The effect of dietary consistency on bone mass and turnover in the growing rat mandible

Archsoral Biol. Vol. 36, No. 2, pp. 129-138, 1991

0003-9969/91 53.00 + 0.00 Copyright 6 1991 Pergamon Press plc

Printed in Great Britain. All rights reserved

THE EFFECT OF DIETARY CONSISTENCY ON BONE MASS AND TURNOVER IN THE GROWING RAT MANDIBLE KAZUHIRO YAMADA*

and DONALD B. KIMMEL

Center for Hard Tissue Research, School of Medicine, Creighton University, Omaha, Nebraska, U.S.A. (Accepted 4 September 1990) Summary-Hard and soft diets were fed to weanling rats for up to 8 weeks. Some animals were switched after 4 weeks to the opposite diet. A histomorphometric study of bone formation activity at the mandibular ramus, body, and condyle was made after in oiuo fluorochrome labelling. Mineral apposition rates at the lateral and inferior periosteal surfaces of the ramus were lower in the soft diet than in the hard diet animals. The rate of bone formation at the lateral pexiosteal surface of the ramus was significantly lower in soft than in hard diet animals. The medial periosteal surface of the ramus sometimes

changed to bone formation in the soft diet groups. Condylar cartilage zones were somewhat thinner in soft diet groups. In the mandibular body, differences due to dietary consistency were less marked than near the gonial angle. Adaptation of periosteal bone and condylar cartilage to a new dietary consistency occurred within 4 weeks of switching. These results suggest that lateral and inferior periosteal bone growth of the ramus and condylar elongation were slowed in rats consuming soft diets. Decreased functional force during rapid mandibular bone growth causes changes in shape. The changes are due to regional decreases in osteoblast function, realignment of bone formation surfaces in the ramus area, and slowed growth in the condylar cartilage. Key words: dietary consistency, mandible, rat, growth, bone histomorphometry.

INTRODUCTION

The incidence of retarded mandibular growth appears to have risen during the past few thousand years. Anthropological studies report a larger gonial angle and narrower mandible in modem than in medieval skulls (Mohlin, Sagne, and Thilander, 1978; Hanihara et al., 1981; Shiono ef al., 1982). Some believe that the softer diet consumed in modem societies is likely to generate reduced masticatory forces. They contend that the soft diet is responsible for the trend toward increased malocclusion from dental crowding, which implies narrower, shorter jaws (Waugh, 1937; Wood, 1971; Inoue, 1980; Shiono et al., 1982; Corrucini and Lee, 1984). Many experiments have shown that feeding soft diets to growing animals creates changes in the condyle and ramus which cause a shorter, narrower mandible. Mandibular size and shape have been investigated in mice, rats, and monkeys fed soft diets (Watt and Williams, 1951; Barber, Green, and Cox, 1962; Kikuta, 1985; Kiliaridis, Endostrom, and Thilander, 1985; McFadden, McFadden, and Precious, 1986; Ito, Mitani and Kim, 1988). These studies show that soft diet animals not only have smaller mandibles with less mineral content, but also a lower ramus. The soft diet thus creates changes that *Address correspondence to: Dr Kazuhiro Yamada, Department of Orthodontics, Niigata University School of Dentistry, Japan. AOB M/Z--c

2-5274 Gakkocho-Dori,

Niigata,

951

mimic the differences between medieval and presentday jaws. Soft diet animals have thinner, more porous cortical bone with poorly developed trabeculae in the ramus and generally slower mineral apposition (Kiliaridis, 1989). Many workers have found smaller condyles and thinner condylar cartilage in rats fed soft diets (Bouvier and Hylander, 1984; Bouvier, 1987; Bouvier, 1988; Hinton and Carlson, 1986). Others have shown less bone remodelling in the cortical bone of the mandibular body (Bouvier and Hylander, 198 1). All the reports point toward decreased mandibular growth in animals fed soft diets. They even suggest that the changes are most prominent in the ramus. However, there are no reports quantifying bone cell activity in various regions of the mandible during feeding of a soft diet. Our purpose now was to examine the effect of soft diets on bone formation activity in the mandibular ramus, body, and condyle in weanling rats. A second purpose was to examine the speed at which bone formation patterns adapt to changes in dietary consistency. MATERIALS

AND METHODS

Thirty-six female rats, aged 21 days, were obtained from SASCO (Omaha, NE). They were caged individually and given free access to water and hard or soft diets. They were randomized by weight into six groups. The experimental design is explained in Text Fig. 1. The rats were weighed each week. The design was approved by the Creighton University

