Local Production Of Inflammatory Cytokines Research Articles

Multifunctional injectable microspheres for osteoarthritis therapy via spatiotemporally modulating macrophage polarization and inflammation

Local injection of anti-inflammatory drugs for osteoarthritis emerged as a promising administration in the clinic, and sustained-release dosage forms have great potential for future therapeutic applications. Controlling the response of patients only in the acute inflammatory phase is currently the focus of therapeutic interventions. To relieve acute pain in patients and to improve the long-term prognosis effect of osteoarthritis treatment, we designed a two-pronged approach in this research: an injectable double-layer microsphere containing a “nonsteroidal anti-inflammatory drug - macrophage polarizing factor” was constructed. The results indicated that microspheres could regulate the intra-articular environment by inhibiting local inflammatory cytokine production, promoting macrophage polarization to the M2-phenotype, and increasing the expression of cartilage repair factors. Polymers chosen could govern the biocompatibility of microspheres and control the release sequence of the two drugs. Injection of microspheres into the degenerative articular cavity of rats leads to suppressed inflammation and well-promoted cartilage regeneration.

Genetic polymorphisms of Endoplasmic reticulum amino peptidase 1 (ERAP1) and Interferon lambda 4 (IFN-λ4) in Egyptian patients with type 1 diabetes mellitus

Different genetic and environmental factors are implicated in type I diabetes (T1DM) pathogenesis. About 50% of the genetic susceptibility for T1DM is related to human leukocyte antigen (HLA) genes. Other non-HLA genes have variable roles in the destruction of pancreatic β cells. A highly variable gene called endoplasmic reticulum associated with antigen processing gene 1(ERAP1) shares in activating autoreactive CD8+ T lymphocytes, peptide trimming, and subsequent pancreatic β cells destruction. Local production of inflammatory cytokines within the cells of islets of Langerhans is linked to T1DM progression. Different viral and autoimmune disorders have been linked to genetic variations in type III interferon (IFNλs). This study aimed to determine genetic polymorphisms of interferon lambda 4 (IFNλ4rs 73555604) and endoplasmic reticulum aminopeptidases 1 (ERAP1 rs26618) in Egyptian patients with T1DM. The study recruited 120 patients with T1DM from Kafrelsheikh University Hospital and 100 normal controls who were age and sex matched with the patients' group. Single-nucleotide polymorphism (SNP) genotyping of ERAP1(rs26618) and IFN-λ-4(rs73555604) was performed using real-time polymerase chain reaction. Patients with CC genotype were less likely to develop T1DM than those with TC and TT genotypes for both genes. In addition, T allele frequency in comparison to C allele frequency was significantly increased in T1DM patients when compared to control group (p < 0.001). There were positive correlations between studied SNPs for both genes, fasting and postprandial blood glucose levels which suggest the association of these genes with T1DM occurrence. We concluded that the studied SNPs of ERAP1gene (rs26618) and IFNλ-4 gene(rs73555604) may be associated with T1DM development. In addition, T alleles for both genes could be considered risk alleles while C alleles would be regarded as a protective allele. Patients with TC and TT genotypes would be at a higher risk for T1DM than those carrying CC genotype.

Praziquantel Treatment of Schistosoma mansoni Infected Mice Renders Them Less Susceptible to Reinfection.

Beyond transient control of the infection, additional benefits of mass drug administration of praziquantel in endemic communities have been suggested in communities but not mechanistically investigated experimentally. The present study sought to evaluate the additional and hitherto unreported benefits of repeated mass drug administration of praziquantel. We used a tractable mouse model of Schistosoma mansoni infection to assess the effects of repeated infection-treatment cycles on the host susceptibility to reinfection. Parasitaemia was assessed by quantification of Schistosoma egg burden in liver tissues and morbidity was followed up by histological observation of liver lesions by microscopy and using biochemical measurement of liver transaminases. Immune responses were further determined by serum probing of schistosoma-specific antibodies, cytokines and quantification of liver cellular and soluble mediator responses by flow cytometry and ELISA, respectively. At similar ages and comparable gender distribution, groups of mice undergoing higher number of infections treatment cycles over a longer period, remained susceptible to reinfection by the parasite, as judged by the presence of eggs and the associated increasing pathology in the liver tissues. However, notably, there was a clear and significantly higher propensity to lower egg burden upon reinfection when compared to counterparts undergoing a lower number of infection-treatment cycles. This relative reduction of susceptibility to infection was paralleled by a more robust humoral response against parasite antigens, elevated serum IL-4 and liver cytokines. Of note, praziquantel treatment of infected mice left them at a higher baseline of serum IL-4, IgE and liver cytokines but lower CD4+ T cell -derived cytokines when compared to infected non-treated mice supporting an immunological treatment-induced advantage of previously infected mice over naïve mice and infected/not treated mice. Notably, repeated infection-treatment cycles did not preclude the infection-driven aggravation of collagen deposition in the livers over time and was corroborated by a more robust local production of inflammatory cytokines in the most exposed livers. Taken together, our data reveal that treatment of S. mansoni-infected hosts with praziquantel rewires the immune system to a conformation less permissive to subsequent reinfection in mice. Provided the data are translatable from mouse to human, our findings may provide mechanistic support to the potential benefits of more frequent MDAs in high transmission areas to allow rapid acquisition of protective immunity against reinfection.

