Downregulation Of IL-6 Research Articles

Prophylactic Administration of Gut Microbiome Metabolites Abrogated Microglial Activation and Subsequent Neuroinflammation in an Experimental Model of Japanese Encephalitis.

Short-chain fatty acids (SCFAs) are gut microbial metabolic derivatives produced during the fermentation of ingested complex carbohydrates. SCFAs have been widely regarded to have a potent anti-inflammatory and neuro-protective role and have implications in several disease conditions, such as, inflammatory bowel disease, type-2 diabetes, and neurodegenerative disorders. Japanese encephalitis virus (JEV), a neurotropic flavivirus, is associated with life threatening neuro-inflammation and neurological sequelae in infected hosts. In this study, we hypothesize that SCFAs have potential in mitigating JEV pathogenesis. Postnatal day 10 BALB/c mice were intraperitoneally injected with either a SCFA mixture (acetate, propionate, and butyrate) or PBS for a period of 7 days, followed by JEV infection. All mice were observed for onset and progression of symptoms. The brain tissue was collected upon reaching terminal illness for further analysis. SCFA-supplemented JEV-infected mice (SCFA + JEV) showed a delayed onset of symptoms, lower hindlimb clasping score, and decreased weight loss and increased survival by 3 days (p < 0.0001) upon infection as opposed to the PBS-treated JEV-infected animals (JEV). Significant downregulation of inflammatory cytokines TNF-α, MCP-1, IL-6, and IFN-Υ in the SCFA + JEV group relative to the JEV-infected control group was observed. Inflammatory mediators, phospho-NF-kB (P-NF-kB) and iba1, showed 2.08 ± 0.1 and 3.132 ± 0.43-fold upregulation in JEV versus 1.19 ± 0.11 and 1.31 ± 0.11-fold in the SCFA + JEV group, respectively. Tissue section analysis exhibited reduced glial activation (JEV group─42 ± 2.15 microglia/ROI; SCFA + JEV group─27.07 ± 1.8 microglia/ROI) in animals that received SCFA supplementation prior to infection as seen from the astrocytic and microglial morphometric analysis. Caspase-3 immunoblotting showed 4.08 ± 1.3-fold upregulation in JEV as compared to 1.03 ± 0.14-fold in the SCFA + JEV group and TUNEL assay showed a reduced cellular death post-JEV infection (JEV-6.4 ± 1.5 cells/ROI and SCFA + JEV-3.7 ± 0.73 cells/ROI). Our study critically contributes to the increasing evidence in support of SCFAs as an anti-inflammatory and neuro-protective agent, we further expand its scope as a potential supplementary intervention in JEV-mediated neuroinflammation.

Anti-Inflammatory Efficacy of Resveratrol-Enriched Rice Callus Extract on Lipopolysaccharide-Stimulated RAW264.7 Macrophages

Resveratrol and its derivative piceid exhibit a wide spectrum of health-promoting bioactivities. A resveratrol-enriched variety of Dongjin rice (DJ526) has been developed by transfection of a resveratrol biosynthesis gene, and increased resveratrol content has been confirmed in seeds following germination. In the current study, these resveratrol-enriched seeds were induced to produce callus, and callus extracts were evaluated for in vitro anti-inflammatory activity. Callus cultures contained greater amounts of resveratrol and piceid than DJ526 seeds, and treatment with DJ526 callus extract significantly reduced the lipopolysaccharide (LPS)-induced production of proinflammatory mediators nitric oxide and prostaglandin E2 by RAW264.7 macrophages. The inflammation-related nuclear factor kappa B and mitogen-activated protein kinase pathways were also inhibited in DJ526 callus extract-treated RAW264.7 cells, resulting in downregulation of proinflammatory factor genes COX-2, iNOS, IL-1β, IL-6, and TNF-α. Expression of the LPS-binding toll-like receptor-4 was also markedly reduced in DJ526 callus extract-treated cells compared to DJ callus extract-treated cells. These findings demonstrate increased resveratrol and piceid content by callus culture of DJ526 rice seeds and the potent anti-inflammatory activity of resveratrol-enriched callus extract.

Chitosan-Loaded Lagenaria siceraria and Thymus vulgaris Potentiate Antibacterial, Antioxidant, and Immunomodulatory Activities against Extensive Drug-Resistant Pseudomonas aeruginosa and Vancomycin-Resistant Staphylococcus aureus: In Vitro and In Vivo Approaches.

