Pharmacological blockade of the interleukin-1 receptor suppressed Escherichia coli lipopolysaccharide-induced neuroinflammation in preterm fetal sheep.

BACKGROUND: Intraamniotic inflammation is associated with preterm birth, especially in cases occurring before 32 weeks' gestation, and is causally linked with an increased risk for neonatal mortality and morbidity. Targeted anti-inflammatory interventions may assist in improving the outcomes for pregnancies impacted by intrauterine inflammation. Interleukin-1 is a central upstream mediator of inflammation. Accordingly, interleukin-1 is a promising candidate target for intervention therapies and has been targeted previously using the interleukin-1 receptor antagonist, anakinra. Recent studies have shown that the novel, noncompetitive, allosteric interleukin-1 receptor inhibitor, rytvela, partially resolved inflammation associated with preterm birth and fetal injury. In this study, we used a preterm sheep model of chorioamnionitis to investigate the anti-inflammatory efficacy of rytvela and anakinra, administered in the amniotic fluid in the setting of intraamniotic Escherichia coli lipopolysaccharide exposure. OBJECTIVE: We hypothesized that both rytvela and anakinra would reduce lipopolysaccharide-induced intrauterine inflammation and protect the fetal brain. STUDY DESIGN: Ewes with a singleton fetus at 105 days of gestation (term is ∼150 days) were randomized to one of the following groups: (1) intraamniotic injections of 2 mL saline at time=0 and time=24 hours as a negative control group (saline group, n=12); (2) intraamniotic injection of 10 mg Escherichia coli lipopolysaccharide in 2 mL saline and intraamniotic injections of 2 mL saline at time=0 hours and time=24 hours as an inflammation positive control group (lipopolysaccharide group, n=11); (3) intraamniotic injection of Escherichia coli lipopolysaccharide in 2 mL saline and intraamniotic injections of 2.5 mg rytvela at time=0 hours and time=24 hours to test the anti-inflammatory efficacy of rytvela (lipopolysaccharide + rytvela group, n=10); or (4) intraamniotic injection of Escherichia coli lipopolysaccharide in 2 mL saline and intraamniotic injections of 100 mg anakinra at time=0 hours and time=24 hours to test the anti-inflammatory efficacy of anakinra (lipopolysaccharide + anakinra group, n=12). Amniotic fluid was sampled at time 0, 24, and 48 hours (ie, at each intervention and at delivery). Fetal umbilical cord blood was collected at delivery for differential blood counts and chemical studies. Inflammation was characterized by the analysis of fetal tissue cytokine and chemokine levels using quantitative polymerase chain reaction, enzyme-linked inmmunosorbent assay, and histology. The primary study outcome of interest was the assessment of anakinra and rytvela brain-protective effects in the setting of Escherichia coli lipopolysaccharide-induced intrauterine inflammation. Secondary outcomes of interest were to assess protection from fetal and intrauterine (ie, amniotic fluid, chorioamnion) inflammation. RESULTS: Intraamniotic administration of lipopolysaccharide caused inflammation of the fetal lung, brain, and chorioamnionitis in preterm fetal sheep. Relative to treatment with saline only in the setting of lipopolysaccharide exposure, intraamniotic administration of both rytvela and anakinra both significantly prevented periventricular white matter injury, microglial activation, and histologic chorioamnionitis. Anakinra showed additional efficacy in inhibiting fetal lung myeloperoxidase activity, but its use was associated with metabolic acidaemia and reduced fetal plasma insulin-like growth factor-1 levels at delivery. CONCLUSION: Intraamniotic administration of rytvela or anakinra significantly inhibited fetal brain inflammation and chorioamnionitis in preterm fetal sheep exposed to intraamniotic lipopolysaccharide. In addition, anakinra treatment was associated with potential negative impacts on the developing fetus.

Effects of high intensity interval training on cardiorespiratory fitness and salivary levels of IL-8, IL-1ra, and IP-10 in adults with asthma and non-asthma controls.

