Effect of ß-Alanine Supplementation on Monocyte Recruitment and Cognition During a 24-Hour Simulated Military Operation.

Wells, AJ, Varanoske, AN, Coker, NA, Kozlowski, GJ, Frosti, CL, Boffey, D, Harat, I, Jahani, S, Gepner, Y, and Hoffman, JR. Effect of ß-alanine supplementation on monocyte recruitment and cognition during a 24-hour simulated military operation. J Strength Cond Res 34(11): 3042-3054, 2020-Sustained military operations (SUSOPs) result in psychological stress and cognitive dysfunction, which may be related to the recruitment of classical monocytes into the brain. This study examined the effect of beta-alanine (BA) on cognition and monocyte recruitment during a simulated 24-hour SUSOP. Nineteen healthy men ingested 12-g/d BA or placebo for 14 days before an SUSOP. Monocyte chemoattractant protein-1 (MCP-1), C-C chemokine receptor-2 (CCR2), and macrophage-1-antigen (CD11b) expression were assessed through multiplex assay and flow cytometry. Psychological stress and cognition were assessed through Automated Neuropsychological Assessment Metrics (ANAM). A composite measure of cognition (COGcomp) was generated from throughput scores extracted from 7 ANAM cognitive tests. Assessments occurred at baseline (0H), 12 hours (12H), 18 hours (18H), and 24 hours (24H). Significance was accepted at p ≤ 0.05. No significant effect of BA was noted for any variable (p's > 0.05). The frequency and severity of symptoms of psychological stress increased significantly at 18 and 24H compared with 0 and 12H (p's < 0.05). COGcomp decreased significantly at 18 and 24H compared with 0 and 12H (p's ≤ 0.001). MCP-1 peaked at 18H was significantly lower at 24H compared with 18H but remained elevated at 24H compared with 0H (p's < 0.001). CCR2 expression was significantly lower at 12 (p = 0.031), 18, and 24H (p's < 0.001). CD11b expression was significantly higher at 12H (p = 0.039) and 24H (p's = 0.003). MCP-1 was negatively associated with COGcomp (ß = -0.395, p = 0.002, r2 = 0.174). Neither CCR2 or CD11b was related to COGcomp (p's > 0.05). Cognitive dysfunction during SUSOPs is related to serum concentrations of MCP-1 but is not influenced by BA supplementation.

Forsythoside A inhibits adhesion and migration of monocytes to type II alveolar epithelial cells in lipopolysaccharide-induced acute lung injury through upregulating miR-124.

Acute lung injury (ALI) is a severe disease for which effective drugs are still lacking at present. Forsythia suspensa is a traditional Chinese medicine commonly used to relieve respiratory symptoms in China, but its functional mechanisms remain unclear. Therefore, forsythoside A (FA), the active constituent of F. suspensa, was studied in the present study. Inflammation models of type II alveolar epithelial MLE-12 cells and BALB/c mice stimulated by lipopolysaccharide (LPS) were established to explore the effects of FA on ALI and the underlying mechanisms. We found that FA inhibited the production of monocyte chemoattractant protein-1 (MCP-1/CCL2) in LPS-stimulated MLE-12 cells in a dose-dependent manner. Moreover, FA decreased the adhesion and migration of monocytes to MLE-12 cells. Furthermore, miR-124 expression was upregulated after FA treatment. The luciferase report assay showed that miR-124 mimic reduced the activity of CCL2 in MLE-12 cells. However, the inhibitory effects of FA on CCL2 expression and monocyte adhesion and migration to MLE-12 cells were counteracted by treatment with a miR-124 inhibitor. Critically, FA ameliorated LPS-induced pathological damage, decreased the serum levels of tumor necrosis factor-α and interleukin-6, and inhibited CCL2 secretion and macrophage infiltration in lungs in ALI mice. Meanwhile, administration of miR-124 inhibitor attenuated the protective effects of FA. The present study suggests that FA attenuates LPS-induced adhesion and migration of monocytes to type II alveolar epithelial cells though upregulating miR-124, thereby inhibiting the expression of CCL2. These findings indicate that the potential application of FA is promising and that miR-124 mimics could also be used in the treatment of ALI.

