Yongping Cao

Raloxifene, estrogen, and alendronate affect the processes of fracture repair differently in ovariectomized rats.

We investigated the effects of inhibitors of bone resorption (estrogen, raloxifene, and alendronate) on the processes of fracture repair in ovariectomized (OVX) rats. One hundred forty female Sprague‐Dawley rats at 3 months of age were either OVX or sham‐operated and divided into five groups: sham control, OVX control, estrogen (17α‐ethynyl estradiol [EE2], 0.1 mg/kg), raloxifene (Rlx, 1.0 mg/kg), and alendronate (Aln, 0.01 mg/kg) groups. Treatment began immediately after the surgery. Four weeks postovariectomy, prefracture controls were killed and bilateral osteotomies were performed on the femoral midshafts and fixed with intramedullary wires. Treatment was continued and fracture calluses were excised at 6 weeks and 16 weeks postfracture for evaluation by X‐ray radiography, quantitative computed tomography (QCT,) biomechanical testing, and histomorphometry. At 6 weeks postfracture, Aln and OVX had larger calluses than other groups. Sham and OVX had higher ultimate load than EE2 and Rlx, with Aln not different from either control. Aln calluses also contained more mineral (bone mineral content [BMC]) than all other groups. By 16 weeks postfracture, OVX calluses were smaller than at 6 weeks and the dimensions for Aln had not changed. Aln had higher BMC and ultimate load than OVX, EE2, and Rlx. EE2 and Rlx had similar biomechanical properties, which were similar to sham. Interestingly, OVX and Aln animals were heavier than other groups at all time points; therefore, ultimate load was normalized by body weight to show no significant differences in strength of the whole callus between groups at either 6 weeks or 16 weeks postfracture. However, Aln strongly suppressed remodeling of the callus, resulting in the highest content of woven bone, persistent visibility of the original fracture line, and lowest content of lamellar bone, compared with other groups. Therefore, the larger Aln callus appeared to be a remarkable, morphological adaptation to secure the fracture with inferior material. In conclusion, OVX‐stimulated bone turnover resulted in the fastest progression of fracture repair that was most delayed with Aln treatment, consistent with marked suppression of bone resorption and formation activity. Estrogen and Rlx had similar effects that were generally similar to sham, indicating that mild suppression of bone turnover with these agents has insignificant effects on the progression of fracture repair.

The effects of suppressed bone remodeling by bisphosphonates on microdamage accumulation and degree of mineralization in the cortical bone of dog rib.

We evaluated the effects of suppressed bone remodeling caused by bisphosphonate on microdamage accumulation and degree of mineralization of bone (DMB) for the dog rib in two independent studies. Study 1: 36 female beagles, 1-2 years old, were treated daily for 1 year with saline vehicle, risedronate at 0.5 mg/kg/day, or alendronate at 1.0 mg/kg/day. Study 2: 29 beagles, 1 year old, were given lactose, or incadronate at 0.3 mg/kg/day or 0.6 mg/kg/day for 3 years. In both studies, the ninth rib was harvested. Intracortical remodeling was significantly suppressed following either 1 year or 3 years of bisphosphonate treatment without impairment of primary mineralization, although the remodeling rate was obviously lower in study 2 than in study 1 because of the aging of animals. Microdamage accumulation was significantly increased following any bisphosphonate treatment in response to the extent of remodeling suppression. One-year treatment with risedronate or alendronate did not significantly affect the mean DMB or osteonal distribution based on DMB. In contrast, mean DMB was significantly increased following 3 years of incadronate treatments, and osteonal distributions based on DMB showed a dose-dependent shift toward the higher values in incadronate-treated animals when compared with controls. Our results demonstrated that DMB was increased following only 3 years but not 1 year of bisphosphonate treatment. This finding suggests that suppressed remodeling induced by long-term bisphosphonate treatment increased DMB by increasing the population of old, highly mineralized osteons; however, the expression of this phenomenon depends on duration of the treatment because the secondary mineralization is a very slow process.

Suppressed Bone Turnover by Long-Term Bisphosphonate Treatment Accumulates Microdamage but Maintains Intrinsic Material Properties in Cortical Bone of Dog Rib†

Effects of long‐term suppression of bone remodeling by bisphosphonate were investigated in cortical bone of dog rib. Although microdamage was accumulated, BMD was increased without increasing cortical bone area. Consequently, the intrinsic material properties were not reduced.

