Inhibition of lipoprotein lipase activity by synthetic peptides of apolipoprotein C-III.

In this study we have examined effects of synthetic polypeptide fragments of apoC-III on the kinetic properties of lipoprotein lipase (LPL) activity. Based on the loss of 79% of LPL-inhibitory activity after CNBr cleavage at the N-terminal portion of apoC-III and a systematic search for synthetic peptides with LPL-inhibitory activity spanning the apoC-III sequence, we concluded that the N-terminal domain is the most important in the modulation of LPL activity. In addition, there are multiple attachment sites in apoC-III for its interaction with LPL and these sites reside in the hydrophilic sequences of apoC-III. Probably for this reason the intact apo-CIII exhibited higher inhibitory potential than its peptide components. Based on the deduced inhibition constants derived for the synthetic apoC-III1-79 we concluded that apoC-III is likely to exhibit a physiological role in regulating LPL activity since the derived dissociation constants for the LPL-apoC-III interaction are within the physiological concentration range of plasma apoC-III. In addition, as the synthetic apoC-III1-79 lacks the carbohydrate moiety, we also concluded that the presence of the oligosaccharide in native apoC-III is not essential for its inhibitory activity on LPL. The fact that the I50 (concentration for inhibition of LPL at 50% activity) decreases for apoC-III-1 when assayed in the presence of apoC-II indicated that the activator actually caused an increased affinity between LPL and apoC-III and demonstrated that apoC-III does not compete for the activator site of apoC-II.     

Role of LCAT in HDL remodeling: investigation of LCAT deficiency states

To better understand the role of LCAT in HDL metabolism, we compared HDL subpopulations in subjects with homozygous (n = 11) and heterozygous (n = 11) LCAT deficiency with controls (n = 22). Distribution and concentrations of apolipoprotein A-I (apoA-I)-, apoA-II-, apoA-IV-, apoC-I-, apoC-III-, and apoE-containing HDL subpopulations were assessed. Compared with controls, homozygotes and heterozygotes had lower LCAT masses (-77% and -13%), and LCAT activities (-99% and -39%), respectively. In homozygotes, the majority of apoA-I was found in small, disc-shaped, poorly lipidated prebeta-1 and alpha-4 HDL particles, and some apoA-I was found in larger, lipid-poor, discoidal HDL particles with alpha-mobility. No apoC-I-containing HDL was noted, and all apoA-II and apoC-III was detected in lipid-poor, prebeta-mobility particles. ApoE-containing particles were more disperse than normal. ApoA-IV-containing particles were normal. Heterozygotes had profiles similar to controls, except that apoC-III was found only in small HDL with prebeta-mobility. Our data are consistent with the concepts that LCAT activity: 1) is essential for developing large, spherical, apoA-I-containing HDL and for the formation of normal-sized apoC-I and apoC-III HDL; and 2) has little affect on the conversion of prebeta-1 into alpha-4 HDL, only slight effects on apoE HDL, and no effect on apoA-IV HDL particles.     

Serum HPL level in maternal vein, umbilical cord artery and vein in term and preterm birth

The serum HPL level in maternal vein and the cord vein and artery in term (n = 34) and preterm (n = 74) birth was tested to classify the hormonal changes and the correlation between the maternal and fetal HPL values. It was found that between the 28th and the 40th weeks of gestation the maternal level tended to increase while in the cord vein and artery it decreased significantly in negative correlation with the weeks of pregnancy. The maternal serum HPL level was significantly higher than that of the cord vein and artery. Between the 28th and 40th week the value in the cord artery divided by the value in the cord vein multiplied by 100 tended to decrease. While the HPL concentration in maternal serum did not correlate with its level in the cord vein and artery the levels in the cord vein and artery were significantly correlated. The characteristic and independent change in the fetal serum HPL concentration raised the possibility of a change in the transport function of the placenta and of an autonomous fetal HPL metabolism.     