KAZUHIROYAMADAand DONALDB. KIMMEL

130 Day

0

56

=P

N

28

1

6

__-__-_-_-_-_-_--0

2 3 4 5 6

6 6 6 6 6

X -_________------------0 X __________x ______---s-0

Fig. 1. Experimental design. Group 1 was fed a soft diet for 4 weeks and then killed. Group 2 was fed a hard diet for 4 weeks and then killed. Group 3 was fed a soft diet for 8 weeks and then killed. Group 4 was fed a soft diet for 8 weeks and then killed. Group 5 was fed a soft diet for 4 weeks, switched to a hard diet for 4 weeks, and then killed. Group 6 was fed a hard diet for 4 weeks, switched to a soft diet for 4 weeks, and then killed. ---: Soft diet; x: 6 rats from hard diet killed; 0: 6 diets from soft diet killed.

Animal Care Committee, and conducted in accordance with all federal guidelines to assure humane treatment of the animals. Rats in the hard diet group were fed a standard solid diet (Ralston-Purina, St Louis, MO). The soft diet animals were given the same diet in a powdered form, with water added. The soft diet was presented in aluminium dishes placed inside the cages. These dishes were cleaned and refilled every morning. Before death, each animal received an intraperitoneal, double label of oxytetracycline (15 mg/kg; Lederle, Pearl River, NJ) and calcein (10 mg/kg; Sigma, St Louis, MO). The labelling sequence was 2262-3, that is 2 days on oxytetracycline, 6 days off, 2 days on calcein, 3 days off, and then killing. The interlabel interval was 8 days. After death, the right mandible was removed and placed in 70% ethanol. Three cuts were made with a diamond disc (Text Fig. 2). First, the condyle was removed from the body [Cut (a)]. Next, the portion of the mandible anterior to the first molar was removed [Cut (b)]. Finally, a third cut was made just posterior to the third molar [Cut (c)l. After 24 h, we moved all the pieces to Villanueva bone stain (Vilianueva, 1974). After 72 h, we returned the pieces to 70% ethanol. We embedded the condyle, body, and ramus regions separately in methyl methacrylate. We then sawed lSOpm-thick frontal sections through the body region (Text Fig. 2B) with a low-speed metallur-

gical saw (Model 650, South Bay Technology, Temple City, CA). We also made frontal sections of the ramus that included the region of the gonial angle (Fig. 2R). We ground them by hand to about 80pm thickness on glass plates with levigated alumina. Finally we cleared them in ethanol and xylene, and mounted them with Permount on glass slides. Condyles were sectioned parasagittally on the Jung Model K microtome. Three micrometre sections were prepared, afixed to glass slides, and stained with modified Goldner’s trichrome. DATA COLLECTION

We did all quantitation with the IBM-PC version of BIOQUANT II semi-automated image analysis system (R&M Biometrics, Nashville, TN). We used a microscope with camera lucida projecting on to a HIPAD graphics pad with lighted cursor. The principles applied during this histomorphometric analysis are discussed in great detail elsewhere (Kimmel and Jee, 1983). Ramus Area

Measurements were taken in the area from the inferior border to 1.5 mm superior (Text Fig. 3). At 40 x magnification, we measured the superiorinferior (si) and mediolateral (ml) dimension of the inferior portion of the ramus (Text Fig. 3). Periosteal surface

The periosteal surfaces were delimited as in Text Fig. 3, into lateral, inferior, mediosuperior, and medial surfaces. For each region, at 40 x magnification, the total length of surface (BS) and length of doublelabelled fluorochrome surfaces (dLS) were measured. The interlabel width of the double fluorochrome labels was measured at 400 x magnification.

r

1.5mm C

m

1

Fig. 2. Lateral view of the rat mandible. The thick lines (a, b, c) indicate the cuts made before embedding Frontal sections from the body area (B) and ramus area (R) were prepared (thin lines). Parasagittal sections were prepared through the condyle area (C).

Fig. 3. Schematic drawing of section from the ramus area. The whole area of the bone studied is delineated by oblique lines. Measurements of width (l-m) and height (s-i) were made. Fluorochrome labelling at the periosteal surfaces on the lateral (L), inferior (I), medial-superior (MS), and medial (M) aspects was also measured.

