Effect of curcumin supplementation on serum expression of select cytokines and chemokines in a female rat model of nonalcoholic steatohepatitis.

OBJECTIVE: We recently reported that curcumin supplementation in a metabolically (i.e., Western diet [WD]) and chemically (i.e., CCl4) induced female rat model of non-alcoholic steatohepatitis (NASH) was associated with lower liver pathology scores and molecular markers of inflammation. This occurred when curcumin was given during induction of disease (preventative arm; 8-week WD with or without curcumin [8WD + C vs. 8WD]) as well as when given after disease development (treatment arm; 12-week WD with or without curcumin during weeks 9-12 [12WD + C vs. 12WD]). Herein, we sought to extend our findings from that study by determining the effects of curcumin supplementation on cytokine/chemokine expression in serum collected from these same rats. RESULTS: 24 cytokines/chemokines were assayed. IL-2 (+ 80%) and IL-13 (+ 83%) were greater with curcumin supplementation in the prevention arm. IL-2 (+ 192%), IL-13 (+ 87%), IL-17A (+ 81%) and fractalkine (+ 121%) were higher while RANTES was lower (- 22%) with curcumin supplementation in the treatment arm (p < 0.05 for all). RANTES concentrations also correlated significantly with hepatic pathology scores of inflammation (r = 0.417, p = 0.008). Select serum cytokines/chemokines were affected with curcumin supplementation in this female rat model of NASH. Moreover, curcumin's effect(s) on RANTES and its association with liver disease pathogenesis and progression may warrant further investigation.

Broadleaf Mahonia attenuates granulomatous lobular mastitis­associated inflammation by inhibiting CCL­5 expression in macrophages.

Granulomatous lobular mastitis (GLM) is a type of chronic mammary inflammation with unclear etiology. Currently systematic corticosteroids and methitrexate are considered as the main drugs for GLM treatment, but a high toxicity and risk of recurrence greatly limit their application. It is therefore an urgent requirement that safe and efficient natural drugs are found to improve the GLM prognosis. Broadleaf Mahonia (BM) is a traditional Chinese herb that is believed to have anti­inflammatory properties according to ancient records of traditional Chinese medicine. The present study investigated this belief and demonstrated that BM significantly inhibited the expression of interleukin­1ß (IL­1ß), IL­6, cyclooxygenase­2 and inducible nitric oxide synthase in RAW264.7 cells, but had little influence on the cell viability, cell cycle and apoptosis. Meanwhile, the lipopolysaccharide­induced elevation of reactive oxygen species and nitric oxide was also blocked following BM treatment, accompanied with decreased activity of nuclear factor­κB and MAPK signaling. A cytokine array further validated that BM exhibited significant inhibitory effects on several chemoattractants, including chemokine (C­C motif) ligand (CCL)­2, CCL­3, CCL­5 and secreted tumor necrosis factor receptor 1, among which CCL­5 exhibited the highest inhibition ratio in cell and clinical GLM specimens. Collectively, the results show that BM is a novel effective anti­inflammatory herb in vitro and ex vivo, and that CCL­5 may be closely associated with GLM pathogenesis.

Increased galectin-3 levels are associated with abdominal aortic aneurysm progression and inhibition of galectin-3 decreases elastase-induced AAA development.

Abdominal aortic aneurysm (AAA) evolution is unpredictable and no specific treatment exists for AAA, except surgery to prevent aortic rupture. Galectin-3 has been previously associated with CVD, but its potential role in AAA has not been addressed. Galectin-3 levels were increased in the plasma of AAA patients (n=225) compared with the control group (n=100). In addition, galectin-3 concentrations were associated with the need for surgical repair, independently of potential confounding factors. Galectin-3 mRNA and protein expression were increased in human AAA samples compared with healthy aortas. Experimental AAA in mice was induced via aortic elastase perfusion. Mice were treated intravenously with the galectin-3 inhibitor modified citrus pectin (MCP, 10 mg/kg, every other day) or saline. Similar to humans, galectin-3 serum and aortic mRNA levels were also increased in elastase-induced AAA mice compared with control mice. Mice treated with MCP showed decreased aortic dilation, as well as elastin degradation, vascular smooth muscle cell (VSMC) loss, and macrophage content at day 14 postelastase perfusion compared with control mice. The underlying mechanism(s) of the protective effect of MCP was associated with a decrease in galectin-3 and cytokine (mainly CCL5) mRNA and protein expression. Interestingly, galectin-3 induced CCL5 expression by a mechanism involving STAT3 activation in VSMC. Accordingly, MCP treatment decreased STAT3 phosphorylation in elastase-induced AAA. In conclusion, increased galectin-3 levels are associated with AAA progression, while galectin-3 inhibition decreased experimental AAA development. Our data suggest the potential role of galectin-3 as a therapeutic target in AAA.

