Guo-Ying Sun

Calcitonin gene-related peptide promotes the wound healing of human bronchial epithelial cells via PKC and MAPK pathways

Calcitonin gene-related peptide (CGRP) is a 37-amino acid neuropeptide derived from the calcitonin gene. CGRP is widely distributed in the central and peripheral neuronal systems. In the lung, CGRP could modulate dendritic cell function, stimulate proliferation of alveolar epithelial cells and mediate lung injury in mice. In this study, we investigated the effect of CGRP on the wound healing of human bronchial epithelial cells (HBECs) in vitro. The results showed that CGRP accelerated the recovery of wound area of monolayer HBECs in a dose-dependent manner. CGRP inhibited the lipopolysaccharide-induced apoptosis in HBECs. The percentage of S phase and G2/M phase was increased in HBECs after CGRP treatment. CGRP upregulated the expression of Ki67 in a dose-dependent manner. Some pathway inhibitors were used to investigate the signal pathway in which CGRP was involved. We found out that PKC pathway inhibitor (H-7) and MAPK pathway inhibitor (PD98059) could partially attenuate the effect of CGRP, which indicated that CGRP might promote the wound healing of HBECs via PKC and/or MAPK dependent pathway by accelerating migration and proliferation, and inhibiting apoptosis.

Soluble Epoxide Hydrolase Inhibitor Attenuates Lipopolysaccharide-Induced Acute Lung Injury and Improves Survival in Mice

ABSTRACT Acute lung injury (ALI) is characterized by rapid alveolar injury, vascular leakage, lung inflammation, neutrophil accumulation, and induced cytokines production leading to lung edema. The mortality rate of patients suffering from ALI remains high. Epoxyeicosatrienoic acids (EETs) are cytochrome P450-dependent derivatives of polyunsaturated fatty acid with antihypertensive, profibrinolytic, and anti-inflammatory functions. EETs are rapidly hydrated by soluble epoxide hydrolase (sEH) to their less potent diols. The aim of this study was to investigate the role of sEH inhibitor trifluoromethoxyphenyl propionylpiperidin urea (TPPU) and EETs in lipopolysaccharide (LPS)-induced ALI of mice. Our studies revealed that inhibition of sEH with TPPU attenuated the morphological changes in mice, decreased the neutrophil infiltration to the lung, pro-inflammatory cytokine levels (IL-1&bgr; and TNF-&agr;) in serum and bronchoalveolar lavage fluid (BALF), and alveolar capillary leakage (lung wet/dry ratio and total protein concentration in BALF). TPPU improved the survival rate of LPS-induced ALI. In addition, in vitro experiments revealed that both TPPU and EETs (11,12-EET and 14,15-EET) suppressed the expression of IL-1&bgr; and TNF-&agr;, and LDH release in RAW264.7 cells. These results indicate that EETs play a role in dampening LPS-induced acute lung inflammation, and suggest that sEH could be a valuable candidate for the treatment of ALI.

Vasoactive intestinal peptide suppresses macrophage-mediated inflammation by downregulating interleukin-17A expression via PKA- and PKC-dependent pathways.

Interleukin (IL)‐17A is a pro‐inflammatory cytokine that markedly enhances inflammatory responses in the lungs by recruiting neutrophils and interacting with other pro‐inflammatory mediators. Reducing the expression of IL‐17A could attenuate inflammation in the lungs. However, whether VIP exerts its anti‐inflammatory effects by regulating the expression of IL‐17A has remained unclear. Here, we show that there is a remarkable increase of IL‐17A in bronchoalveolar lavage fluid (BALF) and lung tissue of mice with acute lung injury (ALI). Moreover, lipopolysaccharides (LPS) stimulated elevated expression of IL‐17A, which was evident by the enhanced levels of mRNA and protein observed. Furthermore, we also found that VIP inhibited LPS‐mediated IL‐17A expression in a time‐ and dose‐dependent manner in an in vitro model of ALI and that this process might be mediated via the phosphokinase A (PKA) and phosphokinase C (PKC) pathways. Taken together, our results demonstrated that VIP might be an effective protector during ALI by suppressing IL‐17A expression.

l‐Arginine promotes DNA repair in cultured bronchial epithelial cells exposed to ozone: involvement of the ATM pathway