Antimicrobial resistance poses considerable issues for current clinical care, so the modified use of antimicrobial agents and public health initiatives, coupled with new antimicrobial approaches, may help to minimize the impact of multidrug-resistant (MDR) bacteria in the future. This study aimed to evaluate the antimicrobial, antioxidant, and immunomodulatory activities of Lagenaria siceraria, Thymus vulgaris, and their chitosan nanocomposites against extensive drug-resistant (XDR) Pseudomonas aeruginosa and vancomycin-resistant Staphylococcus aureus (VRSA) using both in vitro and in vivo assays. The in vitro antimicrobial susceptibilities of P. aeruginosa and VRSA strains revealed 100% sensitivity to imipenem (100%). All P. aeruginosa strains were resistant to cefoxitin, cefepime, trimethoprim + sulfamethoxazole, and fosfomycin. However, S. aureus strains showed a full resistance to cefoxitin, amoxicillin, ampicillin, erythromycin, chloramphenicol, and fosfomycin (100% each). Interestingly, all S. aureus strains were vancomycin-resistant (MIC = 32-512 μg/mL), and 90% of P. aeruginosa and S. aureus strains were XDR. The antimicrobial potential of Lagenaria siceraria and Thymus vulgaris nanocomposites with chitosan nanoparticles demonstrated marked inhibitory activities against XDR P. aeruginosa and VRSA strains with inhibition zones' diameters up to 50 mm and MIC values ranging from 0.125 to 1 μg/mL and 1 to 8 μg/mL, respectively. The results of the in vivo approach in male Sprague Dawley rats revealed that infection with P. aeruginosa and S. aureus displayed significant changes in biochemical, hematological, and histopathological findings compared to the negative control group. These values returned to the normal range after treatment by chitosan nanoparticles, either loaded with Lagenaria siceraria or Thymus vulgaris. Real-time quantitative polymerase chain reaction (RT-qPCR) findings presented significant upregulation of the relative expression of the IL10 gene and downregulation of the IFNG gene throughout the experimental period, especially after treatment with chitosan nanoparticles loaded either with Lagenaria siceraria or Thymus vulgaris in comparison to the positive control groups. In conclusion, this is the first report suggesting the use of Lagenaria siceraria and Thymus vulgaris nanocomposites with chitosan nanoparticles as a promising contender for combating XDR P. aeruginosa and VRSA infections as well as a manager for inflammatory situations and oxidative stress-related disorders.

Myocardial Expression of Pluripotency, Longevity, and Proinflammatory Genes in the Context of Hypercholesterolemia and Statin Treatment.

Background: This study sought to assess the effect of statin therapy on myocardial inflammation in a White New Zealand rabbit model of atherogenesis. Methods: The mRNA expression levels of pro-inflammatory, pluripotency, and aging-related markers were quantified following a controlled feeding protocol and statin treatments. Results: Following high-cholesterol diet induction, we observed significant upregulation in the myocardial mRNA levels of MYD88, NF-κB, chemokines (CCL4, CCL20, and CCR2), IFN-γ, interleukins (IL-1β, IL-2, IL-4, IL-8, IL-10, and IL-18), and novel markers (klotho, KFL4, NANOG, and HIF1α). In contrast, HOXA5 expression was diminished following a hyperlipidemic diet. Both statin treatments significantly influenced the markers studied. Nevertheless, rosuvastatin administration resulted in a greater reduction in MYD88, NF-kB, chemokines (CCL4, CCL20, and CCR2), and interleukins IL-1β, IL-8, KLF4, NANOG, and HIF1α than fluvastatin. Fluvastatin, on the other hand, led to a stronger decrease in IL-4. Downregulation of IL-2 and IL-18 and upregulation of IFNβ and HOXA5 were comparable between the two statins. Notably, rosuvastatin had a stronger effect on the upregulation of klotho and IL-10. Conclusion: Overall, statin therapy significantly attenuated inflammatory, pluripotency, and klotho expression in myocardial tissue under atherogenic conditions. Our findings also highlight the differential efficacy of rosuvastatin over fluvastatin in curtailing proatherogenic inflammation, which could have profound implications for the clinical management of cardiovascular disease.

Green Synthesis of Zinc Oxide Nanoparticles from Althaea officinalis Flower Extract Coated with Chitosan for Potential Healing Effects on Diabetic Wounds by Inhibiting TNF-α and IL-6/IL-1β Signaling Pathways.

Diabetes Mellitus is a multisystem chronic pandemic, wound inflammation, and healing are still major issues for diabetic patients who may suffer from ulcers, gangrene, and other wounds from uncontrolled chronic hyperglycemia. Marshmallows or Althaea officinalis (A.O.) contain bioactive compounds such as flavonoids and phenolics that support wound healing via antioxidant, anti-inflammatory, and antibacterial properties. Our study aimed to develop a combination of eco-friendly formulations of green synthesis of ZnO-NPs by Althaea officinalis extract and further incorporate them into 2% chitosan (CS) gel. First, develop eco-friendly green Zinc Oxide Nanoparticles (ZnO-NPs) and incorporate them into a 2% chitosan (CS) gel. In-vitro study performed by UV-visible spectrum analysis showed a sharp peak at 390 nm, and Energy-dispersive X-ray (EDX) spectrometry showed a peak of zinc and oxygen. Besides, Fourier transforms infrared (FTIR) was used to qualitatively validate biosynthesized ZnO-NPs, and transmission electron microscope (TEM) showed spherical nanoparticles with mean sizes of 76 nm and Zeta potential +30mV. The antibacterial potential of A.O.-ZnO-NPs-Cs was examined by the diffusion agar method against Gram-positive (Staphylococcus aureus and Bacillus subtilis) and Gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa). Based on the zone of inhibition and minimal inhibitory indices (MIC). In addition, an in-silico study investigated the binding affinity of A.O. major components to the expected biological targets that may aid wound healing. Althaea Officinalis, A.O-ZnO-NPs group showed reduced downregulation of IL-6, IL-1β, and TNF-α and increased IL-10 levels compared to the control group signaling pathway expression levels confirming the improved anti-inflammatory effect of the self-assembly method. In-vivo study and histopathological analysis revealed the superiority of the nanoparticles in reducing signs of inflammation and wound incision in rat models. These biocompatible green zinc oxide nanoparticles, by using Althaea Officinalis chitosan gel ensure an excellent new therapeutic approach for quickening diabetic wound healing.

