Adult T cell leukemia (ATL) cells show a mature helper-inducer T cell phenotype and are thought to secrete many kinds of cytokines in vivo, complicating the clinical features in these patients. In an attempt to specify the cytokines produced by ATL cells, we measured the cytokine concentration in the culture supernatants of three ATL cell lines, all of which were confirmed to be true peripheral blood ATL cell in origin. All these cell lines showed the same cytokine production profile, secreting IL1-alpha, IL1-beta, LD78(MIP-l alpha), TNF-alpha, IFN-gamma, and GM-CSF, but not secreting IL-1 alpha, IL-1 beta, IL-1 receptor antagonist (IL-1 Ra), IL-4, IFN-alpha, and G-CSF irrespective of the stimulatory agents used. Such limited cytokine production may indicate the specific origin of ATL cells within the helper-inducer T cell subtypes. Moreover, these results explain some of the unusual clinical features of ATL patients.

The main subtypes of inflammatory myopathies include dermatomyositis (DM), polymyositis (PM), necrotizing autoimmune myositis (NAM) and sporadic inclusion-body myositis (sIBM). The review provides an update on the main clinical characteristics unique to each subset, including fundamental aspects on muscle pathology helpful to assure accurate diagnosis, underlying immunopathomechanisms and therapeutic strategies. DM is a complement-mediated microangiopathy leading to destruction of capillaries, distal hypoperfusion and inflammatory cell stress on the perifascicular regions. NAM is an increasingly recognized subacute myopathy triggered by statins, viral infections, cancer or autoimmunity with macrophages as the final effector cells mediating fiber injury. PM and IBM are characterized by cytotoxic CD8-positive T cells which clonally expand in situ and invade MHC-I-expressing muscle fibers. In IBM, in addition to autoimmunity, there is vacuolization and intrafiber accumulation of degenerative and stressor molecules. Pro-inflammatory mediators, such as gamma interferon and interleukin IL1-β, seem to enhance the accumulation of stressor and amyloid-related misfolded proteins. Current therapies using various immunosuppressive and immunomodulating drugs are discussed for PM, DM and NAM, and the principles for effective treatment strategies in IBM are outlined.

BACKGROUND

Insulin resistance and type 2 Diabetes have been associated to a low grade of inflammation and their prevalence increase with ageing.

OBJECTIVE

To analyse the development of inflammation in adipose tissue, liver, muscle and hypothalamus during ageing and the effects of caloric restriction.

METHODS

We have analysed the expression of inflammatory cytokines (TNFα, IL1-β, IL-12B and IL-6), proteins involved in macrophage recruitment (MCP-1, CCR2), TLR4 and macrophage markers (CD11c, CD11b and arginase1). Immunohistochemistry of macrophages has also been performed.

RESULTS

All studied tissues present signs of inflammation during ageing, but with different pattern and intensity. Caloric restriction decreases the expression of most of inflammatory markers.

CONCLUSIONS

These data indicate a role of adiposity in the development of inflammation and insulin resistance during ageing. Dietetic intervention could be a useful tool to ameliorate the development of inflammation and insulin resistance associated with ageing.

The phytochemical, curcumin, has been reported to play many beneficial roles. However, under diabetic conditions, the detail mechanism of its beneficial action in the glucose homeostasis regulatory organ, pancreas, is poorly understood. The present study has been designed and carried out to explore the role of curcumin in the pancreatic tissue of STZ induced and cellular stress mediated diabetes in eight weeks old male Wistar rats. Diabetes was induced with a single intraperitoneal dose of STZ (65 mg/kg body weight). Post to diabetes induction, animals were treated with curcumin at a dose of 100 mg/kg body weight for eight weeks. Underlying molecular and cellular mechanism was determined using various biochemical assays, DNA fragmentation, FACS, histology, immunoblotting and ELISA. Treatment with curcumin reduced blood glucose level, increased plasma insulin and mitigated oxidative stress related markers. In vivo and in vitro experimental results revealed increased levels of proinflammatory cytokines (TNF-α, IL1-β and IFN-γ), reduced level of cellular defense proteins (Nrf-2 and HO-1) and glucose transporter (GLUT-2) along with enhanced levels of signaling molecules of ER stress dependent and independent apoptosis (cleaved Caspase-12/9/8/3) in STZ administered group. Treatment with curcumin ameliorated all the adverse changes and helps the organ back to its normal physiology. Results suggest that curcumin protects pancreatic beta-cells by attenuating inflammatory responses, and inhibiting ER/mitochondrial dependent and independent pathways of apoptosis and crosstalk between them. This uniqueness and absence of any detectable adverse effect proposes the possibility of using this molecule as an effective protector in the cellular stress mediated diabetes mellitus.

