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Periodontal disease is a common infectious disease that can lead to the loss of teeth. Hower how to effectively suppress the inflammation with medication is unclear. The aim of the present study was to investigate the anti-inflammatory effect of Oroxylin A in periodontitis and its potential role through heme oxygenase-1 (HO-1). Primary rat gingival fibroblasts (RGFs) were cultured using the tissue block method and identified by immunofluorescence. Following lipopolysaccharide (LPS) stimulation of RGFs, Oroxylin A was administered at 50, 100, 200 or 400 µg/ml. Reverse transcription-quantitative PCR was used to assess mRNA expression of cyclooxygenase (COX)-2, TNF-α, RANKL and osteoprotegerin (OPG). Western blotting was used to detect protein expression levels of COX −2, TNF-α, RANKL and OPG. Following HO-1 knockdown, the same treatment was performed. The expression of COX-2 in rat gingival tissue was observed by immunohistochemistry. One-way analysis of variance and Student's t test were used for statistical analysis. Oroxylin A downregulated mRNA expression of COX-2, TNF-α, RANKL and OPG in LPS-induced RGFs. With increase of Oroxylin A dose, the expression of HO-1 was gradually upregulated. When HO-1 was knocked down, Oroxylin A did not downregulate the expression of COX-2, TNF-α, RANKL and OPG in LPS-induced RGFs. Immunohistochemical results showed that expression of COX-2 was downregulated by Oroxylin A, and the expression of TNF-α, RANKL and OPG were also downregulated. Oroxylin A decreased expression of inflammatory cytokines in LPS-induced RGFs and had a good inhibitory effect on periodontitis in rats. [ABSTRACT FROM AUTHOR]

Gastric cancer (GC) is one of the most common malignancies in the world. It is essential to develop novel targets and therapeutic approaches for GC, which requires identification of novel functional molecules. WW-domain containing transcription regulator 1 (WWTR1) may activate many transcriptional factors and exhibit an important role in the development of various tissues in mammals. The results of the present study demonstrated that mRNA and protein levels of WWTR1 are increased in GC tissues and cell lines. The SGC7901 cell line was selected to perform RNA interference (RNAi) targeting WWTR1, and for subsequent study. Compared with control groups (cells without any treatment) and mock groups (cells treated with nonspecific siRNA), cell proliferation of siWWTR1 cells (cells treated with WWTR1 siRNA) was detected using a Cell Counting Kit-8 assay at 12, 24 and 48 h, and decreased in a time-dependent manner. Cell cycle and apoptosis status were determined by flow cytometry, and it was demonstrated that G1/S transition was blocked in the cell cycle and apoptosis promoted in siWWTR1 cells, compared with control and mock cells. Reverse transcription-quantitative polymerase chain reaction and western blotting were performed to detect the mRNA and protein levels of cell cycle and apoptosis-associated factors. The expression of Cyclin D1, cancer Myc and B cell lymphoma/leukemia-2 (Bcl-2) decreased and Bcl-2 associated X protein increased significantly in siWWRT1 cells, at the mRNA and protein level, compared with control and mock cells. With the exception of the Hippo pathway, siWWTR1 regulated downstream factors, including mothers against decapentaplegic homolog family member 3 (SMAD3) and inhibitor of DNA binding 1, HLH protein (ID1), HLH protein in the transforming growth factor (TGF)-β pathway. The expression of asparagine synthetase was decreased whereas ID1, SMAD3 (proteins that participate in intracellular TGF-β transduction) and betacellulin increased notably in siWWRT1 cells. In conclusion, WWTR1 promotes cell proliferation and inhibits apoptosis of GC cells by regulating cell cycle/apoptosis-associated factors, and effectors in the TGF-β pathway. [ABSTRACT FROM AUTHOR]