Radix isatidis Polysaccharides Inhibit Influenza a Virus and Influenza A Virus-Induced Inflammation via Suppression of Host TLR3 Signaling In Vitro.

Influenza remains one of the major epidemic diseases worldwide, and rapid virus replication and collateral lung tissue damage caused by excessive pro-inflammatory host immune cell responses lead to high mortality rates. Thus, novel therapeutic agents that control influenza A virus (IAV) propagation and attenuate excessive pro-inflammatory responses are needed. Polysaccharide extract from Radix isatidis, a traditional Chinese herbal medicine, exerted potent anti-IAV activity against human seasonal influenza viruses (H1N1 and H3N2) and avian influenza viruses (H6N2 and H9N2) in vitro. The polysaccharides also significantly reduced the expression of pro-inflammatory cytokines (IL-6) and chemokines (IP-10, MIG, and CCL-5) stimulated by A/PR/8/34 (H1N1) at a range of doses (7.5 mg/mL, 15 mg/mL, and 30 mg/mL); however, they were only effective against progeny virus at a high dose. Similar activity was detected against inflammation induced by avian influenza virus H9N2. The polysaccharides strongly inhibited the protein expression of TLR-3 induced by PR8, suggesting that they impair the upregulation of pro-inflammatory factors induced by IAV by inhibiting activation of the TLR-3 signaling pathway. The polysaccharide extract from Radix isatidis root therefore has the potential to be used as an adjunct to antiviral therapy for the treatment of IAV infection.

CCL5/RANTES contributes to hypothalamic insulin signaling for systemic insulin responsiveness through CCR5.

Many neurodegenerative diseases are accompanied by metabolic disorders. CCL5/RANTES, and its receptor CCR5 are known to contribute to neuronal function as well as to metabolic disorders such as type 2 diabetes mellitus, obesity, atherosclerosis and metabolic changes after HIV infection. Herein, we found that the lack of CCR5 or CCL5 in mice impaired regulation of energy metabolism in hypothalamus. Immunostaining and co-immunoprecipitation revealed the specific expression of CCR5, associated with insulin receptors, in the hypothalamic arcuate nucleus (ARC). Both ex vivo stimulation and in vitro tissue culture studies demonstrated that the activation of insulin, and PI3K-Akt pathways were impaired in CCR5 and CCL5 deficient hypothalamus. The inhibitory phosphorylation of insulin response substrate-1 at Ser302 (IRS-1S302) but not IRS-2, by insulin was markedly increased in CCR5 and CCL5 deficient animals. Elevating CCR5/CCL5 activity induced GLUT4 membrane translocation and reduced phospho-IRS-1S302 through AMPKα-S6 Kinase. Blocking CCR5 using the antagonist, MetCCL5, abolished the de-phosphorylation of IRS-1S302 and insulin signal activation. In addition, intracerebroventricular delivery of MetCCL5 interrupted hypothalamic insulin signaling and elicited peripheral insulin responsiveness and glucose intolerance. Taken together, our data suggest that CCR5 regulates insulin signaling in hypothalamus which contributes to systemic insulin sensitivity and glucose metabolism.

Ashwagandha attenuates TNF-α- and LPS-induced NF-κB activation and CCL2 and CCL5 gene expression in NRK-52E cells.

BACKGROUND: The aging kidney is marked by a chronic inflammation, which may exacerbate the progression of renal dysfunction, as well as increase the susceptibility to acute injury. The identification of strategies to alleviate inflammation may have translational impact to attenuate kidney disease. METHODS: We tested the potential of ashwaganda, sutherlandia and elderberry on tumor necrosis factor-α (TNF-α) and lipopolysaccharide (LPS) induced chemokine (CCL2 and CCL5) expression in vitro. RESULTS: Elderberry water-soluble extract (WSE) was pro-inflammatory, while sutherlandia WSE only partially attenuated the TNF-α-induced changes in CCL5. However, ashwaganda WSE completely prevented TNF-α-induced increases in CCL5, while attenuating the increase in CCL2 expression and NF-κB activation. The same pattern of ashwagandha protection was seen using LPS as the pro-inflammatory stimuli. CONCLUSIONS: Taken together, these results demonstrate the ashwaganda WSE as a valid candidate for evaluation of therapeutic potential for the treatment of chronic renal dysfunction.

