The Impact of Strontium Ranelate on Metaphyseal Bone Healing in Ovariectomized Rats.

The following questions were addressed: whether therapy with strontium ranelate (SR) should be continued or interrupted if the fractures occur during SR treatment and whether SR could be applied directly after fracture to improve bone healing. Sprague-Dawley rats (3 month old) were ovariectomized (Ovx, n = 48) or left intact (n = 12). After 8 weeks, a bilateral transverse osteotomy of the tibia metaphysis was created in all rats. Ovx rats were divided into four groups: Ovx; SR applied directly after Ovx until osteotomy (prophylaxis, SR pr, 8 weeks); SR applied after osteotomy (therapy, SR th, 5 weeks); SR applied during the whole experiment (pr + th, 13 weeks). SR dosage was 625 mg/kg body weight/day, administered in the feed. Five weeks later, tibiae were analyzed by biomechanical, histological, micro-CT, and gene expression analyses. The SR pr + th treatment increased total bone mineral density (BMD), bone volume fraction, cortical BMD and volume, callus area and density, serum alkaline phosphatase, tartrate-resistant acid phosphatase mRNA, accelerated osteotomy bridging, and callus formation at weeks 2 and 3 of healing and decreased the osteoprotegerin/receptor activator of nuclear factor kB ligand mRNA ratio. SR th enlarged callus area and improved callus formation during the 5th week of healing. SR pr improved cortical BMD preserving bone after SR discontinuation (5-week rest); the bone healing was not affected. SR content in the tibia metaphysis was the highest in SR pr + th group and was not different between SR pr and SR th. SR has a positive effect on osteoporotic bone healing in rat and SR treatment can be continued after the fracture occurs or applied directly after the fracture.

Effect of Urocortin on strength and microarchitecture of osteopenic rat femur.

As yet there is no evidence of the potential antiosteoporotic effect of Urocortin-1 (UCN), a corticotropin releasing factor related peptide, in vivo. In this study, and for the first time, we investigated the effect of UCN in a rat osteopenia model. Sixty female Sprague-Dawley rats were divided into 5 groups: (1) sham-operated, (2) untreated ovariectomized (OVX) rats, (3) and (4) OVX animals treated for 5 weeks with daily subcutaneous low-dose UCN (3 µg/kg of BW) or high-dose UCN (30 µg/kg of BW) 8 weeks after ovariectomy, and (5) OVX rats treated with daily estrogen (0.2 mg/kg of BW p.o) 8 weeks after ovariectomy for 5 weeks (E). After sacrifice, the femurs were reserved for biomechanical, histomorphometric and ash testing. In the biomechanical test, the high-dose UCN rats showed significantly improved mechanical stiffness (341.6 N/mm) compared with the untreated OVX animals (275.9 N/mm). In the histomorphometric evaluation, the high-dose UCN rats demonstrated an improved trabecular microarchitecture especially and significantly at the distal femur (distal femur Tb.Ar = 41.4% and N.Nd/mm(2) = 26.8, proximal femur Tb.Ar = 71.8% and N.Nd/mm(2) = 28.7) compared with untreated OVX rats (distal femur Tb.Ar = 23.3% and N.Nd/mm(2) = 11.7, proximal femur Tb.Ar = 60.2% and N.Nd/mm(2) = 25.2). Our results show that short-term treatment with UCN seems to have a positive effect on the metaphyseal bone structure and strength of the femur in ovariectomized rats.

Do estrogen and alendronate improve metaphyseal fracture healing when applied as osteoporosis prophylaxis?

