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Cystectomy is the removal of all or part of the urinary bladder. It has been observed that there is significant regrowth of the bladder after partial cystectomy and this has been proposed to be through regeneration of the organ. Regrowth of tissue in mammals has been proposed to involve compensatory mechanisms that share many characteristics of true regeneration, like the growth of specialized structures such as blood vessels or nerves. However, the overall structure of the normal organ is not achieved. Here we tested if bladder growth after subtotal cystectomy (STC, removal of 50% of the bladder) was compensatory or regenerative. To do this we subjected adult female mouse bladders to STC and assessed regrowth using several established cellular parameters including histological, gene expression, cytokine accumulation and cell proliferation studies. Bladder function was analyzed using cystometry and the voiding stain on paper (VSOP) technique. We found that STC bladders were able to increase their ability to hold urine with the majority of volume restoration occurring within the first two weeks. Regenerating bladders had thinner walls with less mean muscle thickness, and they showed increased collagen deposition at the incision as well as throughout the bladder wall suggesting that fibrosis was occurring. Cell populations differed in their response to injury with urothelial regeneration complete by day 7, but stromal and detrusor muscle still incomplete after 8wks. Cells incorporated EdU when administered at the time of surgery and tracing of EdU positive cells over time indicated that many newborn cells originate at the incision and move mediolaterally. Basal urothelial cells and bladder mesenchymal stem cells but not smooth muscle cells significantly incorporated EdU after STC. Since anti-inflammatory cytokines play a role in regeneration, we analyzed expressed cytokines and found that no anti-inflammatory cytokines were present in the bladder 1wk after STC. Our findings suggest that bladder regrowth after cystectomy is compensatory and functions to increase the volume that the bladder can hold. This finding sets the stage for understanding how the bladder responds to cystectomy and how this can be improved in patients after suffering bladder injury. [ABSTRACT FROM AUTHOR]

Due to its strong proliferation-reducing effects on keratinocytes, and also anti-inflammatory properties, the isoflavone genistein has already been proposed as a possible antipsoriatic compound. As there is still no detailed information on this topic, we examined the effects of genistein by using an in vitro model of both, normal and “psoriasis-like” keratinocytes at this stage of our work exhaustively testing the selected flavonoid in a mono-treated experimental design. Gene expression studies revealed transcriptional changes that confirms known disease-associated pathways and highlights many psoriasis-related genes. Our results suggested that aberrant expression of genes contributing to the progress of psoriasis could be improved by the action of genistein. Genistein prevented “cytokine mix” as well as TNF-α-induced NF-κB nuclear translocation, with no effect on the PI3K signaling cascade, indicating the luck of turning this pathway into NF-κB activation. It could have attenuated TNF-α and LPS-induced inflammatory responses by suppressing ROS activation. Regardless of the type of keratinocyte stimulation used, reduction of cytokine IL-8, IL-20 and CCL2 production (both at RNA and protein level) following genistein treatment was visible. Because investigations of other groups supported our commentary on potential administration of genistein as a potential weapon in the armamentarium against psoriasis, it is believed that this paper should serve to encourage researchers to conduct further studies on this subject. [ABSTRACT FROM AUTHOR]

Background: Lymphatic filariasis (LF) is a neglected tropical disease, and the Global Program to Eliminate LF delivers mass drug administration (MDA) to 500 million people every year. Adverse events (AEs) are common after LF treatment. Methodology/Principal findings: To better understand the pathogenesis of AEs, we studied LF-patients from a treatment trial. Plasma levels of many filarial antigens increased post-treatment in individuals with AEs, and this is consistent with parasite death. Circulating immune complexes were not elevated in these participants, and the classical complement cascade was not activated. Multiple cytokines increased after treatment in persons with AEs. A transcriptomic analysis was performed for nine individuals with moderate systemic AEs and nine matched controls. Differential gene expression analysis identified a significant transcriptional signature associated with post-treatment AEs; 744 genes were upregulated. The transcriptional signature was enriched for TLR and NF-κB signaling. Increased expression of seven out of the top eight genes upregulated in persons with AEs were validated by qRT-PCR, including TLR2. Conclusions/Significance: This is the first global study of changes in gene expression associated with AEs after treatment of lymphatic filariasis. Changes in cytokines were consistent with prior studies and with the RNAseq data. These results suggest that Wolbachia lipoprotein is involved in AE development, because it activates TLR2-TLR6 and downstream NF-κB. Additionally, LPS Binding Protein (LBP, which shuttles lipoproteins to TLR2) increased post-treatment in individuals with AEs. Improved understanding of the pathogenesis of AEs may lead to improved management, increased MDA compliance, and accelerated LF elimination. [ABSTRACT FROM AUTHOR]

Adipose tissue is highly involved in whole-body metabolism and is the main site for lipid synthesis, storage and mobilization in ruminants. Therefore, knowledge about adipose tissue responses to different diets is important, especially in growing heifers as the feeding regimes of replacement heifers affect their future success as dairy cows. However, at gene expression level such knowledge is limited. As part of a larger feed trial, adipose tissue biopsies from 24 Norwegian Red heifers were collected at 12 months of age (12MO) and at month seven of gestation (PREG) and analyzed by next-generation mRNA sequencing. Between these two sampling points, all heifers had gone through a successful conception and a feed change from four dietary treatments of high or low energy (HE/LE) and protein (HP/LP) content (treatments LPHE, HPHE, LPLE and HPLE) to a low-energy, low-protein pregnancy feed given to all animals. Gene expression differences between different feed treatments at 12MO are described in an earlier publication from our group. The main objectives of this study were to investigate the long-term effects of diets differing in protein and energy density level on gene expression in adipose tissue of growing replacement dairy heifers. To achieve this, we examined the post-treatment effects between the treatment groups at month seven of gestation; 6 months after the termination of experimental feeding, and the long-term gene expression changes occurring in the adipose tissue between 12MO and PREG. Post-treatment group comparisons showed evidence of long-term effects of dietary treatment on adipose gene expression. Differences between protein treatments were smaller than between energy treatments. Adipose gene expression changes from 12MO to PREG were much larger for the HE than the LE treatments and seemed to mostly be explained by the characteristics of the diet change. 97 genes displayed a unidirectional expression change for all groups from 12MO to PREG, and are considered to be treatment-independent, possibly caused by pregnancy or increased age. This study provides candidate genes and key regulators for further studies on pregnancy preservation (TGFB1, CFD) and metabolic regulation and efficiency (PI3K, RICTOR, MAP4K4,) in dairy cattle. [ABSTRACT FROM AUTHOR]

Understanding oxidative stress and HTRA1 locus in abnormal angiogenesis resulting in wet AMD pathology is an important step in developing a novel therapeutic approach. Using subretinal injection of oxLDL into C57BL/6 mice, we observed a lesion resembling the features of choroidal neovascularization (CNV), including macrophage infiltration, increased VEGF expression, and neovascularization. However, incubating ARPE-19 cells with oxLDL–a carrier of oxidized phospholipids–resulted in increased expression of inflammatory cytokines and chemoattractant proteins that recruited monocytes, but no substantial increase in expression of VEGF. Furthermore, incubation of ARPE-19 with oxLDL induced higher expression of HTRA1, which we showed to synergize with oxLDL in elevating the expression of inflammatory cytokines and chemoattractant factors. To investigate the role of macrophage infiltration on these expression changes, we treated cultured J774 macrophages with oxLDL and applied the conditioned medium onto ARPE-19 cells. This treatment was found to greatly enhance the expression of VEGF in ARPE-19, indicating the necessity of macrophage secretory products to induce increased expression of VEGF in retinal pigment epithelium. Gene expression analysis revealed that oxLDL induced the expression of Wnt3A in macrophages, a key activator of canonical Wnt signaling pathways. In addition, western blot analysis showed that the macrophage conditioned media further enhanced the reduction of phosphorylated β-catenin induced by oxLDL. Lastly, we investigated HTRA1 as a potential target for AMD therapeutics. We demonstrated the ability of anti-HTRA1 antibody in vitro to neutralize the protease activity of HTRA1 and reduce the inflammatory and angiogenic response to oxidative stress. Finally, we validated the neutralizing effect of anti-HTRA1 antibody in vivo by evaluating lesion size and protein expression in a laser-photocoagulation murine model of CNV. We found that the combination of oxLDL and HTRA1 enhanced CNV size, which was reversed by the addition of anti-HTRA1 antibody. This study not only provides preliminary evidence that HTRA1 may be a viable target for AMD therapeutics but also elucidates the biochemical mechanisms by which this therapeutic effect may be mediated. [ABSTRACT FROM AUTHOR]

Focal segmental glomerulosclerosis (FSGS) is frequently found in biopsies of patients with steroid resistant nephrotic syndrome (SRNS). The pathogenesis of SRNS/FSGS is often unknown and the disease will recur in up to 50% of patients post-transplant, indicating the presence of circulating podocyte-toxic factor(s). Several studies have reported clinical improvement after anti-TNFα therapy. However, prediction of the clinical outcome in SRNS/FSGS is difficult, and novel predictive biomarkers are needed. An image-based assay, which measures disassembly of focal adhesion complexes in cultured podocytes, was used to ascertain the presence of podocyte toxic activity in SRNS/FSGS sera. Expression of TNFα pathway genes was analysed in the Nephroseq FSGS cohort and in cultured podocytes treated with SRNS/FSGS sera. Podocyte toxic activity was detected in 48/96 SRNS/FSGS patients. It did not correlate with serum TNFα levels, age, sex, ethnicity or glomerular filtration rate. In ~25% of the toxic samples, the toxicity was strongly inhibited by blockade of TNFα signaling. Transcriptional profiling of human FSGS biopsies and podocytes treated with FSGS sera revealed significant increases in expression of TNFα pathway genes. We identified patients with serum podocyte toxic activity who may be at risk for FSGS recurrence, and those patients in whom serum podocyte toxicity may be reversed by TNFα blockade. Activation of TNFα pathway genes occurs in podocytes of FSGS patients suggesting a causative effect of this pathway in response to circulating factor(s). In vitro analyses of patient sera may stratify patients according to prognostic outcomes and potential responses to specific clinical interventions. [ABSTRACT FROM AUTHOR]