Discovery of a Bromodomain and Extraterminal Inhibitor with a Low Predicted Human Dose through Synergistic Use of Encoded Library Technology and Fragment Screening.

The bromodomain and extraterminal (BET) family of bromodomain-containing proteins are important regulators of the epigenome through their ability to recognize N-acetyl lysine (KAc) post-translational modifications on histone tails. These interactions have been implicated in various disease states and, consequently, disruption of BET-KAc binding has emerged as an attractive therapeutic strategy with a number of small molecule inhibitors now under investigation in the clinic. However, until the utility of these advanced candidates is fully assessed by these trials, there remains scope for the discovery of inhibitors from new chemotypes with alternative physicochemical, pharmacokinetic, and pharmacodynamic profiles. Herein, we describe the discovery of a candidate-quality dimethylpyridone benzimidazole compound which originated from the hybridization of a dimethylphenol benzimidazole series, identified using encoded library technology, with an N-methyl pyridone series identified through fragment screening. Optimization via structure- and property-based design led to I-BET469, which possesses favorable oral pharmacokinetic properties, displays activity in vivo, and is projected to have a low human efficacious dose.

Naringenin suppresses neutrophil infiltration into adipose tissue in high-fat diet-induced obese mice.

Recruitment of immune cells to adipose tissue is altered dramatically in obesity, which results in chronic inflammation of the adipose tissue that leads to metabolic disorders, such as insulin resistance and type 2 diabetes mellitus. The regulation of immune cell infiltration into adipose tissue has prophylactic and therapeutic implications for obesity-related diseases. We previously showed that naringenin, a citrus flavonoid, suppressed macrophage infiltration into adipose tissue by inhibiting monocyte chemoattractant protein-1 (MCP-1) expression in the progression phase to high-fat diet (HFD)-induced obesity. In the current study, we evaluated the effects of naringenin on neutrophil infiltration into adipose tissue, because neutrophils also infiltrate into adipose tissue in the progression phase to obesity. Naringenin suppressed neutrophil infiltration into adipose tissue induced by the short-term (2 weeks) feeding of a HFD to mice. Naringenin tended to inhibit the HFD-induced expression of several chemokines, including MCP-1 and MCP-3, in adipose tissue. Naringenin also inhibited MCP-3 expression in 3T3-L1 adipocytes and a co-culture of 3T3-L1 adipocytes and RAW264 macrophages. However, naringenin did not affect the expression of macrophage inflammatory protein-2 (MIP-2), an important chemokine for neutrophil migration and activation, in macrophages or in a co-culture of adipocytes and macrophages. Our results suggest that naringenin suppresses neutrophil infiltration into adipose tissue via the regulation of MCP-3 expression and macrophage infiltration.

CCL2/CCR2 Chemokine System in Embryonic Hypothalamus: Involvement in Sexually Dimorphic Stimulatory Effects of Prenatal Ethanol Exposure on Peptide-Expressing Neurons.

Maternal consumption of ethanol during pregnancy is known to increase the offspring's risk for developing alcohol use disorders and associated behavioral disturbances. Studies in adolescent and adult animals suggest the involvement of neuroimmune and neurochemical systems in the brain that control these behaviors. To understand the origin of these effects during early developmental stages, we examined in the embryo and neonate the effects of maternal intraoral administration of ethanol (2 g/kg/day) from embryonic day 10 (E10) to E15 on the inflammatory chemokine C-C motif ligand 2 (CCL2) and its receptor CCR2 in a specific, dense population of neurons in the lateral hypothalamus (LH), where they are closely related to an orexigenic neuropeptide, melanin-concentrating hormone (MCH), known to promote ethanol consumption and related behaviors. We found that prenatal ethanol exposure increases the expression and density of CCL2 and CCR2 cells along with MCH neurons in the LH and the colocalization of CCL2 with MCH. We also discovered that these effects are sexually dimorphic, consistently stronger in female embryos, and are blocked by maternal administration of a CCL2 antibody (1 and 5 µg/day, i.p., E10-E15) that neutralizes endogenous CCL2 and of a CCR2 antagonist INCB3344 (1 mg/day, i.p., E10-E15) that blocks CCL2's main receptor. These results, which in the embryo anatomically and functionally link the CCL2/CCR2 system to MCH neurons in the LH, suggest an important role for this neuroimmune system in mediating ethanol's sexually dimorphic, stimulatory effect on MCH neurons that may promote higher level of alcohol consumption described in females.