Osteoporosis is accompanied by predominantly metaphyseal fractures with a delayed and qualitatively reduced healing process. This study addressed the question of whether fracture healing in the context of osteoporosis prophylaxis is improved with estrogen (E) or alendronate (ALN). Thirty-six ovariectomized and 12 sham-operated 12-week-old rats received soy-free (osteoporotic C, sham), E-, or ALN- supplemented diets. After 10 weeks, a metaphyseal tibia osteotomy and standardized T-plate fixation were performed. After a 5-week healing process, the fracture callus was evaluated qualitatively by biomechanical bending test and quantitatively in microradiographic sections. The time course of callus formation was examined using fluorochrome-labeled histological sections. Administration of E improved the biomechanical properties of callus (stiffness [N/mm]: sham: 110.2 + or - 76.07, C: 41.28 + or - 33.70, E: 85.72 + or - 47.24, ALN: 72.07 + or - 34.68). The resistance to microfracturing seen in E-treated animals was significantly enhanced and even superior to sham (yield load [N] sham: 27.44 + or - 9.72, C: 21.04 + or - 12.47, E: 42.85 + or - 13.74(Delta), ALN: 25.28 + or - 6.4(.)) (* P < 0.05 vs. sham group, (Delta) P < 0.05 vs. C group, (*) P < 0.05 vs. E group). Trabecular bone in particular was improved, indicating the presence of physiological endosteal bridging (Tr.Dn [%] sham: 10.53 + or - 18.9, C: 1.01 + or - 0.14, E: 24.13 + or - 34.09(Delta), ALN: 3.99 + or - 8.3(.)). ALN did not help bone healing, as shown by mechanical tests. Compared to the C group, statistically, ALN did not show worse properties. The induction of callus formation under ALN treatment was slightly delayed (Tt.Cl [mm(2)] sham: 3.68 + or - 0.66, C: 3.44 + or - 0.42, E: 3.69 + or - 0.58, ALN: 3.06 + or - 0.56). Osteoporotic metaphyseal fracture healing was qualitatively and quantitatively improved by E prophylaxis. The process of fracture healing occurred nearly physiologically (shamlike). Notably, ALN hardly improved metaphyseal callus properties when assessed as osteoporosis prophylaxis, but to a lesser extent than E.

Changes in the histomorphometric and biomechanical properties of the proximal femur of ovariectomized rat after treatment with the phytoestrogens genistein and equol.

The isoflavonoids found in soy have attracted great interest as dietary phytoestrogens that might be effective for postmenopausal hormone replacement therapy. Special attention has been devoted to the hormonal effects of various isoflavonoids, like genistein (GEN) and daidzein's (DAID) potent metabolite, equol (EQ). Here we aimed to investigate the short-term effects of genistein and equol on the proximal femur of ovariectomized (OVX) rats. Forty-eight, 3-month-old female Sprague-Dawley rats were ovarectomized; after eight weeks the bilateral osteotomy and osteosynthesis (OS) of their tibiae was performed and the rats were randomly divided into the following four groups: OVX control group (C), treated with estradiol-17beta (E2) -benzoate (E; daily intake 0.086 mg/d per animal), genistein (GEN; daily intake 12.7 mg/d per animal) and equol (EQ; daily intake 4.65 mg/d per animal). At 5 weeks postoperatively (OS), the breaking test was performed on the trochanteric region of femur. Additionally, histomorphometric assessment, and trabecular and cortical bone microstructure analyses were performed. The relative gain of body weight (BW) in the EQ (24 %) group was significantly (p < 0.05) lower than in the C (33 %) and GEN (30 %) groups. After treatment for 5 weeks, the maximal load (F(max)) and yield load (yL) were higher (p < 0.05 for the weight-adapted results) in the E (188.4 N resp. 113.1 N) and EQ (177.3 N resp. 112 N) groups as compared to C (162.8 N resp. 109.1 N) and GEN (165.7 N resp. 108.8 N). In the histomorphometric tests the E- (trabecular area (Tb.Ar) = 74.93 %, trabecular nodes/mm(2) (N.Nd/mm(2)) = 48.65) and EQ-treated (Tb.Ar = 63.13 %, N.Nd/mm(2) = 43.72) animals showed significant improvement with regard to Tb.Ar and trabecular connectivity (N.Nd./mm(2)) in comparison to C (Tb.Ar = 46.84, N.Nd/mm(2) = 31.86) and GEN (Tb.Ar = 48.22 %, N.Nd/mm(2) = 34.15). There were no differences in relative cortical width (Ct.Wi) among the four groups. The treatment with EQ resulted in improved biomechanical and histomorphometric properties as compared to the treatment with GEN. Thus, of the studied substances, EQ seems to be a possible alternative to hormone replacement therapy, but further studies are needed.

Effects of black cohosh (Cimicifuga racemosa) and estrogen on metaphyseal fracture healing in the early stage of osteoporosis in ovariectomized rats.

