Sequential treatment with zoledronic acid followed by teriparatide or vice versa increases bone mineral density and bone strength in ovariectomized rats

Abstract

Bisphosphonates (BPs) and teriparatide (TPTD) are both effective treatments for osteoporosis, but BP treatment prior to daily TPTD treatment has been shown to impair the effect of TPTD in some clinical studies. In contrast, the loss of bone mineral density (BMD) that occurs after withdrawal of TPTD can be prevented by BP treatment. Although various studies have investigated the combination and/or sequential use of BP and TPTD, there have been no clinical studies investigating sequential treatment with zoledronic acid (ZOL) and TPTD (or vice versa). In this study, we evaluated the effects of sequential treatment with TPTD followed by ZOL, and ZOL followed by TPTD, using ovariectomized (OVX) rats. Two months after OVX, osteopenic rats were treated with ZOL, TPTD, or vehicle for a period of 4 months (first treatment period), and then the treatments were switched and administered for another 4 months (second treatment period). The group treated with ZOL followed by TPTD showed an immediate increase in BMD of the proximal tibia and greater BMD and bone strength of the lumbar vertebral body, femoral diaphysis, and proximal femur than the group treated with ZOL followed by vehicle. Serum osteocalcin, a marker of bone formation, increased rapidly after switching to TPTD from ZOL. The group treated with TPTD followed by ZOL did not lose BMD in the proximal tibia after TPTD was stopped, while the group treated with TPTD followed by vehicle did lose BMD. The BMD and bone strength of the lumbar vertebral body, femoral diaphysis, and proximal femur were greater in the group treated with TPTD followed by ZOL than in the group treated with TPTD followed by vehicle. The increase in serum osteocalcin and urinary CTX after withdrawal of TPTD was prevented by the switch from TPTD to ZOL. In conclusion, our results demonstrate that switching from ZOL to TPTD resulted in a non-attenuated anabolic response in the lumbar spine and femur of OVX rats. In addition, switching from TPTD to ZOL caused BMD to be maintained or further increased. If these results can be reproduced in a clinical setting, the sequential use of ZOL followed by TPTD or vice versa in the treatment of osteoporosis patients would contribute to increases in BMD that, hopefully, would translate into a corresponding decrease in the incidence of vertebral and non-vertebral fractures.

Keywords: Bone metabolism; Bone mineral density; Ovariectomized rat; Sequential treatment; Teriparatide; Zoledronic acid.

Fig. 1

Experimental groups. ZOL to TPTD set: the effects of Z-T sequential treatment were compared with those of the Z-V or V-T therapy. TPTD to ZOL set: the effects of T-Z sequential treatment were compared with those of the T-V or V-Z therapy. All treatments were started (at month 0) 2 months after the OVX operation (at month − 2). 100 μg/kg ZOL was administered as a single intravenous injection. TPTD was administered subcutaneously at 6 μg/kg three times weekly. After 4 months of treatment with one of the drugs or vehicle, the animals started treatment with the other drug or vehicle. Animals in the satellite groups for each treatment (groups B through T4) were sacrificed at months − 2, 0, and 4 to harvest bone tissues. All groups and the treatments they underwent are shown in the chart at the bottom of the figure. The sample size was 20 (n = 20) at the start for the Sham through T-Z groups; the sample sizes provided in the table reflect reductions during the study because of euthanasia due to aging-related symptoms.

Fig. 2

Bone mineral density of the ZOL to TPTD set. (a) BMD at the proximal tibia over time measured by in vivo DXA. (b) Percent changes in BMD at the proximal tibia over time with the percent change at month 0 set as 0%. The sham-operated group has been omitted from the graphs showing percent changes in BMD. BMD at the (c) 4th lumbar vertebral body, (d) femoral diaphysis, and (e) proximal femur at the study conclusion (month 8). See Table 1 for the satellite group results from before the OVX operation (group B), immediately before the first treatment period (groups S0 and V0), and immediately before the second treatment period (groups S4, V4, Z4, and T4). *: p < 0.05, **: p < 0.01, significant difference between the Z-T group and Z-V group during each experimental period (Student's t-test). a: p < 0.05, significant difference between Sham vs V-V (Student's t-test). b: p < 0.05, significant difference vs V-V (Dunnett-test). Data are presented as means ± S.D.

