mRNA Expression Of IL-1β Research Articles

Cyclohexylethanoid derivative and rearranged abietane diterpenoids with anti-inflammatory activities from Clerodendrum bungei and C. inerme

One new dimeric cyclohexylethanoid derivative (1) and one new rearranged abietane diterpenoid (2) were isolated from Clerodendrum bungei and C. inerme, respectively, together with 11 known compounds. The structures of the isolated compounds were elucidatedby detailed spectroscopic and ECD analyses. The absolute configuration of 2 was unequivocally determined using single-crystal X-ray diffraction analysis. Compound 1 showed the greatest inhibitory effect on NO production in RAW 264.7 murine macrophages induced by lipopolysaccharide (LPS) with the IC50 value of 6.328 µM. Compound 1 also suppressed the proinflammatory cytokines IFN-γ, TNF-α, IL-1β, iNOS and IL-6 mRNA expression, while increased the mRNA expression of the anti-inflammatory cytokines IL-4, IL-10 and Arg-1 through NF-κB signaling pathway. The further experiments confirmed that compound 1 demonstrated the strong in vivo anti-inflammatory effect by reducing mortality, the serum TNF-α, IL-6 and IFN-γ levels, and tissue toxicity in CLP-induced septic mice. Collectively, these findings suggested that compound 1might serve as a potential anti-inflammatory agent candidate for the sepsis treatment.

Effects of Enteromorpha prolifera polysaccharides on growth performance, intestinal barrier function and cecal microbiota in yellow-feathered broilers under heat stress

BackgroundGlobal warming leading to heat stress (HS) is becoming a major challenge for broiler production. This study aimed to explore the protective effects of seaweed (Enteromorpha prolifera) polysaccharides (EPS) on the intestinal barrier function, microbial ecology, and performance of broilers under HS. A total of 144 yellow-feathered broilers (male, 56 days old) with 682.59 ± 7.38 g were randomly assigned to 3 groups: 1) TN (thermal neutral zone, 23.6 ± 1.8 °C), 2) HS (heat stress, 33.2 ± 1.5 °C for 10 h/d), and 3) HSE (HS + 0.1% EPS). Each group contained 6 replicates with 8 broilers per replicate. The study was conducted for 4 weeks; feed intake and body weights were measured at the end of weeks 2 and 4. At the end of the feeding trial, small intestine samples were collected for histomorphology, antioxidant, secretory immunoglobulin A (sIgA) content, apoptosis, gene and protein expression analysis; cecal contents were also collected for microbiota analysis based on 16S rDNA sequencing.ResultsDietary EPS promoted the average daily gain (ADG) of broilers during 3–4 weeks of HS (P < 0.05). At the end of HS on broilers, the activity of total superoxide dismutase (T-SOD), glutathione S-transferase (GST), and the content of sIgA in jejunum were improved by EPS supplementation (P < 0.05). Besides, dietary EPS reduced the epithelial cell apoptosis of jejunum and ileum in heat-stressed broilers (P < 0.05). Addition of EPS in HS group broilers' diet upregulated the relative mRNA expression of Occludin, ZO-1, γ-GCLc and IL-10 of the jejunum (P < 0.05), whereas downregulated the relative mRNA expression of NF-κB p65, TNF-α and IL-1β of the jejunum (P < 0.05). Dietary EPS increased the protein expression of Occludin and ZO-1, whereas it reduced the protein expression of NF-κB p65 and MLCK (P < 0.01) and tended to decrease the protein expression of TNF-α (P = 0.094) in heat-stressed broilers. Furthermore, the proportions of Bacteroides and Oscillospira among the three groups were positively associated with jejunal apoptosis and pro-inflammatory cytokine expression (P < 0.05) and negatively correlated with jejunal Occludin level (P < 0.05). However, the proportions of Lactobacillus, Barnesiella, Subdoligranulum, Megasphaera, Collinsella, and Blautia among the three groups were positively related to ADG (P < 0.05).ConclusionsEPS can be used as a feed additive in yellow-feathered broilers. It effectively improves growth performance and alleviates HS-induced intestinal injury by relieving inflammatory damage and improving the tight junction proteins expression. These beneficial effects may be related to inhibiting NF-κB/MLCK signaling pathway activation and regulation of cecal microbiota.

Interleukin-1β triggers muscle-derived extracellular superoxide dismutase expression and protects muscles from doxorubicin-induced atrophy.

