Simple and complex carbohydrates (glycans) have long been known to play major metabolic, structural and physical roles in biological systems. Targeted microbial binding to host glycans has also been studied for decades. But such biological roles can only explain some of the remarkable complexity and organismal diversity of glycans in nature. Reviewing the subject about two decades ago, one could find very few clear-cut instances of glycan-recognition-specific biological roles of glycans that were of intrinsic value to the organism expressing them. In striking contrast there is now a profusion of examples, such that this updated review cannot be comprehensive. Instead, a historical overview is presented, broad principles outlined and a few examples cited, representing diverse types of roles, mediated by various glycan classes, in different evolutionary lineages. What remains unchanged is the fact that while all theories regarding biological roles of glycans are supported by compelling evidence, exceptions to each can be found. In retrospect, this is not surprising. Complex and diverse glycans appear to be ubiquitous to all cells in nature, and essential to all life forms. Thus, >3 billion years of evolution consistently generated organisms that use these molecules for many key biological roles, even while sometimes coopting them for minor functions. In this respect, glycans are no different from other major macromolecular building blocks of life (nucleic acids, proteins and lipids), simply more rapidly evolving and complex. It is time for the diverse functional roles of glycans to be fully incorporated into the mainstream of biological sciences.

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Biological Roles of Glycans

Experimental Neurology By Prof Paul Glees Pp xii + 532 (Oxford: Clarendon Press; London: Oxford University Press, 1961) 75s net

The Nervous System

Glomerular podocytes are highly specialized cells with a complex cytoarchitecture. Their most prominent features are interdigitated foot processes with filtration slits in between. These are bridged by the slit diaphragm, which plays a major role in establishing the selective permeability of the glomerular filtration barrier. Injury to podocytes leads to proteinuria, a hallmark of most glomerular diseases. New technical approaches have led to a considerable increase in our understanding of podocyte biology including protein inventory, composition and arrangement of the cytoskeleton, receptor equipment, and signaling pathways involved in the control of ultrafiltration. Moreover, disturbances of podocyte architecture resulting in the retraction of foot processes and proteinuria appear to be a common theme in the progression of acquired glomerular disease. In hereditary nephrotic syndromes identified over the last 2 years, all mutated gene products were localized in podocytes. This review integrates our recent physiological and molecular understanding of the role of podocytes during the maintenance and failure of the glomerular filtration barrier.

Cell Biology of the Glomerular Podocyte

Chemical Diversity in the Sialic Acids and Related α-Keto Acids: An Evolutionary Perspective

The glycome describes the complete repertoire of glycoconjugates composed of carbohydrate chains, or glycans, that are covalently linked to lipid or protein molecules. Glycoconjugates are formed through a process called glycosylation and can differ in their glycan sequences, the connections between them and their length. Glycoconjugate synthesis is a dynamic process that depends on the local milieu of enzymes, sugar precursors and organelle structures as well as the cell types involved and cellular signals. Studies of rare genetic disorders that affect glycosylation first highlighted the biological importance of the glycome, and technological advances have improved our understanding of its heterogeneity and complexity. Researchers can now routinely assess how the secreted and cell-surface glycomes reflect overall cellular status in health and disease. In fact, changes in glycosylation can modulate inflammatory responses, enable viral immune escape, promote cancer cell metastasis or regulate apoptosis; the composition of the glycome also affects kidney function in health and disease. New insights into the structure and function of the glycome can now be applied to therapy development and could improve our ability to fine-tune immunological responses and inflammation, optimize the performance of therapeutic antibodies and boost immune responses to cancer. These examples illustrate the potential of the emerging field of ‘glycomedicine’. Glycosylation refers to the addition of carbohydrate chains to proteins and lipids. In this Review, the authors discuss the broad role of glycans in immunity, cancer, xenotransplantation and glomerular filtration and the potential of ‘glycomedicine’.

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Glycosylation in health and disease.

