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• Cytokine concentration in finger-prick blood VAMS was more variable than in venous. • Room temperature had the most significant changes in concentration over time. • The optimal storage temperature differed for each analyte. • Incomplete extraction of 6 spiked cytokines may indicate cytokine-VAMS interaction. Dried blood spots (DBS) collected on filter paper such as Guthrie cards are stored for years at room temperature. The assumption is that once dried, the samples remain stable and quantifiable indefinitely since the metabolites these were initially designed to measure, are known for their extended stability. The concentration of other blood proteins such as cytokines, however, are known to vary with storage even in liquid samples stored at −80 °C for extended periods of time. We sought to determine if cytokines are stable for up to 5 months when stored as a dried blood sample using volumetric absorptive microsampling (VAMS) devices. To test this, blood was collected from 4 healthy participants, spiked with recombinant cytokines, and collected into 30 µL VAMS devices. These prepared VAMS devices were stored at room temperature, 4 °C, or −20 °C for up to 5 months and matching VAMS liquid extracts were stored at −80 °C for the same period of time. At each timepoint, the samples were extracted from the VAMS devices and the extracts were analysed by Luminex® for quantification of up to 31 cytokines. These methods were also tested in a remote clinical study over a period of up to 8 months. Cytokine analysis revealed that room temperature, the current standard for DBS and VAMS storage, performed the poorest out of all storage temperatures with significant losses in 13/21 analytes compared to 4 °C at 5 months. Storage at 4 °C or colder performed well for the majority of analytes tested, however out of those, the optimal storage temperature differed for each analyte. There were a small number of analytes that performed poorly regardless of storage conditions and for fractalkine, this was found to be caused by inefficient recovery during extraction. Cytokine concentrations from finger-prick samples were also found to be much more variable that those in venous blood samples. Our results highlight the need to understand the stability of analytes of interest before committing to longitudinal collection and storage of samples in VAMS devices. These data give confidence that storage at 4 °C or colder was beneficial for cytokine stability. Wherein 25/31 cytokines were quantifiably stable at −20 °C when stored for 3 months and 17/21 were quantifiably stable after 5 months when stored at 4 °C. [ABSTRACT FROM AUTHOR]

Abstract In this study, levels of inflammatory protein biomarkers in venous plasma, plasma derived from capillary blood from the earlobe, and capillary plasma stored as dried plasma spots (DPS) were compared. Samples from 12 male individuals were assessed with a panel of 92 inflammation-related proteins using multiplex proximity extension assay. Correlations between sample types varied greatly between analytes. A high correlation of ρ > 0.8 was observed between capillary plasma and DPS for 32 analytes. At this level of correlation, 13 analytes correlated between venous and capillary plasma and 5 analytes in the comparison of venous blood with DPS. Highlights • Measurement of an inflammation protein panel in three sample sources. • Earlobe plasma as a less invasive alternative sample source than venous plasma. • Protein measurement in capillary plasma stored as dried plasma spots (DPS). • Correlation value of ρ > 0.8 between capillary earlobe plasma and DPS for 32 analytes. • Correlation value of ρ > 0.8 between venous and capillary earlobe plasma for 13 analytes. [ABSTRACT FROM AUTHOR]

Encapsulated cell technology (ECT) is a targeted delivery method that uses the genetically engineered cells in semipermeable polymer capsules to deliver cytokines. Thus far, ECT has been extensively utilized in pharmacologic research, and shows enormous potentials in the treatment of posterior segment diseases. Due to the biological barriers within the eyeball, it is difficult to attain effective therapeutic concentration in the posterior segment through topical administration of drug molecules. Encouragingly, therapeutic cytokines provided by ECT can cross these biological barriers and achieve sustained release at the desired location. The encapsulation system uses permeable materials that allow growth factors and cytokines to diffuse efficiently into retinal tissue. Moreover, the ECT based treatment can be terminated timely when we need to retrieve the implant, which makes the therapy reversible and provides a safer alternative for intraocular gene therapy. Meanwhile, we also place special emphasis on optimizing encapsulation materials and enhancing preservation techniques to achieve the stable release of growth factors and cytokines in the eyeball. This technology holds great promise for the treatment of patients with dry AMD, RP, glaucoma and MacTel. These findings would enrich our understandings of ECT and promote its future applications in treatment of degenerative retinopathy. This review comprises articles evaluating the exactness of artificial intelligence-based formulas published from 2000 to March 2024. The papers were identified by a literature search of various databases (PubMed/MEDLINE, Google Scholar, Cochrane Library and Web of Science). [Display omitted] [ABSTRACT FROM AUTHOR]

• There is emerging interest in the role of the immune system in alcohol use disorder (AUD). • We examined the literature regarding cytokines and AUD via systematic review and meta-analysis. • There is sufficient evidence to support an abnormal circulating cytokine profile in AUD. • The cytokine profile of AUD may vary with respect to the different stage of AUD illness. There has been emerging interest in the role of the immune system in the pathophysiology of alcohol use disorder (AUD) given alcohol consumption stimulates immune cells to secrete peripheral pro- and anti-inflammatory cytokines. We conducted a systematic review and meta-analysis to determine whether an abnormal inflammatory cytokine profile exists in AUD patients compared to controls and whether cytokine levels were correlated with behavioural and psychiatric variables. Using the PRISMA (Preferred Reporting Items for Systematic reviews and Meta-analyses) guidelines, a comprehensive search of electronic databases (MEDLINE, EMBASE, Web of Science Core Collection and the Cochrane Library) was conducted, for AUD-related terms in combination with cytokine-related terms. Patients had to meet established criteria for AUD and be compared with healthy controls. A critical appraisal was completed using the Newcastle-Ottawa Scale. Twenty-four papers met the inclusionary criteria with 46 serum or plasma cytokines measured without immune stimulation whereby 17 studies had sufficient data for inclusion in the meta-analysis. Collectively, AUD patients had greater cytokine concentrations than control patients g = 0.85 [ 95% CI 0.42, 1.29]. Differences in cytokine concentrations between AUD patients and controls varied within-study by stage of illness (R (2) 2 = 19.56%). The greatest differences were reported when AUD patients were engaging in active drinking g = 0.96 [0.49, 1.43] or were in alcohol withdrawal g = 1.25 [0.71, 1.80]. Baseline findings were moderated within and between studies by cytokine identity R (2) 2 = 51.10%; R (3) 2 = 44.89%. Cytokine concentrations were not significantly correlated with self-reported craving for alcohol, but were with alcohol consumption r = 0.22 [−0.05, 0.46]. The relationship between cytokine concentration and consumption was moderated by cytokine identity (R (2) 2 = 100.00%; R (3) 2 = 100.00%), and sample age (R (2) 2 = 0.00%; R (3) 2 = 95.76%). There is sufficient evidence to support the presence of an abnormal circulating cytokine profile in AUD which may vary with respect to the different stages of AUD illness. [ABSTRACT FROM AUTHOR]

• Levels of IL-6, IL-4 and IL-17 are slightly upregulated in fibromyalgia (FMS). • Immune-inflammatory and the compensatory (IRS/CIRS) phenotypes are also upregulated. • Research in immune-inflammatory in FMS needs to evaluate cofounding variables. • Immune-inflammatory studies in FMS need to focus on a wider spectrum of biomarkers. The objective was to identify immune alterations in patients with fibromyalgia syndrome (FMS) compared to healthy controls (HC) using meta-analysis and meta-regression. Six electronic databases were searched for suitable original articles investigating immune biomarkers in FMS in comparison to HC. We extracted outcomes and variables of interest, such as mean and SD of peripheral blood immune biomarkers, age or sex. A random-effects model with restricted maximum-likelihood estimator was used to compute effect sizes (standardized mean difference and 95% CI, Hedges' g) and meta-analysis, group meta-analysis and meta-regressions were conducted. Forty-three papers were included in this systematic review, of which 29 were suitable for meta-analysis. Interleukin (IL)-6 (g = 0.36 (0.09–0.63); I 2 = 85.94; p = 0.01), IL-4 (g = 0.50 (0.03–0.98); I 2 = 81.87; p = 0.04), and IL-17A (g = 0.53 (0.00–1.06); I 2 = 87.15; p = 0.05), were significantly higher in FMS compared to HC while also combinations of cytokines into relevant phenotypes were significantly upregulated including M1 macrophage (g = 0.23 (0.03–0.43); I 2 = 77.62; p = 0.02), and immune-regulatory (g = 0.40 (0.09–0.72); I 2 = 84.81; p = 0.01) phenotypes. Heterogeneity levels were very high and subgroup and meta-regression analyses showed that many covariates explained part of the heterogeneity including medication washout, sex, time of blood sampling and exclusion of patients with major depressive disorder. In conclusion, FMS is accompanied by a disbalance between upregulated pro-inflammatory (M1 and Th-17) and immune-regulatory cytokines although effect sizes are small-to-moderate. Based on our results we provide specific methodological suggestions for future research, which should assess Th-1, Th-17, chemokines, and Th-2 phenotypes while controlling for possible confounding variables specified in this study. [ABSTRACT FROM AUTHOR]

