2D or not 2D, and other interesting questions about enamel: reply to

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2D or not 2D, and other interesting questions about enamel:reply to Macho et al. (2003)

Christopher Dean*

Department of Anatomy and Developmental Biology, University College London, Gower Street, London WC1E 6BT, UK

Received 18 September 2003; accepted 1 March 2004

Keywords: Enamel; Decussation; Prisms; Life history; Primates

A clear definition of enamel decussation is hard tofind. Strictly speaking, to decussate means to cross,as signified by the Roman numeral X (decus),but rarely do enamel prisms intersect at near 90degrees, except in certain rodent teeth. Dentalhistologists tend to use the term decussation rathercasually to describe enamel prism deviation from astraight track, either undulating, weaving, or swirl-ing (both within and between adjacent layers ofenamel) as they pass from the enamel dentinejunction (EDJ) to the enamel surface. Whilethere is probably broad consensus that enameldecussation confers crack-stopping properties asteeth are loaded at contact, there is debate abouthow tooth size and either fast or slow secretionrates of enamel matrix may influence the degree ofenamel decussation at various positions within atooth. The purpose of this reply to Macho et al.(2003) is to comment on some of the issues raisedthat focus on the 3D decussation of enamel prismsand how this may complicate 2D studies of enamelformation. One aim is to broaden the literaturecited and to demonstrate that not all the technicalanswers to quantifying the developmental timing

and directional nature of enamel decussation aresimple “2D–3D” issues. A second is to demon-strate that two currently competing views (that allenamel prisms either always or, alternatively, neverincrease in diameter from the EDJ to the enamelsurface) are in fact both wrong. Another is toclarify two apparently different ideas about howtooth development may or may not relate toprimate life history. But here in particular, itwould be premature to champion one or other ofthese ideas as right or wrong.

Macho et al. (2003) and Jiang et al. (2003)present new data that describe differences in thenature and degree of prism decussation in thelateral enamel of humans and several nonhumanprimate taxa. They used low power backscatteredSEM images of two adjacent fractured faces ofenamel blocks, etched for 20 s with 5% HCl, tocompute 3D reconstructions of prism movementswithin the blocks. Previously, besides Osborn(1968), Shellis and Poole (1979), Boyde andFortelius (1986), and Rensberger (1992) have eachadopted different approaches to demonstrating thecomplex inter-relationships between translatoryameloblast cell movement, enamel structure, anddietary adaptation. In particular, Radlanski et al.(2001) have also recently studied lateral enamel

* Corresponding authorE-mail address: [email protected] (C. Dean).

Journal of Human Evolution 46 (2004) 633–640

0047-2484/04/$ - see front matter � 2004 Elsevier Ltd. All rights reserved.doi:10.1016/j.jhevol.2004.03.001

C. Dean / Journal of Human Evolution 46 (2004) 633–640634

prism movements in human deciduous and perma-nent anterior teeth. These authors used a combi-nation of SEM and confocal microscopy to trackthe local movements of one individual prismrelative to another and to image the real changesin geometry and prism cross-sectional area.Radlanski et al. (2001), for the first time in thisway, tracked individual prisms from the EDJ tothe enamel surface. All of this recent researchprovides a much clearer idea about the varia-bility and developmental basis of enamel prismdecussation. Macho et al. (2003) have given adetailed account of prism movements in primatelateral enamel. But lateral enamel is not cuspal orocclusal enamel and primates spend a great manyyears chewing away their occlusal enamel beforethey begin to wear their lateral enamel at the levelof the interproximal contact point (the level on thebuccal and lingual aspects of the teeth at whichMacho et al. [2003] made their observations).Macho et al. (2003) do not comment abouteither cuspal or occlusal enamel decussation in anyprimates, even though this is obviously whereprimary adaptations to diet are likely to beexpressed most clearly. Indeed, Jiang et al. (2003:450) have specifically stated that “Even within asingle tooth, the prism arrangement differs fromregion to region, i.e. from cusp tip to cervical marginand between cusps . This not only confoundscross-species comparisons, but also hampers inter-pretations about biomechanical behaviour of thetissue” [emphasis added]. In other words, thereare grounds for being cautious about definingpatterns of enamel decussation that are supposedly

taxonomically distinct on the basis of smallsamples of teeth (n=1, 2, or 3 nonhuman primatesspecies) and from inconsistent regions within theseteeth. Besides this, as emerges from the literature,low power simulations of prism movements asmodelled by Macho et al. (2003), while perhapsappropriate for exploring gross dietary adap-tations in primate enamel, are not be the best wayof deciding what can or cannot be achieved withother forms of microscopy.