Doxorubicin, a conventional chemotherapeutic agent prescribed for cancer, causes skeletal muscle atrophy and adversely affects mobility and strength. Given that doxorubicin-induced muscle atrophy is attributable primarily to oxidative stress, its effects could be mitigated by antioxidant-focused therapies; however, these protective therapeutic targets remain ambiguous. The aim of this study was to demonstrate that doxorubicin triggers severe muscle atrophy via upregulation of oxidative stress (4-hydroxynonenal and malondialdehyde) and atrogenes (atrogin-1/MAFbx and muscle RING finger-1) in association with decreased expression of the antioxidant enzyme extracellular superoxide dismutase (EcSOD), in cultured C2C12myotubes and mouse skeletal muscle. Supplementation with EcSOD recombinant protein elevated EcSOD levels on the cellular membrane of cultured myotubes, consequently inhibiting doxorubicin-induced oxidative stress and myotube atrophy. Furthermore, doxorubicin treatment reduced interleukin-1β (IL-1β) mRNA expression in cultured myotubes and skeletal muscle, whereas transient IL-1β treatment increased EcSOD protein expression on the myotube membrane. Notably, transient IL-1β treatment of cultured myotubes and local administration in mouse skeletal muscle attenuated doxorubicin-induced muscle atrophy, which was associated with increased EcSOD expression. Collectively, these findings reveal that the regulation of skeletal muscle EcSOD via maintenance of IL-1β signalling is a potential therapeutic approach to counteract the muscle atrophy mediated by doxorubicin and oxidative stress. KEY POINTS: Doxorubicin, a commonly prescribed chemotherapeutic agent for patients with cancer, induces severe muscle atrophy owing to increased expression of oxidative stress; however, protective therapeutic targets are poorly understood. Doxorubicin induced muscle atrophy owing to increased expression of oxidative stress and atrogenes in association with decreased protein expression of extracellular superoxide dismutase (EcSOD) in cultured C2C12myotubes and mouse skeletal muscle. Supplementation with EcSOD recombinant protein increased EcSOD levels on the cellular membrane of cultured myotubes, resulting in inhibition of doxorubicin-induced oxidative stress and myotube atrophy. Doxorubicin treatment decreased interleukin-1β (IL-1β) expression in cultured myotubes and skeletal muscle, whereas transient IL-1β treatment in vivo and in vitro increased EcSOD protein expression and attenuated doxorubicin-induced muscle atrophy. These findings reveal that regulation of skeletal muscle EcSOD via maintenance of IL-1β signalling is a possible therapeutic approach for muscle atrophy mediated by doxorubicin and oxidative stress.

Region-specific remodeling of the enteric nervous system and enteroendocrine cells in the colon of spinal cord injury patients

Patients with spinal cord injury (SCI) suffer from major bowel dysfunction, whose exact pathophysiology, particularly the involvement of the enteric nervous system or epithelial dysfunction is poorly understood. Herein, we aimed to characterize the mucosal biopsies of the right and left colon in SCI patients vs controls (CT): (1) remodeling of key enteric neurotransmitters, (2) remodeling of enteroendocrine cells, and (3) mucosal inflammation compared to those in controls. In SCI, mucosal ACh concentration was lower in the right colon as compared to CT, but no change was observed in the left colon, and AChE expression was lower in both the right and left colons than in CT. While the VIP concentration was similar in the right and left colons, VIP mRNA expression was increased in the right colon and decreased in the left colon, in SCI patients as compared to CT. Interestingly, 5-HT concentration was reduced in the left colon but not in the right colon in SCI patients. Moreover, in SCI patients, as compared to CT, SERT mRNA expression was selectively increased in the left colon while TPH1 mRNA expression was increased in the right and left colons. Although mucosal TNFα and IL-1β mRNA expression did not significantly differ between SCI and CT groups, we identified a significant positive correlation between TNFα and IL-1β mRNA expression and left colon transit time in the SCI group. In conclusion, region-specific changes occur in the enteric neurotransmitter, serotonergic, and inflammatory pathways in the colon of SCI patients. The significant correlations between these pathways and clinical parameters in the left colon further set a scientific basis for designing therapeutic targets to improve colonic motor dysfunction in patients.Biobank information: Spinal cord injury patients: PHRC ConstiCAPE—clinical trial NCT02566746. Controls: Anosain—clinical trial NCT03054415 and biobank of the “Institut des Maladies de l’Appareil Digestif (IMAD)” registered under number DC-2008-402.

The Therapeutic Potential of Panax Ginseng and Ginger on Postsurgical Adhesion Band Formation

Background: Peritoneal adhesions (PA) are a common complication of abdominal operations. Previous studies indicate that inhibition of inflammation and fibrosis at sites of peritoneal damage may prevent the development of intra-abdominal adhesions. Zingiber officinalis Roscoe (ginger) and Panax ginseng (P. ginseng) are herbal products with antioxidant and anti-inflammatory effects that can have restorative properties. Objective: This research aimed to examine the impact of ginger and p. ginseng on prevention of PA in a rat model after surgery. Methods: Following a laparotomy, the wall of the cecum was rubbed to induce intra-abdominal adherence in Wistar rats. Ginger (400mg/kg) and P. ginseng (500mg/kg) were orally adminstered to the animals. The animals were sacrificed on the 10th day after surgery, and the Nair and Leach scoring system was used to assess adhesion. The microscopic histology of the induced cecal adhesions was evaluated. An enzyme-linked immunosorbent assay (ELISA) was used to determined tissue levels of transforming growth factor-beta (TGF-β) on homogenized PA tissue. Real-time PCR was performed to quantify the mRNA expression of IL-1β, TNF-α, Col 1a1, and Col 3a1 in rat tissue. Results: The adhesion score and histopathological rating based on the Nair and Leach scoring criteria showed lower adhesion scores in the group of rats treated with P. ginseng compared to the control group (p&lt;0.05). Treatment with ginger and P. ginseng was associated with reduced tissue level of TGF-β and mRNA expression of TNF-α and IL-1β. The gene expression level of Col 1a1 and Col 3a1 were markedly decreased in the P. ginseng group. Conclusion: The study's outcome recommends that P. ginseng could be an effective agent for preventing the PA and inflammation during the post-operative stage.