There has been much interest in the role of the immune system in the pathophysiology of chronic fatigue syndrome (CFS), as CFS may develop following an infection and cytokines are known to induce acute sickness behaviour, with similar symptoms to CFS. Using the PRISMA (Preferred Reporting Items for Systematic reviews and Meta-analyses) guidelines, a search was conducted on PubMed, Web of Science, Embase and PsycINFO, for CFS related-terms in combination with cytokine-related terms. Cases had to meet established criteria for CFS and be compared with healthy controls. Papers retrieved were assessed for both inclusionary criteria and quality. 38 papers met the inclusionary criteria. The quality of the studies varied. 77 serum or plasma cytokines were measured without immune stimulation. Cases of CFS had significantly elevated concentrations of transforming growth factor-beta (TGF-β) in five out of eight (63%) studies. No other cytokines were present in abnormal concentrations in the majority of studies, although insufficient data were available for some cytokines. Following physical exercise there were no differences in circulating cytokine levels between cases and controls and exercise made no difference to already elevated TGF-β concentrations. The finding of elevated TGF-β concentration, at biologically relevant levels, needs further exploration, but circulating cytokines do not seem to explain the core characteristic of post-exertional fatigue. [ABSTRACT FROM AUTHOR]

Abstract: Objective: Keratinocyte migration from the wound edge is a crucial step in the reepithelization of cutaneous wounds. Growth factors and cytokines, released from cells that invade the wound matrix, play an important role, and several in vitro assays have been performed to elucidate this. The purposes of this study were to review in vitro human studies on keratinocyte migration to identify those growth factors or cytokines that stimulate keratinocyte migration and whether these assays might serve as a screening procedure prior to testing combinations of growth factors or cytokines to promote wound closure in vivo. Methods: Research papers investigating effect of growth factors and cytokines on human keratinocyte migration in vitro were retrieved from library sources, PubMed databases, reference lists of papers, and searches of relevant journals. Results: Fourteen different growth factors and cytokines enhanced migration in scratch wound assay and HGF together with TGF-β, and IGF-1 with EGF, were more stimulatory than either growth factor alone. HGF with TGF-β1 had a greater chemokinetic effect than either growth factor alone in transmigration assay. TGF-β1, FGF-7, FGF-2 and AGF were chemotactic to keratinocytes. EGF, TGF-α, IL-1α, IGF and MGSA enhanced cell migration on ECM proteins. Conclusion: Many growth factors and cytokines enhanced migration of keratinocytes in vitro, and certain combinations of growth factors were more stimulatory than either alone. These and other combinations that stimulate keratinocyte migration in vitro should be tested for effect on wound closure and repair in vivo. The scratch wound assay provides a useful, inexpensive and easy-to-perform screening method for testing individual or combinations of growth factors or cytokines, or growth factors combined with other modalities such as laser irradiation, prior to performing wound healing studies with laboratory animals. [Copyright &y& Elsevier]

Abstract: Multiple sclerosis (MS) is a chronic inflammatory disease characterized by demyelination, remyelination and loss of functions. Even though its etiology is unknown viral, genetic and environmental factors are considered triggers of the disease. MS shows a heterogeneous clinical course, but most patients exhibit exacerbations and remissions from the onset, eventually leading to secondary progressive multiple sclerosis. Systemic inflammatory events are known to signal into the central nervous system (CNS), and can induce a general response known as sickness behavior. Several research papers have demonstrated that a peripheral stimulus can induce the synthesis of cytokines in the brain. In different neurodegenerative diseases peripheral inflammation generates exacerbation to ongoing damage in the brain. In MS, relapsing and remitting episodes are unpredictable; however, peripheral inflammation may exacerbate these events. Clinical studies revealed an association between infections and relapses, which may lead to the worsening of neurological damage. A similar scenario was described in MS animal models demonstrating that peripheral inflammation recrudesced a central ongoing demyelinating lesion. In this paper, we reviewed the existing data on the inflammatory component of MS, with special attention on the effect of peripheral infections in the etiology and progression of MS and its effect on the relapsing and remitting episodes. We also analyzed data concerning the effect of peripheral inflammatory events in MS experimental animal models. This article is part of a Special Issue entitled ''Neuroinflammation in neurodegeneration and neurodysfunction''. [Copyright &y& Elsevier]

Myeloproliferative neoplasm (MPN) is a hematologic malignancy characterized by the clonal outgrowth of hematopoietic cells with a somatically acquired mutation most commonly in JAK2 (JAK2 V 617 F ). This mutation endows upon myeloid progenitors cytokine independent growth and consequently leads to excessive production of myeloid lineage cells. It has been previously suggested that inflammation may play a role in the clonal evolution of JAK2 V 617 F mutants. In particular, it is possible that one or more cellular kinetic parameters of hematopoietic stem cells (HSCs) are affected by inflammation, such as division or death rates of cells, and the probability of HSC differentiation. This suggests a mechanism that can steer the outcome of the cellular competition in favor of the mutants, initiating the disease. In this paper we create a number of mathematical evolutionary models, from very abstract to more concrete, that describe cellular competition in the context of inflammation. It is possible to build a model axiomatically, where only very general assumptions are imposed on the modeling components and no arbitrary (and generally unknown) functional forms are used, and still generate a set of testable predictions. In particular, we show that, if HSC death is negligible, the evolutionary advantage of mutant cells can only be conferred by an increase in differentiation probability of HSCs in the presence of inflammation, and if death plays a significant role in the dynamics, an additional mechanism may be an increase of HSC’s division-to-death ratio in the presence of inflammation. Further, we show that in the presence of inflammation, the wild type cell population is predicted to shrink under inflammation (even in the absence of mutants). Finally, it turns out that if only the differentiation probability is affected by the inflammation, then the resulting steady state population of wild type cells will contain a relatively smaller percentage of HSCs under inflammation. If the division-to-death rate is also affected, then the percentage of HSCs under inflammation can either decrease or increase, depending on other parameters. [ABSTRACT FROM AUTHOR]

One of the major complaints patients who survive cancer often make is chemotherapy induced cognitive impairment (CICI), which survivors often call “chemo brain.” CICI is a side effect of chemotherapy due to the cytotoxicity and neurotoxicity of anti-cancer drugs causing structural and functional changes in brain, even when drugs that do not cross the blood brain barrier (BBB) are used. Diminished cognitive functions including diminution of learning and memory, concentration and attention, processing speed and executive functions that reduce quality of life and ability to work are common signs and symptoms of CICI. There still is not a clarified and complete mechanism for CICI, but researchers have pointed to several biochemical candidates. Chemotherapy-induced, cytokine-mediated involvement in CICI will be mainly discussed in this review paper with emphasis on different types of cytokines, correlated with BBB and epigenetic changes. Mechanisms of ROS-generating, anti-cancer drugs and their relation to cytokine-mediated CICI will be emphasized. [ABSTRACT FROM AUTHOR]

Sickness behavior, a coordinated set of behavioral changes during infection and elicited by the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β), is well studied in non-human animals. Over the last two decades, several papers have expanded this research to include humans. However, these studies use a variety of research designs, and typically focus on a single cytokine and only a few of the many behavioral changes constituting sickness behavior. Therefore, our understanding of human sickness behavior remains equivocal. To generate a more holistic, integrative picture of this phenomenon, a meta-analysis of the human sickness behavior literature was conducted. Full model results show that both IL-6 and IL-1β have significant relationships with sickness behavior, and the strength of these relationships is affected by a number of study parameters, such as type of immune stimulus and inclusion of controls. In addition to research design heterogeneity, other issues to address in future studies include an unequal focus on different cytokines and different sickness behaviors. [ABSTRACT FROM AUTHOR]

