Carbohydrates in Cell Recognition - Exam...

n 1952 Aaron Moscona of the Uni-versity of Chicago separated thecells of a chick embryo by incubat-

ing them in an enzyme solution andswirling them gently. The cells did notremain apart; they coalesced into a newaggregate. Moreover, Moscona saw thatwhen retinal cells and liver cells were al-lowed to coalesce in this way, the reti-nal cells always migrated to the innerpart of the cellular mass. Three yearslater Philip L. Townes and JohannesHoltfreter of the University of Roches-ter performed a similar experiment withcells from amphibian embryos, whichre-sorted themselves into tissue layerslike those from which they had come.

Those experiments and countless oth-er observations testify to the keen abili-ty of cells to recognize one another andto respond accordingly. Sperm, for ex-ample, can distinguish eggs of their ownspecies from those of others, and theywill bind only with the former. Somebacteria settle preferentially in the in-testinal or urinary tract; others fancydi›erent organs.

It is not surprising, then, that decod-ing the language of cellular interactionsholds profound interest for researchersin many areas of biology and medicine.Although we still do not understand

the chemical basis of most cell-recogni-tion phenomena, clear explanations forsome have emerged during the past de-cade. Proteins, which mediate most ofthe chemical reactions inside living or-ganisms, appear on the cell surface aswell, and they certainly play a part.

Yet the accumulating evidence alsosuggests that in many cases carbohy-drates (frequently referred to as sug-ars) are the primary markers for cellrecognition. Discoveries about the in-volvement of specific sugars in recogni-tion will have practical applications tothe prevention and treatment of a vari-ety of ailments, including cancer.

B iologists generally accept that cellsrecognize one another throughpairs of complementary struc-

tures on their surfaces: a structure onone cell carries encoded biological in-formation that the structure on the oth-er cell can decipher. That idea repre-sents an extension of the lock-and-keyhypothesis formulated in 1897 by EmilFischer, the noted German chemist. Heused it to explain the specificity of in-teractions between enzymes and theirsubstrates. Pioneering immunologistPaul Ehrlich extended it in 1900 to ac-count for the highly specific reactionsof the immune system, and in 1914Frank Rattray Lillie of the University ofChicago invoked it to describe recogni-tion between sperm and eggs.

By the 1920s the lock-and-key hypoth-esis had become one of the central the-oretical assumptions of cellular biolo-gy. Yet for many years thereafter, thenature and identity of the molecules in-volved in cellular recognition remaineda complete mystery.

To most biologists, the idea that the molecules might be carbohydratesseemed farfetched. That large class ofcompounds consists of monosaccha-rides (simple sugars such as glucose and

fructose) and of oligosaccharides andpolysaccharides, which are composed oflinked monosaccharides. Until the late1960s, carbohydrates were thought toserve only as energy sources (in theforms of monosaccharides and storagemolecules such as the polysaccharidestarch) and as structural materials (thepolysaccharides cellulose in plants andchitin in the exoskeletons of insects).The two other major classes of biologi-cal materials—nucleic acids, which car-ry genetic information, and proteins—were obviously far more versatile. Bycomparison, carbohydrates looked likedull, second-class citizens.

Interest in carbohydrates was furtherdiscouraged by the extraordinary com-plexity of their structures. In contrastto the nucleotides in nucleic acids andthe amino acids in proteins, which caninterconnect in only one way, the mono-saccharide units in oligosaccharides andpolysaccharides can attach to one an-other at multiple points. Two identi-cal monosaccharides can bond to form 11 di›erent disaccharides, whereas twoamino acids can make only one dipep-tide. Even a small number of monosac-charides can create a staggering diver-sity of compounds, including many withbranching structures. Four di›erent nu-cleotides can make only 24 distinct tet-

82 SCIENTIFIC AMERICAN January 1993

NATHAN SHARON and HALINA LIShave been members of the biophysicsdepartment of the Weizmann Institute ofScience in Rehovot, Israel, for more than30 years. During most of that time, theyhave collaborated closely on the studyof complex carbohydrates and lectins,proteins that bind selectively to carbo-hydrates. In addition to their many jointscientific papers, they have written sev-eral widely cited review articles on thosetopics, as well as the recent book Lectins(Chapman & Hall, London). This is Shar-on’s fifth article for Scientific American.