Spot 14 (S14) protein is primarily expressed in adipogenic tissues. Compared to wild type, S14 knockout mice had better resistance to diet-induced obesity and glucose tolerance. However, the association between serum S14 level and metabolic variables in humans has never been investigated. The objectives of this study were to evaluate the associations between serum S14 concentrations with components of metabolic syndrome (MetS). A total of 327 subjects were recruited in this cross-sectional study and categorized by presence of MetS. The mean serum levels of S14 were significantly lower in subjects with MetS than those without (87.1±26.3 μg/L vs. 107.3±40.2 μg/L, p<0.001). In addition, the subjects with central obesity, low high density lipoprotein-C (HDL-C) or hypertriglyceridemia also had significantly lower S14 levels in comparison to those without. Adjusted with age and sex, diagnosis of MetS (β = -0.227, p<0.001), central obesity (β = -0.176, p = 0.001), low HDL-C (β = -0.149, p = 0.005), and high triglyceride (TG) (β = -0.198, p<0.001) were negatively associated with log transformation of serum S14 levels (logS14). With 25% logS14 increased, the risk of MetS (OR 0.65, 95% CI, 0.51–0.82, p<0.001), central obesity (OR 0.72, 95% CI, 0.58–0.89, p = 0.002), low HDL-C (OR 0.76, 95% CI, 0.61–0.95, p = 0.015) or high TG (OR 0.65, 95% CI, 0.51–0.83, p = 0.001) was reduced with a dose response trend. Our analysis revealed that patients with MetS had lower serum S14 levels than those without. Negative associations existed between MetS, central obesity, high TG, low HDL-C and logS14. [ABSTRACT FROM AUTHOR]

This study investigated the effects of AdipoRon, which is an agonist for adiponectin receptor 1 (AdipoR1) and AdipoR2, on the protein content, myotube diameter, and number of nuclei per myotube of C2C12 cells and skeletal muscle mass in C57BL/6J mice. AdipoRon suppressed the protein content, myotube diameter, and number of nuclei per myotube of C2C12 cells of C2C12 myotubes in a dose-dependent manner. Adiponectin-associated decline of protein content, diameter, and number of nuclei per myotube in C2C12 myotubes was partially rescued by knockdown of AdipoR1 and/or AdipoR2. Phosphorylation level of AMPK showed a trend to be increased by AdipoRon. A significant increase in phosphorylation level of AMPK was observed at 20 μM AdipoRon. Knockdown of AdipoR1 and/or AdipoR2 rescued AdipoRon-associated decrease in protein content of C2C12 myotubes. AdipoRon-associated increase in phosphorylation level of AMPK in C2C12 myotubes was suppressed by knockdown of AdipoR1 and/or AdipoR2. Successive intravenous injections of AdipoRon into mice caused a decrease in the wet weight of plantaris muscle (PLA), but not in soleus muscle (SOL). Mean fiber cross-sectional area of PLA, but not of SOL, was significantly decreased by AdipoRon administration. On the one hand, the expression level of phosphorylated AMPK and ubiquitinated protein in SOL and PLA muscles was upregulated by AdipoRon administration. On the other hand, AdipoRon administration induced no changes in the expression level of puromycin-labeled proteins in both SOL and PLA muscles. Expression level of adiponectin in extensor digitorum longus (EDL) muscle was increased by aging, but not in SOL muscle. Aging had no effect on the expression level of AdipoR1 and AdipoR2 in both muscles. Phosphorylation level of AMPK in EDL was increased by aging, but not SOL muscle. Results from this study suggest that high level of circulating adiponectin may induce skeletal muscle atrophy, especially fast-type muscle. [ABSTRACT FROM AUTHOR]

Aims: Endothelial activation is involved in many chronic inflammatory diseases, such as atherosclerosis, and is often initiated by cytokines. Oncostatin M (OSM) is a relatively unknown cytokine that has been suggested to play a role in both endothelial activation and atherosclerosis. We comprehensively investigated the effect of OSM on endothelial cell activation from different vascular beds and in APOE*3Leiden.CETP mice. Methods and results: Human umbilical vein endothelial cells, human aortic endothelial cells and human microvascular endothelial cells cultured in the presence of OSM express elevated MCP-1, IL-6 and ICAM-1 mRNA levels. Human umbilical vein endothelial cells and human aortic endothelial cells additionally expressed increased VCAM-1 and E-selectin mRNA levels. Moreover, ICAM-1 membrane expression is increased as well as MCP-1, IL-6 and E-selectin protein release. A marked increase was observed in STAT1 and STAT3 phosphorylation indicating that the JAK/STAT pathway is involved in OSM signaling. OSM signals through the LIF receptor alfa (LIFR) and the OSM receptor (OSMR). siRNA knockdown of the LIFR and the OSMR revealed that simultaneous knockdown is necessary to significantly reduce MCP-1 and IL-6 secretion, VCAM-1 and E-selectin shedding and STAT1 and STAT3 phosphorylation after OSM stimulation. Moreover, OSM administration to APOE*3Leiden.CETP mice enhances plasma E-selectin levels and increases ICAM-1 expression and monocyte adhesion in the aortic root area. Furthermore, Il-6 mRNA expression was elevated in the aorta of OSM treated mice. Conclusion: OSM induces endothelial activation in vitro in endothelial cells from different vascular beds through activation of the JAK/STAT cascade and in vivo in APOE*3Leiden.CETP mice. Since endothelial activation is an initial step in atherosclerosis development, OSM may play a role in the initiation of atherosclerotic lesion formation. [ABSTRACT FROM AUTHOR]

We previously found higher level of endothelial cell (EC) activation in patients who suffered from in-stent restenosis after bare-metal stenting compared to subjects who underwent drug-eluting stenting (DES) showing no complications. Here we investigated the potential transcriptional and post-transcriptional regulatory mechanisms by which everolimus attenuated EC activation after DES. We studied the effect of everolimus on E-selectin (SELE) and VCAM1 mRNA levels when human coronary artery (HCAECs) and human umbilical vein ECs were challenged with recombinant TNF-α (100 ng/mL) for 1–24 hours in the presence or absence of everolimus using 0.5 μM concentration locally maintained by DES. EC activation was evaluated via the levels of IL-1β and IL-6 mRNAs with miR-155 expression by RT-qPCR as well as the nuclear translocation of nuclear factor kappa beta (NF-κB) detected by fluorescence microscopy. To investigate the transcriptional regulation of E-selectin and VCAM-1, TNF-α-induced enhancer RNA (eRNA) expression at p65-bound enhancers in the neighboring genomic regions of SELE and VCAM1 genes, including SELE_-11Kb and VCAM1_-10Kb, were measured in HCAECs. Mature and precursor levels of E-selectin and VCAM-1 repressor miR-181b were quantified to analyze the post-transcriptional regulation of these genes in HCAECs. Circulating miR-181b was analyzed in plasma samples of stented subjects by stem-loop RT-qPCR. TNF-α highly elevated E-selectin and VCAM-1 expression at transcriptional level in ECs. Levels of mature, pre- and pri-miR-181b were repressed in ECs by TNF-α, while everolimus acted as a negative regulator of EC activation via inhibited translocation of NF-κB p65 subunit into cell nuclei, lowered eRNA expression at SELE and VCAM1 genes-associated enhancers and modulated expression of their post-transcriptional repressor miR-181b. Significant negative correlation was observed between plasma miR-181b and soluble E-selectin and VCAM-1 in patients. In conclusion, everolimus attenuates EC activation via reduced NF-κB p65 translocation causing decreased E-selectin and VCAM-1 expression at transcriptional and post-transcriptional level after DES. [ABSTRACT FROM AUTHOR]

Gene expression in eukaryotes is controlled by DNA sequences at promoter and enhancer regions, whose accessibility for binding by regulatory proteins dictates their specific patterns of activity. Here, we identify the protein Zbtb7a as a factor required for inducible changes in accessibility driven by transcription factors (TFs). We show that Zbtb7a binds to a significant fraction of genomic promoters and enhancers, encompassing many target genes of nuclear factor kappa B (NFκB) p65 and a variety of other TFs. While Zbtb7a binding is not alone sufficient to directly activate promoters, it is required to enable TF-dependent control of accessibility and normal gene expression. Using p65 as a model TF, we show that Zbtb7a associates with promoters independently of client TF binding. Moreover, the presence of prebound Zbtb7a can specify promoters that are amenable to TF-induced changes in accessibility. Therefore, Zbtb7a represents a widely used promoter factor that transduces signals from other TFs to enable control of accessibility and regulation of gene expression. [ABSTRACT FROM AUTHOR]

Autism spectrum disorder (ASD) is induced by complex hereditary and environmental factors. However, the mechanisms of ASD development are poorly understood. The purpose of this study was to identify standard indicators of this condition by comparing clinical, pathophysiological, and neurobehavioral features in an autism-like animal model. A total of 22 male Sprague-Dawley rats were randomly divided into control and 500 mg/kg propionic acid (PPA)-treated groups. Rats were subjected to behavioral tests, gene expression analyses, and histological analyses to detect pathophysiological and neurobehavioral alterations. Exploratory activity and non-aggressive behavior were significantly reduced in PPA-treated rats, whereas enhanced aggressive behavior during adjacent interactions was observed on day 14 after PPA administration. To evaluate gene expression after PPA administration, we analyzed hippocampal tissue using reverse transcription PCR. Glial fibrillary acidic protein was augmented in the PPA-treated group on day 14 after appearance of ASD-like behaviors by PPA administration, whereas octamer-binding transcription factor 4 expression was significantly decreased in the PPA-treated group. Histological evaluation revealed significantly reduced diameter and layer thickness of granule cells in PPA-treated rats compared with control rats. We conclude that PPA administration induced abnormal neural cell organization, which may have led to autism-like neurobehaviors, including increased aggressive behavior, reduced exploratory activity, and isolative and passive behaviors. [ABSTRACT FROM AUTHOR]