Pathogenesis of osteoarthritis (OA) is multifactorial but interleukin-1β (IL-1β) is known to be an important mediator of cartilage degradation. Autophagy is an essential cellular homeostasis mechanism and has been proposed to protect against cartilage degradation and chondrocyte death under pathological conditions. We investigated the role of autophagy activated by sucrose, a natural disaccharide, in suppressing inflammatory mediator's expression and cell death under pathological conditions in human chondrocytes. Autophagy activation was investigated by Western blotting for LC3 and Beclin-1, immunofluorescence staining for LC3 puncta, and measuring autophagic flux. Activation of mTOR, AKT, and P70S6K was evaluated by Western blotting. Chondrocyte apoptosis was evaluated by propidium iodide (PI) staining using flowcytometry, expression of Bax by Western blotting, gene expression by TaqMan assays and caspase 3/7 activity was measured using a luminescence-based assay. We found that sucrose-induced active autophagy in OA chondrocytes in vitro was dependent on the activation of AKT/mTOR/P70S6K signaling pathways but was independent of reactive oxygen species (ROS) production. Sucrose activated autophagy blocked IL-1β-induced apoptosis and mRNA expression of MMP-13, COX-2, and IL-6 in human OA chondrocytes. Glucose or fructose, the two metabolites of sucrose, failed to induce autophagy indicating that autophagy was specifically mediated by sucrose. In conclusion, sucrose attenuated IL-1β induced apoptosis and the expression of catabolic mediators by inducing autophagy, and the autophagy in part was mediated through the activation of AKT/mTOR/P70S6K signaling pathway in human OA chondrocytes. J. Cell. Biochem. 118: 629-639, 2017. © 2016 Wiley Periodicals, Inc.

We and others have previously shown that IL-17 is elevated in the synovial fluid of patients with reactive arthritis (ReA)/undifferentiated spondyloarthropathy (uSpA) having acute synovitis. Major source for IL-17 is Th17 cells, which differentiate from Th0 cells under the influence of TGF-β and IL-6, IL1-β and are maintained by IL-21 and 23. There is a paucity of data on these cytokines in ReA/uSpA. Thus, we measured the levels of Th-17 differentiating and maintaining cytokines in synovial fluid of patients with ReA and uSpA. Fifty patients with ReA/uSpA (ReA 24, uSpA 26), 19 patients with rheumatoid arthritis (RA) and 11 patients with osteoarthritis (OA) were included in the study. Synovial fluid (SF) were collected from knee joint and stored at -80°C until analysis. Cytokines were assayed using ELISA in SF specimens. The median IL-17A levels were significantly elevated in ReA (48.3 pg/ml) and uSpA (32.5 pg/ml) as compared to non-inflammatory OA controls (<7.8 pg/ml; p < 0.0001), while comparable to RA (57.9 pg/ml). Further, IL-6 median values were higher in ReA (25.2 ng/ml) and uSpA (13.6 ng/ml) as compared to OA (0.76 ng/ml; p < 0.0001), and comparable to RA (15.8 ng/ml). The median levels of IL-1β, IL-21 levels were elevated in ReA, uSpA and RA as compared to OA but were not statistically significant. TGF-β levels in ReA and uSpA were similar to OA but lower than in RA (4340 pg/ml; p < 0.05). IL-23 was not detectable in any synovial fluid sample. However, levels of these cytokines did not correlate with disease activity parameters. Significant positive correlation was observed between IL-17 and IL-1β (r = 0.38, p < 0.005), IL-17 and IL-6 (r = 0.659, p < 0.0001), and IL-1β and IL-6 (r = 0.391, p < 0.0001) in ReA and uSpA group. Inflammatory synovitis in ReA/uSpA is mediated by pro-inflammatory cytokines like IL-17, IL-6, IL-1β, and IL-21. However, IL-23 was not detectable in SF. Good correlation between IL-17, IL-6, and IL 1β suggest that either they are co-regulated or they regulate each other.