Effect of berberine on lipopolysaccharide-induced monocyte chemotactic protein-1 and interleukin-8 expression in a human retinal pigment epithelial cell line.

PURPOSE: In this study, we elucidated the effects of berberine, a major alkaloid component contained in medicinal herbs, such as Phellodendri Cortex and Coptidis Rhizoma, on expression of monocyte chemotactic protein-1 (MCP-1) and interleukin-8 (IL-8) in a human retinal pigment epithelial cell line (ARPE-19) caused by lipopolysaccharide (LPS) stimulation. METHODS: ARPE-19 cells were cultured to confluence. Berberine and LPS were added to the medium. MCP-1 and IL-8 mRNA were measured by real-time polymerase chain reaction. MCP-1 and IL-8 protein concentrations in the media were measured using enzyme-linked immunosorbent assay. RESULTS: After stimulation with LPS, MCP-1 and IL-8 mRNA in ARPE-19 cells reached maximum levels at 3 h, and MCP-1 and IL-8 protein in the culture media reached maximum levels at 24 h. Berberine dose-dependently inhibited MCP-1 and IL-8 mRNA expression of the cells and protein levels in the media stimulated with LPS. CONCLUSIONS: These findings indicate that berberine inhibited the expression of MCP-1 and IL-8 induced by LPS.

Baicalin attenuates non-alcoholic steatohepatitis by suppressing key regulators of lipid metabolism, inflammation and fibrosis in mice.

AIMS: Baicalin (BA), an active flavonoid compound originating from the herb of Scutellaria baicalensis Georgi, has been previously shown to exert anti-inflammation and anti-oxidant effects in liver diseases. However, the potential role of BA in the regulation of non-alcoholic steatohepatitis (NASH) remains elusive. In this study, we newly explored the hepatoprotective effects of BA in MCD diet-induced NASH by ameliorating hepatic steatosis, inflammation, fibrosis and apoptosis. MAIN METHODS: NASH was induced in mice fed a methionine and choline-deficient (MCD) diet for 4weeks. The mice were simultaneously treated with or without BA for 4weeks. Serum liver functional markers and inflammatory indicators were assessed by biochemical and ELISA methods, respectively. The livers were histologically examined using H&E, Oil Red O and Masson's trichrome staining methods. The qRT-PCR, IHC and Western blotting assays were applied to analyze mechanisms underlying BA protection. KEY FINDINGS: BA treatment significantly attenuated MCD diet-induced hepatic lipid accumulation partly through regulating the expression of SREBP-1c, FASN, PPARα and CPT1a. BA treatment dramatically suppressed MCD diet-induced hepatic inflammation, which was associated with decrease in serum TNF-α, IL-1ß and MCP-1 production, macrophage influx and suppression of nuclear factor-κB activation. Additionally, BA was proved to prevent liver fibrosis, which appears to be mediated by inhibition of α-SMA, TGF-ß1 and Col1A1. Furthermore, BA markedly inhibited hepatocyte apoptosis and cleaved caspase-3 protein expression in MCD diet-induced mice. SIGNIFICANCE: These results provide a possible basis of the underlying mechanism for the application of BA in the treatment of NASH.

Therapeutic Strategies of Plant-derived Compounds for Diabetes Via Regulation of Monocyte Chemoattractant Protein-1.