OBJECTIVE: The purpose of this study was to determine whether high intensity interval training (HIIT) would lead to improvements in 1) maximal VO2, VE, VE/VCO2, and VE/MVV, and/or 2) resting salivary concentrations of pro-inflammatory markers Interleukin (IL-8), interferon-gamma-inducible-protein (CXCL10/IP-10)) and anti-inflammatory marker IL-1 receptor antagonist (IL-1ra) in adults with well-controlled asthma compared to non-asthma controls. METHODS: Participants completed a maximal exercise test at the beginning (T1) and end (T2) of a 6-week HIIT intervention; saliva samples were obtained at the beginning and 30 min following the first (T1) and last (T2) exercise session. RESULTS: Adults with asthma (n = 20; age: 21.4 ± 2.4 years) and non-asthma controls (n = 12; age: 22.5 ± 3.4 years) completed the intervention. VO2max increased from T1 to T2 in both groups (asthma T1 32.9 ± 8, T2 38.6 ± 8.2 ml/kg/min; controls T1 34.5 ± 11.8, T2 38.9 ± 12.3 ml/kg/min). VEmax also increased in both groups (asthma T1 97.7, T2 110.8 units, p < 0.001, hp2 = <0.04; control T1 106.3, T2 118.1, p < 0.001, hp2 0.02). An increase in VE/VCO2 (F(1, 10)=22.11, p = 0.001) and VE/MVV (F(1, 10) = 111.30, p < 0.001) was observed in the control group; no differences were observed in the asthma group. No differences in IL-8 or IL-1ra were observed between groups. In the asthma group, resting salivary IP-10 concentrations significantly decreased from T1 (0.025 pg/ug protein) to T2 (0.015 pg/ug protein, p = 0.039, hp2 = 0.3 (moderate effect)). CONCLUSION: A 6-week HIIT intervention led to a similar increase in VO2max and VEmax in those with and without asthma, and a decrease in resting salivary IP-10 levels among adults with asthma.

Investigating the release of inflammatory cytokines in a human model of incontinence-associated dermatitis.

Incontinence-associated dermatitis (IAD) is a painful complication in elderly patients, leading to reduced quality of life. Despite recent attention, its underlying inflammatory mechanisms remain poorly understood. This study was designed to quantify the release of inflammatory cytokines in a human model of IAD. The left volar forearm of ten healthy volunteers was exposed to synthetic urine and synthetic faeces for 2 h, simulating the effects of urinary and faecal incontinence, respectively, and the subsequent cytokine response compared to that of an untreated control site. Inflammatory cytokines were collected using both the Sebutape® absorption method and dermal microdialysis and quantified using immunoassays. Results from the former demonstrated an upregulation in IL-1α, IL-1RA and TNF-α. Synthetic urine caused a higher median increase in IL-1α from baseline compared to synthetic faeces, whereas synthetic faeces were associated with significantly higher median TNF-α levels compared to synthetic urine (p = 0.01). An increase in IL-1α/IL-1RA ratio was also observed with significant differences evident following exposure to synthetic urine (p = 0.047). Additionally, microdialysis revealed a time-dependent increase in IL-1ß and IL-8 following exposure of up to 120 min to synthetic urine and synthetic faeces, respectively. This study demonstrated the suitability of both sampling approaches to recover quantifiable cytokine levels in biofluids for the assessment of skin status following exposure to synthetic fluids associated with incontinence. Findings suggest some differences in the inflammatory mechanisms of IAD, depending on moisture source, and the potential of the cytokines, IL-1α and TNF-α, as responsive markers of early skin damage caused by incontinence.

Evaluating the effects of sedentary behaviour on plantar skin health in people with diabetes.

BACKGROUND: Diabetes-Related Foot Ulcers (DRFUs) are a common and devastating consequence of Diabetes Mellitus and are associated with high morbidity, mortality, social and economic costs. Whilst peak plantar pressures during gait are implicated cited as a major contributory factor, DRFU occurrence has also been associated with increased periods of sedentary behaviour. The present study was designed aimed to assess the effects of sitting postures on plantar tissue health. METHODS: After a period of acclimatisation, transcutaneous oxygen tensions (TCPO2) and inflammatory cytokines (IL-1α and IL-1RA) were measured at the dorsal and plantar aspects of the forefoot before, during and after a 20-min period of seated-weight-bearing in participants with diabetes (n = 11) and no diabetes (n = 10). Corresponding interface pressures at the plantar site were also measured. RESULTS: During weight-bearing, participants with diabetes showed increases in tissue ischaemia which were linearly correlated proportional to plantar pressures (Pearson's r = 0.81; p < 0.05). Within the healthy group, no such correlation was evident (p > 0.05). There were also significant increases in post seated weight-bearing values for ratio for IL-1α and IL-1RA, normalised to total protein, post seated weight-bearing in participants with diabetes compared to healthy controls. CONCLUSION: This study shows that prolonged sitting may be detrimental to plantar skin health. It highlights the need to further examine the effects of prolonged sitting in individuals, who may have a reduced tolerance to loading in the plantar skin and soft tissues.