JianPiYiShen formula prevents cisplatin-induced acute kidney injury in mice by improving necroptosis through MAPK pathway

BackgroundAcute kidney injury (AKI), characterized by necroptosis and activation of MAPK pathway, causes sudden declines in renal function. To date, efficacious treatments are lacking. JianPiYiShen Formula (JPYSF) has a protective effect on the kidneys. The aim of this study is to explore the mechanism of JPYSF in cisplatin-induced AKI.MethodsMale C57/BL6J mice were divided into control group, cisplatin group and cisplatin + JPYSF group. Before establishing the model, the cisplatin + JPYSF group was administered JPYSF (18.35 g/kg/day) by gavage for 5 consecutive days. A single intraperitoneal injection of cisplatin (20 mg/kg) was used to establish AKI model. Measurement of renal function and H&E staining were performed to assess renal damage. WB, PCR, TUNEL staining and immunohistochemistry were used to detect related indicators of mitochondrial function, oxidative stress, necroptosis, inflammation and MAPK pathway. And one-way analysis of variance was used to compare group differences.ResultsCompared with the cisplatin group, JPYSF can attenuate AKI, reflected by the decrease in Scr and BUN levels, the improvement of renal tubular injury, and the downregulation of NGAL and KIM1. Cisplatin can induce mitochondrial dysfunction and oxidative stress, triggering necroptosis. In this study, JPYSF improved mitochondrial dysfunction to enhance oxidative stress, as manifested by upregulation of OPA1, PGC-1α, SOD and CAT, and downregulation of DRP1 and MFF. Then JPYSF showed a significant protective effect in necroptosis, as embodied by reduced number of TUNEL-positive cells, decreased the gene expression of RIPK3 and MLKL, as well as downregulation the proteins expression of P-RIPK1, P-RIPK3, and P-MLKL. Moreover, necroptosis can aggravate inflammation. JPYSF ameliorated inflammation by improving inflammatory and anti-inflammatory indexes, including downregulation of TNF-α, IL-6, MCP-1 and LY6G, and upregulation of IL-10. In addition, JPYSF also inhibited MAPK pathway to improve necroptosis by decreasing the expression of P-JNK and P-ERK.ConclusionOur data showed that JPYSF prevents cisplatin-induced AKI by improving necroptosis through MAPK pathway, which is related to the improvement of mitochondrial dysfunction, oxidative stress, and inflammation.

Magnesium-lithium thin films for neurological applications–An in vitro investigation of glial cytocompatibility and neuroinflammatory response

Lithium (Li), a widely used drug for bipolar disorder management, is associated with many side effects due to systemic exposure. The localized delivery of lithium through implants could be an approach to overcome this challenge, for which biodegradable magnesium (Mg)-based materials are a promising choice. In this study, we focus on Mg-Li thin film alloys as potential Li-releasing implants. Therefore, we investigated the in vitro short-term corrosion behavior and cytocompatibility of two alloys, Mg-1.6wt%Li and Mg-9.5wt%Li. As glial cells are the key players of foreign body responses to implants, we used human glial cell lines for cytocompatibility studies, and a murine brain slice model for a more holistic view at the neuroinflammatory response. We found that Mg-1.6wt%Li corrodes approximately six times slower than Mg-9.5wt%Li. Microscopic analysis showed that the material surface (Mg-1.6wt%Li) is suitable for cell adhesion. The cytocompatibility test with Mg-1.6wt%Li and Mg-9.5wt%Li alloy extracts revealed that both cell types proliferated well up to 10 mM Mg concentration, irrespective of the Li concentration. In the murine brain slice model, Mg-1.6wt%Li and Mg-9.5wt%Li alloy extracts did not provoke a significant upregulation of glial inflammatory/ reactivity markers (IL-1β, IL-6, FN1, TNC) after 24 h of exposure. Furthermore, the gene expression of IL-1β (up to 3-fold) and IL-6 (up to 16-fold) were significantly downregulated after 96 h, and IL-6 downregulation showed a Li concentration dependency. Together, these results indicate the acute cytocompatibility of two Mg-Li thin film alloys and provide basis for future studies to explore promising applications of the material. Statement of significanceWe propose the idea of lithium delivery to the brain via biodegradable implants to reduce systemic side effects of lithium for bipolar disorder therapy and other neurological applications. This is the first in vitro study investigating Mg-xLi thin film degradation under physiological conditions and its influence on cellular responses such as proliferation, viability, morphology and inflammation. Utilizing human brain-derived cell lines, we showed that the material surface of such a thin film alloy is suitable for normal cell attachment. Using murine brain slices, which comprise a multicellular network, we demonstrated that the material extracts did not elicit a pro-inflammatory response. These results substantiate that degradable Mg-Li materials are biocompatible and support the further investigation of their potential as neurological implants.

