Synthesis of ABO histo-blood group type I and II antigens

The ABO histo-blood group system is one of the most clinically important antigen families. As part of our overall goal to prepare the entire set of the A, B and H type I-VI antigens for a range of biochemical investigations, we report herein the synthesis of the type I and II antigens with a 7-octen-1-yl aglycone. This linker was chosen to facilitate not only the future conjugation of the antigens to a protein or solid support but also the synthesis of the H type I and II octyl glycosides for enzyme kinetic studies.     

The distribution of type-2 chain histo-blood group antigens in normal cycling human endometrium

Summary The blood group ABO(H) determinants are major allogenic antigens in both erythrocytes and tissue of man. These antigens and related carbohydrates are markers of cellular maturation and differentiation in many epithelial tissues and have recently attracted great interest as tumor-associated antigens. Previous studies of endometrial tissues have indicated that glycosylation in this tissue may be related to hormonal stimulation. We have investigated the immunohistochemical distribution of type-2 chain histo-blood group-related carbohydrates in specimens of normal, cycling endometria obtained from hysterectomies on women with known ABO/Lewis erythrocyte type and saliva secretor status. N-acetyllactosamine and Le^x were demonstrated to be uninfluenced by the genetic background. A and Ale^y antigens were exclusively demonstrated in endometria from blood group A individuals, while Le^y was expressed in endometria from blood group 0 individuals mainly. The precursor N-acetyllactosamine as well as the terminal H, A, and ALe^y antigens were shown in only a few cells. In contrast, N-acetyllactosamine substituted by sialic acid and/or fucose residues (Le^x, sialosyl-Le^x, Le^y) were demonstrated in epithelial cells of normal, cycling endometrium, but with both quantitative and qualitative differences in staining relating to the menstrual cycle, indicating that type-2 chain antigens are expressed under both genetic and hormonal influence in human cycling endometrium.     

Expression of H type 1 antigen of ABO histo-blood group in normal colon and aberrant expressions of H type 2 and H type 3/4 antigens in colon cancer

We have immunohistochemically examined the distribution of the H antigens of type 1, type 2 and type 3/4 chains of the ABO(H) histo-blood group system in human normal colon and in colon cancer using three monoclonal antibodies specific for each of the H type 1/2, H type 2, and the H type 3/4 chain. We unexpectedly found that mucosa of the normal colon from secretors but not that from nonsecretors expressed only H type 1 and did not express H type 2 or H type 3/4. The H type 1 was expressed in goblet cells. Positive goblet cells expressing H type 1 were decreased in number progressively from the proximal colon to the rectum. In tumors, 4 (57%) of 7 cancer tissues of the proximal colon from secretors expressed no H type 1, whereas all 8 cancer tissues of the distal colon from secretors expressed H type 1. The aberrant expressions of H type 2 and H type 3/4 (47 and 67%, respectively) were found in cancer tissues from both the proximal and the distal colon. Tumors from nonsecretors did not express any H antigens. Our results suggested that the expression of H type 1 in the normal colon and the aberrant expressions of H type 2 and H type 3/4 in colon cancer tissues were regulated by FUT2-encoded Se type (1,2)fucosyltransferase. However, UEA-I-positive substance(s) rather than H type 2 were uniquely expressed throughout the normal colon and in colon cancers from both secretors and nonsecretors.     

ABO血型正反定型及交叉配血实验在外科病人手术输血中应用

目的:探讨ABO血型正反定型及交叉配血实验在外科手术患者输血中的应用效果及影响因素。方法:选取我院自2017年2月-2019年2月收治的80例行ABO正反定型与交叉配血治疗的外科手术患者,记录ABO反定型与交叉配血不合的标本,使用2-Me处理被患者自身冷抗体凝集的红细胞,同时使用微柱凝胶法、凝聚胺法对血型不规则抗体以及特异性进行筛选和鉴定。分析ABO血型反定型不符合以及交叉配血不合的影响因素。结果:对正反定型完全无凝集反应的80例血清标本进行交叉配血实验,其中8例存在凝集反应,配血不合情况;导致外科手术患者输血中ABO血型反定型不符交叉配血不合的主要因素包括自身冷抗体、血型抗原性减弱、血型不规则抗体以及血型抗体效价减弱等。结论:ABO血型正反定型及交叉配血治疗中的患者中,大部分配血一致,少数的交叉配血不合,主要与自身冷抗体、血型抗原性减弱、血型不规则抗体以及血型抗体效价减弱等因素相关。     