Trelagliptin relieved cognitive impairment of diabetes mellitus rats: Involvement of PI3K/Akt/GSK-3β and inflammation pathway

Cognitive impairment frequently coexists with diabetes. Trelagliptin is a once-weekly taking selective dipeptidyl peptidase-4 (DPP-4) inhibitor and a long-term effective hypoglycemic medicine; nonetheless, its effects for the treatment of diabetes-related cognitive impairment have only sometimes been explored. In this study, a DM model was built using streptozotocin (STZ) and a high-fat diet (HFD). The morris water maze test on DM rats revealed a considerably reduced capacity for spatial learning and memory, but trelagliptin was able to restore function. Trelagliptin could lower the mRNA expression of inflammatory factors such IL-1β, TNF-α, and IL-6 in DM rats. It could also reduce the ratio of p-IKKα/IKKα, and the immunofluorescence result of NF-κB also demonstrated a drop. Trelagliptin partially restored dendritic spines and prevented the loss or shrinkage of neurons, respectively, according to the results of Nissl's staining and golgi staining. Furthermore, PI3K/Akt/GSK-3β has been activated, and synaptic plasticity has been modified during this process. In conclusion, trelagliptin improved the cognitive lesion in DM rats by suppressing the activation of the inflammatory route and by activating the PI3K/Akt/GSK-3β pathway at the same time, as well as interacting with the pathways that protect neurons, which still need further research.

NLRP3 neuroinflammatory intervention of Mahuang-Lianqiao-Chixiaodou decoction for mental disorders in atopic dermatitis mice

Ethnopharmacological relevanceMahuang-Lianqiao-Chixiaodou decoction (MLCD) is a traditional Chinese medicinal (TCM) formula recorded in the Treatise on Febrile Diseases. It is commonly used for clinical treatment of atopic dermatitis (AD). However, the potential mechanisms of MLCD intervention in AD combined with mental disorders behaviors such as anxiety and depression remain elusive and deserves further investigation. Aim of the studyThe study aims to observe the effect of MLCD on anxiety- and depression-like behaviors in AD mice and explore the possible neuroinflammatory mechanism of NOD-like receptor 3 (NLRP3) inflammasome. Materials and methodsThe chemical components of MLCD extracts were identified using UHPLC-MS. The AD mice were induced by 2,4-dinitrofluorobenzene and treated with MLCD or mometasone furoate (MF, as a positive control) for 7 days. The pathological changes in their skin tissue and brain hippocampus were observed by hematoxylin-eosin staining. Elevated plus-maze test (EPM), open field test (OFT), and the suspended tail (TST) were used to measure the anxiety- and depressive-like behaviors in AD mice. Expression of NLRP3 inflammasome-related proteins in brain hippocampus were measured by the quantitative real-time polymerase chain reaction (qPCR) and western blotting (WB). ResultsWe found that MLCD contain many active ingredients, including ephedrine, Forsythoside A, phillyrin, glycyrrhizic acid, etc. Both MLCD and MF alleviated skin lesions and promoted positive histopathological changes in the hippocampus of AD mince to varying degrees. MLCD however, could further increase their proportion of open arm entry times (Oentries%) in EPM, residence time in the central area (Ctime) and the proportion of the number of times in the central area (Centries%) in OFT significantly. MLCD also reduces their immobility time in TST considerably. Mechanistically, MLCD downregulated the relative mRNA expression and protein level of NLRP3, Caspase-1, IL-1β, and IL-18 in hippocampal tissue compared to the model group. ConclusionsMLCD can alleviate anxiety-like and depression-like behaviors in AD mice by intervening in the gene and protein expression of NLRP3 inflammasome-related factors, thus treating AD.

Expression of interleukin-1β and its receptor in human granulosa cells and their association with steroidogenesis

This study aimed to investigate whether interleukin 1β (IL-1β) and soluble IL-1 receptor 2 (sIL-1R2) are expressed in human granulosa cells (GCs) and relate to ovarian steroidogenesis. Ninety-six women undergoing in vitro fertilization (IVF) were recruited. RT-PCR and immunocytochemistry were used to detect mRNAs and proteins of IL-1β and IL-1R2, respectively. The steroidogenesis of primary cultured GCs was evaluated following treatment with either IL-1β alone or IL-1β and FSH in combination. There were positive correlations between serum IL-1β and serum progesterone (r = 0.220, p = 0.032) and follicular fluid (FF) estradiol (r = 0.242, p = 0.018). Additionally, serum and FF sIL-1R2 were negatively and positively correlated with FF estradiol (r = −0.376, p = 0.005) and FF progesterone (r = 0.434, p = 0.001), respectively. The mRNA and protein expression of IL-1β and IL-1R2 became evident in GCs. IL-1β alone significantly increased estradiol secretion from GCs, but in the presence of FSH, it could notably promote progesterone secretion in addition to estradiol. In conclusion, IL-1β and sIL-1R2 are expressed in human GCs and substantially contribute to ovarian steroidogenesis, suggesting that the IL-1β system may be a potential target for optimizing ovarian hyperstimulation and steroidogenesis in IVF cycles.