BACKGROUND: Monocyte chemoattractant protein-1 (MCP-1) is a member of the CC chemokine family that plays a key role in the inflammatory process. It has been broadly studied in the aspect of its role in obesity and diabetes related diseases. MCP-1 causes the infiltration of macrophages into obese adipose tissue via binding to the CCR2 receptor and is involved in the development of insulin resistance. METHODS: We reviewed the available literature regarding the importance of plant metabolites that regulate MCP-1 activity and are used in the treatment of diabetic disorders. The characteristics of screened papers were described and the important findings were included in this review. RESULTS: This mini-review provides a summary of functions and therapeutic strategies of this chemokine, with a special focus on plant-derived compounds that possess a putative antidiabetic function via a mechanism of MCP-1 interaction. The highlights of this review include the roles of MCP-1 in development of diabetes, the evaluation of plant metabolites that specifically or non-specifically inhibit MCP-1 overproduction, and the molecular mechanisms of this activity. Among these metabolites, we particularly focused on phenolic acids and their derivatives, flavonoids, stilbenes, anthocyanins, capsaicin, alkaloids, plant sterols, terpenes, saponins, unsaturated fatty acids and plant-derived extracts. CONCLUSION: Regarding the increasing number of diabetic patients yearly, the recent progress in the putative therapies needs to be summarized. This article underlines the significance and involvement of the chemokine MCP-1 in the development of obesity, type 2 diabetes, and diabetic complications, with an emphasis on the role of plant metabolites in the regulation of this chemokine and thus the role in the prevention or therapy of diabetes. We suggest that MCP-1 might be a molecular marker of type 2 diabetes.

Anti-cancer effect of danshen and dihydroisotanshinone I on prostate cancer: targeting the crosstalk between macrophages and cancer cells via inhibition of the STAT3/CCL2 signaling pathway.

Danshen (Salvia miltiorrhiza Bunge) is widely used in traditional Chinese medicine. In our study, the in vivo protective effect of danshen in prostate cancer patients was validated through data from the National Health Insurance Research Database in Taiwan. In vitro, we discovered that dihydroisotanshinone I (DT), a bioactive compound present in danshen, can inhibit the migration of both androgen-dependent and androgen-independent prostate cancer cells. In addition, we noted that DT substantially inhibited the migratory ability of prostate cancer cells in both a macrophage-conditioned medium and macrophage/prostate cancer coculture medium. Mechanistically, DT both diminished the ability of prostate cancer cells to recruit macrophages and reduced the secretion of chemokine (C-C motif) ligand 2 (CCL2) from both macrophages and prostate cancer cells in a dose-dependent manner. Moreover, DT inhibited the protein expression of p-STAT3 and decreased the translocation of STAT3 into nuclear chromatin. DT also suppressed the expression of tumor epithelial-mesenchymal transition genes, including RhoA and SNAI1. In conclusion, danshen can prolong the survival rate of prostate cancer patients in Taiwan. Furthermore, DT can inhibit the migration of prostate cancer cells by interrupting the crosstalk between prostate cancer cells and macrophages via the inhibition of the CCL2/STAT3 axis. These results may provide the basis for a new therapeutic approach toward the treatment of prostate cancer progression.

Dissection of the host-pathogen interaction in human tuberculosis using a bioengineered 3-dimensional model.

Cell biology differs between traditional cell culture and 3-dimensional (3-D) systems, and is modulated by the extracellular matrix. Experimentation in 3-D presents challenges, especially with virulent pathogens. Mycobacterium tuberculosis (Mtb) kills more humans than any other infection and is characterised by a spatially organised immune response and extracellular matrix remodelling. We developed a 3-D system incorporating virulent mycobacteria, primary human blood mononuclear cells and collagen-alginate matrix to dissect the host-pathogen interaction. Infection in 3-D led to greater cellular survival and permitted longitudinal analysis over 21 days. Key features of human tuberculosis develop, and extracellular matrix integrity favours the host over the pathogen. We optimised multiparameter readouts to study emerging therapeutic interventions: cytokine supplementation, host-directed therapy and immunoaugmentation. Each intervention modulates the host-pathogen interaction, but has both beneficial and harmful effects. This methodology has wide applicability to investigate infectious, inflammatory and neoplastic diseases and develop novel drug regimes and vaccination approaches.

Repurposing auranofin for the treatment of cutaneous staphylococcal infections.