Development of a biomarker mortality risk model in acute respiratory distress syndrome.

BACKGROUND: There is a compelling unmet medical need for biomarker-based models to risk-stratify patients with acute respiratory distress syndrome. Effective stratification would optimize participant selection for clinical trial enrollment by focusing on those most likely to benefit from new interventions. Our objective was to develop a prognostic, biomarker-based model for predicting mortality in adult patients with acute respiratory distress syndrome. METHODS: This is a secondary analysis using a cohort of 252 mechanically ventilated subjects with the diagnosis of acute respiratory distress syndrome. Survival to day 7 with both day 0 (first day of presentation) and day 7 sample availability was required. Blood was collected for biomarker measurements at first presentation to the intensive care unit and on the seventh day. Biomarkers included cytokine-chemokines, dual-functioning cytozymes, and vascular injury markers. Logistic regression, latent class analysis, and classification and regression tree analysis were used to identify the plasma biomarkers most predictive of 28-day ARDS mortality. RESULTS: From eight biologically relevant biomarker candidates, six demonstrated an enhanced capacity to predict mortality at day 0. Latent-class analysis identified two biomarker-based phenotypes. Phenotype A exhibited significantly higher plasma levels of angiopoietin-2, macrophage migration inhibitory factor, interleukin-8, interleukin-1 receptor antagonist, interleukin-6, and extracellular nicotinamide phosphoribosyltransferase (eNAMPT) compared to phenotype B. Mortality at 28 days was significantly higher for phenotype A compared to phenotype B (32% vs 19%, p = 0.04). CONCLUSIONS: An adult biomarker-based risk model reliably identifies ARDS subjects at risk of death within 28 days of hospitalization.

Propofol vs desflurane on the cytokine, matrix metalloproteinase-9, and heme oxygenase-1 response during living donor liver transplantation: A pilot study.

BACKGROUND: We investigated the effects of propofol vs desflurane on ischemia and reperfusion injury (IRI)-induced inflammatory responses, especially in matrix metalloproteinase-9 (MMP-9) downregulation and heme oxygenase-1 (HO-1) upregulation, which may result in different clinical outcomes in liver transplant recipients. METHODS: Fifty liver transplant recipients were randomized to receive propofol-based total intravenous anesthesia (TIVA group, n = 25) or desflurane anesthesia (DES group, n = 25). We then measured the following: perioperative serum cytokine concentrations (interleukin 1 receptor antagonist [IL-1RA], IL-6, IL-8, and IL-10); MMP-9 and HO-1 mRNA expression levels at predefined intervals. Further, postoperative outcomes were compared between the 2 groups. RESULTS: The TIVA group showed a significant HO-1 level increase following the anhepatic phase and a significant MMP-9 reduction after reperfusion, in addition to a significant increase in IL-10 levels after the anhepatic phase and IL-1RA levels after reperfusion. Compared to DES patients, TIVA patients showed a faster return of the international normalized ratio to normal values, lower plasma alanine aminotransferase concentrations 24 hours after transplantation, and fewer patients developing acute lung injury. Moreover, compared with DES patients, TIVA patients showed a significant reduction in serum blood lactate levels. However, there were no differences in postoperative outcomes between the two groups. CONCLUSION: Propofol-based TIVA attenuated inflammatory response (elevated IL-1RA and IL-10 levels), downregulated MMP-9 response, and increased HO-1 expression with improved recovery of graft function and better microcirculation compared with desflurane anesthesia in liver transplant recipients.

Changes in Synovial Fluid Biomarker Concentration Before and After ACL Reconstruction.

