scholarly journals Culturing Articular Cartilage Explants in the Presence of Autologous Adipose Tissue Modifies Their Inflammatory Response to Lipopolysaccharide

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
Wendy Pearson ◽  
Anna E. N. Garland ◽  
Ashley Nixon ◽  
John P. Cant ◽  
Mark B. Hurtig
Keyword(s):  
Adipose Tissue ◽  
Inflammatory Response ◽  
Cell Viability ◽  
Inflammatory Responses ◽  
Cartilage Explants ◽  
Cartilage Tissue ◽  
No Production ◽  
Chondrocyte Viability ◽  
Tissue Explants ◽  
Independent Mechanism

The purpose of the current study was to explore the effect of autologous adipose tissue on cartilage responses to lipopolysaccharide (LPS). We hypothesized that LPS elicits an inflammatory response in cartilage, and that response is augmented in the presence of adipose tissue. Furthermore, we hypothesized that this augmented inflammatory response is due, at least in part, to increased exposure of cartilage to adipose tissue-derived c3a. Porcine cartilage explants from market-weight pigs were cultured in the presence or absence of autologous adipose tissue for 96 hours, the final 48 hours of which they were stimulated with LPS (0 or 10 μg/mL). Adipose tissue explants were also cultured alone, in the presence or absence of LPS. Media from all cartilage treatments was assayed for c3a/c3a des Arg, PGE2, GAG, and NO, and the viability of cartilage tissue was determined by differential fluorescent staining. Media from adipose tissue explants was assayed for c3a/c3a des Arg and PGE2. LPS produced a significant increase in PGE2, GAG, and NO production when cartilage was cultured in the absence of adipose tissue. Coculture of adipose tissue prevented a significant increase in PGE2 in cartilage explants. There was no effect of adipose tissue on LPS-induced GAG or NO, but the presence of adipose tissue significantly reduced cell viability in LPS-stimulated cartilage explants. Adipose tissue explants from lean animals reduced inflammatory responses of cartilage to LPS via a c3a/c3a des Arg-independent mechanism and were associated with a significant decline in cell viability. Thus, contrary to our hypothesis, adipose tissue from lean animals does not augment the inflammatory response of cartilage to stimulation by LPS. The mechanism of modulatory effects of adipose tissue on LPS-induced increase in PGE2 and decline in chondrocyte viability requires further research but appears to have occurred via a mechanism that is independent of adipocentric c3a/c3a des Arg.

scholarly journals Electroacupuncture Decreases the Leukocyte Infiltration to White Adipose Tissue and Attenuates Inflammatory Response in High Fat Diet-Induced Obesity Rats

2014 ◽  
Vol 2014 ◽  
pp. 1-11 ◽  
Author(s):  
Chorng-Kai Wen ◽  
Tzung-Yan Lee
Keyword(s):  
Adipose Tissue ◽  
Inflammatory Response ◽  
White Adipose Tissue ◽  
High Fat Diet ◽  
Target Genes ◽  
Inflammatory Responses ◽  
Regulatory Element ◽  
Epididymal Adipose Tissue ◽  
High Fat ◽  
Obese Rats

Suppression of white adipose tissue inflammatory signaling may contribute to the pathogenesis of obesity-induced inflammatory response. However, the precise mechanism of efficacy of acupuncture related to adipose tissue remains poorly understood. In the present study we evaluated the anti-inflammatory activities of 10 Hz electroacupuncture (EA) which was applied at the acupoint Zusanli (ST36) for 20 min per day in high-fat diet- (HFD-) induced obesity model. Treatment lasted for one week. Obese rats treated with EA showed significantly reduced body weight compared with the rats in HFD group. EA decreased the number of F4/80 and CD11b-positive macrophages in epididymal adipose tissue. We found that 10 Hz EA given 7 days/week at ST36 acupoints significantly alleviated macrophage recruitment and then improved the obesity-associated factors of sterol regulatory element-binding protein-1 (SREBP-1) and target genes expression in rats with HFD. Adipose tissue inflammatory responses indicated by tumor necrosis factor-α(TNF-α), IL-6, monocyte chemotactic protein-1 (MCP-1), and CD68 mRNA expression were significantly reduced by EA in obese rats. Additionally, EA was found to significantly reduced serum levels of TNF-α, IL-6, and IL-1 in this model. These results indicated that EA improved adipose tissue inflammatory response in obese rats, at least partly, via attenuation of lipogenesis signaling.

scholarly journals Nonlabeling and quantitative assessment of chondrocyte viability in articular cartilage with intrinsic nonlinear optical signatures