Effect of salvianolic acid A and C compatibility on inflammatory cytokines in rats with unilateral ureteral obstruction.

OBJECTIVE: To investigate the effect of salvianolic acid A and C component molecules, which are involved in drug compatibility, on inflammatory cytokine expression that affects human chemokine ligand 5 (CCL5) and chemokine ligand 10 (CXCL10) levels in rats with unilateral ureteral obstruction (UUO). METHODS: Fifty Sprague Dawley rats were randomly divided into five groups: normal, model, salvianolic acid A, salvianolic acid C and salvianolic acid A and C groups. The normal group was used as the control, and the other groups of rats had a UUO model established. The control group had free access to food and water, and the other groups received the corresponding drugs for 2 weeks. After the last administration, urine ß2-microglobulin (ß 2-MG) and N-acetyl-ß-D-glucosaminidase (NAG) levels were analyzed. After 24 h, all rats were sacrificed and the serum was analyzed for creatinine (Cr) and blood urea nitrogen (BUN) levels. Rat kidneys were removed, and CCL5 and CXCL10 inflammatory cytokine mRNA expression was measured using real-time fluorescent quantitative reverse transcription-polymerase chain reaction (RT-PCR). Kidney fibrosis was observed using hematoxylin-eosin (HE) staining and Masson trichrome staining. RESULTS: In the salvianolic acid A and salvianolic acid C treatment groups, serum Cr and urine NAG levels were significantly lower than in the model group (both P < 0.05). In all treatment groups, urine ß2-MG levels were significantly lower than in the model group (all P < 0.05). Compared with model group, the pathological changes and collagen deposition improved to varying degrees (both P < 0.05). CCL5 and CXCL10 mRNA expression decreased to different degrees compared with the model group (both P < 0.05). CONCLUSION: Salvianolic acid A and C are component molecules of drug compatibility, and they may protect renal function and improve tubular function and renal pathology to a certain degree in UUO. This improvement may be related to a reduction in inflammatory cytokines CCL5 and CXCL10 secretion in the UUO rat kidney.

Chungsimyeonja-eum inhibits inflammatory responses in RAW 264.7 macrophages and HaCaT keratinocytes.

BACKGROUND: Chungsimyeonja-eum (CSYJE) is an herbal prescription used in traditional Oriental medicine for treating cerebral infarction by reducing ischemic damage. However, the effects of CSYJE on inflammation have not been verified scientifically. METHODS: Anti-inflammatory effects of CSYJE was investigated to dertermine the inhibitory effects of CSYJE against inflammation using RAW 264.7 mouse macrophages and HaCaT human keratinocytes. To measure the effects of CSYJE on inflammatory mediators and cytokines/chemokines, we used the following methods: cell viability assay, enzyme-linked immunosorbent assay (ELISA), western blotting, immunocytochemistry. RAW 264.7 cells were pretreated with CSYJE (250, 500, or 1000 µg/mL) for 4 h and treated with lipopolysaccharide (LPS) for additional 20 h. HaCaT cells were stimulated with tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ) (TI), and CSYJE (125, 250, or 500 µg/mL) for 24 h. RESULTS: CSYJE suppressed the production of nitric oxide (NO, IC50 1000 µg/mL), prostaglandin E2 (PGE2, IC50 = 12.1 µg/mL), and interleukin (IL)-6 (IC50 = 248 µg/mL) in LPS-stimulated RAW 264.7 cells. CSYJE suppressed the effects of TI on the production of thymus and activation-regulated chemokine (TARC, IC50 = 330.2 µg/mL), macrophage-derived chemokine (MDC/CCL22, IC50 = 52.5 µg/mL), regulated on activation, normal T-cell expressed and secreted (RANTES/CCL5, IC50 = 372.9 µg/mL), and IL-8 (IC50 = 345.1 µg/mL) in HaCaT cells. CSYJE inhibited TI-stimulated STAT1 phosphorylation in a dose-dependent manner and nuclear translocation at 500 µg/mL in HaCaT cells. CONCLUSION: Our results suggest a possible therapeutic application of CSYJE for treating inflammatory diseases.

