Butyrate Inhibits Osteoclast Activity In Vitro and Regulates Systemic Inflammation and Bone Healing in a Murine Osteotomy Model Compared to Antibiotic-Treated Mice.

Short-chain fatty acids (SCFAs) produced by the gut microbiota have previously been demonstrated to play a role in numerous chronic inflammatory diseases and to be key mediators in the gut-bone signaling axis. However, the role of SCFAs in bone fracture healing and its impact on systemic inflammation during the regeneration process has not been extensively investigated yet. The aim of this study was to first determine the effects of the SCFA butyrate on key cells involved in fracture healing in vitro, namely, osteoclasts and mesenchymal stromal cells (MSCs), and second, to assess if butyrate supplementation or antibiotic therapy impacts bone healing, systemic immune status, and inflammation levels in a murine osteotomy model. Butyrate significantly reduced osteoclast formation and resorption activity in a dose-dependent manner and displayed a trend for increased calcium deposits in MSC cultures. Numerous genes associated with osteoclast differentiation were differentially expressed in osteoclast precursor cells upon butyrate exposure. In vivo, antibiotic-treated mice showed reduced SCFA levels in the cecum, as well as a distinct gut microbiome composition. Furthermore, circulating proinflammatory TNFα, IL-17a, and IL-17f levels, and bone preserving osteoprotegerin (OPG), were increased in antibiotic-treated mice compared to controls. Antibiotic-treated mice also displayed a trend towards delayed bone healing as revealed by reduced mineral apposition at the defect site and higher circulating levels of the bone turnover marker PINP. Butyrate supplementation resulted in a lower abundance of monocyte/macrophages in the bone marrow, as well as reduced circulating proinflammatory IL-6 levels compared to antibiotic- and control-treated mice. In conclusion, this study supports our hypothesis that SCFAs, in particular butyrate, are important contributors to successful bone healing by modulating key cells involved in fracture healing as well as systemic inflammation and immune responses.

Calcium Polyphosphate Nanoparticles Act as an Effective Inorganic Phosphate Source during Osteogenic Differentiation of Human Mesenchymal Stem Cells.

The ability of bone-marrow-derived mesenchymal stem/stromal cells (BM-MSCs) to differentiate into osteoblasts makes them the ideal candidate for cell-based therapies targeting bone-diseases. Polyphosphate (polyP) is increasingly being studied as a potential inorganic source of phosphate for extracellular matrix mineralisation. The aim of this study is to investigate whether polyP can effectively be used as a phosphate source during the in vitro osteogenic differentiation of human BM-MSCs. Human BM-MSCs are cultivated under osteogenic conditions for 28 days with phosphate provided in the form of organic ß-glycerolphosphate (BGP) or calcium-polyP nanoparticles (polyP-NP). Mineralisation is demonstrated using Alizarin red staining, cellular ATP content, and free phosphate levels are measured in both the cells and the medium. The effects of BGP or polyP-NP on alkaline phosphatase (ALP) activity and gene expression of a range of osteogenic-related markers are also assessed. PolyP-NP supplementation displays comparable effects to the classical BGP-containing osteogenic media in terms of mineralisation, ALP activity and expression of osteogenesis-associated genes. This study shows that polyP-NP act as an effective source of phosphate during mineralisation of BM-MSC. These results open new possibilities with BM-MSC-based approaches for bone repair to be achieved through doping of conventional biomaterials with polyP-NP.

The Effect of Krill Oil Supplementation on Exercise Performance and Markers of Immune Function.

BACKGROUND: Krill oil is a rich source of the long-chain n-3 polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which may alter immune function after exercise. The aim of the study was to determine the effects of krill oil supplementation on post exercise immune function and performance. METHODS: Nineteen males and 18 females (age: 25.8 ± 5.3 years; mean ± S.D.) were randomly assigned to 2 g/day of krill oil (n = 18) or placebo (n = 19) supplementation for 6 weeks. A maximal incremental exercise test and cycling time trial (time to complete set amount of work) were performed pre-supplementation with the time trial repeated post-supplementation. Blood samples collected pre- and post- supplementation at rest, and immediately, 1 and 3h post-exercise. Plasma IL-6 and thiobarbituric acid reactive substances (TBARS) concentrations and, erythrocyte fatty acid composition were measured. Natural killer (NK) cell cytotoxic activity and peripheral blood mononuclear cell (PBMC) IL-2, IL-4, IL-10, IL-17 and IFNγ production were also measured. RESULTS: No effects of gender were noted for any variable. PBMC IL-2 and NK cell cytotoxic activity were greater (P < 0.05) 3h post exercise in the krill oil compared to the control group. Plasma IL-6 and TBARS, PBMC IL-4, IL-10, IL-17 and IFNγ production, along with performance and physiological measures during exercise, were not different between groups. CONCLUSION: Six weeks of krill oil supplementation can increase PBMC IL-2 production and NK cell cytotoxic activity 3h post-exercise in both healthy young males and females. Krill oil does not modify exercise performance.

Identification of a bisphosphonate that inhibits isopentenyl diphosphate isomerase and farnesyl diphosphate synthase.

We and others have recently shown that the major molecular target of nitrogen-containing bisphosphonate drugs is farnesyl diphosphate synthase, an enzyme in the mevalonate pathway. In an in vitro screen, we discovered a bisphosphonate, NE21650, that potently inhibited farnesyl diphosphate synthase but, unlike other N-BPs investigated, was also a weak inhibitor of isopentenyl diphosphate isomerase. NE21650 was a more potent inhibitor of protein prenylation in osteoclasts and macrophages, and a more potent inhibitor of bone resorption in vitro, than alendronate, despite very similar IC(50) values for inhibition of farnesyl diphosphate synthase. Our observations show that minor changes to the structure of bisphosphonates allow inhibition of more than one enzyme in the mevalonate pathway and suggest that loss of protein prenylation due to inhibition of more than one enzyme in the mevalonate pathway may lead to an increase in antiresorptive potency compared to bisphosphonates that only inhibit farnesyl diphosphate synthase.