While it is generally agreed that apoptosis of pancreatic β-cells is the most important and final step in the progression of type 1 diabetes without which clinical diabetes does not develop, it has not been elucidated which molecule(s) are the real culprit(s) in type 1 diabetes. Perforin, FasL, TNFα, IL-1, IFNγ, and NO have been claimed as the effector molecules; however, they, as a single agent, might explain only part of β-cell death in type 1 diabetes. While FasL was initially considered as a strong candidate for the most important death effector, following experiments cast doubt on such a hypothesis. Combinations or synergism between IFNγ and TNFα or IL-1β are being revisited as the death effectors, and molecular mechanism explaining such a synergism was addressed in several recent papers. The role of NF-κB for pancreatic β-cell death in type 1 diabetes is also controversial. While NF-κB plays anti-apoptotic roles in most other death models, its role in type 1 diabetes might be different probably due to the involvement of multiple cytokines at different stages of the disease progression and the peculiarity of pancreatic β-cells. Recent papers also suggested a role for Ca2+ in cytokine-mediated pancreatic β-cell death. Such participation of Ca2+ in β-cell death appears to have a close relevance to the mitochondrial events or ER stress that constitutes an important part of cell death machinery recently identified. [Copyright &y& Elsevier]

Recent epidemiological studies have associated obesity with a variety of cancer types including HCC. However, the tumor initiating role of obesity in hepatocarcinogenesis is still unknown. The objective of this paper is to investigate the effect of adipocyte-secreted factors on EpCAM+/CD133+ cancer stem cells and to identify which factors play a role in modulating hepatic cancer stem cell behavior. Our results demonstrated that adipocyte-secreted factors affect motility and drug resistance of EpCAM+/CD133+ cells. When incubated with adipocyte conditioned media, EpCAM+/CD133+ cells exhibited augmented motility and reduced sorafenib-induced apoptosis. Using array-based system, we identified secretion of several cytokines such as IL6, IL8 and MCP1 by cultured adipocytes and activation of c-Met, STAT3 and ERK1/2 signaling pathways in EpCAM+/CD133+ cells incubated with adipocyte conditioned media. Treating EpCAM+/CD133+ cancer stem cells with IL6 receptor blocking antibody or c-Met inhibitor SU11274 both reduced the increase in motility; however SU11274 had greater effect on relieving protection from sorafenib-induced apoptosis. These results indicate that adipocyte-secreted factors might regulate cancer stem cell behavior through several signaling molecules including c-Met, STAT3 and ERK1/2 and inhibition of these signaling pathways offer novel strategies in targeting the effect of adipose-derived cytokines in cancer. [ABSTRACT FROM AUTHOR]

The teleologic link between increased production of pro-inflammatory cytokines resulting from a systemic inflammatory response to a burn injury and consequent stimulation of bone resorption is unclear. While it is known that cytokines can stimulate osteocytic and osteoblastic production of the ligand of the receptor activator of NFκB, or RANKL, it is not certain why this occurs. It was therefore hypothesized that the subsequent osteoclastic bone resorption liberates calcium from the bone matrix and somehow affects the inflammatory response. In this paper we show how the cytokine-mediated inflammatory response following severe burn injury in children results in simultaneous increase in bone resorption and up-regulation of the parathyroid calcium-sensing receptor. The acute bone resorption leads to release of calcium from the bone matrix with consequent calcium accumulation in the circulation. The up-regulation of the parathyroid calcium-sensing receptor suppresses the release of parathyroid hormone resulting in a lowering of blood calcium concentration. The simultaneous occurrences of these processes could regulate blood calcium concentration and if calcium concentration affects the inflammatory response, then the calcium-sensing receptor could, at the very least, modulate the inflammatory response by adjusting the blood calcium concentration. We describe in vitro studies in which we demonstrated that peripheral blood mononuclear cells in culture produce the chemokines MIP-1α and RANTES in proportion to the medium calcium concentration and they produce the chemokine MCP-1 in quantities inversely related to medium calcium concentration. CD14 + monocytes in culture will also produce MIP-1α in direct relationship to medium calcium concentration but the correlation coefficient is markedly reduced compared to that with peripheral blood mononuclear cells. These monocytes, which possess the calcium-sensing receptor, do not produce MCP-1 in either direct or inverse relationship to medium calcium concentration. Therefore, it is possible that other peripheral blood mononuclear cells are primarily responsible for the production of chemokines in relation to calcium concentration but these cells have not yet been defined. [ABSTRACT FROM AUTHOR]

Evidence from numerous animal models shows that vagal activity regulates inflammatory responses by decreasing cytokine release. Heart rate variability (HRV) is a reliable index of cardiac vagal regulation and should be inversely related to levels of inflammatory markers. Inflammation is also regulated by sympathetic inputs, but only one previous paper controlled for this. In a larger and more representative sample, we sought to replicate those results and examine potential sex differences in the relationship between HRV and inflammatory markers. Using data from the MIDUS II study, we analyzed the relationship between 6 inflammatory markers and both HF-HRV and LF-HRV. After controlling for sympathetic effects measured by urinary norepinephrine as well as a host of other factors, LF-HRV was found to be inversely associated with fibrinogen, CRP and IL-6, while HF-HRV was inversely associated with fibrinogen and CRP. We did not observe consistent sex differences. These results support the existence of the vagal anti-inflammatory pathway and suggest that it has similar effects in men and women. [ABSTRACT FROM AUTHOR]

Patients with HIV infection are at increased risk of developing non-Hodgkin lymphoma and Hodgkin lymphoma. While the pathogenesis of these lymphomas is incompletely understood, evidence indicates that immune deregulation, genetic alterations and cytokine production play an important role in HIV lymphomagenesis. The lymphoma microenvironment in this heterogeneous group of lymphomas plays an equally critical role in their development, growth and progression. Important components of the microenvironment in HIV-related lymphomas include EBV and/or HHV-8 coinfection, reactive inflammatory cells, tumor microvasculature, and soluble factors. This paper provides a brief overview of HIV-related lymphomas and focuses on their lymphomagenesis and microenvironment. [ABSTRACT FROM AUTHOR]

Zonula occludens-1 (ZO-1) is essential to the proper assembly of interendothelial junction complexes that control blood–brain barrier (BBB) integrity. The goal of the current paper was to improve our understanding of how proinflammatory cytokines modulate ZO-1 properties within the human BBB microvascular endothelium. In this respect, we investigated the effects of TNF-α and IL-6 on ZO-1 using human brain microvascular endothelial cells (HBMvECs). Following treatment of HBMvECs with either cytokine (0–100 ng/ml, 18 h), we observed significantly decreased ZO-1 expression and ZO-1:occludin co-association, in parallel with increased ZO-1 phosphorylation (pTyr and pThr). All effects were dose-dependent. Either cytokine also caused extensive cell-cell border delocalization of ZO-1 in parallel with elevated HBMvEC permeability. Furthermore, pre-treatment of HBMvECs with antioxidants (superoxide dismutase, catalase, apocynin, N -acetylcysteine), or employing targeted inhibition of NADPH oxidase activation (NSC23766, gp91/p47 siRNA), were all found to comparably attenuate the cytokine-dependent decrease in ZO-1 protein expression. In summary, we present an in vitro model of how different proinflammatory cytokines can dysregulate ZO-1 properties in HBMvECs. A causal role for NADPH oxidase activation and oxidant signalling is also confirmed. Our findings add mechanistic depth to current in vivo models of BBB injury manifesting ZO-1 dysregulation. [ABSTRACT FROM AUTHOR]

