CXC chemokine ligand 12a enhances chondrocyte proliferation and maturation during endochondral bone formation

Authors

G.-W. Kim, M.-S. Han, H.-R. Park, E.-J. Lee, Y.-K. Jung, S.E. Usmani, V. Ulici, S.-W. Han, F. Beier

Abstract

Objective We investigated the roles of CXC chemokine ligand 12a (CXCL12a), also known as stromal cell-derived factor-1α (SDF-1α), in endochondral bone growth, which can give us important clues to understand the role of CXCL12a in osteoarthritis (OA).

Methods Primary chondrocytes and tibial explants from embryonic 15.5 day-old mice were cultured with recombinant mouse CXCL12a. To assess the role of CXCL12a in chondrogenic differentiation, we conducted mesenchymal cell micromass culture.

Results In tibia organ cultures, CXCL12a increased total bone length in a dose-dependent manner through proportional effects on cartilage and bone. In accordance with increased length, CXCL12a increased the protein level of proliferation markers, such as cyclin D1 and proliferating cell nuclear antigen (PCNA), in primary chondrocytes as well as in tibia organ culture. In addition, CXCL12a increased the expression of Runx2, Col10 and MMP13 in primary chondrocytes and tibia organ culture system, implying a role of CXCL12a in chondrocyte maturation. Micromass cultures of limb-bud mesenchymal progenitor cells (MPCs) revealed that CXCL12a has a limited effect on early chondrogenesis, but significantly promoted maturation of chondrocytes. CXCL12a induced the phosphorylation of p38 and Erk1/2 MAP kinases and IκB. The increased expression of cyclin D1 by CXCL12a was significantly attenuated by inhibitors of MEK1 and NF-κB. On the other hand, p38 and Erk1/2 MAP kinase and NF-κB signaling were associated with CXCL12a-induced expression of Runx2 and MMP13, the marker of chondrocyte maturation.

Conclusion CXCL12a promoted the proliferation and maturation of chondrocytes, which strongly suggest that CXCL12a may have a negative effect on articular cartilage and contribute to OA progression.

Link To Article

http://dx.doi.org/10.1016/j.joca.2015.01.016

Decreasing NF-κB Expression Enhances Odontoblastic Differentiation and Collagen Expression in Dental Pulp Stem Cells Exposed to Inflammatory Cytokines

Authors

Neda S. T. Hozhabri, M. Douglas Benson, Michael D. Vu, Rinkesh H. Patel, Rebecca M. Martinez, Fatemeh N. Nakhaie, Harry K. W. Kim, Venu G. Varanasi

Abstract

Inflammatory response in the dental pulp can alter the collagen matrix formation by dental pulp stem cells and lead to a delay or poor healing of the pulp. This inflammatory response is mediated by cytokines, including interleukin-1β and tumor necrosis factor-α. In this study, it is hypothesized that suppressing the actions of these inflammatory cytokines by knocking down the activity of transcription factor Nuclear Factor–κB will lead to dental pulp stem cell differentiation into odontoblasts and the production of collagen. Here, the role of Nuclear Factor–κB signaling and its reduction was examined during odontogenic behavior in the presence of these cytokines. The results showed a significant increase in Nuclear Factor–κB gene expression and p65 protein expression by interleukin-1β and tumor necrosis factor-α. Nuclear Factor–κB activation in the presence of these cytokines decreased significantly in a dose-dependent manner by a Nuclear Factor–κB inhibitor (MG132) and p65 siRNA. Down-regulation of Nuclear Factor–κB activity also enhanced the gene expression of the odontoblastic markers (dentin sialophosphoprotein, Nestin, and alkaline phosphatase) and displayed an odontoblastic cell morphology indicating the promotion of odontogenic differentiation of dental pulp stem cells. Finally, dental pulp stem cells exposed to reduced Nuclear Factor–κB activity resulted in a significant increase in collagen (I)-α1 expression in the presence of these cytokines. In conclusion, a decrease in Nuclear Factor-κB in dental pulp stem cells in the presence of inflammatory cytokines enhanced odontoblastic differentiation and collagen matrix formation.