Ethnic differences in the expression of blood group antigens in the salivary gland secretory cells from German and Japanese non-secretor individuals

Type 1 ABO blood group antigens (peripheral core structure: Gal1-3GlcNAc1-R) are expressed mainly in endodermally-derived tissues, but are not synthesized in mesodermally-derived tissues. In the former tissues, H type 1 antigen is generated largely by -2-l-fucosyltransferase encoded by secretor (Se) gene and acting on the terminal galactose of the type 1 precursor chain. This theory has been generally accepted, and it seems that the expression of ABO blood group antigens is absent, or expressed at a low level, in these tissues from non-secretor individuals. In this immunohistochemical study on the secretory cells of salivary glands, we found ethnic difference between German and Japanese non-secretor individuals in the expression of blood group antigens: i.e. the expression of the type 1 blood group antigens is present in these cells from Japanese non-secretor individuals but absent from German. A possible explanation is that another -2-l-fucosyltransferase, independent of the secretor gene, is present in Japanese non-secretor individuals.     

ABO血型综合实验基因分型技术简化及其群体遗传分析拓展

ABO血型是生活中最常见、运用最广泛的遗传性状之一。人类ABO血型由I ^A、I ^B和i 3个复等位基因决定,它们负责编码不同的糖基转移酶,进而决定3种血红细胞表面抗原。ABO血型涉及复等位基因、基因互作、单核苷酸多态(SNP)、基因演化等多个关键知识内容,是理想的遗传学教学案例。本文以ABO血型为研究对象,对遗传学实验进行了创新与整合。首先,在分子遗传学模块中建立了新颖的ABO血型基因分型方法：基于SNP位点设计特异性引物,通过实时定量PCR鉴定基因型; 其次,在群体遗传学模块中创新了基因演化的实验教学方法,开发群体遗传学软件,利用计算机模拟不同条件下ABO血型决定基因频率的演化趋势。这些教学改革举措旨在丰富遗传学实验内容,拓展教学手段,提高学习效率。     

NMR Characterization of C3H and HCT Down-Regulated Alfalfa Lignin

Independent down-regulation of genes encoding p-coumarate 3-hydroxylase (C3H) and hydroxycinnamoyl CoA:shikimate/quinate hydroxycinnamoyl transferase (HCT) has been previously shown to reduce the recalcitrance of alfalfa and thereby improve the release of fermentable sugars during enzymatic hydrolysis. In this study, ball-milled lignins were isolated from wild-type control, C3H, and HCT gene down-regulated alfalfa plants. One- and two-dimensional nuclear magnetic resonance (NMR) techniques were utilized to determine structural changes in the ball-milled alfalfa lignins resulting from this genetic engineering. After C3H and HCT gene down-regulation, significant structural changes had occurred to the alfalfa ball-milled lignins compared to the wild-type control. A substantial increase in p-hydroxyphenyl units was observed in the transgenic alfalfa ball-milled lignins as well as a concomitant decrease in guaiacyl and syringyl units. Two-dimensional ¹³C–¹H heteronuclear single quantum coherence correlation NMR, one-dimensional distortionless enhancement by polarization transfer-135, and ¹³C NMR measurement showed a noteworthy decrease in methoxyl group and β-O-4 linkage contents in these transgenic alfalfa lignins. ¹³C NMR analysis estimated that C3H gene down-regulation reduced the methoxyl content by ~55–58% in the ball-milled lignin, while HCT down-regulation decreased methoxyl content by ~73%. The gene down-regulated C3H and HCT transgenic alfalfa lignin was largely a p-hydroxyphenyl (H) rich type lignin. Compared to the wild-type plant, the C3H and HCT transgenic lines had an increase in relative abundance of phenylcoumaran and resinol in the ball-milled lignins.     

ABO blood group system

The ABO blood group system in humans has three different carbohydrate antigens named A, B, and O. The A antigen sequence is terminal trisaccharide N-acetylgalactosamine (GalNAc)α1-3[Fucα1-2]Galβ-, B is terminal trisaccharide Galα1-3[Fucα1-2]Galβ-, and O is terminal disaccharide Fucα1-2Galβ-. The single ABO gene locus has three alleles types A, B and O. The A and B genes code A and B glycosyltransferases respectively and O encodes an inactive enzyme. A large allelic diversity has been found for A and B transferases resulting in the genetic subgrouping of each ABO blood type. Genes for both transferases have been cloned and the 3D structure of enzymes with and without substrate has been revealed by NMR and X ray crystallography. The ABO blood group system plays a vital role in transfusion, organ and tissue transplantation, as well as in cellular or molecular therapies.     

