Raloxifene Prevents Skeletal Fragility in Adult Female Zucker Diabetic Sprague-Dawley Rats

Authors

Kathleen M. Hill Gallant, Maxime A. Gallant, Drew M. Brown, Amy Y. Sato, Justin N. Williams, David B. Burr

Abstract

Fracture risk in type 2 diabetes is increased despite normal or high bone mineral density, implicating poor bone quality as a risk factor. Raloxifene improves bone material and mechanical properties independent of bone mineral density. This study aimed to determine if raloxifene prevents the negative effects of diabetes on skeletal fragility in diabetes-prone rats. Adult Zucker Diabetic Sprague-Dawley (ZDSD) female rats (20-week-old, n = 24) were fed a diabetogenic high-fat diet and were randomized to receive daily subcutaneous injections of raloxifene or vehicle for 12 weeks. Blood glucose was measured weekly and glycated hemoglobin was measured at baseline and 12 weeks. At sacrifice, femora and lumbar vertebrae were harvested for imaging and mechanical testing. Raloxifene-treated rats had a lower incidence of type 2 diabetes compared with vehicle-treated rats. In addition, raloxifene-treated rats had blood glucose levels significantly lower than both diabetic vehicle-treated rats as well as vehicle-treated rats that did not become diabetic. Femoral toughness was greater in raloxifene-treated rats compared with both diabetic and non-diabetic vehicle-treated ZDSD rats, due to greater energy absorption in the post-yield region of the stress-strain curve. Similar differences between groups were observed for the structural (extrinsic) mechanical properties of energy-to-failure, post-yield energy-to-failure, and post-yield displacement. These results show that raloxifene is beneficial in preventing the onset of diabetes and improving bone material properties in the diabetes-prone ZDSD rat. This presents unique therapeutic potential for raloxifene in preserving bone quality in diabetes as well as in diabetes prevention, if these results can be supported by future experimental and clinical studies.

Link To Article

http://dx.doi.org/10.1371/journal.pone.0108262

Fufang Kushen injection inhibits sarcoma growth and tumor-induced hyperalgesia via TRPV1 signaling pathways

Authors

Zhizheng Zhao, Huiting Fan, Tim Higgins, Jia Qi, Diana Haines, Anna Trivett, Joost J. Oppenheim, Hou Wei, Jie Li, Hongsheng Lin, O.M. Zack Howard

Abstract

Cancer pain is a deleterious consequence of tumor growth and related inflammation. Opioids and anti-inflammatory drugs provide first line treatment for cancer pain, but both are limited by side effects. Fufang Kushen Injection (FKI) is GMP produced, traditional Chinese medicine used alone or with chemotherapy to reduce cancer-associated pain. FKI limited mouse sarcoma growth both in vivo and in vitro, in part, by reducing the phosphorylation of ERK and AKT kinases and BAD. FKI inhibited TRPV1 mediated capsaicin-induced ERK phosphorylation and reduced tumor-induced proinflammatory cytokine production. Thus, FKI limited cancer pain both directly by blocking TRPV1 signaling and indirectly by reducing tumor growth.

Link To Article

http://dx.doi.org/10.1016/j.canlet.2014.08.037

A novel therapeutic approach with Caviunin-based isoflavonoid that en routes bone marrow cells to bone formation via BMP2/Wnt-β-catenin signaling

Authors

P Kushwaha, V Khedgikar, J Gautam, P Dixit, R Chillara, A Verma, R Thakur, D P Mishra, D Singh1, R Maurya, N Chattopadhyay, P R Mishra and R Trivedi

