51 skeletal strain patterns in relation to exercise training ... - CiteSeerX

J. exp. Biol. 185, 51–69 (1993) Printed in Great Britain © The Company of Biologists Limited 1993

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SKELETAL STRAIN PATTERNS IN RELATION TO EXERCISE TRAINING DURING GROWTH ANDREW A. BIEWENER AND JOHN E. A. BERTRAM* Department of Organismal Biology and Anatomy, The University of Chicago, 1025 East 57th Street, Chicago, IL 60637, USA Accepted 19 July 1993

Summary Bones are believed to alter their shape in response to changes in tissue strains produced by physical activity and the goal of this study is to examine whether modeling responses of a growing bone to changes in physical exercise are adjusted to maintain a uniform distribution of functional strains. We test this idea by comparing in vivo strains recorded in the tibiotarsus of white leghorn chicks during ‘intensive’ treadmill exercise (60% of maximum speed, carrying a weight equal to 20% body weight on the trunk: 60%/L) with strains that had been recorded previously during ‘moderate’ treadmill exercise (35% of maximum speed, unloaded: 35%/UNL) at similar bone sites. Our hypothesis is that modeling adjustments of bones subjected to the intensive load-carrying exercise should re-establish strains recorded in the bones subjected to moderate exercise. At each exercise level, the animals were exercised for 5 days per week (2500 loading cycles per day) from 2 to 12 weeks of age. As in the moderate exercise group studied earlier, strains measured at six functionally equivalent sites on the tibiotarsus of the 60%/L group were consistently maintained during growth from 4 to 12 weeks of age. In addition, the pattern of strain recorded at these sites was uniformly maintained over the full range of speeds recorded (from 0.48 to 2.70 ms 21 at 12 weeks of age). Peak strains measured at 4 weeks of age in the load-carrying exercise group were initially elevated by 57% overall compared with peak strains recorded in the moderate exercise group. At 8 weeks of age, strain levels in the 60%/L group differed by only 4% overall compared with those recorded in the 35%/UNL group. The nature of strain (tensile versus compressive) and the orientation of principal strain at corresponding sites were also similar in the two groups. At 12 weeks of age, however, bone strain levels in the 60%/L group were again elevated (47% overall) compared with those recorded in the 35%/UNL group, although the general pattern and orientation of strains remained similar. This finding suggests a transient modeling response of the bone to the onset of exercise training, which was lost during subsequent growth, possibly because the normal pattern of functional strain was not altered significantly by the faster load-carrying exercise.

*Present address: Department of Anatomy, New York College of Veterinary Medicine, Cornell University, Ithaca, NY 14805, USA. Key words: bone strain, exercise, skeletal growth, adaptive modeling, chick.

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A. A. BIEWENER AND J. E. A. BERTRAM Introduction

It is commonly accepted that bones continually adjust their shape and mass in response to changes in mechanical loading patterns during an animal’s lifetime. The extent to which a bone can alter its form and the extent to which growing and mature bones have similar adaptive capacities, however, remain unclear (Bertram and Swartz, 1991; Biewener and Bertram, 1992). In addition, the precise structural objective of adaptive modeling and the mechanical stimuli that may initiate it are poorly understood. It seems likely that functional adaptation must depend on certain features of a bone’s loading history (Carter et al. 1981; Carter, 1984, 1987), being mediated in some fashion by the cyclic strains developed in the bone associated with its use (Hert et al. 1971; Lanyon, 1984; Rubin, 1984). Rubin and Lanyon (1984) have shown that surprisingly few (>36cyclesday 21), high-magnitude (2000 me, strain3106) loading cycles may be sufficient to elicit a modeling response of a bone. Given that local tissue deformation is the direct, or indirect, agent of this response, it would seem likely that the functional integrity of each bone element in the skeleton could best be achieved and maintained by matching the bone’s form and mass to patterns of strain experienced during normal activity. The consistent pattern of surface midshaft strains recorded in the long bones of a number of species over a range of speed and change of gait supports this view (Lanyon and Baggott, 1976; Rubin and Lanyon, 1982; Biewener et al. 1983, 1988; Biewener and Taylor, 1986). The purpose of this study is to test further the hypothesis that, in response to changes in physical activity, skeletal modeling maintains a uniform distribution of strains at functionally equivalent sites on a growing bone. We found previously (Biewener et al. 1986) that the in vivo pattern of surface strains in the chick tibiotarsus engendered by moderate exercise remained unchanged as the animals grew from 4 to 17 weeks of age (corresponding to a 10-fold increase in mass and a 3.3-fold increase in length of the bone). Though differing among sites, the magnitude, nature (tensile versus compressive) and orientation of principal strains at each site remained largely unaltered during this period of growth. These findings are consistent with those of Lanyon and his colleagues involving studies of older pigs (Goodship et al. 1979) and skeletally mature sheep (Lanyon et al. 1982), in which experimental removal of the ulnar diaphysis stimulated adaptive modeling of the radius to compensate for loss of the ulna in each species. Initially elevated strains at the radial diaphysis returned to normal levels within a 3-month period in the juvenile pigs and within a 1-year period in the mature sheep. A recent study of the tarsometatarsus in skeletally mature roosters (Loitz and Zernicke, 1992), however, suggests that adaptation of mature bone may not be as sensitive to exercise-induced changes in peak strain as that observed in younger, growing bone. Changes in bone mass and shape during growth may be more tightly linked to genetic/epigenetic factors that regulate skeletal differentiation and growth. Accordingly, adjustments to extrinsic loading patterns would be expected to be more limited in growing versus mature bone. To examine this possibility and to test more rigorously our hypothesis concerning the maintenance of functional strain patterns within a bone, we subjected growing chicks to intensive treadmill exercise from 2 to 12 weeks of age. We

