Characteristics of medical students completing an honors program in pathology.

The Honors Program in pathology at Jefferson Medical College provides a voluntary enrichment opportunity for students who have demonstrated a superior ability to cope with the pathology curriculum and who rank in the upper fifth of their class. This study was performed to determine whether honor students possess cognitive and psychosocial attributes that distinguish them from their classmates. Students from five academic years (entering classes 1991 to 1995) were divided into 3 groups: (1) those who completed the Honors Program (n = 85), (2) those in the top 20% of the class who were offered the option but chose not to participate in the Honors Program (n = 128), and (3) students who did not qualify for the program (n = 953). Comparisons between these three groups were made on the basis of selected measures of academic achievement retrieved from the Jefferson Longitudinal Study database and psychosocial data obtained from a questionnaire completed during the first-year orientation. Students who completed the Honors Program in pathology had scored higher on the physical science section of the Medical College Admission Test (MCAT) and had obtained higher first-year grade point averages than students in both of the other groups. Subsequently, they attained higher second-year grade point averages and scored higher on Step 1 and Step 2 of the United States Medical Licensing Examination (USMLE), compared with their peers in the other groups. There were no significant differences in psychosocial measures between honor students and the rest of the cohort (group 3). However, students in the top 20% of the class who declined the invitation to participate in the Honors Program (group 2) showed higher scores on the Taylor Manifest Anxiety Scale and the Eysenck Emotional Instability (Neuroticism) Scale than did their classmates. Despite these differences, students who completed the Honors Program (group 1) and eligible students who declined participation (group 2) selected similar pathways of postgraduate residency training: both groups preferred internal medicine to family practice, and both were more likely than the rest of the cohort to begin residency training at a top-ranked academic/research medical center. Voluntary participation in an Honors Program is a self-selection system that identifies students who are most likely to succeed academically at the highest levels. Residency selection committees may wish to pay dose attention to student involvement in similar programs, because this information may provide insights into student personality and general aptitude.

Effects of monoclonal antibody directed to LeY on implantation in the mouse.

The role of carbohydrates in embryo implantation in the mouse was investigated using an embryo transfer model and a blastocyst-uterine epithelial cell co-culture system. The monoclonal antibody (mAb) AH6 directed to LeY oligosaccharide (Fuc alpha1-2 Gal beta 1-4 [Fuc alpha1-3] GlcNAc) and other three mAbs directed to carbohydrates whose structures are closely related to LeY were used to show the effect of carbohydrate specificity on implantation. In the embryo transfer model, donor blastocysts (4 days post-coitus) were pretreated with mAb AH6 (experimental) or other mAbs (control) and transferred into one uterine horn of a recipient. The implantation rate was checked after 5 days. Implantation was significantly inhibited by mAb AH6 pretreatment, and inhibition was not observed in control groups. In the co-culture system, the attachment and outgrowth rate of blastocysts on the surface of uterine epithelial cells was significantly inhibited when monolayer epithelial cells or blastocysts were pretreated with mAb AH6. The most obvious effect of mAb AH6 was obtained during 2-4 h co-incubation. No inhibition was observed in the control groups. It was, therefore, concluded that oligosaccharide LeY recognized by mAb AH6 plays an essential role at the initial stage of implantation. It may act as a mediator molecule for adhesion between the surface of blastocyst and epithelial cell, and its function is carbohydrate-specific.

Relationship of students' perceptions of faculty to scholastic achievement: are popular instructors better educators?

Student evaluation of the faculty is a standard practice in most medical schools. Implied in these evaluations is the motion that popular instructors (ie, those considered outstanding by the students) are better educators, whose teaching translates into higher scores for their students on examinations. We tested this hypothesis by comparing students' evaluations of the faculty with levels of academic achievement in a second-year pathology course. Objective measures of academic achievement included scores on final comprehensive examinations, final course grade, and performance on the United States Medical Licensing Examination (USMLE). During the 4 years studied (1990 to 1995), students belonging to groups with the highest ratings for their instruction performed no better than those with the poorest ratings. There was no correlation between students' perceptions of quality in teaching and their academic achievement. Our results indicate that students' evaluations of the faculty are subjective and do not correlate with objective results used in the assessment of student knowledge. Popular instructors are not necessarily better educators.