Identification of KMU-3, a novel derivative of gallic acid, as an inhibitor of adipogenesis.

Differentiation of preadipocyte, also called adipogenesis, leads to the phenotype of mature adipocyte. Excessive adipogenesis, however, is largely linked to the development of obesity. Herein we investigated a library of 53 novel chemicals, generated from a number of polyphenolic natural compounds, on adipogenesis. Strikingly, among the chemicals tested, KMU-3, a derivative of gallic acid, strongly suppressed lipid accumulation during the differentiation of 3T3-L1 preadipocytes into adipocytes. On mechanistic levels, KMU-3 inhibited expressions of CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferator-activated receptor-γ (PPAR-γ), and fatty acid synthase (FAS) during adipocyte differentiation. Moreover, KMU-3 reduced expressions of adipokines, including retinol binding protein-4 (RBP-4), leptin, and regulated on activation, normal T cell expressed and secreted (RANTES) during adipocyte differentiation. Of further note, KMU-3 rapidly blocked the phosphorylation of signal transducer and activator of transcription-3 (STAT-3) during the early stage of adipogenesis. Importantly, pharmacological inhibition studies revealed that AG490, a JAK-2/STAT-3 inhibitor suppressed adipogenesis and STAT-3 phosphorylation, implying that early blockage of STAT-3 activity is crucial for the KMU-3-mediated anti-adipogenesis. These findings demonstrate firstly that KMU-3 inhibits adipogenesis by down-regulating STAT-3, PPAR-γ, C/EBP-α, and FAS. This work shows that KMU-3 is an inhibitor of adipogenesis and thus may have therapeutic potential against obesity.

Comparative effects of n-3, n-6 and n-9 unsaturated fatty acid-rich diet consumption on lupus nephritis, autoantibody production and CD4+ T cell-related gene responses in the autoimmune NZBWF1 mouse.

Mortality from systemic lupus erythematosus (SLE), a prototypical autoimmune disease, correlates with the onset and severity of kidney glomerulonephritis. There are both preclinical and clinical evidence that SLE patients may benefit from consumption of n-3 polyunsaturated fatty acids (PUFA) found in fish oil, but the mechanisms remain unclear. Here we employed the NZBWF1 SLE mouse model to compare the effects of dietary lipids on the onset and severity of autoimmune glomerulonephritis after consuming: 1) n-3 PUFA-rich diet containing docosahexaenoic acid-enriched fish oil (DFO), 2) n-6 PUFA-rich Western-type diet containing corn oil (CRN) or 3) n-9 monounsaturated fatty acid (MUFA)-rich Mediterranean-type diet containing high oleic safflower oil (HOS). Elevated plasma autoantibodies, proteinuria and glomerulonephritis were evident in mice fed either the n-6 PUFA or n-9 MUFA diets, however, all three endpoints were markedly attenuated in mice that consumed the n-3 PUFA diet until 34 wk of age. A focused PCR array was used to relate these findings to the expression of 84 genes associated with CD4+ T cell function in the spleen and kidney both prior to and after the onset of the autoimmune nephritis. n-3 PUFA suppression of autoimmunity in NZBWF1 mice was found to co-occur with a generalized downregulation of CD4+ T cell-related genes in kidney and/or spleen at wk 34. These genes were associated with the inflammatory response, antigen presentation, T cell activation, B cell activation/differentiation and leukocyte recruitment. Quantitative RT-PCR of representative affected genes confirmed that n-3 PUFA consumption was associated with reduced expression of CD80, CTLA-4, IL-10, IL-18, CCL-5, CXCR3, IL-6, TNF-α and osteopontin mRNAs in kidney and/or spleens as compared to mice fed n-6 PUFA or n-9 MUFA diets. Remarkably, many of the genes identified in this study are currently under consideration as biomarkers and/or biotherapeutic targets for SLE and other autoimmune diseases.

CCR1 inhibition ameliorates the progression of lupus nephritis in NZB/W mice.