Osteoporosis and its accompanying, predominantly metaphyseal, fractures are a major health problem. Black cohosh (Cimicifuga racemosa) and estrogen positively influence osteoporotic bone. Both substances may improve fracture healing in early osteoporosis as well. In 48 twelve-week-old ovariectomized or, respectively, sham-operated (SHAM) rats, a standardized metaphyseal tibia osteotomy with bridging T-plate fixation was performed. During the healing process of 35 days, rats received soy-free (SHAM, osteopenic C), estrogen- (E) or Cimicifuga racemosa- (CR) supplemented diets. After sacrifice, the callus formation was analyzed with regard to biomechanical quality, morphology, quantity, time course of new bone built and gene expression. CR induced a high rate of metaphyseal callus formation. The biomechanical properties and the amount of new callus formation indicated that fracture healing was still in progress. Therefore, gene expression of osteoblasts was comparatively high. Body weight and the trabecular structure were influenced little by CR. Estrogen improved the biomechanical properties of the callus. Resistance to microfracturing was significantly enhanced in the E group and even superior to SHAM. Remodeling of the callus formation had already begun. The trabecular network and the typical endosteal fracture healing were especially improved. Osteoporotic metaphyseal fracture healing was improved by estrogen more than by Cimicifuga racemosa. The process of fracture healing occurred nearly physiologically. The generation of callus formation was supported by Cimicifuga racemosa as well, but the five-week duration of application was too short for Cimicifuga racemosa to show its complete potential. Already-initiated Cimicifuga racemosa therapy for menopausal symptoms could be continued during fracture healing without hesitation.

Absence of positive effect of black cohosh (Cimicifuga racemosa) on fracture healing in osteopenic rodent model.

The healing of predominantly metaphyseal fractures in postmenopausal osteoporosis is delayed and comparatively poor. Due to the potential side effects of HRT, natural alternatives are appealing. The aim of this study was to determine whether Cimicifuga racemosa extract BNO 1055 improves metaphyseal fracture healing in severe osteopenic bone in rats. Thirty-three 12-week-old female rats developed severe osteopenia during 10 weeks after ovariectomy. After metaphyseal tibial-osteotomy and standardized T-plate-osteosynthesis, the healing periods in ovariectomized rats (C), 17-α-estradiol (E) and Cimicifuga racemosa (CR) supplemented diets were assessed for 35 days. Changes in callus morphology were evaluated qualitatively by biomechanical testing and quantitatively in microradiographies and fluorochrome-labeled histological sections. The CR-supplementation slightly improved callus quality and trabecular bone formation. It significantly enhanced the endosteal callus density compared to C group (Cl.Dn.e C: 59.08 ± 21.89, E: 45.95 ± 18.39, CR: 60.85 ± 18.66*), though most of the other morphological parameters examined showed no improvement. The time course of fracture healing did not change due to CR. Estrogen-supplementation enhanced the biomechanical properties of the fracture site. Trabecular bone was improved indicating the physiological endosteal healing process. The CR-supplementation did not exhibit positive effects in severe (senile) osteopenic fracture healing as seen in early (postmenopausal) osteoporosis in rats. Callus formation was slightly improved under CR. Estrogen improved fracture healing in severe osteopenic bone, while the extent of callus formation played a minor role.

Equol but not genistein improves early metaphyseal fracture healing in osteoporotic rats.

Healing of predominantly metaphyseal fractures in postmenopausal osteoporosis is delayed and comparatively poor. Hormone replacement therapy could improve fracture healing, but, because of its potential side effects, natural alternatives are more appealing. The aim of this study was to determine if the soy metabolite equol and the native isoflavone genistein, in comparison to 17beta-estradiol, improve metaphyseal fracture healing in ovariectomy-induced osteoporotic bone of the rat. Forty-eight 12-week-old female rats developed severe osteoporosis ten weeks after ovariectomy. After metaphyseal tibial osteotomy and standardized stable internal fixation, changes in callus morphology were evaluated biomechanically, qualitatively and quantitatively in fluorochrome-labeled histological sections and microradiographs in ovariectomized rats (C) and under standardized 17beta-estradiol (E), equol (EQ) and genistein (G) supplemented rats over a period of five weeks. Estrogen and equol were able to improve the elasticity of callus formation significantly in postmenopausal osteoporotic bone (stiffness of C: 121.40 +/- 47.08 N/mm, E: 147.90 +/- 39.38 N/mm, EQ: 167.8 +/- 59.90 N/mm). The effects of estrogen were more anabolic than those of equol and were visible in changes to the trabecular bone (N.Nd of E: 6.47 +/- 7.68, EQ: 4.25 +/- 3.96). However, in terms of the whole body, equol seemed to induce less of an adverse reaction than estrogen (body weight of C: 342.20 +/- 19.91 g, E: 280.25 +/- 12.05 g, EQ: 308.75 +/- 24.28 g). Genistein as an osteoclast inhibitor influenced callus stiffness (G: 144.50 +/- 61.52 N/mm) and negatively impacted trabecular structure (N.Nd of G: 0.59 +/- 1.01) in severely osteoporotic bones. Estrogen and equol were able to improve fracture healing in ovariectomy-induced osteoporotic bones, and the extent of callus formation played only a minor role. Genistein rather negatively influenced fracture healing. The metaphyseal osteotomy model in ovariectomized rats allows an accurate study of the therapeutic effects of antiosteoporotic substances on the fracture healing process.