Adult T-cell leukaemia/lymphoma (ATL) arises from chronic non-malignant human T lymphotropic virus type-1 (HTLV-1) infection which is characterized by high plasma pro-inflammatory cytokines whereas ATL is characterized by high plasma anti-inflammatory (IL-10) concentrations. The poor prognosis of ATL is partly ascribed to disease-associated immune suppression. ATL cells have a CD4+CCR4+CD26-CD7- immunophenotype but infected cells with this immunophenotype (‘ATL-like’ cells) are also present in non-malignant HTLV-1 infection. We hypothesized that ‘ATL-like’ and ATL cells have distinct cytokine producing capacity and a switch in the cytokines produced occurs during leukemogenesis. Seventeen asymptomatic carriers (ACs), 28 patients with HTLV-1-associated myelopathy (HAM) and 28 with ATL were studied. Plasma IL-10 concentration and the absolute frequency of IL-10-producing CD4+ T cells were significantly higher in patients with ATL compared to AC. IL-10-producing ATL cells were significantly more frequent than ‘ATL-like’ cells. The cytokine-producing cells were only a small fraction of ATL cells. Clonality analysis revealed that even in patients with ATL the ATL cells were composed not only of a single dominant clone (putative ATL cells) but also tens of non-dominant infected clones (‘ATL-like’ cells). The frequency of cytokine-producing cells showed a strong inverse correlation with the relative abundance of the largest clone in ATL cells suggesting that the putative ATL cells were cytokine non-producing and that the ‘ATL-like’ cells were the primary cytokine producers. These findings were confirmed by RNAseq with cytokine mRNA expression in ATL cells in patients with ATL (confirmed to be composed of both putative ATL and ‘ATL-like’ cells by TCR analysis) significantly lower compared to ‘ATL-like’ cells in patients with non-malignant HTLV-1 infection (confirmed to be composed of hundreds of non-dominant clones by TCR analysis). A significant inverse correlation between the relative abundance of the largest clone and cytokine mRNA expression was also confirmed. Finally, ‘ATL-like’ cells produced less pro- and more anti-inflammatory cytokines than non ‘ATL-like’ CD4+ cells (which are predominantly HTLV uninfected). In summary, HTLV-1 infection of CD4+ T cells is associated with a change in cytokine producing capacity and dominant malignant clonal growth is associated with loss of cytokine producing capacity. Non-dominant clones with ‘ATL-like’ cells contribute to plasma cytokine profile in patients with non-malignant HTLV-1 infection and are also present in patient with ATL. [ABSTRACT FROM AUTHOR]

Background: Obesity is a well-known risk factor of breast cancer in post-menopausal women that also correlates with a diminished therapeutic response. The influence of adipocytes and their secretome, i.e. adipokines, on the efficacy of hormone therapy has yet to be elucidated. Methods: We investigated, ex vivo, whether mature adipocytes, differentiated from adipose stem cells of normal-weight (MA20) or obese (MA30) women, and their secretions, were able to counteract the effects of tamoxifen (Tx) which is known to decrease neoplastic cell proliferation. Results: In a tridimensional model and in a model of co-culture, the anti-proliferative effect of Tx on MCF-7 cancer cells was counteracted by MA30. These two models highlighted two different specific gene expression profiles for genes encoding cytokines or involved in angiogenesis based on the adipocyte microenvironment and the treatment. Thus it notably showed altered expression of genes such as TNFα that correlated with IL-6. In addition, leptin, IL-6 and TNFα, at concentrations reflecting plasma concentrations in obese patients, decreased the anti-proliferative efficacy of 4-hydroxytamoxifen (a major active metabolite of Tx). Conclusions: These findings bring insights on adipocytes and mammary cancer cell interactions in Tx therapy, particularly in overweight/obese people. Indeed, patient’ adipokine status would give valuable information for developing individual strategies and avoid resistance to treatment. [ABSTRACT FROM AUTHOR]

Cytokine signaling is responsible for coordinating conserved epithelial regeneration and immune responses in the digestive tract. In the Drosophila midgut, Upd3 is a major cytokine, which is induced in enterocytes (EC) and enteroblasts (EB) upon oral infection, and initiates intestinal stem cell (ISC) dependent tissue repair. To date, the genetic network directing upd3 transcription remains largely uncharacterized. Here, we have identified the key infection-responsive enhancers of the upd3 gene and show that distinct enhancers respond to various stresses. Furthermore, through functional genetic screening, bioinformatic analyses and yeast one-hybrid screening, we determined that the transcription factors Scalloped (Sd), Mothers against dpp (Mad), and D-Fos are principal regulators of upd3 expression. Our study demonstrates that upd3 transcription in the gut is regulated by the activation of multiple pathways, including the Hippo, TGF-β/Dpp, and Src, as well as p38-dependent MAPK pathways. Thus, these essential pathways, which are known to control ISC proliferation cell-autonomously, are also activated in ECs to promote tissue turnover the regulation of upd3 transcription. [ABSTRACT FROM AUTHOR]

Objective: Genetic P300/CBP-associated factor (PCAF) variation affects restenosis-risk in patients. PCAF has lysine acetyltransferase activity and promotes nuclear factor kappa-beta (NFκB)-mediated inflammation, which drives post-interventional intimal hyperplasia development. We studied the contributing role of PCAF in post-interventional intimal hyperplasia. Methods and results: PCAF contribution to inflammation and intimal hyperplasia was assessed in leukocytes, macrophages and vascular smooth muscle cells (vSMCs) in vitro and in a mouse model for intimal hyperplasia, in which a cuff is placed around the femoral artery. PCAF deficiency downregulate CCL2, IL-6 and TNF-alpha expression, as demonstrated on cultured vSMCs, leukocytes and macrophages. PCAF KO mice showed a 71.8% reduction of vSMC-rich intimal hyperplasia, a 73.4% reduction of intima/media ratio and a 63.7% reduction of luminal stenosis after femoral artery cuff placement compared to wild type (WT) mice. The association of PCAF and vascular inflammation was further investigated using the potent natural PCAF inhibitor garcinol. Garcinol treatment reduced CCL2 and TNF-alpha expression, as demonstrated on cultured vSMCs and leukocytes. To assess the effect of garcinol treatment on vascular inflammation we used hypercholesterolemic ApoE*3-Leiden mice. After cuff placement, garcinol treatment resulted in reduced arterial leukocyte and macrophage adherence and infiltration after three days compared to untreated animals. Conclusions: These results identify a vital role for the lysine acetyltransferase PCAF in the regulation of local inflammation after arterial injury and likely the subsequent vSMC proliferation, responsible for intimal hyperplasia. [ABSTRACT FROM AUTHOR]

Angiopoietin-like protein 2 (ANGPTL2) maintains tissue homeostasis by inducing inflammation and angiogenesis. It is produced in infiltrating immune cells or resident cells, such as adipocytes, vascular endothelial cells, and tumor cells. We hypothesized that ANGPTL2 might play an important role as a unique mediator in both systemic and periodontal disease. We demonstrated an increased ANGPTL2 concentration in gingival crevicular fluid from chronic periodontitis patients. Porphyromonas gingivalis lipopolysaccharide (LPS) treatment strongly induced ANGPTL2 mRNA and protein levels in Ca9-22 human gingival epithelial cells. Recombinant human ANGPTL2 increased interleukin 1β (IL-1β), IL-8, and tumor necrosis factor-α (TNF-α) mRNA and protein levels in Ca9-22 cells. Small-interfering (si)RNA-mediated ANGPTL2 knockdown in Ca9-22 cells reduced IL-1β, IL-8 and TNF-α mRNA and protein levels compared with control siRNA (p<0.01) in P. gingivalis LPS-stimulated Ca9-22 cells. Antibodies against integrin α5β1, an ANGPTL receptor, blocked induction of these inflammatory cytokines in P. gingivalis LPS-treated Ca9-22 cells, suggesting that secreted ANGPTL induces inflammatory cytokines in gingival epithelial cells via an autocrine loop. The classic sequential cascade of P. gingivalis LPS → inflammatory cytokine induction is well established. However, in the current study, we reveal a novel cascade comprising sequential P. gingivalis LPS → ANGPTL2 → integrin α5β1 → inflammatory cytokine induction, which might be responsible for inducing potent periodontal disorganization activity in gingival epithelial cells. Via this pathway, ANGPTL2 functions in the pathogenesis of periodontitis and contributes to prolonging chronic inflammation in patients with systemic disease. [ABSTRACT FROM AUTHOR]

Moringa (Moringa oleifera Lam.) seed extract (MSE) has anti-inflammatory and antioxidant activities. We investigated the effects of MSE enriched in moringa isothiocyanate-1 (MIC-1), its putative bioactive, on ulcerative colitis (UC) and its anti-inflammatory/antioxidant mechanism likely mediated through Nrf2-signaling pathway. Dextran sulfate sodium (DSS)-induced acute (n = 8/group; 3% DSS for 5 d) and chronic (n = 6/group; cyclic rotations of 2.5% DSS/water for 30 d) UC was induced in mice that were assigned to 4 experimental groups: healthy control (water/vehicle), disease control (DSS/vehicle), MSE treatment (DSS/MSE), or 5-aminosalicyic acid (5-ASA) treatment (positive control; DSS/5-ASA). Following UC induction, water (vehicle), 150 mg/kg MSE, or 50 mg/kg 5-ASA were orally administered for 1 or 2 wks. Disease activity index (DAI), spleen/colon sizes, and colonic histopathology were measured. From colon and/or fecal samples, pro-inflammatory biomarkers, tight-junction proteins, and Nrf2-mediated enzymes were analyzed at protein and/or gene expression levels. Compared to disease control, MSE decreased DAI scores, and showed an increase in colon lengths and decrease in colon weight/length ratios in both UC models. MSE also reduced colonic inflammation/damage and histopathological scores (modestly) in acute UC. MSE decreased colonic secretions of pro-inflammatory keratinocyte-derived cytokine (KC), tumor necrosis factor (TNF)-α, nitric oxide (NO), and myeloperoxidase (MPO) in acute and chronic UC; reduced fecal lipocalin-2 in acute UC; downregulated gene expression of pro-inflammatory interleukin (IL)-1, IL-6, TNF-α, and inducible nitric oxide synthase (iNOS) in acute UC; upregulated expression of claudin-1 and ZO-1 in acute and chronic UC; and upregulated GSTP1, an Nrf2-mediated phase II detoxifying enzyme, in chronic UC. MSE was effective in mitigating UC symptoms and reducing UC-induced colonic pathologies, likely by suppressing pro-inflammatory biomarkers and increasing tight-junction proteins. This effect is consistent with Nrf2-mediated anti-inflammatory/antioxidant signaling pathway documented for other isothiocyanates similar to MIC-1. Therefore, MSE, enriched with MIC-1, may be useful in prevention and treatment of UC. [ABSTRACT FROM AUTHOR]