The scourge of multidrug-resistant bacterial infections necessitates the urgent development of novel antimicrobials to address this public health challenge. Drug repurposing is a proven strategy to discover new antimicrobial agents; given that these agents have undergone extensive toxicological and pharmacological analysis, repurposing is an effective method to reduce the time, cost and risk associated with traditional antibiotic innovation. In this study, the in vitro and in vivo antibacterial activities of an antirheumatic drug, auranofin, was investigated against multidrug-resistant Staphylococcus aureus. The results indicated that auranofin possesses potent antibacterial activity against all tested strains of S. aureus, including meticillin-resistant S. aureus (MRSA), vancomycin-intermediate S. aureus (VISA) and vancomycin-resistant S. aureus (VRSA), with minimum inhibitory concentrations (MICs) ranging from 0.0625µg/mL to 0.125µg/mL. In vivo, topical auranofin proved superior to conventional antimicrobials, including fusidic acid and mupirocin, in reducing the mean bacterial load in infected wounds in a murine model of MRSA skin infection. In addition to reducing the bacterial load, topical treatment of auranofin greatly reduced the production of inflammatory cytokines, including tumour necrosis factor-α (TNFα), interleukin-6 (IL-6), interleukin-1 beta (IL-1ß) and monocyte chemoattractant protein-1 (MCP-1), in infected skin lesions. Moreover, auranofin significantly disrupted established in vitro biofilms of S. aureus and Staphylococcus epidermidis, more so than the traditional antimicrobials linezolid and vancomycin. Taken together, these results support that auranofin has potential to be repurposed as a topical antimicrobial agent for the treatment of staphylococcal skin and wound infections.

Intrathecal curcumin attenuates pain hypersensitivity and decreases spinal neuroinflammation in rat model of monoarthritis.

Curcumin is a major component of turmeric and reportedly has anti-inflammatory and anti-oxidant effects. Neuroinflammation has been recognized to play an important role in the pathogenesis of various diseases in the central nervous system. Here we investigated the anti-nociceptive and anti-neuroinflammatory effect of curcumin on arthritic pain in rats. We found that repeated oral treatment with curcumin, either before or after complete Freund's adjuvant (CFA) injection, dose-dependently attenuated CFA-induced mechanical allodynia and thermal hyperalgesia, but had no effect on joint edema. Repeated intrathecal injection of curcumin reversed CFA-induced pain hypersensitivity. Furthermore, such a curcumin treatment reduced CFA-induced activation of glial cells and production of inflammatory mediators [interleukin-1ß (IL-1ß), monocyte chemoattractant protein-1 (MCP-1), and monocyte inflammatory protein-1 (MIP-1α)] in the spinal cord. Curcumin also decreased lipopolysaccharide-induced production of IL-1ß, tumor necrosis factor-α, MCP-1, and MIP-1α in cultured astrocytes and microglia. Our results suggest that intrathecal curcumin attenuates arthritic pain by inhibiting glial activation and the production of inflammatory mediators in the spinal cord, suggesting a new application of curcumin for the treatment of arthritic pain.

Haemanthus coccineus extract and its main bioactive component narciclasine display profound anti-inflammatory activities in vitro and in vivo.

Haemanthus coccineus extracts (HCE) have traditionally been used to treat a variety of diseases, like febrile colds or asthma. Since new therapeutic options against inflammatory processes are still urgently needed, we aimed to pharmacologically characterise the anti-inflammatory potential of HCEin vitro and in vivo and to identify the underlying bioactive component(s). The action of HCE on oedema formation and leucocyte infiltration were analysed in two murine models of inflammation (dermal oedema induced by arachidonic acid and croton oil; kidney injury caused by unilateral ureteral obstruction). The interaction of leucocytes with endothelial cells (ECs) as well as the activation parameters of these two cell types were analysed. Moreover, the nuclear factor κB (NFκB) pathway was investigated in detail in ECs. Using different fractions of HCE, the bioactive principle was identified. In vivo, HCE (450 mg/kg orally or 2 mg/kg intraperitoneally) inhibited oedema formation, leucocyte infiltration and cytokine synthesis. In vitro, HCE (100-300 ng/ml) blocked leucocyte-EC interaction as well as the activation of isolated leucocytes (cytokine synthesis and proliferation) and of primary ECs (adhesion molecule expression). HCE suppressed NFκB-dependent gene transcription in the endothelium, but did not interfere with the NFκB activation cascade (IκB degradation, p65 nuclear translocation and NFκB DNA-binding activity). The alkaloid narciclasine was elucidated as the bioactive compound responsible for the anti-inflammatory action of HCE. Our study highlights HCE and its main alkaloid narciclasine as novel interesting approach for the treatment of inflammation-related disorders.