Xinmaikang (XMK) tablets alleviate atherosclerosis by regulating the SREBP2-mediated NLRP3/ASC/Caspase-1 signaling pathway

Ethnopharmacological relevanceXinmaikang (XMK) tablets, a Chinese patent medicine, have been used for the prevention and treatment of atherosclerosis (AS) clinically. However, the underlying mechanism of XMK is far from completely illustrated. Aim of the studyThis study aimed to determine whether XMK alleviates AS in Apolipoprotein E-knockout (ApoE−/−) mice and to explore the potential mechanism of action in bone marrow-derived macrophages (BMDMs). Materials and methodsXMK decoction was analyzed by an LC‒MS/MS assay. Molecular docking was conducted to determine the interaction of XMK molecular ligands and AS targets. In vivo, 10 ApoE−/− mice were selected as the control group. Fifty ApoE−/− mice were randomly divided into 5 groups: the model group, low-, medium-, and high-dose XMK groups and the simvastatin group. Mice in the control group were fed a chow diet, and the other 5 groups were fed a high-fat diet (HFD) for 12 weeks. After 12 weeks, the treatment groups were administered low-dose XMK (2.28·kg-1·d), medium-dose XMK (4.55·kg-1·d), high-dose XMK (9.1 kg−1 d) and simvastatin (91 mg−1 d) for another 12 weeks. Serum enzymology assays tested AST/ALT, Cr, LDH and CK-MB levels. The atherosclerotic plaques and lipid deposition were measured by Oil red O (ORO) staining and Hematoxylin and Eosin (H&E) staining. Then, we examined the body weight and serum lipids (TC, TG, LDL-C and HDL-C) of the mice. ELISA was performed to determine the levels of inflammatory factors (IL-6, TNF-ɑ, VCAM-1, CXCL8 and CCL2). SREBP2/NLRP3 signaling pathway-related genes (SREBP2, NLRP3, ASC, IL-1β and Caspase-1) were analyzed by RT‒qPCR and western blotting. In vitro, LPS-stimulated BMDMs were treated with different concentrations of XMK (1, 2.5, 5, 10, 20, and 40 μg/ml). Immunofluorescence staining (SREBP2, NLRP3), adenovirus infection and siRNA knockdown (SREBP2, NLRP3, Caspase-1 and ASC) were conducted as complements to the in vivo experiment. ResultsMolecular docking showed a stable interaction between the effective components of XMK and SREBP2 and NLRP3. Serum enzymology assays revealed the medication safety of XMK in cardiac, hepatic and renal function. Studies in vivo indicated that XMK improved serum lipids (TC, TG, LDL-C and HDL-C) and reduced plaque area. Body weight decreased, and the expression of inflammatory cytokines (IL-6, TNF-ɑ and VCAM-1) was inhibited. Then, XMK downregulated the mRNA and protein expression of SREBP2, NLRP3, ASC, IL-1β and Caspase-1. In vitro, the above findings were reinforced in BMDMs, and knocking down SREBP2 restrained the effect of XMK on the NLRP3/ASC/Caspase-1 signaling pathway. ConclusionsXMK restrains AS by improving inflammation through the SREBP2-mediated NLRP3/ASC/Caspase-1 signaling pathway.

A-007 Cardiomyocyte-monocyte Cross-talk Exploits Cellular Stress Pathways Leading to Myocyte Apoptosis in Dilated Cardiomyopathy of Idiopathic Origin

Abstract Background Dilated cardiomyopathy (DCM) is the most prevalent form of cardiovascular diseases (CVDs) results in dilation and disruption of pumping mechanism of heart. Most strikingly, about 50% DCM cases do not have any identified aetiology, hence known as idiopathic DCM (IDCM). The aim of this study is to investigate major inflammatory markers and cellular stress pathways in the circulating leukocytes and cardiomyocytes to find out the cross-talk between these cell types in vitro condition by mimicking the in vivo inflammatory milieu. Methods The study is divided into two parts, first part comprising patient study and the second part is the in vitro cell culture based mechanistic study. 50 IDCM patients and 30 age and sex matched control subjects were recruited from our in-house Ramakrishna Mission Seva Pratishthan (RKMSP) hospital based on their NYHA discomfort criteria followed by intra thoracic echocardiography. Exclusively the IDCM subjects were selected for this study. Blood was collected from the patients for biochemical and inflammatory parameters and to isolate PBMCs. mRNA and protein expressions were studied by real time PCR and immunoblotting respectively. H9C2 cardiomyocyte cells alone or with LPS-activated THP-1 monocyte/macrophage line were co-cultured in presence of the major inflammatory cytokines in the same in vivo concentration as found in the IDCM patients in presence or absence of ER stress agonist or antagonists. Immunofluorescence, RT-PCR and immunoblotting were done for markers specific to myocyte stress, inflammation and apoptosis. Results Inflammatory and cardiac blood biomarkers NLR, hs-CRP, IL-6, pro-BNP and troponin-I were found to be significantly high (P &amp;lt; 0.05) in IDCM. Subsequently, the mRNA expressions of pro-inflammatory genes TNF-α (2.6 fold), IFN-γ (10.2 fold), IL-1β (8.5 fold) and NF-κβ (2.427 fold) were all significantly increased (P &amp;lt; 0.05). Looking into the cellular stress pathways, we further found upregulations of antioxidant genes SOD (2.59 fold) and catalase (2.17 fold) and ER stress markers Grp78 (2.794 fold), CHOP (3.407 fold), IRE1-α (2.104 fold) and ATF-6 (40.618 fold) mRNA expression in the PBMCs. When we stimulated the H9C2 cardiomyocytes with IL-1B and TNF-α at comparable conc. as found in the IDCM patients, the ER stress and inflammation were observed. Further, when we co-cultured these H9C2 with LPS stimulated THP-1 macrophages an exaggerated inflammation and myocyte stress was observed that led to apoptosis of the cells. Interestingly using ER stress inhibitor TUDCA, remarkably blocked the apoptosis and promotes cell survival. Conclusion Our results showed that a systemic inflammation caused by stressed PBMCs leads to cardiac deterioration and dilation in the unknown DCM cases. So far we have established that an exaggerated inflammation induce the circulating PBMCs to migrate and infiltrate the cardiac musculature where they induce oxidative stress and ER stress to the cardiomyocytes leading to apoptosis of the later. In long run these unknown forms of dilated cardiomyopathy patients suffer from a dilated heart due to a decline in the myocyte population. Our findings may warrant for a treatment strategy that alleviate the intracellular stress of the circulating leukocytes and cardiomyocytes to minimize the inflammatory changes of the heart.

