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Chinese herbal Jin-Ying-Tang attenuates the inflammatory response by inhibiting the activation of TLR4/MyD88/TRAF-6/NIK pathway at the mRNA level in LPS-stimulated mouse mammary epithelial cells
2016
Yi, Qiong | Li, Xin | Li, Yuan-Fang | Yang, Hang | Zhang, Xiao-Yi | Ma, Zhe | Wang, Lu
Introduction: The effects of Jin-Ying-Tang (JYT) on Toll-like Receptor 4 (TLR4) signalling transduction of lipopolysaccharide (LPS)-stimulated mouse mammary epithelial cells (MECs) in vitro were examined. Material and Methods: The cytotoxicity of JYT (0.06-62.50 mg/mL) on mouse MECs was determined by MTT assay. The MECs were co-cultured with LPS in the presence or absence of JYT (39.10 μg/mL, 391 μg/mL, 3910 μg/mL). The concentrations of interleukin-6 (IL-6) and tumour necrosis factor-α (TNF-α) in the culture supernatants were detected by ELISA. The mRNA expression of TLR4 and downstream TLR4 signalling molecules such as myeloid differentiation factor 88 (MyD88), tumour necrosis factor receptor associated factor 6 (TRAF-6), inhibitor κB (IκB), and nuclear factor κB inducing kinase (NIK) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Results: The results showed that the IC₅₀ of JYT on MECs was 12.25 mg/mL and JYT could significantly decrease the concentrations of IL-6 and TNF-α in LPS-stimulated MECs (P < 0.05). The mRNA expression of TLR4, MyD88, TRAF-6, IκB, and NIK was also significantly decreased when the LPS-stimulated MECs were cocultured at appropriate concentrations of JYT (P < 0.05, P < 0.01). Conclusion: These observations indicate a potential mechanism through which JYT attenuates the systemic inflammatory response to LPS-stimulated mouse mammary epithelial cells by inhibiting the activation of TLR4/MyD88/ TRAF-6/NIK pathway at the mRNA level.
Показать больше [+] Меньше [-]Role of mast cells in cow metritis
2016
Wang, Guo-Qing | Hou, Jin-Long | Huang, Huanyou | Yuan, Chao-Wen
Introduction: Bovine postpartum metritis causes great losses. Mast cell (MC)-released mediators participate in uterine inflammation and immune response, but their role in postpartum metritis in cows has not been reported. This study investigated the effect of endometrial MC on the disorder.Material and Methods: Ten dairy cows, at 6 to 10 days postpartum and with acute purulent metritis made up the experimental group, and 10 comparable healthy cows the control group. Endometrial histamine and IgE levels were determined by ELISA, and the MC particle state and expression of histamine H₁ (H₁R) and H₂ (H₂R) mRNA receptors were examined by transmission electron microscope and real-time quantitative PCR, respectively.Results: Endometrial histamine and IgE levels were significantly higher in the experimental group. In the control group, homogenously distributed size-varied granules were seen in MC cytoplasm of endometrium of lamina propria. In the experimental group however, these showed degranulation with features of reduction. The level of H₁R mRNA was lower in the experimental group, but that of H₂R mRNA was higher.Conclusion: The results suggest MC type I hypersensitivity characteristics during metritis, and histamine provocation of local inflammation. High expression of H₂R and low expression of H₁R inhibited the inflammatory response and prevented excessive uterine tissue damage.