TRAM/TICAM-2 is used by Toll-like receptor 4 (TLR4) as a bridging adaptor during the mammalian innate immune response. It recruits TRIF, another TIR domain-containing adaptor protein, to TLR4 via TIR domain interactions, which leads to the activation of transcription factors responsible for the production of type-1 interferon and cytokines. The molecular mechanisms of these dual interactions mediated by the TRAM TIR domain are not clear. To understand the molecular basis of TIR:TIR domain interactions, structural and biochemical studies of TRAM TIR domain are necessary, and require a functional soluble protein. In this paper, we report a successful purification and characterization of full-length TRAM. Because full-length TRAM likely contains unstructured regions that may be disadvantageous for structural studies, we also carried out a systematic construct design to determine the boundaries of the TRAM TIR domain. The truncated TRAM constructs were designed based on secondary structure predictions and screened by small-scale expression. Selected constructs were subjected to biophysical analyses. We show that the expressed TRAM TIR domain is functional using in vitro GST pull-down assays that demonstrate a physical interaction with the TLR4 TIR domain. We further show, by site-directed mutagenesis, that the “BB loop” regions of both the TRAM TIR domain and the TLR4 TIR domain are crucial for this physical interaction. [ABSTRACT FROM AUTHOR]

Malignant tumors of ectodermal or endodermal origin may metastasize to the sentinel lymph node, the first lymph node encountered by tumor cells that enter lymphatics in the organ of origin. This pathway is enabled by the anatomy of the disease and the causes of metastasis are the result of complex interactions that include mechanical forces within the tumor and host tissues, and molecular factors initiated by tumor cell proliferation, elaboration of cytokines and changes in the tumor microenvironment. Mechanical stresses may influence complex biochemical, genetic and other molecular events and enhance the likelihood of metastasis. This paper summarizes our understanding of interacting molecular, anatomical and mechanical processes which facilitate metastasis to SLNs. Our understanding of these interacting events is based on a combination of clinical and basic science research, in vitro and in vivo, including studies in lymphatic embryology, anatomy, micro-anatomy, pathology, physiology, molecular biology and mechanobiology. The presence of metastatic tumor in the SLN is now more accurately identifiable and, based upon prospective clinical trials, paradigm-changing SLN biopsy has become the standard of clinical practice in breast cancer and melanoma. [ABSTRACT FROM AUTHOR]

Thrombomodulin (TM), an important determinant of blood vessel homeostasis, is expressed on the luminal vascular endothelial cell surface and is released into serum in response to circulatory signals. This includes the cerebrovascular endothelium, where the anti-coagulant and anti-inflammatory properties of TM are thought to be critical to the brain microcirculation and blood–brain barrier (BBB) integrity. Much is still unknown however about how circulatory stimuli may regulate TM activity within the brain microvasculature. To address this, the current short paper investigated the effects of opposing regulatory signals, namely cytokines (TNF-α, IL-6) and laminar shear stress, on the cellular levels and release of TM in cultured human brain microvascular endothelial cells (HBMvECs). Treatment of confluent HBMvECs with either TNF-α or IL-6 (100 ng/ml, 18 h) reduced TM protein levels by up to 70%, whilst inducing TM release into media by up to 4.4 and 5.5 fold, respectively. The effects of either cytokine (0–100 ng/ml) on TM protein levels (6 or 18 h) and release (0–18 h) were also found to be concentration- and time-dependent. Either cytokine (100 ng/ml, 24–72 h) also reduced TM mRNA levels by > 50%. When exposed to laminar shear stress for 24 h at 8 dyn/cm 2 (SI unit equivalent = 0.8 Pa), TM protein levels were upregulated by 65% in parallel with a 2-fold increase in TM mRNA levels. Shear stress also proved to be a much more potent stimulus for TM release from HBMvECs, yielding media TM levels of 1000 pg/10 5 cells, when compared to 175 and 210 pg/10 5 cells for TNF-α and IL-6, respectively, after parallel 18 h treatments. Finally, shear-conditioned media was found to completely block thrombin-induced permeabilization of HBMvECs, confirming the functional efficacy of released TM. In summary, our data indicate that TM is differentially regulated within cultured HBMvECs by humoral and biomechanical signals. [ABSTRACT FROM AUTHOR]

The practice of burning sugarcane obtained by non-mechanized harvesting exposes workers and the people of neighboring towns to high concentrations of particulate matter (PM) that is harmful to health, and may trigger a series of cardiorespiratory diseases. The aim of this study was to analyze the chemical composition of the micro-particles coming from sugarcane burning residues and to verify the effects of this micro-particulate matter on lung and tracheal tissues. Micro-particulate matter (PM 10 ) was obtained by dissolving filter paper containing burnt residues in NaCl solution. This material was instilled into the Wistar rats׳ nostrils. Histological analyses (hematoxylin and eosin – HE) of cardiac, lung and tracheal tissues were performed. Inflammatory mediators were measured in lung tissues by using ELISA. The chemical composition of the particulate material revealed a large quantity of the phthalic acid ester, high concentrations of phenolic compounds, anthracene and polycyclic aromatic hydrocarbons (PAH). Histological analysis showed a reduction in subjacent conjunctive tissue in the trachea, lung inflammation with inflammatory infiltrate formation and reduction of alveolar spaces and a significant increase ( p <0.05) in the release of IL-1α, IL-1β, IL-6, and INF-γ in the group treated with PM 10 when compared to the control group. We concluded that the burning sugarcane residues release many particles, which have toxic chemical compounds. The micro-particulate matter can induce alterations in the respiratory system. [ABSTRACT FROM AUTHOR]

The immune system of cancer patients is often suppressed. Accumulating evidence suggests that exosomes released from tumor cells may play an essential role in this process but the mechanisms are not fully understood. Here we review recent papers showing that exosomes trigger the release of cytokines/chemokines from immune cells. We suggest that this process will either result in the stimulation of anti-tumor immune reactions or in a systemic immunosuppression. The direction appears to be largely dependent on the duration of interactions between immune cells and exosomes leading to the accumulation of inflammatory factors, i.e. on the length of the exposure to these factors. We propose that a long-term interaction of the immune system with elevated levels of tumor exosomes contributes to the development of immunosuppression in cancer patients. [ABSTRACT FROM AUTHOR]

Abstract: Immune evasion strategies are important for the onset and the maintenance of viral infections. Many viruses have evolved mechanisms to counteract or suppress the host immune response. We have previously characterized two syncytial (syn) variants of Herpes simplex 1 (HSV-1) strain F, syn14-1 and syn17-2, obtained by selective pressure with a natural carrageenan. These variants showed a differential pathology in vaginal and respiratory mucosa infection in comparison with parental strain. In this paper, we evaluated the modulation of immune response in respiratory mucosa by these HSV-1 variants. We observed altered levels of Tumor Necrosis Factor-α and Interleukin-6 in lungs of animals infected with the syn14-1 and syn17-2 variants compared with the parental strain. Also, we detected differences in the recruitment of immune cells to the lung in syn variants infected mice. Both variants exhibit one point mutation in the sequence of the gene of glycoprotein D detected in the ectodomain of syn14-1 and the cytoplasmic tail of syn17-2. Results obtained in the present study contribute to the characterization of HSV-1 syn variants and the participation of the cellular inflammatory response in viral pathogenesis. [Copyright &y& Elsevier]

Highlights: [•] We observe that miR-467b protects apoE−/− mice from atherosclerosis. [•] miR-467b reducing lipid accumulation and inflammatory by targeting LPL in vivo. [•] miR-467b inhibits LPL in aorta and macrophages of apoE−/− mice. [•] The paper explains the regulation mechanism of LPL in vivo. [Copyright &y& Elsevier]

Abstract: The identification of inflammatory mediators in the tear fluid have been extensively used in ocular allergy to find either a ‘disease marker’, to better understand the immune mechanisms involved in the ocular surface inflammation, or to identify potential targets for therapeutic interventions. While the clinical characteristics allow a relatively convincing diagnosis of ocular allergic diseases, in the initial, non active phases, or in the chronic stages, the diagnosis may not be clear. Although not highly specific, total tear IgE can be measured with local tests by inserting a paper strip in the lower meniscus. The measurement of tear specific inflammatory markers, such as histamine, tryptase, ECP, IL-4, IL-5 and eotaxin, may be useful for the diagnosis or monitoring ocular allergy. New technologies such as multiplex bead assays, membrane-bound antibody array and proteomic techniques can characterize the distribution of a wide range of bioactive trace proteins in tears. Dozens of mediators, cytokines, chemokines, growth factors, angiogenic modulators, enzymes and inhibitors were thus identified in small tear samples using these techniques, providing the possible identification of specific biomarker for either specific disease or disease activity. However, to date, there is no a single specific laboratory test suitable for the diagnosis and monitoring of allergic conjunctivitis. [Copyright &y& Elsevier]