The virtues of extended matching and uncued tests as alternatives to multiple choice questions.

The objectives of this study were to compare the reliability and validity of written test formats that are widely used in medical education (multiple choice, uncued, extended matching, and true/false) and evaluate the effects of uncued examinations on long-term retention of medical knowledge. Uncued tests were introduced into a traditional course in general and systemic pathology (six interim tests). In the following year, students were given eight tests written in the four formats, each being used twice. The academic achievement of students in these 2 years was compared with that of students in 2 previous years, in which multiple choice tests were used. Measures of academic achievement included performance on a final comprehensive examination and the United States Medical Licensing Examination (USMLE). Student performance on uncued tests was consistent over time (i.e., there was no learning curve). Mean scores ranged from 77% to 84%, and coefficient alpha reliability estimates on 100-item tests were excellent (0.79 to 0.90). Extended matching tests were also reliable, with a mean coefficient alpha of 0.90. There was no significant relationship between test format and student performance on subsequent comprehensive examinations. Our results indicate that extended matching and uncued tests have considerable advantages over multiple choice and true/false examinations. They are more reliable, better able to discriminate the well-prepared from the marginal student, and well suited for tested core knowledge. Contrary to our expectation, extended matching questions with 20 choices presented to the student were as statistically reliable and valid as uncued queries with several hundred choices.

Glycolipids of human primary testicular germ cell tumours.

The glycolipid content of human non-seminomatous germ cell tumour cell lines correlates with their differentiation lineage. To analyse whether this reflects the situation in primary tumours, we studied five embryonal carcinomas, five yolk sac tumours and nine (mixed) non-seminomas, using thin-layer chromatography and carbohydrate immunostaining. We also analysed the glycolipid content of 19 seminomas to reveal their relationship with non-seminomas. Lactosylceramide (CDH) was detected in all embryonal carcinomas, but in fewer than half of the seminomas. Seminomas and embryonal carcinomas contained globoseries glycolipids, including globotriosylceramide (Gb3), globoside (Gb4), galactosy globoside (Gb5) and sialy1 galactosyl globoside (GL7). The lacto-series glycolipid Le(x) was found in all embryonal carcinomas, but only in one seminoma. Gangliosides GD3 and GT3 were detected in many seminomas, but rarely in embryonal carcinomas. Yolk sac tumours displayed a heterogeneous glycolipid profile. Compared with seminomas and pure embryonal carcinomas, differentiated non-seminomas had reduced levels of globo-series glycolipids, especially Gb3 and Gb5, whereas CDH, Le(x), GD3 and GT3 were found in the majority of cases. Thus, the glycolipid content of non-seminoma cell lines reflects the situation in primary tumours. Globo-series glycolipids are similarly expressed in seminomas and embryonal carcinomas. The expression of Gb3 and Gb5 is reduced in non-seminomas upon differentiation. Le(x) expression in non-seminomas, including embryonal carcinomas, allows discrimination from seminomas. Expression of gangliosides in seminomas might indicate their maturation from ganglioside-negative precursor cells. Reprogramming of these precursors would result in the formation of Le(x)-expressing embryonal carcinomas.

Testing of medical students with open-ended, uncued questions.

A written examination based on material covered in the first year of medical school (anatomy, physiology, biochemistry, and neuroscience) was administered to medical students immediately before they began the pathology course to assess their knowledge of the basic science content that is important for the study of pathology. In alternate years the questions were presented in the standard multiple choice question (MCQ) format or in an open-ended, uncued question (Un-Q) format. The students' mean scores obtained by these two testing methods were comparable. The discrimination indices, which measure the ability of the test questions to distinguish between students of varying ability, were comparable. The Kuder-Richardson Formula 20 (KR-20), an estimate of the precision of the test given, was 0.53 for MCQ and 0.63 for Un-Q. We conclude that the Un-Q format is an acceptable alternative to the MCQ format and it has several advantages over MCQ. Because the answer sheets for Un-Q tests can be scanned optically and graded by computers, we recommend them as an alternative to MCQ tests.

