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Objective: The objective of this systematic review and meta-analysis was to report the safety of intra-articular hyaluronic acid (IAHA) in patients with symptomatic knee osteoarthritis (OA)., Methods: We identified randomized controlled trials reporting the safety of IAHA versus IA saline in adults with symptomatic knee OA. Main safety outcomes were adverse events (AEs), local AEs, serious adverse events (SAEs), study withdrawals, and AE-related study withdrawals., Results: A total of 35 randomized controlled trials with 38 group comparisons comprising 8,078 unique patients (IAHA: 4,295, IA saline: 3,783) were included in the meta-analysis. Comparing IAHA with IA saline over a median of 6 months follow-up, there were no differences in the risk of AEs (42.4% vs. 39.7%, risk ratio [RR] = 1.01, 95% CI = 0.96-1.07, P = 0.61), SAEs (1.8% vs. 1.2%, RR = 1.44, 95% CI = 0.91-2.26, P =0.12), study withdrawals (12.3% vs. 12.7%, RR = 0.99, 95% CI = 0.87-1.12, P = 0.83), or AE-related study withdrawals (2.7% vs. 2.1%, RR = 1.37, 95% CI = 0.97-1.93, P = 0.08). Local AEs, all of which were nonserious, were more common with IAHA vs. IA saline (14.5% vs. 11.7%, RR = 1.21, 95% CI = 1.07-1.36, P = 0.003) and typically resolved within days., Conclusion: IAHA was shown to be safe for use in patients with symptomatic knee OA. Compared with IA saline, IAHA is associated with an increased risk of nonserious, transient local reactions. There was no evidence to suggest any additional safety risks of IAHA.

Macrophage cytokine production is regulated by neural signals, for example in the inflammatory reflex. Signals in the vagus and splenic nerves are relayed by choline acetyltransferase + T cells that release acetylcholine, the cognate ligand for alpha7 nicotinic acetylcholine subunit-containing receptors (α7nAChR), and suppress TNF release in macrophages. Here, we observed that electrical vagus nerve stimulation with a duration of 0.1-60 s significantly reduced systemic TNF release in experimental endotoxemia. This suppression of TNF was sustained for more than 24 h, but abolished in mice deficient in the α7nAChR subunit. Exposure of primary human macrophages and murine RAW 264.7 macrophage-like cells to selective ligands for α7nAChR for 1 h in vitro attenuated TNF production for up to 24 h in response to endotoxin. Pharmacological inhibition of adenylyl cyclase (AC) and knockdown of adenylyl cyclase 6 (AC6) or c-FOS abolished cholinergic suppression of endotoxin-induced TNF release. These findings indicate that action potentials in the inflammatory reflex trigger a change in macrophage behavior that requires AC and phosphorylation of the cAMP response element binding protein (CREB). These observations further our mechanistic understanding of neural regulation of inflammation and may have implications for development of bioelectronic medicine treatment of inflammatory diseases.

Aim: To determine the efficacy of platelet-rich plasma (PRP) injections for symptomatic tendinopathy., Design: Systematic review of randomised, injection-controlled trials with meta-analysis., Data Sources: Systematic searches of MEDLINE and EMBASE, supplemented by manual searches., Eligibility Criteria for Selecting Studies: Randomised controlled trials with 3 months minimum follow-up that evaluated pain reduction with PRP versus control (saline, local anaesthetic, corticosteroid) injections in patients with symptomatic tendinopathy., Results: A total of 16 randomised controlled trials (18 groups) of PRP versus control were included. Median sample size was 35 patients, a study size that would require an effect size ≥1.0 to achieve statistical significance. PRP was more efficacious than control in reducing tendinopathy pain, with an effect size of 0.47 (95% CI 0.22 to 0.72, p<0.001), signifying a moderate treatment effect. Heterogeneity among studies was moderate (I 2 =67%, p<0.001). In subgroup analysis and meta-regression, studies with a higher proportion of female patients were associated with greater treatment benefits with PRP., Conclusions: Injection of PRP is more efficacious than control injections in patients with symptomatic tendinopathy., Competing Interests: Competing interests: None declared.

