It has been reported that Ca deficiency reduces bone mineral density [18, 19] and bone strength [20] in growing rats. The results of the present study are consistent with these previous reports. On the other hand, in mature male and female rats, the Ca-deficient diet decreased the femoral weight and width of the central axis and reduced bone strength, as assessed by the rupture energy and stiffness. These results suggest that Ca deficiency weakens bones during the growing stage, but not during the mature stage.
Bone mineral density, bone rupture energy, and stiffness are believed to be correlated. The ash weights of the femur and whole body were reduced by the Ca-deficient diet in both growing and mature rats. However, during the growth stage, the weight of ash in the deficient diet group was approximately 50% of that in the sufficient diet group, whereas in the mature stage, it was approximately 90% or more. Therefore, during the mature stage, the decrease in bone Ca due to the Ca-deficient diet was not large; therefore, the rupture energy and stiffness did not decrease.
The reason for the lack of a large decrease in bone Ca due to the Ca-deficient diet in mature rats is that, in growing rats, Ca was deficient during the active period of bone formation. In contrast, in mature rats, Ca had accumulated in the bones by 12 weeks of age; therefore, the Ca-deficient diet did not cause a significant decrease in bone Ca.
Saville et al. reported that applying a weight load to the hindlimb increased the bone Ca concentration in the hindlimb of rats [20]. Welten et al. showed that both Ca intake and exercise load contribute to maximum bone mass acquisition [21]. Lovelady et al. reported that exercise during breastfeeding reduced bone density loss without increasing Ca intake [10]. Frost suggested that bones adapt to loads, such as body weight and muscle forces, and increase their mineral content and strength [22]. Welch et al. stated that mechanical stimulation caused by exercise increases bone mass, and that mechanical stimulation above a certain threshold (minimum effective strain) is necessary during the bone mass acquisition phase [23].
In the present study, no effects of exercise were observed in the femur, except for a lower rupture energy in growing males and increased stiffness in mature females. Dalsky et al. observed that the endocrine system is involved in the effect of exercise load on bone metabolism [24]. Estrogen has the function of transmitting the stimulation caused by exercise to osteoblasts, and it has been suggested that when ovarian function declines, even with exercise, sufficient effects on the bones cannot be obtained [24]. The effect of exercise on stiffness observed only in adult female rats in the present study could be attributed to an estrogen-mediated effect.
The exercise protocol used in this study has been reported to increase FHL in growing male rats [16]. In this study, an increase in the skeletal muscle weight due to exercise was observed in the FHL and the FHL per 100 g of body weight and gastrocnemius muscle in growing males, whereas it was only observed in the FHL per 100 g of body weight in growing females. Therefore, muscle hypertrophy due to exercise was thus suggested to be greater in males than in females. On the other hand, in mature rats, no increase in skeletal muscle weight due to exercise was observed in rats of either sex. In the exercise regimen employed in this study, the heavier the weight, the higher the load on the body. However, even in males whose muscle weight increased with exercise during the growth stage, no increase in muscle weight due to exercise was observed in the mature stage. These results suggest that exercise-induced muscle hypertrophy is less likely to occur after maturation.
Okano et al. [7] suggested that exercise-induced muscle hypertrophy may be partially responsible for the increase in bone mineral density and content observed in rats subjected to squat training. Fujii et al. [16] reported that the exercise used in the present study increased muscle mass, increased aminolevulinic acid dehydratase activity (the rate-limiting enzyme in hemoglobin synthesis), and mitigated anemia in rats fed an iron-deficient diet. In the present study, we assumed that resistance exercise promoted the synthesis of bone collagen, which is synthesized in the body in the same way as hemoglobin, resulting in the accumulation of Ca in the bones and a reduction in mineral content and bone fragility in rats fed a Ca-deficient diet. In the present study, the collagen content of the bones was not measured to prioritize their ash weight. However, the difference between the weight of the femur shown in Table 4 and the weight of the ash of the femur shown in Table 5 can be considered to represent the weight of the organic matter in the femur. The main components of organic matter are collagen. Thus, this can be an indicator of collagen weight. No effect of exercise was observed when weight was calculated. Therefore, bone collagen was unlikely to increase with exercise in the present study.
In this study, we investigated the effects of a Ca-deficient diet and resistance exercise on rat bones in growing and mature male and female rats. As a result, the bones of both male and female rats were weakened by a Ca-deficient diet in growing rats but not in mature rats. Furthermore, no obvious effects of exercise on bones were observed in male or female rats, in either growing or mature animals. A possible reason why exercise did not reduce bone fragility due to the Ca-deficient diet in growing rats could be that the Ca level in the diet was too low or that the effect of exercise was insufficient.
In conclusion, bone weakness caused by a Ca-deficient diet was observed in growing rats; however, exercise did not alleviate this weakness.