Carbohydratesin Cell RecognitionTelltale surface sugars enable cells to identify

and interact with one another. New drugs aimed at thosecarbohydrates could stop infection and inflammation

by Nathan Sharon and Halina Lis

HORMONE

GLYCOPROTEIN

Copyright 1992 Scientific American, Inc.

ranucleotides, but four di›erent mono-saccharides can make 35,560 uniquetetrasaccharides.

This potential for structural diver-sity is the bane of the carbohy-drate chemist, but it is a boon to

cells: it makes sugar polymers superblye›ective carriers of information. Carbo-hydrates can carry much more informa-tion per unit weight than do either nu-cleic acids or proteins. Monosaccharidescan therefore serve as letters in a vo-cabulary of biological specificity; thecarbohydrate words are spelled out byvariations in the monosaccharides, dif-ferences in the links between them andthe presence or absence of branches.

Scattered reports that carbohydratescould define specificity began to appearquite early in the scientific literature, al-though they often went unnoticed. Bythe 1950s, for example, it was well es-tablished that injected polysaccharidescould stimulate the production of an-tibodies in animals. Researchers alsoknew that the major ABO blood typesare determined by sugars on blood cellsand that the influenza virus binds to a red blood cell through a sugar, sialicacid. Yet not until the 1960s did sugarscome into their own.

Two major developments promptedthat change. The first was the realiza-

tion that all cells carry a sugar coat. Thiscoat consists for the most part of glyco-proteins and glycolipids, two types ofcomplex carbohydrates in which sugarsare linked to proteins and lipids (fats),respectively. Several thousands of glyco-protein and glycolipid structures havebeen identified, and their number growsalmost daily. This diversity is surely sig-nificant: the repertoire of surface struc-tures on a cell changes characteristicallyas it develops, di›erentiates or sickens.The array of carbohydrates on cancercells is strikingly di›erent from that onnormal ones.

An additional stimulus came fromthe study of lectins—a class of proteinsthat can combine with sugars rapidly,selectively and reversibly. Biologists oncethought lectins were found only inplants, but in fact they are ubiquitous innature. Lectins frequently appear on thesurfaces of cells, where they are strate-gically positioned to combine with car-bohydrates on neighboring cells. Theydemonstrate exquisite specificity: lectinsdistinguish not only between di›erentmonosaccharides but also between dif-ferent oligosaccharides.

A landmark discovery about the roleof lectin-carbohydrate interactions incell recognition came from the work ofG. Gilbert Ashwell of the National In-stitutes of Health and Anatol Morell ofthe Albert Einstein College of Medicine.In 1968 they enzymatically removed afew sialic acid molecules from certain

SCIENTIFIC AMERICAN January 1993 83

SURFACE CARBOHYDRATES on a cell serve as points of attachment for other cells,infectious bacteria, viruses, toxins, hormones and many other molecules. In thisway, carbohydrates mediate the migration of cells during embryo development,the process of infection and other phenomena. Compounds consisting of carbohy-drates that are chemically linked to proteins are called glycoproteins; those inwhich the carbohydrates are linked to fats are glycolipids.

TOXIN

VIRUS

BACTERIUM

CELL


blood plasma glycoproteins, then in-jected the glycoproteins into rabbits. Or-dinarily, such molecules would persistin the animals’ circulation for some time,but the sialic acid–deficient moleculesquickly disappeared.

Ashwell and Morell found that the gly-coproteins ended up in the liver. The re-moval of the sialic acids had unmaskedgalactose in the glycoproteins, and theexposed galactoses had attached to alectin on the liver cells. Subsequently,the researchers learned that if they re-moved both the sialic acids and the un-covered galactoses from the glycopro-teins, the rate at which the moleculeswere eliminated from the blood re-turned to normal. From those results,Ashwell and Morell concluded that car-bohydrate side chains on proteins mayserve as markers for identifying whichones should be removed from the circu-lation and eventually degraded.

Like the surface carbohydrates, thesurface lectins go through changes thatcoincide with a cell’s physiological andpathological states. For instance, in 1981Reuben Lotan and Abraham Raz of theWeizmann Institute of Science showedthat tumor cells from mice and humanscarry a surface lectin not found on nor-mal cells. They and other researcherslater proved that this lectin is involvedin the development of metastases.