We examined whether feeding pregnant and lactating rats with hydrogenated vegetable fats rich in trans fatty acids led to an increase in serum endotoxin levels and inflammation and to impaired satiety-sensing pathways in the hypothalamus of 90-day-old offspring. Pregnant and lactating Wistar rats were fed either a standard chow (Control) or one enriched with hydrogenated vegetable fat (Trans). Upon weaning, the male offspring were divided in two groups: Control-Control (CC), mothers and offspring fed the control diet; and Trans-Control (TC), mothers fed the trans diet, and offspring fed the control diet. The offspring's food intake and body weight were quantified weekly and the offspring were killed on the 90th day of life by decapitation. The blood and hypothalamus were collected from the offspring. Food intake and body weight were higher in the TC rats than in the CC rats. TC rats had increased serum endotoxin levels and increased hypothalamic cytokines, IL-6, TNF-α and IL1-β, concentrations (P<.05). TLR4, NFκBp65 and MyD88 were higher (P<.05) in the TC rats than in the CC rats. AdipoR1 was lower in the TC rats than in the CC rats. Thus, the present study shows that the mothers' hydrogenated vegetable fat intake during pregnancy and lactation led to hypothalamic inflammation and impaired satiety-sensing, which promotes deleterious metabolic consequences such as obesity, even after the withdrawal of the causal factor. In other words, the effect remains after the consumption of the standard chow by offspring.

As a cellular degradation mechanism, autophagy exerts crucial and complicated effects on HCC development. Liver non-parenchymal cells, including hepatic resident macrophage Kupffer cells, also play important roles in this process. However, most associated studies have focused on the influence of the autophagy level in hepatic cells and HCC cells, but not liver non-parenchymal cells. Based on our previous study, we confirmed that Atg5 silence in the liver during the preneoplastic stage facilitated liver fibrosis, inflammation and, ultimately, tumorigenesis. We further found that autophagy deficiency promotes the production of inflammatory and fibrogenic factors in macrophages. Moreover, Kupffer cell depletion rescued the tumor-promoting effect of autophagy deficiency during the preneoplastic stage. In autophagy-deficient macrophages, mitochondrial ROS mediated inflammation- and fibrosis-promoting effects by increasing IL1α/β production via enhancing NF-κB-associated pathways. Both blocking of mitochondrial ROS and blocking the IL1 receptor stopped the promotion of fibrosis, inflammation and tumorigenesis resulting from Atg5 knockdown during the preneoplastic stage. In conclusion, autophagy-deficient Kupffer cells promote liver fibrosis, inflammation and, finally, hepatocarcinogenesis during the preneoplastic stage by enhancing mitochondrial ROS- NF-κB-IL1α/β pathways.

Four human breast cancer cell lines (T47D, ZR-75-1, MCF7D and HS578T) were examined for the effects of cytokines on expression of cell surface antigens. Interferon (IFN)gamma up-regulated the expression of ICAM-I (CD54) in all the cell lines and coordinately up-regulated both CD54 and CD40 expression in T47D. Tumour necrosis factor (TNF)alpha, interleukin (IL)1 alpha, IL1 beta and IL6 also up-regulated the expression of CD54 in all the cell lines but CD40 was unaffected. Levels of expression of CD11a, CD18, CD49b, CD58 and CD71 were unaltered by these cytokines. Conditioned medium (CM) generated from human fibroblasts, and in particular from foetal cells, was highly effective in up-regulating expression of ICAM-1 but not of CD40 in the breast cancer cell lines. ICAM-1 induction correlated with IL6 bioactivity in these CMs. Combinations of IL6 with other cytokines, such as IL1, resulted in further increases in ICAM-1 expression. Our observations suggest that IL6 is involved in intercellular signalling between mesenchyme and breast cancer epithelium.

Exposure of quiescent cultures of human gingival fibroblasts (HuGi) and porcine synovicocytes (PSF) to human recombinant interleukin 1 alpha or -beta (IL1 alpha and -beta) enhanced the rate of glycolysis as judged by increased lactate production. The cytokines also increased uptake of [3H]2-deoxyglucose (DG) in a time- and dose-dependent manner. Stimulation of DG uptake was first evident 6-8 h following addition of IL1 and was maximal by 24-30 h. IL1 alpha and -beta were equipotent. Half-maximal stimulation occurred at approximately 1 pM IL1; maximal stimulation (2.5-4.5-fold in HuGi, 3-7-fold in PSF) was obtained with approximately 80 pM IL1. The dose-response curves for lactate production and DG uptake were similar. Increased DG uptake was blocked by specific antisera to IL1 and by inhibitors of protein and RNA synthesis but not by indomethacin, an inhibitor of prostaglandin production. DG uptake was enhanced by IL1 in serum-starved cells in the presence of neutralizing anti-platelet-derived growth factor serum. The effect was therefore not secondary to prostaglandin or platelet-derived growth factor production. No increase in cell cycling was detected in IL1-treated cells under the experimental conditions. Kinetic analysis revealed that the Vmax for DG uptake was increased by IL1 (from 36 to 144 pmol/min/mg of cell protein), whereas the Km was unchanged. HuGi cells were pulse-labeled with [35S]methionine following exposure to IL1. Cell lysates were immunoprecipitated using a specific antiserum raised against human erythrocyte glucose transporter. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis/autoradiography of these immunoprecipitates revealed dose- and time-dependent increases in the net rate of glucose transporter synthesis which mirrored the changes in DG uptake.

