Metabonomics And Metallomics Study On Birth Defects Using Combined Chromatography-Mass Spectrometry Technology

Birth defects (BDs) are the developmental anomaly statuses before the fetus was born, which including morphological, structural, functional, metabolic and psychoneurotic abnormalities of fetus during the fetal development process. BDs are becoming one of inducements that lead to the death of fetus and disability of child and adults along with the rapid population growth and transformation of disease. In China, there are 51 millions of disabled person and 22 millions of inborn error of metabolism patients and BDs are the main causes of these disabilities, which imposed great financial burden upon our country and society. As multi-factorial and polygenic diseases, BDs have extreme complex etiologies which included not only hereditary defects such as chromosomal and gene abnormality, but also environmental factor based malformation such as poor nutrition, drugs and environmental pollution, and therefore, to find out the BDs related specific biomarkers is the highlight of the diagnosis and therapy of these formidable diseases. To date, it is hard to find out effective prevention and therapy of NTDs except of folic acid supplement during the before and after first trimester of pregnancy. The current prenatal diagnosis methods included imageological results such as transonogram and computed tomography, which are not suitable for most BDs due to these imagings are often late deformity results. There are no specific biomarkers or metabolic profiles in the current clinical biochemistry detection which also limited its usage in BDs diagnosis.Metabonomics or metabolomics is the developing"omics"which quantitative measurement of metabolites, as an integral part of the systems biology encompassing other omics sciences such as genomics, transcriptomics and proteomics. Metabonomics is the quantitative and qualitative measurement of the multiparametric time-related metabolic responses of a complex system to a pathophysiological intervention or genetic modification. The main purpose of metabonomics is to elucidate endogenous metabolic pathway on different biological levels such as system, organ, tissue or cell level. The main characteristics of metabonomics are the high-throughput chemical analysis and multivariate data analysis. Metabonomics based technologies can find out the specific biomarkers or metabolic profiling of complex diseases at nosogenesis, development and prognosis. Metabonomics has become an important highlight for the research of etiology, early diagnosis and drug intervention. Metallomics is another newly developed"omics"science emerging from genomics, transcriptomics, proteomics and metabonomics. The main scopes of metallomics are not only to depict chemical composition of metal and functional protein combination, but also to elucidate the function of metalloprotein and metal enzyme in biosystems. Nowadays, metal based biochemical reaction can not be fully understood on molecular level and the mechanism of metal coenzyme is also a secret. The emphasis and nodus of metallomics is to elucidate the function of metal and its related compound in normal vital movement, etiology and development of complex diseases.In the current dissertation, we tried to establish chromatography in hyphenation with mass spectrometry based metabonomic and metallomic technology to investigate metabolic and trace elemental variation in prenatal malnutrition pregnant rats and pregnant women who were pregnant with BDs fetus and to investigate the applicability of such metabonomics and metallomics method in BDs diagnosis and mechanism research.We used trimethyl silane derivatization, gas chromatography-mass spectrometry (GC/MS) and inductively coupled plasma mass spectrometry (ICP/MS) based metabonomic and metallomic analysis of amniotic fluid (AF) from prenatal malnutrition pregnant rats. PCA scores plot of both GC/MS and ICP/MS data showed similar and unique metabolic signatures of AF in response to the different diets. Rats in the famine group released decreased amounts of methionine, dopa, glutamine, zinc, cobalt, and selenium in the AF. These discriminable variations in the AF may indicate the abnormality of a number of metabolic pathways in fetal rats including the folate cycle and methionine pathway, the monoamine pathway and necessary trace elements metabolism. This study demonstrates the potential of combining profiling of small-molecule metabolites and trace elements to broaden the understanding of biological variations associated with fetal neurodevelopment induced by environmental perturbation.GC/MS and GC/TOFMS based metabonomic approach was used to investigate the BDs related metabolites profiling variation of the urinary and serum samples from pregnant women who were pregnant with neural tube defects (NTDs) and embryo stasimorphy fetus. OPLS-DA model of both GC/MS and GC/TOFMS data showed significant separation tendency between urinary and serum samples on case and control groups. Urine sample from embryo stasimorphy group released increased amounts of methylmalonic acid and decreased amounts of necessary amino acid (MMA), serum sample from NTDs group released decreased amounts of methionine. The urinary and serum metabolites variations in BDs pregnant women may indicate the abnormality of a number of metabolic pathways including methionine cycle, folic acid cycle, homocysteine metabolism, MMA metabolism and TCA. This study demonstrates the potential of metabolic profiling of small-molecule metabolites as a early diagnosis tool to distinguish BDs from healthy control and broaden the understanding of the etiology and mechanism of BDs.UPLC/QTOFMS metabonomic approach was used to investigate the BDs related high molecular weight metabolites variation of the urinary and serum samples from pregnant women who were pregnant with neural tube defects (NTDs) and embryo stasimorphy fetus. OPLS-DA model of UPLC/QTOFMS data showed separation tendency between urinary samples on case and control groups. Serum sample from NTDs group released decreased level of folic acid, vitaminB6 and vitaminB12 as compared with healthy pregnant women.This study broaden the metabolic profiling analytical window in the BDs related diseases.The nitric acid digestion ICP/MS method based metallomics strategy was used to analysis serum trace elements profiling of healthy and BDs pregnant women. OPLS-DA model of ICP/MS data showed separation tendency between NTDs and healthy pregnant women. Serum sample from NTDs group released decreased level of cobalt and increased level of heavy metal such as lead, mercury and chromium as compared with healthy pregnant women. This study demonstrates the potential of trace elements profiling to broaden the understanding of the mechanism of neural tube defects...