To investigate the effects of catalpol on corneal neovascularization (CNV) and associated inflammation, eye drops (5 mM catalpol or PBS) were administered four times daily to alkali-burn rat models of CNV and inflammation. Clinical evaluations of CNV and the degree of inflammation were performed on days 0, 4, 7, 10 and 14 under slit lamp microscopy. Eyes were collected on day 14 and prepared for hematoxylin and eosin, and immunofluorescence staining; corneal cell apoptosis was investigated via terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) staining. Protein expression levels of angiogenic and proinflammatory factors, including vascular endothelial growth factor (VEGF), pigment epithelium-derived factor (PEDF), tumor necrosis factor-α (TNF-α) and necrosis factor-κB (NF-κB) were determined by western blotting. The effects of catalpol on cell proliferation were investigated in vitro using human umbilical vein endothelial cells (HUVECs) and a Cell Counting kit-8 (CCK-8); alterations in migration and tube formation were investigated via HUVEC wound closure and tube formation assays. HUVEC viability and proliferative ability were inhibited in a dose-dependent manner; catalpol also decreased HUVEC cell migration and tube forming ability. Within alkali-burn rat models, decreased inflammation and CNV was associated with catalpol administration; as demonstrated with TUNEL, corneal cell apoptosis was decreased in response to catalpol. Western blot analysis revealed reduced protein expression levels of VEGF and TNF-α; however, PEDF and phosphorylated-NF-κB p65 were increased due to catalpol administration. The present study demonstrated the inhibitory effects exerted by catalpol on CNV and inflammation within alkali-burned rat models. Topical application of catalpol in vivo was associated with reduced CNV and inflammation; therefore, catalpol may be considered an anti-inflammatory agent for the clinical treatment of CNV. [ABSTRACT FROM AUTHOR]

Lymph node metastasis (LNM) is one of the major causes of cancer-associated morbidity and mortality in patients with lung cancer following radical pulmonary carcinoma resection. The present study aimed to investigate the relationship between the expression of collagen and calcium-binding epidermal growth factor domain-containing protein 1 (CCBE1) and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1) in tumor tissue with the clinical prognosis of lung cancer. The present study included 40 patients with lung cancer that underwent pulmonary carcinoma resection, including 10 patients with LNM, and 10 control patients who underwent pulmonary bullae resection. CCBE1 and LYVE1 expression was assessed in samples from normal and tumor tissue using polymerase chain reaction, western blot analysis and immunohistochemistry. CCBE1 expression appeared to be decreased in lung tumor tissue and further downregulated in samples from patients with LNM, and was revealed to be correlated with poor clinical outcome. Conversely, LYVE1 expression appeared to be upregulated in lung cancer tissue. In conclusion, the present results suggested that CCBE1 and LYVE1 may have potential as biomarkers for the identification of lung cancer patients at a high risk of LNM. [ABSTRACT FROM AUTHOR]

Type 1 diabetes (T1D) is an autoimmune disease characterized by a selective destruction of insulin‑secreting β‑cells. Both T cells and B cells serve a crucial role in pathogenesis and development of T1D. CD20 is a specific membrane antigen of B lymphocytes, while interleukin (IL)‑10 is an important cytokine secreted by T helper 2 cells and has a short half‑life in vivo. The combined effect of anti‑CD20 and IL‑10 on immune function of mice with T1D remains unknown. In the present study, 30 non‑obese diabetic (NOD) mice were treated with anti‑CD20 and adenoviral vector‑mediated interleukin‑10 (Ad‑mIL‑10) therapy. Alterations in CD4+, CD8+, CD4+CD25+Foxp3+ T cells, T‑box expressed in T‑cells (T‑bet), GATA‑binding protein‑3 (GATA‑3) interferon‑γ (IFN‑γ) and IL‑4 were detected by flow cytometry, reverse transcription‑quantitative polymerase chain reaction in NOD mice spleen tissue. The present results suggested that anti‑CD20 and IL‑10 treatment in NOD mice can modulate the immune functions by upregulating GATA‑3 and IL‑4 expression as well as downregulating T‑bet and IFN‑γ expression, which are involved in the pathogenesis of T1D. The current findings may provide a potential method for T1D treatment and a novel preventive therapy for T1D. Combination of anti‑CD20 and Ad‑mIL‑10 treatment had not only immune regulatory effects but also protective effects on islet β‑cells in NOD mice with T1DM at the early stages, by regulating T‑bet/GATA‑3 expression and Th1/Th2 cell differentiation, which has the potential for diabetes prevention and therapy. [ABSTRACT FROM AUTHOR]

