Platelet‐Rich Plasma Increases Growth and Motility of Adipose Tissue‐Derived Mesenchymal Stem Cells and Controls Adipocyte Secretory Function

Adipose tissue‐derived mesenchymal stem cells (Ad‐MSC) and platelet derivatives have been used alone or in combination to achieve regeneration of injured tissues. We have tested the effect of platelet‐rich plasma (PRP) on Ad‐MSC and adipocyte function. PRP increased Ad‐MSC viability, proliferation rate and G1‐S cell cycle progression, by at least 7‐, 2‐, and 2.2‐fold, respectively, and reduced caspase 3 cleavage. Higher PRP concentrations or PRPs derived from individuals with higher platelet counts were more effective in increasing Ad‐MSC growth. PRP also accelerated cell migration by at least 1.5‐fold. However, PRP did not significantly affect mature adipocyte viability, differentiation and expression levels of PPAR‐γ and AP‐2 mRNAs, while it increased leptin production by 3.5‐fold. Interestingly, PRP treatment of mature adipocytes also enhanced the release of Interleukin (IL)‐6, IL‐8, IL‐10, Interferon‐γ, and Vascular Endothelial Growth Factor. Thus, data are consistent with a stimulatory effect of platelet derivatives on Ad‐MSC growth and motility. Moreover, PRP did not reduce mature adipocyte survival and increased the release of pro‐angiogenic factors, which may facilitate tissue regeneration processes. © 2015 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc. J. Cell. Biochem. 116: 2408–2418, 2015. © 2015 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.     

IL-17 mediates estrogen-deficient osteoporosis in an Act1-dependent manner

Estrogen-deficient osteoporosis may be an inflammatory disorder and we therefore asked if IL-17 participates in its pathogenesis. Deletion of the principal IL-17 receptor (IL-17RA) protects mice from ovariectomy (OVX)-induced bone loss. Further supporting a central role of IL-17 in its pathogenesis, OVX-induced osteoporosis is prevented by a blocking antibody targeting the cytokine. IL-17 promotes osteoclastogenesis by stimulating RANK ligand (RANKL) expression by osteoblastic cells, mediated by the IL-17RA SEFIR/TILL domain. Estrogen deprivation, however does not enhance IL-17RA mRNA expression by osteoblasts or in bone, but augments that of Act1, an IL-17RA-interacting protein and signaling mediator. Similar to IL-17RA(-/-) mice, those lacking Act1 are protected from OVX-induced bone loss. Also mirroring IL-17RA-deficiency, absence of Act1 in osteoblasts, but not osteoclasts, impairs osteoclastogenesis via dampened RANKL expression. Transduction of WT Act1 into Act1(-/-) osteoblasts substantially rescues their osteoclastogenic capacity. The same construct, however, lacking its E3 ligase U-box or its SEFIR domain, which interacts with its counterpart in IL-17RA, fails to do so. Estrogen deprivation, therefore, promotes RANKL expression and bone resorption in association with upregulation of the IL-17 effector, Act1, supporting the concept that post-menopausal osteoporosis is a disorder of innate immunity.     

Antioxidant supplementation lowers circulating IGF-1 but not F2-isoprostanes immediately following anterior cruciate ligament surgery

Abstract

Interleukin (IL)-10 is an anti-inflammatory cytokine that suppresses pro-inflammatory cytokines. We previously demonstrated that supplementation with vitamins E and C ameliorated the increase in IL-10 immediately following anterior cruciate ligament (ACL) surgery in the absence of other cytokine perturbations. Since both oxidative stress and insulin-like growth factor-1 (IGF-1) can modulate IL-10 concentrations, the mechanisms for these changes warranted further investigation. Our objective was to evaluate the mechanism for the IL-10 decrease following ACL surgery. This study consisted of randomized, double-blind, placebo-controlled experimental design. Subjects were randomly assigned to daily supplementation with either: (i) antioxidants (AO; vitamins E [α-tocopherol] and C [ascorbic acid]; n = 10); or (ii) matching placebos (PL; n = 10). Supplementation started ～2 weeks prior to surgery (baseline) and concluded 3 months after surgery. Subjects provided six fasting blood samples at: (i) baseline; (ii) immediately pre-surgery (Pre); (iii) 90 min; (iv) 72 h; (v) 7 days; and (vi) 3 months post-surgery. α-Tocopherol, ascorbic acid, F₂-isoprostane and IGF-1 concentrations were measured in each blood sample. At 90 min relative to other times, plasma F₂-isoprostane concentrations were significantly (P < 0.05) elevated in both groups, while at 90 min IGF-1 was significantly (P < 0.05) lower in the AO compared to the PL group. The changes in IGF-1 at 90 min relative to baseline were correlated (P < 0.0001) with the changes in IL-10. The decrease in IL-10 observed in the AO group is likely dependent on the decrease IGF-1 since lipid peroxidation was unchanged between the two groups.     