Diet consistency and mandibular growth Total bone tissue in ramus

First, at 40 x magnification, the whole area of the bone inside the periosteal envelope was traced (TV) (area of Text Fig. 3 with oblique lines). Next, the bone marrow was outlined (MV), a movement that gave both bone surface (BS) and marrow area (MV). Next, at 160 x magnification, the length of double labelled fluorochrome surfaces (dLS) was traced. The interlabel width of double fluorochrome labels (IrL.Wi) was measured at 400x magnification.

(a)

s

i Irnnl

1

Body Region (Beneath Distal Root of First Molar)

The cancellous bone inferior to the mandibular nerve and lateral to the mandibular incisor was measured (Fig. 4).

tbi

Periosteal bone

The interlabel width of the double fluorochrome labels (IrL.Wi) at the periosteum was measured at the lateral, inferior, and medial cortices. Cancellous bone

First, at 40 x magnification, the whole bone-tissue area (TV) was traced. Next, the bone marrow (MV) was outlined. Next, at 160x magnification, the length of double labelled fluorochrome surfaces (dLS) was traced. The interlabel width of double fluorochrome labels (IrL.Wi) was measured at 400 x magnification. Other areas

The interlabel width of double fluorochrome labels was measured at four places in the incisor dentine. The interlabel width in the apical bone beneath the first molar was also measured. Condyle Region

Bone volume (BV) measurements were taken inside a box whose maximum inferior distance was 1 mm from the surface ‘of the articular cartilage (Text Fig. Sa). It was very similar to the subcondylar area of Bouvier (1988). The superior boundary of the counting box was the same as used by Bouvier (1988). Its width was two-thirds the total width of the section. Data were collected separately for the pri-

Fig. 5. (a) Schematic drawing of section from condyle area. The cancellous bone inside the thick lines was measured. Note the sampling box in the cartilage which is shown in more detail in Text Fig. 5(b). (A-anterior; P-posterior; S-superior.) (b) Schematic drawing of portion of cartilage from condyle area. Measurements of thickness were made% the moliferative Ip). transitional CT). and hvnertrophv (H) zones. Bone volnme in the primary spongibsa (PS) and secondary spongiosa (SS) was measured. mary and secondary giosa (Bloom and

spongiosa.

The primary

spon-

Fawcett, 1975) began at the superior boundary of subcondylar region and included the area of the subcondylar region that contained predominantly bone cells. The remainder was the secondary spongiosa. The subcondylar bone area was counted separately at 40 x magnification. In the cartilage, the following measurements of thickness were made at the superior aspect of the condyle: (1) proliferative zone, (2) transitional zone, and (3) hy pertrophic zone (Text Fig. Sb and Plate Fig. 6). We followed the example of Bouvier (1988) in defining the zones of the cartilage. CALCULATIONS

Ramus Region

For the periosteal surfaces, we calculated, individually, for the lateral, medial, medial-superior, and inferior surfaces. We calculated the mineral apposition rate by measuring the interlabel width at many places in each region. We used equations to calculate: Percentage of surface with double label; Mineral apposition rate; and Surface-based bone formation rate. Fig. 4. Schematic drawing of section from body area. The cancellous bone between the two parallel thick lines was measured. The apical bone in the black square was also studied, as was the mineral apposition rate at the lateral (L), inferior (I), and medial (M) surfaces. Dentine apposition rate was studied by 4 evenly-spaced measurements around the incisor.

For the total bone (inside the periosteal envelope) of the ramus, we calculated: Bone volume; Percentage of surface with double label; Mineral apposition rate; and Surface-based bone formation rate.

KAZUHIRO YAMADA

132 W/TV

=

(TV - MV)* 100 ’

TV

dLS* 100 dLS/BS = BS ZIrL.Wi MAR=---N * IrLT’ BFR/BS =

MAR * dLS/BS 100

’

where: BV(%), bone volume; TV(mm2), tissue volume; MV(mm2), marrow volume; dLS(mm), double label surface; BS(mm), bone surface; IrLT(day), interlabel time period; IrL.Wi(%), interlabel width; MAR@m/day), mineral apposition rate; BFR/BS(mm2/mm/yr), bone formation rate. Body Region

For the cancellous bone, we calculated the same end-points as in the ramus area. In addition, we calculated the dentine apposition rate (DAR) in the incisor, and the mineral apposition rate of the apical bone beneath the first molar. For DAR, we used an equation similar to MAR, except that we used the interlabel width of the dentine. For the periosteal surfaces, we calculated only the mineral apposition rate (MAR). Condyle Region