Monoclonal antibody directed to Le(y) oligosaccharide inhibits implantation in the mouse.

We investigated the role of carbohydrates in blastocyst attachment to the uterine epithelium. Le(y) (Fuc alpha 1-->2Gal beta 1-->4[Fuc alpha 1-->3] GlcNAc) was localized by indirect immunofluorescence to the surface of the mouse blastocyst and uterine epithelium. Western blot analysis showed that Le(y) is carried on many uterine glycoproteins in both pregnant and nonpregnant females; however, new species were detected on Day 4 postcoitum (p.c.) coincident with the onset of uterine receptivity. The function of Le(y) in implantation was tested by injecting monoclonal antibody (mAb) directly into the uterine lumen on Days 3-5 p.c. The effects of intrauterine injections on implantation were scored by comparing the number of viable embryos to the number of CL on Day 10 p.c. Injection of purified anti-Le(y) IgM into the uterine lumen on the afternoon of Day 4 significantly inhibited implantation. This effect was dose-dependent and was obtained during a narrow time window, from 87 to 93 h p.c. Inhibition of implantation was not observed in contralateral uterine horns injected with saline, nor was it observed in uterine horns injected with other anti-carbohydrate mAbs. We conclude that binding of anti-Le(y) to the blastocyst or luminal epithelium masks a ligand involved in implantation. Although the mechanism of inhibition is unknown, we show that Le(y) can interact with another oligosaccharide (H) that has been described as a possible uterine ligand for blastocyst attachment. We hypothesize that Le(y) and H form carbohydrate-carbohydrate interactions that promote close apposition of cell surface membranes during an early step in implantation.

Globo-series carbohydrate antigens are expressed in different forms on human and murine teratocarcinoma-derived cells.

The glycolipids of human teratocarcinoma-derived cell line NCCIT were compared with those of 5 murine teratocarcinoma-derived cell lines. Glycolipid antigens were identified by cell surface immunofluorescence and high-performance thin-layer chromatography (HPTLC) immunostaining with a panel of monoclonal anti-carbohydrate antibodies. Human NCCIT embryonal carcinoma (EC) cells contained extended globo-series glycolipids Gb5 (galactosyl globoside) and GL7 (sialyl galactosyl globoside) recognized by antibodies to stage-specific embryonic antigens 3 and 4 (SSEA-3 and -4). SSEA-4 was not detected by immunofluorescence on the surface of any of the 5 murine teratocarcinoma-derived cell lines examined; however, SSEA-3 was detected on the surface of murine cell lines resembling primitive endoderm (JC44, NF-PE) and trophectoderm (E6496D). HPTLC analysis revealed a large amount of globoside (Gb4) in these differentiated cells, which may account for their labeling with anti-SSEA-3 antibody. Globo-series glycolipids were also detected in murine EC cells; however, differences were noted between the 2 cell lines examined. F9 cells contained primarily Gb4 and Forssman glycolipid, whereas NF-1 cells contained only minor amounts of Gb4 and lacked Forssman glycolipid entirely. Our results, coupled with the known distribution of Forssman antigen in the egg cylinder-stage mouse embryo, suggest that F9 and NF-1 murine EC cells are replicas of cells at different stages of development of the embryonic ectoderm. Glycolipids of normal mouse embryos were examined for comparison. Gb4 and Forssman glycolipid were presents in both embryonic and extra-embryonic tissues, whereas Gb5 and GL7 were restricted to visceral yolk sac and placenta. Our results demonstrate that human and murine teratocarcinoma-derived cells both synthesize extended globo-series glycolipids; however, oligosaccharide chain elongation takes different pathways in the 2 species. These differences reflect species-related and cell type-specific patterns of glycosylation.

Embryonal carcinoma cells differentiate into parietal endoderm via an intermediate stage corresponding to primitive endoderm.