Role of Probiotics against Human Cancers, Inflammatory Diseases, and Other Complex Malignancies

Probiotics have growing medical importance as a result of their potential in the prevention and therapeutic support of several complex diseases, including different types of cancers. The anticarcinogenic properties of probiotics are attributed to various mechanisms, including alterations in the composition of the intestinal microbiota, suppression of cell proliferation, stimulation of apoptosis, inhibition of NF-kB, reduction in levels of H2AX, 8-hydroxy-deoxyguanosine, RIG-I, downregulation of IL-17, and TNF signaling pathway. Furthermore, probiotics have demonstrated significant advantages in the prevention and management of other complex diseases, including diabetes, obesity, and cardiovascular diseases. Probiotics had a considerable effect in reducing inflammatory infiltration and the occurrence of precancerous lesions. Additionally, the administration of probiotics led to a decrease in the appearance level of genes related to proinflammatory pathways, including NF-κB, IL-17, and TNF signaling pathways. However, further research studies are required to comprehend the processes via which probiotics exert their effects and to authenticate their potential as alternative therapeutic interventions.

In silico, anti-inflammatory and acute toxicological evaluation of an indigenous medicinal plant Pterospermum rubiginosum using Sprague-Dawley rats

BackgroundPterospermum rubiginosum has been traditionally used by the tribal inhabitants of Southern India for treating bone fractures and as a local anti-inflammatory agent; however, experimental evidence to support this traditional usage is lacking. The present study aimed to investigate the phytochemical characterization, in silico and in vitro anti-inflammatory evaluation, followed by in vivo toxicological screening of P. rubiginosum methanolic bark extract (PRME).ResultsThe LCMS evaluation revealed the presence of 80 significant peaks; nearly 50 molecules were identified using the LCMS database. In silico analysis showed notable interactions with inducible nitric oxide synthase (iNOS) and interleukin-6 (IL-6). In vitro gene expression study supported the docking results with significant down-regulation of iNOS, IL-6, and IL-10. PRME was administered orally to the SD rats and was found to be non-toxic up to 1000 mg/kg body weight for 14 days. The antioxidant enzymes catalase and sodium dismutase exhibited an increased value in PRME-administered groups, possibly due to the diverse phytochemical combinations in bark extract.ConclusionsPRME administration significantly downregulated the gene expression of inflammatory markers, such as iNOS, IL-6, and IL-10. The molecular docking analysis of iNOS and IL-6 supports the in vitro study. In vivo toxicological study of PRME in SD rats was found to be non-toxic up to a concentration of 1000 mg/kg body weight for 14 days.Graphical abstract

Lactiplantibacillus plantarum postbiotic protects against Salmonella infection in broilers via modulating NLRP3 inflammasome and gut microbiota

Salmonella infection is a major concern in poultry production which poses potential risks to food safety. Our previous study confirmed that Lactiplantibacillus plantarum (LP) postbiotic exhibited a strong antibacterial capacity on Salmonella in vitro. This study aimed to investigate the beneficial effects and underlying mechanism of LP postbiotic on Salmonella-challenged broilers. A total of 240 one-day-old male yellow-feathered broilers were pretreated with 0.8% deMan Rogosa Sharpe (MRS) medium or 0.8% LP postbiotic (LP cell-free culture supernatant, LPC) in drinking water for 28 d, and then challenged with 1×109 CFU Salmonella enterica serovar Enteritidis (SE). Birds were sacrificed 3 d postinfection. Results showed that LPC maintained the growth performance by increasing body weight (BW), average daily gain (ADG), and average daily feed intake (ADFI) in broilers under SE challenge. LPC significantly attenuated SE-induced intestinal mucosal damage. Specifically, it decreased the intestinal injury score, increased villus length and villus/crypt, regulated the expression of intestinal injury-related genes (Villin, matrix metallopeptidase 3 [MMP3], intestinal fatty acid-binding protein [I-FABP]), and enhanced tight junctions (zona occludens-1 [ZO-1] and Claudin-1). SE infection caused a dramatic inflammatory response, as indicated by the up-regulated concentrations of interleukin (IL)-1β, IL-6, TNF-α, and the downregulation of IL-10, while LPC pretreatment markedly reversed this trend. We then found that LPC inhibited the activation of NOD-like receptor thermal protein domain associated protein 3 (NLRP3) inflammasome by decreasing the gene expression of Caspase-1, IL-lβ, and IL-18. Furthermore, LPC suppressed NLRP3 inflammasome activation by inhibiting nuclear factor-kappa B (NF-κB) signaling pathway (the reduced levels of toll-like receptor 4 [TLR4], myeloid differentiation factor 88 [MyD88], and NF-κB). Finally, our results showed that LPC regulated gut microbiota by enhancing the percentage of Ligilactobacillus and decreasing Alistipes and Barnesiella. In summary, we found that LP postbiotic was effective to protect broilers against Salmonella infection, possibly through suppressing NLRP3 inflammasome and optimizing gut microbiota. Our study provides the potential of postbiotics on prevention of Salmonella infection in poultry.