Skeletal strain in relation to exercise during growth

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achieved this both by increasing the speed of exercise and by having the animals carry loads proportional to their weight on their backs. While increases in running speed and body weight (1.2 times in the present case) each contribute to an overall increase in the magnitude of load that a bone must support, neither is likely to alter significantly the general distribution of strain within a bone (Rubin and Lanyon, 1982; Biewener et al. 1983; Biewener and Taylor, 1986). Consequently, the increase in exercise intensity achieved using this approach was designed to increase strain magnitude without disrupting the normal distribution of strain in the bone. We reasoned that elevated strain levels produced by the increase in exercise intensity would elicit an adaptive modeling response, beyond that of normal bone growth, to return strain levels near to those recorded previously in animals that had experienced only moderate treadmill exercise while growing. A non-invasive exercise approach such as this has the advantage of subjecting the bone to an elevated, but physiologically normal, strain pattern. Measurements of in vivo strain on the growing chick tibiotarsus during heavy exercise are reported here and compared with measurements of bone strain made previously in the moderate exercise group at the same functional sites. Changes in bone cortical geometry and shape elicited by the exercise regimens are reported elsewhere (Biewener and Bertram, 1993).

Materials and methods The training and experimental protocols, the surgical methods and bone strain recording procedures used in the present experiments and described below are similar to those used in Biewener et al. (1986). Because growth slows considerably from 12 to 17 weeks of age, we limited our comparison of in vivo strain recordings (and associated modeling responses of the bone) to chicks aged 4–12 weeks. Animals and exercise training Fifteen male white leghorn chicks were divided randomly into three exercise groups (five animals per group) and exercised at 60% of their maximum running speed, carrying an additional 20% of their body weight (60%/L) in weights distributed by a cloth cummerbund about their trunk. By exercising the animals at a percentage of their maximum speed, the animals maintained a uniform exercise level as they grew to a larger size. At all ages, exercise speed (0.90 ms 21 at 4 weeks; 1.29ms 21 at 8 weeks; 1.54ms 21 at 12 weeks) corresponded to a constant stride frequency of 2.96±0.13strides s21 (N=15). Load-carrying was used to increase further the magnitude of bone loading. The weights that the animals carried were adjusted every few days to account for increases in body weight as the animals grew. The weight-packs were designed to fit over the cloth harness that the animals wore later during the recordings of in vivo strain. The animals were trained from 1 to 3 weeks of age, with regular exercise (5 days per week) being maintained at 60%/L thereafter. The animals generally ran well with the additional load carried on their backs by 2 weeks of age. Postural differences associated with carrying the load were observed in some animals, which tended to increase the variability in the pattern of strains recorded among individual animals compared with that