Characterization of the lipid-binding properties and lipoprotein lipase inhibition of a novel apolipoprotein C-III variant Ala23Thr

We have identified a G-to-A transition in exon 3 of the APOC3 gene resulting in a novel Ala23Thr apolipoprotein (apo) C-III variant, associated with apoC-III deficiency in three unrelated Yucatan Indians. The Ala23Thr substitution modifies the hydrophobic/hydrophilic repartition of the helical N-terminal peptide and hence could disturb the lipid association. In vitro expression in Escherichia coli of wild-type and mutant apoC-III enabled the characterization of the variant. Compared with wild-type apoC-III-Ala23, the mutant apoC-III-Thr23 showed reduced affinity for dimyristoylphosphatidylcholine (DMPC) multilamellar vesicles with higher amounts of free apoC-III. Displacement of apoE from discoidal apoE:dipalmitoylphosphatidycholine (DPPC) complex by apoC-III-Thr23 was comparable to wild type but the less efficient binding of the apoC-III-Thr23 to the discoidal complex resulted in a higher apoE/apoC-III (mol/mol) ratio (34%) than with wild-type/apoE:DPPC mixtures. The inhibition of lipoprotein lipase (LPL) by apoC-III-Thr23 was comparable to that of wild type, and therefore effects on LPL activity could not explain the lower triglyceride (Tg) levels in Thr-23 carriers. Thus, these in vitro results suggest that in vivo the less efficient lipid binding of apoC-III-Thr23 might lead to a faster catabolism of free apoC-III, reflected in the reduced plasma apoC-III levels identified in Thr-23 carriers, and poorer competition with apoE, which might enhance clearance of Tg-rich lipoproteins and lower plasma Tg levels seen in Thr-23 carriers.     

Interaction of the apoproteins of very low density and high density lipoproteins with synthetic phospholipids.

The interaction of synthetic dimyristoyl phosphatidylcholine (lecithin) liposomes with isolated apoC-I and apoC-III proteins from very low density lipoproteins has been studied by microcalorimetry. Complex formation is a highly exothermal process characterized by a maximal enthalpy of -130 kcal/mol (-544 kJ) apoC-III-1 and -65 kcal/mol apoC-I proteins (-272 kJ). The complex composition determined after its isolation by ultracentrifugal flotation agrees with the value derived from the enthalpy binding curves. The binding of a constant amount of dimyristoyl lecithin to apoprotein mixtures containing various proportions of apoA-I and apoC-III failed to demonstrate the existence of any preferential association between the two apoproteins, in contrast with results obtained previously with apoA-I/apoA-II protein mixtures. Finally the various contributions to the enthalpy of binding such as that arising from an increase in apoprotein helicity have been evaluated. A classification of the apolipoproteins according to their lipid-binding affinity is proposed as: apoA-II congruent to apoC-III greater than apoC-I greater than apoA-I proteins.     

Epitope analysis of GAD65 binding in both cord blood and at the time of clinical diagnosis of childhood type 1 diabetes.

The GAD65 epitope immunoglobulin binding pattern in cord blood of children (n=37), who later developed type 1 diabetes at 3.2-14.9 years of age, was analyzed. First, the binding at diagnosis was compared with that in the cord blood serum. The next comparison was between the cord blood serum and the mothers' serum taken at delivery. Basal GAD65 binding levels were determined in Protein A Sepharose-based radiobinding assays with (35)S-labeled human and rat GAD65, rat GAD67 and GAD65/67 fusion proteins representing N-terminal (N), middle (M) and C-terminal (C) epitopes. In the first comparison, 28/37 children had GAD65 binding above 2.44 relative units (RU) (upper three quartiles), representing a marked increase from birth in the binding to human GAD65 (p<0.0001), rat GAD65 (p<0.0001), N- (p=0.04), M- (p<0.0001), C- (p=0.001), and M + C-epitopes (p<0.0001), but not to rat GAD67. At birth, 9/37 had GAD65 binding above 1.56 RU (upper quartile) demonstrating that their binding of human (35)S-GAD65 was higher in cord blood than in the mother (p=0.008). Higher cord blood binding was also observed for the N- (p=0.02) terminal epitope but not for rat GAD65, rat GAD67, and the remaining epitopes. These data suggest that differences in the epitope GAD65 binding between mother and child at birth are limited. In contrast, the epitope pattern at diagnosis differed from that at birth, supporting the view that disease-associated epitopes develop between birth and diagnosis.     