Flavokawain B alleviates LPS-induced acute lung injury via targeting myeloid differentiation factor 2.

Acute lung injury (ALI) is a sudden onset systemic inflammatory response. ALI causes severe morbidity and death and currently no effective pharmacological therapies exist. Natural products represent an excellent resource for discovering new drugs. Screening anti-inflammatory compounds from the natural product bank may offer viable candidates for molecular-based therapies for ALI. In this study, 165 natural compounds were screened for anti-inflammatory activity in lipopolysaccharide (LPS)-challenged macrophages. Among the screened compounds, flavokawain B (FKB) significantly reduced LPS-induced pro-inflammatory IL-6 secretion in macrophages. FKB also reduced the formation of LPS/TLR4/MD2 complex by competitively binding to MD2, suppressing downstream MAPK and NF-κB signaling activation. Finally, FKB treatment of mice reduced LPS-induced lung injury, systemic and local inflammatory cytokine production, and macrophage infiltration in lungs. These protective activities manifested as increased survival in the ALI model, and reduced mortality upon bacterial infection. In summary, we demonstrate that the natural product FKB protects against LPS-induced lung injury and sepsis by interacting with MD2 and inhibiting inflammatory responses. FKB may potentially serve as a therapeutic option for the treatment of ALI.

Attenuation of Murine Collagen-Induced Arthritis by Targeting CD6.

CD6 is an important regulator of T cell function that interacts with the ligands CD166 and CD318. To further clarify the significance of CD6 in rheumatoid arthritis (RA), we examined the effects of targeting CD6 in the mouse model of collagen-induced arthritis (CIA), using CD6-knockout (CD6-KO) mice and CD6-humanized mice that express human CD6 in lieu of mouse CD6 on their T cells. We immunized wild-type (WT) and CD6 gene-KO mice with a collagen emulsion to induce CIA. For treatment studies using CD6-humanized mice, mice were immunized similarly and a mouse anti-human CD6 IgG (UMCD6) or control IgG was injected on days 7, 14, and 21. Joint tissues were evaluated for tissue damage, leukocyte infiltration, and local inflammatory cytokine production. Collagen-specific Th1, Th9, and Th17 responses and serum levels of collagen-specific IgG subclasses were also evaluated in WT and CD6-KO mice with CIA. The absence of CD6 reduced 1) collagen-specific Th9 and Th17, but not Th1 responses, 2) the levels of many proinflammatory joint cytokines, and 3) serum levels of collagen-reactive total IgG and IgG1, but not IgG2a and IgG3. Joint homogenate hemoglobin content was significantly reduced in CD6-KO mice with CIA compared to WT mice with CIA (P < 0.05) (reduced angiogenesis). Moreover, treating CD6-humanized mice with mouse anti-human CD6 monoclonal antibody was similarly effective in reducing joint inflammation in CIA. Taken together, these data suggest that interaction of CD6 with its ligands is important for the perpetuation of CIA and other inflammatory arthritides that are T cell driven.

Accelerated degradation of collagen membranes in type 1 diabetic rats is associated with increased expression and production of several inflammatory molecules.