Показать больше [+] Меньше [-]Expression of T helper cell–associated inflammatory mediator mRNAs in cells of bronchoalveolar lavage fluid samples and oxygen concentration in arterial blood samples from healthy horses exposed to hyperbaric oxygen
2016
Looijen, Maty G. P. | New, Dallas J. | Fischer, Carrie D. | Dardari, Rkia | Irwin, Karyn M. | Berezowski, Chrisotpher J. | Bond, Stephanie L. | Leguillette, Renaud
OBJECTIVE To evaluate the mRNA expression of T helper (Th)1, Th2, and Th17 cell–associated inflammatory mediators in cells of bronchoalveolar lavage fluid samples collected from healthy horses exposed to hyperbaric oxygen (HBO) and to monitor blood oxygen concentration during and following HBO therapy. ANIMALS 8 healthy horses. PROCEDURES In a randomized controlled crossover design study, each horse was exposed (beginning day 1) to 100% oxygen at a maximum of 3 atmospheres absolute (304 kPa) daily for 10 days or ambient air at atmospheric pressure in the HBO chamber for an equivalent amount of time (control). Bronchoalveolar lavage fluid samples were collected on days 0 and 10. After validation of candidate reference genes, relative mRNA expressions of various innate inflammatory, Th1 cell–derived, Th2 cell–derived (including eotaxin-2), Th17 cell–derived, and regulatory cytokines were measured by quantitative PCR assays. For 3 horses, arterial blood samples were collected for blood gas analysis during a separate HBO session. RESULTS The optimal combination of reference genes was glyceraldehyde-3-phosphate dehydrogenase, hypoxanthine ribosyltransferase, and ribosomal protein L32. Compared with day 0 findings, expression of eotaxin-2 mRNA was significantly lower (0.12-fold reduction) and the percentage of neutrophils in bronchoalveolar lavage fluid samples was significantly lower on day 10 when horses received HBO therapy. Values of Pao2 rapidly increased (> 800 mm Hg) but immediately decreased to pretreatment values when HBO sessions ended. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that HBO therapy does not increase mRNA expression of inflammatory cytokines, but reduces eotaxin-2 mRNA transcription. The Pao2 increase was transient with no cumulative effects of HBO.
Показать больше [+] Меньше [-]Effects of the −791(C→T) mutation in the promoter for tumor necrosis factor alpha on gene expression and resistance of Large White pigs to enterotoxigenic Escherichia coli F18
2016
Liu, Ying | Dai, Chaohui | Sun, Li | Zhu, Guoqiang | Wu, Shenglong | Bao, Wenbin
Tumor necrosis factor alpha (TNF-α) plays an important role in the immune system. In this study, TNF-α expression was analyzed in 11 tissues of 8 piglets resistant to enterotoxigenic Escherichia coli (ETEC) F18 and 8 ETEC F18-susceptible piglets from the Large White breed. The expression levels of TNF-α were high in immune organs (spleen, lung, thymus, and lymph nodes). The levels were higher in ETEC F18-resistant piglets than in ETEC F18-susceptible piglets, with significant differences in spleen, kidney, thymus, lymph node, and duodenum (P < 0.05). The mutation TNF-α −791(C→T) and 3 genotypes (CC, CT, and TT) were identified. The TNF-α expression levels in the spleen, kidney, lymph nodes, and duodenum were significantly higher in the TT pigs than in the CC pigs (P < 0.05). Thus, TNF-α −791(C→T) has significant effects on mRNA expression and may regulate ETEC F18 resistance of weaning piglets. Therefore, the −791(C→T) mutation of the TNF-α gene could be considered an important potential genetic marker of ETEC F18 resistance.
Показать больше [+] Меньше [-]Effect of granulocyte colony–stimulating factor administration on renal regeneration after experimentally induced acute kidney injury in dogs
2016
Lim, Chae-Young | Han, Jae-ik | Kim, Seung-Gon | Yi, Chʻang-min | Park, Hee-Myung
OBJECTIVE To evaluate the effects of granulocyte colony–stimulating factor (GCSF) administration in dogs with experimentally induced acute kidney injury. ANIMALS 6 healthy dogs. PROCEDURES After induction of kidney injury (day 0) with cisplatin (5 mg/kg, IV), the dogs were randomly assigned into 2 groups (n = 3 dogs/group). Then dogs immediately received GCSF (10 μg/kg) or 1 mL of saline (0.9% NaCl) solution (control group) SC; this treatment was repeated once daily for 4 additional days (days 1 through 4). A once-daily CBC (day 0 to 4), serum biochemical analysis (day 0 to 3), and urinalysis (day 0 to 3) were performed for each dog; samples were collected before administration of cisplatin (day 0) and before treatment with GCSF or saline solution (days 1 through 4). After sample collection and treatment on day 4, all dogs were euthanized; kidney tissue samples underwent histologic evaluation, immunohistochemical analyses, and cytokine profiling via reverse transcriptase PCR assay. RESULTS In the GCSF-treated group, the histologic evaluation and immunohistochemical analyses of kidney tissue revealed less fibrotic change and greater proliferation of renal tubular epithelial cells, compared with findings in the control group. The mRNA profiles of kidney tissue from the GCSF-treated group indicated lower expression of tumor necrosis factor-α and tumor growth factor-β, compared with findings in the control group; however, concentrations of factors related to renal regeneration were not greater in the GCSF-treated group. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that GCSF treatment can impede renal fibrosis and increase proliferation of renal tubules after experimentally induced acute kidney injury in dogs.