Reduction in lipoprotein-associated apoC-III levels following volanesorsen therapy: phase 2 randomized trial results

Elevated apoC-III levels predict increased cardiovascular risk when present on LDL and HDL particles. We developed novel high-throughput chemiluminescent ELISAs that capture apoB, lipoprotein (a) [Lp(a)], and apoA-I in plasma and then detect apoC-III on these individual lipoproteins as apoCIII-apoB, apoCIII-Lp(a), and apoCIII-apoAI complexes, respectively. We assessed the effects on these complexes of placebo or 100–300 mg volanesorsen, a generation 2.0+ antisense drug that targets apoC3 mRNA in patients with hypertriglyceridemia, including familial chylomicronemia syndrome (n = 3), volanesorsen monotherapy (n = 51), and as add-on to fibrate (n = 26), treated for 85 days and followed for 176 days. Compared with placebo, volanesorsen was associated with an 82.3 ± 11.7%, 81.3 ± 15.7%, and 80.8 ± 13.6% reduction in apoCIII-apoB, apoCIII-Lp(a), and apoCIII-apoA-I, respectively (300 mg dose; P < 0.001 for all), at day 92. Strong correlations in all assay measures were noted with total plasma apoC-III, chylomicron-apoC-III, and VLDL-apoC-III. In conclusion, novel high-throughput ELISAs were developed to detect lipoprotein-associated apoC-III, including for the first time on Lp(a). Volanesorsen uniformly lowers apoC-III on apoB-100, Lp(a), and apoA-I lipoproteins, and may be a potent agent to reduce triglycerides and cardiovascular risk mediated by apoC-III.     

Administration of a PPARalpha agonist increases serum apolipoprotein A-V levels and the apolipoprotein A-V/apolipoprotein C-III ratio

Apolipoprotein A-V (apoA-V) first gained attention as a regulator of triglycerides through transgenic mouse studies. Furthermore, peroxisome proliferator-activated receptor alpha (PPARalpha) agonists such as fenofibrate increase apoA-V mRNA expression. Our group recently developed the first assay to quantitate serum apoA-V levels. Therefore, we sought to determine whether administration of a PPARalpha agonist would increase circulating apoA-V. Cynomolgus monkeys were dosed for 14 days with 0.3 mg/kg/day LY570977 L-lysine, a potent and selective PPARalpha agonist. Blood samples were drawn throughout the treatment period and after a 2 week washout. Administration of the PPARalpha agonist caused a 50% decrease in triglycerides that reversed at washout. Serum apoA-V concentrations increased 2-fold, correlated inversely with triglycerides, and were reversible at washout. The apoA-V/apoC-III ratio increased >2-fold, with this increase also reversible at washout. These data demonstrate for the first time that a PPARalpha agonist increases circulating apoA-V protein levels and the apoA-V/apoC-III ratio.     

Molecular cloning of a human apoC-III variant: Thr 74----Ala 74 mutation prevents O-glycosylation

Apolipoprotein C-III (apoC-III) is a major protein of very low density lipoprotein (VLDL). The apoC-III polypeptide contains a carbohydrate chain containing galactosamine, galactose, and sialic acid attached in O-linkage to a threonine residue at position 74. We have cloned the apoC-III gene from a subject whose serum contained unusually high amounts of apoC-III lacking the carbohydrate moiety (C-III-0). DNA sequence analysis of the cloned gene revealed a single nucleotide substitution (A----G) that encodes an alanine at position 74 instead of the normal threonine. As a result of this amino acid replacement, the mutant apoC-III polypeptide is not glycosylated. The mutation in the apoC-III gene creates a novel AluI site that permits diagnosis of the change by Southern blotting of genomic DNA.     

