Down Syndrome: Development Of A Non-Invasive Prenatal Dna Screening Test Using Superoxide Dismutase 1 Gene In Maternal Blood And Detection Of Cystathionine P-Synthase Gene Mutations

Down syndrome or Trisomy 21, is the most commonly occurring genetic disorder that stems from the failure of ·chromosome 21 to segregate normally during meiosis, resulting in an individual carrying an extra copy of chromosome 21. The main aims of this study were to develop a relatively non-invasive prenatal DNA screening method using maternal blood and to detect mutations on cystathionine J3-synthase (CBS) gene, a folate pathway gene located on chromosome 21. As an initial step, the presence of foetal cells and DNA in the maternal blood was firstly determined by foetal haemoglobin (HbF) staining and polymerase chain reaction (PeR). It was found that the ratio ofthe nucleated foetal cell to maternal cell increased from 2 in 106 to 3 in 106 and 5 in 106 at the first, second and third trimester, respectively. By using Y chromosome specific primers, DNA from male foetuses could be detected as early as 6 weeks of gestation in 200 Jil maternal blood obtained from fingertip. This is in line with the current technology in non-invasive screening methods of foetal aneuploidies which is focused on detecting Y chromosomal sequences which is impossible to be used for female foetus pregnancies. Therefore, the superoxide dismutase 1 (SOD'!) gene sequence, which is located on the Down Syndrome Critical Region, was used to overcome this situation by using real-time quantitative PCR The level of SOD1 sequences in maternal blood was found to be significantly elevated in the third trimester normal pregnancies (mean = 11728 copies/Ill) when compared to the second trimester (mean = 5705.6 copies/Ill), p<0.OO5 and non-pregnant normal women (mean = 3580.2 copies/Ill), p<O.OOOl. Down syndrome pregnancies have the greatest elevation compared to all the three trimesters of normal singleton pregnancies and twin pregnancies,p<0.05. The traditional approach ofprenatal chromosomal diagnosis using amniotic fluid was found to be cumbersome and time-consuming compared to the newly developed method. The mutation detection on CBS gene was carried out using DNA sequencer and denaturing high performance liquid chromatography (DHPLC). This study revealed that the Down syndrome patients have four mutations, which are in intron 1 (A9231C), exon 10 (C20628T) and exon 17 (T277%C and C27817T). The Down syndrome children were found to have the same genotype as their mothers. The number ofmothers and children having the substitutions in the CBS gene was twice the number ofmothers and children with normal genotype, suggesting that the mothers who have these substitutions are at higher risk of having a child with Down syndrome. In conclusion, non-invasive prenatal diagnosis at first trimester using Y chromosomal sequence IS feasible for diagnosis of foetal-derived paternally-inherited polymorphism/mutations or genes. Quantitative analysis using gene associated with a disorder has a potentially significant advantage over the invasive techniques currently used widely for prenatal diagnosis. Finally, the discovery of the mutations in the CBS gene of Down syndrome patients and mothers will help contribute to new knowledge and the future studies on the folate pathway genes mutation and occurrence of Down syndrome. It may also suggest an opportunity to improve public health strategies for the primary prevention ofDown syndrom

Nutritional Composition, Antioxidant Activity And Anticarcinogenic Effect Of Typhonium Flagelliforme (Nicholson 1029) Extract In Rat

Typhonium flagelliforme from the family Araceae is locally known as rodent tuber. It is being used traditionally in Malaysia to treat cancer. The major aim of this study was to investigate the effect of Typhonium flagelliforme crude extract on hepatocarcinogenesis in rats induced by diethylnitrosamine (DEN) and 2- acetylaminofluorene (AAF). Besides the in vivo study, the plant's nutrient and non-nutrient composition, and its antioxidant activity were also determined. Typhonium flagelliforme has carbohydrate (0.5% in leaves-stalks and 27.5% in tubers-roots) as its main constituent. It has high content of vitamin C (106.5 mg/100g in leaves-stalks and 11.6 mg/100g in tubers-roots) and potassium (1276.8 mg/100g in leaves-stalks and 534 mg/100g in tubers-roots). Alkaloid (0.4 mg/100g in leaves-stalks and 0.9 mg/100g in tubers-roots) and catechin (1.5% in leaves-stalks and 0.7% in tubers-roots) were found in this plant. The phytochemicals of Typhonium flagelliforme showed a greater antioxidant activity than vitamin E