Role of Inhibiting Inflammation of LC3-Associated Phagocytosis in Dry Eye Disease

Purpose To confirm the expression and investigate the role of LC3-associated phagocytosis (LAP) in dry eye disease (DED). Methods The DED model of mice was established by scopolamine subcutaneous injection in a low-humidity environment chamber. Tear secretion test and corneal fluorescein sodium staining were used to evaluate the severity of DED. Expression levels of Rubicon, microtubule-associated protein light chain 3-II (LC3-II), Beclin-1 and autophagy-related gene-7 (Atg-7) in corneas of mice with DED were tested by western blot. Cell Counting Kit-8 (CCK-8) assay was used to detect the effects of different concentrations of hypertonic solutions on the proliferation activity of human corneal epithelial cells (HCECs). The expression levels of Dectin-1, IL-6 and IL-1β in HCECs after stimulation with different concentrations of hypertonic solutions were tested. The expressions of Rubicon, LC3-II, Beclin-1 and ATG-7 in HCECs were detected by reverse transcription polymerase chain reaction (RT-PCR). After being pretreated with 10 μM si-Rubicon, the severity of the disease was documented by corneal fluorescein sodium staining. And the expression levels of IL-6 and IL-1β were also tested by RT-PCR. Results Compared with the normal control group, the corneal fluorescein sodium staining scores and tear secretion were significantly reduced. Rubicon, LC3-II, Beclin-1 and ATG-7 were significantly elevated. CCK-8 showed that the 400 and 450 mOsM hypertonic solutions did not affect the proliferation activity of HCECs. The expression of Dectin-1, IL-1β and IL-6 were elevated after stimulation with 450 mOsM solution. LC3-II, Rubicon, ATG-7 and Beclin-1 increased after stimulation with 450 mOsM hyperosmolar solution in HCECs. Corneal fluorescein staining showed that si-Rubicon increased the severity of DED in mice. Moreover, the mRNA expressions of inflammatory factors IL-1β and IL-6 in the cornea of mice were significantly increased. Conclusion DED increased the expression of proteins associated with LAP. LAP could play an anti-inflammatory effect in DED.

GLUT1 Mediates the Metabolic Reprogramming and Inflammation of CCR2$^+$ Monocytes/Macrophages from Patients with DCM.

Macrophages expressing CC chemokine receptor 2 (CCR2) possess characteristics and performance akin to M1 polarized macrophages, which promote inflammation. Advanced heart failure (HF) patients with higher abundance of CCR2+ macrophages are more likely to experience adverse remodeling. The precise mechanism of CCR2+ macrophages in how they affect the progression of dilated cardiomyopathy remains unknown. Cardiac biopsy samples from dilated cardiomyopathy patients (DCM) were used for immunohistochemistry and immunofluorescence staining. PCR is employed to identify the IL-1β, IL-6, TNF-α, TGF-β, MMP2, MMP9, PKM1, PKM1, GLUT1, GLUT2, GLUT3, GLUT4, PDK1, PFKFB3, PFK1 and HK2 mRNA expression of CCR2+ monocytes/macrophages from the peripheral blood of DCM patients. Seahorse was used to evaluate the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of CCR2+ monocytes/macrophages. 2-DG was used to simulate a lack of glucose. Lentivirus containing GLUT1 inhibitory sequence was used to knockdown GLUT1 gene expression of CCR2+ monocytes/macrophages. Western Blot and immunofluorescence staining was used to evaluate the expression of NLRP3. Immunostaining results of cardiac biopsy tissue from dilated cardiomyopathy (DCM) patients demonstrated that the progression to HF was associated with an increase in the number of CCR2+ macrophages. PCR results demonstrated that CCR2 monocytes and macrophages derived from the blood of DCM patients expressed elevated levels of inflammatory factors and up regulation of glycolysis related genes. In addition, OCR and glucose uptake experiments confirmed that increased glucose uptake of these cells was associated with greater inflammation and correlated with a worsening of cardiac function. limiting the glucose supply to CCR2+ monocytes and macrophages, or suppressing the activity of glucose transporter 1 (GLUT1) could reduce inflammation levels. These results suggest that CCR2+ monocytes and macrophages rely on metabolic reprogramming to trigger inflammatory response and contribute to myocardial injury and the progression of DCM.

