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Blood groups can be divided into those consisting of carbohydrate antigens and those that are protein in nature. Carbohydrate blood group systems include ABO, H, Lewis, P1Pk, and Globoside. Protein blood group systems may be single-pass proteins (e.g., MNS and Kell), multipass proteins (e.g., Rh, Duffy, Kidd, and Diego), and glycosylphosphatidylinositol (GPI)-linked proteins (e.g., Cromer, Cartwright, and Dombrock), integral proteins (e.g., Lutheran), or water channel proteins/aquaporins (e.g., Colton and GIL). At the heart of immunohematology is detecting these antigens on red blood cells (RBCs) and the corresponding antibodies individuals may produce. As a general rule, those who lack carbohydrate antigens may produce “naturally” occurring or nonred cell stimulated IgM antibodies without exposure to foreign RBC antigens. In contrast, those individuals who lack protein antigens may produce IgG alloantibodies when exposed to foreign RBC antigens, typically via transfusion or pregnancy. Of course, the immune system “does not read the books” and there are examples of IgG antibodies to carbohydrate antigens and IgM antibodies to protein antigens, albeit unusual. This chapter provides an overview of blood groups, their unique characteristics, and the problems that arise during testing.

Sphingosine is an 18-carbon amino alcohol with a long unsaturated hydrocarbon chain. Ceramide is a long-chain fatty acid amide derivative of sphingosine or, put simply, it is the combination of sphingosine alcohol with a fatty acid. Derivatives of ceramide are formed by attaching an OH group to C1. Ceramide plays an important role in signal transduction in programmed cell death (apoptosis), the cell cycle, and cell differentiation and senescence. Ceramide is the core molecule of sphingolipids. Ceramide links with other molecules to yield derivatives, such as sulfatides, globosides, glucosylceramide, sphingomyelin, and gangliosides. These compounds are vital for brain and nervous system development.

There are increased levels of stage-specific embryonic antigens-3 and -1 (SSEA-3 and SSEA-1) globo-series glycolipids in male versus female DBA/2 and C57BL/6 kidneys, respectively. To determine what enzymatic steps may be responsible for these differences, the activity and properties of UDP-galactose:globoside galactosyltransferase were studied in male and female mouse kidney microsomes. This enzyme participates in the biosynthesis of galactosylgloboside, SSEA-3 glycolipid; the reaction product was identified by high performance thin-layer chromatography (HPTLC) immunostaining. In C57BL/6 mice, the specific activity of the enzyme, in the presence of CHAPS, was 2-fold greater in the male than that in the female. Optimum pH for the enzyme from both sexes was about 5.6, and Mn2+ was essential for maximal activity. Fifty percent of the male and female enzyme activity was lost after preincubating the microsomes for 1 min at 55 degrees C; thereafter, the enzyme from female microsomes had a slower rate of denaturation. The Km for globoside in presence of sodium cholate for both male and female was 0.035 mM, but it was approximately 2-fold greater for the female in presence of CHAPS. The enzyme in male and female microsomes was differentially activated by CHAPS and cholate. The results suggest the presence of an enzyme modulator in these membranes. In DBA/2 mice, the enzyme activity was about 2-fold greater in males than that in the female. The specific activity of the enzyme in the two strains was of a similar magnitude.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiokeratoma corporis diffusum (Fabry's disease) is an X-linked disorder of glycosphingolipid catabolism. Heterozygous females, although usually asymptomatic, are occasionally as severely afflicted as hemizygous males; recently we identified a heterozygous patient with cardiomyopathy and severe pain in the extremities. In order to elucidate the difference of the clinical features, we analyzed the glycolipid composition of the heart, liver, and kidney obtained from the patient and from a hemizygote. Gas-liquid chromatography revealed that globotriaosylceramide (Gb3) was markedly increased in the heart (32.4 times higher than control) and increased to a lesser extent in the liver and kidney (3.74 and 6.79 times, respectively). The pattern of Gb3 accumulation in the heterozygote, where the highest increases were seen in the heart, was distinct from that in the hemizygote, where elevated levels of Gb3 and Ga2 were found in the kidney. Furthermore, the alpha-galactosidase activity in the heart, liver, and kidney of the heterozygote was 17%, 26%, and 36%, respectively, of normal controls, which correlated well with the accumulation of glycosphingolipid in the heart and with the disease's clinical manifestations. Two other hemizygotic patients, who were identified by low alpha-galactosidase activities, demonstrated the cardiac involvement.