Long-Term Treatment of Incadronate Disodium Accumulates Microdamage but Improves the Trabecular Bone Microarchitecture in Dog Vertebra

This study aimed to investigate the effect of long‐term suppression of bone resorption by bisphosphonate on the microstructure, accumulation of microdamage, and mechanical properties of trabecular bone. Twenty‐nine 1‐year‐old beagles (15 males, 14 females) were divided into three groups. The control group (CNT) was treated daily with vehicle, and the other two groups were treated with incadronate at a dose of 0.3 mg/kg/day (LOW) or 0.6 mg/kg/day (HIGH) orally for 3 years. After death, the second thoracic vertebra was scanned with microcomputed tomography (micro‐CT) and assigned to histomorphometric and microdamage measurements. The fourth lumbar vertebra was mechanically tested by compression. Incadronate concentration in bone was measured in the 11th thoracic vertebra. Micro‐CT analysis demonstrated a platelike trabecular structure and increased concave surface of trabeculae in the thoracic vertebra of incadronate‐treated groups. Three‐year incadronate treatment significantly suppressed trabecular activation rates by 56% in LOW and 67% in HIGH without impairment of mineralization, and increased microdamage accumulation in both incadronate‐treated groups. Trabecular bone volume was significantly increased in both LOW and HIGH groups, and vertebral strength was significantly increased in the HIGH group compared with the CNT group. However, intrinsic material properties such as normalized ultimate stress and normalized toughness were reduced in incadronate‐treated groups. Incadronate concentration in bone was dose‐dependent. This study suggests that long‐term suppression of bone remodeling increases microdamage accumulation, but this is not necessarily associated with vertebral fragility because of compensated increase of bone mass and improved microarchitecture.

Incadronate disodium inhibits joint destruction and periarticular bone loss only in the early phase of rat adjuvant-induced arthritis

Destruction of articular cartilage and subchon-dral bone loss in the affected joints of rat adjuvant arthritis have never been quantified histologically. This study aimed to evaluate the effect of incadronate disodium on joint destruction and periarticular bone loss, using histomorphometric measurements. Seven-week-old female Lewis rats were injected with 0.1 mg of heat-killed Mycobacterium butyricum into the tail base. Immediately after sensitization, vehicle, or incadronate at 10 or 100 µg/kg per day, was administered subcutaneously, three times per week. Hind-paw volume was measured weekly and the animals were killed at 2, 4, 6, and 10 weeks after sensitization. After taking X-rays, decalcified sagittal sections of the ankle joint were prepared and stained with toluidine blue and tartarate-resistant acid phosphatase. Articular cartilage destruction and subchondral bone loss were evaluated histomorphometrically. At 2 weeks after sensitization, no radiographic or histologic changes were observed. However, at 4 weeks, severe articular cartilage destruction and subchondral bone loss were found in the arthritic control group, while these changes were inhibited dose-dependently by incadronate treatment. At 6 and 10 weeks, both the destructive changes and the bone loss had further progressed, and they were not inhibited by incadronate treatment. Incadronate dose-dependently inhibited articular cartilage destruction and subchondral bone loss at 4 weeks after sensitization in this adjuvant arthritis model. However, the suppressive effects of incadronate did not continue until 6 and 10 weeks.

Fracture Callus Under Anti-resorptive Agent Treatment Evaluated by pQCT

Effects of estrogen, raloxifene, and alendronate on fracture healing were evaluated by a peripheral quantitative computed tomography (pQCT) in an osteoporotic fracture rat model. Three-month-old ovariectomized (OVX; except sham-operated controls) Sprague-Dawley rats were pretreated with vehicle (sham and OVX controls), 0.1 mg/kg day−1 estrogen (17α-ethynyl estradiol), 1 mg/kg day−1 raloxifene, or 0.01 mg/kg day−1 alendronate for 4 weeks before fracture induction. At this point, the pre-fracture groups were killed while transverse osteotomy was performed at the midshaft of both femora in the remaining animals and kept for 6 weeks with drug treatment, and then killed 16 weeks after fracture induction. Excised femora and fracture calluses were analyzed by high-resolution pQCT. At 6 weeks after fracture, the alendronate and OVX groups showed larger calluses at a larger cross-sectional moment of inertia (CSMI) than that of other groups. At 6 weeks after fracture, the calluses in OVX rats were significantly smaller than those observed at 6 weeks, whereas the calluses treated with alendronate did not change in size; therefore, calluses in OVX rats without drug treatment remodeled towards the original geometry in the femoral midshaft faster than drug-treated rats, and on the contrary, the fracture calluses in rats treated with alendronate were the slowest. In conclusion, OVX-induced higher bone turnover and resulted in the fastest remodeling of fracture callus, which was, however, delayed under alendronate treatment. Estrogen and raloxifene treatment showed intermediate callus remodeling between OVX and sham.