The microRNA miR-17 regulates lung FoxA1 expression during lipopolysaccharide-induced acute lung injury

Acute lung injury (ALI) is a severe pulmonary disease that causes a high number of fatalities worldwide. Studies have shown that FoxA1 expression is upregulated during ALI and may play an important role in ALI by promoting the apoptosis of alveolar type II epithelial cells. However, the mechanism of FoxA1 overexpression in ALI is unclear. In this study, an in vivo murine model of ALI and alveolar type II epithelial cells injury was induced using lipopolysaccharide (LPS). LPS upregulated FoxA1 in the lung tissue of the in vivo ALI model and in LPS-challenged type II epithelial cells. In contrast, miR-17 was significantly downregulated in these models. After miR-17 antagomir injection, the expression of FoxA1 was significantly increased in ALI mice. MiR-17 mimics could significantly inhibit FoxA1 mRNA and protein expression, whereas the miR-17 inhibitor could significantly increase FoxA1 mRNA and protein expression in LPS-induced type II epithelial cells. Thus, our results suggest that the downregulation of miR-17 expression could lead to FoxA1 overexpression in ALI.     

Anti-inflammatory effects of anethole in lipopolysaccharide-induced acute lung injury in mice

Aims

Anethole, the major component of the essential oil of star anise, has been reported to have antioxidant, antibacterial, antifungal, anti-inflammatory, and anesthetic properties. In this study, we investigated the anti-inflammatory effects of anethole in a mouse model of acute lung injury induced by lipopolysaccharide (LPS).

Main methods

BALB/C mice were intraperitoneally administered anethole (62.5, 125, 250, or 500 mg/kg) 1 h before intratracheal treatment with LPS (1.5 mg/kg) and sacrificed after 4 h. The anti-inflammatory effects of anethole were assessed by measuring total protein and cell levels and inflammatory mediator production and by histological evaluation and Western blot analysis.

Key findings

LPS significantly increased total protein levels; numbers of total cells, including macrophages and neutrophils; and the production of inflammatory mediators such as matrix metalloproteinase 9 (MMP-9), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and nitric oxide (NO) in bronchoalveolar lavage fluid. Anethole (250 mg/kg) decreased total protein concentrations; numbers of inflammatory cells, including neutrophils and macrophages; and the inflammatory mediators MMP-9, TNF-α and NO. In addition, pretreatment with anethole decreased LPS-induced histopathological changes. The anti-inflammatory mechanism of anethole in LPS-induced acute lung injury was assessed by investigating the effects of anethole on NF-κB activation. Anethole suppressed the activation of NF-κB by blocking IκB-α degradation.

Significance

These results, showing that anethole prevents LPS-induced acute lung inflammation in mice, suggest that anethole may be therapeutically effective in inflammatory conditions in humans.     

Silencing of Paralemmin-3 Protects Mice from lipopolysaccharide-induced acute lung injury

Excessive inflammatory response induced by lipopolysaccharide (LPS) plays a critical role in the development of acute lung injury (ALI). Paralemmin-3 (PALM3) is a novel protein that can modulate LPS-stimulated inflammatory responses in alveolar epithelial A549 cells. However, it remains unclear whether it is involved in the progression of ALI in vivo. Therefore, we studied the role of PALM3 in the pathogenesis of ALI induced by LPS. ALI was induced by LPS peritoneal injection in C57BL/6J mice. Lentivirus-mediated small interfering RNA (siRNA) targeting the mouse PALM3 gene and a negative control siRNA were intranasally administered to the mice. We found that the expression of PALM3 was up-regulated in the lung tissues obtained from the mouse model of LPS-induced ALI. The LPS-evoked inflammatory response (neutrophils and the concentrations of proinflammatory cytokines [IL-6, IL-1β, TNF-α, MIP-2] in the bronchoalveolar lavage fluid [BALF]), histologic lung injury (lung injury score), permeability of the alveolar capillary barrier (lung wet/dry weight ratio and BALF protein concentration) and mortality rates were attenuated in the PALM3 siRNA-treated mice. These results indicate that PALM3 contributes to the development of ALI in mice challenged with LPS. Inhibiting PALM3 through the intranasal application of specific siRNA protected against LPS-induced ALI.     