Publisher Summary This chapter discusses the terminology, history, occurrence, expression, and clinical significance of Pk antigen and LKE antigen. The molecular basis associated with Pk antigen, effect of enzymes/chemicals on Pk antigen on intact RBCs, in vitro characteristics of alloanti-Pk, and clinical significance of alloanti-Pk are described in the chapter. The molecular basis associated with LKE antigen, effect of enzymes/chemicals on LKE antigen on intact RBCs, in vitro characteristics of alloanti-LKE, and clinical significance of alloanti-LKE are discussed in the chapter.

We describe a method for isolating milligram quantities of the four neutral glycosphingolipids, glucocerebroside, lactosylceramide, traiosylceramide, and globoside, from human placental tissue. This procedure is carried out on a silicic acid column eluted with a continuous chloroform-methanol gradient (19:1 to 4:1); the four glycosphingolipids elute as separate fractions with no need for further separation. The method is simple, rapid, and yields sufficient material to use as analytical standards for several hundred runs. The lipids have been identified by NMR spectroscopy. Placental tissue is freely available in most centers and is an excellent untapped source for these compounds. Given that lactosylceramide is not commercially available and that triaosylceramide (ceramide trihexoside) cannot be obtained in a reliable state, this technique represents an effective solution to this dilemma.

We have developed a high-performance liquid chromatographic (HPLC) procedure for separation of O-acetyl-N-p-nitrobenzoyl derivatives of six neutral glycosphingolipids: glucosylceramide, lactosylceramide, globotriaosylceramide, lactotriaosylceramide, globotetraosylceramide, and neolactotetraosylceramide. The recoveries of glucosylceramide and globotetraosylceramide for the derivatization procedure and HPLC analysis were approximately 75%, and one nanomole of glycolipid could be detected. The procedure was used for analysis of human erythrocyte neutral glycolipids.

Five glycosphingolipids (GSL), glucosylceramide, lactosylceramide, trihexosylceramide, globoside, and hematoside (GM3) were studied in serum from normal human subjects and patients with dyslipoproteinemia and found to be exclusively associated with the various classes of serum lipoproteins. Based on a unit weight of lipoprotein protein, the total amount of GSL in serum from normal subjects was twice as high in very low density lipoprotein (VLDL) (d < than 1.006 g/ml) and low density lipoprotein (LDL) (d 1.019-1.063 g/ml) as in high density lipoproteins HDL2 (d 1.063-1.125 g/ml) or HDL3 (d 1.125-1.21 g/ml). In abetalipoproteinemia the levels of serum GSL were slightly reduced when compared to normal serum and were all found in the only existing lipoprotein, HDL; this contained 2-3 moles of GSL/ mole of lipoprotein as compared to 0.5 GSL/mole in normal HDL. In hypobetalipoproteinemia and Tangier disease, the serum glycosphingolipids were 10 to 30% reduced in concentration compared to the 75% reduction in other lipids, and were again found to be associated only with the serum lipoproteins. The relative proportions of GSL did not vary substantially in the normo- and hypolipidemic subjects studied. Only in patients with homozygous familial hypercholesterolemia was there a significant (3-4-fold) elevation of all of the five GSL species and this elevation correlated well with that of the circulating cholesterol and LDL. On a molar basis the LDL of these patients contained the same amount of GSL as normal subjects (5 moles GSL/mole protein). It is concluded that: (1) glycosphingolipids are associated only with the major lipoprotein classes in both normal and dyslipoproteinemic serum; (2) the relative proportions of the five glycosphingolipids are not significantly affected by dyslipoproteinemia; (3) only in severe hypolipoproteinemia do the remaining serum lipoproteins carry a complement of glycosphingolipids greater than normal. Although our results establish that glycosphingolipids are intimately associated with serum lipoproteins, the mode of association or the structural and functional significance of such an association remains undetermined.