The pro-inflammatory cytokine interleukin-1beta (IL-1beta) is strongly expressed during brain injury and is able to induce severe cellular brain damage via the production of soluble factors. Different processes regulate IL-1 biological activities, like the production of anti-inflammatory cytokines such as interleukin-4 (IL-4) and interleukin-10 (IL-10). In this report, we describe the sequential effects of IL-4 and IL-10 on the production of interleukin-6 (IL-6) induced by IL-1beta in mouse primary astrocytes and compare these effects to those of the synthetic glucocorticoid agonist, dexamethasone. IL-6 secretion and IL-6 mRNA expression were determined by ELISA assay and a comparative RT-PCR method, respectively. Incubation of mouse astrocytes in primary culture simultaneously with IL-1beta (10 ng/ml) + IL-10 (10 ng/ml) or IL-1beta + dexamethasone (10(-6) M) markedly reduced IL-1beta induced IL-6 secretion and IL-6 mRNA expression, respectively, whereas simultaneous addition of IL-4 (10 ng/ml) did not alter the induction of IL-6 by IL-1beta. In contrast, after 24 h of IL-1beta treatment, the level of IL-6 was decreased below constitutive levels, and this change was reversed by addition of IL-4. IL-6 production in IL-1beta pretreated cells was also increased by addition of IL-4, whereas IL-10 and dexamethasone had no effects. The delayed time dependent effect of IL-4 might be partially explained by the induction of IL-4 receptor alpha-chain mRNA expression by IL-1beta. Therefore, we conclude that IL-10 and dexamethasone have rapid immunosuppressive effects on the astrocyte response to IL-1beta stimulation, whereas IL-4, which has a delayed action, acts as an immune inducer.

Therapeutics options for rheumatoid arthritis (RA) have increased tremendously in the past decade with the introduction of biological agents in 1999. Several different cellular and cytokine targets have been identified, with specific inhibitors now approved to treat RA, including the tumor necrosis factor (TNF) antagonists (adalimumab, etanercept, infliximab), an interleukin 1 (IL1) antagonist (anakinra), an inhibitor of T cell co-stimulation (abatacept), and a selective depleter of B cells (rituximab). As research has progressed, additional promising targets have been identified. Results from RA studies using several new agents have been reported in the last year. Some of these compounds are similar to agents already available, with additional TNF inhibitors (certolizumab pegol, golimumab) and agents targeting CD20 (ocrelizumab, ofatumumab, TRU-015) in development. Other agents are directed toward new cytokine targets, including IL-6 (tocilizumab), and lymphotoxin pathways (briobacept), as well as other B-cell targets, to include BLyS and APRIL (belimumab, atacicept). Additional small molecule therapies have been studied that are directed against intracellular kinases, including JAK-3 and Syk. This article provides a brief update of data from selected clinical trials in RA, highlighting efficacy, and mechanism-based safety concerns.