BACKGROUND: Synovial fluid biomarkers can highlight the molecular milieu associated with knee pathology and have been shown to be significantly different in patients with anterior cruciate ligament (ACL) injuries compared to uninjured controls. The purpose of the current study was to establish how synovial fluid biomarker concentrations change in patients undergoing ACL reconstruction between the immediate preoperative period to the acute postoperative period. METHODS: Patients were prospectively enrolled at the time of surgery from September 2016 to March 2017. Patients who had an operative knee synovial fluid sample obtained at the time of ACL reconstruction and provided a synovial fluid sample at their first postoperative appointment were included. The concentrations of 10 biomarkers were determined using a multiplex magnetic bead immunoassay. Biomarker concentrations before and after surgery were compared using a paired sample t-test. RESULTS: Eight patients with mean age of 33.4 years who underwent isolated ACL reconstruction using a bonepatellar tendon-bone autograft were included. The mean time between surgery and postoperative office visit was 10.4 days. There was a statistically significant increase in the concentrations of interleukin-6 (IL-6, p = 0.014), monocyte chemoattractant protein-1 (MCP-1, p = 0.024), human matrix metalloproteinase 3 (MMP-3, p = 0.00002), macrophage inflammatory protein-1 beta (MIP-1ß, p = 0.006), human interleukin-1 receptor antagonist (IL-1Ra, p = 0.017), and vascular endothelial growth factor (VEGF, p = 0.023) between the time of surgery and the first postoperative visit and a decrease in the concentration of tissue inhibitor of metalloproteinase-2 (p = 0.050). CONCLUSION: The molecular profile of the synovial fluid changes in the early postoperative period following arthroscopic ACL reconstruction. The concentration of proinflammatory markers (such as IL-6, MCP-1, MMP-3, and MIP-1ß) and growth factors including VEGF increases. The concentration of the anti-inflammatory marker tissue inhibitor of metalloproteinase-2 (TIMP-2) appears to decrease postoperatively.

Evaluation of the effect of experimentally induced cartilage defect and intra-articular hyaluronan on synovial fluid biomarkers in intercarpal joints of horses.

BACKGROUND: Inflammatory and degenerative activity inside the joint can be studied in vivo by analysis of synovial fluid biomarkers. In addition to pro-inflammatory mediators, several anabolic and anti-inflammatory substances are produced during the disease process. They counteract the catabolic effects of the pro-inflammatory cytokines and thus diminish the cartilage damage. The response of synovial fluid biomarkers after intra-articular hyaluronan injection, alone or in combination with other substances, has been examined only in a few equine studies. The effects of hyaluronan on some pro-inflammatory mediators, such as prostaglandin E2, have been documented but especially the effects on synovial fluid anti-inflammatory mediators are less studied. In animal models hyaluronan has been demonstrated to reduce pain via protecting nociceptive nerve endings and by blocking pain receptor channels. However, the results obtained for pain-relief of human osteoarthritis are contradictory. The aim of the study was to measure the synovial fluid IL-1ra, PDGF-BB, TGF-ß1 and TNF-α concentrations before and after surgically induced cartilage defect, and following intra-articular hyaluronan injection in horses. Eight Standardbred horses underwent bilateral arthroscopic surgeries of their intercarpal joints under general anaesthesia, and cartilage defect was created on the dorsal edge of the third carpal bone of one randomly selected intercarpal joint of each horse. Five days post-surgery, one randomly selected intercarpal joint was injected intra-articular with 3 mL HA (20 mg/mL). RESULTS: Operation type had no significant effect on the synovial fluid IL-1ra, PDGF-BB, TGF-ß1 and TNF-α concentrations but compared with baseline, synovial fluid IL-1ra and TNF-α concentrations increased. Intra-articular hyaluronan had no significant effect on the biomarker concentrations but a trend of mild improvement in the clinical signs of intra-articular inflammation was seen. CONCLUSIONS: Creation of the cartilage defect and sham-operation lead to an increase of synovial fluid IL-1ra and TNF-α concentrations but changes in concentrations of anabolic growth factors TGF-ß1 and PDGF-BB could not be documented 5 days after the arthroscopy. Intra-articular hyaluronan was well tolerated. Further research is needed to document possible treatment effects of intra-articular hyaluronan on the synovial fluid biomarkers of inflammation and cartilage metabolism.

Dietary flaxseed and tamoxifen affect the inflammatory microenvironment in vivo in normal human breast tissue of postmenopausal women.