We calculated the mean thickness of each cartilage zone and also cancellous bone volume. RESULTS

There were no significant differences in weight gain among the groups. Ramus

There were no significant differences in superiorinferior and mediolateral dimensions in the ramus region among the groups (Table 1). Periosteal bone (Table 2) Double label surface (dLS/BS). Rats eating hard diets at the end of the experiment had higher lateral double-label surfaces (dLS/BS) than rats eating soft

and

DONALD B. KIMMEL

diets (Plate Fig. 7). However, the difference between hard and soft diet animals was less at 8 weeks than it had been at 4 weeks. In fact, lateral dLS/BS was significantly higher at 4 weeks in the hard than in the soft diet groups (p < O.OOS),but not significantly higher at eight weeks. However, just 4 weeks after switching diets, the labelling changed dramatically. It increased from 2.7 to 63.0% in rats switched from soft to hard diets, and fell from 58.7 to 15.2% in rats switched from hard to soft. All inferior surfaces in all groups were 100% labelled. There were no significant differences in double labelling at the mediosuperior surface. Double labelling at the medial surface was usually seen only in the soft diet groups. Mineral apposition rate (MAR). Rats eating hard diets near the end of the experiment had higher lateral mineral apposition rates (MAR) than rats eating soft diets. The mineral apposition rate at the lateral surface was significantly higher in the 4-week, hard diet groups than in soft diet groups. Moreover, 4 weeks after switching diets, the lateral mineral apposition rate changed dramatically. It climbed from 0.5 to 2.7 pm/day in rats switched from soft to hard diets, and fell from 2.7 to 1.1 pm/day in rats switched from hard to soft diets. Inferior MAR was significantly higher in the 8-week, hard diet groups than in the I-week, soft diet group (p < 0.05). The differences were less marked than those at the lateral surface. At 4 weeks, the inferior MAR had changed little in rats switched from soft to hard diets, but markedly in those switched from hard to soft. Mediosuperior MAR was the same in all groups. In the hard diet groups, there was usually no fluorochrome labelling at the medial surface. In contrast, half of the animals in the soft diet groups showed medial labelling at this site. Therefore, the MAR at the medial surface was less in hard than in soft diet groups. Bone formation rate (BFRIBS). Rats eating hard diets at the end of the experiment had higher lateral bone formation rates (BFR/BS) than those eating soft diets. Lateral BFR/BS was significantly higher at 4 weeks in the hard than in the soft diet groups (p < 0.005). Four weeks after switching diets, the lateral bone formation rate was much different. It had increased from 0.016 to 0.604 in rats switched from soft to hard diets, and dropped from 0.586 to 0.122 in rats switched from hard to soft diets. Mediosuperior BFR/BS was not significantly different. Summary of bone cell activity. Bone formation at the ramus periosteum was lower at the lateral and

Plate 1 Fig. 6. Light photomicrograph of condylar cartilage. The condyiar cartilage and adjoining primary spongiosa from a rat of Group 4, fed a hard diet for 8 weeks, is shown (a) together with the same area from a rat of Group 3, fed a soft diet for 8 weeks (b). All zones of the cartilage are smaller in the soft diet animal. The proliferative (p), transitional (t), ‘and hypertrophic (h) zones are labelled. Modified Goldner’s trichrome, 500 x . Fig. 7. Ultraviolet photomicrograph of ramus area. The ramus area from a rat of Group 4, fed a hard diet for 8 weeks (a) and the same area from a rat of Group 3, fed a soft diet for 8 weeks (b). The lateral periosteal surface of the soft diet group bears less fluorochrome label than that of the hard diet group (arrows). The lateral (L) and inferior (I), aspects are labelled. Villanueva stain, 37 x ; Villanueva (1974).

