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THE RELATIONS OF ENDOGENOUS AND EXOGENOUS
FACTORS IN BONE AND TOOTH DEVELOPMENT

THE TEETH OF THE GREY-LETHAL MOUSE

By HANS GRUNEBERG
Department of Zoology, University College, London

INTRODUCTION

‘Tue grey-lethal mutation in the mouse (Griineberg, 1935, 1936 a, b) is characterized by a complex of symptoms which is unparalleled in human pathology.
The fur lacks the yellow pigment. No tooth ever erupts. Without artificial
feeding death invariably occurs between the 22nd and the 30th day of life.
Liquid food prolongs the life for a fortnight or so, but does not materially
change the condition. Death is thus not due to mechanical troubles in feeding,
but its actual cause remains unknown.

The main feature of the skeleton is the entire lack of all secondary absorption processes. Thus bone substance once laid down in development is
preserved in its entirety. The geometrical proportions of bones can only be.
maintained during growth by secondary remodelling processes by which some
of the bone temporarily laid down is eventually removed. This has been
thoroughly worked out in detail by means of madder feeding in the pig by
Brash (1984). If, however, these secondary modelling processes fail to act,
very characteristic anomalies in bone shape are bound to develop. The greylethal bones correspond excellently to these expectations. There is, therefore,
no doubt that the grey-lethal complex of symptoms is in fact largely due to
a failure of the secondary resorption processes on the bones. In addition there
is incomplete calcification, both in bone and dentine. This leads to failure of
eruption of the teeth as has been described in detail elsewhere.

In this paper a more thorough description of the crown shape of the greylethal molars will be given. The anomalies to be described are constantly
found. Generally, the grey-lethals are remarkably constant in the abnormal
conditions they show: the differences between individuals other than those
due to age are practically negligible. It will be discussed below whether and
to what extent the anomalies in the shape of teeth can be explained in terms
of the peculiarities already well established in the grey-lethal mouse. Finally,
the bearing of these facts on the general conception of the action of exogenous
stimuli and the response of: the bones will be considered.
Factors in Bone and Tooth Development 237

MORPHOLOGY OF MOLARS

For comparison, developmental stages of normal teeth must be used in
which the shape of the cusps has not yet been altered by wearing off. The
illustrations (Text-figs. 1-4) show camera lucida drawings of a 14 days old
grey-lethal mouse and a normal litter-mate. The roots of first molars in the
normal are still incomplete, while the uncalcified vestiges of roots in the greylethal, dried up and shrivelled and largely lost during the preparation, are
here of no concern.

As already mentioned in the first paper (1985), the lower first molar is
strongly compressed laterally in the grey-lethal. At the same time it is
noticeably longer than the corresponding tooth of the normal. The median
and lateral surfaces are considerably flattened, while those of the normal are
bulging and bulbous. This applies particularly to the fissures between the

 

Text-fig. 1. Left lower first molar of grey-lethal (c, d) and normal litter-mate (a, b), 14 days old.
a and c, median view; 6 and d, lateral view.

cusps, which are shallow grooves or impressions only, while they are deep
and wide in the normal. The cusps themselves are a little more erect and
project slightly more over the tooth neck. The flattened surfaces are not
smooth and regular, but show minute and irregular depressions, the majority
of which are directed parallel to the axis of the mandible. All these anomalies
are more strongly developed on the lateral surface which as a whole is considerably lower. In lateral view a good deal of the inside cavity of the tooth
is therefore visible (Text-figs. 1-2).

In the upper first molars, the most conspicuous anomaly is that all the
cusps are erected, though to a varying extent. In a normal first upper molar,
all the cusps have their axes practically parallel to each other, including angles
of between 40 and 45° with the plane of the neck of the tooth. In the greylethal tooth the axes of the cusps have an inclination of between 55 and 75°.
This difference is most clearly seen in lateral view, since-the lateral surface is
238 Hans Griineberg

comparatively flat while the median surface in both types bulges towards the
sagittal plane (Text-fig. 8). Comparison of such teeth strongly suggests that

AD

 

b
Text-fig. 2. Top view of same teeth as in Text-fig. 1. a, normal; b, grey-lethal. The arrows indicate
the approximate planes of the sections on Pl. I carrying corresponding numbers.