Membrane durability is critical for regenerative procedures. We reported previously that type 1-like diabetes in rats accelerates the degradation of collagen membranes and we tested here whether this is associated with increased local production of inflammatory molecules as part of a diabetes-induced chronic inflammation around and within the membranes. Collagen membrane discs were implanted under the scalp in diabetic (streptozotocin-induced) and control rats, which were sacrificed after 2 or 3weeks. Total RNA and proteins were isolated from the membrane and its surrounding tissues and the expression and production of six inflammatory molecules (interleukin-6 [IL-6], tumor necrosis factor alpha [TNFα], matrix metalloproteinase [MMP]-9, macrophage migration inhibitory factor [MIF], MIP-1α, and MIP-2α) was measured using real-time PCR and western blotting, respectively. Minimal histological analysis of the membranes was conducted to conform to previous studies. Hyperglycemia resulted in reduced membrane thickness (by 10% to 25%) and increased mononuclear infiltrate inside the membrane. mRNA and protein levels of IL-6, TNFα, and MMP-9 were elevated in diabetic rats both 2 and 3weeks post-surgery. The levels (both mRNA and protein) of MIF were increased at 2weeks post-surgery and those of MIP-1α and MIP-2α at 3weeks. There was a very good match in the temporal changes of all examined genes between the mRNA and protein levels. Elevated local production of inflammatory cytokines and MMPs, together with apparent mononuclear infiltrate and increased collagenolysis confirm that hyperglycemia leads to a chronic inflammation in and around the implanted collagen membranes, which reduces membrane longevity.

Correlation between adiponectin, chemerin, vascular endothelial growth factor and epicardial fat volume in patients with coronary artery disease.

Epicardial fat, a local visceral fat depot surrounded by visceral pericardial sac, surrounds the coronary arteries for most of their course and may contribute to the development of coronary atherosclerosis by local production of inflammatory cytokines. Some studies on non-invasive measurement of epicardial fat mass have shown that epicardial fat mass is associated with the increased incidence of coronary artery disease (CAD), onset and progression of coronary plaque, mainly including major adverse cardiovascular events, myocardial ischemia, and atrial fibrillation. In the present study the correlation of adiponectin, chemerin, and vascular endothelial growth factor (VEGF) with the epicardial fat volume in patients with coronary artery disease was explored, and the risk factors for vascular remodeling of CAD patients were analyzed. A total of 50 CAD patients, treated in Chongzuo People's Hospital from August 2017 to September 2018, were enrolled as the observation group, and further 50 healthy individuals, who underwent physical examinations in the hospital at the same period, were enrolled as the control group. RT-qPCR was adopted to detect the expression levels of adiponectin, chemerin and VEGF in the two groups, a 64-slice dual-source CT to detect epicardial fat volume, and Pearson's correlation to analyze adiponectin, chemerin, VEGF and epicardial fat volume. Logistic regression analysis was performed to analyze the risk factors for vascular remodeling in CAD patients, and a receiver operating characteristic (ROC) curve analysis was used to analyze the value of indexes with multifactorial significance in vascular remodeling. The observation group showed obviously larger epicardial fat volume than the control group (P<0.001), lower adiponectin expression than the control group (P<0.001), and higher chemerin and VEGF expression than the control group (P<0.001). In the observation group, adiponectin expression decreased with the increase of epicardial fat volume (P<0.001), while the expression of chemerin and VEGF increased with the increase of epicardial fat volume (P<0.001). Remodeling occurred in 27 of the 50 patients. ROC curve analysis showed that the areas under the curves of adiponectin, chemerin, VEGF and epicardial fat volume were 0.697, 0.652, 0.696 and 0.689, respectively. Epicardial fat volume, adiponectin, chemerin and VEGF are independent risk factors for vascular remodeling and the expression of adiponectin, chemerin and VEGF can reflect epicardial fat volume.

Nicotine exerts neuroprotective effects by attenuating local inflammatory cytokine production following crush injury to rat sciatic nerves.

Recent studies have demonstrated that nicotine exhibited anti-inflammatory and neuroprotective properties by interacting with the alpha 7 nicotinic acetylcholine receptor (α7nAChR). However, the role of nicotine in regeneration during peripheral nerve injury has not been elucidated. The aim of this study was to investigate whether nicotine down-regulated production of proinflammatory cytokines and promoted peripheral nerve regeneration in rats. Rats challenged with sciatic nerve crush injury were treated with nicotine (1.5 mg/kg), three times per day. The expression of the proinflammatory cytokines tumor necrosis factor alpha (TNF-α) and interleukin (IL-1β), pinch test results, growth-associated protein 43 (GAP-43) expression, morphometric analyses, and the sciatic functional indexes were determined in sciatic nerves. Treatment with nicotine decreased local levels of TNF-α and IL-1β, and increased the expression of GAP-43. Nicotine also improved nerve regeneration and functional recovery. The overall protective effects of nicotine were reversed by concomitant treatment with α7nACHR antagonist methyllycaconitine, indicating that nicotine exerted its specific anti-inflammatory and neuroprotective effects through the α7nAChR. These findings show that nicotine administration can provide a potential therapeutic pathway for the treatment of peripheral nerve injury, by a direct protective effect through the α7nAChR-mediated cholinergic anti-inflammatory pathway.