Diet consistency

and mandibular

Plate

I

growth

133

KAZCHIKO Y.AMADA and

134

Table

I. Drmenstons

[IONALU B. KIMMEI

(mm) of ramus

inferior

border

I

2

3

4wkS

4wkH

8wkS

4 8wkH

5 SiH

6 H/S

1.16 0.13 0.47 0.07

1.16 0.12 0.47 0.06

I.17 0.19 0.56 0.12

I.11 0.13 0.44

I .03

Width (.C SD) Hetght $ (.C + SD)

0.13 0.53 0.08

1.14 0.08 0.57 0.04

0.11

H: Hard diet group. S: Soft diet group. H/S: Switching diet group from hard to soft. S/H: Switching diet group from soft to hard. inferior surfaces of animals consuming soft diets. These changes are shown diagrammatically in Text Fig. 8. Over time, they would cause a shorter ramus and narrower mandible than in animals on a normal diet. Total bone

(inside periosteal

envelope)

(Table

3)

There were no intergroup differences in bone volume. Mineral apposition rate, double-label surface, and bone formation rate were significantly higher in the 4-week, hard diet than in 4-week soft diet groups. MAR was also lower in rats switched to soft diets than in those that had always been given the hard diets. Body Region Periosteal

mineral

apposition

rate (Table

4)

The mineral apposition rate at both lateral and inferior periosteal surfaces was somewhat lower in soft than in hard diet rats at 8 weeks. It also decreased after animals changed from hard to soft diets. However, the change was less prominent than that in the ramus area. There were no differences in the apical bone MAR or dentine apposition rate. Cancellow

bone

(Table

5)

Bone volume and mineral apposition rate did not differ among the groups. The double-label surface and bone formation rate did not vary in the animals that ate all one type of diet. The double-label surface and bone formation rate in the group that switched from hard to soft diets was significantly below all others. Table

2 4wk H

dLS/BS (%) (.T k SD) MAR (pm/day) (.i + SD) BFR (%/yr) (s + SD)

2.7 4.1 0.5 I.0 0.016 0.028

58.7d 13.2 2.7b 0.4 0.586‘ 0.178

MAR (pm/day) (.: + SD)

4.8 1.5

Abbreviations

as in Table

Cartilage

thirkness

(Table

Region 6)

All zones of the condylar cartilage tended to be shorter in animals on soft diets than in those on hard (Plate Fig. 6). The differences between the hard and soft diet animals were significant at 4 weeks for the hypertrophic zone, and at 8 weeks for the proliferating zone. In the animals who switched from soft to hard diets, the thickness of the condyle cartilage returned to normal. Bone aolume

(Table

7)

The bone volume in the primary spongiosa was significantly higher in the 4-week, hard diet groups than in the 4-week, soft diet group. However, there was no difference between the groups at 8 weeks. In the secondary spongiosa, bone volume was greater in hard than in soft diet groups. In the animals who switched from hard to soft diets, no significant changes occurred. DISCUSSION

We studied bone-forming activity in various parts of the mandible in growing rats fed hard and soft diets. The major effect of the soft diet was to reduce bone apposition at the lateral and inferior aspects of the periosteal surface. This reduction was most significant near the gonial angle, but also showed somewhat in the body. It was both a reduced mineral apposition rate and a reduced extent of formation surface. Soft diets also caused a slower rate of

2. Ramus periosteal bone formation

I 4wk S

Symbols for group comparisons

Condyle

6.2 0.7 I versus 2 “p < 0.05 bp < 0.02 ‘p < 0.01 $I < 0.005 ep < 0.001 I.

3 8wk S

4 8wk H

Lateral border 50.5 37.3 30.6 20.2 3.1 I.9 I.1 0.6 0.662 0.292 0.646 0.225 Inferior border 5.8’ 4.3 0.9 0.8 3 versus 4 ‘p c 0.05 p? < 0.02 hp < 0.01 ‘p < 0.005 ‘jr < 0.001

5

6

S/H

HIS

63.0 21.7 2.7 0.5 0.604 0.153

15.2Q 17.3 l.lP I.2 0.122 0.146

5.2 1.3

2.7’ 1.1

4 versus 5 “p < 0.05 ‘p < 0.02 “p < 0.01 “p < 0.005 “p < 0.001

4 versus 6 pp < 0.05 qp < 0.02 ‘p < 0.01 “p < 0.005 ‘p < 0.001

Diet consistency and mandibular growth

135

Fig. 8. Schematic drawing of periosteal bone formation in ramus region with (a) hard or (b) soft diets. Bone formation is prominent at the lateral and inferior borders of the hard diet group (a). Bone formation is, in general, less prominent in the soft diet group (b), and is restricted mainly to the inferior border. However, some formation occurs at the medial surface, which was usually not seen in