Fluid pressure induces osteoclast differentiation comparably to titanium particles but through a molecular pathway only partly involving TNFα

In contrast to the well-understood inflammatory pathway driven by TNFα, by which implant-derived particles induce bone resorption, little is known about the process in which loosening is generated as a result of force-induced mechanical stimulus at the bone-implant interface. Specifically, there is no knowledge as to what cells or signaling pathways couple mechanical stimuli to bone resorption in context of loosening. We hypothesized that different stimuli, i.e., fluid flow versus wear particles, act through different cytokine networks for activation and localization of osteoclasts. By using an animal model in which osteoclasts and bone resorption were induced by fluid pressure or particles, we were able to detect distinct differences in osteoclast localization and inflammatory gene expression between fluid pressure and titanium particles. Fluid pressure recruits and activates osteoclasts with bone marrow contact away from the fluid pressure exposure zone, whereas titanium particles recruit and activate osteoclasts in areas in direct contact to particles. Fluid pressure induced weaker expression of the selected inflammatory related genes, although the eventual degree of osteoclast induction was similar in both models. Using TNFαRa (4 mg/kg) (Enbrel) and dexamethasone (2 mg/kg) as specific and more general suppressors of inflammation we showed that the TNFαRa failed to generate statistically impaired osteoclast generation while dexamethasone was much more potent. These results demonstrate that fluid pressure induces osteoclasts at a different localization than titanium particles by a molecular pathway less associated with TNFα and the innate system, which open up for other pathways controlling pressure induced osteoclastogenesis.     

The inhibition of hyaluronan degradation reduced pro-inflammatory cytokines in mouse synovial fibroblasts subjected to collagen-induced arthritis

Hyaluronan (HA) degradation produces small oligosaccharides that are able to increase pro-inflammatory cytokines in rheumatoid arthritis synovial fibroblasts (RASF) by activating both CD44 and the toll-like receptor 4 (TLR-4). CD44 and TLR-4 stimulation in turn activate the NF-kB that induces the production of pro-inflammatory cytokines. Degradation of HA occurs via two mechanisms: one exerted by reactive oxygen species (ROS) and one controlled by different enzymes in particular hyaluronidases (HYALs). We aimed to investigate the effects of inhibiting HA degradation (which prevents the formation of small HA fragments) on synovial fibroblasts obtained from normal DBA/J1 mice (NSF) and on synovial fibroblasts (RASF) obtained from mice subjected to collagen induced arthritis (CIA), both fibroblast types stimulated with tumor necrosis factor alpha (TNF-α). TNF-α stimulation produced high mRNA expression and the related protein production of CD44 and TLR-4 in both NSF and RASF, and activation of NF-kB was also found in all fibroblasts. TNF-α also up-regulated the inflammatory cytokines, interleukin-1beta (IL-1beta) and interleukin-6 (IL-6), and other pro-inflammatory mediators, such as matrix metalloprotease-13 (MMP-13), inducible nitric oxide synthase (iNOS), as well as HA levels and small HA fragment production. Treatment of RASF with antioxidants and specific HYAL1, HYAL2, and HYAL3 small interference RNA (siRNAs) significantly reduced TLR-4 and CD44 increase in the mRNA expression and the related protein synthesis, as well as the release of inflammatory mediators up-regulated by TNF-α. These data suggest that the inhibition of HA degradation during arthritis may contribute to reducing TLR-4 and CD44 activation and the inflammatory mediators response.