The conjugation of the protein transduction domain (PTD) from the HIV-1 Tat protein to shell cross-linked (SCK) nanoparticles is a method to facilitate cell surface binding and transduction. In the previous report, the preparation, derivatization, and characterization of peptide-functionalized SCK nanoparticles were reported in detail. Following assembly, the constructs were evaluated in vitro and in vivo to obtain a preliminary biocompatibility assessment. The effects of SCK exposure on cell viability were evaluated using a metabolic 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and a fluorescent apoptosis assay. Furthermore, stages of apoptosis were quantified by flow cytometry. Although higher levels of peptide functionalization resulted in decreased metabolic function as measured by MTT assay, significant apoptosis was not observed below 500 mg/L for all the samples. To evaluate the potential immunogenic response of the peptide-derivatized constructs, a real-time polymerase chain reaction (RT-PCR) system that allows for the in vitro analysis and quantification of the cellular inflammatory responses tumor necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL1-beta) was utilized. The inflammatory response to the peptide-functionalized SCK nanoparticles as measured by RT-PCR show statistically significant increases in the levels of both TNF-alpha and IL1-beta relative to tissue culture polystyrene (TCPS). However, the measured cytokine levels did not preclude the further testing of SCKs in an in vivo mouse immunization protocol. In this limited assay, measured increases in immunoglobulin G (IgG) concentration in the sera were minimal with no specific interactions being isolated, and more importantly, none of the mice (>50) subjected to the three 100 microg immunization protocol have died. Additionally, no gross morphological changes were observed in postmortem organ histology examinations.

Engagement of the interleukin-1 (IL1) and Toll receptors, which are part of the innate immune system, facilitates activation of the transcription factors NF-kappaB, Elk-1 and AP-1. Pellino1 and Pellino2 have recently been shown to be intermediates in the pathway leading to activation of NF-kappaB. Here we demonstrate for the first time that human Pellino2 interacts with TRAF6 (tumor necrosis factor receptor associated factor 6) and that both Pellino1 and Pellino2 also interact with TAK1 (transforming growth factor-beta activated kinase 1). We also show that Pellino2, but not Pellino1, can activate the mitogen activated protein kinase pathway leading to activation of AP-1 and Elk-1. These observations suggest a role for the Pellino proteins in the IL1/Toll signaling cascades as scaffold proteins that may regulate signaling branch-points.

Liver carcinogenesis is associated with striking changes in the integrin repertoire of hepatocytes, including the overexpression of the laminin and collagen receptors alpha1<em>beta</em>1 and the de novo induction of the laminin receptor alpha6<em>beta</em>1. Our aim was to analyze the role of pro-inflammatory cytokines, interferons and fibrogenic cytokines TGF-<em>beta</em> and FGF2 in the regulation of the expression of <em>beta</em>1 integrins by neoplastic hepatocytes. The 2 human hepatocellular cell lines HepG2 and Hep3B were used as models. Integrin expression was assessed by qualitative methods (immunocytochemistry, Western blotting) and semi-quantitative techniques (FACS, cellular ELISA), before and after stimulation by TNFalpha, <em>IL1</em>-<em>beta</em>, TGF-<em>beta</em>, FGF2, interferon gamma and interferon alpha-2b. HepG2 and Hep3B constitutively expressed alpha1, alpha2, alpha6 and <em>beta</em>1 chains. A 24 to 48-hr stimulation with pro-inflammatory cytokines, TGF-<em>beta</em> and FGF2 induced a significant increase in the concentrations of all integrin chains. The maximum induction was registered for <em>beta</em>1 chain, which presented increases amounting up to 3, 4 and 7 times the control values in the presence of, respectively, TNF alpha/<em>IL1</em>-<em>beta</em>, TGF-<em>beta</em> and FGF2. Interferons had no direct effect on integrin expression and partially antagonized the effects of TNF alpha and TGF-<em>beta</em>. The increased concentrations of integrin chains were associated with an increased membrane expression of the corresponding dimers and with an increased adhesion of stimulated hepatocytes to laminin, which was antagonized by neutralizing anti-<em>beta</em>1 and anti-alpha6 antibodies. Finally, anti-alpha6 antibody inhibited the migration of HepG2 and Hep3B cells in reconstituted basement membrane. Our results suggest that the stimulation of alpha6<em>beta</em>1 integrin expression in hepatocarcinoma cells is essential for cell adhesion and migration.

The directed generation of pure astrocyte cultures from pluripotent stem cells has proven difficult. Generation of defined pluripotent-stem-cell derived astrocytes would allow new approaches to the investigation of plasticity and heterogeneity of astrocytes. We here describe a two-step differentiation scheme resulting in the generation of murine embryonic stem cell (mESC) derived astrocytes (MEDA), as characterized by the upregulation of 19 astrocyte-associated mRNAs, and positive staining of most cells for GFAP (glial fibrillary acidic protein), aquaporin-4 or glutamine synthetase. The MEDA cultures could be cryopreserved, and they neither contained neuronal, nor microglial cells. They also did not react to the microglial stimulus lipopolysaccharide, while inflammatory activation by a complete cytokine mix (CCM) or its individual components (TNF-α, IL1-β, IFN-γ) was readily observed. MEDA, stimulated by CCM, became susceptible to CD95 ligand-induced apoptosis and produced NO and IL-6. This was preceded by NF-kB activation, and up-regulation of relevant mRNAs. Also GFAP-negative astrocytes were fully inflammation-competent. Neurotrophic support by MEDA was found to be independent of GFAP expression. In summary, we described here the generation and functional characterization of microglia-free murine astrocytes, displaying phenotypic heterogeneity as is commonly observed in brain astrocytes.