Astriking recent illustration of therole of surface sugars and themolecules that bind to them

comes from studies of embryo forma-tion by Senitiroh Hakomori of the FredHutchinson Cancer Research Center inSeattle and by Ten Feizi of the ClinicalResearch Center in Harrow, England.Working with mouse embryos, theyhave shown that as a fertilized egg di-vides, the carbohydrate structures on

the resulting embryonic cells change incharacteristic ways. One of the carbo-hydrates is a trisaccharide known bothas stage-specific embryonic antigen 1(SSEA-1) and as Lewisx (Lex ). It appearsat the eight- to 16-cell stage, just as theembryo compacts from a group of loosecells into a smooth ball.

Hakomori’s group has shown that asoluble compound carrying multipleunits of the same trisaccharide inhib-its the compaction process and dis-rupts embryogenesis. Closely related butstructurally di›erent carbohydrates haveno e›ect. Thus, the Lex trisaccharideappears to play a part in compaction.

Adhesive carbohydrates are there-fore essential to embryonic develop-ment. As research continues, their rolein that process will become more de-tailed. Today the two best-understoodphenomena of that type are microbialadhesion to host cells and the adhesion


The Complexity of Carbohydrate Structuresarbohydrates, nucleic acids and proteins all carry bi-ological information in their structures. Yet carbohy-

drates offer the highest capacity for carrying informationbecause they have the greatest potential for structuralvariety. Their component molecules, monosaccharides,can interconnect at several points to form a wide varietyof branched or linear structures; in the example below,

the branching carbohydrate is only one of many possiblestructures that can be made from four identical glucosemolecules. The amino acids in proteins as well as the nu-cleotides in nucleic acids can form only linear assemblies,which restricts their diversity. The peptide (protein frag-ment) shown here is the only one possible made from fourmolecules of the amino acid glycine.

AMINO ACID (GLYCINE) PEPTIDE (TETRAGLYCINE)

MONOSACCHARIDE (GLUCOSE) OLIGOSACCHARIDE (BRANCHED TETRAGLUCOSE)

CARBON OXYGEN NITROGEN HYDROGEN

C


of white blood cells to blood vessels.The more thoroughly characterized ofthese interactions is microbial adhe-sion, which has been studied for nearlytwo decades and serves as a model forother forms of carbohydrate-mediatedcell recognition.

To cause disease, viruses, bacteria or protozoa must be able to stick to atleast one tissue surface in a suscepti-ble host. Infectious agents lacking thatability are swept away from potentialsites of infection by the body’s normalcleansing mechanisms. Microorganismsin the upper respiratory tract, for ex-ample, may be swallowed and eventu-ally destroyed by stomach acid; thosein the urinary tract may be flushed outin the urine.

The first clues about the mechanismof bacterial adhesion sprang from a series of pioneering studies by J. P. Duguid of Ninewells Hospital MedicalSchool in Dundee that began in the1950s. Duguid demonstrated that manystrains of Escherichia coli (a bacterialdenizen of the intestines that can alsocolonize other tissues) and related bac-teria adhered to cells from the epitheliallining of tissues and to erythrocytes, orred blood cells. In the presence of stickybacteria, the erythrocytes would clumptogether—a phenomenon called hemag-glutination. (Researchers still routinelyuse hemagglutination as a simple testfor the adhesion of bacteria to animalcells.) To learn how the bacteria boundto the cells, Duguid exposed them to awide range of compounds. He foundthat only the monosaccharide mannoseand very similar sugars could inhibithemagglutination.

Duguid also made the important ob-servation that the bacterial strains re-sponsible for mannose-sensitive hemag-glutination had submicroscopic, hair-like appendages on their surfaces. Thesestructures were five to 10 nanometersin diameter and several hundreds ofnanometers long. He called them fim-briae, from the Latin word for fringe.Almost simultaneously, Charles C. Brin-ton, Jr., of the University of Pittsburghdescribed the same structures andnamed them pili, from the Latin wordfor hairs. Both terms are still in use.