Regulation of Na,K-ATPase during acute lung injury

A hallmark of acute lung injury is the accumulation of a protein rich edema which impairs gas exchange and leads to hypoxemia. The resolution of lung edema is effected by active sodium transport, mostly contributed by apical Na⁺ channels and the basolateral located Na,K-ATPase. It has been reported that the decrease of Na,K-ATPase function seen during lung injury is due to its endocytosis from the cell plasma membrane into intracellular pools. In alveolar epithelial cells exposed to severe hypoxia, we have reported that increased production of mitochondrial reactive oxygen species leads to Na,K-ATPase endocytosis and degradation. We found that this regulated process follows what is referred as the Phosphorylation–Ubiquitination–Recognition–Endocytosis–Degradation (PURED) pathway. Cells exposed to hypoxia generate reactive oxygen species which activate PKCζ which in turn phosphorylates the Na,K-ATPase at the Ser18 residue in the N-terminus of the α1-subunit leading the ubiquitination of any of the four lysines (K16, K17, K19, K20) adjacent to the Ser18 residue. This process promotes the α1-subunit recognition by the μ2 subunit of the adaptor protein-2 and its endocytosis trough a clathrin dependent mechanism. Finally, the ubiquitinated Na,K-ATPase undergoes degradation via a lysosome/proteasome dependent mechanism.     

胃饥饿素对小鼠急性肺损伤的保护作用及其机制研究

目的 探讨胃饥饿素对小鼠急性肺损伤的保护作用和机制.方法 将60只小鼠采用随机数字表法分为6组:对照组、模型组、胃饥饿素低、中、高剂量组和地塞米松组.对照组和模型组腹腔注射0.2 mL生理盐水,胃饥饿素各组分别注射400、200、100 μg/kg溶液,地塞米松组注射2 mg/kg.给药后1h,对照组滴注等体积生理盐水...     

Protective effects of recombinant 53-kDa protein of Trichinella spiralis on acute lung injury in mice via alleviating lung pyroptosis by promoting M2 macrophage polarization

Trichinella spiralis represents an effective treatment for autoimmune and inflammatory diseases. The effects of recombinant T. spiralis (TS) 53-kDa protein (rTsP53) on acute lung injury (ALI) remain unclear. Here, mice were divided randomly into a control group, LPS group, and rTsP53 + LPS group. ALI was induced in BALB/c mice by LPS (10 mg/kg) injected via the tail vein. rTsP53 (200 µl; 0.4 μg/μl) was injected subcutaneously three times at an interval of 5 d before inducing ALI in the rTsP53+LPS group. Lung pathological score, the ratio and markers of classic activated macrophages (M1) and alternatively activated macrophages (M2), cytokine profiles in alveolar lavage fluid, and pyroptosis protein expression in lung tissue were investigated. RTsP53 decreased lung pathological score. Furthermore, rTsP53 suppressed inflammation by increasing IL-4, IL-10, and IL-13. There was an increase in alveolar M2 macrophage numbers, with an increase in CD206 and arginase-1-positive cells and a decrease in alveolar M1 markers such as CD197 and iNOS. In addition, the polarization of M2 macrophages induced by rTsP53 treatment could alleviate ALI by suppressing lung pyroptosis. RTsP53 was identified as a potential agent for treating LPS-induced ALI via alleviating lung pyroptosis by promoting M2 macrophage polarization.     