A ganglioside fraction isolated from pooled intestines from newborn to 4-week-old piglets, which we previously partially characterized and showed to specifically inhibit the binding of porcine rotavirus (OSU strain) to host cells (M. D. Rolsma, H. B. Gelberg, and M. S. Kuhlenschmidt, J. Virol. 68:258-268, 1994), was further purified and found to contain two major monosialogangliosides. Each ganglioside was purified to apparent homogeneity, and their carbohydrate structure was examined by high-pH anion-exchange chromatography coupled with pulsed amperometric detection and fast atom bombardment mass spectroscopy. Both gangliosides possessed a sialyllactose oligosaccharide moiety characteristic of GM3 gangliosides. Compositional analyses indicated that each ganglioside was composed of sialic acid, galactose, glucose, and sphingosine in approximately a 1:1:1:1 molar ratio. Each ganglioside differed, however, in the type of sialic acid residue it contained. An N-glycolylneuraminic acid (NeuGc) moiety was found in the more polar porcine GM3, whereas the less polar GM3 species contained N-acetylneuraminic acid (NeuAc). Both NeuGcGM3 and NeuAcGM3 displayed dose-dependent inhibition of virus binding to host cells. NeuGcGM3 was approximately two to three times more effective than NeuAcGM3 in blocking virus binding. Inhibition of binding occurred with as little as 400 pmol of NeuGcGM3/50 ng of virus (approximately 2 x 10(7) virions) and 2 x 10(6) cells/ml. Fifty percent inhibition of binding was achieved with 0.64 and 1.5 microM NeuGcGM3 and NeuAcGM3, respectively. The free oligosaccharides 3'- and 6'-sialyllactose inhibited binding 50% at millimolar concentrations, which were nearly 1,000 times the concentration of intact gangliosides required for the same degree of inhibition. Direct binding of infectious, triple-layer rotavirus particles, but not noninfectious, double-layered rotavirus particles, to NeuGcGM3 and NeuAcGM3 was demonstrated by using a thin-layer chromatographic overlay assay. NeuGcGM3 and NeuAcGM3 inhibited virus infectivity of MA-104 cells by 50% at concentrations of 3.97 and 9. 84 microM, respectively. NeuGcGM3 (700 nmol/g [dry weight] of intestine) was found to be the predominant enterocyte ganglioside (comprising 75% of the total lipid-bound sialic acid) in neonatal piglets, followed by NeuAcGM3 (200 nmol/g [dry weight] of intestine). NeuGcGM3 and NeuAcGM3 together comprised nearly 100% of the lipid-bound sialic acid in the neonatal intestine, but their quantities rapidly diminished during the first 5 weeks of life. These data support the hypothesis that porcine NeuGcGM3 and NeuAcGM3 are physiologically relevant receptors for porcine rotavirus (OSU strain). Further support for this hypothesis was obtained from virus binding studies using mutant or neuraminidase-treated cell lines. Lec-2 cells, a mutant clone of CHO cells characterized by a 90% reduction in sialyllation of its glycoconjugates, bound less than 5% of the virus compared to control cell binding. In contrast, Lec-1 cells, a mutant CHO clone characterized by a deficiency in glycosylation of N-linked oligosaccharides, still bound rotavirus. Furthermore, exogenous addition of NeuGcGM3 to the Lec-2 mutant cells restored their ability to bind rotavirus in amounts equivalent to that of their parent (CHO) cell line. In the virus-permissive MA-104 cell line, NeuGcGM3 was also able to partially restore rotavirus infectivity in neuraminidase-treated cells. These data suggest that gangliosides play a major role in recognition of host cells by porcine rotavirus (OSU strain).

Structure and Function of a Ganglioside Receptor for Porcine Rotavirus

In vivo enzymatic removal of alpha 2-->6-linked sialic acid from the glomerular filtration barrier results in podocyte charge alteration and glomerular injury.