Macho et al. (2003) raise a number of issues,many of which relate to studies of cuspal and/orocclusal enamel but not to the lateral enamel theystudied. Cuspal enamel (sometimes described asgnarled enamel directly over the dentine horn) iscomposed of prisms that are said to spiral or swirloutwards to the surface in a vortex (although it isnot clear that all prisms in all cusps move in thisway). Occlusal enamel immediately adjacent tocusps, as well as incisal occlusal enamel in anteriorteeth, both at and between mamelons, is thoughtto be different again to the lateral enamel. Here,using confocal microscopy, prisms can often beseen running for very long distances within theplane of 2D ground sections. While it is absolutelythe case that 2D studies of prism decussation arelimited, a number of important observations havebeen made using conventional ground sections ofenamel and various forms of microscopy. Oneimportant observation is that enamel decussationis always greatest close to the EDJ (where rates ofenamel formation are slowest) and least, if notalmost absent, in outer enamel. Second, it is clearthat not all prisms move in the way described by

Fig. 1. On the left of the figure is a transmitted light montage of buccal lateral enamel in an orangutan first premolar containingS-shaped striae (the same specimen as illustrated in Dean and Shellis, 1998). The EDJ is at the bottom and the enamel surface is atthe top (cuspal is towards the left). The bottom two colour-enhanced images on the right of the figure are higher power confocalimages of the same specimen at the EDJ. The broader strip is one of roughly two hundred 0.2 µm thick slices from a stack ofconsecutive images used to reconstruct the thin strip beneath it, which is an orthogonal reconstruction of prisms in the stack viewed“end on,” in their c axis. The middle two images are of a similar pair of confocal micrographs from the outer third of the montage,again with an orthogonal reconstruction of prisms sectioned in their c axis. The top two images on the right are from the tip of thebuccal cusp of the same tooth close the enamel surface. Three things can be noted: First, it is clear that prisms in this section of lateralenamel run all the way through from the EDJ to the enamel surface within the plane of section. While they turn cervically in the outerenamel (towards the left of the figure), i.e., within the z–y plane of Macho et al. (2003), there is no weaving in and out of this plane,i.e., into and out of the x axis of Macho et al. (2003: 85, Figure 3d). Second, daily cross striations are easily visible along the wholeof the prism lengths in both transmitted and confocal images. Third, prism diameters are identical in the inner and outer lateral enamelbut much greater in the outer cuspal region (see text for average measurements). Three thin white lines, in the cuspal orthogonalreconstruction, run centre to centre across seven hexagonal close-packed prisms to show how measurements of prism diameters weremade (see text for details).

C. Dean / Journal of Human Evolution 46 (2004) 633–640 635

Macho et al. (2003), and while there is no questionthat prisms do move in three dimensions, some runfor very long distances within the plane of groundsections (Fig. 1; see also Radlanski et al., 2001:Figures 5, 6, and 7).

It is interesting to compare the variability of theapparent prism decussation in 2D with the new 3Ddata presented by Macho et al. (2003). Forexample, ten sections of Pan teeth and ten sectionsof modern Homo sapiens teeth (taken from thesample of n=20 used by Dean et al., 2001) con-tained cuspal/occlusal enamel prism tracks, imagedusing polarising light microscopy and measuredin 2D (but avoiding the gnarled enamel), thatexceeded a straight line distance between thedentine horn and the enamel surface by a meanvalue of 2.5% (s.d.=1.7, range=1.0–6.3) in Pan,and 4.7% (s.d.=2.3, range=2.3–9.7) in Homo. Eachof our samples is small, but see the comparablefindings measured in 3D by Macho et al. (2003:86, Figure 4). It follows that Macho et al. (2003)may have stumbled on evidence that answersa paradox that has been debated for sometime(FitzGerald, 1995; Antoine, 2000; Smith et al.,2003). Question: How, when prisms obviouslymove in three dimensions, can a two-dimensionalcount of daily cross striations in modern humansover some 2000 days between birth anddeath match the expected count to within �2%(Antoine, 2000)? Answer: It seems the observedmean deviation of several prism tracks imagedin 2D may closely match the observed meandeviation of several prism tracks imaged in 3D incarefully sectioned, well chosen samples of cuspaland occlusal enamel where the gnarled enamel canbe avoided. Dean et al. (2001) sampled 20 2Docclusal enamel prism tracks in African great apesand 20 in modern humans and assumed that abest fit line of the cumulative enamel formationtrajectories would incorporate the effects of thetrue average 3D deviation of occlusal prisms ineach taxon. Macho et al. (2003) are, therefore,mistaken in their assumption that these methodswere described in Dean (1998), but are prob-ably correct that the values for enamel prismdecussation for Pongo, estimated therein using adifferent approach, are probably too low. It doesseem, however, that good new 3D data are going

to improve our confidence in adopting smallercorrection factors for prism decussation than webelieved were necessary in the past.

