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

Musculoskeletal Response to Whole-Body Vibration During Fracture Healing in Intact and Ovariectomized Rats

This study investigated the effect of vibration on bone healing and muscle in intact and ovariectomized rats. Thirty ovariectomized (at 3 months of age) and 30 intact 5-month old female Sprague-Dawley rats underwent bilateral metaphyseal osteotomy of tibia. Five days later, half of the ovariectomized and of the intact rats were exposed to whole-body vertical vibration (90 Hz, 0.5 mm, 4 × g acceleration) for 15 min twice a day during 30 days. The other animals did not undergo vibration. After decapitation of rats, one tibia was used for computed tomographic, biomechanical, and histological analyses; the other was used for gene expression analyses of alkaline phosphatase (Alp), osteocalcin (Oc), tartrate-resistant acid phosphatase 1, and insulinlike growth factor 1. Serum Alp and Oc were measured. Mitochondrial activity, fiber area and distribution, and capillary densities were analyzed in M. gastrocnemius and M. longissimus. We found that vibration had no effect on body weight and food intake, but it improved cortical and callus densities (97 vs. 99%, 72 vs. 81%), trabecular structure (9 vs. 14 trabecular nodes), blood supply (1.7 vs. 2.1 capillaries/fiber), and oxidative metabolism (17 vs. 23 pmol O2/s/mg) in ovariectomized rats. Vibration generally increased muscle fiber size. Tibia biomechanical properties were diminished after vibration. Oc gene expression was higher in vibrated rats. Serum Alp was increased in ovariectomized rats. In ovariectomized rats, vibration resulted in an earlier bridging; in intact rats, callus bridging occurred later after vibration. The chosen vibration regimen (90 Hz, 0.5 mm, 4 × g acceleration, 15 min twice a day) was effective in improving musculoskeletal tissues in ovariectomized rats but was not optimal for fracture healing

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

Evaluation of twelve vibration regimes applied to improve spine properties in ovariectomized rats

While whole-body vibration (WBV) has recently been introduced as a non-pharmacological therapy for osteoporosis, studies have shown that it has no significant effect on the lumbar spine in older women. However, the vibration protocols differed among studies, and the major factor influencing the outcomes is unclear. The intention of the present study was to evaluate the effect of WBV—vertical (v) or horizontal (h) and of different frequencies and application regimes (1× or 2×/d)—on lumbar spine properties in ovariectomized rats (Ovx). Three experiments were conducted. Thirteen-week old female Sprague–Dawley rats were Ovx or left intact (Non-Ovx). After eight weeks, all of the rats underwent metaphyseal osteotomy of the tibiae. Five days later, the rats were divided into six groups (n = 15): 1) intact, 2) Ovx, and 3–6) Ovx exposed to WBV. In Experiment 1, groups 3–6 underwent 35 Hz-v, 50 Hz-v, 70 Hz-v, and 90 Hz-v, respectively. In Experiment 2, groups 3–6 underwent 30 Hz-h, 50 Hz-h, 70 Hz-h, and 90 Hz-h, respectively. In Experiment 3, groups 3–6 underwent 35 Hz-v, 70 Hz-v, 35 Hz-h, and 70 Hz-h, respectively. Vibration exposure was 15 min 1×/d in Experiment 1 and 2 and 2×/d in Experiment 3 for up to 30 days. Vertebral bodies were used in micro-computed tomography, biomechanical, ashing, and gene expression analyses. Vertical vibrations applied once a day favorably affected bone volume fraction (BV/TV) and Ca2+/PO43− and decreased Rankl gene expression. When applied twice a day, v-vibrations diminished mineral content. Horizontal vibrations (1×/d) reduced Ca2+/PO43− ratio and Opg mRNA level, whereas h-vibration (2×/d) normalized OC serum levels. Many of the other measured parameters did not reveal any significant differences between the vibrated groups and the untreated Ovx group. The effect of ovariectomy was confirmed by atrophied uterus, impaired biomechanical properties, and bone mineral density and BV/TV of the vertebral body. The findings of the present study indicate that application frequency rate and direction of vibration might influence spine response differently. However, we were unable to find any clearly beneficial or harmful effect of vibration regimes on the osteopenic lumbar spine in rats