Later, starting around 1970, RonaldJ. Gibbons of the Forsyth Dental Centerin Boston and his colleagues began re-porting on the selective adhesion ofbacteria to niches within the oral cavi-ty. Gibbons observed that Actinomycesnaeslundii colonizes both the epithelialsurfaces of infants without teeth andthe teeth of children and adults. Con-versely, the related bacterium A. visco-sus does not appear in the mouth untilthe teeth erupt from the gums; it exhib-

its a preference for teeth rather thanoral epithelial surfaces.

Today it is clear that the tissuespecificity of bacterial adhesionis a general phenomenon. For ex-

ample, E. coli, the most common causeof urinary tract infections, is abundantin tissues surrounding the ducts thatconnect the kidneys and the bladder,yet it is seldom found in the upper res-piratory tract. In contrast, group Astreptococci, which colonize only theupper respiratory tract and skin, rarelycause urinary tract infections.

Bacterial adhesion varies not only be-tween tissues but also between speciesand sometimes between individuals ofthe same species, depending on theirage, genetic makeup and health. In theearly 1970s R. Sellwood and Richard A.Gibbons and their colleagues at the In-stitute for Research on Animal Diseasesin Compton, England, studied the infec-tivity of the K88 strain of E. coli. Be-cause those bacteria cause diarrhea inpiglets, they are a costly nuisance forfarmers. Gibbons’s group found thatthe K88 bacteria adhered to the intesti-nal cells of susceptible piglets but notto those of adult pigs or of humans,which the bacteria cannot infect. Bacte-rial mutants that had lost the ability tobind to intestinal cells proved unableto infect the animals.

Moreover, as Gibbons’s work showed,some piglets had a genetic resistanceto K88 bacteria: even potentially viru-lent bacteria could not bind to cellsfrom their intestines. By selecting ge-netically immune piglets for breeding,farmers were able to obtain K88-resis-tant progeny.

The gonorrhea organism, Neisseriagonorrhoeae, serves as another exampleof species and tissue specificity. It ad-heres to human cells of the genital andoral epithelia but not to cells from otherorgans or other animal species. That factexplains why humans are the exclusivehost for N. gonorrhoeae and why otheranimals do not contract gonorrhea.

A strong impetus to the study of bac-terial adhesion was a proposal made in1977 by Itzhak Ofek of Tel Aviv Uni-versity, David Mirelman of our depart-ment at the Weizmann Institute and oneof us (Sharon). We suggested that bac-terial adhesion is mediated by surfacelectins on bacteria that bind to comple-mentary sugars on host cells. That ideahas proved to be generally valid. Workin many laboratories has shown thatbacteria produce lectins specific for cer-tain carbohydrates and that the bacte-ria depend on those lectins for adher-ing to a host’s tissue as the first step inthe process of infection.

Bacterial lectins have already beenthe focus of much study, although far


BACTERIA ADHERE to tissues selectively. Hairlike protrusions called fimbriae onthe bacteria bind exclusively to certain surface carbohydrates. These interactionsdetermine which tissues are susceptible to bacterial invasion. Rod-shaped Esche-richia coli bacteria are shown here on tissue from the urinary tract.


more remains to be done. The best-char-acterized lectins are the type 1 fimbriaeof E. coli , which bind preferentially tosurface glycoproteins containing man-nose. Other research on E. coli during the past decade, primarily by CatharinaSvanborg-Edén and her colleagues whileworking at the University of Göteborg,has described in detail the P fimbriae.Those fimbriae interact specifically withthe P blood-group substance, an ex-tremely common glycolipid containingthe disaccharide galabiose. Researchgroups led by Karl-Anders Karlsson ofthe University of Göteborg and VictorGinsburg of the NIH have mapped thespecificities of lectins from a wide rangeof other bacterial species and strains.

These studies have shown that bacte-ria do not bind solely to the ends ofsurface carbohydrates—they can alsosometimes bind to sugars located with-in the structure. Furthermore, di›erentbacteria may bind to di›erent parts ofthe same carbohydrate. Occasionally,only one face of an oligosaccharide maybe exposed on a particular cell, and asa result the cell will bind bacteria of onekind and not other ones. The ability ofcell-surface sugars to serve as attach-ment sites therefore depends not onlyon the presence of these sugars but alsoon their accessibility and their mode ofpresentation.