 

Text-fig. 3. Left upper first molar of grey-lethal (c, d) and normal litter-mate (a, 6), 14 days old.
a and c, lateral view; b and d, median view.

 

Text-fig. 4. Top view of same teeth as in Text-fig. 3. a, normal; 6, grey-lethal. The arrows
indicate the approximate planes of the sections on Pl. I carrying corresponding numbers.

pressure from the front has acted on the grey-lethal tooth so as strongly to
erect the cusps.

Another striking difference is found in the outline of the crown when seen
in top view (Text-fig. 4). Whereas the normal tooth bulges broadly towards
Factors in Bone and Tooth Development 239

the middle line of the body, this protrusion is very much less pronounced in
the grey-lethal. Still more pronounced is the lack of curvature in the region
of the lateral front cusp which has a very irregular shape, is wrinkled and
merges irregularly into its surroundings. The cingulum is completely suppressed. Evidently this is the most abnormal region. Again, as in the lower
first molars of the grey-lethal, the surface is not smooth and regular, but shows
small depressions, chiefly on the lateral surface and the front parts. The
irregular surface of the lateral front cusp has already been mentioned.

All these anomalies of shape suggest that the grey-lethal teeth have grown
under pressure. What has happened seems to be this. At the time when the
then uncalcified tooth germ gets enclosed into the socket, it has not yet reached
its definite size. Now, when it grows, in a normal animal the surrounding
bone responds by absorption to fit the requirements of the growing germ.
Under normal conditions, therefore, shape and size of the socket is always in
harmony with the dimensions of the tooth. In other words, the growth tendencies of the tooth determine size and shape of the socket.

This situation seems to be reversed in the grey-lethal. Here the bone of
the socket does not yield to the growth pressure of the tooth germ. Accordingly
the tooth germ has to adapt itself to the space available. It does not respond
by stopping growth altogether. The growing germ, chiefly the enamel organ,
is therefore bound to get deformed. In the lower first molars, the socket does
not allow for a transverse expansion, but there is no obstacle to a longitudinal
growth. The tooth, therefore, gets compressed in bucco-lingual direction, but
compensates by longitudinal elongation. In the upper first molar, too, there
are obstacles to a transverse expansion. In the present case, however, the
tooth cannot compensate for this by longitudinal evasion. Under such circumstances, the only possible answer can be erection of the cusps. When calcification starts, the cusps are petrified in these positions.

The upper second molars of the grey-lethal show no anomalies of crown
shape whatever. The lower second molars and the wisdom teeth are very
nearly normal, the differences being almost negligible. In particular, there
are no irregularities on the surface. That would suggest that their growth
tendencies were practically exhausted at the time they were enclosed in the
socket. Their development was therefore not hampered by too narrow walls
of unyielding bone.

HISTOLOGICAL EVIDENCE

If this interpretation of the facts is correct, it should be demonstrable
histologically. Suitable stages of development are those in which the tooth
has already formed some enamel and dentine, but in which the tooth of the
normal has not yet started erupting. For the first molars this stage is found
at about the 9th or 10th day after birth. Longitudinal and transverse sections
were therefore cut serially through grey-lethals and normal litter-mates of
240 Hans Griineberg

that age. The results are shown on Pl. I. Fig. 1 represents a longitudinal
section through all the upper molars of a normal. As is clearly seen, the first
molar is everywhere clear of bone, there being an interval of normal width
all round it. Fig. 2, a corresponding section through a grey-lethal jaw, exhibits
a totally different situation. A big mass of cancellous bone presses hard
against the tooth on its front part, i.e. at the expected place. The erection of
the cusps which results from this is clearly seen, and it is further apparent
that the effect is strongest in the neighbourhood of bone pressure. Single
spicules of the spongy bone mass project towards the tooth, leaving here
impressions on the surface mentioned previously in this paper. Furthermore,
it is seen that the pressure, under which the tooth grew, could not be passed
on to the second molar since a bony interdental septum closes the socket from
behind. This offers the explanation why that tooth is not at all affected by
the pressure under which its neighbour developed. In fact it develops under
perfectly normal space conditions, as might have been predicted from its
normal shape.