Abstract A039: Silencing MICAL2 expression in pancreatic cancer cells reduces IL-1A expression and inhibits tumor growth through a CD8+ T cell dependent mechanism

Abstract Introduction: Pancreatic cancer is characterized by desmoplastic, fibroinflammatory stroma. The crosstalk between cancer cells and their surrounding tumor microenvironment (TME) promotes disease progression, metastasis, and chemoresistance. We identified MICAL2 as a super-enhancer-associated gene in pancreatic cancer. MICAL2 (molecule interacting with Cas-L) proteins are flavin monooxygenases that promote actin depolymerization and indirectly regulate SRF transcription. In other work, we have found that MICAL2 promotes pancreatic cancer growth and progression. This study evaluated how MICAL2 pancreatic cancer cell expression modulates the PDAC tumor microenvironment. Methods: RNA-Seq analysis performed on human pancreas cancer cells (AsPC-1) before and after MICAL2 KD revealed differential regulation of IL-1α. We validated the expression of IL-1α in multiple human and mouse pancreas cancer cell lines by q-PCR. We next exposed human (h) and mouse (m) PSCs to conditioned media (CM) from cancer cells before and after MICAL2 KD and checked the expression of a-SMA, fibronectin, and Col1A and IL-6 genes by qPCR. KrasG12D/+; Trp53R172H/+; Pdx1-cre (KPC) cells with and without MICAL2 were orthotopically injected to assess the in vivo tumor growth and metastasis. Sc-RNA seq and Immunophenotyping were performed on Shcontrol (ShCNT) and MICAL2 (M2KD) KD tumors. CD8+ T cell depletion was done using an antibody-mediated approach. The adoptive transfer of CD8+ T cells extracted from MICAL2 KD tumors was done by tail vein injection. Results: MICAL2 KD resulted in the downregulation of the IL-1α gene in both human and mouse pancreas cancer cell lines (50% reduction, p&amp;lt;0.05). Human and mouse pancreatic stellate cells (hPSCs and mPSCs) co-cultured with cancer cells CM showed significant downregulation of a-SMA, fibronectin, Col1A, and IL-6 upon MICAL2 KD as compared to hPSCs and mPSCs co-cultured with ShCNT cells. Orthotopic injections of KPC MICAL2 KD cells led to decreased tumor growth as compared to ShCNT (0.26 gm vs. 1 gm, p = 0.008). Immunofluorescence (IF) revealed less α-SMA, PDGF-α, and IL-6 expression and reduced deposition of collagen and fibronectin in MICAL2 KD tumors. Single-cell RNA seq and flow cytometry studies of tumors harvested from both groups showed significantly increased infiltration of activated cytotoxic CD8+ T cells in MICAL2 KD tumors which was further confirmed by IF. Antibody-mediated depletion of CD8+ T cells in MICAL2 KD tumors abrogated their reduced tumor growth in both immune-hot and immune-cold KPC cell lines. Adoptive transfer of CD8+ T cells extracted from M2KD tumors significantly reduced ShCNT flank tumor growth. Conclusion: These data reveal that MICAL2 expression mediates tumor-stromal crosstalk through IL1A and creates an immunosuppressive environment in PDAC and suggests MICAL2 may be an attractive immunomodulatory target for pancreatic cancer therapy. Ongoing work is focused on dissecting the role of MICAL2 in priming the metastatic niche through the remodeling of the TME. Citation Format: Bharti Garg, EvanGeline Mose, Edgar Esparza, Jay Patel, Sarah Sass, Alexei Martsinkovskiy, Dawn Jacquish, Herve Tiriac, Andrew M. Lowy. Silencing MICAL2 expression in pancreatic cancer cells reduces IL-1A expression and inhibits tumor growth through a CD8+ T cell dependent mechanism [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A039.

Free heme induces neuroinflammation and cognitive impairment by microglial activation via the TLR4/MyD88/NF-κB signaling pathway