Abstract 665: Perivascular Adipose Tissue near Aortic Arch is a Major Site for Atheroprotective IgM Producing B-1 Cells

Background: Perivascular adipose tissue (PVAT) regulates artery physiology and pathology, such as promoting atherosclerosis development through local production of inflammatory cytokines. The phenotype of PVAT is region-specific and PVAT composed of brown adipose tissue (near the aortic arch and thoracic aorta) and white adipose tissue (near the abdominal aorta). B-1 cells limit adipose tissue inflammation and atherosclerosis through the production of IgM antibodies. PVAT harbors high numbers of B cells. Id3 is a basic helix-loop-helix protein and important for B cell development. B cell-specific Id3 deficiency (Id3 BKO ) increases B-1b cell numbers and provides atheroprotection. However, the location and regulation of IgM producing B-1 cells in the PVAT are unknown. Methods and Results: Flow cytometry analysis of PVAT of normal chow diet fed young ApoE -/- mice (n=5) demonstrated that the abundant CD19 + B cells (per gram fat) harbored in PVAT around aortic arch (2.7 + 0.69 x10 5 cells) compared to PVAT near the thoracic (0.9 + 0.29 x10 5 cells) and abdominal aorta (1.1 + 0.51 x10 5 cells). Interestingly, a large proportion (40%) of these B cells are belong to the CD19 hi B220 low B-1 subset in PVAT near aortic arch. CXCL13 is a chemokine, which is important for B-1 cell recruitment to omental fat. Real time-PCR data confirmed that high numbers of B-1 cell recruitment to PVAT near aortic arch is due to high expression of CXCL13 in the PVAT near the aortic arch compared to PVAT near thoracic aorta and abdominal aorta. Moreover, Flow cytometry and ELISPOT data in normal chow fed young ApoE -/- mice with Id3 BKO demonstrated that Id3 BKO increased B-1b cells (Id3 BKO : 1.2 + 0.16 x10 3 ; Id3 WT : 0.4 + 0.06 x10 3 ; p-value: &lt;0.01; n=6 mice/group) and IgM secreting cells (Id3 BKO : 2.0 + 0.42 x10 3 ; Id3 WT : 0.78 + 0.23 x10 3 ; p-value: &lt;0.05; n=6 mice/group) respectively in PVAT near aortic arch compared to Id3 WT control mice. Conclusion: Results provide the first evidence that atheroprotective B-1 cell profile in PVAT is region-specific and identify Id3 and CXCL13 as regulators of aortic arch PVAT B-1b cell numbers and IgM production.

Mycolactone displays anti-inflammatory effects on the nervous system.

BackgroundMycolactone is a macrolide produced by the skin pathogen Mycobacterium ulcerans, with cytotoxic, analgesic and immunomodulatory properties. The latter were recently shown to result from mycolactone blocking the Sec61-dependent production of pro-inflammatory mediators by immune cells. Here we investigated whether mycolactone similarly affects the inflammatory responses of the nervous cell subsets involved in pain perception, transmission and maintenance. We also investigated the effects of mycolactone on the neuroinflammation that is associated with chronic pain in vivo.Methodology/ Principle findingsSensory neurons, Schwann cells and microglia were isolated from mice for ex vivo assessment of mycolactone cytotoxicity and immunomodulatory activity by measuring the production of proalgesic cytokines and chemokines. In all cell types studied, prolonged (>48h) exposure to mycolactone induced significant cell death at concentrations >10 ng/ml. Within the first 24h treatment, nanomolar concentrations of mycolactone efficiently suppressed the cell production of pro-inflammatory mediators, without affecting their viability. Notably, mycolactone also prevented the pro-inflammatory polarization of cortical microglia. Since these cells critically contribute to neuroinflammation, we next tested if mycolactone impacts this pathogenic process in vivo. We used a rat model of neuropathic pain induced by chronic constriction of the sciatic nerve. Here, mycolactone was injected daily for 3 days in the spinal canal, to ensure its proper delivery to spinal cord. While this treatment failed to prevent injury-induced neuroinflammation, it decreased significantly the local production of inflammatory cytokines without inducing detectable cytotoxicity.Conclusion/ SignificanceThe present study provides in vitro and in vivo evidence that mycolactone suppresses the inflammatory responses of sensory neurons, Schwann cells and microglia, without affecting the cell viability. Together with previous studies using peripheral blood leukocytes, our work implies that mycolactone-mediated analgesia may, at least partially, be explained by its anti-inflammatory properties.