There has been continuing debate as to whetherenamel prisms expand in diameter between theEDJ and the enamel surface. One view (Skobe andStern, 1980, expressed here by Macho et al., 2003)is that because the outer enamel surface is greaterin area than the area of the EDJ, geometry dictatesprisms must expand towards the surface. Thus, thecomputer simulation by Macho et al. (2003) oflateral enamel in S-shaped striae where prisms areheld at constant diameter develops cracks in theouter enamel and is implausible. (Interestingly, noone has ever asked if prisms decrease in diametertowards the surface in regions of a tooth where theEDJ is greater in length or area than the overlyingenamel surface, e.g., the occlusal intercuspalregions of some monkey teeth.) The alternativeview (Radlanski et al., 1995) is that prisms ofconstant diameter are able to make up the volumerequired by their increasingly meandering path asthey travel outwards (rather like spaghetti held ina conical container). This view, however, runscounter to the observation that prisms in the outerthird of the enamel generally run straighter than inthe inner enamel.

Using serial confocal images obtained fromlongitudinal ground sections (Fig. 1) it is nowpossible to make 3D reconstructions of groups ofprisms (which can be made at any point betweenthe enamel dentine junction and the enamel sur-face). Even the geometry of prisms cut orthogonalto their c axis can be imaged and average prismdiameters measured to observe how they do or donot change in various locations (Fig. 1). A goodapproximation of average prism diameter, which isto some extent independent of the complex geo-metric differences in prism/interprism proportions,can be made from stacks of confocal images (typi-cally 200 images in a z series, each with a depth offield of 0.2 µm) reconstructed orthogonal to theoriginal plane of section. A measurement madeacross three hexagonal close-packed prisms, wherethe line measured passes from the estimated centreof one prism head, through the centre of thecentrally positioned prism head, and on to thecentre of the prism head directly opposite, is two


prism diameters long (Fig. 1). An average of eachof the three possible different diameters measuredfrom the same group of seven prisms divided bytwo, is equal to the measurement of average prismdiameter.

Striae of Retzius are incremental markings thatrepresent the former position of the developingameloblast cell sheet. Very rarely, in Pongo andHylobates, these striae run occlusally, first nearlyparallel to the EDJ before swinging outwards, thenalmost at right angles to the EDJ, before emergingat the surface enamel close to their originalorientation. These unusual striae have becomeknown as S-shaped striae. At the moment, thereare no clear answers as to how they arise, or as towhether they arise in more than one way and even,perhaps, as to whether they are simply pathologi-cal structures associated with some mild formsof enamel hypoplasia (Dean and Shellis, 1998).Macho et al. (2003) have proposed that the prismorientations they have described are constantwithin a given taxon and that the curved pathsthey describe in Pongo explain how S-shaped striaeform. They do not, however, explain why theorang specimens in their study do not showS-shaped striae (as described by Dean and Shellis[1998: 405, Figure 2]), even though the prismpaths and striae angles are supposedly identical.Something is clearly lacking in this explan-ation. As elegant as the explanation in Machoet al.’s (2003: 88) Figure 5 is, it fails becauseprisms in the lateral enamel of real teeth, withor without S-shaped striae, do not increase indiameter, and do not split apart at the enamelsurface (see also Radlanski et al., 2001, andreferences therein). In fact, when average prismdiameters are estimated from hexagonal close-packed groups of prisms (Fig. 1), there is noevidence that any permanent or deciduous prismdiameters (at least in Pan, Pongo, and Homosapiens) fall outside the range of 4.5–5.5 µm,except for those in the very outer 100–200 µm ofenamel in permanent (but not deciduous) cusptips. In this position, larger average diameters ofbetween 6.0–6.8 µm can be sometimes be measured(Fig. 1).