Abstract

Recently, we reported that extract of Dalbergia sissoo made from leaves and pods have antiresorptive and bone-forming effects. The positive skeletal effect attributed because of active molecules present in the extract of Dalbergia sissoo. Caviunin 7-O-[β-d-apiofuranosyl-(1-6)-β-d-glucopyranoside] (CAFG), a novel isoflavonoid show higher percentage present in the extract. Here, we show the osteogenic potential of CAFG as an alternative for anabolic therapy for the treatment of osteoporosis by stimulating bone morphogenetic protein 2 (BMP2) and Wnt/β-catenin mechanism. CAFG supplementation improved trabecular micro-architecture of the long bones, increased biomechanical strength parameters of the vertebra and femur and decreased bone turnover markers better than genistein. Oral administration of CAFG to osteopenic ovariectomized mice increased osteoprogenitor cells in the bone marrow and increased the expression of osteogenic genes in femur and show new bone formation without uterine hyperplasia. CAFG increased mRNA expression of osteoprotegerin in bone and inhibited osteoclast activation by inhibiting the expression of skeletal osteoclastogenic genes. CAFG is also an effective accelerant for chondrogenesis and has stimulatory effect on the repair of cortical bone after drill-hole injury at the tissue, cell and gene level in mouse femur. At cellular levels, CAFG stimulated osteoblast proliferation, survival and differentiation. Signal transduction inhibitors in osteoblast demonstrated involvement of p-38 mitogen-activated protein kinase pathway stimulated by BMP2 to initiate Wnt/β-catenin signaling to reduce phosphorylation of GSK3-β and subsequent nuclear accumulation of β-catenin. Osteogenic effects were abrogated by Dkk1, Wnt-receptor blocker and FH535, inhibitor of TCF-complex by reduction in β-catenin levels. CAFG modulated MSC responsiveness to BMP2, which promoted osteoblast differentiation via Wnt/β-catenin mechanism. CAFG at 1 mg/kg/day dose in ovariectomy mice (human dose ~0.081 mg/kg) led to enhanced bone formation, reduced bone resorption and bone turnover better than well-known phytoestrogen genistein. Owing to CAFG’s inherent properties for bone, it could be positioned as a potential drug, food supplement, for postmenopausal osteoporosis and fracture repair.

Link To Article

http://dx.doi.org/10.1038/cddis.2014.350

Local Administration of Bone Morphogenetic Protein-2 and Bisphosphonate During Non-Weight-Bearing Treatment of Ischemic Osteonecrosis of the Femoral Head

Authors

Harry K.W. Kim, MD, MS; Olumide Aruwajoye, MS; Justin Du ; Nobuhiro Kamiya, MD, PhD

Abstract

Background: Non-weight-bearing decreases the femoral head deformity but increases bone resorption without increasing bone formation in an experimental animal model of Legg-Calvé-Perthes disease. We sought to determine if local administration of bone morphogenetic protein (BMP)-2 with or without bisphosphonate can increase the bone formation during the non-weight-bearing treatment in the large animal model of Legg-Calvé-Perthes disease.

Methods: Eighteen piglets were surgically induced with femoral head ischemia. Immediately following the surgery, all animals received an above-the-knee amputation to enforce local non-weight-bearing (NWB). One to two weeks later, six animals received local BMP-2 to the necrotic head (BMP group), six received local BMP-2 and ibandronate (BMP+IB group), and the remaining six received no treatment (NWB group). All animals were killed at eight weeks after the induction of ischemia. Radiographic, microcomputed tomography (micro-CT), and histomorphometric assessments were performed.

Results: Radiographic assessment showed that the femoral heads in the NWB, BMP, and BMP+IB groups had a decrease of 20%, 14%, and 10%, respectively, in their mean epiphyseal quotient in comparison with the normal control group. Micro-CT analyses showed significantly higher femoral head bone volume in the BMP+IB group than in the BMP group (p = 0.02) and the NWB group (p < 0.001). BMP+IB and BMP groups had a significantly higher trabecular number (p < 0.01) and lower trabecular separation (p < 0.02) than the NWB group. In addition, the osteoclast number per bone surface was significantly lower in the BMP+IB group compared with the NWB group. Calcein labeling showed significantly higher bone formation in the BMP and BMP+IB groups than in the NWB group (p < 0.05). Heterotopic ossification was found in the capsule of four hips in the BMP+IB group but not in the BMP group.

Conclusions: Administration of BMP-2 with bisphosphonate best decreased bone resorption and increased new bone formation during non-weight-bearing treatment of ischemic osteonecrosis in a pig model, but heterotopic ossification is a concern.