Perivascular Adipose Tissue Harbors Atheroprotective IgM-Producing B Cells.

Adipose tissue surrounding major arteries (Perivascular adipose tissue or PVAT) has long been thought to exist to provide vessel support and insulation. Emerging evidence suggests that PVAT regulates artery physiology and pathology, such as, promoting atherosclerosis development through local production of inflammatory cytokines. Yet the immune subtypes in PVAT that regulate inflammation are poorly characterized. B cells have emerged as important immune cells in the regulation of visceral adipose tissue inflammation and atherosclerosis. B cell-mediated effects on atherosclerosis are subset-dependent with B-1 cells attenuating and B-2 cells aggravating atherosclerosis. While mechanisms whereby B-2 cells aggravate atherosclerosis are less clear, production of immunoglobulin type M (IgM) antibodies is thought to be a major mechanism whereby B-1 cells limit atherosclerosis development. B-1 cell-derived IgM to oxidation specific epitopes (OSE) on low density lipoproteins (LDL) blocks oxidized LDL-induced inflammatory cytokine production and foam cell formation. However, whether PVAT contains B-1 cells and whether atheroprotective IgM is produced in PVAT is unknown. Results of the present study provide clear evidence that the majority of B cells in and around the aorta are derived from PVAT. Interestingly, a large proportion of these B cells belong to the B-1 subset with the B-1/B-2 ratio being 10-fold higher in PVAT relative to spleen and bone marrow. Moreover, PVAT contains significantly greater numbers of IgM secreting cells than the aorta. ApoE−/− mice with B cell-specific knockout of the gene encoding the helix-loop-helix factor Id3, known to have attenuated diet-induced atherosclerosis, have increased numbers of B-1b cells and increased IgM secreting cells in PVAT relative to littermate controls. Immunostaining of PVAT on human coronary arteries identified fat associated lymphoid clusters (FALCs) harboring high numbers of B cells, and flow cytometry demonstrated the presence of T cells and B cells including B-1 cells. Taken together, these results provide evidence that murine and human PVAT harbor B-1 cells and suggest that local IgM production may serve to provide atheroprotection.

Experimental intermittent ischemia augments exercise-induced inflammatory cytokine production.

Acute exercise-induced inflammation is implicated in mediating the beneficial adaptations to regular exercise. Evidence suggests that reduced oxygen and/or blood flow to contracting muscle alters cytokine appearance. However, the acute inflammatory responses to hypoxic/ischemic exercise have been documented with inconsistent results and may not accurately reflect the ischemia produced during exercise in patients with ischemic cardiovascular diseases. Therefore, we determined the extent to which local inflammation is involved in the response to ischemic exercise. Fourteen healthy males performed unilateral isometric forearm contractions for 30 min with and without experimental ischemia. Blood was drawn at baseline, 5 and 10 min into exercise, at the end of exercise, and 30, 60, and 120 min after exercise. Oxygen saturation levels, as measured by near-infrared spectroscopy, were reduced by 10% and 41% during nonischemic and ischemic exercise, respectively. Nonischemic exercise did not affect cytokine values. Ischemia enhanced concentrations of basic fibroblast growth factor, interleukin (IL)-6, IL-10, tumor necrosis factor-alpha, and vascular endothelial growth factor during exercise, but IL-8 was not influenced by ischemic exercise. In conclusion, the present study demonstrates that ischemic, small-muscle endurance exercise elicits local inflammatory cytokine production compared with nonischemic exercise.NEW & NOTEWORTHY We demonstrate that ischemic, small-muscle endurance exercise elicits local inflammatory cytokine production compared with nonischemic exercise. The present study advances our knowledge of the inflammatory response to exercise in a partial ischemic state, which may be relevant for understanding the therapeutic effects of exercise training for people with ischemic cardiovascular disease-associated comorbidities.