2020 ◽  
Vol 245 (4) ◽  
pp. 348-359 ◽  
Author(s):  
Yang Li ◽  
Xun Chen ◽  
Beckham Watkins ◽  
Neal Saini ◽  
Steven Gannon ◽  
...  
Keyword(s):  
Articular Cartilage ◽  
Second Harmonic Generation ◽  
Cell Viability ◽  
Harmonic Generation ◽  
Second Harmonic ◽  
Cartilage Tissue ◽  
Imaging Method ◽  
Chondrocyte Viability ◽  
Two Photon

Chondrocyte viability is a crucial factor for evaluating cartilage health. Most prevalent cell viability assays rely on dyes and are not applicable for in vivo or longitudinal studies. Here we demonstrated that the two-photon excited autofluorescence and second harmonic generation microscopy provided high-resolution imaging of cartilage tissue and distinguished live/dead chondrocytes by visual assessment. Furthermore, the normalized autofluorescence ratio was proposed as a quantitative indicator to determine chondrocyte viability. Based on the indicator, a curve fitting and simulated receiver operating characteristic method was proposed to identify the live/dead cell populations as well as the indicator threshold without dye labeling. Thus, it established the label-free imaging method for chondrocyte viability assay in cartilage tissue. Impact statement Chondrocytes are the only cellular component found in the cartilage, playing a critical role in maintaining the homeostasis of articular cartilage. The viability of chondrocytes is a crucial factor for evaluating cartilage health. However, the current prevalent cell viability assays rely on dye staining and thereby are not applicable in vivo or in longitudinal assessments. In this study, we demonstrate that the intrinsic signals such as two-photon excited autofluorescence and second harmonic generation can be used to classify live and dead chondrocytes in cartilage tissue. A quantitative measure is also proposed allowing development of automated assessment algorithms. The nonlabeling nature of this method suggests the potential applicability to nondestructive and in vivo assessment of cartilage health.

Effects of Shiranuhi flower extracts and fractions on lipopolysaccharide-induced inflammatory responses in murine RAW 264.7 cells

2018 ◽  
Vol 43 (4) ◽  
pp. 375-384 ◽  
Author(s):  
Chang-Gu Hyun ◽  
Min-Jin Kim ◽  
Sang Suk Kim ◽  
Ji Hye Ko ◽  
Young Il Moon ◽  
...  
Keyword(s):  
Cell Viability ◽  
Inflammatory Responses ◽  
Antioxidant Activities ◽  
Mapk Signaling ◽  
Raw 264.7 Cells ◽  
No Production ◽  
Raw 264.7 ◽  
Dependent Manner ◽  
Anti Inflammatory ◽  
Inflammatory Effect

Abstract Objective In this study, we evaluated the anti-inflammatory effect of Shiranuhi flower in RAW 264.7 cells. Methods The effects of the extracts and solvent fractions on cell viability and LPS-induced inflammatory responses were investigated in RAW 264.7 cells. Results The results showed that the ethyl acetate fraction (HEF) significantly decreased NO production in RAW 264.7 cells; however, cell viability was not affected. In addition, ELISA assay revealed that HEF significantly inhibited the productions of PGE2, TNF-α, and IL-6. As well, using Western blot analysis, it was observed that HEF significantly reduced the expression levels of iNOS and COX-2 in a dose dependent manner. Furthermore, we detected a reduced phosphorylation of mitogen-activated protein kinases such as p38, JNK, and ERK1/2. This indicates that HEF regulates LPS-induced inflammatory responses, at least in part, via suppressing the MAPK signaling pathway. Correlation analysis also showed that anti-inflammatory activities were highly correlated to antioxidant activities in this study. Characterization of the Shiranuhi flowers for flavonoid contents using HPLC showed varied quantity of narirutin and hesperidin. Conclusion Overall, the results demonstrate that HEF may be a potential anti-inflammatory agent. In addition, our findings contribute to understanding the molecular mechanism underlying the anti-inflammatory effect of Shiranuhi flower.

scholarly journals Folic Acid Is Able to Polarize the Inflammatory Response in LPS Activated Microglia by Regulating Multiple Signaling Pathways

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Antonia Cianciulli ◽  
Rosaria Salvatore ◽  
Chiara Porro ◽  
Teresa Trotta ◽  
Maria Antonietta Panaro
Keyword(s):  
Folic Acid ◽  
Inflammatory Response ◽  
Signaling Pathways ◽  
Inflammatory Responses ◽  
Interleukin 1 ◽  
Cytokine Signaling ◽  
No Production ◽  
Acid Pretreatment ◽  
Anti Inflammatory ◽  
Multiple Signaling