Maternal BMI Associations with Maternal and Cord Blood Vitamin D Levels in a North American Subset of Hyperglycemia and Adverse Pregnancy Outcome (HAPO) Study Participants

ObjectiveObesity in pregnancy may be associated with reduced placental transfer of 25-hydroxyvitamin D (25-OHD). The objective of this study was to examine associations between maternal BMI and maternal and cord blood levels of 25-OHD in full term neonates born to a single racial cohort residing at similar latitude. Secondary objectives were to examine associations between maternal glucose tolerance with maternal levels of 25-OHD and the relationship between cord blood 25-OHD levels and neonatal size.MethodsThis study was conducted among participants of the Hyperglycemia and Adverse Pregnancy Outcomes (HAPO) Study meeting the following criteria: residing at latitudes 41–43°, maternal white race, and gestational age 39–41 weeks. Healthy pregnant women underwent measures of height, weight, and a 75-g fasting oral glucose tolerance test (OGTT) at approximately 28 weeks gestation. Maternal and cord blood sera were analyzed for total 25-OHD by HPLC tandem mass spectrometry. Statistical analyses included ANOVA and linear regression models.ResultsMaternal and cord blood (N = 360) mean levels (sd) of 25-OHD were 37.2 (11.2) and 23.4 (9.2) ng/ml, respectively, and these levels were significantly different among the 3 field centers (ANOVA p< 0.001). Maternal serum 25-OHD was lower by 0.40 ng/ml for BMI higher by 1 kg/m² (p<0.001) in an adjusted model. Maternal fasting plasma glucose, insulin sensitivity, and presence of GDM were not associated with maternal serum 25-OHD level when adjusted for maternal BMI. Cord blood 25-OHD was lower by 0.26 ng/ml for maternal BMI higher by 1 kg/m² (p<0.004). With adjustment for maternal age, field center, birth season and maternal serum 25-OHD, the association of cord blood 25-OHD with maternal BMI was attenuated. Neither birth weight nor neonatal adiposity was significantly associated with cord blood 25-OHD levels.ConclusionThese results suggest that maternal levels of 25-OHD are associated with maternal BMI. The results also suggest that interpretation of neonatal 25-OHD levels may need to incorporate specific maternal factors in addition to season of birth and latitude.     

Electrostatically mediated interactions between cationic lipid-DNA particles and an anionic surface.

In an effort to model the interaction of lipid-based DNA delivery systems with anionic surfaces, such as a cell membrane, we have utilized microelectrophoresis to characterize how electrokinetic measurements can provide information on surface charge and binding characteristics. We have established that cationic lipids, specifically N-N-dioleoyl-N,N-dimethylammonium chloride (DODAC), incorporated into liposomes prepared with 1, 2-dioleoyl-i-glycero-3-phosphoethanolamine (DOPE) or 1, 2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) at 50 mol%, change the inherent electrophoretic mobility of anionic latex polystyrene beads. Self-assembling lipid-DNA particles (LDPs), prepared at various cationic lipid to negative DNA phosphate charge ratios, effected no changes in bead mobility when the LDP charge ratio (+/-) was equal to or less than 1. Increasing the LDP concentration in a solution of 0.1% (w/v) anionic beads resulted in a charge reversal effect when a net charge of LDP to total bead charge ratio (+/-) of 1:1 was observed. LDP formulations, utilizing either DOPE or DOPC, showed similar titration profiles with a charge reversal observed at a 1:1 net LDP to bead charge ratio (+/-). It was confirmed through centrifugation studies that the DNA in the LDP was associated with the anionic latex beads through electrostatic interactions. LDP binding, rather than the binding of dissociated cationic lipids, resulted in the observed electrophoretic mobility changes of the anionic latex beads.     