Sex differences in the development of the normal heart and the prevalence of cardiomyopathies have been reported. The molecular basis of these differences remains unclear. Sex differences in the human heart might be related to patterns of gene expression. Recent studies have shown that sex specific differences in gene expression in tissues including the brain, kidney, skeletal muscle, and liver. Similar data is limited for the heart. Herein we address this issue by analyzing donor and post-mortem adult human heart samples originating from 46 control individuals to study whole-genome gene expression in the human left ventricle. Using data from the genotype tissue expression (GTEx) project, we compared the transcriptome expression profiles of male and female hearts. We found that genes located on sex chromosomes were the most abundant ones among the sexually dimorphic genes. The majority of differentially expressed autosomal genes were those involved in the regulation of inflammation, which has been found to be an important contributor to left ventricular remodeling. Specifically, genes on autosomal chromosomes encoding chemokines with inflammatory functions (e.g. CCL4, CX3CL1, TNFAIP3) and a gene that regulates adhesion of immune cells to the endothelium (e.g., VCAM1) were identified with sex-specific expression levels. This study underlines the relevance of sex as an important modifier of cardiac gene expression. These results have important implications in the understanding of the differences in the physiology of the male and female heart transcriptome and how they may lead to different sex specific difference in human cardiac health and its control. [ABSTRACT FROM AUTHOR]

Intracellular DNA- and RNA-sensing receptors, such as the IFN-inducible protein Absent in Melanoma 2 (AIM2), serve as host sensors against a wide range of infections. Immune sensing and inflammasome activation by AIM2 has been implicated in innate antiviral recognition in many experimental systems using cell-lines and animal models. However, little is known about the expression and function of AIM2 in freshly isolated human cells. In this study we investigated the expression of AIM2 in different cell types derived from human cord and adult peripheral blood, in steady state and following in vitro-activation. Adult but not cord blood B-cells expressed high levels of AIM2 mRNA at steady state. In adults, AIM2 was primarily expressed in mature memory CD27+ B-cells. Both adult and cord blood derived B-cells could induce their transcription of AIM2 mRNA in response to type II IFN but not type I IFN or the AIM2 ligand poly dA:dT. Upon B-cell receptor stimulation, B-cells from adult blood expressed reduced levels of AIM2 mRNA. In addition, we show that adult B-cells were able to release IL-1β upon stimulation with synthetic DNA. We conclude that functional AIM2 is preferentially expressed in adult human CD27+ B-cells, but is absent in cord blood mononuclear cells. [ABSTRACT FROM AUTHOR]

Adiponectin plays important roles in metabolic function, inflammation and multiple biological activities in various tissues. However, evidence for adiponectin signaling in sebaceous glands is lacking, and its role remains to be clarified. This study investigated the role of adiponectin in lipid production in sebaceous glands in an experimental study of human sebocytes. We demonstrated that human sebaceous glands in vivo and sebocytes in vitro express adiponectin receptor and that adiponectin increased cell proliferation. Moreover, based on a lipogenesis study using Oil Red O, Nile red staining and thin layer chromatography, adiponectin strongly upregulated lipid production in sebocytes. In three-dimensional culture of sebocytes, lipid synthesis was markedly enhanced in sebocytes treated with adiponectin. This study suggested that adiponectin plays a significant role in human sebaceous gland biology. Adiponectin signaling is a promising target in the clinical management of barrier disorders in which sebum production is decreased, such as in atopic dermatitis and aged skin. [ABSTRACT FROM AUTHOR]

This study was carried out to investigate protective effect of chlorogenic acid against lipopolysaccharide-induced inflammation and oxidative stress in intestinal epithelial cells. As a marker of inflammatory response, IL-6, IL-8, TNF-α mRNA and protein levels, furthermore, COX-2 mRNA level were followed up. Intracellular redox status and extracellular H2O2 level were also monitored by two fluorescent assays (DCFH-DA, Amplex Red). Moreover, the effect of gut microbiota metabolites in the above mentioned processes was taken into account in our model using Lactobacillus plantarum 2142 bacterial strain. Our data revealed that chlorogenic acid had significant lowering effect on the inflammatory response. Treatment with chlorogenic acid (25–50 μM) significantly decreased gene expression and concentration of proinflammatory cytokines IL-6 and IL-8 compared to LPS-treated cells. COX-2 and TNF-α mRNA levels were also reduced. Furthermore, chlorogenic acid reduced the level of reactive oxygen species in IPEC-J2 cells. Simultaneous application of chlorogenic acid and Lactobacillus plantarum 2142 supernatant resulted protective effect against LPS-induced inflammation and oxidative stress as well. [ABSTRACT FROM AUTHOR]

Fusobacterium nucleatum (F. nucleatum) plays a critical role in gastrointestinal inflammation. However, the exact mechanism by which F. nucleatum contributes to inflammation is unclear. In the present study, it was revealed that F. nucleatum could induce the production of proinflammatory cytokines (IL-8, IL-1β and TNF-α) and reactive oxygen species (ROS) in Caco-2 colorectal) adenocarcinoma cells. Furthermore, ROS scavengers (NAC or Tiron) could decrease the production of proinflammatory cytokines during F. nucleatum infection. In addition, we observed that autophagy is impaired in Caco-2 cells after F. nucleatum infection. The production of proinflammatory cytokines and ROS induced by F. nucleatum was enhanced with either autophagy pharmacologic inhibitors (3-methyladenine, bafilomycin A1) or RNA interference in essential autophagy genes (ATG5 or ATG12) in Caco-2 cells. Taken together, these results indicate that F. nucleatum-induced impairment of autophagic flux enhances the expression of proinflammatory cytokines via ROS in Caco-2 Cells. [ABSTRACT FROM AUTHOR]

Sperm are allogenic to the female genital tract; however, oviducts provide optimal conditions for survival and capacitation of these non-self cells until fertilization. Recently, we showed that oviduct-conditioned media and prostaglandin E2 (PGE2) suppress sperm phagocytosis by polymorphonuclear neutrophils (PMNs) under physiological conditions. We hypothesized that sperm binding to bovine oviduct epithelial cells (BOECs) could change the local innate immunity via PGE2. As the first step to obtain basic information, sub-confluent BOEC monolayers were co-cultured with swim-up sperm for 2 h. BOECs with viable bound sperm were cultured for an additional 3, 6, 12, or 24 h. Then, we confirmed the impact of the sperm-BOEC binding on both BOECs and PMN gene expression. Immunohistochemistry revealed that BOECs strongly express TGFB1 and IL10 in the oviduct. Sperm binding to BOECs in culture induced the anti-inflammatory cytokines (TGFB1 and IL10) and PGE2 production by BOECs. Exogenous PGE2 in vitro suppressed pro-inflammatory cytokine expression (TNF and IL1B) in BOECs. Moreover, pre-exposure of PMNs to BOEC-conditioned media suppressed the TNF expression, but the BOEC media co-cultured with sperm stimulated PMNs to express TGFB1 and IL10, with increasing PGE2 secretion. Of note, exogenous PGE2 led PMNs in vitro to decrease their TNF expression and increase anti-inflammatory cytokines expression. Our findings strongly suggest that BOECs provide an anti-inflammatory environment under physiological conditions and the sperm-BOEC binding further strengthens this milieu thus suppresses PMNs in the bovine oviduct. PGE2 is likely to drive this stable anti-inflammatory environment in the oviduct. [ABSTRACT FROM AUTHOR]

Aminopeptidase N/CD13 is highly expressed by fibroblast like synoviocytes (FLS) and may play a role in rheumatoid arthritis (RA). CD13 was previously detected in human synovial fluid where it was significantly increased in RA compared to osteoarthritis. In this study we found that CD13 in biological fluids (plasma, synovial fluid, FLS culture supernatant) is present as both a soluble molecule and on extracellular vesicles, including exosomes, as assessed by differential ultracentrifugation and density gradient separation. Having determined CD13 could be released as a soluble molecule from FLS, we examined potential mechanisms by which CD13 might be shed from the FLS membrane. The use of protease inhibitors revealed that CD13 is cleaved from the FLS surface by metalloproteinases. siRNA treatment of FLS revealed one of those proteases to be MMP14. We determined that pro-inflammatory cytokines (TNFα, IFNγ, IL-17) upregulated CD13 mRNA in FLS, which may contribute to the increased CD13 in RA synovium and synovial fluid. Inhibition of CD13 function by either inhibitors of enzymatic activity or anti-CD13 antibodies resulted in decreased growth and diminished migration of FLS. This suggests that CD13 may be involved in the pathogenic hyperplasia of RA FLS. This data expands potential roles for CD13 in the pathogenesis of RA. [ABSTRACT FROM AUTHOR]

Butyrate in the gut of animals has potential properties including regulating the innate immune, modulating the lipid metabolism, and protecting gut healthy. So far, only limited information on the impact of butyrate on the neonatal is available. This study aimed to investigate effects of oral administration of sodium butyrate (SB) on gut microbiota and the expression of inflammatory cytokine in neonatal piglets. Ten litters of crossbred newborn piglets were randomly allocated to the SB and control (CO) groups, each group consisted of five litters (replicates). Piglets in the SB group were orally administrated with 7 to 13 ml sodium butyrate solution (150 mmol/l) per day from the age of 1 to 7 days, respectively; piglets in the CO group were treated with the same dose of physiological saline. On days 8 and 21 (of age), gut digesta and tissues were collected for the analysis of microbiota, butyrate concentration and gene expression of inflammatory cytokine. Results showed that there was no difference in the butyrate concentration in the gut of piglets on days 8 and 21 between two groups. Real-time PCR assay showed that SB had no effect on the numbers of total bacteria in the stomach, ileum, and colon. MiSeq sequencing of the V3-V4 region of the 16S rRNA gene revealed that SB increased the richness in the stomach and colon, and the diversity of colonic microbiota on day 8 (P < 0.05). Genera Acinetobacter, Actinobacillus, Facklamia, Globicatella, Kocuria, Rothia, unclassified Leptotrichiaceae, unclassified Neisseriaceae, and unclassified Prevotellaceae in the stomach were increased in relative abundance by SB treatment, whereas the abundances of Lactobacillus decreased on day 8 (P < 0.05). At the genus and operational taxonomic unit (OTU) levels, SB had low impact on bacterial community in the ileum and colon on days 8 and 21. SB treatment decreased the expression of IL-6, IL-8, IFN-γ, IL-10, TGF-β, and histone deacetylase 1 (HDAC1) in the ileum of piglets on day 8 (P < 0.05). SB treatment down-regulated the expression of IL-8, IFN-γ, and IL-1β on day 21 (P < 0.05). Correlation analysis on the combined datasets revealed some potential relationships between gut microbiota and the expression of inflammatory cytokines. The results show that early intervention with sodium butyrate can modulate the ileum inflammatory cytokine in neonatal piglets with low impact on intestinal microbial structure, which suggests oral administration of SB may have a benefit role in the health of neonatal piglets. [ABSTRACT FROM AUTHOR]