Link To Article

http://dx.doi.org/10.1371/journal.pone.0113334

Preliminary evaluation of local drug delivery of amphotericin B and in vivo degradation of chitosan and polyethylene glycol blended sponges

Authors

Ashley Cox Parker, Cheyenne Rhodes, Jessica Amber Jennings, Lauren Hittle, Mark Shirtliff, Joel D. Bumgardner and Warren O. Haggard

Abstract

This research investigated the combination of polyethylene glycol with chitosan in point-of-care loaded sponges made by one or two lyophilizations for adjunctive local antifungal delivery in musculoskeletal wounds. Blended and control chitosan sponges were evaluated in vitro for antifungal release and activity, degradation, cytocompatibility, and characterized for spectroscopic, crystallinity, thermal, and morphologic material properties. In vivo biocompatibility and degradation of sponges were also evaluated in a rat intramuscular pouch model 4 and 10 days after implantation. Blended sponges released amphotericin B active against Candida albicans (>0.25 µg/mL) over 72 h and did not elicit cytotoxicity response of fibroblasts. Blended sponges exhibited decreases in surface roughness, decreased thermal decomposition temperatures, as well as small Fourier transform infrared spectroscopy and crystallinity differences, compared with chitosan-only sponges. Three of the four blended sponge formulations exhibited 31%–94% increases in in vitro degradation from the chitosan sponges after 10 days, but did not demonstrate the same increase in in vivo degradation. Low inflammatory in vivo tissue response to blended and chitosan-only sponges was similar over 10 days. These results demonstrated that adding polyethylene glycol to chitosan sponges does improve local antifungal release, cytocompatibility, and in vitro degradation, but does not increase in vivo degradation.

Link To Article

http://dx.doi.org/10.1002/jbm.b.33356

Sim1 Inhibits Bone Formation by Enhancing the Sympathetic Tone in Male Mice

Authors

Xunde Wang, Wei Wei, Andrew R. Zinn, and Yihong Wan

Abstract

Sim1 is a bHLH-PAS transcription factor that is important for neuronal development in the hypothalamus. Loss-of-function mutation of Sim1 causes early onset obesity. However, it is unknown whether and how Sim1 regulates bone remodeling. In this study, we found that adult-onset Sim1 deletion increases bone formation, leading to high bone mass. In contrast, Sim1 over-expressing transgenic mice exhibit decreased bone formation and low bone mass. Sim1 does not directly regulate osteoblastogenesis because bone marrow mesenchymal stem cells from Sim1 mutant mice display a normal capacity for osteoblast differentiation. Instead, Sim1 inhibits bone formation via stimulating the sympathetic nervous system because sympathetic tone is decreased by Sim1 deletion but increased by Sim1 overexpression. Treatment with the β-adrenergic agonist isoproterenol effectively reverses the high bone mass in Sim1 KO mice. These findings reveal Sim1 as a critical yet previously unrecognized modulator of skeletal homeostasis that functions through a central relay.

Link To Article

http://dx.doi.org/10.1210/en.2014-1872

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 Singh, 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

Hypohalous Acids Contribute to Renal Extracellular Matrix Damage in Experimental Diabetes

Authors

Kyle L. Brown, Carl Darris, Kristie Lindsey Rose, Otto A. Sanchez, Hartman Madu, Josh Avance, Nickolas Brooks, Ming-Zhi Zhang, Agnes Fogo, Raymond Harris, Billy G. Hudson and Paul Voziyan

Abstract

In diabetes, toxic oxidative pathways are triggered by persistent hyperglycemia and contribute to diabetic complications. A major proposed pathogenic mechanism is accumulation of protein modifications called advanced glycation end products (AGEs). However, other non-enzymatic post-translational modifications may also contribute to pathogenic protein damage in diabetes. We demonstrate that hypohalous acid-derived modifications of renal tissues and extracellular matrix (ECM) proteins are significantly elevated in experimental diabetic nephropathy. Moreover, diabetic renal ECM shows diminished binding of α1β1 integrin consistent with modification of collagen IV by hypochlorous (HOCl) and hypobromous (HOBr) acids. NC1 hexamers, key connection modules of collagen IV networks, are modified via oxidation and chlorination of tryptophan and bromination of tyrosine residues. Chlorotryptophan, a relatively minor modification, has not been previously found in proteins. In the NC1 hexamers isolated from diabetic kidneys, levels of HOCl-derived oxidized and chlorinated tryptophan residues W28 and W192 are significantly elevated compared to non-diabetic controls. Molecular dynamics simulations predicted more relaxed NC1 hexamer tertiary structure and diminished assembly competence in diabetes; this was confirmed using limited proteolysis and denaturation/refolding. Our results suggest that hypohalous acid-derived modifications of renal ECM and specifically collagen IV networks contribute to functional protein damage in diabetes.

Link To Article

http://dx.doi.org/10.2337/db14-1001