Objective: Investigation of osteoarthritis (OA) risk alleles suggests that reduced levels of growth and differentiation factor-5 (GDF5) may be a precipitating factor in OA. We hypothesized that intra-articular recombinant human GDF5 (rhGDF5) supplementation to the OA joint may alter disease progression., Methods: A rat medial meniscus transection (MMT) joint instability OA model was used. Animals received either one intra-articular injection, or two or three bi-weekly intra-articular injections of either 30 μg or 100 μg of rhGDF5 beginning on day 21 post surgery after structural pathology had been established. Nine weeks after MMT surgery, joints were processed for histological analysis following staining with toluidine blue. Control groups received intra-articular vehicle injections, comprising a glycine-buffered trehalose solution. OA changes in the joint were evaluated using histopathological end points that were collected by a pathologist who was blinded to treatment., Results: Intra-articular rhGDF5 supplementation reduced cartilage lesions on the medial tibial plateau in a dose-dependent manner when administered therapeutically to intercept OA disease progression. A single 100 μg rhGDF5 injection on day 21 slowed disease progression at day 63. A similar effect was achieved with two bi-weekly injections of 30 μg. Two bi-weekly injections of 100 μg or three bi-weekly injections of 30 μg stopped progression of cartilage lesions. Importantly, three biweekly injections of 100 μg rhGDF5 stimulated significant cartilage repair., Conclusions: Intra-articular rhGDF5 supplementation can prevent and even reverse OA disease progression in the rat MMT OA model. Collectively, these results support rhGDF5 supplementation as an intra-articular disease modifying OA therapy., (Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Therapeutic success of platelet-rich plasma (PRP) may vary based on the composition and preparation method. The objective of this study was to evaluate the cellular components of platelet concentrates produced by a leucocyte-rich (LR-PRP) and a leucocyte-poor PRP systems (LP-PRP)., Methods: Parameters evaluated included platelet recovery, platelet concentration, red blood cell (RBC) and white blood cell (WBC) composition, platelet growth factor release and stimulation of human tendon cell proliferation in vitro., Results: Platelet recoveries were 52% for LP-PRP and 89% for LR-PRP. LR-PRP demonstrated greater reproducibility with a 4.2% coefficient of variation (CV) compared with 19.4% for LP-PRP (p<0.001). LR-PRP demonstrated a greater increase in platelet concentration (7.9-fold) than LP-PRP (2.2-fold; p<0.001). LP-PRP showed 5.0-fold reductions in WBCs, while LR-PRP showed a 4.0-fold increase (p<0.001). LP-PRP reduced RBCs to a haematocrit of 0.25, while LR-PRP reduced haematocrit to 11.8. LP-PRP did not coagulate robustly on reactivation with CaCl 2 , and released significantly lower levels of epidermal growth factor (EGF) and transforming growth factor β1 (TGF-β1) than whole blood (p<0.03). LP-PRP also did not stimulate tendon cell proliferation greater than whole blood. In contrast, LR-PRP showed increases in each growth factor on activation with CaCl 2 (p<0.01) and stimulated greater proliferation (p<0.05) compared with whole blood. Forced activation of LP-PRP with exogenous thrombin rescued the coagulation deficiency and induced greater growth factor release than comparable whole blood (p<0.03)., Conclusions: These data suggest that non-platelet cellular components in platelet concentrates are important for proper platelet function, including thrombin generation, growth factor release and clot retraction.

The excessive release of cytokines by the immune system contributes importantly to the pathogenesis of inflammatory diseases. Recent advances in understanding the biology of cytokine toxicity led to the discovery of the "cholinergic anti-inflammatory pathway," defined as neural signals transmitted via the vagus nerve that inhibit cytokine release through a mechanism that requires the alpha7 subunit-containing nicotinic acetylcholine receptor (alpha7nAChR). Vagus nerve regulation of peripheral functions is controlled by brain nuclei and neural networks, but despite considerable importance, little is known about the molecular basis for central regulation of the vagus nerve-based cholinergic anti-inflammatory pathway. Here we report that brain acetylcholinesterase activity controls systemic and organ specific TNF production during endotoxemia. Peripheral administration of the acetylcholinesterase inhibitor galantamine significantly reduced serum TNF levels through vagus nerve signaling, and protected against lethality during murine endotoxemia. Administration of a centrally-acting muscarinic receptor antagonist abolished the suppression of TNF by galantamine, indicating that suppressing acetylcholinesterase activity, coupled with central muscarinic receptors, controls peripheral cytokine responses. Administration of galantamine to alpha7nAChR knockout mice failed to suppress TNF levels, indicating that the alpha7nAChR-mediated cholinergic anti-inflammatory pathway is required for the anti-inflammatory effect of galantamine. These findings show that inhibition of brain acetylcholinesterase suppresses systemic inflammation through a central muscarinic receptor-mediated and vagal- and alpha7nAChR-dependent mechanism. Our data also indicate that a clinically used centrally-acting acetylcholinesterase inhibitor can be utilized to suppress abnormal inflammation to therapeutic advantage.