Abstract: In this paper, we sought to identify the CD4+ T-cell dynamics in the course of HIV infection in response to continuous and intermittent intravenous courses of interleukin-2 (IL-2), the principal cytokine responsible for progression of CD4+ T-lymphocytes from the G1 to the S phase of the cell cycle. Based on multivariate regression models, previous literature has concluded that the increase in survival of CD4+ T-cell appears to be the critical mechanism leading to sustained CD4+ T-cell levels in HIV-infected patients receiving intermittent IL-2 therapy. Underscored by comprehensive mathematical modeling, a major finding of the present work is related to the fact that, rather than due to any increase in survival of CD4+ T-cells, the expressive, selective and sustained CD4+ T-cell expansions following IL-2 administration may be related to the role of IL-2 in modulating the dynamics of Fas-dependent apoptotic pathways, such as activation-induced cell death (AICD) or HIV-specific apoptotic routes triggered by viral proteins. [Copyright &y& Elsevier]

Abstract: The role for dysregulation of the immune system in the pathogenesis of depressive disorder is well established, and emerging research suggests the role of an underlying genetic vulnerability. The purpose of this review is to summarize the existing literature on the genetic variants involved in neurobiological pathways associated with both immune activation and depression. Using PubMed, Scopus, The Cochrane Library, Embase, Ovid of Medline, PsycINFO and ISI web of Knowledge, we selected 52 papers which are relevant for this literature review. Findings across the literature suggest that functional allelic variants of genes for interleukin-1beta (IL)-1β, tumor necrosis factor-alpha (TNF-α) and C-reactive protein (CRP), as well as genetic variations affecting T-cell function, may increase the risk for depression. Moreover, single nucleotide polymorphisms (SNPs) in the IL-1β, IL-6 and IL-11 genes, and in those regulating T-cell function may be associated with reduced responsiveness to antidepressant therapy. There is also some evidence indicative of a role of genetic variants of the enzymes, Cyclo-oxygenase2 (COX-2) and Phospholipase2 (PLA2), in the aetiology of depression. Finally, SNPs in genes related to the serotonin pathway may play a fundamental role in the shared genetic liability to both immune activation and depressive symptoms. Our review confirms that genetic variants influence the biological mechanisms by which the innate immune system contributes to the development of depression. However, future studies are necessary to identify the molecular mechanisms underlying these associations. [Copyright &y& Elsevier]

Abstract: Cytokines are important in the understanding of the immune process in health and disease and are valuable indicators in diagnostics. Measurements of cytokines are based on immunometric methods, and it is important to understand possible pitfalls in these methods to produce reliable cytokine data. This paper focuses on obtaining optimal measurements when applying enzyme-linked immunosorbent assay (ELISA) or multiplex immunoassays (MIA). Cytokines are measured in serum or plasma, as well as in various other body fluids, all containing a series of antibodies and the possibility of interference from these. Some antibodies, such as heterophilic and human anti-animal antibodies, are able to interfere with all immunoassays, but the immunometric techniques are most prone to serious interference from this source. Another type, rheumatoid factor (RF) is a composite of different autoimmune antibodies which can be present in both blood and synovial fluid. RF is present in some arthritic diseases as well as in some other medical conditions. When present, especially RF IgM is known to interfere with the immunometric measurements. A possible and affordable solution to diminish this interference is PEG precipitation, but other efficient, but more expensive, methods, such as precipitation using Protein L or commercially available blocking agents, are also available. Interference of RF is at present not tested in all cytokine assays, but degree of interference from RF, human anti-animal and heterophilic antibodies, as well as from other possible disease-specific antibodies, must always be considered when developing and applying new assays for cytokine measurements. [Copyright &y& Elsevier]

Abstract: Objective: The present paper investigated possible correlations between the clinical presentation of hepatitis B and the TNF-α −308G/A, IFN-γ +874A/T, TGF-beta1 −509C/T, and IL-10 −1081A/G polymorphisms and associated serum levels of these cytokines. Methods: Fifty-three hepatitis patients were selected and divided into two groups: A – inactive (n =30) and B – chronic hepatitis/cirrhosis (n =23). The control group consisted of 100 subjects who were positive for anti-HBc and anti-HBs. The serum concentrations of the cytokines were determined by immunoenzymatic assays. The polymorphisms of the cytokines genes were assessed by PCR and PCR-SSP. Results: The mean serum levels of IFN-γ of the control group were significantly higher than those of groups A and B, whereas the mean levels TGF-beta1 were significantly higher in groups A and B in comparison with the control. In the case of IL-10, the mean serum level recorded in the control group was significantly higher than that of group B. The TNF-α −308AG genotype was considerably more frequent in group B (43.3%) than the control (14.4%). Conclusion: Higher serum levels of IFN-γ and TGF-beta1 were associated with chronic hepatitis B, and lower serum levels of IL-10 were found in patients with the active disease. Furthermore the presence of allele A of the TNF-α −308 polymorphism suggest a risk of the progressive disease. [Copyright &y& Elsevier]

Abstract: This paper investigates serum levels of interleukin 10 (IL-10) and interleukin 6 (IL-6) in patients with chronic hepatitis C genotype 1 (CHC-GT1), the relation of each with clinical and virological characteristics, how they affect the response to combined therapy and their relation with the IL28B polymorphisms rs12979860. Serum level expression and the polymorphism of IL-10, IL-6 and IL28B were determined in 138 CHC-GT1 patients, treated with pegylated interferon/ribavirin (pegIFN-α/RBV) for 48weeks, in the following samples: baseline, week-12 (during treatment) and week-72 (post-treatment). 77 patients (56%) presented Sustained Virological Response (SVR) and 61 (44%) were non-SVR. Multivariate logistic regression showed that age⩽40years (aOR=3.7, 95%CI=1.5–8.9, P =0.004), low activity of gamma glutamyl transferase (GGT) (aOR=0.9, 95%CI=0.98–0.99, P =0.028), CC genotype of IL28B polymorphim (aOR=2.7, 95%CI=1.0–7.2, P =0.044) and low IL-6 (aOR=0.5, 95%CI=0.3–1.0, P =0.038) were predictor factors of virological response. In all patients, following treatment, IL-6 decreased at week-12 (P =0.004) from baseline and had returned to basal values at week-72. Serum IL-10 concentration was significantly decreased at week-72 only in SVR patients (P ⩽0.001). When patients were stratified by IL28B polymorphisms rs12979860 CC vs non-CC patients, a statistically significant decrease in IL-10 at week-72 in both groups was observed (P =0.003 and P ⩽0.001, respectively). None of the polymorphisms of IL-10 or IL-6 studied were associated with SVR. Conclusions: CC genotype of IL28B and low IL-6 serum concentration are factors associated independently with SVR. Moreover, decreased IL-10 at week-72 is associated with SVR in both CC and non-CC patients, and both factors are important to determine the effectiveness of treatment. [Copyright &y& Elsevier]

Abstract: Ro52 is a common target of circulating autoantibodies in autoimmune disease. Data indicate that anti-Ro52 antibodies are associated with distinct clinical manifestations. It is therefore of major interest to understand how it becomes an antigenic target and what cells express this protein under what conditions and what cellular function it has. Ro52 contains a RING and a B-box motif, followed by a coiled-coil domain and a B30.2 (or PRYSPRY) region in the C-terminal end. This molecular structure places Ro52 within the family of tripartite motif proteins (TRIM), and it is also denoted TRIM21. Like several other TRIM proteins, Ro52 has E3 ligase activity and functions in the process of ubiquitination. Ro52 is expressed in the immune system as a predominantly cytoplasmic protein that can be upregulated and translocate to the nucleus in a proinflammatory environment. Reported substrates for Ro52-mediated ubiquitination include IRF3, IRF5, IRF7 and IRF8, and via these transcription factors Ro52 regulates type 1 interferon and cytokine production. Ro52 is upregulated at the site of autoimmune inflammation, such as cutaneous lupus lesions. This implies that Ro52 may have an important role in the pathogenesis of autoimmunity, and this paper will review the available data on the role of Ro52 in immune responses and autoimmune pathogenesis. [Copyright &y& Elsevier]