BACKGROUND: Anti-oestrogens such as tamoxifen, decrease the risk of breast cancer but are unsuitable for prevention because of their side-effects. Diet modifications may be a breast cancer prevention strategy. Here, we investigated if a diet addition of flaxseed, which can be converted to the phytoestrogen enterolactone by the gut microbiota, exhibited similar effects as tamoxifen on normal human breast tissue in vivo, with special emphasis on inflammatory mediators implicated in cancer progression. SUBJECTS: A total of 28 postmenopausal women were included. Thirteen women added 25 g of ground flaxseed per day and 15 were treated with tamoxifen as an adjuvant for early breast cancer for 6 weeks. Microdialysis of normal breast tissue and, as a control, in subcutaneous abdominal fat was performed for sampling of extracellular proteins in vivo before and after exposures. RESULTS: Enterolactone levels increased significantly after flaxseed. IL-1Ra and IL-1Ra/IL-1ß ratio in the breast increased in a similar fashion after the two different treatments. Flaxseed also increased breast specific levels of IL-1RT2, IL-18 and sST2 and an overall increase of MMP-9. These changes correlated significantly with enterolactone levels. Tamoxifen decreased breast tissue levels of IL-8 and IL-18. None of the treatments induced any changes of IL-1ß, IL-1RT1, IL-18BP, IL-33, IL-6, IL-6RA, MMP-1, MMP-2 and MMP-3. CONCLUSIONS: We conclude that dietary flaxseed and tamoxifen exert both similar and different effects, as listed above, on normal breast tissue in vivo and that a relatively modest diet change can induce significant effects on the breast microenvironment.

Effect of cytomegalovirus reactivation on the time course of systemic host response biomarkers in previously immunocompetent critically ill patients with sepsis: a matched cohort study.

BACKGROUND: Cytomegalovirus (CMV) reactivation in previously immunocompetent critically ill patients is associated with increased mortality, which has been hypothesized to result from virus-induced immunomodulation. Therefore, we studied the effects of CMV reactivation on the temporal course of host response biomarkers in patients with sepsis. METHODS: In this matched cohort study, each sepsis patient developing CMV reactivation between day 3 and 17 (CMV+) was compared with one CMV seropositive patient without reactivation (CMVs+) and one CMV seronegative patient (CMVs-). CMV serostatus and plasma loads were determined by enzyme-linked immunoassays and real-time polymerase chain reaction, respectively. Systemic interleukin-6 (IL-6), IL-8, IL-18, interferon-gamma-induced protein-10 (IP-10), neutrophilic elastase, IL-1 receptor antagonist (RA), and IL-10 were measured at five time points by multiplex immunoassay. The effects of CMV reactivation on sequential concentrations of these biomarkers were assessed in multivariable mixed models. RESULTS: Among 64 CMV+ patients, 45 could be matched to CMVs+ or CMVs- controls or both. The two baseline characteristics and host response biomarker levels at viremia onset were similar between groups. CMV+ patients had increased IP-10 on day 7 after viremia onset (symmetric percentage difference +44% versus -15% when compared with CMVs+ and +37% versus +4% when compared with CMVs-) and decreased IL-1RA (-41% versus 0% and -49% versus +10%, respectively). However, multivariable analyses did not show an independent association between CMV reactivation and time trends of IL-6, IP-10, IL-10, or IL-1RA. CONCLUSION: CMV reactivation was not independently associated with changes in the temporal trends of host response biomarkers in comparison with non-reactivating patients. Therefore, these markers should not be used as surrogate clinical endpoints for interventional studies evaluating anti-CMV therapy.

Kineret protein solution survives ten years.

Proteins are labile and their shelf lives are usually months to few years. Herein we report that human interleukin-1 receptor antagonist (IL-1Ra) solution survives ten years. This provides an interesting case that protein biopharmaceutical aqueous solution could be stored for a decade without losing its quality, which could meet the national security needs for 10 years shelf life of biopharmaceuticals.

Interleukin-1ß, interleukin-1 receptor antagonist, interleukin-6, interleukin-10, and tumor necrosis factor-α levels in CSF and serum in relation to the clinical diversity of Parkinson's disease.

Several parameters representing the clinical diversity of Parkinson's disease (PD), including severity, phenotypes, cognitive decline, anxiety and depression were analyzed to examine the link with interleukin-1ß (IL-1ß), the interleukin-1 receptor antagonist (IL-1RA), IL-6, IL-10, and tumor necrosis factor-α (TNFα) and also to determine the relationship between levels of these factors in serum and cerebrospinal fluid (CSF). Significantly elevated serum IL-1ß and IL-6 and reduced IL-1RA levels were found in the PD group. In CSF and serum, inflammatory factors behaved differently, with increased CSF TNFα indicating rapid PD progression, and increased IL-1ß in serum. A low level of IL-6 was associated with a longer duration of PD. Anxiety, depression, non-tremor phenotype and late-onset PD correlated with a high serum level of IL-10. The serum TNFα level was lower in PD patients with mild cognitive impairment compared to controls. Serum IL-1ß, IL-6 and IL-10 levels correlated with CSF markers.