Crocus sativus L. Extract (Saffron) Effectively Reduces Arthritic and Inflammatory Parameters in Monotherapy and in Combination with Methotrexate in Adjuvant Arthritis.

Rheumatoid arthritis (RA), an autoimmune disease, is characterized by inflammation that affects not only the liver but also other organs and the musculoskeletal system. The standard therapy for RA is methotrexate (MTX), which has safety limitations. The extract from Crocus sativus L. (saffron-SF) is also known for its anti-inflammatory effects. Therefore, we decided to investigate the potential benefit of SF in monotherapy via two doses (SF1-25 mg/kg of b.w.; SF2-50 mg/kg of b.w.) and in combination with MTX (0.3 mg/kg of b.w., twice a week) using adjuvant arthritis in rats. To evaluate these therapeutic settings, we used biometric, immunological, and biochemical parameters, as well as the relative gene expression of the mRNA in the liver. Our results showed a statistically significant increase in the experimental animals' body weight and the arthritic score (AS) on day 14 for monotherapy with SF1 and SF2. The change of hind paw volume (CHPV) was significant only for SF2 monotherapy on the 14th day of the experiment. A combination of SF1 and SF2 with MTX significantly modulated all the biometric parameters during the experimental period. Additionally, AS and CHPV improved considerably compared to MTX monotherapy on day 21. Furthermore, all monotherapies and combination therapies were significant for the biochemical parameter γ-glutamyl transferase (GGT) in the joint. GGT activity in the spleen was less pronounced; only MTX in combination with SF1 significantly modified this parameter. The higher dose of SF monotherapy (SF2) was similarly significant with respect to immunological parameters, such as plasmatic IL-17A, IL-1β, and MMP-9 on day 21. The combination of both doses of SF with MTX significantly improved these immunological parameters, except for C-reactive protein (CRP), which was influenced only by the higher dose of SF2 in combination with MTX in plasma at the end of the experiment. A different effect was found for the relative expression of CD36 mRNA, where only SF1 significantly decreased gene expression in the liver. However, the relative gene mRNA expression of IL-1β in the liver was significantly reduced by the SF monotherapies and the combination of both SF doses with MTX. Our findings showed SF's partial antiarthritic and anti-inflammatory potential in monotherapy, but the effect was stronger in combination with MTX.

The Effects of an Osteoarthritic Joint Environment on ACL Damage and Degeneration: A Yucatan Miniature Pig Model.

Posttraumatic osteoarthritis (PTOA) arises secondary to joint injuries and is characteristically driven by inflammatory mediators. PTOA is often studied in the setting of ACL tears. However, a wide range of other injuries also lead to PTOA pathogenesis. The purpose of this study was to characterize the morphological changes in the uninjured ACL in a PTOA inflammatory environment. We retrospectively reviewed 14 ACLs from 13 Yucatan minipigs, 7 of which had undergone our modified intra-articular drilling (mIAD) procedure, which induced PTOA through inflammatory mediators. Seven ACLs were harvested from mIAD minipigs (PTOA) and seven ACLs from control minipigs with no cartilage degeneration (non-PTOA). ACL degeneration was evaluated using histological scoring systems. IL-1β, NF-κB, and TNF-α mRNA expression in the synovium was measured using qRT-PCR. PTOA minipigs demonstrated significant ACL degeneration, marked by a disorganized extracellular matrix, increased vascularity, and changes in cellular shape, density, and alignment. Furthermore, IL-1β, NF-κB, and TNF-α expression was elevated in the synovium of PTOA minipigs. These findings demonstrate the potential for ACL degeneration in a PTOA environment and emphasize the need for anti-inflammatory disease-modifying therapies following joint injury.

Glycyrrhizin inhibits LPS-induced inflammatory responses in goat ruminal epithelial cells in vitro

Inflammation plays a crucial role in the progression of Subacute Ruminal Acidosis (SARA). The experiment was designed to investigate anti-inflammatory effects of glycyrrhizin on goats ruminal epithelial cells (GREC) which were induced SARA by Lipopolysaccharide (LPS) in vitro. The GREC were induced SARA by adding LPS at the concentration of 5 μm and glycyrrhizin was added at different concentration of 0, 60, 90, 120, 150 μm. The structural integrity of LPS-induced GREC with the treatment of glycyrrhizin were observed by electron microscope; The levels of inflammatory factors TNF-α, IL-1β, IL-6, IL-8 and IL-12 were measured by ELISA; The number of Zo-1 and Occludin were measured, the expression of tight junction protein Occludin were measured by Western blot, and the mRNA expression of NF-κB, TNF-α, IL-1β, IL-6, IL-8 and IL-12 were measured in vitro. The results showed that higher concentration treatment of glycyrrhizin led to better morphology in LPS-induced GREC. Glycyrrhizin inhibited the growth of inflammatory factors TNF-α, IL-1β, IL-6, IL-8 and IL-12 in a dose-dependent manner. The number of ZO-1 and Occludin increased with the increase of adding of glycyrrhizin. Western blot analysis showed that the expression of tight junction protein Occludin in LPS-induced GREC increased with the adding of glycyrrhizin in a dose-dependent manner. Furthermore, the mRNA expression of NF-κB, TNF-α, IL-1β, IL-6, IL-8 and IL-12 decreased significantly with the increase treatment of glycyrrhizin. Glycyrrhizin significantly inhibits LPS-induced inflammatory mediators in GREC and the effects are better with the increase treatment of glycyrrhizin in vitro.