Inflammation-associated cachexia is associated with multiple chronic diseases and involves activation of appetite regulating centers in the arcuate nucleus of the hypothalamus (ARH). The nucleus of the solitary tract (NTS) in the brainstem has also been implicated as an important nucleus involved in appetite regulation. We set out to determine whether the NTS may be involved in inflammation-associated anorexia by injecting IL-1 beta into the 4th ventricle and assessing food intake and NTS neuronal activation. Injection of IL-1 beta produced a decrease in food intake at 3 and 12h after injection which was ameliorated at the 12h time point by a sub-threshold dose of agouti-related peptide (AgRP). Investigation into neuron types in the NTS revealed that IL-1 beta injection was associated with an increase in c-Fos activity in NTS neurons expressing tyrosine hydroxylase (TH). Additionally, injection of IL-1 beta into the 4th ventricle did not produce c-Fos activation of neurons expressing pro-opiomelanocortin (POMC) in the ARH, cells known to be involved in producing anorexia in response to systemic inflammation. Double-label in situ hybridization revealed that TH neurons did not express IL-1 receptor I (IL1-RI) transcript, demonstrating that c-Fos activation of TH neurons in this setting was not via direct stimulation of IL-1 beta on TH neurons themselves. We conclude that IL-1 beta injection into the 4th ventricle produces anorexia and is accompanied by an increase in activation in TH neurons in the NTS. This provides evidence that the brainstem may be an important mediator of anorexia in the setting of inflammation.

OBJECTIVE

Trichinella spiralis infection in the rat produces mucosal injury and inflammation and causes changes in muscle function and neurotransmitter release in the longitudinal muscle myenteric plexus (LMMP). Whether these changes in LMMP reflect the production of inflammatory mediators in the LMMP remains unknown. The objective of this study was to investigate the profile of interleukins (IL) 1-alpha, 1-beta, and 6 and tumor necrosis factor (TNF) alpha in the LMMP of infected rats. Because muscle changes are T-cell dependent, we also studied the cytokine profile in infected athymic rats.

METHODS

Cytokine messenger RNA (mRNA) was examined using semiquantitative polymerase chain reaction, and protein was measured by enzyme-linked immunosorbent assay, radioimmunoassay, or bioassay.

RESULTS

There was increased expression of IL1-alpha and -beta, IL-6, and TNF-alpha mRNA and protein in the LMMP of both euthymic and athymic infected rats. Increased expression of each cytokine was observed by 24 hours postinfection and, in the case of IL1-beta and TNF-alpha, remained elevated by day 6 postinfection.

CONCLUSIONS

Mucosal injury following T. spiralis infection is accompanied by enhanced expression of proinflammatory cytokines in the LMMP. The expression of IL1-, IL-6, and TNF-alpha in infected athymic rats suggests that these cytokines do not contribute to the changes in muscle growth or contraction but may mediate the previously described changes in neurotransmitter release in this model.

OBJECTIVE

Contributions to the understanding of acute renal failure (ARF) pathogenesis have not been translated into an effective clinical therapy. We studied the effects of pretreatment with the angiotensin II type 1 (AT1) receptor blocker, losartan, on renal function, tissue injury, inflammatory response and serum aldosterone levels in a model of ischemic ARF.

METHODS

Rats underwent unilateral renal ischemia followed by 24 h of reperfusion (IR), and were pretreated or not with 8 (IRL8) or 80 (IRL80) mg/kg/day of losartan for 3 days.

RESULTS

IR kidneys showed marked renal dysfunction, epithelial damage, capillary congestion, increased myeloperoxidase (MPO) activity and increased TNF-alpha, IL1-beta and IL-6 mRNA levels. IRL80 kidneys showed protection against dysfunction and tissue injury, associated with normal MPO activity and cytokine mRNA levels. The lower dose was not able to achieve the same degree of functional renoprotection and could not prevent an increase of MPO or proinflammatory cytokine mRNA levels. The high losartan dose completely prevented an increase of serum aldosterone levels induced by IR.