the hard diet group. condyle growth. Both effects, that is a lower rate of periosteal bone formation in the ramus, and a slower rate of growth of condylar cartilage, are consistent with a reduced ramus height. A decreased rate of lateral periosteal bone formation in the ramus is consistent with the development of a narrow mandible. These findings are consistent with the trend toward decreasing mandible size discovered in anthropological research (Hanihara et al., 1981; Mohlin et al., 1978; Shiono et al., 1982). As hard and soft diet groups gained weight at equal rates, it is unlikely that the differences are due to nutritional deprivation of the soft diet group. The changes in inferior and lateral periosteal bone formation patterns in the mandible are most likely due to decreased forces applied during mastication. Our results are similar, but less dramatic, than those seen when hard diets are fed to animals with uni-

lateral or bilateral loss of masticatory muscle function (Horowits and Shapiro, 1955; Avis, 1961; Kikuchi et al., 1978; Fukazawa and Sakamoto, 1982). When that function is completely lost, the gonial angle is severely underdeveloped. When function is only decreased, as with soft diets (Maeda et al., 1987; Kiliaridis, Endostrom and Thilander, 1988; Kiliaridis and Shyu, 1988; Maeda et al., 1988), the ramus area becomes more porous (Kiliaridis, 1989). Our data from soft diet animals show changes in bone cell function at the ramus periosteal surface consistent with diminished external growth of the mandible. We conclude that there is a dose-effect relationship of muscle function to osteoblast function at the lateral and inferior surfaces of the gonial angle. When it is mildly decreased, as in our study, mild underdevelopment occurs. When it is moderately decreased, as with nerve resection of one of the masticatory

Table 3. Ramus internal bone formation

dLS/BS (%) (f f SD) MAR (JIm/day) (a f SD) BFR (%/yr) (5 f SD)

1 4wk S

2 4wk H

3 8wk S

4 8wk H

30.3 1.7 3.7 0.7 0.407 0.072

41.0d 2.1 5.6 1.0 0.84b 0.166

37.4 12.2 4.1 2.1 0.546 0.264

36.4 3.5 4.1 1.0 0.552 0.175

SjH

H6/S

35.6 10.0 4.0 :::97

50.5 33.4 3.op 0.3 0.531 0.355

0.096

Abbreviations as in Table 1. Symbols for group comparisons as in Table 2. Table 4. Mineral apposition rate (Dm/day) at body periosteal surface

Lateral (a f SD) Inferior (2 f SD) Medial (R f SD)

1 4wk S

2 4wk H

3 8wk S

4 8wk H

3.2 1.7 6.0 0.8 3.6 1.8

3.5 0.9 5.8 0.9 4.6 1.2

1.7 0.5 2.8 0.7 2.4 0.7

2.0 0.1 3.8’ 0.7 2.6 0.7

Abbreviations as in Table 1. Symbols for group comparisons as in Table 2.

H;S 2.0 0.4 3.8 0.5 3.0 0.3

1.4s 0.4 2.0’ 0.2 1.9 0.2

136

and

KAZUHIROYAMADA

DONALD

B. KIMMEL

Table 5. Body cancellous bone formation

MAR 01m/day) (a + SD) dLS/BS (%) (m + SD) BFR (%/yr) (m k SD)

1 4wk S

2 4wk H

3 8wk S

4 8wk H

S;H

6 HIS

3.5 0.3 34.0 6.6 0.434 0.125

3.2 0.9 27.6 9.5 0.309 0.099

2.1 0.7 22.3 12.9 0.190 0.159

2.6 0.7 28.0 9.6 0.271 0.127

2.2 0.3 25.3 8.9 0.197 0.063

2.0 0.7 10.7’ 4.0 0.084’ 0.051

Abbreviations as in Table 1. Symbols for group comparisons as in Table 2. Table 6. Condylar cartilage thickness by zone brn) 4wk S

2 4wk H

3 8wk S

4 8wk H

S/5H

6 H/S

39.9 16.3 41.2 8.0 44.9 11.9

43.0 9.0 48.2 11.2 68.9b 11.7

22.Or 6.8 28.7 11.9 35.1 7.6

37.0 4.9 39.1 9.7 47.4 11.6

36.7 2.1 41.1 5.3 40.9 5.2

41.0 13.3 40.0 4.2 42.0 7.3

1

Proliferative (a k SD) Transitional (a k SD) Hypertrophy (f t: SD)

Abbreviations as in Table 1. Symbols for group comparisons as in Table 2. muscles,

moderate

underdevelopment

occurs.