F9 embryonal carcinoma (EC) cells were used as a model system to study endoderm formation during mammalian embryogenesis. F9 cells treated with retinoic acid (RA) or RA plus dibutyryl cyclic AMP (cAMP) were examined for the expression of stage-specific embryonic antigen-3 (SSEA-3), a cell surface marker of primitive and visceral endoderm. SSEA-3 was not detected by indirect immunofluorescence on the surface of undifferentiated stem cells; however, a subset of SSEA-3-positive cells appeared with time in culture, amounting to 20% of cells 10 days after plating. When cultured in the presence of RA, the percentage of SSEA-3-positive cells increased to 70% of cells 10 days after plating. In contrast, treatment of cells with RA plus cAMP yielded differentiated cells that were SSEA-3-negative. These SSEA-3-negative cells exhibited ultrastructural features of parietal yolk sac endoderm. In contrast, SSEA-3-positive cells appearing in cultures treated with RA alone exhibited ultrastructural features of primitive endoderm on day 3, switching to ultrastructural features of parietal endoderm on day 10. Cells with hybrid features, resembling both visceral and parietal yolk sac, were also seen. We suggest that differentiation of F9 EC cells into parietal yolk sac-like cells can occur along two distinct pathways: 1) direct under the combined influence of RA and cAMP; and 2) indirect, under the influence of RA alone, in which cells first differentiate into primitive endoderm. Parietal yolk sac-like cells induced through the latter pathway continue to express SSEA-3, a cell surface marker of primitive endoderm that is not normally found on parietal endodermal cells in vivo.

Glycolipids of germ cell tumors: extended globo-series glycolipids are a hallmark of human embryonal carcinoma cells.

Glycolipids of human germ cell tumor lines were analyzed to define the most common immunohistochemical profiles of embryonal carcinoma (EC), differentiated derivatives of EC, yolk sac carcinoma (YC) and choriocarcinoma (CC). Glycolipid composition was examined by high-performance thin-layer chromatography (HPTLC) combined with immunostaining with a panel of anti-carbohydrate monoclonal antibodies (MAbs). All EC cell lines were found to contain high levels of globo-series glycolipids, including globotriosylceramide (Gb3), globoside (Gb4), Gb5 (Gal beta 1-->3Gb4) and GL7 (sialyl Gal beta 1-->3Gb4). Somatic differentiated derivatives (e.g., EC cells treated with retinoic acid) contained decreased levels of globo-series glycolipids and increased levels of lacto- and ganglio-series glycolipids, including GD3, GT3 and GD2. CC cell lines contained relatively large amounts of Gb3 but did not contain extended globo-series glycolipids Gb5 and GL7. CC cell lines also contained a macroglycolipid reactive with the antibody to SSEA-1 (Lex). Glycolipids were not detected in two YC cell lines, while other YC cell lines contained globo-series core glycolipids (Gb3 and Gb4) and gangliosides. We conclude that EC, YC and CC have distinct patterns of membrane glycolipid expression that can be identified by HPTLC and immunostaining. Our results indicate that globo-series glycolipids Gb5 and GL7, which carry stage-specific embryonic antigens 3 and 4 (SSEA-3 and SSEA-4), are a hallmark of human EC cells. Cell lines derived from human germ cell tumors that do not express Gb5 and GL7 deserve to be re-evaluated, since they may represent different stem cells, most likely equivalent to somatic cells and their developmentally committed precursors (e.g., neuroblasts).

Further studies on cell adhesion based on Le(x)-Le(x) interaction, with new approaches: embryoglycan aggregation of F9 teratocarcinoma cells, and adhesion of various tumour cells based on Le(x) expression.