Upregulation of hypoxia-induced mitogenic factor in bacterial lipopolysaccharide-induced acute lung injury

Hypoxia-induced mitogenic factor (HIMF), also known as FIZZ1 (found in inflammatory zone), plays important roles in lung inflammation. We found that intraperitoneal injection of lipopolysaccharide (LPS) induced intensive HIMF production exclusively in mouse lung, but not in the heart, liver, spleen or kidney. This HIMF production, at least partly, contributes to LPS-induced vascular cell adhesion molecule-1 (VCAM-1) upregulation and mononuclear cell sequestration to lung parenchyma, while protecting alveolar type II cells from LPS-resulted decrease in surfactant protein-C production and cell death. These data indicate that HIMF participates in LPS-induced acute lung injury and inflammation through modulating VCAM-1 and SP-C expression.     

血必净对大鼠急性肺损伤的保护作用

目的：研究血必净对急性肺损伤大鼠的保护作用。方法：60只Wistar大鼠随机分为正常组、模型组、地塞米松（10 mg/kg）组与血必净低、中、高剂量（5、10、15 ml/kg）组,每组10只。通过腹腔注射5 mg/kg内毒素建立大鼠急性肺损伤模型,模型成功4 h后腹腔注射给药,每天1次,连续7 d;正常对照组和ALI模型组静脉注射等体积的生理盐水。7 d后采集动脉血,检测动脉血氧分压（PaO₂）、血清丙二醛（MDA）浓度和超氧化物歧化酶（SOD）的活性;取肺组织,检测肺系数（LI）、左肺湿/干质量比（W/D）、肺含水率[（W-D）/W],检测肿瘤坏死因子-α（TNF-α）、白细胞介素-10（IL-10）和高迁移率族蛋白-1（HMGB1）蛋白的表达。结果：与正常对照组比较,模型组大鼠LI、W/D、（W-D）/W,TNF-α和HMGB1蛋白表达以及血清MDA含量升高,PaO₂,IL-10蛋白表达和血清SOD活性减弱,差异有统计学意义（P < 0.01）;与模型组比较,血必净低、中、高剂量组大鼠LI、W/D及（W-D）/W,TNF-α和HMGB1蛋白表达以及血清MDA含量降低,PaO₂,IL-10蛋白表达和血清SOD活性增强,差异有统计学意义（P < 0.01）,其中血必净高剂量组效果较好,与中、低剂量组比较,差异有统计学意义（P < 0.05,P < 0.05）。结论：血必净能减轻对内毒素诱导的急性肺损伤,其药理机制可能与下调TNF-α和HMGB1蛋白表达和血清MDA水平和上调IL-10蛋白表达和血清SOD活性有关,且以高剂量组效果较好。     

ST2 protein induced by inflammatory stimuli can modulate acute lung inflammation

We have investigated gene and protein expression of ST2/ST2L in a murine alveolar macrophage (AM) cell line, MH-S, reacting to inflammatory stimuli in vitro and in the lung tissue of an acute lung injury model in vivo. We have also analyzed the effect of soluble ST2 protein on inflammatory response of MH-S cells. Lipopolysaccharide (LPS) and proinflammatory cytokines such as IL-1beta, IL-6, and TNF-alpha induced ST2 mRNA expression in MH-S cells. In an acute lung injury model, protein and mRNA expression levels of ST2 increased to the maximal level at 24-72h after the LPS challenge. Furthermore, pretreatment with ST2 protein significantly reduced the protein production and gene expression of IL-1alpha, IL-6, and TNF-alpha in LPS-stimulated MH-S cells in vitro. These results suggest that increases in endogenous ST2 protein in AM, which is induced by inflammatory stimuli, such as LPS and proinflammatory cytokines, may modulate acute lung inflammation.     