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observed earlier in the moderate exercise, non-load-carrying group (35%/UNL; Biewener et al. 1986). As in our earlier study, exercise duration was set at 15min corresponding to about 2500 loading cycles per day at the faster speed. At all other times, the animals’ activity was restricted to sedentary weight support by housing them in confined conditions in 0.5 m30.5m cages (e.g. three animals weighing 1.0kg each were housed per cage at 12 weeks of age). Surgery and in vivo bone strain recordings Each group of five animals was exercised to 4, 8 or 12 weeks of age. At each of these ages, three animals (the other animals were used for structural and histological analyses of bone modeling changes; Biewener and Bertram, 1993) underwent aseptic surgery to attach strain gauges on the proximal-medial (PM), medial-midshaft (MM), cranialmidshaft (CRM) and caudal-midshaft (CAM) cortices of the left tibiotarsus and on the cranial-distal (CRD) and caudal-distal (CAD) cortices of the right tibiotarsus (Fig. 1). Only one gauge could be attached medially at the bone’s proximal level because of attachments of anterior and posterior muscles and the fibula laterally. All bone sites were surgically exposed from the medial side of the limb. At each site, the overlying musculature was retracted away from the bone and a small patch (6mm36 mm) of the periosteum was cut out and removed using a scalpel. The underlying mineralized surface of the bone was then lightly scraped with a small periosteal elevator, defatted and dried by applying ether to the bone surface using a cotton-tipped applicator. After ensuring a dry,

Fig. 1. Medial and caudal views of the chick tibiotarsus, showing the strain gauge recording sites. Proximal, midshaft and distal levels were determined as a percentage of the bone’s overall length. (PM, proximal-medial; MM, midshaft-medial; CRM, cranial-midshaft; CAM, caudal-midshaft; CRD, cranial-distal; CAD, caudal-distal).

Skeletal strain in relation to exercise during growth

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clean surface, the strain gauge was bonded with a self-catalyzing cyanoacrylate adhesive with pressure applied to the gauge for 90s. The strain recording sites were defined with respect to a fixed percentage of the bone’s length (proximal 30%, midshaft 50%, distal 70%) and were considered to be functionally equivalent locations on the bone, consistent with the constant linear relationship of muscle and ligamentous attachment sites to a bone during its growth (Grant et al. 1980). The lead wires from the strain gauges (36 gauge, etched Teflon-insulated: Micromeasurements) were sutured to surrounding fascia for strain relief and passed subcutaneously to an opening in the animal’s skin situated over its back, between the wings. The lead wires were then soldered to a multipin plastic connector (Amphenol, 222 series) and all wounds were sutured close. The connector plug was securely wrapped with an elastic bandage (Vet-Wrap) and held in place within a cloth cummerbund wrapped over the animal’s back and under its wings. Because of the small size of the bone at 4 weeks of age, strain recordings were made using single-element strain gauges (type FLE-05, Tokyo Sokki Kenkyujo). These strain recordings were made with the gauges aligned along the bone’s longitudinal axis. At 8 and 12 weeks of age, strain recordings were made using rectangular rosette strain gauges (type FRA-1) at three of the bone sites in each animal. Single-element gauges were used at the other three sites. Sites of rosette strain recordings were alternated to determine the orientation and magnitude of principal strains in four of the six sites overall (excluding the MM and CAD sites). The longitudinal strains recorded at the proximal-medial (PM), cranial-midshaft (CRM), caudal-midshaft (CAM) and cranial-distal (CRD) sites were used to estimate the maximum (absolute magnitude) principal strains at these sites at 4 weeks of age, based on the axis of maximum principal strain determined from rosette strain gauge recordings made at 8 and 12 weeks of age. This procedure assumes a uniaxial planar state of strain at the bone’s surface to construct a Mohr’s circle of strain at each site (Dally and Riley, 1978) based on the magnitude of shear strain determined from the rosette strain recordings at the later ages. The principal strains are then computed on the basis of the magnitude of the longitudinal strain recorded from the single-element gauge made at 4 weeks of age and its angle to the assumed principal strain axes. Estimates of maximum principal strain determined from longitudinal strains measured at 4 weeks of age averaged +18% at the PM site (corresponding to a mean 22˚ deviation from the principal strain axis, see Table 2), +48% at the CRM site (mean 39˚ deviation), +28% at the CAM site (mean 30˚ deviation) and +23% at the CRD site (mean 26˚ deviation). Because this procedure oversimplifies the true state of strain by assuming a uniaxial planar state of strain at the bone’s surface, interpretations based on comparisons of corrected single-element strain recordings at 4 weeks of age should be treated with some caution. By recording bone strain from both hindlimb bones, equal weight-bearing between the two limbs is favored. In addition, we observed little sign of lameness resulting from the surgery. The animals were allowed 24h to recover from surgery, before in vivo strains were recorded on each of the following 2 days. In vivo bone strains were digitally sampled at 200Hz and entered into a microcomputer for calculation of the principal strains and their orientation. Peak strain levels were referenced to a zero level established

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during the swing phase of the limb. These levels were consistent with those recorded while the animal was resting with its limb held in a relaxed state. Following the bone strain recordings, the animals were immediately killed and their bones removed for structural analyses of bone modeling changes (Biewener and Bertram, 1993). Least-squares regressions of the data (based on mean maximum strain levels within each individual animal at each site) were carried out to test for significant shifts in strain magnitude versus age at each site. Additional comparisons of strain magnitude and orientation between groups were performed using Student’s t-tests.