A non-invasive prenatal DNA screening test for Down Syndrome

Prenatal diagnosis still depends on invasive methods using cells contained in the amniotic fluid or villus cells (Suzumori, 1999). These procedures carry a small but important rate of miscarriage (Vyas, 1994). The discovery on the presence of foetal cells in maternal circulation was a turning point for the development of a safe, accurate and reliable prenatal diagnosis, which could be offered to as a routine test to any pregnant women regardless their age

A plethora of human pluripotent stem cells

At the early stages of mammalian development, a number of developmentally plastic cells appear that possess the ability to give rise to all of the differentiated cell types normally derived from the three primary germ layers - unique character known as pluripotency. To date, embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have been shown to be truly pluripotent. However, recent studies have revealed a variety of other cells that demonstrate pluripotentiality, including very small embryonic-like stem cells (VSELs), amniotic fluid stem cells (AFSCs), marrow-isolated adult multilineage inducible cells (MIAMI) and multipotent adult precursor cells (MAPCs). This review summarises key features of these six kinds of pluripotent and potentially pluripotent stem cells (ESCs, iPSCs, VSELs, AFSCs, MIAMI and MAPCs) and the evidence for their pluripotency properties

Differentially expressed proteins in ER+ MCF7 and ER- MDA- MB-231 human breast cancer cells by RhoGDI-α silencing and overexpression

Background: The consequence of Rho GDP dissociation inhibitor alpha (RhoGDIα) activity on migration and invasion of estrogen receptor positive (ER+) and negative (ER-) breast cancer cells has not been studied using the proteomic approach. Changes in expression of RhoGDIα and other proteins interacting directly or indirectly with RhoGDIα in MCF7 and MDA-MB-231, with different metastatic potentials is of particular interest. Materials and Methods: ER+ MCF7 and ER- MDA-MB-231 cell lines were subjected to two-dimensional electrophoresis (2-DE) and spots of interest were identified by matrix-assisted laser desorption/ionization time of- flight/timeof- flight (MALDI-TOF/TOF) mass spectrometry (MS) analysis after downregulation of RhoGDIα using short interfering RNA (siRNA) and upregulated using GFP-tagged ORF clone of RhoGDIα. Results: The results showed a total of 35 proteins that were either up- or down-regulated in these cells. Here we identifed 9 and 15 proteins differentially expressed with silencing of RhoGDIα in MCF-7 and the MDA-MB-231 cells, respectively. In addition, 10 proteins were differentially expressed in the upregulation of RhoGDIα in MCF7, while only one protein was identified in the upregulation of RhoGDIα in MDA-MB-231. Based on the biological functions of these proteins, the results revealed that proteins involved in cell migration are more strongly altered with RhoGDI-α activity. Although several of these proteins have been previously indicated in tumorigenesis and invasiveness of breast cancer cells, some ohave not been previously reported to be involved in breast cancer migration. Hence, these proteins may serve as useful candidate biomarkers for tumorigenesis and invasiveness of breast cancer cells. Conclusions: Future studies are needed to determine the mechanisms by which these proteins regulate cell migration. The combination of RhoGDIα with other potential biomarkers may be a more promising approach in the inhibition of breast cancer cell migration

Spontaneous Unexplained Preterm Labor with Intact Membrane: Finding Protein Biomarkers through Placenta Proteome