Salivary biomarker combination prediction model for the diagnosis of periodontitis in a Taiwanese population.

BACKGROUND/PURPOSE: This study aimed at screening the diagnostic potential of salivary biomarkers and pairing them to establish a prediction model with higher accuracy in diagnosing periodontitis in the Taiwanese population. METHODS: Fifty-seven participants were divided into a non-periodontitis group and a periodontitis group. Salivary biomarkers CRP, IL-6, IL-8, IL-1ß, IL-1ra, lactoferrin, MMP-8, MMP-9, PDGF-BB, TNF-α, and VEGF, were analyzed. The potential and reliability of the biomarkers for diagnosing periodontitis were analyzed dichotomously. The correlation of individual biomarkers with periodontitis was assessed using the Spearman rank correlation coefficient with logistic regression. The combinational prediction model was evaluated using the area under the receiver operating characteristic curve (AUC). RESULTS: Regarding individual biomarkers, IL-1ß and MMP-9 levels were significantly higher, and TNF-α was significantly lower in the periodontitis group. IL-1ß, MMP-8, and MMP-9 exhibited a greater odds ratio with statistical significance in the dichotomous table. The combination of three biomarkers yielded AUCs of 0.821-0.853, and the combination of IL-1ß, IL-1ra, and MMP-9 exhibited the highest AUC (0.853), with high sensitivity (73.3%) and specificity (88.9%). CONCLUSION: Regarding individual biomarkers, IL-1ß, MMP-8, and MMP-9 showed potential for identifying patients with periodontitis. The combination of IL-1ß, IL-1ra, and MMP-9 might be feasible for developing a future point-of-care device for diagnosing periodontitis.

Salusin-α Attenuates Inflammatory Responses in Vascular Endothelial Cells.

Atherosclerosis accounts for numerous cardiovascular diseases, and cytokines have a critical role in acceleration or suppression of disease. Salusin-α presents a new class of bioactive peptides that can have anti-atherogenic properties. Therefore, the effects of salusin-α on the expression of some pro- and anti-inflammatory cytokines and on TNF-α-induced inflammatory responses in human umbilical vein endothelial cells (HUVECs) were examined. The involvement of the NF-κB pathway in effects of salusin-α in HUVECs was checked using Bay 11-7082 as an NF-κB inhibitor. The mRNA expression of pro-inflammatory cytokines including IL-6, IL-8, and IL-18 and anti-inflammatory cytokine IL-1Ra was assessed by real-time PCR. The protein levels of cytokines were measured by the ELISA method. Salusin-α suppressed both mRNA and protein expression of pro-inflammatory cytokines and induced mRNA and protein expression of IL-1Ra in HUVECs. Salusin-α suppressed TNF-α-induced inflammatory responses in HUVECs. The down-regulatory or up-regulatory effects of salusin-α on expression of cytokines could not be influenced by Bay 11-7082 pretreatment. Our findings indicate anti-inflammatory effects of salusin-α and suggest a novel peptide-based therapeutic strategy for atherosclerosis.

Cytokine profile in gingival crevicular fluid and plasma of patients with aggressive periodontitis.

OBJECTIVE: This study aimed to determine the content of cytokines in gingival crevicular fluid (GCF) as well as in plasma of Sudanese patients with aggressive periodontitis (AgP) and healthy controls (HC). MATERIALS AND METHODS: Nineteen AgP patients and 19 HC were included. The mean probing pocket depth and clinical attachment level of the GCF sampled sites in patients were both ≥5 mm. The GCF and plasma levels of 27 cytokines were determined using 27-multiplex fluorescent bead-based immunoassays. Ratios were calculated among cytokines of the T-helper cell subsets Th1 and Th2. Descriptive statistics, the Mann-Whitney U-test and Spearman's rho rank correlation coefficient analysis were used. RESULTS: Interferon-γ was the only cytokine found in significantly lower levels in GCF of patients compared with HC. Levels of interleukin (IL)-10, IL-13, IL-1Ra, monocyte chemoattractant protein-1 (MCP-1), regulated on activation normal T-cell expressed and secreted (RANTES), granulocyte-colony-stimulating factor (G-CSF), and granulocyte-macrophage-CSF (GM-CSF) were significantly lower in plasma of AgP compared with HC. The ratios of Th1:Th2 in GCF and Treg:Th17 in plasma were significantly lower in AgP. CONCLUSIONS: The lower levels of cytokines detected systemically in plasma of AgP patients may have an impact on the immune response. The lower ratio of Th1:Th2 cytokines in GCF samples of AgP patients suggests a role for Th2 at the local site of disease.