一氧化碳吸入对脂多糖诱导大鼠急性肺损伤的保护作用

血红素氧合酶(heme oxygenase,HO)降解血红素的主要代谢产物一氧化碳(carbon monoxide,CO)具有抗氧化、抗炎症和抑制细胞凋亡作用,而脂多糖(lipopolysaccharide,LPS)诱导的肺组织过氧化、炎症性损伤及大量肺泡上皮和血管内皮细胞凋亡正是导致肺损伤(lung injury,LI)的关键.由此我们猜想,CO有可能通过上述机制对LI起保护作用.通过静脉注入LPS(5 mg/kg体重)诱导大鼠LI,观察吸入室内空气或2.5×10-4(V/V)CO 3 h后,肺氧化酶学、炎症细胞因子、细胞凋亡、HO-1表达及组织形态学变化.结果显示,静脉注入LPS诱导LI后,CO吸入组大鼠肺肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)、白细胞介素6(interlukin-6,IL-6)、丙二醛(maleic dialdehyde,MDA)、髓过氧化物酶(myeloperoxidase,MPO)和细胞凋亡分别为(0.91±0.25)pg/mg蛋白、(0.64±0.05)pg/mg蛋白、(1.02±0.23)nmol/mg蛋白、(7.18±1.62)U/mg蛋白、(1.60±0.34)%,均显著低于LI组的(1.48±0.23)pg/mg蛋白、(1.16±0.26)pg/mg蛋白、(1.27+0.33)nmol/mg蛋白、(8.16+1.49)U/mg蛋白、(3.18±0.51)%(P＜0.05).CO吸入组HO-1、白细胞介素10(interlukin-10,IL-10)表达和超氧化物歧化酶(superoxide dismutase,SOD)活性分别为(5.43±0.92)、(0.26±0.07)pg/mg蛋白、(60.09±10.21)U/mg蛋白,它们均显著高于LI组的(3.08±0.82)、(0.15±0.03)pg/mg蛋白、(50.98±6.88)U/mg蛋白(P＜0.05).与LI组相比,CO吸入组肺损伤减轻.研究结果表明,低浓度CO吸入通过抗氧化、抗炎症、抑制细胞凋亡、上调HO-1表达而减轻LPS诱导的肺损伤.     

Anti-inflammatory effects of novel curcumin analogs in experimental acute lung injury

Background

Acute lung injury (ALI) and its most severe form acute respiratory distress syndrome (ARDS) have been the leading cause of morbidity and mortality in intensive care units (ICU). Currently, there is no effective pharmacological treatment for acute lung injury. Curcumin, extracted from turmeric, exhibits broad anti-inflammatory properties through down-regulating inflammatory cytokines. However, the instability of curcumin limits its clinical application.

Methods

A series of new curcumin analogs were synthesized and screened for their inhibitory effects on the production of TNF-α and IL-6 in mouse peritoneal macrophages by ELISA. The evaluation of stability and mechanism of active compounds was determined using UV-assay and Western Blot, respectively. In vivo, SD rats were pretreatment with c26 for seven days and then intratracheally injected with LPS to induce ALI. Pulmonary edema, protein concentration in BALF, injury of lung tissue, inflammatory cytokines in serum and BALF, inflammatory cell infiltration, inflammatory cytokines mRNA expression, and MAPKs phosphorylation were analyzed. We also measured the inflammatory gene expression in human pulmonary epithelial cells.

Results

In the study, we synthesized 30 curcumin analogs. The bioscreeening assay showed that most compounds inhibited LPS-induced production of TNF-α and IL-6. The active compounds, a17, a18, c9 and c26, exhibited their anti-inflammatory activity in a dose-dependent manner and exhibited greater stability than curcumin in vitro. Furthermore, the active compound c26 dose-dependently inhibited ERK phosphorylation. In vivo, LPS significantly increased protein concentration and number of inflammatory cells in BALF, pulmonary edema, pathological changes of lung tissue, inflammatory cytokines in serum and BALF, macrophage infiltration, inflammatory gene expression, and MAPKs phosphorylation . However, pretreatment with c26 attenuated the LPS induced increase through ERK pathway in vivo. Meanwhile, compound c26 reduced the LPS-induced inflammatory gene expression in human pulmonary epithelial cells.

Conclusions

These results suggest that the novel curcumin analog c26 has remarkable protective effects on LPS-induced ALI in rat. These effects may be related to its ability to suppress production of inflammatory cytokines through ERK pathway. Compound c26, with improved chemical stability and bioactivity, may have the potential to be further developed into an anti-inflammatory candidate for the prevention and treatment of ALI.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0199-1) contains supplementary material, which is available to authorized users.     