In Pongo, the linear secretion rate in innerlateral enamel containing S-shaped striae is

w3.6 µm per day (Dean and Shellis, 1998) but thisthen doubles to a maximum of w7.4 µm per day,i.e., as fast as, if not faster than, that measured inany other primate enamel. If we model prisms assimple cylinders of constant w4.5 µm diameter inthis region, then the volume of enamel matrixsecreted per day by each ameloblast (�r2h, where hin this instance is the measurement between twoadjacent daily cross striations) would, like thedaily linear rate, approximately double fromw56 µm3 to w118 µm3. In permanent cuspalenamel, however, where average linear rates ofsecretion vary between w3.0 and w5.5 µm per dayin Pongo, but where prism diameter might welleventually increase (Fig. 1) from w4.5 tow6.8 µm, then the volume of enamel matrixsecreted in a day might rise fourfold, fromw48 µm3 to w200 µm3. One thing we can there-fore say about S-shaped striae is that unusuallyfast daily rates of enamel matrix secretion inprisms associated with them are not then matchedby unusual volumes of matrix secretion becauseprism diameters do not increase.

Implicit in Macho et al. (2003) is the notion thatthe ideas about life history and teeth (and inparticular crown formation times and brainsize) presented in Macho (2001) are universallyaccepted and essentially those adopted by Dean etal. (2001). Macho (2001) and Macho et al. (2003)have implied a causal embryonic (presumablytrophic) relationship between brain growth andenamel formation that in some way relates to thelifespan of ameloblasts and is, therefore, addition-ally bound up with brain/body size allometry.Macho and Wood (1995), Jiang et al. (2003: 450),and Macho et al. (2003) continue to state thatameloblasts die at the end of enamel matrixsecretion. In fact, they do not. They lose theirTomes’ process, switch their polarity, and trans-form into maturational ameloblasts with a precise8 hour functional cycle (Takano, 1994) that maycontinue for years in humans. These maturationalameloblasts raise the enamel mineral content andreduce the organic content over an extendedperiod of time. Thus, true ameloblast lifespan isquite different to their sometimes relatively shortsecretory lifespan even though a proportion ofameloblasts undergoes apoptosis at the late


maturation stage (Matalova et al., 2004). It isnonetheless, quite correct that in order to test theseideas further, a better understanding of amelo-blast secretory function is needed (including thecumulative volume of enamel matrix secreted perunit time in relation to some measure of braingrowth, and the energetic costs of both). Onceagain, 3D reconstructions of enamel prismgeometry are certainly the way to approach thisproblem. All of this, however, forms part of anentirely different idea to that proposed by Deanet al. (2001).

An alternative, and perhaps more straight-forward, point of view is that the secretory lifespanof an ameloblast relates to enamel thickness (Deanet al., 2001). We presumed only that there is a cost

to growing a brain, big or small, and that this costis to some extent time dependent. The dentitionalso grows in a time dependent manner at a ratethat reflects the pace of life history. Teeth, byvirtue of the circadian nature of ameloblast andodontoblast function, happen to contain a recordof the time it takes them to grow. It follows thatthe most likely reason that tooth emergence times,or time to grow the component tissues of a tooth,correlate tightly with brain size (across largesamples of primates) is that this results fromplotting one time dependent variable againstanother.

Fig. 2 illustrates plots for human deciduousenamel formation trajectories alongside humanpermanent enamel formation trajectories. They

Fig. 2. Plots comparing the cuspal enamel trajectories in ten human deciduous teeth and twenty human permanent teeth. Thecumulative rate of enamel formation is greater in the deciduous teeth. The deciduous teeth also have much thinner enamel.


demonstrate how enamel formation rates, likeseveral other components of tooth formation,reflect the time available to grow each dentition.Both dentitions share the same kinds of brains andbodies, but develop within different time scales forobvious reasons.

Daily enamel cross striations are a perfectlygood measure of time and are easily seen, counted,and measured in ground sections of enamel.Knowing what volume of matrix is secreted in aday tells us no more or less about how long enameltakes to form than knowing the daily linear rate ofsecretion, or indeed, just the number of crossstriations present. Body size and enamel decussa-tion can indeed be accounted for by making the

theoretical assumption (in 2D) that, in any largeenough sample, we are able to calculate an averagesecretory trajectory typical of one ameloblast fromeach individual represented. A comparison ofcuspal enamel trajectories in Victoriapithecus andGigantopithecus (Fig. 3) describes their enamelgrowth in a way that is independent of theirobvious differences in body size and tooth size.The plots reveal how the secretory lifespan ofameloblasts relates to cuspal enamel thickness.While we might all agree that this kind of datareveals little, if anything, about brain size or bodysize, it is certainly not biologically “meaningless”(Macho et al., 2003; Jiang et al., 2003: footnote onp. 456).