We previously reported that mouse bone marrow-derived macrophages (BMDMs) that had been co-cultured with platelets exhibited lower susceptibility to bacterial lipopolysaccharide (LPS) and produced lower levels of nitric oxide (NO) and inflammatory cytokines including TNF-α and IL-6. The suppression of macrophage responses was mediated, at least in part, by platelet supernatant. In the present study, we assessed phenotypic changes of BMDMs induced by incubation with the supernatant from thrombin-activated platelets (PLT-sup) and found that BMDMs cultured with PLT-sup (PLT-BMDMs) expressed a lower level of inducible NO synthase (iNOS) and a higher level of arginase-1, both of which are involved in the L-arginine metabolism, upon stimulation with LPS or zymosan. We also examined possible modulation of the NF-κB signaling pathway and observed suppression of IκBα phosphorylation and a decrease of NF-κB p65 expression in LPS-stimulated PLT-BMDMs. These results suggest that PLT-sup suppresses inflammatory responses of BMDMs via negative regulation of NF-κB signaling leading to lowered expression of iNOS and enhanced L-arginine catabolism by arginase-1. [ABSTRACT FROM AUTHOR]

Inflammation in the mammary gland (mastitis) is the most common disease in dairy herds worldwide, often caused by the pathogens Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). Little is known about the effects of mastitis on drug transporters and the impact on transporter-mediated excretion of drugs into milk. We used murine mammary epithelial HC11 cells, after lactogenic differentiation into a secreting phenotype, and studied gene expressions of ABC- and SLC- transporters after treatment of cells with S. aureus and lipopolysaccharide, an endotoxin secreted by E. coli. The studied transporters were Bcrp, Mdr1, Mrp1, Oatp1a5, Octn1 and Oct1. In addition, Csn2, the gene encoding β-casein, was analyzed. As biomarkers of the inflammatory response, gene expressions of the cytokines Il6 and Tnfα and the chemokine Cxcl2 were determined. Our results show that S. aureus and LPS treatment of cells, at non-cytotoxic concentrations, induced an up-regulation of Mdr1 and of the inflammatory biomarkers, except that Tnfα was not affected by lipopolysaccharide. By simple regression analysis we could demonstrate statistically significant positive correlations between each of the transporters with each of the inflammatory biomarkers in cells treated with S. aureus. The coefficients of determination (R2) were 0.7–0.9 for all but one correlation. After treatment of cells with lipopolysaccharide, statistically significant correlations were only found between Mdr1 and the two parameters Cxcl2 and Il6. The expression of Csn2 was up-regulated in cells treated with S. aureus, indicating that the secretory function of the cells was not impaired. The strong correlation in gene expressions between transporters and inflammatory biomarkers may suggest a co-regulation and that the transporters have a role in the transport of cytokines and chemokines. Our results demonstrate that transporters in mammary cells can be affected by infection, which may have an impact on transport of essential compounds and contaminants into milk. [ABSTRACT FROM AUTHOR]

Cancer-associated fibroblasts (CAFs), the most common constituent of the tumor stoma, are known to promote tumor initiation, progression and metastasis. However, the mechanism of how cancer cells transform normal fibroblasts (NFs) into CAFs is largely unknown. In this study, we determined the contribution of miRNAs in the transformation of NFs into CAFs. We found that miR-1 and miR-206 were down-regulated, whereas miR-31 was up-regulated in lung CAFs when compared with matched NFs. Importantly, modifying the expression of these three deregulated miRNAs induced a functional conversion of NFs into CAFs and vice versa. When the miRNA-reprogrammed NFs and CAFs were co-cultured with lung cancer cells (LCCs), a similar pattern of cytokine expression profiling were observed between two groups. Using a combination of cytokine expression profiling and miRNAs algorithms, we identified VEGFA/CCL2 and FOXO3a as direct targets of miR-1, miR-206 and miR-31, respectively. Importantly, systemic delivery of anti-VEGFA/CCL2 or pre-miR-1, pre-miR-206 and anti-miR-31 significantly inhibited tumor angiogenesis, TAMs accumulation, tumor growth and lung metastasis. Our results show that miRNAs-mediated FOXO3a/VEGF/CCL2 signaling plays a prominent role in LCCs-mediated NFs into CAFs, which may have clinical implications for providing novel biomarker(s) and potential therapeutic target(s) of lung cancer in the future. [ABSTRACT FROM AUTHOR]

Asthma is a heterogeneous disease characterized by chronic airway inflammation. CD4(+) T-helper 9 (Th9) cells are closely linked to asthma, helping to regulate inflammation and immunity. Epidemiological studies showed that mycobacteria infections are negatively associated with asthma. Our previous research showed that inactivated Mycobacterium phlei nebulization alleviated the airway hyperresponsiveness and inflammation of asthma. However, the relationship between Th9 cells and mycobacteria remains unknown. Here, we evaluated the relationship between Mycobacterium vaccae nebulization and Th9 cells in asthmatic mice. Eighteen Balb/c mice were randomized into 3 groups of 6 mice each (normal control group, asthma control group, and nebulization asthma group [Neb. group]). The Neb. group was nebulized with M. vaccae one month before establishment of the asthmatic model with ovalbumin (OVA) sensitization, and the normal and asthma control groups were nebulized with phosphate-buffered saline. The hyperresponsiveness of the mouse airways was assessed using a non-invasive lung function machine. Lung airway inflammation was evaluated by hematoxylin and eosin and periodic acid-Schiff staining. Cytokine interlukin-9 (IL-9) concentration and OVA-specific IgE in the bronchoalveolar lavage fluid were measured by enzyme-linked immunosorbent assays. The percentages of γδTCR+ CD3+, IL-9+CD3+, IL-10+CD3+ lymphocytes, and IL9+γδT and IL-10+γδT cells were detected by flow cytometry. The airway inflammation and concentration of IL-9 and OVA-specific IgE were significantly reduced in the Neb. group compared to the asthma control group. The Neb. group had lower airway hyperresponsiveness, percentages of γδTCR+CD3+ and IL-9+CD3+ lymphocytes, and IL9+γδT cells, and higher percentages of IL-10+CD3+ lymphocytes and IL-10+γδT cells compared to the asthma control group. Thus, mouse bronchial asthma could be prevented by M. vaccae nebulization. The mechanism could involve M. vaccae-mediated effects on induction of IL-9 secretion and suppression of IL-10 secretion from γδT cells. γδT cells showed prominent IL-10 expression, indicating that they possibly belong to the Th9 family. [ABSTRACT FROM AUTHOR]

Infections with mycobacteria, including Mycobacterium tuberculosis (Mtb) and Mycobacterium bovis (M. bovis) BCG, are a leading cause of morbidity and mortality for HIV-infected persons. In contrast to HIV, nonpathogenic SIV infections of sooty mangabeys are characterized by a lack of clinical disease including an absence of opportunistic infections. The goal of this study was to identify innate immune responses to M. bovis BCG maintained during nonpathogenic lentiviral infections through a comparison of functional responses during pathogenic HIV or nonpathogenic SIV infections. Monocytes were evaluated for their ability to express key anti-mycobacterial cytokines TNF-α and IL-12 following a six-hour ex vivo BCG exposure. While HIV-infection was associated with a decreased percentage of IL-12-producing monocytes, nonpathogenic SIV-infection was associated with an increased percentage of monocytes producing both cytokines. Gene expression analysis of PBMC following ex vivo BCG exposure identified differential expression of NK cell-related genes and several cytokines, including IFN-γ and IL-23, between HIV-infected and control subjects. In contrast, SIV-infected and uninfected-control mangabeys exhibited no significant differences in gene expression after BCG exposure. Finally, differential gene expression patterns were identified between species, with mangabeys exhibiting lower IL-6 and higher IL-17 in response to BCG when compared to humans. Overall, this comparison of immune responses to M. bovis BCG identified unique immune signatures (involving cytokines IL-12, TNF-α, IL-23, IL-17, and IL-6) that are altered during HIV, but maintained or increased during nonpathogenic SIV infections. These unique cytokine and transcriptome signatures provide insight into the differential immune responses to Mycobacteria during pathogenic HIV-infection that may be associated with an increased incidence of mycobacterial co-infections. [ABSTRACT FROM AUTHOR]

Weaning of dairy calves is an early life husbandry management practice which involves the changeover from a liquid to a solid feed based diet. The objectives of the study were to use RNA-seq technology to examine the effect of (i) breed and (ii) gradual weaning, on the whole blood mRNA transcriptome of artificially reared Holstein-Friesian and Jersey calves. The calves were gradually weaned over 14 days (day (d) -13 to d 0) and mRNA transcription was examined one day before gradual weaning was initiated (d -14), one day after weaning (d 1), and 8 days after weaning (d 8). On d -14, 550 genes were differentially expressed between Holstein-Friesian and Jersey calves, while there were 490 differentially expressed genes (DEG) identified on d 1, and 411 DEG detected eight days after weaning (P < 0.05; FDR < 0.1). No genes were differentially expressed within breed, in response to gradual weaning (P > 0.05). The pathways, gene ontology terms, and biological functions consistently over-represented among the DEG between Holstein-Friesian and Jersey were associated with the immune response and immune cell signalling, specifically chemotaxis. Decreased transcription of several cytokines, chemokines, immunoglobulin-like genes, phagocytosis-promoting receptors and g-protein coupled receptors suggests decreased monocyte, natural killer cell, and T lymphocyte, chemotaxis and activation in Jersey compared to Holstein-Friesian calves. Knowledge of breed-specific immune responses could facilitate health management practices better tailored towards specific disease sensitivities of Holstein-Friesian and Jersey calves. Gradual weaning did not compromise the welfare of artificially-reared dairy calves, evidenced by the lack of alterations in the expression of any genes in response to gradual weaning. [ABSTRACT FROM AUTHOR]