We previously proposed specific interaction of Le(x) (Gal beta 1-->4 [Fuc alpha 1-->3]-GlcNAc beta 1-->3Gal) with Le(x) as a basis of cell adhesion in pre-implantation embryos and in aggregation of F9 teratocarcinoma cells, based on several lines of evidence (Eggens et al., J. Biol Chem (1989) 264:9476-9484). We now present additional evidence for this concept, based on autoaggregation studies of plastic beads coated with glycosphingolipids (GSLs) bearing Le(x) or other epitopes, and affinity chromatography on Le(x)-columns of multivalent lactofucopentaose III (Le(x) oligosaccharide) conjugated with lysyllysine. Comparative adhesion studies of Le(x)-expressing tumour cells vs their Le(x)-non-expressing variants showed that only Le(x)-expressing cells adhere to Le(x)-coated plates and are involved in tumour cell aggregation, in analogy to F9 cell aggregation. The major carrier of Le(x) determinant in F9 cells is not GSL but rather polylactosaminoglycan ('embryoglycan'), and we demonstrated autoaggregation of purified embryoglycan in the presence of Ca2+, and reversible dissociation in the absence of Ca2+ (addition of EDTA). Defucosylated embryoglycan did not show autoaggregation under the same conditions. Thus, Le(x)-Le(x) interaction has been demonstrated on a lactosaminoglycan basis as well as a GSL basis. A molecular model of Le(x)-Le(x) interaction based on minimum energy conformation with involvement of Ca2+ is presented.

Localization of hyaluronan in mouse embryos during implantation, gastrulation and organogenesis.

Hyaluronan was localized in postimplantation mouse embryos using CD44, the principal hyaluronan receptor. The specificity of CD44 receptor-globulin labelling was confirmed using Streptomyces hyaluronidase, anti-chondroitin sulfate antibody, and other receptor globulins. Our major findings are summarized as follows: 1. Implantation of the blastocyst into the uterine wall triggers a rapid loss of hyaluronan from the extracellular matrix of decidual cells on the anti-mesometrial side of the uterus. 2. Hyaluronan appears early in development in the yolk cavity, and the basement membranes of primitive ectoderm and primitive endoderm. 3. During gastrulation, mesodermal cells enter a hyaluronan-rich environment, but lack a pericellular hyaluronan coat themselves. 4. In limb bud embryos, hyaluronan is present throughout the cranial mesenchyme, but is generally not present in the branchial bars, somites, or limb buds. 5. At mid-gestation, hyaluronan is present in the axial skeleton, craniofacial mesenchyme, endocardial cushions of the heart, smooth muscle of the gastrointestinal tract, and connective tissue throughout the body. The pattern of hyaluronan expression in the day 13 fetus is nearly identical to the published distribution of transforming growth factor beta (TGF beta), suggesting a close functional relationship between these molecules. Together, the results suggest that hyaluronan is involved in the formation of early mesoderm, differentiation of craniofacial mesenchyme, and morphogenesis of the axial skeleton.

Hearing and glycoconjugates: localization of Le(y), Le(x) and sialosyl-Le(x) in guinea pig cochlea, particularly at the tectorial membrane and sensory epithelia of the organ of Corti.

Immunohistological examination of guinea pig cochleas was performed using a panel of 25 monoclonal antibodies directed to various lacto-, ganglio- and globo-series carbohydrate epitopes as well as mucin-type epitopes. Lacto-series structures were found to be localized at specific sites of the tectorial membrane (TM) and Corti's organ, i.e. alpha 1-->3 fucosyl type 2 chain (Le(x)) at Kimura's membrane, marginal band and covering net of TM; alpha 1-->2, alpha 1-->3 difucosyl type 2 chain (Le(y)) at covering net; and sialosyl-Le(x) and sialosyl-i at Kimura's membrane and sensory epithelia, particularly sensory tips of hair cells of Corti's organ. In striking contrast, ganglio-series structures (GM3, GD3, GD2, 9-O-Ac-GD3) were detected at spiral ganglion cells, neuronal fibres and stria vascularis, but were completely absent from Corti's organ and most of the TM. Other epitope structures defined by various antibodies were not detectable at any location. The functional roles of lacto-series carbohydrate epitopes expressed at TM and Corti's organ remain unknown. However, the expression of Le(y) (but not other structures) in association with developmental deficiency of TM induced by 6-N-propyl-2-thiouracil in rats suggests that Le(y) plays some role in normal TM development. The presence of Le(x) at Kimura's membrane and sialosyl-Le(x) at hair cell sensory tips of Corti's organ suggests the intriguing possibility that these fucosylated/sialosylated carbohydrate structures play some role in interactions (either attractive or repulsive) of these inner ear components, which have been implicated in the physiology of hearing, i.e. the conversion of sound waves to nerve impulses.