Experimental Intermittent Ischemia Augments Exercise-Induced Inflammatory Cytokine Production

Acute exercise-induced transient increases in inflammatory cytokines are linked to the beneficial vascular effects of exercise, but the underlying mechanisms that promote appearance are contingent on numerous factors (e.g. muscle mass recruited, exercise intensity/duration, etc.). Evidence suggests that a lack of oxygen and/or blood flow to working muscle modifies cytokine appearance. However, little is known about the inflammatory response to intermittent ischemia in working muscle. PURPOSE: Determine the extent to which local inflammation is involved in the response to ischemic exercise by reproducing the peripheral arterial disease (PAD)-associated phenomenon of intermittent claudication without the presence of potential confounding comorbidities frequently exhibited by patients with PAD. METHODS: 14 healthy males performed unilateral isometric forearm contractions for 30 minutes with and without experimental ischemia. Blood was drawn at baseline, 5 and 10 minutes into exercise, at the end of exercise, and 30, 60, and 120 minutes after exercise. RESULTS: Oxygen saturation levels, as measured by near-infrared spectroscopy, were reduced by 10% and 41% during non-ischemic and ischemic exercise, respectively (P < 0.001). Non-ischemic exercise did not affect cytokine values during exercise (all P > 0.05). Ischemic exercise enhanced concentrations of basic fibroblast growth factor, interleukin (IL)-6, IL-10, tumor necrosis factor-alpha, and vascular endothelial growth factor at the end of exercise by 148%, 197%, 129%, 154%, and 164% (P < 0.05), respectively, but IL-8 was not influenced by ischemic exercise (P > 0.05). CONCLUSION: In conclusion, the present study demonstrates that ischemic, small muscle endurance exercise elicits local inflammatory cytokine production, compared to non-ischemic exercise. The effect of ischemic exercise with PAD-associated comorbidities may impact the inflammatory response during and after exercise. FUNDING: This study was supported by funds from the Office of the Vice President for Research and the College of Education at the University of Georgia (to N.T.J.). DISCLOSURES: J.R.M. is Chief Operating Officer and K.K.M. is President of Infrared Rx. The other authors report no conflicts of interest, financial or otherwise.

Adipose Tissue-Derived IGFBP1 Facilitates Progression of Leukemia

We recently reported that adipose tissue functions as a reservoir for leukemia stem cells (LSCs) using a murine model of leukemia (Ye et al., Cell Stem Cell, 2016). Intriguingly, the presence of leukemic cells in adipose tissue induces increased lipolysis and the release of free fatty acids (FFA) which in turn fuels the growth of LSCs. Further, adipose tissue protects resident LSCs from chemotherapy. These findings indicate that the unique characteristics of adipose tissue may provide key insights on the growth and survival of leukemic cells. Thus, in the present study we focus on the endocrine function of adipose tissue and explore its role in leukemia development.First, to evaluate secreted factors, we applied adipokine arrays which detect 38 adipokines to leukemia serum collected at different stages of leukemia pathogenesis. Interestingly, we observed a significantly elevated level of serum IGFBP1 as soon as leukemic disease became evident at low levels (~0.5%) in marrow. IGFBP1 increased to levels approximate 200X normal at peak stages of disease burden. ELISA analyses further confirmed these observations.IGFBP1 is normally considered as a liver-derived protein. However, we did not find any changes of IGFBP1 expression in leukemic liver. Rather, we observed a significant increase in the expression of IGFBP1 in adipose tissue. IGFBP1 in conditioned medium (CM) from leukemic gonadal adipose tissue (GAT) is approximate 100X higher than naive GAT. Together, these findings suggest adipose tissue-derived IGFBP1 contributes to the increased serum IGFBP1 we detected in leukemic animals.We next examined the role of IGFBP1 in leukemia development using a neutralizing antibody. Treatment of experimental animals with anti-IGFBP1 antibody significantly decreased leukemic burden in GAT (~40% IgG treated VS. 20% Anti-IGFBP1 treated) while bone marrow (BM) and spleen leukemic engraftment remained unchanged (~40%). Further, less atrophy of adipose tissue as well as less body weight loss was seen in the IGFBP1 neutralizing antibody treated group. Consistent with this observation, serum FFA level was also reduced, suggesting less lipolysis in the IGFBP1 antagonized group. Together, these results indicate that IGFBP1 is involved in the regulation of leukemic cells homing to adipose tissue and consequently leukemia-induced lipolysis.Next we explored the mechanisms for the increased expression of IGFBP1 in adipose tissue. Studies have shown that both FGF21 and hypoxia induce IGFBP1 expression. We did not find any changes of FGF21 expression in adipose tissue or in liver, suggesting FGF21 was not involved in IGFBP1 regulation in our system. In contrast, we observed a five-fold elevation in IGFBP1 mRNA in primary adipose tissue cultured under hypoxic conditions. Studies have shown that both BM and spleen in leukemia mice are already hypoxic in early stages of leukemic development (Benito et al., Plos One, 2011). Thus, we hypothesize that tissue hypoxia may at least partially regulate IGFBP1 in leukemia. Ongoing experiments are testing this hypothesis. Additionally, inflammatory cytokines have been reported to increase IGFBP1 expression. We previously reported significantly increased levels of inflammatory cytokines in the adipose tissue of leukemic mice including TNF-α, IL1 and CSF2 (1.5X normal adipose tissue). Therefore, we hypothesize the local inflammatory cytokine production may also contribute to increased expression of IGFBP1, a theory that is also currently under investigation.IGFBP1 has recently been reported to activate osteoclasts and thus promotes bone metabolism (Wang et al., Cell Metabolism, 2015). Studies have found an increased number of osteoclasts and thus bone loss in our leukemic model (Frisch et al., Blood, 2012). We hypothesize that adipose tissue-derived IGFBP1 contributes to bone loss in leukemic mice. Ongoing experiments are testing whether antagonization of IGFBP1 in leukemic mice will rescue bone loss.Collectively, these findings suggest that adipose tissue-derived IGFBP1 facilitates progression of leukemia through regulation of adipose tissue lipolysis and may promote bone marrow colonization by leukemia cells through activation of osteoclasts. DisclosuresNo relevant conflicts of interest to declare.

