower transcript levels of BMAL1, PER3, PPARD and CRY2 compared to control women (p < 0.05). No significant difference was documented between GDM women maintained either under insulin treatment or diet. A positive correlation was found between the expression of BMAL1 versus CRY2 (r = 0.45, p = 0.003) and BMAL1 versus PPARD (r = 0.43, p = 0.004). Further investigation on the functional relevance of these clock genes, disclosed that expression of PER3 correlated negatively with HbA(1c) levels (r = -0.36, p = 0.022). These data document for the first time that the expression of BMAL1, PER3, PPARD and CRY2 genes is altered in GDM compared to normal pregnant women and support the notion that deranged expression of clock genes may play a pathogenic role in GDM