Differentiation antigens of human germ cell tumours: distribution of carbohydrate epitopes on glycolipids and glycoproteins analyzed using PDMP, an inhibitor of glycolipid synthesis.

Human and mouse embryonal carcinoma (EC) cells are characterized by their expression of cell surface carbohydrate antigens, present in both glycolipids and glycoproteins. These antigens disappear upon differentiation and are replaced with other antigens. We have used the inhibitor of glucosyl ceramide synthetase, 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), to study the distribution of carbohydrate epitopes between glycolipids and glycoproteins. PDMP inhibited the expression of glycolipid antigens, but not glycoprotein antigens assayed by immunofluorescence and thin layer chromatography. In the case of SSEA1, we observed expression on both glycolipids and glycoproteins. Resistance to PDMP inhibition suggests that glycoproteins carry the immunodominant form of SSEA1 on the cell surface of differentiated human EC cells and undifferentiated murine EC cells.

Glycosphingolipids of rabbit endometrium and their changes during pregnancy.

The glycolipids of nonpregnant and pregnant rabbit endometrium were characterized using a combination of biochemical and immunochemical techniques. Quantitative analyses indicated a 70% decline in acidic glycolipid (ganglioside) content during early pregnancy (day 6), and a 2.5-fold increase in neutral glycolipid content during later pregnancy (day 26). The major gangliosides of rabbit endometrium were identified by thin-layer chromatography as GM3 and GD3, with minor amounts of GM1, GD1a and GT1b. The major neutral glycolipids were identified similarly as globo-series structures Gb3 and Gb4. Monoclonal antibodies (mAbs) directed to glycolipid antigens permitted the detection of additional glycolipid species, including sialylated, sulfated and fucosylated lacto-series structures. Difucosyl Ley structure (defined by mAb AH-6) and sulfated-galactosyl structure (defined by mAb VESP 6.2) were identified by indirect immunofluorescence along the luminal surface of the endometrium during the implantation period. Rapid changes in the glycolipid composition of endometrial cells during early pregnancy may facilitate embryo adhesion and trophectoderm outgrowth during implantation.

Glycolipid depletion using a ceramide analogue (PDMP) alters growth, adhesion, and membrane lipid organization in human A431 cells.

Glycolipids were depleted from the membranes of human A431 cells using 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), an inhibitor of glucosylceramide synthetase. After 6 days of culture in the presence of 5 microM D-threo-PDMP, glycolipid content was reduced to approximately 5% of control levels. By contrast, synthesis per cell of phosphatidylcholine, sphingomyelin, triglycerides, and glycoprotein was relatively unchanged in PDMP-treated cells. In parallel with glycolipid depletion, PDMP-treated cells exhibited a rapid loss of epithelial cell morphology, a reduced rate of cell growth, and inhibition of cell-substrate adhesion. The effects of D-threo-PDMP on cell morphology and substrate adhesion were blocked by exogenous GM3 addition and were not observed with L-threo-PDMP (a relatively inactive enantiomer). Fluorescence photobleaching and recovery (FPR) was used to investigate the hypothesis that glycolipids influence cell behavior, in part, by changing the diffusion characteristics of membrane proteins and lipids. Diffusion coefficients and mobile fractions of two integral membrane proteins, the EGF receptor and a class I MHC antigen, did not differ significantly between control and PDMP-treated cells. Diffusion coefficients of lipid probes, NBD-PC and fluorescent GM1 ganglioside, were similarly unaffected by glycolipid depletion. However, lipid probes did show a significant increase in mobile fraction (the fraction of lipids that are free to diffuse) in PDMP-treated cells. This increase was blocked by culturing cells in the presence of exogenous GM3 ganglioside. The results suggest that glycolipids play a role in the formation of lipid domains in A431 cell membranes. Glycolipid-mediated changes in membrane lipid organization may influence receptor activation and transmembrane signaling, leading to changes in cell growth, morphology, and adhesion.