The fatty acid composition of cerebrosides and sulfatides from frontal lobe gray and white matter was determined for five fresh and four formalinized adult brains and for eight infants. Fatty acid patterns were unaffected by formalinization, but varied considerably from one another in the proportion of saturated to unsaturated fatty acids. The percentages of 24:0 and 24:1 increased with age. Cerebrosides obtained from areas such as the brainstem and cerebellum, where myelination was more advanced, tended to have a larger proportion of long-chain fatty acids than samples extracted from frontal or parietal lobe white matter. Hydroxy fatty acids showed an adult pattern in all instances in which amounts sufficient for accurate quantification could be isolated. Lipid hexose, cerebroside + sulfatide hexose, and methanoleluted hexose were measured in the brains of 12 infants ranging in age from a 4 month fetus to 2 yr. In the most immature, the majority of lipid hexose was in the form of glycolipids more polar than cerebrosides and sulfatides. These have tentatively been identified as hematosides and globosides. With maturation, cerebrosides and sulfatides increased progressively, but the amounts of the more polar glycolipids remained constant in relation to the total lipid content of tissue.

Concentrations of four neutral glycosyl ceramides were determined in plasma and erythrocytes from nine hemizygous patients with Fabry's disease (a hereditary glycolipid lipidosis), from the sister of one of the patients, and from the heterozygous mother of another one. The concentration of a trihexosyl ceramide, galactosylgalactosylglucosyl ceramide, was elevated in plasma from the patients about threefold above the normal mean level, and the amount of this lipid was also increased in plasma from the two female relatives. The concentrations of glucosyl ceramide and lactosyl ceramide in plasma were slightly less than normal in the affected males, while globoside or a similar tetrahexosyl ceramide was slightly higher than normal. All but one of the Fabry patients had significantly less globoside in the red cells, as compared with the normal range, although the red cell concentration of trihexosyl ceramide was normal in all of the patients. Thus the concentration of the lipid that accumulates in the tissues in Fabry's disease is elevated in plasma but not in red cells, whereas in Gaucher's disease the accumulating lipid (glucosyl ceramide) is elevated in both plasma and red cells.

Several amino-oligosaccharides including fragments of gangliosides and Tay-Sachs globoside have been synthesized. The synthesis of hematoside is discussed.

Publisher Summary This chapter discusses Fabry–Anderson disease (FAD), which is an X-linked glycolipid lysosomal storage disorder that is caused by an alpha-galactosidase deficiency. The unusual array of mostly nonspecific symptoms by which FAD may express itself explains why these historical difficulties are still extant for the clinician who needs to distinguish a rare disorder under commonplace manifestations. FAD is a chronic malady with lifelong, mostly nonspecific manifestations. The first symptom of FAD is generally a bout of excruciating burning pain in the extremities. Clinical signs of peripheral neuropathy are not a feature of FAD. Because globoside, the major erythrocyte glycolipid, is an important source of ceramide trihexoside during the degradation of aging cells, one would expect that the reticuloendothelial system is heavily involved, as is the rule in the multisystemic storage disorders.

The carbohydrate portion of mammalian glycosphingolipids varies in the number of glycose residues, their composition, and the sequence, linkages and anomeric configuration between the sugars. This structural diversity makes possible a very large number of substances; this and also evidence for the occurrence of closely related isomers with different immunological specificities suggests an urgent need for additional fractionation procedures and methods of structural analysis. Such techniques as high performance liquid chromatography, and field desorption and chemical ionization mass spectrometry are reviewed and the results of studies using fuco-pentaose I and glucosylceramide as model compounds are presented. The 90.5 MHz, proton decoupled, natural abundance carbon-13 nuclear magnetic resonance spectra have been obtained for glucosylceramide, lactosylceramide, globotriglycosylceramide and globoside. Single lines were observed for most of the sugar, sphingosine and fatty acyl carbon components. A comparison of their resonances with those of simpler model compounds and other glycolipids allowed the determination of the sugar anomeric configuration, the intersugar linkage position and the configuration of the sugar ring structure. These studies also permitted the estimation of fatty acyl composition in terms of degree of unsaturation and chain length from chemical shift assignments and integration values of specific nuclei. Preliminary results of studies involving the uptake and conversion of 1-13C-mannose into virally transformed hamster (NIL-HSV) cell metabolites are also discussed.