Effect of maternal nutrient restriction in early to mid gestation and thyrotrophin-releasing hormone on lamb survival following Caesarean section delivery near to term

We investigated the influence of restricted maternal nutrition between 28 and 77 days gestation on survival and thermoregulatory adaptation following Caesarean section delivery near to term. This study was designed to examine the hypothesis that adaptation after birth would be compromised in those lambs born to nutrient restricted ewes. We further hypothesised that this would be due in part to inadequate hypothalamic-pituitary-thyroidal function. Lambs born to nutrient restricted ewes were untreated (RU) or treated with thyrotrophin-releasing hormone (TRH; RT) immediately prior to umbilical cord clamping. Single bearing ewes consumed either 6.60 MJ x day(-1) (controls, n = 4) or 3.00 MJ x day(-1) (nutrient restricted, n = 15) from 28-77 days gestation, after which all ewes consumed 7.20 MJ x day(-1). All lambs born to control ewes commenced continuous breathing and began to shiver following Caesarean section delivery and survived to 6 h after birth. Only 4 out of 9 RU lambs established continuous breathing and survived to 6 h after birth compared with all RT lambs. Six hours after birth, RT lambs possessed perirenal brown adipose tissue with a higher thermogenic activity than 6 h old RU or control lambs. Lamb birth weight was similar in all groups. In conclusion, near-term lambs born to ewes nutrient restricted in early to mid gestation are at increased risk of death following Caesarean section delivery. Survival after birth can be significantly enhanced if TRH is administered to the lambs immediately before delivery.     

Serum apolipoproteins C-I and C-III are reduced in stomach cancer patients: results from MALDI-based peptidome and immuno-based clinical assays

Finding new peptide biomarkers for stomach cancer in human sera that can be implemented into a clinically practicable prediction method for monitoring of stomach cancer. We studied the serum peptidome from two different biorepositories. We first employed a C8-reverse phase liquid chromatography approach for sample purification, followed by mass-spectrometry analysis. These were applied onto serum samples from cancer-free controls and stomach cancer patients at various clinical stages. We then created a bioinformatics analysis pipeline and identified peptide signature discriminating stomach adenocarcinoma patients from cancer-free controls. Matrix Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) results from 103 samples revealed 9 signature peptides; with prediction accuracy of 89% in the training set and 88% in the validation set. Three of the discriminating peptides discovered were fragments of Apolipoproteins C-I and C-III (apoC-I and C-III); we further quantified their serum levels, as well as CA19-9 and CRP, employing quantitative commercial-clinical assays in 142 samples. ApoC-I and apoC-III quantitative results correlated with the MS results. We then employed apoB-100-normalized apoC-I and apoC-III, CA19-9 and CRP levels to generate rules set for stomach cancer prediction. For training, we used sera from one repository, and for validation, we used sera from the second repository. Prediction accuracies of 88.4% and 74.4% were obtained in the training and validation sets, respectively. Serum levels of apoC-I and apoC-III combined with other clinical parameters can serve as a basis for the formulation of a diagnostic score for stomach cancer patients.     

Initiation of pulmonary gas exchange by fetal sheep in utero

The role of umbilical cord occlusion in the initiation of breathing at birth was investigated using unanesthetized fetal sheep that were provided with access to a tracheal supply of hyperoxic air. Near-term fetuses were studied in utero to eliminate extraneous sensory stimuli. Gasping movements began 1.4 +/- 0.1 min after cord occlusion. Breathing was irregular for several minutes before continuous breathing (greater than or equal to 40 min-1) began 6 +/- 1 min after cord occlusion (n = 10). Arterial PO2 rose significantly from 18 +/- 2 mmHg before occlusion and was 115 +/- 15 mmHg immediately before cord release at 15 or 30 min. Breathing continued even during high-voltage electrocortical activity. Cord release caused the breathing rate to decrease from 77 +/- 13 min-1 during the last 5 min of cord occlusion to 5 +/- 3 min-1 10 min after cord release (P less than 0.002; n = 7). Results indicate the change from placental to lung gas exchange can occur in the absence of sensory and thermal changes normally present at birth and that the transition is reversible.     