CONCLUSIONS

Renoprotection of the high losartan dose would be mainly mediated by its anti-inflammatory actions. Our results show a potential pathophysiological role of AT1 activation in promoting renal dysfunction, structural injury, inflammation and aldosterone elevation after IR injury.

The consumption of diets rich in saturated fat largely contributes to the development of obesity in modern societies. A diet high in saturated fats can induce inflammation and impair leptin signaling in the hypothalamus. However, the role of saturated fatty acids on hypothalamic leptin signaling, and hepatic glucose and lipid metabolism remains largely undiscovered. In this study, we investigated the effects of intracerebroventricular (icv) administration of a saturated fatty acid, palmitic acid (PA, C16:0), on central leptin sensitivity, hypothalamic leptin signaling, inflammatory molecules and hepatic energy metabolism in C57BL/6J male mice. We found that the icv administration of PA led to central leptin resistance, evidenced by the inhibition of central leptin's suppression of food intake. Central leptin resistance was concomitant with impaired hypothalamic leptin signaling (JAK2-STAT3, PKB/Akt-FOXO1) and a pro-inflammatory response (TNF-α, IL1-β, IL-6 and pIκBa) in the mediobasal hypothalamus and paraventricular hypothalamic nuclei. Furthermore, the pre-administration of icv PA blunted the effect of leptin-induced decreases in mRNA expression related to gluconeogenesis (G6Pase and PEPCK), glucose transportation (GLUT2) and lipogenesis (FAS and SCD1) in the liver of mice. Therefore, elevated central PA concentrations can induce pro-inflammatory responses and leptin resistance, which are associated with disorders of energy homeostasis in the liver as a result of diet-induced obesity.

Intravenous treatment of male rats with recombinant human interleukin-6 (rhIL6) at 50, 100 and 200 micrograms/kg (corresponding to 4, 8 and 16 x 10(4) U/animal, respectively) reduced the activities of hepatic microsomal cytochrome P450-dependent monoxygenases to varying degrees. Ethylmorphine-N-demethylase activity fell to 53% of control values, an effect similar to that induced by 2.5 mg/kg Escherichia coli lipopolysaccharide (LPS). Ethoxycoumarin-O-deethylase activity was also sensitive to inhibition, whereas IL6 had little effect on the activities of other P450-dependent enzymes, including ethoxyresorufin-O-deethylase. Pentoxyresorufin dealkylase activity, which is representative of the cytochrome P450 IIB 1/2 subfamily, was unaffected by IL6 whereas LPS reduced it to 33.7% of control values. Another hepatocyte-related parameter, serum concentration of alpha 1-acid glycoprotein (AGP), was increased by up to 3.5-fold over baseline by IL6 and 10-fold by LPS. Recombinant human interleukin-1 beta (rhIL1 beta) (10 micrograms/kg, corresponding to 5 x 10(4) U/rat) and recombinant human tumor necrosis factor alpha (rhTNF) (150 micrograms/kg corresponding to 24 x 10(4) U/rat) were both as potent as LPS (2.5 mg/kg) in increasing serum AGP levels and reducing hepatic microsomal monoxygenase activities. IL6 did not potentiate the effects of rhIL1 beta. Hepatic microsomal glucuronyltransferase activities were little affected by LPS and unaffected by rhIL6. Finally, rhIL6 was more potent after i.p. injection than after i.v. or s.c. injection. These results suggest that the effects of LPS, TNF and IL1 on the mixed-function oxidase system in vivo may be due partly to an induction of IL6 in vivo. The different sensitivities of the enzymes to IL6 but not to IL1 or TNF may be due to the involvement of two distinct mechanisms.

The requirements for lectin-induced interleukin 2 (IL2) production by human T cells have been investigated. With two different types of T cells, the Jurkat T cell lymphoma and highly purified HLA class II- peripheral T cells, the amount of IL2 produced was strongly dependent on the lectin concentration used. Addition of accessory cells caused a shift in the dose-response curve, resulting in strongly enhanced IL2 production at low concentrations. Thus, the (absolute) accessory cell dependency for T cells to produce IL2 is defined by experimental conditions. Only at lectin concentrations that were found to be optimal in the presence of accessory cells, removal of these cells abrogates IL2 production. Furthermore, after depletion of monocytes IL 2 production by peripheral T cells became almost completely dependent on the presence of thiols in the culture medium. In contrast, the IL2 production by the Jurkat line was not influenced by addition of thiols. The Jurkat model was used to study the nature of accessory cell because this cell line does not show any reactivity to allogeneic cells. Various myeloid and B lymphoid cell lines were tested as accessory cells. The capacity to function as accessory cell was not related to the monocytic origin of the cell. B cell lines were far more effective than monocytes, as two HLA class II- monocytic cell lines were not active. Even after HLA class II determinants were induced on these cells by incubation with an interferon-gamma-containing conditioned medium, they failed to act as as accessory cells. These experiments question the importance of HLA class II molecules and monokines, such as IL1, for lectin-induced IL2 production.