1H NMR metabonomics approach to the disease continuum of diabetic complications and premature death

Subtle metabolic changes precede and accompany chronic vascular complications, which are the primary causes of premature death in diabetes. To obtain a multimetabolite characterization of these high‐risk individuals, we measured proton nuclear magnetic resonance (¹H NMR) data from the serum of 613 patients with type I diabetes and a diverse spread of complications. We developed a new metabonomics framework to visualize and interpret the data and to link the metabolic profiles to the underlying diagnostic and biochemical variables. Our results indicate complex interactions between diabetic kidney disease, insulin resistance and the metabolic syndrome. We illustrate how a single ¹H NMR protocol is able to identify the polydiagnostic metabolite manifold of type I diabetes and how its alterations translate to clinical phenotypes, clustering of micro‐ and macrovascular complications, and mortality during several years of follow‐up. This work demonstrates the diffuse nature of complex vascular diseases and the limitations of single diagnostic biomarkers. However, it also promises cost‐effective solutions through high‐throughput analytics and advanced computational methods, as applied here in a case that is representative of the real clinical situation.     

Complete H and C NMR chemical shift assignments of mono-, di-, and trisaccharides as basis for NMR chemical shift predictions of polysaccharides using the computer program casper

The computer program casper uses ¹H and ¹³C NMR chemical shift data of mono- to trisaccharides for the prediction of chemical shifts of oligo- and polysaccharides. In order to improve the quality of these predictions the ¹H and ¹³C, as well as ³¹P when applicable, NMR chemical shifts of 30 mono-, di-, and trisaccharides were assigned. The reducing sugars gave two distinct sets of NMR resonances due to the α- and β-anomeric forms. In total 35 ¹H and ¹³C NMR chemical shift data sets were obtained from the oligosaccharides. One- and two-dimensional NMR experiments were used for the chemical shift assignments and special techniques were employed in some cases such as 2D ¹H,¹³C-HSQC Hadamard Transform methodology which was acquired approximately 45 times faster than a regular t₁ incremented ¹H,¹³C-HSQC experiment and a 1D ¹H,¹H-CSSF-TOCSY experiment which was able to distinguish spin-systems in which the target protons were only 3.3 Hz apart. The ¹H NMR chemical shifts were subsequently refined using total line-shape analysis with the PERCH NMR software. The acquired NMR data were then utilized in the casper program (http://www.casper.organ.su.se/casper/) for NMR chemical shift predictions of the O-antigen polysaccharides from Klebsiella O5, Shigella flexneri serotype X, and Salmonella arizonae O62. The data were compared to experimental data of the polysaccharides from the two former strains and the lipopolysaccharide of the latter strain showing excellent agreement between predicted and experimental ¹H and ¹³C NMR chemical shifts.     

B para-Bombay phenotype in China caused by homozygous mutation for site 328 G>A of FUT1 gene: a case report

The aim of this paper is to accurately identify a case of B para-Bombay and to analyze the genetic mutation. ABO and Lewis blood groups were identified by standard serological methods, and trace antigens on RBCs were detected by adsorption-elution test, while blood group substances in the saliva were detected by agglutination inhibition test. The ABO gene exons 6-7, FUT1 gene exon 4 and FUT2 gene exon 2 were directly sequenced. Serological results showed that there were B antigens on RBCs without H antigens, anti-A and anti-HI antibodies in serum, and B and H blood group substances in the saliva. The Lewis phenotype was Le (a-b+). According to gene sequencing analysis, ABO, FUT1 and FUT2 genotypes were B101/O02, h^328G/Ah^328G/A and Se^357C/TSe^357C/T, respectively. This rare phenotype can be mislabeled as "O" if any of the detailed investigations are not performed. Therefore, in order to ensure the safety of blood transfusion, genetic and serological tests are necessary for the correct identification of difficult blood groups.     

1H NMR Spectroscopy of pyrrolizidine alkaloids

The ¹H NMR spectra of three pyrrolizidine alkaloids of the macrocyclic diester type, retrorsine, seneciphylline and senecionine, plus their three N-oxides have been assigned. Previous ¹H NMR studies of these pyrrolizidine alkaloids have stressed the difficulties of spectral intrepretation. The results reported here will provide a useful resource for analysis of tertiary structure in these and related compounds.     