环孢菌素A对脂多糖诱导的小鼠急性肺损伤的保护作用

目的：探讨线粒体渗透性转换孔道抑制剂环孢菌素A（CsA）对脂多糖（LPS）诱导的小鼠急性肺损伤可能的保护作用。方法：LPS 4 mg/kg气管内滴入复制小鼠急性肺损伤模型,实验随机分为5组（n=24）：分别为正常对照组、LPS组、地塞米松组、CsA组和CsA＋苍术苷组。6 h后小鼠处死,测定各组支气管肺泡灌洗液乳酸脱氢酶（LDH）的含量,酶联免疫吸附法测定肺组织匀浆液TNF-α浓度,测定肺组织湿/干重比和肺毛细血管通透性指数。结果：气管内滴入LPS 6 h后,CsA组与LPS组相比,肺泡灌洗液中LDH活性降低,肺组织匀浆液TNF-α浓度下降,肺组织湿/干重比、肺毛细血管通透性指数均明显下降,但CsA＋Atr组与LPS组相比无明显区别。结论：环孢菌素A对脂多糖诱导的小鼠急性肺损伤有保护作用,其机制可能与其抑制线粒体渗透性转换孔道的开放有关。     

褪黑素对内毒素致大鼠急性肺损伤的保护作用

目的:研究内毒素(LPS)致大鼠急性肺损伤(ALI)时,p-p38蛋白激酶(p-p38MAPK)在肺组织的表达及褪黑素(MT)对肺组织的保护作用及其机制。方法:将72只健康雄性SD大鼠随机分为3组,每组24只,对照组(Control)、模型组(LPS)和褪黑素干预组(LPS+MT),采用气管内滴注LPS的方法建立大鼠ALI的模型,通过免疫组织化学染色和Western blot技术检测大鼠肺组织中p-p38蛋白激酶的表达变化,并在光镜下观察大鼠肺组织形态学变化。结果:Control组气道和肺组织可见反应极弱的p-p38蛋白激酶阳性细胞,散在分布于气道上皮细胞和肺泡上皮细胞;LPS组p-p38蛋白激酶阳性细胞较对照组明显增多(P0.05或P0.01),主要分布于浸润的炎症细胞、气道上皮细胞、肺泡上皮细胞和血管内皮细胞;LPS+MT组气道和肺组织中阳性细胞数较LPS组明显减少(P0.05或P0.01),Western blot结果与免疫组织化学一致。结论:LPS致大鼠急性肺损伤模型中,肺内炎性、非炎性细胞均有p38MAPK信号通路的激活;MT对急性肺损伤的保护机制可能与其抑制p38 MAPK信号通路的过度激活有关。     

Losartan, an antagonist of AT1 receptor for angiotensin II, attenuates lipopolysaccharide-induced acute lung injury in rat

Angiotensin II (Ang II) plays an important role in inflammatory process. Acute lung injury (ALI), an inflammatory disorder of the lung, is commonly associated with endotoxemia; however, the mechanism that endotoxin (lipopolysaccharide, LPS) induces the inflammatory response in ALI is not well defined. Here, we showed, in LPS-induced ALI rat model, that Ang II and Ang II type 1 (AT₁) receptor were significantly increased in lung tissues, compared with those in controls. Meanwhile, nuclear factor (NF)-κB-DNA-binding activity, tumor necrosis factor (TNF)-α mRNA, and pneumocytic apoptosis were significantly increased. Moreover, pretreatment of rats with losartan, an antagonist of AT₁ receptor for Ang II, improved the inflammation, reduced the elevation of Ang II and AT₁ receptor, and inhibited NF-κB-DNA-binding activity, expression of TNF-α mRNA, and pneumocytic apoptosis. The data indicate that Ang II may mediate the inflammatory process in LPS-induced ALI through AT₁ receptor, which can be blocked by losartan.     