Chitin is produced in large amounts by fungi, insects, and other organisms and has been implicated in the pathogenesis of asthma. Airway epithelial cells are in direct contact with environmental particles and serve as the first line of defense against inhaled allergens and pathogens. The potential contributions of airway epithelial cells to chitin-induced asthma remain poorly understood. We hypothesized that chitin directly stimulates airway epithelial cells to release cytokines that promote type 2 immune responses and to induce expression of molecules which are important in innate immune responses. We found that chitin exposure rapidly induced the expression of three key type 2-promoting cytokines, IL-25, IL-33 and TSLP, in BEAS-2B transformed human bronchial epithelial cells and in A549 and H292 lung carcinoma cells. Chitin also induced the expression of the key pattern recognition receptors TLR2 and TLR4. Chitin induced the expression of miR-155, miR-146a and miR-21, each of which is known to up-regulate the expression of pro-inflammatory cytokines. Also the expression of SOCS1 and SHIP1 which are known targets of miR-155 was repressed by chitin treatment. The monoterpene phenol carvacrol (Car) and its isomer thymol (Thy) are found in herbal essential oils and have been shown to inhibit allergic inflammation in asthma models. We found that Car/Thy inhibited the effects of chitin on type 2-promoting cytokine release and on the expression of TLRs, SOCS1, SHIP1, and miRNAs. Car/Thy could also efficiently reduce the protein levels of TLR4, inhibit the increase in TLR2 protein levels in chitin plus Car/Thy-treated cells and increase the protein levels of SHIP1 and SOCS1, which are negative regulators of TLR-mediated inflammatory responses. We conclude that direct effects of chitin on airway epithelial cells are likely to contribute to allergic airway diseases like asthma, and that Car/Thy directly inhibits epithelial cell pro-inflammatory responses to chitin. [ABSTRACT FROM AUTHOR]

Mutations of the transglutaminase 1 gene (TGM1) are a major cause of autosomal recessive congenital ichthyoses (ARCIs) that are associated with defects in skin barrier structure and function. However, the molecular processes induced by the transglutaminase 1 deficiency are not fully understood. The aim of the present study was to uncover those processes by analysis of cutaneous molecular signatures. Gene expression profiles of wild-type and Tgm1–/–epidermis were assessed using microarrays. Gene ontology analysis of the data showed that genes for innate defense responses were up-regulated in Tgm1–/–epidermis. Based on that result, the induction of Il1b and antimicrobial peptide genes, S100a8, S100a9, Defb14, Camp, Slpi, Lcn2, Ccl20 and Wfdc12, was confirmed by quantitative real-time PCR. A protein array revealed that levels of IL-1β, G-CSF, GM-CSF, CXCL1, CXCL2, CXCL9 and CCL2 were increased in Tgm1–/–skin. Epidermal growth factor receptor (EGFR) ligand genes, Hbegf, Areg and Ereg, were activated in Tgm1–/–epidermis. Furthermore, the antimicrobial activity of an epidermal extract from Tgm1–/–mice was significantly increased against both Escherichia coli and Staphylococcus aureus. In the epidermis of ichthyosiform skins from patients with TGM1 mutations, S100A8/9 was strongly positive. The expression of those antimicrobial and defense response genes was also increased in the lesional skin of an ARCI patient with TGM1 mutations. These results suggest that the up-regulation of molecular signatures for antimicrobial and innate defense responses is characteristic of skin with a transglutaminase 1 deficiency, and this autonomous process might be induced to reinforce the defective barrier function of the skin. [ABSTRACT FROM AUTHOR]

Tumor necrosis factor alpha (TNF-α) is a pro-inflammatory cytokine important in normal and pathological biological processes. Newly synthesized pro-TNF-α is expressed on the plasma membrane and cleaved to release soluble TNF-α protein: both are biologically active. Secreted TNF-α signals through TNF receptors and the membrane-bound TNF-α acts by cell contact-dependent signaling. Anti-TNF-α antibodies have been used effectively for treatment of chronic inflammation, however with adverse side effects. Thus, there is a need for new anti-TNF-α small molecule compounds. Anti-TNF-α activity assays involve treatment of keratinocytes with exogenous TNF-α before or after anti-TNF-α incubation. However, this model fails to address the dual signaling of TNF-α. Here we describe a Doxycycline (Dox)-inducible TNF-α (HaCaT-TNF-α) expression system in keratinocytes. Using this in-vitro model, we show cell inhibition and induced expression of pro-inflammatory cytokines and markers, including IL-1β, IL-6, IL-8, NF-κB1, and KRT-16, similar to cells treated with exogenous TNF-α. Sufficient secreted TNF-α produced also activated IL-1β and IL-8 expression in wt HaCaT cells. Importantly, stimulated expression of IL-1β and IL-8 in HaCaT-TNF-α were blocked by Quercetin, a flavanol shown to possess anti-TNF-α activities. This novel in vitro cell model provides an efficient tool to investigate the dual signaling of TNF-α. Importantly, this model provides an effective, fast, and simple screening for compounds with anti-TNF-α activities for chronic inflammatory disease therapies. [ABSTRACT FROM AUTHOR]

Background: Human Campylobacter jejuni infections are progressively rising worldwide. Information about the molecular mechanisms underlying campylobacteriosis, however, are limited. In the present study we investigated whether cytokines such as IL-23, IL-22 and IL-18, which share pivotal functions in host immunity, were involved in mediating intestinal and systemic immunopathological responses upon C. jejuni infection. Methodology/Principal Findings: To assure stable infection, gnotobiotic (i.e. secondary abiotic) IL-23p19-/-, IL-22-/- and IL-18-/- mice were generated by broad-spectrum antibiotic treatment. Following peroral C. jejuni strain 81–176 infection, mice of all genotypes harbored comparably high pathogenic loads in their intestines. As compared to wildtype controls, however, IL-18-/- mice displayed less distinct C. jejuni induced sequelae as indicated by less pronounced large intestinal shrinkage and lower numbers of apoptotic cells in the colonic epithelial layer at day 8 postinfection (p.i.). Furthermore, lower colonic numbers of adaptive immune cells including regulatory T cells and B lymphocytes were accompanied by less distinct secretion of pro-inflammatory cytokines such as TNF and IFN-γ and lower IL-17A mRNA expression levels in colonic ex vivo biopsies of infected IL-18-/- as compared to wildtype mice. Upon C. jejuni infection, colonic IL-23p19 expression was up-regulated in IL-18-/- mice only, whereas IL-22 mRNA levels were lower in uninfected and infected IL-23p19-/- as well as infected IL-18-/- as compared to respective wildtype control mice. Remarkably, not only intestinal, but also systemic infection-induced immune responses were less pronounced in IL-18-/- mice as indicated by lower TNF, IFN-γ and IL-6 serum levels as compared to wildtype mice. Conclusion/Significance: We here show for the first time that IL-18 is essentially involved in mediating C. jejuni infection in the gnotobiotic mouse model. Future studies need to further unravel the underlying regulatory mechanisms orchestrating pathogen-host interaction. [ABSTRACT FROM AUTHOR]

Severe Staphylococcus aureus (S. aureus) infections pose an immense threat to population health and constitute a great burden for the health care worldwide. Inter alia, S. aureus septic arthritis is a disease with high mortality and morbidity caused by destruction of the infected joints and systemic bone loss, osteoporosis. Toll-Like receptors (TLRs) are innate immune cell receptors recognizing a variety of microbial molecules and structures. S. aureus recognition via TLR2 initiates a signaling cascade resulting in production of various cytokines, but the mechanisms by which S. aureus causes rapid and excessive bone loss are still unclear. We, therefore, investigated how S. aureus regulates periosteal/endosteal osteoclast formation and bone resorption. S. aureus stimulation of neonatal mouse parietal bone induced ex vivo bone resorption and osteoclastic gene expression. This effect was associated with increased mRNA and protein expression of receptor activator of NF-kB ligand (RANKL) without significant change in osteoprotegerin (OPG) expression. Bone resorption induced by S. aureus was abolished by OPG. S. aureus increased the expression of osteoclastogenic cytokines and prostaglandins in the parietal bones but the stimulatory effect of S. aureus on bone resorption and Tnfsf11 mRNA expression was independent of these cytokines and prostaglandins. Stimulation of isolated periosteal osteoblasts with S. aureus also resulted in increased expression of Tnfsf11 mRNA, an effect lost in osteoblasts from Tlr2 knockout mice. S. aureus stimulated osteoclastogenesis in isolated periosteal cells without affecting RANKL-stimulated resorption. In contrast, S. aureus inhibited RANKL-induced osteoclast formation in bone marrow macrophages. These data show that S. aureus enhances bone resorption and periosteal osteoclast formation by increasing osteoblast RANKL production through TLR2. Our study indicates the importance of using different in vitro approaches for studies of how S. aureus regulates osteoclastogenesis to obtain better understanding of the complex mechanisms of S. aureus induced bone destruction in vivo. [ABSTRACT FROM AUTHOR]

Gfi1 plays an important role in the development and maintenance of many hematopoietic linage cells. However, the impact of Gfi1-deficiency on the iNKT cell differentiation remains unclear. We herein demonstrate a critical role of Gfi1 in regulating the development of iNKT cell subsets. In the thymus of T cell-specific Gfi1-deficient mice, iNKT cells normally developed up to stage 2, while the number of stage 3 NK1.1pos iNKT cells was significantly reduced. Furthermore, CD4pos iNKT cells were selectively reduced in the peripheral organs of T cell-specific Gfi1-deficient mice. The α-GalCer-dependent production of IFN-γand Th2 cytokines, but not IL-17A, was severely reduced in T cell-specific Gfi1-deficient mice. In addition, a reduction of the α-GalCer-induced anti-tumor activity was observed in Gfi1-deficient mice. These findings demonstrate the important role of Gfi1 in regulating the development and function of NKT1- and NKT2-type iNKT cell subsets. [ABSTRACT FROM AUTHOR]