B para-Bombay phenotype in China caused by homozygous mutation for site 328 G > A of FUT1 gene: a case report

The aim of this paper is to accurately identify a case of B para-Bombay and to analyze the genetic mutation. ABO and Lewis blood groups were identified by standard serological methods, and trace antigens on RBCs were detected by adsorption-elution test, while blood group substances in the saliva were detected by agglutination inhibition test. The ABO gene exons 6-7, FUT1 gene exon 4 and FUT2 gene exon 2 were directly sequenced. Serological results showed that there were B antigens on RBCs without H antigens, anti-A and anti-HI antibodies in serum, and B and H blood group substances in the saliva. The Lewis phenotype was Le (a-b+). According to gene sequencing analysis, ABO, FUT1 and FUT2 genotypes were B101/O02, h^328G/Ah3^28G/A and Se^357C/TSe^357C/T, respectively. This rare phenotype can be mislabeled as "O" if any of the detailed investigations are not performed. Therefore, in order to ensure the safety of blood transfusion, genetic and serological tests are necessary for the correct identification of difficult blood groups.     

基于全外显子组数据的ABO血型的基因判定

全外显子组测序研究已经应用在疾病、药物等方面,是临床研究中一种辅助分子诊断方法。该方法也逐步应用在临床ABO血型的判定中,由于目前临床血型判定的主要方法是血清学,疑难样本判定等问题无法解决,因此分子水平的基因测序方法提高了血型判定的准确性,比如PCR方法和基因芯片方法。为进一步提高ABO血型精细分型的准确性,通过分析全外显子组测序数据,开发了相关的VBA程序,能够快速自动化判定ABO精细分型,并且初步判定结果与临床判定结果一致,可以作为临床血型精确判定的辅助手段。     

Towards miniaturization of a structural genomics pipeline using micro-expression and microcoil NMR

In structural genomics centers, nuclear magnetic resonance (NMR) screening is in increasing use as a tool to identify folded proteins that are promising targets for three-dimensional structure determination by X-ray crystallography or NMR spectroscopy. The use of 1D ¹H NMR spectra or 2D [¹H,¹⁵N]-correlation spectroscopy (COSY) typically requires milligram quantities of unlabeled or isotope-labeled protein, respectively. Here, we outline ways towards miniaturization of a structural genomics pipeline with NMR screening for folded globular proteins, using a high-density micro-fermentation device and a microcoil NMR probe. The proteins are micro-expressed in unlabeled or isotope-labeled media, purified, and then subjected to 1D ¹H NMR and/or 2D [¹H,¹⁵N]-COSY screening. To demonstrate that the miniaturization is functioning effectively, we processed nine mouse homologue protein targets and compared the results with those from the “macro-scale” Joint Center of Structural Genomics (JCSG) high-throughput pipeline. The results from the two pipelines were comparable, illustrating that the data were not compromised in the miniaturized approach.     

Molecular analysis to FUT-1, 2, 3 and ABO genotyping may instead of serological typing to diagnose para-Bombay in Chinese

The aim of this study was to evaluate the consistency between serotyping and molecular analysis in Chinese with para-Bombay. The molecular analysis of gene fragments in FUT-1, FUT-2, FUT-3 and ABO genotyping and serotyping were used including a saliva test to examine the A, B, H substance and an absorption elution test to examine the A, B, H; and further routine tests including ABO, H and Lewis phenotype. From eleven samples with anti-H negative, 10 samples were confirmed with para-Bombay by sequencing to FUT-1, from which six samples were 547-548delAG, three samples were 880TT deletion, one sample was 35C>T and one sample was 649G>T heterozygous (h7, China) as carrier. The sequencing to FUT-2 confirmed 357C>T in 11 samples, meaning H, A and B substance was secreted in saliva except for one sample which occurred 385A>T (I129F) heterozygous, which is a weak secretor. The FUT-3 sequence result demonstrated four samples with heterozygous mutations to 59T>G (L20R) combined with 508G>A (G170S) and seven samples without mutations in FUT-3 gene fragment same as reference. The consistency between sequencing with FUT-1/FUT-2 and serotyping by anti-H reported an identical result, except for one sample, which interestingly showed the H/h7 carrier with serotyping negative to anti-H. The result of sequencing with FUT-2/FUT-3 and Lewis phenotyping also reported a complete consistency. The saliva test to A, B, H substance and absorption elution test examining the A, B, H antigens on the surface of red blood cells completely matched the ABO exon 6, 7 sequence results. The sequencing of FUT-1, FUT-2, FUT-3 and ABO exon 6, 7 may become a useful tool to confirm the para-Bombay blood type.     