Considerable experimental evi-dence now greatly strengthensthe conclusion that the binding

of bacteria to host cell-surface sugarsinitiates infection. For example, uroep-ithelial cells from those rare individu-als who lack the P blood-group sub-stance do not bind to P-fimbriated E.coli. Such individuals are much less sus-ceptible to infections from those bacte-ria than the rest of the population is.

Experiments have shown, however, thatthe bacteria will bind if the epithelialcells are first coated with a synthetic gly-colipid containing galabiose.

Similarly, intestinal cells from pigletsthat are resistant to the diarrhea-caus-ing K88 E. coli lack the large carbohy-drate to which the bacteria bind. Al-though the exact structure of this car-bohydrate has not yet been elucidated,it is known to be present in susceptiblepiglets but absent in adult pigs. This ex-plains why the bacteria were unable toattach to and colonize the intestines ofthe adult pigs, while they caused infec-tion in the young piglets.

Another interesting case is that of theK99 strain of E. coli. Like the K88 strain,K99 causes diarrhea in farm animalsbut not in humans. It is less specific thanK88, however, because it infects youngcalves and lambs as well as piglets. TheK99 bacteria bind specifically to an un-usual glycolipid that contains N-glycol-oylneuraminic acid (a special type ofsialic acid) linked to lactosylceramide.This glycolipid, which is present in pig-lets, calves and lambs, is absent fromthe cells of adult pigs and humans,which instead contain N-acetylneura-minic acid, a nonbinding analogue ofsialic acid. Here a small di›erence be-tween two highly similar sugars—the re-placement of an acetyl group by a gly-coloyl group—is readily detected by thebacteria and explains the host range ofinfection by the organism.

Further confirmation of the above con-clusions was recently obtained in experi-ments on two fimbrial lectins from E.coli that infect the urinary tracts of ei-ther humans or dogs. Both lectins rec-

ognize galabiose, yet one binds only tothe human uroepithelial cells and theother only to canine cells; the galabiose-bearing glycolipids on the surface ofthe cells are presented in subtly differ-ent ways. Those lectin-binding patternsaccord with the host specificities of theE. coli strains.

Because bacterial adhesion is socritical to infection, medical re-searchers are seriously consider-

ing the use of sugars for prevention andtreatment. Sugars that selectively inhib-ited adhesion could act as molecular de-coys, intercepting pathogenic bacteriabefore they reached their tissue targets.Urinary tract infections have been thefocus of particular attention becausethey are second only to respiratory in-fections in frequency.

In collaboration with Ofek, MosheAronson of Tel Aviv University and Mi-relman, we performed the first studyalong those lines in 1979. We injected a mannose-specific strain of E. coli intothe urinary bladder of mice. In some animals, we also injected methyl alpha-mannoside, a sugar that in the test tubeinhibited bacterial adhesion to epithelialcells. The presence of the sugar reducedthe colonization of the urinary tract bybacteria.

Svanborg-Edén has performed analo-gous experiments with P-fimbriated E.coli that infect the kidneys of mice. Sheincubated the bacteria in solutions ofglobotetraose, a sugar found in the gly-colipid of kidney cells. When she sub-sequently injected those bacteria into


WHITE BLOOD CELLS of the immune system, such as lymphocytes, defend the bodyagainst infection by leaving the general circulation and migrating into the tissues.The first step involves selective adhesion between the white blood cells and thewalls of blood vessels called high endothelial venules (photograph). Adhesion de-pends on surface molecules called selectins, which bind to carbohydrates on other

LYMPHOCYTE

HOMINGRECEPTOR(L-SELECTIN)

LYMPHOCYTE MIGRATINGOUT OF VENULE

CARBOHYDRATES

LYMPHOCYTEWITH DIFFERENT

HOMING RECEPTORS

ENDOTHELIAL

CELL

PEYER’S PATCH

PERIPHERAL LYMPH NODE


mice, they persisted in the kidneys forless time than untreated bacteria did.James A. Roberts of Tulane Universityobtained similar results in experimentson monkeys: incubation of P-fimbriatedE. coli with a galabioselike sugar signif-icantly delayed the onset of urinary tractinfections.