Protective effects of Sparstolonin B, a selective TLR2 and TLR4 antagonist, on mouse endotoxin shock.

Sepsis is characterized by an overwhelming systemic inflammation and multiple organ injury. Toll-like receptors (TLRs) 2 and 4 mediate these inflammatory responses. Sparstolonin B (SsnB), isolated from Chinese herb Scirpus yagara, is a new selective TLR2/4 antagonist. Herein, we report that SsnB inhibited the expression of various inflammatory mediators such as tumor necrosis factor (TNF-α), interleukin (IL)-1ß, IL-6, and chemokine (C-C motif) ligand 2 (CCL-2) in lipopolysaccharide (LPS)- or Pam3csk4-stimulated macrophages. Moreover, in LPS-stimulated macrophages, the downregulation of peroxisome proliferator-activated receptor γ (PPAR-γ) was reversed by SsnB dose-dependently; and SsnB had synergistic inhibitory effects with rosiglitazone, a PPAR-γ agonist, on TNF-α and IL-6 expression in LPS-stimulated macrophages. The effects of SsnB were further evaluated in a mouse endotoxin shock model. When intraperitoneal injected in mice 2 days before or 1-2h after LPS challenge, SsnB attenuated the body temperature reduction and decreased the mortality. SsnB pre-treatment significantly suppressed LPS-induced increase of TNF-α and IL-6 in serum, lungs and livers, and substantially attenuated lung dysfunction in mice. In vivo toxicity test showed that at doses as high as 500 mg/kg, SsnB did not cause death of mice. These results suggest that SsnB protects mice against endotoxin shock by inhibiting production of multiple cytokines and lung dysfunction. In conclusion, our findings indicate that SsnB may be used in the prevention and treatment of endotoxin shock.

Tomato extract suppresses the production of proinflammatory mediators induced by interaction between adipocytes and macrophages.

Obese adipose tissue is characterized by enhanced macrophage infiltration. A loop involving monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNFα) between adipocytes and macrophages establishes a vicious cycle that augments inflammatory changes and insulin resistance in obese adipose tissue. Tomatoes, one of the most popular crops worldwide, contain many beneficial phytochemicals that improve obesity-related diseases such as diabetes. Some of them have also been reported to have anti-inflammatory properties. In this study, we focused on the potential protective effects of phytochemicals in tomatoes on inflammation. We screened fractions of tomato extract using nitric oxide (NO) assay in lipopolysaccharide (LPS)-stimulated RAW264 macrophages. One fraction, RF52, significantly inhibited NO production in LPS-stimulated RAW264 macrophages. Furthermore, RF52 significantly decreased MCP-1 and TNFα productions. The coculture of 3T3-L1 adipocytes and RAW264 macrophages markedly enhanced MCP-1, TNFα, and NO productions compared with the control cultures; however, the treatment with RF52 inhibited the production of these proinflammatory mediators. These results suggest that RF52 from tomatoes may have the potential to suppress inflammation by inhibiting the production of NO or proinflammatory cytokines during the interaction between adipocytes and macrophages.