Platelet-Rich Plasma Powder: A New Preparation Method for the Standardization of Growth Factor Concentrations.

BACKGROUND: Platelet-rich plasma (PRP) is widely used in sports medicine. Available PRP preparations differ in white blood cell, platelet, and growth factor concentrations, making standardized research and clinical application challenging. PURPOSE: To characterize a newly standardized procedure for pooled PRP that provides defined growth factor concentrations. STUDY DESIGN: Controlled laboratory study. METHODS: A standardized growth factor preparation (lyophilized PRP powder) was prepared using 12 pooled platelet concentrates (PCs) derived from different donors via apheresis. Blood samples and commercially available PRP (SmartPrep-2) served as controls (n = 5). Baseline blood counts were analyzed. Additionally, single PCs (n = 5) were produced by standard platelet apheresis. The concentrations of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor AB (PDGF-AB), transforming growth factor ß1 (TGF-ß1), insulin-like growth factor 1 (IGF-1), interleukin (IL)-1α, IL-1ß, and IL-1 receptor agonist (IL-1RA) were analyzed by enzyme-linked immunosorbent assay, and statistical analyses were performed using descriptive statistics, mean differences, 95% CIs, and P values (analysis of variance). RESULTS: All growth factor preparation methods showed elevated concentrations of the growth factors VEGF, bFGF, PDGF-AB, and TGF-ß1 compared with those of whole blood. Large interindividual differences were found in VEGF and bFGF concentrations. Respective values (mean ± SD in pg/mL) for whole blood, SmartPrep-2, PC, and PRP powder were as follows: VEGF (574 ± 147, 528 ± 233, 1087 ± 535, and 1722), bFGF (198 ± 164, 410 ± 259, 151 ± 99, and 542), PDGF-AB (2394 ± 451, 17,846 ± 3087, 18,461 ± 4455, and 23,023), and TGF-ß1 (14,356 ± 4527, 77,533 ± 13,918, 68,582 ± 7388, and 87,495). IGF-1 was found in SmartPrep-2 (1539 ± 348 pg/mL). For PC (2266 ± 485 pg/mL), IGF-1 was measured at the same levels of whole blood (2317 ± 711 pg/mL) but was not detectable in PRP powder. IL-1α was detectable in whole blood (111 ± 35 pg/mL) and SmartPrep-2 (119 ± 44 pg/mL). CONCLUSION: Problems with PRP such as absent standardization, lack of consistency among studies, and black box dosage could be solved by using characterized PRP powder made by pooling and lyophilizing multiple PCs. The new PRP powder opens up new possibilities for PRP research as well as for the treatment of patients. CLINICAL RELEVANCE: The preparation of pooled PRP by means of lyophilization may allow physicians to apply a defined amount of growth factors by using a defined amount of PRP powder. Moreover, PRP powder as a dry substance with no need for centrifugation could become ubiquitously available, thus saving time and staff resources in clinical practice. However, before transferring the results of this basic science study to clinical application, regulatory issues have to be cleared.

Particle Formation and Aggregation of a Therapeutic Protein in Nanobubble Suspensions.

The generation of nanobubbles following reconstitution of lyophilized trehalose formulations has recently been reported. Here, we characterize particle formation and aggregation of recombinant human interleukin-1 receptor antagonist (rhIL-1ra) in reconstituted formulations of lyophilized trehalose. Particle characterization methods including resonant mass measurement and nanoparticle tracking analysis were used to count and size particles generated upon reconstitution of lyophilized trehalose formulations. In addition, accelerated degradation studies were conducted to monitor rhIL-1ra aggregation in solutions containing various concentrations of suspended nanobubbles. Reconstitution of lyophilized trehalose formulations with solutions containing rhIL-1ra reduced nanobubble concentrations and generated negatively buoyant particles attributed to aggregated rhIL-1ra. Furthermore, levels of rhIL-1ra aggregation following incubation in aqueous solution correlated with concentrations of suspended nanobubbles. The results of this study suggest that nanobubbles may be a contributor to protein aggregation and particle formation in reconstituted, lyophilized therapeutic protein formulations.