A ceramide analogue (PDMP) inhibits glycolipid synthesis in fish embryos.

Glycolipids were depleted from medaka embryos using 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), an inhibitor of glucosylceramide synthetase. Embryos cultured in the presence of 20 microM PDMP exhibited a dramatic decline in glycolipid synthesis and cell surface expression. Metabolic labeling of glucosylceramide declined by 87% on Days 3-6 of development and 72% on Days 7-10 (hatching occurred on Day 10). In parallel, PDMP-treated embryos exhibited a striking loss of several tissue-specific glycolipid antigens, including 9-O-acetyl GD3 from brain and retina, GT3/GQ1C from brain, neural tube, and retina, and sulfated glycolipid from skin and gut. Despite these changes in glycolipid expression, PDMP-treated embryos were fully viable with no evidence of developmental abnormality. PDMP appears to provide a useful tool for identifying glycolipid antigens in embryos and investigating their role in development.

Carbohydrate antigens of embryonal carcinoma cells: changes upon differentiation.

Embryonal carcinoma (EC) cells, the malignant stem cells of teratocarcinomas, resemble early embryonic cells morphologically, developmentally, and with respect to several cell surface characteristics. EC cells often differentiate into a variety of cell types when treated with chemical agents such as retinoic acid, or when placed in a normal embryonic environment. Developmentally regulated surface antigens of EC cells and their differentiated derivatives have been defined using monoclonal antibodies. Many of these "differentiation antigens" have proved to be oligosaccharide chains carried on membrane glycolipids and/or glycoproteins. Our analyses of glycolipid composition in a pluripotent human EC cell line, NTERA-2, have revealed a complex set of glycoslylation changes that occur during cellular differentiation. Like early embryos, undifferentiated NTERA-2 EC cells express predominantly globo-series glycolipids. However, once differentiation is initiated by addition of retinoic acid there is a marked shift of cellular glycolipids from globo-series to lacto- and ganglio-series. Distinct subsets of differentiated cells are characterized by their expression of different patterns of glycolipid antigens. These changes in cell surface phenotype may serve to mark cellular identity and facilitate morphogenetic cell interactions during embryonic development.

A ceramide analogue inhibits T cell proliferative response through inhibition of glycosphingolipid synthesis and enhancement of N,N-dimethylsphingosine synthesis.

The ceramide analogue 1-phenyl-2-(decanoylamino)-3-morpholino-1-propanol (PDMP) (particularly the D-threo isomer, D-PDMP) caused inhibition of cell growth in some types of cells, and this growth-inhibitory effect has been attributed to inhibition of UDP-Glc:Cer beta-Glc transferase, resulting in reduced glycolipid synthesis and increased free ceramide [Inokuch, J., & Radin, N. S. (1987) J. Lipid Res. 28, 565-571; Okada, Y., et al. (1988) FEBS Lett. 235, 25-29]. In view of increasing evidence that the T cell proliferative immune response is modulated by glycosphingolipids (GSLs), the reagent D-PDMP was used to evaluate the role of GSLs in this respect. Con A induced or PHA-induced mitogenesis of C3H/HeJ mouse splenocytes, as well as IL2-dependent CTLL cell growth, were strongly inhibited in a dose-dependent manner when cells were preincubated in the presence of 5-10 microM D-PDMP, but not with its stereoisomer L-PDMP. Closely associated with this growth-inhibitory effect in the presence of D-PDMP, levels of essentially all GSLs, including GM3 and other gangliosides, were greatly reduced, whereas ceramide accumulated. Importantly, metabolically labeled radioactive bands, corresponding to free sphingosine and N-monomethylsphingosine, were found to be present in very small quantities (5-12%) relative to the band corresponding to N,N-dimethylsphingosine (DMS), which showed significant accumulation in D-PDMP-treated lymphocytes. The quantity of IL2 receptors and their affinity to IL2 on T cells did not change, but IL2-dependent tyrosine phosphorylation was greatly stimulated, following D-PDMP treatment.(ABSTRACT TRUNCATED AT 250 WORDS)