Abstract: This paper deals with the development of a mathematical model for the in vitro dynamics of malignant hepatocytes exposed to anti-cancer therapies. The model consists of a set of integro-differential equations describing the dynamics of tumor cells under the effects of mutation and competition phenomena, interactions with cytokines regulating cell proliferation as well as the action of cytotoxic drugs and targeted therapeutic agents. Asymptotic analysis and simulations, developed with an exploratory aim, are addressed to enlighten the role played by the biological phenomena under consideration in the dynamics of hepatocellular carcinoma, with particular reference to the intra-tumor heterogeneity and the response to therapies. The obtained results suggest that cancer progression selects for highly proliferative clones. Moreover, it seems that intra-tumor heterogeneity makes targeted therapeutic agents to be less effective than cytotoxic drugs and a joint action of these two classes of agents may mutually increase their efficacy. Finally, it is highlighted how targeted therapeutic agents might act as an additional selective pressure leading to the selection for the most fitting, and then most resistant, cancer clones. [Copyright &y& Elsevier]

Abstract: Periodontitis, one of the most common chronic inflammatory diseases afflicting man, is increasingly being recognized as a risk factor for atherosclerotic cardiovascular disease (ASCVD). Non-antimicrobial tetracyclines are known to have inhibitory effects on inflammatory mediators and effector molecules, including cytokines and matrix metalloproteinases (MMPs), associated with both diseases. In this paper, we discuss the evidence that doxycycline and related non-antibiotic chemically modified tetracyclines (e.g., CMT-3) can effectively reduce cytokine (TNF-α, IL-6, and MCP-1) production by human mononuclear inflammatory cells when stimulated either by endotoxin (LPS) or by a complex of C-reactive protein/oxidized LDL cholesterol relevant to the pathogenesis of periodontal disease and ASCVD, respectively. This inhibition by tetracycline compounds appears to be mediated at least in part by a suppression of the phosphorylation/activation of the NFκB cell signaling pathway. We are currently conducting clinical trials on patients who exhibit both diseases, and our preliminary data suggest that virtually all acute coronary syndrome (ACS) patients exhibit moderate-to-severe periodontitis, a higher incidence of this oral inflammatory disease than that seen in the population at large. In other studies, a non-antimicrobial formulation of doxycycline (SDD) has been found to dramatically reduce hsCRP, IL-6 and MMP-9 levels in plasma of ACS patients, and SDD has also been found to significantly increase serum levels of both cardio-protective HDL cholesterol and its core molecule apolipoprotein A-I in ASCVD-vulnerable patients with periodontitis. Our current research suggests that one mechanism involved may be the ability of SDD to inhibit MMP-mediated HDL loss by protecting apolipoprotein A-I from proteinase attack. These pleiotropic mechanisms of non-antimicrobial tetracyclines provide significant therapeutic potential to treat chronic inflammatory diseases including both periodontitis and ASCVD. [Copyright &y& Elsevier]

Abstract: Some self-reactive immature T cells escape negative selection in the thymus and may cause autoimmune diseases later. In the periphery, if T cells are stimulated insufficiently by peptide-major histocompatibility complex, they become inactive and their production of cytokines changes, a phenomenon called “T cell anergy”. In this paper, we explore the hypothesis that T cell anergy may function to reduce the risk of autoimmunity. The underlying logic is as follows: Since those self-reactive T cells that receive strong stimuli from self-antigens are eliminated in the thymus, T cells that receive strong stimuli in the periphery are likely to be non-self-reactive. As a consequence, when a T cell receives a weak stimulus, the likelihood that the cell is self-reactive is higher than in the case that it receives a strong stimulus. Therefore, inactivation of the T cell may reduce the danger of autoimmunity. We consider the formalism in which each T cell chooses its response depending on the strength of stimuli in order to reduce the risk of autoimmune diseases while maintaining its ability to attack non-self-antigens effectively. The optimal T cell responses to a weak and a strong stimulus are obtained both when the cells respond in a deterministic manner and when they respond in a probabilistic manner. We conclude that T cell anergy is the optimal response when a T cell meets with antigen-presenting cells many times in its lifetime, and when the product of the autoimmunity risk and the number of self-reactive T cells has an intermediate value. [Copyright &y& Elsevier]

Abstract: Aotus and Saimiri are non-human primate models recommended by the World Health Organization for experimental studies in malaria, especially for vaccine pre-clinical trials. However, research using these primates is hindered by the lack of specific reagents to evaluate immune responses to infection or vaccination. As a step toward developing molecular tools for cytokine expression studies in these species, primer pairs for 18 cytokine gene fragments were designed based on human DNA sequences and used to amplify the corresponding genes in Aotus infulatus and Saimiri sciureus genomic DNA samples. IFNγ, TNFα, LTA, IL2, IL3, IL4, IL5, IL6, IL10, IL12, IL13, CSF2 and TGFβ2 gene fragments were amplified and sequenced. Primer pairs for IL8, IL17, IL18, IL27 and MIF failed to generate amplification products. When compared to the available corresponding human and non-human primate sequences, most – except IL3 and IL4 – showed identity degrees above 90%. Small variations in sequence can help to explain the failure to amplify certain genes or the amplification only at lower annealing temperatures as compared to human DNA samples for several primer pairs. The sequences made available provide the basis for designing molecular tools such as primers for real time PCR specific for A. infulatus and/or S. sciureus. The nucleotide sequences reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned accession numbers DQ985386 to DQ985389, DQ989356 to DQ989369, FJ89020 to FJ89024 and FJ89029. [Copyright &y& Elsevier]

Abstract: Dendritic cells (DCs) are widely distributed throughout the lymphoid and nonlymphoid tissues, and are important initiators of acquired immunity and also serve as regulators by inducing self-tolerance. However, it has not been thoroughly clarified whether DCs are involved in the termination of immune responses. In this paper, we have examined the kinetical movement of dendritic cells (DCs) in the lamina propria using the 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced ileitis model (an animal model for Crohn''s disease). Increased numbers of DCs were recruited to the inflammatory sites from day 1 to day 3 at which time the inflammatory responses was clearly observed, then gradually decreased to a steady-state level on day 7 along with the cessation of responses. Three subsets of DCs, PIR-A/Bhigh, PIR-A/Bmed, and PIR-A/B− DCs in the CD11c+/B220− conventional DCs (cDCs) were noted on day 3; the number of PIR-A/Bmed cDCs increased when the inflammatory responses ceased on day 7. The expression of costimulatory molecules such as CD86 and CD54 was lower in the PIR-A/Bmed DCs compared with the other two cDC subsets or splenic DCs. Furthermore, the stimulatory activity of PIR-A/Bmed cDCs was lower than those of PIR-A/Bhigh or PIR-A/B− cDCs, and far lower than that of splenic DCs. In addition, an increase in the message level of IL-10 was clearly observed in the PIR-A/Bmed cDCs on day 7 while that of proinflammatory cytokines such as IL-6 and IL-12 was low. These data demonstrate that PIR-A/Bmed cDCs which increase at the final stage of inflammation may be involved in the termination of the TNBS-induced ileitis by the delivery of anergic signals to effector T cells due to the lower expressions of costimulatory molecules and the production of immunoregulatory cytokine. [Copyright &y& Elsevier]

Abstract: Few papers have investigated the cytokine profiles of multiple cytokines in cord blood. We obtained cord blood samples from 224 infants admitted to our neonatal intensive care unit. Cytokine profiles of 17 cytokines were investigated using cytometric bead array technology. We found a wide variety of cytokines of various levels which ranged from 0.59pg/ml (in Interleukin (IL)-4) to 222.0pg/ml (in macrophage inflammatory protein-1β. Pro-inflammatory cytokines were highly correlated with each other and with granulocyte-colony stimulating factor and IL-8. On the contrary, IL-5, IL-13, and IL-17 did not show any significant correlation with other cytokines. Several maternal factors were strongly related to several cytokines in cord blood. IL-6, IL-8 and monocyte chemotactic protein-1 were closely related to certain neonatal diseases in preterm neonates. Some cytokines may be regulated independently of each other, while others appear to work as a network affecting physiological and pathological conditions in the fetus. [Copyright &y& Elsevier]