BackgroundRed blood cells (RBCs) transfusion is related to perioperative neurocognitive disorders. The toxic effect of free heme has been identified in many pathologies. However, the underlying mechanisms of RBCs transfusion or free heme in cognitive impairment have not been clearly explored. Therefore, this research was conducted to determine the mechanism of free heme-induced neuroinflammation and cognitive impairment.MethodsRats were received intraperitoneal injection of hemin alone or combined with intracerebroventricular injection of Hemopexin (HPX), and MWM test was conducted to measure cognitive function. The amount of heme-HPX complexes was evaluated by flow cytometry for CD91 + cells. The microglial inflammatory response in rat brain was observed by immunofluorescence staining of Iba-1, and the inflammatory factors of TNF-α, IL-1β and IL-6 in rat brain and BV2 cells were detected by ELISA analysis. Furthermore, neuronal apoptosis in HT22 cells alone and in HT22 + BV2 coculture system was detected by flow cytometry and immunofluorescence staining. Finally, western blot was conducted to detect TLR4/MyD88/NF-κB proteins in rat brain and BV2 cells treated with hemin or combined with pathway inhibitors. Additionally, the M1 surface marker CD86 was observed in BV2 cells to further confirm neuroinflammation.ResultsIntraperitoneal injection of hemin induced cognitive impairment, increase of CD91 + cells, up-regulation of TNF-α and IL-1β, down-regulation of IL-6, activation of microglia, and activation of the TLR4/MyD88/NF-κB signaling pathway in rat brain. Significantly, intracerebroventricular injection of HPX reduced the above effects. Hemin induced boost of TNF-α, IL-1β and IL-6 in BV2 cells, as well as apoptosis in HT22 cells. Notably, when HT22 cells were cocultured with BV2 cells, apoptosis was significantly increased. Hemin also induced activation of the TLR4/MyD88/NF-κB signaling pathway and increased the M1 surface marker CD86 in BV2 cells, and inhibiting this pathway reduced the inflammatory responses.ConclusionsFree heme induces cognitive impairment, and the underlying mechanism may involve neuronal apoptosis and microglial inflammation via the TLR4/MyD88/NF-κB signaling pathway. HPX may have potential therapeutic effects.4ne3ar5vcagQ2fAD43rWMLVideo

Resveratrol promotes the differentiation of human umbilical cord mesenchymal stem cells into esophageal fibroblasts via AKT signaling pathway.

Objectives: Resveratrol has been implicated in the differentiation and development of human umbilical cord mesenchymal stem cells. The differentiation of into esophageal fibroblasts is a promising strategy for esophageal tissue engineering. However, the pharmacological effect and underlying mechanism of resveratrol on human umbilical cord mesenchymal stem cells differentiation are unknown. Here, we investigated the effects and mechanism of resveratrol on the differentiation of human umbilical cord mesenchymal stem cells. Methods: Using a transwell-membrane coculture system to culture human umbilical cord mesenchymal stem cells and esophageal fibroblasts, we examined how resveratrol act on the differentiation of human umbilical cord mesenchymal stem cells. Immunocytochemistry, Sirius red staining, quantitative real-time PCR, and Western blotting were performed to examine collagen synthesis and possible signaling pathways in human umbilical cord mesenchymal stem cells. Results: We found that resveratrol promoted collagen synthesis and AKT phosphorylation. However, co-treatment of cells with resveratrol and the PI3K inhibitor LY294002 inhibited collagen synthesis and AKT phosphorylation. We demonstrated that resveratrol down-regulated the expression of IL-6, TGF-β, caspase-9, and Bax by activating the AKT pathway in human umbilical cord mesenchymal stem cell. Furthermore, resveratrol inhibited phosphorylated NF-ĸB in human umbilical cord mesenchymal stem cells. Conclusion: Our data suggest that resveratrol promotes the differentiation of human umbilical cord mesenchymal stem cells into fibroblasts. The underlying mechanism is associated with the downregulation of IL-6 and TGF-β via the AKT pathway and by inhibiting the NF-ĸB pathway. Resveratrol may be useful for esophageal tissue engineering.

Anoikis regulator GLI2 promotes NC cell immunity escape by TGF-β-mediated non-classic hedgehog signaling in colorectal cancer: based on artificial intelligence and big data analysis.

Anoikis is a speed-limited procedure to inhibit tumor metastasis during epithelial-mesenchymal transition (EMT). Previous studies have explored anoikis-related genes (ARG) in predicting prognosis and distinguishing tumoral immunity in many types of cancer. However, the role of ARGs in regulating NK cell exhaustion (NKE) and in predicting chemotherapy sensitivity is not clear. Therefore, it is necessary to work on it. Gene expression profiles and clinical features are collected from TCGA and GEO, and data analysis is performed in R4.2.0. The ARGs-based no-supervised learning algorithm identifies three ARG subgroups, amongst which the prognosis is different. WCGNA and Artificial intelligence (AI) are applied to construct an NKE-related drug sensitivity stratification and prognosis identification model in digestive system cancer. Pathways association analysis screens out GLI2 is a key gene in regulating NKE by non-classic Hedgehog signaling (GLI2/TGF-β/IL6). In vitro experiments show that down-regulation of GLI2 enhances the CAPE-mediated cell toxicity and accompanies with down-regulation of PD-L1, tumor-derive IL6, and snial1 whereas the expression of cleaved caspas3, cleaved caspase4, cleaved PARP, and E-cadherin are up-regulated in colorectal cancer. Co-culture experiments show that GLI2- decreased colorectal tumor cells lead to down-regulation of TIM-3 and PD1 in NK cells, which are restored by TGF-bate active protein powder. Besides, the Elisa assay shows that GLI2-decreased colorectal tumor cells lead to up-regulation of IFN-gamma in NK cells.

Amyloidogenic and Neuroinflammatory Molecular Pathways Are Contrasted Using Menaquinone 4 (MK4) and Reduced Menaquinone 7 (MK7R) in Association with Increased DNA Methylation in SK-N-BE Neuroblastoma Cell Line.