We investigated the ability of folic acid to modulate the inflammatory responses of LPS activated BV-2 microglia cells and the signal transduction pathways involved. To this aim, the BV-2 cell line was exposed to LPS as a proinflammatory response inducer, in presence or absence of various concentrations of folic acid. The production of nitric oxide (NO) was determined by the Griess test. The levels of tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and IL-10 were determined by ELISA. Inducible NO synthase (iNOS), nuclear transcription factor-kappa B (NF-κB) p65, MAPKs protein, and suppressors of cytokine signaling (SOCS)1 and SOCS3 were analyzed by western blotting. TNF-αand IL-1β, as well as iNOS dependent NO production, resulted significantly inhibited by folic acid pretreatment in LPS-activated BV-2 cells. We also observed that folic acid dose-dependently upregulated both SOCS1 and SOCS3 expression in BV-2 cells, leading to an increased expression of the anti-inflammatory cytokine IL-10. Finally, p-IκBα, which indirectly reflects NF-κB complex activation, and JNK phosphorylation resulted dose-dependently downregulated by folic acid pretreatment of LPS-activated cells, whereas p38 MAPK phosphorylation resulted significantly upregulated by folic acid treatment. Overall, these results demonstrated that folic acid was able to modulate the inflammatory response in microglia cells, shifting proinflammatory versus anti-inflammatory responses through regulating multiple signaling pathways.

scholarly journals LPS promotes the progression of sepsis by activation of lncRNA HULC/miR-204-5p/TRPM7 network in HUVECs

2020 ◽  
Vol 40 (6) ◽  
Author(s):  
Xinghai Chen ◽  
Debiao Song
Keyword(s):  
Oxidative Stress ◽  
Inflammatory Response ◽  
Cell Viability ◽  
Transient Receptor Potential ◽  
Inflammatory Responses ◽  
Umbilical Vein ◽  
Receptor Potential ◽  
Luciferase Reporter ◽  
Inflammatory Injury ◽  
And Oxidative Stress

Abstract Sepsis is a systemic inflammatory response syndrome caused by infection. Lipopolysaccharide (LPS) has been reported to induce inflammatory responses, and long non-coding RNA highly up-regulated in liver cancer (HULC) expression was associated with the progression of sepsis. But the role and underlying mechanism of HULC in LPS-induced sepsis remain unclear. Cell viability and apoptosis were measured by methyl thiazolyl tetrazolium (MTT) and flow cytometry assays, respectively. The levels of apoptosis-related proteins, inflammatory cytokines and transient receptor potential melastatin7 (TRPM7) were detected by western blot. Reactive oxygen species (ROS), superoxide dismutase (SOD) and malondialdehyde (MDA) levels were detected by dichloro-dihydro-fluorescein diacetate (DCFH-DA) method using commercial kit. HULC, microRNA-204-5p (miR-204-5p) and TRPM7 expressions in serum of sepsis patients and human umbilical vein endothelial cells (HUVECs) were examined by quantitative real-time polymerase chain reaction (qRT-PCR). Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were used to confirm the interaction between HULC and miR-204-5p, miR-204-5p and TRPM7. LPS stimulation restrained cell viability and facilitated apoptosis, inflammatory injury and oxidative stress in HUVECs. HULC and TRPM7 were increased and accompanied with decreased miR-204-5p expression in serum of sepsis patients. A significant negative correlation between miR-204-5p and HULC or TRPM7 was observed, and there was a positive relationship between expressions of HULC and TRPM7. Importantly, LPS inhibited the cell viability and induced apoptosis, inflammatory injury and oxidative stress of HUVECs by up-regulating the expressions of HULC and TRPM7, and down-modulating miR-204-5p expression. Mechanically, HULC positively regulated TRPM7 expression by sponging miR-204-5p in HUVECs. LPS impaired cell viability, and promoted cell apoptosis, inflammatory response and oxidative stress in HUVECs by regulating HULC/miR-204-5p/TRPM7 axis.

scholarly journals Differences in the Inflammatory Response of White Adipose Tissue and Adipose-Derived Stem Cells

2020 ◽  
Vol 21 (3) ◽  
pp. 1086
Author(s):  
Sara Taha ◽  
Elias Volkmer ◽  
Elisabeth Haas ◽  
Paolo Alberton ◽  
Tobias Straub ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Adipose Tissue ◽  
Inflammatory Response ◽  
Gene Expression Profile ◽  
Expression Profile ◽  
White Adipose Tissue ◽  
Inflammatory Responses ◽  
System Response ◽  
Adipose Derived Stem Cells