Systemic lupus erythematosus is a chronic inflammatory autoimmune disease, the development of which is characterized by a progressive loss of renal function. Such dysfunction is associated with leukocyte infiltration in the glomerular and tubulointerstitial compartments in both human and experimental lupus nephritis. In this study, we investigated the role of the Ccr1 chemokine receptor in this infiltration process during the progression of nephritis in the lupus-prone New Zealand Black/New Zealand White (NZB/W) mouse model. We found that peripheral T cells, mononuclear phagocytes, and neutrophils, but not B cells, from nephritic NZB/W mice were more responsive to Ccr1 ligands than the leukocytes from younger prenephritic NZB/W mice. Short-term treatment of nephritic NZB/W mice with the orally available Ccr1 antagonist BL5923 decreased renal infiltration by T cells and macrophages. Longer Ccr1 blockade decreased kidney accumulation of effector/memory CD4(+) T cells, Ly6C(+) monocytes, and both M1 and M2 macrophages; reduced tubulointerstitial and glomerular injuries; delayed fatal proteinuria; and prolonged animal lifespan. In contrast, renal humoral immunity was unaffected in BL5923-treated mice, which reflected the unchanged numbers of infiltrated B cells in the kidneys. Altogether, these findings define a pivotal role for Ccr1 in the recruitment of T and mononuclear phagocyte cells to inflamed kidneys of NZB/W mice, which in turn contribute to the progression of renal injury.

Cannabidiol provides long-lasting protection against the deleterious effects of inflammation in a viral model of multiple sclerosis: a role for A2A receptors.

Inflammation in the central nervous system (CNS) is a complex process that involves a multitude of molecules and effectors, and it requires the transmigration of blood leukocytes across the blood-brain barrier (BBB) and the activation of resident immune cells. Cannabidiol (CBD), a non-psychotropic cannabinoid constituent of Cannabis sativa, has potent anti-inflammatory and immunosuppressive properties. Yet, how this compound modifies the deleterious effects of inflammation in TMEV-induced demyelinating disease (TMEV-IDD) remains unknown. Using this viral model of multiple sclerosis (MS), we demonstrate that CBD decreases the transmigration of blood leukocytes by downregulating the expression of vascular cell adhesion molecule-1 (VCAM-1), chemokines (CCL2 and CCL5) and the proinflammatory cytokine IL-1ß, as well as by attenuating the activation of microglia. Moreover, CBD administration at the time of viral infection exerts long-lasting effects, ameliorating motor deficits in the chronic phase of the disease in conjunction with reduced microglial activation and pro-inflammatory cytokine production. Adenosine A2A receptors participate in some of the anti-inflammatory effects of CBD, as the A2A antagonist ZM241385 partially blocks the protective effects of CBD in the initial stages of inflammation. Together, our findings highlight the anti-inflammatory effects of CBD in this viral model of MS and demonstrate the significant therapeutic potential of this compound for the treatment of pathologies with an inflammatory component.

Anti-inflammatory activity of Cymbopogon citratus leaves infusion via proteasome and nuclear factor-κB pathway inhibition: contribution of chlorogenic acid.

ETHNOPHARMACOLOGICAL RELEVANCE: Cymbopogon citratus (DC.) Stapf leaves infusion is used in traditional medicine for the treatment of inflammatory conditions, however little is known about their bioactive compounds. AIM OF THE STUDY: Investigate the compounds responsible for anti-inflammatory potential of Cymbopogon citratus (Cy) on cytokines production induced by lipopolysaccharide (LPS) in human and mouse macrophages, and the action mechanisms involved. MATERIALS AND METHODS: An essential oil-free infusion of Cy was prepared and polyphenol-rich fractions (PFs) were obtained from it by column chromatography. Chlorogenic acid (CGA) was identified, by HPLC/PDA/ESI-MS(n). The expression of cytokines, namely TNF-α and CCL5, was analyzed by real-time RT-PCR, on LPS-stimulated human macrophages. Activation of nuclear factor (NF)-κB, a master regulator of inflammation, was investigated by western blot and gene reporter assay. Proteasome activity was assessed using a fluorogenic peptide. RESULTS: Cymbopogon citratus extract and its polyphenols inhibited the cytokine production on human macrophages. This supports the anti-inflammatory activity of Cy polyphenols in physiologically relevant cells. Concerning the effect on the activation of NF-κB pathway, the results pointed to an inhibition of LPS-induced NF-κB activation by Cy and PFs. CGA was identified, by HPLC/PDA/ESI-MS(n), as the main phenolic acid of the Cy infusion, and it demonstrated to be, at least in part, responsible by that effect. Additionally, it was verified for the first time that Cy and PFs inhibited the proteasome activity, a complex that controls NF-κB activation, having CGA a strong contribution. CONCLUSIONS: The results evidenced, for the first time, the anti-inflammatory properties of Cymbopogon citratus through proteasome inhibition and, consequently NF-κB pathway and cytokine expression. Additionally, Cy polyphenols, in particular chlorogenic acid, were highlighted as bioactive compounds.