The obesity-asthma phenotype is characterized by increased asthma severity and decreased glucocorticoid responsiveness. To date, the mechanism underlying the association between obesity and asthma remain to be fully elucidated. The present study investigated the correlation between oxidative stress and the nuclear factor (NF)-?B pathway in obese asthmatic mice. The animals were divided into the following groups: Control (n=8), comprising C57BL/6J mice without exposure to a high-fat diet; non-obese asthma group (n=8), comprising mice of a normal weight subjected to the induction of asthma; obese control group (n=8), comprising C57BL/6J mice subjected to a high-fat diet; and obese asthmatic group (n=8), comprising obese mice subject to the induction of asthma. The levels of the malondialdehyde (MDA) oxidant and glutathione (GSH) antioxidant in the lungs and bronchoalveolar lavage fluid (BALF) were measured using ELISA. The expression levels of inhibitory ?B kinase-β (IKK-β) and the inhibitor of ?Bα (I?B-α) in the pulmonary tissues was determined using western blot analysis. An electrophoretic mobility shift assay was performed to determine the transcription activity of NF-?B. The levels of MDA in the BALF and lung tissues increased significantly in the two asthmatic groups, compared with the control groups (P<0.01). The asthmatic mice showed significantly lower concentrations of GSH in the BALF and lung tissues, compared with the control groups (P<0.01). In the asthmatic animals, the expression of I?B kinase (IKK)-β and activation of NF-?B were upregulated in the pulmonary tissues, compared with those in the control groups (P<0.01). The expression of IKK-β and transcriptional activity of NF-?B were significantly higher the in obese asthmatic mice, compared with the non-obese asthmatic mice (P<0.01). On examining the expression levels of I?B-α in the pulmonary tissues, a significant reduction was found in the asthmatic animals, compared with the controls (P<0.01). In addition, the level of I?B-α was significantly lower in the obese asthmatics, compared with the non-obese asthmatics (P<0.01). MDA was positively correlated with NF-?B in the obese asthmatic group (R=0.83; P<0.05) and non-obese asthmatic group (R=0.82; P<0.05). Oxidative stress was upregulated in the pulmonary tissues of the asthmatic mice. This upregulation was more marked in the obese asthmatic mice, and was positively correlated with activation of the NF-?B signaling pathway in the pulmonary tissues. The results in the present study indicated that higher oxidative stress and activation of the NF-?B signaling pathway were observed in the lung tissues of the obese asthmatics. Furthermore, a positive correlation was identified between oxidative stress and NF-?B. [ABSTRACT FROM AUTHOR]

Type 1 diabetes is an autoimmune disorder, which occurs due to β cell damage. Interleukin (IL)-10, a pleotropic cytokine, has been reported to have anti-inflammatory, immunosuppressive and immunostimulatory properties. Administration of IL-10 is known to prevent autoimmune diabetes in non-obese diabetic (NOD) mice. However, the mechanism of IL-10-induced protection in NOD mice requires further investigation. The aim of the present study was to evaluate the protective effect of transgenic IL-10 expression in pancreatic β cells against autoimmune damage in NOD mice and to elucidate its mechanism of action. Female NOD mice (9 weeks old) were intraperitoneally injected with an adenovirus carrying either IL-10 (Adv-IL-10) or green fluorescent protein (Adv-GFP). Blood glucose was monitored weekly and the expression of IL-10 was evaluated using reverse transcription quantitative polymerase chain reaction. IL-10 and interferon (IFN)-γ expression levels in serum and splenocytes were analyzed. CD4+CD25+FoxP3+ T regulatory (Treg) cells were determined by flow cytometry. Apoptosis of pancreatic β cells was determined using a terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling assay and expression levels of Fas and caspase-3 were estimated by immunohistochemistry analysis. The results revealed that mice treated with IL-10 showed less severe insulitis and a lower incidence of diabetes compared with the saline control and Adv-GFP groups. In addition, compared with the control group, IFN-γ levels were decreased in sera and splenocytes, while IL-10 expression was increased in sera only. The number of CD4+CD25+FoxP3+ Treg cells was increased in IL-10-injected mice. Furthermore, the expression levels of Fas and caspase-3 were decreased in IL-10-injected mice compared with that of GFP-injected and control mice, which was concomitant with a reduction in β cell apoptosis. In conclusion, the present study demonstrated that IL-10 gene transfer reduced the expression of the inflammatory cytokines, attenuated pancreatic insulitis and inhibited β cell apoptosis. This therefore indicated that IL-10 reduced the incidence of diabetes in female NOD mice. [ABSTRACT FROM AUTHOR]