Tibial shaft fracture and ankle joint injury.

OBJECTIVE: Detection of tibial fractures in which a concomitant ankle injury may exist. DESIGN: Prospective study. SETTING: Department of Trauma Surgery, University Hospital. PATIENTS: 43 (20.1%) of 214 patients with a tibial fracture were found to have an associated injury of the ankle joint. INTERVENTION: Analysis of all patients with tibial fractures regarding typical mechanisms of injuries and typical radiographic criteria for concomitant injuries of the ankle joint. MAIN OUTCOME MEASURES: Primary x-rays were analyzed looking for spiral fractures of the tibia or proximal fibular fractures or an intact fibula, typically associated with syndesmotic injury. The assessment of patients was based on radiological findings and functional recovery. RESULTS: 45 ankle injuries in 43 patients were found. There were distal fibular fractures in 14, Maisonneuve fractures in 13, isolated ruptures of the syndesmosis in 3, fractures of the posterior malleolus in 8, and fractures of the medial malleolus in 7 of the cases. In 38 of the 43 patients, the distal tibiofibular syndesmosis was ruptured, and 88.4% of the tibia injuries were spiral fractures located in the distal third. Of the 38 patients who could be followed, 31 were categorized according to the Phillip's Score as very good, 3 as good, 2 as satisfactory, and 2 as unsatisfactory after an average of 19.8 months (12-26). CONCLUSION: Due to the obvious injury of the tibia, the potential instability of the ankle joint is often overlooked, and the risk of development of secondary osteoarthritis is often consequently underestimated. Added attention should be paid to the ankle in the following tibial fracture cases: pronation-eversion trauma, spiral fracture of the tibia, proximal fibular fracture, or intact fibula. Using these markers, we were able to diagnose 20.1% of combined injuries compared to our retrospective study in 1999, in which only 13.6% of these injuries could be detected (Pearson r=0.1305, not significant).

Effects of 8-Prenylnaringenin and Whole-Body Vibration Therapy on a Rat Model of Osteopenia.

Background. 8-Prenylnaringenin (8-PN) is the phytoestrogen with the highest affinity for estrogen receptor-α (ER-α), which is required to maintain BMD. The osteoprotective properties of 8-PN have been demonstrated previously in tibiae. We used a rat osteopenia model to perform the first investigation of 8-PN with whole-body vertical vibration (WBVV). Study Design. Ovariectomy was performed on 52 of 64 Sprague-Dawley rats. Five weeks after ovariectomy, one group received daily injections (sc) of 8-PN (1.77 mg/kg) for 10 weeks; a second group was treated with both 8-PN and WBVV (twice a day, 15 min, 35 Hz, amplitude 0.47 mm). Other groups received either only WBVV or no treatment. Methods. The rats were sacrificed 15 weeks after ovariectomy. Lumbar vertebrae and femora were removed for biomechanical and morphological assessment. Results. 8-PN at a cancer-safe dose did not cause fundamental improvements in osteoporotic bones. Treatment with 8-PN caused a slight increase in uterine wet weight. Combined therapy using WBVV and 8-PN showed no significant improvements in bone structure and biomechanical properties. Conclusion. We cannot confirm the osteoprotective effects of 8-PN at a cancer-safe dose in primary affected osteoporotic bones. Higher concentrations of 8-PN are not advisable for safety reasons. Adjunctive therapy with WBVV demonstrates no convincing effects on bones.

Differentiation dependent expression of urocortin's mRNA and peptide in human osteoprogenitor cells: influence of BMP-2, TGF-beta-1 and dexamethasone.