Inhibitory effects of teuclatriol, a sesquiterpene from salvia mirzayanii, on nuclear factor-κB activation and expression of inflammatory mediators.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia mirzayanii Rech. f. & Esfand. is an endemic plant, which is only distributed in the south of Iran. In traditional Iranian medicine, the aerial parts of Salvia mirzayanii have been used for infections, inflammatory diseases, and as a tonic. From this plant, the sesquiterpene teuclatriol was isolated by bioactivity-guided fractionation due to its anti-proliferative actions on human lymphocytes. The guaiane sesquiterpene is lacking the methylene-γ-lactone function that is typically involved in the inhibiting properties of sesquiterpenes on NF-κB, a pivotal transcription factor in inflammatory processes. We here investigated anti-inflammatory effects of teuclatriol on human macrophage-like and endothelial cells. MATERIALS AND METHODS: Non-toxic doses of teuclatriol were determined for both cell types by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)-assay. The effect of teuclatriol on the activity of NF-κB in LPS-stimulated human monocytic THP-1 cells was studied using infrared electrophoretic mobility shift assay (IR-EMSA) using curcumin as positive control (32µM). THP-1 were differentiated into macrophage-like cells and evaluated for TNF-α secretion by L929 bioassay following stimulation with LPS and treatment with teuclatriol. Inflammatory gene expression in human umbilical vein endothelial cells (HUVEC), modeling target cells for TNF-α-induced inflammatory gene activation, was investigated by real-time RT-PCR. RESULTS: The LPS-induced DNA binding activity of NF-κB in THP-1 was significantly decreased by non-toxic doses of teuclatriol (312 and 390µM). Teuclatriol reduced the production of TNF-α in a dose-dependent manner. mRNA levels of both monocyte chemoattractant protein (MCP)-1 and toll-like receptor (TLR)2 were decreased in TNF-α-activated HUVEC. CONCLUSION: These data show an inhibitory effect of teuclatriol on NF-κB signaling at doses of 312µM and higher, validating the traditional use of Salvia mirzayanii in the treatment of inflammatory diseases. Future work on the mode of action of teuclatriol may provide new lead structures with NF-κB inhibiting properties, lacking possible side effects mediated via alkylating centers of sesquiterpene lactones.

Green tea polyphenol epigallocatechin-3-gallate inhibits TNF-α-induced production of monocyte chemoattractant protein-1 in human umbilical vein endothelial cells.

AIMS: Epigallocatechin-3-gallate (EGCG), a major catechin found in green tea, displays a variety of pharmacological properties and recently received attention as a prospective dietary intervention in cardiovascular diseases (CVD). This study was conducted to test the hypothesis that EGCG was able to inhibit tumor necrosis factor-α (TNF-α)-induced production of monocyte chemoattractant protein-1 (MCP-1) in human umbilical vein endothelial cells (HUVECs) and investigated the underlying molecular mechanisms. METHODS: The inhibitory effect of EGCG on TNF-α-induced expression of MCP-1 was measured using ELISA and RT-qPCR. The effect of EGCG on TNF-α-induced nuclear factor-kappa B (NF-κB) activation was investigated by western blot and luciferase assays. Monocyte adhesion assay was detected by microscope. RESULTS: EGCG significantly suppressed the TNF-α-induced protein and mRNA expression of MCP-1. Investigation of the mechanism suggested that EGCG suppressed the TNF-α-mediated NF-κB activation. In addition, the 67-kD laminin receptor (67LR) was involved in EGCG-mediated suppression of MCP-1 generation. Furthermore, EGCG potently inhibited monocyte adhesion to activated HUVECs. CONCLUSION: EGCG suppresses TNF-α-induced MCP-1 expression in HUVECs. This effect was mediated by 67LR and was via the inhibition of NF-κB activation. Our results demonstrated that EGCG might be a possible medicine for CVD prevention and treatment.

The effects of dexpanthenol in streptozotocin-induced diabetic rats: histological, histochemical and immunological evidences.

This study was designed to investigate the effects of Dexpanthenol (Dxp) on liver and pancreas histology and cytokine levels in streptozotocine (STZ)-induced diabetic rats. Twenty-four Wistar albino male rats were divided into four groups: control, Dxp, STZ-induced diabetic (STZ) and diabetic treatment with Dexpanthenol (STZ-Dxp) groups. Experimental diabetes was induced by single dose STZ (50 mg/kg) intraperitoneally (i.p.). After administration of STZ, the STZ-Dxp group began to receive a 300 mg/kg/day i.p. dose of Dxp for 6 weeks. Liver and pancreas tissues of the control group were in normal morphology. Liver tissue of STZ group showed vacuolisation of hepatocytes in the liver parenchyma with enlargement of sinusoidal spaces and increasing amounts of connective tissue in the portal area. Pancreatic section of STZ group displayed ß-cells with of cytoplasmic mass, reduction of islet size, and atrophy. The STZ-Dxp group that received Dxp treatment exhibit partially normal hepatic parenchyma. Histochemical examinations revealed that the diabetes-induced glycogen depletion markedly improved with the Dxp treatment (p⟨0.001). The severity of degenerative alteration was lessened by Dxp supplementation in the STZ-Dxp group. Induction of STZ presented a significant increase both in interleukin-1α (IL-1α) (p=0.033) and monocyte chemotactic protein-1 (MCP-1) (p=0.011) levels, when compared with the control rats. DXP-treated diabetic rats' IL-1α and MCP-1 levels were similar to control value. This evidence suggests that Dxp is effective in reducing STZ-induced, diabetic-related complications and may be beneficial for the treatment of diabetic patients.