Abstract: The semilinear in-slide models (SLIMs) have been shown to be effective methods for normalizing microarray data [J. Fan, P. Tam, G. Vande Woude, Y. Ren, Normalization and analysis of cDNA micro-arrays using within-array replications applied to neuroblastoma cell response to a cytokine, Proceedings of the National Academy of Science (2004) 1135–1140]. Using a backfitting method, [J. Fan, H. Peng, T. Huang, Semilinear high-dimensional model for normalization of microarray data: a theoretical analysis and partial consistency, Journal of American Statistical Association, 471, (2005) 781–798] proposed a profile least squares (PLS) estimation for the parametric and nonparametric components. The general asymptotic properties for their estimator is not developed. In this paper, we consider a new approach, two-stage estimation, which enables us to establish the asymptotic normalities for both of the parametric and nonparametric component estimators. We further propose a plug-in bandwidth selector using the asymptotic normality of the nonparametric component estimator. The proposed method allow for the modeling of the aggregated SLIMs case where we can explicitly show that taking the aggregated information into account can improve both of the parametric and nonparametric component estimator by the proposed two-stage approach. Some simulation studies are conducted to illustrate the finite sample performance of the proposed procedures. [Copyright &y& Elsevier]

Abstract: With only few exceptions that include Hes-1 p53, and IκB, the expression of genes has never been shown to be oscillatory. Here, we show that the inflammatory cytokine TNF triggers oscillations in >5000 genes. We utilize microarrays at 30-min intervals to analyze the pattern of global gene expression in murine macrophages. We find that 15% of genes in the genome underwent a significant >3-fold increase in expression, with 89% of these displaying oscillations at frequencies as low as every 50min. We analyze further two sub-clusters of genes that either began oscillating early or after a lag phase. Through the use of quantitative PCR, we confirm the oscillations and show that the oscillations are continuous. Moreover, we show that these continuous oscillations are not unique to TNF, but that related cytokines such as RANK-L produces oscillations with a unique induction profile. In the two papers accompanying this one, we analyze the mechanism of these oscillations and find that TNF also triggers oscillations in the phosphorylation of MAP kinases, and that these oscillations combine to recruit transcription factors to promoters in a cyclical fashion. The results presented here suggest that gene transcription is a highly dynamic processes, with thousands of genes displaying rapid (<60min) oscillations over time. Considering this dynamism, time-resolved measurements of gene transcription should become the experimental norm. [Copyright &y& Elsevier]

Abstract: Opioids were originally discovered because of their ability to induce analgesia, but further investigation has shown that the opioids regulate the function of cells involved in the immune response. We suggest that the regulation of cytokine, chemokine, and cytokine receptor expression is a critical component of the immunomodulatory activity of the opioids. In this paper we review the literature dealing with the regulation of cytokine and cytokine receptor expression by agonists for the three major opioid receptor types (μ, κ, and δ), and nociceptin, the natural agonist for the orphanin FQ/nociceptin receptor. Although the opioid receptors share a high degree of sequence homology, opposing roles between the kappa opioid receptor (KOR) and the mu opioid receptor (MOR) have become apparent. We suggest that activation of the KOR induces an anti-inflammatory response through the down-regulation of cytokine, chemokine and chemokine receptor expression, while activation of the MOR favors a pro-inflammatory response. Investigation into the opioid receptor-like (ORL1)/nociceptin system also suggests a role for this receptor as a down-regulator of immune function. These effects suggest a broad role for opioids in the modulation of the function of the immune system, and suggest possible targets for the development of new therapeutics for inflammatory and infectious diseases. [Copyright &y& Elsevier]

Abstract: The epithelial-to-mesenchymal transition (EMT) is a crucial process, occurring both during development and tumor progression, by which an epithelial cell undergoes a conversion to a mesenchymal phenotype, dissociates from initial contacts and migrates to secondary sites. We recently reported that in hepatocytes the multifunctional cytokine TGFβ induces a full EMT characterized by (i) Snail induction, (ii) E-cadherin delocalization and down-regulation, (iii) down-regulation of the hepatocyte transcriptional factor HNF4α and (iv) up-regulation of mesenchymal and invasiveness markers. In particular, we showed that Snail directly causes the transcriptional down-regulation of E-cadherin and HNF4, while it is not sufficient for the up-regulation of mesenchymal and invasiveness EMT markers. In this paper, we show that in hepatocytes TGFβ induces a Src-dependent activation of the focal adhesion protein FAK. More relevantly, we gathered results indicating that FAK signaling is required for (i) transcriptional up-regulation of mesenchymal and invasiveness markers and (ii) delocalization of membrane-bound E-cadherin. Our results provide the first evidence of FAK functional role in TGFβ-mediated EMT in hepatocytes. [Copyright &y& Elsevier]

Abstract: Glycoproteins can be cleared from circulation if they carry oligosaccharide structures that are recognized by specific receptors. High-mannose type and asialo complex oligosaccharides are cleared by the mannose and asialoglycoprotein receptors, respectively. This paper presents the protein and terminal saccharide characterization for nine batches of a glycoprotein developed for pharmaceutical use. Each of these batches was evaluated in human pharmacokinetic (PK) studies, and had similar terminal elimination half-lives, but the initial clearance of this glycoprotein varied between batches. The protein is lenercept, an immunoadhesin comprising the Fc domain of human IgG1 and two tumor necrosis factor (TNF) binding domains derived from the extracellular portion of the TNFR1(p55). Lenercept is manufactured in Chinese hamster ovary (CHO) cells and is extensively N-glycosylated but is devoid of high-mannose glycans. The pharmacokinetic variability between these lots only correlated with terminal N-acetylglucosamine and not with terminal galactose, sialic acid or any polypeptide related parameter. The data emphasize the need for appropriate analytical methods for the characterization of glycoproteins, especially those designed for long half-lives, and show that assessment of the content of all three terminal saccharides is sufficient to ensure consistency of their PK performance properties. [Copyright &y& Elsevier]

Abstract: Studies investigating the influence of psychosocial factors on immunity played a critical and formative role in the field of psychoneuroimmunology (PNI), and have been a major component of articles published in Brain, Behavior and Immunity (BBI). An analysis of papers during the first two decades of BBI from 1987–2006 revealed three behavior-related topics were most prominent: (1) stress-induced changes in immune responses, (2) immune correlates of psychopathology and personality, and (3) behavioral conditioning of immunity. Important subthemes included the effect of early rearing conditions on immune maturation in the developing infant and, subsequently, psychosocial influences affecting the decline of immunity in the senescent host. The responsiveness of cell functioning in the young and elderly helped to validate the view that our immune competence is malleable. Many technical advances in immune methods were also evident. Initially, there was a greater reliance on in vitro proliferative and cytolytic assays, while later studies were more likely to use cell subset enumerations, cytokine quantification, and indices of latent virus reactivation. The reach of PNI extended from the traditional clinical entities of infection, autoimmunity, and cancer to attain a broader relevance to inflammatory physiology, and thus to asthma, cardiovascular and gastrointestinal disease. There continue to be many theoretical and applied ramifications of these seminal findings. Fortunately, the initial controversies about whether psychological processes could really impinge upon and modify immune responses have now receded into the pages of history under the weight of the empirical evidence. [Copyright &y& Elsevier]

Abstract: Aim: A growing number of mutations mapped in the receptor gene for fibroblast growth factor have been implicated in several cranial development disorders including the Apert and Crouzon syndromes. The present paper investigated cellular mechanisms underlying Apert phenotype, by analyzing the effects of FGF2 in primary cultures of Apert periosteal fibroblasts carrying the FGFR2 Pro253Arg mutation. Results: FGF2 administration significantly decreased extracellular matrix production in mutant cells by stimulating degradative enzymatic activities. Gene expression analysis revealed that decorin and biglycan, two proteoglycans involved in collagen fibrillogenesis, were more expressed in mutant cells and down-regulated by FGF2. FGF2 receptor binding showed little differences in high affinity receptor counts between mutant and wild-type cells, while we showed for the first time that low affinity receptors are significantly fewer in mutant cells. Differences were found in Crouzon syndrome, where both high and low affinity receptor counts were up-regulated. Conclusions: The different mutation and low affinity receptor regulation in mutant receptors support the hypothesis that the impact on the activity of the ligand–receptor complex could allow distinct modes of FGF2 activation in Apert and Crouzon syndromes, which interfere with the FGFR2 signalling cascade. [Copyright &y& Elsevier]