Design and Synthesis of l-1'-Homologated Adenosine Derivatives as Potential Anti-inflammatory Agents.

Inflammatory responses are fundamental protective warning mechanisms. However, in certain instances, they contribute significantly to the development of several chronic diseases such as cancer. Based on previous studies of truncated 1'-homologated adenosine derivatives, l-nucleosides and their nucleobase-modified quinolone analogues were designed, synthesized, and evaluated for anti-inflammatory activities. The target molecules were synthesized via the key intramolecular cyclization of monotosylate and Mitsunobu condensation from the natural product, d-ribose. All compounds tested and showed potent anti-inflammatory activities, as indicated by their inhibition of LPS-induced IL-1β secretion from the RAW 264.7 macrophages. Gene expressions of pro-inflammatory cytokines showed that all compounds, except 3a and 3b, significantly reduced LPS-induced IL-1β and IL-6 mRNA expressions. The half-maximal inhibitory concentrations (IC50) of 2g and 2h against IL-1β were 1.08 and 2.28 μM, respectively. In contrast, only 2d, 2g, and 3d effectively reversed LPS-induced TNFα mRNA expression. Our mechanistic study revealed that LPS-induced phosphorylation of NF-κB was significantly downregulated by all compounds tested, providing evidence that the NF-κB signaling pathway is involved in their anti-inflammatory activities. Among the compounds tested, 2g and 2h had the most potent anti-inflammatory effects, as shown by the extent of decrease in pro-inflammatory gene expression, protein secretion, and NF-κB phosphorylation. These findings suggest that the l-truncated 1'-homologated adenosine skeleton and its nucleobase-modified analogues have therapeutic potential as treatments for various human diseases by mediating inflammatory processes.

Erythritol Can Inhibit the Expression of Senescence Molecules in Mouse Gingival Tissues and Human Gingival Fibroblasts.

Oral aging causes conditions including periodontal disease. We investigated how the sugar alcohol erythritol, which has anti-caries effects, impacts aging periodontal tissues and gingival fibroblasts in mice and humans in vivo and in vitro. Mice were classified into three groups: control groups of six-week-old (YC) and eighteen-month-old mice (AC) and a group receiving 5% w/w erythritol water for 6 months (AE). After rearing, RNA was extracted from the gingiva, and the levels of aging-related molecules were measured using PCR. Immunostaining was performed for the aging markers p21, γH2AX, and NF-κB p65. p16, p21, γH2AX, IL-1β, and TNFα mRNA expression levels were higher in the gingiva of the AC group than in the YC group, while this enhanced expression was significantly suppressed in AE gingiva. NF-κB p65 expression was high in the AC group but was strongly suppressed in the AE group. We induced senescence in cultured human gingival fibroblasts using H2O2 and lipopolysaccharide before erythritol treatment, which reduced elevated senescence-related marker (p16, p21, SA-β-gal, IL-1β, and TNFα) expression levels. Knockdown of PFK or PGAM promoted p16 and p21 mRNA expression, but erythritol subsequently rescued pyruvate production. Overall, intraoral erythritol administration may prevent age-related oral mucosal diseases.

Interactive effect of dietary vitamin D3 and zinc (Zn) on growth performance, Zn metabolism, and intestinal health of yellow catfish Pelteobagrus fulvidraco

Both vitamin D3 (VD3) and zinc (Zn) are essential micronutrients to fish, and VD3 can regulate Zn uptake and metabolism in various animals. However, the interactive effect of VD3 and Zn on growth performance, Zn metabolism, and intestinal health remains largely unknown in fish. The study formulated four experimental diets to culture yellow catfish, i.e., basic VD3 addition without extra Zn addition (control), basic VD3 addition with extra Zn addition (Zn group), high VD3 addition without extra Zn addition (VD3 group), and high VD3 and Zn addition (Zn + VD3 group). Four experimental groups contain Zn and VD3 contents at 41.15 mg/kg + 1027 IU/kg (control), 94.77 mg/kg + 1033 IU/kg (Zn group), 41.04 mg/kg + 16,047 IU/kg (VD3 group), and 94.89 mg/kg + 16,109 IU/kg (Zn + VD3 group), respectively. The experiment continued for 10 weeks. Dietary Zn and VD3 interacted to affect specific growth rate (SGR), Zn content and mRNA expression of Zn metabolism (zip4, zip9, zip14, znt1, znt2, mtf-1, and mt), SUMOylation-relevant genes (sae1, ubc9, pias1, sumo1, senp2, and senp5), tight junction-relevant genes (claudin-4, zo1, zo2, zo3, and jam2a), antioxidant (nrf2 and cat) and inflammatory genes (il1b and il8). Compared to the control, high dietary Zn addition increased intestinal mRNA expression of zip4, znt1, znt2, mt, sae1, ubc9, pias1, sumo1, claudin4, zo1, zo2, and jam2a and il1b, and Zn contents, but decreased mRNA expression of zip9, senp2, senp5, nrf2 and cat. Compared to high Zn addition in the diets, Zn and VD3 co-addition increased SGR, Zn contents, and mRNA expression of zip4, znt1, znt2, mt, mtf-1, senp2, senp5, claudin4, zo1, zo2, zo3, jam2a, nrf2 and cat, but decreased mRNA expression of zip9, zip14, sae1, ubc9, pias1, sumo1 and il1b. Dietary Zn and VD3 co-addition interacted to influence Cu/Zn-SOD, CAT and GPx activities, T-AOC, MDA content. Compared to control, high dietary Zn addition decreased intestinal Cu/Zn-SOD, CAT, GPx activities and T-AOC but increased MDA content. Compared to extra Zn addition in the diets, Zn and VD3 co-addition increased Cu/Zn-SOD, CAT and GPx activities as well as T-AOC but decreased MDA content. Based on these observations, our study indicated that high dietary Zn addition influenced Zn metabolism and SUMOylation modification, damaged the tight junction, induced oxidative stress and inflammatory responses in the intestine, and dietary vitamin D3 addition promoted growth performance and alleviated high dietary Zn-induced negative effects on the intestine of yellow catfish. Significance of relevanceOur study elucidated the innovative insights into the interaction of Zn and VD3 in fish, which provided the important references for dietary Zn and VD3 addition in aquafeeds.