Clinical Relevance: This preclinical study provides new evidence that BMP-2 with bisphosphonate can effectively prevent the extreme bone loss associated with the non-weight-bearing treatment and increase new bone formation in the femoral head in this animal model of ischemic osteonecrosis.

Link To Article

http://dx.doi.org/10.2106/JBJS.M.01361

Dietary Stearic Acid Leads to a Reduction of Visceral Adipose Tissue in Athymic Nude Mice

Authors

Ming-Che Shen, Xiangmin Zhao, Gene P. Siegal, Renee Desmond, Robert W. Hardy

Abstract

Stearic acid (C18:0) is a long chain dietary saturated fatty acid that has been shown to reduce metastatic tumor burden. Based on preliminary observations and the growing evidence that visceral fat is related to metastasis and decreased survival, we hypothesized that dietary stearic acid may reduce visceral fat. Athymic nude mice, which are used in models of human breast cancer metastasis, were fed a stearic acid, linoleic acid (safflower oil), or oleic acid (corn oil) enriched diet or a low fat diet ad libitum. Total body weight did not differ significantly between dietary groups over the course of the experiment. However visceral fat was reduced by ~70% in the stearic acid fed group compared to other diets. In contrast total body fat was only slightly reduced in the stearic acid diet fed mice when measured by dual-energy x-ray absorptiometry and quantitative magnetic resonance. Lean body mass was increased in the stearic acid fed group compared to all other groups by dual-energy x-ray absorptiometry. Dietary stearic acid significantly reduced serum glucose compared to all other diets and increased monocyte chemotactic protein-1 (MCP-1) compared to the low fat control. The low fat control diet had increased serum leptin compared to all other diets. To investigate possible mechanisms whereby stearic acid reduced visceral fat we used 3T3L1 fibroblasts/preadipocytes. Stearic acid had no direct effects on the process of differentiation or on the viability of mature adipocytes. However, unlike oleic acid and linoleic acid, stearic acid caused increased apoptosis (programmed cell death) and cytotoxicity in preadipocytes. The apoptosis was, at least in part, due to increased caspase-3 activity and was associated with decreased cellular inhibitor of apoptosis protein-2 (cIAP2) and increased Bax gene expression. In conclusion, dietary stearic acid leads to dramatically reduced visceral fat likely by causing the apoptosis of preadipocytes.

Link To Article

http://dx.doi.org/10.1371/journal.pone.0104083

Activated protein C (APC) can increase bone anabolism via a protease-activated receptor (PAR)1/2 dependent mechanism

Authors

Kaitlin Shen, Ciara M. Murphy, Ben Chan, Mille Kolind, Tegan L. Cheng, Kathy Mikulec, Lauren Peacock, Meilang Xue, Sang-Youel Park, David G. Little, Chris J. Jackson, and Aaron Schindeler

Abstract

Activated Protein C (APC) is an anticoagulant with strong cytoprotective properties that has been shown to promote wound healing. In this study APC was investigated for its potential orthopedic application using a Bone Morphogenetic Protein 2 (rhBMP-2) induced ectopic bone formation model. Local co-administration of 10 µg rhBMP-2 with 10 µg or 25 µg APC increased bone volume at 3 weeks by 32% (N.S.) and 74% (p < 0.01) compared to rhBMP-2 alone. This was associated with a significant increase in CD31+ and TRAP+ cells in tissue sections of ectopic bone, consistent with enhanced vascularity and bone turnover. The actions of APC are largely mediated by its receptors endothelial protein C receptor (EPCR) and protease-activated receptors (PARs). Cultured pre-osteoblasts and bone nodule tissue sections were shown to express PAR1/2 and EPCR. When pre-osteoblasts were treated with APC, cell viability and phosphorylation of ERK1/2, Akt, and p38 were increased. Inhibition with PAR1 and sometimes PAR2 antagonists, but not with EPCR blocking antibodies, ameliorated the effects of APC on cell viability and kinase phosphorylation. These data indicate that APC can affect osteoblast viability and signaling, and may have in vivo applications with rhBMP-2 for bone repair.

Link To Article

http://dx.doi.org/10.1002/jor.22726