Urocortin-1 (UCN) a corticotropin releasing-factor (CRF) related peptide, has been found to be expressed in many different tissues like the central nervous system, the cardiovascular system, adipose tissue, and skeletal muscle. The effects of UCN are mediated via stimulation of CRF-receptors 1 and 2 (CRFR1 and 2, CRFR's) with a high affinity for CRFR2. It has been shown that the CRF-related peptides and CRFR's are involved in the regulation of stress-related endocrine, autonomic and behavioural responses. Using immunocytochemistry, immunohistochemistry and RT-PCR, we now can show the differentiation dependent expression of UCN mRNA and peptide in human mesenchymal progenitor cells (MSCs) directed to the osteoblastic phenotype for the first time. UCN expression was down regulated by TGF-beta and BMP-2 in the early proliferation phase of osteoblast development, whereas dexamethasone (dex) minimally induced UCN gene expression during matrix maturation after 24 h stimulation. Stimulation of MSCs for 28 days with ascorbate/beta-glycerophosphate (asc/bGp) induced UCN gene expression at day 14. This effect was prevented when using 1,25-vitamin D3 or dex in addition. There was no obvious correlation to osteocalcin (OCN) gene expression in these experiments. In MSCs from patients with metabolic bone disease (n = 9) UCN gene expression was significantly higher compared to MSCs from normal controls (n = 6). Human MSCs did not express any of the CRFR's during differentiation to osteoblasts. Our results indicate that UCN is produced during the development of MSCs to osteoblasts and differentially regulated during culture as well as by differentiation factors. The expression is maximal between proliferation and matrix maturation phase. However, UCN does not seem to act on the osteoblast itself as shown by the missing CRFR's. Our results suggest new perspectives on the role of urocortin in human skeletal tissue in health and disease.

Effect of daidzein, 4-methylbenzylidene camphor or estrogen on gastrocnemius muscle of osteoporotic rats undergoing tibia healing period.

The effect of daidzein (D), 4-methylbenzylidene camphor (4-MBC) or estradiol-17beta-benzoate (E(2)) on muscle of osteoporotic rats during fracture healing was studied. After performing a metaphyseal tibia osteotomy in 96 osteoporotic 5-month-old female Sprague-Dawley rats, they received daily 50 mg D, 200 mg 4-MBC or 0.4 mg E(2) per kg body weight, or soy free (SF) diet up to 36 and 72 days. Mitochondrial activity, fiber area, and capillary density were analyzed in M. gastrocnemius. Osseous callus bridging of fracture was observed in half of the rats after 36 days. By day 72, fracture was healed in most of the animals. State 3 mitochondrial respiration significantly enhanced in E(2), 4-MBC and D groups versus SF after 36 days (30, 32 and 32 vs 23 pmol O(2)/s per mg). It declined after 72 days, however, in E(2) group it was still at a higher level versus SF (25, 23 and 21 vs 20 pmol O(2)/s per mg). Size of fast oxidative glycolytic (FOG) and fast glycolytic (FG) fibers, capillary density did not differ significantly between the groups, however, at day 36 an increase in D and 4-MBC groups was detectable. FOG diameter was 64, 66, 68, and 58 microm and FG diameter was 88, 98, 95, and 89 microm in SF, D, 4-MBC, and E(2) groups. The ratio of capillaries to muscle fiber was 1.1, 1.4, 1.3, and 1.1 in SF, D, 4-MBC and E(2) groups by day 36. D and 4-MBC react similar to estrogen thereby improving oxidative cell metabolism in severe osteoporotic rats. The level of mitochondrial activity was higher, though no significant morphological differences could be shown.

The Elastic Bridge Plating of the Forearm Fracture: A Prospective Study.

Rigid plate osteosynthesis with compression is still the treatment of choice for forearm fractures to gain anatomic reposition, provide proper rotation and avoid a bridging callus. Due to necessary operative dissection there is a serious risk for infection and malunion. Based on good clinical results with elastic bridge plating at femur, humerus and tibia, this technique was also started to be used for forearm fractures in our clinic in 1995. In a prospective study, 86 of 124 consecutive patients at the age of 35.2 ± 14.7 years with 129 diaphyseal fractures of the radius or ulna (AO: 37 type A, 36 type B, 13 type C) were analyzed between January 1998 and December 2003. All fractures were stabilized by bridge plating. Radiographic union and clinical outcome were documented. Of the 129, 122 diaphyseal fractures (94.5%) healed within 10.2 ± 3.4 weeks without complications (no nerve lesions, nonunion, synostosis callus). One re-osteosynthesis, one secondary lag screw, and five cancellous bone grafts were necessary before final healing. About 79.1% of the patients had a perfect clinical outcome; 17.4% had additional severe injuries of the same arm. Bridge plating without interfragmentary compression is a reliable surgical procedure even for forearm fractures with low risk of infection and nonunion.