The probiotic roles of Lactiplantibacillus plantarum E2 as a dietary supplement in growth promotion and disease resistance of juvenile large yellow croaker (Larimichthys crocea)

Bacterial diseases are the most important limiting factors for the development of large yellow croaker aquaculture. Probiotics are considered to be a promising alternative approach for the control of bacterial diseases in aquaculture. However, the studies on probiotics used in farmed large yellow croakers were very limited. In this study, we isolated and identified a Lactiplantibacillus plantarum E2 from the intestinal tract of large yellow croaker. L. plantarum E2 showed significant antibacterial activities against several aquaculture pathogenic bacteria, intestinal environmental tolerance, and biosafety. After 7 weeks of feeding, the E2 supplementation of dietary significantly improved the growth and the survival rates of large yellow croakers after Pseudomonas plecoglossicida PQLYC4 challenge. Further analysis showed that E2 effectively improved the intestinal integrity, and increased the intestinal α-amylase, trypsin and lipase activities. Moreover, the E2 supplementation also significantly suppressed the mRNA expression of IL-10 and increased the mRNA expression of IL-1β, IL-12α, IL-17D, IFN-γ, and TNF-α-R. Gut microbiota analysis showed that E2 significantly affected gut microbial community composition by decreasing the relative abundance of Sphingomonas and increasing the relative abundance of Lactobacillus and Pseudomonas. Finally, E2 could improve resistance of large yellow croaker against P. plecoglossicida PQLYC4 infection. Therefore, our findings showed that L. plantarum E2 has potential application as a probiotic in large yellow croaker, which may provide a new strategy of preventing and controlling bacterial diseases in this species.

Systemic and local vascular inflammation and arterial reactive oxygen species generation in patients with advanced cardiovascular diseases.

Systemic inflammation may cause endothelial activation, mediate local inflammation, and accelerate progression of atherosclerosis. We examined whether the levels of circulating inflammatory cytokines reflect local vascular inflammation and oxidative stress in two types of human arteries. Human internal mammary artery (IMA) was obtained in 69 patients undergoing coronary artery bypass graft (CABG) surgery and left anterior descending (LAD) artery was obtained in 17 patients undergoing heart transplantation (HTx). Plasma levels of tumor necrosis factor α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) were measured using ELISA, high-sensitivity C-reactive protein (hs-CRP) was measured using Luminex, and mRNA expression of proinflammatory cytokines in the vascular tissues was assessed. Furthermore, formation of superoxide anion was measured in segments of IMA using 5 uM lucigenin-dependent chemiluminescence. Vascular reactivity was measured using tissue organ bath system. TNF-α, IL-6 and IL-1β mRNAs were expressed in all studied IMA and LAD segments. Plasma levels of inflammatory cytokines did not correlate with vascular cytokine mRNA expression neither in IMA nor in LAD. Plasma TNF-α and IL-6 correlated with hs-CRP level in CABG group. Hs-CRP also correlated with TNF-α in HTx group. Neither vascular TNF-α, IL-6 and IL-1β mRNA expression, nor systemic levels of either TNF-α, IL-6 and IL-1β were correlated with superoxide generation in IMAs. Interestingly, circulating IL-1β negatively correlated with maximal relaxation of the internal mammary artery (r = -0.37, p = 0.004). At the same time the mRNA expression of studied inflammatory cytokines were positively associated with each other in both IMA and LAD. The positive correlations were observed between circulating levels of IL-6 and TNF-α in CABG cohort and IL-6 and IL-1β in HTx cohort. This study shows that peripheral inflammatory cytokine measurements may not reflect local vascular inflammation or oxidative stress in patients with advanced cardiovascular disease (CVD). Circulating pro-inflammatory cytokines generally correlated positively with each other, similarly their mRNA correlated in the arterial wall, however, these levels were not correlated between the studied compartments.