Background: Pulmonary arterial hypertension, also known as Ascites syndrome (AS), remains a clinically challenging disease with a large impact on both humans and broiler chickens. Pulmonary arterial remodeling presents a key step in the development of AS. The precise molecular mechanism of pulmonary artery remodeling regulating AS progression remains unclear. Methodology/Principal Findings: We obtained pulmonary arteries from two positive AS and two normal broilers for RNA sequencing (RNA-seq) analysis and pathological observation. RNA-seq analysis revealed a total of 895 significantly differentially expressed genes (DEGs) with 437 up-regulated and 458 down-regulated genes, which were significantly enriched to 12 GO (Gene Ontology) terms and 4 KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways (Padj<0.05) regulating pulmonary artery remodeling and consequently occurrence of AS. These GO terms and pathways include ribosome, Jak-STAT and NOD-like receptor signaling pathways which regulate pulmonary artery remodeling through vascular smooth cell proliferation, inflammation and vascular smooth cell proliferation together. Some notable DEGs within these pathways included downregulation of genes like RPL 5, 7, 8, 9, 14; upregulation of genes such as IL-6, K60, STAT3, STAT5 Pim1 and SOCS3; IKKα, IkB, P38, five cytokines IL-6, IL8, IL-1β, IL-18, and MIP-1β. Six important regulators of pulmonary artery vascular remodeling and construction like CYP1B1, ALDH7A1, MYLK, CAMK4, BMP7 and INOS were upregulated in the pulmonary artery of AS broilers. The pathology results showed that the pulmonary artery had remodeled and become thicker in the disease group. Conclusions/Significance: Our present data suggested some specific components of the complex molecular circuitry regulating pulmonary arterial remodeling underlying AS progression in broilers. We revealed some valuable candidate genes and pathways that involved in pulmonary artery remodeling further contributing to the AS progression. [ABSTRACT FROM AUTHOR]

Sphingosine-1-phosphate receptor 2 (S1PR2) couples with the Gi, Gq, and G12/13 group of proteins, which modulate an array of cellular signaling pathways and affect immune responses to multiple stimuli. In this study, we demonstrated that knockdown of S1PR2 by a specific S1PR2 shRNA lentiviral vector significantly inhibited IL-1β, IL-6, and TNF-α protein levels induced by oral pathogen Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) in murine bone marrow-derived monocytes and macrophages (BMMs) compared with controls. In addition, knockdown of S1PR2 by the S1PR2 shRNA lentiviral vector suppressed p-PI3K, p-ERK, p-JNK, p-p38, and p-NF-κBp65 protein expressions induced by A. actinomycetemcomitans. Furthermore, bone marrow cells treated with the S1PR2 shRNA lentiviral vector inhibited osteoclastogenesis induced by RANKL compared with controls. The S1PR2 shRNA suppressed the mRNA levels of six osteoclastogenic factors including nuclear factor of activated T-cells cytoplasmic calcineurin-dependent 1 (NFATc1), cathepsin K (Ctsk), acid phosphatase 5 (Acp5), osteoclast-associated receptor (Oscar), dendritic cells specific transmembrane protein (Dcstamp), and osteoclast stimulatory transmembrane protein (Ocstamp) in bone marrow cells. We conclude that S1PR2 plays an essential role in modulating proinflammatory cytokine production and osteoclastogenesis. Blocking S1PR2 signaling might be a novel therapeutic strategy to treat inflammatory bone loss diseases. [ABSTRACT FROM AUTHOR]

Epidemiological differences exist between Mycobacterium africanum (Maf)- and Mycobacterium tuberculosis (Mtb)-infected patients, but to date, contributing host factors have not been characterised. We analysed clinical outcomes, as well as soluble markers and gene expression profiles in unstimulated, and ESAT6/CFP-10-, whole-Maf- and Mtb-stimulated blood samples of 26 Maf- and 49 Mtb-HIV-negative tuberculosis patients before, and after 2 and 6 months of anti-tuberculosis therapy. Before treatment, both groups had similar clinical parameters, but differed in few cytokines concentration and gene expression profiles. Following treatment the body mass index, skinfold thickness and chest X-ray scores showed greater improvement in the Mtb- compared to Maf-infected patients, after adjusting for age, sex and ethnicity (p = 0.02; 0.04 and 0.007, respectively). In addition, in unstimulated blood, IL-12p70, IL12A and TLR9 were significantly higher in Maf-infected patients, while IL-15, IL-8 and MIP-1α were higher in Mtb-infected patients. Overnight stimulation with ESAT-6/CFP-10 induced significantly higher levels of IFN-γ and TNF-α production, as well as gene expression of CCL4, IL1B and TLR4 in Mtb- compared to Maf-infected patients. Our study confirms differences in clinical features and immune genes expression and concentration of proteins associated with inflammatory processes between Mtb- and Maf-infected patients following anti-tuberculosis treatment These findings have public health implications for treatment regimens, and biomarkers for tuberculosis diagnosis and susceptibility. [ABSTRACT FROM AUTHOR]

Adiponectin is the most abundantly produced human adipokine with anti-inflammatory, anti-oxidative, and insulin-sensitizing properties. Evidence from in vitro studies has indicated that adiponectin has a potential role in reproduction because it reduces the production of androstenedione in bovine theca cells in vitro. However, this effect on androgen production has not yet been observed in vivo. The current study evaluated the effect of adiponectin on androstenedione secretion and oxidative stress parameters in a rodent model. Seven-week-old female Balb/c mice (n = 33), previously treated with equine gonadotropin chorionic, were assigned to one of four different treatments: Group 1, control (phosphate-buffered saline); Group 2, adiponectin 0.1 μg/mL; Group 3, adiponectin 1.0 μg/mL; Group 4, adiponectin 5.0 μg/mL. After 24 h, all animals were euthanized and androstenedione levels were measured in the serum while oxidative stress markers were quantified in whole ovary tissue. Female mice treated with adiponectin exhibited a significant reduction (about 60%) in serum androstenedione levels in comparison to controls. Androstenedione levels decreased from 0.78 ± 0.4 ng/mL (mean ± SD) in controls to 0.28 ± 0.06 ng/mL after adiponectin (5 μg/mL) treatment (P = 0.01). This change in androgen secretion after 24 hours of treatment was associated with a significant reduction in the expression of CYP11A1 and STAR (but not CYP17A1). In addition, ovarian AOPP product levels, a direct product of protein oxidation, decreased significantly in adiponectin-treated mice (5 μg/mL); AOPP (mean ± SD) decreased to 4.3 ± 2.1 μmol/L in comparison with that of the controls (11.5 ± 1.7 μmol/L; P = 0.0003). Our results demonstrated for the first time that acute treatment with adiponectin reduced the levels of a direct oxidative stress marker in the ovary as well as decreased androstenedione serum levels in vivo after 24 h. [ABSTRACT FROM AUTHOR]

Mares that fail to conceive or lose their embryos, without showing typical signs of clinical endometritis, should be suspected of subclinical endometritis (SE). In this study, the question was addressed: does SE fully activate selected mechanisms of innate immunity in mares? For this aim, expression of mRNAs for Toll-like Receptor 2 and 4 (TLR 2/4), interleukin 1β (IL-1β), interleukin 6 (IL-6) and tumor necrosis factor α (TNF) was examined in control mares versus either mares suffering from chronic endometritis (ChE) or subacute suppurative endometritis (SSE). The concentrations of IL-1β, IL-6 and TNF-α in supernatants from endometrial tissue cultures after 4 h incubation were measured using the enzyme immunoassay (EIA) method. Eighty-two warmblood mares, of known breeding history, were enrolled in this study. Based on histopathological assessment, mares were classified as suffering from ChE, SSE or as being healthy. In addition, immuno-localization of both TLR2 and TLR4 as well as TNF-α was investigated in the equine endometria. The mRNA expression of TLR2 (P < 0.01), IL-1β (P < 0.0001), IL-6 (P < 0.0001) and TLR4 and TNF (P < 0.05) was up-regulated in endometria of mares suffering from SSE compared with unaffected mares. Concentrations of IL-6 and TNF-α were increased only in mares exhibiting SSE, compared with unaffected (P < 0.01 for both) and ChE mares (P < 0.05 for both). Immuno-localization of TNF-α and TLRs was confirmed, both in unaffected and SE-affected endometria, and was present in the luminal and glandular epithelia and stromal cells. The severity of inflammation impacts the immune response and fosters activation of innate immunity mechanisms, as observed in the endometria of mares. The intracellular localization of TLRs and TNF-α in the endometria indicates a key role of endometrial epithelial and stromal cells in the immune response and inflammation. [ABSTRACT FROM AUTHOR]

Excessive mechanical ventilation exerts pathologic mechanical strain on lung vascular endothelium and promotes endothelial cell (EC) inflammatory activation; however, the specific mechanisms underlying EC inflammatory response caused by mechanical ventilation related cyclic stretch (CS) remain unclear. This study investigated the effects of chronic exposure to CS at physiologic (5%) and pathologic (18%) magnitude on pulmonary EC inflammatory status in control conditions and bacterial lipopolysacharide (LPS)-stimulated conditions. EC exposure to high or low CS magnitudes for 28–72 hrs had distinct effects on EC inflammatory activation. 18% CS increased surface expression of endothelial adhesion molecule ICAM1 and release of its soluble form (sICAM1) and inflammatory cytokine IL-8 by CS-stimulated pulmonary endothelial cells (EC). EC inflammatory activation was not observed in EC exposed to 5% CS. Chronic exposure to 18% CS, but not to 5% CS, augmented ICAM1 and IL-8 production and EC monolayer barrier disruption induced by LPS. 18% CS, but not 5% CS, stimulated expression of RhoA GTPase-specific guanine nucleotide exchange factor GEF-H1. GEF-H1 knockdown using gene-specific siRNA abolished 18% CS-induced ICAM1 expression and sICAM1 and IL-8 release by EC. GEF-H1 knockdown also prevented disruption of EC monolayer integrity and attenuated sICAM1 and IL-8 release in the two-hit model of EC barrier dysfunction caused by combined stimulation with 18% CS and LPS. These data demonstrate that exacerbation of inflammatory response by pulmonary endothelium exposed to excessive mechanical stretch is mediated by CS-induced induction of Rho activating protein GEF-H1. [ABSTRACT FROM AUTHOR]