4ß-Hydroxywithanolide E isolated from Physalis pruinosa calyx decreases inflammatory responses by inhibiting the NF-κB signaling in diabetic mouse adipose tissue.

BACKGROUND: Chronic inflammation in adipose tissue together with obesity induces insulin resistance. Inhibitors of chronic inflammation in adipose tissue can be a potent candidate for the treatment of diabetes; however, only a few compounds have been discovered so far. The objective of this study was to find a novel inhibitor that can suppress the inflammatory response in adipose tissue and to elucidate the intracellular signaling mechanisms of the compound. METHODS: To find the active compounds, we established an assay system to evaluate the inhibition of induced MCP-1 production in adipocyte/macrophage coculture in a plant extract library. The active compound was isolated by performing high-performance liquid chromatography (HPLC) and was determined as 4ß-hydroxywithanolide E (4ßHWE) by nuclear magnetic resonance (NMR) and mass spectroscopy (MS) spectral analyses. The effect of 4ßHWE on inflammation in adipose tissue was assessed with adipocyte culture and db/db mice. RESULTS: During the screening process, Physalis pruinosa calyx extract was found to inhibit production of MCP-1 in coculture strongly. 4ßHWE belongs to the withanolide family of compounds, and it has the strongest MCP-1 production inhibitory effect and lowest toxicity than any other withanolides in coculture. Its anti-inflammatory effect was partially dependent on the attenuation of NF-κB signaling in adipocyte. Moreover, in vivo experiments showed that the oral administration of 4ßHWE to db/db mice resulted in the inhibition of macrophage invasion and cytokine expression in adipose tissue after 2 weeks of treatment; improved the plasma adiponectin, non-esterified fatty acids and MCP-1 concentrations; and increased glucose tolerance after 3 to 4 weeks of treatment. CONCLUSIONS: These results suggest that 4ßHWE has anti-inflammatory effect via inhibition of NF-κB activation in adipocyte. Moreover, the attenuation of inflammation in adipocyte has an effect on the inhibition of macrophage accumulation in obese adipose tissue. Consequently, 4ßHWE improves impaired glucose tolerance. Thus, 4ßHWE is a useful natural anti-inflammatory compound to attenuate progression of diabetes and obesity.

Topical hypochlorite ameliorates NF-κB-mediated skin diseases in mice.

Nuclear factor-κB (NF-κB) regulates cellular responses to inflammation and aging, and alterations in NF-κB signaling underlie the pathogenesis of multiple human diseases. Effective clinical therapeutics targeting this pathway remain unavailable. In primary human keratinocytes, we found that hypochlorite (HOCl) reversibly inhibited the expression of CCL2 and SOD2, two NF-κB-dependent genes. In cultured cells, HOCl inhibited the activity of inhibitor of NF-κB kinase (IKK), a key regulator of NF-κB activation, by oxidizing cysteine residues Cys114 and Cys115. In NF-κB reporter mice, topical HOCl reduced LPS-induced NF-κB signaling in skin. We further evaluated topical HOCl use in two mouse models of NF-κB-driven epidermal disease. For mice with acute radiation dermatitis, topical HOCl inhibited the expression of NF-κB-dependent genes, decreased disease severity, and prevented skin ulceration. In aged mice, topical HOCl attenuated age-dependent production of p16INK4a and expression of the DNA repair gene Rad50. Additionally, skin of aged HOCl-treated mice acquired enhanced epidermal thickness and proliferation, comparable to skin in juvenile animals. These data suggest that topical HOCl reduces NF-κB-mediated epidermal pathology in radiation dermatitis and skin aging through IKK modulation and motivate the exploration of HOCl use for clinical aims.