Glycopeptides can also interfere withthe binding of bacteria to host tis-sues. In 1990 Michelle Mouricout of theUniversity of Limoges in France andher co-workers showed that injectionsof glycopeptides taken from the bloodplasma of cows can protect newborncalves from lethal doses of E. coli. Theglycopeptides, which contain sugars forwhich the bacteria have affinities, de-crease the adhesion of the bacteria tothe intestines of treated animals.

Indeed, to interfere with bacterial ad-hesion, one need not even use a carbo-hydrate—any agent that competitivelybinds to either the bacterial lectin or thehost cell’s surface carbohydrate will do.For example, Edwin H. Beachey and hiscolleagues at the Veterans Administra-tion Medical Center and the Universityof Tennessee at Memphis have used an-tibodies against mannose to preventcertain mannose-specific E. coli frominfecting mice. The antibodies bind tomannose on the cells, thereby blockingthe sites of bacterial attachment.

Those successful experiments makea clear case for antiadhesive therapies

against microbial diseases. The applica-tion of this approach in humans is nowthe subject of intense research. Furtherstudies of the sugars on host cells andof bacterial lectins should lead to thedesign of better adhesion inhibitors.One point about the approach is certain:because di›erent infectious agents—even di›erent bacteria within the samestrain—can have a wide variety of carbo-hydrate specificities, a cocktail of inhib-itors will undoubtedly be necessary toprevent or treat the diseases.

Carbohydrate-directed interactionsbetween cells are not restrictedto pathological phenomena; they

are also crucially important to thehealthy operation of the immune sys-tem. The immune system has manyparts, but its most important soldiersare the cells called leukocytes. Thisgroup includes an array of diverse whiteblood cells—lymphocytes, monocytesand neutrophils—that act jointly toeliminate bacteria and other intrudersand to mediate the inflammation re-sponse in injured tissues. All these cellscirculate in the blood, but they accom-plish their major functions in the ex-travascular spaces.

The picture emerging from researchis that the inner lining of blood vessels,called the endothelium, actively snareswhite blood cells and guides them towhere they are needed. This process re-

quires an exquisitely regulated recogni-tion between the circulating leukocytesand the endothelial cells.

Such recognition seems to be mediat-ed by a family of structurally relatedlectins. Because this field of research isso new and because di›erent laborato-ries often identify the same adhesionmolecules simultaneously, the nomen-clature is still in a somewhat chaoticstate. Most researchers refer to thesemolecules as selectins because they me-diate the selective contact between cells.Another name in vogue is LEC-CAMs, anacronym for leukocyte-cell (or lectin)adhesion molecules.

The selectins are highly asymmetriccomposite proteins, with an unusualmosaic architecture. They consist ofthree types of functional domains: onedomain anchors the selectin in the cellmembrane, and a second makes upmost of the body of the molecule. Thethird domain, located at the extracellu-lar tip of the molecule, structurally re-sembles animal lectins that work onlyin the presence of calcium ions. Thebinding of carbohydrate ligands to thatdomain is central to the function of se-lectins in interactions between cells.

About 10 years ago Eugene C. Butch-er and Irving L. Weissman of Stan-ford University laid the foundation forour current understanding of how se-lectins (which were then unknown) di-rect lymphocyte tra¤c. Lymphocytesare unique among leukocytes in thatthey continuously patrol the body insearch of foreign antigens (immunolog-ically significant molecules) from bac-teria, viruses and the like. For that pur-pose, lymphocytes leave the blood ves-sels and migrate through the lymphnodes, the tonsils, the adenoids, thePeyer’s patches in the intestines or oth-er secondary lymphoid organs. Di›er-ent lymphocytes migrate selectively, orhome, toward particular organs. To exitfrom the bloodstream, lymphocytesmust first bind to specialized submi-croscopic blood vessels less than 30microns in diameter, known as high en-dothelial venules.