Besides its role in coagulation, vitamin K seems to be involved in various other mechanisms, including inflammation and age-related diseases, also at the level of gene expression. This work examined the roles of two vitamin K2 (menaquinones) vitamers, namely, menaquinone-4 (MK4) and reduced menaquinone-7 (MK7R), as gene modulator compounds, as well as their potential role in the epigenetic regulation of genes involved in amyloidogenesis and neuroinflammation. The SK-N-BE human neuroblastoma cells provided a "first-line" model for screening the neuroinflammatory and neurodegenerative molecular pathways. MK7R, being a new vitamin K form, was first tested in terms of solubilization, uptake and cell viability, together with MK4 as an endogenous control. We assessed the expression of key factors in amyloidogenesis and neuroinflammation, observing that the MK7R treatment was associated with the downregulation of neurodegeneration- (PSEN1 and BACE1) and neuroinflammation- (IL-1β and IL-6) associated genes, whereas genes retaining protective roles toward amiloidogenesis were upregulated (ADAM10 and ADAM17). By profiling the DNA methylation patterns of genes known to be epigenetically regulated, we observed a correlation between hypermethylation and the downregulation of PSEN1, IL-1β and IL-6. These results suggest a possible role of MK7R in the treatment of cognitive impairment, giving a possible base for further preclinical experiments in animal models of neurodegenerative disease.

Covalent conjugation with polyphenol reduced the sensitization of walnut and ameliorated allergy by enhancing intestinal epithelial barrier in mice

In order to reduce the sensitization of walnut protein (WP), the effects of the interaction between WP and (−)-Epigallocatechin gallate (EGCG), quercetin, trans-ferulic acid, and resveratrol were investigated. Covalent and non-covalent conjugations were compared. The results suggested that covalent conjugation reduced the free amino acid content, sulfhydryl content, and surface hydrophobicity. When compared to non-covalent conjugation, covalent modification showed a lower IgE binding capacity, accompanied by changes in protein conformation. Moreover, animal experiments revealed that there were up-regulation of transforming growth factor-β, T-box expressed in t cells, and forkhead transcription factor Foxp3 mRNA expression, and down-regulation of IL-4, IL-17, GATA binding protein 3 and retinoid-related orphan nuclear receptor γt mRNA expression in the conjugate groups. These results suggested that covalent conjugation of polyphenols, especially EGCG, likely ameliorated allergy by promoting Th1/Th2 and Treg/Th17 balance and alleviating allergy-induced intestinal barrier damage, which might be a support in reducing the allergenicity of WP.

Longitudinal dynamics of gut microbiota in the pathogenesis of acute graft-versus-host disease.

The gut microbiota has been reported to be associated with acute graft-versus-host disease (aGvHD) in hematopoietic stem cell transplantation (HSCT). Dynamic surveillance of the microbiota is required to understand the detailed pathogenesis involved in the process of aGvHD. Fecal samples were collected prospectively at four timepoints, including pre-HSCT (T1), graft infusion (T2), neutrophil engraftment (T3), and 30 days after transplantation (T4). Fecal samples were profiled by 16S ribosomal RNA gene sequencing to assess the microbiota composition. From the T1 to T4 timepoint, the diversity of the gut microbiota decreased, and the dominant species also changed, with a decrease in the obligate anaerobic bacteria and a shift toward a "pathogenic community". Compared with non-aGvHD patients, aGvHD patients had a lower abundance of Roseburia at T1 and a higher abundance of Acinetobacter johnsonii at T2. Furthermore, Acinetobacter johnsonii was negatively correlated with the secretion of IL-4 and TNF-α. At T3, Rothia mucilaginos was demonstrated to be linked with a decreased risk of aGvHD, which was accompanied by decreased secretion of IL-8. At T4, higher abundances of Lactobacillus paracasei and Acinetobacter johnsonii were identified to be related with aGvHD. Lactobacillus paracasei was associated with the downregulation of IL-10, and Acinetobacter johnsonii was associated with the downregulation of IL-2 and TNF-α. Dynamic changes in gut microbiota composition and related cytokines were found to be related to aGvHD, including pathogenic or protective changes. These findings suggested that manipulation of gut microbiota at different timepoints might be a promising avenue for preventing or treating this common complication.

Arid5a/IL-6/PAI-1 Signaling Is Involved in the Pathogenesis of Lipopolysaccharide-Induced Kidney Injury.