Protective effect of Apremilast against LPS-induced acute lung injury via modulation of oxidative stress and inflammation

Lung injuries are attributed due to exposure to Drugs or chemicals. One of the important challenging situations for the clinicians is to manage treatments of different diseases with acute lung injury (ALI). The objective of this study was to investigate the possible protective mechanisms and action of a novel Phosphodiesterase-4 inhibitor “Apremilast” (AP) in lipopolysaccharide (LPS)-induced lung injury. Blood sample from each animals were collected in a vacuum blood collection tube. The rat lungs were isolated for oxidative stress assessment, western blot analysis and their mRNA expressions using RT-PCR. Exposure of LPS in rats causes significant increase in oxidative stress, activates the pro-inflammatory cytokines release like tissue necrotic factor-alpha (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6), modulated gene expression, protein expression and histopathological changes which were reversed by administration of AP. Finding of the research enlighten the protective role of AP against LPS-induced ALI.     

复方清下汤对脓毒症大鼠细胞因子白介素-1、白介素-6调控及肺损伤的作用

目的盲肠结扎穿孔导致大肠埃希菌腹膜炎进而建立脓毒症肺损伤大鼠模型,检测炎性反应时,细胞因子的调控变化,探讨肺水肿的形成机制。经复方清下汤处理后检测上述变化。方法将健康SD大鼠随机分为4组,每组10只:假手术组(SHAM组),只翻动盲肠,不做其他处理;脓毒血症肺损伤组(模型组),盲肠结扎穿孔诱发急性肺损伤(ALI)模型;盲肠结扎穿孔+复方清下汤组(造模后立即灌胃给药,造模后8 h再次灌胃1次,剂量:10 ml/kg);盲肠结扎穿孔+头孢哌酮舒巴坦组(抗生素舒普深)(造模后立即静脉注射1次,造模后8 h再次静脉注射1次,剂量:0.2 g/kg),造模24 h后收集标本。分别观察大鼠的一般状态,留取下腔静脉血清进行白介素-1(IL-1)、白介素-6(IL-6)的测定。镜下观察肺组织病理形态学改变,测量肺湿/干比值的变化。结果与SHAM组比较,模型组IL-1、IL-6水平明显升高(P0.01),肺间质和肺泡内水肿,伴大量红细胞渗出(出血)和纤维素沉积,肺泡间隔毛细血管内皮细胞高度肿胀。肺湿/干比值明显增加(P0.01),抗生素及中药处理组与模型组比较,IL-1、IL-6水平明显降低(P0.01),肺湿/干比值明显降低(P0.01),肺组织镜下表现:中药处理组及抗生素组组较模型组肺泡间隔变窄,毛细血管内皮细胞肿胀减轻,出血减轻,纤维素渗出明显减少。结论实验应用放免检测、显微镜观察以及称量肺湿/干比值等手段,进一步证实了脓毒血症大鼠肺损伤时血清中主要的炎性细胞因子IL-1、IL-6过度表达的情况,并从病理学角度,证实炎性介质的过度表达是造成脓毒症肺损伤的重要原因。经复方清下汤处理的动物模型得到相反结论,为治疗脓毒血症大鼠肺损伤提供一个可能的新的手段。     

Inhibition of glycolysis alleviates lipopolysaccharide-induced acute lung injury in a mouse model