Local Vitamin D Metabolism is Differentially Regulated by MKP‐1 in Osteoclasts Stimulated with RANKL vs. LPS

Recent reports indicate that the substrate level activity of Vitamin D (VD) regulates local inflammatory cytokine production, including mediators that regulate osteoclast (OC) formation. MKP‐1, a phosphatase which controls p38 and JNK MAPK signaling has been shown to be dependent on VD. Conversely, VD‐dependent genes involved in VD metabolism (Cyp27b1, Cyp24a1) and bone turnover (RANKL) are regulated by MKP‐1, resulting in abnormal VD activity and reduced OC. Interestingly, we reported that MKP‐1 positively regulates RANKL‐induce OC formation, but also reported that it negatively regulates OC formation in pre‐OC stimulated with pathogenic Lipopolysaccaride (LPS). Given the recent reports of VD's regulation of the inflammatory microenvironment, the objective of these experiments was to investigate the role of MKP‐1 on VDR‐dependent gene expression in RANKL vs. LPS‐stimulated pre‐OC. WT and MKP‐1‐/‐ bone marrow cells were obtained and isolated for pre‐OC using magnetic sorting for CD11bloCD115+ monocytes, stimulated with MCSF and RANKL (50ng/ml) or LPS (100ng/ml) for 96hrs. mRNA was and measured for expression of VD metabolism using qPCR. By adding LPS, Vdr expression was significantly up‐regulated in the MKP‐1‐/‐ population (P&lt;0.01), however, expression of Cyp27b1 was significantly blunted in MKP‐1‐/‐ OC (P&lt;0.05). Cyp24a1 was not induced by LPS. In conclusion, MKP‐1 neg. regulates VDR and + regulates Cyp27b1 in OC in response to LPS, not RANKL, contrary to what has been observed in osteoblasts. In all, MKP‐1 does not inhibit VDR nuclear activity in OC cells stimulated with LPS, but may play a role in the regulation of enzymes involved in local VD activation.

MyD88 signalling in myeloid cells is sufficient to prevent chronic mycobacterial infection

Tuberculosis is a chronic infectious disease caused by Mycobacterium tuberculosis that is responsible for almost 1.5 million deaths per year. Sensing of mycobacteria by the host's immune system relies on different families of receptors present on innate immune cells. Amongst them, several members of the TLR family are involved in the activation of immune cells by mycobacteria, yet the in vivo contribution of individual TLRs to the protective immune response remains controversial. On the contrary, MyD88, the adaptor molecule for most TLRs, plays a non-redundant role in the protection against tuberculosis and mice with a complete germline deletion of MyD88 succumb very early to infection. MyD88 is expressed in both immune and non-immune cells, but it is not clear whether control of mycobacteria requires ubiquitous or cell-type specific MyD88 expression. Therefore, using novel conditional switch-on mouse models, we aimed to investigate the importance of MyD88 signalling in DCs and macrophages for the induction of protective effector mechanisms against mycobacterial infection. We conclude that specific reactivation of MyD88 signalling in CD11c- or lysozyme M-expressing myeloid cells during Mycobacterium bovis Bacille Calmette-Guerin infection is sufficient to restore systemic and local inflammatory cytokine production and to control pathogen burden.