A systemic inflammatory response leads to widespread organ dysfunction, such as kidney dysfunction. Plasminogen activator inhibitor-1 (PAI-1) is involved in the pathogenesis of inflammatory kidney injury; however, the regulatory mechanism of PAI-1 in injured kidneys remains unclear. PAI-1 is induced by interleukin (IL)-6 in patients with sepsis. In addition, the stabilization of IL-6 is regulated by the adenine-thymine-rich interactive domain-containing protein 5a (Arid5a). Therefore, the aim of the present study was to examine the involvement of Arid5a/IL-6/PAI-1 signaling in lipopolysaccharide (LPS)-induced inflammatory kidney injury. LPS treatment to C57BL/6J mice upregulated Pai-1 mRNA in the kidneys. Enzyme-linked immunosorbent assay (ELISA) revealed that PAI-1 expression was induced in the culture supernatants of LPS-treated human umbilical vein endothelial cells, but not in those of LPS-treated human kidney 2 (HK-2) cells, a tubular cell line. Combined with single-cell analysis, endothelial cells were found to be responsible for PAI-1 elevation in LPS-treated kidneys. Administration of TM5441, a PAI-1 inhibitor, reduced the urinary albumin/creatinine ratio, concomitant with downregulation of Il-6 and Arid5a mRNA expressions. IL-6 treatment in LPS model mice further upregulated Pai-1 mRNA expression compared with LPS alone, accompanied by renal impairment. Furthermore, the expression of Il-6 and Pai-1 mRNA was lower in Arid5a knockout mice than in wild-type mice after LPS treatment. Taken together, the vicious cycle of Arid5a/IL-6/PAI-1 signaling is involved in LPS-induced kidney injury.

The enhancement of bioactive phytochemicals in agarwood leaves by post-harvest application using yeast extract elicitors and evaluation of their bioactivities

ABSTRACT Agarwood (Aquilaria crassna) leaves are commonly used as an herbal tea for health supplements. The present study aims to develop the post-harvest process of agarwood leaves using yeast extract (YE) as an elicitor and evaluate its anti-inflammatory and anti-diabetic effects. The results showed that 1% of YE elicitation for 120 min significantly increased genkwanin 5-O-β-primevoside (1.8-fold), mangiferin (1.3-fold), and total benzophenones (1.2-fold) contents over the control group. The phenylalanine ammonia-lyase (PAL) activity increased in maximal at 30 min after YE treatment. The agarwood leaf elicitation with 1% YE for 120 min showed a higher anti-inflammation effect by down-regulation of iNOS, IL-6, and COX-2 in LPS-stimulated RAW264.7 cells and an anti-diabetic effect by enhanced AMPK-α1, AMPK-α2, and GLUT4 in L6 cells over the non-treatment. Our results suggest that YE could be a biotic elicitor to enhance the phytochemical contents and increase the benefit of agarwood leaves as a health supplement.

PSII-11 Effects of Cannabidiol on Viability and Inflammatory Response in Bovine Primary Ruminal Epithelial Cells

Abstract Phytochemicals derived from hemp (Cannabis sativa) have been noted for their possible beneficial effects, including immunomodulation. Cannabidiol (CBD) may be a promising target as a therapy for gastrointestinal inflammation; however, the effects have not been explored in ruminants. The objective of this study was to investigate immunomodulatory properties of CBD in primary ruminal epithelial cells (REC). REC were isolated from ruminal papillae of yearling Holstein steers (n = 7). A preliminary experiment was performed to determine cytotoxicity of CBD using alamarBlue. Cells were exposed to either 10 or 50 µM CBD for 6 or 24 h. CBD at 10 µM did not affect viability. Growth was inhibited by over 60% at 50 µM for 24 h (P &amp;lt; 0.05), and therefore, 10 µM was selected for use. Two experiments, each 24h in duration, were conducted to explore various responses to CBD when inflammation was induced in REC with lipopolysaccharide (LPS). In Experiment 1, LPS was added prior to CBD exposure, simulating a therapy for established inflammation. In Experiment 2, LPS was added after CBD exposure, simulating a preventative strategy. Treatment groups for both experiments are described in Table 1. Total RNA was extracted from REC and relative gene expression of cytokines TNF, IL1B, IL6, CXCL8 and CXCL10 was evaluated using real time qPCR. Expression was normalized to the geometric mean of two stable housekeeping genes, GAPDH and STX5. All statistical analyses were performed using the lme4 and emmeans packages of R with exposure treatment as the fixed effect. In Experiment 1, expression of TNF, IL1B and CXCL8 was greater for LPS8 compared with CON by 5-, 29- and 14-fold, respectively, but less than LPS24 (P &amp;lt; 0.05). The LPS24+CBD16 group resulted in an upregulation of IL1B compared with CON, but not to the same magnitude as LPS24 (27- vs 93-fold; P = 0.002). A 3-fold down-regulation of IL6 was observed with the addition of CBD alone compared with CON (P &amp;lt; 0.001). Experiment 1 results suggest that CBD may reduce cytokine transcription during LPS-induced inflammation. For Experiment 2, compared with LPS8, there was less expression of IL6 and CXCL10 in CBD24+LPS8 (P &amp;lt; 0.05). There was a 9-fold upregulation of IL6 for CBD16-LPS8 (P &amp;lt; 0.001), but no difference between CON and CBD16-LPS8 for CXCL10 expression. Compared with CON, CBD24 resulted in a 2- and 6-fold down-regulation of IL6 (P = 0.003) and CXCL10 (P = 0.002). Experiment 2 results suggest that IL6 expression was influenced by CBD, but only while present in the media, while CXCL10 expression was suppressed by CBD, even after its removal. Overall, this study provides evidence of immunomodulation by CBD during a pro-inflammatory response in primary REC in culture.