Gluconic metabolic reprogramming, immune response, and inflammation are intimately linked. Glycolysis involves in the pathologic progress in acute and chronic inflammatory diseases. However, the involvement of glycolysis in the acute lung injury (ALI) is still unclear. This study investigated the role of glycolysis in an animal model of ALI. First, we found that lactate content in serum was remarkably increased in ALI patients and a murine model induced by intratracheal administration of lipopolysaccharide (LPS). The key proteins involving in glycolysis were robustly elevated, including HK2, PKM2, and HIF-1α. Intriguingly, inhibition of glycolysis by 2-deoxyglucose (2-DG) pronouncedly attenuated the lung tissue pathological injury, accumulation of neutrophil, oxidative stress, expression of proinflammatory factors in the lung of ALI mice induced by LPS. The 2-DG treatment also strongly suppressed the activation of the NOD-like receptor (NLR) family and pyrin domain-containing protein 3 (NLRP3) inflammasome. Furthermore, we investigated the role of glycolysis in the inflammatory response of primary murine macrophages activated by LPS in vitro. We found that the 2-DG treatment remarkably reduced the expression of proinflammatory factors induced by LPS, including tumor necrosis factor-α messenger RNA (mRNA), pro-interleukin (IL)-1β mRNA, pro-IL-18 mRNA, NLRP3 mRNA, caspase-1 mRNA, and IL-1β protein. Altogether, these data provide a novel link between gluconic metabolism reprogramming and uncontrolled inflammatory response in ALI. This study suggests glycolytic inhibition as an effective anti-inflammatory strategy in treating ALI.     

Toll-like receptor 4 mediates lipopolysaccharide-induced collagen secretion by phosphoinositide3-kinase-akt pathway in fibroblasts during acute lung injury

Gram-negative bacillus infection is an important risk factor of acute lung injury (ALI). Previous experiments have revealed that lipopolysaccharide (LPS), a primary component of endotoxin of gram-negative bacilli, stimulated the inflammatory reactions that contribute to ALI and pulmonary interstitial fibrosis, but the mechanisms were not well understood. We reported that LPS was able to directly induce secretion of collagen in mouse lung fibroblasts via activation of phosphoinositide3-kinase-Akt (PI3K-Akt) pathway through toll-like receptor 4 (TLR4) in vitro. We found that overexpression of TLR4, type I procollagen, alpha smooth muscle actin (alpha-SMA), and p-AKT in primary cultured mouse lung fibroblast stimulated by LPS were detected by real-time PCR or Western blots, and the contents of C-terminal propeptide of type I procollagen (PICP) in cell culture supernatants were increased simultaneously. The activation of TLR4 stimulated by LPS could also up-regulate the expression of integrin beta1 and TLR4 in mouse lung fibroblast, which could accelerate ALI and pulmonary interstitial fibrosis processes. All these changes could be inhabited by transfection of Lentivirus-TLR4-siRNA or application of PI3K inhibitor LY294002. Therefore, we infer that besides pulmonary macrophage, lung fibroblasts are also important target cells directly influenced by LPS, which may play an important role in ALI and pulmonary interstitial fibrosis.     

Beneficial effects of ApoA-I on LPS-induced acute lung injury and endotoxemia in mice

High density lipoprotein (HDL) binds lipopolysaccharide (LPS) and neutralizes its toxicity. The aim of our study was to investigate the effects of Apolipoprotein (ApoA-I), the major apolipoprotein of HDL, on LPS-induced acute lung injury (ALI) and endotoxemia. BALB/c mice were challenged with LPS, followed by ApoA-I or saline administration for 24h. The mice were then sacrificed and histopathological analysis of the lung was performed. We found that ApoA-I could attenuate LPS-induced acute lung injury and inflammation. To investigate the mechanisms, we measured tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) levels in the serum and bronchoalveolar lavage (BAL) fluid and found that ApoA-I could significantly inhibit LPS-induced increases in the IL-1beta and TNF-alpha levels in serum (P<0.05, respectively), as well as in the IL-1beta, TNF-alpha, and IL-6 levels in BAL fluid (P<0.01 and P<0.05, P<0.05, respectively). Moreover, we evaluated the effect of ApoA-I on the mortality of L-929 cells which were attacked by LPS-activated peritoneal macrophages. We found that ApoA-I could significantly inhibit the LPS-induced cell death in a dose-dependent fashion. Furthermore, we investigated in vivo the effects of ApoA-I on the mortality rate and survival time after LPS administration and found that ApoA-I significantly decreased the mortality (P<0.05) and increased the survival time (P<0.05). In summary, the results suggest that ApoA-I could effectively protect against LPS-induced endotoxemia and acute lung damage. The mechanism might be related to inhibition of inflammatory cytokine release from macrophages.