Altered proteinogram in short term portal vein stenosed rats

The electrophoretic pattern of serum proteins has been studied in short-term prehepatic portal hypertensive rats since atrophy is produced in the liver, which is the main origin of most of these proteins, during this postoperative period. After 28 days of evolution, rats (n = 9) with triple stenosing ligated portal vein showed hypoalbuminemia, hypo-alpha-globulinemia, hyper-alpha2-globulinemia and hyper-gamma-globulinemia, the albumin/globulin ratio decreased with respect to the control animals (n = 8). These alterations are associated with hepatic atrophy, portosystemic and portohepatic (44.4%) collateral circulation. The proteinogram alterations found in rats with short-term prehepatic portal hypertension suggest that hepatic failure exists in spite of potential portohepatic revascularization which is frequently originated by the development of portohepatic collateral circulation.     

不同时间采集的小牛血清对CHO-C28细胞培养及表达的影响

通过研究小牛血清对CHO-C28细胞培养及HBsAg表达的影响,探讨小牛血清的不同采集时间对血清质量的影响。采集出生后4、8、12h(未进食)小牛的血清,对CHO-C28细胞进行传代、换液培养,并检测乙肝表面抗原(HBsAg)表达量。结果可见:①在细胞传代4次时,4h采集的血清细胞生长状态良好,8h采集的血清细胞出现明显的衰老,12h采集的血清细胞大面积死亡;②在细胞维持换液方面,4h采集的血清可维持细胞换液25次以上,8h采集的血清可勉强维持20次,12h采集的血清维持细胞换液10次时已大部分死亡;③乙肝表面抗原(HBsAg)表达量的检测结果,同批培养上清,4h采集的血清培养细胞表达量最高。可见,小牛出生后采集血清时间越早越好。     

Concentrations of major plasma proteins in serum and whole-tissue extracts of porcine fetuses during development.

In extracts from fetuses up to 32 days of gestation, the major serum proteins were fetuin, alpha-fetoprotein and alpha 1-antitrypsin, but albumin was not detected. The concentration of all proteins rose with age until 40-50 days of gestation; and then the serum concentration of alpha-fetoprotein (2.9 mg ml-1), alpha 1-antitrypsin (4.4 mg ml-1) and transferrin (2.6 mg ml-1) fell progressively to about 1 mg ml-1 at birth, whereas those of fetuin, albumin and alpha 1-acid glycoprotein increased. The patterns of serum proteins in fetuses at about the middle of gestation were similar in extracts and sera. At birth, the major proteins were alpha 1-acid glycoprotein and fetuin, which accounted for 45 and 18% of serum proteins, respectively. Albumin represented only 7% of serum proteins at this age. For most of the second gestational period, the six quantified proteins accounted for about 85% of total serum proteins. In early gestation, a significant proportion of serum proteins was intracellular.     