Figs. 3 and 4, Pl. I, represent transverse sections through the front part
of the first upper molar, in the region of the deformed lateral front cusp.
Here again, the reason for the deformity in the grey-lethal is clearly a mass
of cancellous bone which in the normal course of events is removed by resorption. Figs. 5 and 6 show transverse sections through the tooth farther
back. Here there is hardly any lateral pressure on the grey-lethal tooth, while
a spicule hindering its median expansion is obvious.

Figs. 7 and 8 represent transverse sections through the lower first molar.
The compression of the grey-lethal tooth acts mainly from the lateral side as
seen by the position of projecting spongiosa. This situation corresponds exactly
with Brash’s conclusions about the growth of the mandible.

The same type of causation is found throughout. Where there is a deformity in any of the teeth, its cause, in the form of persistent spicules, is
readily found. Histological evidence, therefore, leaves no doubt of the correctness of the interpretation that the tooth shape anomalies are due to lack of
bone absorption.

THE UPPER INCISOR

A few words may be added about the upper incisors. In a normal mouse,
this tooth has an outline which is almost exactly a segment of a circle. This
applies already to the original tip, which is soon worn off by use. The axis of
the tooth lies exactly in one plane. The corresponding tooth of the grey-lethal
has an outline like an eagle’s beak. The tip is strongly turned downwards so
that the dorsal surface has two different curvatures meeting each other in an
obtuse and rounded angle. It seems that the extreme tip immediately after
the first dentine and enamel is laid down is subjected to some pressure from
behind. This pressure is caused by the fact that the growing back end of the
tooth finds resistance against unyielding bone. That results in an angle being
Factors in Bone and Tooth Development 241

formed between the tip which is already calcified or at least stiffened by
dentine and enamel, and the rest of the tooth which is still flexible. Actually,
this pressure does not act exactly along the axis of the tooth. In consequence
the tooth is forced into a very shallow spiral, the tip pointing somewhat
towards the middle plane of the body whereas the back opening faces laterally.

The position of the bending on the tooth allows for an estimate of the time
at which it occurs during development. If calcification of this tooth follows
the same course as in the rat, this should have happened during the first day
after birth (Addison & Appleton, 1915).

A very similar situation to that described here for the upper incisor sometimes occurs in human teeth due to an external trauma. If the calcified parts
of a developing tooth are forcibly dislocated as compared with the uncalcified
parts, but both continue their development, a bend with a distinct angle
between the two parts may result. This condition, chiefly found in the most
exposed front teeth, is called “‘dilaceration” in Dental Surgery (Bennett, 1931).

When comparing the width of the dentine and of the pulp cavity of upper
incisors in older animals, one is struck by the fact that the grey-lethal has, at
least in the front parts of the tooth, a considerably thicker layer of dentine,
but a much narrower pulp cavity. This, however, is not yet so in young
animals of about 10 days of age which do not exhibit any appreciable difference.
The reason for this is very simple. In a normal tooth which is worn off by
use at a regular rate, the single odontoblast has only a limited period during
which it may lay down dentine. This period lasts from the time it starts
functioning at the back end of the tooth until it approaches the tip where its
activities are automatically stopped by the formation of secondary dentine
by which the pulp cavity is obliterated. In the grey-lethal, however, the tooth
is not cut and therefore not worn off. No secondary dentine is therefore
formed near the tooth tip. The odontoblasts are therefore able to go on laying
down dentine as long as the animal lives and their physiological condition
allows. This results in a very thick layer of dentine, but a very narrow pulp
cavity.

DISCUSSION

The histological investigation leaves no doubt that the explanation deduced
from the shape of the grey-lethal molars is essentially correct. We have, therefore, a complete reversal of normal development. Whereas in the normal mouse
or any other mammal size and shape of socket is determined by the growth
of the tooth, the size and shape of the tooth in the grey-lethal is limited and
influenced by that of the socket at the time it becomes enclosed in it. Teeth
which have still to grow after that time are deformed, while teeth whose
growth was completed then develop normally.