The application of liposuctioned white adipose tissue (L-WAT) and adipose-derived stem cells (ADSCs) as a novel immunomodulatory treatment option is the currently subject of various clinical trials. Because it is crucial to understand the underlying therapeutic mechanisms, the latest studies focused on the immunomodulatory functions of L-WAT or ADSCs. However, studies that examine the specific transcriptional adaptation of these treatment options to an extrinsic inflammatory stimulus in an unbiased manner are scarce. The aim of this study was to compare the gene expression profile of L-WAT and ADSCs, when subjected to tumor necrosis factor alpha (TNFα), and to identify key factors that might be therapeutically relevant when using L-WAT or ADSCs as an immuno-modulator. Fat tissue was harvested by liposuction from five human donors. ADSCs were isolated from the same donors and shortly subjected to expansion culture. L-WAT and ADSCs were treated with human recombinant TNFα, to trigger a strong inflammatory response. Subsequently, an mRNA deep next-generation sequencing was performed to evaluate the different inflammatory responses of L-WAT and ADSCs. We found significant gene expression changes in both experimental groups after TNFα incubation. However, ADSCs showed a more homogenous gene expression profile by predominantly expressing genes involved in immunomodulatory processes such as CCL19, CCL5, TNFSF15 and IL1b when compared to L-WAT, which reacted rather heterogeneously. As RNA sequencing between L-WAT and ADSCS treated with TNFα revealed that L-WAT responded very heterogeneously to TNFα treatment, we therefore conclude that ADSCs are more reliable and predictable when used therapeutically. Our study furthermore yields insight into potential biological processes regarding immune system response, inflammatory response, and cell activation. Our results can help to better understand the different immunomodulatory effects of L-WAT and ADSCs.

Anti-Inflammatory Effects of Novel Standardized Platelet Rich Plasma Releasates on Knee Osteoarthritic Chondrocytes and Cartilage in vitro

2019 ◽  
Vol 20 (11) ◽  
pp. 920-933 ◽  
Author(s):  
Lucía Gato-Calvo ◽  
Tamara Hermida-Gómez ◽  
Cristina R. Romero ◽  
Elena F. Burguera ◽  
Francisco J. Blanco
Keyword(s):  
Gene Expression ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Cartilage Explants ◽  
Ex Vivo Model ◽  
Chondrocyte Viability ◽  
Platelet Concentration ◽  
The Absolute ◽  
Anti Inflammatory

Background: Platelet Rich Plasma (PRP) has recently emerged as a potential treatment for osteoarthritis (OA), but composition heterogeneity hampers comparison among studies, with the result that definite conclusions on its efficacy have not been reached. Objective: 1) To develop a novel methodology to prepare a series of standardized PRP releasates (PRP-Rs) with known absolute platelet concentrations, and 2) To evaluate the influence of this standardization parameter on the anti-inflammatory properties of these PRP-Rs in an in vitro and an ex vivo model of OA. Methods: A series of PRPs was prepared using the absolute platelet concentration as the standardization parameter. Doses of platelets ranged from 0% (platelet poor plasma, PPP) to 1.5·105 platelets/µl. PRPs were then activated with CaCl2 to obtain releasates (PRP-R). Chondrocytes were stimulated with 10% of each PRP-R in serum-free culture medium for 72 h to assess proliferation and viability. Cells were co-stimulated with interleukin (IL)-1β (5 ng/ml) and 10% of each PRP-R for 48 h to determine the effects on gene expression, secretion and intra-cellular content of common markers associated with inflammation, catabolism and oxidative stress in OA. OA cartilage explants were co-stimulated with IL-1β (5 ng/ml) and 10% of either PRP-R with 0.75·105 platelets/µl or PRP-R with 1.5·105 platelets/µl for 21 days to assess matrix inflammatory degradation. Results: Chondrocyte viability was not affected, and proliferation was dose-dependently increased. The gene expression of all pro-inflammatory mediators was significantly and dose-independently reduced, except for that of IL-1β and IL-8. Immunoblotting corroborated this effect for inducible NO synthase (NOS2). Secreted matrix metalloproteinase-13 (MMP-13) was reduced to almost basal levels by the PRP-R from PPP. Increasing platelet dosage led to progressive loss to this anti-catabolic ability. Safranin O and toluidine blue stains supported the beneficial effect of low platelet dosage on cartilage matrix preservation. Conclusion: We have developed a methodology to prepare PRP releasates using the absolute platelet concentration as the standardization parameter. Using this approach, the composition of the resulting PRP derived product is independent of the donor initial basal platelet count, thereby allowing the evaluation of its effects objectively and reproducibly. In our OA models, PRP-Rs showed antiinflammatory, anti-oxidant and anti-catabolic properties. Platelet enrichment could favor chondrocyte proliferation but is not necessary for the above effects and could even be counter-productive.