Since inflammatory bowel diseases (IBD) represent significant morbidity and mortality in the US, the need for defining novel drug targets and inflammatory mechanisms would be of considerable benefit. Although protein tyrosine kinase 6 (PTK6, also known as breast tumor kinase BRK) has been primarily studied in an oncogenic context, it was noted that PTK6 null mice exhibited significantly enhanced colonic epithelial barrier function. Considering that the inflammatory functions of PTK6 have not yet been explored, we hypothesized that cytokines responsible for mediating IBD, such as TNFα/IFNγ, may solicit the action of PTK6 to alter barrier function. After first assessing critical mediators of TNFα/IFNγ driven epithelial barrier dysfunction, we further explored the possibility of PTK6 in this inflammatory context. In this report, we showed that PTK6 siRNA and PTK6 null young adult mouse colonic epithelial cells (YAMC) exhibited significant attenuation of TNFα/IFNγ induced barrier dysfunction as measured by electric cell-substrate impedance sensing (ECIS) assay and permeability assays. In addition, PTK6 null cells transfected with PTK6 cDNA displayed restored barrier dysfunction in response to TNFα/IFNγ, while the cells transfected with vector alone showed similar attenuation of barrier dysfunction. Furthermore, using subcellular fractionation and immunocytochemistry experiments, we found that PTK6 plays a role in FoxO1 nuclear accumulation leading to down-regulation of claudin-3, a tight junction protein. Moreover, we searched for relevant miRNA candidates putative for targeting PTK6 in order to identify and assess the impact of microRNA that target PTK6 with respect to TNFα/IFNγ induced barrier dysfunction. Subsequently, we assayed likely targets and determined their effectiveness in attenuating PTK6 expression as well as cytokine induced barrier dysfunction. Results showed that miR-93 reduced PTK6 expression and attenuated TNFα/IFNγ imposed decrease in transepithelial electrical resistance (TER), as well as excluded FoxO1 from the nucleus. Our results indicate that PTK6 may act as a novel mediator of intestinal epithelial permeability during inflammatory injury, and miR-93 may protect intestinal epithelial barrier function, at least in part, by targeting PTK6. [ABSTRACT FROM AUTHOR]

Background & Aims: We previously demonstrated that short-term oral administration of the probiotic Lactobacillus reuteri 6475 enhanced bone density in male but not female mice. We also established that L. reuteri 6475 enhanced bone health and prevented bone loss in estrogen-deficient female mice. In this study, we tested whether a mild inflammatory state and/or a long-term treatment with the probiotic was required to promote a positive bone effect in estrogen-sufficient female mice. Methods: A mild inflammatory state was induced in female mice by dorsal surgical incision (DSI). Following DSI animals were orally supplemented with L. reuteri or vehicle control for a period of 8 weeks. Gene expression was measured in the intestine and bone marrow by qPCR. Distal femoral bone density and architecture was analyzed by micro-CT. Results: We report that 8 weeks after DSI there is a significant increase in the weight of spleen, thymus and visceral (retroperitoneal) fat pads. Expression of intestinal cytokines and tight junction proteins are also altered 8 weeks post-DSI. Interestingly, L. reuteri treatment was found to display both intestinal region- and inflammation-dependent effects. Unexpectedly we identified that 1) L. reuteri treatment increased bone density in females but only in those that underwent DSI and 2) DSI benefited cortical bone parameters. In the bone marrow, dorsal surgery induced CD4+ T cell numbers, a response that was unaffected by L. reuteri treatment, whereas expression of RANKL, OPG and IL-10 were significantly affected by L. reuteri treatment. Conclusion: Our data reveals a previously unappreciated effect of a mild surgical procedure causing a long-lasting effect on inflammatory gene expression in the gut and the bone. Additionally, we demonstrate that in intact female mice, the beneficial effect of L. reuteri on bone requires an elevated inflammatory status. [ABSTRACT FROM AUTHOR]

The cytokine IL-33 is constitutively expressed in epithelial cells and it augments Th2 cytokine-mediated inflammatory responses by regulating innate immune cells. We aimed to determine the role of the periodontal pathogen, Porphyromonas gingivalis, in the enhanced expression of IL-33 in human gingival epithelial cells. We detected IL-33 in inflamed gingival epithelium from patients with chronic periodontitis, and found that P. gingivalis increased IL-33 expression in the cytoplasm of human gingival epithelial cells in vitro. In contrast, lipopolysaccharide, lipopeptide, and fimbriae derived from P. gingivalis did not increase IL-33 expression. Specific inhibitors of P. gingivalis proteases (gingipains) suppressed IL-33 mRNA induction by P. gingivalis and the P. gingivalis gingipain-null mutant KDP136 did not induce IL-33 expression. A small interfering RNA for protease-activated receptor-2 (PAR-2) as well as inhibitors of phospholipase C, p38 and NF-κB inhibited the expression of IL-33 induced by P. gingivalis. These results indicate that the PAR-2/IL-33 axis is promoted by P. gingivalis infection in human gingival epithelial cells through a gingipain-dependent mechanism. [ABSTRACT FROM AUTHOR]

Inflammation has an important role in the development of liver fibrosis in general and the activation of hepatic stellate cells (HSCs) in particular. It is known that HSCs are themselves able to produce cytokines and chemokines, and that this production may be a key event in the initiation of fibrogenesis. However, the direct involvement of cytokines and chemokines in HSC (self-)activation remains uncertain. In this study, the effects of pro-inflammatory cytokines IL-1α and β, TNF-α, and IL-8 on the activation state of HSCs were examined, in comparison to the pro-fibrogenic mediator TGF-β1. LX-2 cells were stimulated for 24 or 48 hours with recombinant human form of the pro-inflammatory cytokines IL-1α and β, TNF-α, and IL-8, and also the pro-fibrogenic mediator TGF-β1. Two drugs were also evaluated, the anti-TNF-α monoclonal antibody infliximab and the IL-1 receptor antagonist anakinra, regarding their inhibitory effects. In LX-2 human HSC, treatment with TGF-β1 are associated with downregulation of the metalloproteinase (MMP)-1 and MMP-3, with upregulation of tissue inhibitor of metalloproteinase (TIMP)-1, collagen type I α1, collagen type IV α1, α-SMA, endothelin-1 and PDGF-BB. Cytokines and chemokines expression were found to be downregulated, excepting IL-6. In contrast, we observed that LX-2 exposure to IL-1, TNF-α and IL-8 can reverse the phenotype of pro-fibrogenic activated cells. Indeed, MMP-1, MMP-3 and MMP-9 were found elevated, associated with downregulation of α-SMA and/or PDGF-BB, and a greater expression of IL-1β, IL-6, IL-8, CXCL1 and CCL2. Lastly, we found that infliximab and anakinra successfully inhibits effects of TNF-α and IL-1 respectively in LX-2 cells. Infliximab and anakinra may be of value in preclinical trials in chronic liver disease. Overall, our results suggest that (i) pro-inflammatory mediators exert complex effects in HSCs via an MMP/TIMP imbalance, and (ii) targeting IL-1 signaling may be a potentially valuable therapeutic strategy in chronic liver diseases. [ABSTRACT FROM AUTHOR]

An important epigenetic modification in Huntington’s disease (HD) research is histone acetylation, which is regulated by histone acetyltransferase and histone deacetylase (HDAC) enzymes. HDAC inhibitors have proven effective in HD model systems, and recent work is now focused on functional dissection of the individual HDAC enzymes in these effects. Histone deacetylase 3 (HDAC3), a member of the class I subfamily of HDACs, has previously been implicated in neuronal toxicity and huntingtin-induced cell death. Hence, we tested the effects of RGFP966 ((E)-N-(2-amino-4-fluorophenyl)-3-(1-cinnamyl-1H-pyrazol-4-yl)acrylamide), a benzamide-type HDAC inhibitor that selectively targets HDAC3, in the N171-82Q transgenic mouse model of HD. We found that RGFP966 at doses of 10 and 25 mg/kg improves motor deficits on rotarod and in open field exploration, accompanied by neuroprotective effects on striatal volume. In light of previous studies implicating HDAC3 in immune function, we measured gene expression changes for 84 immune-related genes elicited by RGFP966 using quantitative PCR arrays. RGFP966 treatment did not cause widespread changes in cytokine/chemokine gene expression patterns, but did significantly alter the striatal expression of macrophage migration inhibitory factor (Mif), a hormone immune modulator associated with glial cell activation, in N171-82Q transgenic mice, but not WT mice. Accordingly, RGFP966-treated mice showed decreased glial fibrillary acidic protein (GFAP) immunoreactivity, a marker of astrocyte activation, in the striatum of N171-82Q transgenic mice compared to vehicle-treated mice. These findings suggest that the beneficial actions of HDAC3 inhibition could be related, in part, with lowered Mif levels and its associated downstream effects. [ABSTRACT FROM AUTHOR]

White adipose tissue (WAT) plays a fundamental role in maintaining energy balance and important endocrine functions. The loss of WAT modifies adipokine secretion and disrupts homeostasis, potentially leading to severe metabolic effects and a reduced quality of life. Doxorubicin is a chemotherapeutic agent used clinically because of its good effectiveness against various types of cancer. However, doxorubicin has deleterious effects in many healthy tissues, including WAT, liver, and skeletal and cardiac muscles. Our objective was to investigate the effects of doxorubicin on white adipocytes through in vivo and in vitro experiments. Doxorubicin reduced the uptake of glucose by retroperitoneal adipocytes and 3T3-L1 cells via the inhibition of AMP-activated protein kinase Thr172 phosphorylation and glucose transporter 4 content. Doxorubicin also reduced the serum level of adiponectin and, to a greater extent, the expression of genes encoding lipogenic (Fas and Acc) and adipogenic factors (Pparg, C/ebpa, and Srebp1c) in retroperitoneal adipose tissue. In addition, doxorubicin inhibited both lipogenesis and lipolysis and reduced the hormone-sensitive lipase and adipose tissue triacylglycerol lipase protein levels. Therefore, our results demonstrate the impact of doxorubicin on WAT. These results are important to understand some side effects observed in patients receiving chemotherapy and should encourage new adjuvant treatments that aim to inhibit these side effects. [ABSTRACT FROM AUTHOR]