血清瘦素和胰岛素与犊牛初生重相关性

本实验旨在研究分娩奶牛(Bos taurus)、犊牛和脐静脉血瘦素、胰岛素之间的相关性及与犊牛初生重的相关性,为探究瘦素、胰岛素对犊牛初生重的影响机理提供理论数据。实验选取规模化养殖场正常分娩奶牛54头,按犊牛初生重划分为A组(≤40 kg)9头、B组(40~45 kg)25头、C组(≥45 kg)20头共3组,分别采集分娩奶牛、犊牛和脐静脉血,ELISA法检测血清瘦素、胰岛素含量。多组间采用单因素方差分析和双变量Pearson分析瘦素和胰岛素在各部位静脉血中表达的相关性及与犊牛初生重的相关性。结果表明:(1)奶牛静脉血瘦素、胰岛素含量极显著高于犊牛静脉血和脐静脉血(P0.01),奶牛静脉血瘦素、胰岛素含量与犊牛静脉血和脐静脉血瘦素、胰岛素相关性均不显著(P0.05);(2)瘦素、胰岛素含量在犊牛静脉血与脐静脉血之间均差异不显著(P0.05),犊牛静脉血瘦素、胰岛素含量与脐静脉血瘦素、胰岛素均分别呈极显著正相关(P0.01);(3)奶牛静脉血和脐静脉血中瘦素与胰岛素含量均呈显著正相关(P0.05),而犊牛静脉血中瘦素与胰岛素相关性不显著(P0.05);(4)犊牛初生重与奶牛静脉血瘦素、胰岛素含量均不相关(P0.05),但与犊牛静脉血和脐静脉血瘦素含量显著正相关(P0.05),与犊牛静脉血和脐静脉血胰岛素含量极显著正相关(P0.01);(5)奶牛静脉血、犊牛静脉血和脐静脉血瘦素和胰岛素含量在公犊牛与母犊牛间均无显著差异(P0.05)。可见,奶牛静脉血、犊牛静脉血和脐静脉血均有瘦素、胰岛素表达,且其含量奶牛静脉血显著高于犊牛静脉血和脐静脉血。奶牛静脉血瘦素、胰岛素含量与犊牛初生重相关性不显著,而犊牛静脉血和脐静脉血瘦素、胰岛素含量与犊牛初生重显著正相关性。     

Patients with unsolved congenital disorders of glycosylation type II can be subdivided in six distinct biochemical groups

Defects in the biosynthesis of N- and core 1 O-glycans may be found by isoelectric focusing (IEF) of plasma transferrin and apolipoprotein C-III (apoC-III). We hypothesized that IEF of transferrin and apoC-III in combination with sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of apoC-III may provide a classification for congenital disorders of glycosylation (CDG) patients. We analyzed plasma from 22 patients with eight different and well-characterized CDG subtypes and 19 cases with unsolved CDG. Transferrin IEF (TIEF) has been used to distinguish between N-glycan assembly (type 1 profile) and processing (type 2 profile) defects. We differentiated two different CDG type 2 TIEF profiles: The "asialo profile" characterized by elevated levels of asialo- and monosialotransferrin and the "disialo profile" characterized by increased levels of disialo- and trisialotransferrin. ApoC-III IEF gave two abnormal profiles ("apoC-III(0)" and "apoC-III(1)" profiles). The results for the eight established CDG forms exactly matched the theoretical expectations, providing a validation for the study approach. The combination of the three electrophoretic techniques was not additionally informative for the CDG-Ix patients as they had normal apoC-III IEF patterns. However, the CDG-IIx patients could be further subdivided into six biochemical subgroups. The robustness of the methodology was supported by the fact that three patients with similar clinical features ended in the same subgroup and that another patient, classified in the "CDG-IIe subgroup," turned out to have a similar defect. Dividing the CDG-IIx patients in six subgroups narrows down drastically the options of the primary defect in each of the subgroups and will be helpful to define new CDG type II defects.     

新生儿巨细胞病毒感染的检测

建立了用ELISA检测巨细胞病毒(HCMV)IgA抗体的方法,並用于检测北京地区100对母婴的HCMV抗体,母血、脐带血、母乳中HCMV-IgG抗体的阳性率分别为83％,75％和38％,HCMV-IgA抗体的阳性率分别为19％,15％和58％,对其中的16名婴儿半年后追踪观察,5名出生时母、脐血全为阴性的,有2名抗体阳转。8名出生时母、脐血均阳性的,有1名IgA仍阳性並检查发现肝大肋下二指。另1名IgG持续阳性,其他6名婴儿抗体转阴。3名出生时母血HCMV-IgG阳性者中,1名婴儿IgA和‘gG转阳,此时母亲IgA也阳转。随访的16名婴儿中有3名可能是生后半年内受HCMV感染。