scholarly journals Comparing the protective effects of resveratrol, curcumin and sulforaphane against LPS/IFN-γ-mediated inflammation in doxorubicin-treated macrophages

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Haidy A. Saleh ◽  
Eman Ramdan ◽  
Mohey M. Elmazar ◽  
Hassan M. E. Azzazy ◽  
Anwar Abdelnaser
Keyword(s):  
Nitric Oxide ◽  
Inflammatory Response ◽  
Raw 264.7 Cells ◽  
Inos Mrna ◽  
No Production ◽  
Raw 264.7 ◽  
Mrna Levels ◽  
Protective Effects ◽  
Tnf Α ◽  
Ifn Γ

AbstractDoxorubicin (DOX) chemotherapy is associated with the release of inflammatory cytokines from macrophages. This has been suggested to be, in part, due to DOX-mediated leakage of endotoxins from gut microflora, which activate Toll-like receptor 4 (TLR4) signaling in macrophages, causing severe inflammation. However, the direct function of DOX on macrophages is still unknown. In the present study, we tested the hypothesis that DOX alone is incapable of stimulating inflammatory response in macrophages. Then, we compared the anti-inflammatory effects of curcumin (CUR), resveratrol (RES) and sulforaphane (SFN) against lipopolysaccharide/interferon-gamma (LPS/IFN-γ)-mediated inflammation in the absence or presence of DOX. For this purpose, RAW 264.7 cells were stimulated with LPS/IFN-γ (10 ng/mL/10 U/mL) in the absence or presence of DOX (0.1 µM). Our results showed that DOX alone is incapable of stimulating an inflammatory response in RAW 264.7 macrophages. Furthermore, after 24 h of incubation with LPS/IFN-γ, a significant increase in tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and inducible nitric oxide synthase (iNOS) mRNA levels was observed. Similarly, nitric oxide (NO) production and TNF-α and IL-6 protein levels were significantly upregulated. Moreover, in LPS/IFN-γ-treated macrophages, the microRNAs (miRNAs) miR-146a, miR-155, and miR-21 were significantly overexpressed. Interestingly, upon testing CUR, RES, and SFN against LPS/IFN-γ-mediated inflammation, only SFN was able to significantly reverse the LPS/IFN-γ-mediated induction of iNOS, TNF-α and IL-6 and attenuate miR-146a and miR-155 levels. In conclusion, SFN, at the transcriptional and posttranscriptional levels, exhibits potent immunomodulatory action against LPS/IFN-γ-stimulated macrophages, which may indicate SFN as a potential treatment for DOX-associated inflammation.

scholarly journals Histatin-1 Attenuates LPS-Induced Inflammatory Signaling in RAW264.7 Macrophages

2021 ◽  
Vol 22 (15) ◽  
pp. 7856
Author(s):  
Sang Min Lee ◽  
Kyung-No Son ◽  
Dhara Shah ◽  
Marwan Ali ◽  
Arun Balasubramaniam ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Cytokine Production ◽  
Inflammatory Responses ◽  
Critical Role ◽  
Mapk Signaling ◽  
Response To Treatment ◽  
No Production ◽  
Inflammatory Signaling ◽  
Raw264.7 Macrophages ◽  
Inflammatory Mediator Production

Macrophages play a critical role in the inflammatory response to environmental triggers, such as lipopolysaccharide (LPS). Inflammatory signaling through macrophages and the innate immune system are increasingly recognized as important contributors to multiple acute and chronic disease processes. Nitric oxide (NO) is a free radical that plays an important role in immune and inflammatory responses as an important intercellular messenger. In addition, NO has an important role in inflammatory responses in mucosal environments such as the ocular surface. Histatin peptides are well-established antimicrobial and wound healing agents. These peptides are important in multiple biological systems, playing roles in responses to the environment and immunomodulation. Given the importance of macrophages in responses to environmental triggers and pathogens, we investigated the effect of histatin-1 (Hst1) on LPS-induced inflammatory responses and the underlying molecular mechanisms in RAW264.7 (RAW) macrophages. LPS-induced inflammatory signaling, NO production and cytokine production in macrophages were tested in response to treatment with Hst1. Hst1 application significantly reduced LPS-induced NO production, inflammatory cytokine production, and inflammatory signaling through the JNK and NF-kB pathways in RAW cells. These results demonstrate that Hst1 can inhibit LPS-induced inflammatory mediator production and MAPK signaling pathways in macrophages.

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