A meta-analysis of blood cytokine network alterations in psychiatric patients: comparisons between schizophrenia, bipolar disorder and depression.

Schizophrenia, bipolar disorder and major depressive disorder (MDD) have all been associated with aberrant blood cytokine levels; however, neither the pattern of cytokine alterations nor the impact of clinical status have been compared across disorders. We performed a meta-analysis of blood cytokines in acutely and chronically ill patients with these major psychiatric disorders. Articles were identified by searching the PubMed, PsycInfo and Web of Science, and the reference lists of these studies. Sixty-eight studies met the inclusion criteria (40 schizophrenia, 10 bipolar disorder and 18 MDD) for acutely ill patients. Forty-six studies met the inclusion criteria (18 schizophrenia, 16 bipolar disorder and 12 MDD) for chronically ill patients. Levels of two cytokines (interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α)), one soluble cytokine receptor (sIL-2R), and one cytokine receptor antagonist (IL-1RA) were significantly increased in acutely ill patients with schizophrenia, bipolar mania and MDD compared with controls (P<0.01). Following treatment of the acute illness, IL-6 levels significantly decreased in both schizophrenia and MDD (P<0.01); sIL-2R levels increased in schizophrenia; and IL-1RA levels in bipolar mania decreased. In chronically ill patients, the levels of IL-6 were significantly increased in schizophrenia, euthymic (but not depressed) bipolar disorder and MDD compared with controls (P<0.01). The levels of IL-1ß and sIL-2R were significantly increased in both chronic schizophrenia and euthymic bipolar disorder. Overall, there were similarities in the pattern of cytokine alterations in schizophrenia, bipolar disorder and MDD during acute and chronic phases of illness, raising the possibility of common underlying pathways for immune dysfunction. Effects of treatment on cytokines were more robust for schizophrenia and MDD, but were more frequently studied than for acute mania. These findings have important implications for our understanding of the pathophysiology and treatment of major psychiatric disorders.

Protein and messenger RNA expression of interleukin 1 system members in bovine ovarian follicles and effects of interleukin 1ß on primordial follicle activation and survival in vitro.

This study aimed to investigate the expression of interleukin 1 (IL-1) system members (proteins and messenger RNA of ligands and receptors) and its distribution in ovarian follicles of cyclic cows and to evaluate the effects of IL-1ß on the survival and activation of primordial follicles in vitro. The ovaries were processed for localization of IL-1 system in preantral and antral follicles by immunohistochemical, real-time polymerase chain reaction, and Western blot analysis. For in vitro studies, ovarian fragments were cultured in α-MEM(+) supplemented with IL-1ß (0, 1, 10, 50, or 100 ng/mL), and after 6 d, the cultured tissues were processed for histologic analysis. Immunohistochemical results showed that the IL-1 system proteins IL-1ß, IL-1RA, IL-1RI, and IL-1RII were detected in the cytoplasm of oocytes and granulosa cells from all follicular categories and theca cells of antral follicles. Variable levels of messenger RNA for the IL-1 system members were observed at different stages of development. After 6 d of culture, the presence of IL-1ß (10 or 50 ng/mL) was effective in maintaining the percentage of normal follicles and in promoting primordial follicle activation. In conclusion, IL-1 system members are differentially expressed in ovarian follicles according to their stage of development. Moreover, IL-1ß promotes the development of primordial follicles. These results indicate an important role of the IL-1 system in the regulation of bovine folliculogenesis.

The detection of a synthetic Interleukin-1 receptor antagonist peptide in a seized product from a racing stable.

A synthetic Interleukin-1 receptor antagonist peptide with the sequence Acetyl-Phe-Glu-Trp-Thr-Pro-Gly-Tyr-Trp-Gln-Pro-Tyr-Ala-Leu-Pro-Leu-OH has been identified in a vial seized during a stable inspection. The use of peptide-based Interleukin-1 receptor antagonists as anti-inflammatory agents has not been previously reported, making this peptide the first in a new class of sports doping peptides. The peptide has been characterized by high-resolution mass spectrometry and a detection method developed based on solid-phase extraction and liquid chromatography - triple quadrupole mass spectrometry. Using in vitro and in vivo models to study the properties of the peptide after administration, the peptide was shown to be highly unstable in plasma and was not detected in urine after administration in a rat. The poor stability of the peptide makes detection challenging but also suggests that it has limited effectiveness as an anti-inflammatory drug. Copyright © 2015 John Wiley & Sons, Ltd.