Search

To further characterize the skeletal role of vitamin K (K), markers of bone turnover, density, and strength were evaluated in rats with diet- or warfarin (W)-induced K insufficiency. One hundred two, 7-week-old, female rats were randomly assigned to low K (phylloquinone [K1], 20 microg/kg diet), control K (K1, 1300 microg/kg diet), low-dose W (W, 1.5 mg/kg control diet), or high-dose W plus K (W/K1, 10/100 mg/kg diet). Femur bone mineral content (BMC) and bone mineral density (BMD), plasma prothrombin time (PT) and prothrombin concentration (PC), and serum total alkaline phosphatase (ALP) and skeletal alkaline phosphatase (sALP) were measured at baseline and days 20, 40, 60, and 80. Serum total osteocalcin (OC) and undercarboxylated osteocalcin (ucOC) and femur length (FL) were measured at baseline and day 80. Left femur OC was measured and biomechanical testing of the right femur and third lumbar vertebral body was performed at day 80. Low dietary K elevated circulating ucOC (17% higher than control; p < 0.0001) at day 80. Furthermore, in both W groups, essentially all circulating OC was undercarboxylated and femur OC was lower than control (p < 0.0001). However, there was no change in femur percent ucOC, suggesting deposition of less newly synthesized OC. No between group differences were observed in PT, ALP, sALP, FL, BMC, BMD, or bone strength. In conclusion, skeletal K insufficiency can be induced by W or diet manipulation. This does not hinder peak bone mass attainment in female rats; however, W causes less newly synthesized OC to be deposited in bone.

This study was undertaken to find whether the bone response to increased external loading (EL) of the tibia in rats is affected by estrogen depletion. Female Sprague-Dawley rats 6 months were randomly assigned to four groups of 10 each: sham ovariectomy without loading (Shm-XL), ovariectomy without loading (OVX-XL), sham ovariectomy with external loading (Shm-EL), and ovariectomy with external loading (OVX-EL). In vivo external loading by a four-point bending device was initiated 4 weeks after surgery. The right lower leg of each EL rat was loaded at 31.4 +/- 0.2 N for 36 cycles at 2 Hz every other day for 21 days (11 loading days). Mean in vivo induced strain was 1305 microstrain (mu epsilon) for Shm-EL rats and 1280 mu epsilon for OVX-EL rats. With external loading of the tibia, periosteal bone formation rose equally in Shm and OVX rats. Woven bone was present around the tibia or fibula in 60% of the loaded rats and none of the control rats. No loading response occurred either at the endocortical surface or in the cancellous bone of the proximal tibial metaphysis. After OVX, cancellous bone area in the proximal metaphysis declined and formation surface rose compared to Shm rats. Although periosteal and endocortical bone formation rose after OVX, no cortical bone loss occurred. We conclude that ovariectomy and attendant loss of endogenous estrogen do not change the cortical bone response to an external load of about 1300 mu epsilon in rats. However, these results may not predict the cortical bone response to loading in animals with Haversian remodeling that display estrogen-related loss of cortical bone.

The rat tibia four-point bending model is a new mechanical loading model in which force is applied through external pads to the rat lower limb. The advantages of the model are controlled force application to a well-defined bone, noninvasive external loading, and the addition of loads to normal daily activity. A disadvantage of the model is that the pads create local pressure on the leg at the contact sites. This study examined the differences in tibial response to bending strains and to local pressure under the pads. A total of 30 adult Sprague-Dawley rats were randomized into three external loading groups: bending, cyclic pressure, and static pressure. The right leg of each rat was externally loaded to create either bending or local pressure without bending; the left leg served as a control. Strains on the lateral surface averaged 1200 mu epsilon in compression during bending load application and < 200 mu epsilon in compression during pressure loading. Histomorphometric data were collected from three regions: the maximal bending region, under the loading pads, and outside the maximal bending region. In the maximal bending region, bending loads created greater mineral apposition rate (MAR) on the lateral surface and greater MAR and formation surface on the medial surface of loaded than control tibiae. The region under the bending pad was exposed to similar bending strains and showed the same pattern of increased MAR as sections from the maximal bending region. Cyclic pressure had no effect on periosteal MAR or formation surface. Static pressure increased MAR only on the lateral tibial surface. Bending stimulates bone formation in regions with the highest bending strains. Similar forces applied only in the form of pressure loading do not stimulate tibial formation either at the contact site or between loading pads. These results suggest that externally applied forces of moderate magnitude stimulate bone formation primarily as a result of increased bending strains, not local pressure at the contact site.

Mechanical loading of the living skeleton influences bone formation, mass, and strength. The primary purpose of the present study was to examine the influence of different loading schedules (days/week) on the bone response to external loading using an in vivo rat tibia four-point bending model. Three studies were conducted to (1) characterize the loaded region, (2) examine the variation of the response within the loaded region, and (3) test the response to different loading schedules. In all studies adult female retired breeder Sprague-Dawley rats were used (6 months, 285 g). First, the location of the loaded region during four-point bending was determined by radiogrammetry of 7 rats. Second, 5 rats were externally loaded for 8 of 10 days at 31 N, 36 cycles, and 2 Hz (1349 ± 244 μϵ). The extent of labeled (forming) periosteal and endocortical surface in the loaded region was compared both among four serial sections from the same tibia and between the loaded and the contralateral tibiae. Finally, 50 rats were randomized into five groups: two nonloaded, control and sham, and three loaded, alternate day, Monday, Wednesday, and Friday, and daily. The rats were externally loaded for 3 weeks at 35 N, 36 cycles, and 2 Hz (1533 ± 308 μϵ). The tibia and fibula were studied for labeled surfaces and mineral apposition rate. For adult female rats with tibial length 39 mm, the loaded region was located 3.5–14 (±0.7) mm proximal to the tibia-fibula junction (TFJ). With multiple repositionings in the same rat, the location of the distal inner pad varied from 3.2 to 3.8 mm proximal to the TFJ (95% confidence interval). The response to loading was consistent within the loaded region and reliably sampled by two cross sections from the loaded region. The bone response under the loading pads was not different from that at other points within the loaded region. There were no differences in the response to external mechanical loading among the three loading schedules (daily, alternate day, and Monday-Wednesday-Friday). Loading created greater formation surface on the medial periosteum of the tibia (70%) and on the fibula (92%) of externally loaded legs than in nonloaded legs. Loading had no effect on the lateral periosteal formation surface or on the endocortical surface of the tibia. Loading created greater mineral apposition rate in loaded than nonloaded tibia (28–72% greater) at all periosteal surfaces. The pattern of response to loading was the same in both studies 2 and 3. In an adult rat, this model produces a predictable bone response to external mechanical loading in a defined region of the tibial diaphysis. External loading 3 or 4 days/week is as effective as daily loading for increasing periosteal bone formation surface and mineral apposition rate.