Actually, a similar situation was described in the grey-lethal previously
but not fully understood then. The lower incisors of these animals, instead of
growing in the normal way backwards till they reach the basis of the condyloid

Anatomy LxXxI 16
242 Hans Griineberg

process, find a way out of the mandible through the mental foramen. This
phenomenon is essentially the same as the much less obvious anomalies
described in this paper. Here again, the bone does not yield to the pressure
of the backward-growing end of the tooth.

This situation is of some importance for the understanding of the bone
anomaly of the grey-lethal. As mentioned above, there is no secondary bone
absorption. Now it was doubtful whether this was due to a lack of stimulus
for absorption or to some other reason. What the actual stimulus is which
causes bone absorption, for example, at the surface of a long bone, is unknown.
It was therefore doubtful whether the unknown stimulus was absent for one
reason or another, or whether the stimulus acted normally, but the bone could
not respond to it. Now the situation is clear. For, in the case of a socket, the
stimulus for absorption is known to be the growing tooth germ. If that germ
grows vigorously, but the socket does not respond to its proper stimulus, it is
evident that the lack of secondary bone absorption is due to the inability of
the bone to respond, not to the absence of the stimulus.

This conclusion is of some importance for the general conception of the
relations of external and internal factors in bone development. It is well
known that bone responds to pressure by absorption. That has been worked
out for the relation of tooth and socket in great detail by Gottlieb & Orban
(1931). From work of this kind it was concluded that there is a direct response
of bone to pressure, a conclusion which was fully justified by the facts then
known. In the light of the facts described in this paper, this relationship
turns out to be more complex than then assumed. If the bone is to respond
to pressure by absorption, it must be able to do so. The ability of bone to
react, however, is evidently rooted in hereditary factors. If these factors act
normally, as is the case in all known animals with the exception of the greylethal mouse, the relationship between pressure and absorption appears to be
direct. If, however, this hereditary mechanism is upset, as in the grey-lethal,
the stimulus by itself is unable to induce absorption in the bone. The old
scheme was:

STIMULUS -—-> RESPONSE
(Pressure) (Absorption)

Now we have to fit in a new link:

STIMULUS —-> HEREDITARY BASIS FOR RESPONSE -——-> RESPONSE
(Pressure) (Absorption)

This link is disturbed in the grey-lethal. No response, therefore, can take
place. The physiological mechanism of the hereditary basis is unknown as
yet and is the subject of further studies.

In the light of these facts, certain suggestions previously made by the
author need revision. It was shown that in the grey-lethal the infra-orbital
foramen is considerably narrower than in a normal mouse. Incidentally, the
Factors in Bone and Tooth Development 243

anterior part of the Masseter which originates in that neighbourhood is very
poorly developed. As a tentative suggestion the explanation was put forward
that this is due to the comparative inactivity of the.muscle. Since the greylethal does not cut its teeth it does not nibble, and consequently uses its
masticatory muscles to a much lesser extent than a normal mouse. The explanation given then amounted to the assumption that in that case the
narrowness of the foramen was due to the lack of a proper stimulus for
absorption. It seems now that that assumption is untenable. The causal
relationship is probably reversed. Not: the hole is small since there is only
a weakly working muscle which does not stimulate enlargement. But: the
muscle cannot grow larger since the foramen is narrow. In this connexion
it is irrelevant that the muscle is little used anyhow.

As has been shown in this paper, all the anomalies in tooth shape in the
grey-lethal are secondary consequences of the lack of bone absorption, and
they all develop after birth. In the hierarchy of causes, therefore, the bone
anomaly is nearer the original gene action than are the tooth shape anomalies
dependent on it.

SUMMARY

In normal development, size and shape of socket are determined by the
growth tendencies of the tooth germ. In the grey-lethal mouse the growth
tendencies of the tooth are limited by the width and size the socket had when
the tooth became enclosed in it. This leads to the conclusion that bone reacts
by absorption to pressure only if the hereditary basis for doing so is undisturbed. The relation between stimulus and response is therefore not direct,
as assumed hitherto, but subject to conditions based in the hereditary constitution of an animal.

It is shown that all the anomalies of tooth shape in the grey-lethal are
caused by the lack of secondary bone absorption which is therefore a step
nearer to the original gene action.