Fig. 3. Plots comparing the cuspal enamel trajectories in Homo sapiens compared with plots from a section of Victoriapithecus(KNM-MB 19841) and two plots from a section of Gigantopithecus.


Macho et al. (2003) rightly highlight howcritical it is to think of enamel in three dimensionsand how difficult it is, because of its complexanatomy, to make ground sections of enamel thatare able to provide a reliable temporal record.Enamel decussation patterns certainly hold manyclues about dietary adaptation in primates, butthey need not obscure the incremental record ofenamel growth in any significant way. Macho et al.(2003: 89) are right to conclude that, with respectto estimates of enamel formation times, differencesin enamel decussation patterns “may not mattertoo much in broad interspecies comparisons.”

References

Antoine, D., 2000. Evaluating the periodicity of incrementalstructures in dental enamel as a means of studying growthin children from past populations. Ph.D. Dissertation,University of London.

Boyde, A., Fortelius, M., 1986. Development, structure andfunction of rhinoceros enamel. Zool. J. Lin. Soc. 87,181–214.

Dean, M.C., 1998. A comparative study of cross striationspacings in cuspal enamel and four methods of estimatingthe time taken to grow molar cuspal enamel in Pan, Pongoand Homo. J. Hum. Evol. 35, 449–462.

Dean, M.C., Shellis, R.P., 1998. Observations on striamorphology in the lateral enamel of Pongo, Hylobates andProconsul teeth. J. Hum. Evol. 35, 401–410.

Dean, C., Leakey, M.G., Reid, D., Schrenk, F., Schwartz,G.T., Stringer, C., Walker, A., 2001. Growth processes inteeth distinguish modern humans from Homo erectus andearlier hominins. Nature 414, 628–631.

FitzGerald, C.M., 1995. Tooth crown formation and thevariation of enamel microstructural growth markers inmodern humans. Ph.D. Dissertation, University ofCambridge.

Jiang, Y., Spears, I.R., Macho, G.A., 2003. An investigationinto fractured surfaces of enamel of modern human teeth: a

combined SEM and computer visualisation study. Arch.Oral Biol. 48, 449–457.

Macho, G.A., 2001. Primate molar crown formation times andlife history evolution revisited. Am. J. Primatol. 55,189–201.

Macho, G.A., Wood, B.A., 1995. The role of time and timing inhominoid dental evolution. Evol. Anthrop. 4, 17–31.

Macho, G.A., Jiang, J., Spears, I.R., 2003. Enamelmicrostructure—a truly three-dimensional structure. J.Hum. Evol. 45, 81–90.

Matalova, E., Tucker, A.S., Sharpe, P.T., 2004. Death in thelife of a tooth. J. Dent. Res. 83, 11–16.

Osborn, J.W., 1968. Directions and interrelationships of prismsin cuspal and cervical enamel of human teeth. J. Dent. Res.47, 395–402.

Radlanski, R.J., Seidl, W., Steding, G., 1995. Prism arrange-ment in human dental enamel. In: Moggi-Cecchi, J. (Ed.),Aspects of Dental Biology: Palaeontology, Anthropologyand Evolution. International Institute for the Study of Man,Florence, pp. 33–49.

Radlanski, R.J., Renz, H., Willersinn, U., Cordis, C.A.,Duschner, H., 2001. Outline and arrangement of enamelrods in human deciduous and permanent enamel. 3D-reconstructions obtained from CLSM and SEM imagesbased on serial ground sections. European Journal of OralSciences 109, 409–411.

Rensberger, J.M., 1992. Relationship of chewing stress andenamel microstructure in rhinocerotoid cheek teeth. In:Smith, P., Tchernov, E. (Eds.), Structure, Function andEvolution of Teeth. Freund Publishing House Ltd, London,pp. 163–183.

Shellis, R.P., Poole, D.G.F., 1979. The arrangement of prismsin the enamel of the anterior teeth of the aye-aye. ScanningElectron Microscopy 2, 497–505.

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Takano, Y., 1994. Histochemical aspects of calcium regulationby enamel forming cells during matrix formation andmaturation. Acta Anat. Nippon 69, 106–122.


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