Ozone may lead to DNA breaks in airway epithelial cells. p‐ATM (phosphorylated ataxia telangiectasia mutated) plays a pivotal role in DNA repair. Derivatives of NO (nitric oxide) are regulators of the phosphorylation, and NO is increased under oxidative stress. The present study was aimed to study the effect of NO donor l‐arg (l‐arginine) on DNA damage repair in human bronchial epithelial cells exposed to ozone and the potential mechanisms involved. HBECs (human bronchial epithelial cells) were cultured with or without ozone (1.5 ppm, 30 min), DNA breaks were measured with a comet assay and agarose gel electrophoresis, cell cycling was determined by flow cytometry and p‐ATM was measured by immunofluorescence and Western blot. Data were analysed by ANOVA (analysis of variance). P<0.05 was considered as significant. Ozone induced marked DNA breaks, G1‐phase arrest and increased expression of p‐ATM in HBECs, while wortmannin reduced the levels of p‐ATM induced by ozone; the NO donor, l‐arg, minimized the effects of ozone‐induced DNA breaks and increased the level of p‐ATM, while the NO synthase inhibitor, l‐NMMA [N(G)‐minomethyl‐l‐arginine], restrained those effects of l‐arg. The effect of l‐arg on DNA repair is NO‐mediated, and p‐ATM is implicated in the processes of DNA repair.

Vasoactive intestinal peptide overexpression mediated by lentivirus attenuates lipopolysaccharide-induced acute lung injury in mice by inhibiting inflammation

HIGHLIGHTSWe firstly found intratracheal injection of Lenti‐VIP for 7days significantly increased intrapulmonary VIP content in mice.VIP alleviates ALI induced by LPS in vivo.VIP inhibits pro‐inflammatory TNF‐&agr; expression in murine macrophages stimulated by LPS via PKC and PKA pathways. ABSTRACT Vasoactive intestinal peptide (VIP) is one of the most abundant neuropeptides in the lungs with various biological characters. We have reported that VIP inhibited the expressions of TREM‐1 and IL‐17A, which are involved in the initiation and amplification of inflammation in acute lung injury (ALI). However, the overall effect of VIP on ALI remains unknown. The aim of this study is to investigate the therapeutic effect of VIP mediated by lentivirus (Lenti‐VIP) on lipopolysaccharide (LPS)‐induced murine ALI. We found that the expression of intrapulmonary VIP peaked at day7 after the intratracheal injection of Lenti‐VIP. Lenti‐VIP increased the respiratory rate, lung compliance, and tidal volume, while decreased airway resistance in ALI mice, detected by Buxco system. Lenti‐VIP significantly reduced inflammatory cell infiltration and maintained the integrity of the alveolar septa. Lenti‐VIP also remarkably decreased the total protein level, the number of neutrophil and lactate dehydrogenase activity in the bronchoalveolar lavage fluid of LPS‐induced ALI mice. In addition, Lenti‐VIP down‐regulated pro‐inflammatory tumor necrosis factor (TNF)‐&agr; mRNA and protein expression, while up‐regulated anti‐inflammatory interleukin‐10 mRNA and protein expression in lungs of ALI mice. Furthermore, we observed that VIP reduced the TNF‐&agr; expression in murine macrophages under LPS stimulation through protein kinase C and protein kinase A pathways. Together, our findings show that in vivo administration of lentivirus expressing VIP exerts a potent therapeutic effect on LPS‐induced ALI in mice via inhibiting inflammation.

Effect of EETs on proliferation and apoptosis of lipopolysaccharide-treatment HBECs#

(1 mg/kg/day) was gavage dosing for 21 days. Then the mice were sacrificed. The lift lung lobes were fixed for HE and Masson staining, and the right lobes were used to observe the procollagen I and III expressions by RT-PCR. Results: Histological examination showed that TPPU treatment alleviated the bleomycin-induced inflammation, maintained the alveolar structure. Masson staining indicated that TPPU could attenuate the expression of bleomycin-induced collagen. Furthermore, the expression of procollagen I and III mRNAs were significantly reduced with TPPU treatment. Discussion: TPPU could increase the concentration of EETs in lung, which alleviated the bleomycin-induced inflammatory and collagen deposition, and inhibited the PF in mice. This work was supported by The Innovation Fund for Institution of Higher Education of Hunan Province (11K077), Hunan Provincial Innovation Foundation for postgraduate (CX2011B048), and the Open-End Fund for the Valuable and Precision Instrument of Central South University (CSUZC2012003).