Using an assay technique developedby Hugh B. Stamper, Jr., and Judith J.Woodru› of the State University of NewYork in Brooklyn, Butcher and Weiss-man observed that the homing specific-ity of mouse lymphocytes is dictatedby their selective interaction with thehigh endothelial venules in their tar-geted organs. Butcher and Weissmanthen developed a monoclonal antibody,MEL-14, that bound only to mouse lym-phocytes that went to the peripherallymph nodes. On slices of tissue, theantibody blocked the attachment of thelymphocytes to high endothelial venules


cells. The L-selectins, or homing receptors, on lymphocytes determine the endothe-lial cells to which a lymphocyte will stick : for example, some adhere only in periph-eral lymph nodes or to the Peyer’s patches in the intestines. After a lymphocyte hasattached to the endothelium, it can migrate out of the blood vessel.


from those tissues but not from oth-er lymph organs. When injected intomice, MEL-14 inhibited the migration oflymphocytes into the peripheral lymphnodes.

Butcher and Weissman went on toshow that their antibody bound on thelymphocyte membrane to a single gly-coprotein, now known as L-selectin. Be-cause that glycoprotein is responsiblefor the specific binding of the lympho-cytes to the high endothelial venules, itis also known as the homing receptor.

If high endothelial venules from thelymph nodes are exposed to solutionsof L-selectin, lymphocytes cannot bindto them: the L-selectin molecules occu-py all the potential attachment sites onthe endothelial cells. Conversely, as Ste-ven D. Rosen of the University of Cali-

fornia at San Francisco has shown, cer-tain small sugars and larger polysac-charides can also block interactions be-tween lymphocytes and endothelialvenules. In those cases, the sugars arebinding to the L-selectin.

In 1989 separate experiments byWeissman and by Laurence A. Lasky ofGenentech in South San Francisco incollaboration with Rosen proved con-clusively that the homing receptor me-diates the adhesion of lymphocytes toendothelial cells. The structure of theendothelial carbohydrate to which itbinds is still unknown.

In contrast to the homing receptor,the two other known selectins are foundmainly on endothelial cells, and thenonly when they are actively attractingleukocytes. One of these, E-selectin

(ELAM-1), was discovered in 1987 byMichael P. Bevilacqua of Harvard Medi-cal School. The third member of thegroup, P-selectin (previously known asGMP-140 and PADGEM), was indepen-dently discovered a couple of years lat-er by Rodger P. McEver of the Oklaho-ma Medical Research Foundation andby Bruce and Barbara Furie of the TuftsUniversity School of Medicine.

Research has clarified how tissuesuse selectins to steer white blood cellswhere they are needed. When a tissueis infected, it defensively secretes pro-teins called cytokines, such as interleu-kin-1 and tumor necrosis factor. Thecytokines stimulate endothelial cells inthe venules to express P- and E-selectinson their surfaces. Passing white bloodcells adhere to these protruding mole-

BACTERIUM

LECTIN

SURFACECARBOHYDRATES

CARBOHYDRATEDRUG

a b c

LECTINDRUG

CELL


BLOCKING BACTERIAL ATTACHMENT is one strategy for com-bating infections. As a prelude to infection, bacterial surfaceproteins called lectins attach to surface carbohydrates onsusceptible host cells (a). Drugs containing similar carbohy-

drates could prevent the attachment by binding to the lectins(b). Alternatively, drugs consisting of lectinlike moleculescould have the same e›ect by innocuously occupying thebinding sites on the carbohydrates (c).

SELECTIVE EFFECTS of carbohydrates on bacteria are il-lustrated in these photographs. These E. coli have a lectin for the P glycolipid. Bacteria incubated in the sugar mannose

can still cling to epithelial tissue (left ). A constituent of the P glycolipid binds to the bacteria’s lectin and prevents ad-hesion (right ).


cules because their carbohydrate coatcontains complementary structures.Once attached to the wall of a venule, aleukocyte can leave the bloodstream bysqueezing between adjacent endothe-lial cells.

These two selectins appear on endo-thelial cells at di›erent times and re-cruit di›erent types of white blood cells.Endothelial cells have an internal stock-pile of P-selectin that they can mobilizeto their surface within minutes after aninfection begins. P-selectin can there-fore draw leukocytes that act duringthe earliest phases of the immunologicdefense. In contrast, endothelial cellssynthesize E-selectin only when it is required, so it takes longer to appear.That selectin seems to be most impor-tant about four hours after the start ofan infection, after which it graduallyfades away.

The mechanism that helps leuko-cytes to breech the endothelial barrieris indispensable to the fulfillment oftheir infection-fighting duties. Yet act-ing inappropriately, that same mecha-nism also allows leukocytes to accumu-late in tissues where they do not be-long, thereby causing tissue damage,swelling and pain.

