Quantification of human amniotic fluid constituents by high resolution proton nuclear magnetic resonance (NMR) spectroscopy

We have investigated the ability of high-resolution proton NMR spectroscopy to provide a biochemical constituent screening of human amniotic fluid (AF). Proton NMR spectra were obtained at 300 MHz on AF from patients undergoing amniocentesis in the mid-trimester. Only AF from normal pregnancies (normal fetal karyotype, normal a-fetoprotein levels, normal birth outcome) was used in this study. The AF supernatant was lyophilized and resuspended in deuterated water containing 0.1 mm phosphate buffer and 6.02 mm disodium maleate. Identification of low molecular weight compounds was confirmed by two-dimensional NMR spectra (primarily correlated spectroscopy, or COSY) and standard addition techniques. A broad profile of compounds were ‘NMR visible’ in a single proton spectrum, including creatinine, glucose, organic acids (acetate, citrate, and lactate) and several amino acids (alanine, histidine, leucine, phenylalanine, tyrosine and valine). The proton spectrum was unaffected by prior freezing/thawing of AF samples. We were able to quantify compounds by comparison with an added concentration standard (maleate) at concentrations as low as 30 μm. Good agreement with literature values based on other analytical techniques was obtained.     

High resolution proton NMR spectroscopy of human amniotic fluid

Human amniotic fluid (HAF) is a dynamic system whose characteristics depend on continuous interchanges between fetal and maternal circulations. HAF reflects not only the environment of the fetus but may also provide information about fetal development or pathology. The concentration of HAF constituents varies with gestational age and pathological states. The number of the compounds currently implicated in fetal developmental pathology are relatively few. Currently used assay methods are not adequate to totally explain or predict the complex biochemistry of the fetus. The purpose of this work was to investigate HAF with NMR spectroscopy. In the present study HAF was obtained from 47 women undergoing routine amniocentesis. Cells were separated for karyological analysis and the supernatant was acid-extracted, lyophilized and re-suspended in D₂0 resulting in a concentration increase over native fluid. ¹H NMR spectra were obtained at 360 MHz and 60 MHz. Eighteen compounds including several amino acids, were identified using parallel reference and standard addition protocols. NMR spectroscopy detected compounds of known clinical importance including glucose, leucine, isoleucine, lactate and creatinine. In conclusion, we have demonstrated that a number of physiologically relevant compounds are readily observable in HAF using ¹H NMR spectroscopy. This technique can currently provide valuable information regarding HAF composition and has the potential of being used in vivo in the future.     

Prenatal diagnosis of the infantile type of neuronal ceroid lipofuscinosis by electron microscopic investigation of human chorionic villi

Chorionic villus biopsy specimens were studied electron microscopically in six pregnancies at risk of the infantile type of neuronal ceroid lipofuscinosis (INCL). The biopsy was performed in all cases in the first trimester of pregnancy (8–10 gestation weeks) by the transcervical route. In one case, the biopsy was repeated at 17 weeks by the transabdominal procedure. In two pregnancies, the endothelial cells and, to a lesser extent, the mesenchymal cells of the chorionic villi contained unit membrane-bound inclusions typical of INCL. In both cases, the pregnancy was terminated and in one of them identical inclusions were found in the brains and kidneys of the fetus at 20 weeks of gestational age. The children from the remaining four pregnancies are healthy and have shown no signs of the disease.     

五氯酚钠及二(口恶)(口英)残留毒物对人外周血淋巴细胞的影响

采用彗星试验和微核试验,协同探索了五氯酚钠(Na-PCP)、二(口恶)(口英)类物质联合作用对血吸虫病流行区和正常生活区的人体外周血淋巴细胞影响效应.结果表明,实际用药区居民的外周血淋巴细胞以及用五氯酚钠与二(口恶)(口英)类物质联合作用处理过的正常人外周血淋巴细胞,其彗星率与微核率均高于对照组,在一定浓度范围内有剂量效应关系;基本的联合毒性作用推测为Na-PCP中的杂质PCDs与PCDFs等多种化学物质会损伤人体外周血淋巴细胞的DNA,其毒性作用比单一的二(口恶)(口英)类物质作用强.     

应用~(32)P-后标记技术对苯醌和氢醌处理人血淋巴细胞形成DNA加合物的研究

应用核酸酶Ｐ１促进的３２Ｐ－后标记技术对苯醌和氢醌处理人血淋巴细胞后形成的ＤＮＡ加合物进行了研究。结果表明，人血淋巴细胞被苯醌和氢醌处理２４ｈ之后，均形成了同一种ＤＮＡ加合物。两种化合物在不同浓度下处理人血淋巴细胞每１０７核苷酸中有０．０１～１０个核苷酸形成加合物。为了达到同等ＤＮＡ加合物水平，氢醌比苯醌需要更高的浓度。小牛胸腺ＤＮＡ与苯醌、氢醌反应可生成５种ＤＮＡ加合物。用苯醌或氢醌处理淋巴细胞所形成的加合物种类与上述纯ＤＮＡ与苯醌，氢醌形成的５种加合物在双向薄层层折图谱上完全不相符合。这些结果意味着苯醌、氢醌与ＤＮＡ的加合作用在细胞内存在着修饰机制。     

Mucolipidosis IV: Prenatal diagnosis by electron microscopy

Mucolipidosis IV (ML 1V) is a lysosomal storage disease presenting in infancy with cloudy cornea and psychomotor retardation. Our experience with 12 pregnancies at risk for ML IV, monitored by transmission electron microscopy (TEM) studies of cultured amniotic fluid cells, is presented. The prenatal diagnoses were confirmed in the 3 affected and the 8 un- affected pregnancies. In the one pregnancy where no definite diagnosis was reached the pregnancy was terminated. TEM examination of fetal tissues from this pregnancy showed no abnormal lysosomal storage bodies and a review of the cultured amniotic fluid cell sections revealed that the diagnosis of a normal fetus could have been made.