ACKNOWLEDGEMENTS

The author wishes to express his thanks to Prof. J. B. S. Haldane, F.R.S.,
and Prof. D. M. S. Watson, F.R.S., for the continued interest they took in
the work. The microphotographs were taken by Mr J. R. Thomas of this
department. Particular thanks are due to Dr E. W. Fish (Royal Dental
Hospital, London) who very kindly put the technical experience in tooth
histology of his laboratory at the author’s disposal, and to his assistant,
Mr W. P. Pereira.

REFERENCES

Appison, W. H. F. & Appteton, J. L., jr. (1915). “The structure and growth of the incisor
teeth of the albino rat.” J. Morphol. vol. xxvi, pp. 43-96.

Bennett, Sir Norman (1931). The Science and Practice of Dental Surgery, 2nd ed. Oxford
Medical Publications. London: Oxford Univ. Press.

16—2
244 Hans Griineberg

Brasu, J. C. (1934). “Some problems in the growth and developmental mechanics of bone.”
Edinb. med. J. N.S. (tvth), vol. Xx.

Gorriies, B. & Orzan, B. (1931). Die Verdnderungen der Gewebe bei iibermassiger Beanspruchung
der Zéhne. Leipzig: Georg Thieme. .

GrtneBerea, Hans (1935). “A new sublethal colour mutation in the house mouse.
Soc. B, No. 809, vol. cxvi, pp. 321-42.

—— (1936 a). “Grey-lethal, a new mutation in the house mouse.” J. Heredity, vol. xxvu,
pp. 105-9.

—— (1936 b). ““Komplette Retention des Gebisses, ein neuer Erbfaktor bei der Hausmaus.”
Schweiz. Mschr. f. Zahnheilk. (in the press).

ey

Proc. roy.

DESCRIPTION OF PLATE I

The technique is the same for all the preparations. Suza fixation. Haematoxylin, van Gieson;
7p; 35 times enlarged.

The animals from which Figs. 1 and 2 were made were litter-mates, 9 days old. Figs. 3, 5
and 7, and 4, 6 and 8 respectively, were taken from another pair of animals (litter-mates) of
10 days. Despite this age difference, they represent practically the same stage of tooth development (just before the eruption of the incisors), since the older animals came from a large litter
which develops more slowly.

In these young stages, the enamel freshly laid down is not dissolved entirely by the process
of decalcification. It is seen as a darkly staining layer in all the sections with the exception of
Figs. 7 and 8, where it fades away towards the tips of the cusps. The approximate planes of the
sections are indicated in Text-figs. 2 and 4.

Figs. 1 and 2. Longitudinal sections through the upper molars of normal and grey-lethal. The
first molar of the normal is clear of bone all round. In the grey-lethal dense spongiosa is
pressing against the front parts of the tooth, leaving marked impressions on the surface and
leading to erection of the cusps. No pressure acts upon the second molar of the grey-lethal.

Figs. 3 and 4. Transverse sections through front parts of right upper first molar of normal and
grey-lethal. In the grey-lethal tooth spongiosa spicules press into the tooth mainly from
the lateral side where there are considerably more in the actual preparation than are brought
out in the photograph. One spicule presses strongly on the median side. On the lateral side
the enamel organ is complexly invaginated by spicules; such spicules are covered with a
layer of adamantoblasts which have deposited enamel on the bone surface.

Figs. 5 and 6. Transverse sections through left upper first molars of normal and grey-lethal,
farther backwards than the last two sections. There is little direct pressure in this region
acting on the grey-lethal tooth except for one spicule pressing from the median side, strongly
affecting the curvature in that neighbourhood. Note, too, the steeper shape of the middle
cusp.

Figs. 7 and 8. Transverse sections through left lower first molar of normal and grey-lethal. In
the abnormal tooth the resistance to growth is exclusively on the lateral side where there
are many more spicules than are brought out in the photograph. They form a strong obstacle
particularly for the downward growth of the lateral wall (compare Text-fig. 1).
Journal of Anatomy, Vol LX XI, Part 2

 

GRUNEBERG—Facrors 1x Bonz anp Tooty DEVELOPMENT