Pentosan polysulfate: a novel therapy for the mucopolysaccharidoses.

BACKGROUND: Pentosan polysulfate (PPS) is an FDA-approved, oral medication with anti-inflammatory and pro-chondrogenic properties. We have previously shown that animal models of the mucopolysaccharidoses (MPS) exhibit significant inflammatory disease, contributing to cartilage degeneration. Enzyme replacement therapy (ERT) only partly reduced inflammation, and anti-TNF-alpha antibody therapy significantly enhanced clinical and pathological outcomes. Here we describe the use of PPS for the treatment of MPS type VI rats. METHODOLOGY/PRINCIPAL FINDINGS: Treatment began during prenatal development and at 1 and 6 months of age. All animals were treated until they were 9 months old. Significant reductions in the serum and tissue levels of several inflammatory markers (e.g., TNF-alpha, MIP-1alpha and RANTES/CCL5) were observed, as was reduced expression of inflammatory markers in cultured articular chondrocytes. ADAMTS-5/aggrecanase-2 levels also were reduced in chondrocytes, consistent with an elevation of serum tissue inhibitor of metalloproteinase 1. Marked improvements in motility and grooming behavior occurred, along with a reduction in eye and nasal secretions and a lessening of the tracheal deformities. MicroCT and radiographic analyses further revealed that the treated MPS skulls were longer and thinner, and that the teeth malocclusions, misalignments and mineral densities were improved. MicroCT analysis of the femurs and vertebrae revealed improvements in trabecular bone mineral densities, number and spacing in a subset of treated MPS animals. Biomechanical assessments of PPS-treated spines showed partially restored torsional behaviors, suggesting increased spinal stability. No improvements were observed in cortical bone or femur length. The positive changes in the PPS-treated MPS VI rats occurred despite glycosaminoglycan accumulation in their tissues. CONCLUSIONS: Based on these findings we conclude that PPS could be a simple and effective therapy for MPS that might provide significant clinical benefits alone and in combination with other therapies.

Effects of neferine on CCL5 and CCR5 expression in SCG of type 2 diabetic rats.

Chemokines and their receptors have the key role in inflammatory responses. The phenomenon of low grade inflammation is associated with the development of type 2 diabetes. Postprandial hyperglycemia increases the systemic inflammatory responses, which promotes the development of type 2 diabetic associating autonomic nervous injuries or cardiovascular disease. Neferine is a bisbenzylisoquinline alkaloid isolated from a Chinese medicinal herb. The objectives of this study will examine the CCL5 and CCR5 expression in the superior cervical ganglion (SCG) of type 2 diabetic rats. The effects of neferine on the expression of CCL5 and CCR5 mRNA and protein in the superior cervical ganglion (SCG) of type 2 diabetic rats will also be observed. The studies showed that in type 2 diabetic rats, body weight, blood pressure, heart rates, fasting blood glucose, insulin, total cholesterol and triglyceride were enhanced and high density lipoprotein was decreased, and CCL5 and CCR5 expression levels in the SCG of type 2 diabetic rats were up-regulated. In type 2 diabetic rats treated with neferine, body weight, blood pressure, fasting blood glucose, insulin, total cholesterol and triglyceride were decreased and high density lipoprotein was increased. The elevated expressions of CCL5 and CCR5 in SCG were decreased after type 2 diabetic rats treated with neferine. The motor nerve conduction velocity (MNCV) in diabetic rats treated with neferine group showed a significantly increment in comparison with that in type 2 diabetic group. Neferine can decrease the expression of CCL5 and CCR5 in the SCG and reduce the SCG neuronal signaling mediated by CCL5 and CCR5 in regulating diabetic cardiovascular autonomic complications.

Reduction of RANTES expression in lesional psoriatic skin after narrow band ultraviolet therapy: a possible marker of therapeutic efficacy.