The use of flat panel volumetric computed tomography (fpVCT) in osteoporosis research.

RATIONALE AND OBJECTIVES: Improvements in imaging technology have led to the increased use of computed tomography (CT). For example, micro-CT and quantitative CT (QCT) are now often used in osteoporosis research, in which micro-CT is able to analyze small bones or bone samples with high spatial resolution. In contrast, QCT is able to investigate large samples with low spatial resolution. The aim of this study was to test the usefulness of flat-panel volumetric CT (fpVCT) in a rat model of osteopenia. MATERIAL AND METHODS: Twenty-two 3-month-old rats underwent ovariectomy and were either left untreated or supplemented with estradiol for 15 weeks. After sacrificing, the rats' second lumbar vertebral body bone mineral density (BMD) was analyzed using fpVCT and ashing. The results were compared to those of a microstructural analysis of the first lumbar vertebrae and a biomechanical evaluation of the fourth lumbar vertebrae. RESULTS: BMD measurements using both fpVCT (0.39 vs 0.35 mg/cm(3)) and ashing (0.52 vs 0.48 mg/cm(3)) demonstrated a significant improvement after estradiol supplementation. The correlation coefficient of the two methods was 0.858. After estradiol supplementation, the bone microstructural and bone biomechanical parameters were improved, compared to no treatment. The correlations of both the microstructural and the biomechanical evaluations were closer for BMD measured using fpVCT (r = 0.482-0.769) than on the basis of ashing (r = 0.345-0.573). FpVCT was not able to display the trabecular microstructure of the rat lumbar vertebrae. CONCLUSION: The use of fpVCT demonstrated a close relationship between morphologic and biomechanical evaluations in a rat model of osteopenia. Because of its different proportions, fpVCT might be able to bridge the gap between micro-CT and QCT in analyzing larger animals.

Comparison of the phytohormones genistein, resveratrol and 8-prenylnaringenin as agents for preventing osteoporosis.

As the average age of society increases, identifying and preventing osteoporosis becomes more important. According to the results of the Women's Health Initiative study, substitution of estradiol is not recommended in hormone replacement therapy (HRT), although phytoestrogens might be a safe alternative. In this study, the osteoprotective effects of genistein (Gen), resveratrol (Res) and 8-prenylnaringenin (8PN) were evaluated by analysing bone biomechanical strength and bone mineral density. After ovariectomy, 88 female rats received soy-free food (C), and according to their grouping, were fed estradiol (E), GEN, RES or 8PN for 12 weeks. The phytohormones were given in two dosages. To analyse the osteoprotective effects of the tested substances, bone biomechanical properties and bone mineral density (BMD) were evaluated on the upper tibial metaphysis. Bone biomechanical properties were significantly improved after treatment with E (F (max): 90.6 N) and 8PN (85.0 N) compared to GEN (76.0 N), RES (72.6 N) and C (76.6 N). Bone biomechanical properties with 8PN (yL: 55.7 N) supplementation reached a level similar to that seen after E (49.3 N) supplementation. Treatment with GEN (38.5 N) was not as effective as E and 8PN, but demonstrated improved biomechanical properties compared to C (40.1 N) and RES (36.3 N). E (Cn.Dn. 217 mg/cm (3)) and 8PN (165 mg/cm3) showed superior results in the analysis of bone mineral density compared to C (112 mg/cm (3)). GEN (164 mg/cm (3)) also demonstrated superior results, though not as good as E and 8PN. RES (124 mg/cm (3)) revealed no effect on bone density. Treatment with 8PN resulted in very good biomechanical properties and showed an increased BMD. GEN had a smaller effect on bone biomechanical strength, while RES did not have an effect on bone biomechanical strength or BMD. Therefore, 8PN might be a safe alternative for HRT, but further studies are needed.