Results Strain pattern versus growth Strains recorded at each of the six sites when the animals ran at 60% of their maximum speed, carrying an additional 20% body weight load, increased slightly with age (Fig. 2). At no site did the magnitude of strain decrease at 8 or 12 weeks compared with levels measured at 4 weeks of age. Only at the CAM site was the increase in strain with age significant (Table 1). At two sites (MM and CRD), no change in strain magnitude was evident. At all sites, the nature (tensile versus compressive) of strain remained unchanged. Consequently, the overall pattern of locomotor strains recorded at the six sites changed very little from 4 to 12 weeks of age. Though more variable, the distribution of strain recorded in the intensive, load-carrying exercise animals was similar to that observed previously in animals exercised at 35% of their maximum speed, unloaded (35%/UNL; Biewener et al. 1986). In addition to the uniform distribution of strain magnitude and sign, the orientation of principal strains recorded at three of four (rosette strain gauge) sites also remained similar among animals at 8 and 12 weeks of age (Table 2). Although the orientation of principal strain differed significantly at the proximal-medial (PM) site, rosette strain recordings were obtained from only one animal at each of these ages, rendering this finding of questionable significance. In no case did carrying a load substantially alter the orientation of peak principal strain compared with when the same animals ran unloaded (Table 2). Table 1. Least-squares regression statistics for maximum principal strain versus age (weeks) at six recording sites on the tibiotarsus Site

N

Slope (±95% CI)

PM MM CRM CAM CRD CAD

8 8 8 8 8 7

−76.8 (±79.7) −12.3 (±52.6) 31.7 (±52.4) −57.7 (±46.6) −69.8 (±86.0) 10.5 (±41.9)

r 0.653 0.204 0.475 0.742 0.587 0.218

S.E. of

slope

33.7 22.2 22.1 19.7 36.4 17.7

P 0.069 0.759 0.122 0.031 0.126 0.673

For all but the CAM site, the slope of the regression is not significantly different from zero, indicating no age-related change in the magnitude or nature of strain at each of the other sites. CI, confidence interval.

Skeletal strain in relation to exercise during growth

Fig. 2. Peak strains recorded on the chick tibiotarsus when the animals ran at 60% of their maximum speed, carrying 20% of their body mass (60%/L) at 4, 8 and 12 weeks of age (A, proximal-medial, PM, and medial midshaft, MM, sites; B, cranial, CRM, and caudal, CAM, midshaft sites; C, cranial, CRD, and caudal, CAD, distal sites). The dashed and solid lines represent the least-squares regressions of peak strain versus age for each pair of sites. Error bars denote ± 1 S.D. based on a series of strides (N>12) at the exercise speed (see Table 1 for regression statistics).

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Table 2. Orientation (u) of maximum principal strain versus age at those sites for which rosette strain recordings were made during loaded versus unloaded exercise at 60% maximum speed (positive denotes a proximo-medial orientation for the CRM and CRD sites, proximo-lateral for the CAM site and proximo-caudal for the PM site) u (degrees) Age 8 weeks Unloaded Loaded 12 weeks Unloaded Loaded

PM

CRM

CAM

CAD

+30±5 +33±7 (1, 11)

−30±13 −33±15 (2, 19)

+25±3 +24±5 (2, 19)

−28±15 −30±16 (3, 28)

+9±3 +10±3 (1, 8)

−40+3 −46±4 (2, 17)

+33±5 +35±7 (2, 17)

−23±10 −21±9 (3, 26)

Values represent the mean ± S.D. The number of individuals and total number of strides analyzed are given in parentheses.