BACKGROUND: The regulated upon activation, normal T cell expressed and secreted (RANTES) production in psoriatic lesions may amplify the inflammation in these lesions. Narrow band ultraviolet B (NB-UVB), a therapeutic modality for psoriasis, affects the expression of inflammatory cytokines and chemokines. OBJECTIVE: Our aim was to evaluate RANTES mRNA expression in skin lesions of psoriasis before and after NB-UVB phototherapy. METHODS: This study included 25 psoriatic patients who received 24 sessions of NB-UVB. Skin biopsies were taken before and after phototherapy for real time PCR evaluation of RANTES mRNA. RESULTS: The relative quantitation values (RQ) of RANTES mRNA expression was significantly reduced after treatment. A significant negative correlation was found between pre-treatment RQ RANTES mRNA expression and post-treatment PASI score. We found a significant negative correlation between dRQ RANTES mRNA expression (difference between RQ RANTES mRNA expression before and after phototherapy) and PASI score after phototherapy. We found significant negative correlations between pre-treatment RQ RANTES mRNA expression and both initial response session number and total NB-UVB dose at the end of phototherapy. CONCLUSION: NB-UVB reduces RANTES mRNA expression in psoriatic lesions. Pre-treatment RQ RANTES mRNA expression could be considered as a marker for clinical improvement and NB-UVB phototherapy efficacy.

Inhibition of TNF-α/IFN-γ induced RANTES expression in HaCaT cell by naringin.

CONTEXT: Naringin is a bioflavonoid derivative and is predominantly found in Citrus paradisi Macf., Citrus sinensis (Linn.) Osbeck, Citrus unshiu Marc., Citrus reticulata Blanco cv. Nobilis, Citrus tachibana (Makino) Tanaka, Citrus junos Sieb. ex Tanaka (Rutaceae), and related citrus species. It has anti-inflammatory effects that have been well-documented, but the mechanism is poorly characterized. OBJECTIVE: The effect of naringin on production of RANTES (regulated upon activation normal T-cell expressed and secreted) in human HaCaT cells was investigated here for the first time. MATERIALS AND METHODS: The HaCaT cells were cultured in Dulbecco's modified Eagle's medium (DMEM) and the proliferation of cell was determined by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). The cells were divided into three groups including control group, tumor necrosis factor alpha (TNF-α)/interferon gamma (IFN-γ)-stimulated group, and naringin pretreatment group (first incubated in the presence of naringin and then exposed to TNF-α/IFN-γ). The concentration of RANTES in the supernatants was determined by enzyme-linked immunosorbent assay (ELISA). The expression of RANTES mRNA was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). The expression of nuclear factor kappa B (NF-κB) P65 protein was detected with immunocytochemical method and western blot method. RESULTS: Naringin hardly inhibits HaCaT cells growth at concentrations rising from 0.25 to 1 mmol/L. However, RANTES expression detected in supernatant stimulated with TNF-α/IFN-γ reduced 15 and 16%, respectively, when cultured with 0.25, 0.5 mmol/L naringin. Furthermore, 1 mmol/L naringin significantly decreased RANTES mRNA level. Finally, naringin decreased the expression of NF-κB P65 protein in nuclei. DISCUSSION AND CONCLUSION: Naringin can inhibit the increased production of RANTES, which is partially via NF-κB-dependent signal pathway.

S100B protein stimulates microglia migration via RAGE-dependent up-regulation of chemokine expression and release.

The Ca(2+)-binding protein of the EF-hand type, S100B, is abundantly expressed in and secreted by astrocytes, and release of S100B from damaged astrocytes occurs during the course of acute and chronic brain disorders. Thus, the concept has emerged that S100B might act an unconventional cytokine or a damage-associated molecular pattern protein playing a role in the pathophysiology of neurodegenerative disorders and inflammatory brain diseases. S100B proinflammatory effects require relatively high concentrations of the protein, whereas at physiological concentrations S100B exerts trophic effects on neurons. Most if not all of the extracellular (trophic and toxic) effects of S100B in the brain are mediated by the engagement of RAGE (receptor for advanced glycation end products). We show here that high S100B stimulates murine microglia migration in Boyden chambers via RAGE-dependent activation of Src kinase, Ras, PI3K, MEK/ERK1/2, RhoA/ROCK, Rac1/JNK/AP-1, Rac1/NF-κB, and, to a lesser extent, p38 MAPK. Recruitment of the adaptor protein, diaphanous-1, a member of the formin protein family, is also required for S100B/RAGE-induced migration of microglia. The S100B/RAGE-dependent activation of diaphanous-1/Rac1/JNK/AP-1, Ras/Rac1/NF-κB and Src/Ras/PI3K/RhoA/diaphanous-1 results in the up-regulation of expression of the chemokines, CCL3, CCL5, and CXCL12, whose release and activity are required for S100B to stimulate microglia migration. Lastly, RAGE engagement by S100B in microglia results in up-regulation of the chemokine receptors, CCR1 and CCR5. These results suggests that S100B might participate in the pathophysiology of brain inflammatory disorders via RAGE-dependent regulation of several inflammation-related events including activation and migration of microglia.