CT Quantification of Epicardial Fat: Implications for Cardiovascular Risk Assessment

Epicardial fat, a local visceral fat depot around the heart, surrounds the coronary arteries for most of their course and may contribute to the development of coronary atherosclerosis through local production of inflammatory cytokines. Several studies which measured epicardial fat volume noninvasively from CT have shown a relationship of increased epicardial fat volume with coronary artery disease, the presence and progression of coronary plaque, major adverse cardiovascular events, myocardial ischemia, and atrial fibrillation. Noninvasive quantitative measurement of epicardial fat volume from CT is feasible, and may play a clinical role in cardiovascular risk assessment. The scientific evidence, to date, warrants larger studies with longer follow-up to further investigate the role of epicardial fat as an imaging marker with prognostic importance.

Epicardial and thoracic fat - Noninvasive measurement and clinical implications.

Epicardial fat, the local visceral fat depot enclosed by the visceral pericardial sac, surrounds the coronary arteries for most of their course, and may contribute to the development of coronary atherosclerosis through local production of inflammatory cytokines. Several studies which measured epicardial fat volume noninvasively have shown a relationship of increased epicardial fat volume with coronary artery disease, with the presence and progression of coronary plaque, major adverse cardiovascular events, myocardial ischemia and atrial fibrillation. Quantitative measurement of epicardial fat volume from noninvasive imaging modalities such as CT and MRI are feasible, and may play a clinical role in cardiovascular risk assessment. The evidence to date warrants larger studies with follow-up to further investigate the role of epicardial fat as an imaging marker with prognostic importance.

Effect of anti-rheumatic agents on periodontal parameters and biomarkers of inflammation: a systematic review and meta-analysis

PurposeAnti-rheumatic agents target common molecular pathways of inflammation in rheumatoid arthritis (RA) and periodontitis. The purpose of this study was to determine the relative effect of anti-rheumatic agents on the levels of inflammatory biomarkers and periodontal inflammation in RA patients with periodontitis.MethodsA systematic review and meta-analysis were conducted of studies comparing periodontal parameters of inflammation, such as bleeding on probing, and biomarkers of inflammation in RA patients with periodontitis and healthy adults with and without periodontitis. The search included the electronic databases MEDLINE, Cochrane Database of Systematic Reviews, and Google Scholar, inclusive through October 2011, with no language restrictions. Hand searches were conducted of the bibliographies of related journals and systematic reviews. Observational and interventional studies assessing the effects of antirheumatic therapy qualified for inclusion. Two reviewers performed independent data extraction and risk-of-bias assessment. Of the 187 identified publications, 13 studies fulfilled the inclusion criteria.ResultsWhen compared to healthy adults without periodontitis, RA subjects were found to have significantly higher levels of bleeding on probing and limited evidence of higher levels of interleukin-1β and tumor necrosis factor-α (TNF-α) in gingival crevicular fluid and saliva. No consistent differences were found in periodontal parameters and inflammatory biomarkers between RA subjects and adults with periodontitis. Studies evaluating the effect of anti-TNF-α therapy in RA subjects with periodontitis have yielded inconsistent results.ConclusionsThere are limited data, however, to suggest that anti-TNF-α agents can reduce local production of inflammatory cytokines and periodontal inflammation in RA patients with periodontitis.

A novel strategy for inducing enhanced mucosal HIV-1 antibody responses in an anti-inflammatory environment.

Prophylactic vaccination against HIV-1 sexual transmission will probably require antibody elicitation at genital mucosal surfaces. However, HIV-1 envelope glycoprotein (Env)-based antigens are weakly immunogenic, particularly when applied mucosally. The polyanion PRO 2000 is safe for human vaginal application, and thus may represent a potential formulating agent for vaginal delivery of experimental vaccine immunogens. Based upon its biochemical properties, we hypothesized that PRO 2000 might enhance mucosal immunogenicity of HIV-1 envelope glycoprotein (Env)-based antigens, promoting local and systemic immune responses. Vaginal immunization with Env-PRO 2000 resulted in significantly increased titres of Env-specific mucosal IgA and IgG in mice and rabbits, respectively, compared to Env alone, revealing modest but significant mucosal adjuvant activity for PRO 2000. In vitro, PRO 2000 associated with Env, protecting the glycoprotein from proteolytic degradation in human vaginal lavage. Unexpectedly, PRO 2000 antagonized TLR4 activation, suppressing local production of inflammatory cytokines. Since inflammation-mediated recruitment of viral target cells is a major risk factor in HIV-1 transmission, the immune modulatory and anti-inflammatory activities of PRO 2000 combined with its intravaginal safety profile suggests promise as an HIV-1 mucosal vaccine formulating agent.