Spontaneous unexplained preterm labor with intact membrane (sPTL-IM) remains as an unresolved challenge in obstetrics due to the complex syndromes involved during preterm birth. Two dimensional-gel electrophoresis (2D-GE) coupled with matrix-assisted laser desorption/ionization-time of flight/time of flight (MALDI TOF/TOF) mass spectrometry has become an alternative in screening for potential novel protein-based biomarkers and revealing the pathophysiology of sPTL-IM. To achieve this objective, protein extracted from fetal and maternal sides of the placenta obtained from sPTL-IM (n = 5) and the respective control (n = 10) groups were separated and compared using 2D-gel electrophoresis. MALDI-TOF/TOF mass spectrometry was utilized to identify the differentially expressed proteins between both groups, and the molecular functions of these proteins were studied. A total of 12 proteins were significantly differentiated in sPTL-IM over the control. Differentially expressed proteins were identified to have involved in structural/cytoskeletal components, immune responses, fetal and placenta development, and anticoagulation cascade. More proteins were found to be differentially expressed in the fetal side compared to the maternal side of the placenta. This postulates that the influence of sPTL-IM from fetus is greater than that of the mother. Ultimately, these results might lead to further investigations in elucidating the potential of these proteins as biomarkers and/or drug targets

Characteristics of Full-Term Amniotic Fluid-Derived Mesenchymal Stem Cells in Different Culture Media

Amniotic fluid contains precious therapeutic stem cells with ideal features such as they are broadly multipotent, genetically stable, and non-tumorigenic. One of the stem cells that is abundantly found in amniotic fluid is mesenchymal stem cells. Human amniotic fluid mesenchymal stem cells (hAFMSCs) had been successfully isolated from amniotic fluid obtained from second or third trimester amniocentesis. However, studies on hAFMSCs obtained during full-term delivery are still lacking. Furthermore, suitable culture media to propagate hAFMSCs for therapeutic purposes have not been fully established. Basal medium supplemented with fetal bovine serum is commonly used, and unfortunately, this condition has been associated with the risk of transmission of animal pathogens and xenogenic immune reaction. An efficient isolation and expansion method together with suitable culture conditions is essential in establishing a specific homogenous cell population, such as full-term hAFMSCs, of clinical grade. In this chapter we briefly describe the feasibility of generating hAFMSCs from full-term amniotic fluid obtained during cesarean section using serum-free medium as opposed to the conventional serum containing media. These findings would be very useful in utilizing stem cells for bench side application from a source that is accessible and devoid of ethical and safety concerns

Mesenchymal stem cells: from stem cells to sarcomas

Mesenchymal stem cells (MSCs) have garnered vast interests in clinical settings, especially in regenerative medicine due to their unique properties—they are reliably isolated and expanded from various tissue sources; they are able to differentiate into mesodermal tissues such as bones, cartilages, adipose tissues, and muscles; and they have unique immunosuppressive properties. However, there are some concerns pertaining to the role of MSCs in the human body. On one hand, they are crucial component in the regeneration and repair of the human body. On the contrary, they are shown to transform into sarcomas. Although the exact mechanisms are still unknown, many new leads have pointed to the belief that MSCs do play a role in sarcomagenesis. This review focuses on the current updates and findings of the role of MSCs in their transformation process into sarcomas

Determination of polycyclic aromatic hydrocarbons in human blood samples using solid phase extraction and gas chromatography mass spectrometry- a review

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous pollutants with toxic effects and adverse health impacts on general population. Several methods of extraction had been applied to extract PAHs from human blood samples such as solid phase extraction (SPE). The SPE represents one of the most common techniques for extraction and clean-up procedures as it needs low quantity of solvents with less manual efforts. Similarly, various analytical instruments like gas chromatography coupled to mass spectrometry (GC-MS) was used to measure the PAHs levels. Gas chromatography is a simple, fast, and very efficient method for solvents and small organic molecules. This review provides an overview of the measured concentrations of PAHs in human blood samples through the application of SPE and GC-MS during the last ten years. While these studies used various solvents, their application of SPE method and GC-MS revealed rewarding results about the determination of PAHs levels in the human samples