Показать больше [+] Меньше [-]Assessment of the expression of biomarkers of uremic inflammation in dogs with renal disease
2016
Nentwig, Alice | Schweighauser, Ariane | Maissen-Villiger, Carla | Bruckmaier, Rupert M. | Zurbriggen, Andreas | Dorland, H Anette van | Francey, Thierry
OBJECTIVE To assess the expression of inflammatory cytokines and enzymes in venous whole blood of dogs with impaired renal function attributable to various causes. ANIMALS 46 dogs with acute kidney injury (AKI), 8 dogs with chronic kidney disease (CKD), and 10 healthy dogs. PROCEDURES Dogs with AKI and CKD were prospectively enrolled during 2010 if they met inclusion criteria. Demographic and laboratory characteristics were evaluated for each dog, and expression of inflammatory cytokines (interleukin [IL]-1α, IL-1β, IL-8, tumor necrosis factor [TNF]-α, IL-10, and transforming growth factor [TGF]-β) and enzymes (inducible nitric oxide synthase [iNOS] and 5-lipoxygenase [5-LO]) was measured in venous whole blood obtained at initial evaluation. RESULTS Dogs with impaired renal function had markedly higher expression of the cytokines IL-1α, IL-1β, and TGF-β and the enzyme 5-LO, compared with expression in healthy dogs. Additionally, 17 of 46 AKI dogs (but none of the CKD dogs) had higher IL-8 mRNA expression and 3 of 8 CKD dogs (but only 2/46 AKI dogs) had higher TNF-α expression, compared with results for healthy dogs. No significant difference between renal disease groups was detected for inflammatory markers and laboratory variables, degree of azotemia, or cause of impaired renal function. CONCLUSIONS AND CLINICAL RELEVANCE In this study, expression of the cytokines IL-1α, IL-1β, and TGF-β and the enzyme 5-LO was clearly increased in dogs with renal disease, which suggested that these markers were part of an inflammatory response in animals with AKI or CKD.
Показать больше [+] Меньше [-]Production of serum amyloid A in equine articular chondrocytes and fibroblast-like synoviocytes treated with proinflammatory cytokines and its effects on the two cell types in culture
2016
Jacobsen, Stine | Ladefoged, Soren | Berg, Lisa C.
OBJECTIVE To investigate the role of the major equine acute phase protein serum amyloid A (SAA) in inflammation of equine intraarticular tissues. SAMPLE Articular chondrocytes and fibroblast-like synoviocytes (FLSs) from 8 horses (4 horses/cell type). PROCEDURES Chondrocytes and FLSs were stimulated in vitro for various periods up to 48 hours with cytokines (recombinant interleukin [IL]-1β, IL-6, tumor necrosis factor-α, or a combination of all 3 [IIT]) or with recombinant SAA. Gene expression of SAA, IL-6, matrix metalloproteinases (MMP)-1 and −3, and cartilage-derived retinoic acid-sensitive protein were assessed by quantitative real-time PCR assay; SAA protein was evaluated by immunoturbidimetry and denaturing isoelectric focusing and western blotting. RESULTS All cytokine stimulation protocols increased expression of SAA mRNA and resulted in detectable SAA protein production in chondrocytes and FLSs. Isoforms of SAA in lysed chondrocytes and their culture medium corresponded to those previously detected in synovial fluid from horses with joint disease. When exposed to SAA, chondrocytes and FLSs had increased expression of IL-6, SAA, and MMP3, and chondrocytes had increased expression of MMP-1. Chondrocytes had decreased expression of cartilage-derived retinoic acid-sensitive protein. CONCLUSIONS AND CLINICAL RELEVANCE Upregulation of SAA in chondrocytes and FLSs stimulated with proinflammatory cytokines and the proinflammatory effects of SAA suggested that SAA may be involved in key aspects of pathogenesis of the joint inflammation in horses.
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