A botanical extract from channel flow inhibits cell proliferation, induces apoptosis, and suppresses CCL5 in human endometriotic stromal cells.

Growing evidence suggests that medicinal herbs have direct actions on endometrial cells. By screening multiple herbs using an in vitro model of endometriosis, we found that a commonly used herbal formula exerted considerable antiproliferative effects. Our purpose was to investigate the effects of this antiendometriosis herbal mixture on cell proliferation, apoptosis, and CCL5 expression and secretion in endometriotic stromal cells in vitro. Isolated normal endometrial, eutopic, and ectopic endometriotic stromal cells were cultured under established conditions. Cell proliferation, apoptosis, and CCL5 gene expression protein secretion was evaluated after incubation with different concentrations of an antiendometriosis herbal mixture extract. Cell proliferation was assessed by cell counting, (3)H-thymidine incorporation, and MTS assays. Apoptosis was determined by blotting using anti-cleaved caspase 3 antibodies and by a TUNEL assay. CCL5 gene expression and protein secretion were determined by transient transfection of gene promoter reporters and ELISAs in cell supernatants. Extracts of a traditional herbal mixture dose-dependently decreased cell proliferation in normal, eutopic, and ectopic endometriotic stromal cells. (3)H-Thymidine uptake and MTS confirmed these findings. The herbal extracts induced apoptosis, as evidenced by activation of caspase 3 and the presence of TUNEL-positive cells after treatment. The herbal extracts also suppressed CCL5 gene transcription and protein secretion in endometriotic stromal cells, even when corrected for cell number. Extracts from a medicinal herbal mixture have direct effects on cell proliferation, apoptosis, and CCL5 production in endometriotic stromal cells. Our findings support the further investigation of novel, potentially safe and well-tolerated botanical products as future endometriosis treatments.

Reducing the activity and secretion of microbial antioxidants enhances the immunogenicity of BCG.

BACKGROUND: In early clinical studies, the live tuberculosis vaccine Mycobacterium bovis BCG exhibited 80% protective efficacy against pulmonary tuberculosis (TB). Although BCG still exhibits reliable protection against TB meningitis and miliary TB in early childhood it has become less reliable in protecting against pulmonary TB. During decades of in vitro cultivation BCG not only lost some genes due to deletions of regions of the chromosome but also underwent gene duplication and other mutations resulting in increased antioxidant production. METHODOLOGY/PRINCIPAL FINDINGS: To determine whether microbial antioxidants influence vaccine immunogenicity, we eliminated duplicated alleles encoding the oxidative stress sigma factor SigH in BCG Tice and reduced the activity and secretion of iron co-factored superoxide dismutase. We then used assays of gene expression and flow cytometry with intracellular cytokine staining to compare BCG-specific immune responses in mice after vaccination with BCG Tice or the modified BCG vaccine. Compared to BCG, the modified vaccine induced greater IL-12p40, RANTES, and IL-21 mRNA in the spleens of mice at three days post-immunization, more cytokine-producing CD8+ lymphocytes at the peak of the primary immune response, and more IL-2-producing CD4+ lymphocytes during the memory phase. The modified vaccine also induced stronger secondary CD4+ lymphocyte responses and greater clearance of challenge bacilli. CONCLUSIONS/SIGNIFICANCE: We conclude that antioxidants produced by BCG suppress host immune responses. These findings challenge the hypothesis that the failure of extensively cultivated BCG vaccines to prevent pulmonary tuberculosis is due to over-attenuation and suggest instead a new model in which BCG evolved to produce more immunity-suppressing antioxidants. By targeting these antioxidants it may be possible to restore BCG's ability to protect against pulmonary TB.