The present study investigated the underlying mechanism of the induction of regulatory T cells (Tregs) by retinal pigment epithelial (RPE) cells and the characteristics of these Tregs. Human RPE cells were cultured in the presence or absence of transforming growth factor-β 2 (TGF-β2), and reverse-transcription quantitative PCR was performed to determine the mRNA expression of indoleamine 2,3-dioxygenase (IDO) and nuclear factor erythroid 2-related factor (Nrf2). Supernatants of RPE cell cultures were added to CD4+ T cells to induce Tregs. The RPE-induced Tregs were purified by two-step magnetic cell sorting. The natural Tregs were isolated from the peripheral blood mononuclear cells of healthy volunteers. Purified CD4+ CD25- T cells (2 x 10(5)/well) were cultured alone or with Tregs (various densities, natural or RPE-induced). The proliferation of CD4+ CD25- T cells was determined by 3H-thymidine incorporation. After 24 h of stimulation with TGF-β2, the mRNA expression of IDO in RPE cells was upregulated. The highest level of IDO mRNA expression was reached after 72 h of stimulation with TGF-β2. However, the Nrf2 mRNA expression was slightly decreased after 24 h of stimulation with TGF-β2 and significantly increased after 48-72 h of TGF-β2 stimulation. Increased levels of CD25 expression were observed on CD4+ T cells exposed to supernatants of RPE cell cultures treated with TGF-β2 and recombinant interleukin-2. The RPE-induced Tregs were more effective at suppressing the proliferation of CD4+ CD25- T cells compared with native Tregs. These findings suggested that IDO may be a signaling protein in RPE cells which is implicated in the induction of Tregs. RPE-induced Tregs have the potential to be applied for immunotherapy for ocular inflammatory diseases.

Pharmacological studies have shown that the active components in Dendranthema morifolium exhibit protective effects against ischemia/reperfusion injury; however, its pharmacological action on blood vessels has not yet been investigated. The purpose of the present study was to assess the effects of the total flavones extracted from D. morifolium (Ramat.) Tzvel. cv. Hangju (FDM) on the vasocontraction and proliferation of vascular smooth muscle cells (VSMCs). The tension of rat thoracic aortic rings was measured using a mechanical force transducer attached to a recording system. FDM induced a dose‑dependent relaxation of rings with endothelium pre‑contracted by either phenylephrine (PE; 10(‑6) mol/l) or a high concentration of potassium chloride (KCl; 60 mmol/l). FDM did not significantly affect the vasorelaxant effects on mechanically removed endothelium. In endothelium‑denuded aortic rings depolarized by 60 mmol/l KCl, FDM inhibited the contraction induced by Ca2+. FDM reduced the transient contraction caused by PE in a Ca2+‑free solution, but did not affect the contraction induced by phorbol ester. Furthermore, FDM inhibited the proliferation of VSMCs with or without growth stimulation by insulin. In conclusion, that the vasorelaxation induced by FDM in rat aortic rings is not dependent on the endothelium but is mediated via a reduction of the influx of extracellular Ca2+ through the voltage‑dependent and receptor‑operated channels and via the inhibition of the release of intracellular Ca2+ in VSMCs. The anti‑proliferative activity of FDM suggests that it may be beneficial in inhibiting atherosclerosis.

A water‑soluble polysaccharide fraction (PPSB) was fractionated from Physalis alkekengi var. francheti and purified using DEAE‑52 Cellulose and Sephadex G‑200 chromatography. The physicochemical properties of PPSB and its molecular activities involved in the improvement of intestinal microflora balance were evaluated. PPSB (12.5‑25.0 mg/ml) was shown to promote the growth of Lactobacillus delbrueckii and to inhibit the growth of Escherichia coli. Denaturing gradient gel electrophoresis analysis identified Bacteroides, Lactobacillus, Helicobacter, Prevotella, Enterococcus, Odoribacter, Alistipes and Akkermansia as dominant organisms in the intestinal tract of mice. PPSB exhibited stimulatory effects on the growth of probiotic bacteria but inhibitory effects on the growth of pathogenic bacteria in vitro. The number of species and quantity of the Lactobacillus genus increased significantly with increasing concentrations of PPSB in vivo. Experimental results of this study suggest that PPSB may improve intestinal microflora imbalances and therefore has strong potential as a natural agent for restoring the intestinal microflora balance.