Fig. 3

Mechanical properties of the bones resected at the study conclusion from the ZOL to TPTD set. (a) maximum compression test loads withstood by the L4 vertebral body, (b) maximum three-point bending test loads withstood by the femoral diaphysis, and (c) maximum mechanical test loads withstood by the proximal femur. See Table 1 for the satellite group results before the OVX operation (group B), immediately before the first treatment period (groups S0 and V0), and immediately before the second treatment period (groups S4, V4, Z4, and T4).**: p < 0.01, significant difference between the Z-T group and Z-V group (Student's t-test). a: p < 0.05, significant difference between Sham vs V-V (Student's t-test). b: p < 0.05, significant difference vs V-V (Dunnett-test). Data are presented as means ± S.D.

Fig. 4

Change in markers of bone metabolism for the ZOL to TPTD set. (a) serum osteocalcin concentration, (b) urinary CTX concentration corrected for creatinine. **: p < 0.01, significant difference between the Z-T group and Z-V group during each experimental period (Student's t-test). Data are presented as means ± S.D.

Fig. 5

Results of histomorphometry of the L5 vertebral body for the ZOL to TPTD set. (a) bone volume (BV/TV), (b) trabecular thickness (Tb.Th), (c) osteoblast surface (Ob.S/BS), (d) mineralizing surface (MS/BS), (e) osteoclast surface (Oc.S/BS), (f) eroded surface (ES/BS), (g) representative fluorescent images of each group.**: p < 0.01, significant difference between the Z-T group and Z-V group (Student's t-test). a: p < 0.05, significant difference between Sham vs V-V (Student's t-test), b: p < 0.05, significant difference vs V-V (Dunnett-test). Data are presented as means ± S.D. Scale bar = 200 μm

Fig. 6

Bone mineral density of the TPTD to ZOL set. (a) BMD values at the proximal tibia over time measured by in vivo DXA. (b) Percent change in BMD at the proximal tibia over time, with the percent change at month 0 set as 0%. The sham-operated group has been omitted from the graphs showing percent changes in BMD. BMD at the (c) L4 vertebral body, (d) femoral diaphysis, and (e) proximal femur at the study conclusion (month 8). See Table 1 for the satellite group results from before the OVX operation (group B), immediately before the first treatment period (groups S0 and V0), and immediately before the second treatment period (groups S4, V4, Z4, and T4). ^††: p < 0.01, significant difference between the T-Z group and T-V group during each experimental period (Student's t-test). a: p < 0.05, significant difference between Sham vs V-V (Student's t-test). b: p < 0.05, significant difference vs V-V (Dunnett-test). Data are presented as means ± S.D.

Fig. 7

Mechanical properties of the bones resected at the study conclusion from the TPTD to ZOL set. (a) maximum compression test loads withstood by the L4 vertebral body, (b) maximum three-point bending test loads withstood by the femoral diaphysis, and (c) maximum mechanical testing loads withstood by the proximal femur. See Table 1 for the satellite groups results before the OVX operation (group B), immediately before the first treatment period (groups S0 and V0), and immediately before the second treatment period (groups S4, V4, Z4, and T4). ^†: p < 0.05, ^††: p < 0.01, significant difference between the T-Z group and T-V group during each experimental period (Student's t-test). a: p < 0.05, significant difference between Sham vs V—V (Student's t-test). b: p < 0.05, significant difference vs V-V (Dunnett-test). Data are presented as means ± S.D.

Fig. 8

Changes in markers of bone metabolism for the TPTD to ZOL set. (a) serum osteocalcin concentration, (b) urinary CTX concentration corrected for creatinine.^††: p < 0.01, significant difference between the T-Z group and T-V group during each experimental period (Student's t-test). Data are presented as means ± S.D.

Fig. 9

Results of histomorphometry of the L5 vertebral body for the TPTD to ZOL set. (a) bone volume (BV/TV), (b) trabecular thickness (Tb.Th), (c) osteoblast surface (Ob.S/BS), (d) mineralizing surface (MS/BS), (e) osteoclast surface (Oc.S/BS), (f) eroded surface (ES/BS), and (g) representative fluorescent images of each group. ^†: p < 0.05, ^††: p < 0.01, significant difference between the T-Z group and T-V group during each experimental period (Student's t-test). a: p < 0.05, significant difference between Sham vs V-V (Student's t-test). b: p < 0.05, significant difference vs V-V (Dunnett-test). Data are presented as means ± S.D. Scale bar = 200 μm