The orientation of principal strain determined at each site was also generally consistent with that observed previously in the moderate exercise group (Biewener et al. 1986). Angles of principal strain relative to the bone’s longitudinal axis at the midshaft (CRM 246˚ and CAM +35˚ at 12 weeks) indicate a significant component of torsional loading during the support phase of the stride. Load-carrying and strain magnitude Carrying a load equal to 20% of their body weight resulted in an overall 15% increase in strain magnitude at 8 and 12 weeks of age, compared with when the animals ran unloaded at the same speed (Table 3). The increase was variable among sites, however, ranging at 8 weeks from as high as +32% at the CRD site to as low as +5% at the MM site. Because strain levels recorded at the CAD site were low under all exercise conditions, the potential for error associated with establishing a zero strain level (when balancing the bridge circuits, see Materials and methods) relative to the peak strain recorded is high. Consequently, this site was excluded from comparisons between the two exercise groups and loaded versus unloaded exercise conditions. (The possibility that the low compressive strains recorded at the CAD site were due to gauge loosening was excluded by direct post-mortem loading of the bone to produce artificially high strains at this site.) Bone strain patterns versus speed Although the pooled data show no significant differences in strain magnitude associated with load-carrying at the exercise speed, differences in strain magnitude were generally significant when compared at a given speed within a given animal (Fig. 3). As a result, the elevation of strain magnitude associated with load-carrying at most sites was consistent over each animal’s measured range of running speed (Fig. 3). At no site

Skeletal strain in relation to exercise during growth

Fig. 3. Peak strains recorded at each site on the tibiotarsus as a function of speed in one individual at (A) 8 weeks of age and another individual at (B) 12 weeks of age. Filled symbols denote strains recorded when the animals ran carrying a load (20% body mass); open symbols correspond to strains recorded when the animals ran unloaded. Arrows designate the daily exercise speed (60% maximum) that was used in the present experiments at 8 and at 12 weeks of age. Error bars denote ± 1 S.D. for a series of strides (N>12) at each speed.

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Table 3. Peak principal strains (me or × 106) recorded when the animals ran unloaded versus loaded at 60% of their maximum running speed Bone recording sites Age

PM

MM

CRM

CAM

CRD

CAD

8 weeks 60%/unloaded 60%/loaded Ratio (mean)

−654±342 −763±367 1.17 (1.16)†

−838±174 −877±196 1.05

+511±87 +589±153 1.15

−954±230 −1044±261 1.09

−1206±294 −1593±301 1.32

−204±53 −128±83 0.63

12 weeks 60%/unloaded 60%/loaded Ratio (mean)

−1113±228 −1245±230 1.12 (1.13)†

−860±273 −926±303 1.08

+884±116 +987±250 1.12

−1251±166 −1430±168 1.14

−1217±279 −1442±298 1.18

−181±38 −203±38 1.12

†Values for the CAD site were excluded in calculating an overall mean strain ratio, as the potential for error in establishing the percentage change in strain at this site is high owing to the error inherent in establishing a zero strain level during the limb cycle relative to the low strains recorded at this site.

(excluding the CAD site) did the tensile or compressive nature of strain change as a result of carrying a load. Overall, the pattern of locomotor strain recorded at the six sites on the tibiotarsus remained remarkably consistent as the animals increased speed, both when they were unloaded and when they carried a load. Strain magnitude versus age and exercise level At 4 weeks of age, peak strains recorded at the intensive exercise level (60%/L) were initially 57±14% greater than those recorded when the same three animals ran at the moderate exercise level (35%/UNL) (Fig. 4A and Table 4). Except for the CAD site, the Table 4. Strains (me or × 106) recorded at six sites on the tibiotarsus of three chicks at 4 weeks of age while exercising at the moderate (35%/UNL) and intensive load-carrying (60%/L) exercise conditions Bone sites

35%/UNL 60%/L Ratio

PM

MM

CRM

CAM

CRD

CAD

−410±201 −606±248

−522±69 −931±136

+465±95 +685±152

−612±64 −895±134

−472±52 −774±96

−142±97 −148±42

1.48

1.78

1.47

1.46

1.64

1.04†

Mean

1.57

Values are the pooled means ± S.D. (total number of strides > 24). The ratio indicates the initial strain differential established at each site by the load-carrying exercise regimen compared with the moderate exercise regimen. †This site was excluded in our calculation of a mean strain ratio and in comparisons among groups and exercise levels.

Skeletal strain in relation to exercise during growth

Fig. 4. Histograms of peak strain recorded at (A) 4, (B) 8 and (C) 12 weeks of age for animals running at 35% maximum speed/unloaded (hatched columns) versus 60% maximum speed/loaded (filled columns). The ratio of strain between 35%/UNL versus 60%/L conditions is indicated in each case. The comparison at 4 weeks of age is based on the same three individuals recorded when they ran at the 60%/L versus 35%/UNL exercise level. At 8 and 12 weeks of age, peak strains recorded from the 60%/L exercise animals (three animals at each age) are compared with those reported previously at the same sites for animals subjected to the moderate (35%/UNL) exercise training (Biewener et al. 1986). Error bars are ± 1 S.D. Asterisks denote significantly different strain levels at P