Severe sepsis is the leading cause of mortality in intensive care units. The limited ability of current therapies to reduce sepsis mortality rates has fueled research efforts for the development of novel treatment strategies. Through the close collaboration between clinicians and scientists, progress can be seen in the struggle to develop effective therapeutic approaches for the treatment of sepsis and other immune and inflammatory disorders. Indeed, significant advances in intensive care, such as lung protective mechanical ventilation, improved antibiotics, and superior monitoring of systemic perfusion, are improving patient survival. Nonetheless, specific strategies that target the pathophysiological disorders in sepsis patients are essential to further improve clinical outcomes. This article reviews current clinical management approaches and experimental interventions that target pleiotropic or late-acting inflammatory mediators like caspases, C5a, MIF, and HMGB1, or the body's endogenous inflammatory control mechanisms such as the cholinergic anti-inflammatory pathway. These inflammatory mediators and anti-inflammatory mechanisms, respectively, show significant potential for the development of new experimental therapies for the treatment of severe sepsis and other infectious and inflammatory disorders.

The alpha7 subunit-containing nicotinic acetylcholine receptor (alpha7nAChR) is an essential component in the vagus nerve-based cholinergic anti-inflammatory pathway that regulates the levels of TNF, high mobility group box 1 (HMGB1), and other cytokines during inflammation. Choline is an essential nutrient, a cell membrane constituent, a precursor in the biosynthesis of acetylcholine, and a selective natural alpha7nAChR agonist. Here, we studied the anti-inflammatory potential of choline in murine endotoxemia and sepsis, and the role of the alpha7nAChR in mediating the suppressive effect of choline on TNF release. Choline (0.1-50 mM) dose-dependently suppressed TNF release from endotoxin-activated RAW macrophage-like cells, and this effect was associated with significant inhibition of NF-kappaB activation. Choline (50 mg/kg, intraperitoneally [i.p.]) treatment prior to endotoxin administration in mice significantly reduced systemic TNF levels. In contrast to its TNF suppressive effect in wild type mice, choline (50 mg/kg, i.p.) failed to inhibit systemic TNF levels in alpha7nAChR knockout mice during endotoxemia. Choline also failed to suppress TNF release from endotoxin-activated peritoneal macrophages isolated from alpha7nAChR knockout mice. Choline treatment prior to endotoxin resulted in a significantly improved survival rate as compared with saline-treated endotoxemic controls. Choline also suppressed HMGB1 release in vitro and in vivo, and choline treatment initiated 24 h after cecal ligation and puncture (CLP)-induced polymicrobial sepsis significantly improved survival in mice. In addition, choline suppressed TNF release from endotoxin-activated human whole blood and macrophages. Collectively, these data characterize the anti-inflammatory efficacy of choline and demonstrate that the modulation of TNF release by choline requires alpha7nAChR-mediated signaling.

The autonomic nervous system maintains homeostasis through its sympathetic and parasympathetic divisions. During infection, cells of the immune system release cytokines and other mediators that cause fever, hypotension, and tissue injury. Although the effect of cytokines on the nervous system has been known for decades, only recently has it become evident that the autonomic nervous system, in turn, regulates cytokine production through neural pathways. We have previously shown that efferent vagus nerve signals regulate cytokine production through the nicotinic acetylcholine receptor subunit alpha7, a mechanism termed "the cholinergic antiinflammatory pathway." Here, we show that vagus nerve stimulation during endotoxemia specifically attenuates TNF production by spleen macrophages in the red pulp and the marginal zone. Administration of nicotine, a pharmacological agonist of alpha7, attenuated TNF immunoreactivity in these specific macrophage subpopulations. Synaptophysin-positive nerve endings were observed in close apposition to red pulp macrophages, but they do not express choline acetyltransferase or vesicular acetylcholine transporter. Surgical ablation of the splenic nerve and catecholamine depletion by reserpine indicate that these nerves are catecholaminergic and are required for functional inhibition of TNF production by vagus nerve stimulation. Thus, the cholinergic antiinflammatory pathway regulates TNF production in discrete macrophage populations via two serially connected neurons: one preganglionic, originating in the dorsal motor nucleus of the vagus nerve, and the second postganglionic, originating in the celiac-superior mesenteric plexus, and projecting in the splenic nerve.