Quantitative analysis of metabolite concentrations in human urine samples using 13C{1H} NMR spectroscopy

Targeted profiling is a library-based method of using mathematically modeled reference spectra for quantification of metabolite concentrations in NMR mixture analysis. Metabolomics studies of biofluids, such as urine, represent a highly complex problem in this area, and for this reason targeted profiling of ¹H NMR spectra can be hampered. A number of the issues relating to ¹H NMR spectroscopy can be overcome using ¹³C{¹H} NMR spectroscopy. In this work, a ¹³C{¹H} NMR database was created using Chenomx NMR Suite, incorporating 120 metabolites. The ¹³C{¹H} NMR database was standardized through the analysis of a series of metabolite solutions containing varying concentrations of 19 distinct metabolites, where the metabolite concentrations were varied across a range of values including biological ranges. Subsequently, the NMR spectra of urine samples were collected using ¹³C{¹H} NMR spectroscopy and profiled using the ¹³C{¹H} NMR library. In total, about 30 metabolites were conclusively identified and quantified in the urine samples using ¹³C{¹H} NMR targeted profiling. The proton decoupling and larger spectral window provided easier identification and more accurate quantification for specific classes of metabolites, such as sugars and amino acids with overlap in the aliphatic region of the ¹H NMR spectrum. We discuss potential application areas in which ¹³C{¹H} NMR targeted profiling may be superior to ¹H NMR targeted profiling.     

The ABO, Lewis and related blood group antigens; a review of structure and biosynthesis

Abstract Numerous studies have shown that the antigenic determinants of the ABO blood group system are closely related in biochemical terms to the antigenic determinants of the Hh, P, Lewis and Ii blood group systems. The blood group antigens of each of these systems are formed by the addition of specific sugars to an oligosaccharide precursor chain which may be bound through sphingosine to fatty acids (glycolipid) or through serine or threonine to a peptide chain (glycoproteins). The direct gene products of each of these blood group systems are the glycosyltransferase enzymes which catalyse the addition of the specific sugar thus conferring the specified blood group activity to the glycolipid or glycoprotein molecule. The antigenic determinants of the ABO and Lewis systems in addition to red cells also exist in the body secretions in soluble form when the relevant genes are expressed in the phenotype. The antigens expressed on both the red cells and in the secretions are determined by the interaction of Hh, Sese, ABO and Lele genes.     

PA-I and PA-II lectin interactions with the ABO(H) and P blood group glycosphingolipid antigens may contribute to the broad spectrum adherence ofPseudomonas aeruginosa to human tissues in secondary infections

Pseudomonas aeruginosa may cause serious infections in most human tissues/organs. Its adherence to them is mediated by a battery of adhesins including the PA-I and PA-II lectins, which are produced in this bacterium in high quantities. PA-I binds to thed-galactose of the erythrocyte glycosphingolipids exhibiting highest affinities for B and P^k (followed by P₁) antigens, while PA-II preferentially binds to thel-fucose of H, A and B antigens. IntactP. aeruginosa cells also exhibit a clear P^k and P₁ over p preference. Such affinities for the most common human ABH and P system antigens may underlie the widespread tissue infectivity and pathogenicity of this bacterium.     

Recognition of the blood group H type 2 trisaccharide epitope by 28 monoclonal antibodies and three lectins

The patterns of cross-reaction of 30 monoclonal antibodies and three lectins were determined by ELISA with 21 ABH, Ii or Lewis related synthetic oligosaccharides coupled to bovine serum albumin. At least seven main groups of cross-reactive patterns were identified among the antibodies, plus several intermediate patterns between two of the main antibody groups. The three lectins had different cross-reaction patterns,Galactia tenuiflora was different from all the antibodies,Ulex europaeus lectin 1 andLotus tetragonolobus were similar, but not identical to groups III and V of antibodies respectively. The anti-H antibodies cross-reacting with A type 2 gave similar agglutination scores with all the normal ABO erythrocytes, while the anti-H antibodies not cross-reacting with A type 2 reacted with different scores: O>A₂>A₂B>B>A₁>A₁B>O_h, suggesting that these antibodies react better with the free H epitopes and do not recognize the H in A or B epitopes. Based on the ELISA and agglutination results and the lowest energy conformations of each oligosaccharide obtained by computer modelling, the most probable oligosaccharide surface areas recognized by each antibody main group are illustrated.