The inflammation of rheumatoid ar-thritis, for instance, occurs when whiteblood cells enter the joints and releaseprotein-chopping enzymes, oxygen rad-icals and other toxic factors. Anotherexample is reperfusion injury, a disor-der that occurs after the flow of bloodis temporarily cut o› from a tissue,such as during a heart attack. When theblood flow resumes, the white bloodcells destroy tissues damaged by lackof oxygen.

The development of pharmaceuticalreagents that would inhibit adverse in-flammatory reactions holds major in-terest for the academic, clinical and in-dustrial sectors. In theory, any drug thatinterferes with the adhesion of whiteblood cells to the endothelium, and con-sequently with their exit from the bloodvessel, should be anti-inflammatory. Thekey to developing such drugs is theshape of the binding regions of the se-lectin molecules and the shape of thecarbohydrates that fit into them. Workon determining those shapes is pro-ceeding at a ferocious pace. In parallelresearch, intensive attempts are beingmade to synthesize carbohydrate in-hibitors of the P- and E-selectins.

For an antiadhesive therapy to be suc-cessful, the drugs must simultaneous-ly accomplish two seemingly incompat-ible ends. On the one hand, they muststop white blood cells from leaving thebloodstream inappropriately; on theother hand, they must still allow the

cells to go where they are needed. Thosegoals may be achievable because thespecificities of adhesion molecules varyin di›erent tissues. One can envision,for example, a drug that keeps whitecells from entering the joints but notother parts of the body.

Aside from their involvement in in-flammation, cell-adhesion molecules may play a role in other

diseases, such as the spread of cancercells from the main tumor throughoutthe body. For example, the carbohydraterecognized by E-selectin is expressedon cells from diverse tumors, includingsome cancers. Bevilacqua has recentlyreported that at least one type of hu-man cancer cell binds specifically to E-selectin expressed on activated endo-thelium. Perhaps to promote their ownmetastasis, some malignant cells re-cruit the adhesion molecules that arepart of the body’s defenses.

If so, antiadhesive drugs may also turnout to be antimetastatic. One hopefulsign in that direction recently came fromHakomori’s group, which was studyinghighly metastatic mouse melanoma cellsthat carry a lectin for lactose, the sugarin milk. The researchers found that byexposing the melanoma cells to com-pounds containing lactose before in-jecting them into mice, they could re-duce the metastatic spread of the cellsalmost by half.

Although the importance of carbohy-drates in cell recognition is immense,other modes of recognition that rely ona peptide language do exist. Some forms

of attachment, for instance, involve sur-face proteins called integrins and com-plementary peptides. The existence ofmore than one system for binding activ-ities lends greater flexibility to a cell’srepertoire of interactions.

Biomedical researchers are still striv-ing for a better understanding of thesugar structures on cell surfaces and ofthe specificities that lectins have forthose structures. As they learn more,they will be in a better position to de-sign highly selective, extremely power-ful inhibitors of cell interactions. Theday may not be far o› when antiadhe-sive drugs, possibly in the form of pillsthat are both sugar-coated and sugar-loaded, will be used to prevent and treatinfections, inflammations, the conse-quences of heart attacks and perhapseven cancer.

CANCER CELLS have unusual carbohydrates on their surface, which may accountfor many of their invasive properties. Drugs that interfere with the adhesiveness ofabnormal cells may someday be used in cancer therapies.


FURTHER READINGLECTINS AS CELL RECOGNITION MOLE-CULES. N. Sharon and H. Lis in Science,Vol. 246, pages 227–234; October 13,1989.

GLYCOBIOLOGY: A GROWING FIELD FORDRUG DESIGN. Karl-Anders Karlsson inTrends in Pharmacological Sciences,Vol. 12, No. 7, pages 265–272; July1991.

CARBOHYDRATES AND GLYCOCONJU-GATES: UPWARDLY MOBILE SUGARS GAINSTATUS AS INFORMATION-BEARING MOL-ECULES. K. Drickamer and J. Carver in Current Opinion in Structural Biolo-gy, Vol. 2, No. 5, pages 653–654; Octo-ber 1992.


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