High-mobility group box (HMGB)-1 was recently identified as a lethal mediator of severe sepsis and represents a novel group of intracellular proteins that function as inflammatory cytokines when released into the extracellular milieu. From a clinical perspective, extracellular HMGB-1 can cause multiple organ failure and contribute to the pathogenesis of diverse disorders including sepsis, cardiovascular shock, rheumatoid arthritis, diabetes, and cancer. HMGB-1 has been proven to be a successful therapeutic target in experimental models of diverse infectious and inflammatory diseases, and these findings have renewed the clinical interest of specific cytokine inhibitors. However, little is known about the molecular mechanisms underlying the cytokine activity of HMGB-1 and its contribution to infection and inflammation. This article analyzes the value of HMGB-1 as a therapeutic target for the treatment of diverse infectious and inflammatory disorders and its interest for human clinical trials.

In the budding yeast Saccharomyces cerevisiae, the regulation of phosphatidylinositol 3-phosphate [PtdIns(3)P] is an essential function shared by the myotubularin-related phosphatase Ymr1p and the synaptojanin-like phosphatases Sjl2p and Sjl3p. The aim of this study was to gain further insight into the mechanisms underlying the toxicity of PtdIns(3)P accumulation in ymr1Delta sjl2Delta sjl3Delta mutant cells. We conducted a genetic screen to isolate genes that, when overexpressed, would rescue the conditional lethality of ymr1Delta sjl2Delta sjl3Delta triple-mutant cells expressing YMR1 from the dextrose-repressible GAL1 promoter. This approach identified 17 genes that promoted growth of the triple mutant on media containing dextrose. Interestingly, the most frequently isolated gene product was a truncated form of PKC1 (Pkc1-T615) that lacked the C-terminal kinase domain. This Pkc1-T615 fragment also rescued the lethality of ymr1ts sjl2Delta sjl3Delta cells at restrictive temperature, and further mapping of the rescuing activity showed that the N-terminal Rho1-GTP-interacting HR1 domains (Pkc1-T242) were sufficient. This indicated that the PKC1 fragments might act by interfering with Rho1-GTP signal propagation. Consistent with this, deletion of the ROM2 gene, which encodes a major Rho1p guanine-nucleotide exchange factor, bypassed the lethal effect of PtdIns(3)P accumulation in ymr1Delta sjl2Delta sjl3Delta triple-mutant cells. Furthermore, cells deficient in phosphoinositide 3-phosphatase (PI 3-phosphatase) activity were defective for Rho1p/Pkc1p pathway regulation, which included an inability of these cells to adapt to heat stress. Taken together, the results of this study indicated that aberrant Rho1p/Pkc1p signaling contributes to the lethal effects of PtdIns(3)P accumulation in cells deficient in PI 3-phosphatase activity.

The requirement of Vps34p, the sole phosphatidylinositol (PI) 3-kinase in Saccharomyces cerevisiae, for protein sorting to the vacuole in yeast has exemplified the essential role for phosphoinositides, phosphorylated derivatives of PI, in membrane trafficking. To better understand mechanisms that regulate PI 3-phosphate [PI(3)P]-mediated signaling, the role of the yeast myotubularin-related PI(3)P phosphatase Ymr1p was investigated. We found that Ymr1p and the synaptojanin-like phosphatase Sjl3p function as key regulators of the localization and levels of PI(3)P. Our data indicated that the ymr1Delta sjl3Delta double mutant aberrantly accumulated PI(3)P and demonstrated a steady-state redistribution of this lipid that leads to enrichment on the vacuolar membrane. This resulted in vacuole protein sorting defects, vacuolar fragmentation, and the misregulation of PI(3)P-specific effectors. Triple deletion of YMR1, SJL2, and SJL3 was lethal, suggesting an essential requirement for phosphatase-mediated PI(3)P regulation. Consistent with this, growth was restored to a ymr1Delta sjl2Delta sjl3Delta triple mutant by a PI(3)P-targeted Sac1p domain chimera (GFP-Sac1DeltaC-FYVE(EEA1)) that returned PI(3)P to levels comparable with wild-type cells. Together, this study demonstrated that Ymr1p, a myotubularin phosphatase family member, functions in the control of PI(3)P-dependent signaling and the maintenance of endosomal system integrity. In addition, this work defined an essential overlapping role for lipid phosphatases in the regulation of 3' phosphoinositides in yeast.