Abstract: Impaired appetite and weight changes are commonly seen in patients with depression, but the pathophysiology behind this imbalance between energy intake and energy expenditure remains largely unknown. The aim of this paper is to review the literature regarding a possible role for cytokines in the regulation of appetite and body weight, with special emphasis on depression. There now exists a substantial amount of evidence that depressed patients show signs of immune activation including increased levels of proinflammatory cytokines. Cytokines, which by themselves have anorectic properties, stimulate the release of the cytokine-like anorexogenic peptide leptin. In addition to their anorectic properties, both proinflammatory cytokines and leptin interact with the hypothalamic–pituitary–adrenal (HPA) axis, the sympathetic nervous system (SNS) and the immune system. In turn, these systems regulate energy balance as well as they are dysfunctional in depression. Furthermore, both proinflammatory cytokines and leptin can induce anhedonia, one of the cardinal symptoms of depression. In view of the different effects on appetite and/or body weight observed in melancholic versus atypical depression, we suggest that cytokines are differentially altered in these subtypes of depression, and that this may explain some of the inconsistency in the reported findings of cytokine as well as leptin levels in depressed patients. Finally, we propose that the immune system uses the interoceptive pathway projecting to the insular cortex, a brain region where cytokine-induced changes in appetite could be partly mediated, and that this pathway is activated in depression. [Copyright &y& Elsevier]

Abstract: Understanding how and why the immune and nervous systems communicate in a bidirectional pathway has been fundamental to the development of the psychoneuroimmunology (PNI) field. This review will discuss some of the pivotal results that found the nervous and immune systems use a common chemical language for intra and inter-system communication. Specifically the nervous and immune systems produce a common set of peptide and nonpeptide neurotransmitters and cytokines that provides a common repertoire of receptors and ligands between the two systems. These studies led to the concept that through the sharing of ligands and receptors the immune system could serve as a sixth sense to detect things the body cannot otherwise hear, see, smell, taste or touch. Pathogens, tumors, and allergens are detected with great sensitivity and specificity by the immune system. As a sixth sense the immune system is a means to signal and mobilize the body to respond to these types of challenges. The paper will also review in a chronological manner some of the PNI-related studies important to validating the sixth sense concept. Finally, the review will suggest ways to apply the new found knowledge of the sixth sense to understanding a placebo effect and developing new therapeutic approaches for treatment of human diseases. [Copyright &y& Elsevier]

Abstract: The host response to adenovirus (Ad) infection involves induction of cytokines in lung epithelia. We have demonstrated induction of the lung neutrophil chemokine interleukin-8 (IL-8) by Ad7, a major lung pathogen, in A549 lung epithelial cells and lung tissue through activation of the Erk signaling pathway. However, the mechanism of IL-8 induction is still unclear. In this paper, we first showed that Ad7 viral gene expression is not essential for IL-8 induction as psoralen-UV inactivation of Ad7 did not affect IL-8 mRNA induction or IL-8 protein induction in A549 cells. We then inhibited internalization of Ad7 by treatment of A549 cells with EGTA in calcium-free medium during exposure to Ad7. We verified that this treatment inhibited Ad internalization by confocal microscopy, FACS analysis and Ad E1A and fiber mRNA expression. Preventing internalization by calcium depletion did not inhibit Erk activation by Ad7. However, calcium-dependent internalization was required for IL-8 protein production in Ad7 exposed cells. This is not likely due to an effect of calcium depletion on downstream Erk signaling or IL-8 protein production since calcium depletion did not block IL-8 protein production stimulated by PMA, and because addition of EGTA subsequent to Ad7 internalization also did not prevent Ad induction of IL-8. These studies indicate that Ad7 internalization is calcium-dependent and is required for IL-8 protein induction upon Ad7 infection. Ad7 induction of Erk is independent of calcium and does not require virus internalization. [Copyright &y& Elsevier]

Abstract: Angiopoietin-related growth factor (AGF) is a newly identified member of angiopoietin-related proteins (ARPs)/angiopoietin-like proteins (Angptls). AGF has been considered as a novel growth factor in accelerating cutaneous wound healing, as it is capable of stimulating keratinocytes proliferation as well as angiogenesis. But in our paper, we demonstrate that AGF stimulates keratinocytes proliferation only at high protein concentration, however, it can potently promote adhesion, spreading, and migration of keratinocytes, fibroblasts, and endothelial cells. Furthermore, we confirm that the adhesion and migration cellular events are mediated by RGD-binding integrins, most possibly the αv-containing integrins, by in vitro inhibition assays using synthetic competitive peptides. Our results strongly suggest that AGF is an integrin ligand as well as a mitogenic growth factor and theoretically participates in cutaneous wound healing in a more complex mechanism. [Copyright &y& Elsevier]

Abstract: Renal and cardiac fibrosis leading to organ failure are complications of both diabetes and hypertension. These disease processes, when combined, exacerbate development of fibrotic complications. Control of latent transforming growth factor (TGF)-β activation is a potential determinant of fibrotic progression. Both glucose and angiotensin II (Ang II) upregulate thrombospondin-1 (TSP1), a major activator of latent TGF-β, and stimulate increased TGF-β activity. We previously showed that high glucose stimulated TSP1-dependent TGF-β activation in rat mesangial cells (RMCs). In this paper, we examined whether Ang II similarly upregulates TSP1 production and TSP1-dependent TGF-β activation alone or in combination with high glucose concentrations. Ang II and high glucose stimulated increases in TSP1 protein levels in the conditioned media of both rat cardiac fibroblasts (RCFs) and rat mesangial cells (RMCs). Meanwhile, Ang II stimulated increases in both TGF-β activity and protein by RMCs, whereas, RCFs responded to both Ang II and high glucose with increased TGF-β activity in the absence of altered TGF-β protein levels. A combination of Ang II and high glucose induced synergistic TGF-β activation by RCFs. Moreover, Ang II induction of TSP1 and increased TGF-β activity were blocked by losartan, an antagonist of the Ang II type 1 (AT1) receptor. The increase in TSP1 expression leads to increased TGF-β activity upon Ang II and/or glucose treatment, since peptide antagonists of TSP1-mediated TGF-β activation blocked Ang II and glucose-induced TGF-β activation. Our data support a role for TSP1 in the development and progression of renal and cardiac fibrosis in hypertension and diabetes. [Copyright &y& Elsevier]

Abstract: Monocyte chemoattractant protein-1 (MCP-1) is a chemokine whose circulating levels have been detected in the lesions of several diseases such as pulmonary fibrosis, rheumatoid arthritis and atherosclerosis. However, the factors involved in the regulation of its production remain largely unknown. The main aim of the present paper was to ascertain the contribution of the familial/genetic factors on the production of MCP-1 in apparently healthy individuals. We also tested the possible relationships between the plasma levels of MCP-1 and other cytokines involved in bone metabolism (receptor activator NF-kB ligand (RANKL), osteoprotegerin (OPG), interleukin-6, macrophage-colony stimulating factor, tumor necrosis factor-α). Using ELISA assays the cytokine levels were measured in 570 apparently healthy individuals belonging to ethnically homogeneous Caucasian families. We found that MCP-1 levels were significantly (P <0.01) correlated with RANKL (in both sexes) and with OPG only in women. The study showed that adjusted for potential covariates, 72% of the MCP-1 variance, was attributable to familial effects. About 49% was due to potential genetic factors and the rest was explained by common environmental sources shared by spouses within each family. In conclusion, our data provide reliable evidence for the substantial role of genetic factors in the determination of the phenotypic variability of MCP-1 plasma levels. The association between the osteoclastogenic cytokines and MCP-1 levels in healthy pedigrees is of special interest and might shed light on MCP-1 involvement in bone remodeling. [Copyright &y& Elsevier]