Infectious diarrheal diseases and protein-energy malnutrition (PEM) are major causes of child morbidity and mortality worldwide. In the present study, PEM was superimposed on rotavirus infection in neonatal pigs to simulate chronic small intestinal stress in malnourished infants with viral gastroenteritis. Two-day-old cesarean-derived pigs (n = 39) were allotted to three treatment groups: 1) noninfected, full-fed; 2) infected, full-fed; and 3) infected, malnourished. Two days postinfection, severe diarrhea and weight loss (11%) were accompanied by reductions in villus height (60%) and lactase activity (78%) and increased crypt depth (32%) in infected full-fed compared with noninfected pigs (P < 0.05). Malnutrition blunted (P < 0.05) increases in crypt depth elicited by rotavirus. By 9 d postinfection, body weight was 59% less, villus height and lactase activity remained lower (50%), and crypt depth remained greater (62%) in infected full-fed compared with noninfected pigs (P < 0.05). However, diarrhea began to clear in infected full-fed, but not in infected malnourished pigs. Plasma insulin-like growth factor-I (IGF-I) was reduced 68% and crypt depth was reduced 19% in infected-malnourished compared with infected full-fed pigs (P < 0.05). Sixteen days postinfection, full-fed pigs had recovered from rotaviral infection; however, in infected-malnourished pigs, diarrhea and growth stasis persisted, and plasma IGF-I, villus height and alkaline phosphatase activity remained reduced compared with infected full-fed pigs (P < 0.05). Overall, PEM prolonged diarrhea and delayed small-intestinal recovery, indicating that nutritional status during diarrhea is essential for recovery from rotaviral enteritis.

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Protein-Energy Malnutrition Delays Small-Intestinal Recovery in Neonatal Pigs Infected with Rotavirus

Summary The effect of oral epidermal growth factor (EGF) on histological and biochemical changes in epithelium in the small intestine was studied in colostrum-deprived neonatal pigs. Forty-eight pigs were infected at 4 days of age with 2 ± 107 plaque-forming units of porcine group A rotavirus and orally fed a simulated sow-milk diet supplemented with 0.0, 0.5, or 1.0 mg/L recombinant human EGF. Sixteen noninfected pigs were fed a diet without EGF supplementation. Infected pigs developed severe diarrhea; they also consumed 25% less food and gained 60% less weight than noninfected pigs. Pigs were killed 8 days postinfection to collect samples at seven equidistant points in the small intestine. Rotavirus infection decreased villus height by 37% and reduced specific activity of lactase by 54%, of leucine aminopeptidase by 43%, and of alkaline phosphatase by 54% in the small intestine, compared with noninfected pigs. Only the supraphysiological dose of EGF (1.0 mg/L) consistently increased villus height in the proximal and mid-small intestine and lactase-specific activity in the mid-small intestine of rotavirus-infected pigs. However, this dose was only partially effective in restoring intestinal mucosal dimensions and enzyme activities. Supplemental EGF did not hasten the resolution of diarrhea. These data indicate that high physiological levels of EGF are beneficial in stimulating recovery of epithelium in the small intestine following rotavirus infection.

Effect of orally administered epidermal growth factor on intestinal recovery of neonatal pigs infected with rotavirus.

Two-hundred-and-eighty-nine veterinary students from all four years of the University of Tennessee, College of Veterinary Medicine (UTCVM) were invited to complete the Derogatis Stress Profile (DSP)1 and an original Demographic Data Profile (DDP). The DSP assessed the students' current experiences of perceived stress, and the DDP was designed to gather information about students' academic year, their living situations, their financial situations, their interest area within the veterinary medical profession, and their current methods of coping with stress. These data were gathered as a baseline measure of veterinary medical students' perceived level of stress and quality of life. In an earlier study, data were also collected from faculty and staff about the perceived quality of the climate and culture of the veterinary college. The results of the DSP and DDP suggest that, although veterinary students at UTCVM do not experience significant levels of stress overall, they do report higher levels of subjective stress, time pressure, and depression than the general population. The more companion animals that veterinary students cared for in their personal lives, the more likely they were to report higher levels of perceived stress. Lastly, there were significant differences between genders, with female veterinary students reporting higher levels of perceived stress than their male counterparts. The preliminary results of the climate and culture data suggest that faculty and staff of the veterinary college individually feel that they are cared for in the work environment and collectively believe that the college strives for excellence.

https://jvme.utpjournals.press/doi/pdf/10.3138/jvme.32.2.173

Howard B. Gelberg

Papers

Structure and Function of a Ganglioside Receptor for Porcine Rotavirus

In vivo enzymatic removal of alpha 2-->6-linked sialic acid from the glomerular filtration barrier results in podocyte charge alteration and glomerular injury.

Protein-Energy Malnutrition Delays Small-Intestinal Recovery in Neonatal Pigs Infected with Rotavirus

Effect of orally administered epidermal growth factor on intestinal recovery of neonatal pigs infected with rotavirus.

Stress management interventions for veterinary students.