OBJECTIVE

To investigate both the biochemical and the potential morphological changes in bovine cartilage explants following treatment with glucosamine HCl, and to evaluate the capability of glucosamine to counteract the degradation of cartilage induced by catabolic agents such as interleukin-1beta (IL-1beta) and the bacterial lipopolysaccharide (LPS).

METHODS

Bovine articular cartilage explants were treated with increasing doses of glucosamine HCl (0.25-25mg/ml) in the absence or in the presence of IL-1beta or LPS. The release of matrix proteoglycans in the medium, as well as variations in nitric oxide and lactate production were evaluated by standard assays. Proteoglycan synthesis was determined by incorporation of Na(2)-(35)SO(4). Ultrastructural analysis was performed by transmission electron microscopy.

RESULTS

Increasing doses of glucosamine (2.5, 6.5, 25mg/ml) induced a dose-dependent decrease in proteoglycan synthesis and in lactate production after 24h treatment. The biochemical changes induced by IL1-beta or LPS appeared to be inhibited by 6.5 and 25mg/ml glucosamine. At these concentrations a decrease in cell viability was observed, which reached over 90% at 25mg/ml.

CONCLUSIONS

This study shows that pharmacological doses of glucosamine induce a broad impairment in the metabolic activity of bovine chondrocytes, leading to cell death. The inhibition of the catabolic effects induced by IL1-beta and LPS appears related to glucosamine toxicity. In other experimental models, the same or similar doses of glucosamine have previously been used, without showing any adverse effect. We conclude that, in studying the effects of glucosamine, particular attention should be addressed to the experimental model, the doses and the length of treatment. Published by Elsevier Science Ltd. All rights reserved.

The neuropathogenesis of HIV-associated neurocognitive disorders (HAND) remains puzzling. We interrogated several levels of data (host genetic, histopathology, brain viral load, and neurocognitive) to identify histopathological changes most relevant to HAND. The design of the study is a clinicopathological study employing genetic association analyses. Data and brain tissue from 80 HIV-infected adults were used. Markers in monocyte chemoattractant protein-1 (MCP-1), interleukin 1-alpha (IL1-α), macrophage inflammatory protein 1-alpha (MIP1-α), DRD3, DRD2, and apolipoprotein E (ApoE) were genotyped. Microtubule associated protein 2 (MAP2), synaptophysin (SYP), human leukocyte antigen-DR (HLA-DR), glial fibrillary acidic protein (GFAP), amyloid beta (A-Beta), and ionized calcium-binding adaptor molecule-1 (Iba-1) immunoreactivity were quantified in the frontal cortex, putamen, and hippocampus. A composite score for each marker (mean of the three brain regions) was used. Neurocognitive functioning and other clinical variables were determined within 1 year of death. Brain HIV RNA viral load was available for a subset of cases. MAP2 and SYP proved most relevant to neurocognitive functioning. Immunoreactivity of these markers, as well as A-Beta and Iba-1, was correlated with brain HIV RNA viral load. Several genetic markers in combination with other factors predicted histopathology: HIV blood viral load, MIP1-α genotype, and DRD3 genotype predicted Iba-1 immunoreactivity; the duration of infection and IL1-α genotype predicted GFAP immunoreactivity; ApoE genotype and age at death predicted A-Beta immunoreactivity. These data indicate that HIV replication in the brain is the primary driving force leading to neuroinflammation and dysfunctional protein clearance, as reflected by A-Beta and Iba-1. Downstream to these changes are synaptodendritic degeneration, which is the immediate histopathological substrate of the neurocognitive impairment characteristic of HAND. These intermediate histopathological phenotypes are influenced by host genetic polymorphisms in genes encoding cytokines/chemokines, neuronal protein clearance pathways, and dopaminergic factors.