When

it is severely decreased, as with surgical removal of the masseter muscle, severe underdevelopment occurs. We found that the periosteal surfaces in the body region were less responsive to the soft diet than those of the ramus. This agrees with the earlier, morphological work on rats and mice fed soft diets (Kikuta, 1985; Kiliaridis et al., 1985; Ito et al., 1988; McFadden er al., 1986). Muscle resection (Horowits and Shapiro, 1955; Avis, 1961; Fukazawa and Sakamoto, 1982) and experiments with masseteric neurectomy (Kikuchi, 1978) also produced less change in the anterior area of mandibular body than in the ramus. The difference may occur because the insertions of major masticatory muscles are located mostly in the ramus rather than in the body. It seems logical that changes in muscle usage would have their most profound effects on the area near the insertion where forces are greatest. This difference between the periosteal surfaces in the body and ramus regions seems consistent with the pattern of bone strain generated during mastication. In dogs, the use of multiple strain gauges showed that mandibular strains during chewing are greater in the region posterior to the molars than in the subincisor region (Sugimura et al., 1984). An in vitro analysis of photoelastic models of human mandibles in masticatory motion showed that stress patterns were much more prominent

in the ramus area than in the molar

region (Standlee, Caputo and Ralph, 1977). We infer from this that causing reduced masticatory forces will cause a greater relative change in forces in the posterior area of the mandible than in the mid-body region. Our data, which show a greater decline in bone formation activity in the ramus area than in the body area, imply that it is the relative reduction in local strain that dictates the response. Moyers and Enlow (1988) stated that growth and shaping of the tooth-bearing area of the mandible are controlled more by tooth eruption than by intrinsic osteogenic factors or extrinsic muscle function. Kiliaridis (1989) suggested that the continuously erupting incisor in the rat, which occupies a large part of the mandibular corpus, may play a role in the reduced changes seen in the mandibular body of a soft diet group. Our findings also leave open the possibility that lower levels of strain at the periosteal surface, due to few nearby muscle insertions, may also play some role in the lower rates of bone formation seen at the body than at the ramus. We found that the internal bone of the ramus and the cancellous bone of the body responded with only transient declines in their formation. Bone volume was never different. Moreover, by 8 weeks, no difference persisted between the different diet groups. It is likely that internal or cancellous bone, being in the interior of the mandible, is more isolated from muscle function than is the periosteal surface. Changes in muscle function affect the periosteal cells more than

Table 7. Bone volume in condyle spongiosa (%)

Primary (*i f SD) Secondary (f zt SD)

1 4wk S

2 4wk H

3 8wk S

4 8wk H

S:H

6 H/S

17.8 3.3 41.4 4.6

29.1b 6.9 51.5b 6.3

26.5 10.7 42.7 6.5

28.5 2.3 56.0 8.7

18.6k 6.0 51.7 5.4

24.2 7.8 45.2 7.1

Abbreviations as in Table I. Symbols for group comparisons

as in Table 2.

Diet consistency and mandibular growth

the endosteal cells. It is possible than the cancellous bone of the mandible is more affected by systemic factors, like low calcium diets, than by muscle function (Kiliaridis, 1989). All zones of the condylar cartilage were affected somewhat by changes in dietary consistency. Significant decreases were Seen in the soft diet animals at 4 weeks in the proliferative zone and 8 weeks in the hypertrophic zone. The trabecular bone volume of secondary spongiosa decreased in soft diet groups. These findings suggest mildly decreased growth at the condylar cartilage during decreased muscle function.

This agrees with the findings of Bouvier and Hylander (1984), Hinton and Carlson (1986) and Bouvier (1988). It also agrees with data from experimental muscle resection (Fukazawa and Sakamoto, 1982). Others have shown that the condyle is the destination of the major trajectories of masticatory stress within the mandible (Standlee er al., 1977). In that case, even small declines in masticatory forces would be expected to generate changes in condylar growth patterns. Bouvier (1987, 1988) and Bouvier and Hylander (1984) showed that 2 weeks after a return to a hard diet from a soft diet, recovery of cartilage thickness, condylar bone volume, matrix synthesis, and enzymatic activity of condylar cartilage cells occurred. Our study suggests that adaptation of both condylar cartilage and periosteal bone to a new dietary consistency occurred by 4 weeks after switching. The normal mechanical loading of normal mastication pattern with normal diet seems important for periosteal bone cell and cartilage cell activity in the growing mandible. Acknowhdgemenr-We thank Toni Coble for her help and advice during dissection and data-gathering in this experiment.

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